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Sample records for biofilms reveals distinct

  1. Achromobacter Species Isolated from Cystic Fibrosis Patients Reveal Distinctly Different Biofilm Morphotypes

    DEFF Research Database (Denmark)

    Nielsen, Signe M; Nørskov-Lauritsen, Niels; Bjarnsholt, Thomas

    2016-01-01

    findings were visualized in sputum samples from cystic fibrosis patients using an Achromobacter specific peptide nucleic acid fluorescence in situ hybridization (PNA-FISH) probe, confirming the presence of Achromobacter biofilms in the CF lung. High antibiotic tolerance was associated with the biofilm...

  2. Dynamic remodeling of microbial biofilms by functionally distinct exopolysaccharides.

    Science.gov (United States)

    Chew, Su Chuen; Kundukad, Binu; Seviour, Thomas; van der Maarel, Johan R C; Yang, Liang; Rice, Scott A; Doyle, Patrick; Kjelleberg, Staffan

    2014-08-05

    Biofilms are densely populated communities of microbial cells protected and held together by a matrix of extracellular polymeric substances. The structure and rheological properties of the matrix at the microscale influence the retention and transport of molecules and cells in the biofilm, thereby dictating population and community behavior. Despite its importance, quantitative descriptions of the matrix microstructure and microrheology are limited. Here, particle-tracking microrheology in combination with genetic approaches was used to spatially and temporally study the rheological contributions of the major exopolysaccharides Pel and Psl in Pseudomonas aeruginosa biofilms. Psl increased the elasticity and effective cross-linking within the matrix, which strengthened its scaffold and appeared to facilitate the formation of microcolonies. Conversely, Pel reduced effective cross-linking within the matrix. Without Psl, the matrix becomes more viscous, which facilitates biofilm spreading. The wild-type biofilm decreased in effective cross-linking over time, which would be advantageous for the spreading and colonization of new surfaces. This suggests that there are regulatory mechanisms to control production of the exopolysaccharides that serve to remodel the matrix of developing biofilms. The exopolysaccharides were also found to have profound effects on the spatial organization and integration of P. aeruginosa in a mixed-species biofilm model of P. aeruginosa-Staphylococcus aureus. Pel was required for close association of the two species in mixed-species microcolonies. In contrast, Psl was important for P. aeruginosa to form single-species biofilms on top of S. aureus biofilms. Our results demonstrate that Pel and Psl have distinct physical properties and functional roles during biofilm formation. Importance: Most bacteria grow as biofilms in the environment or in association with eukaryotic hosts. Removal of biofilms that form on surfaces is a challenge in clinical

  3. Distinct roles of extracellular polymeric substances in Pseudomonas aeruginosa biofilm development

    DEFF Research Database (Denmark)

    Yang, Liang; Hu, Yifan; Liu, Yang

    2011-01-01

    Bacteria form surface attached biofilm communities as one of the most important survival strategies in nature. Biofilms consist of water, bacterial cells and a wide range of self‐generated extracellular polymeric substances (EPS). Biofilm formation is a dynamic self‐assembly process and several d...... polysaccharide is more important than Pel polysaccharide in P. aeruginosa PAO1 biofilm formation and antibiotic resistance. Our study thus suggests that different EPS materials play distinct roles during bacterial biofilm formation.......Bacteria form surface attached biofilm communities as one of the most important survival strategies in nature. Biofilms consist of water, bacterial cells and a wide range of self‐generated extracellular polymeric substances (EPS). Biofilm formation is a dynamic self‐assembly process and several...

  4. In Situ Biomineralization and Particle Deposition Distinctively Mediate Biofilm Susceptibility to Chlorine.

    Science.gov (United States)

    Li, Xiaobao; Chopp, David L; Russin, William A; Brannon, Paul T; Parsek, Matthew R; Packman, Aaron I

    2016-05-15

    Microbial biofilms and mineral precipitation commonly co-occur in engineered water systems, such as cooling towers and water purification systems, and both decrease process performance. Microbial biofilms are extremely challenging to control and eradicate. We previously showed that in situ biomineralization and the precipitation and deposition of abiotic particles occur simultaneously in biofilms under oversaturated conditions. Both processes could potentially alter the essential properties of biofilms, including susceptibility to biocides. However, the specific interactions between mineral formation and biofilm processes remain poorly understood. Here we show that the susceptibility of biofilms to chlorination depends specifically on internal transport processes mediated by biomineralization and the accumulation of abiotic mineral deposits. Using injections of the fluorescent tracer Cy5, we show that Pseudomonas aeruginosa biofilms are more permeable to solutes after in situ calcite biomineralization and are less permeable after the deposition of abiotically precipitated calcite particles. We further show that biofilms are more susceptible to chlorine killing after biomineralization and less susceptible after particle deposition. Based on these observations, we found a strong correlation between enhanced solute transport and chlorine killing in biofilms, indicating that biomineralization and particle deposition regulate biofilm susceptibility by altering biocide penetration into the biofilm. The distinct effects of in situ biomineralization and particle deposition on biocide killing highlight the importance of understanding the mechanisms and patterns of biomineralization and scale formation to achieve successful biofilm control.

  5. Dispersed cells represent a distinct stage in the transition from bacterial biofilm to planktonic lifestyles

    DEFF Research Database (Denmark)

    Chua, Song Lin; Liu, Yang; Yam, Joey Kuok Hoong

    2014-01-01

    Bacteria assume distinct lifestyles during the planktonic and biofilm modes of growth. Increased levels of the intracellular messenger c-di-GMP determine the transition from planktonic to biofilm growth, while a reduction causes biofilm dispersal. It is generally assumed that cells dispersed from...... biofilms immediately go into the planktonic growth phase. Here we use single-nucleotide resolution transcriptomic analysis to show that the physiology of dispersed cells from Pseudomonas aeruginosa biofilms is highly different from those of planktonic and biofilm cells. In dispersed cells, the expression...... of the small regulatory RNAs RsmY and RsmZ is downregulated, whereas secretion genes are induced. Dispersed cells are highly virulent against macrophages and Caenorhabditis elegans compared with planktonic cells. In addition, they are highly sensitive towards iron stress, and the combination of a biofilm...

  6. Transcriptomic and proteomic analyses of Desulfovibrio vulgaris biofilms: Carbon and energy flow contribute to the distinct biofilm growth state

    Directory of Open Access Journals (Sweden)

    Clark Melinda E

    2012-04-01

    extracellular fraction from biofilm cells. Conclusions Even though both the planktonic and biofilm cells were oxidizing lactate and reducing sulfate, the biofilm cells were physiologically distinct compared to planktonic growth states due to altered abundances of genes/proteins involved in carbon/energy flow and extracellular structures. In addition, average expression values for multiple rRNA transcripts and respiratory activity measurements indicated that biofilm cells were metabolically more similar to exponential-phase cells although biofilm cells are structured differently. The characterization of physiological advantages and constraints of the biofilm growth state for sulfate-reducing bacteria will provide insight into bioremediation applications as well as microbially-induced metal corrosion.

  7. Transcriptomic and proteomic analyses of Desulfovibrio vulgaris biofilms: carbon and energy flow contribute to the distinct biofilm growth state.

    Science.gov (United States)

    Clark, Melinda E; He, Zhili; Redding, Alyssa M; Joachimiak, Marcin P; Keasling, Jay D; Zhou, Jizhong Z; Arkin, Adam P; Mukhopadhyay, Aindrila; Fields, Matthew W

    2012-04-16

    . Even though both the planktonic and biofilm cells were oxidizing lactate and reducing sulfate, the biofilm cells were physiologically distinct compared to planktonic growth states due to altered abundances of genes/proteins involved in carbon/energy flow and extracellular structures. In addition, average expression values for multiple rRNA transcripts and respiratory activity measurements indicated that biofilm cells were metabolically more similar to exponential-phase cells although biofilm cells are structured differently. The characterization of physiological advantages and constraints of the biofilm growth state for sulfate-reducing bacteria will provide insight into bioremediation applications as well as microbially-induced metal corrosion.

  8. Patterned biofilm formation reveals a mechanism for structural heterogeneity in bacterial biofilms.

    Science.gov (United States)

    Gu, Huan; Hou, Shuyu; Yongyat, Chanokpon; De Tore, Suzanne; Ren, Dacheng

    2013-09-03

    Bacterial biofilms are ubiquitous and are the major cause of chronic infections in humans and persistent biofouling in industry. Despite the significance of bacterial biofilms, the mechanism of biofilm formation and associated drug tolerance is still not fully understood. A major challenge in biofilm research is the intrinsic heterogeneity in the biofilm structure, which leads to temporal and spatial variation in cell density and gene expression. To understand and control such structural heterogeneity, surfaces with patterned functional alkanthiols were used in this study to obtain Escherichia coli cell clusters with systematically varied cluster size and distance between clusters. The results from quantitative imaging analysis revealed an interesting phenomenon in which multicellular connections can be formed between cell clusters depending on the size of interacting clusters and the distance between them. In addition, significant differences in patterned biofilm formation were observed between wild-type E. coli RP437 and some of its isogenic mutants, indicating that certain cellular and genetic factors are involved in interactions among cell clusters. In particular, autoinducer-2-mediated quorum sensing was found to be important. Collectively, these results provide missing information that links cell-to-cell signaling and interaction among cell clusters to the structural organization of bacterial biofilms.

  9. Biofilm formation assessment in Sinorhizobium meliloti reveals interlinked control with surface motility

    OpenAIRE

    Amaya-Gomez, CV; Hirsch, AM; Soto, MJ

    2015-01-01

    Background Swarming motility and biofilm formation are opposite, but related surface-associated behaviors that allow various pathogenic bacteria to colonize and invade their hosts. In Sinorhizobium meliloti, the alfalfa endosymbiont, these bacterial processes and their relevance for host plant colonization are largely unexplored. Our previous work demonstrated distinct swarming abilities in two S. meliloti strains (Rm1021 and GR4) and revealed that both environmental cues (iron concentration)...

  10. Distinct gene expression profile of Xanthomonas retroflexus engaged in synergistic multispecies biofilm formation

    DEFF Research Database (Denmark)

    Hansen, Lea Benedicte Skov; Ren, Dawei; Burmølle, Mette;

    2017-01-01

    It is well known that bacteria often exist in naturally formed multispecies biofilms. Within these biofilms, interspecies interactions seem to have an important role in ecological processes. Little is known about the effects of interspecies interactions on gene expression in these multispecies...... biofilms. This study presents a comparative gene expression analysis of the Xanthomonas retroflexus transcriptome when grown in a single-species biofilm and in dual-and four-species consortia with Stenotrophomonas rhizophila, Microbacterium oxydans and Paenibacillus amylolyticus. The results revealed...... complex interdependent interaction patterns in the multispecies biofilms. Many of the regulated functions are related to interactions with the external environment and suggest a high phenotypic plasticity in response to coexistence with other species. Furthermore, the changed expression of genes involved...

  11. Transcriptomics Analysis Reveals Putative Genes Involved in Biofilm Formation and Biofilm-associated Drug Resistance of Enterococcus faecalis.

    Science.gov (United States)

    Seneviratne, Chaminda J; Suriyanarayanan, Tanujaa; Swarup, Sanjay; Chia, Kuan Hui Burton; Nagarajan, Niranjan; Zhang, Chengfei

    2017-06-01

    Enterococcus faecalis is a gram-positive bacterium associated with endodontic infections and is capable of forming biofilms that can confer drug resistance to the bacterium, resulting in treatment failure. Current knowledge on E. faecalis drug resistance is of a limited and conflicting nature. The present study examined the genetic basis of E. faecalis biofilm formation and drug resistance using a RNA sequencing (RNA-Seq)-based transcriptome approach. Eighteen clinical isolates of E. faecalis were screened for their biofilm formation abilities using the crystal violet assay, colony counting, and confocal imaging. Selected isolates were then evaluated for antibiotic susceptibility in planktonic and biofilm growth modes followed by RNA-Seq analysis of E. faecalis planktonic, biofilm, and vancomycin-treated biofilm samples and Kyoto Encyclopedia of Genes and Genomes mapping in order to identify genes associated with biofilm formation and drug resistance of E. faecalis. All 18 clinical isolates retained biofilm formation ability and were classified as strong, weak, or laboratory American Type Culture Collection strainlike biofilm formers. Interestingly, both the strong and weak biofilm-forming isolates were uniformly resistant to ampicillin and vancomycin at the treated concentrations (256-4096 μg/mL). RNA-Seq analysis of these isolates identified a total of 163 and 101 differentially regulated genes in planktonic versus biofilm and vancomycin-treated biofilm versus biofilm comparisons, respectively, with significant differences in arsenic resistance operon genes arsR and arsD, sporulation regulatory gene paiA, ABC drug transporter classes, and penicillin-binding proteins. The present transcriptomic study revealed putative genes associated with E. faecalis biofilm formation and drug resistance, which will provide a foundation for improved therapeutic strategies against E. faecalis infections in the future. Copyright © 2017 American Association of Endodontists

  12. The transcriptional programme of Salmonella enterica serovar Typhimurium reveals a key role for tryptophan metabolism in biofilms

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    Cochrane Brett

    2009-12-01

    was restored by exogenous tryptophan or indole. Conclusions Biofilm growth of S. Typhimurium causes distinct changes in gene and protein expression. Our results show that aromatic amino acids make an important contribution to biofilm formation and reveal a link between SPI2 expression and surface-associated growth in S. Typhimurium.

  13. The transcriptional programme of Salmonella enterica serovar Typhimurium reveals a key role for tryptophan metabolism in biofilms.

    LENUS (Irish Health Repository)

    Hamilton, Shea

    2009-12-11

    exogenous tryptophan or indole. Conclusions Biofilm growth of S. Typhimurium causes distinct changes in gene and protein expression. Our results show that aromatic amino acids make an important contribution to biofilm formation and reveal a link between SPI2 expression and surface-associated growth in S. Typhimurium.

  14. Vibrio cholerae biofilm growth program and architecture revealed by single-cell live imaging.

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    Yan, Jing; Sharo, Andrew G; Stone, Howard A; Wingreen, Ned S; Bassler, Bonnie L

    2016-09-01

    Biofilms are surface-associated bacterial communities that are crucial in nature and during infection. Despite extensive work to identify biofilm components and to discover how they are regulated, little is known about biofilm structure at the level of individual cells. Here, we use state-of-the-art microscopy techniques to enable live single-cell resolution imaging of a Vibrio cholerae biofilm as it develops from one single founder cell to a mature biofilm of 10,000 cells, and to discover the forces underpinning the architectural evolution. Mutagenesis, matrix labeling, and simulations demonstrate that surface adhesion-mediated compression causes V. cholerae biofilms to transition from a 2D branched morphology to a dense, ordered 3D cluster. We discover that directional proliferation of rod-shaped bacteria plays a dominant role in shaping the biofilm architecture in V. cholerae biofilms, and this growth pattern is controlled by a single gene, rbmA Competition analyses reveal that the dense growth mode has the advantage of providing the biofilm with superior mechanical properties. Our single-cell technology can broadly link genes to biofilm fine structure and provides a route to assessing cell-to-cell heterogeneity in response to external stimuli.

  15. Pseudomonas chlororaphis Produces Two Distinct R-Tailocins That Contribute to Bacterial Competition in Biofilms and on Roots.

    Science.gov (United States)

    Dorosky, Robert J; Yu, Jun Myoung; Pierson, Leland S; Pierson, Elizabeth A

    2017-08-01

    R-type tailocins are high-molecular-weight bacteriocins that resemble bacteriophage tails and are encoded within the genomes of many Pseudomonas species. In this study, analysis of the P. chlororaphis 30-84 R-tailocin gene cluster revealed that it contains the structural components to produce two R-tailocins of different ancestral origins. Two distinct R-tailocin populations differing in length were observed in UV-induced lysates of P. chlororaphis 30-84 via transmission electron microscopy. Mutants defective in the production of one or both R-tailocins demonstrated that the killing spectrum of each tailocin is limited to Pseudomonas species. The spectra of pseudomonads killed by the two R-tailocins differed, although a few Pseudomonas species were either killed by or insusceptible to both tailocins. Tailocin release was disrupted by deletion of the holin gene within the tailocin gene cluster, demonstrating that the lysis cassette is required for the release of both R-tailocins. The loss of functional tailocin production reduced the ability of P. chlororaphis 30-84 to compete with an R-tailocin-sensitive strain within biofilms and rhizosphere communities. Our study demonstrates that Pseudomonas species can produce more than one functional R-tailocin particle sharing the same lysis cassette but differing in their killing spectra. This study provides evidence for the role of R-tailocins as determinants of bacterial competition among plant-associated Pseudomonas in biofilms and the rhizosphere.IMPORTANCE Recent studies have identified R-tailocin gene clusters potentially encoding more than one R-tailocin within the genomes of plant-associated Pseudomonas but have not demonstrated that more than one particle is produced or the ecological significance of the production of multiple R-tailocins. This study demonstrates for the first time that Pseudomonas strains can produce two distinct R-tailocins with different killing spectra, both of which contribute to bacterial

  16. Proteomic profile of Cryptococcus neoformans biofilm reveals changes in metabolic processes.

    Science.gov (United States)

    Santi, Lucélia; Beys-da-Silva, Walter O; Berger, Markus; Calzolari, Diego; Guimarães, Jorge A; Moresco, James J; Yates, John R

    2014-03-07

    Cryptococcus neoformans, a pathogenic yeast, causes meningoencephalitis, especially in immunocompromised patients, leading in some cases to death. Microbes in biofilms can cause persistent infections, which are harder to treat. Cryptococcal biofilms are becoming common due to the growing use of brain valves and other medical devices. Using shotgun proteomics we determine the differences in protein abundance between biofilm and planktonic cells. Applying bioinformatic tools, we also evaluated the metabolic pathways involved in biofilm maintenance and protein interactions. Our proteomic data suggest general changes in metabolism, protein turnover, and global stress responses. Biofilm cells show an increase in proteins related to oxidation-reduction, proteolysis, and response to stress and a reduction in proteins related to metabolic process, transport, and translation. An increase in pyruvate-utilizing enzymes was detected, suggesting a shift from the TCA cycle to fermentation-derived energy acquisition. Additionally, we assign putative roles to 33 proteins previously categorized as hypothetical. Many changes in metabolic enzymes were identified in studies of bacterial biofilm, potentially revealing a conserved strategy in biofilm lifestyle.

  17. Distinct roles of long/short fimbriae and gingipains in homotypic biofilm development by Porphyromonas gingivalis

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    Tribble Gena D

    2009-05-01

    Full Text Available Abstract Background Porphyromonas gingivalis, a periodontal pathogen, expresses a number of virulence factors, including long (FimA and short (Mfa fimbriae as well as gingipains comprised of arginine-specific (Rgp and lysine-specific (Kgp cysteine proteinases. The aim of this study was to examine the roles of these components in homotypic biofilm development by P. gingivalis, as well as in accumulation of exopolysaccharide in biofilms. Results Biofilms were formed on saliva-coated glass surfaces in PBS or diluted trypticase soy broth (dTSB. Microscopic observation showed that the wild type strain formed biofilms with a dense basal monolayer and dispersed microcolonies in both PBS and dTSB. A FimA deficient mutant formed patchy and small microcolonies in PBS, but the organisms proliferated and formed a cohesive biofilm with dense exopolysaccharides in dTSB. A Mfa mutant developed tall and large microcolonies in PBS as well as dTSB. A Kgp mutant formed markedly thick biofilms filled with large clumped colonies under both conditions. A RgpA/B double mutant developed channel-like biofilms with fibrillar and tall microcolonies in PBS. When this mutant was studied in dTSB, there was an increase in the number of peaks and the morphology changed to taller and loosely packed biofilms. In addition, deletion of FimA reduced the autoaggregation efficiency, whereas autoaggregation was significantly increased in the Kgp and Mfa mutants, with a clear association with alteration of biofilm structures under the non-proliferation condition. In contrast, this association was not observed in the Rgp-null mutants. Conclusion These results suggested that the FimA fimbriae promote initial biofilm formation but exert a restraining regulation on biofilm maturation, whereas Mfa and Kgp have suppressive and regulatory roles during biofilm development. Rgp controlled microcolony morphology and biovolume. Collectively, these molecules seem to act coordinately to regulate

  18. Surface Polysaccharide Mutants Reveal that Absence of O Antigen Reduces Biofilm Formation of Actinobacillus pleuropneumoniae.

    Science.gov (United States)

    Hathroubi, S; Hancock, M A; Bossé, J T; Langford, P R; Tremblay, Y D N; Labrie, J; Jacques, M

    2015-10-19

    Actinobacillus pleuropneumoniae is a Gram-negative bacterium belonging to the Pasteurellaceae family and the causative agent of porcine pleuropneumonia, a highly contagious lung disease causing important economic losses. Surface polysaccharides, including lipopolysaccharides (LPS) and capsular polysaccharides (CPS), are implicated in the adhesion and virulence of A. pleuropneumoniae, but their role in biofilm formation is still unclear. In this study, we investigated the requirement for these surface polysaccharides in biofilm formation by A. pleuropneumoniae serotype 1. Well-characterized mutants were used: an O-antigen LPS mutant, a truncated core LPS mutant with an intact O antigen, a capsule mutant, and a poly-N-acetylglucosamine (PGA) mutant. We compared the amount of biofilm produced by the parental strain and the isogenic mutants using static and dynamic systems. Compared to the findings for the biofilm of the parental or other strains, the biofilm of the O antigen and the PGA mutants was dramatically reduced, and it had less cell-associated PGA. Real-time PCR analyses revealed a significant reduction in the level of pgaA, cpxR, and cpxA mRNA in the biofilm cells of the O-antigen mutant compared to that in the biofilm cells of the parental strain. Specific binding between PGA and LPS was consistently detected by surface plasmon resonance, but the lack of O antigen did not abolish these interactions. In conclusion, the absence of the O antigen reduces the ability of A. pleuropneumoniae to form a biofilm, and this is associated with the reduced expression and production of PGA. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  19. Molecular Techniques Revealed Highly Diverse Microbial Communities in Natural Marine Biofilms on Polystyrene Dishes for Invertebrate Larval Settlement

    KAUST Repository

    Lee, On On

    2014-01-09

    Biofilm microbial communities play an important role in the larval settlement response of marine invertebrates. However, the underlying mechanism has yet to be resolved, mainly because of the uncertainties in characterizing members in the communities using traditional 16S rRNA gene-based molecular methods and in identifying the chemical signals involved. In this study, pyrosequencing was used to characterize the bacterial communities in intertidal and subtidal marine biofilms developed during two seasons. We revealed highly diverse biofilm bacterial communities that varied with season and tidal level. Over 3,000 operational taxonomic units with estimates of up to 8,000 species were recovered in a biofilm sample, which is by far the highest number recorded in subtropical marine biofilms. Nineteen phyla were found, of which Cyanobacteria and Proteobacteria were the most dominant one in the intertidal and subtidal biofilms, respectively. Apart from these, Actinobacteria, Bacteroidetes, and Planctomycetes were the major groups recovered in both intertidal and subtidal biofilms, although their relative abundance varied among samples. Full-length 16S rRNA gene clone libraries were constructed for the four biofilm samples and showed similar bacterial compositions at the phylum level to those revealed by pyrosequencing. Laboratory assays confirmed that cyrids of the barnacle Balanus amphitrite preferred to settle on the intertidal rather than subtidal biofilms. This preference was independent of the biofilm bacterial density or biomass but was probably related to the biofilm community structure, particularly, the Proteobacterial and Cyanobacterial groups. © 2014 Springer Science+Business Media New York.

  20. A novel assay of biofilm antifungal activity reveals that amphotericin B and caspofungin lyse Candida albicans cells in biofilms.

    Science.gov (United States)

    DiDone, Louis; Oga, Duana; Krysan, Damian J

    2011-08-01

    The ability of Candida albicans to form drug-resistant biofilms is an important factor in its contribution to human disease. Assays to identify and characterize molecules with activity against fungal biofilms are crucial for the development of drugs with improved anti-biofilm activity. Here we report the application of an adenylate kinase (AK)-based cytotoxicity assay of fungal cell lysis to the characterization of agents active against C. albicans biofilms. We have developed three protocols for the AK assay. The first measures AK activity in the supernatants of biofilms treated with antifungal drugs and can be performed in parallel with a standard 2,3-bis-(2-methoxy-4-nitro-5-sulphophenyl)-2H-tetrazolium-5-caboxanilide-based biofilm susceptibility assay; a second, more sensitive protocol measures the AK activity present within the biofilm matrix; and a third procedure allows the direct visualization of lytic activity toward biofilms formed on catheter material. Amphotericin B and caspofungin, the two most effective anti-biofilm drugs currently used to treat fungal infections, both directly lyse planktonic C. albicans cells in vitro, leading to the release of AK into the culture medium. These studies serve to validate the AK-based lysis assay as a useful addition to the methods for the characterization of antifungal agents active toward biofilms and provide insights into the mode of action of amphotericin B and caspofungin against C. albicans biofilms. Copyright © 2011 John Wiley & Sons, Ltd.

  1. Comparative genome and transcriptome analysis reveals distinctive surface characteristics and unique physiological potentials of Pseudomonas aeruginosa ATCC 27853

    KAUST Repository

    Cao, Huiluo

    2017-06-12

    Pseudomonas aeruginosa ATCC 27853 was isolated from a hospital blood specimen in 1971 and has been widely used as a model strain to survey antibiotics susceptibilities, biofilm development, and metabolic activities of Pseudomonas spp.. Although four draft genomes of P. aeruginosa ATCC 27853 have been sequenced, the complete genome of this strain is still lacking, hindering a comprehensive understanding of its physiology and functional genome.Here we sequenced and assembled the complete genome of P. aeruginosa ATCC 27853 using the Pacific Biosciences SMRT (PacBio) technology and Illumina sequencing platform. We found that accessory genes of ATCC 27853 including prophages and genomic islands (GIs) mainly contribute to the difference between P. aeruginosa ATCC 27853 and other P. aeruginosa strains. Seven prophages were identified within the genome of P. aeruginosa ATCC 27853. Of the predicted 25 GIs, three contain genes that encode monoxoygenases, dioxygenases and hydrolases that could be involved in the metabolism of aromatic compounds. Surveying virulence-related genes revealed that a series of genes that encode the B-band O-antigen of LPS are lacking in ATCC 27853. Distinctive SNPs in genes of cellular adhesion proteins such as type IV pili and flagella biosynthesis were also observed in this strain. Colony morphology analysis confirmed an enhanced biofilm formation capability of ATCC 27853 on solid agar surface compared to Pseudomonas aeruginosa PAO1. We then performed transcriptome analysis of ATCC 27853 and PAO1 using RNA-seq and compared the expression of orthologous genes to understand the functional genome and the genomic details underlying the distinctive colony morphogenesis. These analyses revealed an increased expression of genes involved in cellular adhesion and biofilm maturation such as type IV pili, exopolysaccharide and electron transport chain components in ATCC 27853 compared with PAO1. In addition, distinctive expression profiles of the

  2. Distinct loading conditions reveal various patterns of right ventricular adaptation

    NARCIS (Netherlands)

    Borgdorff, Marinus A. J.; Bartelds, Beatrijs; Dickinson, Michael G.; Steendijk, Paul; de Vroomen, Maartje; Berger, Rolf M. F.

    2013-01-01

    Right ventricular (RV) failure due to chronically abnormal loading is a main determinant of outcome in pulmonary hypertension (PH) and congenital heart disease. However, distinct types of RV loading have been associated with different outcomes. To determine whether the adaptive RV response depends o

  3. A novel technique using potassium permanganate and reflectance confocal microscopy to image biofilm extracellular polymeric matrix reveals non-eDNA networks in Pseudomonas aeruginosa biofilms.

    Science.gov (United States)

    Swearingen, Matthew C; Mehta, Ajeet; Mehta, Amar; Nistico, Laura; Hill, Preston J; Falzarano, Anthony R; Wozniak, Daniel J; Hall-Stoodley, Luanne; Stoodley, Paul

    2016-02-01

    Biofilms are etiologically important in the development of chronic medical and dental infections. The biofilm extracellular polymeric substance (EPS) determines biofilm structure and allows bacteria in biofilms to adapt to changes in mechanical loads such as fluid shear. However, EPS components are difficult to visualize microscopically because of their low density and molecular complexity. Here, we tested potassium permanganate, KMnO4, for use as a non-specific EPS contrast-enhancing stain using confocal laser scanning microscopy in reflectance mode. We demonstrate that KMnO4 reacted with EPS components of various strains of Pseudomonas, Staphylococcus and Streptococcus, yielding brown MnO2 precipitate deposition on the EPS, which was quantifiable using data from the laser reflection detector. Furthermore, the MnO2 signal could be quantified in combination with fluorescent nucleic acid staining. COMSTAT image analysis indicated that KMnO4 staining increased the estimated biovolume over that determined by nucleic acid staining alone for all strains tested, and revealed non-eDNA EPS networks in Pseudomonas aeruginosa biofilm. In vitro and in vivo testing indicated that KMnO4 reacted with poly-N-acetylglucosamine and Pseudomonas Pel polysaccharide, but did not react strongly with DNA or alginate. KMnO4 staining may have application as a research tool and for diagnostic potential for biofilms in clinical samples.

  4. Network analysis reveals distinct clinical syndromes underlying acute mountain sickness.

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    David P Hall

    Full Text Available Acute mountain sickness (AMS is a common problem among visitors at high altitude, and may progress to life-threatening pulmonary and cerebral oedema in a minority of cases. International consensus defines AMS as a constellation of subjective, non-specific symptoms. Specifically, headache, sleep disturbance, fatigue and dizziness are given equal diagnostic weighting. Different pathophysiological mechanisms are now thought to underlie headache and sleep disturbance during acute exposure to high altitude. Hence, these symptoms may not belong together as a single syndrome. Using a novel visual analogue scale (VAS, we sought to undertake a systematic exploration of the symptomatology of AMS using an unbiased, data-driven approach originally designed for analysis of gene expression. Symptom scores were collected from 292 subjects during 1110 subject-days at altitudes between 3650 m and 5200 m on Apex expeditions to Bolivia and Kilimanjaro. Three distinct patterns of symptoms were consistently identified. Although fatigue is a ubiquitous finding, sleep disturbance and headache are each commonly reported without the other. The commonest pattern of symptoms was sleep disturbance and fatigue, with little or no headache. In subjects reporting severe headache, 40% did not report sleep disturbance. Sleep disturbance correlates poorly with other symptoms of AMS (Mean Spearman correlation 0.25. These results challenge the accepted paradigm that AMS is a single disease process and describe at least two distinct syndromes following acute ascent to high altitude. This approach to analysing symptom patterns has potential utility in other clinical syndromes.

  5. Genetic and Modeling Approaches Reveal Distinct Components of Impulsive Behavior.

    Science.gov (United States)

    Nautiyal, Katherine M; Wall, Melanie M; Wang, Shuai; Magalong, Valerie M; Ahmari, Susanne E; Balsam, Peter D; Blanco, Carlos; Hen, René

    2017-01-18

    Impulsivity is an endophenotype found in many psychiatric disorders including substance use disorders, pathological gambling, and attention deficit hyperactivity disorder. Two behavioral features often considered in impulsive behavior are behavioral inhibition (impulsive action) and delayed gratification (impulsive choice). However, the extent to which these behavioral constructs represent distinct facets of behavior with discrete biological bases is unclear. To test the hypothesis that impulsive action and impulsive choice represent statistically independent behavioral constructs in mice, we collected behavioral measures of impulsivity in a single cohort of mice using well-validated operant behavioral paradigms. Mice with manipulation of serotonin 1B receptor (5-HT1BR) expression were included as a model of disordered impulsivity. A factor analysis was used to characterize correlations between the measures of impulsivity and to identify covariates. Using two approaches, we dissociated impulsive action from impulsive choice. First, the absence of 5-HT1BRs caused increased impulsive action, but not impulsive choice. Second, based on an exploratory factor analysis, a two-factor model described the data well, with measures of impulsive action and choice separating into two independent factors. A multiple-indicator multiple-causes analysis showed that 5-HT1BR expression and sex were significant covariates of impulsivity. Males displayed increased impulsivity in both dimensions, whereas 5-HT1BR expression was a predictor of increased impulsive action only. These data support the conclusion that impulsive action and impulsive choice are distinct behavioral phenotypes with dissociable biological influences that can be modeled in mice. Our work may help inform better classification, diagnosis, and treatment of psychiatric disorders, which present with disordered impulsivity.Neuropsychopharmacology advance online publication, 18 January 2017; doi:10.1038/npp.2016.277.

  6. Anode biofilm transcriptomics reveals outer surface components essential for high density current production in Geobacter sulfurreducens fuel cells.

    Directory of Open Access Journals (Sweden)

    Kelly P Nevin

    Full Text Available The mechanisms by which Geobacter sulfurreducens transfers electrons through relatively thick (>50 microm biofilms to electrodes acting as a sole electron acceptor were investigated. Biofilms of Geobacter sulfurreducens were grown either in flow-through systems with graphite anodes as the electron acceptor or on the same graphite surface, but with fumarate as the sole electron acceptor. Fumarate-grown biofilms were not immediately capable of significant current production, suggesting substantial physiological differences from current-producing biofilms. Microarray analysis revealed 13 genes in current-harvesting biofilms that had significantly higher transcript levels. The greatest increases were for pilA, the gene immediately downstream of pilA, and the genes for two outer c-type membrane cytochromes, OmcB and OmcZ. Down-regulated genes included the genes for the outer-membrane c-type cytochromes, OmcS and OmcT. Results of quantitative RT-PCR of gene transcript levels during biofilm growth were consistent with microarray results. OmcZ and the outer-surface c-type cytochrome, OmcE, were more abundant and OmcS was less abundant in current-harvesting cells. Strains in which pilA, the gene immediately downstream from pilA, omcB, omcS, omcE, or omcZ was deleted demonstrated that only deletion of pilA or omcZ severely inhibited current production and biofilm formation in current-harvesting mode. In contrast, these gene deletions had no impact on biofilm formation on graphite surfaces when fumarate served as the electron acceptor. These results suggest that biofilms grown harvesting current are specifically poised for electron transfer to electrodes and that, in addition to pili, OmcZ is a key component in electron transfer through differentiated G. sulfurreducens biofilms to electrodes.

  7. Two distinct microbial communities revealed in the sponge Cinachyrella

    Directory of Open Access Journals (Sweden)

    Marie Laure Cuvelier

    2014-11-01

    Full Text Available Marine sponges are vital components of benthic and coral reef ecosystems, providing shelter and nutrition for many organisms. In addition, sponges act as an essential carbon and nutrient link between the pelagic and benthic environment by filtering large quantities of seawater. Many sponge species harbor a diverse microbial community (including Archaea, Bacteria and Eukaryotes, which can constitute up to 50% of the sponge biomass. Sponges of the genus Cinachyrella are common in Caribbean and Floridian reefs and their archaeal and bacterial microbiomes were explored here using 16S rDNA tag pyrosequencing. Cinachyrella specimens and seawater samples were collected from the same South Florida reef at two different times of year. In total, 639 OTUs (12 archaeal and 627 bacterial belonging to 2 archaeal and 21 bacterial phyla were detected in the sponges. Based on their microbiomes, the six sponge samples formed two distinct groups, namely sponge group 1 (SG1 with low diversity (Shannon-Weiner index: 3.73 ± 0.22 and SG2 with higher diversity (Shannon-Weiner index: 5.95 ± 0.25. Hosts’ 28S rDNA sequences further confirmed that the sponge specimens were composed of two taxa closely related to Cinachyrella kuekenthalli. Both sponge groups were dominated by Proteobacteria, but Alphaproteobacteria were significantly more abundant in SG1. SG2 harbored many bacterial phyla (>1% of sequences present in low abundance or below detection limits (<0.07% in SG1 including: Acidobacteria, Chloroflexi, Gemmatimonadetes, Nitrospirae, PAUC34f, Poribacteria and Verrucomicrobia. Furthermore, SG1 and SG2 only had 95 OTUs in common, representing 30.5% and 22.4% of SG1 and SG2’s total OTUs, respectively. These results suggest that the sponge host may exert a pivotal influence on the nature and structure of the microbial community and may only be marginally affected by external environment parameters.

  8. Solar Radiation Stress in Natural Acidophilic Biofilms of Euglena mutabilis Revealed by Metatranscriptomics and PAM Fluorometry.

    Science.gov (United States)

    Puente-Sánchez, Fernando; Olsson, Sanna; Gómez-Rodriguez, Manuel; Souza-Egipsy, Virginia; Altamirano-Jeschke, Maria; Amils, Ricardo; Parro, Victor; Aguilera, Angeles

    2016-02-01

    The daily photosynthetic performance of a natural biofilm of the extreme acidophilic Euglena mutabilis from Río Tinto (SW, Spain) under full solar radiation was analyzed by means of pulse amplitude-modulated (PAM) fluorescence measurements and metatrascriptomic analysis. Natural E. mutabilis biofilms undergo large-scale transcriptomic reprogramming during midday due to a dynamic photoinhibition and solar radiation stress. Photoinhibition is due to UV radiation and not to light intensity, as revealed by PAM fluorometry analysis. In order to minimize the negative effects of solar radiation, our data supports the presence of a circadian rhythm in this euglenophyte that increases their opportunity to survive. Differential gene expression throughout the day (at 12:00, 20:00 and night) was monitored by massive Illumina parallel sequencing of metatranscriptomic libraries. The transcription pattern was altered in genes involved in Photosystem II stability and repair, UV damaged DNA repair, non-photochemical quenching and oxidative stress, supporting the photoinhibition detected by PAM fluorometry at midday.

  9. Distinct SagA from Hospital-Associated Clade A1 Enterococcus faecium Strains Contributes to Biofilm Formation.

    Science.gov (United States)

    Paganelli, F L; de Been, M; Braat, J C; Hoogenboezem, T; Vink, C; Bayjanov, J; Rogers, M R C; Huebner, J; Bonten, M J M; Willems, R J L; Leavis, H L

    2015-10-01

    Enterococcus faecium is an important nosocomial pathogen causing biofilm-mediated infections. Elucidation of E. faecium biofilm pathogenesis is pivotal for the development of new strategies to treat these infections. In several bacteria, extracellular DNA (eDNA) and proteins act as matrix components contributing to biofilm development. In this study, we investigated biofilm formation capacity and the roles of eDNA and secreted proteins for 83 E. faecium strains with different phylogenetic origins that clustered in clade A1 and clade B. Although there was no significant difference in biofilm formation between E. faecium strains from these two clades, the addition of DNase I or proteinase K to biofilms demonstrated that eDNA is essential for biofilm formation in most E. faecium strains, whereas proteolysis impacted primarily biofilms of E. faecium clade A1 strains. Secreted antigen A (SagA) was the most abundant protein in biofilms from E. faecium clade A1 and B strains, although its localization differed between the two groups. sagA was present in all sequenced E. faecium strains, with a consistent difference in the repeat region between the clades, which correlated with the susceptibility of biofilms to proteinase K. This indicates an association between the SagA variable repeat profile and the localization and contribution of SagA in E. faecium biofilms.

  10. Diversity and functions of bacterial community in drinking water biofilms revealed by high-throughput sequencing

    Science.gov (United States)

    Chao, Yuanqing; Mao, Yanping; Wang, Zhiping; Zhang, Tong

    2015-01-01

    The development of biofilms in drinking water (DW) systems may cause various problems to water quality. To investigate the community structure of biofilms on different pipe materials and the global/specific metabolic functions of DW biofilms, PCR-based 454 pyrosequencing data for 16S rRNA genes and Illumina metagenomic data were generated and analysed. Considerable differences in bacterial diversity and taxonomic structure were identified between biofilms formed on stainless steel and biofilms formed on plastics, indicating that the metallic materials facilitate the formation of higher diversity biofilms. Moreover, variations in several dominant genera were observed during biofilm formation. Based on PCA analysis, the global functions in the DW biofilms were similar to other DW metagenomes. Beyond the global functions, the occurrences and abundances of specific protective genes involved in the glutathione metabolism, the SoxRS system, the OxyR system, RpoS regulated genes, and the production/degradation of extracellular polymeric substances were also evaluated. A near-complete and low-contamination draft genome was constructed from the metagenome of the DW biofilm, based on the coverage and tetranucleotide frequencies, and identified as a Bradyrhizobiaceae-like bacterium according to a phylogenetic analysis. Our findings provide new insight into DW biofilms, especially in terms of their metabolic functions. PMID:26067561

  11. Physiology of Pseudomonas aeruginosa in biofilms as revealed by transcriptome analysis

    Directory of Open Access Journals (Sweden)

    Parker Albert

    2010-11-01

    Full Text Available Abstract Background Transcriptome analysis was applied to characterize the physiological activities of Pseudomonas aeruginosa grown for three days in drip-flow biofilm reactors. Conventional applications of transcriptional profiling often compare two paired data sets that differ in a single experimentally controlled variable. In contrast this study obtained the transcriptome of a single biofilm state, ranked transcript signals to make the priorities of the population manifest, and compared ranki ngs for a priori identified physiological marker genes between the biofilm and published data sets. Results Biofilms tolerated exposure to antibiotics, harbored steep oxygen concentration gradients, and exhibited stratified and heterogeneous spatial patterns of protein synthetic activity. Transcriptional profiling was performed and the signal intensity of each transcript was ranked to gain insight into the physiological state of the biofilm population. Similar rankings were obtained from data sets published in the GEO database http://www.ncbi.nlm.nih.gov/geo. By comparing the rank of genes selected as markers for particular physiological activities between the biofilm and comparator data sets, it was possible to infer qualitative features of the physiological state of the biofilm bacteria. These biofilms appeared, from their transcriptome, to be glucose nourished, iron replete, oxygen limited, and growing slowly or exhibiting stationary phase character. Genes associated with elaboration of type IV pili were strongly expressed in the biofilm. The biofilm population did not indicate oxidative stress, homoserine lactone mediated quorum sensing, or activation of efflux pumps. Using correlations with transcript ranks, the average specific growth rate of biofilm cells was estimated to be 0.08 h-1. Conclusions Collectively these data underscore the oxygen-limited, slow-growing nature of the biofilm population and are consistent with antimicrobial tolerance due

  12. Single particle tracking reveals spatial and dynamic organization of the E. coli biofilm matrix.

    Science.gov (United States)

    Birjiniuk, Alona; Billings, Nicole; Nance, Elizabeth; Hanes, Justin; Ribbeck, Katharina; Doyle, Patrick S

    2014-08-27

    Biofilms are communities of surface-adherent bacteria surrounded by secreted polymers known as the extracellular polymeric substance (EPS). Biofilms are harmful in many industries, and thus it is of great interest to understand their mechanical properties and structure to determine ways to destabilize them. By performing single particle tracking with beads of varying surface functionalization it was found that charge interactions play a key role in mediating mobility within biofilms. With a combination of single particle tracking and microrheological concepts, it was found that Escherichia coli biofilms display height dependent charge density that evolves over time. Statistical analyses of bead trajectories and confocal microscopy showed inter-connecting micron scale channels that penetrate throughout the biofilm, which may be important for nutrient transfer through the system. This methodology provides significant insight into a particular biofilm system and can be applied to many others to provide comparisons of biofilm structure. The elucidation of structure provides evidence for the permeability of biofilms to microscale objects, and the ability of a biofilm to mature and change properties over time.

  13. Candida-streptococcal mucosal biofilms display distinct structural and virulence characteristics depending on growth conditions and hyphal morphotypes.

    Science.gov (United States)

    Bertolini, M M; Xu, H; Sobue, T; Nobile, C J; Del Bel Cury, A A; Dongari-Bagtzoglou, A

    2015-08-01

    Candida albicans and streptococci of the mitis group form communities in multiple oral sites, where moisture and nutrient availability can change spatially or temporally. This study evaluated structural and virulence characteristics of Candida-streptococcal biofilms formed on moist or semidry mucosal surfaces, and tested the effects of nutrient availability and hyphal morphotype on dual-species biofilms. Three-dimensional models of the oral mucosa formed by immortalized keratinocytes on a fibroblast-embedded collagenous matrix were used. Infections were carried out using Streptococcus oralis strain 34, in combination with a C. albicans wild-type strain, or pseudohyphal-forming mutant strains. Increased moisture promoted a homogeneous surface biofilm by C. albicans. Dual biofilms had a stratified structure, with streptococci growing in close contact with the mucosa and fungi growing on the bacterial surface. Under semidry conditions, Candida formed localized foci of dense growth, which promoted focal growth of streptococci in mixed biofilms. Candida biofilm biovolume was greater under moist conditions, albeit with minimal tissue invasion, compared with semidry conditions. Supplementing the infection medium with nutrients under semidry conditions intensified growth, biofilm biovolume and tissue invasion/damage, without changing biofilm structure. Under these conditions, the pseudohyphal mutants and S. oralis formed defective superficial biofilms, with most bacteria in contact with the epithelial surface, below a pseudohyphal mass, resembling biofilms growing in a moist environment. The presence of S. oralis promoted fungal invasion and tissue damage under all conditions. We conclude that moisture, nutrient availability, hyphal morphotype and the presence of commensal bacteria influence the architecture and virulence characteristics of mucosal fungal biofilms.

  14. Distinct SagA from hospital-associated clade A1 Enterococcus faecium strains contributes to biofilm formation

    NARCIS (Netherlands)

    F.L. Paganelli; M. de Been; J.C. Braat (Johanna); T.A. Hoogenboezem (Thomas); C. Vink (Cornelis); J. Bayjanov; M.R.C. Rogers; J. Huebner; M.J.M. Bonten (Marc); R.J.L. Willems (Rob J.); H.L. Leavis

    2015-01-01

    textabstractEnterococcus faecium is an important nosocomial pathogen causing biofilm-mediated infections. Elucidation of E. faecium biofilm pathogenesis is pivotal for the development of new strategies to treat these infections. In several bacteria, extracellular DNA (eDNA) and proteins act as matri

  15. Distinct SagA from Hospital-Associated Clade A1 Enterococcus faecium Strains Contributes to Biofilm Formation

    NARCIS (Netherlands)

    Paganelli, F. L.; de Been, M.; Braat, J. C.; Hoogenboezem, T.; Vink, C.; Bayjanov, J.; Rogers, M. R. C.; Huebner, J.; Bonten, M. J. M.; Willems, R. J. L.; Leavis, H. L.

    2015-01-01

    Enterococcus faecium is an important nosocomial pathogen causing biofilm-mediated infections. Elucidation of E. faecium biofilm pathogenesis is pivotal for the development of new strategies to treat these infections. In several bacteria, extracellular DNA (eDNA) and proteins act as matrix components

  16. Combination of microscopic techniques reveals a comprehensive visual impression of biofilm structure and composition.

    Science.gov (United States)

    Alhede, Morten; Qvortrup, Klaus; Liebrechts, Ramon; Høiby, Niels; Givskov, Michael; Bjarnsholt, Thomas

    2012-07-01

    Bacterial biofilms are imaged by various kinds of microscopy including confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM). One limitation of CLSM is its restricted magnification, which is resolved by the use of SEM that provides high-magnification spatial images of how the single bacteria are located and interact within the biofilm. However, conventional SEM is limited by the requirement of dehydration of the samples during preparation. As biofilms consist mainly of water, the specimen dehydration might alter its morphology. High magnification yet authentic images are important to understand the physiology of biofilms. We compared conventional SEM, Focused Ion Beam (FIB)-SEM and CLSM with SEM techniques [cryo-SEM and environmental-SEM (ESEM)] that do not require dehydration. In the case of cryo-SEM, the biofilm is not dehydrated but kept frozen to obtain high-magnification images closer to the native state of the sample. Using the ESEM technique, no preparation is needed. Applying these methods to biofilms of Pseudomonas aeruginosa showed us that the dehydration of biofilms substantially influences its appearance and that a more authentic biofilm image emerges when combining all methods. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  17. The metabolically active subpopulation in Pseudomonas aeruginosa biofilms survives exposure to membrane-targeting antimicrobials via distinct molecular mechanisms

    DEFF Research Database (Denmark)

    Chiang, Wen-Chi; Pamp, Sünje Johanna; Nilsson, Martin

    2012-01-01

    Biofilms are reported to be inherently refractory toward antimicrobial attack and, therefore, cause problems in industrial and medical settings. Pseudomonas aeruginosa biofilms contain subpopulations that exhibit high metabolic activity and subpopulations that exhibit low metabolic activity. We......-targeting compounds colistin, EDTA, SDS, and chlorhexidine resulted in the same spatial distribution of live and dead bacteria, we investigated whether tolerance to these compounds originated from the same molecular mechanisms. Development of colistin-tolerant subpopulations was found to depend on the pmr genes...

  18. Whole transcriptome analysis of Acinetobacter baumannii assessed by RNA-sequencing reveals different mRNA expression profiles in biofilm compared to planktonic cells.

    Directory of Open Access Journals (Sweden)

    Soraya Rumbo-Feal

    Full Text Available Acinetobacterbaumannii has emerged as a dangerous opportunistic pathogen, with many strains able to form biofilms and thus cause persistent infections. The aim of the present study was to use high-throughput sequencing techniques to establish complete transcriptome profiles of planktonic (free-living and sessile (biofilm forms of A. baumannii ATCC 17978 and thereby identify differences in their gene expression patterns. Collections of mRNA from planktonic (both exponential and stationary phase cultures and sessile (biofilm cells were sequenced. Six mRNA libraries were prepared following the mRNA-Seq protocols from Illumina. Reads were obtained in a HiScanSQ platform and mapped against the complete genome to describe the complete mRNA transcriptomes of planktonic and sessile cells. The results showed that the gene expression pattern of A. baumannii biofilm cells was distinct from that of planktonic cells, including 1621 genes over-expressed in biofilms relative to stationary phase cells and 55 genes expressed only in biofilms. These differences suggested important changes in amino acid and fatty acid metabolism, motility, active transport, DNA-methylation, iron acquisition, transcriptional regulation, and quorum sensing, among other processes. Disruption or deletion of five of these genes caused a significant decrease in biofilm formation ability in the corresponding mutant strains. Among the genes over-expressed in biofilm cells were those in an operon involved in quorum sensing. One of them, encoding an acyl carrier protein, was shown to be involved in biofilm formation as demonstrated by the significant decrease in biofilm formation by the corresponding knockout strain. The present work serves as a basis for future studies examining the complex network systems that regulate bacterial biofilm formation and maintenance.

  19. Metagenome analyses of corroded concrete wastewater pipe biofilms reveal a complex microbial system

    Directory of Open Access Journals (Sweden)

    Gomez-Alvarez Vicente

    2012-06-01

    Full Text Available Abstract Background Concrete corrosion of wastewater collection systems is a significant cause of deterioration and premature collapse. Failure to adequately address the deteriorating infrastructure networks threatens our environment, public health, and safety. Analysis of whole-metagenome pyrosequencing data and 16S rRNA gene clone libraries was used to determine microbial composition and functional genes associated with biomass harvested from crown (top and invert (bottom sections of a corroded wastewater pipe. Results Taxonomic and functional analysis demonstrated that approximately 90% of the total diversity was associated with the phyla Actinobacteria, Bacteroidetes, Firmicutes and Proteobacteria. The top (TP and bottom pipe (BP communities were different in composition, with some of the differences attributed to the abundance of sulfide-oxidizing and sulfate-reducing bacteria. Additionally, human fecal bacteria were more abundant in the BP communities. Among the functional categories, proteins involved in sulfur and nitrogen metabolism showed the most significant differences between biofilms. There was also an enrichment of genes associated with heavy metal resistance, virulence (protein secretion systems and stress response in the TP biofilm, while a higher number of genes related to motility and chemotaxis were identified in the BP biofilm. Both biofilms contain a high number of genes associated with resistance to antibiotics and toxic compounds subsystems. Conclusions The function potential of wastewater biofilms was highly diverse with level of COG diversity similar to that described for soil. On the basis of the metagenomic data, some factors that may contribute to niche differentiation were pH, aerobic conditions and availability of substrate, such as nitrogen and sulfur. The results from this study will help us better understand the genetic network and functional capability of microbial members of wastewater concrete biofilms.

  20. Distinct Cortical Pathways for Music and Speech Revealed by Hypothesis-Free Voxel Decomposition.

    Science.gov (United States)

    Norman-Haignere, Sam; Kanwisher, Nancy G; McDermott, Josh H

    2015-12-16

    The organization of human auditory cortex remains unresolved, due in part to the small stimulus sets common to fMRI studies and the overlap of neural populations within voxels. To address these challenges, we measured fMRI responses to 165 natural sounds and inferred canonical response profiles ("components") whose weighted combinations explained voxel responses throughout auditory cortex. This analysis revealed six components, each with interpretable response characteristics despite being unconstrained by prior functional hypotheses. Four components embodied selectivity for particular acoustic features (frequency, spectrotemporal modulation, pitch). Two others exhibited pronounced selectivity for music and speech, respectively, and were not explainable by standard acoustic features. Anatomically, music and speech selectivity concentrated in distinct regions of non-primary auditory cortex. However, music selectivity was weak in raw voxel responses, and its detection required a decomposition method. Voxel decomposition identifies primary dimensions of response variation across natural sounds, revealing distinct cortical pathways for music and speech.

  1. Combination of microscopic techniques reveals a comprehensive visual impression of biofilm structure and composition

    DEFF Research Database (Denmark)

    Alhede, Morten; Qvortrup, Klaus; Liebrechts, Ramon

    2012-01-01

    Bacterial biofilms are imaged by various kinds of microscopy including confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM). One limitation of CLSM is its restricted magnification, which is resolved by the use of SEM that provides high-magnification spatial images of h...... combining all methods....

  2. Distinct genetic lineages of Bactrocera caudata (Insecta: Tephritidae revealed by COI and 16S DNA sequences.

    Directory of Open Access Journals (Sweden)

    Phaik-Eem Lim

    Full Text Available The fruit fly Bactrocera caudata is a pest species of economic importance in Asia. Its larvae feed on the flowers of Cucurbitaceae such as Cucurbita moschata. To-date it is distinguished from related species based on morphological characters. Specimens of B. caudata from Peninsular Malaysia and Indonesia (Bali and Lombok were analysed using the partial DNA sequences of cytochrome c oxidase subunit I (COI and 16S rRNA genes. Both gene sequences revealed that B. caudata from Peninsular Malaysia was distinctly different from B. caudata of Bali and Lombok, without common haplotype between them. Phylogenetic analysis revealed two distinct clades, indicating distinct genetic lineage. The uncorrected 'p' distance for COI sequences between B. caudata of Malaysia-Thailand-China and B. caudata of Bali-Lombok was 5.65%, for 16S sequences from 2.76 to 2.99%, and for combined COI and 16S sequences 4.45 to 4.46%. The 'p' values are distinctly different from intraspecific 'p' distance (0-0.23%. Both the B. caudata lineages are distinctly separated from related species in the subgenus Zeugodacus - B. ascita, B. scutellata, B. ishigakiensis, B. diaphora, B. tau, B. cucurbitae, and B. depressa. Molecular phylogenetic analysis indicates that the B. caudata lineages are closely related to B. ascita sp. B, and form a clade with B. scutellata, B. ishigakiensis, B. diaphora and B. ascita sp. A. This study provides additional baseline for the phylogenetic relationships of Bactrocera fruit flies of the subgenus Zeugodacus. Both the COI and 16S genes could be useful markers for the molecular differentiation and phylogenetic analysis of tephritid fruit flies.

  3. Distinct genetic lineages of Bactrocera caudata (Insecta: Tephritidae) revealed by COI and 16S DNA sequences.

    Science.gov (United States)

    Lim, Phaik-Eem; Tan, Ji; Suana, I Wayan; Eamsobhana, Praphathip; Yong, Hoi Sen

    2012-01-01

    The fruit fly Bactrocera caudata is a pest species of economic importance in Asia. Its larvae feed on the flowers of Cucurbitaceae such as Cucurbita moschata. To-date it is distinguished from related species based on morphological characters. Specimens of B. caudata from Peninsular Malaysia and Indonesia (Bali and Lombok) were analysed using the partial DNA sequences of cytochrome c oxidase subunit I (COI) and 16S rRNA genes. Both gene sequences revealed that B. caudata from Peninsular Malaysia was distinctly different from B. caudata of Bali and Lombok, without common haplotype between them. Phylogenetic analysis revealed two distinct clades, indicating distinct genetic lineage. The uncorrected 'p' distance for COI sequences between B. caudata of Malaysia-Thailand-China and B. caudata of Bali-Lombok was 5.65%, for 16S sequences from 2.76 to 2.99%, and for combined COI and 16S sequences 4.45 to 4.46%. The 'p' values are distinctly different from intraspecific 'p' distance (0-0.23%). Both the B. caudata lineages are distinctly separated from related species in the subgenus Zeugodacus - B. ascita, B. scutellata, B. ishigakiensis, B. diaphora, B. tau, B. cucurbitae, and B. depressa. Molecular phylogenetic analysis indicates that the B. caudata lineages are closely related to B. ascita sp. B, and form a clade with B. scutellata, B. ishigakiensis, B. diaphora and B. ascita sp. A. This study provides additional baseline for the phylogenetic relationships of Bactrocera fruit flies of the subgenus Zeugodacus. Both the COI and 16S genes could be useful markers for the molecular differentiation and phylogenetic analysis of tephritid fruit flies.

  4. The Biofilm Challenge

    DEFF Research Database (Denmark)

    Alhede, Maria; Alhede, Morten

    2014-01-01

    The concept of biofilms has emerged in the clinical setting during the last decade. Infections involving biofilms have been documented in all parts of the human body, and it is currently believed that the presence of biofilm-forming bacteria is equivalent to chronic infection. A quick Pubmed search...... reveals the significance of biofilms, as evidenced by a dramatic increase in scientific publications on the topic, as well as in publications concerning wounds with biofilms, which reached 600 publications in 2013. Judged from the number of publications, it appears that biofilms play a significant role...... in wounds. However, the impact of biofilms is often debated, because infected wounds were also treated before the concept of biofilms was coined. In this short review, we will address the significance of biofilms and their role in wounds, and discuss the future tasks of the biofilm challenge....

  5. Coral transcriptome and bacterial community profiles reveal distinct Yellow Band Disease states in Orbicella faveolata

    KAUST Repository

    Closek, Collin J.

    2014-06-20

    Coral diseases impact reefs globally. Although we continue to describe diseases, little is known about the etiology or progression of even the most common cases. To examine a spectrum of coral health and determine factors of disease progression we examined Orbicella faveolata exhibiting signs of Yellow Band Disease (YBD), a widespread condition in the Caribbean. We used a novel combined approach to assess three members of the coral holobiont: the coral-host, associated Symbiodinium algae, and bacteria. We profiled three conditions: (1) healthy-appearing colonies (HH), (2) healthy-appearing tissue on diseased colonies (HD), and (3) diseased lesion (DD). Restriction fragment length polymorphism analysis revealed health state-specific diversity in Symbiodinium clade associations. 16S ribosomal RNA gene microarrays (PhyloChips) and O. faveolata complimentary DNA microarrays revealed the bacterial community structure and host transcriptional response, respectively. A distinct bacterial community structure marked each health state. Diseased samples were associated with two to three times more bacterial diversity. HD samples had the highest bacterial richness, which included components associated with HH and DD, as well as additional unique families. The host transcriptome under YBD revealed a reduced cellular expression of defense- and metabolism-related processes, while the neighboring HD condition exhibited an intermediate expression profile. Although HD tissue appeared visibly healthy, the microbial communities and gene expression profiles were distinct. HD should be regarded as an additional (intermediate) state of disease, which is important for understanding the progression of YBD. © 2014 International Society for Microbial Ecology. All rights reserved.

  6. ‘Get in Early’; Biofilm and Wax Moth (Galleria mellonella) Models Reveal New Insights into the Therapeutic Potential of Clostridium difficile Bacteriophages

    Science.gov (United States)

    Nale, Janet Y.; Chutia, Mahananda; Carr, Philippa; Hickenbotham, Peter T.; Clokie, Martha R. J.

    2016-01-01

    Clostridium difficile infection (CDI) is a global health threat associated with high rates of morbidity and mortality. Conventional antibiotic CDI therapy can result in treatment failure and recurrent infection. C. difficile produces biofilms which contribute to its virulence and impair antimicrobial activity. Some bacteriophages (phages) can penetrate biofilms and thus could be developed to either replace or supplement antibiotics. Here, we determined the impact of a previously optimized 4-phage cocktail on C. difficile ribotype 014/020 biofilms, and additionally as adjunct to vancomycin treatment in Galleria mellonella larva CDI model. The phages were applied before or after biofilm establishment in vitro, and the impact was analyzed according to turbidity, viability counts and topography as observed using scanning electron and confocal microscopy. The infectivity profiles and efficacies of orally administered phages and/or vancomycin were ascertained by monitoring colonization levels and larval survival rates. Phages prevented biofilm formation, and penetrated established biofilms. A single phage application reduced colonization causing extended longevity in the remedial treatment and prevented disease in the prophylaxis group. Multiple phage doses significantly improved the larval remedial regimen, and this treatment is comparable to vancomycin and the combined treatments. Taken together, our data suggest that the phages significantly reduce C. difficile biofilms, and prevent colonization in the G. mellonella model when used alone or in combination with vancomycin. The phages appear to be highly promising therapeutics in the targeted eradication of CDI and the use of these models has revealed that prophylactic use could be a propitious therapeutic option. PMID:27630633

  7. 'Get in Early'; Biofilm and Wax Moth (Galleria mellonella) Models Reveal New Insights into the Therapeutic Potential of Clostridium difficile Bacteriophages.

    Science.gov (United States)

    Nale, Janet Y; Chutia, Mahananda; Carr, Philippa; Hickenbotham, Peter T; Clokie, Martha R J

    2016-01-01

    Clostridium difficile infection (CDI) is a global health threat associated with high rates of morbidity and mortality. Conventional antibiotic CDI therapy can result in treatment failure and recurrent infection. C. difficile produces biofilms which contribute to its virulence and impair antimicrobial activity. Some bacteriophages (phages) can penetrate biofilms and thus could be developed to either replace or supplement antibiotics. Here, we determined the impact of a previously optimized 4-phage cocktail on C. difficile ribotype 014/020 biofilms, and additionally as adjunct to vancomycin treatment in Galleria mellonella larva CDI model. The phages were applied before or after biofilm establishment in vitro, and the impact was analyzed according to turbidity, viability counts and topography as observed using scanning electron and confocal microscopy. The infectivity profiles and efficacies of orally administered phages and/or vancomycin were ascertained by monitoring colonization levels and larval survival rates. Phages prevented biofilm formation, and penetrated established biofilms. A single phage application reduced colonization causing extended longevity in the remedial treatment and prevented disease in the prophylaxis group. Multiple phage doses significantly improved the larval remedial regimen, and this treatment is comparable to vancomycin and the combined treatments. Taken together, our data suggest that the phages significantly reduce C. difficile biofilms, and prevent colonization in the G. mellonella model when used alone or in combination with vancomycin. The phages appear to be highly promising therapeutics in the targeted eradication of CDI and the use of these models has revealed that prophylactic use could be a propitious therapeutic option.

  8. Set potential regulation reveals additional oxidation peaks of Geobacter sulfurreducens anodic biofilms

    KAUST Repository

    Zhu, Xiuping

    2012-08-01

    Higher current densities produced in microbial fuel cells and other bioelectrochemical systems are associated with the presence of various Geobacter species. A number of electron transfer components are involved in extracellular electron transfer by the model exoelectrogen, Geobacter sulfurreducens. It has previously been shown that 5 main oxidation peaks can be identified in cyclic voltammetry scans. It is shown here that 7 separate oxidation peaks emerged over relatively long periods of time when a larger range of set potentials was used to acclimate electroactive biofilms. The potentials of oxidation peaks obtained with G. sulfurreducens biofilms acclimated at 0.60 V (vs. Ag/AgCl) were different from those that developed at - 0.46 V, and both of their peaks were different from those obtained for biofilms incubated at - 0.30 V, 0 V, and 0.30 V. These results expand the known range of potentials for which G. sulfurreducens produces identifiable oxidation peaks that could be important for extracellular electron transfer. © 2012 Elsevier B.V.

  9. Pyrosequencing reveals a core community of anodic bacterial biofilms in bioelectrochemical systems from China

    Directory of Open Access Journals (Sweden)

    Yong eXiao

    2015-12-01

    Full Text Available Bioelectrochemical systems (BESs are promising technologies for energy and product recovery coupled with wastewater treatment, and the core microbial community in electrochemically active biofilm in BESs remains controversy. In the present study, 7 anodic communities from 6 bioelectrochemical systems in 4 labs in southeast, north and south-central of China are explored by 454 pyrosequencing. A total of 251,225 effective sequences are obtained for 7 electrochemically active biofilm samples at 3% cutoff level. While Alpha-, Beta- and Gamma-proteobacteria are the most abundant classes (averaging 16.0-17.7%, Bacteroidia and Clostridia are the two sub-dominant and commonly shared classes. Six commonly shared genera i.e. Azospira, Azospirillum, Acinetobacter, Bacteroides, Geobacter, Pseudomonas and Rhodopseudomonas dominate the electrochemically active communities and are defined as core genera. A total of 25 OTUs with average relative abundance >0.5% were selected and designated as core OTUs, and some species relating to these OTUs have been reported electrochemically active. Furthermore, cyclic voltammetry and chronoamperometry tests show that two strains from Acinetobacter guillouiae and Stappia indica, bacteria relate to two core OTUs, are electrochemically active. Using randomly selected bioelectrochemical systems, the study presented extremely diverse bacterial communities in anodic biofilms, though, we still can suggest some potential microbes for investigating the electrochemical mechanisms in bioelectrochemical systems.

  10. Lipidomics of Candida albicans biofilms reveals phase-dependent production of phospholipid molecular classes and role for lipid rafts in biofilm formation.

    Science.gov (United States)

    Lattif, Ali Abdul; Mukherjee, Pranab K; Chandra, Jyotsna; Roth, Mary R; Welti, Ruth; Rouabhia, Mahmoud; Ghannoum, Mahmoud A

    2011-11-01

    Candida albicans-associated bloodstream infections are linked to the ability of this yeast to form biofilms. In this study, we used lipidomics to compare the lipid profiles of C. albicans biofilms and planktonic cells, in early and mature developmental phases. Our results showed that significant differences exist in lipid composition in both developmental phases. Biofilms contained higher levels of phospholipid and sphingolipids than planktonic cells (nmol per g biomass, Pbiofilms compared to planktonic cells (P≤0.05). The ratio of phosphatidylcholine to phosphatidylethanolamine was lower in biofilms compared to planktonic cells in both early (1.17 vs 2.52, P≤0.001) and late (2.34 vs 3.81, P≤0.001) developmental phases. The unsaturation index of phospholipids decreased with time, with this effect being particularly strong for biofilms. Inhibition of the biosynthetic pathway for sphingolipid [mannosyl diinositolphosphoryl ceramide, M(IP)₂C] by myriocin or aureobasidin A, and disruption of the gene encoding inositolphosphotransferase (Ipt1p), abrogated the ability of C. albicans to form biofilms. The differences in lipid profiles between biofilms and planktonic Candida cells may have important implications for the biology and antifungal resistance of biofilms.

  11. Dynamic Changes in Amygdala Psychophysiological Connectivity Reveal Distinct Neural Networks for Facial Expressions of Basic Emotions

    Science.gov (United States)

    Diano, Matteo; Tamietto, Marco; Celeghin, Alessia; Weiskrantz, Lawrence; Tatu, Mona-Karina; Bagnis, Arianna; Duca, Sergio; Geminiani, Giuliano; Cauda, Franco; Costa, Tommaso

    2017-01-01

    The quest to characterize the neural signature distinctive of different basic emotions has recently come under renewed scrutiny. Here we investigated whether facial expressions of different basic emotions modulate the functional connectivity of the amygdala with the rest of the brain. To this end, we presented seventeen healthy participants (8 females) with facial expressions of anger, disgust, fear, happiness, sadness and emotional neutrality and analyzed amygdala’s psychophysiological interaction (PPI). In fact, PPI can reveal how inter-regional amygdala communications change dynamically depending on perception of various emotional expressions to recruit different brain networks, compared to the functional interactions it entertains during perception of neutral expressions. We found that for each emotion the amygdala recruited a distinctive and spatially distributed set of structures to interact with. These changes in amygdala connectional patters characterize the dynamic signature prototypical of individual emotion processing, and seemingly represent a neural mechanism that serves to implement the distinctive influence that each emotion exerts on perceptual, cognitive, and motor responses. Besides these differences, all emotions enhanced amygdala functional integration with premotor cortices compared to neutral faces. The present findings thus concur to reconceptualise the structure-function relation between brain-emotion from the traditional one-to-one mapping toward a network-based and dynamic perspective. PMID:28345642

  12. How Cyanobacterial Distributions Reveal Flow and Irradiance Conditions of Photosynthetic Biofilm Formation

    Science.gov (United States)

    Prufert-Bebout, Lee

    2001-01-01

    Microbial life on Earth is enormously abundant at sediment-water interfaces. The fossil record in fact contains abundant evidence of the preservation of life on such surfaces. It is therefore critical to our interpretation of early Earth history, and potentially to history of life on other planets, to be able to recognize life forms at these interfaces. On Earth this life often occurs as organized structures of microbes and their extracellular exudates known as biofilms. When such biofilms occur in areas receiving sunlight photosynthetic biofilms are the dominant form in natural ecosystems due to selective advantage inherent in their ability to utilize solar energy. Cyanobacteria are the dominant phototrophic microbes in most modern and ancient photosynthetic biofilms, microbial mats and stromatolites. Due to their long (3.5 billion year) evolutionary history, this group has extensively diversified resulting in an enormous array of morphologies and physiological abilities. This enormous diversity and specialization results in very specific selection for a particular cyanobacterium in each available photosynthetic niche. Furthermore these organisms can alter their spatial orientation, cell morphology, pigmentation and associations with heterotrophic organisms in order to fine tune their optimization to a given micro-niche. These adaptations can be detected, and if adequate knowledge of the interaction between environmental conditions and organism response is available, the detectable organism response can be used to infer the environmental conditions causing that response. This presentation will detail two specific examples which illustrate this point. Light and water are essential to photosynthesis in cyanobacteria and these organisms have specific detectable behavioral responses to these parameters. We will present cyanobacterial responses to quantified flow and irradiance to demonstrate the interpretative power of distribution and orientation information. This

  13. Relationship between Distinct African Cholera Epidemics Revealed via MLVA Haplotyping of 337 Vibrio cholerae Isolates.

    Science.gov (United States)

    Moore, Sandra; Miwanda, Berthe; Sadji, Adodo Yao; Thefenne, Hélène; Jeddi, Fakhri; Rebaudet, Stanislas; de Boeck, Hilde; Bidjada, Bawimodom; Depina, Jean-Jacques; Bompangue, Didier; Abedi, Aaron Aruna; Koivogui, Lamine; Keita, Sakoba; Garnotel, Eric; Plisnier, Pierre-Denis; Ruimy, Raymond; Thomson, Nicholas; Muyembe, Jean-Jacques; Piarroux, Renaud

    2015-01-01

    Since cholera appeared in Africa during the 1970s, cases have been reported on the continent every year. In Sub-Saharan Africa, cholera outbreaks primarily cluster at certain hotspots including the African Great Lakes Region and West Africa. In this study, we applied MLVA (Multi-Locus Variable Number Tandem Repeat Analysis) typing of 337 Vibrio cholerae isolates from recent cholera epidemics in the Democratic Republic of the Congo (DRC), Zambia, Guinea and Togo. We aimed to assess the relationship between outbreaks. Applying this method, we identified 89 unique MLVA haplotypes across our isolate collection. MLVA typing revealed the short-term divergence and microevolution of these Vibrio cholerae populations to provide insight into the dynamics of cholera outbreaks in each country. Our analyses also revealed strong geographical clustering. Isolates from the African Great Lakes Region (DRC and Zambia) formed a closely related group, while West African isolates (Togo and Guinea) constituted a separate cluster. At a country-level scale our analyses revealed several distinct MLVA groups, most notably DRC 2011/2012, DRC 2009, Zambia 2012 and Guinea 2012. We also found that certain MLVA types collected in the DRC persisted in the country for several years, occasionally giving rise to expansive epidemics. Finally, we found that the six environmental isolates in our panel were unrelated to the epidemic isolates. To effectively combat the disease, it is critical to understand the mechanisms of cholera emergence and diffusion in a region-specific manner. Overall, these findings demonstrate the relationship between distinct epidemics in West Africa and the African Great Lakes Region. This study also highlights the importance of monitoring and analyzing Vibrio cholerae isolates.

  14. Global analysis of gene expression in pulmonary fibrosis reveals distinct programs regulating lung inflammation and fibrosis

    Science.gov (United States)

    Kaminski, Naftali; Allard, John D.; Pittet, Jean F.; Zuo, Fengrong; Griffiths, Mark J. D.; Morris, David; Huang, Xiaozhu; Sheppard, Dean; Heller, Renu A.

    2000-02-01

    The molecular mechanisms of pulmonary fibrosis are poorly understood. We have used oligonucleotide arrays to analyze the gene expression programs that underlie pulmonary fibrosis in response to bleomycin, a drug that causes lung inflammation and fibrosis, in two strains of susceptible mice (129 and C57BL/6). We then compared the gene expression patterns in these mice with 129 mice carrying a null mutation in the epithelial-restricted integrin 6 subunit (6/-), which develop inflammation but are protected from pulmonary fibrosis. Cluster analysis identified two distinct groups of genes involved in the inflammatory and fibrotic responses. Analysis of gene expression at multiple time points after bleomycin administration revealed sequential induction of subsets of genes that characterize each response. The availability of this comprehensive data set should accelerate the development of more effective strategies for intervention at the various stages in the development of fibrotic diseases of the lungs and other organs.

  15. Source-based morphometry reveals distinct patterns of aberrant brain volume in delusional infestation.

    Science.gov (United States)

    Wolf, Robert Ch; Huber, Markus; Lepping, Peter; Sambataro, Fabio; Depping, Malte S; Karner, Martin; Freudenmann, Roland W

    2014-01-03

    Little is known about the neural correlates of delusional infestation (DI), the delusional belief to be infested with pathogens. So far, evidence comes mainly from case reports and case series. We investigated brain morphology in 16 DI patients and 16 healthy controls using structural magnetic resonance imaging and a multivariate data analysis technique, i.e. source-based morphometry (SBM). In addition, we explored differences in brain structure in patient subgroups based on disease aetiology. SBM revealed two patterns exhibiting significantly (pdisorder) and "organic" DI (DI due to a medical condition). In contrast, aberrant white matter volume was only confirmed for the "organic" DI patient subgroup. These results suggest prefrontal, temporal, parietal, insular, thalamic and striatal dysfunction underlying DI. Moreover, the data suggest that aetiologically distinct presentations of DI share similar patterns of abnormal grey matter volume, whereas aberrant white matter volume appears to be restricted to organic cases. © 2013.

  16. Genome-wide mutagenesis of Xanthomonas axonopodis pv. citri reveals novel genetic determinants and regulation mechanisms of biofilm formation.

    Directory of Open Access Journals (Sweden)

    Jinyun Li

    Full Text Available Xanthomonas axonopodis pv. citri (Xac causes citrus canker disease, a major threat to citrus production worldwide. Accumulating evidence suggests that the formation of biofilms on citrus leaves plays an important role in the epiphytic survival of this pathogen prior to the development of canker disease. However, the process of Xac biofilm formation is poorly understood. Here, we report a genome-scale study of Xac biofilm formation in which we identified 92 genes, including 33 novel genes involved in biofilm formation and 7 previously characterized genes, colR, fhaB, fliC, galU, gumD, wxacO, and rbfC, known to be important for Xac biofilm formation. In addition, 52 other genes with defined or putative functions in biofilm formation were identified, even though they had not previously reported been to be associated with biofilm formation. The 92 genes were isolated from 292 biofilm-defective mutants following a screen of a transposon insertion library containing 22,000 Xac strain 306 mutants. Further analyses indicated that 16 of the novel genes are involved in the production of extracellular polysaccharide (EPS and/or lipopolysaccharide (LPS, 7 genes are involved in signaling and regulatory pathways, and 5 genes have unknown roles in biofilm formation. Furthermore, two novel genes, XAC0482, encoding a haloacid dehalogenase-like phosphatase, and XAC0494 (designated as rbfS, encoding a two-component sensor protein, were confirmed to be biofilm-related genes through complementation assays. Our data demonstrate that the formation of mature biofilm requires EPS, LPS, both flagellum-dependent and flagellum-independent cell motility, secreted proteins and extracellular DNA. Additionally, multiple signaling pathways are involved in Xac biofilm formation. This work is the first report on a genome-wide scale of the genetic processes of biofilm formation in plant pathogenic bacteria. The report provides significant new information about the genetic

  17. Xenon and iodine reveal multiple distinct exotic xenon components in Efremovka "nanodiamonds"

    Science.gov (United States)

    Gilmour, J. D.; Holland, G.; Verchovsky, A. B.; Fisenko, A. V.; Crowther, S. A.; Turner, G.

    2016-03-01

    We identify new xenon components in a nanodiamond-rich residue from the reduced CV3 chondrite Efremovka. We demonstrate for the first time that these, and the previously identified xenon components Xe-P3 and Xe-P6, are associated with elevated I/Xe ratios. The 129I/127I ratio associated with xenon loss from these presolar compositions during processing on planetesimals in the early solar system was (0.369 ± 0.019) × 10-4, a factor of 3-4 lower than the canonical value. This suggests either incorporation of iodine into carbonaceous grains before the last input of freshly synthesized 129I to the solar system's precursor material, or loss of noble gases during processing of planetesimals around 30 Myr after solar system formation. The xenon/iodine ratios and model closure ages were revealed by laser step pyrolysis analysis of a neutron-irradiated, coarse-grained nanodiamond separate. Three distinct low temperature compositions are identified by characteristic I/Xe ratios and 136Xe/132Xe ratios. There is some evidence of multiple compositions with distinct I/Xe ratios in the higher temperature releases associated with Xe-P6. The presence of iodine alongside Q-Xe and these components in nanodiamonds constrains the pathway by which extreme volatiles entered the solid phase and may facilitate the identification of their carriers. There is no detectable iodine contribution to the presolar Xe-HL component, which is released at intermediate temperatures; this suggests a distinct trapping process. Releases associated with the other components all include significant contributions of 128Xe produced from iodine by neutron capture during reactor irradiation. We propose a revised model relating the origin of Xe-P3 (which exhibits an s-process deficit) through a ;Q-process; to the Q component (which makes the dominant contribution to the heavy noble gas budget of primitive material). The Q-process incorporates noble gases and iodine into specific carbonaceous phases with mass

  18. Systematic analysis of reportedly distinct populations of multipotent bone marrow-derived stem cells reveals a lack of distinction.

    Science.gov (United States)

    Lodie, Tracey A; Blickarz, Courtney E; Devarakonda, Tara J; He, Chufa; Dash, Ajeeta B; Clarke, Jennifer; Gleneck, Kristen; Shihabuddin, Lamya; Tubo, Ross

    2002-10-01

    Adult human bone marrow-derived stem cells, having the ability to differentiate into cells of multiple lineages, have been isolated and propagated by varied protocols, including positive (CD105(+))/negative (CD45(-)GlyA(-)) selection with immunomagnetic beads, or direct plating into selective culture media. Each substratum-adherent cell population was subjected to a systematic analysis of their cell surface markers and differentiation potential. In the initial stages of culture, each cell population proliferated slowly, reaching confluence in 10-14 days. Adherent cells proliferated at similar rates whether cultured in serum-free medium supplemented with basic fibroblast growth factor, medium containing 2% fetal bovine serum (FBS) supplemented with epidermal growth factor and platelet-derived growth factor, or medium containing 10% FBS alone. Cell surface marker analysis revealed that more than 95% of the cells were positive for CD105/endoglin, a putative mesenchymal stem cell marker, and negative for CD34, CD31, and CD133, markers of hematopoietic, endothelial, and neural stem cells, respectively, regardless of cell isolation and propagation method. CD44 expression was variable, apparently dependent on serum concentration. Functional similarity of the stem cell populations was also observed, with each different cell population expressing the cell type-specific markers beta-tubulin, type II collagen, and desmin, and demonstrating endothelial tube formation when cultured under conditions favoring neural, cartilage, muscle, and endothelial cell differentiation, respectively. On the basis of these data, adult human bone marrow-derived stem cells cultured in adherent monolayer are virtually indistinguishable, both physically and functionally, regardless of the method of isolation or proliferative expansion.

  19. Genome Scale Mutational Analysis of Geobacter sulfurreducens Reveals Distinct Molecular Mechanisms for Respiration and Sensing of Poised Electrodes versus Fe(III) Oxides.

    Science.gov (United States)

    Chan, Chi Ho; Levar, Caleb E; Jiménez-Otero, Fernanda; Bond, Daniel R

    2017-10-01

    metals using the same molecular mechanisms used to form biofilms on electrodes and produce electricity. However, by simultaneously comparing thousands of Geobacter sulfurreducens transposon mutants undergoing electrode-dependent respiration, we discovered new cytochromes and chemosensory proteins supporting growth with electrodes that are not required for metal respiration. This supports an emerging model where G. sulfurreducens recognizes surfaces and forms conductive biofilms using mechanisms distinct from those used for growth with metal oxides. These findings provide a possible explanation for studies that correlate electricity generation with syntrophic interspecies electron transfer by Geobacter and reveal many previously unrecognized targets for engineering this useful capability in other organisms. Copyright © 2017 Chan et al.

  20. Pyrosequencing reveals bacterial communities in unchlorinated drinking water distribution system: an integral study of bulk water, suspended solids, loose deposits, and pipe wall biofilm.

    Science.gov (United States)

    Liu, G; Bakker, G L; Li, S; Vreeburg, J H G; Verberk, J Q J C; Medema, G J; Liu, W T; Van Dijk, J C

    2014-05-20

    The current understanding of drinking water distribution system (DWDS) microbiology is limited to pipe wall biofilm and bulk water; the contributions of particle-associated bacteria (from suspended solids and loose deposits) have long been neglected. Analyzing the composition and correlation of bacterial communities from different phases helped us to locate where most of the bacteria are and understand the interactions among these phases. In the present study, the bacteria from four critical phases of an unchlorinated DWDS, including bulk water, pipe wall biofilm, suspended solids, and loose deposits, were quantified and identified by adenosine triphosphate analysis and pyrosequencing, respectively. The results showed that the bulk water bacteria (including the contribution of suspended solids) contributed less than 2% of the total bacteria. The bacteria associated with loose deposits and pipe wall biofilm that accumulated in the DWDS accounted for over 98% of the total bacteria, and the contributions of bacteria in loose deposits and pipe wall biofilm were comparable. Depending on the amount of loose deposits, its contribution can be 7-fold higher than the pipe wall biofilm. Pyrosequencing revealed relatively stable bacterial communities in bulk water, pipe wall biofilm, and suspended solids throughout the distribution system; however, the communities present in loose deposits were dependent on the amount of loose deposits locally. Bacteria within the phases of suspended solids, loose deposits, and pipe wall biofilm were similar in phylogenetic composition. The bulk water bacteria (dominated by Polaromonas spp.) were clearly different from the bacteria from the other three phases (dominated by Sphingomonas spp.). This study highlighted that the integral DWDS ecology should include contributions from all of the four phases, especially the bacteria harbored by loose deposits. The accumulation of loose deposits and the aging process create variable microenvironments

  1. Pyrosequencing Reveals Bacterial Communities in Unchlorinated Drinking Water Distribution System: An Integral Study of Bulk Water, Suspended Solids, Loose Deposits, and Pipe Wall Biofilm

    KAUST Repository

    Liu, G.

    2014-05-20

    The current understanding of drinking water distribution system (DWDS) microbiology is limited to pipe wall biofilm and bulk water; the contributions of particle-associated bacteria (from suspended solids and loose deposits) have long been neglected. Analyzing the composition and correlation of bacterial communities from different phases helped us to locate where most of the bacteria are and understand the interactions among these phases. In the present study, the bacteria from four critical phases of an unchlorinated DWDS, including bulk water, pipe wall biofilm, suspended solids, and loose deposits, were quantified and identified by adenosine triphosphate analysis and pyrosequencing, respectively. The results showed that the bulk water bacteria (including the contribution of suspended solids) contributed less than 2% of the total bacteria. The bacteria associated with loose deposits and pipe wall biofilm that accumulated in the DWDS accounted for over 98% of the total bacteria, and the contributions of bacteria in loose deposits and pipe wall biofilm were comparable. Depending on the amount of loose deposits, its contribution can be 7-fold higher than the pipe wall biofilm. Pyrosequencing revealed relatively stable bacterial communities in bulk water, pipe wall biofilm, and suspended solids throughout the distribution system; however, the communities present in loose deposits were dependent on the amount of loose deposits locally. Bacteria within the phases of suspended solids, loose deposits, and pipe wall biofilm were similar in phylogenetic composition. The bulk water bacteria (dominated by Polaromonas spp.) were clearly different from the bacteria from the other three phases (dominated by Sphingomonas spp.). This study highlighted that the integral DWDS ecology should include contributions from all of the four phases, especially the bacteria harbored by loose deposits. The accumulation of loose deposits and the aging process create variable microenvironments

  2. ‘Get in early’; biofilm and wax moth (Galleria mellonella models reveal new insights into the therapeutic potential of Clostridium difficile bacteriophages

    Directory of Open Access Journals (Sweden)

    Janet Yakubu Nale

    2016-08-01

    Full Text Available Clostridium difficile infection (CDI is a global health threat associated with high rates of morbidity and mortality. Conventional antibiotic CDI therapy can result in treatment failure and recurrent infection. C. difficile produces biofilms which contribute to its virulence and impair antimicrobial activity. Some bacteriophages (phages can penetrate biofilms and thus could be developed to either replace or supplement antibiotics. Here, we determined the impact of a previously optimized 4-phage cocktail on C. difficile ribotype 014/020 biofilms, and additionally as adjunct to vancomycin treatment in Galleria mellonella larva CDI model. The phages were applied before or after biofilm establishment in vitro, and the impact was analyzed according to turbidity, viability counts and topography as observed using scanning electron and confocal microscopy. The infectivity profiles and efficacies of orally administered phages and/or vancomycin were ascertained by monitoring colonization levels and larval survival rates. Phages prevented biofilm formation, and penetrated established biofilms. A single phage application reduced colonization causing extended longevity in the remedial treatment and prevented disease in the prophylaxis group. Multiple phage doses significantly improved the larval remedial regimen, and this treatment is comparable to vancomycin and the combined treatments. Taken together, our data suggest that the phages significantly reduce C. difficile biofilms, and prevent colonization in the G. mellonella model when used alone or in combination with vancomycin. The phages appear to be highly promising therapeutics in the targeted eradication of CDI and the use of these models has revealed that prophylactic use could be a propitious therapeutic option.

  3. A systems approach reveals distinct metabolic strategies among the NCI-60 cancer cell lines

    Science.gov (United States)

    Aurich, Maike K.; Fleming, Ronan M. T.; Thiele, Ines

    2017-01-01

    The metabolic phenotype of cancer cells is reflected by the metabolites they consume and by the byproducts they release. Here, we use quantitative, extracellular metabolomic data of the NCI-60 panel and a novel computational method to generate 120 condition-specific cancer cell line metabolic models. These condition-specific cancer models used distinct metabolic strategies to generate energy and cofactors. The analysis of the models’ capability to deal with environmental perturbations revealed three oxotypes, differing in the range of allowable oxygen uptake rates. Interestingly, models based on metabolomic profiles of melanoma cells were distinguished from other models through their low oxygen uptake rates, which were associated with a glycolytic phenotype. A subset of the melanoma cell models required reductive carboxylation. The analysis of protein and RNA expression levels from the Human Protein Atlas showed that IDH2, which was an essential gene in the melanoma models, but not IDH1 protein, was detected in normal skin cell types and melanoma. Moreover, the von Hippel-Lindau tumor suppressor (VHL) protein, whose loss is associated with non-hypoxic HIF-stabilization, reductive carboxylation, and promotion of glycolysis, was uniformly absent in melanoma. Thus, the experimental data supported the predicted role of IDH2 and the absence of VHL protein supported the glycolytic and low oxygen phenotype predicted for melanoma. Taken together, our approach of integrating extracellular metabolomic data with metabolic modeling and the combination of different network interrogation methods allowed insights into the metabolism of cells. PMID:28806730

  4. Investigation at the atomic level of homologous enzymes reveals distinct reaction paths

    Science.gov (United States)

    Zoi, Ioanna; Schwartz, Steven D.

    2015-03-01

    Bacterial enzymes Escherichia coli and Vibrio cholerae 5' -Methylthioadenosine nucleosidases (MTANs) have different binding affinities for the same transition state analogue. This was surprising as these enzymes share 60% sequence identity, have almost identical active sites and act under the same mechanism. We performed Transition Path Sampling simulations of both enzymes to reveal the atomic details of the catalytic chemical step, to explain the inhibitor affinity differences. Unlike EcMTAN, VcMTAN has multiple distinct transition states, which is an indication that multiple sets of coordinated protein motions can reach a transition state. We also identified the important residues that participate in each enzyme's reaction coordinate and explained their contribution. Subtle dynamic differences manifest in difference of reaction coordinate and transition state structure and also suggest that MTANs differ from most ribosyl transferases. As experimental approaches report averages regarding reaction coordinate information, this study offers, previously unavailable, detailed knowledge to the explanation of bacterial MTANs catalytic mechanism, and could have a significant impact on pharmaceutical design. We acknowledge the support of the National Institutes of Health through Grant GM068036.

  5. Genome comparison of Candida orthopsilosis clinical strains reveals the existence of hybrids between two distinct subspecies.

    Science.gov (United States)

    Pryszcz, Leszek P; Németh, Tibor; Gácser, Attila; Gabaldón, Toni

    2014-05-01

    The Candida parapsilosis species complex comprises a group of emerging human pathogens of varying virulence. This complex was recently subdivided into three different species: C. parapsilosis sensu stricto, C. metapsilosis, and C. orthopsilosis. Within the latter, at least two clearly distinct subspecies seem to be present among clinical isolates (Type 1 and Type 2). To gain insight into the genomic differences between these subspecies, we undertook the sequencing of a clinical isolate classified as Type 1 and compared it with the available sequence of a Type 2 clinical strain. Unexpectedly, the analysis of the newly sequenced strain revealed a highly heterozygous genome, which we show to be the consequence of a hybridization event between both identified subspecies. This implicitly suggests that C. orthopsilosis is able to mate, a so-far unanswered question. The resulting hybrid shows a chimeric genome that maintains a similar gene dosage from both parental lineages and displays ongoing loss of heterozygosity. Several of the differences found between the gene content in both strains relate to virulent-related families, with the hybrid strain presenting a higher copy number of genes coding for efflux pumps or secreted lipases. Remarkably, two clinical strains isolated from distant geographical locations (Texas and Singapore) are descendants of the same hybrid line, raising the intriguing possibility of a relationship between the hybridization event and the global spread of a virulent clone.

  6. Methylome sequencing in triple-negative breast cancer reveals distinct methylation clusters with prognostic value.

    Science.gov (United States)

    Stirzaker, Clare; Zotenko, Elena; Song, Jenny Z; Qu, Wenjia; Nair, Shalima S; Locke, Warwick J; Stone, Andrew; Armstong, Nicola J; Robinson, Mark D; Dobrovic, Alexander; Avery-Kiejda, Kelly A; Peters, Kate M; French, Juliet D; Stein, Sandra; Korbie, Darren J; Trau, Matt; Forbes, John F; Scott, Rodney J; Brown, Melissa A; Francis, Glenn D; Clark, Susan J

    2015-02-02

    Epigenetic alterations in the cancer methylome are common in breast cancer and provide novel options for tumour stratification. Here, we perform whole-genome methylation capture sequencing on small amounts of DNA isolated from formalin-fixed, paraffin-embedded tissue from triple-negative breast cancer (TNBC) and matched normal samples. We identify differentially methylated regions (DMRs) enriched with promoters associated with transcription factor binding sites and DNA hypersensitive sites. Importantly, we stratify TNBCs into three distinct methylation clusters associated with better or worse prognosis and identify 17 DMRs that show a strong association with overall survival, including DMRs located in the Wilms tumour 1 (WT1) gene, bi-directional-promoter and antisense WT1-AS. Our data reveal that coordinated hypermethylation can occur in oestrogen receptor-negative disease, and that characterizing the epigenetic framework provides a potential signature to stratify TNBCs. Together, our findings demonstrate the feasibility of profiling the cancer methylome with limited archival tissue to identify regulatory regions associated with cancer.

  7. Metagenomic investigation of the geologically unique Hellenic Volcanic Arc reveals a distinctive ecosystem with unexpected physiology.

    Science.gov (United States)

    Oulas, Anastasis; Polymenakou, Paraskevi N; Seshadri, Rekha; Tripp, H James; Mandalakis, Manolis; Paez-Espino, A David; Pati, Amrita; Chain, Patrick; Nomikou, Paraskevi; Carey, Steven; Kilias, Stephanos; Christakis, Christos; Kotoulas, Georgios; Magoulas, Antonios; Ivanova, Natalia N; Kyrpides, Nikos C

    2016-04-01

    Hydrothermal vents represent a deep, hot, aphotic biosphere where chemosynthetic primary producers, fuelled by chemicals from Earth's subsurface, form the basis of life. In this study, we examined microbial mats from two distinct volcanic sites within the Hellenic Volcanic Arc (HVA). The HVA is geologically and ecologically unique, with reported emissions of CO2 -saturated fluids at temperatures up to 220°C and a notable absence of macrofauna. Metagenomic data reveals highly complex prokaryotic communities composed of chemolithoautotrophs, some methanotrophs, and to our surprise, heterotrophs capable of anaerobic degradation of aromatic hydrocarbons. Our data suggest that aromatic hydrocarbons may indeed be a significant source of carbon in these sites, and instigate additional research into the nature and origin of these compounds in the HVA. Novel physiology was assigned to several uncultured prokaryotic lineages; most notably, a SAR406 representative is attributed with a role in anaerobic hydrocarbon degradation. This dataset, the largest to date from submarine volcanic ecosystems, constitutes a significant resource of novel genes and pathways with potential biotechnological applications.

  8. Highly distinct chromosomal structures in cowpea (Vigna unguiculata), as revealed by molecular cytogenetic analysis.

    Science.gov (United States)

    Iwata-Otsubo, Aiko; Lin, Jer-Young; Gill, Navdeep; Jackson, Scott A

    2016-05-01

    Cowpea (Vigna unguiculata (L.) Walp) is an important legume, particularly in developing countries. However, little is known about its genome or chromosome structure. We used molecular cytogenetics to characterize the structure of pachytene chromosomes to advance our knowledge of chromosome and genome organization of cowpea. Our data showed that cowpea has highly distinct chromosomal structures that are cytologically visible as brightly DAPI-stained heterochromatic regions. Analysis of the repetitive fraction of the cowpea genome present at centromeric and pericentromeric regions confirmed that two retrotransposons are major components of pericentromeric regions and that a 455-bp tandem repeat is found at seven out of 11 centromere pairs in cowpea. These repeats likely evolved after the divergence of cowpea from common bean and form chromosomal structure unique to cowpea. The integration of cowpea genetic and physical chromosome maps reveals potential regions of suppressed recombination due to condensed heterochromatin and a lack of pairing in a few chromosomal termini. This study provides fundamental knowledge on cowpea chromosome structure and molecular cytogenetics tools for further chromosome studies.

  9. Quantitative proteomics reveals distinct differences in the protein content of outer membrane vesicle vaccines.

    Science.gov (United States)

    van de Waterbeemd, Bas; Mommen, Geert P M; Pennings, Jeroen L A; Eppink, Michel H; Wijffels, René H; van der Pol, Leo A; de Jong, Ad P J M

    2013-04-05

    At present, only vaccines containing outer membrane vesicles (OMV) have successfully stopped Neisseria meningitidis serogroup B epidemics. These vaccines however require detergent-extraction to remove endotoxin, which changes immunogenicity and causes production difficulties. To investigate this in more detail, the protein content of detergent-extracted OMV is compared with two detergent-free alternatives. A novel proteomics strategy has been developed that allows quantitative analysis of many biological replicates despite inherent multiplex restrictions of dimethyl labeling. This enables robust statistical analysis of relative protein abundance. The comparison with detergent-extracted OMV reveales that detergent-free OMV are enriched with membrane (lipo)proteins and contain less cytoplasmic proteins due to a milder purification process. These distinct protein profiles are substantiated with serum blot proteomics, confirming enrichment with immunogenic proteins in both detergent-free alternatives. Therefore, the immunogenic protein content of OMV vaccines depends at least partially on the purification process. This study demonstrates that detergent-free OMV have a preferred composition.

  10. Col-OSSOS: z-Band Photometry Reveals Three Distinct TNO Surface Types

    Science.gov (United States)

    Pike, Rosemary E.; Fraser, Wesley C.; Schwamb, Megan E.; Kavelaars, J. J.; Marsset, Michael; Bannister, Michele T.; Lehner, Matthew J.; Wang, Shiang-Yu; Alexandersen, Mike; Chen, Ying-Tung; Gladman, Brett J.; Gwyn, Stephen; Petit, Jean-Marc; Volk, Kathryn

    2017-09-01

    Several different classes of trans-Neptunian objects (TNOs) have been identified based on their optical and near-infrared colors. As part of the Colours of the Outer Solar System Origins Survey (Col-OSSOS), we have obtained g-, r-, and z-band photometry of 26 TNOs using Subaru and Gemini Observatories. Previous color surveys have not utilized z-band reflectance, and the inclusion of this band reveals significant surface reflectance variations between sub-populations. The colors of TNOs in g - r and r - z show obvious structure, and appear consistent with the previously measured bi-modality in g - r. The distribution of colors of the two dynamically excited surface types can be modeled using the two-component mixing models from Fraser & Brown. With the combination of g - r and r - z, the dynamically excited classes can be separated cleanly into red and neutral surface classes. In g - r and r - z, the two dynamically excited surface groups are also clearly distinct from the cold classical TNO surfaces, which are red, with g-r≳ 0.85 and r - z ≲ 0.6, while all dynamically excited objects with similar g - r colors exhibit redder r - z colors. The z-band photometry makes it possible for the first time to differentiate the red excited TNO surfaces from the red cold classical TNO surfaces. The discovery of different r - z colors for these cold classical TNOs makes it possible to search for cold classical surfaces in other regions of the Kuiper Belt and to completely separate cold classical TNOs from the dynamically excited population, which overlaps in orbital parameter space.

  11. ASCL1 and NEUROD1 Reveal Heterogeneity in Pulmonary Neuroendocrine Tumors and Regulate Distinct Genetic Programs

    Directory of Open Access Journals (Sweden)

    Mark D. Borromeo

    2016-08-01

    Full Text Available Small cell lung carcinoma (SCLC is a high-grade pulmonary neuroendocrine tumor. The transcription factors ASCL1 and NEUROD1 play crucial roles in promoting malignant behavior and survival of human SCLC cell lines. Here, we find that ASCL1 and NEUROD1 identify heterogeneity in SCLC, bind distinct genomic loci, and regulate mostly distinct genes. ASCL1, but not NEUROD1, is present in mouse pulmonary neuroendocrine cells, and only ASCL1 is required in vivo for tumor formation in mouse models of SCLC. ASCL1 targets oncogenic genes including MYCL1, RET, SOX2, and NFIB while NEUROD1 targets MYC. ASCL1 and NEUROD1 regulate different genes that commonly contribute to neuronal function. ASCL1 also regulates multiple genes in the NOTCH pathway including DLL3. Together, ASCL1 and NEUROD1 distinguish heterogeneity in SCLC with distinct genomic landscapes and distinct gene expression programs.

  12. A bioluminescent Pseudomonas aeruginosa wound model reveals increased mortality of type 1 diabetic mice to biofilm infection.

    Science.gov (United States)

    Agostinho Hunt, A M; Gibson, J A; Larrivee, C L; O'Reilly, S; Navitskaya, S; Needle, D B; Abramovitch, R B; Busik, J V; Waters, C M

    2017-07-01

    To examine how bacterial biofilms, as contributing factors in the delayed closure of chronic wounds in patients with diabetes, affect the healing process. We used daily microscopic imaging and the IVIS Spectrum in vivo imaging system to monitor biofilm infections of bioluminescent Pseudomonas aeruginosa and evaluate healing in non-diabetic and streptozotocin-induced diabetic mice. Our studies determined that diabetes alone did not affect the rate of healing of full-depth murine back wounds compared with non-diabetic mice. The application of mature biofilms to the wounds significantly decreased the rate of healing compared with non-infected wounds for both non-diabetic as well as diabetic mice. Diabetic mice were also more severely affected by biofilms displaying elevated pus production, higher mortality rates and statistically significant increase in wound depth, granulation/fibrosis and biofilm presence. Introduction of a mutant Pseudomonas aeruginosa capable of producing high concentrations of cyclic di-GMP did not result in increased persistence in either diabetic or non-diabetic animals compared with the wild type strain. Understanding the interplay between diabetes and biofilms may lead to novel treatments and better clinical management of chronic wounds.

  13. Laboratory-Cultured Strains of the Sea Anemone Exaiptasia Reveal Distinct Bacterial Communities

    KAUST Repository

    Herrera Sarrias, Marcela

    2017-05-02

    Exaiptasia is a laboratory sea anemone model system for stony corals. Two clonal strains are commonly used, referred to as H2 and CC7, that originate from two genetically distinct lineages and that differ in their Symbiodinium specificity. However, little is known about their other microbial associations. Here, we examined and compared the taxonomic composition of the bacterial assemblages of these two symbiotic Exaiptasia strains, both of which have been cultured in the laboratory long-term under identical conditions. We found distinct bacterial microbiota for each strain, indicating the presence of host-specific microbial consortia. Putative differences in the bacterial functional profiles (i.e., enrichment and depletion of various metabolic processes) based on taxonomic inference were also detected, further suggesting functional differences of the microbiomes associated with these lineages. Our study contributes to the current knowledge of the Exaiptasia holobiont by comparing the bacterial diversity of two commonly used strains as models for coral research.

  14. The structure of a conserved piezo channel domain reveals a topologically distinct β sandwich fold.

    Science.gov (United States)

    Kamajaya, Aron; Kaiser, Jens T; Lee, Jonas; Reid, Michelle; Rees, Douglas C

    2014-10-07

    Piezo has recently been identified as a family of eukaryotic mechanosensitive channels composed of subunits containing over 2,000 amino acids, without recognizable sequence similarity to other channels. Here, we present the crystal structure of a large, conserved extramembrane domain located just before the last predicted transmembrane helix of C. elegans PIEZO, which adopts a topologically distinct β sandwich fold. The structure was also determined of a point mutation located on a conserved surface at the position equivalent to the human PIEZO1 mutation found in dehydrated hereditary stomatocytosis patients (M2225R). While the point mutation does not change the overall domain structure, it does alter the surface electrostatic potential that may perturb interactions with a yet-to-be-identified ligand or protein. The lack of structural similarity between this domain and any previously characterized fold, including those of eukaryotic and bacterial channels, highlights the distinctive nature of the Piezo family of eukaryotic mechanosensitive channels.

  15. Scanning Electron Microscopy Reveals Two Distinct Classes of Erythroblastic Island Isolated from Adult Mammalian Bone Marrow.

    Science.gov (United States)

    Yeo, Jia Hao; McAllan, Bronwyn M; Fraser, Stuart T

    2016-04-01

    Erythroblastic islands are multicellular clusters in which a central macrophage supports the development and maturation of red blood cell (erythroid) progenitors. These clusters play crucial roles in the pathogenesis observed in animal models of hematological disorders. The precise structure and function of erythroblastic islands is poorly understood. Here, we have combined scanning electron microscopy and immuno-gold labeling of surface proteins to develop a better understanding of the ultrastructure of these multicellular clusters. The erythroid-specific surface antigen Ter-119 and the transferrin receptor CD71 exhibited distinct patterns of protein sorting during erythroid cell maturation as detected by immuno-gold labeling. During electron microscopy analysis we observed two distinct classes of erythroblastic islands. The islands varied in size and morphology, and the number and type of erythroid cells interacting with the central macrophage. Assessment of femoral marrow isolated from a cavid rodent species (guinea pig, Cavis porcellus) and a marsupial carnivore species (fat-tailed dunnarts, Sminthopsis crassicaudata) showed that while the morphology of the central macrophage varied, two different types of erythroblastic islands were consistently identifiable. Our findings suggest that these two classes of erythroblastic islands are conserved in mammalian evolution and may play distinct roles in red blood cell production.

  16. Evolutionary adaptations of biofilms infecting cystic fibrosis lungs promote mechanical toughness by adjusting polysaccharide production.

    Science.gov (United States)

    Kovach, Kristin; Davis-Fields, Megan; Irie, Yasuhiko; Jain, Kanishk; Doorwar, Shashvat; Vuong, Katherine; Dhamani, Numa; Mohanty, Kishore; Touhami, Ahmed; Gordon, Vernita D

    2017-01-01

    Biofilms are communities of microbes embedded in a matrix of extracellular polymeric substances, largely polysaccharides. Multiple types of extracellular polymeric substances can be produced by a single bacterial strain. The distinct polymer components of biofilms are known to provide chemical protection, but little is known about how distinct extracellular polysaccharides may also protect biofilms against mechanical stresses such as shear or phagocytic engulfment. Decades-long infections of Pseudomonas. aeruginosa biofilms in the lungs of cystic fibrosis patients are natural models for studies of biofilm fitness under pressure from antibiotics and the immune system. In cystic fibrosis infections, production of the extracellular polysaccharide alginate has long been known to increase with time and to chemically protect biofilms. More recently, it is being recognized that chronic cystic fibrosis infections also evolve to increase production of another extracellular polysaccharide, Psl; much less is known about Psl's protective benefits to biofilms. We use oscillatory bulk rheology, on biofilms grown from longitudinal clinical isolates and from genetically-manipulated lab strains, to show that increased Psl stiffens biofilms and increases biofilm toughness, which is the energy cost to cause the biofilm to yield mechanically. Further, atomic force microscopy measurements reveal greater intercellular cohesion for higher Psl expression. Of the three types of extracellular polysaccharides produced by P. aeruginosa, only Psl increases the stiffness. Stiffening by Psl requires CdrA, a protein that binds to mannose groups on Psl and is a likely cross-linker for the Psl components of the biofilm matrix. We compare the elastic moduli of biofilms to the estimated stresses exerted by neutrophils during phagocytosis, and infer that increased Psl could confer a mechanical protection against phagocytic clearance.

  17. Reconstructing dynamic mental models of facial expressions in prosopagnosia reveals distinct representations for identity and expression.

    Science.gov (United States)

    Richoz, Anne-Raphaëlle; Jack, Rachael E; Garrod, Oliver G B; Schyns, Philippe G; Caldara, Roberto

    2015-04-01

    The human face transmits a wealth of signals that readily provide crucial information for social interactions, such as facial identity and emotional expression. Yet, a fundamental question remains unresolved: does the face information for identity and emotional expression categorization tap into common or distinct representational systems? To address this question we tested PS, a pure case of acquired prosopagnosia with bilateral occipitotemporal lesions anatomically sparing the regions that are assumed to contribute to facial expression (de)coding (i.e., the amygdala, the insula and the posterior superior temporal sulcus--pSTS). We previously demonstrated that PS does not use information from the eye region to identify faces, but relies on the suboptimal mouth region. PS's abnormal information use for identity, coupled with her neural dissociation, provides a unique opportunity to probe the existence of a dichotomy in the face representational system. To reconstruct the mental models of the six basic facial expressions of emotion in PS and age-matched healthy observers, we used a novel reverse correlation technique tracking information use on dynamic faces. PS was comparable to controls, using all facial features to (de)code facial expressions with the exception of fear. PS's normal (de)coding of dynamic facial expressions suggests that the face system relies either on distinct representational systems for identity and expression, or dissociable cortical pathways to access them. Interestingly, PS showed a selective impairment for categorizing many static facial expressions, which could be accounted for by her lesion in the right inferior occipital gyrus. PS's advantage for dynamic facial expressions might instead relate to a functionally distinct and sufficient cortical pathway directly connecting the early visual cortex to the spared pSTS. Altogether, our data provide critical insights on the healthy and impaired face systems, question evidence of deficits

  18. Feeding characteristics reveal functional distinctions among browsing herbivorous fishes on coral reefs

    Science.gov (United States)

    Streit, Robert P.; Hoey, Andrew S.; Bellwood, David R.

    2015-12-01

    The removal of macroalgal biomass by fishes is a key process on coral reefs. Numerous studies have identified the fish species responsible for removing mature macroalgae, and have identified how this varies spatially, temporally, and among different algal types. None, however, have considered the behavioural and morphological traits of the browsing fishes and how this may influence the removal of macroalgal material. Using video observations of fish feeding on the brown macroalga Sargassum polycystum, we quantified the feeding behaviour and morphology of the four dominant browsing species on the Great Barrier Reef ( Kyphosus vaigiensis, Naso unicornis, Siganus canaliculatus, and Siganus doliatus). The greatest distinction between species was the algal material they targeted. K. vaigiensis and N. unicornis bit on the entire macroalgal thallus in approximately 90 % of bites. In contrast, Si. canaliculatus and Si. doliatus avoided biting the stalks, with 80-98 % of bites being on the macroalgal leaves only. This distinctive grouping into `entire thallus-biters' versus `leaf-biters' was not supported by size-standardized measures of biting morphology. Rather, species-specific adult body sizes, tooth shape, and feeding behaviour appear to underpin this functional distinction, with adults of the two larger fish species ( N. unicornis and K. vaigiensis) eating the entire macroalgal thallus, while the two smaller species ( Si. canaliculatus and Si. doliatus) bite only leaves. These findings caution against assumed homogeneity within this, and potentially other, functional groups on coral reefs. As functional redundancy within the macroalgal browsers is limited, the smaller `leaf-biting' species are unlikely to be able to compensate functionally for the loss of larger `entire thallus-biting' species.

  19. Distinct populations of innate CD8+ T cells revealed in a CXCR3 reporter mouse.

    Science.gov (United States)

    Oghumu, Steve; Dong, Ran; Varikuti, Sanjay; Shawler, Todd; Kampfrath, Thomas; Terrazas, Cesar A; Lezama-Davila, Claudio; Ahmer, Brian M M; Whitacre, Caroline C; Rajagopalan, Sanjay; Locksley, Richard; Sharpe, Arlene H; Satoskar, Abhay R

    2013-03-01

    CXCR3, expressed mainly on activated T and NK cells, is implicated in a host of immunological conditions and can contribute either to disease resolution or pathology. We report the generation and characterization of a novel CXCR3 internal ribosome entry site bicistronic enhanced GFP reporter (CIBER) mouse in which enhanced GFP expression correlates with surface levels of CXCR3. Using CIBER mice, we identified two distinct populations of innate CD8(+) T cells based on constitutive expression of CXCR3. We demonstrate that CXCR3(+) innate CD8(+) T cells preferentially express higher levels of Ly6C and CD122, but lower levels of CCR9 compared with CXCR3(-) innate CD8(+) T cells. Furthermore, we show that CXCR3(+) innate CD8(+) T cells express higher transcript levels of antiapoptotic but lower levels of proapoptotic factors, respond more robustly to IL-2 and IL-15, and produce significantly more IFN-γ and granzyme B. Interestingly, CXCR3(+) innate CD8(+) T cells do not respond to IL-12 or IL-18 alone, but produce significant amounts of IFN-γ on stimulation with a combination of these cytokines. Taken together, these findings demonstrate that CXCR3(+) and CXCR3(-) innate CD8(+) T cells are phenotypically and functionally distinct. These newly generated CIBER mice provide a novel tool for studying the role of CXCR3 and CXCR3-expressing cells in vivo.

  20. Proteomic analyses reveal distinct chromatin-associated and soluble transcription factor complexes.

    Science.gov (United States)

    Li, Xu; Wang, Wenqi; Wang, Jiadong; Malovannaya, Anna; Xi, Yuanxin; Li, Wei; Guerra, Rudy; Hawke, David H; Qin, Jun; Chen, Junjie

    2015-01-21

    The current knowledge on how transcription factors (TFs), the ultimate targets and executors of cellular signalling pathways, are regulated by protein-protein interactions remains limited. Here, we performed proteomics analyses of soluble and chromatin-associated complexes of 56 TFs, including the targets of many signalling pathways involved in development and cancer, and 37 members of the Forkhead box (FOX) TF family. Using tandem affinity purification followed by mass spectrometry (TAP/MS), we performed 214 purifications and identified 2,156 high-confident protein-protein interactions. We found that most TFs form very distinct protein complexes on and off chromatin. Using this data set, we categorized the transcription-related or unrelated regulators for general or specific TFs. Our study offers a valuable resource of protein-protein interaction networks for a large number of TFs and underscores the general principle that TFs form distinct location-specific protein complexes that are associated with the different regulation and diverse functions of these TFs.

  1. Cortical Connectivity Maps Reveal Anatomically Distinct Areas in the Parietal Cortex of the Rat

    Directory of Open Access Journals (Sweden)

    Aaron eWilber

    2015-01-01

    Full Text Available A central feature of theories of spatial navigation involves the representation of spatial relationships between objects in complex environments. The parietal cortex has long been linked to the processing of spatial visual information and recent evidence from single unit recording in rodents suggests a role for this region in encoding egocentric and world-centered frames. The rat parietal cortex can be subdivided into up to four distinct rostral-caudal and medial-lateral regions, which includes a zone previously characterized as secondary visual cortex. At present, very little is known regarding the relative connectivity of these parietal subdivisions. Thus, we set out to map the connectivity of the entire anterior-posterior and medial-lateral span of this region. To do this we used anterograde and retrograde tracers in conjunction with open source neuronal segmentation and tracer detection tools to generate whole brain connectivity maps of parietal inputs and outputs. Our present results show that inputs to the parietal cortex varied significantly along the medial-lateral, but not the rostral-caudal axis. Specifically, retrosplenial connectivity is greater medially, but connectivity with visual cortex, though generally sparse, is more significant laterally. Finally, based on connection density, the connectivity between parietal cortex and hippocampus is indirect and likely achieved largely via dysgranular retrosplenial cortex. Thus, similar to primates, the parietal cortex of rats exhibits a difference in connectivity along the medial-lateral axis, which may represent functionally distinct areas.

  2. An overexpression screen of Toxoplasma gondii Rab-GTPases reveals distinct transport routes to the micronemes.

    Directory of Open Access Journals (Sweden)

    Katrin Kremer

    2013-03-01

    Full Text Available The basic organisation of the endomembrane system is conserved in all eukaryotes and comparative genome analyses provides compelling evidence that the endomembrane system of the last common eukaryotic ancestor (LCEA is complex with many genes required for regulated traffic being present. Although apicomplexan parasites, causative agents of severe human and animal diseases, appear to have only a basic set of trafficking factors such as Rab-GTPases, they evolved unique secretory organelles (micronemes, rhoptries and dense granules that are sequentially secreted during invasion of the host cell. In order to define the secretory pathway of apicomplexans, we performed an overexpression screen of Rabs in Toxoplasma gondii and identified Rab5A and Rab5C as important regulators of traffic to micronemes and rhoptries. Intriguingly, we found that not all microneme proteins traffic depends on functional Rab5A and Rab5C, indicating the existence of redundant microneme targeting pathways. Using two-colour super-resolution stimulated emission depletion (STED we verified distinct localisations of independent microneme proteins and demonstrate that micronemal organelles are organised in distinct subsets or subcompartments. Our results suggest that apicomplexan parasites modify classical regulators of the endocytic system to carryout essential parasite-specific roles in the biogenesis of their unique secretory organelles.

  3. The Danger Signal Extracellular ATP Is an Inducer of Fusobacterium nucleatum Biofilm Dispersal

    Science.gov (United States)

    Ding, Qinfeng; Tan, Kai Soo

    2016-01-01

    Plaque biofilm is the primary etiological agent of periodontal disease. Biofilm formation progresses through multiple developmental stages beginning with bacterial attachment to a surface, followed by development of microcolonies and finally detachment and dispersal from a mature biofilm as free planktonic bacteria. Tissue damage arising from inflammatory response to biofilm is one of the hallmark features of periodontal disease. A consequence of tissue damage is the release of ATP from within the cell into the extracellular space. Extracellular ATP (eATP) is an example of a danger associated molecular pattern (DAMP) employed by mammalian cells to elicit inflammatory and damage healing responses. Although, the roles of eATP as a signaling molecule in multi-cellular organisms have been relatively well studied, exogenous ATP also influences bacteria biofilm formation. Since plaque biofilms are continuously exposed to various stresses including exposure to the host damage factors such as eATP, we hypothesized that eATP, in addition to eliciting inflammation could potentially influence the biofilm lifecycle of periodontal associated bacteria. We found that eATP rather than nutritional factors or oxidative stress induced dispersal of Fusobacterium nucleatum, an organism associated with periodontal disease. eATP induced biofilm dispersal through chelating metal ions present in biofilm. Dispersed F. nucleatum biofilm, regardless of natural or induced dispersal by exogenous ATP, were more adhesive and invasive compared to planktonic or biofilm counterparts, and correspondingly activated significantly more pro-inflammatory cytokine production in infected periodontal fibroblasts. Dispersed F. nucleatum also showed higher expression of fadA, a virulence factor implicated in adhesion and invasion, compared to planktonic or biofilm bacteria. This study revealed for the first time that periodontal bacterium is capable of co-opting eATP, a host danger signaling molecule to detach

  4. Biofilm Risks

    DEFF Research Database (Denmark)

    Wirtanen, Gun Linnea; Salo, Satu

    2016-01-01

    This chapter on biofilm risks deals with biofilm formation of pathogenic microbes, sampling and detection methods, biofilm removal, and prevention of biofilm formation. Several common pathogens produce sticky and/or slimy structures in which the cells are embedded, that is, biofilms, on various s...

  5. Phenotypic and metabolic profiling of colony morphology variants evolved from Pseudomonas fluorescens biofilms.

    Science.gov (United States)

    Workentine, Matthew L; Harrison, Joe J; Weljie, Aalim M; Tran, Vy A; Stenroos, Pernilla U; Tremaroli, Valentina; Vogel, Hans J; Ceri, Howard; Turner, Raymond J

    2010-06-01

    Colony morphology variants isolated from natural and laboratory-grown biofilms represent subpopulations of biofilm cells that may be important for multiple aspects of the sessile lifestyle, from surface colonization to stress resistance. There are many genetic and environmental factors that determine the frequency at which colony morphology variants are recovered from biofilms. One of these factors involves an increased selection for variants in biofilms of Pseudomonas species bearing inactivating mutations in the global activator of cyanide biosynthesis/regulator of secondary metabolism (gac/rsm) signal transduction pathway. Here we characterize two distinct colony morphology variants isolated from biofilms of Pseudomonas fluorescens missing the gacS sensor kinase. These variants produced more biofilm cell mass, and in one case, this was likely due to overproduction of the exopolysaccharide cellulose. Nuclear magnetic resonance (NMR) metabolomics revealed distinct metabolic changes for each of the two phenotypic variants, and these changes involved amino acids and metabolites produced through glutathione biochemistry. Some of these metabolites are hypothesized to play a role in redox and metal homeostasis, and corresponding to this, we show that biofilm populations grown from each of these variants had a different ability to survive when exposed to toxic doses of metal ions. These data suggest that colony morphology variants that evolve during growth of P. fluorescens as a biofilm may have distinct metabolic capacities that contribute to their individual abilities to withstand environmental stress.

  6. An eye-to-hand magnet effect reveals distinct spatial interference in motor planning and execution.

    Science.gov (United States)

    Richardson, Brian A; Cluff, Tyler; Lyons, James; Balasubramaniam, Ramesh

    2013-03-01

    An important question in oculomanual control is whether motor planning and execution modulate interference between motion of the eyes and hands. Here we investigated oculomanual interference using a novel paradigm that required saccadic eye movements and unimanual finger tapping. We examined finger trajectories for spatial interference caused by concurrent saccades. The first experiment used synchronous cues so that saccades and taps shared a common timekeeping goal. We found that finger trajectories showed bilateral interference where either finger was attracted in the direction of the accompanying saccade. The second experiment avoided interference due to shared planning resources by examining interference caused by reactive saccades. Here, we observed a lesser degree of execution-dependent coupling where the finger trajectory deviated only when reactive saccades were directed toward the hemifield of the responding hand. Our results show that distinct forms of eye-to-hand coupling emerge according to the demands of the task.

  7. Functional chromatography reveals three natural products that target the same protein with distinct mechanisms of action

    Science.gov (United States)

    Kang, MinJin; Wu, Tongde; Wijeratne, E. M. Kithsiri; Lau, Eric C.; Mason, Damian J.; Mesa, Celestina; Tillotson, Joseph; Zhang, Donna D.; Gunatilaka, A. A. Leslie; La Clair, James J.

    2014-01-01

    Access to lead compounds with defined molecular targets continues to be a barrier to the translation of natural product resources. As a solution, we have developed a system that uses discreet, recombinant proteins as the vehicles for natural product isolation. Here, we describe the use of this functional chromatographic method to identify natural products that bind to the AAA+ chaperone, p97, a promising cancer target. Application of this method to a panel of fungal and plant extracts identified rheoemodin, 1-hydroxydehydroherbarin and phomapyrrolidone A as distinct p97 modulators. Excitingly, each of these molecules displayed a unique mechanism of p97 modulation. This discovery provides strong support for the application of functional chromatography to the discovery of protein modulators that would likely escape traditional high-throughput or phenotypic screening platforms. PMID:25125376

  8. Distinctive anatomical and physiological features of migraine aura revealed by 18 years of recording

    DEFF Research Database (Denmark)

    Hansen, Jakob Møller; Baca, Serapio Michael; Vanvalkenburgh, Paul

    2013-01-01

    or scotoma in the visual field at one minute intervals. These drawings were digitized and the spatial and temporal features of auras were quantified and analysed. Consistent patterns of aura initiation, propagation and termination were observed in both right and left visual fields. Most aura attacks...... originated centrally (within 10° eccentricity), but there were also other distinct sites of initiation in the visual field. Auras beginning centrally preferentially propagated first through lower nasal field (69-77% of all auras) before travelling to upper and temporal fields, on both sides. Some auras...... propagated from peripheral to central regions of the visual field-these typically followed the reverse path of those travelling in the opposite direction. The mean velocity of the perceived visual phenomenon did not differ between attacks starting peripherally and centrally. The estimated speed...

  9. Identifying Two Groups of Entitled Individuals: Cluster Analysis Reveals Emotional Stability and Self-Esteem Distinction.

    Science.gov (United States)

    Crowe, Michael L; LoPilato, Alexander C; Campbell, W Keith; Miller, Joshua D

    2016-12-01

    The present study hypothesized that there exist two distinct groups of entitled individuals: grandiose-entitled, and vulnerable-entitled. Self-report scores of entitlement were collected for 916 individuals using an online platform. Model-based cluster analyses were conducted on the individuals with scores one standard deviation above mean (n = 159) using the five-factor model dimensions as clustering variables. The results support the existence of two groups of entitled individuals categorized as emotionally stable and emotionally vulnerable. The emotionally stable cluster reported emotional stability, high self-esteem, more positive affect, and antisocial behavior. The emotionally vulnerable cluster reported low self-esteem and high levels of neuroticism, disinhibition, conventionality, psychopathy, negative affect, childhood abuse, intrusive parenting, and attachment difficulties. Compared to the control group, both clusters reported being more antagonistic, extraverted, Machiavellian, and narcissistic. These results suggest important differences are missed when simply examining the linear relationships between entitlement and various aspects of its nomological network.

  10. Physical versus psychological social stress in male rats reveals distinct cardiovascular, inflammatory and behavioral consequences

    Science.gov (United States)

    Padi, Akhila R.; Moffitt, Casey M.; Wilson, L. Britt; Wood, Christopher S.; Wood, Susan K.

    2017-01-01

    Repeated exposure to social stress can precipitate the development of psychosocial disorders including depression and comorbid cardiovascular disease. While a major component of social stress often encompasses physical interactions, purely psychological stressors (i.e. witnessing a traumatic event) also fall under the scope of social stress. The current study determined whether the acute stress response and susceptibility to stress-related consequences differed based on whether the stressor consisted of physical versus purely psychological social stress. Using a modified resident-intruder paradigm, male rats were either directly exposed to repeated social defeat stress (intruder) or witnessed a male rat being defeated. Cardiovascular parameters, behavioral anhedonia, and inflammatory cytokines in plasma and the stress-sensitive locus coeruleus were compared between intruder, witness, and control rats. Surprisingly intruders and witnesses exhibited nearly identical increases in mean arterial pressure and heart rate during acute and repeated stress exposures, yet only intruders exhibited stress-induced arrhythmias. Furthermore, re-exposure to the stress environment in the absence of the resident produced robust pressor and tachycardic responses in both stress conditions indicating the robust and enduring nature of social stress. In contrast, the long-term consequences of these stressors were distinct. Intruders were characterized by enhanced inflammatory sensitivity in plasma, while witnesses were characterized by the emergence of depressive-like anhedonia, transient increases in systolic blood pressure and plasma levels of tissue inhibitor of metalloproteinase. The current study highlights that while the acute cardiovascular responses to stress were identical between intruders and witnesses, these stressors produced distinct differences in the enduring consequences to stress, suggesting that witness stress may be more likely to produce long-term cardiovascular

  11. Large-scale tectonic cycles in Europe revealed by distinct Pb isotope provinces

    Science.gov (United States)

    Blichert-Toft, Janne; Delile, Hugo; Lee, Cin-Ty; Stos-Gale, Zofia; Billström, Kjell; Andersen, Tom; Hannu, Huhma; Albarède, Francis

    2016-10-01

    Lead isotopic systematics of U-poor minerals, such as sulfides and feldspars, can provide unique insights into the origin and evolution of continents because these minerals "freeze in" the Pb isotopic composition of the crust during major tectonothermal events, allowing the history of a continent to be told through Pb isotopes. Lead model ages constrain the timing of crust formation while time-integrated U/Pb, Th/Pb, and Th/U ratios shed light onto key geochemical processes associated with continent formation. Using ˜6800 Pb isotope measurements of primarily lead ores and minor K-feldspar, we mapped out the Pb isotope systematics across Europe and the Mediterranean. Lead model ages define spatially distinct age provinces, consistent with major tectonic events ranging from the Paleozoic to the Proterozoic and latest Archean. However, the regions defined by time-integrated U/Pb and Th/Pb ratios cut across the boundaries of age provinces, with high U/Pb systematics characterizing most of southern Europe. Magmatic influx, followed by segregation of dense sulfide-rich mafic cumulates, resulted in foundering of U- and Th-poor lower crust, thereby changing the bulk composition of the continental crust and leading to distinct time-integrated U-Th/Pb provinces. We show that the tectonic assembly of small crustal fragments leaves the crust largely undifferentiated, whereas the formation of supercontinents results in fundamental changes in the composition of the crust, identifiable in time and space by means of Pb isotope systematics. Observations based on Pb isotopes open up a new perspective on possible relationships between crustal thickness and geodynamic processes, in particular the role of crustal foundering into the mantle and the mechanisms responsible for the existence of cratons.

  12. Phylogenetic variation of Aggregatibacter actinomycetemcomitans serotype e reveals an aberrant distinct evolutionary stable lineage

    NARCIS (Netherlands)

    van der Reijden, Wil A.; Brunner, Jorg; Bosch-Tijhof, Carolien J.; van Trappen, Stefanie; Rijnsburger, Martine C.; de Graaff, Marcel P. W.; van Winkelhoff, Arie J.; Cleenwerck, Ilse; de Vos, Paul

    2010-01-01

    The periodontal pathogen Aggregatibacter actinomycetemcomitans that comprises six serotypes (a-f), is often identified by PCR-based techniques targeting the 16S rRNA gene. In this study, 16S rRNA gene sequence analysis revealed an aberrant cluster of 19 strains within serotype e, denoted as serotype

  13. Comparative genomics in acid mine drainage biofilm communities reveals metabolic and structural differentiation of co-occurring archaea

    Science.gov (United States)

    2013-01-01

    Background Metal sulfide mineral dissolution during bioleaching and acid mine drainage (AMD) formation creates an environment that is inhospitable to most life. Despite dominance by a small number of bacteria, AMD microbial biofilm communities contain a notable variety of coexisting and closely related Euryarchaea, most of which have defied cultivation efforts. For this reason, we used metagenomics to analyze variation in gene content that may contribute to niche differentiation among co-occurring AMD archaea. Our analyses targeted members of the Thermoplasmatales and related archaea. These results greatly expand genomic information available for this archaeal order. Results We reconstructed near-complete genomes for uncultivated, relatively low abundance organisms A-, E-, and Gplasma, members of Thermoplasmatales order, and for a novel organism, Iplasma. Genomic analyses of these organisms, as well as Ferroplasma type I and II, reveal that all are facultative aerobic heterotrophs with the ability to use many of the same carbon substrates, including methanol. Most of the genomes share genes for toxic metal resistance and surface-layer production. Only Aplasma and Eplasma have a full suite of flagellar genes whereas all but the Ferroplasma spp. have genes for pili production. Cryogenic-electron microscopy (cryo-EM) and tomography (cryo-ET) strengthen these metagenomics-based ultrastructural predictions. Notably, only Aplasma, Gplasma and the Ferroplasma spp. have predicted iron oxidation genes and Eplasma and Iplasma lack most genes for cobalamin, valine, (iso)leucine and histidine synthesis. Conclusion The Thermoplasmatales AMD archaea share a large number of metabolic capabilities. All of the uncultivated organisms studied here (A-, E-, G-, and Iplasma) are metabolically very similar to characterized Ferroplasma spp., differentiating themselves mainly in their genetic capabilities for biosynthesis, motility, and possibly iron oxidation. These results indicate that

  14. Distinctive anatomical and physiological features of migraine aura revealed by 18 years of recording.

    Science.gov (United States)

    Hansen, Jakob Møller; Baca, Serapio Michael; Vanvalkenburgh, Paul; Charles, Andrew

    2013-12-01

    The mechanisms underlying the initiation and propagation of the migraine aura, and the visual percept that is produces, remain uncertain. The objective of this study was to characterize and quantify a large number of visual auras recorded by a single individual over nearly two decades to gain insight into basic aura mechanisms. An individual made detailed drawings of his visual percept of migraine aura in real time during more than 1000 attacks of migraine aura without headache over 18 years. Drawings were made in a consistent fashion documenting the shape and location of the aura wavefront or scotoma in the visual field at one minute intervals. These drawings were digitized and the spatial and temporal features of auras were quantified and analysed. Consistent patterns of aura initiation, propagation and termination were observed in both right and left visual fields. Most aura attacks originated centrally (within 10° eccentricity), but there were also other distinct sites of initiation in the visual field. Auras beginning centrally preferentially propagated first through lower nasal field (69-77% of all auras) before travelling to upper and temporal fields, on both sides. Some auras propagated from peripheral to central regions of the visual field-these typically followed the reverse path of those travelling in the opposite direction. The mean velocity of the perceived visual phenomenon did not differ between attacks starting peripherally and centrally. The estimated speed of the underlying cortical event (2-3 mm/min) was in the same range as has been previously reported by others. Some auras had limited propagation and spontaneously 'aborted' after a few minutes, despite being initiated in similar locations to those that spread throughout the entire visual field. The visual percept of the aura changed corresponding with the presumed propagation from the V1 to the V2 region of the occipital cortex. In some cases the visual percept disappeared for several minutes

  15. Extensive cargo identification reveals distinct biological roles of the 12 importin pathways

    Science.gov (United States)

    Kimura, Makoto; Morinaka, Yuriko; Imai, Kenichiro; Kose, Shingo; Horton, Paul; Imamoto, Naoko

    2017-01-01

    Vast numbers of proteins are transported into and out of the nuclei by approximately 20 species of importin-β family nucleocytoplasmic transport receptors. However, the significance of the multiple parallel transport pathways that the receptors constitute is poorly understood because only limited numbers of cargo proteins have been reported. Here, we identified cargo proteins specific to the 12 species of human import receptors with a high-throughput method that employs stable isotope labeling with amino acids in cell culture, an in vitro reconstituted transport system, and quantitative mass spectrometry. The identified cargoes illuminated the manner of cargo allocation to the receptors. The redundancies of the receptors vary widely depending on the cargo protein. Cargoes of the same receptor are functionally related to one another, and the predominant protein groups in the cargo cohorts differ among the receptors. Thus, the receptors are linked to distinct biological processes by the nature of their cargoes. DOI: http://dx.doi.org/10.7554/eLife.21184.001 PMID:28117667

  16. Recombination analysis of Soybean mosaic virus sequences reveals evidence of RNA recombination between distinct pathotypes

    Directory of Open Access Journals (Sweden)

    Babu Mohan

    2008-11-01

    Full Text Available Abstract RNA recombination is one of the two major factors that create RNA genome variability. Assessing its incidence in plant RNA viruses helps understand the formation of new isolates and evaluate the effectiveness of crop protection strategies. To search for recombination in Soybean mosaic virus (SMV, the causal agent of a worldwide seed-borne, aphid-transmitted viral soybean disease, we obtained all full-length genome sequences of SMV as well as partial sequences encoding the N-terminal most (P1 protease and the C-terminal most (capsid protein; CP viral protein. The sequences were analyzed for possible recombination events using a variety of automatic and manual recombination detection and verification approaches. Automatic scanning identified 3, 10, and 17 recombination sites in the P1, CP, and full-length sequences, respectively. Manual analyses confirmed 10 recombination sites in three full-length SMV sequences. To our knowledge, this is the first report of recombination between distinct SMV pathotypes. These data imply that different SMV pathotypes can simultaneously infect a host cell and exchange genetic materials through recombination. The high incidence of SMV recombination suggests that recombination plays an important role in SMV evolution. Obtaining additional full-length sequences will help elucidate this role.

  17. Mass Spectrometry-Based Quantitative Metabolomics Revealed a Distinct Lipid Profile in Breast Cancer Patients

    Directory of Open Access Journals (Sweden)

    Yun Yen

    2013-04-01

    Full Text Available Breast cancer accounts for the largest number of newly diagnosed cases in female cancer patients. Although mammography is a powerful screening tool, about 20% of breast cancer cases cannot be detected by this method. New diagnostic biomarkers for breast cancer are necessary. Here, we used a mass spectrometry-based quantitative metabolomics method to analyze plasma samples from 55 breast cancer patients and 25 healthy controls. A number of 30 patients and 20 age-matched healthy controls were used as a training dataset to establish a diagnostic model and to identify potential biomarkers. The remaining samples were used as a validation dataset to evaluate the predictive accuracy for the established model. Distinct separation was obtained from an orthogonal partial least squares-discriminant analysis (OPLS-DA model with good prediction accuracy. Based on this analysis, 39 differentiating metabolites were identified, including significantly lower levels of lysophosphatidylcholines and higher levels of sphingomyelins in the plasma samples obtained from breast cancer patients compared with healthy controls. Using logical regression, a diagnostic equation based on three metabolites (lysoPC a C16:0, PC ae C42:5 and PC aa C34:2 successfully differentiated breast cancer patients from healthy controls, with a sensitivity of 98.1% and a specificity of 96.0%.

  18. Iron isotopes reveal distinct dissolved iron sources and pathways in the intermediate versus deep Southern Ocean.

    Science.gov (United States)

    Abadie, Cyril; Lacan, Francois; Radic, Amandine; Pradoux, Catherine; Poitrasson, Franck

    2017-01-31

    As an essential micronutrient, iron plays a key role in oceanic biogeochemistry. It is therefore linked to the global carbon cycle and climate. Here, we report a dissolved iron (DFe) isotope section in the South Atlantic and Southern Ocean. Throughout the section, a striking DFe isotope minimum (light iron) is observed at intermediate depths (200-1,300 m), contrasting with heavier isotopic composition in deep waters. This unambiguously demonstrates distinct DFe sources and processes dominating the iron cycle in the intermediate and deep layers, a feature impossible to see with only iron concentration data largely used thus far in chemical oceanography. At intermediate depths, the data suggest that the dominant DFe sources are linked to organic matter remineralization, either in the water column or at continental margins. In deeper layers, however, abiotic non-reductive release of Fe (desorption, dissolution) from particulate iron-notably lithogenic-likely dominates. These results go against the common but oversimplified view that remineralization of organic matter is the major pathway releasing DFe throughout the water column in the open ocean. They suggest that the oceanic iron cycle, and therefore oceanic primary production and climate, could be more sensitive than previously thought to continental erosion (providing lithogenic particles to the ocean), particle transport within the ocean, dissolved/particle interactions, and deep water upwelling. These processes could also impact the cycles of other elements, including nutrients.

  19. Nonpolarized signaling reveals two distinct modes of 3D cell migration.

    Science.gov (United States)

    Petrie, Ryan J; Gavara, Núria; Chadwick, Richard S; Yamada, Kenneth M

    2012-04-30

    We search in this paper for context-specific modes of three-dimensional (3D) cell migration using imaging for phosphatidylinositol (3,4,5)-trisphosphate (PIP3) and active Rac1 and Cdc42 in primary fibroblasts migrating within different 3D environments. In 3D collagen, PIP3 and active Rac1 and Cdc42 were targeted to the leading edge, consistent with lamellipodia-based migration. In contrast, elongated cells migrating inside dermal explants and the cell-derived matrix (CDM) formed blunt, cylindrical protrusions, termed lobopodia, and Rac1, Cdc42, and PIP3 signaling was nonpolarized. Reducing RhoA, Rho-associated protein kinase (ROCK), or myosin II activity switched the cells to lamellipodia-based 3D migration. These modes of 3D migration were regulated by matrix physical properties. Specifically, experimentally modifying the elasticity of the CDM or collagen gels established that nonlinear elasticity supported lamellipodia-based migration, whereas linear elasticity switched cells to lobopodia-based migration. Thus, the relative polarization of intracellular signaling identifies two distinct modes of 3D cell migration governed intrinsically by RhoA, ROCK, and myosin II and extrinsically by the elastic behavior of the 3D extracellular matrix.

  20. Myf5 haploinsufficiency reveals distinct cell fate potentials for adult skeletal muscle stem cells.

    Science.gov (United States)

    Gayraud-Morel, Barbara; Chrétien, Fabrice; Jory, Aurélie; Sambasivan, Ramkumar; Negroni, Elisa; Flamant, Patricia; Soubigou, Guillaume; Coppée, Jean-Yves; Di Santo, James; Cumano, Ana; Mouly, Vincent; Tajbakhsh, Shahragim

    2012-04-01

    Skeletal muscle stem cell fate in adult mice is regulated by crucial transcription factors, including the determination genes Myf5 and Myod. The precise role of Myf5 in regulating quiescent muscle stem cells has remained elusive. Here we show that most, but not all, quiescent satellite cells express Myf5 protein, but at varying levels, and that resident Myf5 heterozygous muscle stem cells are more primed for myogenic commitment compared with wild-type satellite cells. Paradoxically however, heterotypic transplantation of Myf5 heterozygous cells into regenerating muscles results in higher self-renewal capacity compared with wild-type stem cells, whereas myofibre regenerative capacity is not altered. By contrast, Pax7 haploinsufficiency does not show major modifications by transcriptome analysis. These observations provide a mechanism linking Myf5 levels to muscle stem cell heterogeneity and fate by exposing two distinct and opposing phenotypes associated with Myf5 haploinsufficiency. These findings have important implications for how stem cell fates can be modulated by crucial transcription factors while generating a pool of responsive heterogeneous cells.

  1. Internal Transcribed Spacer 1 (ITS1 based sequence typing reveals phylogenetically distinct Ascaris population

    Directory of Open Access Journals (Sweden)

    Koushik Das

    2015-01-01

    Full Text Available Taxonomic differentiation among morphologically identical Ascaris species is a debatable scientific issue in the context of Ascariasis epidemiology. To explain the disease epidemiology and also the taxonomic position of different Ascaris species, genome information of infecting strains from endemic areas throughout the world is certainly crucial. Ascaris population from human has been genetically characterized based on the widely used genetic marker, internal transcribed spacer1 (ITS1. Along with previously reported and prevalent genotype G1, 8 new sequence variants of ITS1 have been identified. Genotype G1 was significantly present among female patients aged between 10 to 15 years. Intragenic linkage disequilibrium (LD analysis at target locus within our study population has identified an incomplete LD value with potential recombination events. A separate cluster of Indian isolates with high bootstrap value indicate their distinct phylogenetic position in comparison to the global Ascaris population. Genetic shuffling through recombination could be a possible reason for high population diversity and frequent emergence of new sequence variants, identified in present and other previous studies. This study explores the genetic organization of Indian Ascaris population for the first time which certainly includes some fundamental information on the molecular epidemiology of Ascariasis.

  2. Phosphoproteomics Reveals Distinct Modes of Mec1/ATR Signaling During DNA Replication

    Science.gov (United States)

    de Oliveira, Francisco Meirelles Bastos; Kim, Dongsung; Cussiol, Jose Renato; Das, Jishnu; Jeong, Min Cheol; Doerfler, Lillian; Schmidt, Kristina Hildegard; Yu, Haiyuan; Smolka, Marcus Bustamante

    2015-01-01

    SUMMARY The Mec1/Tel1 kinases (human ATR/ATM) play numerous roles in the DNA replication stress response. Despite the multi-functionality of these kinases, studies of their in vivo action have mostly relied on a few well-established substrates. Here we employed a combined genetic-phosphoproteomic approach to monitor Mec1/Tel1 signaling in a systematic, unbiased and quantitative manner. Unexpectedly, we find that Mec1 is highly active during normal DNA replication, at levels comparable or higher than Mec1’s activation state induced by replication stress. This “replication-correlated” mode of Mec1 action requires the 9-1-1 clamp and the Dna2 lagging-strand factor, and is distinguishable from Mec1’s action in activating the downstream kinase Rad53. We propose that Mec1/ATR performs key functions during ongoing DNA synthesis that are distinct from their canonical checkpoint role during replication stress. PMID:25752575

  3. Distinct configurations of protein complexes and biochemical pathways revealed by epistatic interaction network motifs

    LENUS (Irish Health Repository)

    Casey, Fergal

    2011-08-22

    Abstract Background Gene and protein interactions are commonly represented as networks, with the genes or proteins comprising the nodes and the relationship between them as edges. Motifs, or small local configurations of edges and nodes that arise repeatedly, can be used to simplify the interpretation of networks. Results We examined triplet motifs in a network of quantitative epistatic genetic relationships, and found a non-random distribution of particular motif classes. Individual motif classes were found to be associated with different functional properties, suggestive of an underlying biological significance. These associations were apparent not only for motif classes, but for individual positions within the motifs. As expected, NNN (all negative) motifs were strongly associated with previously reported genetic (i.e. synthetic lethal) interactions, while PPP (all positive) motifs were associated with protein complexes. The two other motif classes (NNP: a positive interaction spanned by two negative interactions, and NPP: a negative spanned by two positives) showed very distinct functional associations, with physical interactions dominating for the former but alternative enrichments, typical of biochemical pathways, dominating for the latter. Conclusion We present a model showing how NNP motifs can be used to recognize supportive relationships between protein complexes, while NPP motifs often identify opposing or regulatory behaviour between a gene and an associated pathway. The ability to use motifs to point toward underlying biological organizational themes is likely to be increasingly important as more extensive epistasis mapping projects in higher organisms begin.

  4. Ultradeep Human Phosphoproteome Reveals a Distinct Regulatory Nature of Tyr and Ser/Thr-Based Signaling

    Directory of Open Access Journals (Sweden)

    Kirti Sharma

    2014-09-01

    Full Text Available Regulatory protein phosphorylation controls normal and pathophysiological signaling in eukaryotic cells. Despite great advances in mass-spectrometry-based proteomics, the extent, localization, and site-specific stoichiometry of this posttranslational modification (PTM are unknown. Here, we develop a stringent experimental and computational workflow, capable of mapping more than 50,000 distinct phosphorylated peptides in a single human cancer cell line. We detected more than three-quarters of cellular proteins as phosphoproteins and determined very high stoichiometries in mitosis or growth factor signaling by label-free quantitation. The proportion of phospho-Tyr drastically decreases as coverage of the phosphoproteome increases, whereas Ser/Thr sites saturate only for technical reasons. Tyrosine phosphorylation is maintained at especially low stoichiometric levels in the absence of specific signaling events. Unexpectedly, it is enriched on higher-abundance proteins, and this correlates with the substrate KM values of tyrosine kinases. Our data suggest that P-Tyr should be considered a functionally separate PTM of eukaryotic proteomes.

  5. Comparative physiological and proteomic analysis reveal distinct regulation of peach skin quality traits by altitude

    Directory of Open Access Journals (Sweden)

    Evangelos Karagiannis

    2016-11-01

    Full Text Available The role of environment in fruit physiology has been established; however, knowledge regarding the effect of altitude in fruit quality traits is still lacking. Here, skin tissue quality characters were analysed in peach fruit (cv. June Gold, harvested in 16 orchards located in low (71.5 m mean or high (495. m mean altitutes sites. Data indicated that soluble solids concentration and fruit firmness at commercial harvest stage were unaffected by alitute. Peach grown at high-altitude environment displayed higher levels of pigmentation and specific antioxidant-related activity in their skin at the commercial harvest stage. Skin extracts from distinct developmental stages and growing altitudes exhibited different antioxidant ability against DNA strand-scission. The effects of altitude on skin tissue were further studied using a proteomic approach. Protein expression analysis of the mature fruits depicted altered expression of 42 proteins that are mainly involved in the metabolic pathways of defense, primary metabolism, destination/storage and energy. The majority of these proteins were up-regulated at the low-altitude region. High-altitude environment increased the accumulation of several proteins, including chaperone ClpC, chaperone ClpB, pyruvate dehydrogenase E1, TCP domain class transcription factor and lipoxygenase. We also discuss the altitude-affected protein variations, taking into account their potential role in peach ripening process. This study provides the first characterization of the peach skin proteome and helps to improve our understanding of peach’s response to altitude.

  6. Natural resource landscapes of a marine bacterium reveal distinct fitness-determining genes across the genome.

    Science.gov (United States)

    Takemura, Alison F; Corzett, Christopher H; Hussain, Fatima; Arevalo, Philip; Datta, Manoshi; Yu, Xiaoqian; Le Roux, Frederique; Polz, Martin F

    2017-06-01

    Heterotrophic bacteria exploit diverse microhabitats in the ocean, from particles to transient gradients. Yet the degree to which genes and pathways can contribute to an organism's fitness on such complex and variable natural resource landscapes remains poorly understood. Here, we determine the gene-by-gene fitness of a generalist saprophytic marine bacterium (Vibrio sp. F13 9CS106) on complex resources derived from its natural habitats - copepods (Apocyclops royi) and brown algae (Fucus vesiculosus) - and as reference substrates, glucose and the polysaccharide alginate, derived from brown algal cell walls. We find that resource complexity strongly buffers fitness costs of mutations, and that anabolic rather than catabolic pathways are more stringently required, likely due to functional redundancy in the latter. Moreover, while carbohydrate-rich algae requires several synthesis pathways, protein-rich Apocyclops does not, suggesting this ancestral habitat for Vibrios is a replete medium with metabolically redundant substrates. We also identify a candidate fitness trade-off for algal colonization: deletion of mshA increases mutant fitness. Our results demonstrate that gene fitness depends on habitat composition, and suggest that this generalist uses distinct resources in different natural habitats. The results further indicate that substrate replete conditions may lead to relatively relaxed selection on catabolic genes. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

  7. Iron isotopes reveal distinct dissolved iron sources and pathways in the intermediate versus deep Southern Ocean

    Science.gov (United States)

    Abadie, Cyril; Lacan, Francois; Radic, Amandine; Pradoux, Catherine; Poitrasson, Franck

    2017-01-01

    As an essential micronutrient, iron plays a key role in oceanic biogeochemistry. It is therefore linked to the global carbon cycle and climate. Here, we report a dissolved iron (DFe) isotope section in the South Atlantic and Southern Ocean. Throughout the section, a striking DFe isotope minimum (light iron) is observed at intermediate depths (200–1,300 m), contrasting with heavier isotopic composition in deep waters. This unambiguously demonstrates distinct DFe sources and processes dominating the iron cycle in the intermediate and deep layers, a feature impossible to see with only iron concentration data largely used thus far in chemical oceanography. At intermediate depths, the data suggest that the dominant DFe sources are linked to organic matter remineralization, either in the water column or at continental margins. In deeper layers, however, abiotic non-reductive release of Fe (desorption, dissolution) from particulate iron—notably lithogenic—likely dominates. These results go against the common but oversimplified view that remineralization of organic matter is the major pathway releasing DFe throughout the water column in the open ocean. They suggest that the oceanic iron cycle, and therefore oceanic primary production and climate, could be more sensitive than previously thought to continental erosion (providing lithogenic particles to the ocean), particle transport within the ocean, dissolved/particle interactions, and deep water upwelling. These processes could also impact the cycles of other elements, including nutrients.

  8. Quantification of cell edge velocities and traction forces reveals distinct motility modules during cell spreading.

    Directory of Open Access Journals (Sweden)

    Benjamin J Dubin-Thaler

    Full Text Available Actin-based cell motility and force generation are central to immune response, tissue development, and cancer metastasis, and understanding actin cytoskeleton regulation is a major goal of cell biologists. Cell spreading is a commonly used model system for motility experiments -- spreading fibroblasts exhibit stereotypic, spatially-isotropic edge dynamics during a reproducible sequence of functional phases: 1 During early spreading, cells form initial contacts with the surface. 2 The middle spreading phase exhibits rapidly increasing attachment area. 3 Late spreading is characterized by periodic contractions and stable adhesions formation. While differences in cytoskeletal regulation between phases are known, a global analysis of the spatial and temporal coordination of motility and force generation is missing. Implementing improved algorithms for analyzing edge dynamics over the entire cell periphery, we observed that a single domain of homogeneous cytoskeletal dynamics dominated each of the three phases of spreading. These domains exhibited a unique combination of biophysical and biochemical parameters -- a motility module. Biophysical characterization of the motility modules revealed that the early phase was dominated by periodic, rapid membrane blebbing; the middle phase exhibited continuous protrusion with very low traction force generation; and the late phase was characterized by global periodic contractions and high force generation. Biochemically, each motility module exhibited a different distribution of the actin-related protein VASP, while inhibition of actin polymerization revealed different dependencies on barbed-end polymerization. In addition, our whole-cell analysis revealed that many cells exhibited heterogeneous combinations of motility modules in neighboring regions of the cell edge. Together, these observations support a model of motility in which regions of the cell edge exhibit one of a limited number of motility modules

  9. Distinct levels in the nanoscale organization of DNA-histone complex revealed by its mechanical unfolding

    Science.gov (United States)

    Soni, G. V.; Brar, Loveleen; Hameed, Feroz M.; Raychaudhuri, A. K.; Shivashankar, G. V.

    2007-04-01

    Mechanical unfolding of nanoscale DNA-histone complex, using an atomic force microscope, shows a stepwise disassembly of histones from the nucleosome. A quantitative analysis of the rupture jump statistics and the length released per jump reveals insights into the possible histone contacts within the octamer complex. The measured ruptures correlate with the breakage of multiple contacts that stabilize the histone octamer. These results provide a mechanistic basis by which stepwise disassembly of histone proteins may result from an external force exerted by the adenosinetriphosphate (ATP) dependent chromatin remodeling machines to access regulatory sites on DNA.

  10. Large-scale experimental landscapes reveal distinctive effects of patch shape and connectivity on arthropod communities.

    Energy Technology Data Exchange (ETDEWEB)

    Orrock, John, L.; Curler, Gregory, R.; Danielson, Brent, J.; Coyle, David. R.

    2011-09-14

    The size, shape, and isolation of habitat patches can affect organism behavior and population dynamics, but little is known about the relative role of shape and connectivity in affecting ecological communities at large spatial scales. Using six sampling sessions from July 2001 until August 2002, we collected 33,685 arthropods throughout seven 12-ha experimental landscapes consisting of clear-cut patches surrounded by a matrix of mature pine forest. Patches were explicitly designed to manipulate connectivity (via habitat corridors) independently of area and edge effects. We found that patch shape, rather than connectivity, affected ground-dwelling arthropod richness and beta diversity (i.e. turnover of genera among patches). Arthropod communities contained fewer genera and exhibited less turnover in high-edge connected and high-edge unconnected patches relative to low-edge unconnected patches of similar area. Connectivity, rather than patch shape, affected the evenness of ground-dwelling arthropod communities; regardless of patch shape, high-edge connected patches had lower evenness than low- or high-edge unconnected patches. Among the most abundant arthropod orders, increased richness in low-edge unconnected patches was largely due to increased richness of Coleoptera, whereas Hymenoptera played an important role in the lower evenness in connected patches and patterns of turnover. These findings suggest that anthropogenic habitat alteration can have distinct effects on ground-dwelling arthropod communities that arise due to changes in shape and connectivity. Moreover, this work suggests that corridors, which are common conservation tools that change both patch shape and connectivity, can have multiple effects on arthropod communities via different mechanisms, and each effect may alter components of community structure.

  11. Genomic analysis of the TRIM family reveals two groups of genes with distinct evolutionary properties

    Directory of Open Access Journals (Sweden)

    Fontanella Bianca

    2008-08-01

    Full Text Available Abstract Background The TRIM family is composed of multi-domain proteins that display the Tripartite Motif (RING, B-box and Coiled-coil that can be associated with a C-terminal domain. TRIM genes are involved in ubiquitylation and are implicated in a variety of human pathologies, from Mendelian inherited disorders to cancer, and are also involved in cellular response to viral infection. Results Here we defined the entire human TRIM family and also identified the TRIM sets of other vertebrate (mouse, rat, dog, cow, chicken, tetraodon, and zebrafish and invertebrate species (fruitfly, worm, and ciona. By means of comparative analyses we found that, after assembly of the tripartite motif in an early metazoan ancestor, few types of C-terminal domains have been associated with this module during evolution and that an important increase in TRIM number occurred in vertebrate species concomitantly with the addition of the SPRY domain. We showed that the human TRIM family is split into two groups that differ in domain structure, genomic organization and evolutionary properties. Group 1 members present a variety of C-terminal domains, are highly conserved among vertebrate species, and are represented in invertebrates. Conversely, group 2 is absent in invertebrates, is characterized by the presence of a C-terminal SPRY domain and presents unique sets of genes in each mammal examined. The generation of independent sets of group 2 genes is also evident in the other vertebrate species. Comparing the murine and human TRIM sets, we found that group 1 and 2 genes evolve at different speeds and are subject to different selective pressures. Conclusion We found that the TRIM family is composed of two groups of genes with distinct evolutionary properties. Group 2 is younger, highly dynamic, and might act as a reservoir to develop novel TRIM functions. Since some group 2 genes are implicated in innate immune response, their evolutionary features may account for

  12. The Crystal Structure of Streptococcus pyogenes Uridine Phosphorylase Reveals a Distinct Subfamily of Nucleoside Phosphorylases

    Energy Technology Data Exchange (ETDEWEB)

    Tran, Timothy H.; Christoffersen, S.; Allan, Paula W.; Parker, William B.; Piskur, Jure; Serra, I.; Terreni, M.; Ealick, Steven E. (Cornell); (Pavia); (Lund); (Southern Research)

    2011-09-20

    Uridine phosphorylase (UP), a key enzyme in the pyrimidine salvage pathway, catalyzes the reversible phosphorolysis of uridine or 2'-deoxyuridine to uracil and ribose 1-phosphate or 2'-deoxyribose 1-phosphate. This enzyme belongs to the nucleoside phosphorylase I superfamily whose members show diverse specificity for nucleoside substrates. Phylogenetic analysis shows Streptococcus pyogenes uridine phosphorylase (SpUP) is found in a distinct branch of the pyrimidine subfamily of nucleoside phosphorylases. To further characterize SpUP, we determined the crystal structure in complex with the products, ribose 1-phosphate and uracil, at 1.8 {angstrom} resolution. Like Escherichia coli UP (EcUP), the biological unit of SpUP is a hexamer with an ?/? monomeric fold. A novel feature of the active site is the presence of His169, which structurally aligns with Arg168 of the EcUP structure. A second active site residue, Lys162, is not present in previously determined UP structures and interacts with O2 of uracil. Biochemical studies of wild-type SpUP showed that its substrate specificity is similar to that of EcUP, while EcUP is {approx}7-fold more efficient than SpUP. Biochemical studies of SpUP mutants showed that mutations of His169 reduced activity, while mutation of Lys162 abolished all activity, suggesting that the negative charge in the transition state resides mostly on uracil O2. This is in contrast to EcUP for which transition state stabilization occurs mostly at O4.

  13. Distinct and complex bacterial profiles in human periodontitis and health revealed by 16S pyrosequencing.

    Science.gov (United States)

    Griffen, Ann L; Beall, Clifford J; Campbell, James H; Firestone, Noah D; Kumar, Purnima S; Yang, Zamin K; Podar, Mircea; Leys, Eugene J

    2012-06-01

    Periodontitis has a polymicrobial etiology within the framework of a complex microbial ecosystem. With advances in sequencing technologies, comprehensive studies to elucidate bacterial community differences have recently become possible. We used 454 sequencing of 16S rRNA genes to compare subgingival bacterial communities from 29 periodontally healthy controls and 29 subjects with chronic periodontitis. Amplicons from both the V1-2 and V4 regions of the 16S gene were sequenced, yielding 1,393,579 sequences. They were identified by BLAST against a curated oral 16S database, and mapped to 16 phyla, 106 genera, and 596 species. 81% of sequences could be mapped to cultivated species. Differences between health- and periodontitis-associated bacterial communities were observed at all phylogenetic levels, and UniFrac and principal coordinates analysis showed distinct community profiles in health and disease. Community diversity was higher in disease, and 123 species were identified that were significantly more abundant in disease, and 53 in health. Spirochaetes, Synergistetes and Bacteroidetes were more abundant in disease, whereas the Proteobacteria were found at higher levels in healthy controls. Within the phylum Firmicutes, the class Bacilli was health-associated, whereas the Clostridia, Negativicutes and Erysipelotrichia were associated with disease. These results implicate a number of taxa that will be targets for future research. Some, such as Filifactor alocis and many Spirochetes were represented by a large fraction of sequences as compared with previously identified targets. Elucidation of these differences in community composition provides a basis for further understanding the pathogenesis of periodontitis.

  14. γ-Secretase modulator (GSM) photoaffinity probes reveal distinct allosteric binding sites on presenilin.

    Science.gov (United States)

    Pozdnyakov, Nikolay; Murrey, Heather E; Crump, Christina J; Pettersson, Martin; Ballard, T Eric; Am Ende, Christopher W; Ahn, Kwangwook; Li, Yue-Ming; Bales, Kelly R; Johnson, Douglas S

    2013-04-05

    γ-Secretase is an intramembrane aspartyl protease that cleaves the amyloid precursor protein to produce neurotoxic β-amyloid peptides (i.e. Aβ42) that have been implicated in the pathogenesis of Alzheimer disease. Small molecule γ-secretase modulators (GSMs) have emerged as potential disease-modifying treatments for Alzheimer disease because they reduce the formation of Aβ42 while not blocking the processing of γ-secretase substrates. We developed clickable GSM photoaffinity probes with the goal of identifying the target of various classes of GSMs and to better understand their mechanism of action. Here, we demonstrate that the photoaffinity probe E2012-BPyne specifically labels the N-terminal fragment of presenilin-1 (PS1-NTF) in cell membranes as well as in live cells and primary neuronal cultures. The labeling is competed in the presence of the parent imidazole GSM E2012, but not with acid GSM-1, allosteric GSI BMS-708163, or substrate docking site peptide inhibitor pep11, providing evidence that these compounds have distinct binding sites. Surprisingly, we found that the cross-linking of E2012-BPyne to PS1-NTF is significantly enhanced in the presence of the active site-directed GSI L-685,458 (L458). In contrast, L458 does not affect the labeling of the acid GSM photoprobe GSM-5. We also observed that E2012-BPyne specifically labels PS1-NTF (active γ-secretase) but not full-length PS1 (inactive γ-secretase) in ANP.24 cells. Taken together, our results support the hypothesis that multiple binding sites within the γ-secretase complex exist, each of which may contribute to different modes of modulatory action. Furthermore, the enhancement of PS1-NTF labeling by E2012-BPyne in the presence of L458 suggests a degree of cooperativity between the active site of γ-secretase and the modulatory binding site of certain GSMs.

  15. Neuropeptidomics Mass Spectrometry Reveals Signaling Networks Generated by Distinct Protease Pathways in Human Systems

    Science.gov (United States)

    Hook, Vivian; Bandeira, Nuno

    2015-12-01

    Neuropeptides regulate intercellular signaling as neurotransmitters of the central and peripheral nervous systems, and as peptide hormones in the endocrine system. Diverse neuropeptides of distinct primary sequences of various lengths, often with post-translational modifications, coordinate and integrate regulation of physiological functions. Mass spectrometry-based analysis of the diverse neuropeptide structures in neuropeptidomics research is necessary to define the full complement of neuropeptide signaling molecules. Human neuropeptidomics has notable importance in defining normal and dysfunctional neuropeptide signaling in human health and disease. Neuropeptidomics has great potential for expansion in translational research opportunities for defining neuropeptide mechanisms of human diseases, providing novel neuropeptide drug targets for drug discovery, and monitoring neuropeptides as biomarkers of drug responses. In consideration of the high impact of human neuropeptidomics for health, an observed gap in this discipline is the few published articles in human neuropeptidomics compared with, for example, human proteomics and related mass spectrometry disciplines. Focus on human neuropeptidomics will advance new knowledge of the complex neuropeptide signaling networks participating in the fine control of neuroendocrine systems. This commentary review article discusses several human neuropeptidomics accomplishments that illustrate the rapidly expanding diversity of neuropeptides generated by protease processing of pro-neuropeptide precursors occurring within the secretory vesicle proteome. Of particular interest is the finding that human-specific cathepsin V participates in producing enkephalin and likely other neuropeptides, indicating unique proteolytic mechanisms for generating human neuropeptides. The field of human neuropeptidomics has great promise to solve new mechanisms in disease conditions, leading to new drug targets and therapeutic agents for human

  16. Functional Analysis of GLRX5 Mutants Reveals Distinct Functionalities of GLRX5 Protein.

    Science.gov (United States)

    Liu, Gang; Wang, Yongwei; Anderson, Gregory J; Camaschella, Clara; Chang, Yanzhong; Nie, Guangjun

    2016-01-01

    Glutaredoxin 5 (GLRX5) is a 156 amino acid mitochondrial protein that plays an essential role in mitochondrial iron-sulfur cluster transfer. Mutations in this protein were reported to result in sideroblastic anemia and variant nonketotic hyperglycinemia in human. Recently, we have characterized a Chinese congenital sideroblastic anemia patient who has two compound heterozygous missense mutations (c. 301 A>C and c. 443 T>C) in his GLRX5 gene. Herein, we developed a GLRX5 knockout K562 cell line and studied the biochemical functions of the identified pathogenic mutations and other conserved amino acids with predicted essential functions. We observed that the K101Q mutation (due to c. 301 A>C mutation) may prevent the binding of [Fe-S] to GLRX5 protein, while L148S (due to c. 443 T>C mutation) may interfere with [Fe-S] transfer from GLRX5 to iron regulatory protein 1 (IRP1), mitochondrial aconitase (m-aconitase) and ferrochelatase. We also demonstrated that L148S is functionally complementary to the K51del mutant with respect to Fe/S-ferrochelatase, Fe/S-IRP1, Fe/S-succinate dehydrogenase, and Fe/S-m-aconitase biosynthesis and lipoylation of pyruvate dehydrogenase complex and α-ketoglutarate dehydrogenase complex. Furthermore, we demonstrated that the mutations of highly conserved amino acid residues in GLRX5 protein can have different effects on downstream Fe/S proteins. Collectively, our current work demonstrates that GLRX5 protein is multifunctional in [Fe-S] protein synthesis and maturation and defects of the different amino acids of the protein will lead to distinct effects on downstream Fe/S biosynthesis.

  17. Genetic diversity within Schistosoma haematobium: DNA barcoding reveals two distinct groups.

    Directory of Open Access Journals (Sweden)

    Bonnie L Webster

    Full Text Available BACKGROUND: Schistosomiasis in one of the most prevalent parasitic diseases, affecting millions of people and animals in developing countries. Amongst the human-infective species S. haematobium is one of the most widespread causing urogenital schistosomiasis, a major human health problem across Africa, however in terms of research this human pathogen has been severely neglected. METHODOLOGY/PRINCIPAL FINDINGS: To elucidate the genetic diversity of Schistosoma haematobium, a DNA 'barcoding' study was performed on parasite material collected from 41 localities representing 18 countries across Africa and the Indian Ocean Islands. Surprisingly low sequence variation was found within the mitochondrial cytochrome oxidase subunit I (cox1 and the NADH-dehydrogenase subunit 1 snad1. The 61 haplotypes found within 1978 individual samples split into two distinct groups; one (Group 1 that is predominately made up of parasites from the African mainland and the other (Group 2 that is made up of samples exclusively from the Indian Ocean Islands and the neighbouring African coastal regions. Within Group 1 there was a dominance of one particular haplotype (H1 representing 1574 (80% of the samples analyzed. Population genetic diversity increased in samples collected from the East African coastal regions and the data suggest that there has been movement of parasites between these areas and the Indian Ocean Islands. CONCLUSIONS/SIGNIFICANCE: The high occurrence of the haplotype (H1 suggests that at some point in the recent evolutionary history of S. haematobium in Africa the population may have passed through a genetic 'bottleneck' followed by a population expansion. This study provides novel and extremely interesting insights into the population genetics of S. haematobium on a large geographic scale, which may have consequence for control and monitoring of urogenital schistosomiasis.

  18. Comparative genome analyses reveal distinct structure in the saltwater crocodile MHC.

    Science.gov (United States)

    Jaratlerdsiri, Weerachai; Deakin, Janine; Godinez, Ricardo M; Shan, Xueyan; Peterson, Daniel G; Marthey, Sylvain; Lyons, Eric; McCarthy, Fiona M; Isberg, Sally R; Higgins, Damien P; Chong, Amanda Y; John, John St; Glenn, Travis C; Ray, David A; Gongora, Jaime

    2014-01-01

    The major histocompatibility complex (MHC) is a dynamic genome region with an essential role in the adaptive immunity of vertebrates, especially antigen presentation. The MHC is generally divided into subregions (classes I, II and III) containing genes of similar function across species, but with different gene number and organisation. Crocodylia (crocodilians) are widely distributed and represent an evolutionary distinct group among higher vertebrates, but the genomic organisation of MHC within this lineage has been largely unexplored. Here, we studied the MHC region of the saltwater crocodile (Crocodylus porosus) and compared it with that of other taxa. We characterised genomic clusters encompassing MHC class I and class II genes in the saltwater crocodile based on sequencing of bacterial artificial chromosomes. Six gene clusters spanning ∼452 kb were identified to contain nine MHC class I genes, six MHC class II genes, three TAP genes, and a TRIM gene. These MHC class I and class II genes were in separate scaffold regions and were greater in length (2-6 times longer) than their counterparts in well-studied fowl B loci, suggesting that the compaction of avian MHC occurred after the crocodilian-avian split. Comparative analyses between the saltwater crocodile MHC and that from the alligator and gharial showed large syntenic areas (>80% identity) with similar gene order. Comparisons with other vertebrates showed that the saltwater crocodile had MHC class I genes located along with TAP, consistent with birds studied. Linkage between MHC class I and TRIM39 observed in the saltwater crocodile resembled MHC in eutherians compared, but absent in avian MHC, suggesting that the saltwater crocodile MHC appears to have gene organisation intermediate between these two lineages. These observations suggest that the structure of the saltwater crocodile MHC, and other crocodilians, can help determine the MHC that was present in the ancestors of archosaurs.

  19. Comparative genome analyses reveal distinct structure in the saltwater crocodile MHC.

    Directory of Open Access Journals (Sweden)

    Weerachai Jaratlerdsiri

    Full Text Available The major histocompatibility complex (MHC is a dynamic genome region with an essential role in the adaptive immunity of vertebrates, especially antigen presentation. The MHC is generally divided into subregions (classes I, II and III containing genes of similar function across species, but with different gene number and organisation. Crocodylia (crocodilians are widely distributed and represent an evolutionary distinct group among higher vertebrates, but the genomic organisation of MHC within this lineage has been largely unexplored. Here, we studied the MHC region of the saltwater crocodile (Crocodylus porosus and compared it with that of other taxa. We characterised genomic clusters encompassing MHC class I and class II genes in the saltwater crocodile based on sequencing of bacterial artificial chromosomes. Six gene clusters spanning ∼452 kb were identified to contain nine MHC class I genes, six MHC class II genes, three TAP genes, and a TRIM gene. These MHC class I and class II genes were in separate scaffold regions and were greater in length (2-6 times longer than their counterparts in well-studied fowl B loci, suggesting that the compaction of avian MHC occurred after the crocodilian-avian split. Comparative analyses between the saltwater crocodile MHC and that from the alligator and gharial showed large syntenic areas (>80% identity with similar gene order. Comparisons with other vertebrates showed that the saltwater crocodile had MHC class I genes located along with TAP, consistent with birds studied. Linkage between MHC class I and TRIM39 observed in the saltwater crocodile resembled MHC in eutherians compared, but absent in avian MHC, suggesting that the saltwater crocodile MHC appears to have gene organisation intermediate between these two lineages. These observations suggest that the structure of the saltwater crocodile MHC, and other crocodilians, can help determine the MHC that was present in the ancestors of archosaurs.

  20. Optical Coherence Tomography Reveals Distinct Patterns of Retinal Damage in Neuromyelitis Optica and Multiple Sclerosis.

    Directory of Open Access Journals (Sweden)

    Elisa Schneider

    Full Text Available Neuromyelitis optica (NMO and relapsing-remitting multiple sclerosis (RRMS are difficult to differentiate solely on clinical grounds. Optical coherence tomography (OCT studies investigating retinal changes in both diseases focused primarily on the retinal nerve fiber layer (RNFL while rare data are available on deeper intra-retinal layers.To detect different patterns of intra-retinal layer alterations in patients with NMO spectrum disorders (NMOSD and RRMS with focus on the influence of a previous optic neuritis (ON.We applied spectral-domain OCT in eyes of NMOSD patients and compared them to matched RRMS patients and healthy controls (HC. Semi-automatic intra-retinal layer segmentation was used to quantify intra-retinal layer thicknesses. In a subgroup low contrast visual acuity (LCVA was assessed.NMOSD-, MS- and HC-groups, each comprising 17 subjects, were included in analysis. RNFL thickness was more severely reduced in NMOSD compared to MS following ON. In MS-ON eyes, RNFL thinning showed a clear temporal preponderance, whereas in NMOSD-ON eyes RNFL was more evenly reduced, resulting in a significantly lower ratio of the nasal versus temporal RNFL thickness. In comparison to HC, ganglion cell layer thickness was stronger reduced in NMOSD-ON than in MS-ON, accompanied by a more severe impairment of LCVA. The inner nuclear layer and the outer retinal layers were thicker in NMOSD-ON patients compared to NMOSD without ON and HC eyes while these differences were primarily driven by microcystic macular edema.Our study supports previous findings that ON in NMOSD leads to more pronounced retinal thinning and visual function impairment than in RRMS. The different retinal damage patterns in NMOSD versus RRMS support the current notion of distinct pathomechanisms of both conditions. However, OCT is still insufficient to help with the clinically relevant differentiation of both conditions in an individual patient.

  1. Metabolomic Profiles of Body Mass Index in the Framingham Heart Study Reveal Distinct Cardiometabolic Phenotypes.

    Directory of Open Access Journals (Sweden)

    Jennifer E Ho

    Full Text Available Although obesity and cardiometabolic traits commonly overlap, underlying pathways remain incompletely defined. The association of metabolite profiles across multiple cardiometabolic traits may lend insights into the interaction of obesity and metabolic health. We sought to investigate metabolic signatures of obesity and related cardiometabolic traits in the community using broad-based metabolomic profiling.We evaluated the association of 217 assayed metabolites and cross-sectional as well as longitudinal changes in cardiometabolic traits among 2,383 Framingham Offspring cohort participants. Body mass index (BMI was associated with 69 of 217 metabolites (P<0.00023 for all, including aromatic (tyrosine, phenylalanine and branched chain amino acids (valine, isoleucine, leucine. Additional metabolic pathways associated with BMI included the citric acid cycle (isocitrate, alpha-ketoglutarate, aconitate, the tryptophan pathway (kynurenine, kynurenic acid, and the urea cycle. There was considerable overlap in metabolite profiles between BMI, abdominal adiposity, insulin resistance [IR] and dyslipidemia, modest overlap of metabolite profiles between BMI and hyperglycemia, and little overlap with fasting glucose or elevated blood pressure. Metabolite profiles were associated with longitudinal changes in fasting glucose, but the involved metabolites (ornithine, 5-HIAA, aminoadipic acid, isoleucine, cotinine were distinct from those associated with baseline glucose or other traits. Obesity status appeared to "modify" the association of 9 metabolites with IR. For example, bile acid metabolites were strongly associated with IR among obese but not lean individuals, whereas isoleucine had a stronger association with IR in lean individuals.In this large-scale metabolite profiling study, body mass index was associated with a broad range of metabolic alterations. Metabolite profiling highlighted considerable overlap with abdominal adiposity, insulin resistance

  2. Combating biofilms

    DEFF Research Database (Denmark)

    Yang, Liang; Liu, Yang; Wu, Hong;

    2012-01-01

    Biofilms are complex microbial communities consisting of microcolonies embedded in a matrix of self-produced polymer substances. Biofilm cells show much greater resistance to environmental challenges including antimicrobial agents than their free-living counterparts. The biofilm mode of life...... is believed to significantly contribute to successful microbial survival in hostile environments. Conventional treatment, disinfection and cleaning strategies do not proficiently deal with biofilm-related problems, such as persistent infections and contamination of food production facilities. In this review......, strategies to control biofilms are discussed, including those of inhibition of microbial attachment, interference of biofilm structure development and differentiation, killing of biofilm cells and induction of biofilm dispersion....

  3. Feasibility study of spectral pattern recognition reveals distinct classes of volcanic tremor

    Science.gov (United States)

    Unglert, K.; Jellinek, A. M.

    2017-04-01

    Systematic investigations of the similarities and differences among volcanic tremor at a range of volcano types may hold crucial information about the plausibility of inferred source mechanisms, which, in turn, may be important for eruption forecasting. However, such studies are rare, in part because of an intrinsic difficulty with identifying tremor signals within very long time series of volcano seismic data. Accordingly, we develop an efficient tremor detection algorithm and identify over 12,000h of volcanic tremor on 24 stations at Kīlauea, Okmok, Pavlof, and Redoubt volcanoes. We estimate spectral content over 5-minute tremor windows, and apply a novel combination of Principal Component Analysis (PCA) and hierarchical clustering to identify patterns in the tremor spectra. Analyzing several stations from a given volcano together reveals different styles of tremor within individual volcanic settings. In addition to identifying tremor properties common to all stations in a given network, we find localized tremor signals including those related to processes such as lahars or dike intrusions that are only observed on some of the stations within a network. Subsequent application of our analysis to a combination of stations from the different volcanoes reveals that at least three main tremor classes can be detected across all settings. Whereas a regime with a ridge of high power distributed over 1-2Hz and a gradual decay of spectral power towards higher frequencies is observed dominantly at three volcanoes (Kīlauea, Okmok, Redoubt) with magma reservoirs centered at less than 5km below sea level (b.s.l.), a spectrum with a steeper slope and a narrower peak at 1-2Hz is observed only in association with open vents (Kīlauea and Pavlof). A third regime with a peak at approximately 3Hz is confined to two stratovolcanoes (Pavlof and Redoubt). These observations suggest generic relationships between the spectral character of the observed signals and volcano

  4. SINE polymorphism reveals distinct strains of Entamoeba histolytica from North India.

    Science.gov (United States)

    Sharma, Shraddha; Banyal, Neha; Singh, Mukul; Mandal, A K; Bhattacharya, Sudha; Paul, Jaishree

    2017-04-01

    Entamoeba histolytica is an intestinal parasite causing significant morbidity and mortality in the developing world. More tools are needed to understand the epidemiology and molecular pathogenesis of amebiasis. Virulence pattern of E. histolytica could be linked with the genotype of a strain. Several loci showing insertion polymorphism of retrotransposable short interspersed nuclear elements EhSINE1 and EhSINE2 have been reported among laboratory strains of E. histolytica. The present study was undertaken to validate this observation in clinical isolates from north India. Our results indicate that the Indian samples show a different propensity of SINE retention or loss at two of the polymorphic loci compared with non-Indian samples. Statistical analysis of different loci revealed Locus 17 of EhSINE1as a potential geographical marker for distinguishing Indian isolates from non Indian isolates. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Intact-Brain Analyses Reveal Distinct Information Carried by SNc Dopamine Subcircuits.

    Science.gov (United States)

    Lerner, Talia N; Shilyansky, Carrie; Davidson, Thomas J; Evans, Kathryn E; Beier, Kevin T; Zalocusky, Kelly A; Crow, Ailey K; Malenka, Robert C; Luo, Liqun; Tomer, Raju; Deisseroth, Karl

    2015-07-30

    Recent progress in understanding the diversity of midbrain dopamine neurons has highlighted the importance--and the challenges--of defining mammalian neuronal cell types. Although neurons may be best categorized using inclusive criteria spanning biophysical properties, wiring of inputs, wiring of outputs, and activity during behavior, linking all of these measurements to cell types within the intact brains of living mammals has been difficult. Here, using an array of intact-brain circuit interrogation tools, including CLARITY, COLM, optogenetics, viral tracing, and fiber photometry, we explore the diversity of dopamine neurons within the substantia nigra pars compacta (SNc). We identify two parallel nigrostriatal dopamine neuron subpopulations differing in biophysical properties, input wiring, output wiring to dorsomedial striatum (DMS) versus dorsolateral striatum (DLS), and natural activity patterns during free behavior. Our results reveal independently operating nigrostriatal information streams, with implications for understanding the logic of dopaminergic feedback circuits and the diversity of mammalian neuronal cell types.

  6. Proteomic Stable Isotope Probing Reveals Taxonomically Distinct Patterns in Amino Acid Assimilation by Coastal Marine Bacterioplankton.

    Science.gov (United States)

    Bryson, Samuel; Li, Zhou; Pett-Ridge, Jennifer; Hettich, Robert L; Mayali, Xavier; Pan, Chongle; Mueller, Ryan S

    2016-01-01

    . IMPORTANCE An estimated 50 gigatons of carbon is annually fixed within marine systems, of which heterotrophic microbial populations process nearly half. These communities vary in composition and activity across spatial and temporal scales, so understanding how these changes affect global processes requires the delineation of functional roles for individual members. In a step toward ascertaining these roles, we applied proteomic stable isotope probing to quantify the assimilation of organic carbon from DFAAs into microbial protein biomass, since the turnover of DFAAs accounts for a substantial fraction of marine microbial carbon metabolism that is directed into biomass production. We conducted experiments at two coastal North Pacific locations and found taxonomically distinct responses. This approach allowed us to compare amino acid assimilation by specific bacterioplankton populations and characterize their allocation of this substrate among cellular functions.

  7. Spatial Patterns of Carbonate Biomineralization in Biofilms

    Science.gov (United States)

    Li, Xiaobao; Chopp, David L.; Russin, William A.; Brannon, Paul T.; Parsek, Matthew R.

    2015-01-01

    Microbially catalyzed precipitation of carbonate minerals is an important process in diverse biological, geological, and engineered systems. However, the processes that regulate carbonate biomineralization and their impacts on biofilms are largely unexplored, mainly because of the inability of current methods to directly observe biomineralization within biofilms. Here, we present a method for in situ, real-time imaging of biomineralization in biofilms and use it to show that Pseudomonas aeruginosa biofilms produce morphologically distinct carbonate deposits that substantially modify biofilm structures. The patterns of carbonate biomineralization produced in situ were substantially different from those caused by accumulation of particles produced by abiotic precipitation. Contrary to the common expectation that mineral precipitation should occur at the biofilm surface, we found that biomineralization started at the base of the biofilm. The carbonate deposits grew over time, detaching biofilm-resident cells and deforming the biofilm morphology. These findings indicate that biomineralization is a general regulator of biofilm architecture and properties. PMID:26276112

  8. Biogeography of Oenococcus oeni Reveals Distinctive but Nonspecific Populations in Wine-Producing Regions.

    Science.gov (United States)

    El Khoury, Mariette; Campbell-Sills, Hugo; Salin, Franck; Guichoux, Erwan; Claisse, Olivier; Lucas, Patrick M

    2017-02-01

    Understanding the mechanisms behind the typicity of regional wines inevitably brings attention to microorganisms associated with their production. Oenococcus oeni is the main bacterial species involved in wine and cider making. It develops after the yeast-driven alcoholic fermentation and performs the malolactic fermentation, which improves the taste and aromatic complexity of most wines. Here, we have evaluated the diversity and specificity of O. oeni strains in six regions. A total of 235 wines and ciders were collected during spontaneous malolactic fermentations and used to isolate 3,212 bacterial colonies. They were typed by multilocus variable analysis, which disclosed a total of 514 O. oeni strains. Their phylogenetic relationships were evaluated by a second typing method based on single nucleotide polymorphism (SNP) analysis. Taken together, the results indicate that each region holds a high diversity of strains that constitute a unique population. However, strains present in each region belong to diverse phylogenetic groups, and the same groups can be detected in different regions, indicating that strains are not genetically adapted to regions. In contrast, greater strain identity was seen for cider, white wine, or red wine of Burgundy, suggesting that genetic adaptation to these products occurred. This study reports the isolation, genotyping, and geographic distribution analysis of the largest collection of O. oeni strains performed to date. It reveals that there is very high diversity of strains in each region, the majority of them being detected in a single region. The study also reports the development of an SNP genotyping method that is useful for analyzing the distribution of O. oeni phylogroups. The results show that strains are not genetically adapted to regions but to specific types of wines. They reveal new phylogroups of strains, particularly two phylogroups associated with white wines and red wines of Burgundy. Taken together, the results shed

  9. NMR based serum metabolomics reveals a distinctive signature in patients with Lupus Nephritis

    Science.gov (United States)

    Guleria, Anupam; Pratap, Avadhesh; Dubey, Durgesh; Rawat, Atul; Chaurasia, Smriti; Sukesh, Edavalath; Phatak, Sanat; Ajmani, Sajal; Kumar, Umesh; Khetrapal, Chunni Lal; Bacon, Paul; Misra, Ramnath; Kumar, Dinesh

    2016-01-01

    Management of patient with Lupus Nephritis (LN) continues to remain a challenge for the treating physicians because of considerable morbidity and even mortality. The search of biomarkers in serum and urine is a focus of researchers to unravel new targets for therapy. In the present study, the utility of NMR-based serum metabolomics has been evaluated for the first time in discriminating LN patients from non-nephritis lupus patients (SLE) and further to get new insights into the underlying disease processes for better clinical management. Metabolic profiling of sera obtained from 22 SLE patients, 40 LN patients and 30 healthy controls (HC) were performed using high resolution 1D 1H-CPMG and diffusion edited NMR spectra to identify the potential molecular biomarkers. Using multivariate analysis, we could distinguish SLE and LN patients from HC and LN from SLE patients. Compared to SLE patients, the LN patients had increased serum levels of lipid metabolites (including LDL/VLDL lipoproteins), creatinine and decreased levels of acetate. Our results revealed that metabolic markers especially lipids and acetate derived from NMR spectroscopy has high sensitivity and specificity to distinguish LN among SLE patients and has the potential to be a useful adjunctive tool in diagnosis and clinical management of LN. PMID:27739464

  10. Phosphoproteomic dynamics of chickpea (Cicer arietinum L.) reveals shared and distinct components of dehydration response.

    Science.gov (United States)

    Subba, Pratigya; Barua, Pragya; Kumar, Rajiv; Datta, Asis; Soni, Kamlesh Kumar; Chakraborty, Subhra; Chakraborty, Niranjan

    2013-11-01

    Reversible protein phosphorylation is a ubiquitous regulatory mechanism that plays critical roles in transducing stress signals to bring about coordinated intracellular responses. To gain better understanding of dehydration response in plants, we have developed a differential phosphoproteome in a food legume, chickpea (Cicer arietinum L.). Three-week-old chickpea seedlings were subjected to progressive dehydration by withdrawing water, and the changes in the phosphorylation status of a large repertoire of proteins were monitored. The proteins were resolved by 2-DE and stained with phosphospecific fluorescent Pro-Q Diamond dye. Mass spectrometric analysis led to the identification of 91 putative phosphoproteins, presumably involved in a variety of functions including cell defense and rescue, photosynthesis and photorespiration, molecular chaperones, and ion transport, among others. Multiple sites of phosphorylation were predicted on several key elements, which include both the regulatory as well as the functional proteins. A critical survey of the phosphorylome revealed a DREPP (developmentally regulated plasma membrane protein) plasma membrane polypeptide family protein, henceforth designated CaDREPP1. The transcripts of CaDREPP1 were found to be differentially regulated under dehydration stress, further corroborating the proteomic results. This work provides new insights into the possible phosphorylation events triggered by the conditions of progressive water-deficit in plants.

  11. Different human gut models reveal the distinct fermentation patterns of Arabinoxylan versus inulin.

    Science.gov (United States)

    Van den Abbeele, Pieter; Venema, Koen; Van de Wiele, Tom; Verstraete, Willy; Possemiers, Sam

    2013-10-16

    Different in vitro models have been developed to assess how food compounds affect the human gut microbiota. Using two such models (SHIME(R) and TIM-2), we compared how long-chain arabinoxylan (LC-AX), a wheat-derived potentially prebiotic fiber, and inulin (IN), a well-established prebiotic compound, modulate SCFA production and bifidobacteria composition. While both the SHIME and TIM-2 differ in experimental design, they both demonstrated that LC-AX and IN specifically increased the health-promoting metabolites propionate and butyrate, respectively. Furthermore, LC-AX stimulated Bifidobacterium longum, while IN stimulated other bifidobacteria including Bifidobacterium adolescentis. The SHIME experiment also revealed that effects of LC-AX were more persistent during the 2-week wash-out period. These results confirm a recent in vivo study, during which humanized rats were treated with the same LC-AX/IN. In conclusion, results from different human gut models suggest that, besides IN, LC-AX are promising prebiotic candidates with high specificity toward Bifidobacterium longum and a selective propionate increase.

  12. Comprehensive RNA Polymerase II Interactomes Reveal Distinct and Varied Roles for Each Phospho-CTD Residue

    Directory of Open Access Journals (Sweden)

    Kevin M. Harlen

    2016-06-01

    Full Text Available Transcription controls splicing and other gene regulatory processes, yet mechanisms remain obscure due to our fragmented knowledge of the molecular connections between the dynamically phosphorylated RNA polymerase II (Pol II C-terminal domain (CTD and regulatory factors. By systematically isolating phosphorylation states of the CTD heptapeptide repeat (Y1S2P3T4S5P6S7, we identify hundreds of protein factors that are differentially enriched, revealing unappreciated connections between the Pol II CTD and co-transcriptional processes. These data uncover a role for threonine-4 in 3′ end processing through control of the transition between cleavage and termination. Furthermore, serine-5 phosphorylation seeds spliceosomal assembly immediately downstream of 3′ splice sites through a direct interaction with spliceosomal subcomplex U1. Strikingly, threonine-4 phosphorylation also impacts splicing by serving as a mark of co-transcriptional spliceosome release and ensuring efficient post-transcriptional splicing genome-wide. Thus, comprehensive Pol II interactomes identify the complex and functional connections between transcription machinery and other gene regulatory complexes.

  13. Transmission electron microscopy reveals distinct macrophage- and tick cell-specific morphological stages of Ehrlichia chaffeensis.

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    Sarah E Dedonder

    Full Text Available BACKGROUND: Ehrlichia chaffeensis is an emerging tick-borne rickettsial pathogen responsible for human monocytic ehrlichiosis. Despite the induction of an active host immune response, the pathogen has evolved to persist in its vertebrate and tick hosts. Understanding how the organism progresses in tick and vertebrate host cells is critical in identifying effective strategies to block the pathogen transmission. Our recent molecular and proteomic studies revealed differences in numerous expressed proteins of the organism during its growth in different host environments. METHODOLOGY/PRINCIPAL FINDINGS: Transmission electron microscopy analysis was performed to assess morphological changes in the bacterium within macrophages and tick cells. The stages of pathogen progression observed included the attachment of the organism to the host cells, its engulfment and replication within a morulae by binary fission and release of the organisms from infected host cells by complete host cell lysis or by exocytosis. E. chaffeensis grown in tick cells was highly pleomorphic and appears to replicate by both binary fission and filamentous type cell divisions. The presence of Ehrlichia-like inclusions was also observed within the nucleus of both macrophages and tick cells. This observation was confirmed by confocal microscopy and immunoblot analysis. CONCLUSIONS/SIGNIFICANCE: Morphological differences in the pathogen's progression, replication, and processing within macrophages and tick cells provide further evidence that E. chaffeensis employs unique host-cell specific strategies in support of adaptation to vertebrate and tick cell environments.

  14. Biofilms and the food industry

    Directory of Open Access Journals (Sweden)

    Nathanon Trachoo

    2003-11-01

    Full Text Available In the past, interest in biofilms was limited to research related to water distribution systems, waste water treatment and dental plaques. Biofilm has become a more popular research topic in many other areas in recent years including food safety. Biofilm formation can compromise the sanitation of food surfaces and environmental surfaces by spreading detached organisms to other areas of processing plants. Unfortunately, these detached organisms are not similar to normal microorganisms suspended in an aquatic environment but are more resistant to several stresses or microbial inactivation including some food preservation methods. Microstructures of biofilms as revealed by different types of microscopic techniques showed that biofilms are highly complex and consist of many symbiotic organisms, some of which are human pathogens. This article reviewed the process of biofilm formation, the significance of biofilms on food or food contact surfaces, their ability to protect foodborne pathogens from environmental stresses and recent methods for the study of biofilms on food contact surfaces.

  15. Glutathione-Disrupted Biofilms of Clinical Pseudomonas aeruginosa Strains Exhibit an Enhanced Antibiotic Effect and a Novel Biofilm Transcriptome.

    Science.gov (United States)

    Klare, William; Das, Theerthankar; Ibugo, Amaye; Buckle, Edwina; Manefield, Mike; Manos, Jim

    2016-08-01

    Pseudomonas aeruginosa infections result in high morbidity and mortality rates for individuals with cystic fibrosis (CF), with premature death often occurring. These infections are complicated by the formation of biofilms in the sputum. Antibiotic therapy is stymied by antibiotic resistance of the biofilm matrix, making novel antibiofilm strategies highly desirable. Within P. aeruginosa biofilms, the redox factor pyocyanin enhances biofilm integrity by intercalating with extracellular DNA. The antioxidant glutathione (GSH) reacts with pyocyanin, disrupting intercalation. This study investigated GSH disruption by assaying the physiological effects of GSH and DNase I on biofilms of clinical CF isolates grown in CF artificial sputum medium (ASMDM+). Confocal scanning laser microscopy showed that 2 mM GSH, alone or combined with DNase I, significantly disrupted immature (24-h) biofilms of Australian epidemic strain (AES) isogens AES-1R and AES-1M. GSH alone greatly disrupted mature (72-h) AES-1R biofilms, resulting in significant differential expression of 587 genes, as indicated by RNA-sequencing (RNA-seq) analysis. Upregulated systems included cyclic diguanylate and pyoverdine biosynthesis, the type VI secretion system, nitrate metabolism, and translational machinery. Biofilm disruption with GSH revealed a cellular physiology distinct from those of mature and dispersed biofilms. RNA-seq results were validated by biochemical and quantitative PCR assays. Biofilms of a range of CF isolates disrupted with GSH and DNase I were significantly more susceptible to ciprofloxacin, and increased antibiotic effectiveness was achieved by increasing the GSH concentration. This study demonstrated that GSH, alone or with DNase I, represents an effective antibiofilm treatment when combined with appropriate antibiotics, pending in vivo studies.

  16. Improved flow cytometric assessment reveals distinct microvesicle (cell-derived microparticle signatures in joint diseases.

    Directory of Open Access Journals (Sweden)

    Bence György

    Full Text Available INTRODUCTION: Microvesicles (MVs, earlier referred to as microparticles, represent a major type of extracellular vesicles currently considered as novel biomarkers in various clinical settings such as autoimmune disorders. However, the analysis of MVs in body fluids has not been fully standardized yet, and there are numerous pitfalls that hinder the correct assessment of these structures. METHODS: In this study, we analyzed synovial fluid (SF samples of patients with osteoarthritis (OA, rheumatoid arthritis (RA and juvenile idiopathic arthritis (JIA. To assess factors that may confound MV detection in joint diseases, we used electron microscopy (EM, Nanoparticle Tracking Analysis (NTA and mass spectrometry (MS. For flow cytometry, a method commonly used for phenotyping and enumeration of MVs, we combined recent advances in the field, and used a novel approach of differential detergent lysis for the exclusion of MV-mimicking non-vesicular signals. RESULTS: EM and NTA showed that substantial amounts of particles other than MVs were present in SF samples. Beyond known MV-associated proteins, MS analysis also revealed abundant plasma- and immune complex-related proteins in MV preparations. Applying improved flow cytometric analysis, we demonstrate for the first time that CD3(+ and CD8(+ T-cell derived SF MVs are highly elevated in patients with RA compared to OA patients (p=0.027 and p=0.009, respectively, after Bonferroni corrections. In JIA, we identified reduced numbers of B cell-derived MVs (p=0.009, after Bonferroni correction. CONCLUSIONS: Our results suggest that improved flow cytometric assessment of MVs facilitates the detection of previously unrecognized disease-associated vesicular signatures.

  17. Analysis of gene expression during parabolic flights reveals distinct early gravity responses in Arabidopsis roots.

    Science.gov (United States)

    Aubry-Hivet, D; Nziengui, H; Rapp, K; Oliveira, O; Paponov, I A; Li, Y; Hauslage, J; Vagt, N; Braun, M; Ditengou, F A; Dovzhenko, A; Palme, K

    2014-01-01

    Plant roots are among most intensively studied biological systems in gravity research. Altered gravity induces asymmetric cell growth leading to root bending. Differential distribution of the phytohormone auxin underlies root responses to gravity, being coordinated by auxin efflux transporters from the PIN family. The objective of this study was to compare early transcriptomic changes in roots of Arabidopsis thaliana wild type, and pin2 and pin3 mutants under parabolic flight conditions and to correlate these changes to auxin distribution. Parabolic flights allow comparison of transient 1-g, hypergravity and microgravity effects in living organisms in parallel. We found common and mutation-related genes differentially expressed in response to transient microgravity phases. Gene ontology analysis of common genes revealed lipid metabolism, response to stress factors and light categories as primarily involved in response to transient microgravity phases, suggesting that fundamental reorganisation of metabolic pathways functions upstream of a further signal mediating hormonal network. Gene expression changes in roots lacking the columella-located PIN3 were stronger than in those deprived of the epidermis and cortex cell-specific PIN2. Moreover, repetitive exposure to microgravity/hypergravity and gravity/hypergravity flight phases induced an up-regulation of auxin responsive genes in wild type and pin2 roots, but not in pin3 roots, suggesting a critical function of PIN3 in mediating auxin fluxes in response to transient microgravity phases. Our study provides important insights towards understanding signal transduction processes in transient microgravity conditions by combining for the first time the parabolic flight platform with the transcriptome analysis of different genetic mutants in the model plant, Arabidopsis. © 2013 German Botanical Society and The Royal Botanical Society of the Netherlands.

  18. Salmonella enterica serotypes isolated from squabs reveal multidrug resistance and a distinct pathogenicity gene repertoire.

    Science.gov (United States)

    Osman, K M; Marouf, S H; Mehana, O A; AlAtfeehy, N

    2014-12-01

    The consumption of squab (young unfledged pigeons) as part of the cuisine of many countries, together with the observation that squabs are vectors of zoonotic agents, may make them a public health risk. This study was designed to determine the serotypes, distribution of 11 virulence genes (invA, avrA, ssaQ, mgtC, siiD, sopB, gipA, sodC1, sopE1, spvC, bcfC) and the antimicrobial resistance profiles of salmonellae recovered from squabs. Six isolates were identified from among 45 (13.3%) squabs sampled. Three serotypes were identified according to the Kauffmann-White serotyping scheme: Salmonella Typhimurium (4/6; 66.7%), S. Braenderup (1/6; 16.7%) and S. Lomita (1/6; 16.7%). Polymerase chain reaction analyses revealed the presence of invA, sopB and bcfC in all six isolates, whereas sopE1 and gipA were absent. All six isolates were resistant to lincomycin and streptomycin, but all were susceptible to ciprofloxacin, colistin sulphate and gentamicin. Among the S. Typhimurium isolates, seven resistance profiles were identified: penicillins,aminoglycosides,fluoroquinolones, lincosamides,phenicols, tetracyclines and sulphonamides; four resistance profiles were identified in the isolates of S. Braenderup and S. Lomita: aminoglycosides, fluoroquinolones, lincosamides and polymyxin. Thus, the distribution of resistance to the antibiotics was largely dependent on serotype identity. The presence of invA, avrA, ssaQ, mgtC, siiD, sopB and bcfC was associated with resistance to chloramphenicol; invA, sopB and bcfC with resistance to streptomycin and lincosamide; and invA and sodC1 with resistance to trimethoprim-sulfamethoxazole. The identification of serotypes S. Typhimurium, S. Braenderup and S. Lomita in the squab samples has important implications because these serotypes are significant causes of food poisoning and enteric fever in humans.

  19. Mistletoe lectin I in complex with galactose and lactose reveals distinct sugar-binding properties

    Energy Technology Data Exchange (ETDEWEB)

    Mikeska, Ruth [Institute of Biochemistry and Food Chemistry, University of Hamburg, c/o DESY, Notkestrasse 85, Building 22a, 22603 Hamburg (Germany); Wacker, Roland [Institute of Physiological Chemistry, University of Tübingen, Hoppe-Seyler-Strasse 4, 72076 Tübingen (Germany); Arni, Raghuvir [Department of Physics, IBILCE/UNESP, São Jose do Rio Preto, São Paul (Brazil); Singh, Tej P. [Department of Biophysics, All India Institute of Medical Sciences, New Delhi (India); Mikhailov, Albert; Gabdoulkhakov, Azat [Institute of Crystallography of Russian Academy of Sciences, Leninsky Prospect 59, 117333 Moscow (Russian Federation); Voelter, Wolfgang [Institute of Physiological Chemistry, University of Tübingen, Hoppe-Seyler-Strasse 4, 72076 Tübingen (Germany); Betzel, Christian, E-mail: betzel@unisgi1.desy.de [Institute of Biochemistry and Food Chemistry, University of Hamburg, c/o DESY, Notkestrasse 85, Building 22a, 22603 Hamburg (Germany)

    2005-01-01

    The structures of mistletoe lectin I in complex with lactose and galactose reveal differences in binding by the two known sites in subdomains α1 and γ2 and suggest the presence of a third low-affinity site in subdomain β1. The structures of mistletoe lectin I (ML-I) from Viscum album complexed with lactose and galactose have been determined at 2.3 Å resolution and refined to R factors of 20.9% (R{sub free} = 23.6%) and 20.9 (R{sub free} = 24.6%), respectively. ML-I is a heterodimer and belongs to the class of ribosome-inactivating proteins of type II, which consist of two chains. The A-chain has rRNA N-glycosidase activity and irreversibly inhibits eukaryotic ribosomes. The B-chain is a lectin and preferentially binds to galactose-terminated glycolipids and glycoproteins on cell membranes. Saccharide binding is performed by two binding sites in subdomains α1 and γ2 of the ML-I B-chain separated by ∼62 Å from each other. The favoured binding of galactose in subdomain α1 is achieved via hydrogen bonds connecting the 4-hydroxyl and 3-hydroxyl groups of the sugar moiety with the side chains of Asp23B, Gln36B and Lys41B and the main chain of 26B. The aromatic ring of Trp38B on top of the preferred binding pocket supports van der Waals packing of the apolar face of galactose and stabilizes the sugar–lectin complex. In the galactose-binding site II of subdomain γ2, Tyr249B provides the hydrophobic stacking and the side chains of Asp235B, Gln238B and Asn256B are hydrogen-bonding partners for galactose. In the case of the galactose-binding site I, the 2-hydroxyl group also stabilizes the sugar–protein complex, an interaction thus far rarely detected in galactose-specific lectins. Finally, a potential third low-affinity galactose-binding site in subunit β1 was identified in the present ML-I structures, in which a glycerol molecule from the cryoprotectant buffer has bound, mimicking the sugar compound.

  20. Analysis of Candida albicans mutants defective in the Cdk8 module of mediator reveal links between metabolism and biofilm formation.

    Directory of Open Access Journals (Sweden)

    Allia K Lindsay

    2014-10-01

    Full Text Available Candida albicans biofilm formation is a key virulence trait that involves hyphal growth and adhesin expression. Pyocyanin (PYO, a phenazine secreted by Pseudomonas aeruginosa, inhibits both C. albicans biofilm formation and development of wrinkled colonies. Using a genetic screen, we identified two mutants, ssn3Δ/Δ and ssn8Δ/Δ, which continued to wrinkle in the presence of PYO. Ssn8 is a cyclin-like protein and Ssn3 is similar to cyclin-dependent kinases; both proteins are part of the heterotetrameric Cdk8 module that forms a complex with the transcriptional co-regulator, Mediator. Ssn3 kinase activity was also required for PYO sensitivity as a kinase dead mutant maintained a wrinkled colony morphology in the presence of PYO. Furthermore, similar phenotypes were observed in mutants lacking the other two components of the Cdk8 module-Srb8 and Srb9. Through metabolomics analyses and biochemical assays, we showed that a compromised Cdk8 module led to increases in glucose consumption, glycolysis-related transcripts, oxidative metabolism and ATP levels even in the presence of PYO. In the mutant, inhibition of respiration to levels comparable to the PYO-treated wild type inhibited wrinkled colony development. Several lines of evidence suggest that PYO does not act through Cdk8. Lastly, the ssn3 mutant was a hyperbiofilm former, and maintained higher biofilm formation in the presence of PYO than the wild type. Together these data provide novel insights into the role of the Cdk8 module of Mediator in regulation of C. albicans physiology and the links between respiratory activity and both wrinkled colony and biofilm development.

  1. Changes in gene expression of Actinobacillus pleuropneumoniae in response to anaerobic stress reveal induction of central metabolism and biofilm formation.

    Science.gov (United States)

    Li, Lu; Zhu, Jiawen; Yang, Kui; Xu, Zhuofei; Liu, Ziduo; Zhou, Rui

    2014-06-01

    Actinobacillus pleuropneumoniae is an important porcine respiratory pathogen causing great economic losses in the pig industry worldwide. Oxygen deprivation is a stress that A. pleuropneumoniae will encounter during both early infection and the later, persistent stage. To understand modulation of A. pleuropneumoniae gene expression in response to the stress caused by anaerobic conditions, gene expression profiles under anaerobic and aerobic conditions were compared in this study. The microarray results showed that 631 genes (27.7% of the total ORFs) were differentially expressed in anaerobic conditions. Many genes encoding proteins involved in glycolysis, carbon source uptake systems, pyruvate metabolism, fermentation and the electron respiration transport chain were up-regulated. These changes led to an increased amount of pyruvate, lactate, ethanol and acetate in the bacterial cells as confirmed by metabolite detection. Genes encoding proteins involved in cell surface structures, especially biofilm formation, peptidoglycan biosynthesis and lipopolysaccharide biosynthesis were up-regulated as well. Biofilm formation was significantly enhanced under anaerobic conditions. These results indicate that induction of central metabolism is important for basic survival of A. pleuropneumoniae after a shift to an anaerobic environment. Enhanced biofilm formation may contribute to the persistence of this pathogen in the damaged anaerobic host tissue and also in the early colonization stage. These discoveries give new insights into adaptation mechanisms of A. pleuropneumoniae in response to environmental stress.

  2. An autoinhibited conformation of LGN reveals a distinct interaction mode between GoLoco motifs and TPR motifs.

    Science.gov (United States)

    Pan, Zhu; Zhu, Jinwei; Shang, Yuan; Wei, Zhiyi; Jia, Min; Xia, Caihao; Wen, Wenyu; Wang, Wenning; Zhang, Mingjie

    2013-06-01

    LGN plays essential roles in asymmetric cell divisions via its N-terminal TPR-motif-mediated binding to mInsc and NuMA. This scaffolding activity requires the release of the autoinhibited conformation of LGN by binding of Gα(i) to its C-terminal GoLoco (GL) motifs. The interaction between the GL and TPR motifs of LGN represents a distinct GL/target binding mode with an unknown mechanism. Here, we show that two consecutive GL motifs of LGN form a minimal TPR-motif-binding unit. GL12 and GL34 bind to TPR0-3 and TPR4-7, respectively. The crystal structure of a truncated LGN reveals that GL34 forms a pair of parallel α helices and binds to the concave surface of TPR4-7, thereby preventing LGN from binding to other targets. Importantly, the GLs bind to TPR motifs with a mode distinct from that observed in the GL/Gα(i)·GDP complexes. Our results also indicate that multiple and orphan GL motif proteins likely respond to G proteins with distinct mechanisms.

  3. Characterization of structures in biofilms formed by a Pseudomonas fluorescens isolated from soil

    Directory of Open Access Journals (Sweden)

    Wu Siva

    2009-05-01

    Full Text Available Abstract Background Microbial biofilms represent an incompletely understood, but fundamental mode of bacterial growth. These sessile communities typically consist of stratified, morphologically-distinct layers of extracellular material, where numerous metabolic processes occur simultaneously in close proximity. Limited reports on environmental isolates have revealed highly ordered, three-dimensional organization of the extracellular matrix, which may hold important implications for biofilm physiology in vivo. Results A Pseudomonas spp. isolated from a natural soil environment produced flocculent, nonmucoidal biofilms in vitro with unique structural features. These mature biofilms were made up of numerous viable bacteria, even after extended culture, and contained up to 50% of proteins and accumulated 3% (by dry weight calcium, suggesting an important role for the divalent metal in biofilm formation. Ultrastructurally, the mature biofilms contained structural motifs consisting of dense, fibrillary clusters, nanofibers, and ordered, honeycomb-like chambers enveloped in thin sheets. Conclusion Mature biofilms contained living bacteria and were structurally, chemically, and physiologically heterogeneous. The principal architectural elements observed by electron microscopy may represent useful morphological clues for identifying bacterial biofilms in vivo. The complexity and reproducibility of the structural motifs observed in bacterial biofilms appear to be the result of organized assembly, suggesting that this environmental isolate may possess ecological advantages in its natural habitat.

  4. Metagenome survey of a multispecies and alga-associated biofilm revealed key elements of bacterial-algal interactions in photobioreactors.

    Science.gov (United States)

    Krohn-Molt, Ines; Wemheuer, Bernd; Alawi, Malik; Poehlein, Anja; Güllert, Simon; Schmeisser, Christel; Pommerening-Röser, Andreas; Grundhoff, Adam; Daniel, Rolf; Hanelt, Dieter; Streit, Wolfgang R

    2013-10-01

    Photobioreactors (PBRs) are very attractive for sunlight-driven production of biofuels and capturing of anthropogenic CO2. One major problem associated with PBRs however, is that the bacteria usually associated with microalgae in nonaxenic cultures can lead to biofouling and thereby affect algal productivity. Here, we report on a phylogenetic, metagenome, and functional analysis of a mixed-species bacterial biofilm associated with the microalgae Chlorella vulgaris and Scenedesmus obliquus in a PBR. The biofilm diversity and population dynamics were examined through 16S rRNA phylogeny. Overall, the diversity was rather limited, with approximately 30 bacterial species associated with the algae. The majority of the observed microorganisms were affiliated with Alphaproteobacteria, Betaproteobacteria, and Bacteroidetes. A combined approach of sequencing via GS FLX Titanium from Roche and HiSeq 2000 from Illumina resulted in the overall production of 350 Mbp of sequenced DNA, 165 Mbp of which was assembled in larger contigs with a maximum size of 0.2 Mbp. A KEGG pathway analysis suggested high metabolic diversity with respect to the use of polymers and aromatic and nonaromatic compounds. Genes associated with the biosynthesis of essential B vitamins were highly redundant and functional. Moreover, a relatively high number of predicted and functional lipase and esterase genes indicated that the alga-associated bacteria are possibly a major sink for lipids and fatty acids produced by the microalgae. This is the first metagenome study of microalga- and PBR-associated biofilm bacteria, and it gives new clues for improved biofuel production in PBRs.

  5. Environmental factors that shape biofilm formation.

    Science.gov (United States)

    Toyofuku, Masanori; Inaba, Tomohiro; Kiyokawa, Tatsunori; Obana, Nozomu; Yawata, Yutaka; Nomura, Nobuhiko

    2015-01-01

    Cells respond to the environment and alter gene expression. Recent studies have revealed the social aspects of bacterial life, such as biofilm formation. Biofilm formation is largely affected by the environment, and the mechanisms by which the gene expression of individual cells affects biofilm development have attracted interest. Environmental factors determine the cell's decision to form or leave a biofilm. In addition, the biofilm structure largely depends on the environment, implying that biofilms are shaped to adapt to local conditions. Second messengers such as cAMP and c-di-GMP are key factors that link environmental factors with gene regulation. Cell-to-cell communication is also an important factor in shaping the biofilm. In this short review, we will introduce the basics of biofilm formation and further discuss environmental factors that shape biofilm formation. Finally, the state-of-the-art tools that allow us investigate biofilms under various conditions are discussed.

  6. CILAIR-Based Secretome Analysis of Obese Visceral and Subcutaneous Adipose Tissues Reveals Distinctive ECM Remodeling and Inflammation Mediators

    Science.gov (United States)

    Roca-Rivada, Arturo; Belen Bravo, Susana; Pérez-Sotelo, Diego; Alonso, Jana; Isabel Castro, Ana; Baamonde, Iván; Baltar, Javier; Casanueva, Felipe F.; Pardo, María

    2015-01-01

    In the context of obesity, strong evidences support a distinctive pathological contribution of adipose tissue depending on its anatomical site of accumulation. Therefore, subcutaneous adipose tissue (SAT) has been lately considered metabolically benign compared to visceral fat (VAT), whose location is associated to the risk of developing cardiovascular disease, insulin resistance, and other associated comorbidities. Under the above situation, the chronic local inflammation that characterizes obese adipose tissue, has acquired a major role on the pathogenesis of obesity. In this work, we have analyzed for the first time human obese VAT and SAT secretomes using an improved quantitative proteomic approach for the study of tissue secretomes, Comparison of Isotope-Labeled Amino acid Incorporation Rates (CILAIR). The use of double isotope-labeling-CILAIR approach to analyze VAT and SAT secretomes allowed the identification of location-specific secreted proteins and its differential secretion. Additionally to the very high percentage of identified proteins previously implicated in obesity or in its comorbidities, this approach was revealed as a useful tool for the study of the obese adipose tissue microenvironment including extracellular matrix (ECM) remodeling and inflammatory status. The results herein presented reinforce the fact that VAT and SAT depots have distinct features and contribute differentially to metabolic disease. PMID:26198096

  7. Single Particle Tracking reveals two distinct environments for CD4 receptors at the surface of living T lymphocytes

    Energy Technology Data Exchange (ETDEWEB)

    Mascalchi, Patrice; Lamort, Anne Sophie; Salome, Laurence [CNRS, IPBS (Institut de Pharmacologie et de Biologie Structurale), 205 route de Narbonne, BP 64182, F-31077 Toulouse (France); Universite de Toulouse, UPS, IPBS (Institut de Pharmacologie et de Biologie Structurale), F-31077 Toulouse (France); Dumas, Fabrice, E-mail: fabrice.dumas@ipbs.fr [CNRS, IPBS (Institut de Pharmacologie et de Biologie Structurale), 205 route de Narbonne, BP 64182, F-31077 Toulouse (France); Universite de Toulouse, UPS, IPBS (Institut de Pharmacologie et de Biologie Structurale), F-31077 Toulouse (France)

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer We studied the diffusion of single CD4 receptors on living lymphocytes. Black-Right-Pointing-Pointer This study reveals that CD4 receptors have either a random or confined diffusion. Black-Right-Pointing-Pointer The dynamics of unconfined CD4 receptors was accelerated by a temperature raise. Black-Right-Pointing-Pointer The dynamics of confined CD4 receptors was unchanged by a temperature raise. Black-Right-Pointing-Pointer Our results suggest the existence of two different environments for CD4 receptors. -- Abstract: We investigated the lateral diffusion of the HIV receptor CD4 at the surface of T lymphocytes at 20 Degree-Sign C and 37 Degree-Sign C by Single Particle Tracking using Quantum Dots. We found that the receptors presented two major distinct behaviors that were not equally affected by temperature changes. About half of the receptors showed a random diffusion with a diffusion coefficient increasing upon raising the temperature. The other half of the receptors was permanently or transiently confined with unchanged dynamics on raising the temperature. These observations suggest that two distinct subpopulations of CD4 receptors with different environments are present at the surface of living T lymphocytes.

  8. Patch clamp studies of human sperm under physiological ionic conditions reveal three functionally and pharmacologically distinct cation channels.

    Science.gov (United States)

    Mansell, S A; Publicover, S J; Barratt, C L R; Wilson, S M

    2014-05-01

    Whilst fertilizing capacity depends upon a K(+) conductance (GK) that allows the spermatozoon membrane potential (Vm) to be held at a negative value, the characteristics of this conductance in human sperm are virtually unknown. We therefore studied the biophysical/pharmacological properties of the K(+) conductance in spermatozoa from normal donors held under voltage/current clamp in the whole cell recording configuration. Our standard recording conditions were designed to maintain quasi-physiological, Na(+), K(+) and Cl(-) gradients. Experiments that explored the effects of ionic substitution/ion channel blockers upon membrane current/potential showed that resting Vm was dependent upon a hyperpolarizing K(+) current that flowed via channels that displayed only weak voltage dependence and limited (∼7-fold) K(+) versus Na(+) selectivity. This conductance was blocked by quinidine (0.3 mM), bupivacaine (3 mM) and clofilium (50 µM), NNC55-0396 (2 µM) and mibefradil (30 µM), but not by 4-aminopyridine (2 mM, 4-AP). Progesterone had no effect upon the hyperpolarizing K(+) current. Repolarization after a test depolarization consistently evoked a transient inward 'tail current' (ITail) that flowed via a second population of ion channels with poor (∼3-fold) K(+) versus Na(+) selectivity. The activity of these channels was increased by quinidine, 4-AP and progesterone. Vm in human sperm is therefore dependent upon a hyperpolarizing K(+) current that flows via channels that most closely resemble those encoded by Slo3. Although 0.5 µM progesterone had no effect upon these channels, this hormone did activate the pharmacologically distinct channels that mediate ITail. In conclusion, this study reveals three functionally and pharmacologically distinct cation channels: Ik, ITail, ICatSper.

  9. Permeabilizing biofilms

    Science.gov (United States)

    Soukos, Nikolaos S.; Lee, Shun; Doukas,; Apostolos G.

    2008-02-19

    Methods for permeabilizing biofilms using stress waves are described. The methods involve applying one or more stress waves to a biofilm, e.g., on a surface of a device or food item, or on a tissue surface in a patient, and then inducing stress waves to create transient increases in the permeability of the biofilm. The increased permeability facilitates delivery of compounds, such as antimicrobial or therapeutic agents into and through the biofilm.

  10. Biofilm Formation and Detachment in Gram-Negative Pathogens Is Modulated by Select Bile Acids.

    Science.gov (United States)

    Sanchez, Laura M; Cheng, Andrew T; Warner, Christopher J A; Townsley, Loni; Peach, Kelly C; Navarro, Gabriel; Shikuma, Nicholas J; Bray, Walter M; Riener, Romina M; Yildiz, Fitnat H; Linington, Roger G

    2016-01-01

    Biofilms are a ubiquitous feature of microbial community structure in both natural and host environments; they enhance transmission and infectivity of pathogens and provide protection from human defense mechanisms and antibiotics. However, few natural products are known that impact biofilm formation or persistence for either environmental or pathogenic bacteria. Using the combination of a novel natural products library from the fish microbiome and an image-based screen for biofilm inhibition, we describe the identification of taurine-conjugated bile acids as inhibitors of biofilm formation against both Vibrio cholerae and Pseudomonas aeruginosa. Taurocholic acid (1) was isolated from the fermentation broth of the fish microbiome-derived strain of Rhodococcus erythropolis and identified using standard NMR and MS methods. Screening of the twelve predominant human steroidal bile acid components revealed that a subset of these compounds can inhibit biofilm formation, induce detachment of preformed biofilms under static conditions, and that these compounds display distinct structure-activity relationships against V. cholerae and P. aeruginosa. Our findings highlight the significance of distinct bile acid components in the regulation of biofilm formation and dispersion in two different clinically relevant bacterial pathogens, and suggest that the bile acids, which are endogenous mammalian metabolites used to solubilize dietary fats, may also play a role in maintaining host health against bacterial infection.

  11. Biofilm Formation and Detachment in Gram-Negative Pathogens Is Modulated by Select Bile Acids.

    Directory of Open Access Journals (Sweden)

    Laura M Sanchez

    Full Text Available Biofilms are a ubiquitous feature of microbial community structure in both natural and host environments; they enhance transmission and infectivity of pathogens and provide protection from human defense mechanisms and antibiotics. However, few natural products are known that impact biofilm formation or persistence for either environmental or pathogenic bacteria. Using the combination of a novel natural products library from the fish microbiome and an image-based screen for biofilm inhibition, we describe the identification of taurine-conjugated bile acids as inhibitors of biofilm formation against both Vibrio cholerae and Pseudomonas aeruginosa. Taurocholic acid (1 was isolated from the fermentation broth of the fish microbiome-derived strain of Rhodococcus erythropolis and identified using standard NMR and MS methods. Screening of the twelve predominant human steroidal bile acid components revealed that a subset of these compounds can inhibit biofilm formation, induce detachment of preformed biofilms under static conditions, and that these compounds display distinct structure-activity relationships against V. cholerae and P. aeruginosa. Our findings highlight the significance of distinct bile acid components in the regulation of biofilm formation and dispersion in two different clinically relevant bacterial pathogens, and suggest that the bile acids, which are endogenous mammalian metabolites used to solubilize dietary fats, may also play a role in maintaining host health against bacterial infection.

  12. Role of the luxS gene in initial biofilm formation by Streptococcus mutans.

    Science.gov (United States)

    He, Zhiyan; Liang, Jingping; Tang, Zisheng; Ma, Rui; Peng, Huasong; Huang, Zhengwei

    2015-01-01

    Quorum sensing (QS) is a process by which bacteria communicate with each other by secreting chemical signals called autoinducers (AIs). Among Gram-negative and Gram-positive bacteria, AI-2 synthesized by the LuxS enzyme is widespread. The aim of this study was to evaluate the effect of QS luxS gene on initial biofilm formation by Streptococcus mutans. The bacterial cell surface properties, including cell hydrophobicity (bacterial adherence to hydrocarbons) and aggregation, which are important for initial adherence during biofilm development, were investigated. The biofilm adhesion assay was evaluated by the MTT method. The structures of the 5-hour biofilms were observed by using confocal laser scanning microscopy, and QS-related gene expressions were investigated by real-time PCR. The luxS mutant strain exhibited higher biofilm adherence and aggregation, but lower hydrophobicity than the wild-type strain. The confocal laser scanning microscopy images revealed that the wild-type strain tended to form smaller aggregates with uniform distribution, whereas the luxS mutant strain aggregated into distinct clusters easily discernible in the generated biofilm. Most of the genes examined were downregulated in the biofilms formed by the luxS mutant strain, except the gtfB gene. QS luxS gene can affect the initial biofilm formation by S. mutans.

  13. Macromolecular fingerprinting of sulfolobus species in biofilm: a transcriptomic and proteomic approach combined with spectroscopic analysis.

    Science.gov (United States)

    Koerdt, Andrea; Orell, Alvaro; Pham, Trong Khoa; Mukherjee, Joy; Wlodkowski, Alexander; Karunakaran, Esther; Biggs, Catherine A; Wright, Phillip C; Albers, Sonja-Verena

    2011-09-02

    Microorganisms in nature often live in surface-associated sessile communities, encased in a self-produced matrix, referred to as biofilms. Biofilms have been well studied in bacteria but in a limited way for archaea. We have recently characterized biofilm formation in three closely related hyperthermophilic crenarchaeotes: Sulfolobus acidocaldarius, S. solfataricus, and S. tokodaii. These strains form different communities ranging from simple carpet structures in S. solfataricus to high density tower-like structures in S. acidocaldarius under static condition. Here, we combine spectroscopic, proteomic, and transcriptomic analyses to describe physiological and regulatory features associated with biofilms. Spectroscopic analysis reveals that in comparison to planktonic life-style, biofilm life-style has distinctive influence on the physiology of each Sulfolobus spp. Proteomic and transcriptomic data show that biofilm-forming life-style is strain specific (eg ca. 15% of the S. acidocaldarius genes were differently expressed, S. solfataricus and S. tokodaii had ~3.4 and ~1%, respectively). The -omic data showed that regulated ORFs were widely distributed in basic cellular functions, including surface modifications. Several regulated genes are common to biofilm-forming cells in all three species. One of the most striking common response genes include putative Lrs14-like transcriptional regulators, indicating their possible roles as a key regulatory factor in biofilm development.

  14. Characterization of traumatic brain injury in human brains reveals distinct cellular and molecular changes in contusion and pericontusion.

    Science.gov (United States)

    Harish, Gangadharappa; Mahadevan, Anita; Pruthi, Nupur; Sreenivasamurthy, Sreelakshmi K; Puttamallesh, Vinuth N; Keshava Prasad, Thottethodi Subrahmanya; Shankar, Susarla Krishna; Srinivas Bharath, Muchukunte Mukunda

    2015-07-01

    Traumatic brain injury (TBI) contributes to fatalities and neurological disabilities worldwide. While primary injury causes immediate damage, secondary events contribute to long-term neurological defects. Contusions (Ct) are primary injuries correlated with poor clinical prognosis, and can expand leading to delayed neurological deterioration. Pericontusion (PC) (penumbra), the region surrounding Ct, can also expand with edema, increased intracranial pressure, ischemia, and poor clinical outcome. Analysis of Ct and PC can therefore assist in understanding the pathobiology of TBI and its management. This study on human TBI brains noted extensive neuronal, astroglial and inflammatory changes, alterations in mitochondrial, synaptic and oxidative markers, and associated proteomic profile, with distinct differences in Ct and PC. While Ct displayed petechial hemorrhages, thrombosis, inflammation, neuronal pyknosis, and astrogliosis, PC revealed edema, vacuolation of neuropil, axonal loss, and dystrophic changes. Proteomic analysis demonstrated altered immune response, synaptic, and mitochondrial dysfunction, among others, in Ct, while PC displayed altered regulation of neurogenesis and cytoskeletal architecture, among others. TBI brains displayed oxidative damage, glutathione depletion, mitochondrial dysfunction, and loss of synaptic proteins, with these changes being more profound in Ct. We suggest that analysis of markers specific to Ct and PC may be valuable in the evaluation of TBI pathobiology and therapeutics. We have characterized the primary injury in human traumatic brain injury (TBI). Contusions (Ct) - the injury core displayed hemorrhages, inflammation, and astrogliosis, while the surrounding pericontusion (PC) revealed edema, vacuolation, microglial activation, axonal loss, and dystrophy. Proteomic analysis demonstrated altered immune response, synaptic and mitochondrial dysfunction in Ct, and altered regulation of neurogenesis and cytoskeletal architecture in

  15. The ``Swiss cheese'' instability of bacterial biofilms

    Science.gov (United States)

    Jang, Hongchul; Rusconi, Roberto; Stocker, Roman

    2012-11-01

    Bacteria often adhere to surfaces, where they develop polymer-encased communities (biofilms) that display dramatic resistance to antibiotic treatment. A better understanding of cell detachment from biofilms may lead to novel strategies for biofilm disruption. Here we describe a new detachment mode, whereby a biofilm develops a nearly regular array of ~50-100 μm holes. Using surface-treated microfluidic devices, we create biofilms of controlled shape and size. After the passage of an air plug, the break-up of the residual thin liquid film scrapes and rearranges bacteria on the surface, such that a ``Swiss cheese'' pattern is left in the residual biofilm. Fluorescent staining of the polymeric matrix (EPS) reveals that resistance to cell dislodgement correlates with local biofilm age, early settlers having had more time to hunker down. Because few survivors suffice to regrow a biofilm, these results point at the importance of considering microscale heterogeneity in assessing the effectiveness of biofilm removal strategies.

  16. Modeling autosomal recessive cutis laxa type 1C in mice reveals distinct functions for Ltbp-4 isoforms

    Directory of Open Access Journals (Sweden)

    Insa Bultmann-Mellin

    2015-04-01

    Full Text Available Recent studies have revealed an important role for LTBP-4 in elastogenesis. Its mutational inactivation in humans causes autosomal recessive cutis laxa type 1C (ARCL1C, which is a severe disorder caused by defects of the elastic fiber network. Although the human gene involved in ARCL1C has been discovered based on similar elastic fiber abnormalities exhibited by mice lacking the short Ltbp-4 isoform (Ltbp4S−/−, the murine phenotype does not replicate ARCL1C. We therefore inactivated both Ltbp-4 isoforms in the mouse germline to model ARCL1C. Comparative analysis of Ltbp4S−/− and Ltbp4-null (Ltbp4−/− mice identified Ltbp-4L as an important factor for elastogenesis and postnatal survival, and showed that it has distinct tissue expression patterns and specific molecular functions. We identified fibulin-4 as a previously unknown interaction partner of both Ltbp-4 isoforms and demonstrated that at least Ltbp-4L expression is essential for incorporation of fibulin-4 into the extracellular matrix (ECM. Overall, our results contribute to the current understanding of elastogenesis and provide an animal model of ARCL1C.

  17. A chemical-genetic strategy reveals distinct temporal requirements for SAD-1 kinase in neuronal polarization and synapse formation

    Directory of Open Access Journals (Sweden)

    Shokat Kevan M

    2008-09-01

    Full Text Available Abstract Background Neurons assemble into a functional network through a sequence of developmental processes including neuronal polarization and synapse formation. In Caenorhabditis elegans, the serine/threonine SAD-1 kinase is essential for proper neuronal polarity and synaptic organization. To determine if SAD-1 activity regulates the establishment or maintenance of these neuronal structures, we examined its temporal requirements using a chemical-genetic method that allows for selective and reversible inactivation of its kinase activity in vivo. Results We generated a PP1 analog-sensitive variant of SAD-1. Through temporal inhibition of SAD-1 kinase activity we show that its activity is required for the establishment of both neuronal polarity and synaptic organization. However, while SAD-1 activity is needed strictly when neurons are polarizing, the temporal requirement for SAD-1 is less stringent in synaptic organization, which can also be re-established during maintenance. Conclusion This study reports the first temporal analysis of a neural kinase activity using the chemical-genetic system. It reveals that neuronal polarity and synaptic organization have distinct temporal requirements for SAD-1.

  18. Distinct and shared functions of ALS-associated proteins TDP-43, FUS and TAF15 revealed by multisystem analyses.

    Science.gov (United States)

    Kapeli, Katannya; Pratt, Gabriel A; Vu, Anthony Q; Hutt, Kasey R; Martinez, Fernando J; Sundararaman, Balaji; Batra, Ranjan; Freese, Peter; Lambert, Nicole J; Huelga, Stephanie C; Chun, Seung J; Liang, Tiffany Y; Chang, Jeremy; Donohue, John P; Shiue, Lily; Zhang, Jiayu; Zhu, Haining; Cambi, Franca; Kasarskis, Edward; Hoon, Shawn; Ares, Manuel; Burge, Christopher B; Ravits, John; Rigo, Frank; Yeo, Gene W

    2016-07-05

    The RNA-binding protein (RBP) TAF15 is implicated in amyotrophic lateral sclerosis (ALS). To compare TAF15 function to that of two ALS-associated RBPs, FUS and TDP-43, we integrate CLIP-seq and RNA Bind-N-Seq technologies, and show that TAF15 binds to ∼4,900 RNAs enriched for GGUA motifs in adult mouse brains. TAF15 and FUS exhibit similar binding patterns in introns, are enriched in 3' untranslated regions and alter genes distinct from TDP-43. However, unlike FUS and TDP-43, TAF15 has a minimal role in alternative splicing. In human neural progenitors, TAF15 and FUS affect turnover of their RNA targets. In human stem cell-derived motor neurons, the RNA profile associated with concomitant loss of both TAF15 and FUS resembles that observed in the presence of the ALS-associated mutation FUS R521G, but contrasts with late-stage sporadic ALS patients. Taken together, our findings reveal convergent and divergent roles for FUS, TAF15 and TDP-43 in RNA metabolism.

  19. Four tropical, closely related fern species belonging to the genus Adiantum L. are genetically distinct as revealed by ISSR fingerprinting.

    Science.gov (United States)

    Korpelainen, Helena; de Britto, John; Doublet, Jérémy; Pravin, Sahaya

    2005-11-01

    The level and pattern of genetic variation was analyzed in four species of the fern genus Adiantum L., A. hispidulum Sw., A. incisum Forrsk., A. raddianum C.Presl, and A. zollingeri Mett. ex Kuhn, originating from South India, using the ISSR fingerprinting method. The populations of Adiantum possessed a considerable level of genetic variation, the diversity indices ranging from 0.284 to 0.464. Only 12% of the ISSR markers found were restricted to one species only, and 54% were detected in all four species. The analysis of molecular variance revealed that 71.1% of variation was present within populations. The proportion of variation detected among species was only 18.5% while the proportion of variation among populations within species equalled 10.4%. Despite the low level of intrageneric differentiation, the discriminant analysis and clustering of genetic distances indicated that the four Adiantum species are genetically distinct. The F(ST) values calculated for the species were low, varying from 0.089 to 0.179. No linkage disequilibrium was detected between the loci. Such low level of differentiation among populations and the presence of linkage equilibrium reflect that the life history of Adiantum ferns apparently involves common or relatively common sexuality, effective wind-dispersal of spores and outcrossing.

  20. Comparative analysis of ITS1 nucleotide sequence reveals distinct genetic difference between Brugia malayi from Northeast Borneo and Thailand.

    Science.gov (United States)

    Fong, Mun-Yik; Noordin, Rahmah; Lau, Yee-Ling; Cheong, Fei-Wen; Yunus, Muhammad Hafiznur; Idris, Zulkarnain Md

    2013-01-01

    Brugia malayi is one of the parasitic worms which causes lymphatic filariasis in humans. Its geographical distribution includes a large part of Asia. Despite its wide distribution, very little is known about the genetic variation and molecular epidemiology of this species. In this study, the internal transcribed spacer 1 (ITS1) nucleotide sequences of B. malayi from microfilaria-positive human blood samples in Northeast Borneo Island were determined, and compared with published ITS1 sequences of B. malayi isolated from cats and humans in Thailand. Multiple alignment analysis revealed that B. malayi ITS1 sequences from Northeast Borneo were more similar to each other than to those from Thailand. Phylogenetic trees inferred using Neighbour-Joining and Maximum Parsimony methods showed similar topology, with 2 distinct B. malayi clusters. The first cluster consisted of Northeast Borneo B. malayi isolates, whereas the second consisted of the Thailand isolates. The findings of this study suggest that B. malayi in Borneo Island has diverged significantly from those of mainland Asia, and this has implications for the diagnosis of B. malayi infection across the region using ITS1-based molecular techniques.

  1. Neurodegenerative disease mutations in TREM2 reveal a functional surface and distinct loss-of-function mechanisms

    Energy Technology Data Exchange (ETDEWEB)

    Kober, Daniel L.; Alexander-Brett, Jennifer M.; Karch, Celeste M.; Cruchaga, Carlos; Colonna, Marco; Holtzman, Michael J.; Brett, Thomas J. (WU-MED)

    2016-12-20

    Genetic variations in the myeloid immune receptor TREM2 are linked to several neurodegenerative diseases. To determine how TREM2 variants contribute to these diseases, we performed structural and functional studies of wild-type and variant proteins. Our 3.1 Å TREM2 crystal structure revealed that mutations found in Nasu-Hakola disease are buried whereas Alzheimer’s disease risk variants are found on the surface, suggesting that these mutations have distinct effects on TREM2 function. Biophysical and cellular methods indicate that Nasu-Hakola mutations impact protein stability and decrease folded TREM2 surface expression, whereas Alzheimer’s risk variants impact binding to a TREM2 ligand. Additionally, the Alzheimer’s risk variants appear to epitope map a functional surface on TREM2 that is unique within the larger TREM family. These findings provide a guide to structural and functional differences among genetic variants of TREM2, indicating that therapies targeting the TREM2 pathway should be tailored to these genetic and functional differences with patient-specific medicine approaches for neurodegenerative disorders.

  2. Extinction of cocaine self-administration reveals functionally and temporally distinct dopaminergic signals in the nucleus accumbens.

    Science.gov (United States)

    Stuber, Garret D; Wightman, R Mark; Carelli, Regina M

    2005-05-19

    While Pavlovian and operant conditioning influence drug-seeking behavior, the role of rapid dopamine signaling in modulating these processes is unknown. During self-administration of cocaine, two dopaminergic signals, measured with 100 ms resolution, occurred immediately before and after the lever press (termed pre- and post-response dopamine transients). Extinction of self-administration revealed that these two signals were functionally distinct. Pre-response transients, which could reflect the motivation to obtain the drug, did not decline during extinction. Remarkably, post-response dopamine transients attenuated as extinction progressed, suggesting that they encode the learned association between environmental cues and cocaine. A third type of dopamine transient, not time locked to overt stimuli, decreased in frequency during extinction and correlated with calculated cocaine concentrations. These results show that dopamine release transients involved in different aspects of cocaine self-administration are highly plastic--differentially governed by motivation, learned associations linked with environmental stimuli, and the pharmacological actions of cocaine.

  3. Characterization of CLL exosomes reveals a distinct microRNA signature and enhanced secretion by activation of BCR signaling.

    Science.gov (United States)

    Yeh, Yuh-Ying; Ozer, Hatice Gulcin; Lehman, Amy M; Maddocks, Kami; Yu, Lianbo; Johnson, Amy J; Byrd, John C

    2015-05-21

    Multiple studies show that chronic lymphocytic leukemia (CLL) cells are heavily dependent on their microenvironment for survival. Communication between CLL cells and the microenvironment is mediated through direct cell contact, soluble factors, and extracellular vesicles. Exosomes are small particles enclosed with lipids, proteins, and small RNAs that can convey biological materials to surrounding cells. Our data herein demonstrate that CLL cells release significant amounts of exosomes in plasma that exhibit abundant CD37, CD9, and CD63 expression. Our work also pinpoints the regulation of B-cell receptor (BCR) signaling in the release of CLL exosomes: BCR activation by α-immunoglobulin (Ig)M induces exosome secretion, whereas BCR inactivation via ibrutinib impedes α-IgM-stimulated exosome release. Moreover, analysis of serial plasma samples collected from CLL patients on an ibrutinib clinical trial revealed that exosome plasma concentration was significantly decreased following ibrutinib therapy. Furthermore, microRNA (miR) profiling of plasma-derived exosomes identified a distinct exosome microRNA signature, including miR-29 family, miR-150, miR-155, and miR-223 that have been associated with CLL disease. Interestingly, expression of exosome miR-150 and miR-155 increases with BCR activation. In all, this study successfully characterized CLL exosomes, demonstrated the control of BCR signaling in the release of CLL exosomes, and uncovered a disease-relevant exosome microRNA profile.

  4. Proteomics Analysis Reveals Distinct Corona Composition on Magnetic Nanoparticles with Different Surface Coatings: Implications for Interactions with Primary Human Macrophages.

    Science.gov (United States)

    Vogt, Carmen; Pernemalm, Maria; Kohonen, Pekka; Laurent, Sophie; Hultenby, Kjell; Vahter, Marie; Lehtiö, Janne; Toprak, Muhammet S; Fadeel, Bengt

    2015-01-01

    Superparamagnetic iron oxide nanoparticles (SPIONs) have emerged as promising contrast agents for magnetic resonance imaging. The influence of different surface coatings on the biocompatibility of SPIONs has been addressed, but the potential impact of the so-called corona of adsorbed proteins on the surface of SPIONs on their biological behavior is less well studied. Here, we determined the composition of the plasma protein corona on silica-coated versus dextran-coated SPIONs using mass spectrometry-based proteomics approaches. Notably, gene ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed distinct protein corona compositions for the two different SPIONs. Relaxivity of silica-coated SPIONs was modulated by the presence of a protein corona. Moreover, the viability of primary human monocyte-derived macrophages was influenced by the protein corona on silica-coated, but not dextran-coated SPIONs, and the protein corona promoted cellular uptake of silica-coated SPIONs, but did not affect internalization of dextran-coated SPIONs.

  5. Proteomics Analysis Reveals Distinct Corona Composition on Magnetic Nanoparticles with Different Surface Coatings: Implications for Interactions with Primary Human Macrophages.

    Directory of Open Access Journals (Sweden)

    Carmen Vogt

    Full Text Available Superparamagnetic iron oxide nanoparticles (SPIONs have emerged as promising contrast agents for magnetic resonance imaging. The influence of different surface coatings on the biocompatibility of SPIONs has been addressed, but the potential impact of the so-called corona of adsorbed proteins on the surface of SPIONs on their biological behavior is less well studied. Here, we determined the composition of the plasma protein corona on silica-coated versus dextran-coated SPIONs using mass spectrometry-based proteomics approaches. Notably, gene ontology (GO enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG pathway analysis revealed distinct protein corona compositions for the two different SPIONs. Relaxivity of silica-coated SPIONs was modulated by the presence of a protein corona. Moreover, the viability of primary human monocyte-derived macrophages was influenced by the protein corona on silica-coated, but not dextran-coated SPIONs, and the protein corona promoted cellular uptake of silica-coated SPIONs, but did not affect internalization of dextran-coated SPIONs.

  6. Community structure of microbial biofilms associated with membrane-based water purification processes as revealed using a polyphasic approach

    Energy Technology Data Exchange (ETDEWEB)

    Chen, C.L.; Chong, M.L.; Wong, M.T.; Ong, S.L.; Ng, W.J. [Dept. of Civil Engineering, National Univ. of Singapore (Singapore); Liu, W.T. [Dept. of Civil Engineering, National Univ. of Singapore (Singapore); Nanoscience and Nanotechnology Initiative, National Univ. of Singapore (Singapore); Seah, H. [Public Utilities Board (Singapore)

    2004-07-01

    The microbial communities of membrane biofilms occurring in two full-scale water purification processes employing microfiltration (MF) and reverse osmosis (RO) membranes were characterized using a polyphasic approach that employed bacterial cultivation, 16S rDNA clone library and fluorescence in situ hybridization techniques. All methods showed that the {alpha}-proteobacteria was the largest microbial fraction in the samples, followed by the {gamma}-proteobacteria. This suggested that members of these two groups could be responsible for the biofouling on the membranes studied. Furthermore, the microbial community structures between the MF and RO samples were considerably different in composition of the most predominant 16S rDNA clones and bacterial isolates from the {alpha}-proteobacteria and only shared two common groups (Bradyrhizobium, Bosea) out of more than 17 different bacterial groups observed. The MF and RO samples further contained Planctomycetes and Fibroacter/Acidobacteria as the second predominant bacterial clones, respectively, and differed in minor bacterial clones and isolates. The community structure differences were mainly attributed to differences in feed water, process configurations and operating environments, such as the pressure and hydrodynamic conditions present in the water purification systems. (orig.)

  7. Beneficial biofilms

    Directory of Open Access Journals (Sweden)

    Sara R Robertson

    2015-10-01

    Full Text Available Surface-adherent biofilm growth is a common trait of bacteria and other microorganisms in nature. Within biofilms, organisms are present in high density and are enmeshed in an organic matrix containing polysaccharides and other molecules. The close proximity of organisms within biofilms facilitates microbial interactions and signaling, including many metabolic processes in which consortia rather than individual organisms participate. Biofilm growth also enables microorganisms to withstand chemical and biological stresses. Here, we review some current literature and document representative beneficial aspects of biofilms using examples from wastewater treatment, microbial fuel cells, biological repair (biocementation of stonework, and biofilm protection against Candida albicans infections. Finally, we address a chemical ecology strategy whereby desired microbial succession and beneficial biofilm formation can be encouraged via manipulation of culture conditions and bacterial signaling.

  8. Biofilms in chronic wounds.

    Science.gov (United States)

    James, Garth A; Swogger, Ellen; Wolcott, Randall; Pulcini, Elinor deLancey; Secor, Patrick; Sestrich, Jennifer; Costerton, John W; Stewart, Philip S

    2008-01-01

    Chronic wounds including diabetic foot ulcers, pressure ulcers, and venous leg ulcers are a worldwide health problem. It has been speculated that bacteria colonizing chronic wounds exist as highly persistent biofilm communities. This research examined chronic and acute wounds for biofilms and characterized microorganisms inhabiting these wounds. Chronic wound specimens were obtained from 77 subjects and acute wound specimens were obtained from 16 subjects. Culture data were collected using standard clinical techniques. Light and scanning electron microscopy techniques were used to analyze 50 of the chronic wound specimens and the 16 acute wound specimens. Molecular analyses were performed on the remaining 27 chronic wound specimens using denaturing gradient gel electrophoresis and sequence analysis. Of the 50 chronic wound specimens evaluated by microscopy, 30 were characterized as containing biofilm (60%), whereas only one of the 16 acute wound specimens was characterized as containing biofilm (6%). This was a statistically significant difference (p<0.001). Molecular analyses of chronic wound specimens revealed diverse polymicrobial communities and the presence of bacteria, including strictly anaerobic bacteria, not revealed by culture. Bacterial biofilm prevalence in specimens from chronic wounds relative to acute wounds observed in this study provides evidence that biofilms may be abundant in chronic wounds.

  9. The hidden anatomy of paranasal sinuses reveals biogeographically distinct morphotypes in the nine-banded armadillo (Dasypus novemcinctus

    Directory of Open Access Journals (Sweden)

    Guillaume Billet

    2017-08-01

    second medialmost pair of caudal frontal sinuses constitutes an unexpected morphological diagnostic feature for this potentially distinct species. Our results demonstrate the benefits of studying overlooked internal morphological structures in supposedly cryptic species revealed by molecular data. It also illustrates the under-exploited potential of the highly variable paranasal sinuses of armadillos for systematic studies.

  10. Pattern formation in Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Parsek, Matthew R.; Tolker-Nielsen, Tim

    2008-01-01

    Bacteria are capable of forming elaborate multicellular communities called biofilms. Pattern formation in biofilms depends on cell proliferation and cellular migration in response to the available nutrients and other external cues, as well as on self-generated intercellular signal molecules...... and the production of an extracellular matrix that serves as a structural 'scaffolding' for the biofilm cells. Pattern formation in biofilms allows cells to position themselves favorably within nutrient gradients and enables buildup and maintenance of physiologically distinct subpopulations, which facilitates...... survival of one or more subpopulations upon environmental insult, and therefore plays an important role in the innate tolerance displayed by biofilms toward adverse conditions....

  11. Numerical Investigation of Evolution of the Biofilm Streamers

    Science.gov (United States)

    Karimi, Alireza

    2015-11-01

    Filamentous bacterial structures, called streamers, start to form when there is a sustained hydrodynamic flow over a biofilm. Recent experimental studies have reported formation of biofilm streamers in microfluidic chambers. It is speculated that development of an invisible array of extracellular polymeric substances (EPS) gives rise to aggregation of swimming cells and formation of bacterial filaments. In order to investigate this phenomenon, we employ a multiphase biofilm model which treats the bacterial cells, EPS, and background solvent as distinct phases of a complex fluid. Numerical simulations conducted using this theoretical framework reveals the impact of the viscoelasticity of the polymeric substances on the characteristics of the streamers and the complex interplay of shear flow and the bacterial filaments.

  12. Current understanding of multi-species biofilms

    DEFF Research Database (Denmark)

    Yang, Liang; Liu, Yang; Wu, Hong

    2011-01-01

    Direct observation of a wide range of natural microorganisms has revealed the fact that the majority of microbes persist as surface-attached communities surrounded by matrix materials, called biofilms. Biofilms can be formed by a single bacterial strain. However, most natural biofilms are actuall...

  13. Current understanding of multi-species biofilms

    DEFF Research Database (Denmark)

    Yang, Liang; Liu, Yang; Wu, Hong

    2011-01-01

    Direct observation of a wide range of natural microorganisms has revealed the fact that the majority of microbes persist as surface-attached communities surrounded by matrix materials, called biofilms. Biofilms can be formed by a single bacterial strain. However, most natural biofilms are actuall...

  14. Staphylococcus aureus Biofilm and Planktonic cultures differentially impact gene expression, mapk phosphorylation, and cytokine production in human keratinocytes

    Directory of Open Access Journals (Sweden)

    Olerud John E

    2011-06-01

    Full Text Available Abstract Background Many chronic diseases, such as non-healing wounds are characterized by prolonged inflammation and respond poorly to conventional treatment. Bacterial biofilms are a major impediment to wound healing. Persistent infection of the skin allows the formation of complex bacterial communities termed biofilm. Bacteria living in biofilms are phenotypically distinct from their planktonic counterparts and are orders of magnitude more resistant to antibiotics, host immune response, and environmental stress. Staphylococcus aureus is prevalent in cutaneous infections such as chronic wounds and is an important human pathogen. Results The impact of S. aureus soluble products in biofilm-conditioned medium (BCM or in planktonic-conditioned medium (PCM on human keratinocytes was investigated. Proteomic analysis of BCM and PCM revealed differential protein compositions with PCM containing several enzymes involved in glycolysis. Global gene expression of keratinocytes exposed to biofilm and planktonic S. aureus was analyzed after four hours of exposure. Gene ontology terms associated with responses to bacteria, inflammation, apoptosis, chemotaxis, and signal transduction were enriched in BCM treated keratinocytes. Several transcripts encoding cytokines were also upregulated by BCM after four hours. ELISA analysis of cytokines confirmed microarray results at four hours and revealed that after 24 hours of exposure, S. aureus biofilm induced sustained low level cytokine production compared to near exponential increases of cytokines in planktonic treated keratinocytes. The reduction in cytokines produced by keratinocytes exposed to biofilm was accompanied by suppressed phosphorylation of MAPKs. Chemical inhibition of MAPKs did not drastically reduce cytokine production in BCM-treated keratinocytes suggesting that the majority of cytokine production is mediated through MAPK-independent mechanisms. Conclusions Collectively the results indicate that S

  15. Dynamics of Dual Infection with Campylobacter jejuni Strains in Chickens Reveals Distinct Strain-to-Strain Variation in Infection Ecology

    Science.gov (United States)

    Wigley, Paul; Humphrey, Suzanne; Kemmett, Kirsty; Lacharme-Lora, Lizeth; Humphrey, Tom; Williams, Nicola

    2014-01-01

    Although multiple genotypes of Campylobacter jejuni may be isolated from the same commercial broiler flock, little is known about the infection dynamics of different genotypes within individuals or their colonization sites within the gut. Single experimental infections with C. jejuni M1 (sequence type 137, clonal complex 45) and C. jejuni 13126 (sequence type 21, clonal complex 21) revealed that 13126 colonized the ceca at significantly higher levels. The dissemination and colonization sites of the two C. jejuni strains then were examined in an experimental broiler flock. Two 33-day-old broiler chickens were infected with M1 and two with 13126, and 15 birds were left unchallenged. Cloacal swabs were taken postinfection to determine the colonization and shedding of each strain. By 2 days postinfection (dpi), 8/19 birds were shedding M1 whereas none were shedding 13126. At 8 dpi, all birds were shedding both strains. At 18 dpi, liver and cecal levels of each isolate were quantified, while in 10 birds they also were quantified at nine sites throughout the gastrointestinal (GI) tract. 13126 was found throughout the GI tract, while M1 was largely restricted to the ceca and colon. The livers of 7/19 birds were culture positive for 13126 only. These data show that 13126 has a distinctly different infection biology than strain M1. It showed slower colonization of the lower GI tract but was more invasive and able to colonize at a high level throughout the GI tract. The finding that C. jejuni strains have markedly different infection ecologies within the chicken has implications for control in the poultry industry and suggests that the contamination risk of edible tissues is dependent on the isolate involved. PMID:25107966

  16. Promiscuous binding of invariant chain-derived CLIP peptide to distinct HLA-I molecules revealed in leukemic cells.

    Science.gov (United States)

    van Luijn, Marvin M; van de Loosdrecht, Arjan A; Lampen, Margit H; van Veelen, Peter A; Zevenbergen, Adri; Kester, Michel G D; de Ru, Arnoud H; Ossenkoppele, Gert J; van Hall, Thorbald; van Ham, S Marieke

    2012-01-01

    Antigen presentation by HLA class I (HLA-I) and HLA class II (HLA-II) complexes is achieved by proteins that are specific for their respective processing pathway. The invariant chain (Ii)-derived peptide CLIP is required for HLA-II-mediated antigen presentation by stabilizing HLA-II molecules before antigen loading through transient and promiscuous binding to different HLA-II peptide grooves. Here, we demonstrate alternative binding of CLIP to surface HLA-I molecules on leukemic cells. In HLA-II-negative AML cells, we found plasma membrane display of the CLIP peptide. Silencing Ii in AML cells resulted in reduced HLA-I cell surface display, which indicated a direct role of CLIP in the HLA-I antigen presentation pathway. In HLA-I-specific peptide eluates from B-LCLs, five Ii-derived peptides were identified, of which two were from the CLIP region. In vitro peptide binding assays strikingly revealed that the eluted CLIP peptide RMATPLLMQALPM efficiently bound to four distinct HLA-I supertypes (-A2, -B7, -A3, -B40). Furthermore, shorter length variants of this CLIP peptide also bound to these four supertypes, although in silico algorithms only predicted binding to HLA-A2 or -B7. Immunization of HLA-A2 transgenic mice with these peptides did not induce CTL responses. Together these data show a remarkable promiscuity of CLIP for binding to a wide variety of HLA-I molecules. The found participation of CLIP in the HLA-I antigen presentation pathway could reflect an aberrant mechanism in leukemic cells, but might also lead to elucidation of novel processing pathways or immune escape mechanisms.

  17. Promiscuous binding of invariant chain-derived CLIP peptide to distinct HLA-I molecules revealed in leukemic cells.

    Directory of Open Access Journals (Sweden)

    Marvin M van Luijn

    Full Text Available Antigen presentation by HLA class I (HLA-I and HLA class II (HLA-II complexes is achieved by proteins that are specific for their respective processing pathway. The invariant chain (Ii-derived peptide CLIP is required for HLA-II-mediated antigen presentation by stabilizing HLA-II molecules before antigen loading through transient and promiscuous binding to different HLA-II peptide grooves. Here, we demonstrate alternative binding of CLIP to surface HLA-I molecules on leukemic cells. In HLA-II-negative AML cells, we found plasma membrane display of the CLIP peptide. Silencing Ii in AML cells resulted in reduced HLA-I cell surface display, which indicated a direct role of CLIP in the HLA-I antigen presentation pathway. In HLA-I-specific peptide eluates from B-LCLs, five Ii-derived peptides were identified, of which two were from the CLIP region. In vitro peptide binding assays strikingly revealed that the eluted CLIP peptide RMATPLLMQALPM efficiently bound to four distinct HLA-I supertypes (-A2, -B7, -A3, -B40. Furthermore, shorter length variants of this CLIP peptide also bound to these four supertypes, although in silico algorithms only predicted binding to HLA-A2 or -B7. Immunization of HLA-A2 transgenic mice with these peptides did not induce CTL responses. Together these data show a remarkable promiscuity of CLIP for binding to a wide variety of HLA-I molecules. The found participation of CLIP in the HLA-I antigen presentation pathway could reflect an aberrant mechanism in leukemic cells, but might also lead to elucidation of novel processing pathways or immune escape mechanisms.

  18. Coexpression network analysis in chronic hepatitis B and C hepatic lesions reveals distinct patterns of disease progression to hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    Danning He; Zhi-Ping Liu; Masao Honda; Shuichi Kaneko; Luonan Chen

    2012-01-01

    Chronic infections with the hepatitis B virus (HBV) and hepatitis C virus (HCV) are the major risks of hepatocellular carcinoma (HCC),and great efforts have been made towards the understanding of the different mechanisms that link the viral infection of hepatic lesions to HCC development.In this work,we developed a novel framework to identify distinct patterns of gene coexpression networks and inflammation-related modules from genome-scale microarray data upon viral infection,and further classified them into oncogenic and dysfunctional ones.The core of our framework lies in the comparative study on viral infection modules across different disease stages and disease types--the module preservation during disease progression is evaluated according to the change of network connectivity in different stages,while the similarity and difference in HBV and HCV are evaluated by comparing the overlap of gene compositions and functional annotations in HBV and HCV modules.In particular,we revealed two types of driving modules related to infection for carcinogenesis in HBV and HCV,respectively,i.e.pro-apoptosis modules that are oncogenic in HBV,and antiapoptosis and inflammation modules that are oncogenic in HCV,which are in concordance with the results of previous differential expression-based approaches.Moreover,we found that intracellular protein transmembrane transportation and the transmembrane receptor protein tyrosine kinase signaling pathway act as oncogenic factors in HBV-HCC.Our findings provide novel insights into viral hepatocarcinogenesis and disease progression,and also demonstrate the advantages of an integrative and comparative network analysis over the existing differential expression-based approach and virus-host interactome-based approach.

  19. High-throughput SHAPE analysis reveals structures in HIV-1 genomic RNA strongly conserved across distinct biological states.

    Directory of Open Access Journals (Sweden)

    Kevin A Wilkinson

    2008-04-01

    Full Text Available Replication and pathogenesis of the human immunodeficiency virus (HIV is tightly linked to the structure of its RNA genome, but genome structure in infectious virions is poorly understood. We invent high-throughput SHAPE (selective 2'-hydroxyl acylation analyzed by primer extension technology, which uses many of the same tools as DNA sequencing, to quantify RNA backbone flexibility at single-nucleotide resolution and from which robust structural information can be immediately derived. We analyze the structure of HIV-1 genomic RNA in four biologically instructive states, including the authentic viral genome inside native particles. Remarkably, given the large number of plausible local structures, the first 10% of the HIV-1 genome exists in a single, predominant conformation in all four states. We also discover that noncoding regions functioning in a regulatory role have significantly lower (p-value < 0.0001 SHAPE reactivities, and hence more structure, than do viral coding regions that function as the template for protein synthesis. By directly monitoring protein binding inside virions, we identify the RNA recognition motif for the viral nucleocapsid protein. Seven structurally homologous binding sites occur in a well-defined domain in the genome, consistent with a role in directing specific packaging of genomic RNA into nascent virions. In addition, we identify two distinct motifs that are targets for the duplex destabilizing activity of this same protein. The nucleocapsid protein destabilizes local HIV-1 RNA structure in ways likely to facilitate initial movement both of the retroviral reverse transcriptase from its tRNA primer and of the ribosome in coding regions. Each of the three nucleocapsid interaction motifs falls in a specific genome domain, indicating that local protein interactions can be organized by the long-range architecture of an RNA. High-throughput SHAPE reveals a comprehensive view of HIV-1 RNA genome structure, and further

  20. Large-scale expression analysis reveals distinct microRNA profiles at different stages of human neurodevelopment.

    Directory of Open Access Journals (Sweden)

    Brandon Smith

    Full Text Available BACKGROUND: MicroRNAs (miRNAs are short non-coding RNAs predicted to regulate one third of protein coding genes via mRNA targeting. In conjunction with key transcription factors, such as the repressor REST (RE1 silencing transcription factor, miRNAs play crucial roles in neurogenesis, which requires a highly orchestrated program of gene expression to ensure the appropriate development and function of diverse neural cell types. Whilst previous studies have highlighted select groups of miRNAs during neural development, there remains a need for amenable models in which miRNA expression and function can be analyzed over the duration of neurogenesis. PRINCIPAL FINDINGS: We performed large-scale expression profiling of miRNAs in human NTera2/D1 (NT2 cells during retinoic acid (RA-induced transition from progenitors to fully differentiated neural phenotypes. Our results revealed dynamic changes of miRNA patterns, resulting in distinct miRNA subsets that could be linked to specific neurodevelopmental stages. Moreover, the cell-type specific miRNA subsets were very similar in NT2-derived differentiated cells and human primary neurons and astrocytes. Further analysis identified miRNAs as putative regulators of REST, as well as candidate miRNAs targeted by REST. Finally, we confirmed the existence of two predicted miRNAs; pred-MIR191 and pred-MIR222 associated with SLAIN1 and FOXP2, respectively, and provided some evidence of their potential co-regulation. CONCLUSIONS: In the present study, we demonstrate that regulation of miRNAs occurs in precise patterns indicative of their roles in cell fate commitment, progenitor expansion and differentiation into neurons and glia. Furthermore, the similarity between our NT2 system and primary human cells suggests their roles in molecular pathways critical for human in vivo neurogenesis.

  1. Distinct genetic diversity of Oncomelania hupensis, intermediate host of Schistosoma japonicum in mainland China as revealed by ITS sequences.

    Directory of Open Access Journals (Sweden)

    Qin Ping Zhao

    Full Text Available BACKGROUND: Oncomelania hupensis is the unique intermediate host of Schistosoma japonicum, which causes schistosomiasis endemic in the Far East, and especially in mainland China. O. hupensis largely determines the parasite's geographical range. How O. hupensis's genetic diversity is distributed geographically in mainland China has never been well examined with DNA sequence data. METHODOLOGY/PRINCIPAL FINDINGS: In this study we investigate the genetic variation among O. hupensis from different geographical origins using the combined complete internal transcribed spacer 1 (ITS1 and ITS2 regions of nuclear ribosomal DNA. 165 O. hupensis isolates were obtained in 29 localities from 7 provinces across mainland China: lake/marshland and hill regions in Anhui, Hubei, Hunan, Jiangxi and Jiangsu provinces, located along the middle and lower reaches of Yangtze River, and mountainous regions in Sichuan and Yunnan provinces. Phylogenetic and haplotype network analyses showed distinct genetic diversity and no shared haplotypes between populations from lake/marshland regions of the middle and lower reaches of the Yangtze River and populations from mountainous regions of Sichuan and Yunnan provinces. The genetic distance between these two groups is up to 0.81 based on Fst, and branch time was estimated as 2-6 Ma. As revealed in the phylogenetic tree, snails from Sichuan and Yunnan provinces were also clustered separately. Geographical separation appears to be an important factor accounting for the diversification of the two groups of O. hupensis in mainland China, and probably for the separate clades between snails from Sichuan and Yunnan provinces. In lake/marshland and hill regions along the middle and lower reaches of the Yangtze River, three clades were identified in the phylogenetic tree, but without any obvious clustering of snails from different provinces. CONCLUSIONS: O. hupensis in mainland China may have considerable genetic diversity, and a more

  2. Molecular Analysis of Shower Curtain Biofilm Microbes

    OpenAIRE

    Kelley, Scott T.; Theisen, Ulrike; Angenent, Largus T.; Amand, Allison St.; Pace, Norman R.

    2004-01-01

    Households provide environments that encourage the formation of microbial communities, often as biofilms. Such biofilms constitute potential reservoirs for pathogens, particularly for immune-compromised individuals. One household environment that potentially accumulates microbial biofilms is that provided by vinyl shower curtains. Over time, vinyl shower curtains accumulate films, commonly referred to as “soap scum,” which microscopy reveals are constituted of lush microbial biofilms. To dete...

  3. Biofilm Infections

    DEFF Research Database (Denmark)

    Bjarnsholt, Thomas; Jensen, Peter Østrup; Moser, Claus Ernst

    A still increasing interest and emphasis on the sessile bacterial lifestyle biofilms has been seen since it was realized that the vast majority of the total microbial biomass exists as biofilms. Aggregation of bacteria was first described by Leeuwenhoek in 1677, but only recently recognized...... as being important in chronic infection. In 1993 the American Society for Microbiology (ASM) recognized that the biofilm mode of growth was relevant to microbiology. This book covers both the evidence for biofilms in many chronic bacterial infections as well as the problems facing these infections...... such as diagnostics, pathogenesis, treatment regimes and in vitro and in vivo models for studying biofilms. This is the first scientific book on biofilm infections, chapters written by the world leading scientist and clinicians. The intended audience of this book is scientists, teachers at university level as well...

  4. Biofilms in wounds

    DEFF Research Database (Denmark)

    Cooper, R A; Bjarnsholt, Thomas; Alhede, M

    2014-01-01

    Following confirmation of the presence of biofilms in chronic wounds, the term biofilm became a buzzword within the wound healing community. For more than a century pathogens have been successfully isolated and identified from wound specimens using techniques that were devised in the nineteenth...... extracellular polymeric substances (EPS). Cells within such aggregations (or biofilms) display varying physiological and metabolic properties that are distinct from those of planktonic cells, and which contribute to their persistence. There are many factors that influence healing in wounds and the discovery...... century by Louis Pasteur and Robert Koch. Although this approach still provides valuable information with which to help diagnose acute infections and to select appropriate antibiotic therapies, it is evident that those organisms isolated from clinical specimens with the conditions normally used...

  5. The characterization of functions involved in the establishment and maturation of Klebsiella pneumoniae in vitro biofilm reveals dual roles for surface exopolysaccharides

    DEFF Research Database (Denmark)

    Balestrino, D.; Ghigo, J.M.; Charbonnel, N.

    2008-01-01

    The ability to form biofilm is seen as an increasingly important colonization strategy among both pathogenic and environmental Klebsiella pneumoniae strains. The aim of the present study was to identify abiotic surface colonization factors of K. pneumoniae using different models at different phases...... of biofilm development. A 2200 K. pneumoniae mutant library previously obtained by signature-tagged mutagenesis was screened in static and dynamic culture models to detect clones impaired at early and/or mature stages of biofilm formation. A total of 28 mutants were affected during late phases of biofilm...... that LPS is involved in initial adhesion on both glass and polyvinyl-chloride and the capsule required for the appropriate initial coverage of substratum and the construction of mature biofilm architecture. These results give new insight into the bacterial factors sequentially associated with the ability...

  6. The Pseudomonas aeruginosa transcriptome in planktonic cultures and static biofilms using RNA sequencing.

    Directory of Open Access Journals (Sweden)

    Andreas Dötsch

    Full Text Available In this study, we evaluated how gene expression differs in mature Pseudomonas aeruginosa biofilms as opposed to planktonic cells by the use of RNA sequencing technology that gives rise to both quantitative and qualitative information on the transcriptome. Although a large proportion of genes were consistently regulated in both the stationary phase and biofilm cultures as opposed to the late exponential growth phase cultures, the global biofilm gene expression pattern was clearly distinct indicating that biofilms are not just surface attached cells in stationary phase. A large amount of the genes found to be biofilm specific were involved in adaptation to microaerophilic growth conditions, repression of type three secretion and production of extracellular matrix components. Additionally, we found many small RNAs to be differentially regulated most of them similarly in stationary phase cultures and biofilms. A qualitative analysis of the RNA-seq data revealed more than 3000 putative transcriptional start sites (TSS. By the use of rapid amplification of cDNA ends (5'-RACE we confirmed the presence of three different TSS associated with the pqsABCDE operon, two in the promoter of pqsA and one upstream of the second gene, pqsB. Taken together, this study reports the first transcriptome study on P. aeruginosa that employs RNA sequencing technology and provides insights into the quantitative and qualitative transcriptome including the expression of small RNAs in P. aeruginosa biofilms.

  7. Novel entries in a fungal biofilm matrix encyclopedia.

    Science.gov (United States)

    Zarnowski, Robert; Westler, William M; Lacmbouh, Ghislain Ade; Marita, Jane M; Bothe, Jameson R; Bernhardt, Jörg; Lounes-Hadj Sahraoui, Anissa; Fontaine, Joël; Sanchez, Hiram; Hatfield, Ronald D; Ntambi, James M; Nett, Jeniel E; Mitchell, Aaron P; Andes, David R

    2014-08-05

    Virulence of Candida is linked with its ability to form biofilms. Once established, biofilm infections are nearly impossible to eradicate. Biofilm cells live immersed in a self-produced matrix, a blend of extracellular biopolymers, many of which are uncharacterized. In this study, we provide a comprehensive analysis of the matrix manufactured by Candida albicans both in vitro and in a clinical niche animal model. We further explore the function of matrix components, including the impact on drug resistance. We uncovered components from each of the macromolecular classes (55% protein, 25% carbohydrate, 15% lipid, and 5% nucleic acid) in the C. albicans biofilm matrix. Three individual polysaccharides were identified and were suggested to interact physically. Surprisingly, a previously identified polysaccharide of functional importance, β-1,3-glucan, comprised only a small portion of the total matrix carbohydrate. Newly described, more abundant polysaccharides included α-1,2 branched α-1,6-mannans (87%) associated with unbranched β-1,6-glucans (13%) in an apparent mannan-glucan complex (MGCx). Functional matrix proteomic analysis revealed 458 distinct activities. The matrix lipids consisted of neutral glycerolipids (89.1%), polar glycerolipids (10.4%), and sphingolipids (0.5%). Examination of matrix nucleic acid identified DNA, primarily noncoding sequences. Several of the in vitro matrix components, including proteins and each of the polysaccharides, were also present in the matrix of a clinically relevant in vivo biofilm. Nuclear magnetic resonance (NMR) analysis demonstrated interaction of aggregate matrix with the antifungal fluconazole, consistent with a role in drug impedance and contribution of multiple matrix components. Importance: This report is the first to decipher the complex and unique macromolecular composition of the Candida biofilm matrix, demonstrate the clinical relevance of matrix components, and show that multiple matrix components are needed

  8. Salmonella biofilms

    NARCIS (Netherlands)

    Castelijn, G.A.A.

    2013-01-01

    Biofilm formation by Salmonellaspp. is a problem in the food industry, since biofilms may act as a persistent source of product contamination. Therefore the aim of this study was to obtain more insight in the processes involved and the factors contributing to Salmonellabiofilm formation. A collectio

  9. Phylogenetic relationships and protein modelling revealed two distinct subfamilies of group II HKT genes between crop and model grasses.

    Science.gov (United States)

    Ariyarathna, H A Chandima K; Francki, Michael G

    2016-07-01

    Molecular evolution of large protein families in closely related species can provide useful insights on structural functional relationships. Phylogenetic analysis of the grass-specific group II HKT genes identified two distinct subfamilies, I and II. Subfamily II was represented in all species, whereas subfamily I was identified only in the small grain cereals and possibly originated from an ancestral gene duplication post divergence from the coarse grain cereal lineage. The core protein structures were highly analogous despite there being no more than 58% amino acid identity between members of the two subfamilies. Distinctly variable regions in known functional domains, however, indicated functional divergence of the two subfamilies. The subsets of codons residing external to known functional domains predicted signatures of positive Darwinian selection potentially identifying new domains of functional divergence and providing new insights on the structural function and relationships between protein members of the two subfamilies.

  10. Auditioning the distinctiveness account: Expanding the production effect to the auditory modality reveals the superiority of writing over vocalising.

    Science.gov (United States)

    Mama, Yaniv; Icht, Michal

    2016-01-01

    The production effect (PE) documents the advantage in memory performance for words that are read aloud during study, rather than words that are read silently. Until now, the PE was examined in the visual modality, as the participants read the study words. In the present study, we extended the PE phenomenon and used the auditory modality at study. This novel methodology provides a critical test of the distinctiveness account. Accordingly, the participants heard the study words and learned them by vocal production (saying aloud) or by writing, followed by a free recall test. The use of the auditory modality yielded a memory advantage for words that were written during study over words that were vocally produced. We explain this result in light of the encoding distinctiveness account, suggesting that the PE is determined by the number of different encoding processes involved in learning, emphasising the essential role of active production.

  11. Three-dimensional motion aftereffects reveal distinct direction-selective mechanisms for binocular processing of motion through depth

    Science.gov (United States)

    Czuba, Thaddeus B.; Rokers, Bas; Guillet, Kyle; Huk, Alexander C.; Cormack, Lawrence K.

    2013-01-01

    Motion aftereffects are historically considered evidence for neuronal populations tuned to specific directions of motion. Despite a wealth of motion aftereffect studies investigating 2D (frontoparallel) motion mechanisms, there is a remarkable dearth of psychophysical evidence for neuronal populations selective for the direction of motion through depth (i.e., tuned to 3D motion). We compared the effects of prolonged viewing of unidirectional motion under dichoptic and monocular conditions and found large 3D motion aftereffects that could not be explained by simple inheritance of 2D monocular aftereffects. These results (1) demonstrate the existence of neurons tuned to 3D motion as distinct from monocular 2D mechanisms, (2) show that distinct 3D direction selectivity arises from both interocular velocity differences and changing disparities over time, and (3) provide a straightforward psychophysical tool for further probing 3D motion mechanisms. PMID:21945967

  12. Time to Go Our Separate Ways: Opposite Effects of Study Duration on Priming and Recognition Reveal Distinct Neural Substrates

    OpenAIRE

    Voss, Joel L.; Gonsalves, Brian D.

    2010-01-01

    Amnesic patients have difficulties recognizing when stimuli are repeated, even though their responses to stimuli can change as a function of repetition in indirect tests of memory – a pattern known as priming without recognition. Likewise, experimental manipulations can impair recognition in healthy individuals while leaving priming relatively unaffected, and priming and recognition have been associated with distinct neural correlates in these circumstances. Does this evidence necessarily ind...

  13. Biofilm development

    DEFF Research Database (Denmark)

    Tolker-Nielsen, Tim

    2015-01-01

    During the past decade we have gained much knowledge about the molecular mechanisms that are involved in initiation and termination of biofilm formation. In many bacteria, these processes appear to occur in response to specific environmental cues and result in, respectively, induction or terminat......During the past decade we have gained much knowledge about the molecular mechanisms that are involved in initiation and termination of biofilm formation. In many bacteria, these processes appear to occur in response to specific environmental cues and result in, respectively, induction...... or termination of biofilm matrix production via the second messenger molecule c-di-GMP. In between initiation and termination of biofilm formation we have defined specific biofilm stages, but the currently available evidence suggests that these transitions are mainly governed by adaptive responses......, and not by specific genetic programs. It appears that biofilm formation can occur through multiple pathways and that the spatial structure of the biofilms is species dependent as well as dependent on environmental conditions. Bacterial subpopulations, e.g., motile and nonmotile subpopulations, can develop...

  14. Coupling genetic and chemical microbiome profiling reveals heterogeneity of archaeome and bacteriome in subsurface biofilms that are dominated by the same archaeal species.

    Directory of Open Access Journals (Sweden)

    Alexander J Probst

    Full Text Available Earth harbors an enormous portion of subsurface microbial life, whose microbiome flux across geographical locations remains mainly unexplored due to difficult access to samples. Here, we investigated the microbiome relatedness of subsurface biofilms of two sulfidic springs in southeast Germany that have similar physical and chemical parameters and are fed by one deep groundwater current. Due to their unique hydrogeological setting these springs provide accessible windows to subsurface biofilms dominated by the same uncultivated archaeal species, called SM1 Euryarchaeon. Comparative analysis of infrared imaging spectra demonstrated great variations in archaeal membrane composition between biofilms of the two springs, suggesting different SM1 euryarchaeal strains of the same species at both aquifer outlets. This strain variation was supported by ultrastructural and metagenomic analyses of the archaeal biofilms, which included intergenic spacer region sequencing of the rRNA gene operon. At 16S rRNA gene level, PhyloChip G3 DNA microarray detected similar biofilm communities for archaea, but site-specific communities for bacteria. Both biofilms showed an enrichment of different deltaproteobacterial operational taxonomic units, whose families were, however, congruent as were their lipid spectra. Consequently, the function of the major proportion of the bacteriome appeared to be conserved across the geographic locations studied, which was confirmed by dsrB-directed quantitative PCR. Consequently, microbiome differences of these subsurface biofilms exist at subtle nuances for archaea (strain level variation and at higher taxonomic levels for predominant bacteria without a substantial perturbation in bacteriome function. The results of this communication provide deep insight into the dynamics of subsurface microbial life and warrant its future investigation with regard to metabolic and genomic analyses.

  15. Comparison of mitochondrial and nucleolar RNase MRP reveals identical RNA components with distinct enzymatic activities and protein components.

    Science.gov (United States)

    Lu, Qiaosheng; Wierzbicki, Sara; Krasilnikov, Andrey S; Schmitt, Mark E

    2010-03-01

    RNase MRP is a ribonucleoprotein endoribonuclease found in three cellular locations where distinct substrates are processed: the mitochondria, the nucleolus, and the cytoplasm. Cytoplasmic RNase MRP is the nucleolar enzyme that is transiently relocalized during mitosis. Nucleolar RNase MRP (NuMRP) was purified to homogeneity, and we extensively purified the mitochondrial RNase MRP (MtMRP) to a single RNA component identical to the NuMRP RNA. Although the protein components of the NuMRP were identified by mass spectrometry successfully, none of the known NuMRP proteins were found in the MtMRP preparation. Only trace amounts of the core NuMRP protein, Pop4, were detected in MtMRP by Western blot. In vitro activity of the two enzymes was compared. MtMRP cleaved only mitochondrial ORI5 substrate, while NuMRP cleaved all three substrates. However, the NuMRP enzyme cleaved the ORI5 substrate at sites different than the MtMRP enzyme. In addition, enzymatic differences in preferred ionic strength confirm these enzymes as distinct entities. Magnesium was found to be essential to both enzymes. We tested a number of reported inhibitors including puromycin, pentamidine, lithium, and pAp. Puromycin inhibition suggested that it binds directly to the MRP RNA, reaffirming the role of the RNA component in catalysis. In conclusion, our study confirms that the NuMRP and MtMRP enzymes are distinct entities with differing activities and protein components but a common RNA subunit, suggesting that the RNA must be playing a crucial role in catalytic activity.

  16. Targeted gene knockdown in zebrafish reveals distinct intraembryonic functions for insulin-like growth factor II signaling.

    Science.gov (United States)

    White, Yvonne A R; Kyle, Joshua T; Wood, Antony W

    2009-09-01

    IGF-II is the predominant IGF ligand regulating prenatal growth in all vertebrates, including humans, but its central role in placental development has confounded efforts to fully elucidate its functions within the embryo. Here we use a nonplacental model vertebrate (zebrafish) to interrogate the intraembryonic functions of IGF-II signaling. The zebrafish genome contains two coorthologs of mammalian IGF2 (igf2a, igf2b), which exhibit distinct patterns of expression during embryogenesis. Expression of igf2a mRNA is restricted to the notochord, primarily during segmentation/neurulation. By contrast, igf2b mRNA is expressed in midline tissues adjacent to the notochord, with additional sites of expression in the ventral forebrain, and the pronephros. To identify their intraembryonic functions, we suppressed the expression of each gene with morpholino oligonucleotides. Knockdown of igf2a led to defects in dorsal midline development, characterized by delayed segmentation, notochord undulations, and ventral curvature. Similarly, suppression of igf2b led to defects in dorsal midline development but also induced ectopic fusion of the nephron primordia, and defects in ventral forebrain development. Subsequent onset of severe body edema in igf2b, but not igf2a morphants, further suggested a distinct role for igf2b in development of the embryonic kidney. Simultaneous knockdown of both genes increased the severity of dorsal midline defects, confirming a conserved role for both genes in dorsal midline development. Collectively, these data provide evidence that the zebrafish orthologs of IGF2 function in dorsal midline development during segmentation/neurulation, whereas one paralog, igf2b, has evolved additional, distinct functions during subsequent organogenesis.

  17. Distinctive Drug-resistant Mutation Profiles and Interpretations of HIV-1 Proviral DNA Revealed by Deep Sequencing in Reverse Transcriptase

    Institute of Scientific and Technical Information of China (English)

    YIN Qian Qian; SHAO Yi Ming; MA Li Ying; LI Zhen Peng; ZHAO Hai; PAN Dong; WANG Yan; XU Wei Si; XING Hui; FENGYi; JIANG Shi Bo

    2016-01-01

    ObjectiveTo investigate distinctive features in drug-resistant mutations(DRMs) and interpretations for reverse transcriptase inhibitors (RTIs) between proviral DNA and paired viral RNA in HIV-1-infected patients. MethodsForty-three HIV-1-infected individuals receiving first-line antiretroviral therapy were recruited to participate in a multicenter AIDS Cohort Study in Anhui and Henan Provinces in China in 2004. Drug resistance genotyping was performed by bulk sequencing and deep sequencing on the plasma and whole blood of 77 samples, respectively. Drug-resistance interpretation was compared between viral RNA and paired proviral DNA. ResultsCompared with bulk sequencing, deep sequencing could detect more DRMs and samples with DRMs in both viral RNA and proviral DNA. The mutations M184I and M230I were more prevalent in proviral DNA than in viral RNA (Fisher’s exact test,P ConclusionCompared with viral RNA, the distinctive information of DRMsand drug resistance interpretations for proviral DNA could be obtained by deep sequencing, which could provide more detailed and precise information for drug resistance monitoring and the rational design of optimal antiretroviral therapy regimens.

  18. Comparative linkage meta-analysis reveals regionally-distinct, disparate genetic architectures: application to bipolar disorder and schizophrenia.

    Directory of Open Access Journals (Sweden)

    Brady Tang

    Full Text Available New high-throughput, population-based methods and next-generation sequencing capabilities hold great promise in the quest for common and rare variant discovery and in the search for "missing heritability." However, the optimal analytic strategies for approaching such data are still actively debated, representing the latest rate-limiting step in genetic progress. Since it is likely a majority of common variants of modest effect have been identified through the application of tagSNP-based microarray platforms (i.e., GWAS, alternative approaches robust to detection of low-frequency (1-5% MAF and rare (<1% variants are of great importance. Of direct relevance, we have available an accumulated wealth of linkage data collected through traditional genetic methods over several decades, the full value of which has not been exhausted. To that end, we compare results from two different linkage meta-analysis methods--GSMA and MSP--applied to the same set of 13 bipolar disorder and 16 schizophrenia GWLS datasets. Interestingly, we find that the two methods implicate distinct, largely non-overlapping, genomic regions. Furthermore, based on the statistical methods themselves and our contextualization of these results within the larger genetic literatures, our findings suggest, for each disorder, distinct genetic architectures may reside within disparate genomic regions. Thus, comparative linkage meta-analysis (CLMA may be used to optimize low-frequency and rare variant discovery in the modern genomic era.

  19. Transgenic Mouse Lines Subdivide External Segment of the Globus Pallidus (GPe) Neurons and Reveal Distinct GPe Output Pathways

    Science.gov (United States)

    Mastro, Kevin J.; Bouchard, Rachel S.; Holt, Hiromi A. K.

    2014-01-01

    Cell-type diversity in the brain enables the assembly of complex neural circuits, whose organization and patterns of activity give rise to brain function. However, the identification of distinct neuronal populations within a given brain region is often complicated by a lack of objective criteria to distinguish one neuronal population from another. In the external segment of the globus pallidus (GPe), neuronal populations have been defined using molecular, anatomical, and electrophysiological criteria, but these classification schemes are often not generalizable across preparations and lack consistency even within the same preparation. Here, we present a novel use of existing transgenic mouse lines, Lim homeobox 6 (Lhx6)–Cre and parvalbumin (PV)–Cre, to define genetically distinct cell populations in the GPe that differ molecularly, anatomically, and electrophysiologically. Lhx6–GPe neurons, which do not express PV, are concentrated in the medial portion of the GPe. They have lower spontaneous firing rates, narrower dynamic ranges, and make stronger projections to the striatum and substantia nigra pars compacta compared with PV–GPe neurons. In contrast, PV–GPe neurons are more concentrated in the lateral portions of the GPe. They have narrower action potentials, deeper afterhyperpolarizations, and make stronger projections to the subthalamic nucleus and parafascicular nucleus of the thalamus. These electrophysiological and anatomical differences suggest that Lhx6–GPe and PV–GPe neurons participate in different circuits with the potential to contribute to different aspects of motor function and dysfunction in disease. PMID:24501350

  20. Environmental transcriptome analysis reveals physiological differences between biofilm and planktonic modes of life of the iron oxidizing bacteria Leptospirillum spp. in their natural microbial community

    Directory of Open Access Journals (Sweden)

    Parro Víctor

    2010-06-01

    Full Text Available Abstract Background Extreme acidic environments are characterized by their high metal content and lack of nutrients (oligotrophy. Macroscopic biofilms and filaments usually grow on the water-air interface or under the stream attached to solid substrates (streamers. In the Río Tinto (Spain, brown filaments develop under the water stream where the Gram-negative iron-oxidizing bacteria Leptospirillum spp. (L. ferrooxidans and L. ferriphilum and Acidithiobacillus ferrooxidans are abundant. These microorganisms play a critical role in bioleaching processes for industrial (biominery and environmental applications (acid mine drainage, bioremediation. The aim of this study was to investigate the physiological differences between the free living (planktonic and the sessile (biofilm associated lifestyles of Leptospirillum spp. as part of its natural extremely acidophilic community. Results Total RNA extracted from environmental samples was used to determine the composition of the metabolically active members of the microbial community and then to compare the biofilm and planktonic environmental transcriptomes by hybridizing to a genomic microarray of L. ferrooxidans. Genes up-regulated in the filamentous biofilm are involved in cellular functions related to biofilm formation and maintenance, such as: motility and quorum sensing (mqsR, cheAY, fliA, motAB, synthesis of cell wall structures (lnt, murA, murB, specific proteases (clpX/clpP, stress response chaperons (clpB, clpC, grpE-dnaKJ, groESL, etc. Additionally, genes involved in mixed acid fermentation (poxB, ackA were up-regulated in the biofilm. This result, together with the presence of small organic acids like acetate and formate (1.36 mM and 0.06 mM respectively in the acidic (pH 1.8 water stream, suggests that either L. ferrooxidans or other member of the microbial community are producing acetate in the acidophilic biofilm under microaerophilic conditions. Conclusions Our results indicate that the

  1. The immune system vs. Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Jensen, Peter Østrup; Givskov, Michael; Bjarnsholt, Thomas

    2010-01-01

    revealed both innate as well as adaptive immune responses to biofilms. On the other hand, measures launched by biofilm bacteria to achieve protection against the various immune responses have also been demonstrated. Whether particular immune responses to biofilm infections exist remains to be firmly...... established. However, because biofilm infections are often persistent (or chronic), an odd situation appears with the simultaneous activation of both arms of the host immune response, neither of which can eliminate the biofilm pathogen, but instead, in synergy, causes collateral tissue damage. Although...... the present review on the immune system vs. biofilm bacteria is focused on Pseudomonas aeruginosa (mainly because this is the most thoroughly studied), many of the same mechanisms are also seen with biofilm infections generated by other microorganisms....

  2. Molecular Basis for Saccharomyces cerevisiae Biofilm Development

    DEFF Research Database (Denmark)

    Andersen, Kaj Scherz

    In this study, I sought to identify genes regulating the global molecular program for development of sessile multicellular communities, also known as biofilm, of the eukaryotic microorganism, Saccharomyces cerevisiae (yeast). Yeast biofilm has a clinical interest, as biofilms can cause chronic...... infections in humans. Biofilm is also interesting from an evolutionary standpoint, as an example of primitive multicellularity. By using a genome-wide screen of yeast deletion mutants, I show that 71 genes are essential for biofilm formation. Two-thirds of these genes are required for transcription of FLO11......, but only a small subset is previously described as regulators of FLO11. These results reveal that the regulation of biofilm formation and FLO11 is even more complex than what has previously been described. I find that the molecular program for biofilm formation shares many essential components with two...

  3. Molecular Basis for Saccharomyces cerevisiae Biofilm Development

    DEFF Research Database (Denmark)

    Andersen, Kaj Scherz

    of translation of FLO11. In conclusion, I have conducted the first global study of the genetic program for yeast biofilm formation on polystyrene. This work provide several target genes as good basis for further research of biofilm, that I believe can contribute to fields such as cell biology, genetics, system......In this study, I sought to identify genes regulating the global molecular program for development of sessile multicellular communities, also known as biofilm, of the eukaryotic microorganism, Saccharomyces cerevisiae (yeast). Yeast biofilm has a clinical interest, as biofilms can cause chronic......, but only a small subset is previously described as regulators of FLO11. These results reveal that the regulation of biofilm formation and FLO11 is even more complex than what has previously been described. I find that the molecular program for biofilm formation shares many essential components with two...

  4. Analysis of the Complete Mycoplasma hominis LBD-4 Genome Sequence Reveals Strain-Variable Prophage Insertion and Distinctive Repeat-Containing Surface Protein Arrangements

    OpenAIRE

    2015-01-01

    The complete genome sequence of Mycoplasma hominis LBD-4 has been determined and the gene content ascribed. The 715,165-bp chromosome contains 620 genes, including 14 carried by a strain-variable prophage genome related to Mycoplasma fermentans MFV-1 and Mycoplasma arthritidis MAV-1. Comparative analysis with the genome of M. hominis PG21T reveals distinctive arrangements of repeat-containing surface proteins.

  5. Analysis of the Complete Mycoplasma hominis LBD-4 Genome Sequence Reveals Strain-Variable Prophage Insertion and Distinctive Repeat-Containing Surface Protein Arrangements.

    Science.gov (United States)

    Calcutt, Michael J; Foecking, Mark F

    2015-02-26

    The complete genome sequence of Mycoplasma hominis LBD-4 has been determined and the gene content ascribed. The 715,165-bp chromosome contains 620 genes, including 14 carried by a strain-variable prophage genome related to Mycoplasma fermentans MFV-1 and Mycoplasma arthritidis MAV-1. Comparative analysis with the genome of M. hominis PG21(T) reveals distinctive arrangements of repeat-containing surface proteins.

  6. OCT2, SSX and SAGE1 reveal the phenotypic heterogeneity of spermatocytic seminoma reflecting distinct subpopulations of spermatogonia

    Science.gov (United States)

    Lim, Jasmine; Goriely, Anne; Turner, Gareth DH; Ewen, Katherine A; Jacobsen, Grete Krag; Graem, Niels; Wilkie, Andrew OM; Rajpert-De Meyts, Ewa

    2011-01-01

    Spermatocytic seminoma (SS) is a rare testicular neoplasm that occurs predominantly in older men. In this study, we aimed to shed light on the histogenesis of SS by investigating the developmental expression of protein markers that identify distinct subpopulations of human spermatogonia in the normal adult testis. We analysed the expression pattern of OCT2, SSX2-4, and SAGE1 in 36 SS cases and four intratubular SS (ISS) as well as a series of normal testis samples throughout development. We describe for the first time two different types of SS characterized by OCT2 or SSX2-4 immunoexpression. These findings are consistent with the mutually exclusive antigenic profile of these markers during different stages of testicular development and in the normal adult testis. OCT2 was expressed predominantly in Adark spermatogonia, SSX2-4 was present in Apale and B spermatogonia and leptotene spermatocytes, whilst SAGE1 was exclusively present in a subset of post-pubertal germ cells, most likely B spermatogonia. The presence of OCT2 and SSX2-4 in distinct subsets of germ cells implies that these markers represent germ cells at different maturation stages. Analysis of SAGE1 and SSX2-4 in ISS showed spatial differences suggesting ongoing maturation of germ cells during progression of SS tumourigenesis. We conclude that the expression pattern of OCT2, SSX2-4, and SAGE1 supports the origin of SS from spermatogonia and provides new evidence for heterogeneity of this tumour, potentially linked either to the cellular origin of SS or to partial differentiation during tumour progression, including a hitherto unknown OCT2-positive variant of the tumour likely derived from Adark spermatogonia. Copyright © 2011 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. PMID:21706474

  7. K-shell Analysis Reveals Distinct Functional Parts in an Electron Transfer Network and Its Implications for Extracellular Electron Transfer

    Science.gov (United States)

    Ding, Dewu; Li, Ling; Shu, Chuanjun; Sun, Xiao

    2016-01-01

    Shewanella oneidensis MR-1 is capable of extracellular electron transfer (EET) and hence has attracted considerable attention. The EET pathways mainly consist of c-type cytochromes, along with some other proteins involved in electron transfer processes. By whole genome study and protein interactions inquisition, we constructed a large-scale electron transfer network containing 2276 interactions among 454 electron transfer related proteins in S. oneidensis MR-1. Using the k-shell decomposition method, we identified and analyzed distinct parts of the electron transfer network. We found that there was a negative correlation between the ks (k-shell values) and the average DR_100 (disordered regions per 100 amino acids) in every shell, which suggested that disordered regions of proteins played an important role during the formation and extension of the electron transfer network. Furthermore, proteins in the top three shells of the network are mainly located in the cytoplasm and inner membrane; these proteins can be responsible for transfer of electrons into the quinone pool in a wide variety of environmental conditions. In most of the other shells, proteins are broadly located throughout the five cellular compartments (cytoplasm, inner membrane, periplasm, outer membrane, and extracellular), which ensures the important EET ability of S. oneidensis MR-1. Specifically, the fourth shell was responsible for EET and the c-type cytochromes in the remaining shells of the electron transfer network were involved in aiding EET. Taken together, these results show that there are distinct functional parts in the electron transfer network of S. oneidensis MR-1, and the EET processes could achieve high efficiency through cooperation through such an electron transfer network. PMID:27148219

  8. Distinct immunological mechanisms of CTLA-4 and PD-1 blockade revealed by analyzing TCR usage in blood lymphocytes

    Science.gov (United States)

    Robert, Lidia; Harview, Christina; Emerson, Ryan; Wang, Xiaoyan; Mok, Stephen; Homet, Blanca; Comin-Anduix, Begonya; Koya, Richard C; Robins, Harlan; Tumeh, Paul C; Ribas, Antoni

    2014-01-01

    Targeting immune inhibitory receptors has brought excitement, innovation and hope to cancer patients. Our recent work revealed the immunological effects of blocking the CTLA4 and PD-1 immune checkpoints on T cell receptor usage among peripheral blood cells, and further uncovers how the expansion of the T cell repertoire matches the immunotoxicity profile of the therapy. PMID:25083336

  9. HD-GYP domain proteins regulate biofilm formation and virulence in Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Ryan, Robert P.; Lucey, Jean; O'Donovan, Karen

    2009-01-01

    2572 had a negative influence on swarming that was cryptic and was revealed only after removal of an uncharacterized C-terminal domain. Mutation of PA4108, PA4781 and PA2572 had distinct effects on biofilm formation and architecture of P. aeruginosa. All three proteins contributed to virulence of P...... residues (YN-GYP). Here we have investigated the role of these proteins in biofilm formation, virulence factor synthesis and virulence of P. aeruginosa. Mutation of PA4108 and PA4781 led to an increase in the level of cyclic-di-GMP in P. aeruginosa, consistent with the predicted activity of the encoded...

  10. Development of a simplified biofilm model

    Science.gov (United States)

    Sarkar, Sushovan; Mazumder, Debabrata

    2017-07-01

    A simplified approach for analyzing the biofilm process in deriving an easy model has been presented. This simplified biofilm model formulated correlations between substrate concentration in the influent/effluent and at biofilm-liquid interface along with substrate flux and biofilm thickness. The model essentially considered the external mass transport according to Fick's Law, steady state substrate as well as biomass balance for attached growth microorganisms. In substrate utilization, Monod growth kinetics has been followed incorporating relevant boundary conditions at the liquid-biofilm interface and at the attachment surface. The numerical solution of equations was accomplished using Runge-Kutta method and accordingly an integrated computer program was developed. The model has been successfully applied in a distinct set of trials with varying range of representative input variables. The model performance was compared with available existing methods and it was found an easy, accurate method that can be used for process design of biofilm reactor.

  11. Identification of Genes Controlled by the Essential YycFG Two-Component System Reveals a Role for Biofilm Modulation in Staphylococcus epidermidis.

    Science.gov (United States)

    Xu, Tao; Wu, Yang; Lin, Zhiwei; Bertram, Ralph; Götz, Friedrich; Zhang, Ying; Qu, Di

    2017-01-01

    Biofilms play a crucial role in the pathogenicity of Staphylococcus epidermidis, while little is known about whether the essential YycFG two-component signal transduction system (TCS) is involved in biofilm formation. We used antisense RNA (asRNA) to silence the yycFG TCS in order to study its regulatory functions in S. epidermidis. Strain 1457 expressing asRNA yycF exhibited a significant delay (~4-5 h) in entry to log phase, which was partially complemented by overexpressing ssaA. The expression of asRNA yycF and asRNA yycG resulted in a 68 and 50% decrease in biofilm formation at 6 h, respectively, while they had no significant inhibitory effect on 12 h biofilm formation. The expression of asRNA yycF led to a ~5-fold increase in polysaccharide intercellular adhesion (PIA) production, but it did not affect the expression of accumulation-associated protein (Aap) or the release of extracellular DNA. Consistently, quantitative real-time PCR showed that silencing yycF resulted in an increased transcription of biofilm-related genes, including icaA, arlR, sarA, sarX, and sbp. An in silico search of the YycF regulon for the conserved YycF recognition pattern and a modified motif in S. epidermidis, along with additional gel shift and DNase I footprinting assays, showed that arlR, sarA, sarX, and icaA are directly regulated by YycF. Our data suggests that YycFG modulates S. epidermidis biofilm formation in an ica-dependent manner.

  12. Tissue Doppler echocardiography reveals distinct patterns of impaired myocardial velocities in different degrees of coronary artery disease

    DEFF Research Database (Denmark)

    Hoffmann, Soren; Mogelvang, Rasmus; Olsen, Niels Thue

    2010-01-01

    Aim To determine how the left ventricular wall motion assessed by echocardiographic Tissue Doppler Imaging (TDI) is affected by increasing severity of coronary artery disease (CAD) among patients with stable angina pectoris and preserved ejection fraction. METHODS AND RESULTS: This study comprises.......86 +/- 0.24 vs. 1.00 +/- 0.28, P preserved ejection fraction reveals both diastolic and systolic dysfunction and the nature...

  13. Transcriptional profiling reveals developmental relationship and distinct biological functions of CD16+ and CD16- monocyte subsets

    Directory of Open Access Journals (Sweden)

    Zhou Xiaobo

    2009-08-01

    Full Text Available Abstract Background Human peripheral blood monocytes (Mo consist of subsets distinguished by expression of CD16 (FCγRIII and chemokine receptors. Classical CD16- Mo express CCR2 and migrate in response to CCL2, while a minor CD16+ Mo subset expresses CD16 and CX3CR1 and migrates into tissues expressing CX3CL1. CD16+ Mo produce pro-inflammatory cytokines and are expanded in certain inflammatory conditions including sepsis and HIV infection. Results To gain insight into the developmental relationship and functions of CD16+ and CD16- Mo, we examined transcriptional profiles of these Mo subsets in peripheral blood from healthy individuals. Of 16,328 expressed genes, 2,759 genes were differentially expressed and 228 and 250 were >2-fold upregulated and downregulated, respectively, in CD16+ compared to CD16- Mo. CD16+ Mo were distinguished by upregulation of transcripts for dendritic cell (DC (SIGLEC10, CD43, RARA and macrophage (MΦ (CSF1R/CD115, MafB, CD97, C3aR markers together with transcripts relevant for DC-T cell interaction (CXCL16, ICAM-2, LFA-1, cell activation (LTB, TNFRSF8, LST1, IFITM1-3, HMOX1, SOD-1, WARS, MGLL, and negative regulation of the cell cycle (CDKN1C, MTSS1, whereas CD16- Mo were distinguished by upregulation of transcripts for myeloid (CD14, MNDA, TREM1, CD1d, C1qR/CD93 and granulocyte markers (FPR1, GCSFR/CD114, S100A8-9/12. Differential expression of CSF1R, CSF3R, C1QR1, C3AR1, CD1d, CD43, CXCL16, and CX3CR1 was confirmed by flow cytometry. Furthermore, increased expression of RARA and KLF2 transcripts in CD16+ Mo coincided with absence of cell surface cutaneous lymphocyte associated antigen (CLA expression, indicating potential imprinting for non-skin homing. Conclusion These results suggest that CD16+ and CD16- Mo originate from a common myeloid precursor, with CD16+ Mo having a more MΦ – and DC-like transcription program suggesting a more advanced stage of differentiation. Distinct transcriptional programs, together

  14. Structure of N-Terminal Domain of NPC1 Reveals Distinct Subdomains for Binding and Transfer of Cholesterol

    Energy Technology Data Exchange (ETDEWEB)

    Kwon, Hyock Joo; Abi-Mosleh, Lina; Wang, Michael L.; Deisenhofer, Johann; Goldstein, Joseph L.; Brown, Michael S.; Infante, Rodney E.; (UTSMC)

    2010-09-21

    LDL delivers cholesterol to lysosomes by receptor-mediated endocytosis. Exit of cholesterol from lysosomes requires two proteins, membrane-bound Niemann-Pick C1 (NPC1) and soluble NPC2. NPC2 binds cholesterol with its isooctyl side chain buried and its 3{beta}-hydroxyl exposed. Here, we describe high-resolution structures of the N-terminal domain (NTD) of NPC1 and complexes with cholesterol and 25-hydroxycholesterol. NPC1(NTD) binds cholesterol in an orientation opposite to NPC2: 3{beta}-hydroxyl buried and isooctyl side chain exposed. Cholesterol transfer from NPC2 to NPC1(NTD) requires reorientation of a helical subdomain in NPC1(NTD), enlarging the opening for cholesterol entry. NPC1 with point mutations in this subdomain (distinct from the binding subdomain) cannot accept cholesterol from NPC2 and cannot restore cholesterol exit from lysosomes in NPC1-deficient cells. We propose a working model wherein after lysosomal hydrolysis of LDL-cholesteryl esters, cholesterol binds NPC2, which transfers it to NPC1(NTD), reversing its orientation and allowing insertion of its isooctyl side chain into the outer lysosomal membranes.

  15. Meta-analysis of several gene lists for distinct types of cancer: A simple way to reveal common prognostic markers

    Directory of Open Access Journals (Sweden)

    Sun Xiao

    2007-04-01

    Full Text Available Abstract Background Although prognostic biomarkers specific for particular cancers have been discovered, microarray analysis of gene expression profiles, supported by integrative analysis algorithms, helps to identify common factors in molecular oncology. Similarities of Ordered Gene Lists (SOGL is a recently proposed approach to meta-analysis suitable for identifying features shared by two data sets. Here we extend the idea of SOGL to the detection of significant prognostic marker genes from microarrays of multiple data sets. Three data sets for leukemia and the other six for different solid tumors are used to demonstrate our method, using established statistical techniques. Results We describe a set of significantly similar ordered gene lists, representing outcome comparisons for distinct types of cancer. This kind of similarity could improve the diagnostic accuracies of individual studies when SOGL is incorporated into the support vector machine algorithm. In particular, we investigate the similarities among three ordered gene lists pertaining to mesothelioma survival, prostate recurrence and glioma survival. The similarity-driving genes are related to the outcomes of patients with lung cancer with a hazard ratio of 4.47 (p = 0.035. Many of these genes are involved in breakdown of EMC proteins regulating angiogenesis, and may be used for further research on prognostic markers and molecular targets of gene therapy for cancers. Conclusion The proposed method and its application show the potential of such meta-analyses in clinical studies of gene expression profiles.

  16. Resting state functional magnetic resonance imaging reveals distinct brain activity in heavy cannabis users - a multi-voxel pattern analysis.

    Science.gov (United States)

    Cheng, H; Skosnik, P D; Pruce, B J; Brumbaugh, M S; Vollmer, J M; Fridberg, D J; O'Donnell, B F; Hetrick, W P; Newman, S D

    2014-11-01

    Chronic cannabis use can cause cognitive, perceptual and personality alterations, which are believed to be associated with regional brain changes and possible changes in connectivity between functional regions. This study aims to identify the changes from resting state functional magnetic resonance imaging scans. A two-level multi-voxel pattern analysis was proposed to classify male cannabis users from normal controls. The first level analysis works on a voxel basis and identifies clusters for the input of a second level analysis, which works on the functional connectivity between these regions. We found distinct clusters for male cannabis users in the middle frontal gyrus, precentral gyrus, superior frontal gyrus, posterior cingulate cortex, cerebellum and some other regions. Based on the functional connectivity of these clusters, a high overall accuracy rate of 84-88% in classification accuracy was achieved. High correlations were also found between the overall classification accuracy and Barrett Barrett Impulsiveness Scale factor scores of attention and motor. Our result suggests regional differences in the brains of male cannabis users that span from the cerebellum to the prefrontal cortex, which are associated with differences in functional connectivity. © The Author(s) 2014.

  17. Structural comparison of the Caenorhabditis elegans and human Ndc80 complexes bound to microtubules reveals distinct binding behavior

    Science.gov (United States)

    Wilson-Kubalek, Elizabeth M.; Cheeseman, Iain M.; Milligan, Ronald A.

    2016-01-01

    During cell division, kinetochores must remain tethered to the plus ends of dynamic microtubule polymers. However, the molecular basis for robust kinetochore–microtubule interactions remains poorly understood. The conserved four-subunit Ndc80 complex plays an essential and direct role in generating dynamic kinetochore–microtubule attachments. Here we compare the binding of the Caenorhabditis elegans and human Ndc80 complexes to microtubules at high resolution using cryo–electron microscopy reconstructions. Despite the conserved roles of the Ndc80 complex in diverse organisms, we find that the attachment mode of these complexes for microtubules is distinct. The human Ndc80 complex binds every tubulin monomer along the microtubule protofilament, whereas the C. elegans Ndc80 complex binds more tightly to β-tubulin. In addition, the C. elegans Ndc80 complex tilts more toward the adjacent protofilament. These structural differences in the Ndc80 complex between different species may play significant roles in the nature of kinetochore–microtubule interactions. PMID:26941333

  18. OCT2, SSX and SAGE1 reveal the phenotypic heterogeneity of spermatocytic seminoma reflecting distinct subpopulations of spermatogonia

    DEFF Research Database (Denmark)

    Lim, Jasmine; Goriely, Anne; Turner, Gareth Dh;

    2011-01-01

    in the normal adult testis. We analysed the expression pattern of OCT2, SSX2-4, and SAGE1 in 36 SS cases and four intratubular SS (ISS) as well as a series of normal testis samples throughout development. We describe for the first time two different types of SS characterized by OCT2 or SSX2-4 immunoexpression......, whilst SAGE1 was exclusively present in a subset of post-pubertal germ cells, most likely B spermatogonia. The presence of OCT2 and SSX2-4 in distinct subsets of germ cells implies that these markers represent germ cells at different maturation stages. Analysis of SAGE1 and SSX2-4 in ISS showed spatial...... differences suggesting ongoing maturation of germ cells during progression of SS tumourigenesis. We conclude that the expression pattern of OCT2, SSX2-4, and SAGE1 supports the origin of SS from spermatogonia and provides new evidence for heterogeneity of this tumour, potentially linked either to the cellular...

  19. Cells Respond to Distinct Nanoparticle Properties with Multiple Strategies As Revealed by Single-Cell RNA-Seq

    Energy Technology Data Exchange (ETDEWEB)

    Mitchell, Hugh D.; Markillie, Lye Meng; Chrisler, William B.; Gaffrey, Matthew J.; Hu, Dehong; Szymanski, Craig J.; Xie, Yumei; Melby, Eric S.; Dohnalkova, Alice; Taylor, Ronald C.; Grate, Eva K.; Cooley, Scott K.; McDermott, Jason E.; Heredia-Langner, Alejandro; Orr, Galya

    2016-11-22

    The impact of distinct nanoparticle (NP) properties on cellular response and ultimately human health is unclear. This gap is partially due to experimental difficulties in achieving uniform NP loads in the studied cells, creating heterogeneous populations with some cells “overloaded” while other cells are loaded with few or no NPs. Yet gene expression studies have been conducted in the population as a whole, identifying generic responses, while missing unique responses due to signal averaging across many cells, each carrying different loads. Here we applied single-cell RNA-Seq to alveolar epithelial cells carrying defined loads of aminated or carboxylated quantum dots (QDs), showing higher or lower toxicity, respectively. Interestingly, cells carrying lower loads responded with multiple strategies, mostly with upregulated processes, which were nonetheless coherent and unique to each QD type. In contrast, cells carrying higher loads responded more uniformly, with mostly downregulated processes that were shared across QD types. Strategies unique to aminated QDs showed strong upregulation of stress responses, coupled in some cases with regulation of cell cycle, protein synthesis and organelle activities. In contrast, strategies unique to carboxylated QDs showed upregulation of DNA repair and RNA activities, and decreased regulation of cell division, coupled in some cases with upregulation of stress responses and ATP related functions. Together, our studies suggest scenarios where higher NP loads lock cells into uniform responses, mostly shutdown of cellular processes, whereas lower loads allow for unique responses to each NP type that are more diversified, proactive defenses or repairs of the NP insults.

  20. Analysis of grain characters in temperate grasses reveals distinctive patterns of endosperm organization associated with grain shape

    Science.gov (United States)

    Drea, Sinéad

    2012-01-01

    Members of the core pooids represent the most important crops in temperate zones including wheat, barley, and oats. Their importance as crops is largely due to the grain, particularly the storage capabilities of the endosperm. In this study, a comprehensive survey of grain morphology and endosperm organization in representatives of wild and cultivated species throughout the core pooids was performed. As sister to the core pooid tribes Poeae, Aveneae, Triticeae, and Bromeae within the Pooideae subfamily, Brachypodium provides a taxonomically relevant reference point. Using macroscopic, histological, and molecular analyses distinct patterns of grain tissue organization in these species, focusing on the peripheral and modified aleurone, are described. The results indicate that aleurone organization is correlated with conventional grain quality characters such as grain shape and starch content. In addition to morphological and organizational variation, expression patterns of candidate gene markers underpinning this variation were examined. Features commonly associated with grains are largely defined by analyses on lineages within the Triticeae and knowledge of grain structure may be skewed as a result of the focus on wheat and barley. Specifically, the data suggest that the modified aleurone is largely restricted to species in the Triticeae tribe. PMID:23081982

  1. Multi-Omics of Tomato Glandular Trichomes Reveals Distinct Features of Central Carbon Metabolism Supporting High Productivity of Specialized Metabolites.

    Science.gov (United States)

    Balcke, Gerd U; Bennewitz, Stefan; Bergau, Nick; Athmer, Benedikt; Henning, Anja; Majovsky, Petra; Jiménez-Gómez, José M; Hoehenwarter, Wolfgang; Tissier, Alain

    2017-05-01

    Glandular trichomes are metabolic cell factories with the capacity to produce large quantities of secondary metabolites. Little is known about the connection between central carbon metabolism and metabolic productivity for secondary metabolites in glandular trichomes. To address this gap in our knowledge, we performed comparative metabolomics, transcriptomics, proteomics, and (13)C-labeling of type VI glandular trichomes and leaves from a cultivated (Solanum lycopersicum LA4024) and a wild (Solanum habrochaites LA1777) tomato accession. Specific features of glandular trichomes that drive the formation of secondary metabolites could be identified. Tomato type VI trichomes are photosynthetic but acquire their carbon essentially from leaf sucrose. The energy and reducing power from photosynthesis are used to support the biosynthesis of secondary metabolites, while the comparatively reduced Calvin-Benson-Bassham cycle activity may be involved in recycling metabolic CO2 Glandular trichomes cope with oxidative stress by producing high levels of polyunsaturated fatty acids, oxylipins, and glutathione. Finally, distinct mechanisms are present in glandular trichomes to increase the supply of precursors for the isoprenoid pathways. Particularly, the citrate-malate shuttle supplies cytosolic acetyl-CoA and plastidic glycolysis and malic enzyme support the formation of plastidic pyruvate. A model is proposed on how glandular trichomes achieve high metabolic productivity. © 2017 American Society of Plant Biologists. All rights reserved.

  2. Distinct signal transduction pathways downstream of the (PRR revealed by microarray and ChIP-chip analyses.

    Directory of Open Access Journals (Sweden)

    Daniela Zaade

    Full Text Available The (prorenin receptor ((PRR signaling is involved in different pathophysiologies ranging from cardiorenal end-organ damage via diabetic retinopathy to tumorigenesis. We have previously shown that the transcription factor promyelocytic leukemia zinc finger (PLZF is an adaptor protein of the (PRR. Furthermore, recent publications suggest that major functions of the (PRR are mediated ligand-independently by its transmembrane and intracellular part, which acts as an accessory protein of V-ATPases. The transcriptome and recruitmentome downstream of the V-ATPase function and PLZF in the context of the (PRR are currently unknown. Therefore, we performed a set of microarray and chromatin-immunoprecipitation (ChIP-chip experiments using siRNA against the (PRR, stable overexpression of PLZF, the PLZF translocation inhibitor genistein and the specific V-ATPase inhibitor bafilomycin to dissect transcriptional pathways downstream of the (PRR. We were able to identify distinct and overlapping genetic signatures as well as novel real-time PCR-validated target genes of the different molecular functions of the (PRR. Moreover, bioinformatic analyses of our data confirm the role of (PRŔs signal transduction pathways in cardiovascular disease and tumorigenesis.

  3. De novo Transcriptome Analysis Reveals Distinct Defense Mechanisms by Young and Mature Leaves of Hevea brasiliensis (Para Rubber Tree).

    Science.gov (United States)

    Fang, Yongjun; Mei, Hailiang; Zhou, Binhui; Xiao, Xiaohu; Yang, Meng; Huang, Yacheng; Long, Xiangyu; Hu, Songnian; Tang, Chaorong

    2016-09-13

    Along with changes in morphology in the course of maturation, leaves of Hevea brasiliensis become more resistant to leaf diseases, including the South American Leaf Blight (SALB), a devastating fungal disease of this economically important tree species. To understand the underlying mechanisms of this defense, and to identify the candidate genes involved, we sequenced the Hevea leaf transcriptome at four developmental stages (I to IV) by Illumina sequencing. A total of 62.6 million high-quality reads were generated, and assembled into 98,796 unique transcripts. We identified 3,905 differentially expressed genes implicated in leaf development, 67.8% (2,651) of which were during the transition to leaf maturation. The genes involved in cyanogenic metabolism, lignin and anthocyanin biosynthesis were noteworthy for their distinct patterns of expression between developing leaves (stages I to III) and mature leaves (stage IV), and the correlation with the change in resistance to SALB and the Oidium/Colletotrichum leaf fall. The results provide a first profile of the molecular events that relate to the dynamics of leaf morphology and defense strategies during Hevea leaf development. This dataset is beneficial to devising strategies to engineer resistance to leaf diseases as well as other in-depth studies in Hevea tree.

  4. Genome-wide location analysis reveals distinct transcriptional circuitry by paralogous regulators Foxa1 and Foxa2.

    Science.gov (United States)

    Bochkis, Irina M; Schug, Jonathan; Ye, Diana Z; Kurinna, Svitlana; Stratton, Sabrina A; Barton, Michelle C; Kaestner, Klaus H

    2012-01-01

    Gene duplication is a powerful driver of evolution. Newly duplicated genes acquire new roles that are relevant to fitness, or they will be lost over time. A potential path to functional relevance is mutation of the coding sequence leading to the acquisition of novel biochemical properties, as analyzed here for the highly homologous paralogs Foxa1 and Foxa2 transcriptional regulators. We determine by genome-wide location analysis (ChIP-Seq) that, although Foxa1 and Foxa2 share a large fraction of binding sites in the liver, each protein also occupies distinct regulatory elements in vivo. Foxa1-only sites are enriched for p53 binding sites and are frequently found near genes important to cell cycle regulation, while Foxa2-restricted sites show only a limited match to the forkhead consensus and are found in genes involved in steroid and lipid metabolism. Thus, Foxa1 and Foxa2, while redundant during development, have evolved divergent roles in the adult liver, ensuring the maintenance of both genes during evolution.

  5. Genome-wide location analysis reveals distinct transcriptional circuitry by paralogous regulators Foxa1 and Foxa2.

    Directory of Open Access Journals (Sweden)

    Irina M Bochkis

    Full Text Available Gene duplication is a powerful driver of evolution. Newly duplicated genes acquire new roles that are relevant to fitness, or they will be lost over time. A potential path to functional relevance is mutation of the coding sequence leading to the acquisition of novel biochemical properties, as analyzed here for the highly homologous paralogs Foxa1 and Foxa2 transcriptional regulators. We determine by genome-wide location analysis (ChIP-Seq that, although Foxa1 and Foxa2 share a large fraction of binding sites in the liver, each protein also occupies distinct regulatory elements in vivo. Foxa1-only sites are enriched for p53 binding sites and are frequently found near genes important to cell cycle regulation, while Foxa2-restricted sites show only a limited match to the forkhead consensus and are found in genes involved in steroid and lipid metabolism. Thus, Foxa1 and Foxa2, while redundant during development, have evolved divergent roles in the adult liver, ensuring the maintenance of both genes during evolution.

  6. Genome-wide analysis of immune activation in human T and B cells reveals distinct classes of alternatively spliced genes.

    Directory of Open Access Journals (Sweden)

    Yevgeniy A Grigoryev

    Full Text Available Alternative splicing of pre-mRNA is a mechanism that increases the protein diversity of a single gene by differential exon inclusion/exclusion during post-transcriptional processing. While alternative splicing is established to occur during lymphocyte activation, little is known about the role it plays during the immune response. Our study is among the first reports of a systematic genome-wide analysis of activated human T and B lymphocytes using whole exon DNA microarrays integrating alternative splicing and differential gene expression. Purified human CD2(+ T or CD19(+ B cells were activated using protocols to model the early events in post-transplant allograft immunity and sampled as a function of time during the process of immune activation. Here we show that 3 distinct classes of alternatively spliced and/or differentially expressed genes change in an ordered manner as a function of immune activation. We mapped our results to function-based canonical pathways and demonstrated that some are populated by only one class of genes, like integrin signaling, while other pathways, such as purine metabolism and T cell receptor signaling, are populated by all three classes of genes. Our studies augment the current view of T and B cell activation in immunity that has been based exclusively upon differential gene expression by providing evidence for a large number of molecular networks populated as a function of time and activation by alternatively spliced genes, many of which are constitutively expressed.

  7. Mining the mouse transcriptome of receptive endometrium reveals distinct molecular signatures for the luminal and glandular epithelium.

    Science.gov (United States)

    Niklaus, Andrea L; Pollard, Jeffrey W

    2006-07-01

    Epithelia coat most tissues where they sense and respond to the environment and participate in innate immune responses. In the adult mouse uterus, columnar epithelium lines the central lumen and the glands that penetrate the underlying stroma. A nidatory surge of estrogen causes differentiation of the luminal epithelium to the receptive state that permits blastocyst attachment and allows subsequent implantation. Here, using laser-capture microdissection to isolate the luminal and glandular epithelia separately, we have profiled gene expression 2 h before embryo attachment to determine whether there are unique roles for these two epithelial structures in this process. Although most genes were expressed in both compartments, there was greater expression of 153 and 118 genes in the lumen and glands, respectively. In the luminal epithelium, there is enrichment in lipid, metal-ion binding, and carbohydrate-metabolizing enzymes, whereas in the glands, immune response genes are emphasized. In situ hybridization to uterine sections obtained from mice during the preimplantation period validated these data and indicated an array of previously undocumented genes expressed with unique patterns in these epithelia. The data show that each epithelial compartment has a distinct molecular signature and that they act differentially and synergistically to permit blastocyst implantation.

  8. Wound biofilms: lessons learned from oral biofilms.

    Science.gov (United States)

    Mancl, Kimberly A; Kirsner, Robert S; Ajdic, Dragana

    2013-01-01

    Biofilms play an important role in the development and pathogenesis of many chronic infections. Oral biofilms, more commonly known as dental plaque, are a primary cause of oral diseases including caries, gingivitis, and periodontitis. Oral biofilms are commonly studied as model biofilm systems as they are easily accessible; thus, biofilm research in oral diseases is advanced with details of biofilm formation and bacterial interactions being well elucidated. In contrast, wound research has relatively recently directed attention to the role biofilms have in chronic wounds. This review discusses the biofilms in periodontal disease and chronic wounds with comparisons focusing on biofilm detection, biofilm formation, the immune response to biofilms, bacterial interaction, and quorum sensing. Current treatment modalities used by both fields and future therapies are also discussed.

  9. Ultrastructural and molecular distinctions between the porcine inner cell mass and epiblast reveal unique pluripotent cell states

    DEFF Research Database (Denmark)

    Hall, V. J.; Jacobsen, Janus Valentin; Rasmussen, M. A.

    2010-01-01

    pluripotent cell population as it develops from the ICM to the late epiblast. The ultrastructural observations revealed that the outer cells of the ICM have a high nuclear:cytoplasmic ratio but are transcriptionally inactive and contain mitochondria with few cristae. In contrast, the epiblast cells have......Characterization of the pluripotent cell populations within the porcine embryo is essential for understanding pluripotency and self-renewal regulation in the inner cell mass (ICM) and epiblast. In this study, we perform detailed ultrastructural and molecular characterization of the developing...

  10. Curli fimbriae are conditionally required in Escherichia coli O157:H7 for initial attachment and biofilm formation.

    Science.gov (United States)

    Carter, Michelle Qiu; Louie, Jacqueline W; Feng, Doris; Zhong, Wayne; Brandl, Maria T

    2016-08-01

    Several species of enteric pathogens produce curli fimbriae, which may affect their interaction with surfaces and other microbes in nonhost environments. Here we used two Escherichia coli O157:H7 outbreak strains with distinct genotypes to understand the role of curli in surface attachment and biofilm formation in several systems relevant to fresh produce production and processing. Curli significantly enhanced the initial attachment of E. coli O157:H7 to spinach leaves and stainless steel surfaces by 5-fold. Curli was also required for E. coli O157:H7 biofilm formation on stainless steel and enhanced biofilm production on glass by 19-27 fold in LB no-salt broth. However, this contribution was not observed when cells were grown in sterile spinach lysates. Furthermore, both strains of E. coli O157:H7 produced minimal biofilms on polypropylene in LB no-salt broth but considerable amounts in spinach lysates. Under the latter conditions, curli appeared to slightly increase biofilm production. Importantly, curli played an essential role in the formation of mixed biofilm by E. coli O157:H7 and plant-associated microorganisms in spinach leaf washes, as revealed by confocal microscopy. Little or no E. coli O157:H7 biofilms were detected at 4 °C, supporting the importance of temperature control in postharvest and produce processing environments.

  11. Multiple analytical approaches reveal distinct gene-environment interactions in smokers and non smokers in lung cancer.

    Directory of Open Access Journals (Sweden)

    Rakhshan Ihsan

    Full Text Available Complex disease such as cancer results from interactions of multiple genetic and environmental factors. Studying these factors singularly cannot explain the underlying pathogenetic mechanism of the disease. Multi-analytical approach, including logistic regression (LR, classification and regression tree (CART and multifactor dimensionality reduction (MDR, was applied in 188 lung cancer cases and 290 controls to explore high order interactions among xenobiotic metabolizing genes and environmental risk factors. Smoking was identified as the predominant risk factor by all three analytical approaches. Individually, CYP1A1*2A polymorphism was significantly associated with increased lung cancer risk (OR = 1.69;95%CI = 1.11-2.59,p = 0.01, whereas EPHX1 Tyr113His and SULT1A1 Arg213His conferred reduced risk (OR = 0.40;95%CI = 0.25-0.65,p<0.001 and OR = 0.51;95%CI = 0.33-0.78,p = 0.002 respectively. In smokers, EPHX1 Tyr113His and SULT1A1 Arg213His polymorphisms reduced the risk of lung cancer, whereas CYP1A1*2A, CYP1A1*2C and GSTP1 Ile105Val imparted increased risk in non-smokers only. While exploring non-linear interactions through CART analysis, smokers carrying the combination of EPHX1 113TC (Tyr/His, SULT1A1 213GG (Arg/Arg or AA (His/His and GSTM1 null genotypes showed the highest risk for lung cancer (OR = 3.73;95%CI = 1.33-10.55,p = 0.006, whereas combined effect of CYP1A1*2A 6235CC or TC, SULT1A1 213GG (Arg/Arg and betel quid chewing showed maximum risk in non-smokers (OR = 2.93;95%CI = 1.15-7.51,p = 0.01. MDR analysis identified two distinct predictor models for the risk of lung cancer in smokers (tobacco chewing, EPHX1 Tyr113His, and SULT1A1 Arg213His and non-smokers (CYP1A1*2A, GSTP1 Ile105Val and SULT1A1 Arg213His with testing balance accuracy (TBA of 0.6436 and 0.6677 respectively. Interaction entropy interpretations of MDR results showed non-additive interactions of tobacco chewing with

  12. Genomic and phenotypic characterization of myxoma virus from Great Britain reveals multiple evolutionary pathways distinct from those in Australia

    Science.gov (United States)

    Kerr, Peter J.; Cattadori, Isabella M.; Fitch, Adam; Geber, Adam; Liu, June; Sim, Derek G.; Boag, Brian; Ghedin, Elodie

    2017-01-01

    The co-evolution of myxoma virus (MYXV) and the European rabbit occurred independently in Australia and Europe from different progenitor viruses. Although this is the canonical study of the evolution of virulence, whether the genomic and phenotypic outcomes of MYXV evolution in Europe mirror those observed in Australia is unknown. We addressed this question using viruses isolated in the United Kingdom early in the MYXV epizootic (1954–1955) and between 2008–2013. The later UK viruses fell into three distinct lineages indicative of a long period of separation and independent evolution. Although rates of evolutionary change were almost identical to those previously described for MYXV in Australia and strongly clock-like, genome evolution in the UK and Australia showed little convergence. The phenotypes of eight UK viruses from three lineages were characterized in laboratory rabbits and compared to the progenitor (release) Lausanne strain. Inferred virulence ranged from highly virulent (grade 1) to highly attenuated (grade 5). Two broad disease types were seen: cutaneous nodular myxomatosis characterized by multiple raised secondary cutaneous lesions, or an amyxomatous phenotype with few or no secondary lesions. A novel clinical outcome was acute death with pulmonary oedema and haemorrhage, often associated with bacteria in many tissues but an absence of inflammatory cells. Notably, reading frame disruptions in genes defined as essential for virulence in the progenitor Lausanne strain were compatible with the acquisition of high virulence. Combined, these data support a model of ongoing host-pathogen co-evolution in which multiple genetic pathways can produce successful outcomes in the field that involve both different virulence grades and disease phenotypes, with alterations in tissue tropism and disease mechanisms. PMID:28253375

  13. Distinct Cellular Assembly Stoichiometry of Polycomb Complexes on Chromatin Revealed by Single-molecule Chromatin Immunoprecipitation Imaging*♦

    Science.gov (United States)

    Tatavosian, Roubina; Zhen, Chao Yu; Duc, Huy Nguyen; Balas, Maggie M.; Johnson, Aaron M.; Ren, Xiaojun

    2015-01-01

    Epigenetic complexes play an essential role in regulating chromatin structure, but information about their assembly stoichiometry on chromatin within cells is poorly understood. The cellular assembly stoichiometry is critical for appreciating the initiation, propagation, and maintenance of epigenetic inheritance during normal development and in cancer. By combining genetic engineering, chromatin biochemistry, and single-molecule fluorescence imaging, we developed a novel and sensitive approach termed single-molecule chromatin immunoprecipitation imaging (Sm-ChIPi) to enable investigation of the cellular assembly stoichiometry of epigenetic complexes on chromatin. Sm-ChIPi was validated by using chromatin complexes with known stoichiometry. The stoichiometry of subunits within a polycomb complex and the assembly stoichiometry of polycomb complexes on chromatin have been extensively studied but reached divergent views. Moreover, the cellular assembly stoichiometry of polycomb complexes on chromatin remains unexplored. Using Sm-ChIPi, we demonstrated that within mouse embryonic stem cells, one polycomb repressive complex (PRC) 1 associates with multiple nucleosomes, whereas two PRC2s can bind to a single nucleosome. Furthermore, we obtained direct physical evidence that the nucleoplasmic PRC1 is monomeric, whereas PRC2 can dimerize in the nucleoplasm. We showed that ES cell differentiation induces selective alteration of the assembly stoichiometry of Cbx2 on chromatin but not other PRC1 components. We additionally showed that the PRC2-mediated trimethylation of H3K27 is not required for the assembly stoichiometry of PRC1 on chromatin. Thus, these findings uncover that PRC1 and PRC2 employ distinct mechanisms to assemble on chromatin, and the novel Sm-ChIPi technique could provide single-molecule insight into other epigenetic complexes. PMID:26381410

  14. Large-scale atlas of microarray data reveals the distinct expression landscape of different tissues in Arabidopsis.

    Science.gov (United States)

    He, Fei; Yoo, Shinjae; Wang, Daifeng; Kumari, Sunita; Gerstein, Mark; Ware, Doreen; Maslov, Sergei

    2016-06-01

    Transcriptome data sets from thousands of samples of the model plant Arabidopsis thaliana have been collectively generated by multiple individual labs. Although integration and meta-analysis of these samples has become routine in the plant research community, it is often hampered by a lack of metadata or differences in annotation styles of different labs. In this study, we carefully selected and integrated 6057 Arabidopsis microarray expression samples from 304 experiments deposited to the Gene Expression Omnibus (GEO) at the National Center for Biotechnology Information (NCBI). Metadata such as tissue type, growth conditions and developmental stage were manually curated for each sample. We then studied the global expression landscape of the integrated data set and found that samples of the same tissue tend to be more similar to each other than to samples of other tissues, even in different growth conditions or developmental stages. Root has the most distinct transcriptome, compared with aerial tissues, but the transcriptome of cultured root is more similar to the transcriptome of aerial tissues, as the cultured root samples lost their cellular identity. Using a simple computational classification method, we showed that the tissue type of a sample can be successfully predicted based on its expression profile, opening the door for automatic metadata extraction and facilitating the re-use of plant transcriptome data. As a proof of principle, we applied our automated annotation pipeline to 708 RNA-seq samples from public repositories and verified the accuracy of our predictions with sample metadata provided by the authors. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

  15. Immunogold electron microscopy and confocal analyses reveal distinctive patterns of histone H3 phosphorylation during mitosis in MCF-7 cells.

    Science.gov (United States)

    Yan, Yitang; Cummings, Connie A; Sutton, Deloris; Yu, Linda; Castro, Lysandra; Moore, Alicia B; Gao, Xiaohua; Dixon, Darlene

    2016-04-01

    Histone phosphorylation has a profound impact on epigenetic regulation of gene expression, chromosome condensation and segregation, and maintenance of genome integrity. Histone H3 Serine 10 is evolutionally conserved and heavily phosphorylated during mitosis. To examine Histone H3 Serine 10 phosphorylation (H3S10ph) dynamics in mitosis, we applied immunogold labeling and confocal microscopy to visualize H3S10ph expression in MCF-7 cells. Confocal observations showed that MCF-7 cells had abundant H3S10ph expression in prophase and metaphase. In anaphase, the H3S10ph expression was significantly decreased and displayed only sparsely localized staining that mainly associated with the chromatid tips. We showed that immunogold bead density distribution followed the H3S10ph expression patterns observed in confocal analysis. At a higher magnification in metaphase, the immunogold beads were readily visible and the bead distribution along the condensed chromosomes was distinctive, indicating the specificity and reliability of the immunogold staining procedure. In anaphase, the beads were found to distribute focally in specific regions of chromatids, reinforcing the confocal observations of differential H3 phosphorylation. To our knowledge, this is the first report to show the specific H3S10ph expression with an immunogold technique and transmission electron microscopy. Additionally, with confocal microscopy, we analyzed H3S10ph expression in an immortalized cell line derived from benign uterine smooth muscle tumor cells. H3S10ph epitope was expressed more abundantly during anaphase in the benign tumor cells, and there was no dramatic differential expression within the condensed chromatid clusters as observed in MCF-7 cells. The differences in H3S10ph expression pattern and dynamics may contribute to the differential proliferative potential between benign tumor cells and MCF-7 cells.

  16. Genome sequencing reveals diversification of virulence factor content and possible host adaptation in distinct subpopulations of Salmonella enterica

    Directory of Open Access Journals (Sweden)

    Rodriguez-Rivera Lorraine D

    2011-08-01

    Full Text Available Abstract Background Divergence of bacterial populations into distinct subpopulations is often the result of ecological isolation. While some studies have suggested the existence of Salmonella enterica subsp. enterica subclades, evidence for these subdivisions has been ambiguous. Here we used a comparative genomics approach to define the population structure of Salmonella enterica subsp. enterica, and identify clade-specific genes that may be the result of ecological specialization. Results Multi-locus sequence analysis (MLSA and single nucleotide polymorphisms (SNPs data for 16 newly sequenced and 30 publicly available genomes showed an unambiguous subdivision of S. enterica subsp. enterica into at least two subpopulations, which we refer to as clade A and clade B. Clade B strains contain several clade-specific genes or operons, including a β-glucuronidase operon, a S-fimbrial operon, and cell surface related genes, which strongly suggests niche specialization of this subpopulation. An additional set of 123 isolates was assigned to clades A and B by using qPCR assays targeting subpopulation-specific SNPs and genes of interest. Among 98 serovars examined, approximately 20% belonged to clade B. All clade B isolates contained two pathogenicity related genomic islands, SPI-18 and a cytolethal distending toxin islet; a combination of these two islands was previously thought to be exclusive to serovars Typhi and Paratyphi A. Presence of β-glucuronidase in clade B isolates specifically suggests an adaptation of this clade to the vertebrate gastrointestinal environment. Conclusions S. enterica subsp. enterica consists of at least two subpopulations that differ specifically in genes involved in host and tissue tropism, utilization of host specific carbon and nitrogen sources and are therefore likely to differ in ecology and transmission characteristics.

  17. Morphological and behavioral differences in the gastropod Trophon geversianus associated to distinct environmental conditions, as revealed by a multidisciplinary approach

    Science.gov (United States)

    Márquez, Federico; Nieto Vilela, Rocío Aimé; Lozada, Mariana; Bigatti, Gregorio

    2015-01-01

    The gastropod Trophon geversianus exhibits shell polymorphisms along its distribution in subtidal and intertidal habitats. Our hypothesis is that morphological and behavioral patterns of T. geversianus represent habitat-specific constrains; subsequently we expect an association between shell morphology, attachment behavior, and habitat. In order to test this hypothesis we compared individuals from intertidal and subtidal habitats, at three sites in Golfo Nuevo (Argentina). We analyzed shell morphology using classic morphometric variables, 3D geometric morphometrics and computing tomography scan. The results were complemented with field observations of attachment to substrate and turning time behavior, as well as of the number of shell scars produced by crab predation. Our results showed differences in shell size and shape between intertidal and subtidal-collected individuals. Centroid size, total weight and shell weight, as well as shell density and thickness were significantly lower in intertidal individuals than in subtidal ones. Gastropods from intertidal habitats presented a low-spired shell and an expanded aperture which might allow better attachment to the bottom substrate, while subtidal individuals presented a slender and narrower shell shape. The number of crab scars was significantly higher in shells from subtidal individuals. Observations of the behavior of gastropods placed at the intertidal splash zone showed 100% of attachment to the bottom in the intertidal individuals, while subtidal specimens only attached in average in 32% of the cases. These latter took 12 times longer to re-attach to the bottom when faced up. Phylogenetic analysis of COI gene fragments showed no consistent differences among individuals sampled in both habitats. All these results suggest that T. geversianus has developed two ecomorphs with distinct morphological and behavioral responses to physically stressful conditions registered in north Patagonian intertidals, as opposed to

  18. Multiplex and genome-wide analyses reveal distinctive properties of KIR+ and CD56+ T cells in human blood.

    Science.gov (United States)

    Chan, Wing Keung; Rujkijyanont, Piya; Neale, Geoffrey; Yang, Jie; Bari, Rafijul; Das Gupta, Neha; Holladay, Martha; Rooney, Barbara; Leung, Wing

    2013-08-15

    Killer cell Ig-like receptors (KIRs) on NK cells have been linked to a wide spectrum of health conditions such as chronic infections, autoimmune diseases, pregnancy complications, cancers, and transplant failures. A small subset of effector memory T cells also expresses KIRs. In this study, we use modern analytic tools including genome-wide and multiplex molecular, phenotypic, and functional assays to characterize the KIR(+) T cells in human blood. We find that KIR(+) T cells primarily reside in the CD56(+) T population that is distinctively DNAM-1(high) with a genome-wide quiescent transcriptome, short telomere, and limited TCR excision circles. During CMV reactivation in bone marrow transplant recipients, KIR(+)CD56(+) T cells rapidly expanded in real-time but not KIR(+)CD56(-) T cells or KIR(+) NK cells. In CMV(+) asymptomatic donors, as much as 50% of CD56(+) T cells are KIR(+), and most are distinguishably KIR2DL2/3(+)NKG2C(+)CD57(+). Functionally, the KIR(+)CD56(+) T cell subset lyses cancer cells and CMVpp65-pulsed target cells in a dual KIR-dependent and TCR-dependent manner. Analysis of metabolic transcriptome confirms the immunological memory status of KIR(+)CD56(+) T cells in contrast to KIR(-)CD56(+) T cells that are more active in energy metabolism and effector differentiation. KIR(-)CD56(+) T cells have >25-fold higher level of expression of RORC than the KIR(+) counterpart and are a previously unknown producer of IL-13 rather than IL-17 in multiplex cytokine arrays. Our data provide fundamental insights into KIR(+) T cells biologically and clinically.

  19. Integrative genomics analyses reveal molecularly distinct subgroups of B-cell chronic lymphocytic leukemia patients with 13q14 deletion.

    Science.gov (United States)

    Mosca, Laura; Fabris, Sonia; Lionetti, Marta; Todoerti, Katia; Agnelli, Luca; Morabito, Fortunato; Cutrona, Giovanna; Andronache, Adrian; Matis, Serena; Ferrari, Francesco; Gentile, Massimo; Spriano, Mauro; Callea, Vincenzo; Festini, Gianluca; Molica, Stefano; Deliliers, Giorgio Lambertenghi; Bicciato, Silvio; Ferrarini, Manlio; Neri, Antonino

    2010-12-01

    Chromosome 13q14 deletion occurs in a substantial number of chronic lymphocytic leukemia (CLL) patients and it is believed to play a pathogenetic role. The exact mechanisms involved in this lesion have not yet been fully elucidated because of its heterogeneity and the imprecise knowledge of the implicated genes. This study was addressed to further contribute to the molecular definition of this lesion in CLL. We applied single-nucleotide polymorphism (SNP)-array technology and gene expression profiling data to investigate the 13q14 deletion occurring in a panel of 100 untreated, early-stage (Binet A) patients representative of the major genetics, molecular, and biological features of the disease. Concordantly with FISH analysis, SNP arrays identified 44 patients with del(13)(q14) including 11 cases with a biallelic deletion. The shorter monoallelic deletion was 635-kb long. The loss of the miR-15a/16-1 cluster occurred in all del(13)(q14) cases except in 2 patients with a monoallelic deletion, who retained both copies. MiR-15a/16 expression was significantly downregulated only in patients with the biallelic loss of the miRNA cluster compared to 13q normal cases. Finally, the natural grouping of SNP profiles by nonnegative matrix factorization algorithm showed that patients could be classified into 2 separate clusters, mainly characterized by short/biallelic versus wide/monoallelic 13q14 deletions. Supervised analyses of expression data showed that specific transcriptional profiles are correlated with these 2 genomic subgroups. Overall, our data highlight the presence of 2 distinct molecular types of 13q14 deletions, which may be of clinical relevance in CLL. ©2010 AACR.

  20. Distinct Cellular Assembly Stoichiometry of Polycomb Complexes on Chromatin Revealed by Single-molecule Chromatin Immunoprecipitation Imaging.

    Science.gov (United States)

    Tatavosian, Roubina; Zhen, Chao Yu; Duc, Huy Nguyen; Balas, Maggie M; Johnson, Aaron M; Ren, Xiaojun

    2015-11-20

    Epigenetic complexes play an essential role in regulating chromatin structure, but information about their assembly stoichiometry on chromatin within cells is poorly understood. The cellular assembly stoichiometry is critical for appreciating the initiation, propagation, and maintenance of epigenetic inheritance during normal development and in cancer. By combining genetic engineering, chromatin biochemistry, and single-molecule fluorescence imaging, we developed a novel and sensitive approach termed single-molecule chromatin immunoprecipitation imaging (Sm-ChIPi) to enable investigation of the cellular assembly stoichiometry of epigenetic complexes on chromatin. Sm-ChIPi was validated by using chromatin complexes with known stoichiometry. The stoichiometry of subunits within a polycomb complex and the assembly stoichiometry of polycomb complexes on chromatin have been extensively studied but reached divergent views. Moreover, the cellular assembly stoichiometry of polycomb complexes on chromatin remains unexplored. Using Sm-ChIPi, we demonstrated that within mouse embryonic stem cells, one polycomb repressive complex (PRC) 1 associates with multiple nucleosomes, whereas two PRC2s can bind to a single nucleosome. Furthermore, we obtained direct physical evidence that the nucleoplasmic PRC1 is monomeric, whereas PRC2 can dimerize in the nucleoplasm. We showed that ES cell differentiation induces selective alteration of the assembly stoichiometry of Cbx2 on chromatin but not other PRC1 components. We additionally showed that the PRC2-mediated trimethylation of H3K27 is not required for the assembly stoichiometry of PRC1 on chromatin. Thus, these findings uncover that PRC1 and PRC2 employ distinct mechanisms to assemble on chromatin, and the novel Sm-ChIPi technique could provide single-molecule insight into other epigenetic complexes.

  1. Genomic and phenotypic characterization of myxoma virus from Great Britain reveals multiple evolutionary pathways distinct from those in Australia.

    Science.gov (United States)

    Kerr, Peter J; Cattadori, Isabella M; Rogers, Matthew B; Fitch, Adam; Geber, Adam; Liu, June; Sim, Derek G; Boag, Brian; Eden, John-Sebastian; Ghedin, Elodie; Read, Andrew F; Holmes, Edward C

    2017-03-01

    The co-evolution of myxoma virus (MYXV) and the European rabbit occurred independently in Australia and Europe from different progenitor viruses. Although this is the canonical study of the evolution of virulence, whether the genomic and phenotypic outcomes of MYXV evolution in Europe mirror those observed in Australia is unknown. We addressed this question using viruses isolated in the United Kingdom early in the MYXV epizootic (1954-1955) and between 2008-2013. The later UK viruses fell into three distinct lineages indicative of a long period of separation and independent evolution. Although rates of evolutionary change were almost identical to those previously described for MYXV in Australia and strongly clock-like, genome evolution in the UK and Australia showed little convergence. The phenotypes of eight UK viruses from three lineages were characterized in laboratory rabbits and compared to the progenitor (release) Lausanne strain. Inferred virulence ranged from highly virulent (grade 1) to highly attenuated (grade 5). Two broad disease types were seen: cutaneous nodular myxomatosis characterized by multiple raised secondary cutaneous lesions, or an amyxomatous phenotype with few or no secondary lesions. A novel clinical outcome was acute death with pulmonary oedema and haemorrhage, often associated with bacteria in many tissues but an absence of inflammatory cells. Notably, reading frame disruptions in genes defined as essential for virulence in the progenitor Lausanne strain were compatible with the acquisition of high virulence. Combined, these data support a model of ongoing host-pathogen co-evolution in which multiple genetic pathways can produce successful outcomes in the field that involve both different virulence grades and disease phenotypes, with alterations in tissue tropism and disease mechanisms.

  2. Genomic and phenotypic characterization of myxoma virus from Great Britain reveals multiple evolutionary pathways distinct from those in Australia.

    Directory of Open Access Journals (Sweden)

    Peter J Kerr

    2017-03-01

    Full Text Available The co-evolution of myxoma virus (MYXV and the European rabbit occurred independently in Australia and Europe from different progenitor viruses. Although this is the canonical study of the evolution of virulence, whether the genomic and phenotypic outcomes of MYXV evolution in Europe mirror those observed in Australia is unknown. We addressed this question using viruses isolated in the United Kingdom early in the MYXV epizootic (1954-1955 and between 2008-2013. The later UK viruses fell into three distinct lineages indicative of a long period of separation and independent evolution. Although rates of evolutionary change were almost identical to those previously described for MYXV in Australia and strongly clock-like, genome evolution in the UK and Australia showed little convergence. The phenotypes of eight UK viruses from three lineages were characterized in laboratory rabbits and compared to the progenitor (release Lausanne strain. Inferred virulence ranged from highly virulent (grade 1 to highly attenuated (grade 5. Two broad disease types were seen: cutaneous nodular myxomatosis characterized by multiple raised secondary cutaneous lesions, or an amyxomatous phenotype with few or no secondary lesions. A novel clinical outcome was acute death with pulmonary oedema and haemorrhage, often associated with bacteria in many tissues but an absence of inflammatory cells. Notably, reading frame disruptions in genes defined as essential for virulence in the progenitor Lausanne strain were compatible with the acquisition of high virulence. Combined, these data support a model of ongoing host-pathogen co-evolution in which multiple genetic pathways can produce successful outcomes in the field that involve both different virulence grades and disease phenotypes, with alterations in tissue tropism and disease mechanisms.

  3. Biomarker Analysis Revealed Distinct Profiles of Innate and Adaptive Immunity in Infants with Ocular Lesions of Congenital Toxoplasmosis

    Science.gov (United States)

    Machado, Anderson Silva; Carneiro, Ana Carolina Aguiar Vasconcelos; Béla, Samantha Ribeiro; Andrade, Gláucia Manzan Queiroz; Vasconcelos-Santos, Daniel Vitor; Januário, José Nélio; Coelho-dos-Reis, Jordana G.; Ferro, Eloisa Amália Vieira; Teixeira-Carvalho, Andréa; Vitor, Ricardo Wagner Almeida; Martins-Filho, Olindo Assis; —UFMG-CTBG, UFMG Congenital Toxoplasmosis Brazilian Group

    2014-01-01

    Toxoplasma gondii is the main infectious cause of human posterior retinochoroiditis, the most frequent clinical manifestation of congenital toxoplasmosis. This investigation was performed after neonatal screening to identify biomarkers of immunity associated with immunopathological features of the disease by flow cytometry. The study included infected infants without NRL and with retinochoroidal lesions (ARL, ACRL, and CRL) as well as noninfected individuals (NI). Our data demonstrated that leukocytosis, with increased monocytes and lymphocytes, was a relevant hematological biomarker of ARL. Immunophenotypic analysis also revealed expansion of CD14+CD16+HLA-DRhigh monocytes and CD56dim cytotoxic NK-cells in ARL. Moreover, augmented TCRγδ+ and CD8+ T-cell counts were apparently good biomarkers of morbidity. Biomarker network analysis revealed that complex and intricated networks underscored the negative correlation of monocytes with NK- and B-cells in NRL. The remarkable lack of connections involving B-cells and a relevant shift of NK-cell connections from B-cells toward T-cells observed in ARL were outstanding. A tightly connected biomarker network was observed in CRL, with relevant connections of NK- and CD8+ T-cells with a broad range of cell subsets. Our findings add novel elements to the current knowledge on the innate and adaptive immune responses in congenital toxoplasmosis. PMID:25328286

  4. Identification of bacterial community composition in freshwater aquaculture system farming of Litopenaeus vannamei reveals distinct temperature-driven patterns.

    Science.gov (United States)

    Tang, Yuyi; Tao, Peiying; Tan, Jianguo; Mu, Haizhen; Peng, Li; Yang, Dandan; Tong, Shilu; Chen, Lanming

    2014-08-07

    Change in temperature is often a major environmental factor in triggering waterborne disease outbreaks. Previous research has revealed temporal and spatial patterns of bacterial population in several aquatic ecosystems. To date, very little information is available on aquaculture environment. Here, we assessed environmental temperature effects on bacterial community composition in freshwater aquaculture system farming of Litopenaeus vannamei (FASFL). Water samples were collected over a one-year period, and aquatic bacteria were characterized by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and 16S rDNA pyrosequencing. Resulting DGGE fingerprints revealed a specific and dynamic bacterial population structure with considerable variation over the seasonal change, suggesting that environmental temperature was a key driver of bacterial population in the FASFL. Pyrosequencing data further demonstrated substantial difference in bacterial community composition between the water at higher (WHT) and at lower (WLT) temperatures in the FASFL. Actinobacteria, Proteobacteria and Bacteroidetes were the highest abundant phyla in the FASFL, however, a large number of unclassified bacteria contributed the most to the observed variation in phylogenetic diversity. The WHT harbored remarkably higher diversity and richness in bacterial composition at genus and species levels when compared to the WLT. Some potential pathogenenic species were identified in both WHT and WLT, providing data in support of aquatic animal health management in the aquaculture industry.

  5. Analysis of segmental duplications reveals a distinct pattern of continuation-of-synteny between human and mouse genomes.

    Science.gov (United States)

    Mehan, Michael R; Almonte, Maricel; Slaten, Erin; Freimer, Nelson B; Rao, P Nagesh; Ophoff, Roel A

    2007-03-01

    About 5% of the human genome consists of large-scale duplicated segments of almost identical sequences. Segmental duplications (SDs) have been proposed to be involved in non-allelic homologous recombination leading to recurrent genomic variation and disease. It has also been suggested that these SDs are associated with syntenic rearrangements that have shaped the human genome. We have analyzed 14 members of a single family of closely related SDs in the human genome, some of which are associated with common inversion polymorphisms at chromosomes 8p23 and 4p16. Comparative analysis with the mouse genome revealed syntenic inversions for these two human polymorphic loci. In addition, 12 of the 14 SDs, while absent in the mouse genome, occur at the breaks of synteny; suggesting a non-random involvement of these sequences in genome evolution. Furthermore, we observed a syntenic familial relationship between 8 and 12 breakpoint-loci, where broken synteny that ends at one family member resumes at another, even across different chromosomes. Subsequent genome-wide assessment revealed that this relationship, which we named continuation-of-synteny, is not limited to the 8p23 family and occurs 46 times in the human genome with high frequency at specific chromosomes. Our analysis supports a non-random breakage model of genomic evolution with an active involvement of segmental duplications for specific regions of the human genome.

  6. Identification of Bacterial Community Composition in Freshwater Aquaculture System Farming of Litopenaeus vannamei Reveals Distinct Temperature-Driven Patterns

    Directory of Open Access Journals (Sweden)

    Yuyi Tang

    2014-08-01

    Full Text Available Change in temperature is often a major environmental factor in triggering waterborne disease outbreaks. Previous research has revealed temporal and spatial patterns of bacterial population in several aquatic ecosystems. To date, very little information is available on aquaculture environment. Here, we assessed environmental temperature effects on bacterial community composition in freshwater aquaculture system farming of Litopenaeus vannamei (FASFL. Water samples were collected over a one-year period, and aquatic bacteria were characterized by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE and 16S rDNA pyrosequencing. Resulting DGGE fingerprints revealed a specific and dynamic bacterial population structure with considerable variation over the seasonal change, suggesting that environmental temperature was a key driver of bacterial population in the FASFL. Pyrosequencing data further demonstrated substantial difference in bacterial community composition between the water at higher (WHT and at lower (WLT temperatures in the FASFL. Actinobacteria, Proteobacteria and Bacteroidetes were the highest abundant phyla in the FASFL, however, a large number of unclassified bacteria contributed the most to the observed variation in phylogenetic diversity. The WHT harbored remarkably higher diversity and richness in bacterial composition at genus and species levels when compared to the WLT. Some potential pathogenenic species were identified in both WHT and WLT, providing data in support of aquatic animal health management in the aquaculture industry.

  7. Biomarker Analysis Revealed Distinct Profiles of Innate and Adaptive Immunity in Infants with Ocular Lesions of Congenital Toxoplasmosis

    Directory of Open Access Journals (Sweden)

    Anderson Silva Machado

    2014-01-01

    Full Text Available Toxoplasma gondii is the main infectious cause of human posterior retinochoroiditis, the most frequent clinical manifestation of congenital toxoplasmosis. This investigation was performed after neonatal screening to identify biomarkers of immunity associated with immunopathological features of the disease by flow cytometry. The study included infected infants without NRL and with retinochoroidal lesions (ARL, ACRL, and CRL as well as noninfected individuals (NI. Our data demonstrated that leukocytosis, with increased monocytes and lymphocytes, was a relevant hematological biomarker of ARL. Immunophenotypic analysis also revealed expansion of CD14+CD16+HLA-DRhigh monocytes and CD56dim cytotoxic NK-cells in ARL. Moreover, augmented TCRγδ+ and CD8+ T-cell counts were apparently good biomarkers of morbidity. Biomarker network analysis revealed that complex and intricated networks underscored the negative correlation of monocytes with NK- and B-cells in NRL. The remarkable lack of connections involving B-cells and a relevant shift of NK-cell connections from B-cells toward T-cells observed in ARL were outstanding. A tightly connected biomarker network was observed in CRL, with relevant connections of NK- and CD8+ T-cells with a broad range of cell subsets. Our findings add novel elements to the current knowledge on the innate and adaptive immune responses in congenital toxoplasmosis.

  8. Comparative transcriptional profiling reveals differential expression of pathways directly and indirectly influencing biofilm formation in Escherichia coli O157:H7

    Science.gov (United States)

    Introduction: Escherichia coli O157:H7 (O157) is a frequent cause of foodborne disease outbreaks. O157 encodes virulence factors for colonizing and survival in reservoir animals and the environment. For example, genetic factors promoting biofilm formation are linked to survival of O157 in and outsid...

  9. A narrow amide I vibrational band observed by sum frequency generation spectroscopy reveals highly ordered structures of a biofilm protein at the air/water interface.

    Science.gov (United States)

    Wang, Zhuguang; Morales-Acosta, M Daniela; Li, Shanghao; Liu, Wei; Kanai, Tapan; Liu, Yuting; Chen, Ya-Na; Walker, Frederick J; Ahn, Charles H; Leblanc, Roger M; Yan, Elsa C Y

    2016-02-18

    We characterized BslA, a bacterial biofilm protein, at the air/water interface using vibrational sum frequency generation spectroscopy and observed one of the sharpest amide I bands ever reported. Combining methods of surface pressure measurements, thin film X-ray reflectivity, and atomic force microscopy, we showed extremely ordered BslA at the interface.

  10. Biofilm-specific extracellular matrix proteins of nontypeable Haemophilus influenzae.

    Science.gov (United States)

    Wu, Siva; Baum, Marc M; Kerwin, James; Guerrero, Debbie; Webster, Simon; Schaudinn, Christoph; VanderVelde, David; Webster, Paul

    2014-12-01

    Nontypeable Haemophilus influenzae (NTHi), a human respiratory tract pathogen, can form colony biofilms in vitro. Bacterial cells and the amorphous extracellular matrix (ECM) constituting the biofilm can be separated using sonication. The ECM from 24- and 96-h NTHi biofilms contained polysaccharides and proteinaceous components as detected by nuclear magnetic resonance (NMR) and Fourier transform infrared spectroscopy (FTIR) spectroscopy. More conventional chemical assays on the biofilm ECM confirmed the presence of these components and also DNA. Proteomics revealed eighteen proteins present in biofilm ECM that were not detected in planktonic bacteria. One ECM protein was unique to 24-h biofilms, two were found only in 96-h biofilms, and fifteen were present in the ECM of both 24- and 96-h NTHi biofilms. All proteins identified were either associated with bacterial membranes or cytoplasmic proteins. Immunocytochemistry showed two of the identified proteins, a DNA-directed RNA polymerase and the outer membrane protein OMP P2, associated with bacteria and biofilm ECM. Identification of biofilm-specific proteins present in immature biofilms is an important step in understanding the in vitro process of NTHi biofilm formation. The presence of a cytoplasmic protein and a membrane protein in the biofilm ECM of immature NTHi biofilms suggests that bacterial cell lysis may be a feature of early biofilm formation.

  11. Microbial community composition and dynamics of moving bed biofilm reactor systems treating municipal sewage.

    Science.gov (United States)

    Biswas, Kristi; Turner, Susan J

    2012-02-01

    Moving bed biofilm reactor (MBBR) systems are increasingly used for municipal and industrial wastewater treatment, yet in contrast to activated sludge (AS) systems, little is known about their constituent microbial communities. This study investigated the community composition of two municipal MBBR wastewater treatment plants (WWTPs) in Wellington, New Zealand. Monthly samples comprising biofilm and suspended biomass were collected over a 12-month period. Bacterial and archaeal community composition was determined using a full-cycle community approach, including analysis of 16S rRNA gene libraries, fluorescence in situ hybridization (FISH) and automated ribosomal intergenic spacer analysis (ARISA). Differences in microbial community structure and abundance were observed between the two WWTPs and between biofilm and suspended biomass. Biofilms from both plants were dominated by Clostridia and sulfate-reducing members of the Deltaproteobacteria (SRBs). FISH analyses indicated morphological differences in the Deltaproteobacteria detected at the two plants and also revealed distinctive clustering between SRBs and members of the Methanosarcinales, which were the only Archaea detected and were present in low abundance (MBBR systems and indicates that this process selects for distinctive biofilm and planktonic communities, both of which differ from those found in conventional AS systems.

  12. Disruption of aminergic signalling reveals novel compounds with distinct inhibitory effects on mosquito reproduction, locomotor function and survival

    Science.gov (United States)

    Fuchs, Silke; Rende, Ermelinda; Crisanti, Andrea; Nolan, Tony

    2014-07-01

    Insecticide resistance amongst disease vectors is a growing problem and novel compounds are needed. Biogenic amines are important for neurotransmission and we have recently shown a potential role for these in mosquito fertility. Here, we dissected the relative contribution of different aminergic signalling pathways to biological processes essential for vectorial capacity such as fertility, locomotion and survival by injecting agonists and antagonists and showed that octopaminergic/tyraminergic signalling is essential for oviposition and hatching rate. We show that egg melanisation is regulated by adrenergic signalling, whose disruption causes premature melanisation specifically through the action of tyramine. In addition to this, co-injection of tyramine with DOPA, the precursor of melanin, had a strong cumulative negative effect on mosquito locomotion and survival. Dopaminergic and serotonergic antagonists such as amitriptyline and citalopram recapitulate this effect. Together these results reveal potential new target sites for the development of future mosquito sterilants and insecticides.

  13. Metagenome Sequence Analysis of Filamentous Microbial Communities Obtained from Geochemically Distinct Geothermal Channels Reveals Specialization of Three Aquificales Lineages

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    Cristina eTakacs-vesbach

    2013-05-01

    Full Text Available The Aquificales are thermophilic microorganisms that inhabit hydrothermal systems worldwide and are considered one of the earliest lineages of the domain Bacteria. We analyzed metagenome sequence obtained from six thermal ‘filamentous streamer’ communities (~40 Mbp per site, which targeted three different groups of Aquificales found in Yellowstone National Park (YNP. Unassembled metagenome sequence and PCR-amplified 16S rRNA gene libraries revealed that acidic, sulfidic sites were dominated by Hydrogenobaculum (Aquificaceae populations, whereas the circumneutral pH (6.5 - 7.8 sites containing dissolved sulfide were dominated by Sulfurihydrogenibium spp. (Hydrogenothermaceae. Thermocrinis (Aquificaceae populations were found primarily in the circumneutral sites with undetectable sulfide, and to a lesser extent in one sulfidic system at pH 8. Phylogenetic analysis of assembled sequence containing 16S rRNA genes as well as conserved protein-encoding genes revealed that the composition and function of these communities varied across geochemical conditions. Each Aquificales lineage contained genes for CO2 fixation by the reverse TCA cycle, but only the Sulfurihydrogenibium populations perform citrate cleavage using ATP citrate lyase (Acl. The Aquificaceae populations use an alternative pathway catalyzed by two separate enzymes, citryl CoA synthetase (Ccs and citryl CoA lyase (Ccl. All three Aquificales lineages contained evidence of aerobic respiration, albeit due to completely different types of heme Cu oxidases (subunit I involved in oxygen reduction. The distribution of Aquificales populations and differences among functional genes involved in energy generation and electron transport is consistent with the hypothesis that geochemical parameters (e.g., pH, sulfide, H2, O2 have resulted in niche specialization among members of the Aquificales.

  14. RNA-Seq Reveals Enhanced Sugar Metabolism in Streptococcus mutans Co-cultured with Candida albicans within Mixed-Species Biofilms

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    Jinzhi He

    2017-06-01

    Full Text Available Early childhood caries (ECC, which can lead to rampant tooth-decay that is painful and costly to treat, is one of the most prevalent infectious diseases affecting children worldwide. Previous studies support that interactions between Streptococcus mutans and Candida albicans are associated with the pathogenesis of ECC. The presence of Candida enhances S. mutans growth, fitness and accumulation within biofilms in vitro, although the molecular basis for these behaviors is undefined. Using an established co-cultivation biofilm model and RNA-Seq, we investigated how C. albicans influences the transcriptome of S. mutans. The presence of C. albicans dramatically altered gene expression in S. mutans in the dual-species biofilm, resulting in 393 genes differentially expressed, compared to mono-species biofilms of S. mutans. By Gene Ontology analysis, the majority of up-regulated genes were related to carbohydrate transport and metabolic/catabolic processes. KEGG pathway impact analysis showed elevated pyruvate and galactose metabolism, suggesting that co-cultivation with C. albicans influences carbohydrate utilization by S. mutans. Analysis of metabolites confirmed the increases in carbohydrate metabolism, with elevated amounts of formate in the culture medium of co-cultured biofilms. Moreover, co-cultivation with C. albicans altered transcription of S. mutans signal transduction (comC and ciaRH genes associated with fitness and virulence. Interestingly, the expression of genes for mutacins (bacteriocins and CRISPR were down-regulated. Collectively, the data provide a comprehensive insight into S. mutans transcriptomic changes induced by C. albicans, and offer novel insights into how bacterial–fungal interactions may enhance the severity of dental caries.

  15. Detection of microbial biofilms on food processing surfaces: hyperspectral fluorescence imaging study

    Science.gov (United States)

    Jun, Won; Kim, Moon S.; Chao, Kaunglin; Lefcourt, Alan M.; Roberts, Michael S.; McNaughton, James L.

    2009-05-01

    We used a portable hyperspectral fluorescence imaging system to evaluate biofilm formations on four types of food processing surface materials including stainless steel, polypropylene used for cutting boards, and household counter top materials such as formica and granite. The objective of this investigation was to determine a minimal number of spectral bands suitable to differentiate microbial biofilm formation from the four background materials typically used during food processing. Ultimately, the resultant spectral information will be used in development of handheld portable imaging devices that can be used as visual aid tools for sanitation and safety inspection (microbial contamination) of the food processing surfaces. Pathogenic E. coli O157:H7 and Salmonella cells were grown in low strength M9 minimal medium on various surfaces at 22 +/- 2 °C for 2 days for biofilm formation. Biofilm autofluorescence under UV excitation (320 to 400 nm) obtained by hyperspectral fluorescence imaging system showed broad emissions in the blue-green regions of the spectrum with emission maxima at approximately 480 nm for both E. coli O157:H7 and Salmonella biofilms. Fluorescence images at 480 nm revealed that for background materials with near-uniform fluorescence responses such as stainless steel and formica cutting board, regardless of the background intensity, biofilm formation can be distinguished. This suggested that a broad spectral band in the blue-green regions can be used for handheld imaging devices for sanitation inspection of stainless, cutting board, and formica surfaces. The non-uniform fluorescence responses of granite make distinctions between biofilm and background difficult. To further investigate potential detection of the biofilm formations on granite surfaces with multispectral approaches, principal component analysis (PCA) was performed using the hyperspectral fluorescence image data. The resultant PCA score images revealed distinct contrast between

  16. Spatial and temporal variability of biomarkers and microbial diversity reveal metabolic and community flexibility in Streamer Biofilm Communities in the Lower Geyser Basin, Yellowstone National Park.

    Science.gov (United States)

    Schubotz, F; Meyer-Dombard, D R; Bradley, A S; Fredricks, H F; Hinrichs, K-U; Shock, E L; Summons, R E

    2013-11-01

    Detailed analysis of 16S rRNA and intact polar lipids (IPLs) from streamer biofilm communities (SBCs), collected from geochemically similar hot springs in the Lower Geyser Basin, Yellowstone National Park, shows good agreement and affirm that IPLs can be used as reliable markers for the microbial constituents of SBCs. Uncultured Crenarchaea are prominent in SBS, and their IPLs contain both glycosidic and mixed glyco-phospho head groups with tetraether cores, having 0-4 rings. Archaeal IPL contributions increase with increasing temperature and comprise up to one-fourth of the total IPL inventory at >84 °C. At elevated temperatures, bacterial IPLs contain abundant glycosidic glycerol diether lipids. Diether and diacylglycerol (DAG) lipids with aminopentanetetrol and phosphatidylinositol head groups were identified as lipids diagnostic of Aquificales, while DAG glycolipids and glyco-phospholipids containing N-acetylgycosamine as head group were assigned to members of the Thermales. With decreasing temperature and concomitant changes in water chemistry, IPLs typical of phototrophic bacteria, such as mono-, diglycosyl, and sulfoquinovosyl DAG, which are specific for cyanobacteria, increase in abundance, consistent with genomic data from the same samples. Compound-specific stable carbon isotope analysis of IPL breakdown products reveals a large isotopic diversity among SBCs in different hot springs. At two of the hot springs, 'Bison Pool' and Flat Cone, lipids derived from Aquificales are enriched in (13) C relative to biomass and approach values close to dissolved inorganic carbon (DIC) (approximately 0‰), consistent with fractionation during autotrophic carbon fixation via the reversed tricarboxylic acid pathway. At a third site, Octopus Spring, the same Aquificales-diagnostic lipids are 10‰ depleted relative to biomass and resemble stable carbon isotope values of dissolved organic carbon (DOC), indicative of heterotrophy. Other bacterial and archaeal lipids show

  17. Genetic Elimination of GABAergic Neurotransmission Reveals Two Distinct Pacemakers for Spontaneous Waves of Activity in the Developing Mouse Cortex

    Science.gov (United States)

    Easton, Curtis R.; Weir, Keiko; Scott, Adina; Moen, Samantha P.; Barger, Zeke; Folch, Albert; Hevner, Robert F.

    2014-01-01

    Many structures of the mammalian CNS generate propagating waves of electrical activity early in development. These waves are essential to CNS development, mediating a variety of developmental processes, such as axonal outgrowth and pathfinding, synaptogenesis, and the maturation of ion channel and receptor properties. In the mouse cerebral cortex, waves of activity occur between embryonic day 18 and postnatal day 8 and originate in pacemaker circuits in the septal nucleus and the piriform cortex. Here we show that genetic knock-out of the major synthetic enzyme for GABA, GAD67, selectively eliminates the picrotoxin-sensitive fraction of these waves. The waves that remain in the GAD67 knock-out have a much higher probability of propagating into the dorsal neocortex, as do the picrotoxin-resistant fraction of waves in controls. Field potential recordings at the point of wave initiation reveal different electrical signatures for GABAergic and glutamatergic waves. These data indicate that: (1) there are separate GABAergic and glutamatergic pacemaker circuits within the piriform cortex, each of which can initiate waves of activity; (2) the glutamatergic pacemaker initiates waves that preferentially propagate into the neocortex; and (3) the initial appearance of the glutamatergic pacemaker does not require preceding GABAergic waves. In the absence of GAD67, the electrical activity underlying glutamatergic waves shows greatly increased tendency to burst, indicating that GABAergic inputs inhibit the glutamatergic pacemaker, even at stages when GABAergic pacemaker circuitry can itself initiate waves. PMID:24623764

  18. Crystal structure of TAZ-TEAD complex reveals a distinct interaction mode from that of YAP-TEAD complex.

    Science.gov (United States)

    Kaan, Hung Yi Kristal; Chan, Siew Wee; Tan, Siew Kim Joyce; Guo, Fusheng; Lim, Chun Jye; Hong, Wanjin; Song, Haiwei

    2017-05-17

    The Hippo pathway is a tumor suppressor pathway that is implicated in the regulation of organ size. The pathway has three components: the upstream regulatory factors, the kinase core, and the downstream transcriptional machinery, which consists of YAP, TAZ (transcription co-activators) and TEAD (transcription factor). Formation of YAP/TAZ-TEAD complexes leads to the transcription of growth-promoting genes. Herein, we report the crystal structure of TAZ-TEAD4 complex, which reveals two binding modes. The first is similar to the published YAP-TEAD structure. The second is a unique binding mode, whereby two molecules of TAZ bind to and bridge two molecules of TEAD4. We validated the latter using cross-linking and multi-angle light scattering. Using siRNA, we showed that TAZ knockdown leads to a decrease in TEAD4 dimerization. Lastly, results from luciferase assays, using YAP/TAZ transfected or knockdown cells, give support to the non-redundancy of YAP/TAZ co-activators in regulating gene expression in the Hippo pathway.

  19. Molecular characterization of HCV in a Swedish county over 8 years (2002–2009 reveals distinct transmission patterns

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    Josefine Ederth

    2016-02-01

    Full Text Available Background: Hepatitis C virus (HCV is a major public health concern and data on its molecular epidemiology in Sweden is scarce. We carried out an 8-year population-based study of newly diagnosed HCV cases in one of Sweden's centrally situated counties, Södermanland (D-county. The aim was to characterize the HCV strains circulating, analyze their genetic relatedness to detect networks, and in combination with demographic data learn more about transmission. Methods: Molecular analyses of serum samples from 91% (N=557 of all newly notified cases in D-county, 2002–2009, were performed. Phylogenetic analysis (NS5B gene, 300 bp was linked to demographic data from the national surveillance database, SmiNet, to characterize D-county transmission clusters. The linear-by-linear association test (LBL was used to analyze trends over time. Results: The most prevalent subtypes were 1a (38% and 3a (34%. Subtype 1a was most prevalent among cases transmitted via sexual contact, via contaminated blood, or blood products, while subtype 3a was most prevalent among people who inject drugs (PWIDs. Phylogenetic analysis revealed that the subtype 3a sequences formed more and larger transmission clusters (50% of the sequences clustered, while the 1a sequences formed smaller clusters (19% of the sequences clustered, possibly suggesting different epidemics. Conclusion: We found different transmission patterns in D-county which may, from a public health perspective, have implications for how to control virus infections by targeted interventions.

  20. Quantitative morphometry of electrophysiologically identified CA3b interneurons reveals robust local geometry and distinct cell classes.

    Science.gov (United States)

    Ascoli, Giorgio A; Brown, Kerry M; Calixto, Eduardo; Card, J Patrick; Galván, E J; Perez-Rosello, T; Barrionuevo, Germán

    2009-08-20

    The morphological and electrophysiological diversity of inhibitory cells in hippocampal area CA3 may underlie specific computational roles and is not yet fully elucidated. In particular, interneurons with somata in strata radiatum (R) and lacunosum-moleculare (L-M) receive converging stimulation from the dentate gyrus and entorhinal cortex as well as within CA3. Although these cells express different forms of synaptic plasticity, their axonal trees and connectivity are still largely unknown. We investigated the branching and spatial patterns, plus the membrane and synaptic properties, of rat CA3b R and L-M interneurons digitally reconstructed after intracellular labeling. We found considerable variability within but no difference between the two layers, and no correlation between morphological and biophysical properties. Nevertheless, two cell types were identified based on the number of dendritic bifurcations, with significantly different anatomical and electrophysiological features. Axons generally branched an order of magnitude more than dendrites. However, interneurons on both sides of the R/L-M boundary revealed surprisingly modular axodendritic arborizations with consistently uniform local branch geometry. Both axons and dendrites followed a lamellar organization, and axons displayed a spatial preference toward the fissure. Moreover, only a small fraction of the axonal arbor extended to the outer portion of the invaded volume, and tended to return toward the proximal region. In contrast, dendritic trees demonstrated more limited but isotropic volume occupancy. These results suggest a role of predominantly local feedforward and lateral inhibitory control for both R and L-M interneurons. Such a role may be essential to balance the extensive recurrent excitation of area CA3 underlying hippocampal autoassociative memory function.

  1. Cap analog substrates reveal three clades of cap guanine-N2 methyltransferases with distinct methyl acceptor specificities.

    Science.gov (United States)

    Benarroch, Delphine; Jankowska-Anyszka, Marzena; Stepinski, Janusz; Darzynkiewicz, Edward; Shuman, Stewart

    2010-01-01

    The Tgs proteins are structurally homologous AdoMet-dependent eukaryal enzymes that methylate the N2 atom of 7-methyl guanosine nucleotides. They have an imputed role in the synthesis of the 2,2,7-trimethylguanosine (TMG) RNA cap. Here we exploit a collection of cap-like substrates to probe the repertoire of three exemplary Tgs enzymes, from mammalian, protozoan, and viral sources, respectively. We find that human Tgs (hTgs1) is a bona fide TMG synthase adept at two separable transmethylation steps: (1) conversion of m(7)G to m(2,7)G, and (2) conversion of m(2,7)G to m(2,2,7)G. hTgs1 is unable to methylate G or m(2)G, signifying that both steps require an m(7)G cap. hTgs1 utilizes a broad range of m(7)G nucleotides, including mono-, di-, tri-, and tetraphosphate derivatives as well as cap dinucleotides with triphosphate or tetraphosphate bridges. In contrast, Giardia lamblia Tgs (GlaTgs2) exemplifies a different clade of guanine-N2 methyltransferase that synthesizes only a dimethylguanosine (DMG) cap structure and cannot per se convert DMG to TMG under any conditions tested. Methylation of benzyl(7)G and ethyl(7)G nucleotides by hTgs1 and GlaTgs2 underscored the importance of guanine N7 alkylation in providing a key pi-cation interaction in the methyl acceptor site. Mimivirus Tgs (MimiTgs) shares with the Giardia homolog the ability to catalyze only a single round of methyl addition at guanine-N2, but is distinguished by its capacity for guanine-N2 methylation in the absence of prior N7 methylation. The relaxed cap specificity of MimiTgs is revealed at alkaline pH. Our findings highlight both stark and subtle differences in acceptor specificity and reaction outcomes among Tgs family members.

  2. Genome-wide transcriptional profiling reveals two distinct outcomes in central Nervous system infections of rabies virus

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    Daiting eZhang

    2016-05-01

    Full Text Available Rabies remains a major public health concern in many developing countries. The precise neuropathogenesis of rabies is unknown, though it is hypothesized to be due to neuronal death or dysfunction. Mice that received intranasal inoculation of an attenuated rabies virus (RABV strain HEP-Flury exhibited subtle clinical signs, and eventually recovered, which is different from the fatal encephalitis caused by the virulent RABV strain CVS-11. To understand the neuropathogenesis of rabies and the mechanisms of viral clearance, we applied RNA sequencing (RNA-Seq to compare the brain transcriptomes of normal mice versus HEP-Flury or CVS-11 intranasally inoculated mice. Our results revealed that both RABV strains altered positively and negatively the expression levels of many host genes, including genes associated with innate and adaptive immunity, inflammation and cell death. It is found that HEP-Flury infection can activate the innate immunity earlier through the RIG-I/MDA-5 signaling, and the innate immunity pre-activated by HEP-Flury or Newcastle disease virus (NDV infection can effectively prevent the CVS-11 to invade central nervous system (CNS, but fails to clear the CVS-11 after its entry into the CNS. In addition, following CVS-11 infection, genes implicated in cell adhesion, blood vessel morphogenesis and coagulation were mainly up-regulated, while the genes involved in synaptic transmission and ion transport were significantly down-regulated. On the other hand, several genes involved in the MHC class II-mediated antigen presentation pathway were activated to a greater extent after the HEP-Flury infection as compared with the CVS-11 infection suggesting that the collaboration of CD4+ T cells and MHC class II-mediated antigen presentation is critical for the clearance of attenuated RABV from the CNS. The differentially regulated genes reported here are likely to include potential therapeutic targets for expanding the postexposure treatment window

  3. Yeast DNA ligase IV mutations reveal a nonhomologous end joining function of BRCT1 distinct from XRCC4/Lif1 binding.

    Science.gov (United States)

    Chiruvella, Kishore K; Renard, Brian M; Birkeland, Shanda R; Sunder, Sham; Liang, Zhuobin; Wilson, Thomas E

    2014-12-01

    LIG4/Dnl4 is the DNA ligase that (re)joins DNA double-strand breaks (DSBs) via nonhomologous end joining (NHEJ), an activity supported by binding of its tandem BRCT domains to the ligase accessory protein XRCC4/Lif1. We screened a panel of 88 distinct ligase mutants to explore the structure–function relationships of the yeast Dnl4 BRCT domains and inter-BRCT linker in NHEJ. Screen results suggested two distinct classes of BRCT mutations with differential effects on Lif1 interaction as compared to NHEJ completion. Validated constructs confirmed that D800K and GG(868:869)AA mutations, which target the Lif1 binding interface, showed a severely defective Dnl4–Lif1 interaction but a less consistent and often small decrease in NHEJ activity in some assays, as well as nearly normal levels of Dnl4 accumulation at DSBs. In contrast, mutants K742A and KTT(742:744)ATA, which target the β3-α2 region of the first BRCT domain, substantially decreased NHEJ function commensurate with a large defect in Dnl4 recruitment to DSBs, despite a comparatively greater preservation of the Lif1 interaction. Together, these separation-of-function mutants indicate that Dnl4 BRCT1 supports DSB recruitment and NHEJ in a manner distinct from Lif1 binding and reveal a complexity of Dnl4 BRCT domain functions in support of stable DSB association.

  4. Environmental transcriptome analysis reveals physiological differences between biofilm and planktonic modes of life of the iron oxidizing bacteria Leptospirillum spp. in their natural microbial community

    OpenAIRE

    Parro Víctor; Arcas Aida; Gómez Manuel J; Moreno-Paz Mercedes

    2010-01-01

    Abstract Background Extreme acidic environments are characterized by their high metal content and lack of nutrients (oligotrophy). Macroscopic biofilms and filaments usually grow on the water-air interface or under the stream attached to solid substrates (streamers). In the Río Tinto (Spain), brown filaments develop under the water stream where the Gram-negative iron-oxidizing bacteria Leptospirillum spp. (L. ferrooxidans and L. ferriphilum) and Acidithiobacillus ferrooxidans are abundant. Th...

  5. Electroactive biofilms of sulphate reducing bacteria

    Energy Technology Data Exchange (ETDEWEB)

    Cordas, Cristina M.; Guerra, L. Tiago; Xavier, Catarina [Requimte-CQFB, Departamento de Quimica, Faculdade de Ciencias e Tecnologia, Universidade Nova de Lisboa, 2829-516 Caparica (Portugal); Moura, Jose J.G. [Requimte-CQFB, Departamento de Quimica, Faculdade de Ciencias e Tecnologia, Universidade Nova de Lisboa, 2829-516 Caparica (Portugal)], E-mail: jose.moura@dq.fct.unl.pt

    2008-12-01

    Biofilms formed from a pure strain of Desulfovibrio desulfuricans 27774 on stainless steel and graphite polarised surfaces were studied. The polarisation conditions applied were -0.4 V vs. SCE for different times. A cathodic current related with the biofilms growth was observed with a maximum intensity of -270 mA m{sup -2} that remained stable for several days using graphite electrodes. These sulphate reducing bacteria biofilms present electrocatalytic activity towards hydrogen and oxygen reduction reactions. Electrode polarisation has a selective effect on the catalytic activity. The biofilms were also observed by scanning electronic microscopy revealing the formation of homogeneous films on the surfaces.

  6. Characterization of the TolB-Pal trans-envelope complex from Xylella fastidiosa reveals a dynamic and coordinated protein expression profile during the biofilm development process.

    Science.gov (United States)

    Santos, Clelton A; Janissen, Richard; Toledo, Marcelo A S; Beloti, Lilian L; Azzoni, Adriano R; Cotta, Monica A; Souza, Anete P

    2015-10-01

    The intriguing roles of the bacterial Tol-Pal trans-envelope protein complex range from maintenance of cell envelope integrity to potential participation in the process of cell division. In this study, we report the characterization of the XfTolB and XfPal proteins of the Tol-Pal complex of Xylella fastidiosa. X. fastidiosa is a major plant pathogen that forms biofilms inside xylem vessels, triggering the development of diseases in important cultivable plants around the word. Based on functional complementation experiments in Escherichia coli tolB and pal mutant strains, we confirmed the role of xftolB and xfpal in outer membrane integrity. In addition, we observed a dynamic and coordinated protein expression profile during the X. fastidiosa biofilm development process. Using small-angle X-ray scattering (SAXS), the low-resolution structure of the isolated XfTolB-XfPal complex in solution was solved for the first time. Finally, the localization of the XfTolB and XfPal polar ends was visualized via immunofluorescence labeling in vivo during bacterial cell growth. Our results highlight the major role of the components of the cell envelope, particularly the TolB-Pal complex, during the different phases of bacterial biofilm development. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Revealing the relationship between microbial community structure in natural biofilms and the pollution level in urban rivers: a case study in the Qinhuai River basin, Yangtze River Delta.

    Science.gov (United States)

    Cai, Wei; Li, Yi; Wang, Peifang; Niu, Lihua; Zhang, Wenlong; Wang, Chao

    River pollution is one of the most challenging environmental issues, but the effect of river pollution levels on the biofilm communities has not been well-studied. Spatial and temporal distribution characteristics of environmental parameters and the biofilm communities were investigated in the Qinhuai River basin, Nanjing, China. Water samples were grouped into three clusters reflecting their varying pollution levels of relatively slight pollution, moderated pollution, and high pollution by hierarchical cluster analysis. In different clusters, the biofilm communities mainly differed in the proportion of Actinobacteria, Firmicutes, and Proteobacteria. As the dominant classes of Proteobacteria, Alpha-, Beta- and Gammaproteobacteria seemed to show an upward trend followed by a small fluctuation in the abundance with the escalation of water pollution level. Results of redundancy analysis demonstrated that temperature, total nitrogen to total phosphorus ratios (TN/TP) and concentrations of ammonia nitrogen (NH3-N) and TN were mainly responsible for the variation in bacterial community structure. The occurrences of Alpha-, Beta- and Gammaproteobacteria were closely associated with higher temperature, higher concentrations of NH3-N and TN and a lower TN/TP ratio. This study may provide a theoretical basis for the water pollution control and ecological restoration in urban rivers under different pollution levels.

  8. Oogonial biometry and phylogenetic analyses of the Pythium vexans species group from woody agricultural hosts in South Africa reveal distinct groups within this taxon.

    Science.gov (United States)

    Spies, Christoffel F J; Mazzola, Mark; Botha, Wilhelm J; Van Der Rijst, Marieta; Mostert, Lizel; Mcleod, Adéle

    2011-02-01

    Pythium vexans fits into the internal transcribed spacer (ITS) clade K sensu Lévesque & De Cock (2004). Within clade K, P. vexans forms a distinct clade containing two enigmatic species, Pythium indigoferae and Pythium cucurbitacearum of which no ex-type strains are available. In South Africa, as well as in other regions of the world, P. vexans isolates are known to be heterogeneous in their ITS sequences and may consist of more than one species. This study aimed to investigate the diversity of South African P. vexans isolates, mainly from grapevines, but also citrus and apple using (i) phylogenetic analyses of the ITS, cytochrome c oxidase (cox) I, cox II, and β-tubulin regions and (ii) seven biometric oogonial parameters. Each of the phylogenies clustered P. vexans isolates into a single well-supported clade, distinct from other clade K species. The β-tubulin region was phylogenetically uninformative regarding the P. vexans group. The ITS phylogeny and combined cox I and II phylogenies, although each revealing several P. vexans subclades, were incongruent. One of the most striking incongruences was the presence of one cox subclade that contained two distinct ITS subclades (Ib and IV). Three groups (A-C) were subjectively identified among South African P. vexans isolates using (i) phylogenetic clades (ITS and cox), (ii) univariate analysis of oogonial diameters, and (iii) multivariate analyses of biometric oogonial parameters. Group A is considered to be P. vexans s. str. since it contained the P. vexans CBS reference strain from Van der Plaats-Niterink (1981). This group had significantly smaller oogonial diameters than group B and C isolates. Group B contained the isolates from ITS subclades Ib and IV, which formed a single cox subclade. The ITS subclade IV isolates were all sexually sterile or produced mainly abortive oospores, as opposed to the sexually fertile subclade Ib isolates, and may thus represent a distinct assemblage within group B. Although ITS

  9. Comparative Genomic Analyses of Multiple Pseudomonas Strains Infecting Corylus avellana Trees Reveal the Occurrence of Two Genetic Clusters with Both Common and Distinctive Virulence and Fitness Traits.

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    Simone Marcelletti

    Full Text Available The European hazelnut (Corylus avellana is threatened in Europe by several pseudomonads which cause symptoms ranging from twig dieback to tree death. A comparison of the draft genomes of nine Pseudomonas strains isolated from symptomatic C. avellana trees was performed to identify common and distinctive genomic traits. The thorough assessment of genetic relationships among the strains revealed two clearly distinct clusters: P. avellanae and P. syringae. The latter including the pathovars avellanae, coryli and syringae. Between these two clusters, no recombination event was found. A genomic island of approximately 20 kb, containing the hrp/hrc type III secretion system gene cluster, was found to be present without any genomic difference in all nine pseudomonads. The type III secretion system effector repertoires were remarkably different in the two groups, with P. avellanae showing a higher number of effectors. Homologue genes of the antimetabolite mangotoxin and ice nucleation activity clusters were found solely in all P. syringae pathovar strains, whereas the siderophore yersiniabactin was only present in P. avellanae. All nine strains have genes coding for pectic enzymes and sucrose metabolism. By contrast, they do not have genes coding for indolacetic acid and anti-insect toxin. Collectively, this study reveals that genomically different Pseudomonas can converge on the same host plant by suppressing the host defence mechanisms with the use of different virulence weapons. The integration into their genomes of a horizontally acquired genomic island could play a fundamental role in their evolution, perhaps giving them the ability to exploit new ecological niches.

  10. AFM Structural Characterization of Drinking Water Biofilm ...

    Science.gov (United States)

    Due to the complexity of mixed culture drinking water biofilm, direct visual observation under in situ conditions has been challenging. In this study, atomic force microscopy (AFM) revealed the three dimensional morphology and arrangement of drinking water relevant biofilm in air and aqueous solution. Operating parameters were optimized to improve imaging of structural details for a mature biofilm in liquid. By using a soft cantilever (0.03 N/m) and slow scan rate (0.5 Hz), biofilm and individual bacterial cell’s structural topography were resolved and continuously imaged in liquid without loss of spatial resolution or sample damage. The developed methodology will allow future in situ investigations to temporally monitor mixed culture drinking water biofilm structural changes during disinfection treatments. Due to the complexity of mixed culture drinking water biofilm, direct visual observation under in situ conditions has been challenging. In this study, atomic force microscopy (AFM) revealed the three dimensional morphology and arrangement of drinking water relevant biofilm in air and aqueous solution. Operating parameters were optimized to improve imaging of structural details for a mature biofilm in liquid. By using a soft cantilever (0.03 N/m) and slow scan rate (0.5 Hz), biofilm and individual bacterial cell’s structural topography were resolved and continuously imaged in liquid without loss of spatial resolution or sample damage. The developed methodo

  11. Genome-wide evaluation of the interplay between Caenorhabditis elegans and Yersinia pseudotuberculosis during in vivo biofilm formation.

    Science.gov (United States)

    Joshua, George W P; Atkinson, Steve; Goldstone, Robert J; Patrick, Hannah L; Stabler, Richard A; Purves, Joanne; Cámara, Miguel; Williams, Paul; Wren, Brendan W

    2015-01-01

    The formation of an incapacitating biofilm on Caenorhabditis elegans by Yersinia pseudotuberculosis represents a tractable model for investigating the genetic basis for host-pathogen interplay during the biofilm-mediated infection of a living surface. Previously we established a role for quorum sensing (QS) and the master motility regulator, FlhDC, in biofilm formation by Y. pseudotuberculosis on C. elegans. To obtain further genome-wide insights, we used transcriptomic analysis to obtain comparative information on C. elegans in the presence and absence of biofilm and on wild-type Y. pseudotuberculosis and Y. pseudotuberculosis QS mutants. Infection of C. elegans with the wild-type Y. pseudotuberculosis resulted in the differential regulation of numerous genes, including a distinct subset of nematode C-lectin (clec) and fatty acid desaturase (fat) genes. Evaluation of the corresponding C. elegans clec-49 and fat-3 deletion mutants showed delayed biofilm formation and abolished biofilm formation, respectively. Transcriptomic analysis of Y. pseudotuberculosis revealed that genes located in both of the histidine utilization (hut) operons were upregulated in both QS and flhDC mutants. In addition, mutation of the regulatory gene hutC resulted in the loss of biofilm, increased expression of flhDC, and enhanced swimming motility. These data are consistent with the existence of a regulatory cascade in which the Hut pathway links QS and flhDC. This work also indicates that biofilm formation by Y. pseudotuberculosis on C. elegans is an interactive process during which the initial attachment/recognition of Yersinia to/by C. elegans is followed by bacterial growth and biofilm formation.

  12. Resistance of non-typeable Haemophilus influenzae biofilms is independent of biofilm size.

    Science.gov (United States)

    Reimche, Jennifer L; Kirse, Daniel J; Whigham, Amy S; Swords, W Edward

    2017-02-01

    The inflammatory middle ear disease known as otitis media can become chronic or recurrent in some cases due to failure of the antibiotic treatment to clear the bacterial etiological agent. Biofilms are known culprits of antibiotic-resistant infections; however, the mechanisms of resistance for non-typeable Haemophilus influenzae biofilms have not been completely elucidated. In this study, we utilized in vitro static biofilm assays to characterize clinical strain biofilms and addressed the hypothesis that biofilms with greater biomass and/or thickness would be more resistant to antimicrobial-mediated eradication than thinner and/or lower biomass biofilms. Consistent with previous studies, antibiotic concentrations required to eliminate biofilm bacteria tended to be drastically higher than concentrations required to kill planktonic bacteria. The size characterizations of the biofilms formed by the clinical isolates were compared to their minimum biofilm eradication concentrations for four antibiotics. This revealed no correlation between biofilm thickness or biomass and the ability to resist eradication by antibiotics. Therefore, we concluded that biofilm size does not play a role in antibiotic resistance, suggesting that reduction of antibiotic penetration may not be a significant mechanism for antibiotic resistance for this bacterial opportunist. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  13. Role of biofilm roughness and hydrodynamic conditions in Legionella pneumophila adhesion to and detachment from simulated drinking water biofilms.

    Science.gov (United States)

    Shen, Yun; Monroy, Guillermo L; Derlon, Nicolas; Janjaroen, Dao; Huang, Conghui; Morgenroth, Eberhard; Boppart, Stephen A; Ashbolt, Nicholas J; Liu, Wen-Tso; Nguyen, Thanh H

    2015-04-07

    Biofilms in drinking water distribution systems (DWDS) could exacerbate the persistence and associated risks of pathogenic Legionella pneumophila (L. pneumophila), thus raising human health concerns. However, mechanisms controlling adhesion and subsequent detachment of L. pneumophila associated with biofilms remain unclear. We determined the connection between L. pneumophila adhesion and subsequent detachment with biofilm physical structure characterization using optical coherence tomography (OCT) imaging technique. Analysis of the OCT images of multispecies biofilms grown under low nutrient condition up to 34 weeks revealed the lack of biofilm deformation even when these biofilms were exposed to flow velocity of 0.7 m/s, typical flow for DWDS. L. pneumophila adhesion on these biofilm under low flow velocity (0.007 m/s) positively correlated with biofilm roughness due to enlarged biofilm surface area and local flow conditions created by roughness asperities. The preadhered L. pneumophila on selected rough and smooth biofilms were found to detach when these biofilms were subjected to higher flow velocity. At the flow velocity of 0.1 and 0.3 m/s, the ratio of detached cell from the smooth biofilm surface was from 1.3 to 1.4 times higher than that from the rough biofilm surface, presumably because of the low shear stress zones near roughness asperities. This study determined that physical structure and local hydrodynamics control L. pneumophila adhesion to and detachment from simulated drinking water biofilm, thus it is the first step toward reducing the risk of L. pneumophila exposure and subsequent infections.

  14. The salmochelin receptor IroN itself, but not salmochelin-mediated iron uptake promotes biofilm formation in extraintestinal pathogenic Escherichia coli (ExPEC).

    Science.gov (United States)

    Magistro, Giuseppe; Hoffmann, Christiane; Schubert, Sören

    2015-01-01

    The key to success of extraintestinal pathogenic Escherichia coli (ExPEC) to colonize niches outside the intestinal tract and to establish infection is the coordinated action of numerous virulence and fitness factors. Intense research revealed not only an arsenal of unique virulence determinants with specific action, but also the multi-functionality of single elements. Especially iron uptake systems of ExPEC proved to be of prime importance. Apart from iron acquisition they optimize certain virulence properties. Here we analyzed the contribution of the salmochelin siderophore system to the ability of ExPEC to form biofilms. In the same iron limited environment, ExPEC displayed a distinct transcriptional profile of siderophore systems. During biofilm formation the iroN gene coding for the specific receptors of the siderophore salmochelin was highly upregulated. Almost no induction was observed during planctonic growth. Disruption of iroN resulted in a reduction of almost 50% in biofilm production. Efficient biofilm formation was not affected in a salmochelin synthesis mutant. Thus, the contribution of IroN is independent from the ability to produce salmochelin. Enhanced expression of IroN did not increase significantly the capacity to form biofilms in ExPEC. Interestingly, the additional expression of IroN or even the acquisition of the entire salmochelin system was not able to improve biofilm formation in a poor biofilm producer like a laboratory E. coli K12 strain. However, complementation with only IroN in an ExPEC iroA deletion mutant was able to restore biofilm formation. The contribution of IroN to biofilm formation appears to require a certain background found in ExPEC, but not in E. coli K12. This study identified the contribution of IroN to biofilm formation and highlights the multi-functional role of iron uptake systems in ExPEC.

  15. How we know it hurts: item analysis of written narratives reveals distinct neural responses to others' physical pain and emotional suffering.

    Directory of Open Access Journals (Sweden)

    Emile Bruneau

    Full Text Available People are often called upon to witness, and to empathize with, the pain and suffering of others. In the current study, we directly compared neural responses to others' physical pain and emotional suffering by presenting participants (n = 41 with 96 verbal stories, each describing a protagonist's physical and/or emotional experience, ranging from neutral to extremely negative. A separate group of participants rated "how much physical pain", and "how much emotional suffering" the protagonist experienced in each story, as well as how "vivid and movie-like" the story was. Although ratings of Pain, Suffering and Vividness were positively correlated with each other across stories, item-analyses revealed that each scale was correlated with activity in distinct brain regions. Even within regions of the "Shared Pain network" identified using a separate data set, responses to others' physical pain and emotional suffering were distinct. More broadly, item analyses with continuous predictors provided a high-powered method for identifying brain regions associated with specific aspects of complex stimuli - like verbal descriptions of physical and emotional events.

  16. Staphylokinase Control of Staphylococcus aureus Biofilm Formation and Detachment Through Host Plasminogen Activation.

    Science.gov (United States)

    Kwiecinski, Jakub; Peetermans, Marijke; Liesenborghs, Laurens; Na, Manli; Björnsdottir, Halla; Zhu, Xuefeng; Jacobsson, Gunnar; Johansson, Bengt R; Geoghegan, Joan A; Foster, Timothy J; Josefsson, Elisabet; Bylund, Johan; Verhamme, Peter; Jin, Tao

    2016-01-01

    Staphylococcus aureus biofilms, a leading cause of persistent infections, are highly resistant to immune defenses and antimicrobial therapies. In the present study, we investigated the contribution of fibrin and staphylokinase (Sak) to biofilm formation. In both clinical S. aureus isolates and laboratory strains, high Sak-producing strains formed less biofilm than strains that lacked Sak, suggesting that Sak prevents biofilm formation. In addition, Sak induced detachment of mature biofilms. This effect depended on plasminogen activation by Sak. Host-derived fibrin, the main substrate cleaved by Sak-activated plasminogen, was a major component of biofilm matrix, and dissolution of this fibrin scaffold greatly increased susceptibility of biofilms to antibiotics and neutrophil phagocytosis. Sak also attenuated biofilm-associated catheter infections in mouse models. In conclusion, our results reveal a novel role for Sak-induced plasminogen activation that prevents S. aureus biofilm formation and induces detachment of existing biofilms through proteolytic cleavage of biofilm matrix components.

  17. Distinct summer and winter bacterial communities in the active layer of Svalbard permafrost revealed by DNA- and RNA-based analyses

    DEFF Research Database (Denmark)

    Schostag, Morten; Stibal, Marek; Jacobsen, Carsten S.

    2015-01-01

    N) by co-extracting DNA and RNA from 12 soil cores collected monthly over a year. PCR amplicons of 16S rRNA genes (DNA) and reverse transcribed transcripts (cDNA) were quantified and sequenced to test for the effect of low winter temperature and seasonal variation in concentration of easily degradable...... organic matter on the bacterial communities. The copy number of 16S rRNA genes and transcripts revealed no distinct seasonal changes indicating potential bacterial activity during winter despite soil temperatures well below -10ºC. Multivariate statistical analysis of the bacterial diversity data (DNA......The active layer of soil overlaying permafrost in the Arctic is subjected to dramatic annual changes in temperature and soil chemistry, which likely affect bacterial activity and community structure. We studied seasonal variations in the bacterial community of active layer soil from Svalbard (78º...

  18. Molecular study on the carAB operon reveals that carB gene is required for swimming and biofilm formation in Xanthomonas citri subsp. citri.

    Science.gov (United States)

    Zhuo, Tao; Rou, Wei; Song, Xue; Guo, Jing; Fan, Xiaojing; Kamau, Gicharu Gibson; Zou, Huasong

    2015-10-23

    The carA and carB genes code the small and large subunits of carbamoyl-phosphate synthase (CPS) that responsible for arginine and pyrimidine production. The purpose of this work was to study the gene organization and expression pattern of carAB operon, and the biological functions of carA and carB genes in Xanthomonas citri subsp. citri. RT-PCR method was employed to identify the full length of carAB operon transcript in X. citri subsp. citri. The promoter of carAB operon was predicted and analyzed its activity by fusing a GUS reporter gene. The swimming motility was tested on 0.25% agar NY plates with 1% glucose. Biofilm was measured by cell adhesion to polyvinyl chloride 96-well plate. The results indicated that carAB operon was composed of five gene members carA-orf-carB-greA-rpfE. A single promoter was predicted from the nucleotide sequence upstream of carAB operon, and its sensitivity to glutamic acid, uracil and arginine was confirmed by fusing a GUS reporter gene. Deletion mutagenesis of carB gene resulted in reduced abilities in swimming on soft solid media and in forming biofilm on polystyrene microtiter plates. From these results, we concluded that carAB operon was involved in multiple biological processes in X. citri subsp. citri.

  19. In situ biosensing of the nanomechanical property and electrochemical spectroscopy of Streptococcus mutans-containing biofilms

    Science.gov (United States)

    Haochih Liu, Bernard; Li, Kun-Lin; Kang, Kai-Li; Huang, Wen-Ke; Liao, Jiunn-Der

    2013-07-01

    This work presents in situ biosensing approaches to study the nanomechanical and electrochemical behaviour of Streptococcus mutans biofilms under different cultivation conditions and microenvironments. The surface characteristics and sub-surface electrochemistry of the cell wall of S. mutans were measured by atomic force microscopy (AFM) based techniques to monitor the in situ biophysical status of biofilms under common anti-pathogenic procedures such as ultraviolet (UV) radiation and alcohol treatment. The AFM nanoindentation suggested a positive correlation between nanomechanical strength and the level of UV radiation of S. mutans; scanning impedance spectroscopy of dehydrated biofilms revealed reduced electrical resistance that is distinctive from that of living biofilms, which can be explained by the discharge of cytoplasm after alcohol treatment. Furthermore, the localized elastic moduli of four regions of the biofilm were studied: septum (Z-ring), cell wall, the interconnecting area between two cells and extracellular polymeric substance (EPS) area. The results indicated that cell walls exhibit the highest elastic modulus, followed by Z-ring, interconnect and EPS. Our approach provides an effective alternative for the characterization of the viability of living cells without the use of biochemical labelling tools such as fluorescence dyeing, and does not rely on surface binding or immobilization for detection. These AFM-based techniques can be very promising approaches when the conventional methods fall short.

  20. Fungal Biofilms: In Vivo Models for Discovery of Anti-Biofilm Drugs.

    Science.gov (United States)

    Nett, Jeniel E; Andes, David R

    2015-06-01

    During infection, fungi frequently transition to a biofilm lifestyle, proliferating as communities of surface-adherent aggregates of cells. Phenotypically, cells in a biofilm are distinct from free-floating cells. Their high tolerance of antifungals and ability to withstand host defenses are two characteristics that foster resilience. Biofilm infections are particularly difficult to eradicate, and most available antifungals have minimal activity. Therefore, the discovery of novel compounds and innovative strategies to treat fungal biofilms is of great interest. Although many fungi have been observed to form biofilms, the most well-studied is Candida albicans. Animal models have been developed to simulate common Candida device-associated infections, including those involving vascular catheters, dentures, urinary catheters, and subcutaneous implants. Models have also reproduced the most common mucosal biofilm infections: oropharyngeal and vaginal candidiasis. These models incorporate the anatomical site, immune components, and fluid dynamics of clinical niches and have been instrumental in the study of drug resistance and investigation of novel therapies. This chapter describes the significance of fungal biofilm infections, the animal models developed for biofilm study, and how these models have contributed to the development of new strategies for the eradication of fungal biofilm infections.

  1. Investigating biofilm structure developing on carriers from lab-scale moving bed biofilm reactors based on light microscopy and optical coherence tomography.

    Science.gov (United States)

    Li, Chunyan; Felz, Simon; Wagner, Michael; Lackner, Susanne; Horn, Harald

    2016-01-01

    This study focused on characterizing the structure of biofilms developed on carriers used in lab-scale moving bed biofilm reactors. Both light microscopy (2D) and optical coherence tomography (OCT) were employed to track the biofilm development on carriers of different geometry and under different aeration rates. Biofilm structure was further characterized with respect to average biofilm thickness, biofilm growth velocity, biomass volume, compartment filling degree, surface area, etc. The results showed that carriers with a smaller compartment size stimulated a quick establishment of biofilms. Low aeration rates favored fast development of biofilms. Comparison between the results derived from 2D and 3D images revealed comparable results with respect to average biofilm thickness and compartment filling degree before the carrier compartments were fully willed with biomass. However, 3D imaging with OCT was capable of visualizing and quantifying the heterogeneous structure of biofilms, which cannot be achieved using 2D imaging.

  2. Transcriptome comparison of global distinctive features between pollination and parthenocarpic fruit set reveals transcriptional phytohormone cross-talk in cucumber (Cucumis sativus L.).

    Science.gov (United States)

    Li, Ji; Wu, Zhe; Cui, Li; Zhang, Tinglin; Guo, Qinwei; Xu, Jian; Jia, Li; Lou, Qunfeng; Huang, Sanwen; Li, Zhengguo; Chen, Jinfeng

    2014-07-01

    Parthenocarpy is an important trait determining yield and quality of fruit crops. However, the understanding of the mechanisms underlying parthenocarpy induction is limited. Cucumber (Cucumis sativus L.) is abundant in parthenocarpic germplasm resources and is an excellent model organism for parthenocarpy studies. In this study, the transcriptome of cucumber fruits was studied using RNA sequencing (RNA-Seq). Differentially expressed genes (DEGs) of set fruits were compared against aborted fruits. Distinctive features of parthenocarpic and pollinated fruits were revealed by combining the analysis of the transcriptome together with cytomorphological and physiological analysis. Cell division and the transcription of cell division genes were found to be more active in parthenocarpic fruit. The study also indicated that parthenocarpic fruit set is a high sugar-consuming process which is achieved via enhanced carbohydrate degradation through transcription of genes that lead to the breakdown of carbohydrates. Furthermore, the evidence provided by this work supports a hypothesis that parthenocarpic fruit set is induced by mimicking the processes of pollination/fertilization at the transcriptional level, i.e. by performing the same transcriptional patterns of genes inducing pollination and gametophyte development as in pollinated fruit. Based on the RNA-Seq and ovary transient expression results, 14 genes were predicted as putative parthenocarpic genes. The transcription analysis of these candidate genes revealed auxin, cytokinin and gibberellin cross-talk at the transcriptional level during parthenocarpic fruit set.

  3. Analysis of metabolites in plasma reveals distinct metabolic features between Dahl salt-sensitive rats and consomic SS.13(BN) rats.

    Science.gov (United States)

    Wang, Le; Hou, Entai; Wang, Zhengjun; Sun, Na; He, Liqing; Chen, Lan; Liang, Mingyu; Tian, Zhongmin

    2014-07-18

    Salt-sensitive hypertension is a major risk factor for cardiovascular disorders. Our previous proteomic study revealed substantial differences in several proteins between Dahl salt-sensitive (SS) rats and salt-insensitive consomic SS.13(BN) rats. Subsequent experiments indicated a role of fumarase insufficiency in the development of hypertension in SS rats. In the present study, a global metabolic profiling study was performed using gas chromatography/mass spectrometry (GC/MS) in plasma of SS rats (n=9) and SS.13(BN) rats (n=8) on 0.4% NaCl diet, designed to gain further insights into the relationship between alterations in cellular intermediary metabolism and predisposition to hypertension. Principal component analysis of the data sets revealed a clear clustering and separation of metabolic profiles between SS rats and SS.13(BN) rats. 23 differential metabolites were identified (PSS rats. Pyruvate, which connects TCA cycle and glycolysis, was also increased in SS rats. Moreover, lower activity levels of fumarase, aconitase, α-ketoglutarate dehydrogenase and succinyl-CoA synthetase were detected in the heart, liver or skeletal muscles of SS rats. The distinct metabolic features in SS and SS.13(BN) rats indicate abnormalities of TCA cycle in SS rats, which may play a role in predisposing SS rats to developing salt-sensitive hypertension.

  4. Genetic Analyses and Simulations of Larval Dispersal Reveal Distinct Populations and Directional Connectivity across the Range of the Hawaiian Grouper (Epinephelus quernus

    Directory of Open Access Journals (Sweden)

    Malia Ana J. Rivera

    2011-01-01

    Full Text Available Integration of ecological and genetic data to study patterns of biological connectivity can aid in ecosystem-based management. Here we investigated connectivity of the Hawaiian grouper Epinephelus quernus, a species of management concern within the Main Hawaiian Islands (MHI, by comparing genetic analyses with simulated larval dispersal patterns across the species range in the Hawaiian Archipelago and Johnston Atoll. Larval simulations revealed higher dispersal from the MHI to the Northwestern Hawaiian Islands (NWHI than in the opposite direction and evidence for a dispersal corridor between Johnston and the middle of the Hawaiian Archipelago. Genetic analyses using mitochondrial DNA (mtDNA control region sequences and microsatellites revealed relatively high connectivity across the Hawaiian Archipelago, with the exception of genetically distinct populations and higher mtDNA diversity in the mid-Archipelago. These analyses support the preservation of the mid-archipelago as a source of genetic diversity and a region of connectivity with locations outside the Hawaiian Archipelago. Additionally, our evidence for directional dispersal away from the MHI lends caution to any management decisions that would rely on the NWHI replenishing depleted MHI stocks.

  5. RPL30 regulation of splicing reveals distinct roles for Cbp80 in U1 and U2 snRNP cotranscriptional recruitment.

    Science.gov (United States)

    Bragulat, Mireia; Meyer, Markus; Macías, Sara; Camats, Maria; Labrador, Mireia; Vilardell, Josep

    2010-10-01

    Pre-mRNA splicing is catalyzed by the spliceosome, and its control is essential for correct gene expression. While splicing repressors typically interfere with transcript recognition by spliceosomal components, the yeast protein L30 blocks spliceosomal rearrangements required for the engagement of U2 snRNP (small ribonucleoprotein particle) to its own transcript RPL30. Using a mutation in the RPL30 binding site that disrupts this repression, we have taken a genetic approach to reveal that regulation of splicing is restored in this mutant by deletion of the cap-binding complex (CBC) component Cbp80. Indeed, our data indicate that Cbp80 plays distinct roles in the recognition of the intron by U1 and U2 snRNP. It promotes the initial 5' splice site recognition by U1 and, independently, facilitates U2 recruitment, depending on sequences located in the vicinity of the 5' splice site. These results reveal a novel function for CBC in splicing and imply that these molecular events can be the target of a splicing regulator.

  6. The Biofilm Challenge

    DEFF Research Database (Denmark)

    Alhede, Maria; Alhede, Morten

    2014-01-01

    in wounds. However, the impact of biofilms is often debated, because infected wounds were also treated before the concept of biofilms was coined. In this short review, we will address the significance of biofilms and their role in wounds, and discuss the future tasks of the biofilm challenge....

  7. An expanded regulatory network temporally controls Candida albicans biofilm formation.

    Science.gov (United States)

    Fox, Emily P; Bui, Catherine K; Nett, Jeniel E; Hartooni, Nairi; Mui, Michael C; Andes, David R; Nobile, Clarissa J; Johnson, Alexander D

    2015-06-01

    Candida albicans biofilms are composed of highly adherent and densely arranged cells with properties distinct from those of free-floating (planktonic) cells. These biofilms are a significant medical problem because they commonly form on implanted medical devices, are drug resistant and are difficult to remove. C. albicans biofilms are not static structures; rather they are dynamic and develop over time. Here we characterize gene expression in biofilms during their development, and by comparing them to multiple planktonic reference states, we identify patterns of gene expression relevant to biofilm formation. In particular, we document time-dependent changes in genes involved in adhesion and metabolism, both of which are at the core of biofilm development. Additionally, we identify three new regulators of biofilm formation, Flo8, Gal4, and Rfx2, which play distinct roles during biofilm development over time. Flo8 is required for biofilm formation at all time points, and Gal4 and Rfx2 are needed for proper biofilm formation at intermediate time points.

  8. Global phylogenomic analysis of nonencapsulated Streptococcus pneumoniae reveals a deep-branching classic lineage that is distinct from multiple sporadic lineages.

    Science.gov (United States)

    Hilty, Markus; Wüthrich, Daniel; Salter, Susannah J; Engel, Hansjürg; Campbell, Samuel; Sá-Leão, Raquel; de Lencastre, Hermínia; Hermans, Peter; Sadowy, Ewa; Turner, Paul; Chewapreecha, Claire; Diggle, Mathew; Pluschke, Gerd; McGee, Lesley; Köseoğlu Eser, Özgen; Low, Donald E; Smith-Vaughan, Heidi; Endimiani, Andrea; Küffer, Marianne; Dupasquier, Mélanie; Beaudoing, Emmanuel; Weber, Johann; Bruggmann, Rémy; Hanage, William P; Parkhill, Julian; Hathaway, Lucy J; Mühlemann, Kathrin; Bentley, Stephen D

    2014-12-04

    The surrounding capsule of Streptococcus pneumoniae has been identified as a major virulence factor and is targeted by pneumococcal conjugate vaccines (PCV). However, nonencapsulated S. pneumoniae (non-Ec-Sp) have also been isolated globally, mainly in carriage studies. It is unknown if non-Ec-Sp evolve sporadically, if they have high antibiotic nonsusceptiblity rates and a unique, specific gene content. Here, whole-genome sequencing of 131 non-Ec-Sp isolates sourced from 17 different locations around the world was performed. Results revealed a deep-branching classic lineage that is distinct from multiple sporadic lineages. The sporadic lineages clustered with a previously sequenced, global collection of encapsulated S. pneumoniae (Ec-Sp) isolates while the classic lineage is comprised mainly of the frequently identified multilocus sequences types (STs) ST344 (n = 39) and ST448 (n = 40). All ST344 and nine ST448 isolates had high nonsusceptiblity rates to β-lactams and other antimicrobials. Analysis of the accessory genome reveals that the classic non-Ec-Sp contained an increased number of mobile elements, than Ec-Sp and sporadic non-Ec-Sp. Performing adherence assays to human epithelial cells for selected classic and sporadic non-Ec-Sp revealed that the presence of a integrative conjugative element (ICE) results in increased adherence to human epithelial cells (P = 0.005). In contrast, sporadic non-Ec-Sp lacking the ICE had greater growth in vitro possibly resulting in improved fitness. In conclusion, non-Ec-Sp isolates from the classic lineage have evolved separately. They have spread globally, are well adapted to nasopharyngeal carriage and are able to coexist with Ec-Sp. Due to continued use of PCV, non-Ec-Sp may become more prevalent.

  9. The in vivo biofilm

    DEFF Research Database (Denmark)

    Bjarnsholt, Thomas; Alhede, Maria; Alhede, Morten

    2013-01-01

    Bacteria can grow and proliferate either as single, independent cells or organized in aggregates commonly referred to as biofilms. When bacteria succeed in forming a biofilm within the human host, the infection often becomes very resistant to treatment and can develop into a chronic state. Biofilms...... have been studied for decades using various in vitro models, but it remains debatable whether such in vitro biofilms actually resemble in vivo biofilms in chronic infections. In vivo biofilms share several structural characteristics that differ from most in vitro biofilms. Additionally, the in vivo...... experimental time span and presence of host defenses differ from chronic infections and the chemical microenvironment of both in vivo and in vitro biofilms is seldom taken into account. In this review, we discuss why the current in vitro models of biofilms might be limited for describing infectious biofilms...

  10. Spindle assembly checkpoint genes reveal distinct as well as overlapping expression that implicates MDF-2/Mad2 in postembryonic seam cell proliferation in Caenorhabditis elegans.

    Science.gov (United States)

    Tarailo-Graovac, Maja; Wang, Jun; Chu, Jeffrey S C; Tu, Domena; Baillie, David L; Chen, Nansheng

    2010-09-21

    The spindle assembly checkpoint (SAC) delays anaphase onset by inhibiting the activity of the anaphase promoting complex/cyclosome (APC/C) until all of the kinetochores have properly attached to the spindle. The importance of SAC genes for genome stability is well established; however, the roles these genes play, during postembryonic development of a multicellular organism, remain largely unexplored. We have used GFP fusions of 5' upstream intergenic regulatory sequences to assay spatiotemporal expression patterns of eight conserved genes implicated in the spindle assembly checkpoint function in Caenorhabditis elegans. We have shown that regulatory sequences for all of the SAC genes drive ubiquitous GFP expression during early embryonic development. However, postembryonic spatial analysis revealed distinct, tissue-specific expression of SAC genes with striking co-expression in seam cells, as well as in the gut. Additionally, we show that the absence of MDF-2/Mad2 (one of the checkpoint genes) leads to aberrant number and alignment of seam cell nuclei, defects mainly attributed to abnormal postembryonic cell proliferation. Furthermore, we show that these defects are completely rescued by fzy-1(h1983)/CDC20, suggesting that regulation of the APC/CCDC20 by the SAC component MDF-2 is important for proper postembryonic cell proliferation. Our results indicate that SAC genes display different tissue-specific expression patterns during postembryonic development in C. elegans with significant co-expression in hypodermal seam cells and gut cells, suggesting that these genes have distinct as well as overlapping roles in postembryonic development that may or may not be related to their established roles in mitosis. Furthermore, we provide evidence, by monitoring seam cell lineage, that one of the checkpoint genes is required for proper postembryonic cell proliferation. Importantly, our research provides the first evidence that postembryonic cell division is more sensitive to

  11. Sequencing and functional annotation of avian pathogenic Escherichia coli serogroup O78 strains reveal the evolution of E. coli lineages pathogenic for poultry via distinct mechanisms.

    Science.gov (United States)

    Dziva, Francis; Hauser, Heidi; Connor, Thomas R; van Diemen, Pauline M; Prescott, Graham; Langridge, Gemma C; Eckert, Sabine; Chaudhuri, Roy R; Ewers, Christa; Mellata, Melha; Mukhopadhyay, Suman; Curtiss, Roy; Dougan, Gordon; Wieler, Lothar H; Thomson, Nicholas R; Pickard, Derek J; Stevens, Mark P

    2013-03-01

    Avian pathogenic Escherichia coli (APEC) causes respiratory and systemic disease in poultry. Sequencing of a multilocus sequence type 95 (ST95) serogroup O1 strain previously indicated that APEC resembles E. coli causing extraintestinal human diseases. We sequenced the genomes of two strains of another dominant APEC lineage (ST23 serogroup O78 strains χ7122 and IMT2125) and compared them to each other and to the reannotated APEC O1 sequence. For comparison, we also sequenced a human enterotoxigenic E. coli (ETEC) strain of the same ST23 serogroup O78 lineage. Phylogenetic analysis indicated that the APEC O78 strains were more closely related to human ST23 ETEC than to APEC O1, indicating that separation of pathotypes on the basis of their extraintestinal or diarrheagenic nature is not supported by their phylogeny. The accessory genome of APEC ST23 strains exhibited limited conservation of APEC O1 genomic islands and a distinct repertoire of virulence-associated loci. In light of this diversity, we surveyed the phenotype of 2,185 signature-tagged transposon mutants of χ7122 following intra-air sac inoculation of turkeys. This procedure identified novel APEC ST23 genes that play strain- and tissue-specific roles during infection. For example, genes mediating group 4 capsule synthesis were required for the virulence of χ7122 and were conserved in IMT2125 but absent from APEC O1. Our data reveal the genetic diversity of E. coli strains adapted to cause the same avian disease and indicate that the core genome of the ST23 lineage serves as a chassis for the evolution of E. coli strains adapted to cause avian or human disease via acquisition of distinct virulence genes.

  12. Phylogenomic and MALDI-TOF MS analysis of Streptococcus sinensis HKU4T reveals a distinct phylogenetic clade in the genus Streptococcus.

    Science.gov (United States)

    Teng, Jade L L; Huang, Yi; Tse, Herman; Chen, Jonathan H K; Tang, Ying; Lau, Susanna K P; Woo, Patrick C Y

    2014-10-20

    Streptococcus sinensis is a recently discovered human pathogen isolated from blood cultures of patients with infective endocarditis. Its phylogenetic position, as well as those of its closely related species, remains inconclusive when single genes were used for phylogenetic analysis. For example, S. sinensis branched out from members of the anginosus, mitis, and sanguinis groups in the 16S ribosomal RNA gene phylogenetic tree, but it was clustered with members of the anginosus and sanguinis groups when groEL gene sequences used for analysis. In this study, we sequenced the draft genome of S. sinensis and used a polyphasic approach, including concatenated genes, whole genomes, and matrix-assisted laser desorption ionization-time of flight mass spectrometry to analyze the phylogeny of S. sinensis. The size of the S. sinensis draft genome is 2.06 Mb, with GC content of 42.2%. Phylogenetic analysis using 50 concatenated genes or whole genomes revealed that S. sinensis formed a distinct cluster with Streptococcus oligofermentans and Streptococcus cristatus, and these three streptococci were clustered with the "sanguinis group." As for phylogenetic analysis using hierarchical cluster analysis of the mass spectra of streptococci, S. sinensis also formed a distinct cluster with S. oligofermentans and S. cristatus, but these three streptococci were clustered with the "mitis group." On the basis of the findings, we propose a novel group, named "sinensis group," to include S. sinensis, S. oligofermentans, and S. cristatus, in the Streptococcus genus. Our study also illustrates the power of phylogenomic analyses for resolving ambiguities in bacterial taxonomy.

  13. Antimicrobial effect of diallyl sulphide on Campylobacter jejuni biofilms

    Science.gov (United States)

    Lu, Xiaonan; Samuelson, Derrick R.; Rasco, Barbara A.; Konkel, Michael E.

    2012-01-01

    Objectives Bacterial biofilms pose significant food safety risks because of their attachment to fomites and food surfaces, including fresh produce surfaces. The purpose of this study was to systematically investigate the activity of selected antimicrobials on Campylobacter jejuni biofilms. Methods C. jejuni biofilms and planktonic cells were treated with ciprofloxacin, erythromycin and diallyl sulphide and examined using infrared and Raman spectroscopies coupled with imaging analysis. Results Diallyl sulphide eliminated planktonic cells and sessile cells in biofilms at a concentration that was at least 100-fold less than used for either ciprofloxacin or erythromycin on the basis of molarity. Distinct cell lysis was observed in diallyl sulphide-treated planktonic cells using immunoblot analysis and was confirmed by a rapid decrease in cellular ATP. Two phases of C. jejuni biofilm recalcitrance modes against ciprofloxacin and erythromycin were validated using vibrational spectroscopies: (i) an initial hindered adsorption into biofilm extracellular polymeric substance (EPS) and delivery of antibiotics to sessile cells within biofilms; and (ii) a different interaction between sessile cells in a biofilm compared with their planktonic counterparts. Diallyl sulphide destroyed the EPS structure of the C. jejuni biofilm, after which the sessile cells were killed in a similar manner as planktonic cells. Spectroscopic models can predict the survival of sessile cells within biofilms. Conclusions Diallyl sulphide elicits strong antimicrobial activity against planktonic and sessile C. jejuni and may have applications for reducing the prevalence of this microbe in foods, biofilm reduction and, potentially, as an alternative chemotherapeutic agent for multidrug-resistant bacterial strains. PMID:22550133

  14. Mesoscale Elucidation of Biofilm Shear Behavior

    CERN Document Server

    Barai, Pallab; Mukherjee, Partha P

    2015-01-01

    Formation of bacterial colonies as biofilm on the surface/interface of various objects has the potential to impact not only human health and disease but also energy and environmental considerations. Biofilms can be regarded as soft materials, and comprehension of their shear response to external forces is a key element to the fundamental understanding. A mesoscale model has been presented in this article based on digitization of a biofilm microstructure. Its response under externally applied shear load is analyzed. Strain stiffening type behavior is readily observed under high strain loads due to the unfolding of chains within soft polymeric substrate. Sustained shear loading of the biofilm network results in strain localization along the diagonal direction. Rupture of the soft polymeric matrix can potentially reduce the intercellular interaction between the bacterial cells. Evolution of stiffness within the biofilm network under shear reveals two regions: a) initial increase in stiffness due to strain stiffe...

  15. Biofilm formation by the emerging fungal pathogen Trichosporon asahii: development, architecture, and antifungal resistance.

    Science.gov (United States)

    Di Bonaventura, Giovanni; Pompilio, Arianna; Picciani, Carla; Iezzi, Manuela; D'Antonio, Domenico; Piccolomini, Raffaele

    2006-10-01

    Trichosporon asahii is the most common cause of fatal disseminated trichosporonosis, frequently associated with indwelling medical devices. Despite the use of antifungal drugs to treat trichosporonosis, infection is often persistent and is associated with high mortality. This drove our interest in evaluating the capability of T. asahii to form a biofilm on biomaterial-representative polystyrene surfaces through the development and optimization of a reproducible T. asahii-associated biofilm model. Time course analyses of viable counts and a formazan salt reduction assay, as well as microscopy studies, revealed that biofilm formation by T. asahii occurred in an organized fashion through four distinct developmental phases: initial adherence of yeast cells (0 to 2 h), germination and microcolony formation (2 to 4 h), filamentation (4 to 6 h), and proliferation and maturation (24 to 72 h). Scanning electron microscopy and confocal scanning laser microscopy revealed that mature T. asahii biofilms (72-h) displayed a complex, heterogeneous three-dimensional structure, consisting of a dense network of metabolically active yeast cells and hyphal elements completely embedded within exopolymeric material. Antifungal susceptibility testing demonstrated a remarkable rise in the MICs of sessile T. asahii cells against clinically used amphotericin B, caspofungin, voriconazole, and fluconazole compared to their planktonic counterparts. In particular, T. asahii biofilms were up to 16,000 times more resistant to voriconazole, the most active agent against planktonic cells (MIC, 0.06 microg/ml). Our results suggest that the ability of T. asahii to form a biofilm may be a major factor in determining persistence of the infection in spite of in vitro susceptibility of clinical isolates.

  16. Outcome measures for hand function naturally reveal three distinct domains in older adults: strength, coordinated upper extremity function, and sensorimotor processing

    Directory of Open Access Journals (Sweden)

    Emily L Lawrence

    2015-06-01

    Full Text Available Understanding the mapping between individual outcome measures and the latent functional domains of interest is critical to a quantitative evaluation and rehabilitation of hand function. We examined whether and how the associations among six hand-specific outcome measures reveal latent functional domains in elderly individuals. We asked 66 healthy older adult participants (38F, 28M, 66.1±11.6yrs, range: 45-88yrs and 33 older adults (65.8±9.7yrs, 44-81yrs, 51 hands diagnosed with osteoarthritis (OA of the carpometacarpal (CMC joint, to complete six functional assessments: hand strength (Grip, Key and Precision Pinch, Box and Block, Nine Hole Pegboard, and Strength-Dexterity tests. The first three principal components suffice to explain 86% of variance among the six outcome measures in healthy older adults, and 84% of variance in older adults with CMC OA. The composition of these dominant associations revealed three distinct latent functional domains: strength, coordinated upper extremity function, and sensorimotor processing. Furthermore, in participants with thumb CMC OA we found a blurring of the associations between the latent functional domains of strength and coordinated upper extremity function. This motivates future work to understand how the physiological effects of thumb CMC OA lead upper extremity coordination to become strongly associated with strength, while dynamic sensorimotor ability remains an independent functional domain.Thus, when assessing the level of hand function in our growing older adult populations, it is particularly important to acknowledge its multidimensional nature—and explicitly consider how each outcome measure maps to these three latent and fundamental domains of function. Moreover, this ability to distinguish among latent functional domains may facilitate the design of treatment modalities to target the rehabilitation of each of them.

  17. Distinct summer and winter bacterial communities in the active layer of Svalbard permafrost revealed by DNA- and RNA-based analyses

    Energy Technology Data Exchange (ETDEWEB)

    Schostag, Morten; Stibal, Marek; Jacobsen, Carsten S.; Baelum, Jacob; Tas, Neslihan; Elberling, Bo; Jansson, Janet K.; Semenchuk, Phillip; Prieme, Anders

    2015-04-30

    The active layer of soil overlaying permafrost in the Arctic is subjected to dramatic annual changes in temperature and soil chemistry, which likely affect bacterial activity and community structure. We studied seasonal variations in the bacterial community of active layer soil from Svalbard (78°N) by co-extracting DNA and RNA from 12 soil cores collected monthly over a year. PCR amplicons of 16S rRNA genes (DNA) and reverse transcribed transcripts (cDNA) were quantified and sequenced to test for the effect of low winter temperature and seasonal variation in concentration of easily degradable organic matter on the bacterial communities. The copy number of 16S rRNA genes and transcripts revealed no distinct seasonal changes indicating potential bacterial activity during winter despite soil temperatures well below -10°C. Multivariate statistical analysis of the bacterial diversity data (DNA and cDNA libraries) revealed a season-based clustering of the samples, and, e.g., the relative abundance of potentially active Cyanobacteria peaked in June and Alphaproteobacteria increased over the summer and then declined from October to November. The structure of the bulk (DNA-based) community was significantly correlated with pH and dissolved organic carbon, while the potentially active (RNA-based) community structure was not significantly correlated with any of the measured soil parameters. A large fraction of the 16S rRNA transcripts was assigned to nitrogen-fixing bacteria (up to 24% in June) and phototrophic organisms (up to 48% in June) illustrating the potential importance of nitrogen fixation in otherwise nitrogen poor Arctic ecosystems and of phototrophic bacterial activity on the soil surface.

  18. Phylogenetic analysis of spring virema of carp virus reveals distinct subgroups with common origins for recent isolates in North America and the UK.

    Science.gov (United States)

    Miller, O; Fuller, F J; Gebreyes, W A; Lewbart, G A; Shchelkunov, I S; Shivappa, R B; Joiner, C; Woolford, G; Stone, D M; Dixon, P F; Raley, M E; Levine, J F

    2007-07-16

    Genetic relationships between 35 spring viremia of carp virus (SVCV) genogroup Ia isolates were determined based on the nucleotide sequences of the phosphoprotein (P) gene and glycoprotein (G) genes. Phylogenetic analysis based on P gene sequences revealed 2 distinct subgroups within SVCV genogroup Ia, designated SVCV Iai and Iaii, and suggests at least 2 independent introductions of the virus into the USA in 2002. Combined P- and G-sequence data support the emergence of SVCV in Illinois, USA, and in Lake Ontario, Canada, from the initial outbreak in Wisconsin, USA, and demonstrate a close genetic link to viruses isolated during routine import checks on fish brought into the UK from Asia. The data also showed a genetic link between SVCV isolations made in Missouri and Washington, USA, in 2004 and the earlier isolation made in North Carolina, USA, in 2002. However, based on the close relationship to a 2004 UK isolate, the data suggest than the Washington isolate represents a third introduction into the US from a common source, rather than a reemergence from the 2002 isolate. There was strong phylogenetic support for an Asian origin for 9 of 16 UK viruses isolated either from imported fish, or shown to have been in direct contact with fish imported from Asia. In one case, there was 100% nucleotide identity in the G-gene with a virus isolated in China.

  19. Incomplete deletion of IL-4Rα by LysM(Cre reveals distinct subsets of M2 macrophages controlling inflammation and fibrosis in chronic schistosomiasis.

    Directory of Open Access Journals (Sweden)

    Kevin M Vannella

    2014-09-01

    Full Text Available Mice expressing a Cre recombinase from the lysozyme M-encoding locus (Lyz2 have been widely used to dissect gene function in macrophages and neutrophils. Here, we show that while naïve resident tissue macrophages from IL-4Rαf(lox/deltaLysM(Cre mice almost completely lose IL-4Rα function, a large fraction of macrophages elicited by sterile inflammatory stimuli, Schistosoma mansoni eggs, or S. mansoni infection, fail to excise Il4rα. These F4/80(hiCD11b(hi macrophages, in contrast to resident tissue macrophages, express lower levels of Lyz2 explaining why this population resists LysM(Cre-mediated deletion. We show that in response to IL-4 and IL-13, Lyz2(loIL-4Rα(+ macrophages differentiate into an arginase 1-expressing alternatively-activated macrophage (AAM population, which slows the development of lethal fibrosis in schistosomiasis. In contrast, we identified Lyz2(hiIL-4Rα(+ macrophages as the key subset of AAMs mediating the downmodulation of granulomatous inflammation in chronic schistosomiasis. Our observations reveal a limitation on using a LysMCre mouse model to study gene function in inflammatory settings, but we utilize this limitation as a means to demonstrate that distinct populations of alternatively activated macrophages control inflammation and fibrosis in chronic schistosomiasis.

  20. Incomplete deletion of IL-4Rα by LysM(Cre) reveals distinct subsets of M2 macrophages controlling inflammation and fibrosis in chronic schistosomiasis.

    Science.gov (United States)

    Vannella, Kevin M; Barron, Luke; Borthwick, Lee A; Kindrachuk, Kristen N; Narasimhan, Prakash Babu; Hart, Kevin M; Thompson, Robert W; White, Sandra; Cheever, Allen W; Ramalingam, Thirumalai R; Wynn, Thomas A

    2014-09-01

    Mice expressing a Cre recombinase from the lysozyme M-encoding locus (Lyz2) have been widely used to dissect gene function in macrophages and neutrophils. Here, we show that while naïve resident tissue macrophages from IL-4Rαf(lox/delta)LysM(Cre) mice almost completely lose IL-4Rα function, a large fraction of macrophages elicited by sterile inflammatory stimuli, Schistosoma mansoni eggs, or S. mansoni infection, fail to excise Il4rα. These F4/80(hi)CD11b(hi) macrophages, in contrast to resident tissue macrophages, express lower levels of Lyz2 explaining why this population resists LysM(Cre)-mediated deletion. We show that in response to IL-4 and IL-13, Lyz2(lo)IL-4Rα(+) macrophages differentiate into an arginase 1-expressing alternatively-activated macrophage (AAM) population, which slows the development of lethal fibrosis in schistosomiasis. In contrast, we identified Lyz2(hi)IL-4Rα(+) macrophages as the key subset of AAMs mediating the downmodulation of granulomatous inflammation in chronic schistosomiasis. Our observations reveal a limitation on using a LysMCre mouse model to study gene function in inflammatory settings, but we utilize this limitation as a means to demonstrate that distinct populations of alternatively activated macrophages control inflammation and fibrosis in chronic schistosomiasis.

  1. Dissecting the social brain: Introducing the EmpaToM to reveal distinct neural networks and brain-behavior relations for empathy and Theory of Mind.

    Science.gov (United States)

    Kanske, Philipp; Böckler, Anne; Trautwein, Fynn-Mathis; Singer, Tania

    2015-11-15

    Successful social interactions require both affect sharing (empathy) and understanding others' mental states (Theory of Mind, ToM). As these two functions have mostly been investigated in isolation, the specificity of the underlying neural networks and the relation of these networks to the respective behavioral indices could not be tested. Here, we present a novel fMRI paradigm (EmpaToM) that independently manipulates both empathy and ToM. Experiments 1a/b (N=90) validated the task with established empathy and ToM paradigms on a behavioral and neural level. Experiment 2 (N=178) employed the EmpaToM and revealed clearly separable neural networks including anterior insula for empathy and ventral temporoparietal junction for ToM. These distinct networks could be replicated in task-free resting state functional connectivity. Importantly, brain activity in these two networks specifically predicted the respective behavioral indices, that is, inter-individual differences in ToM related brain activity predicted inter-individual differences in ToM performance, but not empathic responding, and vice versa. Taken together, the validated EmpaToM allows separation of affective and cognitive routes to understanding others. It may thus benefit future clinical, developmental, and intervention studies on identifying selective impairments and improvement in specific components of social cognition.

  2. A comparative study on Ca content and distribution in two Gesneriaceae species reveals distinctive mechanisms to cope with high rhizospheric soluble calcium.

    Science.gov (United States)

    Li, Wenlong; Xu, Falun; Chen, Shixuan; Zhang, Zhennan; Zhao, Yan; Jin, Yukuan; Li, Meijing; Zhu, Yan; Liu, Yongxiu; Yang, Yi; Deng, Xin

    2014-01-01

    Excessive Ca is toxic to plants thus significantly affects plant growth and species distribution in Ca-rich karst areas. To understand how plants survive high Ca soil, laboratory experiments were established to compare the physiological responses and internal Ca distribution in organ, tissue, cell, and intracellular levels under different Ca levels for Lysionotus pauciflorus and Boea hygrometrica, two karst habitant Gesneriaceae species in Southwest China. In the controlled condition, L. pauciflorus could survive as high as 200 mM rhizospheric soluble Ca, attributed to a series of physiological responses and preferential storage that limited Ca accumulation in chloroplasts of palisade cells. In contrast, B. hygrometrica could survive only 20 mM rhizospheric soluble Ca, but accumulated a high level of internal Ca in both palisade and spongy cells without disturbance on photosynthetic activity. By phenotype screening of transgenic plants expressing high Ca-inducible genes from B. hygrometrica, the expression of BhDNAJC2 in A. thaliana was found to enhance plant growth and photosynthesis under high soluble Ca stress. BhDNAJC2 encodes a recently reported heat shock protein (HSP) 40 family DnaJ-domain protein. The Ca-resistant phenotype of BhDNAJC2 highlights the important role of chaperone-mediated protein quality control in Ca tolerance in B. hygrometrica. Taken together, our results revealed that distinctive mechanisms were employed in the two Gesneriaceae karst habitants to cope with a high Ca environment.

  3. A comparative study on Ca content and distribution in two Gesneriaceae species reveals distinctive mechanisms to cope with high rhizospheric soluble calcium

    Directory of Open Access Journals (Sweden)

    Wenlong eLi

    2014-11-01

    Full Text Available Excessive Ca is toxic to plants thus significantly affects plant growth and species distribution in Ca-rich karst areas. To understand how plants survive high Ca soil, laboratory experiments were established to compare the physiological responses and internal Ca distribution in organ, tissue, cell and intracellular levels under different Ca levels for Lysionotus pauciflorus and Boea hygrometrica, two karst habitant Gesneriaceae species in Southwest China. In the controlled condition, L. pauciflorus could survive as high as 200 mM rhizospheric soluble Ca, attributed to a series of physiological responses and preferential storage that limited Ca accumulation in chloroplasts of palisade cells. In contrast, B. hygrometrica could survive only 20 mM rhizospheric soluble Ca, but accumulated a high level of internal Ca in both palisade and spongy cells without disturbance on photosynthetic activity. By phenotype screening of transgenic plants expressing high Ca-inducible genes from B. hygrometrica, the expression of BhDNAJC2 in A. thaliana was found to enhance plant growth and photosynthesis under high soluble Ca stress. BhDNAJC2 encodes a recently reported heat shock protein (HSP 40 family DnaJ-domain protein. The Ca-resistant phenotype of BhDNAJC2 highlights the important role of chaperone-mediated protein quality control in Ca tolerance in B. hygrometrica. Taken together, our results revealed that distinctive mechanisms were employed in the two Gesneriaceae karst habitants to cope with a high Ca environment.

  4. Putative Role of β-1,3 Glucans in Candida albicans Biofilm Resistance▿

    OpenAIRE

    Nett, Jeniel; Lincoln, Leslie; Marchillo, Karen; Massey, Randall; Holoyda, Kathleen; Hoff, Brian; VanHandel, Michelle; Andes, David

    2006-01-01

    Biofilms are microbial communities, embedded in a polymeric matrix, growing attached to a surface. Nearly all device-associated infections involve growth in the biofilm life style. Biofilm communities have characteristic architecture and distinct phenotypic properties. The most clinically important phenotype involves extraordinary resistance to antimicrobial therapy, making biofilm infections very difficulty to cure without device removal. The current studies examine drug resistance in Candid...

  5. A new biofilm-associated colicin with increased efficiency against biofilm bacteria

    Science.gov (United States)

    Rendueles, Olaya; Beloin, Christophe; Latour-Lambert, Patricia; Ghigo, Jean-Marc

    2014-01-01

    Formation of bacterial biofilm communities leads to profound physiological modifications and increased physical and metabolic exchanges between bacteria. It was previously shown that bioactive molecules produced within the biofilm environment contribute to bacterial interactions. Here we describe new pore-forming colicin R, specifically produced in biofilms formed by the natural isolate Escherichia coli ROAR029 but that cannot be detected under planktonic culture conditions. We demonstrate that an increased SOS stress response within mature biofilms induces SOS-dependent colicin R expression. We provide evidence that colicin R displays increased activity against E. coli strains that have a reduced lipopolysaccharide length, such as the pathogenic enteroaggregative E. coli LF82 clinical isolate, therefore pointing to lipopolysaccharide size as an important determinant for resistance to colicins. We show that colicin R toxicity toward E. coli LF82 is increased under biofilm conditions compared with planktonic susceptibility and that release of colicin R confers a strong competitive advantage in mixed biofilms by rapidly outcompeting sensitive neighboring bacteria. This work identifies the first biofilm-associated colicin that preferentially targets biofilm bacteria. Furthermore, it indicates that the study of antagonistic molecules produced in biofilm and multispecies contexts could reveal unsuspected, ecologically relevant bacterial interactions influencing population dynamics in natural environments. PMID:24451204

  6. Selective reactivity of monochloramine with extracellular matrix components affects the disinfection of biofilm and detached clusters.

    Science.gov (United States)

    Xue, Zheng; Lee, Woo Hyoung; Coburn, Kimberly M; Seo, Youngwoo

    2014-04-01

    The efficiency of monochloramine disinfection was dependent on the quantity and composition of extracellular polymeric substances (EPS) in biofilms, as monochloramine has a selective reactivity with proteins over polysaccharides. Biofilms with protein-based (Pseudomonas putida) and polysaccharide based EPS (Pseudomonas aeruginosa), as well as biofilms with varied amount of polysaccharide EPS (wild-type and mutant P. aeruginosa), were compared. The different reactivity of EPS components with monochloramine influenced disinfectant penetration, biofilm inactivation, as well as the viability of detached clusters. Monochloramine transport profiling measured by a chloramine-sensitive microelectrode revealed a broader diffusion boundary layer between bulk and biofilm surface in the P. putida biofilm compared to those of P. aeruginosa biofilms. The reaction with proteins in P. putida EPS multiplied both the time and the monochloramine mass required to achieve a full biofilm penetration. Cell viability in biofilms was also spatially influenced by monochloramine diffusion and reaction within biofilms, showing a lower survival in the surface section and a higher persistence in the middle section of the P. putida biofilm compared to the P. aeruginosa biofilms. While polysaccharide EPS promoted biofilm cell viability by obstructing monochloramine reactive sites on bacterial cells, protein EPS hindered monochloramine penetration by reacting with monochloramine and reduced its concentration within biofilms. Furthermore, the persistence of bacterial cells detached from biofilm (over 70% for P. putida and ∼40% for polysaccharide producing P. aeruginosa) suggested that currently recommended monochloramine residual levels may underestimate the risk of water quality deterioration caused by biofilm detachment.

  7. The complete chloroplast DNA sequence of the green alga Oltmannsiellopsis viridis reveals a distinctive quadripartite architecture in the chloroplast genome of early diverging ulvophytes

    Directory of Open Access Journals (Sweden)

    Lemieux Claude

    2006-02-01

    Full Text Available Abstract Background The phylum Chlorophyta contains the majority of the green algae and is divided into four classes. The basal position of the Prasinophyceae has been well documented, but the divergence order of the Ulvophyceae, Trebouxiophyceae and Chlorophyceae is currently debated. The four complete chloroplast DNA (cpDNA sequences presently available for representatives of these classes have revealed extensive variability in overall structure, gene content, intron composition and gene order. The chloroplast genome of Pseudendoclonium (Ulvophyceae, in particular, is characterized by an atypical quadripartite architecture that deviates from the ancestral type by a large inverted repeat (IR featuring an inverted rRNA operon and a small single-copy (SSC region containing 14 genes normally found in the large single-copy (LSC region. To gain insights into the nature of the events that led to the reorganization of the chloroplast genome in the Ulvophyceae, we have determined the complete cpDNA sequence of Oltmannsiellopsis viridis, a representative of a distinct, early diverging lineage. Results The 151,933 bp IR-containing genome of Oltmannsiellopsis differs considerably from Pseudendoclonium and other chlorophyte cpDNAs in intron content and gene order, but shares close similarities with its ulvophyte homologue at the levels of quadripartite architecture, gene content and gene density. Oltmannsiellopsis cpDNA encodes 105 genes, contains five group I introns, and features many short dispersed repeats. As in Pseudendoclonium cpDNA, the rRNA genes in the IR are transcribed toward the single copy region featuring the genes typically found in the ancestral LSC region, and the opposite single copy region harbours genes characteristic of both the ancestral SSC and LSC regions. The 52 genes that were transferred from the ancestral LSC to SSC region include 12 of those observed in Pseudendoclonium cpDNA. Surprisingly, the overall gene organization of

  8. Biofilm Fixed Film Systems

    Directory of Open Access Journals (Sweden)

    Dipesh Das

    2011-09-01

    Full Text Available The work reviewed here was published between 2008 and 2010 and describes research that involved aerobic and anoxic biofilm treatment of water pollutants. Biofilm denitrification systems are covered when appropriate. References catalogued here are divided on the basis of fundamental research area or reactor types. Fundamental research into biofilms is presented in two sections, Biofilm Measurement and Characterization and Growth and Modeling. The reactor types covered are: trickling filters, rotating biological contactors, fluidized bed bioreactors, submerged bed biofilm reactors, biological granular activated carbon, membrane bioreactors, and immobilized cell reactors. Innovative reactors, not easily classified, are then presented, followed by a section on biofilms on sand, soil and sediment.

  9. Characterization of temporal protein production in Pseudomonas aeruginosa biofilms.

    Science.gov (United States)

    Southey-Pillig, Christopher J; Davies, David G; Sauer, Karin

    2005-12-01

    Phenotypic and genetic evidence supporting the notion of biofilm formation as a developmental process is growing. In the present work, we provide additional support for this hypothesis by identifying the onset of accumulation of biofilm-stage specific proteins during Pseudomonas aeruginosa biofilm maturation and by tracking the abundance of these proteins in planktonic and three biofilm developmental stages. The onset of protein production was found to correlate with the progression of biofilms in developmental stages. Protein identification revealed that proteins with similar function grouped within similar protein abundance patterns. Metabolic and housekeeping proteins were found to group within a pattern separate from virulence, antibiotic resistance, and quorum-sensing-related proteins. The latter were produced in a progressive manner, indicating that attendant features that are characteristic of biofilms such as antibiotic resistance and virulence may be part of the biofilm developmental process. Mutations in genes for selected proteins from several protein production patterns were made, and the impact of these mutations on biofilm development was evaluated. The proteins cytochrome c oxidase, a probable chemotaxis transducer, a two-component response regulator, and MexH were produced only in mature and late-stage biofilms. Mutations in the genes encoding these proteins did not confer defects in growth, initial attachment, early biofilm formation, or twitching motility but were observed to arrest biofilm development at the stage of cell cluster formation we call the maturation-1 stage. The results indicated that expression of theses genes was required for the progression of biofilms into three-dimensional structures on abiotic surfaces and the completion of the biofilm developmental cycle. Reverse transcription-PCR analysis confirmed the detectable change in expression of the respective genes ccoO, PA4101, and PA4208. We propose a possible mechanism for the

  10. Essential factors of an integrated moving bed biofilm reactor-membrane bioreactor: Adhesion characteristics and microbial community of the biofilm.

    Science.gov (United States)

    Tang, Bing; Yu, Chunfei; Bin, Liying; Zhao, Yiliang; Feng, Xianfeng; Huang, Shaosong; Fu, Fenglian; Ding, Jiewei; Chen, Cuiqun; Li, Ping; Chen, Qianyu

    2016-07-01

    This work aims at revealing the adhesion characteristics and microbial community of the biofilm in an integrated moving bed biofilm reactor-membrane bioreactor, and further evaluating their variations over time. With multiple methods, the adhesion characteristics and microbial community of the biofilm on the carriers were comprehensively illuminated, which showed their dynamic variation along with the operational time. Results indicated that: (1) the roughness of biofilm on the carriers increased very quickly to a maximum value at the start-up stage, then, decreased to become a flat curve, which indicated a layer of smooth biofilm formed on the surface; (2) the tightly-bound protein and polysaccharide was the most important factor influencing the stability of biofilm; (3) the development of biofilm could be divided into three stages, and Gammaproteobacteria were the most dominant microbial species in class level at the last stage, which occupied the largest ratio (51.48%) among all microbes.

  11. Characteristics of biofilm attaching to carriers in moving bed biofilm reactor used to treat vitamin C wastewater.

    Science.gov (United States)

    Hu, Xiao-bing; Xu, Ke; Wang, Zhao; Ding, Li-li; Ren, Hong-qiang

    2013-01-01

    In order to investigate characteristics of biofilm attaching firmly to carriers in the moving bed biofilm reactor (MBBR) used for vitamin C wastewater treatment, experiments were undertaken with instrumental analysis methods. Scanning electron microscopy (SEM) micrographs of MBBR biofilms revealed that there were rod-shaped microbes and cocci in the biofilm, and microbes were embedded within medium substances and the biofilm matrix adhered firmly to carriers, leading to the formation of a smooth compacted surface at the base of the biofilm. Transmission electron microscopy (TEM) analysis revealed that extracellular polymeric substances (EPS) layer surrounded cell, sequestered inorganics to form a mixed structure, which ensured firm attachment of the biofilm to the carrier. X-ray diffraction (XRD) experiments and thermogravimetry analysis revealed that (i) the biofilm contained many inorganic substances, about 70.5%, and the inorganic substances contained multiple classes of inorganic with a high boiling point; (ii) inorganic elements such as calcium and phosphorous were selectively absorbed and accumulated in the biofilm as insoluble compounds with amorphous phases, rendering the biofilm highly resistant to detachment. Fourier-transform infrared (FTIR) spectroscopy showed carbohydrates were the main EPS.

  12. Influence of ROBO1 and RORA on risk of age-related macular degeneration reveals genetically distinct phenotypes in disease pathophysiology.

    Directory of Open Access Journals (Sweden)

    Gyungah Jun

    Full Text Available ROBO1 is a strong candidate gene for age-related macular degeneration (AMD based upon its location under a linkage peak on chromosome 3p12, its expression pattern, and its purported function in a pathway that includes RORA, a gene previously associated with risk for neovascular AMD. Previously, we observed that expression of ROBO1 and RORA is down-regulated among wet AMD cases, as compared to their unaffected siblings. Thus, we hypothesized that contribution of association signals in ROBO1, and interaction between these two genes may be important for both wet and dry AMD. We evaluated association of 19 single nucleotide polymorphisms (SNPs in ROBO1 with wet and dry stages of AMD in a sibling cohort and a Greek case-control cohort containing 491 wet AMD cases, 174 dry AMD cases and 411 controls. Association signals and interaction results were replicated in an independent prospective cohort (1070 controls, 164 wet AMD cases, 293 dry AMD cases. The most significantly associated ROBO1 SNPs were rs1387665 under an additive model (meta P = 0.028 for wet AMD and rs9309833 under a recessive model (meta P = 6 × 10(-4 for dry AMD. Further analyses revealed interaction between ROBO1 rs9309833 and RORA rs8034864 for both wet and dry AMD (interaction P<0.05. These studies were further supported by whole transcriptome expression profile studies from 66 human donor eyes and chromatin immunoprecipitation assays from mouse retinas. These findings suggest that distinct ROBO1 variants may influence the risk of wet and dry AMD, and the effects of ROBO1 on AMD risk may be modulated by RORA variants.

  13. Genetic Structuration, Demography and Evolutionary History of Mycobacterium tuberculosis LAM9 Sublineage in the Americas as Two Distinct Subpopulations Revealed by Bayesian Analyses.

    Science.gov (United States)

    Reynaud, Yann; Millet, Julie; Rastogi, Nalin

    2015-01-01

    Tuberculosis (TB) remains broadly present in the Americas despite intense global efforts for its control and elimination. Starting from a large dataset comprising spoligotyping (n = 21183 isolates) and 12-loci MIRU-VNTRs data (n = 4022 isolates) from a total of 31 countries of the Americas (data extracted from the SITVIT2 database), this study aimed to get an overview of lineages circulating in the Americas. A total of 17119 (80.8%) strains belonged to the Euro-American lineage 4, among which the most predominant genotypic family belonged to the Latin American and Mediterranean (LAM) lineage (n = 6386, 30.1% of strains). By combining classical phylogenetic analyses and Bayesian approaches, this study revealed for the first time a clear genetic structuration of LAM9 sublineage into two subpopulations named LAM9C1 and LAM9C2, with distinct genetic characteristics. LAM9C1 was predominant in Chile, Colombia and USA, while LAM9C2 was predominant in Brazil, Dominican Republic, Guadeloupe and French Guiana. Globally, LAM9C2 was characterized by higher allelic richness as compared to LAM9C1 isolates. Moreover, LAM9C2 sublineage appeared to expand close to twenty times more than LAM9C1 and showed older traces of expansion. Interestingly, a significant proportion of LAM9C2 isolates presented typical signature of ancestral LAM-RDRio MIRU-VNTR type (224226153321). Further studies based on Whole Genome Sequencing of LAM strains will provide the needed resolution to decipher the biogeographical structure and evolutionary history of this successful family.

  14. Genetic Structuration, Demography and Evolutionary History of Mycobacterium tuberculosis LAM9 Sublineage in the Americas as Two Distinct Subpopulations Revealed by Bayesian Analyses.

    Directory of Open Access Journals (Sweden)

    Yann Reynaud

    Full Text Available Tuberculosis (TB remains broadly present in the Americas despite intense global efforts for its control and elimination. Starting from a large dataset comprising spoligotyping (n = 21183 isolates and 12-loci MIRU-VNTRs data (n = 4022 isolates from a total of 31 countries of the Americas (data extracted from the SITVIT2 database, this study aimed to get an overview of lineages circulating in the Americas. A total of 17119 (80.8% strains belonged to the Euro-American lineage 4, among which the most predominant genotypic family belonged to the Latin American and Mediterranean (LAM lineage (n = 6386, 30.1% of strains. By combining classical phylogenetic analyses and Bayesian approaches, this study revealed for the first time a clear genetic structuration of LAM9 sublineage into two subpopulations named LAM9C1 and LAM9C2, with distinct genetic characteristics. LAM9C1 was predominant in Chile, Colombia and USA, while LAM9C2 was predominant in Brazil, Dominican Republic, Guadeloupe and French Guiana. Globally, LAM9C2 was characterized by higher allelic richness as compared to LAM9C1 isolates. Moreover, LAM9C2 sublineage appeared to expand close to twenty times more than LAM9C1 and showed older traces of expansion. Interestingly, a significant proportion of LAM9C2 isolates presented typical signature of ancestral LAM-RDRio MIRU-VNTR type (224226153321. Further studies based on Whole Genome Sequencing of LAM strains will provide the needed resolution to decipher the biogeographical structure and evolutionary history of this successful family.

  15. Distinct steps of neural induction revealed by Asterix, Obelix and TrkC, genes induced by different signals from the organizer.

    Directory of Open Access Journals (Sweden)

    Sonia Pinho

    Full Text Available The amniote organizer (Hensen's node can induce a complete nervous system when grafted into a peripheral region of a host embryo. Although BMP inhibition has been implicated in neural induction, non-neural cells cannot respond to BMP antagonists unless previously exposed to a node graft for at least 5 hours before BMP inhibitors. To define signals and responses during the first 5 hours of node signals, a differential screen was conducted. Here we describe three early response genes: two of them, Asterix and Obelix, encode previously undescribed proteins of unknown function but Obelix appears to be a nuclear RNA-binding protein. The third is TrkC, a neurotrophin receptor. All three genes are induced by a node graft within 4-5 hours but they differ in the extent to which they are inducible by FGF: FGF is both necessary and sufficient to induce Asterix, sufficient but not necessary to induce Obelix and neither sufficient nor necessary for induction of TrkC. These genes are also not induced by retinoic acid, Noggin, Chordin, Dkk1, Cerberus, HGF/SF, Somatostatin or ionomycin-mediated Calcium entry. Comparison of the expression and regulation of these genes with other early neural markers reveals three distinct "epochs", or temporal waves, of gene expression accompanying neural induction by a grafted organizer, which are mirrored by specific stages of normal neural plate development. The results are consistent with neural induction being a cascade of responses elicited by different signals, culminating in the formation of a patterned nervous system.

  16. Direct Comparison of Physical Properties of Bacillus subtilis NCIB 3610 and B-1 Biofilms.

    Science.gov (United States)

    Kesel, Sara; Grumbein, Stefan; Gümperlein, Ina; Tallawi, Marwa; Marel, Anna-Kristina; Lieleg, Oliver; Opitz, Madeleine

    2016-04-01

    Many bacteria form surface-attached communities known as biofilms. Due to the extreme resistance of these bacterial biofilms to antibiotics and mechanical stresses, biofilms are of growing interest not only in microbiology but also in medicine and industry. Previous studies have determined the extracellular polymeric substances present in the matrix of biofilms formed by Bacillus subtilis NCIB 3610. However, studies on the physical properties of biofilms formed by this strain are just emerging. In particular, quantitative data on the contributions of biofilm matrix biopolymers to these physical properties are lacking. Here, we quantitatively investigated three physical properties of B. subtilis NCIB 3610 biofilms: the surface roughness and stiffness and the bulk viscoelasticity of these biofilms. We show how specific biomolecules constituting the biofilm matrix formed by this strain contribute to those biofilm properties. In particular, we demonstrate that the surface roughness and surface elasticity of 1-day-old NCIB 3610 biofilms are strongly affected by the surface layer protein BslA. For a second strain,B. subtilis B-1, which forms biofilms containing mainly γ-polyglutamate, we found significantly different physical biofilm properties that are also differently affected by the commonly used antibacterial agent ethanol. We show that B-1 biofilms are protected from ethanol-induced changes in the biofilm's stiffness and that this protective effect can be transferred to NCIB 3610 biofilms by the sole addition of γ-polyglutamate to growing NCIB 3610 biofilms. Together, our results demonstrate the importance of specific biofilm matrix components for the distinct physical properties of B. subtilis biofilms.

  17. Coaggregation between freshwater bacteria within biofilm and planktonic communities.

    Science.gov (United States)

    Rickard, A H; McBain, A J; Ledder, R G; Handley, P S; Gilbert, P

    2003-03-14

    The coaggregation ability of bacteria isolated from a freshwater biofilm was compared to those derived from the coexisting planktonic population. Twenty-nine morphologically distinct bacterial strains were isolated from a 6-month-old biofilm, established in a glass tank under high-shear conditions, and 15 distinct strains were isolated from the associated re-circulating water. All 44 strains were identified to genus or species level by 16S rDNA sequencing. The 29 biofilm strains belonged to 14 genera and 23.4% of all the possible pair-wise combinations coaggregated. The 15 planktonic strains belonged to seven genera and only 5.8% of all the possible pair-wise combinations coaggregated. Therefore, compared to the planktonic population, a greater proportion of the biofilm strains coaggregated. It is proposed that coaggregation influences biofilm formation and species diversity in freshwater under high shear.

  18. Fur is a repressor of biofilm formation in Yersinia pestis.

    Directory of Open Access Journals (Sweden)

    Fengjun Sun

    Full Text Available BACKGROUND: Yersinia pestis synthesizes the attached biofilms in the flea proventriculus, which is important for the transmission of this pathogen by fleas. The hmsHFRS operons is responsible for the synthesis of exopolysaccharide (the major component of biofilm matrix, which is activated by the signaling molecule 3', 5'-cyclic diguanylic acid (c-di-GMP synthesized by the only two diguanylate cyclases HmsT, and YPO0449 (located in a putative operonYPO0450-0448. METHODOLOGY/PRINCIPAL FINDINGS: The phenotypic assays indicated that the transcriptional regulator Fur inhibited the Y. pestis biofilm production in vitro and on nematode. Two distinct Fur box-like sequences were predicted within the promoter-proximal region of hmsT, suggesting that hmsT might be a direct Fur target. The subsequent primer extension, LacZ fusion, electrophoretic mobility shift, and DNase I footprinting assays disclosed that Fur specifically bound to the hmsT promoter-proximal region for repressing the hmsT transcription. In contrast, Fur had no regulatory effect on hmsHFRS and YPO0450-0448 at the transcriptional level. The detection of intracellular c-di-GMP levels revealed that Fur inhibited the c-di-GMP production. CONCLUSIONS/SIGNIFICANCE: Y. pestis Fur inhibits the c-di-GMP production through directly repressing the transcription of hmsT, and thus it acts as a repressor of biofilm formation. Since the relevant genetic contents for fur, hmsT, hmsHFRS, and YPO0450-0448 are extremely conserved between Y. pestis and typical Y. pseudotuberculosis, the above regulatory mechanisms can be applied to Y. pseudotuberculosis.

  19. Endodontic Biofilm: Quo Vadis?

    National Research Council Canada - National Science Library

    Tatiana Ramírez Mora

    2016-01-01

    .... However, the evidence of biofilm along the root canal system and the inability of current instruments and irrigants to eliminate bacterial biofilm have built a barrier toward a higher favorable...

  20. Biofilms of Clostridium species.

    Science.gov (United States)

    Pantaléon, Véronique; Bouttier, Sylvie; Soavelomandroso, Anna Philibertine; Janoir, Claire; Candela, Thomas

    2014-12-01

    The biofilm is a microbial community embedded in a synthesized matrix and is the main bacterial way of life. A biofilm adheres on surfaces or is found on interfaces. It protects bacteria from the environment, toxic molecules and may have a role in virulence. Clostridium species are spread throughout both environments and hosts, but their biofilms have not been extensively described in comparison with other bacterial species. In this review we describe all biofilms formed by Clostridium species during both industrial processes and in mammals where biofilms may be formed either during infections or associated to microbiota in the gut. We have specifically focussed on Clostridium difficile and Clostridium perfringens biofilms, which have been studied in vitro. Regulatory processes including sporulation and germination highlight how these Clostridium species live in biofilms. Furthermore, biofilms may have a role in the survival and spreading of Clostridium species. Copyright © 2014 Elsevier Ltd. All rights reserved.

  1. Pseudomonas aeruginosa Biofilm Infections

    DEFF Research Database (Denmark)

    Rybtke, Morten; Hultqvist, Louise Dahl; Givskov, Michael

    2015-01-01

    Studies of biopsies from infectious sites, explanted tissue and medical devises have provided evidence that biofilms are the underlying cause of a variety of tissue-associated and implant-associated recalcitrant human infections. With a need for novel anti-biofilm treatment strategies, research...... in biofilm infection microbiology, biofilm formation mechanisms and biofilm-associated antimicrobial tolerance has become an important area in microbiology. Substantial knowledge about biofilm formation mechanisms, biofilm-associated antimicrobial tolerance and immune evasion mechanisms has been obtained...... through work with biofilms grown in in vitro experimental setups, and the relevance of this information in the context of chronic infections is being investigated by the use of animal models of infection. Because our current in vitro experimental setups and animal models have limitations, new advanced...

  2. Diversification of Gene Expression during Formation of Static Submerged Biofilms by Escherichia coli

    Science.gov (United States)

    Besharova, Olga; Suchanek, Verena M.; Hartmann, Raimo; Drescher, Knut; Sourjik, Victor

    2016-01-01

    Many bacteria primarily exist in nature as structured multicellular communities, so called biofilms. Biofilm formation is a highly regulated process that includes the transition from the motile planktonic to sessile biofilm lifestyle. Cellular differentiation within a biofilm is a commonly accepted concept but it remains largely unclear when, where and how exactly such differentiation arises. Here we used fluorescent transcriptional reporters to quantitatively analyze spatio-temporal expression patterns of several groups of genes during the formation of submerged Escherichia coli biofilms in an open static system. We first confirm that formation of such submerged biofilms as well as pellicles at the liquid-air interface requires the major matrix component, curli, and flagella-mediated motility. We further demonstrate that in this system, diversification of gene expression leads to emergence of at least three distinct subpopulations of E. coli, which differ in their levels of curli and flagella expression, and in the activity of the stationary phase sigma factor σS. Our study reveals mutually exclusive expression of curli fibers and flagella at the single cell level, with high curli levels being confined to dense cell aggregates/microcolonies and flagella expression showing an opposite expression pattern. Interestingly, despite the known σS-dependence of curli induction, there was only a partial correlation between the σS activity and curli expression, with subpopulations of cells having high σS activity but low curli expression and vice versa. Finally, consistent with different physiology of the observed subpopulations, we show striking differences between the growth rates of cells within and outside of aggregates. PMID:27761132

  3. Early canine plaque biofilms: characterization of key bacterial interactions involved in initial colonization of enamel.

    Directory of Open Access Journals (Sweden)

    Lucy J Holcombe

    Full Text Available Periodontal disease (PD is a significant problem in dogs affecting between 44% and 63.6% of the population. The main etiological agent for PD is plaque, a microbial biofilm that colonizes teeth and causes inflammation of the gingiva. Understanding how this biofilm initiates on the tooth surface is of central importance in developing interventions against PD. Although the stages of plaque development on human teeth have been well characterized little is known about how canine plaque develops. Recent studies of the canine oral microbiome have revealed distinct differences between the canine and human oral environments and the bacterial communities they support, particularly with respect to healthy plaque. These differences mean knowledge about the nature of plaque formation in humans may not be directly translatable to dogs. The aim of this study was to identify the bacterial species important in the early stages of canine plaque formation in vivo and then use isolates of these species in a laboratory biofilm model to develop an understanding of the sequential processes which take place during the initial colonization of enamel. Supra-gingival plaque samples were collected from 12 dogs at 24 and 48 hour time points following a full mouth descale and polish. Pyrosequencing of the 16S rDNA identified 134 operational taxonomic units after statistical analysis. The species with the highest relative abundance were Bergeyella zoohelcum, Neisseria shayeganii and a Moraxella species. Streptococcal species, which tend to dominate early human plaque biofilms, had very low relative abundance. In vitro testing of biofilm formation identified five primary colonizer species, three of which belonged to the genus Neisseria. Using these pioneer bacteria as a starting point, viable two and three species communities were developed. Combining in vivo and in vitro data has led us to construct novel models of how the early canine plaque biofilm develops.

  4. Evolution of Ecological Diversity in Biofilms of Pseudomonas aeruginosa by Altered Cyclic Diguanylate Signaling

    Science.gov (United States)

    Flynn, Kenneth M.; Dowell, Gabrielle; Johnson, Thomas M.; Koestler, Benjamin J.; Waters, Christopher M.

    2016-01-01

    ABSTRACT The ecological and evolutionary forces that promote and maintain diversity in biofilms are not well understood. To quantify these forces, three Pseudomonas aeruginosa populations were experimentally evolved from strain PA14 in a daily cycle of attachment, assembly, and dispersal for 600 generations. Each biofilm population evolved diverse colony morphologies and mutator genotypes defective in DNA mismatch repair. This diversity enhanced population fitness and biofilm output, owing partly to rare, early colonizing mutants that enhanced attachment of others. Evolved mutants exhibited various levels of the intracellular signal cyclic-di-GMP, which associated with their timing of adherence. Manipulating cyclic-di-GMP levels within individual mutants revealed a network of interactions in the population that depended on various attachment strategies related to this signal. Diversification in biofilms may therefore arise and be reinforced by initial colonists that enable community assembly. IMPORTANCE How biofilm diversity assembles, evolves, and contributes to community function is largely unknown. This presents a major challenge for understanding evolution during chronic infections and during the growth of all surface-associated microbes. We used experimental evolution to probe these dynamics and found that diversity, partly related to altered cyclic-di-GMP levels, arose and persisted due to the emergence of ecological interdependencies related to attachment patterns. Clonal isolates failed to capture population attributes, which points to the need to account for diversity in infections. More broadly, this study offers an experimental framework for linking phenotypic variation to distinct ecological strategies in biofilms and for studying eco-evolutionary interactions. PMID:27021563

  5. Diversification of gene expression during formation of static submerged biofilms by Escherichia coli

    Directory of Open Access Journals (Sweden)

    Olga Besharova

    2016-10-01

    Full Text Available Many bacteria primarily exist in nature as structured multicellular communities, so called biofilms. Biofilm formation is a highly regulated process that includes the transition from the motile planktonic to sessile biofilm lifestyle. Cellular differentiation within a biofilm is a commonly accepted concept but it remains largely unclear when, where and how exactly such differentiation arises. Here we used fluorescent transcriptional reporters to quantitatively analyze spatio-temporal expression patterns of several groups of genes during the formation of submerged Escherichia coli biofilms in an open static system. We first confirm that formation of such submerged biofilms as well as pellicles at the liquid-air interface requires the major matrix component, curli, and flagella-mediated motility. We further demonstrate that in this system, diversification of gene expression leads to emergence of at least three distinct subpopulations of E. coli, which differ in their levels of curli and flagella expression, and in the activity of the stationary phase sigma factor σS. Our study reveals mutually exclusive expression of curli fibers and flagella at the single cell level, with high curli levels being confined to dense cell aggregates/microcolonies and flagella expression showing an opposite expression pattern. Interestingly, despite the known σS-dependence of curli induction, there was only a partial correlation between the σS activity and curli expression, with subpopulations of cells having high σS activity but low curli expression and vice versa. Finally, consistent with different physiology of the observed subpopulations, we show striking differences between the growth rates of cells within and outside of aggregates.

  6. Biofilm Formation of Pasteurella Multocida on Bentonite Clay

    Directory of Open Access Journals (Sweden)

    Ramachandranpillai Rajagopal

    2013-06-01

    Full Text Available Background and objectives: Biofilms are structural communities of bacterial cells enshrined in a self produced polymeric matrix. The studies on biofilm formation of Pasteurella multocida have become imperative since it is a respiratory pathogen and its biofilm mode could possibly be one of its virulence factors for survival inside a host. The present study describes a biofilm assay for P. multocida on inert hydrophilic material called bentonite clay.Materials and methods: The potential of the organism to form in vitro biofilm was assessed by growing the organism under nutrient restriction along with the inert substrate bentonite clay, which will provide a surface for attachment. For quantification of biofilm, plate count by the spread plate method was employed. Capsule production of the attached bacteria was demonstrated by light microscopic examination following Maneval staining and capsular polysaccharide estimation was done using standard procedures.Results and Conclusion: The biofilm formation peaked on the third day of incubation (1.54 ×106 cfu/g of bentonite clay while the planktonic cells were found to be at a maximum on day one post inoculation (8.10 ×108 cfu/ml of the broth. Maneval staining of late logarithmic phase biofilm cultures revealed large aggregates of bacterial cells, bacteria appearing as chains or as a meshwork. The capsular polysaccharide estimation of biofilm cells revealed a 3.25 times increase over the planktonic bacteria. The biofilm cells cultured on solid media also produced some exclusive colony morphotypes

  7. Biofilm formation of Pasteurella multocida on bentonite clay.

    Science.gov (United States)

    Rajagopal, Ramachandranpillai; Nair, Govindapillai Krishnan; Mini, Mangattumuruppel; Joseph, Leo; Saseendranath, Mapranath Raghavan; John, Koshy

    2013-06-01

    Biofilms are structural communities of bacterial cells enshrined in a self produced polymeric matrix. The studies on biofilm formation of Pasteurella multocida have become imperative since it is a respiratory pathogen and its biofilm mode could possibly be one of its virulence factors for survival inside a host. The present study describes a biofilm assay for P. multocida on inert hydrophilic material called bentonite clay. The potential of the organism to form in vitro biofilm was assessed by growing the organism under nutrient restriction along with the inert substrate bentonite clay, which will provide a surface for attachment. For quantification of biofilm, plate count by the spread plate method was employed. Capsule production of the attached bacteria was demonstrated by light microscopic examination following Maneval staining and capsular polysaccharide estimation was done using standard procedures. The biofilm formation peaked on the third day of incubation (1.54 ×10(6) cfu/g of bentonite clay) while the planktonic cells were found to be at a maximum on day one post inoculation (8.10 ×10(8) cfu/ml of the broth). Maneval staining of late logarithmic phase biofilm cultures revealed large aggregates of bacterial cells, bacteria appearing as chains or as a meshwork. The capsular polysaccharide estimation of biofilm cells revealed a 3.25 times increase over the planktonic bacteria. The biofilm cells cultured on solid media also produced some exclusive colony morphotypes.

  8. Sexual Biofilm Formation in Candida tropicalis Opaque Cells

    Science.gov (United States)

    Jones, Stephen K.; Hirakawa, Matthew P.; Bennett, Richard J.

    2014-01-01

    Summary Candida albicans and Candida tropicalis are opportunistic fungal pathogens that can transition between white and opaque phenotypic states. White and opaque cells differ both morphologically and in their responses to environmental signals. In C. albicans, opaque cells respond to sexual pheromones by undergoing conjugation, while white cells are induced by pheromones to form sexual biofilms. Here, we show that sexual biofilm formation also occurs in C. tropicalis but, unlike C. albicans, biofilms are formed exclusively by opaque cells. C. tropicalis biofilm formation was dependent on the pheromone receptors Ste2 and Ste3, confirming the role of pheromone signaling in sexual biofilm development. Structural analysis of C. tropicalis sexual biofilms revealed stratified communities consisting of a basal layer of yeast cells and an upper layer of filamentous cells, together with an extracellular matrix. Transcriptional profiling showed that genes involved in pheromone signaling and conjugation were upregulated in sexual biofilms. Furthermore, FGR23, which encodes an agglutinin-like protein, was found to enhance both mating and sexual biofilm formation. Together, these studies reveal that C. tropicalis opaque cells form sexual biofilms with a complex architecture, and suggest a conserved role for sexual agglutinins in mediating mating, cell cohesion and biofilm formation. PMID:24612417

  9. Biofilm-specific extracellular matrix proteins of non-typeable Haemophilus influenzae

    Science.gov (United States)

    Wu, Siva; Baum, Marc M.; Kerwin, James; Guerrero-Given, Debbie; Webster, Simon; Schaudinn, Christoph; VanderVelde, David; Webster, Paul

    2014-01-01

    Non-typeable Haemophilus influenzae (NTHi), a human respiratory tract pathogen can form colony biofilms in vitro. Bacterial cells and the amorphous extracellular matrix (ECM) constituting the biofilm can be separated using sonication. The ECM from 24 hr and 96 hr NTHi biofilms contained polysaccharides and proteinaceous components as detected by NMR and FTIR spectroscopy. More conventional chemical assays on the biofilm ECM confirmed the presence of these components and also DNA. Proteomics revealed eighteen proteins present in biofilm ECM that were not detected in planktonic bacteria. One ECM protein was unique to 24 hr biofilms, two were found only in 96 hr biofilms, and fifteen were present in the ECM of both 24 hr and 96 hr NTHi biofilms. All proteins identified were either associated with bacterial membranes or were cytoplasmic proteins. Immunocytochemistry showed two of the identified proteins, a DNA-directed RNA polymerase and the outer membrane protein OMP P2, associated with bacteria and biofilm ECM. Identification of biofilm-specific proteins present in immature biofilms is an important step in understanding the in vitro process of NTHi biofilm formation. The presence of a cytoplasmic protein and a membrane protein in the biofilm ECM of immature NTHi biofilms suggests that bacterial cell lysis may be a feature of early biofilm formation. PMID:24942343

  10. Biofilms: A microbial home

    Science.gov (United States)

    Chandki, Rita; Banthia, Priyank; Banthia, Ruchi

    2011-01-01

    Microbial biofilms are mainly implicated in etiopathogenesis of caries and periodontal disease. Owing to its properties, these pose great challenges. Continuous and regular disruption of these biofilms is imperative for prevention and management of oral diseases. This essay provides a detailed insight into properties, mechanisms of etiopathogenesis, detection and removal of these microbial biofilms. PMID:21976832

  11. Biofilms: A microbial home

    OpenAIRE

    Chandki, Rita; Banthia, Priyank; Banthia, Ruchi

    2011-01-01

    Microbial biofilms are mainly implicated in etiopathogenesis of caries and periodontal disease. Owing to its properties, these pose great challenges. Continuous and regular disruption of these biofilms is imperative for prevention and management of oral diseases. This essay provides a detailed insight into properties, mechanisms of etiopathogenesis, detection and removal of these microbial biofilms.

  12. Treatment of Oral Multispecies Biofilms by an Anti-Biofilm Peptide.

    Science.gov (United States)

    Wang, Zhejun; de la Fuente-Núñez, Cesar; Shen, Ya; Haapasalo, Markus; Hancock, Robert E W

    2015-01-01

    Human oral biofilms are multispecies microbial communities that exhibit high resistance to antimicrobial agents. Dental plaque gives rise to highly prevalent and costly biofilm-related oral infections, which lead to caries or other types of oral infections. We investigated the ability of the recently identified anti-biofilm peptide 1018 to induce killing of bacterial cells present within oral multispecies biofilms. At 10 μg/ml (6.5 μM), peptide 1018 was able to significantly (pbiofilm formation over 3 days. The activity of the peptide on preformed biofilms was found to be concentration-dependent since more than 60% of the total plaque biofilm cell population was killed by 10 μg/ml of peptide 1018 in 3 days, while at 5 μg/ml 50% of cells were dead and at 1 μg/ml the peptide triggered cell death in around 30% of the total bacterial population, as revealed by confocal microscopy. The presence of saliva did not affect peptide activity, since no statistically significant difference was found in the ability of peptide 1018 to kill oral biofilms using either saliva coated and non-saliva coated hydroxyapatite surfaces. Scanning electron microscopy experiments indicated that peptide 1018 induced cell lysis in plaque biofilms. Furthermore, combined treatment using peptide 1018 and chlorhexidine (CHX) increased the anti-biofilm activity of each compound compared to when these were used alone, resulting in >50% of the biofilm being killed and >35% being dispersed in only 3 minutes. Peptide 1018 may potentially be used by itself or in combination with CHX as a non-toxic and effective anti-biofilm agent for plaque disinfection in clinical dentistry.

  13. Sensor kinase RscS induces the production of antigenically distinct outer membrane vesicles that depend on the symbiosis polysaccharide locus in Vibrio fischeri.

    Science.gov (United States)

    Shibata, Satoshi; Visick, Karen L

    2012-01-01

    Robust biofilm formation by Vibrio fischeri depends upon activation of the symbiosis polysaccharide (syp) locus, which is achieved by overexpressing the RscS sensor kinase (RscS(+)). Other than the Syp polysaccharide, however, little is known about V. fischeri biofilm matrix components. In other bacteria, biofilms contain polysaccharides, secreted proteins, and outer membrane vesicles (OMVs). Here, we asked whether OMVs are part of V. fischeri biofilms. Transmission electron microscopy revealed OMV-like particles between cells within colonies. In addition, OMVs could be purified from culture supernatants of both RscS(+) and control cells, with the former releasing 2- to 3-fold more OMVs. The increase depended upon the presence of an intact syp locus, as an RscS(+) strain deleted for sypK, which encodes a putative oligosaccharide translocase, exhibited reduced production of OMVs; it also showed a severe defect in biofilm formation. Western immunoblot analyses revealed that the RscS(+) strain, but not the control strain or the RscS(+) sypK mutant, produced a distinct set of nonproteinaceous molecules that could be detected in whole-cell extracts, OMV preparations, and lipopolysaccharide (LPS) extracts. Finally, deletion of degP, which in other bacteria influences OMV production, decreased OMV production and reduced the ability of the cells to form biofilms. We conclude that overexpression of RscS induces OMV production in a manner that depends on the presence of the syp locus and that OMVs produced under these conditions contain antigenically distinct molecules, possibly representing a modified form of lipopolysaccharide (LPS). Finally, our data indicate a correlation between OMV production and biofilm formation by V. fischeri.

  14. Dynamics of development and dispersal in sessile microbial communities: examples from Pseudomonas aeruginosa and Pseudomonas putida model biofilms

    DEFF Research Database (Denmark)

    Klausen, M.; Gjermansen, Morten; Kreft, J.-U.

    2006-01-01

    in these model biofilms develop characteristic multicellular structures through a series of distinct steps where cellular migration plays an important role. Despite the appearance of these characteristic developmental patterns in the model biofilms the available evidence suggest that the biofilm forming...

  15. MATHEMATICAL MODELING OF EXTRACELLULAR ELECTRON TRANSFER IN BIOFILMS

    Energy Technology Data Exchange (ETDEWEB)

    Renslow, Ryan S.; Babauta, Jerome T.; Kuprat, Andrew P.; Schenk, Jim; Ivory, Cornelius; Fredrickson, Jim K.; Beyenal, Haluk

    2015-09-12

    Electrochemically active biofilms have a unique form of respiration in which they utilize solid external materials as terminal electron acceptors for their metabolism. Currently, two primary mechanisms have been identified for long-range extracellular electron transfer (EET): a diffusion- and a conduction-based mechanism. Evidence in the literature suggests that some biofilms, particularly Shewanella oneidensis, produce the requisite components for both mechanisms. In this study, a generic model is presented that incorporates the diffusion- and the conduction-based mechanisms and allows electrochemically active biofilms to utilize both simultaneously. The model was applied to S. oneidensis and Geobacter sulfurreducens biofilms using experimentally generated data found in the literature. Our simulation results show that 1) biofilms having both mechanisms available, especially if they can interact, may have a metabolic advantage over biofilms that can use only a single mechanism; 2) the thickness of G. sulfurreducens biofilms is likely not limited by conductivity; 3) accurate intrabiofilm diffusion coefficient values are critical for current generation predictions; and 4) the local biofilm potential and redox potential are two distinct parameters and cannot be assumed to have identical values. Finally, we determined that simulated cyclic and squarewave voltammetry based on our model are currently not capable of determining the specific percentages of extracellular electron transfer mechanisms in a biofilm. The developed model will be a critical tool for designing experiments to explain EET mechanisms.

  16. Comparative transcriptomic analysis of Clostridium acetobutylicum biofilm and planktonic cells.

    Science.gov (United States)

    Liu, Dong; Xu, Jiahui; Wang, Yanyan; Chen, Yong; Shen, Xiaoning; Niu, Huanqing; Guo, Ting; Ying, Hanjie

    2016-01-20

    Biofilm-based immobilization of solventogenic Clostridia has been extensively exploited to overcome traditional bottlenecks in biobutanol production like solvent toxicity and low productivities. However, the molecular basis of solventogenic Clostridia biofilm is rarely explored. Here, for the first time, we report DNA array-based study of Clostridium acetobutylicum biofilm cells to elucidate the transcriptional modulation. Results showed that 16.2% of the C. acetobutylicum genome genes within the biofilm cells were differentially expressed, with most genes being up-regulated. The most dramatic changes occurred with amino acid biosynthesis, with sulfur uptake and cysteine biosynthesis being the most up-regulated and histidine biosynthesis being the most down-regulated in the biofilm cells. It was demonstrated that C. acetobutylicum biofilm cells increased metabolic activities probably by up-regulating iron and sulfur uptake and Fe-S cluster biosynthesis genes as well as glycolysis genes. Furthermore, genes involved in sporulation, granulose formation, extracellular polymer degradation, pentose catabolisms, and various other processes were also notably regulated, indicating that the biofilm mode of growth rendered the cells a distinct phenotype. This study provides valuable insights into the transcriptional regulation in C. acetobutylicum biofilm cells and should be highly useful for understanding and developing the biofilm-based processes. Copyright © 2015 Elsevier B.V. All rights reserved.

  17. Modeling biofilms with dual extracellular electron transfer mechanisms

    Science.gov (United States)

    Renslow, Ryan; Babauta, Jerome; Kuprat, Andrew; Schenk, Jim; Ivory, Cornelius; Fredrickson, Jim; Beyenal, Haluk

    2013-01-01

    Electrochemically active biofilms have a unique form of respiration in which they utilize solid external materials as terminal electron acceptors for their metabolism. Currently, two primary mechanisms have been identified for long-range extracellular electron transfer (EET): a diffusion- and a conduction-based mechanism. Evidence in the literature suggests that some biofilms, particularly Shewanella oneidensis, produce the requisite components for both mechanisms. In this study, a generic model is presented that incorporates the diffusion- and the conduction-based mechanisms and allows electrochemically active biofilms to utilize both simultaneously. The model was applied to S. oneidensis and Geobacter sulfurreducens biofilms using experimentally generated data found in the literature. Our simulation results show that 1) biofilms having both mechanisms available, especially if they can interact, may have a metabolic advantage over biofilms that can use only a single mechanism; 2) the thickness of G. sulfurreducens biofilms is likely not limited by conductivity; 3) accurate intrabiofilm diffusion coefficient values are critical for current generation predictions; and 4) the local biofilm potential and redox potential are two distinct parameters and cannot be assumed to have identical values. Finally, we determined that simulated cyclic and squarewave voltammetry based on our model are currently not capable of determining the specific percentages of extracellular electron transfer mechanisms in a biofilm. The developed model will be a critical tool for designing experiments to explain EET mechanisms. PMID:24113651

  18. In vitro study of biofilm growth on biologic prosthetics.

    Science.gov (United States)

    Bellows, Charles; Smith, Alison

    2014-01-01

    Biologic prosthetics are increasingly used for the repair of abdominal wall hernia defects but can become infected as a result of peri- or early post-operative bacterial contamination. Data evaluating biofilm formation on biologic prosthetics is lacking. The aim of this study was to investigate the influence of different biologic prosthetics on the growth behavior of two different bacterial species and their ability to form biofilms. Methicillin resistant Staphylococcus aureus (MRSA) or Pseudomrnonas aeruginosa were incubated on disks of two biologic prosthetics-human acellular dermis (ADM), and porcine small intestinal submucosa (SIS). The bacteria were allowed to attach to the prosthetics and propagate into mature biofilms for 24 hours at 370C. Images of biofilms were obtained using confocal microscopy and scanning electron microscopy (SEM). The number of viable cells and the biofilm biomass were quantified by colony forming units (CFUs) and crystal violet staining respectively. Analysis of variance was performed to compare the mean values for the different prosthetics. Each biologic matrix had a distinct surface characteristic. SEM visualized mature biofilms characterized by highly organized multi-cellular structures on surface of both biologic prosthetics. Quantification of bacterial growth over time showed that ADM had the lowest CFUs and biofilm biomass at 24 hours post-inoculation compared to SIS for both bacterial strains. MRSA and P. aeruginosa can form mature biofilms on biologic prosthetics but the relative abundance of the biofilm varies on different prosthetic constructs. Biologic material composition and manufacturing methods may influence bacterial adherence.

  19. Characterization of biofilm formed on intrauterine devices

    Directory of Open Access Journals (Sweden)

    Pruthi V

    2003-01-01

    Full Text Available PURPOSE: Intrauterine device (IUD is one of the most convenient contraceptive procedures used by women of Asian and African countries. Previous surveys have revealed that 75% of the IUDs recovered from patients suffering from reproductive tract infections (RTIs were covered with a consortium of microbes. This study was designed to characterize these microbes and recommend remedial measures. METHODS: Quantitative measurement of biofilm formation was assessed by a microtitre plate assay on 86 samples of microorganisms dislodged from IUDs of patients with RTIs. Susceptibility of biofilm to various antimicrobial agents was also quantified. Scanning electron microscopy (SEM was used to scrutinize the microorganisms adherent to IUDs. RESULTS: The organisms associated with IUDs were predominantly composed of Staphylococcus aureus (16%, Staphylococcus epidermidis (18%, Pseudomonas aeruginosa (5%, Escherichia coli (27%, Neisseria gonorrhoeae (2%, Candida albicans (20% and Candida dubliniesis (12%. SEM studies indicated that these organisms were organized into biofilms. Studies on the in vitro adherence pattern by crystal violet staining on 96 well microtitre plates revealed that the biofilms were stably established after 60 hours. These biofilms are resistant to an array of antibiotics tested. CONCLUSION: Biofilm formation may be one of the major causes for persistent infection and antibiotic resistance in IUD users.

  20. Streptococcus pneumoniae biofilm formation and dispersion during colonization and disease.

    Science.gov (United States)

    Chao, Yashuan; Marks, Laura R; Pettigrew, Melinda M; Hakansson, Anders P

    2014-01-01

    Streptococcus pneumoniae (the pneumococcus) is a common colonizer of the human nasopharynx. Despite a low rate of invasive disease, the high prevalence of colonization results in millions of infections and over one million deaths per year, mostly in individuals under the age of 5 and the elderly. Colonizing pneumococci form well-organized biofilm communities in the nasopharyngeal environment, but the specific role of biofilms and their interaction with the host during colonization and disease is not yet clear. Pneumococci in biofilms are highly resistant to antimicrobial agents and this phenotype can be recapitulated when pneumococci are grown on respiratory epithelial cells under conditions found in the nasopharyngeal environment. Pneumococcal biofilms display lower levels of virulence in vivo and provide an optimal environment for increased genetic exchange both in vitro and in vivo, with increased natural transformation seen during co-colonization with multiple strains. Biofilms have also been detected on mucosal surfaces during pneumonia and middle ear infection, although the role of these biofilms in the disease process is debated. Recent studies have shown that changes in the nasopharyngeal environment caused by concomitant virus infection, changes in the microflora, inflammation, or other host assaults trigger active release of pneumococci from biofilms. These dispersed bacteria have distinct phenotypic properties and transcriptional profiles different from both biofilm and broth-grown, planktonic bacteria, resulting in a significantly increased virulence in vivo. In this review we discuss the properties of pneumococcal biofilms, the role of biofilm formation during pneumococcal colonization, including their propensity for increased ability to exchange genetic material, as well as mechanisms involved in transition from asymptomatic biofilm colonization to dissemination and disease of otherwise sterile sites. Greater understanding of pneumococcal biofilm

  1. Development of a high-throughput Candida albicans biofilm chip.

    Directory of Open Access Journals (Sweden)

    Anand Srinivasan

    Full Text Available We have developed a high-density microarray platform consisting of nano-biofilms of Candida albicans. A robotic microarrayer was used to print yeast cells of C. albicans encapsulated in a collagen matrix at a volume as low as 50 nL onto surface-modified microscope slides. Upon incubation, the cells grow into fully formed "nano-biofilms". The morphological and architectural complexity of these biofilms were evaluated by scanning electron and confocal scanning laser microscopy. The extent of biofilm formation was determined using a microarray scanner from changes in fluorescence intensities due to FUN 1 metabolic processing. This staining technique was also adapted for antifungal susceptibility testing, which demonstrated that, similar to regular biofilms, cells within the on-chip biofilms displayed elevated levels of resistance against antifungal agents (fluconazole and amphotericin B. Thus, results from structural analyses and antifungal susceptibility testing indicated that despite miniaturization, these biofilms display the typical phenotypic properties associated with the biofilm mode of growth. In its final format, the C. albicans biofilm chip (CaBChip is composed of 768 equivalent and spatially distinct nano-biofilms on a single slide; multiple chips can be printed and processed simultaneously. Compared to current methods for the formation of microbial biofilms, namely the 96-well microtiter plate model, this fungal biofilm chip has advantages in terms of miniaturization and automation, which combine to cut reagent use and analysis time, minimize labor intensive steps, and dramatically reduce assay costs. Such a chip should accelerate the antifungal drug discovery process by enabling rapid, convenient and inexpensive screening of hundreds-to-thousands of compounds simultaneously.

  2. Development of a high-throughput Candida albicans biofilm chip.

    Science.gov (United States)

    Srinivasan, Anand; Uppuluri, Priya; Lopez-Ribot, Jose; Ramasubramanian, Anand K

    2011-04-22

    We have developed a high-density microarray platform consisting of nano-biofilms of Candida albicans. A robotic microarrayer was used to print yeast cells of C. albicans encapsulated in a collagen matrix at a volume as low as 50 nL onto surface-modified microscope slides. Upon incubation, the cells grow into fully formed "nano-biofilms". The morphological and architectural complexity of these biofilms were evaluated by scanning electron and confocal scanning laser microscopy. The extent of biofilm formation was determined using a microarray scanner from changes in fluorescence intensities due to FUN 1 metabolic processing. This staining technique was also adapted for antifungal susceptibility testing, which demonstrated that, similar to regular biofilms, cells within the on-chip biofilms displayed elevated levels of resistance against antifungal agents (fluconazole and amphotericin B). Thus, results from structural analyses and antifungal susceptibility testing indicated that despite miniaturization, these biofilms display the typical phenotypic properties associated with the biofilm mode of growth. In its final format, the C. albicans biofilm chip (CaBChip) is composed of 768 equivalent and spatially distinct nano-biofilms on a single slide; multiple chips can be printed and processed simultaneously. Compared to current methods for the formation of microbial biofilms, namely the 96-well microtiter plate model, this fungal biofilm chip has advantages in terms of miniaturization and automation, which combine to cut reagent use and analysis time, minimize labor intensive steps, and dramatically reduce assay costs. Such a chip should accelerate the antifungal drug discovery process by enabling rapid, convenient and inexpensive screening of hundreds-to-thousands of compounds simultaneously.

  3. Comparative Kinetic Studies and Performance Evaluation of Biofilm and Biomass Characteristics of Pseudomonas fluorescens in Degrading Synthetic Phenolic Effluent in Inverse Fluidized Bed Biofilm Reactor.

    Science.gov (United States)

    Begum, S Sabarunisha; Radha, K V

    2016-05-01

    The bioremediation potential of Pseudomonas fluorescens was studied in an Inverse Fluidized Bed Biofilm Reactor under batch recirculation conditions using synthetic phenolic effluent of various concentrations (400, 600, 800, 1000 and 1200 mg/l). The performance of the reactor was investigated and the characteristics of biomass and biofilm were determined by evaluating biofilm dry density and thickness, bioparticle density, suspended and attached biomass concentration, chemical oxygen demand and phenol removal efficiency. Biodegradation kinetics had been studied for suspended biomass culture and biofilm systems with respect to its specific growth and substrate consumption rates. Suspended biomass followed substrate inhibition kinetics and the experimental data fitted well with the Haldane model. The degradation kinetic behavior of biofilm revealed that a well adapted biofilm system with effective control of biofilm thickness in an inverse fluidized bed biofilm reactor overcomes substrate inhibition effects by tolerating higher phenol concentration and fitted well to the Monod model.

  4. First in-depth analysis of the novel Th2-type cytokines in salmonid fish reveals distinct patterns of expression and modulation but overlapping bioactivities

    Science.gov (United States)

    Wang, Tiehui; Johansson, Petronella; Abós, Beatriz; Holt, Amy; Tafalla, Carolina; Jiang, Youshen; Wang, Alex; Xu, Qiaoqing; Qi, Zhitao; Huang, Wenshu; Costa, Maria M.; Diaz-Rosales, Patricia; Holland, Jason W.; Secombes, Christopher J.

    2016-01-01

    IL-4 and IL-13 are closely related canonical type-2 cytokines in mammals and have overlapping bioactivities via shared receptors. They are frequently activated together as part of the same immune response and are the signature cytokines produced by T-helper (Th)2 cells and type-2 innate lymphoid cells (ILC2), mediating immunity against extracellular pathogens. Little is known about the origin of type-2 responses, and whether they were an essential component of the early adaptive immune system that gave a fitness advantage by limiting collateral damage caused by metazoan parasites. Two evolutionary related type-2 cytokines, IL-4/13A and IL-4/13B, have been identified recently in several teleost fish that likely arose by duplication of an ancestral IL-4/13 gene as a consequence of a whole genome duplication event that occurred at the base of this lineage. However, studies of their comparative expression levels are largely missing and bioactivity analysis has been limited to IL-4/13A in zebrafish. Through interrogation of the recently released salmonid genomes, species in which an additional whole genome duplication event has occurred, four genomic IL-4/13 loci have been identified leading to the cloning of three active genes, IL-4/13A, IL-4/13B1 and IL-4/13B2, in both rainbow trout and Atlantic salmon. Comparative expression analysis by real-time PCR in rainbow trout revealed that the IL-4/13A expression is broad and high constitutively but less responsive to pathogen-associated molecular patterns (PAMPs) and pathogen challenge. In contrast, the expression of IL-4/13B1 and IL-4/13B2 is low constitutively but is highly induced by viral haemorrhagic septicaemia virus (VHSH) infection and during proliferative kidney disease (PKD) in vivo, and by formalin-killed bacteria, PAMPs, the T cell mitogen PHA, and the T-cell cytokines IL-2 and IL-21 in vitro. Moreover, bioactive recombinant cytokines of both IL-4/13A and B were produced and found to have shared but also distinct

  5. Locked Nucleic Acid-Based In Situ Hybridization Reveals miR-7a as a Hypothalamus-Enriched MicroRNA with a Distinct Expression Pattern

    DEFF Research Database (Denmark)

    Herzer, S; Silahtaroglu, A; Meister, B

    2012-01-01

    present in the hypothalamus, miR-7a, was the only miRNA found to be enriched in the hypothalamus, with low or no expression in other parts of the central nervous system (CNS). Within the hypothalamus, strong miR-7a expression was distinct and restricted to some hypothalamic nuclei and adjacent areas. mi...

  6. Distinct Pattern Separation Related Transfer Functions in Human CA3/Dentate and CA1 Revealed Using High-Resolution fMRI and Variable Mnemonic Similarity

    Science.gov (United States)

    Lacy, Joyce W.; Yassa, Michael A.; Stark, Shauna M.; Muftuler, L. Tugan; Stark, Craig E. L.

    2011-01-01

    Producing and maintaining distinct (orthogonal) neural representations for similar events is critical to avoiding interference in long-term memory. Recently, our laboratory provided the first evidence for separation-like signals in the human CA3/dentate. Here, we extended this by parametrically varying the change in input (similarity) while…

  7. Structure, composition, and strength of nitrifying membrane-aerated biofilms

    DEFF Research Database (Denmark)

    Pellicer i Nàcher, Carles; Smets, Barth F.

    2014-01-01

    Membrane-aerated biofilm reactors (MABRs) are a novel technology based on the growth of biofilms on oxygen-permeable membranes. Hereby, MABRs combine all the advantages of biofilm growth with a more flexible and efficient control of the oxygen load. In the present work, flow cell operation...... had a higher content of proteins and a lower level of carbohydrates. Staining analyses revealed that the EPS in the stronger biofilm regions had hydrophilic nature and distributed around dense microbial aggregates, whereas it was homogeneously distributed in the weaker strata. Overall, the obtained...

  8. Effect of proteases on biofilm formation of the plastic-degrading actinomycete Rhodococcus ruber C208.

    Science.gov (United States)

    Gilan, Irit; Sivan, Alex

    2013-05-01

    In most habitats, the vast majority of microbial populations form biofilms on solid surfaces, whether natural or artificial. These biofilms provide either increased physical support and/or a source of nutrients. Further modifications and development of biofilms are regulated by signal molecules secreted by the cells. Because synthetic polymers are not soluble in aqueous solutions, biofilm-producing bacteria may biodegrade such materials more efficiently than planktonic strains. Bacterial biofilms comprise bacterial cells embedded in self-secreted extracellular polymeric substances (EPS). Revealing the roles of each component of the EPS will enable further insight into biofilm development and the EPS structure-function relationship. A strain of Rhodococcus ruber (C208) displayed high hydrophobicity and formed a dense biofilm on the surface of polyethylene films while utilizing the polyolefin as carbon and energy sources. This study investigated the effects of several proteases on C208 biofilm formation and stability. The proteolysis of C208 biofilm gave conflicting results. Trypsin significantly reduced biofilm formation, and the resultant biofilm appeared monolayered. In contrast, proteinase K enhanced biofilm formation, which was robust and multilayered. Presumably, proteinase K degraded self-secreted proteases or quorum-sensing peptides, which may be involved in biofilm detachment processes, leading to a multilayered, nondispersed biofilm.

  9. Shaping the Growth Behaviour of Bacterial Aggregates in Biofilms

    CERN Document Server

    Melaugh, Gavin; Kragh, Kasper Nørskov; Irie, Yasuhiko; Roberts, Aled; Bjarnsholt, Thomas; Diggle, Steve P; Gordon, Vernita; Allen, Rosalind J

    2015-01-01

    Bacterial biofilms are usually assumed to originate from individual cells deposited on a surface. However, many biofilm-forming bacteria tend to aggregate in the planktonic phase meaning it is possible that many natural and infectious biofilms originate wholly or partially from pre-formed cell aggregates. Here, we use agent-based computer simulations to investigate the role of pre-formed aggregates in biofilm development. Focusing on the role of aggregate shape, we find that the degree of spreading of an aggregate on a surface can play a key role in determining its eventual fate during biofilm development. Specifically, initially spread aggregates perform better when competition with surrounding bacterial cells is low, while initially rounded aggregates perform better when competition is high. These contrasting outcomes are governed by a trade-off between aggregate surface area and height. Our results provide new insight into biofilm formation and development, and reveal new factors that may be at play in the...

  10. Production of tyrosol by Candida albicans biofilms and its role in quorum sensing and biofilm development.

    Science.gov (United States)

    Alem, Mohammed A S; Oteef, Mohammed D Y; Flowers, T Hugh; Douglas, L Julia

    2006-10-01

    Tyrosol and farnesol are quorum-sensing molecules produced by Candida albicans which accelerate and block, respectively, the morphological transition from yeasts to hyphae. In this study, we have investigated the secretion of tyrosol by C. albicans and explored its likely role in biofilm development. Both planktonic (suspended) cells and biofilms of four C. albicans strains, including three mutants with defined defects in the Efg 1 and Cph 1 morphogenetic signaling pathways, synthesized extracellular tyrosol during growth at 37 degrees C. There was a correlation between tyrosol production and biomass for both cell types. However, biofilm cells secreted at least 50% more tyrosol than did planktonic cells when tyrosol production was related to cell dry weight. The addition of exogenous farnesol to a wild-type strain inhibited biofilm formation by up to 33% after 48 h. Exogenous tyrosol appeared to have no effect, but scanning electron microscopy revealed that tyrosol stimulated hypha production during the early stages (1 to 6 h) of biofilm development. Experiments involving the simultaneous addition of tyrosol and farnesol at different concentrations suggested that the action of farnesol was dominant, and 48-h biofilms formed in the presence of both compounds consisted almost entirely of yeast cells. When biofilm supernatants were tested for their abilities to inhibit or enhance germ tube formation by planktonic cells, the results indicated that tyrosol activity exceeds that of farnesol after 14 h, but not after 24 h, and that farnesol activity increases significantly during the later stages (48 to 72 h) of biofilm development. Overall, our results support the conclusion that tyrosol acts as a quorum-sensing molecule for biofilms as well as for planktonic cells and that its action is most significant during the early and intermediate stages of biofilm formation.

  11. Spore formation and toxin production in Clostridium difficile biofilms.

    Directory of Open Access Journals (Sweden)

    Ekaterina G Semenyuk

    Full Text Available The ability to grow as a biofilm can facilitate survival of bacteria in the environment and promote infection. To better characterize biofilm formation in the pathogen Clostridium difficile, we established a colony biofilm culture method for this organism on a polycarbonate filter, and analyzed the matrix and the cells in biofilms from a variety of clinical isolates over several days of biofilm culture. We found that biofilms readily formed in all strains analyzed, and that spores were abundant within about 6 days. We also found that extracellular DNA (eDNA, polysaccharide and protein was readily detected in the matrix of all strains, including the major toxins A and/or B, in toxigenic strains. All the strains we analyzed formed spores. Apart from strains 630 and VPI10463, which sporulated in the biofilm at relatively low frequencies, the frequencies of biofilm sporulation varied between 46 and 65%, suggesting that variations in sporulation levels among strains is unlikely to be a major factor in variation in the severity of disease. Spores in biofilms also had reduced germination efficiency compared to spores obtained by a conventional sporulation protocol. Transmission electron microscopy revealed that in 3 day-old biofilms, the outermost structure of the spore is a lightly staining coat. However, after 6 days, material that resembles cell debris in the matrix surrounds the spore, and darkly staining granules are closely associated with the spores surface. In 14 day-old biofilms, relatively few spores are surrounded by the apparent cell debris, and the surface-associated granules are present at higher density at the coat surface. Finally, we showed that biofilm cells possess 100-fold greater resistance to the antibiotic metronidazole then do cells cultured in liquid media. Taken together, our data suggest that C. difficile cells and spores in biofilms have specialized properties that may facilitate infection.

  12. Spore formation and toxin production in Clostridium difficile biofilms.

    Science.gov (United States)

    Semenyuk, Ekaterina G; Laning, Michelle L; Foley, Jennifer; Johnston, Pehga F; Knight, Katherine L; Gerding, Dale N; Driks, Adam

    2014-01-01

    The ability to grow as a biofilm can facilitate survival of bacteria in the environment and promote infection. To better characterize biofilm formation in the pathogen Clostridium difficile, we established a colony biofilm culture method for this organism on a polycarbonate filter, and analyzed the matrix and the cells in biofilms from a variety of clinical isolates over several days of biofilm culture. We found that biofilms readily formed in all strains analyzed, and that spores were abundant within about 6 days. We also found that extracellular DNA (eDNA), polysaccharide and protein was readily detected in the matrix of all strains, including the major toxins A and/or B, in toxigenic strains. All the strains we analyzed formed spores. Apart from strains 630 and VPI10463, which sporulated in the biofilm at relatively low frequencies, the frequencies of biofilm sporulation varied between 46 and 65%, suggesting that variations in sporulation levels among strains is unlikely to be a major factor in variation in the severity of disease. Spores in biofilms also had reduced germination efficiency compared to spores obtained by a conventional sporulation protocol. Transmission electron microscopy revealed that in 3 day-old biofilms, the outermost structure of the spore is a lightly staining coat. However, after 6 days, material that resembles cell debris in the matrix surrounds the spore, and darkly staining granules are closely associated with the spores surface. In 14 day-old biofilms, relatively few spores are surrounded by the apparent cell debris, and the surface-associated granules are present at higher density at the coat surface. Finally, we showed that biofilm cells possess 100-fold greater resistance to the antibiotic metronidazole then do cells cultured in liquid media. Taken together, our data suggest that C. difficile cells and spores in biofilms have specialized properties that may facilitate infection.

  13. BIOFILM DETECTION AND CLINICAL SIGNIFICANCE OF STAPHYLOCOCCUS EPIDERMIDIS ISOLATES

    Directory of Open Access Journals (Sweden)

    Sudha Rani

    2015-04-01

    Full Text Available CONTEXT: Isolates of Staphylococcus epidermidis . AIMS : To detect biofilm among clinical isolates of Staphylococcus epidermidis . MATERIAL AND METHODS : 50 Staphylococcus epidermidis isolates were collected from clinical samples like blood, IV catheter tips, catheterized urine, wound swabs and exudates received from various clinical departments. Biofilm formation was studied in these isolates. The study was carried out over a period o f one year i. e from March 2011 - February 2012. The specimens received were processed by conventional methods. Tissue Culture plate method was used for detection of biofilm. RESULTS: IV catheter tip samples revealed 25%, implant device associated infection s revealed 20%, the Catheterized urine samples showed 16%, blood culture 8%, ventilator associated infections 10%, post - operative wound infections 11% & exudates 5% of Staphylococcus epidermidis isolates. Isolates with O. D. values more than 0. 2 were consi dered as high biofilm producers. 40% of S. epidermidis isolates were weak biofilm producers, 24% were moderate biofilm producers and 36% were high biofilm producers. Isolates from IV catheter tips showed high biofilm formation. CONCLUSIONS: In the modern h ealth care set up, various devices such as IV catheters, urine catheters, shunts, implanted prosthetic devices etc are being increasingly used thereby causing the device associated infections particularly of Staphylococcus epidermidis. Device associated in fections caused by Staphylococcus epidermidis are mainly due to biofilm formation which ultimately makes the treatment difficult.

  14. AFM Structural Characterization of Drinking Water Biofilm under Physiological Conditions

    Science.gov (United States)

    Due to the complexity of mixed culture drinking water biofilm, direct visual observation under in situ conditions has been challenging. In this study, atomic force microscopy (AFM) revealed the three dimensional morphology and arrangement of drinking water relevant biofilm in air...

  15. AFM Structural Characterization of Drinking Water Biofilm under Physiological Conditions

    Science.gov (United States)

    Due to the complexity of mixed culture drinking water biofilm, direct visual observation under in situ conditions has been challenging. In this study, atomic force microscopy (AFM) revealed the three dimensional morphology and arrangement of drinking water relevant biofilm in air...

  16. Pseudomonas aeruginosa biofilm infections

    DEFF Research Database (Denmark)

    Tolker-Nielsen, Tim

    2014-01-01

    Bacteria in natural, industrial and clinical settings predominantly live in biofilms, i.e., sessile structured microbial communities encased in self-produced extracellular matrix material. One of the most important characteristics of microbial biofilms is that the resident bacteria display...... a remarkable increased tolerance toward antimicrobial attack. Biofilms formed by opportunistic pathogenic bacteria are involved in devastating persistent medical device-associated infections, and chronic infections in individuals who are immune-compromised or otherwise impaired in the host defense. Because...... the use of conventional antimicrobial compounds in many cases cannot eradicate biofilms, there is an urgent need to develop alternative measures to combat biofilm infections. The present review is focussed on the important opportunistic pathogen and biofilm model organism Pseudomonas aeruginosa. Initially...

  17. Glucose-specific enzyme IIA has unique binding partners in the vibrio cholerae biofilm.

    Science.gov (United States)

    Pickering, Bradley S; Smith, Daniel R; Watnick, Paula I

    2012-11-06

    Glucose-specific enzyme IIA (EIIA(Glc)) is a central regulator of bacterial metabolism and an intermediate in the phosphoenolpyruvate phosphotransferase system (PTS), a conserved phosphotransfer cascade that controls carbohydrate transport. We previously reported that EIIA(Glc) activates transcription of the genes required for Vibrio cholerae biofilm formation. While EIIA(Glc) modulates the function of many proteins through a direct interaction, none of the known regulatory binding partners of EIIA(Glc) activates biofilm formation. Therefore, we used tandem affinity purification (TAP) to compare binding partners of EIIA(Glc) in both planktonic and biofilm cells. A surprising number of novel EIIA(Glc) binding partners were identified predominantly under one condition or the other. Studies of planktonic cells revealed established partners of EIIA(Glc), such as adenylate cyclase and glycerol kinase. In biofilms, MshH, a homolog of Escherichia coli CsrD, was found to be a dominant binding partner of EIIA(Glc). Further studies revealed that MshH inhibits biofilm formation. This function was independent of the Carbon storage regulator (Csr) pathway and dependent on EIIA(Glc). To explore the existence of multiprotein complexes centered on EIIA(Glc), we also affinity purified the binding partners of adenylate cyclase from biofilm cells. In addition to EIIA(Glc), this analysis yielded many of the same proteins that copurified with EIIA(Glc). We hypothesize that EIIA(Glc) serves as a hub for multiprotein complexes and furthermore that these complexes may provide a mechanism for competitive and cooperative interactions between binding partners. EIIA(Glc) is a global regulator of microbial physiology that acts through direct interactions with other proteins. This work represents the first demonstration that the protein partners of EIIA(Glc) are distinct in the microbial biofilm. Furthermore, it provides the first evidence that EIIA(Glc) may exist in multiprotein complexes with

  18. Biofilm characteristics and evaluation of the sanitation procedures of thermophilic Aeribacillus pallidus E334 biofilms.

    Science.gov (United States)

    Kilic, Tugba; Karaca, Basar; Ozel, Beste Piril; Ozcan, Birgul; Cokmus, Cumhur; Coleri Cihan, Arzu

    2017-04-01

    The ability of Aeribacillus pallidus E334 to produce pellicle and form a biofilm was studied. Optimal biofilm formation occurred at 60 °C, pH 7.5 and 1.5% NaCl. Extra polymeric substances (EPS) were composed of proteins and eDNA (21.4 kb). E334 formed biofilm on many surfaces, but mostly preferred polypropylene and glass. Using CLSM analysis, the network-like structure of the EPS was observed. The A. pallidus biofilm had a novel eDNA content. DNaseI susceptibility (86.8% removal) of eDNA revealed its importance in mature biofilms, but the purified eDNA was resistant to DNaseI, probably due to its extended folding outside the matrix. Among 15 cleaning agents, biofilms could be removed with alkaline protease and sodium dodecyl sulphate (SDS). The removal of cells from polypropylene and biomass on glass was achieved with combined SDS/alkaline protease treatment. Strong A. pallidus biofilms could cause risks for industrial processes and abiotic surfaces must be taken into consideration in terms of sanitation procedures.

  19. Studying bacterial multispecies biofilms

    DEFF Research Database (Denmark)

    Røder, Henriette Lyng; Sørensen, Søren Johannes; Burmølle, Mette

    2016-01-01

    and drawbacks of varying the degree of complexity. This review aims to facilitate multispecies biofilm research in order to expand the current limited knowledge on interspecies interactions. Recent technological advances have enabled total diversity analysis of highly complex and diverse microbial communities...... at the microscale of complex communities, including biofilms.Studies of multispecies biofilms and the interactions shaping these are conducted in traditional approaches used for single-species biofilms with some adjustments; but a crucial point for consideration is which strains to combine and where these should...

  20. Shared and distinct anatomical correlates of semantic and phonemic fluency revealed by lesion-symptom mapping in patients with ischemic stroke.

    Science.gov (United States)

    Biesbroek, J Matthijs; van Zandvoort, Martine J E; Kappelle, L Jaap; Velthuis, Birgitta K; Biessels, Geert Jan; Postma, Albert

    2016-05-01

    Semantic and phonemic fluency tasks are frequently used to test executive functioning, speed and attention, and access to the mental lexicon. In semantic fluency tasks, subjects are required to generate words belonging to a category (e.g., animals) within a limited time window, whereas in phonemic fluency tasks subjects have to generate words starting with a given letter. Anatomical correlates of semantic and phonemic fluency are currently assumed to overlap in left frontal structures, reflecting shared executive processes, and to be distinct in left temporal and right frontal structures, reflecting involvement of distinct memory processes and search strategies. Definite evidence for this assumption is lacking. To further establish the anatomical correlates of semantic and phonemic fluency, we applied assumption-free voxel-based and region-of-interest-based lesion-symptom mapping in 93 patients with ischemic stroke. Fluency was assessed by asking patients to name animals (semantic), and words starting with the letter N and A (phonemic). Our findings indicate that anatomical correlates of semantic and phonemic fluency overlap in the left inferior frontal gyrus and insula, reflecting shared underlying cognitive processes. Phonemic fluency additionally draws on the left rolandic operculum, which might reflect a search through phonological memory, and the middle frontal gyrus. Semantic fluency additionally draws on left medial temporal regions, probably reflecting a search through semantic memory, and the right inferior frontal gyrus, which might reflect the application of a visuospatial mental imagery strategy in semantic fluency. These findings establish shared and distinct anatomical correlates of semantic and phonemic fluency.

  1. MicroRNA transfection and AGO-bound CLIP-seq data sets reveal distinct determinants of miRNA action

    DEFF Research Database (Denmark)

    Wen, Jiayu; Parker, Brian J; Jacobsen, Anders

    2011-01-01

    the predictive effect of target flanking features. We observe distinct target determinants between expression-based and CLIP-based data. Target flanking features such as flanking region conservation are an important AGO-binding determinant-we hypothesize that CLIP experiments have a preference for strongly bound...... miRNP-target interactions involving adjacent RNA-binding proteins that increase the strength of cross-linking. In contrast, seed-related features are major determinants in expression-based studies, but less so for CLIP-seq studies, and increased miRNA concentrations typical of transfection studies...

  2. Hsp90 governs dispersion and drug resistance of fungal biofilms.

    Directory of Open Access Journals (Sweden)

    Nicole Robbins

    2011-09-01

    Full Text Available Fungal biofilms are a major cause of human mortality and are recalcitrant to most treatments due to intrinsic drug resistance. These complex communities of multiple cell types form on indwelling medical devices and their eradication often requires surgical removal of infected devices. Here we implicate the molecular chaperone Hsp90 as a key regulator of biofilm dispersion and drug resistance. We previously established that in the leading human fungal pathogen, Candida albicans, Hsp90 enables the emergence and maintenance of drug resistance in planktonic conditions by stabilizing the protein phosphatase calcineurin and MAPK Mkc1. Hsp90 also regulates temperature-dependent C. albicans morphogenesis through repression of cAMP-PKA signalling. Here we demonstrate that genetic depletion of Hsp90 reduced C. albicans biofilm growth and maturation in vitro and impaired dispersal of biofilm cells. Further, compromising Hsp90 function in vitro abrogated resistance of C. albicans biofilms to the most widely deployed class of antifungal drugs, the azoles. Depletion of Hsp90 led to reduction of calcineurin and Mkc1 in planktonic but not biofilm conditions, suggesting that Hsp90 regulates drug resistance through different mechanisms in these distinct cellular states. Reduction of Hsp90 levels led to a marked decrease in matrix glucan levels, providing a compelling mechanism through which Hsp90 might regulate biofilm azole resistance. Impairment of Hsp90 function genetically or pharmacologically transformed fluconazole from ineffectual to highly effective in eradicating biofilms in a rat venous catheter infection model. Finally, inhibition of Hsp90 reduced resistance of biofilms of the most lethal mould, Aspergillus fumigatus, to the newest class of antifungals to reach the clinic, the echinocandins. Thus, we establish a novel mechanism regulating biofilm drug resistance and dispersion and that targeting Hsp90 provides a much-needed strategy for improving

  3. Fluid dynamic effects on staphylococci bacteria biofilms

    Science.gov (United States)

    Sherman, Erica; Bayles, Kenneth; Endres, Jennifer; Wei, Timothy

    2016-11-01

    Staphylococcus aureus bacteria are able to form biofilms and distinctive tower structures that facilitate their ability to tolerate treatment and to spread within the human body. The formation of towers, which break off, get carried downstream and serve to initiate biofilms in other parts of the body are of particular interest here. It is known that flow conditions play a role in the development, dispersion and propagation of biofilms in general. The influence of flow on tower formation, however, is not at all understood. This work is focused on the effect of applied shear on tower development. The hypothesis being examined is that tower structures form within a specific range of shear stresses and that there is an as yet ill defined fluid dynamic phenomenon that occurs hours before a tower forms. In this study, a range of shear stresses is examined that brackets 0.6 dynes/cm2, the nominal shear stress where towers seem most likely to form. This talk will include µPTV measurements and cell density data indicating variations in flow and biofilm evolution as a function of the applied shear. Causal relations between flow and biofilm development will be discussed.

  4. Sulfate reducing bacteria and their activities in oil sands process-affected water biofilm

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Hong; Yu, Tong, E-mail: tong.yu@ualberta.ca; Liu, Yang, E-mail: yang.liu@ualberta.ca

    2015-12-01

    Biofilm reactors were constructed to grow stratified multispecies biofilm in oil sands process-affected water (OSPW) supplemented with growth medium. The development of sulfate reducing bacteria (SRB) within the biofilm and the biofilm treatment of OSPW were evaluated. The community structure and potential activity of SRB in the biofilm were investigated with H{sub 2}S microsensor measurements, dsrB gene-based denaturing gradient gel electrophoresis (DGGE), and the real time quantitative polymerase chain reaction (qPCR). Multispecies biofilm with a thickness of 1000 μm was successfully developed on engineered biocarriers. H{sub 2}S production was observed in the deeper anoxic zone of the biofilm from around 750 μm to 1000 μm below the bulk water-biofilm interface, revealing sulfate reduction in the deeper zone of the stratified biofilm. The biofilm removed chemical oxygen demand (COD), sulfate, and nitrogen. The study expands current knowledge of biofilm treatment of OSPW and the function of anaerobic SRB in OSPW biofilm, and thus provides information for future bioreactor development in the reclamation of OSPW. - Graphical abstract: The development of sulfate reducing bacteria (SRB) within Oil Sands Process-affected Water (OSPW) biofilm and the biofilm treatment of OSPW were evaluated by Liu and coworkers. Combined microsensor and molecular biology techniques were utilized in this study. Their results demonstrated that multispecies biofilm with a thickness of 1000 μm was successfully developed on engineered biocarriers. H{sub 2}S production was observed in the deeper anoxic zone of the biofilm from around 750 μm to 1000 μm below the bulk water-biofilm interface, revealing sulfate reduction in the deeper zone of the biofilm. The biofilm removed chemical oxygen demand (COD), sulfate, and nitrogen. - Highlights: • Biofilm in oil sands wastewater was developed on engineered biocarriers. • Bacterial community and in situ activity of SRB were studied in the

  5. A triple-emission fluorescent probe reveals distinctive amyloid fibrillar polymorphism of wild-type alpha-synuclein and its familial Parkinson's disease mutants.

    Science.gov (United States)

    Celej, M Soledad; Caarls, Wouter; Demchenko, Alexander P; Jovin, Thomas M

    2009-08-11

    Intracytoplasmic neuronal deposits containing amyloid fibrils of the 140-amino acid presynaptic protein alpha-synuclein (AS) are the hallmark of Parkinson's (PD) disease and related neurodegenerative disorders. Three point mutations (A53T, A30P, and E46K) are linked to early onset PD. Compared to the wild-type (WT) protein, the mutants aggregate faster in vitro, but their fibrillar products are quite similar. Using the extrinsic multiple-emission probe 4'-(diethylamino)-3-hydroxyflavone (FE), we demonstrate unique and distinct spectroscopic signatures for the amyloid fibrils formed by the WT and mutant AS, presumably indicative of subtle differences in supramolecular structure. The two well-separated emission bands of the FE probe originate from a proton transfer reaction in the excited state. The ratiometric response constitutes a sensitive, tunable reporter of microenvironmental properties such as polarity and hydrogen bonding. The very distinctive fluorescence spectra of the FE probe bound to the four AS variants reflect different tautomeric equilibria in the excited state and the existence of at least two different binding sites in the fibrils for the dye. Deconvolution of the two-dimensional excitation-emission spectra leads to estimations of different local dielectric constants and extents of hydration characteristic of the proteins. The sensitivity of such a simple external probe to conformational alterations induced by point mutations is unprecedented and provides new insight into key phenomena related to amyloid fibrils: plasticity, polymorphism, propagation of structural features, and structure-function relationships underlying toxicity.

  6. Prophage spontaneous activation promotes DNA release enhancing biofilm formation in Streptococcus pneumoniae.

    Directory of Open Access Journals (Sweden)

    Margarida Carrolo

    Full Text Available Streptococcus pneumoniae (pneumococcus is able to form biofilms in vivo and previous studies propose that pneumococcal biofilms play a relevant role both in colonization and infection. Additionally, pneumococci recovered from human infections are characterized by a high prevalence of lysogenic bacteriophages (phages residing quiescently in their host chromosome. We investigated a possible link between lysogeny and biofilm formation. Considering that extracellular DNA (eDNA is a key factor in the biofilm matrix, we reasoned that prophage spontaneous activation with the consequent bacterial host lysis could provide a source of eDNA, enhancing pneumococcal biofilm development. Monitoring biofilm growth of lysogenic and non-lysogenic pneumococcal strains indicated that phage-infected bacteria are more proficient at forming biofilms, that is their biofilms are characterized by a higher biomass and cell viability. The presence of phage particles throughout the lysogenic strains biofilm development implicated prophage spontaneous induction in this effect. Analysis of lysogens deficient for phage lysin and the bacterial major autolysin revealed that the absence of either lytic activity impaired biofilm development and the addition of DNA restored the ability of mutant strains to form robust biofilms. These findings establish that limited phage-mediated host lysis of a fraction of the bacterial population, due to spontaneous phage induction, constitutes an important source of eDNA for the S. pneumoniae biofilm matrix and that this localized release of eDNA favors biofilm formation by the remaining bacterial population.

  7. Suspended biofilm carrier and activated sludge removal of acidic pharmaceuticals

    DEFF Research Database (Denmark)

    Falås, Per; Baillon-Dhumez, Aude; Andersen, Henrik Rasmus

    2012-01-01

    Removal of seven active pharmaceutical substances (ibuprofen, ketoprofen, naproxen, diclofenac, clofibric acid, mefenamic acid, and gemfibrozil) was assessed by batch experiments, with suspended biofilm carriers and activated sludge from several full-scale wastewater treatment plants. A distinct...... and attached solids for the carriers) of diclofenac, ketoprofen, gemfibrozil, clofibric acid and mefenamic acid compared to the sludges. Among the target pharmaceuticals, only ibuprofen and naproxen showed similar removal rates per unit biomass for the sludges and biofilm carriers. In contrast...

  8. Natural antigenic differences in the functionally equivalent extracellular DNABII proteins of bacterial biofilms provide a means for targeted biofilm therapeutics.

    Science.gov (United States)

    Rocco, C J; Davey, M E; Bakaletz, L O; Goodman, S D

    2017-04-01

    Bacteria that persist in the oral cavity exist within complex biofilm communities. A hallmark of biofilms is the presence of an extracellular polymeric substance (EPS), which consists of polysaccharides, extracellular DNA (eDNA), and proteins, including the DNABII family of proteins. The removal of DNABII proteins from a biofilm results in the loss of structural integrity of the eDNA and the collapse of the biofilm structure. We examined the role of DNABII proteins in the biofilm structure of the periodontal pathogen Porphyromonas gingivalis and the oral commensal Streptococcus gordonii. Co-aggregation with oral streptococci is thought to facilitate the establishment of P. gingivalis within the biofilm community. We demonstrate that DNABII proteins are present in the EPS of both S. gordonii and P. gingivalis biofilms, and that these biofilms can be disrupted through the addition of antisera derived against their respective DNABII proteins. We provide evidence that both eDNA and DNABII proteins are limiting in S. gordonii but not in P. gingivalis biofilms. In addition, these proteins are capable of complementing one another functionally. We also found that whereas antisera derived against most DNABII proteins are capable of binding a wide variety of DNABII proteins, the P. gingivalis DNABII proteins are antigenically distinct. The presence of DNABII proteins in the EPS of these biofilms and the antigenic uniqueness of the P. gingivalis proteins provide an opportunity to develop therapies that are targeted to remove P. gingivalis and biofilms that contain P. gingivalis from the oral cavity. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  9. Development of Spatial Distribution Patterns by Biofilm Cells

    DEFF Research Database (Denmark)

    Haagensen, Janus Anders Juul; Hansen, Susse Kirkelund; Bak Christensen, Bjarke

    2015-01-01

    Confined spatial patterns of microbial distribution are prevalent in nature, such as in microbial mats, soil communities, and water stream biofilms. The symbiotic two-species consortium of Pseudomonas putida and Acinetobacter sp. C6, originally isolated from a creosote-polluted aquifer, has evolved...... a distinct spatial organization in the laboratory that is characterized by an increased fitness and productivity. In this consortium, P. putida is reliant on microcolonies formed by Acinetobacter sp. C6 — to which it attaches. Here we describe the processes that lead to the microcolony......-pattern by Acinetobacter sp. C6. Ecological spatial pattern analyses revealed that the microcolonies were not entirely randomly distributed, and instead arranged in a uniform pattern. Detailed time-lapse confocal microscopy at the single cell level demonstrated that the spatial pattern was the result of an intriguing self...

  10. Single-atom imino substitutions at A9 and A10 reveal distinct effects on the fold and function of the hairpin ribozyme catalytic core.

    Science.gov (United States)

    Spitale, Robert C; Volpini, Rosaria; Mungillo, Michael V; Krucinska, Jolanta; Cristalli, Gloria; Wedekind, Joseph E

    2009-08-25

    The hairpin ribozyme cleaves a phosphodiester bond within a cognate substrate. Structural and biochemical data indicate the conserved A9 and A10 bases reside close to the scissile bond but make distinct contributions to catalysis. To investigate these residues, we replaced the imino moiety of each base with N1-deazaadenosine. This single-atom change resulted in an 8-fold loss in k(obs) for A9 and displacement of the base from the active site; no effects were observed for A10. We propose that the imino moiety of A9 promotes a key water-mediated contact that favors transition-state formation, which suggests an enhanced chemical repertoire for RNA.

  11. Genome-wide Functional Analysis of CREB/Long-Term Memory-Dependent Transcription Reveals Distinct Basal and Memory Gene Expression Programs

    Science.gov (United States)

    Lakhina, Vanisha; Arey, Rachel N.; Kaletsky, Rachel; Kauffman, Amanda; Stein, Geneva; Keyes, William; Xu, Daniel; Murphy, Coleen T.

    2014-01-01

    SUMMARY Induced CREB activity is a hallmark of long-term memory, but the full repertoire of CREB transcriptional targets required specifically for memory is not known in any system. To obtain a more complete picture of the mechanisms involved in memory, we combined memory training with genome-wide transcriptional analysis of C. elegans CREB mutants. This approach identified 757 significant CREB/memory-induced targets and confirmed the involvement of known memory genes from other organisms, but also suggested new mechanisms and novel components that may be conserved through mammals. CREB mediates distinct basal and memory transcriptional programs at least partially through spatial restriction of CREB activity: basal targets are regulated primarily in nonneuronal tissues, while memory targets are enriched for neuronal expression, emanating from CREB activity in AIM neurons. This suite of novel memory-associated genes will provide a platform for the discovery of orthologous mammalian long-term memory components. PMID:25611510

  12. Genome-wide functional analysis of CREB/long-term memory-dependent transcription reveals distinct basal and memory gene expression programs.

    Science.gov (United States)

    Lakhina, Vanisha; Arey, Rachel N; Kaletsky, Rachel; Kauffman, Amanda; Stein, Geneva; Keyes, William; Xu, Daniel; Murphy, Coleen T

    2015-01-21

    Induced CREB activity is a hallmark of long-term memory, but the full repertoire of CREB transcriptional targets required specifically for memory is not known in any system. To obtain a more complete picture of the mechanisms involved in memory, we combined memory training with genome-wide transcriptional analysis of C. elegans CREB mutants. This approach identified 757 significant CREB/memory-induced targets and confirmed the involvement of known memory genes from other organisms, but also suggested new mechanisms and novel components that may be conserved through mammals. CREB mediates distinct basal and memory transcriptional programs at least partially through spatial restriction of CREB activity: basal targets are regulated primarily in nonneuronal tissues, while memory targets are enriched for neuronal expression, emanating from CREB activity in AIM neurons. This suite of novel memory-associated genes will provide a platform for the discovery of orthologous mammalian long-term memory components.

  13. Locked Nucleic Acid-Based In Situ Hybridization Reveals miR-7a as a Hypothalamus-Enriched MicroRNA with a Distinct Expression Pattern

    DEFF Research Database (Denmark)

    Herzer, S; Silahtaroglu, A; Meister, B

    2012-01-01

    MicroRNAs (miRNAs) are short (22 nucleotides) non-coding ribonucleic acid (RNA) molecules that post-transcriptionally repress expression of protein-coding genes by binding to 3'-untranslated regions of the target mRNAs. In order to identify miRNAs selectively expressed within the hypothalamus...... present in the hypothalamus, miR-7a, was the only miRNA found to be enriched in the hypothalamus, with low or no expression in other parts of the central nervous system (CNS). Within the hypothalamus, strong miR-7a expression was distinct and restricted to some hypothalamic nuclei and adjacent areas. mi......R-7a expression was particularly prominent in the subfornical organ, suprachiasmatic, paraventricular, periventricular, supraoptic, dorsomedial and arcuate nuclei. Identical expression patterns for miR-7a was seen in mouse and rat hypothalamus. By combining LNA-FISH with immunohistochemistry...

  14. Evaluation of intraspecies interactions in biofilm formation by Methylobacterium species isolated from pink-pigmented household biofilms.

    Science.gov (United States)

    Xu, Fang-Fang; Morohoshi, Tomohiro; Wang, Wen-Zhao; Yamaguchi, Yuka; Liang, Yan; Ikeda, Tsukasa

    2014-01-01

    Concern regarding household biofilms has grown due to their widespread existence and potential to threaten human health by serving as pathogen reservoirs. Previous studies identified Methylobacterium as one of the dominant genera found in household biofilms. In the present study, we examined the mechanisms underlying biofilm formation by using the bacterial consortium found in household pink slime. A clone library analysis revealed that Methylobacterium was the predominant genus in household pink slime. In addition, 16 out of 21 pink-pigmented bacterial isolates were assigned to the genus Methylobacterium. Although all of the Methylobacterium isolates formed low-level biofilms, the amount of the biofilms formed by Methylobacterium sp. P-1M and P-18S was significantly increased by co-culturing with other Methylobacterium strains that belonged to a specific phylogenetic group. The single-species biofilm was easily washed from the glass surface, whereas the dual-species biofilm strongly adhered after washing. A confocal laser scanning microscopy analysis showed that the dual-species biofilms were significantly thicker and tighter than the single-species biofilms.

  15. Mycobacterium avium Possesses Extracellular DNA that Contributes to Biofilm Formation, Structural Integrity, and Tolerance to Antibiotics.

    Science.gov (United States)

    Rose, Sasha J; Babrak, Lmar M; Bermudez, Luiz E

    2015-01-01

    Mycobacterium avium subsp. hominissuis is an opportunistic pathogen that is associated with biofilm-related infections of the respiratory tract and is difficult to treat. In recent years, extracellular DNA (eDNA) has been found to be a major component of bacterial biofilms, including many pathogens involved in biofilm-associated infections. To date, eDNA has not been described as a component of mycobacterial biofilms. In this study, we identified and characterized eDNA in a high biofilm-producing strain of Mycobacterium avium subsp. hominissuis (MAH). In addition, we surveyed for presence of eDNA in various MAH strains and other nontuberculous mycobacteria. Biofilms of MAH A5 (high biofilm-producing strain) and MAH 104 (reference strain) were established at 22°C and 37°C on abiotic surfaces. Acellular biofilm matrix and supernatant from MAH A5 7 day-old biofilms both possess abundant eDNA, however very little eDNA was found in MAH 104 biofilms. A survey of MAH clinical isolates and other clinically relevant nontuberculous mycobacterial species revealed many species and strains that also produce eDNA. RAPD analysis demonstrated that eDNA resembles genomic DNA. Treatment with DNase I reduced the biomass of MAH A5 biofilms when added upon biofilm formation or to an already established biofilm both on abiotic surfaces and on top of human pharyngeal epithelial cells. Furthermore, co-treatment of an established biofilm with DNase 1 and either moxifloxacin or clarithromycin significantly increased the susceptibility of the bacteria within the biofilm to these clinically used antimicrobials. Collectively, our results describe an additional matrix component of mycobacterial biofilms and a potential new target to help treat biofilm-associated nontuberculous mycobacterial infections.

  16. Lipopeptide biosurfactant viscosin enhances dispersal of Pseudomonas fluorescens SBW25 biofilms.

    Science.gov (United States)

    Bonnichsen, Lise; Bygvraa Svenningsen, Nanna; Rybtke, Morten; de Bruijn, Irene; Raaijmakers, Jos M; Tolker-Nielsen, Tim; Nybroe, Ole

    2015-12-01

    Pseudomonads produce several lipopeptide biosurfactants that have antimicrobial properties but that also facilitate surface motility and influence biofilm formation. Detailed studies addressing the significance of lipopeptides for biofilm formation and architecture are rare. Hence, the present study sets out to determine the specific role of the lipopeptide viscosin in Pseudomonas fluorescens SBW25 biofilm formation, architecture and dispersal, and to relate viscA gene expression to viscosin production and effect. Initially, we compared biofilm formation of SBW25 and the viscosin-deficient mutant strain SBW25ΔviscA in static microtitre assays. These experiments demonstrated that viscosin had little influence on the amount of biofilm formed by SBW25 during the early stages of biofilm development. Later, however, SBW25 formed significantly less biofilm than SBW25ΔviscA. The indication that viscosin is involved in biofilm dispersal was confirmed by chemical complementation of the mutant biofilm. Furthermore, a fluorescent bioreporter showed that viscA expression was induced in biofilms 4 h prior to dispersal. Subsequent detailed studies of biofilms formed in flow cells for up to 5 days revealed that SBW25 and SBW25ΔviscA developed comparable biofilms dominated by well-defined, mushroom-shaped structures. Carbon starvation was required to obtain biofilm dispersal in this system. Dispersal of SBW25 biofilms was significantly greater than of SBW25ΔviscA biofilms after 3 h and, importantly, carbon starvation strongly induced viscA expression, in particular for cells that were apparently leaving the biofilm. Thus, the present study points to a role for viscosin-facilitated motility in dispersal of SBW25 biofilms.

  17. Phylogenetic analysis and polyphasic characterization of Clavibacter michiganensis strains isolated from tomato seeds reveal that nonpathogenic strains are distinct from C. michiganensis subsp. michiganensis.

    Science.gov (United States)

    Jacques, Marie-Agnès; Durand, Karine; Orgeur, Geoffrey; Balidas, Samuel; Fricot, Céline; Bonneau, Sophie; Quillévéré, Anne; Audusseau, Corinne; Olivier, Valérie; Grimault, Valérie; Mathis, René

    2012-12-01

    The genus Clavibacter comprises one species and five subspecies of plant-pathogenic bacteria, four of which are classified as quarantine organisms due to the high economic threat they pose. Clavibacter michiganensis subsp. michiganensis is one of the most important pathogens of tomato, but the recommended diagnostic tools are not satisfactory due to false-negative and/or -positive results. To provide a robust analysis of the genetic relatedness among a worldwide collection of C. michiganensis subsp. michiganensis strains, relatives (strains from the four other C. michiganensis subspecies), and nonpathogenic Clavibacter-like strains isolated from tomato, we performed multilocus sequence-based analysis and typing (MLSA and MLST) based on six housekeeping genes (atpD, dnaK, gyrB, ppK, recA, and rpoB). We compared this "framework" with phenotypic and genotypic characteristics such as pathogenicity on tomato, reaction to two antisera by immunofluorescence and to five PCR identification tests, and the presence of four genes encoding the main C. michiganensis subsp. michiganensis pathogenicity determinants. We showed that C. michiganensis subsp. michiganensis is monophyletic and is distinct from its closest taxonomic neighbors. The nonpathogenic Clavibacter-like strains were identified as C. michiganensis using 16S rRNA gene sequencing. These strains, while cross-reacting with C. michiganensis subsp. michiganensis identification tools, are phylogenetically distinct from the pathogenic strains but belong to the C. michiganensis clade. C. michiganensis subsp. michiganensis clonal complexes linked strains from highly diverse geographical origins and also strains isolated over long periods of time in the same location. This illustrates the importance of seed transmission in the worldwide dispersion of this pathogen and its survival and adaptation abilities in a new environment once introduced.

  18. Shared as well as distinct roles of EHD proteins revealed by biochemical and functional comparisons in mammalian cells and C. elegans

    Directory of Open Access Journals (Sweden)

    Gao Qingshen

    2007-01-01

    Full Text Available Abstract Background The four highly homologous human EHD proteins (EHD1-4 form a distinct subfamily of the Eps15 homology domain-containing protein family and are thought to regulate endocytic recycling. Certain members of this family have been studied in different cellular contexts; however, a lack of concurrent analyses of all four proteins has impeded an appreciation of their redundant versus distinct functions. Results Here, we analyzed the four EHD proteins both in mammalian cells and in a cross-species complementation assay using a C. elegans mutant lacking the EHD ortholog RME-1. We show that all human EHD proteins rescue the vacuolated intestinal phenotype of C. elegans rme-1 mutant, are simultaneously expressed in a panel of mammalian cell lines and tissues tested, and variably homo- and hetero-oligomerize and colocalize with each other and Rab11, a recycling endosome marker. Small interfering RNA (siRNA knock-down of EHD1, 2 and 4, and expression of dominant-negative EH domain deletion mutants showed that loss of EHD1 and 3 (and to a lesser extent EHD4 but not EHD2 function retarded transferrin exit from the endocytic recycling compartment. EH domain deletion mutants of EHD1 and 3 but not 2 or 4, induced a striking perinuclear clustering of co-transfected Rab11. Knock-down analyses indicated that EHD1 and 2 regulate the exit of cargo from the recycling endosome while EHD4, similar to that reported for EHD3 (Naslavsky et al. (2006 Mol. Biol. Cell 17, 163, regulates transport from the early endosome to the recycling endosome. Conclusion Altogether, our studies suggest that concurrently expressed human EHD proteins perform shared as well as discrete functions in the endocytic recycling pathway and lay a foundation for future studies to identify and characterize the molecular pathways involved.

  19. Absolute quantification of acetylation and phosphorylation of the histone variant H2AX upon ionizing radiation reveals distinct cellular responses in two cancer cell lines.

    Science.gov (United States)

    Matsuda, Shun; Furuya, Kanji; Ikura, Masae; Matsuda, Tomonari; Ikura, Tsuyoshi

    2015-11-01

    Histone modifications change upon the cellular response to ionizing radiation, and their cellular amounts could reflect the DNA damage response activity. We previously reported a sensitive and reliable method for the absolute quantification of γH2AX within cells, using liquid chromatography-tandem mass spectrometry (LC/MS/MS). The technique has broad adaptability to a variety of biological systems and can quantitate different modifications of histones. In this study, we applied it to quantitate the levels of γH2AX and K5-acetylated H2AX, and to compare the radiation responses between two cancer cell lines: HeLa and U-2 OS. The two cell lines have distinct properties in terms of their H2AX modifications. HeLa cells have relatively high γH2AX (3.1 %) against the total H2AX even in un-irradiated cells, while U-2 OS cells have an essentially undetectable level (nearly 0 %) of γH2AX. In contrast, the amounts of acetylated histones are lower in HeLa cells (9.3 %) and higher in U-2 OS cells (24.2 %) under un-irradiated conditions. Furthermore, after ionizing radiation exposure, the time-dependent increases and decreases in the amounts of histone modifications differed between the two cell lines, especially at the early time points. These results suggest that each biological system has distinct kinase/phosphatase and/or acetylase/deacetylase activities. In conclusion, for the first time, we have succeeded in simultaneously monitoring the absolute amounts of phosphorylated and acetylated cellular H2AX after ionizing radiation exposure. This multi-criteria assessment enables precise comparisons of the effects of radiation between any biological systems.

  20. ECoG high gamma activity reveals distinct cortical representations of lyrics passages, harmonic and timbre-related changes in a rock song.

    Science.gov (United States)

    Sturm, Irene; Blankertz, Benjamin; Potes, Cristhian; Schalk, Gerwin; Curio, Gabriel

    2014-01-01

    Listening to music moves our minds and moods, stirring interest in its neural underpinnings. A multitude of compositional features drives the appeal of natural music. How such original music, where a composer's opus is not manipulated for experimental purposes, engages a listener's brain has not been studied until recently. Here, we report an in-depth analysis of two electrocorticographic (ECoG) data sets obtained over the left hemisphere in ten patients during presentation of either a rock song or a read-out narrative. First, the time courses of five acoustic features (intensity, presence/absence of vocals with lyrics, spectral centroid, harmonic change, and pulse clarity) were extracted from the audio tracks and found to be correlated with each other to varying degrees. In a second step, we uncovered the specific impact of each musical feature on ECoG high-gamma power (70-170 Hz) by calculating partial correlations to remove the influence of the other four features. In the music condition, the onset and offset of vocal lyrics in ongoing instrumental music was consistently identified within the group as the dominant driver for ECoG high-gamma power changes over temporal auditory areas, while concurrently subject-individual activation spots were identified for sound intensity, timbral, and harmonic features. The distinct cortical activations to vocal speech-related content embedded in instrumental music directly demonstrate that song integrated in instrumental music represents a distinct dimension in complex music. In contrast, in the speech condition, the full sound envelope was reflected in the high gamma response rather than the onset or offset of the vocal lyrics. This demonstrates how the contributions of stimulus features that modulate the brain response differ across the two examples of a full-length natural stimulus, which suggests a context-dependent feature selection in the processing of complex auditory stimuli.

  1. ECoG high gamma activity reveals distinct cortical representations of lyrics passages, harmonic and timbre-related changes in a rock song

    Directory of Open Access Journals (Sweden)

    Irene eSturm

    2014-10-01

    Full Text Available Listening to music moves our minds and moods, stirring interest in its neural underpinnings. A multitude of compositional features drives the appeal of natural music. How such original music, where a composer's opus is not manipulated for experimental purposes, engages a listener's brain has not been studied until recently. Here, we report an in-depth analysis of two electrocorticographic (ECoG data sets obtained over the left hemisphere in ten patients during presentation of either a rock song or a read-out narrative. First, the time courses of five acoustic features (intensity, presence/absence of vocals with lyrics, spectral centroid, harmonic change, and pulse clarity were extracted from the audio tracks and found to be correlated with each other to varying degrees. In a second step, we uncovered the specific impact of each musical feature on ECoG high-gamma power (70-170 Hz by calculating partial correlations to remove the influence of the other four features. In the music condition, the onset and offset of vocal lyrics in ongoing instrumental music was consistently within the subject group identified as the dominant driver for ECoG high-gamma power changes over temporal auditory areas, while concurrently subject-individual activation spots were identified for sound intensity, timbral and harmonic features. The distinct cortical activations to vocal speech-related content embedded in instrumental music directly demonstrate that song overlaid on instrumental music represents a distinct dimension in complex music. In contrast, in the speech condition, the full sound envelope was reflected in the high gamma response rather than the onset or offset of the vocal lyrics. This demonstrates how the contributions of stimulus features that modulate the brain response differ across the two examples of a full-length natural stimulus, which suggests a context-dependent feature selection in the processing of complex auditory stimuli.

  2. Hydraulic resistance of biofilms

    KAUST Repository

    Dreszer, C.

    2013-02-01

    Biofilms may interfere with membrane performance in at least three ways: (i) increase of the transmembrane pressure drop, (ii) increase of feed channel (feed-concentrate) pressure drop, and (iii) increase of transmembrane passage. Given the relevance of biofouling, it is surprising how few data exist about the hydraulic resistance of biofilms that may affect the transmembrane pressure drop and membrane passage. In this study, biofilms were generated in a lab scale cross flow microfiltration system at two fluxes (20 and 100Lm-2h-1) and constant cross flow (0.1ms-1). As a nutrient source, acetate was added (1.0mgL-1 acetate C) besides a control without nutrient supply. A microfiltration (MF) membrane was chosen because the MF membrane resistance is very low compared to the expected biofilm resistance and, thus, biofilm resistance can be determined accurately. Transmembrane pressure drop was monitored. As biofilm parameters, thickness, total cell number, TOC, and extracellular polymeric substances (EPS) were determined, it was demonstrated that no internal membrane fouling occurred and that the fouling layer actually consisted of a grown biofilm and was not a filter cake of accumulated bacterial cells. At 20Lm-2h-1 flux with a nutrient dosage of 1mgL-1 acetate C, the resistance after 4 days reached a value of 6×1012m-1. At 100Lm-2h-1 flux under the same conditions, the resistance was 5×1013m-1. No correlation of biofilm resistance to biofilm thickness was found; Biofilms with similar thickness could have different resistance depending on the applied flux. The cell number in biofilms was between 4×107 and 5×108 cellscm-2. At this number, bacterial cells make up less than a half percent of the overall biofilm volume and therefore did not hamper the water flow through the biofilm significantly. A flux of 100Lm-2h-1 with nutrient supply caused higher cell numbers, more biomass, and higher biofilm resistance than a flux of 20Lm-2h-1. However, the biofilm thickness

  3. Diffusion retardation by binding of tobramycin in an alginate biofilm model

    OpenAIRE

    Cao, Bao; Christophersen, Lars; Kolpen, Mette; Jensen, Peter Østrup; Sneppen, Kim; Høiby, Niels; Moser, Claus; Sams, Thomas

    2016-01-01

    Microbial cells embedded in a self-produced extracellular biofilm matrix cause chronic infections, e. g. by Pseudomonas aeruginosa in the lungs of cystic fibrosis patients. The antibiotic killing of bacteria in biofilms is generally known to be reduced by 100-1000 times relative to planktonic bacteria. This makes such infections difficult to treat. We have therefore proposed that biofilms can be regarded as an independent compartment with distinct pharmacokinetics. To elucidate this pharmacok...

  4. Recursive Distinctioning

    CERN Document Server

    Isaacson, Joel

    2016-01-01

    Recursive distinctioning (RD) is a name coined by Joel Isaacson in his original patent document describing how fundamental patterns of process arise from the systematic application of operations of distinction and description upon themselves. Recursive distinctioning means just what it says. A pattern of distinctions is given in a space based on a graphical structure (such as a line of print or a planar lattice or given graph). Each node of the graph is occupied by a letter from some arbitrary alphabet. A specialized alphabet is given that can indicate distinctions about neighbors of a given node. The neighbors of a node are all nodes that are connected to the given node by edges in the graph. The letters in the specialized alphabet (call it SA) are used to describe the states of the letters in the given graph and at each stage in the recursion, letters in SA are written at all nodes in the graph, describing its previous state. The recursive structure that results from the iteration of descriptions is called ...

  5. Meningococcal biofilm formation

    DEFF Research Database (Denmark)

    Lappann, M.; Haagensen, Janus Anders Juul; Claus, H.

    2006-01-01

    We show that in a standardized in vitro flow system unencapsulated variants of genetically diverse lineages of Neisseria meningitidis formed biofilms, that could be maintained for more than 96 h. Biofilm cells were resistant to penicillin, but not to rifampin or ciprofloxacin. For some strains......, microcolony formation within biofilms was observed. Microcolony formation in strain MC58 depended on a functional copy of the pilE gene encoding the pilus subunit pilin, and was associated with twitching of cells. Nevertheless, unpiliated pilE mutants formed biofilms showing that attachment and accumulation......X alleles was identified among genetically diverse meningococcal strains. PilX alleles differed in their propensity to support autoaggregation of cells in suspension, but not in their ability to support microcolony formation within biofilms in the continuous flow system....

  6. Meningococcal biofilm formation

    DEFF Research Database (Denmark)

    Lappann, M.; Haagensen, Janus Anders Juul; Claus, H.

    2006-01-01

    We show that in a standardized in vitro flow system unencapsulated variants of genetically diverse lineages of Neisseria meningitidis formed biofilms, that could be maintained for more than 96 h. Biofilm cells were resistant to penicillin, but not to rifampin or ciprofloxacin. For some strains......, microcolony formation within biofilms was observed. Microcolony formation in strain MC58 depended on a functional copy of the pilE gene encoding the pilus subunit pilin, and was associated with twitching of cells. Nevertheless, unpiliated pilE mutants formed biofilms showing that attachment and accumulation......X alleles was identified among genetically diverse meningococcal strains. PilX alleles differed in their propensity to support autoaggregation of cells in suspension, but not in their ability to support microcolony formation within biofilms in the continuous flow system....

  7. Gene-Metabolite Networks of Volatile Metabolism in Airen and Tempranillo Grape Cultivars Revealed a Distinct Mechanism of Aroma Bouquet Production

    Science.gov (United States)

    Rambla, José L.; Trapero-Mozos, Almudena; Diretto, Gianfranco; Rubio-Moraga, Angela; Granell, Antonio; Gómez-Gómez, Lourdes; Ahrazem, Oussama

    2016-01-01

    Volatile compounds are the major determinants of aroma and flavor in both grapes and wine. In this study, we investigated the emission of volatile and non-volatile compounds during berry maturation in two grape varieties (Airén and Tempranillo) throughout 2010 and 2011. HS-SPME coupled to gas chromatography and mass spectrometry was applied for the identification and relative quantitation of these compounds. Principal component analysis was performed to search for variability between the two cultivars and evolution during 10 developmental stages. Results showed that there are distinct differences in volatile compounds between cultivars throughout fruit development. Early stages were characterized in both cultivars by higher levels of some apocarotenoids such as β-cyclocitral or β-ionone, terpenoids (E)-linalool oxide and (Z)-linalool oxide and several furans, while the final stages were characterized by the highest amounts of ethanol, benzenoid phenylacetaldehyde and 2-phenylethanol, branched-amino acid-derived 3-methylbutanol and 2-methylbutanol, and a large number of lipid derivatives. Additionally, we measured the levels of the different classes of volatile precursors by using liquid chromatography coupled to high resolution mass spectrometry. In both varieties, higher levels of carotenoid compounds were detected in the earlier stages, zeaxanthin and α-carotene were only detected in Airén while neoxanthin was found only in Tempranillo; more variable trends were observed in the case of the other volatile precursors. Furthermore, we monitored the expression of homolog genes of a set of transcripts potentially involved in the biosynthesis of these metabolites, such as some glycosyl hydrolases family 1, lipoxygenases, alcohol dehydrogenases hydroperoxide lyases, O-methyltransferases and carotenoid cleavage dioxygenases during the defined developmental stages. Finally, based on Pearson correlation analyses, we explored the metabolite-metabolite fluctuations

  8. Antimicrobial and biofilm inhibiting diketopiperazines.

    Science.gov (United States)

    de Carvalho, M P; Abraham, W-R

    2012-01-01

    Diketopiperazines are the smallest cyclic peptides known. 90% of Gram-negative bacteria produce diketopiperazines and they have also been isolated from Gram-positive bacteria, fungi and higher organisms. Biosynthesis of cyclodipeptides can be achieved by dedicated nonribosomal peptide synthetases or by a novel type of synthetases named cyclopeptide synthases. Since the first report in 1924 a large number of bioactive diketopiperazines was discovered spanning activities as antitumor, antiviral, antifungal, antibacterial, antiprion, antihyperglycemic or glycosidase inhibitor agents. As infections are of increasing concern for human health and resistances against existing antibiotics are growing this review focuses on the antimicrobial activities of diketopiperazines. The antibiotic bicyclomycin is a diketopiperazine and structure activity studies revealed the unique nature of this compound which was finally developed for clinical applications. The antimicrobial activities of a number of other diketopiperazines along with structure activity relationships are discussed. Here a special focus is on the activity-toxicity problem of many compounds setting tight limitations to their application as drugs. Not only these classical antimicrobial activities but also proposed action in modulating bacterial communication as a new target to control biofilms will be evaluated. Pathogens organized in biofilms are difficult to eradicate because of the increase of their tolerance for antibiotics for several orders. Diketopiperazines were reported to modulate LuxR-mediated quorum-sensing systems of bacteria, and they are considered to influence cell-cell signaling offering alternative ways of biofilm control by interfering with microbial communication. Concluding the review we will finally discuss the potential of diketopiperazines in the clinic to erase biofilm infections.

  9. Geomicrobiology and hopanoid content of sulfidic subsurface vent biofilms, Little Salt Spring, Florida

    Science.gov (United States)

    Yang, E.; Schaperdoth, I.; Albrecht, H.; Freeman, K. H.; Macalady, J. L.

    2008-12-01

    Sulfide-rich, oxygen-poor environments are widespread in the subsurface and were prevalent at the earth's surface during critical intervals in the geologic past. Modern microbial communities in sulfidic niches have the potential to shed light on the biogeochemistry and biosignatures of anoxia and euxinia in earth history. Caves and sinkholes provide rare windows into microbially-dominated, sulfidic subsurface environments that are otherwise difficult and expensive to access. Little Salt Spring (Sarasota County, Florida) is a cover-collapse sinkhole lake with oxic surface water and anoxic, sulfidic bottom water (Alvarez Zarikian 2005). The site is famous for excellent preservation of human and animal archaeological remains (Clausen 1979), and its microbiology has never been investigated. Abundant white biofilms develop seasonally at a warm vent that feeds into the anoxic bottom water at 73 m depth below the water surface. The biofilms are of interest both as potential sources of biomarker compounds and because of their likely role in sulfuric acid production and limestone dissolution (speleogenesis). Biofilm samples were collected by expert science divers and investigated using microscopy, nucleic acid, and lipid analytical methods. Microscopy of the live biofilm revealed clusters of microbial filaments with holdfasts and dendritic, sulfur-rich colonial structures similar to those described in the 1960s for Thiobacterium, a sulfur-oxidizing genus with undetermined phylogeny. A 16S rDNA library constructed from the biofilm was split into three main phylotypes, with multiple clones representing (1) a Betaproteobacterial clade with no cultivated representatives, (2) filamentous Epsilonproteobacteria, and (3) a major bacterial lineage without named isolates (OP11/OD2). A full cycle rRNA approach is currently underway to link 16S rDNA phylotypes with specific populations in the biofilm. We confirmed using fluorescence in situ hybridization (FISH) that abundant

  10. Pseudomonas aeruginosa Biofilm Response and Resistance to Cold Atmospheric Pressure Plasma Is Linked to the Redox-Active Molecule Phenazine.

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    Anne Mai-Prochnow

    Full Text Available Pseudomonas aeruginosa is an important opportunistic pathogen displaying high antibiotic resistance. Its resistance is in part due to its outstanding ability to form biofilms on a range of biotic and abiotic surfaces leading to difficult-to-treat, often long-term infections. Cold atmospheric plasma (CAP is a new, promising antibacterial treatment to combat antibiotic-resistant bacteria. Plasma is ionized gas that has antibacterial properties through the generation of a mix of reactive oxygen and nitrogen species (RONS, excited molecules, charged particles and UV photons. Our results show the efficient removal of P. aeruginosa biofilms using a plasma jet (kINPen med, with no viable cells detected after 5 min treatment and no attached biofilm cells visible with confocal microscopy after 10 min plasma treatment. Because of its multi-factorial action, it is widely presumed that the development of bacterial resistance to plasma is unlikely. However, our results indicate that a short plasma treatment (3 min may lead to the emergence of a small number of surviving cells exhibiting enhanced resistance to subsequent plasma exposure. Interestingly, these cells also exhibited a higher degree of resistance to hydrogen peroxide. Whole genome comparison between surviving cells and control cells revealed 10 distinct polymorphic regions, including four belonging to the redox active, antibiotic pigment phenazine. Subsequently, the interaction between phenazine production and CAP resistance was demonstrated in biofilms of transposon mutants disrupted in different phenazine pathway genes which exhibited significantly altered sensitivity to CAP.

  11. Pseudomonas aeruginosa Biofilm Response and Resistance to Cold Atmospheric Pressure Plasma Is Linked to the Redox-Active Molecule Phenazine.

    Science.gov (United States)

    Mai-Prochnow, Anne; Bradbury, Mark; Ostrikov, Kostya; Murphy, Anthony B

    2015-01-01

    Pseudomonas aeruginosa is an important opportunistic pathogen displaying high antibiotic resistance. Its resistance is in part due to its outstanding ability to form biofilms on a range of biotic and abiotic surfaces leading to difficult-to-treat, often long-term infections. Cold atmospheric plasma (CAP) is a new, promising antibacterial treatment to combat antibiotic-resistant bacteria. Plasma is ionized gas that has antibacterial properties through the generation of a mix of reactive oxygen and nitrogen species (RONS), excited molecules, charged particles and UV photons. Our results show the efficient removal of P. aeruginosa biofilms using a plasma jet (kINPen med), with no viable cells detected after 5 min treatment and no attached biofilm cells visible with confocal microscopy after 10 min plasma treatment. Because of its multi-factorial action, it is widely presumed that the development of bacterial resistance to plasma is unlikely. However, our results indicate that a short plasma treatment (3 min) may lead to the emergence of a small number of surviving cells exhibiting enhanced resistance to subsequent plasma exposure. Interestingly, these cells also exhibited a higher degree of resistance to hydrogen peroxide. Whole genome comparison between surviving cells and control cells revealed 10 distinct polymorphic regions, including four belonging to the redox active, antibiotic pigment phenazine. Subsequently, the interaction between phenazine production and CAP resistance was demonstrated in biofilms of transposon mutants disrupted in different phenazine pathway genes which exhibited significantly altered sensitivity to CAP.

  12. Rapid depletion of dissolved oxygen in 96-well microtiter plate Staphylococcus epidermidis biofilm assays promotes biofilm development and is influenced by inoculum cell concentration.

    Science.gov (United States)

    Cotter, John J; O'Gara, James P; Casey, Eoin

    2009-08-01

    Biofilm-related research using 96-well microtiter plates involves static incubation of plates indiscriminate of environmental conditions, making oxygen availability an important variable which has not been considered to date. By directly measuring dissolved oxygen concentration over time we report here that dissolved oxygen is rapidly consumed in Staphylococcus epidermidis biofilm cultures grown in 96-well plates irrespective of the oxygen concentration in the gaseous environment in which the plates are incubated. These data indicate that depletion of dissolved oxygen during growth of bacterial biofilm cultures in 96-well plates may significantly influence biofilm production. Furthermore higher inoculum cell concentrations are associated with more rapid consumption of dissolved oxygen and higher levels of S. epidermidis biofilm production. Our data reveal that oxygen depletion during bacterial growth in 96-well plates may significantly influence biofilm production and should be considered in the interpretation of experimental data using this biofilm model.

  13. Whole transcriptome RNAseq analysis of Oenococcus oeni reveals distinct intra-specific expression patterns during malolactic fermentation, including genes involved in diacetyl metabolism.

    Science.gov (United States)

    Sternes, Peter R; Costello, Peter J; Chambers, Paul J; Bartowsky, Eveline J; Borneman, Anthony R

    2017-09-18

    We report the first whole transcriptome RNAseq analysis of the wine-associated lactic acid bacterium Oenococcus oeni using a combination of reference-based mapping and de novo transcript assembly in three distinct strains during malolactic fermentation in Cabernet Sauvignon wine. Two of the strains (AWRIB551 and AWRIB552) exhibited similar transcriptomes relative to the third strain (AWRIB419) which was dissimilar by comparison. Significant intra-specific variation for genes related to glycolysis/gluconeogenesis, purine metabolism, aminoacyl-tRNA biosynthesis, ABC transporters and phosphotransferase systems was observed. Importantly, thirteen genes associated with the production of diacetyl, a commercially valuable aroma and flavour compound, were also found to be differentially expressed between the strains in a manner that correlated positively with total diacetyl production. This included a key strain-specific gene that is predicted to encode a l-lactate dehydrogenase that may enable l-lactic acid to be utilised as a precursor for the production of diacetyl. In conjunction with previous comparative genomic studies of O. oeni, this study progresses the understanding of genetic variations which contribute to the phenotypes of this industrially-important bacterium. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Serum metabolomics analysis of patients with chikungunya and dengue mono/co-infections reveals distinct metabolite signatures in the three disease conditions

    Science.gov (United States)

    Shrinet, Jatin; Shastri, Jayanthi S.; Gaind, Rajni; Bhavesh, Neel Sarovar; Sunil, Sujatha

    2016-11-01

    Chikungunya and dengue are arboviral infections with overlapping clinical symptoms. A subset of chikungunya infection occurs also as co-infections with dengue, resulting in complications during diagnosis and patient management. The present study was undertaken to identify the global metabolome of patient sera infected with chikungunya as mono infections and with dengue as co-infections. Using nuclear magnetic resonance (NMR) spectroscopy, the metabolome of sera of three disease conditions, namely, chikungunya and dengue as mono-infections and when co-infected were ascertained and compared with healthy individuals. Further, the cohorts were analyzed on the basis of age, onset of fever and joint involvement. Here we show that many metabolites in the serum are significantly differentially regulated during chikungunya mono-infection as well as during chikungunya co-infection with dengue. We observed that glycine, serine, threonine, galactose and pyrimidine metabolisms are the most perturbed pathways in both mono and co-infection conditions. The affected pathways in our study correlate well with the clinical manifestation like fever, inflammation, energy deprivation and joint pain during the infections. These results may serve as a starting point for validations and identification of distinct biomolecules that could be exploited as biomarker candidates thereby helping in better patient management.

  15. Genome-wide occupancy of SREBP1 and its partners NFY and SP1 reveals novel functional roles and combinatorial regulation of distinct classes of genes.

    Directory of Open Access Journals (Sweden)

    Brian D Reed

    2008-07-01

    Full Text Available The sterol regulatory element-binding protein (SREBP family member SREBP1 is a critical transcriptional regulator of cholesterol and fatty acid metabolism and has been implicated in insulin resistance, diabetes, and other diet-related diseases. We globally identified the promoters occupied by SREBP1 and its binding partners NFY and SP1 in a human hepatocyte cell line using chromatin immunoprecipitation combined with genome tiling arrays (ChIP-chip. We find that SREBP1 occupies the promoters of 1,141 target genes involved in diverse biological pathways, including novel targets with roles in lipid metabolism and insulin signaling. We also identify a conserved SREBP1 DNA-binding motif in SREBP1 target promoters, and we demonstrate that many SREBP1 target genes are transcriptionally activated by treatment with insulin and glucose using gene expression microarrays. Finally, we show that SREBP1 cooperates extensively with NFY and SP1 throughout the genome and that unique combinations of these factors target distinct functional pathways. Our results provide insight into the regulatory circuitry in which SREBP1 and its network partners coordinate a complex transcriptional response in the liver with cues from the diet.

  16. Analysis of gene expression data from non-small cell lung carcinoma cell lines reveals distinct sub-classes from those identified at the phenotype level.

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    Andrew R Dalby

    Full Text Available Microarray data from cell lines of Non-Small Cell Lung Carcinoma (NSCLC can be used to look for differences in gene expression between the cell lines derived from different tumour samples, and to investigate if these differences can be used to cluster the cell lines into distinct groups. Dividing the cell lines into classes can help to improve diagnosis and the development of screens for new drug candidates. The micro-array data is first subjected to quality control analysis and then subsequently normalised using three alternate methods to reduce the chances of differences being artefacts resulting from the normalisation process. The final clustering into sub-classes was carried out in a conservative manner such that sub-classes were consistent across all three normalisation methods. If there is structure in the cell line population it was expected that this would agree with histological classifications, but this was not found to be the case. To check the biological consistency of the sub-classes the set of most strongly differentially expressed genes was be identified for each pair of clusters to check if the genes that most strongly define sub-classes have biological functions consistent with NSCLC.

  17. Multilocus sequence typing reveals a lack of diversity among Escherichia coli O157:H7 isolates that are distinct by pulsed-field gel electrophoresis.

    Science.gov (United States)

    Noller, Anna C; McEllistrem, M Catherine; Stine, O Colin; Morris, J Glenn; Boxrud, David J; Dixon, Bruce; Harrison, Lee H

    2003-02-01

    Escherichia coli O157:H7 is a major cause of foodborne illness in the United States. Pulsed-field gel electrophoresis (PFGE) is the molecular epidemiologic method mostly commonly used to identify food-borne outbreaks. Although PFGE is a powerful epidemiologic tool, it has disadvantages that make a DNA sequence-based approach potentially attractive. Multilocus sequence typing (MLST) analyzes the internal fragments of housekeeping genes to establish genetic relatedness between isolates. We sequenced selected portions of seven housekeeping genes and two membrane protein genes (ompA and espA) of 77 isolates that were diverse by PFGE to determine whether there was sufficient sequence variation to be useful as an epidemiologic tool. There was no DNA sequence diversity in the sequenced portions of the seven housekeeping genes and espA. For ompA, all but five isolates had sequence identical to that of the reference strains. E. coli O157:H7 has a striking lack of genetic diversity in the genes we explored, even among isolates that are clearly distinct by PFGE. Other approaches to identify improved molecular subtyping methods for E. coli 0157:H7 are needed.

  18. Distinct abscisic acid signaling pathways for modulation of guard cell versus mesophyll cell potassium channels revealed by expression studies in Xenopus laevis oocytes

    Science.gov (United States)

    Sutton, F.; Paul, S. S.; Wang, X. Q.; Assmann, S. M.; Evans, M. L. (Principal Investigator)

    2000-01-01

    Regulation of guard cell ion transport by abscisic acid (ABA) and in particular ABA inhibition of a guard cell inward K(+) current (I(Kin)) is well documented. However, little is known concerning ABA effects on ion transport in other plant cell types. Here we applied patch clamp techniques to mesophyll cell protoplasts of fava bean (Vicia faba cv Long Pod) plants and demonstrated ABA inhibition of an outward K(+) current (I(Kout)). When mesophyll cell protoplast mRNA (mesophyll mRNA) was expressed in Xenopus laevis oocytes, I(Kout) was generated that displayed similar properties to I(Kout) observed from direct analysis of mesophyll cell protoplasts. I(Kout) expressed by mesophyll mRNA-injected oocytes was inhibited by ABA, indicating that the ABA signal transduction pathway observed in mesophyll cells was preserved in the frog oocytes. Co-injection of oocytes with guard cell protoplast mRNA and cRNA for KAT1, an inward K(+) channel expressed in guard cells, resulted in I(Kin) that was similarly inhibited by ABA. However, oocytes co-injected with mesophyll mRNA and KAT1 cRNA produced I(Kin) that was not inhibited by ABA. These results demonstrate that the mesophyll-encoded signaling mechanism could not substitute for the guard cell pathway. These findings indicate that mesophyll cells and guard cells use distinct and different receptor types and/or signal transduction pathways in ABA regulation of K(+) channels.

  19. Chelator profiling in Deschampsia cespitosa (L.) Beauv. Reveals a Ni reaction, which is distinct from the ABA and cytokinin associated response to Cd.

    Science.gov (United States)

    Hayward, Allison R; Coates, Kahlan E; Galer, Amy L; Hutchinson, Thomas C; Emery, R J Neil

    2013-03-01

    Plant hormones, including abscisic acid (ABA) and cytokinins (CKs), fluctuate as a result of excess metal exposure. Changes in hormonal concentration regulate plant growth and may also signal activation of metal chelators. The grass Deschampsia cespitosa was dosed with either Ni or Cd or pulsed with exogenous ABA. The roots were analyzed for ABA and CKs and for multiple potential metal chelators including: amino acids, nicotianamine (NA), and phytochelatins (PCs). They were quantified after 3 h and after 7 days, using LC-ESI MS/MS. The Ni treatment caused no measurable change in ABA or CK concentration; however, an increase in NA was documented. The Cd treatment resulted in a short-term ABA increase followed by a reduction in CKs and an increase in PC concentration. An exogenous ABA pulse in non-metal challenged plants induced changes in CKs and PCs that followed those of Cd treatment. Ni and Cd stress resulted in distinctly different detoxification responses. Since the reaction of CKs and putative metal chelators to Cd stress can be mimicked by an exogenous ABA pulse, it is suggested that ABA acts as a stress signal, resulting in reduced growth by way of decreased CK concentration and reduced metal toxicity through increased PC production.

  20. Phylogenetic analysis consistent with a clinical history of sexual transmission of HIV-1 from a single donor reveals transmission of highly distinct variants

    Directory of Open Access Journals (Sweden)

    McClure Myra

    2011-07-01

    Full Text Available Abstract Background To combat the pandemic of human immunodeficiency virus 1 (HIV-1, a successful vaccine will need to cope with the variability of transmissible viruses. Human hosts infected with HIV-1 potentially harbour many viral variants but very little is known about viruses that are likely to be transmitted, or even if there are viral characteristics that predict enhanced transmission in vivo. We show for the first time that genetic divergence consistent with a single transmission event in vivo can represent several years of pre-transmission evolution. Results We describe a highly unusual case consistent with a single donor transmitting highly related but distinct HIV-1 variants to two individuals on the same evening. We confirm that the clustering of viral genetic sequences, present within each recipient, is consistent with the history of a single donor across the viral env, gag and pol genes by maximum likelihood and Bayesian Markov Chain Monte Carlo based phylogenetic analyses. Based on an uncorrelated, lognormal relaxed clock of env gene evolution calibrated with other datasets, the time since the most recent common ancestor is estimated as 2.86 years prior to transmission (95% confidence interval 1.28 to 4.54 years. Conclusion Our results show that an effective design for a preventative vaccine will need to anticipate extensive HIV-1 diversity within an individual donor as well as diversity at the population level.

  1. The use of global transcriptional analysis to reveal the biological and cellular events involved in distinct development phases of Trichophyton rubrum conidial germination

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    Ding Guohui

    2007-04-01

    Full Text Available Abstract Background Conidia are considered to be the primary cause of infections by Trichophyton rubrum. Results We have developed a cDNA microarray containing 10250 ESTs to monitor the transcriptional strategy of conidial germination. A total of 1561 genes that had their expression levels specially altered in the process were obtained and hierarchically clustered with respect to their expression profiles. By functional analysis, we provided a global view of an important biological system related to conidial germination, including characterization of the pattern of gene expression at sequential developmental phases, and changes of gene expression profiles corresponding to morphological transitions. We matched the EST sequences to GO terms in the Saccharomyces Genome Database (SGD. A number of homologues of Saccharomyces cerevisiae genes related to signalling pathways and some important cellular processes were found to be involved in T. rubrum germination. These genes and signalling pathways may play roles in distinct steps, such as activating conidial germination, maintenance of isotropic growth, establishment of cell polarity and morphological transitions. Conclusion Our results may provide insights into molecular mechanisms of conidial germination at the cell level, and may enhance our understanding of regulation of gene expression related to the morphological construction of T. rubrum.

  2. Comparative pathogenomic characterization of a non-invasive serotype M71 strain Streptococcus pyogenes NS53 reveals incongruent phenotypic implications from distinct genotypic markers.

    Science.gov (United States)

    Bao, Yun-Juan; Li, Yang; Liang, Zhong; Agrahari, Garima; Lee, Shaun W; Ploplis, Victoria A; Castellino, Francis J

    2017-07-31

    The strains serotyped as M71 from group A Streptococcus are common causes of pharyngeal and skin diseases worldwide. Here we characterize the genome of a unique non-invasive M71 human isolate, NS53. The genome does not contain structural rearrangements or large-scale gene gains/losses, but encodes a full set of non-truncated known virulence factors, thus providing an ideal reference for comparative studies. However, the NS53 genome showed incongruent phenotypic implications from distinct genotypic markers. NS53 is characterized as an emm pattern D and FCT (fibronectin-collagen-T antigen) type-3 strain, typical of skin tropic strains, but is phylogenetically close to emm pattern E strains with preference for both skin and pharyngeal infections. We propose that this incongruence could result from recombination within the emm gene locus, or, alternatively, selection has been against those genetic alterations. Combined with the inability to select for CovS switching, a process is indicated whereby NS53 has been pre-adapted to specific host niches selecting against variations in CovS and many other genes. This may allow the strain to attain successful colonization and long-term survival. A balance between genetic variations and fitness may exist for this bacterium to form a stabilized genome optimized for survival in specific host environments. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  3. Early transcriptome analyses of Z-3-Hexenol-treated zea mays revealed distinct transcriptional networks and anti-herbivore defense potential of green leaf volatiles.

    Science.gov (United States)

    Engelberth, Jurgen; Contreras, Claudia Fabiola; Dalvi, Chinmay; Li, Ting; Engelberth, Marie

    2013-01-01

    Green leaf volatiles (GLV), which are rapidly emitted by plants in response to insect herbivore damage, are now established as volatile defense signals. Receiving plants utilize these molecules to prime their defenses and respond faster and stronger when actually attacked. To further characterize the biological activity of these compounds we performed a microarray analysis of global gene expression. The focus of this project was to identify early transcriptional events elicited by Z-3-hexenol (Z-3-HOL) as our model GLV in maize (Zea mays) seedlings. The microarray results confirmed previous studies on Z-3-HOL -induced gene expression but also provided novel information about the complexity of Z-3-HOL -induced transcriptional networks. Besides identifying a distinct set of genes involved in direct and indirect defenses we also found significant expression of genes involved in transcriptional regulation, Ca(2+)-and lipid-related signaling, and cell wall reinforcement. By comparing these results with those obtained by treatment of maize seedlings with insect elicitors we found a high degree of correlation between the two expression profiles at this early time point, in particular for those genes related to defense. We further analyzed defense gene expression induced by other volatile defense signals and found Z-3-HOL to be significantly more active than methyl jasmonate, methyl salicylate, and ethylene. The data presented herein provides important information on early genetic networks that are activated by Z-3-HOL and demonstrates the effectiveness of this compound in the regulation of typical plant defenses against insect herbivores in maize.

  4. Global MYCN transcription factor binding analysis in neuroblastoma reveals association with distinct E-box motifs and regions of DNA hypermethylation.

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    Derek M Murphy

    Full Text Available BACKGROUND: Neuroblastoma, a cancer derived from precursor cells of the sympathetic nervous system, is a major cause of childhood cancer related deaths. The single most important prognostic indicator of poor clinical outcome in this disease is genomic amplification of MYCN, a member of a family of oncogenic transcription factors. METHODOLOGY: We applied MYCN chromatin immunoprecipitation to microarrays (ChIP-chip using MYCN amplified/non-amplified cell lines as well as a conditional knockdown cell line to determine the distribution of MYCN binding sites within all annotated promoter regions. CONCLUSION: Assessment of E-box usage within consistently positive MYCN binding sites revealed a predominance for the CATGTG motif (p<0.0016, with significant enrichment of additional motifs CATTTG, CATCTG, CAACTG in the MYCN amplified state. For cell lines over-expressing MYCN, gene ontology analysis revealed enrichment for the binding of MYCN at promoter regions of numerous molecular functional groups including DNA helicases and mRNA transcriptional regulation. In order to evaluate MYCN binding with respect to other genomic features, we determined the methylation status of all annotated CpG islands and promoter sequences using methylated DNA immunoprecipitation (MeDIP. The integration of MYCN ChIP-chip and MeDIP data revealed a highly significant positive correlation between MYCN binding and DNA hypermethylation. This association was also detected in regions of hemizygous loss, indicating that the observed association occurs on the same homologue. In summary, these findings suggest that MYCN binding occurs more commonly at CATGTG as opposed to the classic CACGTG E-box motif, and that disease associated over expression of MYCN leads to aberrant binding to additional weaker affinity E-box motifs in neuroblastoma. The co-localization of MYCN binding and DNA hypermethylation further supports the dual role of MYCN, namely that of a classical transcription

  5. Global MYCN transcription factor binding analysis in neuroblastoma reveals association with distinct E-box motifs and regions of DNA hypermethylation.

    LENUS (Irish Health Repository)

    Murphy, Derek M

    2009-01-01

    BACKGROUND: Neuroblastoma, a cancer derived from precursor cells of the sympathetic nervous system, is a major cause of childhood cancer related deaths. The single most important prognostic indicator of poor clinical outcome in this disease is genomic amplification of MYCN, a member of a family of oncogenic transcription factors. METHODOLOGY: We applied MYCN chromatin immunoprecipitation to microarrays (ChIP-chip) using MYCN amplified\\/non-amplified cell lines as well as a conditional knockdown cell line to determine the distribution of MYCN binding sites within all annotated promoter regions. CONCLUSION: Assessment of E-box usage within consistently positive MYCN binding sites revealed a predominance for the CATGTG motif (p<0.0016), with significant enrichment of additional motifs CATTTG, CATCTG, CAACTG in the MYCN amplified state. For cell lines over-expressing MYCN, gene ontology analysis revealed enrichment for the binding of MYCN at promoter regions of numerous molecular functional groups including DNA helicases and mRNA transcriptional regulation. In order to evaluate MYCN binding with respect to other genomic features, we determined the methylation status of all annotated CpG islands and promoter sequences using methylated DNA immunoprecipitation (MeDIP). The integration of MYCN ChIP-chip and MeDIP data revealed a highly significant positive correlation between MYCN binding and DNA hypermethylation. This association was also detected in regions of hemizygous loss, indicating that the observed association occurs on the same homologue. In summary, these findings suggest that MYCN binding occurs more commonly at CATGTG as opposed to the classic CACGTG E-box motif, and that disease associated over expression of MYCN leads to aberrant binding to additional weaker affinity E-box motifs in neuroblastoma. The co-localization of MYCN binding and DNA hypermethylation further supports the dual role of MYCN, namely that of a classical transcription factor affecting the

  6. Impact of early colonizers on in vitro subgingival biofilm formation.

    Directory of Open Access Journals (Sweden)

    Thomas W Ammann

    Full Text Available The aim of this study was to investigate the impact of early colonizing species on the structure and the composition of the bacterial community developing in a subgingival 10-species biofilm model system. The model included Streptococcus oralis, Streptococcus anginosus, Actinomycesoris, Fusobacterium nucleatum subsp. nucleatum, Veillonella dispar, Campylobacter rectus, Prevotella intermedia, Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola. Based on literature, we considered Streptococcus oralis, Streptococcus anginosus, and Actinomyces oris as early colonizers and examined their role in the biofilms by either a delayed addition to the consortium, or by not inoculating at all the biofilms with these species. We quantitatively evaluated the resulting biofilms by real-time quantitative PCR and further compared the structures using confocal laser scanning microscopy following fluorescence in situ hybridisation. The absence of the early colonizers did not hinder biofilm formation. The biofilms reached the same total counts and developed to normal thickness. However, quantitative shifts in the abundances of individual species were observed. In the absence of streptococci, the overall biofilm structure appeared looser and more dispersed. Moreover, besides a significant increase of P. intermedia and a decrease of P. gingivalis , P. intermedia appeared to form filamented long chains that resembled streptococci. A. oris, although growing to significantly higher abundance in absence of streptococci, did not have a visible impact on the biofilms. Hence, in the absence of the early colonizers, there is a pronounced effect on P. intermedia and P. gingivalis that may cause distinct shifts in the structure of the biofilm. Streptococci possibly facilitate the establishment of P. gingivalis into subgingival biofilms, while in their absence P. intermedia became more dominant and forms elongated chains.

  7. Early transcriptome analyses of Z-3-Hexenol-treated zea mays revealed distinct transcriptional networks and anti-herbivore defense potential of green leaf volatiles.

    Directory of Open Access Journals (Sweden)

    Jurgen Engelberth

    Full Text Available Green leaf volatiles (GLV, which are rapidly emitted by plants in response to insect herbivore damage, are now established as volatile defense signals. Receiving plants utilize these molecules to prime their defenses and respond faster and stronger when actually attacked. To further characterize the biological activity of these compounds we performed a microarray analysis of global gene expression. The focus of this project was to identify early transcriptional events elicited by Z-3-hexenol (Z-3-HOL as our model GLV in maize (Zea mays seedlings. The microarray results confirmed previous studies on Z-3-HOL -induced gene expression but also provided novel information about the complexity of Z-3-HOL -induced transcriptional networks. Besides identifying a distinct set of genes involved in direct and indirect defenses we also found significant expression of genes involved in transcriptional regulation, Ca(2+-and lipid-related signaling, and cell wall reinforcement. By comparing these results with those obtained by treatment of maize seedlings with insect elicitors we found a high degree of correlation between the two expression profiles at this early time point, in particular for those genes related to defense. We further analyzed defense gene expression induced by other volatile defense signals and found Z-3-HOL to be significantly more active than methyl jasmonate, methyl salicylate, and ethylene. The data presented herein provides important information on early genetic networks that are activated by Z-3-HOL and demonstrates the effectiveness of this compound in the regulation of typical plant defenses against insect herbivores in maize.

  8. Visualization of Whole-Night Sleep EEG From 2-Channel Mobile Recording Device Reveals Distinct Deep Sleep Stages with Differential Electrodermal Activity.

    Science.gov (United States)

    Onton, Julie A; Kang, Dae Y; Coleman, Todd P

    2016-01-01

    Brain activity during sleep is a powerful marker of overall health, but sleep lab testing is prohibitively expensive and only indicated for major sleep disorders. This report demonstrates that mobile 2-channel in-home electroencephalogram (EEG) recording devices provided sufficient information to detect and visualize sleep EEG. Displaying whole-night sleep EEG in a spectral display allowed for quick assessment of general sleep stability, cycle lengths, stage lengths, dominant frequencies and other indices of sleep quality. By visualizing spectral data down to 0.1 Hz, a differentiation emerged between slow-wave sleep with dominant frequency between 0.1-1 Hz or 1-3 Hz, but rarely both. Thus, we present here the new designations, Hi and Lo Deep sleep, according to the frequency range with dominant power. Simultaneously recorded electrodermal activity (EDA) was primarily associated with Lo Deep and very rarely with Hi Deep or any other stage. Therefore, Hi and Lo Deep sleep appear to be physiologically distinct states that may serve unique functions during sleep. We developed an algorithm to classify five stages (Awake, Light, Hi Deep, Lo Deep and rapid eye movement (REM)) using a Hidden Markov Model (HMM), model fitting with the expectation-maximization (EM) algorithm, and estimation of the most likely sleep state sequence by the Viterbi algorithm. The resulting automatically generated sleep hypnogram can help clinicians interpret the spectral display and help researchers computationally quantify sleep stages across participants. In conclusion, this study demonstrates the feasibility of in-home sleep EEG collection, a rapid and informative sleep report format, and novel deep sleep designations accounting for spectral and physiological differences.

  9. Visualization of Whole-Night Sleep EEG From 2-Channel Mobile Recording Device Reveals Distinct Deep Sleep Stages With Differential Electrodermal Activity

    Directory of Open Access Journals (Sweden)

    Julie A Onton

    2016-11-01

    Full Text Available Brain activity during sleep is powerful marker of overall health, but sleep lab testing is prohibitively expensive and only indicated for major sleep disorders. This report demonstrates that mobile 2-channel in-home electroencephalogram (EEG recording devices provided sufficient information to detect and visualize sleep EEG. Displaying whole-night sleep EEG in a spectral display allowed for quick assessment of general sleep stability, cycle lengths, stage lengths, dominant frequencies, and other indices of sleep quality. By visualizing spectral data down to 0.1 Hz, a differentiation emerged between slow-wave sleep with dominant frequency between 0.1─1 Hz or 1─3 Hz, but rarely both. Thus, we present here the new designations, Hi and Lo Deep sleep, according to the frequency range with dominant power. Simultaneously recorded electrodermal activity (EDA was primarily associated with Lo Deep and very rarely with Hi Deep or any other stage. Therefore, Hi and Lo Deep sleep appear to be physiologically distinct states that may serve unique functions during sleep. We developed an algorithm to classify 5 stages (Awake, Light, Hi Deep, Lo Deep, and REM using a Hidden Markov Model, model fitting with the expectation-maximization algorithm, and estimation of the most likely sleep state sequence by the Viterbi algorithm. The resulting automatically generated sleep hypnogram can help clinicians interpret the spectral display and help researchers computationally quantify sleep stages across participants. In conclusion, this study demonstrates the feasibility of in-home sleep EEG collection, a rapid and informative sleep report format, and novel deep sleep designations accounting for spectral and physiological differences.

  10. Comparative transcriptional network modeling of three PPAR-α/γ co-agonists reveals distinct metabolic gene signatures in primary human hepatocytes.

    Directory of Open Access Journals (Sweden)

    Renée Deehan

    Full Text Available AIMS: To compare the molecular and biologic signatures of a balanced dual peroxisome proliferator-activated receptor (PPAR-α/γ agonist, aleglitazar, with tesaglitazar (a dual PPAR-α/γ agonist or a combination of pioglitazone (Pio; PPAR-γ agonist and fenofibrate (Feno; PPAR-α agonist in human hepatocytes. METHODS AND RESULTS: Gene expression microarray profiles were obtained from primary human hepatocytes treated with EC(50-aligned low, medium and high concentrations of the three treatments. A systems biology approach, Causal Network Modeling, was used to model the data to infer upstream molecular mechanisms that may explain the observed changes in gene expression. Aleglitazar, tesaglitazar and Pio/Feno each induced unique transcriptional signatures, despite comparable core PPAR signaling. Although all treatments inferred qualitatively similar PPAR-α signaling, aleglitazar was inferred to have greater effects on high- and low-density lipoprotein cholesterol levels than tesaglitazar and Pio/Feno, due to a greater number of gene expression changes in pathways related to high-density and low-density lipoprotein metabolism. Distinct transcriptional and biologic signatures were also inferred for stress responses, which appeared to be less affected by aleglitazar than the comparators. In particular, Pio/Feno was inferred to increase NFE2L2 activity, a key component of the stress response pathway, while aleglitazar had no significant effect. All treatments were inferred to decrease proliferative signaling. CONCLUSIONS: Aleglitazar induces transcriptional signatures related to lipid parameters and stress responses that are unique from other dual PPAR-α/γ treatments. This may underlie observed favorable changes in lipid profiles in animal and clinical studies with aleglitazar and suggests a differentiated gene profile compared with other dual PPAR-α/γ agonist treatments.

  11. Distinct effects of Hoxa2 overexpression in cranial neural crest populations reveal that the mammalian hyomandibular-ceratohyal boundary maps within the styloid process.

    Science.gov (United States)

    Kitazawa, Taro; Fujisawa, Kou; Narboux-Nême, Nicolas; Arima, Yuichiro; Kawamura, Yumiko; Inoue, Tsuyoshi; Wada, Youichiro; Kohro, Takahide; Aburatani, Hiroyuki; Kodama, Tatsuhiko; Kim, Ki-Sung; Sato, Takahiro; Uchijima, Yasunobu; Maeda, Kazuhiro; Miyagawa-Tomita, Sachiko; Minoux, Maryline; Rijli, Filippo M; Levi, Giovanni; Kurihara, Yukiko; Kurihara, Hiroki

    2015-06-15

    Most gnathostomata craniofacial structures derive from pharyngeal arches (PAs), which are colonized by cranial neural crest cells (CNCCs). The anteroposterior and dorsoventral identities of CNCCs are defined by the combinatorial expression of Hox and Dlx genes. The mechanisms associating characteristic Hox/Dlx expression patterns with the topology and morphology of PAs derivatives are only partially known; a better knowledge of these processes might lead to new concepts on the origin of taxon-specific craniofacial morphologies and of certain craniofacial malformations. Here we show that ectopic expression of Hoxa2 in Hox-negative CNCCs results in distinct phenotypes in different CNCC subpopulations. Namely, while ectopic Hoxa2 expression is sufficient for the morphological and molecular transformation of the first PA (PA1) CNCC derivatives into the second PA (PA2)-like structures, this same genetic alteration does not provoke the transformation of derivatives of other CNCC subpopulations, but severely impairs their development. Ectopic Hoxa2 expression results in the transformation of the proximal Meckel's cartilage and of the malleus, two ventral PA1 CNCCs derivatives, into a supernumerary styloid process (SP), a PA2-derived mammalian-specific skeletal structure. These results, together with experiments to inactivate and ectopically activate the Edn1-Dlx5/6 pathway, indicate a dorsoventral PA2 (hyomandibular/ceratohyal) boundary passing through the middle of the SP. The present findings suggest context-dependent function of Hoxa2 in CNCC regional specification and morphogenesis, and provide novel insights into the evolution of taxa-specific patterning of PA-derived structures. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Integrated analysis of seed proteome and mRNA oxidation reveals distinct post-transcriptional features regulating dormancy in wheat (Triticum aestivum L.).

    Science.gov (United States)

    Gao, Feng; Rampitsch, Christof; Chitnis, Vijaya R; Humphreys, Gavin D; Jordan, Mark C; Ayele, Belay T

    2013-10-01

    Wheat seeds can be released from a dormant state by after-ripening; however, the underlying molecular mechanisms are still mostly unknown. We previously identified transcriptional programmes involved in the regulation of after-ripening-mediated seed dormancy decay in wheat (Triticum aestivum L.). Here, we show that seed dormancy maintenance and its release by dry after-ripening in wheat is associated with oxidative modification of distinct seed-stored mRNAs that mainly correspond to oxidative phosphorylation, ribosome biogenesis, nutrient reservoir and α-amylase inhibitor activities, suggesting the significance of post-transcriptional repression of these biological processes in regulating seed dormancy. We further show that after-ripening induced seed dormancy release in wheat is mediated by differential expression of specific proteins in both dry and hydrated states, including those involved in proteolysis, cellular signalling, translation and energy metabolism. Among the genes corresponding to these proteins, the expression of those encoding α-amylase/trypsin inhibitor and starch synthase appears to be regulated by mRNA oxidation. Co-expression analysis of the probesets differentially expressed and oxidized during dry after-ripening along with those corresponding to proteins differentially regulated between dormant and after-ripened seeds produced three co-expressed gene clusters containing more candidate genes potentially involved in the regulation of seed dormancy in wheat. Two of the three clusters are enriched with elements that are either abscisic acid (ABA) responsive or recognized by ABA-regulated transcription factors, indicating the association between wheat seed dormancy and ABA sensitivity.

  13. Phylogeny and expression analysis of C-reactive protein (CRP) and serum amyloid-P (SAP) like genes reveal two distinct groups in fish.

    Science.gov (United States)

    Lee, P T; Bird, S; Zou, J; Martin, S A M

    2017-06-01

    The acute phase response (APR) is an early innate immune function that is initiated by inflammatory signals, leading to the release of acute phase proteins to the bloodstream to re-establish homeostasis following microbial infection. In this study we analysed the Atlantic salmon (Salmo salar) whole-genome database and identified five C-reactive protein (CRP)/serum amyloid P component (SAP) like molecules namely CRP/SAP-1a, CRP/SAP-1b, CRP/SAP-1c, CRP/SAP-2 and CRP/SAP-3. These CRP/SAP genes formed two distinct sub-families, a universal group (group I) present in all vertebrates and a fish/amphibian specific group (group II). Salmon CRP/SAP-1a, CRP/SAP-1b and CRP/SAP-1c and CRP/SAP-2 belong to the group I family whilst salmon CRP/SAP-3 is a member of group II. Gene expression analysis showed that the salmon CRP/SAP-1a as well as serum amyloid A-5 (SAA-5), one of the major acute phase proteins, were significantly up-regulated by recombinant cytokines (rIL-1β and rIFNγ) in primary head kidney cells whilst the other four CRP/SAPs remained refractory. Furthermore, SAA-5 was produced as the main acute phase protein (APP) in Atlantic salmon challenged with Aeromonas salmonicida (aroA(-) strain) whilst salmon CRP/SAPs remained unaltered. Overall, these data illustrate the potential different functions of expanded salmon CRP/SAPs to their mammalian homologues. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  14. Analysis of the type II-A CRISPR-Cas system of Streptococcus agalactiae reveals distinctive features according to genetic lineages

    Directory of Open Access Journals (Sweden)

    Clément eLier

    2015-06-01

    Full Text Available CRISPR-Cas systems (clustered regularly interspaced short palindromic repeats/CRISPR-associated proteins are found in 90% of archaea and about 40% of bacteria. In this original system, CRISPR arrays comprise short, almost unique sequences called spacers that are interspersed with conserved palindromic repeats. These systems play a role in adaptive immunity and participate to fight non-self DNA such as integrative and conjugative elements, plasmids, and phages. In Streptococcus agalactiae, a bacterium implicated in colonisation and infections in humans since the 1960s, two CRISPR-Cas systems have been described. A type II-A system, characterised by proteins Cas9, Cas1, Cas2 and Csn2, is ubiquitous, and a type I-C system, with the Cas8c signature protein, is present in about 20% of the isolates. Unlike type I-C, which appears to be non-functional, type II-A appears fully functional. Here we studied type II-A CRISPR-Cas loci from 126 human isolates of S. agalactiae belonging to different clonal complexes that represent the diversity of the species and that have been implicated in colonisation or infection. The CRISPR-Cas locus was analysed both at spacer and repeat levels. Major distinctive features were identified according to the phylogenetic lineages previously defined by multilocus sequence typing, especially for the Sequence Type (ST 17, which is considered hypervirulent. Among other idiosyncrasies, ST-17 shows a significantly lower number of spacers in comparison with other lineages. This characteristic could reflect the peculiar virulence or colonisation specificities of this lineage.

  15. Distinct interaction modes of an AKAP bound to two regulatory subunit isoforms of protein kinase A revealed by amide hydrogen/deuterium exchange

    Science.gov (United States)

    Burns-Hamuro, Lora L.; Hamuro, Yoshitomo; Kim, Jack S.; Sigala, Paul; Fayos, Rosa; Stranz, David D.; Jennings, Patricia A.; Taylor, Susan S.; Woods, Virgil L.

    2005-01-01

    The structure of an AKAP docked to the dimerization/docking (D/D) domain of the type II (RIIα) isoform of protein kinase A (PKA) has been well characterized, but there currently is no detailed structural information of an AKAP docked to the type I (RIα) isoform. Dual-specific AKAP2 (D-AKAP2) binds in the nanomolar range to both isoforms and provided us with an opportunity to characterize the isoform-selective nature of AKAP binding using a common docked ligand. Hydrogen/deuterium (H/D) exchange combined with mass spectrometry (DXMS) was used to probe backbone structural changes of an α-helical A-kinase binding (AKB) motif from D-AKAP2 docked to both RIα and RIIα D/D domains. The region of protection upon complex formation and the magnitude of protection from H/D exchange were determined for both interacting partners in each complex. The backbone of the AKB ligand was more protected when bound to RIα compared to RIIα, suggesting an increased helical stabilization of the docked AKB ligand. This combined with a broader region of backbone protection induced by the AKAP on the docking surface of RIα indicated that there were more binding constraints for the AKB ligand when bound to RIα. This was in contrast to RIIα, which has a preformed, localized binding surface. These distinct modes of AKAP binding may contribute to the more discriminating nature of the RIα AKAP-docking surface. DXMS provides valuable structural information for understanding binding specificity in the absence of a high-resolution structure, and can readily be applied to other protein–ligand and protein–protein interactions. PMID:16260760

  16. Targeted systems biology profiling of tomato fruit reveals coordination of the Yang cycle and a distinct regulation of ethylene biosynthesis during postclimacteric ripening.

    Science.gov (United States)

    Van de Poel, Bram; Bulens, Inge; Markoula, Aikaterina; Hertog, Maarten L A T M; Dreesen, Rozemarijn; Wirtz, Markus; Vandoninck, Sandy; Oppermann, Yasmin; Keulemans, Johan; Hell, Ruediger; Waelkens, Etienne; De Proft, Maurice P; Sauter, Margret; Nicolai, Bart M; Geeraerd, Annemie H

    2012-11-01

    The concept of system 1 and system 2 ethylene biosynthesis during climacteric fruit ripening was initially described four decades ago. Although much is known about fruit development and climacteric ripening, little information is available about how ethylene biosynthesis is regulated during the postclimacteric phase. A targeted systems biology approach revealed a novel regulatory mechanism of ethylene biosynthesis of tomato (Solanum lycopersicum) when fruit have reached their maximal ethylene production level and which is characterized by a decline in ethylene biosynthesis. Ethylene production is shut down at the level of 1-aminocyclopropane-1-carboxylic acid oxidase. At the same time, 1-aminocyclopropane-1-carboxylic acid synthase activity increases. Analysis of the Yang cycle showed that the Yang cycle genes are regulated in a coordinated way and are highly expressed during postclimacteric ripening. Postclimacteric red tomatoes on the plant showed only a moderate regulation of 1-aminocyclopropane-1-carboxylic acid synthase and Yang cycle genes compared with the regulation in detached fruit. Treatment of red fruit with 1-methylcyclopropane and ethephon revealed that the shut-down mechanism in ethylene biosynthesis is developmentally programmed and only moderately ethylene sensitive. We propose that the termination of autocatalytic ethylene biosynthesis of system 2 in ripe fruit delays senescence and preserves the fruit until seed dispersal.

  17. Biofilm Cohesive Strength as a Basis for Biofilm Recalcitrance: Are Bacterial Biofilms Overdesigned?

    Science.gov (United States)

    Aggarwal, Srijan; Stewart, Philip S; Hozalski, Raymond M

    2015-01-01

    Bacterial biofilms are highly resistant to common antibacterial treatments, and several physiological explanations have been offered to explain the recalcitrant nature of bacterial biofilms. Herein, a biophysical aspect of biofilm recalcitrance is being reported on. While engineering structures are often overdesigned with a factor of safety (FOS) usually under 10, experimental measurements of biofilm cohesive strength suggest that the FOS is on the order of thousands. In other words, bacterial biofilms appear to be designed to withstand extreme forces rather than typical or average loads. In scenarios requiring the removal or control of unwanted biofilms, this emphasizes the importance of considering strategies for structurally weakening the biofilms in conjunction with bacterial inactivation.

  18. Role of Extracellular DNA during Biofilm Formation by Listeria monocytogenes

    DEFF Research Database (Denmark)

    Harmsen, Morten; Lappann, Martin; Knøchel, S

    2010-01-01

    Listeria monocytogenes is a food-borne pathogen that is capable of living in harsh environments. It is believed to do this by forming biofilms, which are surface-associated multicellular structures encased in a self-produced matrix. In this paper we show that in L. monocytogenes extracellular DNA...... (eDNA) may be the only central component of the biofilm matrix and that it is necessary for both initial attachment and early biofilm formation for 41 L. monocytogenes strains that were tested. DNase I treatment resulted in dispersal of biofilms, not only in microtiter tray assays but also in flow...... cell biofilm assays. However, it was also demonstrated that in a culture without eDNA, neither Listeria genomic DNA nor salmon sperm DNA by itself could restore the capacity to adhere. A search for additional necessary components revealed that peptidoglycan (PG), specifically N-acetylglucosamine (NAG...

  19. Shell biofilm-associated nitrous oxide production in marine molluscs

    DEFF Research Database (Denmark)

    Heisterkamp, I.M.; Schramm, Andreas; Larsen, Lone Heimann

    2013-01-01

    Emission of the greenhouse gas nitrous oxide (N2O) from freshwater and terrestrial invertebrates has exclusively been ascribed to N2O production by ingested denitrifying bacteria in the anoxic gut of the animals. Our study of marine molluscs now shows that also microbial biofilms on shell surfaces...... are important sites of N2O production. The shell biofilms of Mytilus edulis, Littorina littorea and Hinia reticulata contributed 18-94% to the total animal-associated N2O emission. Nitrification and denitrification were equally important sources of N2O in shell biofilms as revealed by 15N-stable isotope...... experiments with dissected shells. Microsensor measurements confirmed that both nitrification and denitrification can occur in shell biofilms due to a heterogeneous oxygen distribution. Accordingly, ammonium, nitrite and nitrate were important drivers of N2O production in the shell biofilm of the three...

  20. Sulfate reducing bacteria and their activities in oil sands process-affected water biofilm.

    Science.gov (United States)

    Liu, Hong; Yu, Tong; Liu, Yang

    2015-12-01

    Biofilm reactors were constructed to grow stratified multispecies biofilm in oil sands process-affected water (OSPW) supplemented with growth medium. The development of sulfate reducing bacteria (SRB) within the biofilm and the biofilm treatment of OSPW were evaluated. The community structure and potential activity of SRB in the biofilm were investigated with H2S microsensor measurements, dsrB gene-based denaturing gradient gel electrophoresis (DGGE), and the real time quantitative polymerase chain reaction (qPCR). Multispecies biofilm with a thickness of 1000 μm was successfully developed on engineered biocarriers. H2S production was observed in the deeper anoxic zone of the biofilm from around 750 μm to 1000 μm below the bulk water-biofilm interface, revealing sulfate reduction in the deeper zone of the stratified biofilm. The biofilm removed chemical oxygen demand (COD), sulfate, and nitrogen. The study expands current knowledge of biofilm treatment of OSPW and the function of anaerobic SRB in OSPW biofilm, and thus provides information for future bioreactor development in the reclamation of OSPW.

  1. Host intestinal signal-promoted biofilm dispersal induces Vibrio cholerae colonization.

    Science.gov (United States)

    Hay, Amanda J; Zhu, Jun

    2015-01-01

    Vibrio cholerae causes human infection through ingestion of contaminated food and water, leading to the devastating diarrheal disease cholera. V. cholerae forms matrix-encased aggregates, known as biofilms, in the native aquatic environment. While the formation of V. cholerae biofilms has been well studied, little is known about the dispersal from biofilms, particularly upon entry into the host. In this study, we found that the exposure of mature biofilms to physiologic levels of the bile salt taurocholate, a host signal for the virulence gene induction of V. cholerae, induces an increase in the number of detached cells with a concomitant decrease in biofilm mass. Scanning electron microscopy micrographs of biofilms exposed to taurocholate revealed an altered, perhaps degraded, appearance of the biofilm matrix. The inhibition of protein synthesis did not alter rates of detachment, suggesting that V. cholerae undergoes a passive dispersal. Cell-free media from taurocholate-exposed biofilms contains a larger amount of free polysaccharide, suggesting an abiotic degradation of biofilm matrix by taurocholate. Furthermore, we found that V. cholerae is only able to induce virulence in response to taurocholate after exit from the biofilm. Thus, we propose a model in which V. cholerae ingested as a biofilm has coopted the host-derived bile salt signal to detach from the biofilm and go on to activate virulence.

  2. Community analysis of pigment patterns from 37 microalgae strains reveals new carotenoids and porphyrins characteristic of distinct strains and taxonomic groups

    Science.gov (United States)

    Bérard, Jean-Baptiste; Kaas, Raymond; Pasquet, Virginie; Picot, Laurent; Cadoret, Jean-Paul

    2017-01-01

    Phytoplankton, with an estimated 30 000 to 1 000 000 species clustered in 12 phyla, presents a high taxonomic and ecophysiological diversity, reflected by the complex distribution of pigments among the different algal classes. High performance liquid chromatography is the gold standard method for qualitative and quantitative analysis of phytoplankton pigments in seawater and culture samples, but only a few pigments can be used as robust chemotaxonomic markers. A major challenge is thus to identify new ones, characteristic of a strain, species, class or taxon that cannot be currently identified on the basis of its pigment signature. Using an optimized extraction process coupled to a HPLC de-replication strategy, we examined the pigment composition of 37 microalgae strains, representative of the broad taxonomic diversity of marine and freshwater species (excluding cyanobacteria). For each species, the major pigments already described were unambiguously identified. We also observed the presence of several minor unidentified pigments in each chromatogram. The global analysis of pigment compositions revealed a total of 124 pigments, including 98 pigments or derivatives unidentified using the standards. Absorption spectra indicated that 35 corresponded to chlorophyll/porphyrin derivatives, 57 to carotenoids and six to derivatives having both spectral signatures. Sixty-one of these unidentified or new carotenoids and porphyrin derivatives were characteristic of particular strains or species, indicating their possible use as highly specific chemotaxonomic markers capable of identifying one strain out of the 37 selected. We developed a graphical analysis using Gephi software to give a clear representation of pigment communities among the various phytoplankton strains, and to reveal strain-characteristic and shared pigments. This made it possible to reconstruct the taxonomic evolution of microalgae classes, on the basis of the conservation, loss, and/or appearance of

  3. In silico genomic analyses reveal three distinct lineages of Escherichia coli O157:H7, one of which is associated with hyper-virulence

    Directory of Open Access Journals (Sweden)

    Karmali Mohamed A

    2009-06-01

    Full Text Available Abstract Background Many approaches have been used to study the evolution, population structure and genetic diversity of Escherichia coli O157:H7; however, observations made with different genotyping systems are not easily relatable to each other. Three genetic lineages of E. coli O157:H7 designated I, II and I/II have been identified using octamer-based genome scanning and microarray comparative genomic hybridization (mCGH. Each lineage contains significant phenotypic differences, with lineage I strains being the most commonly associated with human infections. Similarly, a clade of hyper-virulent O157:H7 strains implicated in the 2006 spinach and lettuce outbreaks has been defined using single-nucleotide polymorphism (SNP typing. In this study an in silico comparison of six different genotyping approaches was performed on 19 E. coli genome sequences from 17 O157:H7 strains and single O145:NM and K12 MG1655 strains to provide an overall picture of diversity of the E. coli O157:H7 population, and to compare genotyping methods for O157:H7 strains. Results In silico determination of lineage, Shiga-toxin bacteriophage integration site, comparative genomic fingerprint, mCGH profile, novel region distribution profile, SNP type and multi-locus variable number tandem repeat analysis type was performed and a supernetwork based on the combination of these methods was produced. This supernetwork showed three distinct clusters of strains that were O157:H7 lineage-specific, with the SNP-based hyper-virulent clade 8 synonymous with O157:H7 lineage I/II. Lineage I/II/clade 8 strains clustered closest on the supernetwork to E. coli K12 and E. coli O55:H7, O145:NM and sorbitol-fermenting O157 strains. Conclusion The results of this study highlight the similarities in relationships derived from multi-locus genome sampling methods and suggest a "common genotyping language" may be devised for population genetics and epidemiological studies. Future genotyping

  4. Antibody VH and VL recombination using phage and ribosome display technologies reveals distinct structural routes to affinity improvements with VH-VL interface residues providing important structural diversity.

    Science.gov (United States)

    Groves, Maria A T; Amanuel, Lily; Campbell, Jamie I; Rees, D Gareth; Sridharan, Sudharsan; Finch, Donna K; Lowe, David C; Vaughan, Tristan J

    2014-01-01

    In vitro selection technologies are an important means of affinity maturing antibodies to generate the optimal therapeutic profile for a particular disease target. Here, we describe the isolation of a parent antibody, KENB061 using phage display and solution phase selections with soluble biotinylated human IL-1R1. KENB061 was affinity matured using phage display and targeted mutagenesis of VH and VL CDR3 using NNS randomization. Affinity matured VHCDR3 and VLCDR3 library blocks were recombined and selected using phage and ribosome display protocol. A direct comparison of the phage and ribosome display antibodies generated was made to determine their functional characteristics.In our analyses, we observed distinct differences in the pattern of beneficial mutations in antibodies derived from phage and ribosome display selections, and discovered the lead antibody Jedi067 had a ~3700-fold improvement in KD over the parent KENB061. We constructed a homology model of the Fv region of Jedi067 to map the specific positions where mutations occurred in the CDR3 loops. For VL CDR3, positions 94 to 97 carry greater diversity in the ribosome display variants compared with the phage display. The positions 95a, 95b and 96 of VLCDR3 form part of the interface with VH in this model. The model shows that positions 96, 98, 100e, 100f, 100 g, 100h, 100i and 101 of the VHCDR3 include residues at the VH and VL interface. Importantly, Leu96 and Tyr98 are conserved at the interface positions in both phage and ribosome display indicating their importance in maintaining the VH-VL interface. For antibodies derived from ribosome display, there is significant diversity at residues 100a to 100f of the VH CDR3 compared with phage display. A unique deletion of isoleucine at position 102 of the lead candidate, Jedi067, also occurs in the VHCDR3.As anticipated, recombining the mutations via ribosome display led to a greater structural diversity, particularly in the heavy chain CDR3, which in turn

  5. SinR controls enterotoxin expression in Bacillus thuringiensis biofilms.

    Directory of Open Access Journals (Sweden)

    Annette Fagerlund

    Full Text Available The entomopathogen Bacillus thuringiensis produces dense biofilms under various conditions. Here, we report that the transition phase regulators Spo0A, AbrB and SinR control biofilm formation and swimming motility in B. thuringiensis, just as they control biofilm formation and swarming motility in the closely related saprophyte species B. subtilis. However, microarray analysis indicated that in B. thuringiensis, in contrast to B. subtilis, SinR does not control an eps operon involved in exopolysaccharides production, but regulates genes involved in the biosynthesis of the lipopeptide kurstakin. This lipopeptide is required for biofilm formation and was previously shown to be important for survival in the host cadaver (necrotrophism. Microarray analysis also revealed that the SinR regulon contains genes coding for the Hbl enterotoxin. Transcriptional fusion assays, Western blots and hemolysis assays confirmed that SinR controls Hbl expression, together with PlcR, the main virulence regulator in B. thuringiensis. We show that Hbl is expressed in a sustained way in a small subpopulation of the biofilm, whereas almost all the planktonic population transiently expresses Hbl. The gene coding for SinI, an antagonist of SinR, is expressed in the same biofilm subpopulation as hbl, suggesting that hbl transcription heterogeneity is SinI-dependent. B. thuringiensis and B. cereus are enteric bacteria which possibly form biofilms lining the host intestinal epithelium. Toxins produced in biofilms could therefore be delivered directly to the target tissue.

  6. Biofilm formation enhances Helicobacter pylori survivability in vegetables.

    Science.gov (United States)

    Ng, Chow Goon; Loke, Mun Fai; Goh, Khean Lee; Vadivelu, Jamuna; Ho, Bow

    2017-04-01

    To date, the exact route and mode of transmission of Helicobacter pylori remains elusive. The detection of H. pylori in food using molecular approaches has led us to postulate that the gastric pathogen may survive in the extragastric environment for an extended period. In this study, we show that H. pylori prolongs its survival by forming biofilm and micro-colonies on vegetables. The biofilm forming capability of H. pylori is both strain and vegetable dependent. H. pylori strains were classified into high and low biofilm formers based on their highest relative biofilm units (BU). High biofilm formers survived longer on vegetables compared to low biofilm formers. The bacteria survived better on cabbage compared to other vegetables tested. In addition, images captured on scanning electron and confocal laser scanning microscopes revealed that the bacteria were able to form biofilm and reside as micro-colonies on vegetable surfaces, strengthening the notion of possible survival of H. pylori on vegetables for an extended period of time. Taken together, the ability of H. pylori to form biofilm on vegetables (a common food source for human) potentially plays an important role in its survival, serving as a mode of transmission of H. pylori in the extragastric environment. Copyright © 2016 Elsevier Ltd. All rights reserved.

  7. Effects of lactoferricin B against keratitis-associated fungal biofilms.

    Science.gov (United States)

    Sengupta, Jayangshu; Saha, Suman; Khetan, Archana; Sarkar, Sujoy K; Mandal, Santi M

    2012-10-01

    Biofilms are considered as the most important developmental characteristics in ocular infections. Biofilm eradication is a major challenge today to overcome the incidence of drug resistance. This report demonstrates the in vitro ability of biofilm formation on contact lens by three common keratitis-associated fungal pathogens, namely, Aspergillus fumigatus, Fusarium solani, and Candida albicans. Antifungal sensitivity testing performed for both planktonic cells and biofilm revealed the sessile phenotype to be resistant at MIC levels for the planktonic cells and also at higher concentrations. A prototype lens care solution was also found to be partially effective in eradication of the mature biofilm from contact lenses. Lactoferricin B (Lacf, 64 μg/ml), an antimicrobial peptide, exhibited almost no effect on the sessile phenotype. However, the combinatory effect of Lacf with antifungals against planktonic cells and biofilms of three fungal strains that were isolated from keratitis patients exhibited a reduction of antifungal dose more than eightfold. Furthermore, the effect of Lacf in lens care solution against biofilms in which those strains formed was eradicated successfully. These results suggest that lactoferricin B could be a promising candidate for clinical use in improving biofilm susceptibility to antifungals and also as an antibiofilm-antifungal additive in lens care solution.

  8. Analysis of Drosophila p8 and p52 mutants reveals distinct roles for the maintenance of TFIIH stability and male germ cell differentiation

    Science.gov (United States)

    Cruz-Becerra, Grisel; Juárez, Mandy; Valadez-Graham, Viviana

    2016-01-01

    Eukaryotic gene expression is activated by factors that interact within complex machinery to initiate transcription. An important component of this machinery is the DNA repair/transcription factor TFIIH. Mutations in TFIIH result in three human syndromes: xeroderma pigmentosum, Cockayne syndrome and trichothiodystrophy. Transcription and DNA repair defects have been linked to some clinical features of these syndromes. However, how mutations in TFIIH affect specific developmental programmes, allowing organisms to develop with particular phenotypes, is not well understood. Here, we show that mutations in the p52 and p8 subunits of TFIIH have a moderate effect on the gene expression programme in the Drosophila testis, causing germ cell differentiation arrest in meiosis, but no Polycomb enrichment at the promoter of the affected differentiation genes, supporting recent data that disagree with the current Polycomb-mediated repression model for regulating gene expression in the testis. Moreover, we found that TFIIH stability is not compromised in p8 subunit-depleted testes that show transcriptional defects, highlighting the role of p8 in transcription. Therefore, this study reveals how defects in TFIIH affect a specific cell differentiation programme and contributes to understanding the specific syndrome manifestations in TFIIH-afflicted patients.

  9. Molecular characterization of Cucumber mosaic virus infecting Gladiolus, revealing its phylogeny distinct from the Indian isolate and alike the Fny strain of CMV.

    Science.gov (United States)

    Dubey, Vimal Kumar; Aminuddin; Singh, Vijai Pal

    2010-08-01

    The majority of Gladiolus plants growing in the botanical garden at NBRI, Lucknow, India and adjoining areas exhibited symptoms of mosaic, color breaking, stunting of spikes and reduction in flower size. The occurrence of Cucumber mosaic virus (CMV) was suspected in symptomatic Gladiolus plants. Cucumber mosaic virus, the type species of the genus Cucumovirus of the family Bromoviridae, is an important plant virus worldwide, which infects many plants and causes quantity and quality losses. For virus characterization, total RNA was isolated from leaves of infected plants and used in reverse transcriptase polymerase chain reaction with a primer set designed in the Cucumber mosaic virus coat protein region. Viral amplicons of the expected 657 bp size were obtained from infected plants. No viral amplicon was obtained from healthy control plants. Viral amplicons were cloned and sequenced (DQ295914). Molecular characterization was performed and phylogenetic relationship determined by the comparison of coat protein gene nucleotide and amino acid sequences with other Cucumber mosaic virus isolates reported from India and worldwide. The nucleotide and amino acid percentage comparison and phylogenetic tree results revealed that Cucumber mosaic virus infecting Gladiolus show resemblance with the Fny strain, which is not common in the Asian continent.

  10. Bioinformatics Analysis Reveals Distinct Molecular Characteristics of Hepatitis B-Related Hepatocellular Carcinomas from Very Early to Advanced Barcelona Clinic Liver Cancer Stages.

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    Fan-Yun Kong

    Full Text Available Hepatocellular carcinoma (HCCis the fifth most common malignancy associated with high mortality. One of the risk factors for HCC is chronic hepatitis B virus (HBV infection. The treatment strategy for the disease is dependent on the stage of HCC, and the Barcelona clinic liver cancer (BCLC staging system is used in most HCC cases. However, the molecular characteristics of HBV-related HCC in different BCLC stages are still unknown. Using GSE14520 microarray data from HBV-related HCC cases with BCLC stages from 0 (very early stage to C (advanced stage in the gene expression omnibus (GEO database, differentially expressed genes (DEGs, including common DEGs and unique DEGs in different BCLC stages, were identified. These DEGs were located on different chromosomes. The molecular functions and biology pathways of DEGs were identified by gene ontology (GO analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG pathway analysis, and the interactome networks of DEGs were constructed using the NetVenn online tool. The results revealed that both common DEGs and stage-specific DEGs were associated with various molecular functions and were involved in special biological pathways. In addition, several hub genes were found in the interactome networks of DEGs. The identified DEGs and hub genes promote our understanding of the molecular mechanisms underlying the development of HBV-related HCC through the different BCLC stages, and might be used as staging biomarkers or molecular targets for the treatment of HCC with HBV infection.

  11. Bioinformatics Analysis Reveals Distinct Molecular Characteristics of Hepatitis B-Related Hepatocellular Carcinomas from Very Early to Advanced Barcelona Clinic Liver Cancer Stages

    Science.gov (United States)

    Hu, Wei; Kou, Yan-Bo; You, Hong-Juan; Liu, Xiao-Mei; Zheng, Kui-Yang; Tang, Ren-Xian

    2016-01-01

    Hepatocellular carcinoma (HCC)is the fifth most common malignancy associated with high mortality. One of the risk factors for HCC is chronic hepatitis B virus (HBV) infection. The treatment strategy for the disease is dependent on the stage of HCC, and the Barcelona clinic liver cancer (BCLC) staging system is used in most HCC cases. However, the molecular characteristics of HBV-related HCC in different BCLC stages are still unknown. Using GSE14520 microarray data from HBV-related HCC cases with BCLC stages from 0 (very early stage) to C (advanced stage) in the gene expression omnibus (GEO) database, differentially expressed genes (DEGs), including common DEGs and unique DEGs in different BCLC stages, were identified. These DEGs were located on different chromosomes. The molecular functions and biology pathways of DEGs were identified by gene ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, and the interactome networks of DEGs were constructed using the NetVenn online tool. The results revealed that both common DEGs and stage-specific DEGs were associated with various molecular functions and were involved in special biological pathways. In addition, several hub genes were found in the interactome networks of DEGs. The identified DEGs and hub genes promote our understanding of the molecular mechanisms underlying the development of HBV-related HCC through the different BCLC stages, and might be used as staging biomarkers or molecular targets for the treatment of HCC with HBV infection. PMID:27454179

  12. Human exceptional longevity: transcriptome from centenarians is distinct from septuagenarians and reveals a role of Bcl-xL in successful aging

    Science.gov (United States)

    Borras, Consuelo; M. Abdelaziz, Kheira; Gambini, Juan; Serna, Eva; Inglés, Marta; de la Fuente, Monica; Garcia, Idoia; Matheu, Ander; Sanchís, Paula; Belenguer, Angel; Errigo, Alessandra; Avellana, Juan- Antonio; Barettino, Ana; Lloret-Fernández, Carla; Flames, Nuria; Pes, Gianni; Rodriguez-Mañas, Leocadio; Viña, Jose

    2016-01-01

    Centenarians not only enjoy an extraordinary aging, but also show a compression of morbidity. Using functional transcriptomic analysis of peripheral blood mononuclear cells (PMBC) we identified 1721 mRNAs differentially expressed by centenarians when compared with septuagenarians and young people. Sub-network analysis led us to identify Bcl - xL as an important gene up-regulated in centenarians. It is involved in the control of apoptosis, cellular damage protection and also in modulation of immune response, all associated to healthy aging. Indeed, centenarians display lower plasma cytochrome C levels, higher mitochondrial membrane potential and also less cellular damage accumulation than septuagenarians. Leukocyte chemotaxis and NK cell activity are significantly impaired in septuagenarians compared with young people whereas centenarians maintain them. To further ascertain the functional role of Bcl- xL in cellular aging, we found that lymphocytes from septuagenarians transduced with Bcl-xL display a reduction in senescent-related markers. Finally, to demonstrate the role of BcL-xL in longevity at the organism level, C. elegans bearing a gain of function mutation in the BcL-xL ortholog ced-9, showed a significant increase in mean and maximal life span. These results show that mRNA expression in centenarians is unique and reveals that BcL- xL plays an important role in exceptional aging. PMID:27794564

  13. Human 45,X Fibroblast Transcriptome Reveals Distinct Differentially Expressed Genes Including Long Noncoding RNAs Potentially Associated with the Pathophysiology of Turner Syndrome

    Science.gov (United States)

    Patowary, Ashok; Scaria, Vinod; Sivasubbu, Sridhar; Deobagkar, Deepti D.

    2014-01-01

    Turner syndrome is a chromosomal abnormality characterized by the absence of whole or part of the X chromosome in females. This X aneuploidy condition is associated with a diverse set of clinical phenotypes such as gonadal dysfunction, short stature, osteoporosis and Type II diabetes mellitus, among others. These phenotypes differ in their severity and penetrance among the affected individuals. Haploinsufficiency for a few X linked genes has been associated with some of these disease phenotypes. RNA sequencing can provide valuable insights to understand molecular mechanism of disease process. In the current study, we have analysed the transcriptome profiles of human untransformed 45,X and 46,XX fibroblast cells and identified differential expression of genes in these two karyotypes. Functional analysis revealed that these differentially expressing genes are associated with bone differentiation, glucose metabolism and gonadal development pathways. We also report differential expression of lincRNAs in X monosomic cells. Our observations provide a basis for evaluation of cellular and molecular mechanism(s) in the establishment of Turner syndrome phenotypes. PMID:24932682

  14. Depiction of carbohydrate-active enzyme diversity in Caldicellulosiruptor sp. F32 at the genome level reveals insights into distinct polysaccharide degradation features.

    Science.gov (United States)

    Meng, Dong-Dong; Ying, Yu; Zhang, Kun-Di; Lu, Ming; Li, Fu-Li

    2015-11-01

    Thermophilic bacterium Caldicellulosiruptor sp. F32 can utilize cellulose-, hemicellulose-containing biomass, including unpretreated wheat straw. We have conducted a bioinformatics analysis of the carbohydrate-active enzyme (CAZyme) in the genome of Caldicellulosiruptor sp. F32, which reveals a broad substrate range of the strain. Among 2285 predicted open reading frames (ORFs), 73 (3.2%) CAZyme encoding genes, including 44 glycoside hydrolases (GHs) distributing in 22 GH families, 6 carbohydrate esterases (CEs), 3 polysaccharide lyases (PLs), 21 glycosyl transferases (GTs), and 25 carbohydrate-binding modules (CBMs) were found. An in-depth bioinformatics analysis of CAZyme families that target cellulose, hemicellulose, chitin, pectin, starch, and β-1,3-1,4-glucan degradation were performed to highlight specialized polysaccharide degrading abilities of strain F32. A great number of orthologous multimodular CAZymes of Caldicellulosiruptor sp. F32 were found in other strains of genus Caldicellulosiruptor. While, a portion of the CAZymes of Caldicellulosiruptor sp. F32 showed sequence identity with proteins from strains of genus Clostridium. A thermostable β-glucosidase BlgA synergistically facilitated the enzymatic degradation of Avicel by endo-1,4-β-glucanase CelB, which indicated that the synchronous action of synergism between CAZymes enhanced the lignocellulose degradation by Caldicellulosiruptor sp. F32.

  15. Loss of PTB or negative regulation of Notch mRNA reveals distinct zones of Notch and actin protein accumulation in Drosophila embryo.

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    Cedric S Wesley

    Full Text Available Polypyrimidine Tract Binding (PTB protein is a regulator of mRNA processing and translation. Genetic screens and studies of wing and bristle development during the post-embryonic stages of Drosophila suggest that it is a negative regulator of the Notch pathway. How PTB regulates the Notch pathway is unknown. Our studies of Drosophila embryogenesis indicate that (1 the Notch mRNA is a potential target of PTB, (2 PTB and Notch functions in the dorso-lateral regions of the Drosophila embryo are linked to actin regulation but not their functions in the ventral region, and (3 the actin-related Notch activity in the dorso-lateral regions might require a Notch activity at or near the cell surface that is different from the nuclear Notch activity involved in cell fate specification in the ventral region. These data raise the possibility that the Drosophila embryo is divided into zones of different PTB and Notch activities based on whether or not they are linked to actin regulation. They also provide clues to the almost forgotten role of Notch in cell adhesion and reveal a role for the Notch pathway in cell fusions.

  16. Distinctive Structure of the EphA3/Ephrin-A5 Complex Reveals a Dual Mode of Eph Receptor Interaction for Ephrin-A5.

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    Garry Jason Forse

    Full Text Available The Eph receptor tyrosine kinase/ephrin ligand system regulates a wide spectrum of physiological processes, while its dysregulation has been implicated in cancer progression. The human EphA3 receptor is widely upregulated in the tumor microenvironment and is highly expressed in some types of cancer cells. Furthermore, EphA3 is among the most highly mutated genes in lung cancer and it is also frequently mutated in other cancers. We report the structure of the ligand-binding domain of the EphA3 receptor in complex with its preferred ligand, ephrin-A5. The structure of the complex reveals a pronounced tilt of the ephrin-A5 ligand compared to its orientation when bound to the EphA2 and EphB2 receptors and similar to its orientation when bound to EphA4. This tilt brings an additional area of ephrin-A5 into contact with regions of EphA3 outside the ephrin-binding pocket thereby enlarging the size of the interface, which is consistent with the high binding affinity of ephrin-A5 for EphA3. This large variation in the tilt of ephrin-A5 bound to different Eph receptors has not been previously observed for other ephrins.

  17. Human 45,X fibroblast transcriptome reveals distinct differentially expressed genes including long noncoding RNAs potentially associated with the pathophysiology of Turner syndrome.

    Directory of Open Access Journals (Sweden)

    Shriram N Rajpathak

    Full Text Available Turner syndrome is a chromosomal abnormality characterized by the absence of whole or part of the X chromosome in females. This X aneuploidy condition is associated with a diverse set of clinical phenotypes such as gonadal dysfunction, short stature, osteoporosis and Type II diabetes mellitus, among others. These phenotypes differ in their severity and penetrance among the affected individuals. Haploinsufficiency for a few X linked genes has been associated with some of these disease phenotypes. RNA sequencing can provide valuable insights to understand molecular mechanism of disease process. In the current study, we have analysed the transcriptome profiles of human untransformed 45,X and 46,XX fibroblast cells and identified differential expression of genes in these two karyotypes. Functional analysis revealed that these differentially expressing genes are associated with bone differentiation, glucose metabolism and gonadal development pathways. We also report differential expression of lincRNAs in X monosomic cells. Our observations provide a basis for evaluation of cellular and molecular mechanism(s in the establishment of Turner syndrome phenotypes.

  18. Human 45,X fibroblast transcriptome reveals distinct differentially expressed genes including long noncoding RNAs potentially associated with the pathophysiology of Turner syndrome.

    Science.gov (United States)

    Rajpathak, Shriram N; Vellarikkal, Shamsudheen Karuthedath; Patowary, Ashok; Scaria, Vinod; Sivasubbu, Sridhar; Deobagkar, Deepti D

    2014-01-01

    Turner syndrome is a chromosomal abnormality characterized by the absence of whole or part of the X chromosome in females. This X aneuploidy condition is associated with a diverse set of clinical phenotypes such as gonadal dysfunction, short stature, osteoporosis and Type II diabetes mellitus, among others. These phenotypes differ in their severity and penetrance among the affected individuals. Haploinsufficiency for a few X linked genes has been associated with some of these disease phenotypes. RNA sequencing can provide valuable insights to understand molecular mechanism of disease process. In the current study, we have analysed the transcriptome profiles of human untransformed 45,X and 46,XX fibroblast cells and identified differential expression of genes in these two karyotypes. Functional analysis revealed that these differentially expressing genes are associated with bone differentiation, glucose metabolism and gonadal development pathways. We also report differential expression of lincRNAs in X monosomic cells. Our observations provide a basis for evaluation of cellular and molecular mechanism(s) in the establishment of Turner syndrome phenotypes.

  19. Biofilm-associated persistence of food-borne pathogens.

    Science.gov (United States)

    Bridier, A; Sanchez-Vizuete, P; Guilbaud, M; Piard, J-C; Naïtali, M; Briandet, R

    2015-02-01

    Microbial life abounds on surfaces in both natural and industrial environments, one of which is the food industry. A solid substrate, water and some nutrients are sufficient to allow the construction of a microbial fortress, a so-called biofilm. Survival strategies developed by these surface-associated ecosystems are beginning to be deciphered in the context of rudimentary laboratory biofilms. Gelatinous organic matrices consisting of complex mixtures of self-produced biopolymers ensure the cohesion of these biological structures and contribute to their resistance and persistence. Moreover, far from being just simple three-dimensional assemblies of identical cells, biofilms are composed of heterogeneous sub-populations with distinctive behaviours that contribute to their global ecological success. In the clinical field, biofilm-associated infections (BAI) are known to trigger chronic infections that require dedicated therapies. A similar belief emerging in the food industry, where biofilm tolerance to environmental stresses, including cleaning and disinfection/sanitation, can result in the persistence of bacterial pathogens and the recurrent cross-contamination of food products. The present review focuses on the principal mechanisms involved in the formation of biofilms of food-borne pathogens, where biofilm behaviour is driven by its three-dimensional heterogeneity and by species interactions within these biostructures, and we look at some emergent control strategies.

  20. Bacterial community analysis of drinking water biofilms in southern Sweden.

    Science.gov (United States)

    Lührig, Katharina; Canbäck, Björn; Paul, Catherine J; Johansson, Tomas; Persson, Kenneth M; Rådström, Peter

    2015-01-01

    Next-generation sequencing of the V1-V2 and V3 variable regions of the 16S rRNA gene generated a total of 674,116 reads that described six distinct bacterial biofilm communities from both water meters and pipes. A high degree of reproducibility was demonstrated for the experimental and analytical work-flow by analyzing the communities present in parallel water meters, the rare occurrence of biological replicates within a working drinking water distribution system. The communities observed in water meters from households that did not complain about their drinking water were defined by sequences representing Proteobacteria (82-87%), with 22-40% of all sequences being classified as Sphingomonadaceae. However, a water meter biofilm community from a household with consumer reports of red water and flowing water containing elevated levels of iron and manganese had fewer sequences representing Proteobacteria (44%); only 0.6% of all sequences were classified as Sphingomonadaceae; and, in contrast to the other water meter communities, markedly more sequences represented Nitrospira and Pedomicrobium. The biofilm communities in pipes were distinct from those in water meters, and contained sequences that were identified as Mycobacterium, Nocardia, Desulfovibrio, and Sulfuricurvum. The approach employed in the present study resolved the bacterial diversity present in these biofilm communities as well as the differences that occurred in biofilms within a single distribution system, and suggests that next-generation sequencing of 16S rRNA amplicons can show changes in bacterial biofilm communities associated with different water qualities.

  1. Mechanical properties of the superficial biofilm layer determine the architecture of biofilms.

    Science.gov (United States)

    Kundukad, Binu; Seviour, Thomas; Liang, Yang; Rice, Scott A; Kjelleberg, Staffan; Doyle, Patrick S

    2016-06-29

    Cells in biofilms sense and interact with their environment through the extracellular matrix. The physicochemical properties of the matrix, particularly at the biofilm-environment interface, determine how cells respond to changing conditions. In this study we describe the application of atomic force microscopy and confocal imaging to probe in situ the mechanical properties of these interfacial regions and to elucidate how key matrix components can contribute to the physical sensing by the cells. We describe how the Young's modulus of microcolonies differs according to the size and morphology of microcolonies, as well as the flow rate. The Young's modulus increased as a function of microcolony diameter, which was correlated with the production of the polysaccharide Psl at later stages of maturation for hemispherical or mushroom shaped microcolonies. The Young's modulus of the periphery of the biofilm colony was however independent of the hydrodynamic shear. The morphology of the microcolonies also influenced interfacial or peripheral stiffness. Microcolonies with a diffuse morphology had a lower Young's modulus than isolated, circular ones and this phenomenon was due to a deficiency of Psl. In this way, changes in the specific polysaccharide components imbue the biofilm with distinct physical properties that may modulate the way in which bacteria perceive or respond to their environment. Further, the physical properties of the polysaccharides are closely linked to the specific architectures formed by the developing biofilm.

  2. Systematic analysis of phloem-feeding insect-induced transcriptional reprogramming in Arabidopsis highlights common features and reveals distinct responses to specialist and generalist insects.

    Science.gov (United States)

    Foyer, Christine H; Verrall, Susan R; Hancock, Robert D

    2015-02-01

    Phloem-feeding insects (PFIs), of which aphids are the largest group, are major agricultural pests causing extensive damage to crop plants. In contrast to chewing insects, the nature of the plant response to PFIs remains poorly characterized. Scrutiny of the literature concerning transcriptional responses of model and crop plant species to PFIs reveals surprisingly little consensus with respect to the transcripts showing altered abundance following infestation. Nevertheless, core features of the transcriptional response to PFIs can be defined in Arabidopsis thaliana. This comparison of the PFI-associated transcriptional response observed in A. thaliana infested by the generalists Myzus persicae and Bemisia tabaci with the specialist Brevicoryne brassicae highlights the importance of calcium-dependent and receptor kinase-associated signalling. We discuss these findings within the context of the complex cross-talk between the different hormones regulating basal immune response mechanisms in plants. We identify PFI-responsive genes, highlighting the importance of cell wall-associated kinases in plant-PFI interactions, as well as the significant role of kinases containing the domain of unknown function 26. A common feature of plant-PFI interaction is enhanced abundance of transcripts encoding WRKY transcription factors. However, significant divergence was observed with respect to secondary metabolism dependent upon the insect attacker. Transcripts encoding enzymes and proteins associated with glucosinolate metabolism were decreased following attack by the generalist M. persicae but not by the specialist B. brassicae. This analysis provides a comprehensive overview of the molecular patterns associated with the plant response to PFIs and suggests that plants recognize and respond to perturba