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Sample records for mannose-resistant proteus-like fimbriae

  1. Proteus mirabilis fimbriae- and urease-dependent clusters assemble in an extracellular niche to initiate bladder stone formation.

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    Schaffer, Jessica N; Norsworthy, Allison N; Sun, Tung-Tien; Pearson, Melanie M

    2016-04-19

    The catheter-associated uropathogenProteus mirabilisfrequently causes urinary stones, but little has been known about the initial stages of bladder colonization and stone formation. We found thatP. mirabilisrapidly invades the bladder urothelium, but generally fails to establish an intracellular niche. Instead, it forms extracellular clusters in the bladder lumen, which form foci of mineral deposition consistent with development of urinary stones. These clusters elicit a robust neutrophil response, and we present evidence of neutrophil extracellular trap generation during experimental urinary tract infection. We identified two virulence factors required for cluster development: urease, which is required for urolithiasis, and mannose-resistantProteus-like fimbriae. The extracellular cluster formation byP. mirabilisstands in direct contrast to uropathogenicEscherichia coli, which readily formed intracellular bacterial communities but not luminal clusters or urinary stones. We propose that extracellular clusters are a key mechanism ofP. mirabilissurvival and virulence in the bladder.

  2. Distinct Residues Contribute to Motility Repression and Autoregulation in the Proteus mirabilis Fimbria-Associated Transcriptional Regulator AtfJ.

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    Bode, Nadine J; Chan, Kun-Wei; Kong, Xiang-Peng; Pearson, Melanie M

    2016-08-01

    Proteus mirabilis contributes to a significant number of catheter-associated urinary tract infections, where coordinated regulation of adherence and motility is critical for ascending disease progression. Previously, the mannose-resistant Proteus-like (MR/P) fimbria-associated transcriptional regulator MrpJ has been shown to both repress motility and directly induce the transcription of its own operon; in addition, it affects the expression of a wide range of cellular processes. Interestingly, 14 additional mrpJ paralogs are included in the P. mirabilis genome. Looking at a selection of MrpJ paralogs, we discovered that these proteins, which consistently repress motility, also have nonidentical functions that include cross-regulation of fimbrial operons. A subset of paralogs, including AtfJ (encoded by the ambient temperature fimbrial operon), Fim8J, and MrpJ, are capable of autoinduction. We identified an element of the atf promoter extending from 487 to 655 nucleotides upstream of the transcriptional start site that is responsive to AtfJ, and we found that AtfJ directly binds this fragment. Mutational analysis of AtfJ revealed that its two identified functions, autoregulation and motility repression, are not invariably linked. Residues within the DNA-binding helix-turn-helix domain are required for motility repression but not necessarily autoregulation. Likewise, the C-terminal domain is dispensable for motility repression but is essential for autoregulation. Supported by a three-dimensional (3D) structural model, we hypothesize that the C-terminal domain confers unique regulatory capacities on the AtfJ family of regulators. Balancing adherence with motility is essential for uropathogens to successfully establish a foothold in their host. Proteus mirabilis uses a fimbria-associated transcriptional regulator to switch between these antagonistic processes by increasing fimbrial adherence while simultaneously downregulating flagella. The discovery of multiple

  3. Porphyromonas gingivalis fimbriae

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    Morten Enersen

    2013-05-01

    Full Text Available Marginal periodontitis is not a homogeneous disease but is rather influenced by an intricate set of host susceptibility differences as well as diversities in virulence among the harbored organisms. It is likely that clonal heterogeneity of subpopulations with both high and low levels of pathogenicity exists among organisms harbored by individuals with negligible, slight, or even severe periodontal destruction. Therefore, specific virulent clones of periodontal pathogens may cause advanced and/or aggressive periodontitis. Porphyromonas gingivalis is a predominant periodontal pathogen that expresses a number of potential virulence factors involved in the pathogenesis of periodontitis, and accumulated evidence shows that its expression of heterogenic virulence properties is dependent on clonal diversity. Fimbriae are considered to be critical factors that mediate bacterial interactions with and invasion of host tissues, with P. gingivalis shown to express two distinct fimbria-molecules, long and short fimbriae, on the cell surface, both of which seem to be involved in development of periodontitis. Long fimbriae are classified into six types (I to V and Ib based on the diversity of fimA genes encoding FimA (a subunit of long fimbriae. Studies of clones with type II fimA have revealed their significantly greater adhesive and invasive capabilities as compared to other fimA type clones. Long and short fimbriae induce various cytokine expressions such as IL-1α, IL-β, IL-6, and TNF-α, which result in alveolar bone resorption. Although the clonal diversity of short fimbriae is unclear, distinct short fimbria-molecules have been found in different strains. These fimbriae variations likely influence the development of periodontal disease.

  4. Occurrence of mannose resistant hemagglutinins in Escherichia coli strains isolated from porcine colibacillosis.

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    Truszczyński, M; Osek, J

    1987-01-01

    Three-hundred and fifty-eight E. coli strains isolated from piglets were tested for the presence of hemagglutinins by the use of the active hemagglutination test with or without mannose. Additionally 86 strains from the mentioned number of strains were investigated for the presence of common fimbriae using the same method but growing the strains in media especially suited for the development of this kind of fimbriae. These 358 strains and additionally 202 E. coli strains were tested using antisera for 987P and K88 antigens. It was found, using the active hemagglutination test, that 51.4% of the strains were hemagglutinating. The hemagglutinating strains carried the K88 antigen. All these strains were isolated from new-born and weaned piglets with enterotoxic form of colibacillosis, called also E. coli diarrhea. From cases of this form of colibacillosis originated also 26.7% of the strains in which common fimbriae (type 1) were detected. This result was obtained when the BHI medium was used for cultivation. In case of TSA medium only 2.3% of strains were positive. No specific or common fimbriae were found in strains recovered from septic form of colibacillosis and oedema disease (called also enterotoxaemic form of colibacillosis). No strain of 560 examined showed the presence of fimbrial 987P antigen.

  5. Increased Frequency of ColV Plasmids and Mannose-Resistant Hemagglutinating Activity in an Escherichia coli K1 Population

    OpenAIRE

    1984-01-01

    The expression of traits linked to pathogenicity was studied in a population of Escherichia coli K1 strains. It was found that E. coli K1 strains isolated from extraintestinal infection harbor the ColV plasmid and express mannose-resistant hemagglutinating activity type VI with a high frequency. The presence of these properties may play a role in the ability of some E. coli K1 serogroups to invade.

  6. Role of type 1 and type 3 fimbriae in Klebsiella pneumoniae biofilm formation

    DEFF Research Database (Denmark)

    Schroll, C.; Barken, Kim Bundvig; Krogfelt, K.A.

    2010-01-01

    nosocomial infections. Most clinical K. pneumoniae isolates express two types of fimbrial adhesins, type 1 fimbriae and type 3 fimbriae. In this study, we characterized the role of type 1 and type 3 fimbriae in K. pneumoniae biofilm formation. Results: Isogenic fimbriae mutants of the clinical K. pneumoniae...... of planktonic cells. Type 1 fimbriae did not influence biofilm formation and the expression of type 1 fimbriae was found to be down-regulated in biofilm forming cells. In contrast, expression of type 3 fimbriae was found to strongly promote biofilm formation. Conclusion: By use of well defined isogenic mutants...... we found that type 3 fimbriae, but not type 1 fimbriae, strongly promote biofilm formation in K. pneumoniae C3091. As the vast majority of clinical K. pneumoniae isolates express type 3 fimbriae, this fimbrial adhesin may play a significant role in development of catheter associated K. pneumoniae...

  7. Coordinate gene regulation by fimbriae-induced signal transduction

    DEFF Research Database (Denmark)

    Schembri, Mark; Klemm, Per

    2001-01-01

    whether fimbriae expression can affect expression of other genes, Analysis of gene expression in two E.coli strains, differing in the fim locus, indicated the flu gene to be affected. The flu gene encodes the antigen 43 (Ag43) surface protein, specifically involved in bacterial aggregation...

  8. Fimbriae have distinguishable roles in Proteus mirabilis biofilm formation.

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    Scavone, Paola; Iribarnegaray, Victoria; Caetano, Ana Laura; Schlapp, Geraldine; Härtel, Steffen; Zunino, Pablo

    2016-07-01

    Proteus mirabilis is one of the most common etiological agents of complicated urinary tract infections, especially those associated with catheterization. This is related to the ability of P. mirabilis to form biofilms on different surfaces. This pathogen encodes 17 putative fimbrial operons, the highest number found in any sequenced bacterial species so far. The present study analyzed the role of four P. mirabilis fimbriae (MR/P, UCA, ATF and PMF) in biofilm formation using isogenic mutants. Experimental approaches included migration over catheter, swimming and swarming motility, the semiquantitative assay based on adhesion and crystal violet staining, and biofilm development by immunofluorescence and confocal microscopy. Different assays were performed using LB or artificial urine. Results indicated that the different fimbriae contribute to the formation of a stable and functional biofilm. Fimbriae revealed particular associated roles. First, all the mutants showed a significantly reduced ability to migrate across urinary catheter sections but neither swimming nor swarming motility were affected. However, some mutants formed smaller biofilms compared with the wild type (MRP and ATF) while others formed significantly larger biofilms (UCA and PMF) showing different bioarchitecture features. It can be concluded that P. mirabilis fimbriae have distinguishable roles in the generation of biofilms, particularly in association with catheters. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  9. Chaperone-usher fimbriae in a diverse selection of Gallibacterium genomes

    DEFF Research Database (Denmark)

    Kudirkiene, Egle; Bager, Ragnhild Jørgensen; Johnson, Timothy J.

    2014-01-01

    Background Fimbriae are bacterial cell surface organelles involved in the pathogenesis of many bacterial species, including Gallibacterium anatis, in which a F17-like fimbriae of the chaperone-usher (CU) family was recently shown to be an important virulence factor and vaccine candidate. To reveal...... that their expression may require other in vitro or in vivo conditions. Conclusions This is the first approach establishing a systematic fimbria classification system within Gallibacterium spp., which indicates a species-wide distribution of γ4 CU fimbriae among a diverse collection of Gallibacterium isolates...

  10. Do type 1 fimbriae promote inflammation in the human urinary tract?

    DEFF Research Database (Denmark)

    Bergsten, G.; Wullt, B.; Schembri, Mark

    2007-01-01

    Type 1 fimbriae have been implicated as virulence factors in animal models of urinary tract infection (UTI), but the function in human disease remains unclear. This study used a human challenge model to examine if type 1 fimbriae trigger inflammation in the urinary tract. The asymptomatic...

  11. Novel Aggregative Adherence Fimbria Variant of Enteroaggregative Escherichia coli

    DEFF Research Database (Denmark)

    Jønsson, Rie; Struve, Carsten; Boisen, Nadia

    2015-01-01

    the stools of Danish adults with traveler’s diarrhea. We evaluated the presence of the aggregative adherence fimbriae (AAFs) by a multiplex PCR, targeting the four known major subunit variants as well as their usher-encoding genes. Almost one-half (49/118) of the clinical isolates did not possess any known...... AAF major fimbrial subunit, despite the presence of other AggR-related loci. Further investigation revealed the presence of an AAF-related gene encoding a yet-uncharacterized adhesin, termed agg5A. The sequence of the agg5DCBA gene cluster shared fimbrial accessory genes (usher, chaperone, and minor...

  12. The pancreatic zymogen granule membrane protein, GP2, binds Escherichia coli type 1 Fimbriae

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    Lowe Anson W

    2009-07-01

    Full Text Available Abstract Background GP2 is the major membrane protein present in the pancreatic zymogen granule, and is cleaved and released into the pancreatic duct along with exocrine secretions. The function of GP2 is unknown. GP2's amino acid sequence is most similar to that of uromodulin, which is secreted by the kidney. Recent studies have demonstrated uromodulin binding to bacterial Type 1 fimbria. The fimbriae serve as adhesins to host receptors. The present study examines whether GP2 also shares similar binding properties to bacteria with Type 1 fimbria. Commensal and pathogenic bacteria, including E. coli and Salmonella, express type 1 fimbria. Methods An in vitro binding assay was used to assay the binding of recombinant GP2 to defined strains of E. coli that differ in their expression of Type 1 fimbria or its subunit protein, FimH. Studies were also performed to determine whether GP2 binding is dependent on the presence of mannose residues, which is a known determinant for FimH binding. Results GP2 binds E. coli that express Type 1 fimbria. Binding is dependent on GP2 glycosylation, and specifically the presence of mannose residues. Conclusion GP2 binds to Type 1 fimbria, a bacterial adhesin that is commonly expressed by members of the Enterobacteriacae family.

  13. Characterization of 17 chaperone-usher fimbriae encoded by Proteus mirabilis reveals strong conservation

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    Kuan, Lisa; Schaffer, Jessica N.; Zouzias, Christos D.

    2014-01-01

    Proteus mirabilis is a Gram-negative enteric bacterium that causes complicated urinary tract infections, particularly in patients with indwelling catheters. Sequencing of clinical isolate P. mirabilis HI4320 revealed the presence of 17 predicted chaperone-usher fimbrial operons. We classified these fimbriae into three groups by their genetic relationship to other chaperone-usher fimbriae. Sixteen of these fimbriae are encoded by all seven currently sequenced P. mirabilis genomes. The predicted protein sequence of the major structural subunit for 14 of these fimbriae was highly conserved (≥95 % identity), whereas three other structural subunits (Fim3A, UcaA and Fim6A) were variable. Further examination of 58 clinical isolates showed that 14 of the 17 predicted major structural subunit genes of the fimbriae were present in most strains (>85 %). Transcription of the predicted major structural subunit genes for all 17 fimbriae was measured under different culture conditions designed to mimic conditions in the urinary tract. The majority of the fimbrial genes were induced during stationary phase, static culture or colony growth when compared to exponential-phase aerated culture. Major structural subunit proteins for six of these fimbriae were detected using MS of proteins sheared from the surface of broth-cultured P. mirabilis, demonstrating that this organism may produce multiple fimbriae within a single culture. The high degree of conservation of P. mirabilis fimbriae stands in contrast to uropathogenic Escherichia coli and Salmonella enterica, which exhibit greater variability in their fimbrial repertoires. These findings suggest there may be evolutionary pressure for P. mirabilis to maintain a large fimbrial arsenal. PMID:24809384

  14. Role of overexpressed CFA/I fimbriae in bacterial swimming.

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    Cao, Ling; Suo, Zhiyong; Lim, Timothy; Jun, Sangmu; Deliorman, Muhammedin; Riccardi, Carol; Kellerman, Laura; Avci, Recep; Yang, Xinghong

    2012-06-01

    Enterotoxigenic Escherichia coli CFA/I is a protective antigen and has been overexpressed in bacterial vectors, such as Salmonella Typhimurium H683, to generate vaccines. Effects that overexpressed CFA/I may engender on the bacterial host remain largely unexplored. To investigate, we constructed a high CFA/I expression strain, H683-pC2, and compared it to a low CFA/I expression strain, H683-pC, and to a non-CFA/I expression strain, H683-pY. The results showed that H683-pC2 was less able to migrate into semisolid agar (0.35%) than either H683-pC or H683-pY. Bacteria that migrated showed motility halo sizes of H683-pC2 CFA/I fimbriae on bacterial swimming motility.

  15. How type 1 fimbriae help Escherichia coli to evade extracellular antibiotics.

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    Avalos Vizcarra, Ima; Hosseini, Vahid; Kollmannsberger, Philip; Meier, Stefanie; Weber, Stefan S; Arnoldini, Markus; Ackermann, Martin; Vogel, Viola

    2016-01-05

    To survive antibiotics, bacteria use two different strategies: counteracting antibiotic effects by expression of resistance genes or evading their effects e.g. by persisting inside host cells. Since bacterial adhesins provide access to the shielded, intracellular niche and the adhesin type 1 fimbriae increases bacterial survival chances inside macrophages, we asked if fimbriae also influenced survival by antibiotic evasion. Combined gentamicin survival assays, flow cytometry, single cell microscopy and kinetic modeling of dose response curves showed that type 1 fimbriae increased the adhesion and internalization by macrophages. This was caused by strongly decreased off-rates and affected the number of intracellular bacteria but not the macrophage viability and morphology. Fimbriae thus promote antibiotic evasion which is particularly relevant in the context of chronic infections.

  16. AFSC/ABL: Young of the year sablefish (Anoplopoma fimbria) voucher otoliths 1995-2004.

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    National Oceanic and Atmospheric Administration, Department of Commerce — We studied young of the year sablefish Anoplopoma fimbria to collect basic life history information on their abundance, growth, and diet and to determine whether...

  17. Off-pathway assembly of fimbria subunits is prevented by chaperone CfaA of CFA/I fimbriae from enterotoxigenic E. coli.

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    Bao, Rui; Liu, Yang; Savarino, Stephen J; Xia, Di

    2016-12-01

    The assembly of the class 5 colonization factor antigen I (CFA/I) fimbriae of enterotoxigenic E. coli was proposed to proceed via the alternate chaperone-usher pathway. Here, we show that in the absence of the chaperone CfaA, CfaB, the major pilin subunit of CFA/I fimbriae, is able to spontaneously refold and polymerize into cyclic trimers. CfaA kinetically traps CfaB to form a metastable complex that can be stabilized by mutations. Crystal structure of the stabilized complex reveals distinctive interactions provided by CfaA to trap CfaB in an assembly competent state through donor-strand complementation (DSC) and cleft-mediated anchorage. Mutagenesis indicated that DSC controls the stability of the chaperone-subunit complex and the cleft-mediated anchorage of the subunit C-terminus additionally assist in subunit refolding. Surprisingly, over-stabilization of the chaperone-subunit complex led to delayed fimbria assembly, whereas destabilizing the complex resulted in no fimbriation. Thus, CfaA acts predominantly as a kinetic trap by stabilizing subunit to avoid its off-pathway self-polymerization that results in energetically favorable trimers and could serve as a driving force for CFA/I pilus assembly, representing an energetic landscape unique to class 5 fimbria assembly. Published 2016. This article is a U.S. Government work and is in the public domain in the USA. Molecular Microbiology published by John Wiley & Sons Ltd.

  18. Role of overexpressed CFA/I fimbriae in bacterial swimming

    International Nuclear Information System (INIS)

    Cao, Ling; Lim, Timothy; Jun, SangMu; Riccardi, Carol; Yang, Xinghong; Suo, Zhiyong; Deliorman, Muhammedin; Kellerman, Laura; Avci, Recep

    2012-01-01

    Enterotoxigenic Escherichia coli CFA/I is a protective antigen and has been overexpressed in bacterial vectors, such as Salmonella Typhimurium H683, to generate vaccines. Effects that overexpressed CFA/I may engender on the bacterial host remain largely unexplored. To investigate, we constructed a high CFA/I expression strain, H683-pC2, and compared it to a low CFA/I expression strain, H683-pC, and to a non-CFA/I expression strain, H683-pY. The results showed that H683-pC2 was less able to migrate into semisolid agar (0.35%) than either H683-pC or H683-pY. Bacteria that migrated showed motility halo sizes of H683-pC2 < H683-pC < H683-pY. In the liquid culture media, H683-pC2 cells precipitated to the bottom of the tube, while those of H683-pY did not. In situ imaging revealed that H683-pC2 bacilli tended to auto-agglutinate within the semisolid agar, while H683-pY bacilli did not. When the cfaBE fimbrial fiber encoding genes were deleted from pC2, the new plasmid, pC2(-), significantly recovered bacterial swimming capability. Our study highlights the negative impact of overexpressed CFA/I fimbriae on bacterial swimming motility. (paper)

  19. Distinct Mutations Led to Inactivation of Type 1 Fimbriae Expression in Shigella spp.

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    Bravo, Verónica; Puhar, Andrea; Sansonetti, Philippe; Parsot, Claude; Toro, Cecilia S.

    2015-01-01

    Shigella spp. are responsible for bacillary dysentery in humans. The acquisition or the modification of the virulence plasmid encoding factors promoting entry of bacteria into and dissemination within epithelial cells was a critical step in the evolution of these bacteria from their Escherichia coli ancestor(s). Incorporation of genomic islands (GI) and gene inactivation also shaped interactions between these pathogens and their human host. Sequence analysis of the GI inserted next to the leuX tRNA gene in S. boydii, S. dysenteriae, S. flexneri, S. sonnei and enteroinvasive E. coli (EIEC) suggests that this region initially carried the fec, yjhATS and fim gene clusters. The fim cluster encoding type I fimbriae is systematically inactivated in both reference strains and clinical isolates and distinct mutations are responsible for this inactivation in at least three phylogenetic groups. To investigate consequences of the presence of fimbriae on the outcome of the interaction of Shigella with host cells, we used a S. flexneri strain harboring a plasmid encoding the E. coli fim operon. Production of fimbriae by this recombinant strain increased the ability of bacteria to adhere to and enter into epithelial cells and had no effect on their ability to disseminate from cell to cell. The observations that production of type I fimbriae increases invasion of epithelial cells and that independent mutations abolish fimbriae production in Shigella suggest that these mutations correspond to pathoadaptive events. PMID:25811616

  20. Distinct mutations led to inactivation of type 1 fimbriae expression in Shigella spp.

    Directory of Open Access Journals (Sweden)

    Verónica Bravo

    Full Text Available Shigella spp. are responsible for bacillary dysentery in humans. The acquisition or the modification of the virulence plasmid encoding factors promoting entry of bacteria into and dissemination within epithelial cells was a critical step in the evolution of these bacteria from their Escherichia coli ancestor(s. Incorporation of genomic islands (GI and gene inactivation also shaped interactions between these pathogens and their human host. Sequence analysis of the GI inserted next to the leuX tRNA gene in S. boydii, S. dysenteriae, S. flexneri, S. sonnei and enteroinvasive E. coli (EIEC suggests that this region initially carried the fec, yjhATS and fim gene clusters. The fim cluster encoding type I fimbriae is systematically inactivated in both reference strains and clinical isolates and distinct mutations are responsible for this inactivation in at least three phylogenetic groups. To investigate consequences of the presence of fimbriae on the outcome of the interaction of Shigella with host cells, we used a S. flexneri strain harboring a plasmid encoding the E. coli fim operon. Production of fimbriae by this recombinant strain increased the ability of bacteria to adhere to and enter into epithelial cells and had no effect on their ability to disseminate from cell to cell. The observations that production of type I fimbriae increases invasion of epithelial cells and that independent mutations abolish fimbriae production in Shigella suggest that these mutations correspond to pathoadaptive events.

  1. Contribution of the major and minor subunits to fimbria-mediated adherence of Haemophilus influenzae to human epithelial cells and erythrocytes

    NARCIS (Netherlands)

    van Ham, S. M.; van Alphen, L.; Mooi, F. R.; van Putten, J. P.

    1995-01-01

    Fimbriae are colonization factors of the human pathogen Haemophilus influenzae in that they mediate bacterial adherence to human eukaryotic cells. The contribution of the major (HifA) and putative minor (HifD and HifE) subunits of H. influenzae fimbriae to fimbria-specific adherence was studied by

  2. Porcine aminopeptidase N binds to F4+ enterotoxigenic Escherichia coli fimbriae.

    Science.gov (United States)

    Xia, Pengpeng; Wang, Yiting; Zhu, Congrui; Zou, Yajie; Yang, Ying; Liu, Wei; Hardwidge, Philip R; Zhu, Guoqiang

    2016-02-09

    F4(+) enterotoxigenic Escherichia coli (ETEC) strains cause diarrheal disease in neonatal and post-weaned piglets. Several different host receptors for F4 fimbriae have been described, with porcine aminopeptidase N (APN) reported most recently. The FaeG subunit is essential for the binding of the three F4 variants to host cells. Here we show in both yeast two-hybrid and pulldown assays that APN binds directly to FaeG, the major subunit of F4 fimbriae, from three serotypes of F4(+) ETEC. Modulating APN gene expression in IPEC-J2 cells affected ETEC adherence. Antibodies raised against APN or F4 fimbriae both reduced ETEC adherence. Thus, APN mediates the attachment of F4(+) E. coli to intestinal epithelial cells.

  3. P-fimbriae in the presence of anti-PapA antibodies: new insight of antibodies action against pathogens

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    Mortezaei, Narges; Singh, Bhupender; Bullitt, Esther; Uhlin, Bernt Eric; Andersson, Magnus

    2013-12-01

    Uropathogenic strains of Escherichia coli establish urinary tract infections by attaching to host epithelial cells using adhesive organelles called fimbriae. Fimbriae are helix-like structures with a remarkable adaptability, offering safeguarding for bacteria exposed to changing fluid forces in the urinary tract. We challenged this property of P-fimbriae by cross-linking their subunits with shaft-specific antibodies and measuring the corresponding force response at a single organelle level. Our data show compromised extension and rewinding of P-fimbriae in the presence of antibodies and reduced fimbrial elasticity, which are important properties of fimbriae contributing to the ability of bacteria to cause urinary tract infections. The reduced elasticity found by cross-linking fimbrial subunits could thus be another assignment for antibodies; in addition to marking bacteria as foreign, antibodies physically compromise fimbrial function. We suggest that our assay and results will be a starting point for further investigations aimed at inhibiting sustained bacterial adhesion by antibodies.

  4. Authentic display of a cholera toxin epitope by chimeric type 1 fimbriae

    DEFF Research Database (Denmark)

    Stentebjerg-Olesen, Bodil; Pallesen, Lars; Jensen, Lars Bogø

    1997-01-01

    . Several of the chosen positions seemed amenable even for large foreign inserts; the chimeric proteins were exposed on the bacterial surface and the cholera toxin epitope was authentically displayed, i.e. it was recognized on bacteria by specific antiserum. Display of chimeric fimbriae was tested...... with respect to host background in three different Escherichia coli strains, i.e. an isogenic set of K-12 strains, differing in the presence of an indigenous fim gene cluster, as well as a wild-type isolate. Immunization of rabbits with purified chimeric fimbriae resulted in serum which specifically recognized...

  5. Clathrin-mediated endocytosis and transcytosis of enterotoxigenic Escherichia coli F4 fimbriae in porcine intestinal epithelial cells.

    Science.gov (United States)

    Rasschaert, Kristien; Devriendt, Bert; Favoreel, Herman; Goddeeris, Bruno M; Cox, Eric

    2010-10-15

    Enterotoxigenic Escherichia coli (ETEC) cause severe diarrhea in neonatal and recently weaned piglets. Previously, we demonstrated that oral immunization of F4 receptor positive piglets with purified F4 fimbriae induces a protective F4-specific intestinal immune response. However, in F4 receptor negative animals no F4-specific immune response can be elicited, indicating that the induction of an F4-specific mucosal immune response upon oral immunisation is receptor-dependent. Although F4 fimbriae undergo transcytosis across the intestinal epithelium in vivo, the endocytosis pathways used remain unknown. In the present study, we characterized the internalization of F4 fimbriae in the porcine intestinal epithelial cell line IPEC-J2. The results in the present study demonstrate that F4 fimbriae are internalized through a clathrin-dependent pathway. Furthermore, our results suggest that F4 fimbriae are transcytosed across differentiated IPEC-J2 cells. This receptor-dependent transcytosis of F4 fimbriae may explain the immunogenicity of these fimbriae upon oral administration in vivo. (c) 2010 Elsevier B.V. All rights reserved.

  6. Receptor for the F4 fimbriae of enterotoxigenic Escherichia coli (ETEC).

    Science.gov (United States)

    Xia, Pengpeng; Zou, Yajie; Wang, Yiting; Song, Yujie; Liu, Wei; Francis, David H; Zhu, Guoqiang

    2015-06-01

    Infection with F4(+) enterotoxigenic Escherichia coli (ETEC) responsible for diarrhea in neonatal and post-weaned piglets leads to great economic losses in the swine industry. These pathogenic bacteria express either of three fimbrial variants F4ab, F4ac, and F4ad, which have long been known for their importance in host infection and initiating protective immune responses. The initial step in infection for the bacterium is to adhere to host enterocytes through fimbriae-mediated recognition of receptors on the host cell surface. A number of receptors for ETEC F4 have now been described and characterized, but their functions are still poorly understood. The current review summarizes the latest research addressing the characteristics of F4 fimbriae receptors and the interactions of F4 fimbriae and their receptors on host cells. These include observations that as follows: (1) FaeG mediates the binding activities of F4 and is an essential component of the F4 fimbriae, (2) the F4 fimbrial receptor gene is located in a region of chromosome 13, (3) the biochemical properties of F4 fimbrial receptors that form the binding site of the bacterium are now recognized, and (4) specific receptors confer susceptibility/resistance to ETEC F4 infection in pigs. Characterizing the host-pathogen interaction will be crucial to understand the pathogenicity of the bacteria, provide insights into receptor activation of the innate immune system, and develop therapeutic strategies to prevent this illness.

  7. Erythrocyte and porcine intestinal glycosphingolipids recognized by F4 fimbriae of enterotoxigenic Escherichia coli.

    Science.gov (United States)

    Coddens, Annelies; Valis, Erik; Benktander, John; Ångström, Jonas; Breimer, Michael E; Cox, Eric; Teneberg, Susann

    2011-01-01

    Enterotoxigenic F4-fimbriated Escherichia coli is associated with diarrheal disease in neonatal and postweaning pigs. The F4 fimbriae mediate attachment of the bacteria to the pig intestinal epithelium, enabling an efficient delivery of diarrhea-inducing enterotoxins to the target epithelial cells. There are three variants of F4 fimbriae designated F4ab, F4ac and F4ad, respectively, having different antigenic and adhesive properties. In the present study, the binding of isolated F4ab, F4ac and F4ad fimbriae, and F4ab/ac/ad-fimbriated E. coli, to glycosphingolipids from erythrocytes and from porcine small intestinal epithelium was examined, in order to get a comprehensive view of the F4-binding glycosphingolipids involved in F4-mediated hemagglutination and adhesion to the epithelial cells of porcine intestine. Specific interactions between the F4ab, F4ac and F4ad fimbriae and both acid and non-acid glycosphingolipids were obtained, and after isolation of binding-active glycosphingolipids and characterization by mass spectrometry and proton NMR, distinct carbohydrate binding patterns were defined for each fimbrial subtype. Two novel glycosphingolipids were isolated from chicken erythrocytes, and characterized as GalNAcα3GalNAcß3Galß4Glcß1Cer and GalNAcα3GalNAcß3Galß4GlcNAcß3Galß4Glcß1Cer. These two compounds, and lactosylceramide (Galß4Glcß1Cer) with phytosphingosine and hydroxy fatty acid, were recognized by all three variants of F4 fimbriae. No binding of the F4ad fimbriae or F4ad-fimbriated E. coli to the porcine intestinal glycosphingolipids occurred. However, for F4ab and F4ac two distinct binding patterns were observed. The F4ac fimbriae and the F4ac-expressing E. coli selectively bound to galactosylceramide (Galß1Cer) with sphingosine and hydroxy 24:0 fatty acid, while the porcine intestinal glycosphingolipids recognized by F4ab fimbriae and the F4ab-fimbriated bacteria were characterized as galactosylceramide, sulfatide (SO(3)-3Galß1Cer), sulf

  8. The polymeric stability of the Escherichia coli F4 (K88) fimbriae enhances its mucosal immunogenicity following oral immunization.

    Science.gov (United States)

    Verdonck, Frank; Joensuu, Jussi Joonas; Stuyven, Edith; De Meyer, Julie; Muilu, Mikko; Pirhonen, Minna; Goddeeris, Bruno Maria; Mast, Jan; Niklander-Teeri, Viola; Cox, Eric

    2008-10-23

    Only a few vaccines are commercially available against intestinal infections since the induction of a protective intestinal immune response is difficult to achieve. For instance, oral administration of most proteins results in oral tolerance instead of an antigen-specific immune response. We have shown before that as a result of oral immunization of piglets with F4 fimbriae purified from pathogenic enterotoxigenic Escherichia coli (ETEC), the fimbriae bind to the F4 receptor (F4R) in the intestine and induce a protective F4-specific immune response. F4 fimbriae are very stable polymeric structures composed of some minor subunits and a major subunit FaeG that is also the fimbrial adhesin. In the present study, the mutagenesis experiments identified FaeG amino acids 97 (N to K) and 201 (I to V) as determinants for F4 polymeric stability. The interaction between the FaeG subunits in mutant F4 fimbriae is reduced but both mutant and wild type fimbriae behaved identically in F4R binding and showed equal stability in the gastro-intestinal lumen. Oral immunization experiments indicated that a higher degree of polymerisation of the fimbriae in the intestine was correlated with a better F4-specific mucosal immunogenicity. These data suggest that the mucosal immunogenicity of soluble virulence factors can be increased by the construction of stable polymeric structures and therefore help in the development of effective mucosal vaccines.

  9. Binding determinants in the interplay between porcine aminopeptidase N and enterotoxigenic Escherichia coli F4 fimbriae.

    Science.gov (United States)

    Xia, Pengpeng; Quan, Guomei; Yang, Yi; Zhao, Jing; Wang, Yiting; Zhou, Mingxu; Hardwidge, Philip R; Zhu, Jianzhong; Liu, Siguo; Zhu, Guoqiang

    2018-02-26

    The binding of F4 + enterotoxigenic Escherichia coli (ETEC) and the specific receptor on porcine intestinal epithelial cells is the initial step in F4 + ETEC infection. Porcine aminopeptidase N (APN) is a newly discovered receptor for F4 fimbriae that binds directly to FaeG adhesin, which is the major subunit of the F4 fimbriae variants F4ab, F4ac, and F4ad. We used overlapping peptide assays to map the APN-FaeG binding sites, which has facilitated in the identifying the APN-binding amino acids that are located in the same region of FaeG variants, thereby limiting the major binding regions of APN to 13 peptides. To determine the core sequence motif, a panel of FaeG peptides with point mutations and FaeG mutants were constructed. Pull-down and binding reactivity assays using piglet intestines determined that the amino acids G159 of F4ab, N209 and L212 of F4ac, and A200 of F4ad were the critical residues for APN binding of FaeG. We further show using ELISA and confocal microscopy assay that amino acids 553-568, and 652-670 of the APN comprise the linear epitope for FaeG binding in all three F4 fimbriae variants.

  10. F4+ ETEC infection and oral immunization with F4 fimbriae elicits an IL-17-dominated immune response.

    Science.gov (United States)

    Luo, Yu; Van Nguyen, Ut; de la Fe Rodriguez, Pedro Y; Devriendt, Bert; Cox, Eric

    2015-10-21

    Enterotoxigenic Escherichia coli (ETEC) are an important cause of post-weaning diarrhea (PWD) in piglets. Porcine-specific ETEC strains possess different fimbrial subtypes of which F4 fimbriae are the most frequently associated with ETEC-induced diarrhea in piglets. These F4 fimbriae are potent oral immunogens that induce protective F4-specific IgA antibody secreting cells at intestinal tissues. Recently, T-helper 17 (Th17) cells have been implicated in the protection of the host against extracellular pathogens. However, it remains unknown if Th17 effector responses are needed to clear ETEC infections. In the present study, we aimed to elucidate if ETEC elicits a Th17 response in piglets and if F4 fimbriae trigger a similar response. F4(+) ETEC infection upregulated IL-17A, IL-17F, IL-21 and IL-23p19, but not IL-12 and IFN-γ mRNA expression in the systemic and mucosal immune system. Similarly, oral immunization with F4 fimbriae triggered a Th17 signature evidenced by an upregulated mRNA expression of IL-17F, RORγt, IL-23p19 and IL-21 in the peripheral blood mononuclear cells (PBMCs). Intriguingly, IL-17A mRNA levels were unaltered. To further evaluate this difference between systemic and mucosal immune responses, we assayed the cytokine mRNA profile of F4 fimbriae stimulated PBMCs. F4 fimbriae induced IL-17A, IL-17F, IL-22 and IL-23p19, but downregulated IL-17B mRNA expression. Altogether, these data indicate a Th17 dominated response upon oral immunization with F4 fimbriae and F4(+) ETEC infection. Our work also highlights that IL-17B and IL-17F participate in the immune response to protect the host against F4(+) ETEC infection and could aid in the design of future ETEC vaccines.

  11. Hemagglutinin Typing as an Aid in Identification of Biochemically Atypical Escherichia coli Strains

    OpenAIRE

    Crichton, Pamela B.; Ip, S. M.; Old, D. C.

    1981-01-01

    Tests for the presence of mannose-sensitive and mannose-resistant, eluting hemagglutinins and fimbriae were helpful in indicating whether biochemically atypical strains of the tribe Escherichieae might be escherichiae or shigellae.

  12. Hemagglutinin Typing as an Aid in Identification of Biochemically Atypical Escherichia coli Strains

    Science.gov (United States)

    Crichton, Pamela B.; Ip, S. M.; Old, D. C.

    1981-01-01

    Tests for the presence of mannose-sensitive and mannose-resistant, eluting hemagglutinins and fimbriae were helpful in indicating whether biochemically atypical strains of the tribe Escherichieae might be escherichiae or shigellae. PMID:7334072

  13. Type 3 Fimbriae Encoded on Plasmids Are Expressed from a Unique Promoter without Affecting Host Motility, Facilitating an Exceptional Phenotype That Enhances Conjugal Plasmid Transfer

    DEFF Research Database (Denmark)

    Madsen, Jonas Stenlokke; Riber, Leise; Kot, Witold

    2016-01-01

    Horizontal gene transfer (HGT), the transmission of genetic material to a recipient that is not the progeny of the donor, is fundamental in bacterial evolution. HGT is often mediated by mobile genetic elements such as conjugative plasmids, which may be in conflict with the chromosomal elements...... of the genome because they are independent replicons that may petition their own evolutionary strategy. Here we study differences between type 3 fimbriae encoded on wild type plasmids and in chromosomes. Using known and newly characterized plasmids we show that the expression of type 3 fimbriae encoded...... on plasmids is systematically different, as MrkH, a c-di-GMP dependent transcriptional activator is not needed for strong expression of the fimbriae. MrkH is required for expression of type 3 fimbriae of the Klebsiella pneumoniae chromosome, wherefrom the fimbriae operon (mrkABCDF) of plasmids is believed...

  14. Oral Escherichia coli Colonization Factor Antigen I (CFA/I) Fimbriae Ameliorate Arthritis via IL-35, not IL-27

    Science.gov (United States)

    Kochetkova, Irina; Thornburg, Theresa; Callis, Gayle; Holderness, Kathryn; Maddaloni, Massimo; Pascual, David W.

    2014-01-01

    A Salmonella therapeutic expressing enterotoxigenic E. coli colonization factor antigen I (CFA/I) fimbriae protects against collagen-induced arthritis (CIA) by eliciting two regulatory T cell (Treg) subsets: TGF-β-producing Foxp3−CD39+CD4+ and IL-10-producing Foxp3+CD39+CD4+ T cells. However, it is unclear if CFA/I fimbriae alone are protective, and if other regulatory cytokines are involved especially in the context for the EBI3-sharing cytokines, Treg-derived IL-35 and APC-derived IL-27, both capable of suppressing Th17 cells and regulating autoimmune diseases. Subsequent evaluation revealed that a single oral dose of purified, soluble CFA/I fimbriae protected against CIA as effectively as Salmonella-CFA/I, and found Foxp3+CD39+CD4+ T cells as the source of secreted IL-35, whereas IL-27 production by CD11c+ cells was inhibited. Inquiring into their relevance, CFA/I fimbriae-treated IL-27 receptor-deficient (WSX-1−/−) mice were equally protected against CIA as wild-type mice suggesting a limited role for IL-27. In contrast, CFA/I fimbriae-mediated protection was abated in EBI3−/− mice accompanied by the loss of TGF-β- and IL-10-producing Tregs. Adoptive transfer of B6 CD39+CD4+ T cells to EBI3−/− mice with concurrent CFA/I plus IL-35 treatment effectively stimulated Tregs suppressing proinflammatory CII-specific Th cells. Opposingly, recipients co-transferred with B6 and EBI3−/− CD39+CD4+ T cells and treated with CFA/I plus IL-35 failed in protecting mice implicating the importance for endogenous IL-35 to confer CFA/I-mediated protection. Thus, CFA/I fimbriae stimulate IL-35 required for the co-induction of TGF-β and IL-10. PMID:24337375

  15. Novel Zn2+-chelating peptides selected from a fimbria-displayed random peptide library

    DEFF Research Database (Denmark)

    Kjærgaard, Kristian; Schembri, Mark; Klemm, Per

    2001-01-01

    The display of peptide sequences on the surface of bacteria is a technology that offers exciting applications in biotechnology and medical research. Type 1 fimbriae are surface organelles of Escherichia coli which mediate D-mannose-sensitive binding to different host surfaces by virtue of the Fim......H adhesin. FimH is a component of the fimbrial organelle that can accommodate and display a diverse range of peptide sequences on the E. coli cell surface. In this study we have constructed a random peptide library in FimH. The library, consisting of similar to 40 million individual clones, was screened...

  16. Authentic display of a cholera toxin epitope by chimeric type 1 fimbriae: effects of insert position and host background

    DEFF Research Database (Denmark)

    Stentebjerg-Olesen, B; Pallesen, L; Jensen, LB

    1997-01-01

    . Several of the chosen positions seemed amenable even for large foreign inserts; the chimeric proteins were exposed on the bacterial surface and the cholera toxin epitope was authentically displayed, i.e. it was recognized on bacteria by specific antiserum. Display of chimeric fimbriae was tested...... with respect to host background in three different Escherichia coli strains, i.e. an isogenic set of K-12 strains, differing in the presence of an indigenous fim gene cluster, as well as a wild-type isolate. Immunization of rabbits with purified chimeric fimbriae resulted in serum which specifically recognized...

  17. Crystallization of the FaeE chaperone of Escherichia coli F4 fimbriae

    International Nuclear Information System (INIS)

    Van Molle, Inge; Buts, Lieven; Coppens, Fanny; Qiang, Liu; Wyns, Lode; Loris, Remy; Bouckaert, Julie; De Greve, Henri

    2005-01-01

    The periplasmic chaperone FaeE of E. coli F4 fimbriae crystallizes in three crystal forms. F4 (formerly K88) fimbriae from enterotoxigenic Escherichia coli are assembled via the FaeE/FaeD chaperone/usher pathway. The chaperone FaeE crystallizes in three crystal forms, all belonging to space group C2. Crystals of form 1 diffract to 2.3 Å and have unit-cell parameters a = 195.7, b = 78.5, c = 184.6 Å, β = 102.2°. X-ray data for crystal form 2 were collected to 2.7 Å using an SeMet variant of FaeE. The crystals have unit-cell parameters a = 136.4, b = 75.7, c = 69.4 Å, β = 92.8°. Crystals of form 3 were formed in a solution containing the FaeE–FaeG complex and diffract to 2.8 Å. Unit-cell parameters are a = 109.7, b = 78.6, c = 87.8 Å, β = 96.4°

  18. Crystallization of the FaeE chaperone of Escherichia coli F4 fimbriae

    Energy Technology Data Exchange (ETDEWEB)

    Van Molle, Inge, E-mail: ivmolle@vub.ac.be; Buts, Lieven; Coppens, Fanny; Qiang, Liu; Wyns, Lode; Loris, Remy; Bouckaert, Julie; De Greve, Henri [Laboratorium voor Ultrastructuur, Vlaams Interuniversitair Instituut voor Biotechnologie, Vrije Universiteit Brussel, Pleinlaan 2, 1050 Brussel (Belgium)

    2005-04-01

    The periplasmic chaperone FaeE of E. coli F4 fimbriae crystallizes in three crystal forms. F4 (formerly K88) fimbriae from enterotoxigenic Escherichia coli are assembled via the FaeE/FaeD chaperone/usher pathway. The chaperone FaeE crystallizes in three crystal forms, all belonging to space group C2. Crystals of form 1 diffract to 2.3 Å and have unit-cell parameters a = 195.7, b = 78.5, c = 184.6 Å, β = 102.2°. X-ray data for crystal form 2 were collected to 2.7 Å using an SeMet variant of FaeE. The crystals have unit-cell parameters a = 136.4, b = 75.7, c = 69.4 Å, β = 92.8°. Crystals of form 3 were formed in a solution containing the FaeE–FaeG complex and diffract to 2.8 Å. Unit-cell parameters are a = 109.7, b = 78.6, c = 87.8 Å, β = 96.4°.

  19. Stimulation of Escherichia coli F-18Col- Type-1 fimbriae synthesis by leuX

    DEFF Research Database (Denmark)

    Newman, Joseph V.; Burghoff, Robert L.; Pallesen, Lars

    1994-01-01

    Escherichia coli F-18, a normal human fecal isolate, is an excellent colonizer of the streptomycin-treated mouse large intestine. E. coli F-18Col-, a derivative of E. coli F-18 which no longer makes the E. coli F-18 colicin, colonizes the large intestine as well as E. coli F-18 when fed to mice...... alone but is eliminated when fed together with E. coli F-18. Recently we randomly cloned E. coli F-18 DNA into E. coli F-18Col- and let the mouse intestine select the best colonizer. In this way, we isolated a 6.5-kb E. coli F-18 DNA sequence that simultaneously stimulated synthesis of type 1 fimbriae...... and enhanced E. coli F-18Col- colonizing ability. In the present investigation we show that the gene responsible for stimulation of type 1 fimbriae synthesis appears to be leuX, which encodes a tRNA specific for the rare leucine codon UUG. Moreover, it appears that expression of leuX may be regulated by two...

  20. A general procedure for small-scale purification of fimbriae expressed by porcine enterotoxigenic Escherichia coli strains

    Directory of Open Access Journals (Sweden)

    Ana Cristina Campal Espinosa

    2008-01-01

    Full Text Available Fimbriae expression by enterotoxigenic Escherichia coli strains is a complex process which is controlled by global and local transcriptional regulators and post-transcriptional control. It is influenced by factors such as bacterial growth rate, culture medium composition, pH and temperature. Fimbrial expression could thus frequently become lost. Bacterial culture procedures favouring fimbrial expression are thus needed. The fimbriated bacterial population was therefore enriched by static culture in Mueller–Hinton broth. Fimbrial expression was then maintained by making it grow consecutively in agar CFA and Minca or minimal broth according to the fimbrial serotype. Maximum fimbrial expression was reached after 4h or 5h in culture. The fimbriae were extracted by heat -shock treatment and precipitated with 40% ammonium sulphate. Further purification was carried out by molecular exclusion and sodium deoxycholate treatment. This methodology integrates known procedures in a simple and reproducible process for obtaining F4, F5, F6 and F41 fimbriae in sufficient quantities for their subsequent use in producing antibodies, immunoassays and other studies (at laboratory level requiring high-purity preparations (80% to maintain their native structure. Key words: Enterotoxigenic Escherichia coli; fimbriae; Minca; minimal medium; CFA.

  1. The export systems of type 1 and F1C fimbriae are interchangeable but work in parental pairs

    DEFF Research Database (Denmark)

    Klemm, P; Jørgensen, BJ; Kreft, B

    1995-01-01

    , specialized two-component assembly systems. The organization of the fim and foc gene clusters encoding these fimbriae, as well as the structure of the organelles, is very similar; however, the actual sequence homology of the structural elements is not remarkable (34 to 60%). Both gene clusters encode...

  2. CRP-Cyclic AMP Regulates the Expression of Type 3 Fimbriae via Cyclic di-GMP in Klebsiella pneumoniae.

    Directory of Open Access Journals (Sweden)

    Ching-Ting Lin

    Full Text Available Klebsiella pneumoniae is the predominant pathogen isolated from liver abscesses of diabetic patients in Asian countries. However, the effects of elevated blood glucose levels on the virulence of this pathogen remain largely unknown. Type 3 fimbriae, encoded by the mrkABCDF genes, are important virulence factors in K. pneumoniae pathogenesis. In this study, the effects of exogenous glucose and the intracellular cyclic AMP (cAMP signaling pathway on type 3 fimbriae expression regulation were investigated. The production of MrkA, the major subunit of type 3 fimbriae, was increased in glucose-rich medium, whereas cAMP supplementation reversed the effect. MrkA production was markedly increased by cyaA or crp deletion, but slightly decreased by cpdA deletion. In addition, the mRNA levels of mrkABCDF genes and the activity of PmrkA were increased in Δcrp strain, as well as the mRNA levels of mrkHIJ genes that encode cyclic di-GMP (c-di-GMP-related regulatory proteins that influence type 3 fimbriae expression. Moreover, the activities of PmrkHI and PmrkJ were decreased in ΔlacZΔcrp strain. These results indicate that CRP-cAMP down-regulates mrkABCDF and mrkHIJ at the transcriptional level. Further deletion of mrkH or mrkI in Δcrp strain diminished the production of MrkA, indicating that MrkH and MrkI are required for the CRP regulation of type 3 fimbriae expression. Furthermore, the high activity of PmrkHI in the ΔlacZΔcrp strain was diminished in ΔlacZΔcrpΔmrkHI, but increased in the ΔlacZΔcrpΔmrkJ strain. Deletion of crp increased the intracellular c-di-GMP concentration and reduced the phosphodiesterase activity. Moreover, we found that the mRNA levels of multiple genes related to c-di-GMP metabolism were altered in Δcrp strain. These indicate that CRP regulates type 3 fimbriae expression indirectly via the c-di-GMP signaling pathway. In conclusion, we found evidence of a coordinated regulation of type 3 fimbriae expression by the CRP

  3. Transcriptome of Proteus mirabilis in the Murine Urinary Tract: Virulence and Nitrogen Assimilation Gene Expression▿†

    Science.gov (United States)

    Pearson, Melanie M.; Yep, Alejandra; Smith, Sara N.; Mobley, Harry L. T.

    2011-01-01

    The enteric bacterium Proteus mirabilis is a common cause of complicated urinary tract infections. In this study, microarrays were used to analyze P. mirabilis gene expression in vivo from experimentally infected mice. Urine was collected at 1, 3, and 7 days postinfection, and RNA was isolated from bacteria in the urine for transcriptional analysis. Across nine microarrays, 471 genes were upregulated and 82 were downregulated in vivo compared to in vitro broth culture. Genes upregulated in vivo encoded mannose-resistant Proteus-like (MR/P) fimbriae, urease, iron uptake systems, amino acid and peptide transporters, pyruvate metabolism enzymes, and a portion of the tricarboxylic acid (TCA) cycle enzymes. Flagella were downregulated. Ammonia assimilation gene glnA (glutamine synthetase) was repressed in vivo, while gdhA (glutamate dehydrogenase) was upregulated in vivo. Contrary to our expectations, ammonia availability due to urease activity in P. mirabilis did not drive this gene expression. A gdhA mutant was growth deficient in minimal medium with citrate as the sole carbon source, and loss of gdhA resulted in a significant fitness defect in the mouse model of urinary tract infection. Unlike Escherichia coli, which represses gdhA and upregulates glnA in vivo and cannot utilize citrate, the data suggest that P. mirabilis uses glutamate dehydrogenase to monitor carbon-nitrogen balance, and this ability contributes to the pathogenic potential of P. mirabilis in the urinary tract. PMID:21505083

  4. FimH adhesin of Escherichia coli K1 type 1 fimbriae activates BV-2 microglia

    International Nuclear Information System (INIS)

    Lee, Jongseok; Shin, Sooan; Teng, C.-H.; Hong, Suk Jin; Kim, Kwang Sik

    2005-01-01

    The generation of intense inflammation in the subarachnoid space in response to meningitis-causing bacteria contributes to brain dysfunction and neuronal injury in bacterial meningitis. Microglia, the major immune effector cells in the central nervous system (CNS), become activated by bacterial components to produce proinflammatory immune mediators. In this study, we showed that FimH adhesin, a tip component of type 1 fimbriae of meningitis-causing Escherichia coli K1, activated the murine microglial cell line, BV-2, which resulted in the production of nitric oxide and the release of tumor necrosis factor-α. Mitogen-activated protein kinases, ERK and p-38, and nuclear factor-κB were involved in FimH adhesin-mediated microglial activation. These findings suggest that FimH adhesin contributes to the CNS inflammatory response by virtue of activating microglia in E. coli meningitis

  5. Maternal immunity enhances systemic recall immune responses upon oral immunization of piglets with F4 fimbriae.

    Science.gov (United States)

    Nguyen, Ut V; Melkebeek, Vesna; Devriendt, Bert; Goetstouwers, Tiphanie; Van Poucke, Mario; Peelman, Luc; Goddeeris, Bruno M; Cox, Eric

    2015-06-23

    F4 enterotoxigenic Escherichia coli (ETEC) cause diarrhoea and mortality in piglets leading to severe economic losses. Oral immunization of piglets with F4 fimbriae induces a protective intestinal immune response evidenced by an F4-specific serum and intestinal IgA response. However, successful oral immunization of pigs with F4 fimbriae in the presence of maternal immunity has not been demonstrated yet. In the present study we aimed to evaluate the effect of maternal immunity on the induction of a systemic immune response upon oral immunization of piglets. Whereas F4-specific IgG and IgA could be induced by oral immunization of pigs without maternal antibodies and by intramuscular immunization of pigs with maternal antibodies, no such response was seen in the orally immunized animals with maternal antibodies. Since maternal antibodies can mask an antibody response, we also looked by ELIspot assays for circulating F4-specific antibody secreting cells (ASCs). Enumerating the F4-specific ASCs within the circulating peripheral blood mononuclear cells, and the number of F4-specific IgA ASCs within the circulating IgA(+) B-cells revealed an F4-specific immune response in the orally immunized animals with maternal antibodies. Interestingly, results suggest a more robust IgA booster response by oral immunization of pigs with than without maternal antibodies. These results demonstrate that oral immunization of piglets with F4-specific maternal antibodies is feasible and that these maternal antibodies seem to enhance the secondary systemic immune response. Furthermore, our ELIspot assay on enriched IgA(+) B-cells could be used as a screening procedure to optimize mucosal immunization protocols in pigs with maternal immunity.

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

    Directory of Open Access Journals (Sweden)

    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

  7. F9 fimbriae of uropathogenic Escherichia coli are expressed at low temperature and recognise Galβ1-3GlcNAc-containing glycans.

    Directory of Open Access Journals (Sweden)

    Daniël J Wurpel

    Full Text Available Uropathogenic Escherichia coli (UPEC is the leading causative agent of urinary tract infections (UTI in the developed world. Among the major virulence factors of UPEC, surface expressed adhesins mediate attachment and tissue tropism. UPEC strains typically possess a range of adhesins, with type 1 fimbriae and P fimbriae of the chaperone-usher class the best characterised. We previously identified and characterised F9 as a new chaperone-usher fimbrial type that mediates biofilm formation. However, the regulation and specific role of F9 fimbriae remained to be determined in the context of wild-type clinical UPEC strains. In this study we have assessed the distribution and genetic context of the f9 operon among diverse E. coli lineages and pathotypes and demonstrated that f9 genes are significantly more conserved in a UPEC strain collection in comparison to the well-defined E. coli reference (ECOR collection. In the prototypic UPEC strain CFT073, the global regulator protein H-NS was identified as a transcriptional repressor of f9 gene expression at 37°C through its ability to bind directly to the f9 promoter region. F9 fimbriae expression was demonstrated at 20°C, representing the first evidence of functional F9 fimbriae expression by wild-type E. coli. Finally, glycan array analysis demonstrated that F9 fimbriae recognise and bind to terminal Galβ1-3GlcNAc structures.

  8. Authentic display of a cholera toxin epitope by chimeric type 1 fimbriae: effects of insert position and host background

    DEFF Research Database (Denmark)

    Stentebjerg-Olesen, B; Pallesen, L; Jensen, LB

    1997-01-01

    The potential of the major structural protein of type 1 fimbriae as a display system for heterologous sequences was tested. As a reporter-epitope, a heterologous sequence mimicking a neutralizing epitope of the cholera toxin B chain was inserted, in one or two copies, into four different positions...... in the fimA gene. This was carried out by introduction of new restriction sites by PCR-mediated site-directed mutagenesis of fimA in positions predicted to correspond to optimally surface-located regions of the subunit protein. Subsequently, the synthetic cholera-toxin-encoding DNA segment was inserted....... Several of the chosen positions seemed amenable even for large foreign inserts; the chimeric proteins were exposed on the bacterial surface and the cholera toxin epitope was authentically displayed, i.e. it was recognized on bacteria by specific antiserum. Display of chimeric fimbriae was tested...

  9. Erythropoietin improves place learning in fimbria-fornix-transected rats and modifies the search pattern of normal rats

    DEFF Research Database (Denmark)

    Mogensen, Jesper; Miskowiak, Kamilla; Sørensen, Thomas Alrik

    2004-01-01

    administration of EPO significantly improves the posttraumatic functional recovery of the presently studied place learning task after transections of the fimbria-fornix. Additionally, administration of EPO influences the strategy, although not quality, of task solution in normal (sham-operated) rats.......The acquisition of a water-maze-based allocentric place learning task was studied in four groups of rats: two groups subjected to bilateral transections of the fimbria-fornix and two groups undergoing a sham control operation. At the moment of surgery all animals were given one systemic......-associated impairment. The two sham-operated groups did not differ with respect to the proficiency of task acquisition. But administration of EPO to intact animals caused a significant modification of swim patterns-apparently reflecting a somewhat modified strategy of task solution. It is concluded that systemic...

  10. sRNA-Mediated Regulation of P-Fimbriae Phase Variation in Uropathogenic Escherichia coli

    DEFF Research Database (Denmark)

    Khandige, Surabhi; Kronborg, Tina; Uhlin, Bernt Eric

    2015-01-01

    Uropathogenic Escherichia coli (UPEC) are capable of occupying physiologically distinct intracellular and extracellular niches within the urinary tract. This feat requires the timely regulation of gene expression and small RNAs (sRNAs) are known to mediate such rapid adjustments in response to ch...... to changing environmental cues. This study aimed to uncover sRNA-mediated gene regulation in the UPEC strain UTI89, during infection of bladder epithelial cells. Hfq is an RNA chaperone known to facilitate and stabilize sRNA and target mRNA interactions with bacterial cells. The co...... to the discovery of a novel virulence-associated trans-acting sRNA-PapR. Deletion of papR was found to enhance adhesion of UTI89 to both bladder and kidney cell lines in a manner independent of type-1 fimbriae. We demonstrate PapR mediated posttranscriptional repression of the P-fimbriae phase regulator gene pap...

  11. Structural and functional insight into the carbohydrate receptor binding of F4 fimbriae-producing enterotoxigenic Escherichia coli.

    Science.gov (United States)

    Moonens, Kristof; Van den Broeck, Imke; De Kerpel, Maia; Deboeck, Francine; Raymaekers, Hanne; Remaut, Han; De Greve, Henri

    2015-03-27

    Enterotoxigenic Escherichia coli (ETEC) strains are important causes of intestinal disease in humans and lead to severe production losses in animal farming. A range of fimbrial adhesins in ETEC strains determines host and tissue tropism. ETEC strains expressing F4 fimbriae are associated with neonatal and post-weaning diarrhea in piglets. Three naturally occurring variants of F4 fimbriae (F4ab, F4ac, and F4ad) exist that differ in the primary sequence of their major adhesive subunit FaeG, and each features a related yet distinct receptor binding profile. Here the x-ray structure of FaeGad bound to lactose provides the first structural insight into the receptor specificity and mode of binding by the poly-adhesive F4 fimbriae. A small D'-D″-α1-α2 subdomain grafted on the immunoglobulin-like core of FaeG hosts the carbohydrate binding site. Two short amino acid stretches Phe(150)-Glu(152) and Val(166)-Glu(170) of FaeGad bind the terminal galactose in the lactosyl unit and provide affinity and specificity to the interaction. A hemagglutination-based assay with E. coli expressing mutant F4ad fimbriae confirmed the elucidated co-complex structure. Interestingly, the crucial D'-α1 loop that borders the FaeGad binding site adopts a different conformation in the two other FaeG variants and hints at a heterogeneous binding pocket among the FaeG serotypes. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  12. Role of Fimbriae, Flagella and Cellulose on the Attachment of Salmonella Typhimurium ATCC 14028 to Plant Cell Wall Models.

    Directory of Open Access Journals (Sweden)

    Michelle S F Tan

    Full Text Available Cases of foodborne disease caused by Salmonella are frequently associated with the consumption of minimally processed produce. Bacterial cell surface components are known to be important for the attachment of bacterial pathogens to fresh produce. The role of these extracellular structures in Salmonella attachment to plant cell walls has not been investigated in detail. We investigated the role of flagella, fimbriae and cellulose on the attachment of Salmonella Typhimurium ATCC 14028 and a range of isogenic deletion mutants (ΔfliC fljB, ΔbcsA, ΔcsgA, ΔcsgA bcsA and ΔcsgD to bacterial cellulose (BC-based plant cell wall models [BC-Pectin (BCP, BC-Xyloglucan (BCX and BC-Pectin-Xyloglucan (BCPX] after growth at different temperatures (28°C and 37°C. We found that all three cell surface components were produced at 28°C but only the flagella was produced at 37°C. Flagella appeared to be most important for attachment (reduction of up to 1.5 log CFU/cm2 although both cellulose and fimbriae also aided in attachment. The csgD deletion mutant, which lacks both cellulose and fimbriae, showed significantly higher attachment as compared to wild type cells at 37°C. This may be due to the increased expression of flagella-related genes which are also indirectly regulated by the csgD gene. Our study suggests that bacterial attachment to plant cell walls is a complex process involving many factors. Although flagella, cellulose and fimbriae all aid in attachment, these structures are not the only mechanism as no strain was completely defective in its attachment.

  13. CfaE tip mutations in enterotoxigenic Escherichia coli CFA/I fimbriae define critical human intestinal binding sites.

    Science.gov (United States)

    Baker, K K; Levine, M M; Morison, J; Phillips, A; Barry, E M

    2009-05-01

    Enterotoxigenic Escherichia coli (ETEC) use colonization factors to attach to the human intestinal mucosa, followed by enterotoxin expression that induces net secretion and diarrhoeal illness. ETEC strain H10407 expresses CFA/I fimbriae, which are composed of multiple CfaB structural subunits and a CfaE tip subunit. Currently, the contribution of these individual fimbrial subunits in intestinal binding remains incompletely defined. To identify the role of CfaE in attachment in the native ETEC background, an R181A single-amino-acid substitution was introduced by recombination into the H10407 genome. The substitution of R181A eliminated haemagglutination and binding of intestinal mucosa biopsies in in vitro organ culture assays, without loss of CFA/I fimbriae expression. Wild-type in trans plasmid-expressed cfaE restored the binding phenotype. In contrast, in trans expression of cfaE containing amino acid 181 substitutions with similar amino acids, lysine, methionine and glutamine did not restore the binding phenotype, indicating that the loss of the binding phenotype was due to localized areas of epitope disruption. R181 appears to have an irreplaceable role in the formation of a receptor-binding feature on CFA/I fimbriae. The results specifically indicate that the CfaE tip protein is a required binding factor in CFA/I-mediated ETEC colonization, making it a potentially important vaccine antigen. © 2009 Blackwell Publishing Ltd.

  14. Proteus mirabilis uroepithelial cell adhesin (UCA) fimbria plays a role in the colonization of the urinary tract.

    Science.gov (United States)

    Pellegrino, Rafael; Scavone, Paola; Umpiérrez, Ana; Maskell, Duncan J; Zunino, Pablo

    2013-03-01

    Urinary tract infections (UTIs) are among the most common bacterial infections in humans. Proteus mirabilis is an opportunistic pathogen, capable of causing severe UTIs, with serious kidney damage that may even lead to death. Several virulence factors are involved in the pathogenicity of this bacterium. Among these, adherence to the uroepithelium mediated by fimbriae appears to be a significant bacterial attribute related to urovirulence. Proteus mirabilis expresses several types of fimbriae that could be involved in the pathogenesis of UTI, including uroepithelial cell adhesin (UCA). In this report, we used an uropathogenic P. mirabilis wild-type strain and an isogenic ucaA mutant unable to express UCA to study the pathogenic role of this fimbria in UTI. Ability of the mutant to adhere to desquamated uroepithelial cells and to infect mice using different experimental UTI models was significantly impaired. These results allow us to conclude that P. mirabilis UCA plays an important role in the colonization of the urinary tract. © 2013 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  15. Involvement of polyphosphate kinase in virulence and stress tolerance of uropathogenic Proteus mirabilis.

    Science.gov (United States)

    Peng, Liang; Jiang, Qiao; Pan, Jia-Yun; Deng, Cong; Yu, Jing-Yi; Wu, Xiao-Man; Huang, Sheng-He; Deng, Xiao-Yan

    2016-04-01

    Proteus mirabilis (P. mirabilis), a gram-negative enteric bacterium, frequently causes urinary tract infections. Many virulence factors of uropathogenic P. mirabilis have been identified, including urease, flagella, hemolysin and fimbriae. However, the functions of polyphosphate kinase (PPK), which are related to the pathogenicity of many bacteria, remain entirely unknown in P. mirabilis. In this study, a ppk gene encoding the PPK insertional mutant in P. mirabilis strain HI4320 was constructed, and its biological functions were examined. The results of survival studies demonstrated that the ppk mutant was deficient in resistance to oxidative, hyperosmotic and heat stress. The swarming and biofilm formation abilities of P. mirabilis were also attenuated after the ppk interruption. In vitro and in vivo experiments suggested that ppk was required for P. mirabilis to invade the bladder. The negative phenotypes of the ppk mutant could be restored by ppk gene complementation. Furthermore, two-dimensional gel electrophoresis and liquid chromatography-mass spectrometry were used to analyze the proteomes of the wild-type strain and the ppk mutant. Compared with the wild-type strain, seven proteins including TonB-dependent receptor, universal stress protein G, major mannose-resistant/Proteus-like fimbrial protein (MR/P fimbriae), heat shock protein, flagellar capping protein, putative membrane protein and multidrug efflux protein were down-regulated, and four proteins including exported peptidase, repressor protein for FtsI, FKBP-type peptidyl-prolyl cis-trans isomerase and phosphotransferase were up-regulated in the ppk mutant. As a whole, these results indicate that PPK is an important regulator and plays a crucial role in stress tolerance and virulence in uropathogenic P. mirabilis.

  16. Promoting crystallisation of the Salmonella enteritidis fimbriae 14 pilin SefD using deuterium oxide

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Bing; Garnett, James A.; Lee, Wei-chao; Lin, Jing; Salgado, Paula; Taylor, Jonathan; Xu, Yingqi; Lambert, Sebastian; Cota, Ernesto [Centre for Structural Biology and Division of Molecular Biosciences, Imperial College London, South Kensington, London SW7 2AZ (United Kingdom); Matthews, Steve, E-mail: s.j.matthews@imperial.ac.uk [Centre for Structural Biology and Division of Molecular Biosciences, Imperial College London, South Kensington, London SW7 2AZ (United Kingdom)

    2012-05-04

    Highlights: Black-Right-Pointing-Pointer The benefits of D{sub 2}O in screening for crystallisation was explored. Black-Right-Pointing-Pointer The crystal structures of the SefD pilin in both H{sub 2}O and D{sub 2}O reveal differences. Black-Right-Pointing-Pointer Crystallisation improvements are explained by altered interactions in D{sub 2}O crystals. Black-Right-Pointing-Pointer D{sub 2}O is useful additive in sparse-matrix screening for crystallisation. -- Abstract: The use of heavy water (D{sub 2}O) as a solvent is commonplace in many spectroscopic techniques for the study of biological macromolecules. A significant deuterium isotope effect exists where hydrogen-bonding is important, such as in protein stability, dynamics and assembly. Here we illustrate the use of D{sub 2}O in additive screening for the production of reproducible diffraction-quality crystals for the Salmonella enteritidis fimbriae 14 (SEF14) putative tip adhesin, SefD.

  17. Toxic effects of juvenile sablefish, Anoplopoma fimbria by ammonia exposure at different water temperature.

    Science.gov (United States)

    Kim, Jun-Hwan; Park, Hee-Ju; Hwang, In-Ki; Han, Jae-Min; Kim, Do-Hyung; Oh, Chul Woong; Lee, Jung-Sick; Kang, Ju-Chan

    2017-09-01

    Juvenile sablefish, Anoplopoma fimbria (mean length 17.1±2.4cm, and mean weight 75.6±5.7g) were used to evaluate toxic effects on antioxidant systems, immune responses, and stress indicators by ammonia exposure (0, 0.25, 0.75, and 1.25mg/L) at different water temperature (12 and 17°C) in 1 and 2 months. In antioxidant responses, superoxide dismutase (SOD) and catalase (CAT) were significantly increased by ammonia exposure, whereas glutathione (GSH) was decreased. In immune responses, lysozyme and phagocytosis activity were significantly increased by ammonia exposure. In stress indicators, plasma glucose, heat shock protein 70 (HSP 70), and cortisol were significantly increased. At high water temperature (17°C), alterations by ammonia exposure were more distinctly. The results of this study indicated that ammonia exposure can induce toxic effects in the sablefish, and high water temperature can affect the ammonia exposure toxicity. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Alternate thermoregulation and functional binding of Escherichia coli type 1 fimbriae in environmental and animal isolates.

    Science.gov (United States)

    Marshall, Jacqueline; Rossez, Yannick; Mainda, Geoffrey; Gally, David L; Daniell, Tim J; Holden, Nicola J

    2016-11-01

    Type 1 fimbriae (T1F) are well characterised cell surface organelles expressed by Escherichia coli and required for adherence to mannosylated host tissue. They satisfy molecular Koch's postulates as a virulence determinant and a host-adapted role has been reinforced by reports that T1F expression is repressed at submammalian temperatures. Analysis of a group of 136 environmental and animal E. coli isolates that express T1F at 37°C showed that 28% are also capable of expression at 20°C, in a phase variable manner. The heterogeneous proportions varied widely, and although growth temperature impacted the total proportion expressing T1F, there was no direct correlation between growth at 37°C and 20°C, indicative of differences in thermoregulation of the genetic switch (fimS) that controls phase variation. Specificities of the adhesin (FimH) also varied between the isolates: most bound to α-(1-3) mannan and yeast extracts as expected, but some recognised β-(1-4)-mannans and N-linked glycoproteins from plants, and T1F from two of the isolates mediated binding to plant roots. The results expand our view of a well-described adherence factor to show alternative expression profiles and adhesin specificities, which in turn may confer an advantage for certain isolates in alternative hosts and habitats. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  19. A previously uncharacterized gene stm0551 plays a repressive role in the regulation of type 1 fimbriae in Salmonella enterica serotype Typhimurium

    Directory of Open Access Journals (Sweden)

    Wang Ke-Chuan

    2012-06-01

    Full Text Available Abstract Background Salmonella enterica serotype Typhimurium produces surface-associated fimbriae that facilitate adherence of the bacteria to a variety of cells and tissues. Type 1 fimbriae with binding specificity to mannose residues are the most commonly found fimbrial type. In vitro, static-broth culture favors the growth of S. Typhimurium with type 1 fimbriae, whereas non-type 1 fimbriate bacteria are obtained by culture on solid-agar media. Previous studies demonstrated that the phenotypic expression of type 1 fimbriae is the result of the interaction and cooperation of the regulatory genes fimZ, fimY, fimW, and fimU within the fim gene cluster. Genome sequencing revealed a novel gene, stm0551, located between fimY and fimW that encodes an 11.4-kDa putative phosphodiesterase specific for the bacterial second messenger cyclic-diguanylate monophosphate (c-di-GMP. The role of stm0551 in the regulation of type 1 fimbriae in S. Typhimurium remains unclear. Results A stm0551-deleted stain constructed by allelic exchange constitutively produced type 1 fimbriae in both static-broth and solid-agar medium conditions. Quantative RT-PCR revealed that expression of the fimbrial major subunit gene, fimA, and one of the regulatory genes, fimZ, were comparably increased in the stm0551-deleted strain compared with those of the parental strain when grown on the solid-agar medium, a condition that normally inhibits expression of type 1 fimbriae. Following transformation with a plasmid possessing the coding sequence of stm0551, expression of fimA and fimZ decreased in the stm0551 mutant strain in both culture conditions, whereas transformation with the control vector pACYC184 relieved this repression. A purified STM0551 protein exhibited a phosphodiesterase activity in vitro while a point mutation in the putative EAL domain, substituting glutamic acid (E with alanine (A, of STM0551 or a FimY protein abolished this activity. Conclusions The finding that the

  20. Mfa4, an Accessory Protein of Mfa1 Fimbriae, Modulates Fimbrial Biogenesis, Cell Auto-Aggregation, and Biofilm Formation in Porphyromonas gingivalis.

    Science.gov (United States)

    Ikai, Ryota; Hasegawa, Yoshiaki; Izumigawa, Masashi; Nagano, Keiji; Yoshida, Yasuo; Kitai, Noriyuki; Lamont, Richard J; Yoshimura, Fuminobu; Murakami, Yukitaka

    2015-01-01

    Porphyromonas gingivalis, a gram-negative obligate anaerobic bacterium, is considered to be a key pathogen in periodontal disease. The bacterium expresses Mfa1 fimbriae, which are composed of polymers of Mfa1. The minor accessory components Mfa3, Mfa4, and Mfa5 are incorporated into these fimbriae. In this study, we characterized Mfa4 using genetically modified strains. Deficiency in the mfa4 gene decreased, but did not eliminate, expression of Mfa1 fimbriae. However, Mfa3 and Mfa5 were not incorporated because of defects in posttranslational processing and leakage into the culture supernatant, respectively. Furthermore, the mfa4-deficient mutant had an increased tendency to auto-aggregate and form biofilms, reminiscent of a mutant completely lacking Mfa1. Notably, complementation of mfa4 restored expression of structurally intact and functional Mfa1 fimbriae. Taken together, these results indicate that the accessory proteins Mfa3, Mfa4, and Mfa5 are necessary for assembly of Mfa1 fimbriae and regulation of auto-aggregation and biofilm formation of P. gingivalis. In addition, we found that Mfa3 and Mfa4 are processed to maturity by the same RgpA/B protease that processes Mfa1 subunits prior to polymerization.

  1. Diversification of the Salmonella fimbriae: a model of macro- and microevolution.

    Directory of Open Access Journals (Sweden)

    Min Yue

    Full Text Available Bacteria of the genus Salmonella comprise a large and evolutionary related population of zoonotic pathogens that can infect mammals, including humans and domestic animals, birds, reptiles and amphibians. Salmonella carries a plethora of virulence genes, including fimbrial adhesins, some of them known to participate in mammalian or avian host colonization. Each type of fimbria has its structural subunit and biogenesis genes encoded by one fimbrial gene cluster (FGC. The accumulation of new genomic information offered a timely opportunity to better evaluate the number and types of FGCs in the Salmonella pangenome, to test the use of current classifications based on phylogeny, and to infer potential correlations between FGC evolution in various Salmonella serovars and host niches. This study focused on the FGCs of the currently deciphered 90 genomes and 60 plasmids of Salmonella. The analysis highlighted a fimbriome consisting of 35 different FGCs, of which 16 were new, each strain carrying between 5 and 14 FGCs. The Salmonella fimbriome was extremely diverse with FGC representatives in 8 out of 9 previously categorized fimbrial clades and subclades. Phylogenetic analysis of Salmonella suggested macroevolutionary shifts detectable by extensive FGC deletion and acquisition. In addition, microevolutionary drifts were best depicted by the high level of allelic variation in predicted or known adhesins, such as the type 1 fimbrial adhesin FimH for which 67 different natural alleles were identified in S. enterica subsp. I. Together with strain-specific collections of FGCs, allelic variation among adhesins attested to the pathoadaptive evolution of Salmonella towards specific hosts and tissues, potentially modulating host range, strain virulence, disease progression, and transmission efficiency. Further understanding of how each Salmonella strain utilizes its panel of FGCs and specific adhesin alleles for survival and infection will support the

  2. Diversification of the Salmonella Fimbriae: A Model of Macro- and Microevolution

    Science.gov (United States)

    Yue, Min; Rankin, Shelley C.; Blanchet, Ryan T.; Nulton, James D.; Edwards, Robert A.; Schifferli, Dieter M.

    2012-01-01

    Bacteria of the genus Salmonella comprise a large and evolutionary related population of zoonotic pathogens that can infect mammals, including humans and domestic animals, birds, reptiles and amphibians. Salmonella carries a plethora of virulence genes, including fimbrial adhesins, some of them known to participate in mammalian or avian host colonization. Each type of fimbria has its structural subunit and biogenesis genes encoded by one fimbrial gene cluster (FGC). The accumulation of new genomic information offered a timely opportunity to better evaluate the number and types of FGCs in the Salmonella pangenome, to test the use of current classifications based on phylogeny, and to infer potential correlations between FGC evolution in various Salmonella serovars and host niches. This study focused on the FGCs of the currently deciphered 90 genomes and 60 plasmids of Salmonella. The analysis highlighted a fimbriome consisting of 35 different FGCs, of which 16 were new, each strain carrying between 5 and 14 FGCs. The Salmonella fimbriome was extremely diverse with FGC representatives in 8 out of 9 previously categorized fimbrial clades and subclades. Phylogenetic analysis of Salmonella suggested macroevolutionary shifts detectable by extensive FGC deletion and acquisition. In addition, microevolutionary drifts were best depicted by the high level of allelic variation in predicted or known adhesins, such as the type 1 fimbrial adhesin FimH for which 67 different natural alleles were identified in S. enterica subsp. I. Together with strain-specific collections of FGCs, allelic variation among adhesins attested to the pathoadaptive evolution of Salmonella towards specific hosts and tissues, potentially modulating host range, strain virulence, disease progression, and transmission efficiency. Further understanding of how each Salmonella strain utilizes its panel of FGCs and specific adhesin alleles for survival and infection will support the development of new approaches

  3. Structural insight in the inhibition of adherence of F4 fimbriae producing enterotoxigenic Escherichia coli by llama single domain antibodies.

    Science.gov (United States)

    Moonens, Kristof; Van den Broeck, Imke; Okello, Emmanuel; Pardon, Els; De Kerpel, Maia; Remaut, Han; De Greve, Henri

    2015-02-24

    Enterotoxigenic Escherichia coli that cause neonatal and post-weaning diarrhea in piglets express F4 fimbriae to mediate attachment towards host receptors. Recently we described how llama single domain antibodies (VHHs) fused to IgA, produced in Arabidopsis thaliana seeds and fed to piglets resulted in a progressive decline in shedding of F4 positive ETEC bacteria. Here we present the structures of these inhibiting VHHs in complex with the major adhesive subunit FaeG. A conserved surface, distant from the lactose binding pocket, is targeted by these VHHs, highlighting the possibility of targeting epitopes on single-domain adhesins that are non-involved in receptor binding.

  4. Asymptomatic bacteriuria Escherichia coli strain 83972 carries mutations in the foc locus and is unable to express F1C fimbriae

    DEFF Research Database (Denmark)

    Hancock, Viktoria; Schembri, M.A.; Ulett, G.C.

    2006-01-01

    Escherichia coli is the most common organism associated with asymptomatic bacteriuria (ABU). In contrast to uropathogenic E. coli (UPEC), which causes symptomatic urinary tract infection (UTI), very little is known about the mechanisms by which these strains colonize the urinary tract. Bacterial...... status of one of the primary adhesion factors known to be associated with UTI, namely F1C fimbriae, encoded by the foc gene cluster. F1C fimbriae recognize receptors present in the human kidney and bladder. Expression of the foc genes was found to be up-regulated in human urine. It was also shown...

  5. Salmonella enterica serotype Typhimurium Std fimbriae bind terminal α (1,2)fucose residues in the cecal mucosa

    Science.gov (United States)

    Chessa, Daniela; Winter, Maria G.; Jakomin, Marcello; Bäumler, Andreas J.

    2013-01-01

    SUMMARY The std operon encodes a fimbrial adhesin of Salmonella enterica serotype Typhimurium that is required for attachment to intestinal epithelial cells and for cecal colonization in the mouse. To study the mechanism by which this virulence factor contributes to colonization we characterized its binding specificity. Std-mediated binding to human colonic epithelial (Caco-2) cells could be abrogated by removing N-linked glycans. Adherence of Std fimbriated S. Typhimurium to Caco-2 cells could be blocked by co-incubation with H type 2 oligosaccharide (Fucα1-2Galβ1-4GlcNAc) or by pretreatment of cells with α1-2 fucosidase. In contrast, pretreatment of Caco-2 cells with neuraminidase or co-incubation with the type 2 disaccharide precursor (Galβ1-4GlcNAc) did not reduce adherence of Std fimbriated S. Typhimurium. Binding of purified Std fimbriae to Fucα1-2Galβ1-4GlcNAc in a solid phase binding assay was competitively inhibited by Ulex europaeus agglutinin-I (UEA-I), a lectin specific for Fucα1-2 moieties. Purified Std fimbriae and UEA both bound to a receptor localized in the mucus layer of the murine cecum. These data suggest that the std operon encodes an adhesin that binds an α1-2 fucosylated receptor(s) present in the cecal mucosa. PMID:19183274

  6. Synergy between type 1 fimbriae expression and C3 opsonisation increases internalisation of E. coli by human tubular epithelial cells.

    Science.gov (United States)

    Li, Ke; Zhou, Wuding; Hong, Yuzhi; Sacks, Steven H; Sheerin, Neil S

    2009-03-31

    Bacterial infection of the urinary tract is a common clinical problem with E. coli being the most common urinary pathogen. Bacterial uptake into epithelial cells is increasingly recognised as an important feature of infection. Bacterial virulence factors, especially fimbrial adhesins, have been conclusively shown to promote host cell invasion. Our recent study reported that C3 opsonisation markedly increases the ability of E. coli strain J96 to internalise into human proximal tubular epithelial cells via CD46, a complement regulatory protein expressed on host cell membrane. In this study, we further assessed whether C3-dependent internalisation by human tubular epithelial cells is a general feature of uropathogenic E. coli and investigated features of the bacterial phenotype that may account for any heterogeneity. In 31 clinical isolates of E. coli tested, C3-dependent internalisation was evident in 10 isolates. Type 1 fimbriae mediated-binding is essential for C3-dependent internalisation as shown by phenotypic association, type 1 fimbrial blockade with soluble ligand (mannose) and by assessment of a type 1 fimbrial mutant. we propose that efficient internalisation of uropathogenic E. coli by the human urinary tract depends on co-operation between type 1 fimbriae-mediated adhesion and C3 receptor -ligand interaction.

  7. Two novel genes, fanA and fanB, involved in the biogenesis of K99 fimbriae.

    Science.gov (United States)

    Roosendaal, E; Boots, M; de Graaf, F K

    1987-08-11

    The nucleotide sequence of the region located transcriptionally upstream of the K99 fimbrial subunit gene (fanC) was determined. Several putative transcription signals and two open reading frames, designated fanA and fanB, became apparent. Frameshift mutations in fanA and fanB reduced K99 fimbriae expression 8-fold and 16-fold, respectively. Complementation of the mutants in trans restored the K99 expression to about 75% of the wild type level, indicating that fanA and fanB code for transacting polypeptides involved in the biogenesis of K99 fimbriae. The fanA and fanB gene products FanA and FanB were not detectable in minicell preparations, indicating that both polypeptides are synthesized in very small amounts. However, in an in vitro DNA directed translation system FanA and FanB could be identified. The deduced amino acid sequences of FanA and FanB showed that both polypeptides contain no signal peptides, indicating a cytoplasmic location. Furthermore, the polypeptides are very hydrophilic, mainly basic, and exhibit remarkable homology to each other and to a regulatory protein (papB) encoded by the pap-operon (1). Some of these features are characteristics of nucleic acid binding proteins, which suggests that FanA and FanB have a regulatory function in the synthesis of FanC and the auxiliary polypeptides FanD-H.

  8. Escherichia coli F4 fimbriae specific lama single-domain antibody fragments effectively inhibit bacterial adhesion in vitro but poorly protect against diarrhea

    NARCIS (Netherlands)

    Harmsen, M.M.; Solt, van C.B.; Hoogendoorn, A.; Zijderveld, van F.G.; Niewold, T.A.; Meulen, van der J.

    2005-01-01

    Oral administration of polyclonal antibodies directed against enterotoxigenic Escherichia coli (ETEC) F4 fimbriae is used to protect against piglet post-weaning diarrhoea. For cost reasons, we aim to replace these polyclonal antibodies by recombinant llama single-domain antibody fragments (VHHs)

  9. Development and Efficacy Assessment of an Enteric Coated Porous Tablet Loaded With F4 Fimbriae for Oral Vaccination of Piglets against F4+ Escherichia coli Infections.

    Science.gov (United States)

    Srivastava, Atul; Gowda, D V; Madhunapantula, SubbaRao V; Siddaramaiah

    2016-01-01

    Enterotoxigenic Escherichia coli (ETEC) infection is one of the major causes contributing to the development of diarrhoea and mortality in new born, suckling and newly weaned piglets. To date, no preventive/treatment strategy showed promising results, which could be due to the lack of potent vaccines, and/or due to the development of resistance of ETEC to antibiotics. Therefore, in the present investigation, a novel porous sodium alginate (SA) tablet formulation loaded with F4 fimbriae antigen was developed and tested for efficacy against ETEC infections in piglet models. Precompression parameters of the powder mixes and post compression parameters of tablets have been evaluated and results were found to be satisfactory. Loading of F4 fimbrial antigens into the tablets was achieved by inducing pores in the tablets via the sublimation of camphor followed by incubation with purified F4 fimbriae. The loaded tablets have been coated with Eudragit L100 to protect the F4 fimbriae from (a) highly acidic gastric environment; (b) proteolytic cleavage by pepsin; and (c) to promote subsequent release in the intestine. Evaluation of developed F4 fimbrial tablets in a Pig model demonstrated induction of mucosal immunity, and a significant reduction of F4+ E. coli in faeces. Therefore, F4 fimbriae loaded porous tablets could be a novel oral vaccination candidate to induce mucosal and systemic immunity against ETEC infections.

  10. Both flagella and F4 fimbriae from F4ac+ enterotoxigenic Escherichia coli contribute to attachment to IPEC-J2 cells in vitro.

    Science.gov (United States)

    Zhou, Mingxu; Duan, Qiangde; Zhu, Xiaofang; Guo, Zhiyan; Li, Yinchau; Hardwidge, Philip R; Zhu, Guoqiang

    2013-05-13

    The role of flagella in the pathogenesis of F4ac+ Enterotoxigenic Escherichia coli (ETEC) mediated neonatal and post-weaning diarrhea (PWD) is not currently understood. We targeted the reference C83902 ETEC strain (O8:H19:F4ac+ LT+ STa+ STb+), to construct isogenic mutants in the fliC (encoding the major flagellin protein), motA (encoding the flagella motor), and faeG (encoding the major subunit of F4 fimbriae) genes. Both the ΔfliC and ΔfaeG mutants had a reduced ability to adhere to porcine intestinal epithelial IPEC-J2 cells. F4 fimbriae expression was significantly down-regulated after deleting fliC, which revealed that co-regulation exists between flagella and F4 fimbriae. However, there was no difference in adhesion between the ΔmotA mutant and its parent strain. These data demonstrate that both flagella and F4 fimbriae are required for efficient F4ac+ ETEC adhesion in vitro.

  11. Genes involucrados en la biogénesis de fimbrias afectan la formación de biopelículas por parte de Klebsiella pneumoniae

    Directory of Open Access Journals (Sweden)

    Ana Victoria Suescún

    2006-12-01

    Conclusión. Los resultados indicaron que las fimbrias tipo 1 y 3, ésta última ya implicada en este fenómeno en K. pneumoniae, son factores importantes para la adhesión y la formación de agregados multicelulares.

  12. Type 3 fimbriae, encoded by the conjugative plasmid pOLA52, enhance biofilm formation and transfer frequencies in Enterobacteriaceae strains

    DEFF Research Database (Denmark)

    Burmølle, Mette; Bahl, Martin Iain; Jensen, Lars Bogø

    2008-01-01

    pathogenic, members of the family Enterobacteriaceae, including Klebsiella pneumoniae, Salmonella Typhimurium, Kluyvera sp. and Enterobacter aerogenes, pOLA52 facilitated increased biofilm formation. pOLA52 is believed to represent the first example of a conjugative plasmid encoding type 3 fimbriae...

  13. Preliminary X-ray diffraction analysis of CfaA, a molecular chaperone essential for the assembly of CFA/I fimbriae of human enterotoxigenic Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Bao, Rui; Esser, Lothar; Poole, Steven; McVeigh, Annette; Chen, Yu-xing; Savarino, Stephen J.; Xia, Di, E-mail: xiad@mail.nih.gov

    2014-01-21

    The molecular chaperone CfaA plays a critical role in the bioassembly of the surface-adhesive CFA/I fimbriae of enterotoxigenic E. coli. Purified CfaA was crystallized and the phase solution was determined by the multiple isomorphous replacement coupled with anomalous scattering method.

  14. Growth data - Stable isotope analysis as a tool to determine the metabolic fates of dietary carbohydrates from plant-based alternative feed ingredients in the carnivorous sablefish, Anoplopoma fimbria

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Development of specialized feeds for carnivorous species such as sablefish (Anoplopoma fimbria) in which fishmeal and oil from marine sources are replaced by more...

  15. Fish culture data - Stable isotope analysis as a tool to determine the metabolic fates of dietary carbohydrates from plant-based alternative feed ingredients in the carnivorous sablefish, Anoplopoma fimbria

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Development of specialized feeds for carnivorous species such as sablefish (Anoplopoma fimbria) in which fishmeal and oil from marine sources are replaced by more...

  16. Chemical composition of fish and diets - Stable isotope analysis as a tool to determine the metabolic fates of dietary carbohydrates from plant-based alternative feed ingredients in the carnivorous sablefish, Anoplopoma fimbria

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Development of specialized feeds for carnivorous species such as sablefish (Anoplopoma fimbria) in which fishmeal and oil from marine sources are replaced by more...

  17. Altered Regulation of the Diguanylate Cyclase YaiC Reduces Production of Type 1 Fimbriae in a Pst Mutant of Uropathogenic Escherichia coli CFT073.

    Science.gov (United States)

    Crépin, Sébastien; Porcheron, Gaëlle; Houle, Sébastien; Harel, Josée; Dozois, Charles M

    2017-12-15

    The pst gene cluster encodes the phosphate-specific transport (Pst) system. Inactivation of the Pst system constitutively activates the two-component regulatory system PhoBR and attenuates the virulence of pathogenic bacteria. In uropathogenic Escherichia coli strain CFT073, attenuation by inactivation of pst is predominantly attributed to the decreased expression of type 1 fimbriae. However, the molecular mechanisms connecting the Pst system and type 1 fimbriae are unknown. To address this, a transposon library was constructed in the pst mutant, and clones were tested for a regain in type 1 fimbrial production. Among them, the diguanylate cyclase encoded by yaiC ( adrA in Salmonella ) was identified to connect the Pst system and type 1 fimbrial expression. In the pst mutant, the decreased expression of type 1 fimbriae is connected by the induction of yaiC This is predominantly due to altered expression of the FimBE-like recombinase genes ipuA and ipbA , affecting at the same time the inversion of the fim promoter switch ( fimS ). In the pst mutant, inactivation of yaiC restored fim -dependent adhesion to bladder cells and virulence. Interestingly, the expression of yaiC was activated by PhoB, since transcription of yaiC was linked to the PhoB-dependent phoA-psiF operon. As YaiC is involved in cyclic di-GMP (c-di-GMP) biosynthesis, an increased accumulation of c-di-GMP was observed in the pst mutant. Hence, the results suggest that one mechanism by which deletion of the Pst system reduces the expression of type 1 fimbriae is through PhoBR-mediated activation of yaiC , which in turn increases the accumulation of c-di-GMP, represses the fim operon, and, consequently, attenuates virulence in the mouse urinary tract infection model. IMPORTANCE Urinary tract infections (UTIs) are common bacterial infections in humans. They are mainly caused by uropathogenic Escherichia coli (UPEC). We previously showed that interference with phosphate homeostasis decreases the

  18. Flagellin and F4 fimbriae have opposite effects on biofilm formation and quorum sensing in F4ac+ enterotoxigenic Escherichia coli.

    Science.gov (United States)

    Zhou, Mingxu; Guo, Zhiyan; Yang, Yang; Duan, Qiangde; Zhang, Qi; Yao, Fenghua; Zhu, Jun; Zhang, Xinjun; Hardwidge, Philip R; Zhu, Guoqiang

    2014-01-10

    Bacteria that form biofilms are often highly resistant to antibiotics and are capable of evading the host immune system. To evaluate the role of flagellin and F4 fimbriae on biofilm formation by enterotoxigenic Escherichia coli (ETEC), we deleted the fliC (encoding the major flagellin protein) and/or the faeG (encoding the major subunit of F4 fimbriae) genes from ETEC C83902. Biofilm formation was reduced in the fliC mutant but increased in the faeG mutant, as compared with the wild-type strain. The expression of AI-2 quorum sensing associated genes was regulated in the fliC and faeG mutants, consistent with the biofilm formation of these strains. But, deleting fliC and/or faeG also inhibited AI-2 quorum sensing activity. Copyright © 2013 Elsevier B.V. All rights reserved.

  19. Transcytosis of F4 fimbriae by villous and dome epithelia in F4-receptor positive pigs supports importance of receptor-dependent endocytosis in oral immunization strategies.

    Science.gov (United States)

    Snoeck, Veerle; Van den Broeck, Wim; De Colvenaer, Veerle; Verdonck, Frank; Goddeeris, Bruno; Cox, Eric

    2008-07-15

    Very few antigens have been described that induce an intestinal immunity when given orally. Our laboratory demonstrated that oral administration of isolated F4 (K88) fimbriae of Escherichia coli to F4-receptor positive (F4R(+)) pigs induces protective mucosal immunity against challenge infection. However, presence of F4-receptors (F4R) on villous enterocytes is a prerequisite for inducing the immune response, as no F4-specific antibody-secreting cells (ASC) can be induced in F4R(-) pigs. In this study, the in vivo binding of isolated F4 fimbriae (F4) to the gut epithelium was examined in F4R(+) and F4R(-) pigs. It was further investigated whether binding of F4 to the F4R results in endocytosis in and translocation across the gut epithelium using microscopy. F4 did not adhere to the intestinal epithelium of F4R(-) pigs, whereas it strongly adhered to the villous epithelium and the follicle-associated epithelium (FAE) of the jejunum and ileum of F4R(+) pigs. Following binding to F4R, F4 was endocytosed by villous enterocytes, follicle-associated enterocytes and M cells. Transcytosis of F4 across the epithelium resulted in the appearance of F4 in the lamina propria and dome region of the jejunal and ileal PP. This is the first study showing transcytosis of fimbriae across the gut epithelium. This receptor-dependent transcytosis can explain the success of F4 fimbriae as oral immunogen for inducing protective immunity in F4R(+) pigs strengthening the importance of receptor-dependent endocytosis and translocation in oral vaccine strategies. Further identification of the receptor responsible for this transport is in progress.

  20. The plasmid-encoded Ipf and Klf fimbriae display different expression and varying roles in the virulence of Salmonella enterica serovar Infantis in mouse vs. avian hosts.

    Directory of Open Access Journals (Sweden)

    Gili Aviv

    2017-08-01

    Full Text Available Salmonella enterica serovar Infantis is one of the prevalent Salmonella serovars worldwide. Different emergent clones of S. Infantis were shown to acquire the pESI virulence-resistance megaplasmid affecting its ecology and pathogenicity. Here, we studied two previously uncharacterized pESI-encoded chaperone-usher fimbriae, named Ipf and Klf. While Ipf homologs are rare and were found only in S. enterica subspecies diarizonae and subspecies VII, Klf is related to the known K88-Fae fimbria and klf clusters were identified in seven S. enterica subspecies I serovars, harboring interchanging alleles of the fimbria major subunit, KlfG. Regulation studies showed that the klf genes expression is negatively and positively controlled by the pESI-encoded regulators KlfL and KlfB, respectively, and are activated by the ancestral leucine-responsive regulator (Lrp. ipf genes are negatively regulated by Fur and activated by OmpR. Furthermore, induced expression of both klf and ipf clusters occurs under microaerobic conditions and at 41°C compared to 37°C, in-vitro. Consistent with these results, we demonstrate higher expression of ipf and klf in chicks compared to mice, characterized by physiological temperature of 41.2°C and 37°C, respectively. Interestingly, while Klf was dispensable for S. Infantis colonization in the mouse, Ipf was required for maximal colonization in the murine ileum. In contrast to these phenotypes in mice, both Klf and Ipf contributed to a restrained infection in chicks, where the absence of these fimbriae has led to moderately higher bacterial burden in the avian host. Taken together, these data suggest that physiological differences between host species, such as the body temperature, can confer differences in fimbriome expression, affecting Salmonella colonization and other host-pathogen interplays.

  1. Influence of type-I fimbriae and fluid shear stress on bacterial behavior and multicellular architecture of early Escherichia coli biofilms at single-cell resolution.

    Science.gov (United States)

    Wang, Liyun; Keatch, Robert; Zhao, Qi; Wright, John A; Bryant, Clare E; Redmann, Anna L; Terentjev, Eugene M

    2018-01-12

    Biofilm formation on abiotic surfaces in food and medical industry can cause severe contamination and infection, yet how biological and physical factors determine cellular architecture of early biofilms and bacterial behavior of the constituent cells remains largely unknown. In this study we examine the specific role of type-I fimbriae in nascent stages of biofilm formation and the response of micro-colonies to environmental flow shear at single-cell resolution. The results show that type-I fimbriae are not required for reversible adhesion from plankton, but critical for irreversible adhesion of Escherichia coli ( E.coli ) MG1655 forming biofilms on polyethylene terephthalate (PET) surfaces. Besides establishing a firm cell-surface contact, the irreversible adhesion seems necessary to initiate the proliferation of E.coli on the surface. After application of shear stress, bacterial retention is dominated by the 3D architecture of colonies independent of the population and the multi-layered structure could protect the embedded cells from being insulted by fluid shear, while cell membrane permeability mainly depends on the biofilm population and the duration time of the shear stress. Importance Bacterial biofilms could lead to severe contamination problems in medical devices and food processing equipment. However, biofilms are usually studied at a rough macroscopic level, thus little is known about how individual bacterial behavior within biofilms and multicellular architecture are influenced by bacterial appendages (e.g. pili/fimbriae) and environmental factors during early biofilm formation. We apply Confocal Laser Scanning Microscopy (CLSM) to visualize E.coli micro-colonies at single-cell resolution. Our findings suggest that type-I fimbriae are vital to the initiation of bacterial proliferation on surfaces and that the responses of biofilm architecture and cell membrane permeability of constituent bacteria to fluid shear stress are different, which are

  2. Targeting of Escherichia coli F4 fimbriae to Fcgamma receptors enhances the maturation of porcine dendritic cells.

    Science.gov (United States)

    Devriendt, Bert; Verdonck, Frank; Summerfield, Artur; Goddeeris, Bruno M; Cox, Eric

    2010-06-15

    F4(+) enterotoxigenic Escherichia coli (ETEC) infections are an important cause of postweaning diarrhoea in piglets and an oral immunization of piglets with purified F4 fimbriae protects them from a subsequent F4(+) ETEC infection. However, oral immunization of suckling piglets is hampered due to the immature status of their immune system. Targeting of antigens to Fcgamma receptors (FcgammaR) on human and murine dendritic cells (DC) has been shown to enhance DC maturation and both humoral and cellular immune responses. To investigate the effect of F4 fimbriae incorporated in immune complexes (F4-IC) on porcine DC, we used porcine monocytic-derived DC (MoDC) as a model system. The results in this study demonstrate that FcgammaRI, II and III mRNA is expressed by porcine MoDC. Furthermore, we show that FcgammaRII and III are expressed on the cell surface and that F4-IC are internalized by MoDC via FcgammaR. This FcgammaR ligation induced a significantly enhanced expression of Major Histocompatibility complex (MHCII) class II and the costimulatory molecules CD80/86 and CD40 by MoDC compared with immature MoDC. Furthermore, the phagocytic capacity of F4-IC stimulated MoDC was reduced as evidenced by a reduced uptake of DQ-ovalbumin and FITC-dextran. In an allogenic and autologous mixed lymphocyte reaction, these F4-IC-activated MoDC showed an improved T cell stimulatory capacity in comparison with immature MoDC. The F4-IC induced DC maturation correlated with significant higher expression levels of several pro-inflammatory cytokines such as interleukine (IL) 1beta, IL-6 and Tumor necrosis factor alpha, the chemokine IL-8 and IL-12p40 in comparison with immature MoDC. Altogether, these results clearly demonstrate that FcgammaR engagement enhances the maturation of porcine MoDC, which may suggest that antigen targeting to FcgammaR on DC could improve vaccine design against infections. Copyright 2009 Elsevier B.V. All rights reserved.

  3. 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. Published by Elsevier Ltd.

  4. Type 3 fimbriae and biofilm formation are regulated by the transcriptional regulators MrkHI in Klebsiella pneumoniae.

    Science.gov (United States)

    Johnson, Jeremiah G; Murphy, Caitlin N; Sippy, Jean; Johnson, Tylor J; Clegg, Steven

    2011-07-01

    Klebsiella pneumoniae is an opportunistic pathogen which frequently causes hospital-acquired urinary and respiratory tract infections. K. pneumoniae may establish these infections in vivo following adherence, using the type 3 fimbriae, to indwelling devices coated with extracellular matrix components. Using a colony immunoblot screen, we identified transposon insertion mutants which were deficient for type 3 fimbrial surface production. One of these mutants possessed a transposon insertion within a gene, designated mrkI, encoding a putative transcriptional regulator. A site-directed mutant of this gene was constructed and shown to be deficient for fimbrial surface expression under aerobic conditions. MrkI mutants have a significantly decreased ability to form biofilms on both abiotic and extracellular matrix-coated surfaces. This gene was found to be cotranscribed with a gene predicted to encode a PilZ domain-containing protein, designated MrkH. This protein was found to bind cyclic-di-GMP (c-di-GMP) and regulate type 3 fimbrial expression.

  5. Type 3 Fimbriae and Biofilm Formation Are Regulated by the Transcriptional Regulators MrkHI in Klebsiella pneumoniae▿

    Science.gov (United States)

    Johnson, Jeremiah G.; Murphy, Caitlin N.; Sippy, Jean; Johnson, Tylor J.; Clegg, Steven

    2011-01-01

    Klebsiella pneumoniae is an opportunistic pathogen which frequently causes hospital-acquired urinary and respiratory tract infections. K. pneumoniae may establish these infections in vivo following adherence, using the type 3 fimbriae, to indwelling devices coated with extracellular matrix components. Using a colony immunoblot screen, we identified transposon insertion mutants which were deficient for type 3 fimbrial surface production. One of these mutants possessed a transposon insertion within a gene, designated mrkI, encoding a putative transcriptional regulator. A site-directed mutant of this gene was constructed and shown to be deficient for fimbrial surface expression under aerobic conditions. MrkI mutants have a significantly decreased ability to form biofilms on both abiotic and extracellular matrix-coated surfaces. This gene was found to be cotranscribed with a gene predicted to encode a PilZ domain-containing protein, designated MrkH. This protein was found to bind cyclic-di-GMP (c-di-GMP) and regulate type 3 fimbrial expression. PMID:21571997

  6. Preliminary X-ray diffraction analysis of CfaA, a molecular chaperone essential for the assembly of CFA/I fimbriae of human enterotoxigenic Escherichia coli.

    Science.gov (United States)

    Bao, Rui; Esser, Lothar; Poole, Steven; McVeigh, Annette; Chen, Yu Xing; Savarino, Stephen J; Xia, Di

    2014-02-01

    Understanding of pilus bioassembly in Gram-negative bacteria stems mainly from studies of P pili and type 1 fimbriae of uropathogenic Escherichia coli, which are mediated by the classic chaperone-usher pathway (CUP). However, CFA/I fimbriae, a class 5 fimbria and intestinal colonization factor for enterotoxigenic E. coli (ETEC), are proposed to assemble via the alternate chaperone pathway (ACP). Both CUP and ACP fimbrial bioassembly pathways require the function of a periplasmic chaperone, but their corresponding proteins share very low similarity in primary sequence. Here, the crystallization of the CFA/I periplasmic chaperone CfaA by the hanging-drop vapor-diffusion method is reported. X-ray diffraction data sets were collected from a native CfaA crystal to 2 Å resolution and to 1.8 and 2.8 Å resolution, respectively, from a lead and a platinum derivative. These crystals displayed the symmetry of space group C2, with unit-cell parameters a = 103.6, b = 28.68, c = 90.60 Å, β = 119.7°. Initial phases were derived from multiple isomorphous replacement with anomalous scattering experiments using the data from the platinum and lead derivatives. This resulted in an interpretable electron-density map showing one CfaA molecule in an asymmetric unit. Sequence assignments were aided by anomalous signals from the heavy-atom derivatives. Refinement of the atomic model of CfaA is ongoing, which is expected to further understanding of the essential aspects and allowable variations in tertiary structure of the greater family of chaperones involved in chaperone-usher mediated bioassembly.

  7. Crystallization and preliminary X-ray diffraction analyses of several forms of the CfaB major subunit of enterotoxigenic Escherichia coli CFA/I fimbriae

    International Nuclear Information System (INIS)

    Li, Yong-Fu; Poole, Steven; Rasulova, Fatima; McVeigh, Annette L.; Savarino, Stephen J.; Xia, Di

    2009-01-01

    Three fusion proteins were generated in order to resolve the atomic structure of the CFA/I fimbriae of enterotoxigenic E. coli. CfaEB is a fusion of the minor and major CFA/I subunits, while CfaBB and CfaBBB are tandem fusions of two and three repeats, respectively, of the major subunit. Each protein was crystallized and the crystal structures of each of these fusions were determined successively by the molecular-replacement method using the CfaE crystal structure as an initial phasing model. Enterotoxigenic Escherichia coli (ETEC), a major global cause of diarrhea, initiates the pathogenic process via fimbriae-mediated attachment to the small intestinal epithelium. A common prototypic ETEC fimbria, colonization factor antigen I (CFA/I), consists of a tip-localized minor adhesive subunit CfaE and the stalk-forming major subunit CfaB, both of which are necessary for fimbrial assembly. To elucidate the structure of CFA/I at atomic resolution, three recombinant proteins were generated consisting of fusions of the minor and major subunits (CfaEB) and of two (CfaBB) and three (CfaBBB) repeats of the major subunit. Crystals of CfaEB diffracted X-rays to 2.1 Å resolution and displayed the symmetry of space group P2 1 . CfaBB exhibited a crystal diffraction limit of 2.3 Å resolution and had the symmetry of space group P2 1 2 1 2. CfaBBB crystallized in the monoclinic space group C2 and diffracted X-rays to 2.3 Å resolution. These structures were determined using the molecular-replacement method

  8. Induction of Th1 polarized immune responses by thiolated Eudragit-coated F4 and F18 fimbriae of enterotoxigenic Escherichia coli.

    Science.gov (United States)

    Lee, Won-Jung; Cha, Seungbin; Shin, Minkyoung; Islam, Mohammad Ariful; Cho, Chong-su; Yoo, Han Sang

    2011-10-01

    Diarrhea in newborn and weaned piglets is mainly induced by enterotoxigenic Escherichia coli (ETEC) with fimbriae F4 (K88) and F18 (F107). In this study, we evaluated F4 and F18 coated with thiolated Eudragit microspheres (TEMS) as a candidate for an oral vaccine. The average particle sizes of TEMS, F4-loaded TEMS, and F18-loaded TEMS were measured as 4.2±0.75 μm, 4.7±0.50 μm, and 4.5±0.37 μm, respectively. F4 is more efficiently encapsulated than F18 in the loading with TEMS. In the release test, F4 and F18 fimbriae were protected in acidic circumstances, whereas most were released at pH 7.4 of intestine circumstances. Production of TNF-α and NO from RAW 264.7 cells was increased in a time-dependent manner after exposure to all groups, whereas only F4- or F18-loaded TEMS-stimulated IL-6 secretion. The levels of IFN-γ from mouse splenocytes after exposure to F4 or F18 were increased while IL-4 was not detectable. These results suggest that F4- and F18-loaded TEMS may effectively induce immune response with the efficient release of antigens to appropriate target sites. Copyright © 2011 Elsevier B.V. All rights reserved.

  9. Variation in 12 porcine genes involved in the carbohydrate moiety assembly of glycosphingolipids does not account for differential binding of F4 Escherichia coli and their fimbriae.

    Science.gov (United States)

    Goetstouwers, Tiphanie; Van Poucke, Mario; Coddens, Annelies; Nguyen, Van Ut; Melkebeek, Vesna; Deforce, Dieter; Cox, Eric; Peelman, Luc J

    2014-10-03

    Glycosphingolipids (GSLs) are important membrane components composed of a carbohydrate structure attached to a hydrophobic ceramide. They can serve as specific membrane receptors for microbes and microbial products, such as F4 Escherichia coli (F4 ETEC) and isolated F4 fimbriae. The aim of this study was to investigate the hypothesis that variation in genes involved in the assembly of the F4 binding carbohydrate moiety of GSLs (i.e. ARSA, B4GALT6, GAL3ST1, GALC, GBA, GLA, GLB1, GLB1L, NEU1, NEU2, UGCG, UGT8) could account for differential binding of F4 ETEC and their fimbriae. RT-PCR could not reveal any differential expression of the 12 genes in the jejunum of F4 receptor-positive (F4R(+)) and F4 receptor-negative (F4R(-)) pigs. Sequencing the complete open reading frame of the 11 expressed genes (NEU2 was not expressed) identified 72 mutations. Although some of them might have a structural effect, none of them could be associated with a F4R phenotype. We conclude that no regulatory or structural variation in any of the investigated genes is responsible for the genetic susceptibility of pigs towards F4 ETEC.

  10. Crystallization and preliminary X-ray diffraction analysis of CfaE, the adhesive subunit of the CFA/I fimbriae from human enterotoxigenic Escherichia coli

    International Nuclear Information System (INIS)

    Li, Yong-Fu; Poole, Steven; Rasulova, Fatima; Esser, Lothar; Savarino, Stephen J.; Xia, Di

    2006-01-01

    The adhesin CfaE of the CFA/I fimbriae from human enterotoxigenic E. coli has been crystallized. CfaE crystals diffracted X-rays to better than 2.4 Å and phasing was solved by the SIRAS method. Enterotoxigenic Escherichia coli (ETEC) represents a formidable food and waterborne diarrheal disease threat of global importance. The first step in ETEC pathogenesis is bacterial attachment to small-intestine epithelial cells via adhesive fimbriae, many of which are genetically related to the prototype colonization factor antigen I (CFA/I). The minor fimbrial subunit CfaE is required for initiation of CFA/I fimbrial assembly and mediates bacterial attachment to host cell-surface receptors. A donor-strand complemented variant of CfaE (dscCfaE) was expressed with a hexahistidine tag, purified to homogeneity and crystallized using the hanging-drop vapor-diffusion method. X-ray diffraction data sets were collected to 2.4 Å resolution for both native and derivatized crystals and showed the symmetry of space group P6 2 22, with unit-cell parameters a = b = 142.9, c = 231.9 Å. Initial phases were derived from the SIRAS approach and electron density showed two molecules in the crystallographic asymmetric unit. Sequence assignments were aided by anomalous signals from the selenium of an SeMet-derivatized crystal and from S atoms of a native crystal

  11. Construction and expression of immunogenic hybrid enterotoxigenic Escherichia coli CFA/I and CS2 colonization fimbriae for use in vaccines.

    Science.gov (United States)

    Tobias, Joshua; Svennerholm, Ann-Mari; Holmgren, Jan; Lebens, Michael

    2010-07-01

    Enterotoxigenic Escherichia coli (ETEC) are an important cause of diarrheal morbidity in developing countries, especially in children and also of traveler's diarrhea. Colonization factors (CFs) of ETEC, like CFA/I and CS2 which are genetically and structurally related, play a substantial role in pathogenicity, and since intestinal-mucosal immune responses against CFs appear to be protective, much effort has focused on the development of a CF-based ETEC vaccine. We have constructed hybrid operons in which the major CS2 subunit-encoding cotA gene was inserted into the CFA/I operon, either replacing (hybrid I) or being added to the major CFA/I subunit-encoding cfaB gene (hybrid II). Using specific monoclonal antibodies against the major subunits of CFA/I and CS2, high levels of surface expression of both fimbrial subunits were shown in E. coli carrying the hybrid II operon. Oral immunization of mice with formalin-killed bacteria expressing hybrid II fimbriae induced strong CFA/I- and CS2-specific serum IgG + IgM and fecal IgA antibody responses, which were higher than those achieved by similar immunization with the reference strains. Bacteria expressing hybrid fimbriae are potential candidate strains in an oral-killed CF-ETEC vaccine, and the approach represents an attractive and novel means of producing a broad-spectrum ETEC vaccine.

  12. Role of Proteus mirabilis MR/P fimbriae and flagella in adhesion, cytotoxicity and genotoxicity induction in T24 and Vero cells.

    Science.gov (United States)

    Scavone, Paola; Villar, Silvia; Umpiérrez, Ana; Zunino, Pablo

    2015-06-01

    Proteus mirabilis is frequently associated with complicated urinary tract infections (UTI). It is proposed that several virulence factors are associated with P. mirabilis uropathogenicity. The aim of this work was to elucidate genotoxic and cytotoxic effects mediated by MR/P fimbriae and flagella in eukaryotic cells in vitro. Two cell lines (kidney- and bladder-derived) were infected with a clinical wild-type P. mirabilis strain and an MR/P and a flagellar mutant. We evaluated adhesion, genotoxicity and cytotoxicity by microscopy, comet assay and triple staining technique, respectively. Mutant strains displayed lower adhesion rates than the P. mirabilis wild-type strain and were significantly less effective to induce genotoxic and cytotoxic effects compared to the wild type. We report for the first time that P. mirabilis MR/P fimbriae and flagella mediate genotoxic and cytotoxic effects on eukaryotic cells, at least in in vitro conditions. These results could contribute to design new strategies for the control of UTI. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  13. PecS regulates the urate-responsive expression of type 1 fimbriae in Klebsiella pneumoniae CG43.

    Science.gov (United States)

    Wang, Zhe-Chong; Liu, Chia-Jui; Huang, Ying-Jung; Wang, Yu-Seng; Peng, Hwei-Ling

    2015-12-01

    In the Klebsiella pneumoniae CG43 genome, the divergently transcribed genes coding for PecS, the MarR-type transcription factor, and PecM, the drug metabolite transporter, are located between the type 1 and type 3 fimbrial gene clusters. The intergenic sequence pecO between pecS and pecM contains three putative PecS binding sites and a CpxR box. Electrophoretic mobility shift assay revealed that the recombinant PecS and CpxR could specifically bind to the pecO sequence, and the specific interaction of PecS and pecO could be attenuated by urate. The expression of pecS and pecM was negatively regulated by CpxAR and PecS, and was inducible by exogenous urate in the absence of cpxAR. Compared with CG43S3ΔcpxAR, the derived mutants CG43S3ΔcpxARΔpecS and CG43S3ΔcpxARΔpecSΔpecM exerted similar levels of sensitivity to H2O2 or paraquat, but higher levels of mannose-sensitive yeast agglutination activity and FimA production. The promoter activity and transcript levels of fimA in CG43S3ΔcpxAR were also increased by deleting pecS. However, no binding activity between PecS and the fimA promoter could be observed. Nevertheless, PecS deletion could reduce the expression of the global regulator HNS and release the negative effect of HNS on FimA expression. In CG43S3ΔcpxAR, the expression of FimA as well as PecS was inducible by urate, whilst urate-induced FimA expression was inhibited by the deletion of pecS. Taken together, we propose that K. pneumoniae PecS indirectly and negatively regulates the expression of type 1 fimbriae, and the regulation is urate-inducible in the absence of CpxAR.

  14. MrkH, a novel c-di-GMP-dependent transcriptional activator, controls Klebsiella pneumoniae biofilm formation by regulating type 3 fimbriae expression.

    Directory of Open Access Journals (Sweden)

    Jonathan J Wilksch

    2011-08-01

    Full Text Available Klebsiella pneumoniae causes significant morbidity and mortality worldwide, particularly amongst hospitalized individuals. The principle mechanism for pathogenesis in hospital environments involves the formation of biofilms, primarily on implanted medical devices. In this study, we constructed a transposon mutant library in a clinical isolate, K. pneumoniae AJ218, to identify the genes and pathways implicated in biofilm formation. Three mutants severely defective in biofilm formation contained insertions within the mrkABCDF genes encoding the main structural subunit and assembly machinery for type 3 fimbriae. Two other mutants carried insertions within the yfiN and mrkJ genes, which encode GGDEF domain- and EAL domain-containing c-di-GMP turnover enzymes, respectively. The remaining two isolates contained insertions that inactivated the mrkH and mrkI genes, which encode for novel proteins with a c-di-GMP-binding PilZ domain and a LuxR-type transcriptional regulator, respectively. Biochemical and functional assays indicated that the effects of these factors on biofilm formation accompany concomitant changes in type 3 fimbriae expression. We mapped the transcriptional start site of mrkA, demonstrated that MrkH directly activates transcription of the mrkA promoter and showed that MrkH binds strongly to the mrkA regulatory region only in the presence of c-di-GMP. Furthermore, a point mutation in the putative c-di-GMP-binding domain of MrkH completely abolished its function as a transcriptional activator. In vivo analysis of the yfiN and mrkJ genes strongly indicated their c-di-GMP-specific function as diguanylate cyclase and phosphodiesterase, respectively. In addition, in vitro assays showed that purified MrkJ protein has strong c-di-GMP phosphodiesterase activity. These results demonstrate for the first time that c-di-GMP can function as an effector to stimulate the activity of a transcriptional activator, and explain how type 3 fimbriae

  15. Oral immunization with F4 fimbriae and CpG formulated with carboxymethyl starch enhances F4-specific mucosal immune response and modulates Th1 and Th2 cytokines in weaned pigs.

    Science.gov (United States)

    Delisle, Benjamin; Calinescu, Carmen; Mateescu, Mircea Alexandru; Fairbrother, John Morris; Nadeau, Éric

    2012-01-01

    F4 fimbriae are a potential candidate for an oral subunit vaccine for prevention of post-weaning diarrhea in swine due to infection with F4-positive enterotoxigenic Escherichia coli. However, large quantities of F4 fimbriae are required to induce a specific antibody response. The aim of the present study was to evaluate the effect of supplementation of F4 fimbriae with Cytosine-phosphate-Guanosine-oligodeoxynucleotide (CpG-A D19) or with complete cholera toxin (CT) as adjuvants on the F4-specific antibody response and cytokine production in weaned pigs following oral administration of F4 fimbrial antigen formulated with Carboxymethyl Starch (CMS). Oral dosage forms of F4 fimbriae alone or supplemented with CpG-A D19 or with CT were formulated with CMS as monolithic tablets, obtained by direct compression, and administered to weaned pigs. Blood and faecal samples were collected to determine the systemic and mucosal immune status of animals at various times until necropsy. During necropsy, contents of the jejunum and ileum were collected for determination of mucosal F4 specific antibodies. Segments of jejunum and ileum were also used to measure mRNA cytokine production. The presence of CpG in the formulation of the fimbriae significantly increased F4-specific immunoglobulin (Ig) IgM and IgG levels in intestinal secretions, and enhanced Th1 (Interferon-gamma / IFN-γ, Tumour Necrosis Factor-alpha / TNF-α, Interleukin-12p40 / IL-12p40, IL-1β) and Th2 (IL-4, IL-6) cytokine production in intestinal tissues. Supplementation with CT did not result in induction of F4-specific antibodies in secretions, although a significant Th1 response (IFN-α, IFN-γ, IL-18) was detected in tissues. Neither F4-specific systemic antibodies, nor intestinally secreted IgA were detected throughout the immunization trial for all groups. CpG-A D19 appeared to be a promising adjuvant for an oral F4 subunit vaccine formulated with CMS excipient as monolithic tablets. This matrix afforded gastro

  16. Detection of Aggregatibacter actinomycetemcomitans leukotoxin and fimbria-associated protein gene genotypes among periodontitis patients and healthy controls: A case–control study

    Science.gov (United States)

    Mahalakshmi, Krishnan; Krishnan, Padma; Chandrasekaran, S. C.

    2018-01-01

    Background: Aggregatibacter actinomycetemcomitans has been reported in higher proportions in subgingival microbiota of individuals with aggressive periodontitis (AgP) compared with those with chronic periodontitis (ChP) and healthy controls. The major virulence factors are the ones that help in colonization and evasion of host's defenses. Hence, this study was aimed to assess the prevalence of A. actinomycetemcomitans 16S rRNA and its virulent genotypes (leukotoxin [lktA] and fimbria-associated protein [fap]). Materials and Methods: In this case– control study We performed periodontal examination and measured probing depth and clinical attachment level (CAL). Subgingival plaque samples from 200 (ChP: n = 128 and AgP: n = 72) periodontitis patients and 200 healthy controls were screened for the presence of A. actinomycetemcomitans 16S rRNA, lktA, and fap genotypes by polymerase chain reaction. The prevalence of genotypes between periodontitis patients and healthy controls was compared with Pearson's Chi-square test. P periodontitis patients, while A. actinomycetemcomitans fap genotype showed 31.8% prevalence. The prevalence of these genotypes was insignificant in healthy controls. Conclusion: The high odds ratio for A. actinomycetemcomitans prevalence suggests its strong link to periodontitis. Detection of A. actinomycetemcomitans lktA + genotype may be a useful marker for AgP as its prevalence was found to be high in AgP. PMID:29922337

  17. Association of germline mutation in the PTEN tumour suppressor gene and Proteus and Proteus-like syndromes

    NARCIS (Netherlands)

    Zhou, X.; Hampel, H.; Thiele, H.; Gorlin, R. J.; Hennekam, R. C.; Parisi, M.; Winter, R. M.; Eng, C.

    2001-01-01

    The molecular aetiology of Proteus syndrome (PS) remains elusive. Germline mutations in PTEN cause Cowden syndrome and Bannayan-Riley-Ruvalcaba syndrome, which are hereditary hamartoma syndromes. Some features-eg, macrocephaly, lipomatosis, and vascular malformations-can be seen in all three

  18. Inhibitory effects of tocopherols on expression of the cyclooxygenase-2 gene in RAW264.7 cells stimulated by lipopolysaccharide, tumor necrosis factor-α or Porphyromonas gingivalis fimbriae.

    Science.gov (United States)

    Murakami, Yukio; Kawata, Akifumi; Koh, Teho; Seki, Yuya; Tamura, Seiko; Katayama, Tadashi; Fujisawa, Seiichiro

    2013-01-01

    Tocopherols, which include α-, β-, γ-, and δ-tocopherol, protect cells against harmful free radicals and play an important role in preventing many human diseases such as cancer, inflammatory disorders, and ageing itself. However, the causal relationships between periodontal or oral chronic diseases and tocopherols have not been sufficiently studied. The present study investigated the inhibitory effects of these compounds on the expression of cyclooxygenase-2 (COX2) mRNA in RAW264.7 cells stimulated with lipopolysaccharide (LPS), tumor necrosis factor-α (TNFα) or fimbriae of Poryphyromonas gingivalis (Pg), an oral anaerobe. The cytotoxicity (EC₅₀) of tocopherols toward RAW cells was determined using a cell counting kit (CCK-8). The regulatory effect of these compounds on the expression of COX2 mRNA stimulated with LPS, TNFα or Pg fimbriae was investigated using real-time polymerase chain reaction (PCR). Each tocopherol had similarly low cytotoxicity. COX2 gene expression in RAW cells after exposure to the three different macrophage activators was inhibited by the tocopherols (ptocopherol, β-, γ- and δ-tocopherol exhibited greater inhibitory effects (pTocopherols exhibit anti-inflammatory activity, and β-, γ- and δ-tocopherol have particularly more potent anti-inflammatory activity than α-tocopherol. Tocopherols may have potential utility for prevention of periodontal and chronic oral diseases.

  19. Manual segmentation of the fornix, fimbria, and alveus on high-resolution 3T MRI: Application via fully-automated mapping of the human memory circuit white and grey matter in healthy and pathological aging.

    Science.gov (United States)

    Amaral, Robert S C; Park, Min Tae M; Devenyi, Gabriel A; Lynn, Vivian; Pipitone, Jon; Winterburn, Julie; Chavez, Sofia; Schira, Mark; Lobaugh, Nancy J; Voineskos, Aristotle N; Pruessner, Jens C; Chakravarty, M Mallar

    2018-04-15

    Recently, much attention has been focused on the definition and structure of the hippocampus and its subfields, while the projections from the hippocampus have been relatively understudied. Here, we derive a reliable protocol for manual segmentation of hippocampal white matter regions (alveus, fimbria, and fornix) using high-resolution magnetic resonance images that are complementary to our previous definitions of the hippocampal subfields, both of which are freely available at https://github.com/cobralab/atlases. Our segmentation methods demonstrated high inter- and intra-rater reliability, were validated as inputs in automated segmentation, and were used to analyze the trajectory of these regions in both healthy aging (OASIS), and Alzheimer's disease (AD) and mild cognitive impairment (MCI; using ADNI). We observed significant bilateral decreases in the fornix in healthy aging while the alveus and cornu ammonis (CA) 1 were well preserved (all p's<0.006). MCI and AD demonstrated significant decreases in fimbriae and fornices. Many hippocampal subfields exhibited decreased volume in both MCI and AD, yet no significant differences were found between MCI and AD cohorts themselves. Our results suggest a neuroprotective or compensatory role for the alveus and CA1 in healthy aging and suggest that an improved understanding of the volumetric trajectories of these structures is required. Copyright © 2016 Elsevier Inc. All rights reserved.

  20. A video demonstration of preserved piloting by scent tracking but impaired dead reckoning after fimbria-fornix lesions in the rat.

    Science.gov (United States)

    Whishaw, Ian Q; Gorny, Boguslaw P

    2009-04-24

    ; Martin et al., 1997; Maaswinkel and Whishaw, 1999). The objective of the present video demonstrations was to solve the problem of cue specification in order to examine the relative contribution of the hippocampus in the use of these strategies. The rats were trained in a new task in which they followed linear or polygon scented trails to obtain a large food pellet hidden on an open field. Because rats have a proclivity to carry the food back to the refuge, accuracy and the cues used to return to the home base were dependent variables (Whishaw and Tomie, 1997). To force an animal to use a a dead reckoning strategy to reach its refuge with the food, the rats were tested when blindfolded or under infrared light, a spectral wavelength in which they cannot see, and in some experiments the scent trail was additionally removed once an animal reached the food. To examine the relative contribution of the hippocampus, fimbria-fornix (FF) lesions, which disrupt information flow in the hippocampal formation (Bland, 1986), impair memory (Gaffan and Gaffan, 1991), and produce spatial deficits (Whishaw and Jarrard, 1995), were used.

  1. Passive immunity in cattle against enterotoxigenic Escherichia coli: serologic evaluation of a bacterin containing K99 and F41 fimbriae in colostrum of vaccinated females and calf serum Imunidade passiva contra Escherichia coli enterotoxigênica: avaliação sorológica de uma bacterina contendo as fímbrias K99 e F41 no colostro de fêmeas vacinadas e no soro de bezerros

    Directory of Open Access Journals (Sweden)

    H.C.P. Figueiredo

    2004-08-01

    Full Text Available A bacterin from enterotoxigenic Escherichia coli (ETEC, containing fimbriae K99 and F41, was produced and its capacity to induce anti-K99 and anti-F41 antibodies in colostrum of vaccinated cows and in calf serum, and the persistence of these antibodies in neonates were determined. Three experiments were performed on two commercial farms. In all experiments animals were allotted randomly to the blocks, each block consisting of two pregnant females (a vaccinated one and a control one and their respective calves. In experiment A (farm 1, comprised of 18 blocks, the animals received a vaccine dose 30 days before delivery. In experiment B (farm 1, consisted of 26 blocks, the animals received two vaccine doses (60 and 30 days before delivery. In experiment C (farm 2, consisted of 22 blocks, the animals received two vaccine doses (60 and 30 days before delivery. In experiments A and B pregnant cows and heifers were used and colostrum and serum from 24- to 36-hour-old calves were collected. In experiment C, pregnant embryo-recipient heifers were used and colostrum and sera from calves at 7, 14, 28 and 42 days of age were collected. Anti-K99 and anti-F41 antibodies were detected by ELISA using purified K99 and F41 fimbrial antigens. In experiment A no difference between treated and control groups was observed for the concentration of anti-K99 and anti-F41 antibodies in colostrum and calf serum. In experiment B a difference (PProduziu-se uma bacterina de Escherichia coli enterotoxigênica (ETEC contendo as fímbrias K99 e F41 e avaliaram-se a capacidade de indução de anticorpos anti-K99 e anti F-41 no colostro de vacas vacinadas e no soro de bezerros e a persistência dos anticorpos nos neonatos. Três experimentos foram realizados em duas fazendas comerciais. Os animais foram aleatoriamente alocados em blocos, de duas fêmeas prenhes (uma vacinada e outra controle e seus respectivos bezerros. No experimento A (fazenda 1, com 18 blocos, os animais

  2. Nanobody mediated inhibition of attachment of F18 Fimbriae expressing Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Kristof Moonens

    Full Text Available Post-weaning diarrhea and edema disease caused by F18 fimbriated E. coli are important diseases in newly weaned piglets and lead to severe production losses in farming industry. Protective treatments against these infections have thus far limited efficacy. In this study we generated nanobodies directed against the lectin domain of the F18 fimbrial adhesin FedF and showed in an in vitro adherence assay that four unique nanobodies inhibit the attachment of F18 fimbriated E. coli bacteria to piglet enterocytes. Crystallization of the FedF lectin domain with the most potent inhibitory nanobodies revealed their mechanism of action. These either competed with the binding of the blood group antigen receptor on the FedF surface or induced a conformational change in which the CDR3 region of the nanobody displaces the D″-E loop adjacent to the binding site. This D″-E loop was previously shown to be required for the interaction between F18 fimbriated bacteria and blood group antigen receptors in a membrane context. This work demonstrates the feasibility of inhibiting the attachment of fimbriated pathogens by employing nanobodies directed against the adhesin domain.

  3. Identification and Characterization of a Gene Cluster Mediating Enteroaggregative Escherichia Coli Aggregative Adherence Fimbria I Biogenesis

    Science.gov (United States)

    1994-08-01

    microscopy were performed by standard methods with fluoresceinated forward and reverse pUC primers (ABI) with a JOEL JEM 1200 EX 11 transr-ission...MyfB 11 enzrocolinca 41 220 uous plasmid regions required for AAF/I expre:sion and AA. PapD E. cobi 34 200 In this paper. we present a detailed analysis

  4. Composition of fish and feeds - Development and optimization of grow out diets for sablefish (Anoplopoma fimbria)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Our experience with juvenile sablefish and long term rearing of broodstock indicate that salmon grower feeds currently used by commercial sablefish farmers for grow...

  5. Uropathogenic Escherichia coli Express Type 1 Fimbriae Only in Surface Adherent Populations Under Physiological Growth Conditions

    DEFF Research Database (Denmark)

    Stærk, Kristian; Kolmos, Hans Jørn; Khandige, Surabhi

    2016-01-01

    were correlated with the ability to adhere to and invade cultured human bladder cells. RESULTS:  Although inactive during planktonic growth in urine, T1F expression occurs when UPEC settles on and infects bladder epithelial cells or colonizes catheters. As a result, UPEC in these sessile populations...... with increased expression during surface growth adaptation and infection of uroepithelial cells. This leads to separation of UPEC into low-expression planktonic populations and high-expression sessile populations....... enhances bladder cell adhesion and invasion potential. Only T1F-negative UPEC are subsequently released to the urine, thus limiting T1F expression to surface-associated UPEC alone. CONCLUSION:  Our results demonstrate that T1F expression is strictly regulated under physiological growth conditions...

  6. Growth data - Development and optimization of grow out diets for sablefish (Anoplopoma fimbria)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Our experience with juvenile sablefish and long term rearing of broodstock indicate that salmon grower feeds currently used by commercial sablefish farmers for grow...

  7. Fish culture data - Development and optimization of grow out diets for sablefish (Anoplopoma fimbria)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Our experience with juvenile sablefish and long term rearing of broodstock indicate that salmon grower feeds currently used by commercial sablefish farmers for grow...

  8. Chimeric FimH adhesin of type 1 fimbriae: a bacterial surface display system for heterologous sequences

    DEFF Research Database (Denmark)

    Pallesen, L; Poulsen, LK; Christiansen, Gunna

    1995-01-01

    of heterologous DNA segments encoding two reporter sequences. In the selected positions such insertions did not significantly alter the function of the FimH protein with regard to surface location and adhesive ability. The system seemed to be quite flexible, since chimeric versions of the FimH adhesin containing...... as many as 56 foreign amino acids were transported to the bacterial surface as components of the fimbrial organelles. Furthermore, the foreign protein segments were recognized by insert-specific antibodies when expressed within chimeric proteins on the surface of the bacteria. The results from...

  9. Volunteer Challenge With Enterotoxigenic Escherichia coli That Express Intestinal Colonization Factor Fimbriae CS17 and CS19

    Science.gov (United States)

    2011-07-01

    Serotype was determined by classic serological methods at the Universidad Nacional Aut6noma de Mexico [UNAMl. H- indrcates non-motility. b CF...Levine MM, Merson MM. Serologic differentiation between antitoxin responses to infection with Vibrio cholerae and enterotoxin-producing Escherichia coli...prototype cholera B subunit-colonization factor antigen cnterotoxigenic Escherichia coli vaccine. Vaccine 1993; 1[:929-34. 15. Levine MM, Nalin DR

  10. Effect of dietary taurine supplementation on growth, feed efficiency, and nutrient composition of juvenile sablefish (Anoplopoma fimbria)

    Science.gov (United States)

    Juvenile sablefish were fed a low taurine, basal feed with seven graded levels of supplemental taurine to determine taurine requirements for growth and feed efficiency. The basal feed was plant based, formulated primarily with soy and corn proteins with a minimal (9%) amount of fishmeal. The unsuppl...

  11. A novel pAA virulence plasmid encoding toxins and two distinct variants of the fimbriae of enteroaggregative Escherichia coli

    DEFF Research Database (Denmark)

    Jønsson, Rie; Struve, Carsten; Boll, Erik J.

    2017-01-01

    phylogenetically distinct, strains harboring the major pilin subunits from both AAF/III and AAF/V. Whole-genome and plasmid sequencing revealed that in these six strains the agg3A and agg5A genes were located on a novel pAA plasmid variant. Moreover, the plasmid also encoded several other virulence genes including...... some not previously found on pAA plasmids. Thus, this plasmid endows the host strains with a remarkably high number of EAEC associated virulence genes hereby likely promoting strain pathogenicity....

  12. Structural and functional studies of Escherichia coli aggregative adherence fimbriae (AAF/V) reveal a deficiency in extracellular matrix binding

    DEFF Research Database (Denmark)

    Jønsson, Rie; Liu, Bing; Struve, Carsten

    2017-01-01

    A possesses an inserted helix at the beginning of the donor strand, which together with altered surface electrostatics, renders the protein unable to interact with fibronectin. Hence, here we characterize the first AAF variant with a binding mode that varies from previously described AAFs...

  13. Taurine Growth Effects - Determining Optimum Taurine Supplementation Levels for Plant Proteins Incorporated into Marine Finfish Feeds using Juvenile Sablefish Anoplopoma fimbria

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Taurine, an amino sulfonic acid, has important roles in osmoregulation, bile acid conjugation, membrane stabilization and calcium homeostasis in vertebrates. Though...

  14. The PprA-PprB two-component system activates CupE, the first non-archetypal Pseudomonas aeruginosa chaperione-usher pathway system assembling fimbriae

    DEFF Research Database (Denmark)

    Giraud, Caroline; Bernard, Christophe S.; Calderon, Virginie

    2011-01-01

    - and chaperone-encoding genes. This locus, widely conserved in different bacterial species, contains four additional genes encoding non-archetypal fimbrial subunits. We first evidenced that the cupE gene cluster was specifically expressed in biofilm conditions and was responsible for fibre assembly containing...

  15. New vaccine strategies against enterotoxigenic Escherichia coli: II: Enhanced systemic and secreted antibody responses against the CFA/I fimbriae by priming with DNA and boosting with a live recombinant Salmonella vaccine

    Directory of Open Access Journals (Sweden)

    M.O. Lásaro

    1999-02-01

    Full Text Available The induction of systemic (IgG and mucosal (IgA antibody responses against the colonization factor I antigen (CFA/I of enterotoxigenic Escherichia coli (ETEC was evaluated in mice primed with an intramuscularly delivered CFA/I-encoding DNA vaccine followed by two oral immunizations with a live recombinant Salmonella typhimurium vaccine strain expressing the ETEC antigen. The booster effect induced by the oral immunization was detected two weeks and one year after the administration of the DNA vaccine. The DNA-primed/Salmonella-boosted vaccination regime showed a synergistic effect on the induced CFA/I-specific systemic and secreted antibody levels which could not be attained by either immunization strategy alone. These results suggest that the combined use of DNA vaccines and recombinant Salmonella vaccine strains can be a useful immunization strategy against enteric pathogens.

  16. Stimulation of mucosal immune response following oral administration of enterotoxigenic Escherichia coli fimbriae (CFA/I) entrapped in liposomes in conjunction with inactivated whole-cell Vibrio cholerae vaccine.

    Science.gov (United States)

    Dima, V F; Ionescu, M D; Palade, R; Balotescu, C; Becheanu, G; Dima, S V

    2001-01-01

    In this study, we have searched for an effective mucosal vaccine. An oral enterotoxigenic E. coli vaccine containing colonization factor antigen (CFA/I) associated with inactivated whole-cell V. cholerae vaccine (WCV) has been tested for safety and immunogenicity in animals. Five groups of animals were used. The results showed the following: (a) vaccine containing CFA/I antigen entrapped in liposomes and associated with WCV (batch C) had increased titers of specific antibodies to CFA/I antigen in 15 to 18 (83.3%) animals; (b) specific Peyer's patches (PP), lymph nodes (LN) and spleen (SPL) lymphocytes proliferation was detected following in vitro restimulation with CFA/I antigen or WCV. This response gradually increased to the highest value by the 35th postimmunization day. Moreover, lower PP, LN and spleen (SPL) proliferation was observed in rabbits receiving soluble CFA/I antigen (S-CFA/I) or free liposomes (F-L) alone; (c) adhesion of E. coli H10407 strain labelled with 3H-leucine in immunized and control animals revealed the following local effects: (i) protection of rabbit intestinal mucosa against virulent E. coli cells; (ii) inhibition of adhesion of ETEC bacteria to intestinal mucosa and (iii) significantly faster release of E. coli H 10407 strain labelled with 3H-leucine from the intestinal tract of immunized animals. The histopathological and electron microscope findings confirmed the above results. The experimental results point out an efficient protection against infection with E. coli strains (ETEC), after mucosal vaccination with CFA/I antigen entrapped in liposomes associated with inactivated whole-cell Vibrio cholerae as immunological adjuvant.

  17. Construction and evaluation of the immune protection of a recombinant divalent protein composed of the MrpA from MR/P fimbriae and flagellin of Proteus mirabilis strain against urinary tract infection.

    Science.gov (United States)

    Habibi, Mehri; Asadi Karam, Mohammad Reza; Bouzari, Saeid

    2018-04-01

    Urinary tract infections (UTI) caused by Proteus mirabilis are prevalent among the catheterized patients. There is no effective vaccine to reduce the frequency of UTIs caused by P. mirabilis. In the present study, the immune responses and effectiveness of different combinations of MrpA and flagellin (FliC) of P. mirabilis were assessed intranasally in the mice model. The addition of FliC as adjuvant to MrpA in fusion form significantly raised the mucosal IgA and cellular (IFN-γ and IL-17) responses and maintained the serum IgG responses for 180 days after the first vaccination. Furthermore, MrpA in fusion form with FliC significantly increased the systemic, mucosal and IFN-γ responses of the FliC alone. In a bladder challenge assay with P. mirabilis, the fusion MrpA.FliC and the mixture of MrpA and FliC significantly decreased the colony count of the bacteria in the bladder and kidneys of mice in comparison to the control mice. It suggests a complex of the systemic, mucosal and cellular responses are needed for protection of the bladder and kidneys against P. mirabilis UTI. In our knowledge, the adjuvant property of the recombinant P. mirabilis flagellin was evaluated for the first time in a vaccine combination administered by an intranasal route. Our results suggest the recombinant flagellin of P. mirabilis could be used as an intranasal adjuvant in combination with other potential antigens against UTIs. Copyright © 2018 Elsevier Ltd. All rights reserved.

  18. AFSC/ABL: Gulf of Alaska Diel Trawl Survey, 2005-2006

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Diel epipelagic sampling for juvenile Pacific salmon (Oncorhynchus spp.), rockfish (Sebastes spp.), sablefish (Anoplopoma fimbria), and associated species was...

  19. The Escherichia coli K-12 gntP gene allows E. coli F-18 to occupy a distinct nutritional niche in the streptomycin-treated mouse large intestine

    DEFF Research Database (Denmark)

    Sweeney, N.J.; Klemm, Per; McCormick, Beth A.

    1996-01-01

    Escherichia coli F-18 is a human fecal isolate that makes type 1 fimbriae, encoded by the fim gene cluster, and is an excellent colonizer of the streptomycin-treated mouse intestine. E. coli F-18 fimA::tet, lacking type 1 fimbriae, was constructed by bacteriophage P1 transduction of the fim regio...

  20. Functional Analysis of the Chaperone-Usher Fimbrial Gene Clusters of Salmonella enterica serovar Typhi.

    Science.gov (United States)

    Dufresne, Karine; Saulnier-Bellemare, Julie; Daigle, France

    2018-01-01

    The human-specific pathogen Salmonella enterica serovar Typhi causes typhoid, a major public health issue in developing countries. Several aspects of its pathogenesis are still poorly understood. S . Typhi possesses 14 fimbrial gene clusters including 12 chaperone-usher fimbriae ( stg, sth, bcf , fim, saf , sef , sta, stb, stc, std, ste , and tcf ). These fimbriae are weakly expressed in laboratory conditions and only a few are actually characterized. In this study, expression of all S . Typhi chaperone-usher fimbriae and their potential roles in pathogenesis such as interaction with host cells, motility, or biofilm formation were assessed. All S . Typhi fimbriae were better expressed in minimal broth. Each system was overexpressed and only the fimbrial gene clusters without pseudogenes demonstrated a putative major subunits of about 17 kDa on SDS-PAGE. Six of these (Fim, Saf, Sta, Stb, Std, and Tcf) also show extracellular structure by electron microscopy. The impact of fimbrial deletion in a wild-type strain or addition of each individual fimbrial system to an S . Typhi afimbrial strain were tested for interactions with host cells, biofilm formation and motility. Several fimbriae modified bacterial interactions with human cells (THP-1 and INT-407) and biofilm formation. However, only Fim fimbriae had a deleterious effect on motility when overexpressed. Overall, chaperone-usher fimbriae seem to be an important part of the balance between the different steps (motility, adhesion, host invasion and persistence) of S . Typhi pathogenesis.

  1. DNA microarray analysis of fim mutations in Escherichia coli

    DEFF Research Database (Denmark)

    Schembri, Mark; Ussery, David; Workman, Christopher

    2002-01-01

    Bacterial adhesion is often mediated by complex polymeric surface structures referred to as fimbriae. Type I fimbriae of Escherichia coli represent the archetypical and best characterised fimbrial system. These adhesive organelles mediate binding to D-mannose and are directly associated...... we have used DNA microarray analysis to examine the molecular events involved in response to fimbrial gene expression in E. coli K-12. Observed differential expression levels of the fim genes were in good agreement with our current knowledge of the stoichiometry of type I fimbriae. Changes in fim...

  2. Influence of some bacterial and host factors on colonization and invasiveness of Escherichia coli K1 in neonatal rats.

    OpenAIRE

    Wullenweber, M; Beutin, L; Zimmermann, S; Jonas, C

    1993-01-01

    Of 209 healthy infants examined, 44 (21.1%) carried Escherichia coli K1 in their feces. Of these 44 isolates, 36 (81.8%) were attributed to 10 different known clonal groups of E. coli K1 and 4 isolates represented unknown types. The influence of mannose-resistant (MR) adhesins, aerobactin production, and resistance to serum on colonization and invasiveness of E. coli K1 in orally infected inbred LEW baby rats was investigated. Strains expressing MR adhesins had significantly higher colonizati...

  3. Efectos del tratamiento con Compvit B, ácido orótico o la combinación de ambos sobre la recuperación de la memoria espacial en ratas con lesión de fimbria-fornix

    OpenAIRE

    González Gay, Olivia Teresa; Almaguer Melián, William; Ramírez Figueredo, Magalys; Vallejo Morales, Araceli; Bergado Rosado, Jorge Alberto

    2011-01-01

    Las vitaminoterapias han sido ampliamente utilizadas en neurología para el tratamiento de neuritis o la correción de déficit metabólicos. En Cuba, se produce desde hace algunos años el preparado vitamínico Compvit®, que contiene vitaminas B1, B6 y B12. El ácido orótico, también llamado vitamina B13, es un producto natural que ha mostrado acciones como nootrópico en estudios con animales jóvenes y viejos que acusan deterioro cognitivo. En el presente trabajo se reportan los resultados de un es...

  4. California coast sablefish - Reproductive Life History Analysis of Sablefish Populations off the Washington and California Coasts

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Sablefish (Anoplopoma fimbria) have a wide distribution along the Pacific coast, extending from Baja California to Alaska, the Bering Sea and through to the eastern...

  5. Medline Plus

    Full Text Available ... hormones signal the development and release a single egg cell from one of the ovaries. The sweeping motion of the fimbriae draws the egg cell through a very small space in the ...

  6. Type 1 fimbrial expression enhances Escherichia coli virulence for the urinary tract

    DEFF Research Database (Denmark)

    Connell, Hugh; Agace, William; Klemm, Per

    1996-01-01

    of Escherichia coli for the urinary tract by promoting bacterial persistence and enhancing the inflammatory responce to infection. In a clinical study, we observed that disease severity was greater in children infected with E. coli O1:K1:H7 isolates expressing type 1 fimbriae than in those infected with type 1...... negative isolates of the same serotype. The E. coli O1:K1:H7 isolates had the same electrophoretic type, were hemolysin-negative, expressed P fimbriae, and carried the fim DNA sequences. When tested in a mouse urinary tract infection model, the type 1-positive E. coli O1:K1:H7 isolates survived inhigher...... urinary tract infection model. E. coli CN1016 reconstituted with type 1 fimbriae had restored virulence similar to that of the wild-type parent strain. These results show that type 1 fimbriae in the genetic background of a uropathogenic strain contribute to the pathogenesis of E. coli in the urinary tract....

  7. Sablefish GI microbiome & histology - Incorporating Gastrointestinal Microbiome Analysis into Fish Nutritional Assessments

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Sustainable culture of sablefish (Anoplopoma fimbria) is an active research area at NWFSC, with a primary objective of rearing a human food source under healthy,...

  8. Washington coast sablefish - Reproductive Life History Analysis of Sablefish Populations off the Washington and California Coasts

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Sablefish (Anoplopoma fimbria) have a wide distribution along the Pacific coast, extending from Baja California to Alaska, the Bering Sea and through to the eastern...

  9. The F4 fimbrial antigen of Escherichia coli and its receptors.

    NARCIS (Netherlands)

    Van den Broeck, W; Cox, E; Oudega, B.; Goddeeris, B

    2000-01-01

    F4 or K88 fimbriae are long filamentous polymeric surface proteins of enterotoxigenic Escherichia coli (ETEC), consisting of so-called major (FaeG) and minor (FaeF, FaeH, FaeC, and probably FaeI) subunits. Several serotypes of F4 have been described, namely F4ab, F4ac, and F4ad. The F4 fimbriae

  10. Type 1 fimbrial expression enhances Escherichia coli virulence for the urinary tract.

    OpenAIRE

    Connell, I; Agace, W; Klemm, P; Schembri, M; Mărild, S; Svanborg, C

    1996-01-01

    Type 1 fimbriae are adhesion organelles expressed by many Gram-negative bacteria. They facilitate adherence to mucosal surfaces and inflammatory cells in vitro, but their contribution to virulence has not been defined. This study presents evidence that type 1 fimbriae increase the virulence of Escherichia coli for the urinary tract by promoting bacterial persistence and enhancing the inflammatory response to infection. In a clinical study, we observed that disease severity was greater in chil...

  11. Escherichia coli O157:H7 Acid Sensitivity Correlates with Flocculation Phenotype during Nutrient Limitation

    Directory of Open Access Journals (Sweden)

    Kathryn L. Kay

    2017-07-01

    Full Text Available Shiga toxin producing Escherichia coli (STEC strains vary in acid resistance; however, little is known about the underlying mechanisms that result in strain specific differences. Among 25 STEC O157:H7 strains tested, 7 strains flocculated when grown statically for 18 h in minimal salts medium at 37°C, while 18 strains did not. Interestingly, the flocculation phenotype (cells came out of suspension was found to correlate with degree of acid sensitivity in an assay with 400 mM acetic acid solution at pH 3.3 targeting acidified foods. Strains exhibiting flocculation were more acid sensitive and were designated FAS, for flocculation acid sensitive, while the acid resistant strain designated PAR for planktonic acid resistant. Flocculation was not observed for any strains during growth in complex medium (Luria Bertani broth. STEC strains B201 and B241 were chosen as representative FAS (2.4 log reduction and PAR (0.15 log reduction strains, respectively, due to differences in acid resistance and flocculation phenotype. Results from electron microscopy showed evidence of fimbriae production in B201, whereas fimbriae were not observed in B241.Curli fimbriae production was identified through plating on Congo red differential medium, and all FAS strains showed curli fimbriae production. Surprisingly, 5 PAR strains also had evidence of curli production. Transcriptomic and targeted gene expression data for B201 and B241indicated that csg and hde (curli and acid induced chaperone genes, respectively expression positively correlated with the phenotypic differences observed for these strains. These data suggest that FAS strains grown in minimal medium express curli, resulting in a flocculation phenotype. This may be regulated by GcvB, which positively regulates curli fimbriae production and represses acid chaperone proteins. RpoS and other regulatory mechanisms may impact curli fimbriae production, as well. These findings may help elucidate mechanisms

  12. Artificial theta stimulation impairs encoding of contextual fear memory.

    Directory of Open Access Journals (Sweden)

    Arto Lipponen

    Full Text Available Several experiments have demonstrated an intimate relationship between hippocampal theta rhythm (4-12 Hz and memory. Lesioning the medial septum or fimbria-fornix, a fiber track connecting the hippocampus and the medial septum, abolishes the theta rhythm and results in a severe impairment in declarative memory. To assess whether there is a causal relationship between hippocampal theta and memory formation we investigated whether restoration of hippocampal theta by electrical stimulation during the encoding phase also restores fimbria-fornix lesion induced memory deficit in rats in the fear conditioning paradigm. Male Wistar rats underwent sham or fimbria-fornix lesion operation. Stimulation electrodes were implanted in the ventral hippocampal commissure and recording electrodes in the septal hippocampus. Artificial theta stimulation of 8 Hz was delivered during 3-min free exploration of the test cage in half of the rats before aversive conditioning with three foot shocks during 2 min. Memory was assessed by total freezing time in the same environment 24 h and 28 h after fear conditioning, and in an intervening test session in a different context. As expected, fimbria-fornix lesion impaired fear memory and dramatically attenuated hippocampal theta power. Artificial theta stimulation produced continuous theta oscillations that were almost similar to endogenous theta rhythm in amplitude and frequency. However, contrary to our predictions, artificial theta stimulation impaired conditioned fear response in both sham and fimbria-fornix lesioned animals. These data suggest that restoration of theta oscillation per se is not sufficient to support memory encoding after fimbria-fornix lesion and that universal theta oscillation in the hippocampus with a fixed frequency may actually impair memory.

  13. Stx1 prophage excision in Escherichia coli strain PA20 confers strong curli and biofilm formation by restoring native mlrA

    Science.gov (United States)

    Prophage insertions in Escherichia coli O157:H7 mlrA contribute to the low expression of curli fimbriae and biofilm observed in many clinical isolates. Varying levels of CsgD-dependent curli/biofilm expression are restored to strains bearing prophage insertions in mlrA by mutation of regulatory gene...

  14. Gene duplication and promoter mutation expand the range csgD-dependent biofilm responses in a STEC population

    Science.gov (United States)

    Expression of Escherichia coli major biofilm components, curli fimbriae and cellulose, require the CsgD transcription factor. A complex regulatory network allows environmental control of csgD transcription and biofilm formation. However, most clinical serotype O157:H7 strains contain prophage inser...

  15. Marker-assisted selection on E. coli F4ab/ac resistance and the effect on neonatal survival in piglets

    DEFF Research Database (Denmark)

    Jørgensen, Claus Bøttcher; Anderson, Susan I.; Cirera, Susanna

    Enterotoxigenic Escherichia coli (ETEC) that express the F4ab or F4ac fimbriae (formerly known as K88ab/ac) are major causes of diarrhea and death in neonatal and young pigs. A locus controlling susceptibility towards ETEC F4ab/ac has previously been mapped to pig chromosome 13q41. A number...

  16. Prevention of diarrhoea using pathogen specific monoclonal antibodies in an experimental enterotoxigenic E. coli infection in germfree piglets

    NARCIS (Netherlands)

    Geus, de B.; Harmsen, M.; Zijderveld, van F.

    1998-01-01

    In the present study we describe the effect of oral application of mAB specific for ETEC F4(ac) fimbriae in an experimental ETEC challenge model in neonatal germfree piglets. The results show that mAB, specific for different F4(ac) epitopes protect animals against ETEC specific pathology. Moreover,

  17. Reduction of Salmonella Enteritidis in the spleens of hens by bacterins that vary in fimbrial protein SefD

    Science.gov (United States)

    Gene sefD is part of operon sefABCD, and it is required for production of the SEF14 fimbria by Salmonella Enteritidis. We compared strains that varied in SefD content for their ability to reduce recovery of Salmonella Enteritidis from the spleens of hens infected by parenteral challenge. The two bac...

  18. Virulence and metabolic characteristics of Salmonella Enteritidis sefD variants in hens.

    Science.gov (United States)

    Salmonella Enteritidis is one of a few pathogenic Salmonella enterica serotypes that have SEF14 fimbriae encoded by the sef operon, which consists of 4 co-transcribed genes sefABCD that are regulated by sefR. To explore the function of sefD within the infection pathway resulting in egg contamination...

  19. Fulltext PDF

    Indian Academy of Sciences (India)

    2013-07-14

    Jul 14, 2013 ... 5Division of Animal Medicine, Animal Technology Institute Taiwan, Chunan, Miaoli, Taiwan ... Static broth culture favours the production of fimbriae, while solid agar .... study. Antibiotics (100 μg ampicillin/mL and 50 μg kana-.

  20. The Pai-associated leuX specific tRNA5(Leu) affects type 1fimbriation in pathogenic Escherichia coli by control of FimB recombinase expression

    DEFF Research Database (Denmark)

    Ritter, A.; Gally, D.; Olsen, Peter Bjarke

    1997-01-01

    The uropathogenic Escherichia coli strain 536 (06:K15:H31) carries two large chromosomalpathogenicity islands (Pais). Both Pais are flanked by tRNA genes. Spontaneous deletion of Pai IIresults in truncation of the leuX tRNA5Leu gene. This tRNA is required for the expression of type 1fimbriae (Fim...

  1. Antigen-43-mediated autoaggregation of Escherichia coli is blocked by fimbriation

    DEFF Research Database (Denmark)

    Hasman, Henrik; Chakraborty, Trinad; Klemm, Per

    1999-01-01

    that the expression of type 1 fimbriae and the expression of Ag43 are mutually exclusive. In the present report, we show, by use of well-defined mutants, that fimbriation abolishes Ag43-mediated autoaggregation but does not affect Ag43 expression. Autoaggregation is shown to require an intercellular Ag43-Ag43...

  2. Interactions of Neuropathogenic Escherichia coli K1 (RS218) and Its Derivatives Lacking Genomic Islands with Phagocytic Acanthamoeba castellanii and Nonphagocytic Brain Endothelial Cells

    Science.gov (United States)

    Yousuf, Farzana Abubakar; Yousuf, Zuhair; Iqbal, Junaid; Siddiqui, Ruqaiyyah; Khan, Hafsa; Khan, Naveed Ahmed

    2014-01-01

    Here we determined the role of various genomic islands in E. coli K1 interactions with phagocytic A. castellanii and nonphagocytic brain microvascular endothelial cells. The findings revealed that the genomic islands deletion mutants of RS218 related to toxins (peptide toxin, α-hemolysin), adhesins (P fimbriae, F17-like fimbriae, nonfimbrial adhesins, Hek, and hemagglutinin), protein secretion system (T1SS for hemolysin), invasins (IbeA, CNF1), metabolism (D-serine catabolism, dihydroxyacetone, glycerol, and glyoxylate metabolism) showed reduced interactions with both A. castellanii and brain microvascular endothelial cells. Interestingly, the deletion of RS218-derived genomic island 21 containing adhesins (P fimbriae, F17-like fimbriae, nonfimbrial adhesins, Hek, and hemagglutinin), protein secretion system (T1SS for hemolysin), invasins (CNF1), metabolism (D-serine catabolism) abolished E. coli K1-mediated HBMEC cytotoxicity in a CNF1-independent manner. Therefore, the characterization of these genomic islands should reveal mechanisms of evolutionary gain for E. coli K1 pathogenicity. PMID:24818136

  3. Interactions of Neuropathogenic Escherichia coli K1 (RS218 and Its Derivatives Lacking Genomic Islands with Phagocytic Acanthamoeba castellanii and Nonphagocytic Brain Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Farzana Abubakar Yousuf

    2014-01-01

    Full Text Available Here we determined the role of various genomic islands in E. coli K1 interactions with phagocytic A. castellanii and nonphagocytic brain microvascular endothelial cells. The findings revealed that the genomic islands deletion mutants of RS218 related to toxins (peptide toxin, α-hemolysin, adhesins (P fimbriae, F17-like fimbriae, nonfimbrial adhesins, Hek, and hemagglutinin, protein secretion system (T1SS for hemolysin, invasins (IbeA, CNF1, metabolism (D-serine catabolism, dihydroxyacetone, glycerol, and glyoxylate metabolism showed reduced interactions with both A. castellanii and brain microvascular endothelial cells. Interestingly, the deletion of RS218-derived genomic island 21 containing adhesins (P fimbriae, F17-like fimbriae, nonfimbrial adhesins, Hek, and hemagglutinin, protein secretion system (T1SS for hemolysin, invasins (CNF1, metabolism (D-serine catabolism abolished E. coli K1-mediated HBMEC cytotoxicity in a CNF1-independent manner. Therefore, the characterization of these genomic islands should reveal mechanisms of evolutionary gain for E. coli K1 pathogenicity.

  4. Fimbrial adhesins from extraintestinal Escherichia coli

    DEFF Research Database (Denmark)

    Klemm, Per; Hancock, Viktoria; Schembri, Mark A.

    2010-01-01

    Extraintestinal pathogenic Escherichia coli (ExPEC) represent an important subclass of E. coli that cause a wide spectrum of diseases in human and animal hosts. Fimbriae are key virulence factors of ExPEC strains. These long surface located rod-shaped organelles mediate receptor-specific attachment...

  5. Serum Antibodies Protect against Intraperitoneal Challenge with Enterotoxigenic Escherichia coli

    Directory of Open Access Journals (Sweden)

    Xinghong Yang

    2011-01-01

    Full Text Available To assess whether anticolonization factor antigen I (CFA/I fimbriae antibodies (Abs from enterotoxigenic Escherichia coli (ETEC can protect against various routes of challenge, BALB/c mice were immunized with a live attenuated Salmonella vaccine vector expressing CFA/I fimbriae. Vaccinated mice elicited elevated systemic IgG and mucosal IgA Abs, unlike mice immunized with the empty Salmonella vector. Mice were challenged with wild-type ETEC by the oral, intranasal (i.n., and intraperitoneal (i.p. routes. Naïve mice did not succumb to oral challenge, but did to i.n. challenge, as did immunized mice; however, vaccinated mice were protected against i.p. ETEC challenge. Two intramuscular (i.m. immunizations with CFA/I fimbriae without adjuvant conferred 100% protection against i.p. ETEC challenge, while a single 30 μg dose conferred 88% protection. Bactericidal assays showed that ETEC is highly sensitive to anti-CFA/I sera. These results suggest that parenteral immunization with purified CFA/I fimbriae can induce protective Abs and may represent an alternative method to elicit protective Abs for passive immunity to ETEC.

  6. Cellular chain formation in Escherichia coli biofilms

    DEFF Research Database (Denmark)

    Vejborg, Rebecca Munk; Klemm, Per

    2009-01-01

    ; type I fimbriae expression significantly reduced cellular chain formation, presumably by steric hindrance. Cellular chain formation did not appear to be specific to E coli K-12. Although many urinary tract infection (UTI) isolates were found to form rather homogeneous, flat biofilms, three isolates...

  7. Xyloglucan, hibiscus and propolis for the prevention of urinary tract infections: results of in vitro studies.

    Science.gov (United States)

    Fraile, Benito; Alcover, Javier; Royuela, Mar; Rodríguez, David; Chaves, Concepción; Palacios, Ricardo; Piqué, Núria

    2017-06-01

    To assess the properties of a medical device containing xyloglucan, propolis and hibiscus to create a bioprotective barrier to avoid the contact of uropathogenic Escherichia coli strains on cell walls in models of intestinal (CacoGoblet) and uroepithelial (RWPE-1) cells (derived from normal human prostate epithelium). Two uropathogenic E. coli strains (expressing type 1 fimbriae and P fimbriae) were used to assess, by electronic microscopy and ELISA, the barrier properties of the medical device. The antimicrobial activity was assessed in broth dilution assays. The three components (xyloglucan, propolis and hibiscus) did not alter E. coli cell integrity in intestinal and uroepithelial cell models and were devoid of antibacterial activity. The three components avoided bacterial contact in both cell monolayers. The nonpharmacological barrier properties of xyloglucan, propolis and hibiscus confirm the role of the medical device for the management of urinary tract infections.

  8. Impact of cranberry on Escherichia coli cellular surface characteristics

    International Nuclear Information System (INIS)

    Johnson, Brandy J.; Lin Baochuan; Dinderman, Michael A.; Rubin, Robert A.; Malanoski, Anthony P.; Ligler, Frances S.

    2008-01-01

    The anti-adhesive effects of cranberry have been attributed to both interactions of its components with the surface of bacterial cells and to inhibition of p-fimbriae expression. Previous reports also suggested that the presence of cranberry juice changed the Gram stain characteristics of Escherichia coli. Here, we show that the morphology of E. coli is changed when grown in the presence of juice or extract from Vaccinium macrocarpon (cranberry). Gene expression analysis indicates the down regulation of flagellar basal body rod and motor proteins. Consistent with this finding and previous reports, the SEM images indicate a decrease in the visible p-fimbriae. The iodine used in Gram-staining protocols was found to interact differently with the bacterial membrane when cells were cultured in spiked media. Slight alterations in the Gram stain protocol demonstrated that culturing in the presence of cranberry juice does not change the Gram stain characteristics contradicting other reports.

  9. Fimbrial phase variation

    DEFF Research Database (Denmark)

    Khandige, Surabhi; Møller-Jensen, Jakob

    2016-01-01

    Surface fimbriae of pathogenic Escherichia coli facilitate sensing, adhesion and even invasion of host epithelial cells. While it is known that the pathogen has the potential to express a plethora of fimbrial variants susceptible to rapid phase ON/OFF variation, it is an open question if the fimb......Surface fimbriae of pathogenic Escherichia coli facilitate sensing, adhesion and even invasion of host epithelial cells. While it is known that the pathogen has the potential to express a plethora of fimbrial variants susceptible to rapid phase ON/OFF variation, it is an open question...... if the fimbrial diversity seen at the population level is the product of random stochasticity or a concerted effort based on active communication. Here we discuss the possibility of a mechanism alternative to a stochastic fimbrial phase variation model affecting the dynamics of a heterogeneous population....

  10. Ovary and fimbrial stem cells: biology, niche and cancer origins.

    Science.gov (United States)

    Ng, Annie; Barker, Nick

    2015-10-01

    The mammalian ovary is covered by a single-layered epithelium that undergoes rupture and remodelling following each ovulation. Although resident stem cells are presumed to be crucial for this cyclic regeneration, their identity and mode of action have been elusive. Surrogate stemness assays and in vivo fate-mapping studies using recently discovered stem cell markers have identified stem cell pools in the ovary and fimbria that ensure epithelial homeostasis. Recent findings provide insights into intrinsic mechanisms and local extrinsic cues that govern the function of ovarian and fimbrial stem cells. These discoveries have advanced our understanding of stem cell biology in the ovary and fimbria, and lay the foundations for evaluating the contribution of resident stem cells to the initiation and progression of human epithelial ovarian cancer.

  11. The exopolysaccharide gene cluster Bcam1330-Bcam1341 is involved in Burkholderia cenocepacia biofilm formation, and its expression is regulated by c-di-GMP and Bcam1349

    DEFF Research Database (Denmark)

    Fazli, Mustafa; McCarthy, Yvonne; Givskov, Michael

    2013-01-01

    In Burkholderia cenocepacia, the second messenger cyclic diguanosine monophosphate (c-di-GMP) has previously been shown to positively regulate biofilm formation and the expression of cellulose and type-I fimbriae genes through binding to the transcriptional regulator Bcam1349. Here, we provide...... evidence that cellulose and type-I fimbriae are not involved in B. cenocepacia biofilm formation in flow chambers, and we identify a novel Bcam1349/c-di-GMP-regulated exopolysaccharide gene cluster which is essential for B. cenocepacia biofilm formation. Overproduction of Bcam1349 in trans promotes wrinkly...... matrix exopolysaccharide and to be essential for flow-chamber biofilm formation. We demonstrate that Bcam1349 binds to the promoter region of genes in the Bcam1330-Bcam1341 cluster and that this binding is enhanced by the presence of c-di-GMP. Furthermore, we demonstrate that overproduction of both c-di-GMP...

  12. From Catheter to Kidney Stone: The Uropathogenic Lifestyle of Proteus mirabilis.

    Science.gov (United States)

    Norsworthy, Allison N; Pearson, Melanie M

    2017-04-01

    Proteus mirabilis is a model organism for urease-producing uropathogens. These diverse bacteria cause infection stones in the urinary tract and form crystalline biofilms on indwelling urinary catheters, frequently leading to polymicrobial infection. Recent work has elucidated how P. mirabilis causes all of these disease states. Particularly exciting is the discovery that this bacterium forms large clusters in the bladder lumen that are sites for stone formation. These clusters, and other steps of infection, require two virulence factors in particular: urease and MR/P fimbriae. Highlighting the importance of MR/P fimbriae is the cotranscribed regulator, MrpJ, which globally controls virulence. Overall, P. mirabilis exhibits an extraordinary lifestyle, and further probing will answer exciting basic microbiological and clinically relevant questions. Copyright © 2016 Elsevier Ltd. All rights reserved.

  13. Protein translation and cell death: the role of rare tRNAs in biofilm formation and in activating dormant phage killer genes.

    Directory of Open Access Journals (Sweden)

    Rodolfo García-Contreras

    2008-06-01

    Full Text Available We discovered previously that the small Escherichia coli proteins Hha (hemolysin expression modulating protein and the adjacent, poorly-characterized YbaJ are important for biofilm formation; however, their roles have been nebulous. Biofilms are intricate communities in which cell signaling often converts single cells into primitive tissues. Here we show that Hha decreases biofilm formation dramatically by repressing the transcription of rare codon tRNAs which serves to inhibit fimbriae production and by repressing to some extent transcription of fimbrial genes fimA and ihfA. In vivo binding studies show Hha binds to the rare codon tRNAs argU, ileX, ileY, and proL and to two prophage clusters D1P12 and CP4-57. Real-time PCR corroborated that Hha represses argU and proL, and Hha type I fimbriae repression is abolished by the addition of extra copies of argU, ileY, and proL. The repression of transcription of rare codon tRNAs by Hha also leads to cell lysis and biofilm dispersal due to activation of prophage lytic genes rzpD, yfjZ, appY, and alpA and due to induction of ClpP/ClpX proteases which activate toxins by degrading antitoxins. YbaJ serves to mediate the toxicity of Hha. Hence, we have identified that a single protein (Hha can control biofilm formation by limiting fimbriae production as well as by controlling cell death. The mechanism used by Hha is the control of translation via the availability of rare codon tRNAs which reduces fimbriae production and activates prophage lytic genes. Therefore, Hha acts as a toxin in conjunction with co-transcribed YbaJ (TomB that attenuates Hha toxicity.

  14. Presence and Characterization of Extraintestinal Pathogenic Escherichia coli Virulence Genes in F165-Positive E. coli Strains Isolated from Diseased Calves and Pigs

    OpenAIRE

    Dezfulian, Hojabr; Batisson, Isabelle; Fairbrother, John M.; Lau, Peter C. K.; Nassar, Atef; Szatmari, George; Harel, Josée

    2003-01-01

    The virulence genotype profile and presence of a pathogenicity island(s) (PAI) were studied in 18 strains of F165-positive Escherichia coli originally isolated from diseased calves or piglets. On the basis of their adhesion phenotypes and genotypes, these extraintestinal pathogenic strains were classified into three groups. The F165 fimbrial complex consists of at least two serologically and genetically distinct fimbriae: F1651 and F1652. F1651 is encoded by the foo operon (pap-like), and F16...

  15. A Structural Basis for Sustained Bacterial Adhesion – Biomechanical Properties of CFA/I Pili

    OpenAIRE

    Andersson, Magnus; Björnham, Oscar; Svantesson, Mats; Badahdah, Arwa; Uhlin, Bernt Eric; Bullitt, Esther

    2011-01-01

    Enterotoxigenic Escherichia coli (ETEC) are a major cause of diarrheal disease worldwide. Adhesion pili (or fimbriae), such as the CFA/I (colonization factor antigen I) organelles that enable ETEC to attach efficiently to the host intestinal tract epithelium, are critical virulence factors for initiation of infection. We characterized at single organelle level the intrinsic biomechanical properties and kinetics of individual CFA/I pili, demonstrating that weak external forces (7.5 pN) are suf...

  16. Identification and Whole Genome Sequencing of the First Case of Kosakonia radicincitans Causing a Human Bloodstream Infection

    OpenAIRE

    Bhatti, Micah D.; Kalia, Awdhesh; Sahasrabhojane, Pranoti; Kim, Jiwoong; Greenberg, David E.; Shelburne, Samuel A.

    2017-01-01

    The taxonomy of Enterobacter species is rapidly changing. Herein we report a bloodstream infection isolate originally identified as Enterobacter cloacae by Vitek2 methodology that we found to be Kosakonia radicincitans using genetic means. Comparative whole genome sequencing of our isolate and other published Kosakonia genomes revealed these organisms lack the AmpC β-lactamase present on the chromosome of Enterobacter sp. A fimbriae operon primarily found in Escherichia coli O157:H7 isolates ...

  17. Role of yqiC in the pathogenicity of Salmonella and innate immune responses of human intestinal epithelium

    Directory of Open Access Journals (Sweden)

    Ke-Chuan Wang

    2016-10-01

    Full Text Available The yqiC gene of Salmonella enterica serovar Typhimurium (S. Typhimurium regulates bacterial growth at different temperatures and mice survival after infection. However, the role of yqiC in bacterial colonization and host immunity remains unknown. We infected human LS174T, Caco-2, HeLa, and THP-1 cells with S. Typhimurium wild-type SL1344, its yqiC mutant, and its complemented strain. Bacterial colonization and internalization in the four cell lines significantly reduced on yqiC depletion. Postinfection production of interleukin-8 and human β-defensin-3 in LS174T cells significantly reduced because of yqiC deleted in S. Typhimurium. The phenotype of yqiC mutant exhibited few and short flagella, fimbriae on the cell surface, enhanced biofilm formation, upregulated type-1 fimbriae expression, and reduced bacterial motility. Type-1 fimbriae, flagella, SPI-1, and SPI-2 gene expression was quantified using real-time PCR. The data show that deletion of yqiC upregulated fimA and fimZ expression and downregulated flhD, fliZ, invA, and sseB expression. Furthermore, thin-layer chromatography and high-performance liquid chromatography revealed the absence of menaquinone in the yqiC mutant, thus validating the importance of yqiC in the bacterial electron transport chain. Therefore, YqiC can negatively regulate FimZ for type-1 fimbriae expression and manipulate the functions of its downstream virulence factors including flagella, SPI-1, and SPI-2 effectors.

  18. High-throughput sequencing reveals key genes and immune homeostatic pathways activated in myeloid dendritic cells by Porphyromonas gingivalis 381 and its fimbrial mutants.

    Science.gov (United States)

    Arjunan, P; El-Awady, A; Dannebaum, R O; Kunde-Ramamoorthy, G; Cutler, C W

    2016-02-01

    The human microbiome consists of highly diverse microbial communities that colonize our skin and mucosal surfaces, aiding in maintenance of immune homeostasis. The keystone pathogen Porphyromonas gingivalis induces a dysbiosis and disrupts immune homeostasis through as yet unclear mechanisms. The fimbrial adhesins of P. gingivalis facilitate biofilm formation, invasion of and dissemination by blood dendritic cells; hence, fimbriae may be key factors in disruption of immune homeostasis. In this study we employed RNA-sequencing transcriptome profiling to identify differentially expressed genes (DEGs) in human monocyte-derived dendritic cells (MoDCs) in response to in vitro infection/exposure by Pg381 or its isogenic mutant strains that solely express minor-Mfa1 fimbriae (DPG3), major-FimA fimbriae (MFI) or are deficient in both fimbriae (MFB) relative to uninfected control. Our results yielded a total of 479 DEGs that were at least two-fold upregulated and downregulated in MoDCs significantly (P ≤ 0.05) by all four strains and certain DEGs that were strain-specific. Interestingly, the gene ontology biological and functional analysis shows that the upregulated genes in DPG3-induced MoDCs were more significant than other strains and associated with inflammation, immune response, anti-apoptosis, cell proliferation, and other homeostatic functions. Both transcriptome and quantitative polymerase chain reaction results show that DPG3, which solely expresses Mfa1, increased ZNF366, CD209, LOX1, IDO1, IL-10, CCL2, SOCS3, STAT3 and FOXO1 gene expression. In conclusion, we have identified key DC-mediated immune homeostatic pathways that could contribute to dysbiosis in periodontal infection with P. gingivalis. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  19. Cloning, purification, crystallization and preliminary X-ray diffraction studies of Escherichia coli PapD-like protein (EcpD)

    International Nuclear Information System (INIS)

    Pandey, Nishant Kumar; Pal, Ravi Kant; Kashyap, Maruthi; Bhavesh, Neel Sarovar

    2012-01-01

    The Escherichia coli PapD-like protein (EcpD), from uropathogenic Escherichia coli (UPEC), which is a periplasmic chaperon of Yad fimbriae was cloned, overexpressed, purified and crystallized. The crystals obtained diffracted X-rays to 1.67 Å resolution and belonged to space group C222 1 . Many Gram-negative bacteria are characterized by hair-like proteinaceous appendages on their surface known as fimbriae. In uropathogenic strains of Escherichia coli, fimbriae mediate attachment by binding to receptors on the host cell, often contributing to virulence and disease. E. coli PapD-like protein (EcpD) is a periplasmic chaperone that plays an important role in the proper folding and guiding of Yad fimbrial proteins to the outer membrane usher protein in a process known as pilus biogenesis. EcpD is essential for pilus biogenesis in uropathogenic E. coli and plays an important role in virulence. In the present study, EcpD was cloned, overexpressed, purified and crystallized by the hanging-drop vapour-diffusion method. The crystals diffracted to 1.67 Å resolution and belonged to the orthorhombic space group C222 1 , with unit-cell parameters a = 100.3, b = 127.6, c = 45.9 Å. There was a single molecule in the asymmetric unit and the corresponding Matthews coefficient was calculated to be 3.02 Å 3 Da −1 , with 59% solvent content. Initial phases were determined by molecular replacement

  20. Virulence Factors and Antibiotic Resistance in Uropathogenic and Commensal Escherichia coli Isolates

    Directory of Open Access Journals (Sweden)

    Iraj Sedighi

    2016-10-01

    Full Text Available Background: Urinary Tract Infections (UTIs, including cystitis and pyelonephritis, are the most common infectious diseases in childhood. Aim and Objectives: Escherichia coli (E. coli account for as much as 90% of the community-acquired and also 50% of nosocomial UTIs. Therefore, the identification of E. coli strains and antibiotic resistance patterns is important for both clinical and epidemiological implications. Material and Methods: To characterize uropathogenic strains E. coli, we studied 100 strains recovered from both urine samples of children aged less than 7 years with community-acquired UTIs and stool samples of healthy children, respectively. Results: We assessed Virulence Factors (VFs and drug sensitivities of E. coli isolates. Drug sensitivities of the isolates were 94% (amikacin, 90% (nitrofurantoin, 66% (gentamicin, 56% (cefixime, 40% (nalidixic acid and 28% (cotrimoxazol. Laboratory tests showed that the prevalence of virulence factors ranged from 18% for hemolysin and P-fimbriae to 2% for type1-fimbriae. Most drug resistance was cotrimoxazole and amikacin was the lowest. P-fimbriae and hemolysin in uropathogenic E. coli were more frequent than non-pathogen type of E. coli. Conclusion: Although amikacin appeared to be the first choice for UTI in children, but nitrofurantoin seems to be practical and could be considered as the selective choice for uncomplicated lower UTIs.

  1. Invasion of Human Oral Epithelial Cells by Prevotella intermedia

    Science.gov (United States)

    Dorn, Brian R.; Leung, K.-P.; Progulske-Fox, Ann

    1998-01-01

    Invasion of oral epithelial cells by pathogenic oral bacteria may represent an important virulence factor in the progression of periodontal disease. Here we report that a clinical isolate of Prevotella intermedia, strain 17, was found to invade a human oral epithelial cell line (KB), whereas P. intermedia 27, another clinical isolate, and P. intermedia 25611, the type strain, were not found to invade the cell line. Invasion was quantified by the recovery of viable bacteria following a standard antibiotic protection assay and observed by electron microscopy. Cytochalasin D, cycloheximide, monodansylcadaverine, and low temperature (4°C) inhibited the internalization of P. intermedia 17. Antibodies raised against P. intermedia type C fimbriae and against whole cells inhibited invasion, but the anti-type-C-fimbria antibody inhibited invasion to a greater extent than the anti-whole-cell antibody. This work provides evidence that at least one strain of P. intermedia can invade an oral epithelial cell line and that the type C fimbriae and a cytoskeletal rearrangement are required for this invasion. PMID:9826397

  2. Correlation of clinical data with fallopian tube specimen immune cells and tissue culture capacity.

    Science.gov (United States)

    Ramraj, Satish Kumar; Smith, Katie M; Janakiram, Naveena B; Toal, Coralee; Raman, Ankita; Benbrook, Doris Mangiaracina

    2018-06-01

    Human fallopian tube fimbria secretory epithelial cells (hFTSECs) are considered an origin of ovarian cancer and methods for their culture from fallopian tube specimens have been reported. Our objective was to determine whether characteristics of the donors or surgeries were associated with the capacities of fimbria specimens to generate hFTSEC cultures or their immune profiles. There were no surgical complications attributable to fallopian tube removal. Attempts to establish primary hFTSEC cultures were successful in 37 of 55 specimens (67%). Success rates did not differ significantly between specimens grouped by patient or surgery characteristics. Established cultures could be revived after cryopreservation and none became contaminated with microorganisms. Two cultures evaluated for long term growth senesced between passages 10 and 15. M1 macrophages were the predominant cell type, while all other immune cells were present at much lower percentages. IL-10 and TGF-β exhibited opposing trends with M1 and M2 macrophages. Plasma IL-10 levels exhibited significant positive correlation with patient age. In conclusion, fallopian tube fimbria specimens exhibit a pro-inflammatory phenotype and can be used to provide a source of hFTSECs that can be cultured for a limited time regardless of the donor patient age or race, or the type of surgery performed. Copyright © 2018 Elsevier Ltd. All rights reserved.

  3. A low-pH medium in vitro or the environment within a macrophage decreases the transcriptional levels of fimA, fimZ and lrp in Salmonella enterica serovar Typhimurium.

    Science.gov (United States)

    Wang, Ke-Chuan; Hsu, Yuan-Hsun; Huang, Yi-Ning; Chen, Ter-Hsin; Lin, Jiunn-Horng; Hsuan, Shih-Ling; Chien, Maw-Sheng; Lee, Wei-Cheng; Yeh, Kuang-Sheng

    2013-09-01

    Many Salmonella Typhimurium isolates produce type 1 fimbriae and exhibit fimbrial phase variation in vitro. Static broth culture favours the production of fimbriae, while solid agar medium inhibits the generation of these appendages. Little information is available regarding whether S. Typhimurium continues to produce type 1 fimbriae during in vivo growth. We used a type 1 fimbrial phase-variable strain S. Typhimurium LB5010 and its derivatives to infect RAW 264.7 macrophages. Following entry into macrophages, S. Typhimurium LB5010 gradually decreased the transcript levels of fimbrial subunit gene fimA, positive regulatory gene fimZ, and global regulatory gene lrp. A similar decrease in transcript levels was detected by RT-PCRwhen the pH of static brothmediumwas shifted frompH 7 to amore acidic pH 4. A fimA-deleted strain continued to multiply within macrophages as did the parental strain. An lrp deletion strain was unimpaired for in vitro growth at pH 7 or pH 4, while a strain harboring an lrp-containing plasmid exhibited impaired in vitro growth at pH 4. We propose that acidic medium, which resembles one aspect of the intracellular environment in a macrophage, inhibits type 1 fimbrial production by down-regulation of the expression of lrp, fimZ and fimA.

  4. Stress tolerant virulent strains of Cronobacter sakazakii from food

    Directory of Open Access Journals (Sweden)

    Md Fakruddin

    2014-01-01

    Full Text Available BACKGROUND: Cronobacter sakazakii is considered as an emerging foodborne pathogen. The aim of this study was to isolate and characterize virulent strains of Cronobacter sakazakii from food samples of Bangladesh. RESULT: Six (6 Cronobacter sakazakii was isolated and identified from 54 food samples on the basis of biochemical characteristics, sugar fermentation, SDS-PAGE of whole cell protein, plasmid profile and PCR of Cronobacter spp. specific genes (esak, gluA, zpx, ompA, ERIC, BOX-AIR and sequencing. These strains were found to have moderately high antibiotic resistance against common antibiotics and some are ESBL producer. Most of the C. sakazakii isolates were capable of producing biofilm (strong biofilm producer, extracellular protease and siderophores, curli expression, haemolysin, haemagglutinin, mannose resistant haemagglutinin, had high cell surface hydrophobicity, significant resistance to human serum, can tolerate high concentration of salt, bile and DNase production. Most of them produced enterotoxins of different molecular weight. The isolates pose significant serological cross-reactivity with other gram negative pathogens such as serotypes of Salmonella spp., Shigella boydii, Shigella sonnei, Shigella flexneri and Vibrio cholerae. They had significant tolerance to high temperature, low pH, dryness and osmotic stress. CONCLUSION: Special attention should be given in ensuring hygiene in production and post-processing to prevent contamination of food with such stress-tolerant virulent Cronobacter sakazakii.

  5. Porphyromonas gingivalis evasion of autophagy and intracellular killing by human myeloid dendritic cells involves DC-SIGN-TLR2 crosstalk.

    Science.gov (United States)

    El-Awady, Ahmed R; Miles, Brodie; Scisci, Elizabeth; Kurago, Zoya B; Palani, Chithra D; Arce, Roger M; Waller, Jennifer L; Genco, Caroline A; Slocum, Connie; Manning, Matthew; Schoenlein, Patricia V; Cutler, Christopher W

    2015-02-01

    Signaling via pattern recognition receptors (PRRs) expressed on professional antigen presenting cells, such as dendritic cells (DCs), is crucial to the fate of engulfed microbes. Among the many PRRs expressed by DCs are Toll-like receptors (TLRs) and C-type lectins such as DC-SIGN. DC-SIGN is targeted by several major human pathogens for immune-evasion, although its role in intracellular routing of pathogens to autophagosomes is poorly understood. Here we examined the role of DC-SIGN and TLRs in evasion of autophagy and survival of Porphyromonas gingivalis in human monocyte-derived DCs (MoDCs). We employed a panel of P. gingivalis isogenic fimbriae deficient strains with defined defects in Mfa-1 fimbriae, a DC-SIGN ligand, and FimA fimbriae, a TLR2 agonist. Our results show that DC-SIGN dependent uptake of Mfa1+P. gingivalis strains by MoDCs resulted in lower intracellular killing and higher intracellular content of P. gingivalis. Moreover, Mfa1+P. gingivalis was mostly contained within single membrane vesicles, where it survived intracellularly. Survival was decreased by activation of TLR2 and/or autophagy. Mfa1+P. gingivalis strain did not induce significant levels of Rab5, LC3-II, and LAMP1. In contrast, P. gingivalis uptake through a DC-SIGN independent manner was associated with early endosomal routing through Rab5, increased LC3-II and LAMP-1, as well as the formation of double membrane intracellular phagophores, a characteristic feature of autophagy. These results suggest that selective engagement of DC-SIGN by Mfa-1+P. gingivalis promotes evasion of antibacterial autophagy and lysosome fusion, resulting in intracellular persistence in myeloid DCs; however TLR2 activation can overcome autophagy evasion and pathogen persistence in DCs.

  6. F4+ enterotoxigenic Escherichia coli (ETEC) adhesion mediated by the major fimbrial subunit FaeG.

    Science.gov (United States)

    Xia, Pengpeng; Song, Yujie; Zou, Yajie; Yang, Ying; Zhu, Guoqiang

    2015-09-01

    The FaeG subunit is the major constituent of F4(+) fimbriae, associated with glycoprotein and/or glycolipid receptor recognition and majorly contributes to the pathogen attachment to the host cells. To investigate the key factor involved in the fimbrial binding of F4(+) Escherichia coli, both the recombinant E. coli SE5000 strains carrying the fae operon gene clusters that express the different types of fimbriae in vitro, named as rF4ab, rF4ac, and rF4ad, respectively, corresponding to the fimbrial types F4ab, F4ac, and F4ad, and the three isogenic in-frame faeG gene deletion mutants were constructed. The adhesion assays and adhesion inhibition assays showed that ΔfaeG mutants had a significant reduction in the binding to porcine brush border as well as the intestinal epithelial cell lines, while the complemented strain ΔfaeG/pfaeG restored the adhesion function. The recombinant bacterial strains rF4ab, rF4ac, and rF4ad have the same binding property as wild-type F4(+) E. coli strains do and improvement in terms of binding to porcine brush border and the intestinal epithelial cells, and the adherence was blocked by the monoclonal antibody anti-F4 fimbriae. These data demonstrate that the fimbrial binding of F4(+) E. coli is directly mediated by the major FaeG subunit. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. The impact of virulence factors of Porphyromonas gingivalis on wound healing in vitro

    Directory of Open Access Journals (Sweden)

    Alexa M. G. A. Laheij

    2015-07-01

    Full Text Available Background: Porphyromonas gingivalis inhibits oral epithelial wound healing in vitro more strongly than other oral bacteria, but it is unknown why P. gingivalis is such a potent inhibitor of wound healing. Objective: Therefore, the aim of this study was to investigate the influence of major virulence factors of P. gingivalis on wound healing in an in vitro wound-healing model. The influence of the capsular polysaccharide, the Arg- and Lys- gingipains, the major fimbriae and lipopolysaccharide (LPS was investigated. Design: A standardized scratch was made in a confluent layer of human oral epithelial cells HO-1-N-1. The epithelial cells were then challenged with different concentrations of several P. gingivalis wild-type strains and knockout mutants. Closure of the scratch was determined after 17 h and compared to control conditions without bacteria. Results: The P. gingivalis strains ATCC 33277, W83, and W50 significantly inhibited wound healing. The presence of a capsular polysaccharide lowered significantly the inhibition of epithelial cell migration, while gingipain activity significantly increased the inhibition of cell migration. LPS and the major fimbriae did not influence epithelial cell migration. None of the tested P. gingivalis strains completely prevented the inhibition of cell migration, suggesting that other characteristics of P. gingivalis also play a role in the inhibition of wound healing, and that further research is needed. Conclusions: The capsular polysaccharide and the Arg- and Lys- gingipains of P. gingivalis influenced the capacity of P. gingivalis to hinder wound healing, while LPS and the major fimbriae had no effect.

  8. New aspects of RpoE in uropathogenic Proteus mirabilis.

    Science.gov (United States)

    Liu, Ming-Che; Kuo, Kuan-Ting; Chien, Hsiung-Fei; Tsai, Yi-Lin; Liaw, Shwu-Jen

    2015-03-01

    Proteus mirabilis is a common human pathogen causing recurrent or persistent urinary tract infections (UTIs). The underlying mechanisms for P. mirabilis to establish UTIs are not fully elucidated. In this study, we showed that loss of the sigma factor E (RpoE), mediating extracytoplasmic stress responses, decreased fimbria expression, survival in macrophages, cell invasion, and colonization in mice but increased the interleukin-8 (IL-8) expression of urothelial cells and swarming motility. This is the first study to demonstrate that RpoE modulated expression of MR/P fimbriae by regulating mrpI, a gene encoding a recombinase controlling the orientation of MR/P fimbria promoter. By real-time reverse transcription-PCR, we found that the IL-8 mRNA amount of urothelial cells was induced significantly by lipopolysaccharides extracted from rpoE mutant but not from the wild type. These RpoE-associated virulence factors should be coordinately expressed to enhance the fitness of P. mirabilis in the host, including the avoidance of immune attacks. Accordingly, rpoE mutant-infected mice displayed more immune cell infiltration in bladders and kidneys during early stages of infection, and the rpoE mutant had a dramatically impaired ability of colonization. Moreover, it is noteworthy that urea (the major component in urine) and polymyxin B (a cationic antimicrobial peptide) can induce expression of rpoE by the reporter assay, suggesting that RpoE might be activated in the urinary tract. Altogether, our results indicate that RpoE is important in sensing environmental cues of the urinary tract and subsequently triggering the expression of virulence factors, which are associated with the fitness of P. mirabilis, to build up a UTI. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  9. Pathogenesis of Proteus mirabilis Infection

    Science.gov (United States)

    Armbruster, Chelsie E.; Mobley, Harry L. T.; Pearson, Melanie M.

    2017-01-01

    Proteus mirabilis, a Gram-negative rod-shaped bacterium most noted for its swarming motility and urease activity, frequently causes catheter-associated urinary tract infections (CAUTI) that are often polymicrobial. These infections may be accompanied by urolithiasis, development of bladder or kidney stones due to alkalinization of urine from urease-catalyzed urea hydrolysis. Adherence of the bacterium to epithelial and catheter surfaces is mediated by 17 different fimbriae, most notably MR/P fimbriae. Repressors of motility are often encoded by these fimbrial operons. Motility is mediated by flagella encoded on a single contiguous 54 kb chromosomal sequence. On agar plates, P. mirabilis undergoes a morphological conversion to a filamentous swarmer cell expressing hundreds of flagella. When swarms from different strains meet, a line of demarcation, a “Dienes line”, develops due to the killing action of each strain’s type VI secretion system. During infection, histological damage is caused by cytotoxins including hemolysin and a variety of proteases, some autotransported. The pathogenesis of infection, including assessment of individual genes or global screens for virulence or fitness factors has been assessed in murine models of ascending UTI or CAUTI using both single-species and polymicrobial models. Global gene expression studies carried out in culture and in the murine model have revealed the unique metabolism of this bacterium. Vaccines, using MR/P fimbria and its adhesin, MrpH, have been shown to be efficacious in the murine model. A comprehensive review of factors associated with urinary tract infection is presented, encompassing both historical perspectives and current advances. PMID:29424333

  10. Activation of the NLRP3 Inflammasome Pathway by Uropathogenic Escherichia coli Is Virulence Factor-Dependent and Influences Colonization of Bladder Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Isak Demirel

    2018-03-01

    Full Text Available The NLRP3 inflammasome and IL-1β release have recently been suggested to be important for the progression of urinary tract infection (UTI. However, much is still unknown regarding the interaction of UPEC and the NLRP3 inflammasome. The purpose of this study was to elucidate what virulence factors uropathogenic Escherichia coli (UPEC use to modulate NLRP3 inflammasome activation and subsequent IL-1β release and the role of NLRP3 for UPEC colonization of bladder epithelial cells. The bladder epithelial cell line 5637, CRISPR/Cas9 generated NLRP3, caspase-1 and mesotrypsin deficient cell lines and transformed primary bladder epithelial cells (HBLAK were stimulated with UPEC isolates and the non-pathogenic MG1655 strain. We found that the UPEC strain CFT073, but not MG1655, induced an increased caspase-1 activity and IL-1β release from bladder epithelial cells. The increase was shown to be mediated by α-hemolysin activation of the NLRP3 inflammasome in an NF-κB-independent manner. The effect of α-hemolysin on IL-1β release was biphasic, initially suppressive, later inductive. Furthermore, the phase-locked type-1-fimbrial ON variant of CFT073 inhibited caspase-1 activation and IL-1β release. In addition, the ability of CFT073 to adhere to and invade NLRP3 deficient cells was significantly reduced compare to wild-type cells. The reduced colonization of NLRP3-deficient cells was type-1 fimbriae dependent. In conclusion, we found that the NLRP3 inflammasome was important for type-1 fimbriae-dependent colonization of bladder epithelial cells and that both type-1 fimbriae and α-hemolysin can modulate the activity of the NLRP3 inflammasome.

  11. Hemagglutination and biofilm formation as virulence markers of uropathogenic Escherichia coli in acute urinary tract infections and urolithiasis

    Directory of Open Access Journals (Sweden)

    Uma B Maheswari

    2013-01-01

    Full Text Available Introduction: Urinary tract infections (UTI are a major public health concern in developing countries. Most UTIs are caused by E. coli, accounting for up to 90% of community-acquired UTIs (CAUTI. Recurrent UTI is considered as a major risk factor for urolithiasis. Virulence factors like adhesins and biofilm have been extensively studied by authors on UPEC isolated from recurrent UTI.The studies on isolates from infection stones in kidney are scanty . In a prospective study, we aimed to determine the expression of Haemagglutinins, (Type 1 and P fimbriae , Biofilm production and resistance pattern to common antibiotics of Uropathogenic E.coli (UPEC isolates from Community acquired Acute Urinary Tract Infection(CAUTI and Urolithiasis. Materials and Methods: A total of 43 UPEC isolates , 23 mid-stream urine (MSU samples from patients with CAUTI attending Out Patient Departments and 20 from renal calculi of urolithiasis patients at the time of Percutaneous nephrolithostomy (PCNL were included in the study and the expression of Haemagglutinins,(Type 1 and P fimbriae , Biofilm production and resistance pattern to common antibiotics was assessed. Results: A total of 43 UPEC isolates 23 from CAUTI and 20 from renal calculi were tested for production of biofilm and hemagglutinins. In CAUTI, biofilm producers were 56.52% and hemagglutinins were detected in all isolates 100%. In urolithiasis, biofilm producers were 100% but hemagglutinins were detected only in 70% of isolates. All isolates were resistant to multiple antibiotics used. CAUTI isolates were susceptible to 3 rd generation cephalosporins, whereas urolithiasis isolates were resistant to 3 rd generation cephalosporins and 25% were Extended Spectrum Beta Lactamases ESBL producers. Conclusions: HA mediated by type 1 fimbriae plays an important role in CAUTI (P < 0.001 highly significant, whereas, in chronic conditions like urolithiasis, biofilm plays an important role in persistence of infection and

  12. Emergence of a Clonal Lineage of Multidrug-Resistant ESBL-Producing Salmonella Infantis Transmitted from Broilers and Broiler Meat to Humans in Italy between 2011 and 2014

    DEFF Research Database (Denmark)

    Franco, Alessia; Leekitcharoenphon, Pimlapas; Feltrin, Fabiola

    2015-01-01

    We report the spread of a clone of multidrug-resistant (MDR), ESBL-producing (blaCTX-M-1) Salmonella enterica subsp. enterica serovar Infantis, in the Italian broiler chicken industry and along the food-chain. This was first detected in Italy in 2011 and led to human infection in Italy in 2013....... This megaplasmid carried the ESBL gene blaCTX-M-1, and additional genes [tet(A), sul1, dfrA1 and dfrA14] mediating cefotaxime, tetracycline, sulfonamide, and trimethoprim resistance. It also contained genes conferring enhanced colonization capability, virulence (fimbriae, yersiniabactin), resistance and fitness...

  13. Genes associados à virulência e multirresistência de antimicrobianos em linhagens Trueperella Pyogenes isoladas de mastite e outras afecções em animais domésticos

    OpenAIRE

    Risseti, Rafaela Mastrangelo [UNESP

    2015-01-01

    Trueperella pyogenes are opportunistic bacterium characterized by suppurative infections in domestic animals, commonly refractory to conventional therapy. Recently, genes which encode exotoxin pyolysin (plo), and factors that promote adhesion of T. pyogenes to host cells, such as fimbriae (fimA, fimC, fimE, fimG), neuraminidases (nanH, nanP), and collagen-binding protein (cbpA) have been associated to virulence of pathogen. The aim of present study was investigate occurrence of multi-drug res...

  14. Differentiation of F4 receptor profiles in pigs based on their mucin 4 polymorphism, responsiveness to oral F4 immunization and in vitro binding of F4 to villi.

    Science.gov (United States)

    Nguyen, V U; Goetstouwers, T; Coddens, A; Van Poucke, M; Peelman, L; Deforce, D; Melkebeek, V; Cox, E

    2013-03-15

    F4(+) enterotoxigenic Escherichia coli (F4(+) ETEC) are an important cause of diarrhoea and mortality in piglets. F4(+) ETEC use their F4 fimbriae to adhere to specific receptors (F4Rs) on small intestinal brush borders, resulting in colonization of the small intestine. To prevent pigs from post-weaning diarrhoea, pigs should be vaccinated during the suckling period. Previously, we demonstrated that F4acR(+), but not F4acR(-) piglets could be orally immunized with purified F4 fimbriae resulting in a protective immunity against F4(+) ETEC infections, indicating that this immune response was F4R dependent. Recently, aminopeptidase N has been identified as a glycoprotein receptor important for this oral immune response. However, in some oral immunization experiments, a few F4acR(+) piglets did not show an antibody response upon oral immunization, suggesting additional receptors. Therefore, the binding profile of F4 to brush border membrane (glyco)proteins was determined for pigs differing in F4-specific antibody response upon oral immunization, in in vitro adhesion of F4(+)E. coli to small intestinal villi, and in Muc4 genotype. Six groups of pigs could be identified. Only two groups positive in all three assays showed two high molecular weight (MW) glycoprotein bands (>250kDa) suggesting that these high MW bands are linked to the MUC4 susceptible genotype. The fact that these bands were absent in the MUC4 resistant group which showed a positive immune response against F4 and was positive in the adhesion test confirm that at least one or perhaps more other F4Rs exist. Interestingly, two pigs that were positive in the villous adhesion assay did not show an immune response against F4 fimbriae. This suggests that a third receptor category might exist which allows the bacteria to adhere but does not allow effective immunization with soluble F4 fimbriae. Future research will be necessary to confirm or reveal the identity of these receptors. Copyright © 2012 Elsevier B

  15. Influence of l-Leucine and l-Alanine on Lrp Regulation of foo, Coding for F1651, a Pap Homologue

    OpenAIRE

    Berthiaume, Frédéric; Crost, Cécile; Labrie, Vincent; Martin, Christine; Newman, Elaine B.; Harel, Josée

    2004-01-01

    The foo operon encodes F1651 fimbriae that belong to the P-regulatory family and are synthesized by septicemic Escherichia coli. Using an Lrp-deficient host and the lrp gene cloned under the arabinose pBAD promoter, we demonstrated that foo was transcribed proportionally to the amount of Lrp synthesized. l-Leucine and l-alanine decreased drastically the steady-state transcription of foo and modified phase variation, independently of the presence of FooI. Specific mutations in the C-terminal r...

  16. Differential decay of RNA of the CFA/I fimbrial operon and control of relative gene expression.

    OpenAIRE

    Jordi, B J; op den Camp, I E; de Haan, L A; van der Zeijst, B A; Gaastra, W

    1993-01-01

    CFA/I fimbriae on human enterotoxigenic Escherichia coli are composed of the CfaB protein, the product of the second gene of the CFA/I operon. We show here that CfaB is expressed at a higher level than other proteins of the CFA/I operon. mRNA encoding the CfaB protein is much more abundant than mRNA encoding CfaA, the first protein, together with CfaB or mRNA encoding CfaA only. Only one promoter, upstream of cfaA, is present. These data indicate that a primary transcript containing cfaA and ...

  17. Genes associated with pathogenicity of avian Escherichia coli (APEC isolated from respiratory cases of poultry Genes associados à patogenicidade de Escherichia coli patogênica para aves (APEC isoladas de frangos de corte com sintomatologia clínica respiratória

    Directory of Open Access Journals (Sweden)

    Ana C.G.P. Rocha

    2008-03-01

    Full Text Available The virulence mechanisms of avian pathogenic Escherichia coli (APEC have been continually studied and are believed to be multi-factorial. Certain properties are primarily associated with virulent samples and have been identified in avian isolates. In this study a total of 61 E. coli, isolates from chicken flocks with respiratory symptomatology, were probed by Polimerase Chain Reation (PCR for the presence of genes responsible for the adhesion capacity, P fimbria (papC e F11 fimbria (felA, colicin production (cvaC, aerobactin presence (iutA, serum resistance (iss, temperature-sensitive hemagglutinin (tsh, and presence of K1 and K5 capsular antigens (kpsII. The iss gene was detected in 73,8%, tsh in 55,7%, iutA in 45,9%, felA in 39,3%, papC in 24,3%, cvaC in 23% and kpsII in18%.Os mecanismos de virulência das amostras de Escherichia coli potencialmente patogênicas para aves (APEC têm sido continuamente estudados e acredita-se ser multifatorial. Certas propriedades são associadas primariamente a amostras virulentas e vêm sendo identificadas em amostras de E. coli isoladas de aves. Neste estudo um total de 61 amostras de E. coli, isoladas de frangos de corte com problemas respiratórios, foram testadas através da Reação em Cadeia da Polimerase (PCR, para a presença dos genes responsáveis pela capacidade de adesão, fimbria P (papC e fimbria F11 (felA, produção de colicinas (cvaC, presença de aerobactina (iutA, resistência sérica (iss, hemaglutinina temperatura sensível (tsh e presença de dos antígenos capsulares K1 e K5 (kpsII. O gene iss foi detectado em 73,8%, tsh em 55,7%, iutA em 45,9%, felA em 39,3%, papC em 24,3%, cvaC em 23% e kpsII em 18%.

  18. A 48,XXXX female with absence of ovaries.

    Science.gov (United States)

    Collen, R J; Falk, R E; Lippe, B M; Kaplan, S A

    1980-01-01

    A 16 1/2-year-old phenotypic female had primary amenorrhea, mild mental retardation, radioulnar synostosis, and other minor anomalies. Chromosome constitution of leukocytes and skin fibroblasts was 48,XXXX. Plasma levels of gonadotropins were increased, and those of estrogens, decreased consistent with ovarian failure. Laparoscopy showed a small midline uterus, and 2 fallopian tubes, and fimbriae. Neither ovaries nor gonadal streaks were seen on either side. This patient appears to represent the first instance of 48,XXXX aneuploidy with documented absence of ovaries.

  19. Glycan-functionalized diamond nanoparticles as potent E. coli anti-adhesives

    Science.gov (United States)

    Barras, Alexandre; Martin, Fernando Ariel; Bande, Omprakash; Baumann, Jean-Sébastien; Ghigo, Jean-Marc; Boukherroub, Rabah; Beloin, Christophe; Siriwardena, Aloysius; Szunerits, Sabine

    2013-02-01

    Bacterial attachment and subsequent biofilm formation on biotic surfaces or medical devices is an increasing source of infections in clinical settings. A large proportion of these biofilm-related infections are caused by Escherichia coli, a major nosocomial pathogen, in which the major adhesion factor is the FimH adhesin located at the tip of type 1 fimbriae. Inhibition of FimH-mediated adhesion has been identified as an efficient antibiotic-alternative strategy to potentially reduce E. coli-related infections. In this article we demonstrate that nanodiamond particles, covently modified with mannose moieties by a ``click'' chemistry approach, are able to efficiently inhibit E. coli type 1 fimbriae-mediated adhesion to eukaryotic cells with relative inhibitory potency (RIP) of as high as 9259 (bladder cell adhesion assay), which is unprecedented when compared with RIP values previously reported for alternate multivalent mannose-functionalized nanostructures designed to inhibit E. coli adhesion. Also remarkable is that these novel mannose-modified NDs reduce E. coli biofilm formation, a property previously not observed for multivalent glyco-nanoparticles and rarely demonstrated for other multivalent or monovalent mannose glycans. This work sets the stage for the further evaluation of these novel NDs as an anti-adhesive therapeutic strategy against E. coli-derived infections.Bacterial attachment and subsequent biofilm formation on biotic surfaces or medical devices is an increasing source of infections in clinical settings. A large proportion of these biofilm-related infections are caused by Escherichia coli, a major nosocomial pathogen, in which the major adhesion factor is the FimH adhesin located at the tip of type 1 fimbriae. Inhibition of FimH-mediated adhesion has been identified as an efficient antibiotic-alternative strategy to potentially reduce E. coli-related infections. In this article we demonstrate that nanodiamond particles, covently modified with

  20. FimH-mediated autoaggregation of Escherichia coli

    DEFF Research Database (Denmark)

    Schembri, Mark; Christiansen, G.; Klemm, Per

    2001-01-01

    Autoaggregation is a phenomenon thought to contribute to colonization of mammalian hosts by pathogenic bacteria. Type 1 fimbriae are surface organelles of Escherichia coli that mediate D-mannose-sensitive binding to various host surfaces. This binding is conferred by the minor fimbrial component...... FimH. In this study, we have used random mutagenesis to identify variants of the FimH adhesin that confer the ability of E. coli to autoaggregate and settle from liquid cultures. Three separate autoaggregating clones were identified, all of which contained multiple amino acid changes located within...

  1. Adjuvant effect of Gantrez®AN nanoparticles during oral vaccination of piglets against F4+enterotoxigenic Escherichia coli.

    Science.gov (United States)

    Vandamme, Katrien; Melkebeek, Vesna; Vesna, Melkebeek; Cox, Eric; Eric, Cox; Remon, Jean Paul; Paul, Remon Jean; Vervaet, Chris; Chris, Vervaet

    2011-02-15

    In this study, the adjuvanticity of methylvinylether-co-maleic anhydride (Gantrez(®)AN) nanoparticles (NP) was investigated in an oral immunisation experiment of pigs against F4+enterotoxigenic Escherichia coli (F4+ETEC). In addition, Wheat Germ Agglutinin (WGA)-coating of the nanoparticles was tested for enterocyte-targeting. Pigs were either vaccinated with F4 fimbriae, F4 encapsulated in Gantrez(®)AN NP, F4 encapsulated in Gantrez(®)AN NP coated with WGA or F4 fimbriae mixed with empty Gantrez(®)AN NP. Only vaccination with the combination of F4 mixed with empty Gantrez(®)AN NP improved protection against F4+ETEC infection. In addition, vaccination with this formulation also resulted in an F4-specific serum antibody response prior to F4+ETEC challenge. Encapsulation of F4 in Gantrez(®)AN NP only raised the serum antibody response after F4+ETEC challenge compared to soluble F4, but did not improve protection, whereas WGA-coating almost completely abolished the serum antibody response. These data indicate that nanoparticle effects after F4 encapsulation were of lesser importance for the adjuvant effect of Gantrez(®)AN NP, contrarily to the reactivity of the Gantrez(®)AN polymer used to prepare the nanoparticles. Copyright © 2010 Elsevier B.V. All rights reserved.

  2. Distribution of virulence determinants among antimicrobial-resistant and antimicrobial-susceptible Escherichia coli implicated in urinary tract infections.

    Science.gov (United States)

    Stephenson, Sam; Brown, P D

    2016-01-01

    Uropathogenic Escherichia coli (UPEC) rely on the correlation of virulence expression with antimicrobial resistance to persist and cause severe urinary tract infections (UTIs). We assessed the virulence pattern and prevalence among UPEC strains susceptible and resistant to multiple antimicrobial classes. A total of 174 non-duplicate UPEC strains from patients with clinically significant UTIs were analysed for susceptibility to aminoglycoside, antifolate, cephalosporin, nitrofuran and quinolone antibiotics for the production of extended-spectrum β-lactamases and for the presence of six virulence determinants encoding adhesins (afimbrial, Type 1 fimbriae, P and S-fimbriae) and toxins (cytotoxic necrotising factor and haemolysin). Relatively high resistance rates to nalidixic acid, ciprofloxacin, cephalothin and trimethoprim-sulfamethoxazole (82%, 78%, 62% and 59%, respectively) were observed. Fourteen distinct patterns were identified for the virulence determinants such as afaBC, cnfI, fimH, hylA, papEF and sfaDE. The toxin gene, cnfI (75.3%), was the second most prevalent marker to the adhesin, fimH (97.1%). The significant association of sfaDE/hylA (P < 0.01) among antimicrobial resistant and susceptible strains was also observed notwithstanding an overall greater occurrence of virulence factors among the latter. This study provides a snapshot of UPEC complexity in Jamaica and highlights the significant clonal heterogeneity among strains. Such outcomes emphasise the need for evidence-based strategies in the effective management and control of UTIs.

  3. Porphyromonas gingivalis Outer Membrane Vesicles Enter Human Epithelial Cells via an Endocytic Pathway and Are Sorted to Lysosomal Compartments ▿

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    Furuta, Nobumichi; Tsuda, Kayoko; Omori, Hiroko; Yoshimori, Tamotsu; Yoshimura, Fuminobu; Amano, Atsuo

    2009-01-01

    Porphyromonas gingivalis, a periodontal pathogen, secretes outer membrane vesicles (MVs) that contain major virulence factors, including major fimbriae and proteases termed gingipains, although it is not confirmed whether MVs enter host cells. In this study, we analyzed the mechanisms involved in the interactions of P. gingivalis MVs with human epithelial cells. Our results showed that MVs swiftly adhered to HeLa and immortalized human gingival epithelial cells in a fimbria-dependent manner and then entered via a lipid raft-dependent endocytic pathway. The intracellular MVs were subsequently routed to early endosome antigen 1-associated compartments and then were sorted to lysosomal compartments within 90 min, suggesting that intracellular MVs were ultimately degraded by the cellular digestive machinery. However, P. gingivalis MVs remained there for over 24 h and significantly induced acidified compartment formation after being taken up by the cellular digestive machinery. In addition, MV entry was shown to be mediated by a novel pathway for transmission of bacterial products into host cells, a Rac1-regulated pinocytic pathway that is independent of caveolin, dynamin, and clathrin. Our findings indicate that P. gingivalis MVs efficiently enter host cells via an endocytic pathway and survive within the endocyte organelles for an extended period, which provides better understanding of the role of MVs in the etiology of periodontitis. PMID:19651865

  4. Regulation of Expression of Uropathogenic Escherichia coli Nonfimbrial Adhesin TosA by PapB Homolog TosR in Conjunction with H-NS and Lrp

    Science.gov (United States)

    Engstrom, Michael D.

    2016-01-01

    Urinary tract infections (UTIs) are a major burden to human health. The overwhelming majority of UTIs are caused by uropathogenic Escherichia coli (UPEC) strains. Unlike some pathogens, UPEC strains do not have a fixed core set of virulence and fitness factors but do have a variety of adhesins and regulatory pathways. One such UPEC adhesin is the nonfimbrial adhesin TosA, which mediates adherence to the epithelium of the upper urinary tract. The tos operon is AT rich, resides on pathogenicity island aspV, and is not expressed under laboratory conditions. Because of this, we hypothesized that tosA expression is silenced by H-NS. Lrp, based on its prominent function in the regulation of other adhesins, is also hypothesized to contribute to tos operon regulation. Using a variety of in vitro techniques, we mapped both the tos operon promoter and TosR binding sites. We have now identified TosR as a dual regulator of the tos operon, which could control the tos operon in association with H-NS and Lrp. H-NS is a negative regulator of the tos operon, and Lrp positively regulates the tos operon. Exogenous leucine also inhibits Lrp-mediated tos operon positive regulation. In addition, TosR binds to the pap operon, which encodes another important UPEC adhesin, P fimbria. Induction of TosR synthesis reduces production of P fimbria. These studies advance our knowledge of regulation of adhesin expression associated with uropathogen colonization of a host. PMID:26755158

  5. ANIMAL ENTEROTOXIGENIC ESCHERICHIA COLI

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    Dubreuil, J. Daniel; Isaacson, Richard E.; Schifferli, Dieter M.

    2016-01-01

    Enterotoxigenic Escherichia coli (ETEC) is the most common cause of E. coli diarrhea in farm animals. ETEC are characterized by the ability to produce two types of virulence factors; adhesins that promote binding to specific enterocyte receptors for intestinal colonization and enterotoxins responsible for fluid secretion. The best-characterized adhesins are expressed in the context of fimbriae, such as the F4 (also designated K88), F5 (K99), F6 (987P), F17 and F18 fimbriae. Once established in the animal small intestine, ETEC produces enterotoxin(s) that lead to diarrhea. The enterotoxins belong to two major classes; heat-labile toxin that consist of one active and five binding subunits (LT), and heat-stable toxins that are small polypeptides (STa, STb, and EAST1). This chapter describes the disease and pathogenesis of animal ETEC, the corresponding virulence genes and protein products of these bacteria, their regulation and targets in animal hosts, as well as mechanisms of action. Furthermore, vaccines, inhibitors, probiotics and the identification of potential new targets identified by genomics are presented in the context of animal ETEC. PMID:27735786

  6. Biochemical evidence for overlapping neocortical and allocortical glutamate projections to the nucleus accumbens and rostral caudatoputamen in the rat brain

    Energy Technology Data Exchange (ETDEWEB)

    Walaas, I

    1981-01-01

    The high affinity uptake of L-glutamate has been used to investigate the origin and distribution of putative glutamate fibers in restricted parts of the rostral caudatoputamen and the nucleus accumbens of the rat brain. Ablation of the frontal cortex reduced the glutamate uptake heavily (-77%) in the dorsal part of the ipsilateral caudatoputamen, but also led to significant decreases in the ventral parts of the ipsilateral caudatoputamen (-62% and -53%) in the ipsilateral nucleus accumbens (-25% and -18%) and in the contralateral dorsal part of the caudatoputamen (-21%). Lesion of the caudal neocortex reduced the glutamate uptake in the dorsal part of the ipsilateral caudatoputamen only (-23%). Lesions of the fimbria/fornix reduced the glutamate uptake in both parts of the ipsilateral nucleus accumbens (-46% and -34%) and by approximately 20% in the whole dorsoventral extent of the anterior caudatoputamen. The results indicate that the frontal neocortex distributes fibers which may use glutamate as neurotransmitter both to the whole ipsilateral caudatoputamen and to the nucleus accumbens, and also to the dorsal parts of the contralateral caudatoputamen. The caudal neocortex probably sends such fibers to the dorsal ipsilateral caudatoputamen and the caudal allocortex sends such fibers through the fimbria/fornix to the nucleus accumbens and the ventral part of the ipsilateral caudatoputamen. The results thus corroborate previous suggestions of close similarities between the nucleus accumbens and the ventral caudatoputamen.

  7. Frequency of Fimbrial Virulence Genes (fim, pap, sfa in Escherichia Coli Isolated from the Patients with Urinary Tract Infections from selective hospitals of Tehran, Boroujerd and Sanandej City in 2015-2016

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    mohsen mirzaee

    2017-02-01

    Full Text Available Introduction: Urinary tract infection (UTI caused by E coli is one of the most common diseases in community. Colonization of E. coli and its attachment to the uroepithelium are mediated by adhesions such as P, Fim and S fimbriae. The present study was aimed to evaluate the prevalence of fimbrial virulence genes in Escherichia coli (E. coli isolates from the patients with urinary tract infection in Tehran, Boroujerd and Sanandaj cities, Iran. Methods: In this descriptive cross sectional study, 150 clinical isolates of uropathogenic E.coli were collected from the patients with urinary tract infection. All bacterial isolates were identified by standard biochemical laboratory methods; the fim, pap and sfa genes were detected using the PCR and Multiplex-PCR methods. Results: One hundred forty-five (96.66% isolates were positive for fim gene, one hundred forty (93.33% isolates were positive for pap gene and seven (4.66 isolates were positive for sfa gene. Whole of the isolates were possessed at least one of the three virulence genes. Six (4% isolates were positive for all genes. Conclusion: The findings of this study showed the high frequency of fim and P fimbriae among uropathogenic E.coli isolates from the patients with urinary tract infection. Because of the higher prevalence of UTI in the presence of these genes, detection of the genes in urine samples may help in more suspicious and rapid management of UTI.

  8. Macrophages lift off surface-bound bacteria using a filopodium-lamellipodium hook-and-shovel mechanism.

    Science.gov (United States)

    Möller, Jens; Lühmann, Tessa; Chabria, Mamta; Hall, Heike; Vogel, Viola

    2013-10-07

    To clear pathogens from host tissues or biomaterial surfaces, phagocytes have to break the adhesive bacteria-substrate interactions. Here we analysed the mechanobiological process that enables macrophages to lift-off and phagocytose surface-bound Escherichia coli (E. coli). In this opsonin-independent process, macrophage filopodia hold on to the E. coli fimbriae long enough to induce a local protrusion of a lamellipodium. Specific contacts between the macrophage and E. coli are formed via the glycoprotein CD48 on filopodia and the adhesin FimH on type 1 fimbriae (hook). We show that bacterial detachment from surfaces occurrs after a lamellipodium has protruded underneath the bacterium (shovel), thereby breaking the multiple bacterium-surface interactions. After lift-off, the bacterium is engulfed by a phagocytic cup. Force activated catch bonds enable the long-term survival of the filopodium-fimbrium interactions while soluble mannose inhibitors and CD48 antibodies suppress the contact formation and thereby inhibit subsequent E. coli phagocytosis.

  9. Ongoing Horizontal and Vertical Transmission of Virulence Genes and papA Alleles among Escherichia coli Blood Isolates from Patients with Diverse-Source Bacteremia

    Science.gov (United States)

    Johnson, James R.; O'Bryan, Timothy T.; Kuskowski, Michael; Maslow, Joel N.

    2001-01-01

    The phylogenetic distributions of multiple putative virulence factors (VFs) and papA (P fimbrial structural subunit) alleles among 182 Escherichia coli blood isolates from patients with diverse-source bacteremia were defined. Phylogenetic correspondence among these strains, the E. coli Reference (ECOR) collection, and other collections of extraintestinal pathogenic E. coli (ExPEC) was assessed. Although among the 182 bacteremia isolates phylogenetic group B2 predominated, exhibited the greatest concentration of individual VFs, and contained the largest number of familiar virulent clones, other phylogenetic groups exhibited greater concentrations of certain VFs than did group B2 and included several additional virulent clones. Certain of the newly detected VF genes, e.g., fyuA (yersiniabactin; 76%) and focG (F1C fimbriae; 25%), were as prevalent or more prevalent than their more familiar traditional counterparts, e.g., iut (aerobactin; 57%) and sfaS (S fimbriae; 14%), thus possibly offering additional useful targets for preventive interventions. Considerable diversity of VF profiles was observed at every level within the phylogenetic tree, including even within individual lineages. This suggested that many different pathways can lead to extraintestinal virulence in E. coli and that the evolution of ExPEC, which involves extensive horizontal transmission of VFs and continuous remodeling of pathogenicity-associated islands, is a highly active, ongoing process. PMID:11500406

  10. STICS, SCOUTs and p53 signatures; a new language for pelvic serous carcinogenesis.

    Science.gov (United States)

    Mehra, Karishma; Mehrad, Mitra; Ning, Geng; Drapkin, Ronny; McKeon, Frank D; Xian, Wa; Crum, Christopher P

    2011-01-01

    The events leading to the most common and most lethal ovarian carcinoma - high grade serous carcinoma - have been poorly understood. However, the detailed pathologic study of asymptomatic women with germ-line BRCA 1 or BRCA2 (BCRA+) mutations has unearthed an early malignancy, serous tubal intraepithelial carcinomas (STIC), which has linked many peritoneal and ovarian serous carcinomas to the fimbria. The distinction between high-grade serous and endometrioid carcinomas continues to narrow, with shared alterations in expression of pTEN, PAX2 and p53. Moreover, the discovery of clonal alterations in p53 in benign tubal epithelium, - p53 signatures - has established a foundation for a serous cancer precursor in the fimbria. We have expanded this concept to include a generic secretory cell outgrowth (SCOUT) in the fallopian tube that is associated with altered PAX2 expression. As the repertoire of gene alterations is expanded and its link to serous carcinogenesis clarified, a cogent pathway to high-grade Mullerian carcinomas will emerge. This will challenge conventional thinking about ovarian carcinogenesis but will provide a new template for studies of ovarian cancer prevention.

  11. Distinct temporal and anatomical distributions of amyloid-β and tau abnormalities following controlled cortical impact in transgenic mice.

    Directory of Open Access Journals (Sweden)

    Hien T Tran

    Full Text Available Traumatic brain injury (TBI is a major environmental risk factor for Alzheimer's disease. Intracellular accumulations of amyloid-β and tau proteins have been observed within hours following severe TBI in humans. Similar abnormalities have been recapitulated in young 3xTg-AD mice subjected to the controlled cortical impact model (CCI of TBI and sacrificed at 24 h and 7 days post injury. This study investigated the temporal and anatomical distributions of amyloid-β and tau abnormalities from 1 h to 24 h post injury in the same model. Intra-axonal amyloid-β accumulation in the fimbria was detected as early as 1 hour and increased monotonically over 24 hours following injury. Tau immunoreactivity in the fimbria and amygdala had a biphasic time course with peaks at 1 hour and 24 hours, while tau immunoreactivity in the contralateral CA1 rose in a delayed fashion starting at 12 hours after injury. Furthermore, rapid intra-axonal amyloid-β accumulation was similarly observed post controlled cortical injury in APP/PS1 mice, another transgenic Alzheimer's disease mouse model. Acute increases in total and phospho-tau immunoreactivity were also evident in single transgenic Tau(P301L mice subjected to controlled cortical injury. These data provide further evidence for the causal effects of moderately severe contusional TBI on acceleration of acute Alzheimer-related abnormalities and the independent relationship between amyloid-β and tau in this setting.

  12. Molecular simulations of lactose-bound and unbound forms of the FaeG adhesin reveal critical amino acids involved in sugar binding.

    Science.gov (United States)

    Baker, Joseph L; Jafri, Heba

    2016-11-01

    F4 fimbriae are protein filaments found in enterotoxigenic Escherichia coli cells and are implicated in the process of bacterial infection due to their function as bacterial adhesins. These filaments are comprised from several proteins, but the bacterial adhesin FaeG, which is a lactose-binding protein, is the major subunit comprising F4 fimbriae. Crystal structures for three variants of the FaeG protein were recently solved, including the ad variant of FaeG that was crystallized in complex with lactose. However, the dynamics of the FaeG protein bound to lactose have not been explored previously using molecular dynamics simulations. Therefore, in order to study the dynamical interactions between the FaeG ad variant and lactose, we have carried out the first all-atom molecular dynamics simulations of this system. We have also probed the role of crystallographic water molecules on the stability of lactose in the FaeG binding site, and have simulated seven FaeG mutants to probe the influence of amino acid substitutions on the ability of FaeG to bind lactose effectively. Our simulations agree well with experimental results for the influence of mutations on lactose binding, provide dynamical insights into the interactions of FaeG with lactose, and also suggest the possibility of additional regions of the FaeG protein that may act as secondary lactose binding sites. Copyright © 2016 Elsevier Inc. All rights reserved.

  13. A Constitutively Mannose-Sensitive Agglutinating Salmonella enterica subsp. enterica Serovar Typhimurium Strain, Carrying a Transposon in the Fimbrial Usher Gene stbC, Exhibits Multidrug Resistance and Flagellated Phenotypes

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    Kuan-Hsun Wu

    2012-01-01

    Full Text Available Static broth culture favors Salmonella enterica subsp. enterica serovar Typhimurium to produce type 1 fimbriae, while solid agar inhibits its expression. A transposon inserted in stbC, which would encode an usher for Stb fimbriae of a non-flagellar Salmonella enterica subsp. enterica serovar Typhimurium LB5010 strain, conferred it to agglutinate yeast cells on both cultures. RT-PCR revealed that the expression of the fimbrial subunit gene fimA, and fimZ, a regulatory gene of fimA, were both increased in the stbC mutant when grown on LB agar; fimW, a repressor gene of fimA, exhibited lower expression. Flagella were observed in the stbC mutant and this phenotype was correlated with the motile phenotype. Microarray data and RT-PCR indicated that the expression of three genes, motA, motB, and cheM, was enhanced in the stbC mutant. The stbC mutant was resistant to several antibiotics, consistent with the finding that expression of yhcQ and ramA was enhanced. A complementation test revealed that transforming a recombinant plasmid possessing the stbC restored the mannose-sensitive agglutination phenotype to the stbC mutant much as that in the parental Salmonella enterica subsp. enterica serovar Typhimurium LB5010 strain, indicating the possibility of an interplay of different fimbrial systems in coordinating their expression.

  14. Subtypes of GABAergic neurons project axons in the neocortex

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    Shigeyoshi Higo

    2009-11-01

    Full Text Available γ-aminobutyric acid (GABAergic neurons in the neocortex have been regarded as interneurons and speculated to modulate the activity of neurons locally. Recently, however, several experiments revealed that neuronal nitric oxide synthase (nNOS-positive GABAergic neurons project cortico-cortically with long axons. In this study, we illustrate Golgi-like images of the nNOS-positive GABAergic neurons using a nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d reaction and follow the emanating axon branches in cat brain sections. These axon branches projected cortico-cortically with other non-labeled arcuate fibers, contra-laterally via the corpus callosum and anterior commissure. The labeled fibers were not limited to the neocortex but found also in the fimbria of the hippocampus. In order to have additional information on these GABAergic neuron projections, we investigated green fluorescent protein (GFP-labeled GABAergic neurons in GAD67-Cre knock-in / GFP Cre-reporter mice. GFP-labeled axons emanate densely, especially in the fimbria, a small number in the anterior commissure, and very sparsely in the corpus callosum. These two different approaches confirm that not only nNOS-positive GABAergic neurons but also other subtypes of GABAergic neurons project long axons in the cerebral cortex and are in a position to be involved in information processing.

  15. Native flagellin does not protect mice against an experimental Proteus mirabilis ascending urinary tract infection and neutralizes the protective effect of MrpA fimbrial protein.

    Science.gov (United States)

    Scavone, Paola; Umpiérrez, Ana; Rial, Analía; Chabalgoity, José A; Zunino, Pablo

    2014-06-01

    Proteus mirabilis expresses several virulence factors including MR/P fimbriae and flagella. Bacterial flagellin has frequently shown interesting adjuvant and protective properties in vaccine formulations. However, native P. mirabilis flagellin has not been analyzed so far. Native P. mirabilis flagellin was evaluated as a protective antigen and as an adjuvant in co-immunizations with MrpA (structural subunit of MR/P fimbriae) using an ascending UTI model in the mouse. Four groups of mice were intranasally treated with either MrpA, native flagellin, both proteins and PBS. Urine and blood samples were collected before and after immunization for specific antibodies determination. Cytokine production was assessed in immunized mice splenocytes cultures. Mice were challenged with P. mirabilis, and bacteria quantified in kidneys and bladders. MrpA immunization induced serum and urine specific anti-MrpA antibodies while MrpA coadministered with native flagellin did not. None of the animals developed significant anti-flagellin antibodies. Only MrpA-immunized mice showed a significant decrease of P. mirabilis in bladders and kidneys. Instead, infection levels in MrpA-flagellin or flagellin-treated mice showed no significant differences with the control group. IL-10 was significantly induced in splenocytes of mice that received native flagellin or MrpA-flagellin. Native P. mirabilis flagellin did not protect mice against an ascending UTI. Moreover, it showed an immunomodulatory effect, neutralizing the protective role of MrpA. P. mirabilis flagellin exhibits particular immunological properties compared to other bacterial flagellins.

  16. E. coli Nissle 1917 Affects Salmonella adhesion to porcine intestinal epithelial cells.

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    Peter Schierack

    Full Text Available BACKGROUND: The probiotic Escherichia coli strain Nissle 1917 (EcN has been shown to interfere in a human in vitro model with the invasion of several bacterial pathogens into epithelial cells, but the underlying molecular mechanisms are not known. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we investigated the inhibitory effects of EcN on Salmonella Typhimurium invasion of porcine intestinal epithelial cells, focusing on EcN effects on the various stages of Salmonella infection including intracellular and extracellular Salmonella growth rates, virulence gene regulation, and adhesion. We show that EcN affects the initial Salmonella invasion steps by modulating Salmonella virulence gene regulation and Salmonella SiiE-mediated adhesion, but not extra- and intracellular Salmonella growth. However, the inhibitory activity of EcN against Salmonella invasion always correlated with EcN adhesion capacities. EcN mutants defective in the expression of F1C fimbriae and flagellae were less adherent and less inhibitory toward Salmonella invasion. Another E. coli strain expressing F1C fimbriae was also adherent to IPEC-J2 cells, and was similarly inhibitory against Salmonella invasion like EcN. CONCLUSIONS: We propose that EcN affects Salmonella adhesion through secretory components. This mechanism appears to be common to many E. coli strains, with strong adherence being a prerequisite for an effective reduction of SiiE-mediated Salmonella adhesion.

  17. Essential Oils and Eugenols Inhibit Biofilm Formation and the Virulence of Escherichia coli O157:H7

    Science.gov (United States)

    Kim, Yong-Guy; Lee, Jin-Hyung; Gwon, Giyeon; Kim, Soon-Il; Park, Jae Gyu; Lee, Jintae

    2016-01-01

    Enterohemorrhagic Escherichia coli O157:H7 (EHEC) has caused foodborne outbreaks worldwide and the bacterium forms antimicrobial-tolerant biofilms. We investigated the abilities of various plant essential oils and their components to inhibit biofilm formation by EHEC. Bay, clove, pimento berry oils and their major common constituent eugenol at 0.005% (v/v) were found to markedly inhibit EHEC biofilm formation without affecting planktonic cell growth. In addition, three other eugenol derivatives isoeugenol, 2-methoxy-4-propylphenol, and 4-ethylguaiacol had antibiofilm activity, indicating that the C-1 hydroxyl unit, the C-2 methoxy unit, and C-4 alkyl or alkane chain on the benzene ring of eugenol play important roles in antibiofilm activity. Interestingly, these essential oils and eugenol did not inhibit biofilm formation by three laboratory E. coli K-12 strains that reduced curli fimbriae production. Transcriptional analysis showed that eugenol down-regulated 17 of 28 genes analysed, including curli genes (csgABDFG), type I fimbriae genes (fimCDH) and ler-controlled toxin genes (espD, escJ, escR, and tir), which are required for biofilm formation and the attachment and effacement phenotype. In addition, biocompatible poly(lactic-co-glycolic acid) coatings containing clove oil or eugenol exhibited efficient biofilm inhibition on solid surfaces. In a Caenorhabditis elegans nematode model, clove oil and eugenol attenuated the virulence of EHEC. PMID:27808174

  18. RDAR morphotype: A resting stage of some Enteroba cteriaceae

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    Milanov Dubravka S.

    2015-01-01

    Full Text Available Bacteria of the genus Salmonella, some Escherichia coli strains as well as other species of the family Enterobacteriaceae, manifest the ability to express a specific phenotype, which is commonly termed as the 'rdar' (red, dry and rough morphotype. Both rdar and non-rdar phenotypes are commonly found in the nature; however, rdar is of essential importance for bacterial survival in the environment, outside of the host organisms. In that respect, it is comparable with the process of spore formation in Gram-positive bacteria. In laboratory conditions, rdar morphotype demonstrates growth of aforementioned organisms on agar containing diazo dye Congo Red, appearing as dark red (violet, rough and irregularly margined colonies. A coordinated expression of the number of genes mediated by major transcriptional regulators CsgD (curli subunit gene D is the prerequisite for the formation of rdar morphotype colonies. The key point in this process is the activation of csgBAC operon transcription, which encodes the synthesis of curli fimbriae and activates the AgfD regulated gene (AdrA involved in the cellulose biosynthesis. Curli fimbriae and cellulose are two basic structural components of the rdar morphotype. Curli fimbriae enable initial surface adhesion and intercellular aggregation of bacteria, whilst the cellulose (and other exopolysaccharides promotes intercellular interactions. CsgD affects the overall cell physiology towards a 'bacterial multicellular behavior' pattern, such as biofilm formation. The colony growth on Congo Red agar is a widely accepted laboratory method used for bacteria from the family Enterobacteriaceae and highly conserved between Salmonella. However, repeated laboratory sub-passaging results in the substantial loss of rdar morphotype and the formation of smooth mutants, suggests the necessity for using wild isolates. In this article, the characteristics of rdar and non-rdar morphotypes of diverse Salmonella serotypes isolated from

  19. Survival potential of wild type cellulose deficient Salmonella from the feed industry

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    Ballance Simon

    2009-11-01

    Full Text Available Abstract Background Biofilm has been shown to be one way for Salmonella to persist in the feed factory environment. Matrix components, such as fimbriae and cellulose, have been suggested to play an important role in the survival of Salmonella in the environment. Multicellular behaviour by Salmonella is often categorized according to colony morphology into rdar (red, dry and rough expressing curli fimbriae and cellulose, bdar (brown, dry and rough expressing curli fimbriae and pdar (pink, dry and rough expressing cellulose. The aim of the study was to look into the distribution of morphotypes among feed and fish meal factory strains of Salmonella, with emphasis on potential differences between morphotypes with regards to survival in the feed factory environment. Results When screening a total of 148 Salmonella ser. Agona, Salmonella ser. Montevideo, Salmonella ser. Senftenberg and Salmonella ser. Typhimurium strains of feed factory, human clinical and reference collection origin, as many as 99% were able to express rough morphology (rdar or bdar. The dominant morphotype was rdar (74%, however as many as 55% of Salmonella ser. Agona and 19% of Salmonella ser. Senftenberg displayed the bdar morphology. Inconsistency in Calcofluor binding, indicating expression of cellulose, was found among 25% of all the strains tested, however Salmonella ser. Agona showed to be highly consistent in Calcofluor binding (98%. In biofilm, Salmonella ser. Agona strains with bdar mophology was found to be equally tolerant to disinfection treatment as strains with rdar morphotype. However, rdar morphology appeared to be favourable in long term survival in biofilm in a very dry environment. Chemical analysis showed no major differences in polysaccharide content between bdar and rdar strains. Our results indicate that cellulose is not a major component of the Salmonella biofilm matrix. Conclusion The bdar morphotype is common among Salmonella ser. Agona strains isolated

  20. The pilus usher controls protein interactions via domain masking and is functional as an oligomer.

    Science.gov (United States)

    Werneburg, Glenn T; Henderson, Nadine S; Portnoy, Erica B; Sarowar, Samema; Hultgren, Scott J; Li, Huilin; Thanassi, David G

    2015-07-01

    The chaperone-usher (CU) pathway assembles organelles termed pili or fimbriae in Gram-negative bacteria. Type 1 pili expressed by uropathogenic Escherichia coli are prototypical structures assembled by the CU pathway. Biogenesis of pili by the CU pathway requires a periplasmic chaperone and an outer-membrane protein termed the usher (FimD). We show that the FimD C-terminal domains provide the high-affinity substrate-binding site but that these domains are masked in the resting usher. Domain masking requires the FimD plug domain, which serves as a switch controlling usher activation. We demonstrate that usher molecules can act in trans for pilus biogenesis, providing conclusive evidence for a functional usher oligomer. These results reveal mechanisms by which molecular machines such as the usher regulate and harness protein-protein interactions and suggest that ushers may interact in a cooperative manner during pilus assembly in bacteria.

  1. The asymptomatic bacteriuria Escherichia coli strain 83972 outcompetes uropathogenic E. coli strains in human urine

    DEFF Research Database (Denmark)

    Hancock, Viktoria; Ulett, G.C.; Schembri, M.A.

    2006-01-01

    Escherichia coli is the most common organism associated with asymptomatic bacteriuria (ABU). In contrast to uropathogenic E. coli (UPEC), which causes symptomatic urinary tract infections (UTI), very little is known about the mechanisms by which these strains colonize the human urinary tract....... The prototype ABU E. coli strain 83972 was originally isolated from a girl who had carried it asymptomatically for 3 years. Deliberate colonization of UTI-susceptible individuals with E. coli 83972 has been used successfully as an alternative approach for the treatment of patients who are refractory...... to conventional therapy. Colonization with strain 83972 appears to prevent infection with UPEC strains in such patients despite the fact that this strain is unable to express the primary adhesins involved in UTI, viz. P and type 1 fimbriae. Here we investigated the growth characteristics of E. coli 83972 in human...

  2. Molecular diagnostics of foodborne pathogens

    DEFF Research Database (Denmark)

    Hansen, Trine

    be connected to specific genes. Deletion of either of the operons prgor flhDCin S. Typhimurium resulted in lower attachment ability to the pork meat surface. In addition, was it found that a S. Rissen isolate with low attachment ability after immobilized growth lacked two fimbriae genes, safC and lpf...... to be more complicated and for the method to be used for this type of samples, additional optimizations have to be conducted. In conclusion, the work present in this thesis contributes to the better understanding of the behavior of Salmonellain the pork processing and which factors that might influence...... behavior in pork processing environments, and detection of B. cereusin food, feed and water samples without prior cultivation. The persistence of Salmonellain food production chains has been suggested to be a result of bacterial attachment and surface colonization. It was found that the physiological state...

  3. Genome analysis of urease positive Serratia marcescens, co-producing SRT-2 and AAC(6')-Ic with multidrug efflux pumps for antimicrobial resistance.

    Science.gov (United States)

    Srinivasan, Vijaya Bharathi; Rajamohan, Govindan

    2018-04-05

    In this study, we present the genome sequence of Serratia marcescens SM03, recovered from a human gut in India. The final assembly consists of 26 scaffolds (4620 coding DNA sequences, 5.08 Mb, 59.6% G + C ratio) and 79 tRNA genes. Analysis identified novel genes associated with lactose utilization, virulence, P-loop GTPases involved in urease production, CFA/I fimbriae apparatus and Yersinia - type CRISPR proteins. Antibiotic susceptibility testing indicated drug tolerant phenotype and inhibition assays demonstrated involvement of extrusion in resistance. Presence of enzymes SRT-2, AAC(6')-Ic, with additional Ybh transporter and EamA-like efflux pumps signifies the genetic plasticity observed in S. marcescens SM03. Copyright © 2018 Elsevier Inc. All rights reserved.

  4. Rare occurrence of heart lesions in Pacific oysters Crassostrea gigas caused by an unknown bacterial infection.

    Science.gov (United States)

    Meyer, Gary R; Lowe, Geoffrey J; Bower, Susan M

    2017-09-20

    On rare occasions, small cream-coloured cysts have been observed in the heart and pericardial cavity of Pacific oysters Crassostrea gigas from British Columbia, Canada. Histopathology revealed the presence of large colonies of bacteria (up to 800 µm in diameter) causing significant host response and hypertrophy of the heart epithelium. The causative bacteria were characterized as follows: Gram-negative, coccoid to small rod-shaped, typically <1.5 µm in size, cell walls highly endowed with surface fimbriae and division via binary fission. Although these bacteria shared some morphological characteristics with the order Rickettsiales, they did not require an intracellular existence for multiplication. Unfortunately, a cultured isolate was not available, and a retrospective attempt to further characterize the bacteria using DNA sequence analysis of a fragment from the 16S rDNA region proved to be uninformative.

  5. Refined localization of the Escherichia coli F4ab/F4ac receptor locus on pig chromosome 13

    DEFF Research Database (Denmark)

    Joller, D.; Jørgensen, Claus Bøttcher; Bertschinger, H.U.

    2009-01-01

    Diarrhoea in newborn and weaned pigs caused by enterotoxigenic Escherichia coli (ETEC) expressing F4 fimbriae leads to considerable losses in pig production. In this study, we refined the mapping of the receptor locus for ETEC F4ab/F4ac adhesion (F4bcR) by joint analysis of Nordic and Swiss data...... (MUC4-8227) were used to create the linkage map. The region for F4bcR was refined to the interval SW207-S0075 on pig chromosome 13. The most probable position of F4bcR was in the SW207-MUC4 region. The order of six markers was supported by physical mapping on the BAC fingerprint contig from...

  6. Primary structure and subcellular localization of two fimbrial subunit-like proteins involved in the biosynthesis of K99 fibrillae.

    Science.gov (United States)

    Roosendaal, E; Jacobs, A A; Rathman, P; Sondermeyer, C; Stegehuis, F; Oudega, B; de Graaf, F K

    1987-09-01

    Analysis of the nucleotide sequence of the distal part of the fan gene cluster encoding the proteins involved in the biosynthesis of the fibrillar adhesin, K99, revealed the presence of two structural genes, fanG and fanH. The amino acid sequence of the gene products (FanG and FanH) showed significant homology to the amino acid sequence of the fibrillar subunit protein (FanC). Introduction of a site-specific frameshift mutation in fanG or fanH resulted in a simultaneous decrease in fibrillae production and adhesive capacity. Analysis of subcellular fractions showed that, in contrast to the K99 fibrillar subunit (FanC), both the FanH and the FanG protein were loosely associated with the outer membrane, possibly on the periplasmic side, but were not components of the fimbriae themselves.

  7. Adhesion of Escherichia coli under flow conditions reveals potential novel effects of FimH mutations

    DEFF Research Database (Denmark)

    Feenstra, T.; Schmidt Thøgersen, Mariane; Wieser, E.

    2017-01-01

    H mutations on bacterial adhesion using a novel adhesion assay, which models the physiological flow conditions bacteria are exposed to. We introduced 12 different point mutations in the mannose binding pocket of FimH in an E. coli strain expressing type 1 fimbriae only (MSC95-FimH). We compared the bacterial...... adhesion of each mutant across several commonly used adhesion assays, including agglutination of yeast, adhesion to mono- and tri-mannosylated substrates, and static adhesion to bladder epithelial and endothelial cells. We performed a comparison of these assays to a novel method that we developed to study...... mutations abrogated adhesion. We demonstrated that FimH residues E50 and T53 are crucial for adhesion under flow conditions. The coating of endothelial cells on biochips and modelling of physiological flow conditions enabled us to identify FimH residues crucial for adhesion. These results provide novel...

  8. Distribution of Wfs1 protein in the central nervous system of the mouse and its relation to clinical symptoms of the Wolfram syndrome

    DEFF Research Database (Denmark)

    Luuk, H.; Koks, S.; Plaas, M.

    2008-01-01

    enrichment of Wf1 protein in the central extended amygdala and ventral striatum. Prominent Wfs1 expression was seen in the hippocampal CA1 region, parasubiculum, superficial part of the second and third layers of the prefrontal cortex and proisocortical areas, hypothalamic magnocellular neurosecretory system......, alveus, fimbria, dorsal hippocampal commissure; subiculum, and to a lesser extent in the central sublenticular extended amygdala, compact part of substantia nigra, and ventral tegmental area. The neuroanatomical findings suggest that the lack of Wf1 protein function can be related to several neurological...... and psychiatric symptoms found in Wolfram syndrome. Enrichment of Wfs1 protein in the central extended amygdala suggests a role in the modulation of anxiety and fear Udgivelsesdato: 2008/8/20...

  9. Evaluation of components of X-ray irradiated 7-valent pneumococcal conjugate vaccine and pneumococcal vaccine polyvalent and X-ray and gamma-ray irradiated acellular pertussis component of DTaP vaccine products

    International Nuclear Information System (INIS)

    May, J.C.; Rey, L.; Lee, C.-J.; Arciniega, Juan

    2004-01-01

    Samples of pneumococcal vaccine polyvalent, 7-valent pneumococcal conjugate vaccine, and two different diphtheria and tetanus toxoids and acellular pertussis vaccines adsorbed were irradiated with X-rays and/or gamma-rays (Co-60). Mouse IgG and IgM antibody responses (ELISA) for types 9V, 14, 18C, and 19F pneumococcal polysaccharides and conjugates indicated that the polysaccharides were more tolerant of the radiation than the conjugates. The mouse antibody response for the detoxified pertussis toxin (PT) antigen, filamentous hemagglutinin antigen (FHA), pertactin (PRN), and fimbriae types 2 and 3 (FIM) antigens for the appropriate vaccine type indicated that the antibody response was not significantly changed in the 25 kGy X-ray irradiated vaccines frozen in liquid nitrogen compared to the control vaccine

  10. Evaluation of components of X-ray irradiated 7-valent pneumococcal conjugate vaccine and pneumococcal vaccine polyvalent and X-ray and gamma-ray irradiated acellular pertussis component of DTaP vaccine products

    Energy Technology Data Exchange (ETDEWEB)

    May, J.C. E-mail: may@cber.fda.gov; Rey, L. E-mail: louis.rey@bluewin.ch; Lee, C.-J.; Arciniega, Juan

    2004-10-01

    Samples of pneumococcal vaccine polyvalent, 7-valent pneumococcal conjugate vaccine, and two different diphtheria and tetanus toxoids and acellular pertussis vaccines adsorbed were irradiated with X-rays and/or gamma-rays (Co-60). Mouse IgG and IgM antibody responses (ELISA) for types 9V, 14, 18C, and 19F pneumococcal polysaccharides and conjugates indicated that the polysaccharides were more tolerant of the radiation than the conjugates. The mouse antibody response for the detoxified pertussis toxin (PT) antigen, filamentous hemagglutinin antigen (FHA), pertactin (PRN), and fimbriae types 2 and 3 (FIM) antigens for the appropriate vaccine type indicated that the antibody response was not significantly changed in the 25 kGy X-ray irradiated vaccines frozen in liquid nitrogen compared to the control vaccine.

  11. Development and maturation of Escherichia coli K-12 biofilms

    DEFF Research Database (Denmark)

    Reisner, A.; Haagensen, J.A.J.; Schembri, Mark

    2003-01-01

    The development and maturation of E. coli biofilms in flow-chambers was investigated. We found that the presence of transfer constitutive IncF plasmids induced biofilm development forming structures resembling those reported for Pseudomonas aeruginosa . The development occurred in a step...... occurred in conjugation pilus proficient plasmid-carrying strains. The final shapes of the expanding structures in the mature biofilm seem to be determined by the pilus configuration, as various mutants affected in the processing and activity of the transfer pili displayed differently structured biofilms....... We further provide evidence that flagella, type 1 fimbriae, curli and Ag43 are all dispensable for the observed biofilm maturation. In addition, our results indicate that cell-to-cell signalling mediated by autoinducer 2 (AI-2) is not required for differentiation of E. coli within a biofilm community...

  12. Escherichia coli bacteraemia in patients with and without haematological malignancies: a study of strain characters and recurrent episodes

    DEFF Research Database (Denmark)

    Olesen, B; Kolmos, H J; Orskov, F

    1998-01-01

    We compared serotypes, virulence factors and susceptibility to antibiotics of Escherichia coli strains isolated from 282 patients with bacteraemia. Thirty-five of these were neutropenic patients with haematological malignancy and 247 were patients with a normal or raised total white blood cell......-haematological patients had recurrences with a strain different from the strain isolated in a previous episode. A possible connection between shorter intervals and recurrence with identical strains is discussed. We suggest that strains from recurrent E. coli bacteraemia are sent to a reference laboratory for serotyping...... count and no haematological malignancy. Strains isolated from recurrent bacteraemia were also bio- and ribotyped. Overall, no significant difference was found between O serogroups, K antigens, serum sensitivity, production of haemolysin, expression of P-fimbriae and patterns of antibiotic susceptibility...

  13. Modulating bacterial and gut mucosal interactions with engineered biofilm matrix proteins.

    Science.gov (United States)

    Duraj-Thatte, Anna M; Praveschotinunt, Pichet; Nash, Trevor R; Ward, Frederick R; Joshi, Neel S

    2018-02-22

    Extracellular appendages play a significant role in mediating communication between bacteria and their host. Curli fibers are a class of bacterial fimbria that is highly amenable to engineering. We demonstrate the use of engineered curli fibers to rationally program interactions between bacteria and components of the mucosal epithelium. Commensal E. coli strains were engineered to produce recombinant curli fibers fused to the trefoil family of human cytokines. Biofilms formed from these strains bound more mucins than those producing wild-type curli fibers, and modulated mucin rheology as well. When treated with bacteria producing the curli-trefoil fusions mammalian cells behaved identically in terms of their migration behavior as when they were treated with the corresponding soluble trefoil factors. Overall, this demonstrates the potential utility of curli fibers as a scaffold for the display of bioactive domains and an untapped approach to rationally modulating host-microbe interactions using bacterial matrix proteins.

  14. Characteristics of cytotoxic necrotizing factor and cytolethal distending toxin producing Escherichia coli strains isolated from meat samples in Northern Ireland.

    Science.gov (United States)

    Kadhum, H J; Ball, H J; Oswald, E; Rowe, M T

    2006-08-01

    Swabs collected from pig, lamb and beef carcasses and samples of pork, lamb and beef mince were cultured for Escherichia coli strains. Strains harbouring cytotoxic necrotizing factors (CNF1 and 2) and cytolethal distending toxins (CDT-I,-II,-III and -IV) were identified in plate cultures of the isolates by colony hybridization with labelled probes and multiplex PCR assays. Simplex and multiplex PCR assays were used to further characterize the isolates to determine the presence of P, S and F17 fimbriae as well as afimbrial adhesins and haemolysin. The serotype was also determined where possible. Thirty strains with the capacity to code for CNF (4), CDT (24) or both (2) were isolated and characterized, and a wide range of associated factor patterns was observed. The methods utilized were successful in demonstrating the detection of viable strains with potentially significant pathogenic factors from human food sources.

  15. Cytoarchitectonic distribution of zinc in the hippocampus of man and the rat

    International Nuclear Information System (INIS)

    Frederickson, C.J.; Klitenick, M.A.; Manton, W.I.; Kirkpatrick, J.B.

    1983-01-01

    Zinc was measured in whole hippocampus and in hippocampal sub-regions by stable-isotope dilution mass spectrometry. Lyophilized tissues were spiked by a precisely-known amount of zinc-64. The zinc-64/zinc-66 isotope ratio was determined by mass spectrometry. In both man and the rat, the most zinc (102-145 ppm, dry weight) was found in the hilar region, the least (27-35) in the fimbria. The amount of zinc directly associated with mossy-fiber axons was estimated to be approximately 8% of the total zinc in the hippocampus, and the concentration of mossy-fiber zinc was estimated at 220-300 μM. Methodological and theoretical implications of the quantitative findings were discussed. (Auth.)

  16. Enteroaggregative Escherichia coli O78:H10, the cause of an outbreak of urinary tract infection

    DEFF Research Database (Denmark)

    Olesen, Bente; Scheutz, Flemming; Andersen, Rebecca L

    2012-01-01

    to these traits and resistance profiles, are unknown. Accordingly, we extensively characterized 51 archived E. coli O78:H10 isolates (48 human isolates from seven countries, including 19 Copenhagen outbreak isolates, and 1 each of calf, avian, and unknown-source isolates), collected from 1956 through 2000. E...... by molecular methods as enteroaggregative E. coli (EAEC) (P 90% of outbreak isolates included fimH (type 1 fimbriae; ubiquitous in E. coli); fyuA, traT, and iutA (associated with extraintestinal pathogenic E. coli [ExPEC]); and sat, pic, aatA, aggR, aggA, ORF61, aaiC, aap, and ORF3......In 1991, multiresistant Escherichia coli O78:H10 strains caused an outbreak of urinary tract infections in Copenhagen, Denmark. The phylogenetic origin, clonal background, and virulence characteristics of the outbreak isolates, and their relationship to nonoutbreak O78:H10 strains according...

  17. Nitazoxanide Inhibits Pilus Biogenesis by Interfering with Folding of the Usher Protein in the Outer Membrane.

    Science.gov (United States)

    Chahales, Peter; Hoffman, Paul S; Thanassi, David G

    2016-04-01

    Many bacterial pathogens assemble surface fibers termed pili or fimbriae that facilitate attachment to host cells and colonization of host tissues. The chaperone/usher (CU) pathway is a conserved secretion system that is responsible for the assembly of virulence-associated pili by many different Gram-negative bacteria. Pilus biogenesis by the CU pathway requires a dedicated periplasmic chaperone and an integral outer membrane (OM) assembly and secretion platform termed the usher. Nitazoxanide (NTZ), an antiparasitic drug, was previously shown to inhibit the function of aggregative adherence fimbriae and type 1 pili assembled by the CU pathway in enteroaggregativeEscherichia coli, an important causative agent of diarrhea. We show here that NTZ also inhibits the function of type 1 and P pili from uropathogenicE. coli(UPEC). UPEC is the primary causative agent of urinary tract infections, and type 1 and P pili mediate colonization of the bladder and kidneys, respectively. By analysis of the different stages of the CU pilus biogenesis pathway, we show that treatment of bacteria with NTZ causes a reduction in the number of usher molecules in the OM, resulting in a loss of pilus assembly on the bacterial surface. In addition, we determine that NTZ specifically prevents proper folding of the usher β-barrel domain in the OM. Our findings demonstrate that NTZ is a pilicide with a novel mechanism of action and activity against diverse CU pathways. This suggests that further development of the NTZ scaffold may lead to new antivirulence agents that target the usher to prevent pilus assembly. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  18. Temperature control of fimbriation circuit switch in uropathogenic Escherichia coli: quantitative analysis via automated model abstraction.

    Science.gov (United States)

    Kuwahara, Hiroyuki; Myers, Chris J; Samoilov, Michael S

    2010-03-26

    Uropathogenic Escherichia coli (UPEC) represent the predominant cause of urinary tract infections (UTIs). A key UPEC molecular virulence mechanism is type 1 fimbriae, whose expression is controlled by the orientation of an invertible chromosomal DNA element-the fim switch. Temperature has been shown to act as a major regulator of fim switching behavior and is overall an important indicator as well as functional feature of many urologic diseases, including UPEC host-pathogen interaction dynamics. Given this panoptic physiological role of temperature during UTI progression and notable empirical challenges to its direct in vivo studies, in silico modeling of corresponding biochemical and biophysical mechanisms essential to UPEC pathogenicity may significantly aid our understanding of the underlying disease processes. However, rigorous computational analysis of biological systems, such as fim switch temperature control circuit, has hereto presented a notoriously demanding problem due to both the substantial complexity of the gene regulatory networks involved as well as their often characteristically discrete and stochastic dynamics. To address these issues, we have developed an approach that enables automated multiscale abstraction of biological system descriptions based on reaction kinetics. Implemented as a computational tool, this method has allowed us to efficiently analyze the modular organization and behavior of the E. coli fimbriation switch circuit at different temperature settings, thus facilitating new insights into this mode of UPEC molecular virulence regulation. In particular, our results suggest that, with respect to its role in shutting down fimbriae expression, the primary function of FimB recombinase may be to effect a controlled down-regulation (rather than increase) of the ON-to-OFF fim switching rate via temperature-dependent suppression of competing dynamics mediated by recombinase FimE. Our computational analysis further implies that this down

  19. Temperature control of fimbriation circuit switch in uropathogenic Escherichia coli: quantitative analysis via automated model abstraction.

    Directory of Open Access Journals (Sweden)

    Hiroyuki Kuwahara

    2010-03-01

    Full Text Available Uropathogenic Escherichia coli (UPEC represent the predominant cause of urinary tract infections (UTIs. A key UPEC molecular virulence mechanism is type 1 fimbriae, whose expression is controlled by the orientation of an invertible chromosomal DNA element-the fim switch. Temperature has been shown to act as a major regulator of fim switching behavior and is overall an important indicator as well as functional feature of many urologic diseases, including UPEC host-pathogen interaction dynamics. Given this panoptic physiological role of temperature during UTI progression and notable empirical challenges to its direct in vivo studies, in silico modeling of corresponding biochemical and biophysical mechanisms essential to UPEC pathogenicity may significantly aid our understanding of the underlying disease processes. However, rigorous computational analysis of biological systems, such as fim switch temperature control circuit, has hereto presented a notoriously demanding problem due to both the substantial complexity of the gene regulatory networks involved as well as their often characteristically discrete and stochastic dynamics. To address these issues, we have developed an approach that enables automated multiscale abstraction of biological system descriptions based on reaction kinetics. Implemented as a computational tool, this method has allowed us to efficiently analyze the modular organization and behavior of the E. coli fimbriation switch circuit at different temperature settings, thus facilitating new insights into this mode of UPEC molecular virulence regulation. In particular, our results suggest that, with respect to its role in shutting down fimbriae expression, the primary function of FimB recombinase may be to effect a controlled down-regulation (rather than increase of the ON-to-OFF fim switching rate via temperature-dependent suppression of competing dynamics mediated by recombinase FimE. Our computational analysis further implies

  20. Maillard neoglycans as inhibitors for in vitro adhesion of F4+ enterotoxigenic Escherichia coli to piglet intestinal cells.

    Science.gov (United States)

    Sarabia-Sainz, Héctor Manuel; Mata Haro, Verónica; Sarabia Sainz, José Andre-I; Vázquez-Moreno, Luz; Montfort, Gabriela Ramos-Clamont

    2017-01-01

    Adhesion of enterotoxigenic (ETEC) E. coli to host intestinal cells is mediated by lectin-like fimbriae that bind to specific glycan moieties on the surfaces of enterocytes. To prevent in vitro binding of E. coli F4 fimbriae (F4 ETEC + ) to piglet enterocytes, neoglycans were synthesized by the Maillard reaction conjugating lactose (Lac), galacto-oligosaccharides (GOS) or chitin oligosaccharides (Ochit) to porcine serum albumin (PSA). Neoglycans were characterized by SDS-PAGE, intrinsic tryptophan fluorescence and recognition by plant lectins, as well as by F4 ETEC variants. Electrophoretic patterns suggested the binding to PSA of 63, 13 and 2 molecules of Lac, GOS and Ochit, respectively. All neoglycans displayed quenching of tryptophan fluorescence consistent with the degree of glycation estimated by SDS-PAGE. Plant lectins recognized the neoglycans according to their specificity, whereas antigenic variants of F4 ETEC (ab, ac and ad) recognized PSA-Ochit and PSA-Lac with higher affinity than that for GOS. Neoglycans partially hindered the in vitro binding of F4 + ETEC to piglet enterocytes in a dose-dependent manner. The most effective blocking was observed with PSA-Lac that partially inhibited the adhesion of bacteria to enterocytes in a dose dependent manner, as quantified by flow cytometry. Increased production of the cytokines IL-6 and TNF-α was observed in response to F4 + ETEC infection of enterocytes and production was reduced in the presence of PSA-Ochit and PSA-GOS. These results suggest that neoglycans synthesized by the Maillard reaction could be useful in the prophylaxis of diarrhea in piglets.

  1. Evaluation of Antibiotic Resistance and Detection of papC and papG genes in Escherichia coli Strains Isolated from Patients with Urinary Tract Infection

    Directory of Open Access Journals (Sweden)

    Mitra Alishah

    2016-11-01

    Full Text Available Abstract Background and Objectives: Escherichia coli is the most common etiologic factor of urinary tract infection, which its most important virulence factor is P fimbriae. Uropathogenic E. coli expresses various types of adhesins, such as pili adhesins (pyelonephritis-associated pili, Pap that mediates binding to the surface of epithelial cells of the urinary tract. This study aims to identify papC and papG genes and to evaluate antibiotic resistance level in the isolated E. coli samples. Methods: In this descriptive cross-sectional study, 50 samples were collected from patients with urinary tract infection and after isolation of bacteria and DNA extraction, antibiotic susceptibility tests was performed by disc diffusion method using related antibiotics. Presence of papG and papC genes (class I, II, and III was assessed by multiplex PCR method. Statistical data were analyzed using descriptive t-test. Results: The isolated E. coli samples were susceptible to amikacin (100% and cefepime (72% and resistant to ampicillin (100% and nitrofurantoin (94%. Eighteen samples (32.7% had papG gene, of which 17 (30.9% samples had papGII gene and 1 sample (1.8% had papGIII gene; papGI gene was not detected in any of the samples. Conclusion: The results of the present study showed that papC and papGI genes are the most common Pap fimbriae adhesion-encoding genes in E. coli isolated from urinary tract infection. The difference between the results of this study with those of other studies is due to geographic diversity. Keywords: Escherichia coli; Adhesion pap, Disk diffusion antimicrobial tests; Multiplex polymerase chain reaction.

  2. stg fimbrial operon from S. Typhi STH2370 contributes to association and cell disruption of epithelial and macrophage-like cells.

    Science.gov (United States)

    Berrocal, Liliana; Fuentes, Juan A; Trombert, A Nicole; Jofré, Matías R; Villagra, Nicolás A; Valenzuela, Luis M; Mora, Guido C

    2015-07-07

    Salmonella enterica serovar Typhi (S. Typhi) stg operon, encoding a chaperone/usher fimbria (CU), contributes to an increased adherence to human epithelial cells. However, one report suggests that the presence of the Stg fimbria impairs the monocyte--bacteria association, as deduced by the lower level of invasion to macrophage-like cells observed when the stg fimbrial cluster was overexpressed. Nevertheless, since other CU fimbrial structures increase the entry of S. Typhi into macrophages, and considering that transcriptomic analyses revealed that stg operon is indeed expressed in macrophages, we reassessed the role of the stg operon in the interaction between S. Typhi strain STH2370 and human cells, including macrophage-like cells and mononuclear cells directly taken from human peripheral blood. We compared S. Typhi STH2370 WT, a Chilean clinical strain, and the S. Typhi STH2370 Δstg mutant with respect to association and invasion using epithelial and macrophage-like cells. We observed that deletion of stg operon reduced the association and invasion of S. Typhi, in both cellular types. The presence of the cloned stg operon restored the WT phenotype in all the cases. Moreover, we compared Salmonella enterica sv. Typhimurium 14028s (S. Typhimurium, a serovar lacking stg operon) and S. Typhimurium heterologously expressing S. Typhi stg. We found that the latter presents an increased cell disruption of polarized epithelial cells and an increased association in both epithelial and macrophage-like cells. S. Typhi stg operon encodes a functional adhesin that participates in the interaction bacteria-eukaryotic cells, including epithelial cells and macrophages-like cells. The phenotypes associated to stg operon include increased association and consequent invasion in bacteria-eukaryotic cells, and cell disruption.

  3. Fetal hypothalamic transplants into brain irradiated rats: Graft morphometry and host behavioral responses

    International Nuclear Information System (INIS)

    Pearlman, S.H.; Rubin, P.; White, H.C.; Wiegand, S.J.; Gash, D.M.

    1990-01-01

    This study was designed to test the hypothesis that neural implants can ameliorate or prevent some of the long-term changes associated with CNS irradiation. Using a rat model, the initial study focused on establishing motor, regulatory, and morphological changes associated with brain radiation treatments. Secondly, fetal hypothalamic tissue grafts were placed into the third ventricle of rats which had been previously irradiated. Adult male Long Evans rats received one of three radiation doses (15, 22.5, ampersand 30 Gy) or no radiation. Three days after irradiation, 7 animals in each dose group received an embryonic day 17 hypothalamic graft into the third ventricle while the remaining 8-9 animals in each group received injections of vehicle solution (sham). Few changes were observed in the 15 and 22.5 Gy animals, however rats in the 30 Gy treatment group showed stereotypic and ambulatory behavioral hyperactivity 32 weeks after irradiation. Regulatory changes in the high dose group included decreased growth rate and decreased urine osmolalities, but these measures were extremely variable among animals. Morphological results demonstrated that 30 Gy irradiated animals showed extensive necrosis primarily in the fimbria, which extended into the internal capsule, optic nerve, hippocampus, and thalamus. Hemorrhages were found in the hippocampus, thalamus, and fimbria. Defects in the blood-brain barrier also were evident by entry of intravascularly injected horseradish peroxidase into the parenchyma of the brain. Animals in the 30 Gy grafted group showed fewer behavioral changes and less brain damage than their sham grafted counterparts. Specifically, activity measures were comparable to normal levels, and a dilute urine was not found in the 30 Gy implanted rats. Morphological changes support these behavioral results since only two 30 Gy implanted rats showed necrosis

  4. Sequencing of IncX-plasmids suggests ubiquity of mobile forms of a biofilm-promoting gene cassette recruited from Klebsiella pneumoniae.

    Directory of Open Access Journals (Sweden)

    Mette Burmølle

    Full Text Available Plasmids are a highly effective means with which genetic traits that influence human health, such as virulence and antibiotic resistance, are disseminated through bacterial populations. The IncX-family is a hitherto sparsely populated group of plasmids that are able to thrive within Enterobacteriaceae. In this study, a replicon-centric screening method was used to locate strains from wastewater sludge containing plasmids belonging to the IncX-family. A transposon aided plasmid capture method was then employed to transport IncX-plasmids from their original hosts (and co-hosted plasmids into a laboratory strain (Escherichia coli Genehogs® for further study. The nucleotide sequences of the three newly isolated IncX-plasmids (pLN126_33, pMO17_54, pMO440_54 and the hitherto un-sequenced type-plasmid R485 revealed a remarkable occurrence of whole or partial gene cassettes that promote biofilm-formation in Klebsiella pneumonia or E. coli, in all four instances. Two of the plasmids (R485 and pLN126_33 were shown to directly induce biofilm formation in a crystal violet retention assay in E. coli. Sequence comparison revealed that all plasmid-borne forms of the type 3 fimbriae encoding gene cassette mrkABCDF were variations of a composite transposon Tn6011 first described in the E. coli IncX plasmid pOLA52. In conclusion, IncX-plasmids isolated from Enterobacteriaceae over almost 40 years and on three different continents have all been shown to carry a type 3 fimbriae gene cassette mrkABCDF stemming from pathogenic K. pneumoniae. Apart from contributing general knowledge about IncX-plasmids, this study also suggests an apparent ubiquity of a mobile form of an important virulence factor and is an illuminating example of the recruitment, evolution and dissemination of genetic traits through plasmid-mediated horizontal gene transfer.

  5. Sequencing of IncX-plasmids suggests ubiquity of mobile forms of a biofilm-promoting gene cassette recruited from Klebsiella pneumoniae.

    Science.gov (United States)

    Burmølle, Mette; Norman, Anders; Sørensen, Søren J; Hansen, Lars Hestbjerg

    2012-01-01

    Plasmids are a highly effective means with which genetic traits that influence human health, such as virulence and antibiotic resistance, are disseminated through bacterial populations. The IncX-family is a hitherto sparsely populated group of plasmids that are able to thrive within Enterobacteriaceae. In this study, a replicon-centric screening method was used to locate strains from wastewater sludge containing plasmids belonging to the IncX-family. A transposon aided plasmid capture method was then employed to transport IncX-plasmids from their original hosts (and co-hosted plasmids) into a laboratory strain (Escherichia coli Genehogs®) for further study. The nucleotide sequences of the three newly isolated IncX-plasmids (pLN126_33, pMO17_54, pMO440_54) and the hitherto un-sequenced type-plasmid R485 revealed a remarkable occurrence of whole or partial gene cassettes that promote biofilm-formation in Klebsiella pneumonia or E. coli, in all four instances. Two of the plasmids (R485 and pLN126_33) were shown to directly induce biofilm formation in a crystal violet retention assay in E. coli. Sequence comparison revealed that all plasmid-borne forms of the type 3 fimbriae encoding gene cassette mrkABCDF were variations of a composite transposon Tn6011 first described in the E. coli IncX plasmid pOLA52. In conclusion, IncX-plasmids isolated from Enterobacteriaceae over almost 40 years and on three different continents have all been shown to carry a type 3 fimbriae gene cassette mrkABCDF stemming from pathogenic K. pneumoniae. Apart from contributing general knowledge about IncX-plasmids, this study also suggests an apparent ubiquity of a mobile form of an important virulence factor and is an illuminating example of the recruitment, evolution and dissemination of genetic traits through plasmid-mediated horizontal gene transfer.

  6. Serous tubal intraepithelial carcinoma: its potential role in primary peritoneal serous carcinoma and serous cancer prevention.

    Science.gov (United States)

    Carlson, Joseph W; Miron, Alexander; Jarboe, Elke A; Parast, Mana M; Hirsch, Michelle S; Lee, Yonghee; Muto, Michael G; Kindelberger, David; Crum, Christopher P

    2008-09-01

    A diagnosis of primary peritoneal serous carcinoma (PPSC) requires exclusion of a source in other reproductive organs. Serous tubal intraepithelial carcinoma (STIC; stage 0) has been described in asymptomatic women with BRCA mutations and linked to a serous cancer precursor in the fimbria. This study examined the frequency of STIC in PPSC and its clinical outcome in BRCA-positive women. Presence or absence of STIC was recorded in consecutive cases meeting the 2001 WHO criteria for PPSC, including 26 patients with nonuniform sampling of the fallopian tubes (group 1) and 19 patients with complete tubal examination (group 2; sectioning and extensively examining the fimbriated end, or SEE-FIM protocol). In selected cases, STIC or its putative precursor and the peritoneal tumor were analyzed for p53 mutations (exons 1 to 11). Outcome of STIC was ascertained by literature review. Thirteen (50%) of 26 PPSCs in group 1 involved the endosalpinx, with nine STICs (35%). Fifteen (79%) of 19 cases in group 2 contained endosalpingeal involvement, with nine STICs (47%). STIC was typically fimbrial and unifocal, with variable invasion of the tubal wall. In five of five cases, the peritoneal and tubal lesion shared an identical p53 mutation. Of 10 reported STICs in BRCA-positive women, all patients were without disease on follow-up. The fimbria is the source of nearly one half of PPSCs, suggesting serous malignancy originates in the tubal mucosa but grows preferentially at a remote peritoneal site. The generally low risk of recurrence in stage 0 (STIC) disease further underscores STIC as a possible target for early serous cancer detection and prevention.

  7. Genetic exchange of fimbrial alleles exemplifies the adaptive virulence strategy of Porphyromonas gingivalis.

    Directory of Open Access Journals (Sweden)

    Jennifer E Kerr

    Full Text Available Porphyromonas gingivalis is a gram-negative anaerobic bacterium, a member of the human oral microbiome, and a proposed "keystone" pathogen in the development of chronic periodontitis, an inflammatory disease of the gingiva. P. gingivalis is a genetically diverse species, and is able to exchange chromosomal DNA between strains by natural competence and conjugation. In this study, we investigate the role of horizontal DNA transfer as an adaptive process to modify behavior, using the major fimbriae as our model system, due to their critical role in mediating interactions with the host environment. We show that P. gingivalis is able to exchange fimbrial allele types I and IV into four distinct strain backgrounds via natural competence. In all recombinants, we detected a complete exchange of the entire fimA allele, and the rate of exchange varies between the different strain backgrounds. In addition, gene exchange within other regions of the fimbrial genetic locus was identified. To measure the biological implications of these allele swaps we compared three genotypes of fimA in an isogenic background, strain ATCC 33277. We demonstrate that exchange of fimbrial allele type results in profound phenotypic changes, including the quantity of fimbriae elaborated, membrane blebbing, auto-aggregation and other virulence-associated phenotypes. Replacement of the type I allele with either the type III or IV allele resulted in increased invasion of gingival fibroblast cells relative to the isogenic parent strain. While genetic variability is known to impact host-microbiome interactions, this is the first study to quantitatively assess the adaptive effect of exchanging genes within the pan genome cloud. This is significant as it presents a potential mechanism by which opportunistic pathogens may acquire the traits necessary to modify host-microbial interactions.

  8. Genetic exchange of fimbrial alleles exemplifies the adaptive virulence strategy of Porphyromonas gingivalis.

    Science.gov (United States)

    Kerr, Jennifer E; Abramian, Jared R; Dao, Doan-Hieu V; Rigney, Todd W; Fritz, Jamie; Pham, Tan; Gay, Isabel; Parthasarathy, Kavitha; Wang, Bing-yan; Zhang, Wenjian; Tribble, Gena D

    2014-01-01

    Porphyromonas gingivalis is a gram-negative anaerobic bacterium, a member of the human oral microbiome, and a proposed "keystone" pathogen in the development of chronic periodontitis, an inflammatory disease of the gingiva. P. gingivalis is a genetically diverse species, and is able to exchange chromosomal DNA between strains by natural competence and conjugation. In this study, we investigate the role of horizontal DNA transfer as an adaptive process to modify behavior, using the major fimbriae as our model system, due to their critical role in mediating interactions with the host environment. We show that P. gingivalis is able to exchange fimbrial allele types I and IV into four distinct strain backgrounds via natural competence. In all recombinants, we detected a complete exchange of the entire fimA allele, and the rate of exchange varies between the different strain backgrounds. In addition, gene exchange within other regions of the fimbrial genetic locus was identified. To measure the biological implications of these allele swaps we compared three genotypes of fimA in an isogenic background, strain ATCC 33277. We demonstrate that exchange of fimbrial allele type results in profound phenotypic changes, including the quantity of fimbriae elaborated, membrane blebbing, auto-aggregation and other virulence-associated phenotypes. Replacement of the type I allele with either the type III or IV allele resulted in increased invasion of gingival fibroblast cells relative to the isogenic parent strain. While genetic variability is known to impact host-microbiome interactions, this is the first study to quantitatively assess the adaptive effect of exchanging genes within the pan genome cloud. This is significant as it presents a potential mechanism by which opportunistic pathogens may acquire the traits necessary to modify host-microbial interactions.

  9. Genotypic characterization of virulence factors in Escherichia coli strains from patients with cystitis Caracterização genotípica dos fatores de virulência em amostras de Escherichia coli isoladas de pacientes com cistite

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    Monique Ribeiro Tiba

    2008-10-01

    Full Text Available Adhesins (P-fimbriae, S-fimbriae, type 1 fimbriae and afimbrial adhesin, toxins (α-hemolysin and cytotoxic necrotizing factor type 1, iron acquisition systems (aerobactin and host defense avoidance mechanisms (capsule or lipopolysaccharide have been shown to be prevalent in Escherichia coli strains associated with urinary tract infections. In this work, 162 Uropathogenic Escherichia coli (UPEC strains from patients with cystitis were genotypically characterized by polymerase chain reaction (PCR assay. We developed three multiplex PCR assays for virulence-related genes papC, papE/F, papG alleles, fimH, sfa/foc, afaE, hly, cnf-1, usp, cdtB, iucD, and kpsMTII, all of them previously identified in UPEC strains. The PCR assay results identified 158 fimH (97.5%, 86 kpsMTII (53.1%, 53 papC/papEF/papG (32.7%, 45 sfa (27.8%, 42 iucD (25.9%, 41 hly (25.3%, 36 usp (22.2%, 30 cnf-1(18.5% and 10 afa (6.2% strains. No strain was positive for cdtB. In this work, we also demonstrated that adhesins may be multiple within a single strain and that several virulence genes can occur combined in association.Adesinas (Fímbria P, fímbria S, fímbria do tipo 1 e a adesina afimbrial, toxinas (α-hemolisina e o fator necrosante citotóxico do tipo 1, sistemas de captação de ferro (aerobactina, e mecanismos de defesa do hospedeiro (cápsula ou lipopolissacarídeo são prevalentes em amostras de Escherichia coli associadas a infecções do trato urinário. O objetivo deste trabalho foi caracterizar genotipicamente 162 amostras de Escherichia coli uropatogênica (UPEC de pacientes com cistite através do ensaio da reação em cadeia da polimerase. Foram realizados três ensaios de PCR multiplex para os seguintes fatores de virulência: papC, papE/F, alelos de papG, fimH, sfa/foc, afaE, hly, cnf-1, usp, cdtB, iucD, e kpsMTII. Os resultados da PCR identificaram, 158 amostras fimH (97,5%, 86 amostras kpsMTII (53,1%, 53 amostras papC/papEF/papG (32,7%, 45 amostras sfa (27

  10. Efecto del herbicida ácido 2,4-diclorofenoxiacético sobre la fimbriación de bacterias uropatógenas Effect of the herbicide 2,4-dichlorophenoxyacetic acid on fimbriation of uropathogenic bacteria

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    Claudia Balagué

    2006-07-01

    Full Text Available Existen algunos datos epidemiológicos que demuestran que, en poblaciones rurales, la frecuencia de lesiones inflamatorias de origen infeccioso en el riñón es mayor en individuos expuestos a pesticidas. Estudios previos han demostrado que el herbicida 2,4- D altera las propiedades de adherencia bacteriana a riñón, mediada por fimbrias de Escherichia coli uropatógena, si bien aún no se ha estudiado el mecanismo que podría estar involucrado. El 2,4-D demostró capacidad de generar uniones covalentes a proteínas de bacilos gram negativos. Debido a este hallazgo, se hipotetizó que la unión del herbicida a proteínas involucradas en la síntesis, exportación o ensamble de las subunidades fimbriales podía alterar la expresión de las fimbrias. En este trabajo se demuestra una unión covalente del herbicida a proteínas de membrana externa de Escherichia coli uropatógena y paralelamente una disminución de la fimbriación, determinada por tres técnicas diferentes: hemoaglutinación, cuantificación de proteínas de superficie y microscopía electrónica. Una conclusión importante está referida a que la exposición accidental de trabajadores rurales al 2,4-D en muy bajas dosis y durante un corto período de tiempo tendría un efecto protector de la pielonefritis por disminución de la fimbriación; mientras que los altos niveles de exposición predispondrían a la recurrencia del proceso infeccioso en el modelo de la pielonefritis ascendente no obstructiva en ratón (Balagué et al 2002. Probablemente los estudios epidemiológicos que asocian exposición a los herbicidas e infección renal estén relacionados con este último tipo de exposiciones.Epidemiological data demonstrate, on rural populations, that the frequency of inflammatory lesions in renal tissue is higher for individuals exposed to pesticides. Previous studies demonstrated that the herbicide 2,4-D altered the bacterial adherence properties to renal tissue, mediated by

  11. Preoperative automated fibre quantification predicts postoperative seizure outcome in temporal lobe epilepsy.

    Science.gov (United States)

    Keller, Simon S; Glenn, G Russell; Weber, Bernd; Kreilkamp, Barbara A K; Jensen, Jens H; Helpern, Joseph A; Wagner, Jan; Barker, Gareth J; Richardson, Mark P; Bonilha, Leonardo

    2017-01-01

    Approximately one in every two patients with pharmacoresistant temporal lobe epilepsy will not be rendered completely seizure-free after temporal lobe surgery. The reasons for this are unknown and are likely to be multifactorial. Quantitative volumetric magnetic resonance imaging techniques have provided limited insight into the causes of persistent postoperative seizures in patients with temporal lobe epilepsy. The relationship between postoperative outcome and preoperative pathology of white matter tracts, which constitute crucial components of epileptogenic networks, is unknown. We investigated regional tissue characteristics of preoperative temporal lobe white matter tracts known to be important in the generation and propagation of temporal lobe seizures in temporal lobe epilepsy, using diffusion tensor imaging and automated fibre quantification. We studied 43 patients with mesial temporal lobe epilepsy associated with hippocampal sclerosis and 44 healthy controls. Patients underwent preoperative imaging, amygdalohippocampectomy and postoperative assessment using the International League Against Epilepsy seizure outcome scale. From preoperative imaging, the fimbria-fornix, parahippocampal white matter bundle and uncinate fasciculus were reconstructed, and scalar diffusion metrics were calculated along the length of each tract. Altogether, 51.2% of patients were rendered completely seizure-free and 48.8% continued to experience postoperative seizure symptoms. Relative to controls, both patient groups exhibited strong and significant diffusion abnormalities along the length of the uncinate bilaterally, the ipsilateral parahippocampal white matter bundle, and the ipsilateral fimbria-fornix in regions located within the medial temporal lobe. However, only patients with persistent postoperative seizures showed evidence of significant pathology of tract sections located in the ipsilateral dorsal fornix and in the contralateral parahippocampal white matter bundle

  12. Histomorphometric evaluation of intestinal cellular immune responses in pigs immunized with live oral F4ac+ non-enterotoxigenic E. coli vaccine against postweaning colibacillosis

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    A. Kovšca Janjatovic

    2010-02-01

    Full Text Available Enterotoxigenic Escherichia coli (ETEC infection is the most common type of porcine postweaning colibacillosis (PWC. Among fimbriae of porcine ETEC strains the best studied family of fimbriae are the members of F4 adhesins, existing in at least three variants: ab, ac, ad. Active immunization against porcine PWC is difficult due to: i ETEC strains are only one of the essential predisposing factors, ii the success of vaccinal antigen uptake depends on the presence of enterocyte receptors for F4 adhesins, iii the intestinal immune system may react with tolerance or hypersensitivity to the same antigens depending on the dose and form of the vaccinal immunogen, and iv kinetics of the specific immune responses may be different in the case of F4 (earlier and the other ETEC adhesins, particularly F18 (later. The aim of this study was to test the effectiveness of a live attenuated F4ac+ non-ETEC vaccine against porcine PWC by analyzing quantitative differences in the small intestinal lymphoid and myeloid cell subsets of immunized (with or without levamisole given as an adjuvant vs control non-immunized pigs. Four week-old pigs were intragastrically immunized with a vaccine candidate F4ac+ non-ETEC strain 2407 at day 0, challenged 7 days later with a virulent F4ac+ strain ETEC 11-800/1/94, euthanatized at day 13 and sampled for immunohistology. Non-immunized pigs received saline at day 0 and were processed as the principals. Immuno-phenotypes of lymphoid and myeloid cell subsets were demonstrated within jejunal and ileal mucosa by immunohistochemical avidin-biotin complex method and corresponding morphometric data were analyzed using software program Lucia G for digital image analyses. Monoclonal antibodies reactive with surface molecules on porcine immune cells such as CD3, CD45RA, CD45RC, CD21 and SWC3 enabled clear insight into distribution patterns and amount of these cells within the gut-associated lymphoid tissues (GALT examined. The numbers of

  13. A comparative study of biofilm formation by Shiga toxigenic Escherichia coli using epifluorescence microscopy on stainless steel and a microtitre plate method.

    Science.gov (United States)

    Rivas, Lucia; Dykes, Gary A; Fegan, Narelle

    2007-04-01

    Attachment of Shiga toxigenic Escherichia coli (STEC) to surfaces and the formation of biofilms may enhance persistence in a food processing environment and present a risk of contaminating products. Seven strains of STEC and three non-STEC strains were selected to compare two biofilm quantification methods; epifluorescence microscopy on stainless steel (SS) and a microtitre plate assay. The influence of prior growth in planktonic (nutrient broth) and sessile (nutrient agar) culture on biofilm production, as well as expression of surface structures and the possession of antigen 43 (encoded by agn43) on biofilm formation were also investigated. Biofilms were produced in diluted nutrient broth at 25 degrees C for 24 and 48 h. Curli expression was determined using congo red indicator agar, while the presence of agn43 was determined using polymerase chain reaction. No correlation was found between counts for epifluorescence microscopy on SS and the absorbance values obtained with the microtitre plate method for planktonic and sessile grown cultures. Different abilities of individual STEC strains to attach to SS and microtitre plates were found with some strains attaching better to each surface following growth in either planktonic or sessile culture. All O157 STEC strains had low biofilm counts on SS for planktonic and sessile grown cultures; however, one STEC O157:H- strain (EC516) had significantly greater (pbiofilm production on microtitre plates compared to the other O157 STEC strains. EC516 and other STEC (O174:H21 and O91:H21) strains expressing curli fimbriae were found to produce significantly greater (pbiofilms on microtitre plates compared to the non-curli expressing strains. No relationship was found between the production of type-I fimbriae, motility, agn43 and bacterial physicochemical properties (previously determined) and biofilm formation on SS or microtitre plates. Variations between the two biofilm determination methods may suggest that the biofilm

  14. Proliferation in the normal FTE is a hallmark of the follicular phase, not BRCA mutation status.

    Science.gov (United States)

    George, Sophia H L; Milea, Anca; Shaw, Patricia A

    2012-11-15

    Women who have inherited germline mutations of BRCA1/BRCA2 are at increased risk of developing high-grade serous carcinoma, and many of these cancers arise in the distal fimbriated end of the fallopian tube. We have previously shown that the fallopian tube epithelia of BRCA1 mutation carriers (FTE-BRCA) have altered signaling pathways compared to nonmutation carriers. In this study, we sought to determine whether these differences result in a proliferative advantage to the epithelia in this high-risk patient population and to investigate whether the postovulation environment of the FTE-BRCA compared to FTE from nonmutation carriers experiences a differential abundance of immune cells. Immunohistochemistry for Ki67, CD3, CD8, CD20, and CD68 was performed on histologically normal tubal epithelium (ampulla, n = 83), fimbria (n = 18) with known ovarian cycle status and germline mutation status and for Ki67 on fimbrial epithelium from women (n = 144) with and without BRCA1 or BRCA2 mutations who underwent risk-reducing salpingo-oophorectomy (RRSO). Serous tubal intraepithelial carcinomas (STIC) with concomitant cancer (n = 15) were also analyzed for presence of immune infiltrates. All slides were digitized and analyzed using automated image analysis software. There was no significant difference in the proliferative index in histologically normal FTE between BRCA1/BRCA2 and non-BRCA, in 144 fimbriae and 83 ampullae. The FTE-BRCA1 epithelia did not exhibit a differential presence of lymphocytes or macrophages, however more macrophages were present in the luteal phase compared to the follicular phase epithelia. In STICs macrophages were more abundant than lymphocytes with an incremental increase noted with disease progression. BRCA1/2 mutation carriers exhibited no significant increase in proliferation in the fallopian tube epithelial cells either in the ampulla or fimbriated ends of the tube. Rather, a significant proliferative increase was defined in the cases determined

  15. Incidental serous tubal intraepithelial carcinoma and early invasive serous carcinoma in the nonprophylactic setting: analysis of a case series.

    Science.gov (United States)

    Morrison, Jane C; Blanco, Luis Z; Vang, Russell; Ronnett, Brigitte M

    2015-04-01

    A precursor for invasive ovarian/pelvic high-grade serous carcinoma, termed serous tubal intraepithelial carcinoma (STIC), has been identified and characterized through careful analysis of the fallopian tubes in both prophylactic salpingo-oophorectomy specimens obtained from women with either a family history of breast and/or ovarian cancer or germline mutations of BRCA1 and BRCA2 and in cases of pelvic high-grade serous carcinoma. Data on incidental STICs and clinically occult microscopic invasive high-grade serous carcinomas are limited. We analyzed the clinicopathologic features of 22 cases, including 15 pure STICs and 7 STICs associated with microscopic invasive high-grade serous carcinomas, identified incidentally in fallopian tubes removed for nonprophylactic indications. Patient age ranged from 39 to 79 years (mean: 62.7; median: 61), with only 1 patient under the age of 50. No patients were known to carry BRCA1 or BRCA2 mutations. Of the 12 pure STICs for which the location in the fallopian tube could be established, 9 were in the fimbriated portion, 1 was at the junction of the fimbria and infundibulum, and 2 were in the nonfimbriated tube. Of the 7 STICs with associated invasive high-grade serous carcinoma, 3 were located in the fimbriated portion, 2 were at the junction of the fimbria and infundibulum, and 2 were in the nonfimbriated tube. The invasive components were in the fallopian tube in 6 cases, 4 in subepithelial stroma of tubal mucosa, and 2 as an intramucosal (exophytic) luminal lesion without invasion of underlying subepithelial stroma (size range: 1 to 4 mm). The remaining case had a microscopic focus of high-grade serous carcinoma within the ipsilateral ovary (1.3 mm cortical focus) identified only on deeper sections, without an associated invasive component in the fallopian tube. The preferential finding of atypical epithelium with the cytologic features of high-grade serous carcinoma, namely STIC, in the fallopian tubes rather than the

  16. Agentes bacterianos enteropatogênicos em suínos de diferentes faixas etárias e perfil de resistência a antimicrobianos de cepas de Escherichia coli e Salmonella spp Enteropathogenic bacterial agents in pigs of different age groups and profile of resistance in strains of Escherichia coli and Salmonella spp. to antimicrobial agents

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    Álvaro Menin

    2008-09-01

    Full Text Available As enterites infecciosas bacterianas provocam severas perdas para a indústria suína em todo o mundo. Os objetivos deste trabalho foram determinar os agentes bacterianos, associados com a ocorrência de diarréia em suínos, em diferentes faixas etárias, no Estado de Santa Catarina, Brasil, e verificar o perfil de resistência das cepas de Escherichia coli e Salmonella spp, frente aos principais antimicrobianos utilizados em granjas de suínos. Os principais gêneros/espécies bacterianos diagnosticados foram Escherichia coli, Clostridium spp, Salmonella spp Brachyspira hyodysenteriae, Brachyspira pilosicoli e Lawsonia intracellularis. Os fatores de virulência de E. coli mais prevalentes na fase de maternidade foram F5 / (K99 20%, F6 / (987P 16,3%, F42 6,8% e F41 5,7%, já nas fases de creche e terminação, predominaram cepas com fimbrias F4 (K88 11,2% e 5,4%, respectivamente. Para E. coli os maiores índices de resistência foram encontrados para oxitetraciclina (94% e tetraciclina (89,5% e os menores índices de resistência para neomicina (55%, ceftiofur (57,4%. Quanto às amostras de Salmonella spp, estas apresentaram maior resistência à oxitetraciclina (77%, e à tetraciclina (42,1% e menor à gentamicina (3,5% e amoxicilina (4,8%.Infectious bacterial enteritis causes severe losses to the swine industry worldwide. The objective of this study was to determine the epidemiology of bacterial agents that are associated with the occurrence of diarrhea in pigs at different age groups, and to verify the profile of resistance of strains of Escherichia coli and Salmonella spp to the main antimicrobial agents. The main bacterial species diagnosed were Escherichia coli, Clostridium spp, Salmonella spp, Brachyspira hyodysenteriae, Brachyspira pilosicoli and Lawsonia intracellularis. The E. coli virulence factors of higher prevalence in preweaning piglets were F5 / (K99 20%, F6 / (987P 16.3%, F42 6.8% and F41 5.7%, whereas at the nursery and with

  17. The role of Actinobacillus actinomycetemcomitans fimbrial adhesin on MMP-8 activity in aggressive periodontitis pathogenesis

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    Rini Devijanti Ridwan

    2012-12-01

    Full Text Available Background: Actinobacillus actinomycetemcomitans (A. actinomycetemcomitans is Gram negative and a major bacterial agent associated with aggressive periodontitis in young adult, this bacteria was an important factor in pathogenesis of aggressive periodontitis. A. actinomycetemcomitans possesses fimbriae with an adhesin protein that was the first bacterial molecules to make physical contact with host. Purpose: The objective of this research was to analyzed the influence of A. actinomycetemcomitans fimbrial adhesin protein induction on MMP-8 activity. Methods: The research was an experimental laboratory study, the step in this study were isolation and identification A. actinomycetemcomitans, characterize A. actinomycetemcomitans adhesin and study the role of A. actinomycetemcomitans adhesin in Wistar rats. Results: The result of this research on the role of adhesin in Wistar rats after analysis with Analysis of Variance (ANOVA showed significant differences in the control group with group induction with A. actinomycetemcomitans, A. actinomycetemcomitans plus adhesin and adhesin. MMP-8 activity increased with induction A. actinomycetemcomitans and 24 kDa A. actinomycetemcomitans adhesin. This fimbrial adhesin protein showed that A. actinomycetemcomitans has the ability to adhesion, colonization and invasion for host in aggressive periodontitis pathogenesis. Conclusion: A. actinomycetemcomitans fimbrial adhesin protein induction increasing MMP-8 activity for aggressive periodontitis pathogenesis.Latar belakang: A. actinomycetemcomitans merupakan salah satu bakteri Gram negatif yang terkait dengan periodontitis agresif yang menyerang penderita usia muda dan merupakan faktor penting dalam patogenesis periodontitis agresif. A. actimycetemcomitans mempunyai fimbriae dengan protein adhesin yang merupakan molekul pertama dari bakteri untuk melakukan kontak fisik dengan host. Tujuan: Tujuan penelitian ini adalah menganalisis pengaruh induksi adhesin A

  18. Detecção de cepas patogênicas pela PCR multiplex e avaliação da sensibilidade a antimicrobianos de Escherichia coli isoladas de leitões diarréicos Detection of pathogenic strains by multiplex PCR and antimicrobial sensitivity of Escherichia coli isolated from piglets

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    N.R. Macêdo

    2007-10-01

    Full Text Available Avaliou-se a freqüência dos genes de fímbrias (K88, K99, 987P, F18 e F41 e toxinas (LT, Stb, StaP e Stx2e de cepas de E. coli isoladas de leitões com diarréia usando a técnica de PCR multiplex com primers específicos para esses genes, e estudou-se o padrão de sensibilidade das cepas patogênicas pelo método de difusão em disco ao florfenicol, ceftiofur sódico, colistina, fosfomicina, neomicina, norfloxacina, sulfa + trimetoprim, doxiciclina, tetraciclina e lincomicina. Foram utilizadas 144 amostras de E.coli isoladas de leitões com diarréia, provenientes de granjas localizadas no estado de Minas Gerais. Dessas, 42 (29,2% foram positivas para pelo menos um dos fatores de virulência testados. Dentre essas 42 amostras, 23 (54,8% apresentaram genes de fímbria e toxina, sete (16,6% apresentaram somente genes de toxinas e 12 (28,6% amostras somente genes de fímbria. O resultado do teste de sensibilidade aos antimicrobianos demonstrou que o florfenicol (89,5 % e o ceftiofur sódico (84,2% foram as drogas de melhor eficácia in vitro sobre cepas de E. coli com fatores de virulência.The frequency of virulence determinants genes for fimbrial adhesions (K88, K99, 987P, F18 and F41 and toxins (LT, Stb, StaP and Stx2e in E. coli strains isolated from diarrheic piglets using the multiplex polymerase chain reaction assay with specific primers for these genes was studied. The antimicrobial sensitivity pattern of pathogenic isolates for florfenicol, sodium ceftiofur, colistin, fosfomycin, neomycin, norfloxacin, sulfa + trimetoprim, doxycycline, tetracycline and lincomycin was also tested using the disk diffusion method. E. coli were isolated from 144 diarrheic piglets from farms in the state of Minas Gerais. Forty-two out of 144 studied samples (29.2% were positive for at least one tested virulence factor. Out of these 42, 23 samples (54.8% contained fimbria and toxin genes, seven (16.6% samples had genes for toxins only and 12 (28.6% samples

  19. Isolated torsion of the fallopian tube in a menopausal woman and a pre-pubertal girl: two case reports.

    Science.gov (United States)

    Toyoshima, Masafumi; Mori, Hikaru; Kudo, Kei; Yodogawa, Yuki; Sato, Kazuyo; Kudo, Takako; Igeta, Saori; Makino, Hiromitsu; Shima, Takashi; Matsuura, Rui; Ishigaki, Nobuko; Akagi, Kozo; Takeyama, Yoichi; Iwahashi, Hideki; Yoshinaga, Kosuke

    2015-11-17

    Isolated torsion of the fallopian tube without an ovarian abnormality is an uncommon event, with an incidence of approximately 1 in 1,500,000 females. Isolated torsion of the fallopian tube occurs mostly in reproductive-aged women, and is thus extremely rare in menopausal women and pre-pubertal girls. In case 1, 63-year-old Japanese woman presented with a 2-day history of acute lower abdominal pain. Menopause occurred at 53 years of age. Pelvic ultrasonography showed an enlarged mass (73 × 47 mm) on the right side of her uterus. An urgent laparoscopy was performed based on a presumptive diagnosis of right ovarian tumor torsion. During the laparoscopy, we noted a black, necrotic, solid tumor arising from the distal end of her right fimbria. Her right fallopian tube was twisted with the tumor, but her right ovary was normal and not involved. A laparoscopic tumorectomy with a right salpingectomy was performed. Her post-operative course was uneventful. In case 2, a 10-year-old Japanese girl presented with a 1-day history of lower abdominal pain associated with nausea and vomiting. Menarche had occurred 2 months earlier. A computed tomography and magnetic resonance imaging examination demonstrated a dilated tubal cystic mass with a normal uterus and bilateral ovaries. An urgent laparoscopy was performed based on a presumptive diagnosis of right fallopian tube torsion. During laparoscopy, her right fallopian tube was noted to be dark red, dilated, and twisted several times. Her right fimbria was necrotic-appearing and could not be preserved. Therefore, a laparoscopic right salpingectomy was performed. A histologic examination revealed ischemic changes with congestion of her right fallopian tube, which was consistent with tubal torsion. She had an uncomplicated post-operative course. We have presented two very rare cases of isolated fallopian tubal torsion. Radiologic interventions, such as computed tomography and magnetic resonance imaging, in addition to

  20. The influence of adhesin protein from Aggregatibacter actinomycetemcomitans on IL-8 and MMP-8 titre in aggressive periodontitis

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    Rini Revijanti Ridwan

    2015-03-01

    Full Text Available Background: Adhesion can actually be considered as a part of both a powerful survival mechanism and a virulence mechanism for bacterial pathogens. Bacterial adhesin is an instrument for bacteria to do invasion to host. Bacterial adhesin depends on ligand interaction as a signaling mediator that will influence invasion and increase pro and anti-inflammatory because of the influence of the receptors of innate immune response. Aggregatibacter actimycetemcomitans has fimbriae included in type IV pili containing mostly with protein weighed 6.5 kDa and at least with protein weighed 54 kDa. Purpose: The purpose of this research is to analyze the influence of the induction of adhesin protein derived from A. actinomycetemcomitans on IL-8 and MMP-8 titre of Wistar rats. Methods: Adhesin protein derived from A. actinomycetemcomitans weighed 24 kDa was induced on the maxillary first molar sulcus of Wistar rats to prove that adhesin protein could affect IL-8 and MMP-8 titre. Next, to determine its influence, Elisa technique was conducted. Results: It is known that the levels of IL-8 and MMP-8 titre were increased in the group induced with adhesin protein derived from A. actinomycetemcomitans compared with the control group. Conclusion: It can be concluded that adhesin protein derived from A. actinomycetemcomitans can cause alveolar bone damage through the increasing levels of IL-8 and MMP-8 in aggressive periodontitis.

  1. Epigenetic control of virulence gene expression in Pseudomonas aeruginosa by a LysR-type transcription regulator.

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    Keith H Turner

    2009-12-01

    Full Text Available Phenotypic variation within an isogenic bacterial population is thought to ensure the survival of a subset of cells in adverse conditions. The opportunistic pathogen Pseudomonas aeruginosa variably expresses several phenotypes, including antibiotic resistance, biofilm formation, and the production of CupA fimbriae. Here we describe a previously unidentified bistable switch in P. aeruginosa. This switch controls the expression of a diverse set of genes, including aprA, which encodes the secreted virulence factor alkaline protease. We present evidence that bistable expression of PA2432, herein named bexR (bistable expression regulator, which encodes a LysR-type transcription regulator, controls this switch. In particular, using DNA microarrays, quantitative RT-PCR analysis, chromatin immunoprecipitation, and reporter gene fusions, we identify genes directly under the control of BexR and show that these genes are bistably expressed. Furthermore, we show that bexR is itself bistably expressed and positively autoregulated. Finally, using single-cell analyses of a GFP reporter fusion, we present evidence that positive autoregulation of bexR is necessary for bistable expression of the BexR regulon. Our findings suggest that a positive feedback loop involving a LysR-type transcription regulator serves as the basis for an epigenetic switch that controls virulence gene expression in P. aeruginosa.

  2. Salmonella biofilm formation on Aspergillus niger involves cellulose--chitin interactions.

    Directory of Open Access Journals (Sweden)

    Maria T Brandl

    Full Text Available Salmonella cycles between host and nonhost environments, where it can become an active member of complex microbial communities. The role of fungi in the environmental adaptation of enteric pathogens remains relatively unexplored. We have discovered that S. enterica Typhimurium rapidly attaches to and forms biofilms on the hyphae of the common fungus, Aspergillus niger. Several Salmonella enterica serovars displayed a similar interaction, whereas other bacterial species were unable to bind to the fungus. Bacterial attachment to chitin, a major constituent of fungal cell walls, mirrored this specificity. Pre-incubation of S. Typhimurium with N-acetylglucosamine, the monomeric component of chitin, reduced binding to chitin beads by as much as 727-fold and inhibited attachment to A. niger hyphae considerably. A cellulose-deficient mutant of S. Typhimurium failed to attach to chitin beads and to the fungus. Complementation of this mutant with the cellulose operon restored binding to chitin beads to 79% of that of the parental strain and allowed for attachment and biofilm formation on A. niger, indicating that cellulose is involved in bacterial attachment to the fungus via the chitin component of its cell wall. In contrast to cellulose, S. Typhimurium curli fimbriae were not required for attachment and biofilm development on the hyphae but were critical for its stability. Our results suggest that cellulose-chitin interactions are required for the production of mixed Salmonella-A. niger biofilms, and support the hypothesis that encounters with chitinaceous alternate hosts may contribute to the ecological success of human pathogens.

  3. Comparative Genome Analysis of Enterobacter cloacae

    Science.gov (United States)

    Liu, Wing-Yee; Wong, Chi-Fat; Chung, Karl Ming-Kar; Jiang, Jing-Wei; Leung, Frederick Chi-Ching

    2013-01-01

    The Enterobacter cloacae species includes an extremely diverse group of bacteria that are associated with plants, soil and humans. Publication of the complete genome sequence of the plant growth-promoting endophytic E. cloacae subsp. cloacae ENHKU01 provided an opportunity to perform the first comparative genome analysis between strains of this dynamic species. Examination of the pan-genome of E. cloacae showed that the conserved core genome retains the general physiological and survival genes of the species, while genomic factors in plasmids and variable regions determine the virulence of the human pathogenic E. cloacae strain; additionally, the diversity of fimbriae contributes to variation in colonization and host determination of different E. cloacae strains. Comparative genome analysis further illustrated that E. cloacae strains possess multiple mechanisms for antagonistic action against other microorganisms, which involve the production of siderophores and various antimicrobial compounds, such as bacteriocins, chitinases and antibiotic resistance proteins. The presence of Type VI secretion systems is expected to provide further fitness advantages for E. cloacae in microbial competition, thus allowing it to survive in different environments. Competition assays were performed to support our observations in genomic analysis, where E. cloacae subsp. cloacae ENHKU01 demonstrated antagonistic activities against a wide range of plant pathogenic fungal and bacterial species. PMID:24069314

  4. Constraints on lateral gene transfer in promoting fimbrial usher protein diversity and function.

    Science.gov (United States)

    Stubenrauch, Christopher J; Dougan, Gordon; Lithgow, Trevor; Heinz, Eva

    2017-11-01

    Fimbriae are long, adhesive structures widespread throughout members of the family Enterobacteriaceae. They are multimeric extrusions, which are moved out of the bacterial cell through an integral outer membrane protein called usher. The complex folding mechanics of the usher protein were recently revealed to be catalysed by the membrane-embedded translocation and assembly module (TAM). Here, we examine the diversity of usher proteins across a wide range of extraintestinal (ExPEC) and enteropathogenic (EPEC) Escherichia coli , and further focus on a so far undescribed chaperone-usher system, with this usher referred to as UshC. The fimbrial system containing UshC is distributed across a discrete set of EPEC types, including model strains like E2348/67, as well as ExPEC ST131, currently the most prominent multi-drug-resistant uropathogenic E. coli strain worldwide. Deletion of the TAM from a naive strain of E. coli results in a drastic time delay in folding of UshC, which can be observed for a protein from EPEC as well as for two introduced proteins from related organisms, Yersinia and Enterobacter We suggest that this models why the TAM machinery is essential for efficient folding of proteins acquired via lateral gene transfer. © 2017 The Authors.

  5. Ultraestructura del bambú Guadua macclurei (Poaceae: Bambusoideae de Costa Rica

    Directory of Open Access Journals (Sweden)

    Mayra Montiel

    2006-06-01

    Full Text Available Se evaluaron ultraestructuralmente secciones de la lámina de la hoja, vaina de la hoja y rama, de una planta adulta de Guadua macclurei. Las brácteas del culmo carecen de setas orales. En la superficie adaxial de la lámina foliar se observó un patrón celular diferente al de otras especies de este género. Además, hay tricomas tricelulares y bandas de estomas de alto domo muy ordenadas y evidentes, característica que la podría diferenciar de otras especies en el género. Esta es la primera descripción microscópica de esta especie.We studied the ultrastructure of sections of leaf lamina, leaf sheath and branches of an adult plant of Guadua macclurei. Culm sheaths lack fimbriae (oral setae. The adaxial section of the leaf lamina has a particular cellular pattern, previously unreported in this genus. The tricellular trichomes, and the evident and ordered bands of high dome stomata may be useful for taxonomic purposes. To the best of our knowledge, this is the first microscopic description of this species. Rev. Biol. Trop. 54(Suppl. 2: 35-42. Epub 2006 Dec. 01.

  6. The extracellular proteome of Bifidobacterium animalis subsp. lactis BB‐12 reveals proteins with putative roles in probiotic effects

    DEFF Research Database (Denmark)

    Gilad, Ofir; Svensson, Birte; Viborg, Alexander Holm

    2011-01-01

    Probiotics are live microorganisms that exert health‐promoting effects on the human host, as demonstrated for numerous strains of the genus Bifidobacterium. To unravel the proteins involved in the interactions between the host and the extensively used and well‐studied probiotic strain Bifidobacte......Probiotics are live microorganisms that exert health‐promoting effects on the human host, as demonstrated for numerous strains of the genus Bifidobacterium. To unravel the proteins involved in the interactions between the host and the extensively used and well‐studied probiotic strain...... Bifidobacterium animalis subsp. lactis BB‐12, proteins secreted by the bacterium, i.e. belonging to the extracellular proteome present in the culture medium, were identified by 2‐DE coupled with MALDI‐TOF MS. Among the 74 distinct proteins identified, 31 are predicted to carry out their physiological role either...... functions include binding of plasminogen, formation of fimbriae, adhesion to collagen, attachment to mucin and intestinal cells as well as induction of immunomodulative response. These findings suggest a role of the proteins in colonization of the gastrointestinal tract, adhesion to host tissues...

  7. Dual Functional Core-Shell Fluorescent Ag2S@Carbon Nanostructure for Selective Assay of E. coli O157:H7 and Bactericidal Treatment.

    Science.gov (United States)

    Wang, Ning; Wei, Xing; Zheng, An-Qi; Yang, Ting; Chen, Ming-Li; Wang, Jian-Hua

    2017-03-24

    A dual functional fluorescent core-shell Ag 2 S@Carbon nanostructure is prepared by a hydrothermally assisted multi-amino synthesis approach with folic acid (FA), polyethylenimine (PEI), and mannoses (Mans) as carbon and nitrogen sources (FA-PEI-Mans-Ag 2 S nanocomposite shortly as Ag 2 S@C). The nanostructure exhibits strong fluorescent emission at λ ex /λ em = 340/450 nm with a quantum yield of 12.57 ± 0.52%. Ag 2 S@C is bound to E. coli O157:H7 via strong interaction with the Mans moiety in Ag 2 S@C with FimH proteins on the fimbriae tip in E. coli O157:H7. Fluorescence emission from Ag 2 S@C/E. coli conjugate is closely related to the content of E. coli O157:H7. Thus, a novel procedure for fluorescence assay of E. coli O157:H7 is developed, offering a detection limit of 330 cfu mL -1 . Meanwhile, the Ag 2 S@C nanostructure exhibits excellent antibacterial performance against E. coli O157:H7. A 99.9% sterilization rate can be readily achieved for E. coli O157:H7 at a concentration of 10 6 -10 7 cfu mL -1 with 3.3 or 10 μg mL -1 of Ag 2 S@C with an interaction time of 5 or 0.5 min, respectively.

  8. Incidence of Escherichia coli O157:H7 in Thailand

    International Nuclear Information System (INIS)

    Sukhumungoon, P.

    2015-01-01

    Entero hemorrhagic Escherichia coli (EHEC) especially serotype O157:H7 is one of the important food-borne pathogens because it is able to produce crucial toxins Shiga. However, the outbreak of this organism in Thailand has not been reported. Antibody to O157 antigen was detected in some Thai populations and Shiga toxin-producing E. coli were detected in low numbers of clinical specimens. Interestingly, some E. coli that showed positive to O157 fimbriae probe and lack of virulence gene were isolated from certain patients and one isolate of E. coli O157:H7 which possessed stx1, stx2v was detected in a normal child. In addition, the incidence of E. coli O157:H7 strains were monitored by the samples from cattle and retail beef in Thailand although their inability to produce toxins or produce in a low concentration was demonstrated. This review discusses the incidences of E. coli O157 in clinical and environmental samples of Thailand including the transmission possibility of this bacterium across the Thai border through food trade. (author)

  9. Temperature control of molecular circuit switch responsible for virulent phenotype expression in uropathogenic Escherichia coli

    Science.gov (United States)

    Samoilov, Michael

    2010-03-01

    The behavior and fate of biological organisms are to a large extent dictated by their environment, which can be often viewed as a collection of features and constraints governed by physics laws. Since biological systems comprise networks of molecular interactions, one such key physical property is temperature, whose variations directly affect the rates of biochemical reactions involved. For instance, temperature is known to control many gene regulatory circuits responsible for pathogenicity in bacteria. One such example is type 1 fimbriae (T1F) -- the foremost virulence factor in uropathogenic E. coli (UPEC), which accounts for 80-90% of all community-acquired urinary tract infections (UTIs). The expression of T1F is randomly `phase variable', i.e. individual cells switch between virulent/fimbriate and avirulent/afimbriate phenotypes, with rates regulated by temperature. Our computational investigation of this process, which is based on FimB/FimE recombinase-mediated inversion of fimS DNA element, offers new insights into its discrete-stochastic kinetics. In particular, it elucidates the logic of T1F control optimization to the host temperature and contributes further understanding toward the development of novel therapeutic approaches to UPEC-caused UTIs.

  10. Distribution of Escherichia coli F4 adhesion phenotypes in pigs of 15 Chinese and Western breeds and a White DurocxErhualian intercross.

    Science.gov (United States)

    Yan, Xueming; Huang, Xiang; Ren, Jun; Zou, Zhengzhi; Yang, Shujin; Ouyang, Jing; Zeng, Weihong; Yang, Bin; Xiao, Shijun; Huang, Lusheng

    2009-08-01

    Diarrhoea in newborn and weaned piglets is mainly caused by enterotoxigenic Escherichia coli (ETEC) with fimbriae F4. To investigate the prevalence of resistance to three fimbrial strains, F4ab, F4ac and F4ad, among Chinese indigenous pigs and Western commercial pigs introduced into China, we determined the ETEC F4 adhesion phenotypes in 292 pure-bred piglets from three Western commercial breeds and 12 Chinese indigenous breeds, and a total of 1093 adult pigs in a White DurocxErhualian intercross, by an in vitro microscopic adhesion assay. All the Tibet and Lantang pigs and a majority of the Erhualian and Rongchang pigs were resistant (nonadherent) to ETEC F4 whereas all the Laiwu pigs and most of the Jiangquhai and Tongcheng pigs were susceptible (adhesive) to at least one of the F4 strains. Yushan Black pigs were uniformly resistant to F4ab, and Jinhua pigs were predominantly resistant to F4ac. Susceptible and resistant animals were observed in the other breeds, indicating that diarrhoea caused by ETEC F4 could be prevalent in these breeds. This study confirmed the existence of eight previously reported F4 adhesion patterns, and supported the assumption that the three F4 receptors are encoded by distinct loci. Expression of the weakly adherent phenotype was observed in six pure-bred piglets and 90 adult F(2)/F(3) animals, and the inheritance of this phenotype and its correlation with susceptibility to disease are still not known.

  11. F4-related mutation and expression analysis of the aminopeptidase N gene in pigs.

    Science.gov (United States)

    Goetstouwers, T; Van Poucke, M; Nguyen, V U; Melkebeek, V; Coddens, A; Deforce, D; Cox, E; Peelman, L J

    2014-05-01

    Intestinal infections with F4 enterotoxigenic Escherichia coli (ETEC) are worldwide an important cause of diarrhea in neonatal and recently weaned pigs. Adherence of F4 ETEC to the small intestine by binding to specific receptors is mediated by F4 fimbriae. Porcine aminopeptidase N (ANPEP) was recently identified as a new F4 receptor. In this study, 7 coding mutations and 1 mutation in the 3' untranslated region (3' UTR)were identified in ANPEP by reverse transcriptase (RT-) PCR and sequencing using 3 F4 receptor-positive (F4R+) and 2 F4 receptor-negative (F4R-) pigs, which were F4 phenotyped based on the MUC4 TaqMan, oral immunization, and the in vitro villous adhesion assay. Three potential differential mutations (g.2615C > T, g.8214A > G, and g.16875C > G) identified by comparative analysis between the 3 F4R+ and 2 F4R- pigs were genotyped in 41 additional F4 phenotyped pigs. However, none of these 3 mutations could be associated with F4 ETEC susceptibility. In addition, the RT-PCR experiments did not reveal any differential expression or alternative splicing in the small intestine of F4R+ and F4R- pigs. In conclusion, we hypothesize that the difference in F4 binding to ANPEP is due to modifications in its carbohydrate moieties.

  12. Ultrastructural and chemotaxonomic analysis of a xylanolytic strain of Cryptococcus adeliensis isolated from sheep droppings in Spain.

    Science.gov (United States)

    Velázquez, Encarna; del Villar, María; Grondona, Isabel; Monte, Enrique; González-Villa, Tomás

    2006-09-01

    Cryptococcus adeliensis was initially described as a psycrophilic species containing a single strain CBS 8351(T) isolated from decayed algae in Terre Adelie (Antartida). Later, a second strain of this species was isolated from an immunosuppressed patient affected by leukaemia in Germany and recently several strains from this species have been found in human patients and pigeon droppings of the same country. In this study, we isolated from sheep droppings in Spain a xylanolytic strain named LEVX01 that was phenotypically related to the strain CBS 8351(T) and showed a 100% similarity in the D1/D2 domain and 5.8S-ITS region sequences with respect to the remaining described strains of C. adeliensis. These findings suggest that this species has a wide geographical distribution and that the animal faeces are a common habitat for C. adeliensis. The chemotaxonomic analyses showed the absence of detectable amounts of xylose in the cell walls of the strains LEVX01 and CBS8351(T) in contrast to other Cryptococcus species. Interestingly, the ultrastructural study showed the presence of fimbriae in these two strains that could be involved in the attachment to the host cells and, as occurs in Candida albicans, they could also be a pathogenicity factor for the man.

  13. Nucleotide sequence of pOLA52: a conjugative IncX1 plasmid from Escherichia coli which enables biofilm formation and multidrug efflux

    DEFF Research Database (Denmark)

    Norman, Anders; Hansen, Lars H.; She, Qunxin

    2008-01-01

    . The plasmid was also classified as IncX1 with incompatibility testing. The conjugal transfer and plasmid maintenance regions of pOLA52 therefore seem to represent IncX1 orthologues of the well-characterized IncX2 plasmid R6K. Sequence homology searches in GenBank also suggested a considerably higher...... of type 3 fimbriae (mrkABCDF). The plasmid was found to be 51,602 bp long with 68 putative genes. About half of the plasmid constituted a conserved IncX1-type backbone with predicted regions for conjugation, replication and partitioning, as well as a toxin/antitoxin (TA) plasmid addiction system...... prevalence of IncX1 group plasmids than IncX2. The 21 kb 'genetic load' region of pOLA52 was shown to consist of a mosaic, among other things a fragmented Tn3 transposon encoding ampicillin resistance. Most notably the oqxAB and mrkABCDF cassettes were contained within two composite transposons (Tn6010...

  14. Delphi definition of the EADC-ADNI Harmonized Protocol for hippocampal segmentation on magnetic resonance.

    Science.gov (United States)

    Boccardi, Marina; Bocchetta, Martina; Apostolova, Liana G; Barnes, Josephine; Bartzokis, George; Corbetta, Gabriele; DeCarli, Charles; deToledo-Morrell, Leyla; Firbank, Michael; Ganzola, Rossana; Gerritsen, Lotte; Henneman, Wouter; Killiany, Ronald J; Malykhin, Nikolai; Pasqualetti, Patrizio; Pruessner, Jens C; Redolfi, Alberto; Robitaille, Nicolas; Soininen, Hilkka; Tolomeo, Daniele; Wang, Lei; Watson, Craig; Wolf, Henrike; Duvernoy, Henri; Duchesne, Simon; Jack, Clifford R; Frisoni, Giovanni B

    2015-02-01

    This study aimed to have international experts converge on a harmonized definition of whole hippocampus boundaries and segmentation procedures, to define standard operating procedures for magnetic resonance (MR)-based manual hippocampal segmentation. The panel received a questionnaire regarding whole hippocampus boundaries and segmentation procedures. Quantitative information was supplied to allow evidence-based answers. A recursive and anonymous Delphi procedure was used to achieve convergence. Significance of agreement among panelists was assessed by exact probability on Fisher's and binomial tests. Agreement was significant on the inclusion of alveus/fimbria (P = .021), whole hippocampal tail (P = .013), medial border of the body according to visible morphology (P = .0006), and on this combined set of features (P = .001). This definition captures 100% of hippocampal tissue, 100% of Alzheimer's disease-related atrophy, and demonstrated good reliability on preliminary intrarater (0.98) and inter-rater (0.94) estimates. Consensus was achieved among international experts with respect to hippocampal segmentation using MR resulting in a harmonized segmentation protocol. Copyright © 2015 The Alzheimer's Association. Published by Elsevier Inc. All rights reserved.

  15. Effect of intrahippocampal kainic acid injections and surgical lesions on neurotransmitters in hippocampus and septum

    Energy Technology Data Exchange (ETDEWEB)

    Fonnum, F; Walaas, I

    1978-01-01

    Local injection of kainic acid (2 ..mu..g) was accompanied by destruction of intrinsic neurons in the dorsal part of hippocampus. The lesion was accompanied by a 75% reduction in glutamate decarboxylase activity, a 60% reduction in the high affinity uptake of L-glutamate, a 40 to 60% reduction in the endogeneous levels of aspartate, glutamate and GABA and no changes in the activities of choline acetyltransferase or aromatic amino acid decarboxylase in the dorsal hippocampus. Unilateral destruction of neurons in the dorsal hippocampus was followed by a 20 to 40% reduction in the high affinity uptake of glutamate in lateral, but not in medial septum, on both sides. There was no reduction in choline acetyltransferase, glutamate decarboxylase or aromatic amino acid decarboxylase activities in the lateral or medial part of the septum. Transection of fimbria and superior fornix was accompanied by a severe reduction in choline acetyltransferase and aromatic amino acid decarboxylase activity in hippocampus, in the high affinity uptake of glutamate and in the endogenous level of glutamate in the lateral septum. The results are consistent with the concept that in the hippocampus kainic acid destroys intrinsic neurons and not afferent fibres. It seems therefore that all GABAergic fibres in the hippocampus belong to intrinsic neurons whereas glutamergic and aspartergic neurons belong partly to local neurons. The connection from the hippocampus to the lateral septum probably uses glutamate as a transmitter.

  16. An investigation of virulence factors of Legionella pneumophila environmental isolates

    Directory of Open Access Journals (Sweden)

    Elif Özlem Arslan-Aydoğdu

    Full Text Available ABSTRACT Nine Legionella pneumophila strains isolated from cooling towers and a standard strain (L. pneumophila serogroup 1, ATCC 33152, Philadelphia 1 were analyzed and compared in terms of motility, flagella structure, ability to form biofilms, enzymatic activities (hemolysin, nucleases, protease, phospholipase A, phospholipase C, acid phosphatase, alkaline phosphatase and lipase, hemagglutination capabilities, and pathogenicity in various host cells (Acanthamoeba castellanii ATCC 30234, mouse peritoneal macrophages and human peripheral monocytes. All the isolates of bacteria appeared to be motile and polar-flagellated and possessed the type-IV fimbria. Upon the evaluation of virulence factors, isolate 4 was found to be the most pathogenic strain, while 6 out of the 9 isolates (the isolates 1, 2, 3, 4, 5, and 7 were more virulent than the ATCC 33152 strain. The different bacterial strains exhibited differences in properties such as adhesion, penetration and reproduction in the hosts, and preferred host type. To our knowledge, this is the first study to compare the virulence of environmental L. pneumophila strains isolated in Turkey, and it provides important information relevant for understanding the epidemiology of L. pneumophila.

  17. Fungal lectin of Peltigera canina induces chemotropism of compatible Nostoc cells by constriction-relaxation pulses of cyanobiont cytoskeleton.

    Science.gov (United States)

    Díaz, Eva Maria; Vicente-Manzanares, Miguel; Sacristan, Mara; Vicente, Carlos; Legaz, Maria-Estrella

    2011-10-01

    A glycosylated arginase acting as a fungal lectin from Peltigera canina is able to produce recruitment of cyanobiont Nostoc cells and their adhesion to the hyphal surface. This implies that the cyanobiont would develop organelles to motility towards the chemoattractant. However when visualized by transmission electron microscopy, Nostoc cells recently isolated from P. canina thallus do not reveal any motile, superficial organelles, although their surface was covered by small spindles and serrated layer related to gliding. The use of S-(3,4-dichlorobenzyl)isothiourea, blebbistatin, phalloidin and latrunculin A provide circumstantial evidence that actin microfilaments rather than MreB, the actin-like protein from prokaryota, and, probably, an ATPase which develops contractile function similar to that of myosin II, are involved in cell motility. These experimental facts, the absence of superficial elements (fimbriae, pili or flagellum) related to cell movement, and the appearance of sunken cells during of after movement verified by scanning electron microscopy, support the hypothesis that the motility of lichen cyanobionts could be achieved by contraction-relaxation episodes of the cytoskeleton induced by fungal lectin act as a chemoattractant.

  18. Porphyromonas gingivalis Fim-A genotype distribution among Colombians

    Science.gov (United States)

    Jaramillo, Adriana; Parra, Beatriz; Botero, Javier Enrique; Contreras, Adolfo

    2015-01-01

    Introduction: Porphyromonas gingivalis is associated with periodontitis and exhibit a wide array of virulence factors, including fimbriae which is encoded by the FimA gene representing six known genotypes. Objetive: To identify FimA genotypes of P. gingivalis in subjects from Cali-Colombia, including the co-infection with Aggregatibacter actinomycetemcomitans, Treponema denticola, and Tannerella forsythia. Methods: Subgingival samples were collected from 151 people exhibiting diverse periodontal condition. The occurrence of P. gingivalis, FimA genotypes and other bacteria was determined by PCR. Results: P. gingivalis was positive in 85 patients. Genotype FimA II was more prevalent without reach significant differences among study groups (54.3%), FimA IV was also prevalent in gingivitis (13.0%). A high correlation (p= 0.000) was found among P. gingivalis, T. denticola, and T. forsythia co-infection. The FimA II genotype correlated with concomitant detection of T. denticola and T. forsythia. Conclusions: Porphyromonas gingivalis was high even in the healthy group at the study population. A trend toward a greater frequency of FimA II genotype in patients with moderate and severe periodontitis was determined. The FimA II genotype was also associated with increased pocket depth, greater loss of attachment level, and patients co-infected with T. denticola and T. forsythia. PMID:26600627

  19. The Molecular Fingerprint of High Grade Serous Ovarian Cancer Reflects Its Fallopian Tube Origin

    Directory of Open Access Journals (Sweden)

    Thomas Meyer

    2013-03-01

    Full Text Available High grade serous ovarian cancer (HGSC, the most lethal and frequent type of epithelial ovarian cancer (EOC, has poor long term prognosis due to a combination of factors: late detection, great metastatic potential and the capacity to develop resistance to available therapeutic drugs. Furthermore, there has been considerable controversy concerning the etiology of this malignancy. New studies, both clinical and molecular, strongly suggest that HGSC originates not from the surface of the ovary, but from the epithelial layer of the neighboring fallopian tube fimbriae. In this paper we summarize data supporting the central role of fallopian tube epithelium in the development of HGSC. Specifically, we address cellular pathways and regulatory mechanisms which are modulated in the process of transformation, but also genetic changes which accumulate during disease progression. Similarities between fallopian tube mucosa and the malignant tissue of HGSC warrant a closer analysis of homeostatic mechanisms in healthy epithelium in order to elucidate key steps in disease development. Finally, we highlight the importance of the cancer stem cell (CSC identification and understanding of its niche regulation for improvement of therapeutic strategies.

  20. Activation of human gingival epithelial cells by cell-surface components of black-pigmented bacteria: augmentation of production of interleukin-8, granulocyte colony-stimulating factor and granulocyte-macrophage colony-stimulating factor and expression of intercellular adhesion molecule 1.

    Science.gov (United States)

    Sugiyama, A; Uehara, A; Iki, K; Matsushita, K; Nakamura, R; Ogawa, T; Sugawara, S; Takada, H

    2002-01-01

    Black-pigmented anaerobic bacteria, such as Porphyromonas gingivalis and Prevotella intermedia, are amongst the predominant bacteria in periodontal pockets and have been implicated in periodontal diseases. To elucidate the roles of gingival keratinocytes, which are the first cells encountered by oral bacteria in periodontal diseases, human gingival keratinocytes in primary culture were stimulated with cell-surface components of P gingivalis and Pr. intermedia. A glycoprotein fraction from Pr. intermedia (PGP) clearly augmented the release of interleukin-8, granulocyte colony-stimulating factor and granulocyte-macrophage colony-stimulating factor, as determined by enzyme-linked immunosorbent assay. This PGP also induced expression of intercellular adhesion molecule-1 (ICAM-1), as determined by flow cytometry. The augmentation of mRNA expression for these molecules was also confirmed by reverse transcription PCR. In contrast, lipopolysaccharide (LPS) from Pr. intermedia and Escherichia coli was completely inactive in these assays. LPS fraction and purified fimbriae from P gingivalis exhibited weak activities. Cytokine production and ICAM-1 expression by gingival keratinocytes might cause accumulation and activation of neutrophils in the epithelium and, therefore, may be involved in the initiation and development of inflammation in periodontal tissues.

  1. Milk-based nutraceutical for treating autoimmune arthritis via the stimulation of IL-10- and TGF-β-producing CD39+ regulatory T cells.

    Directory of Open Access Journals (Sweden)

    Massimo Maddaloni

    Full Text Available Autoimmune diseases arise from the loss of tolerance to self, and because the etiologies of such diseases are largely unknown, symptomatic treatments rely on anti-inflammatory and analgesic agents. Tolerogenic treatments that can reverse disease are preferred, but again, often thwarted by not knowing the responsible auto-antigens (auto-Ags. Hence, a viable alternative to stimulating regulatory T cells (Tregs is to induce bystander tolerance. Colonization factor antigen I (CFA/I has been shown to evoke bystander immunity and to hasten Ag-specific Treg development independent of auto-Ag. To translate in treating human autoimmune diseases, the food-based Lactococcus was engineered to express CFA/I fimbriae, and Lactococcus-CFA/I fermented milk fed to arthritic mice proved highly efficacious. Protection occurred via CD39+ Tregs producing TGF-β and IL-10 to potently suppress TNF-α production and neutrophil influx into the joints. Thus, these data demonstrate the feasibility of oral nutraceuticals for treating arthritis, and potency of protection against arthritis was improved relative to that obtained with Salmonella-CFA/I.

  2. Distribution of Porphyromonas gingivalis fimA genotypes in primary endodontic infections.

    Science.gov (United States)

    Rôças, Isabela N; Siqueira, José F

    2010-03-01

    Long fimbriae (FimA) are important virulence factors of Porphyromonas gingivalis. Based on the diversity of the fimA gene, this species is classified into 6 genotypes. This study surveyed samples from primary endodontic infections for the presence of these P. gingivalis fimA variants. Genomic DNA isolated from samples taken from 25 root canals of teeth with chronic apical periodontitis and 25 aspirates from acute apical abscess was used as template in polymerase chain reaction (PCR) assays directed toward the detection of the different P. gingivalis fimA genotypes. Porphyromonas gingivalis was detected by a 16S rRNA gene-based PCR in 36% of the total number of cases sampled (44% of chronic apical periodontitis and 28% of abscess aspirates). In cases of chronic apical periodontitis, P. gingivalis variant type IV was the most prevalent (24%), followed by types I (20%), II (16%), and III (8%). In acute abscess samples, variant type II was the most prevalent (12%), followed by types III and IV (8% of each) and type I (4%). Combinations of up to 3 different genotypes were detected in a few cases. No single fimA genotype variant or combination thereof was significantly associated with symptoms. Overall, fimA types IV (16%), II (14%), and I (12%) were the most prevalent. Findings demonstrated that different P. gingivalis fimA genotypes can be present in primary endodontic infections. Copyright 2010 Mosby, Inc. All rights reserved.

  3. A novel Pseudomonas aeruginosa bacteriophage, Ab31, a chimera formed from temperate phage PAJU2 and P. putida lytic phage AF: characteristics and mechanism of bacterial resistance.

    Directory of Open Access Journals (Sweden)

    Libera Latino

    Full Text Available A novel temperate bacteriophage of Pseudomonas aeruginosa, phage vB_PaeP_Tr60_Ab31 (alias Ab31 is described. Its genome is composed of structural genes related to those of lytic P. putida phage AF, and regulatory genes similar to those of temperate phage PAJU2. The virion structure resembles that of phage AF and other lytic Podoviridae (S. enterica Epsilon 15 and E. coli phiv10 with similar tail spikes. Ab31 was able to infect P. aeruginosa strain PA14 and two genetically related strains called Tr60 and Tr162, out of 35 diverse strains from cystic fibrosis patients. Analysis of resistant host variants revealed different phenotypes, including induction of pigment and alginate overproduction. Whole genome sequencing of resistant variants highlighted the existence of a large deletion of 234 kbp in two strains, encompassing a cluster of genes required for the production of CupA fimbriae. Stable lysogens formed by Ab31 in strain Tr60, permitted the identification of the insertion site. During colonization of the lung in cystic fibrosis patients, P. aeruginosa adapts by modifying its genome. We suggest that bacteriophages such as Ab31 may play an important role in this adaptation by selecting for bacterial characteristics that favor persistence of bacteria in the lung.

  4. Increased number of intestinal villous M cells in levamisole - pretreated weaned pigs experimentally infected with F4ac+ enterotoxigenic Escherichia coli strain

    Directory of Open Access Journals (Sweden)

    H. Valpotić

    2010-07-01

    Full Text Available Immunoprophylaxis of porcine postweaning colibacillosis (PWC caused by enterotoxigenic Escherichia coli (ETEC expressing F4 fimbriae is an unsolved problem. Just as ETEC strains can exploit intestinal microfold (M cells as the entry portal for infection, their high transcytotic ability make them an attractive target for mucosally delivered vaccines, adjuvants and therapeutics. We have developed a model of parenteral/oral immunization of 4-weeks-old pigs with either levamisole or vaccine candidate F4ac+ non-ETEC strain to study their effects on de novo differentiation of antigen-sampling M cells. Identification, localization and morphometric quantification of cytokeratin 18 positive M cells in the ileal mucosa of 6-weeks-old pigs revealed that they were: 1 exclusively located within villous epithelial layer, 2 significantly numerous (P< 0.01 in levamisole pretreated/challenged pigs, and 3 only slightly, but not significantly numerous in vaccinated/challenged pigs compared with non-pretreated/challenged control pigs. The fact that levamisole may affect the M cells frequency by increasing their numbers, makes it an interesting adjuvant to study development of an effective M cell-targeted vaccine against porcine PWC.

  5. Healthy versus Entorhinal Cortical Atrophy Identification in Asymptomatic APOE4 Carriers at Risk for Alzheimer's Disease.

    Science.gov (United States)

    Konishi, Kyoko; Joober, Ridha; Poirier, Judes; MacDonald, Kathleen; Chakravarty, Mallar; Patel, Raihaan; Breitner, John; Bohbot, Véronique D

    2018-01-01

    Early detection of Alzheimer's disease (AD) has been challenging as current biomarkers are invasive and costly. Strong predictors of future AD diagnosis include lower volume of the hippocampus and entorhinal cortex, as well as the ɛ4 allele of the Apolipoprotein E gene (APOE) gene. Therefore, studying functions that are critically mediated by the hippocampus and entorhinal cortex, such as spatial memory, in APOE ɛ4 allele carriers, may be key to the identification of individuals at risk of AD, prior to the manifestation of cognitive impairments. Using a virtual navigation task developed in-house, specifically designed to assess spatial versus non-spatial strategies, the current study is the first to differentiate functional and structural differences within APOE ɛ4 allele carriers. APOE ɛ4 allele carriers that predominantly use non-spatial strategies have decreased fMRI activity in the hippocampus and increased atrophy in the hippocampus, entorhinal cortex, and fimbria compared to APOE ɛ4 allele carriers who use spatial strategies. In contrast, APOE ɛ4 allele carriers who use spatial strategies have grey matter levels comparable to non-APOE ɛ4 allele carriers. Furthermore, in a leave-one-out analysis, grey matter in the entorhinal cortex could predict navigational strategy with 92% accuracy.

  6. Healthy versus Entorhinal Cortical Atrophy Identification in Asymptomatic APOE4 Carriers at Risk for Alzheimer’s Disease

    Science.gov (United States)

    Konishi, Kyoko; Joober, Ridha; Poirier, Judes; MacDonald, Kathleen; Chakravarty, Mallar; Patel, Raihaan; Breitner, John; Bohbot, Véronique D.

    2018-01-01

    Early detection of Alzheimer’s disease (AD) has been challenging as current biomarkers are invasive and costly. Strong predictors of future AD diagnosis include lower volume of the hippocampus and entorhinal cortex, as well as the ɛ4 allele of the Apolipoprotein E gene (APOE) gene. Therefore, studying functions that are critically mediated by the hippocampus and entorhinal cortex, such as spatial memory, in APOE ɛ4 allele carriers, may be key to the identification of individuals at risk of AD, prior to the manifestation of cognitive impairments. Using a virtual navigation task developed in-house, specifically designed to assess spatial versus non-spatial strategies, the current study is the first to differentiate functional and structural differences within APOE ɛ4 allele carriers. APOE ɛ4 allele carriers that predominantly use non-spatial strategies have decreased fMRI activity in the hippocampus and increased atrophy in the hippocampus, entorhinal cortex, and fimbria compared to APOE ɛ4 allele carriers who use spatial strategies. In contrast, APOE ɛ4 allele carriers who use spatial strategies have grey matter levels comparable to non-APOE ɛ4 allele carriers. Furthermore, in a leave-one-out analysis, grey matter in the entorhinal cortex could predict navigational strategy with 92% accuracy. PMID:29278888

  7. Determination of the optimum standardised ileal digestible sulphur amino acids to lysine ratio in weaned pigs challenged with enterotoxigenic Escherichia coli

    DEFF Research Database (Denmark)

    Capozzalo, M. M.; Resink, J. W.; Htoo, J. K.

    2017-01-01

    This experiment tested the hypothesis that pigs challenged with an enterotoxigenic strain of E. coli (ETEC) would require a higher sulphur amino acids (SAA) to Lys ratio (SAA:Lys). Pigs (n = 120) weighing 7.4 ± 0.52 kg (mean ± SD) and weaned at 27 d (Pietrain genotype, mixed sex) were stratified...... into 1 of 6 treatments based on weaning weight, sex and genotype for the F4 fimbria receptor (n = 20). Five diets were formulated with increasing ratios of standardised ileal digestible (SID) sulphurSAA:Lys. Pigs were housed in pens of 4 during an adaptation period of 6 d after which time pigs were...... housed individually. Pigs fed different SID SAA:Lys levels were infected with ETEC (5 mL, 1.13 × 108 CFU/mL, serotype O149:K91:K88) on d 8, 9, and 10 after weaning. The sixth diet, which contained 0.55 SID SAA:Lys and corresponded to current NRC recommendations, was allocated to 2 groups of pigs either...

  8. Radiation-Induced Growth Retardation and Microstructural and Metabolite Abnormalities in the Hippocampus.

    Science.gov (United States)

    Rodgers, Shaefali P; Zawaski, Janice A; Sahnoune, Iman; Leasure, J Leigh; Gaber, M Waleed

    2016-01-01

    Cranial radiotherapy (CRT) increases survival in pediatric brain-tumor patients but can cause deleterious effects. This study evaluates the acute and long-term impact of CRT delivered during childhood/adolescence on the brain and body using a rodent model. Rats received CRT, either 4 Gy fractions × 5 d (fractionated) or a cumulative dose of 20 Gy (single dose) at 28 d of age. Animals were euthanized 1 d, 5 d, or 3.5 mo after CRT. The 3.5 mo group was imaged prior to euthanasia. At 3.5 mo, we observed significant growth retardation in irradiated animals, versus controls, and the effects of single dose on brain and body weights were more severe than fractionated. Acutely single dose significantly reduced body weight but increased brain weight, whereas fractionation significantly reduced brain but not body weights, versus controls. CRT suppressed cell proliferation in the hippocampal subgranular zone acutely. Fractional anisotropy (FA) in the fimbria was significantly lower in the single dose versus controls. Hippocampal metabolite levels were significantly altered in the single dose animals, reflecting a heightened state of inflammation that was absent in the fractionated. Our findings indicate that despite the differences in severity between the doses they both demonstrated an effect on cell proliferation and growth retardation, important factors in pediatric CRT.

  9. Manipulation of Neutrophils by Porphyromonas gingivalis in the Development of Periodontitis.

    Science.gov (United States)

    Sochalska, Maja; Potempa, Jan

    2017-01-01

    The pathogenesis of the chronic periodontal disease is associated with a skewed host inflammatory response to periodontal pathogens, such as Porphyromonas gingivalis , that accounts for the majority of periodontal tissue damage. Neutrophils are the most abundant leukocytes in periodontal pockets and depending on the stage of the disease, also plentiful PMNs are present in the inflamed gingival tissue and the gingival crevice. They are the most efficient phagocytes and eliminate pathogens by a variety of means, which are either oxygen-dependent or -independent. However, these secretory lethal weapons do not strictly discriminate between pathogens and host tissue. Current studies describe conflicting findings about neutrophil involvement in periodontal disease. On one hand literature indicate that hyper-reactive neutrophils are the main immune cell type responsible for this observed tissue damage and disease progression. Deregulation of neutrophil survival and functions, such as chemotaxis, migration, secretion of antimicrobial peptides or enzymes, and production of reactive oxygen species, contribute to observed tissue injury and the clinical signs of periodontal disease. On the other hand neutrophils deficiencies in patients and mice also result in periodontal phenotype. Therefore, P. gingivalis represents a periodontal pathogen that manipulates the immune responses of PMNs, employing several virulence factors, such as gingipains, serine proteases, lipid phosphatases, or fimbriae. This review will sum up studies devoted to understanding different strategies utilized by P. gingivalis to manipulate PMNs survival and functions in order to inhibit killing by a granular content, prolong inflammation, and gain access to nutrient resources.

  10. Porphyromonas gingivalis decreases osteoblast proliferation through IL-6-RANKL/OPG and MMP-9/TIMPs pathways

    Directory of Open Access Journals (Sweden)

    Le Xuan

    2009-01-01

    Full Text Available Background: Porphyromonas gingivalis, an important periodontal pathogen, is closely associated with inflammatory alveolar bone resorption. This bacterium exerts its pathogenic effect indirectly through multiple virulence factors, such as lipopolysaccharides, fimbriae, and proteases. Another possible pathogenic path may be through a direct interaction with the host′s soft and hard tissues (e.g., alveolar bone, which could lead to periodontitis. Aims and Objectives: The aim of the present study was to investigate the direct effect of live and heat-inactivated P gingivalis on bone resorption, using an in vitro osteoblast culture model. Results: Optical microscopy and 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide MTT assay revealed that live P gingivalis induced osteoblast detachment and reduced their proliferation. This effect was specific to live bacteria and was dependent on their concentration. Live P gingivalis increased IL-6 mRNA expression and protein production and downregulated RANKL and OPG mRNA expression. The effect of live P gingivalis on bone resorption was strengthened by an increase in MMP-9 expression and its activity. This increase was accompanied by an increase in TIMP-1 and TIMP-2 mRNA expression and protein production by osteoblasts infected with live P gingivalis. Conclusion: Overall, the results suggest that direct contact of P gingivalis with osteoblasts induces bone resorption through an inflammatory pathway that involves IL-6, RANKL/OPG, and MMP-9/TIMPs.

  11. Manipulation of Neutrophils by Porphyromonas gingivalis in the Development of Periodontitis

    Directory of Open Access Journals (Sweden)

    Maja Sochalska

    2017-05-01

    Full Text Available The pathogenesis of the chronic periodontal disease is associated with a skewed host inflammatory response to periodontal pathogens, such as Porphyromonas gingivalis, that accounts for the majority of periodontal tissue damage. Neutrophils are the most abundant leukocytes in periodontal pockets and depending on the stage of the disease, also plentiful PMNs are present in the inflamed gingival tissue and the gingival crevice. They are the most efficient phagocytes and eliminate pathogens by a variety of means, which are either oxygen-dependent or -independent. However, these secretory lethal weapons do not strictly discriminate between pathogens and host tissue. Current studies describe conflicting findings about neutrophil involvement in periodontal disease. On one hand literature indicate that hyper-reactive neutrophils are the main immune cell type responsible for this observed tissue damage and disease progression. Deregulation of neutrophil survival and functions, such as chemotaxis, migration, secretion of antimicrobial peptides or enzymes, and production of reactive oxygen species, contribute to observed tissue injury and the clinical signs of periodontal disease. On the other hand neutrophils deficiencies in patients and mice also result in periodontal phenotype. Therefore, P. gingivalis represents a periodontal pathogen that manipulates the immune responses of PMNs, employing several virulence factors, such as gingipains, serine proteases, lipid phosphatases, or fimbriae. This review will sum up studies devoted to understanding different strategies utilized by P. gingivalis to manipulate PMNs survival and functions in order to inhibit killing by a granular content, prolong inflammation, and gain access to nutrient resources.

  12. Arcanobacterium pyogenes: Virulence factors, importance in mastitis etiology and therapeutic (impossibilities

    Directory of Open Access Journals (Sweden)

    Milanov Dubravka

    2011-01-01

    Full Text Available Arcanobacterium pyogenes is an opportunistic pathogen, a causative agent of suppurative infections of organs and tissues in economically important livestock species. Most frequently this bacteria is isolated from inflamed lung lesions in pigs and cattle, in samples of uterine mucus of cows with endometritis and milk from cows with clinical mastitis. A. pyogenes possesses a number of virulence factors: cholesterol-dependent cytolysin (pyolysin, two neuraminidases, several proteases, extracellular matrix-binding proteins, DNases, fimbriae. The virulence factors are well studied in laboratory conditions, but the role of these factors in the pathogenesis of A. pyogenes infections remains to be elucidated. Lately, the ability of A. pyogenes to form biofilm in vivo has also been implicated as a virulence factor and a possible cause of therapeutic failure. Despite the fact that A. pyogenes milk isolates in cows with mastitis in vitro are very sensitive to β-lactam drugs and tetracycline, experience has shown that therapy is usually ineffective, prognosis is poor and the affected quarter is lost for milk production.

  13. Detection of virulence genes and the phylogenetic groups of Escherichia coli isolated from dogs in Brazil

    Directory of Open Access Journals (Sweden)

    Fernanda Morcatti Coura

    2018-02-01

    Full Text Available ABSTRACT: This study identified the virulence genes, pathovars, and phylogenetic groups of Escherichia coli strains obtained from the feces of dogs with and without diarrhea. Virulence genes and phylogenetic group identification were studied using polymerase chain reaction. Thirty-seven E. coli isolates were positive for at least one virulence factor gene. Twenty-one (57.8% of the positive isolates were isolated from diarrheal feces and sixteen (43.2% were from the feces of non-diarrheic dogs. Enteropathogenic E. coli (EPEC were the most frequently (62.2% detected pathovar in dog feces and were mainly from phylogroup B1 and E. Necrotoxigenic E. coli were detected in 16.2% of the virulence-positive isolates and these contained the cytotoxic necrotizing factor 1 (cnf1 gene and were classified into phylogroups B2 and D. All E. coli strains were negative for the presence of enterotoxigenic E. coli (ETEC enterotoxin genes, but four strains were positive for ETEC-related fimbriae 987P and F18. Two isolates were Shiga toxin-producing E. coli strains and contained the toxin genesStx2 or Stx2e, both from phylogroup B1. Our data showed that EPEC was the most frequent pathovar and B1 and E were the most common phylogroups detected in E. coli isolated from the feces of diarrheic and non-diarrheic dogs.

  14. The three-dimensional structure of CFA/I adhesion pili: traveler's diarrhea bacteria hang on by a spring.

    Science.gov (United States)

    Mu, Xiang-Qi; Savarino, Stephen J; Bullitt, Esther

    2008-02-22

    To survive the harsh environment of a churning intestinal tract, bacteria attach to the host epithelium via thin fibers called pili (or fimbriae). Enterotoxigenic Escherichia coli bacteria expressing colonization factor antigen I (CFA/I) pili and related pili are the most common known bacterial cause of diarrheal disease, including traveler's diarrhea. CFA/I pili, assembled via the alternate chaperone pathway, are essential for binding and colonization of the small bowel by these pathogenic bacteria. Herein, we elucidate unique structural features of CFA/I pili that appear to optimize their function as bacterial tethers in the intestinal tract. Using transmission electron microscopy of negatively stained samples in combination with iterative three-dimensional helical reconstruction methods for image processing, we determined the structure of the CFA/I pilus filament. Our results indicate that strong end-to-end protein interactions and weak interactions between the coils of a sturdy spring-like helix provide the combination of strength, stability, and flexibility required to sustain bacterial adhesion and incite intestinal disease. We propose that CFA/I pili behave like a spring to maintain attachment to the gut lining during vortex mixing and downward flow of the intestinal contents, thereby persisting long enough for these bacteria to colonize the host epithelium and cause enteric disease.

  15. Klebsiella pneumoniae yfiRNB operon affects biofilm formation, polysaccharide production and drug susceptibility.

    Science.gov (United States)

    Huertas, Mónica G; Zárate, Lina; Acosta, Iván C; Posada, Leonardo; Cruz, Diana P; Lozano, Marcela; Zambrano, María M

    2014-12-01

    Klebsiella pneumoniae is an opportunistic pathogen important in hospital-acquired infections, which are complicated by the rise of drug-resistant strains and the capacity of cells to adhere to surfaces and form biofilms. In this work, we carried out an analysis of the genes in the K. pneumoniae yfiRNB operon, previously implicated in biofilm formation. The results indicated that in addition to the previously reported effect on type 3 fimbriae expression, this operon also affected biofilm formation due to changes in cellulose as part of the extracellular matrix. Deletion of yfiR resulted in enhanced biofilm formation and an altered colony phenotype indicative of cellulose overproduction when grown on solid indicator media. Extraction of polysaccharides and treatment with cellulase were consistent with the presence of cellulose in biofilms. The enhanced cellulose production did not, however, correlate with virulence as assessed using a Caenorhabditis elegans assay. In addition, cells bearing mutations in genes of the yfiRNB operon varied with respect to the WT control in terms of susceptibility to the antibiotics amikacin, ciprofloxacin, imipenem and meropenem. These results indicated that the yfiRNB operon is implicated in the production of exopolysaccharides that alter cell surface characteristics and the capacity to form biofilms--a phenotype that does not necessarily correlate with properties related with survival, such as resistance to antibiotics. © 2014 The Authors.

  16. Cell Adhesion Modification of Streptococcus viridians in the Presence of Xylitol

    Science.gov (United States)

    Esmacher, Jason; Vidakovich, Blair; Giangrande, Michael; Hoffmann, Peter

    2012-10-01

    There is scientific documentation that those who chew gum sweetened by the sugar alcohol xylitol report a dramatically lower incident of both dental caries and otitis media compared to those who chew conventional gum sweetened by sucrose. An explanation contends that xylitol interferes with the ability of Streptococcus viridian (SV) to form biofilms which is a necessary precursor to the bacteria's ability to damage human tissues. We have used atomic force microscopy to study the cell wall/fimbria properties at the nanonewton level in both the presence and absence of xylitol. The first set of measurements used varying concentrations of xylitol incorporated within the incubation medium. The second used non-xylitol grown bacteria, the xylitol was added externally at various concentrations. Our study suggests that growing SV with xylitol reduces their ability to adhere together. Additionally, externally added xylitol showed grouping of cell adhesion to a relatively narrow nanonewton spread that is concentration dependent. Measurement of the adhesion properties of the bacterial cell wall have found that there is a dramatic increase in the cell wall's firmness which simultaneously accompanied a decrease in its ability to support adhesion, even at very low concentrations of xylitol.

  17. Differential Atrophy of Hippocampal Subfields: A Comparative Study of Dementia with Lewy Bodies and Alzheimer Disease.

    Science.gov (United States)

    Mak, Elijah; Su, Li; Williams, Guy B; Watson, Rosie; Firbank, Michael; Blamire, Andrew; O'Brien, John

    2016-02-01

    Dementia with Lewy bodies (DLB) is characterized by relative preservation of the medial temporal lobe compared with Alzheimer disease (AD). The differential involvement of the hippocampal subfields in both diseases has not been clearly established, however. We aim to investigate hippocampal subfield differences in vivo in a clinical cohort of DLB and AD subjects. 104 participants (35 DLBs, 36 ADs, and 35 healthy comparison [HC] subjects) underwent clinical assessment and 3T T1-weighted imaging. A Bayesian model implemented in Freesurfer was used to automatically segment the hippocampus and its subfields. We also examined associations between hippocampal subfields and tests of memory function. Both the AD and DLB groups demonstrated significant atrophy of the total hippocampus relative to HC but the DLB group was characterized by preservation of the cornu ammonis 1 (CA1), fimbria, and fissure. In contrast, all the hippocampal subfields except the fissure were significantly atrophied in AD compared with both DLB and HC groups. Among DLB subjects, CA1 was correlated with the Recent Memory score of the CAMCOG and Delayed Recall subscores of the HVLT. DLB is characterized by milder hippocampal atrophy that was accompanied by preservation of the CA1. The CA1 was also associated with memory function in DLB. Our findings highlight the promising role of hippocampal subfield volumetry, particularly that of the CA1, as a biomarker for the distinction between AD and DLB. Copyright © 2016 American Association for Geriatric Psychiatry. Published by Elsevier Inc. All rights reserved.

  18. Occurrence of Proteus mirabilis associated with two species of venezuelan oysters.

    Science.gov (United States)

    Fernández-Delgado, Milagro; Contreras, Monica; García-Amado, María Alexandra; Gueneau, Pulchérie; Suárez, Paula

    2007-01-01

    The fecal contamination of raw seafood by indicators and opportunistic pathogenic microorganisms represents a public health concern. The objective of this study was to investigate the presence of enteric bacteria colonizing oysters collected from a Venezuelan touristic area. Oyster samples were collected at the northwestern coast of Venezuela and local salinity, pH, temperature, and dissolved oxygen of seawater were recorded. Total and fecal coliforms were measured for the assessment of the microbiological quality of water and oysters, using the Multiple Tube Fermentation technique. Analyses were made using cultures and 16S rRNA gene sequencing. Diverse enrichment and selective culture methods were used to isolate enteric bacteria. We obtained pure cultures of Gram-negative straight rods with fimbriae from Isognomon alatus and Crassostrea rhizophorae. Our results show that P. mirabilis was predominant under our culture conditions. We confirmed the identity of the cultures by biochemical tests, 16S rRNA gene sequencing, and data analysis. Other enterobacteria such as Escherichia coli, Morganella morganii and Klebsiella pneumoniae were also isolated from seawater and oysters. The presence of pathogenic bacteria in oysters could have serious epidemiological implications and a potential human health risk associated with consumption of raw seafood.

  19. Effects of surgical and chemical lesions on neurotransmitter candidates in the nucleus accumbens of the rat

    Energy Technology Data Exchange (ETDEWEB)

    Walaas, I; Fonnum, F

    1979-01-01

    The origin of fibers containing different neurotransmitter candidates in the nucleus accumbens of rat brain has been studied with surgical and chemical lesion techniques. Destruction of the medial forebrain bundle decreased the activity of aromatic amino acid decarboxylase by 80% in the nucleus. Cutting of the fornix or a hemitransection decreased the high affinity uptake of glutamate by 45% and the endogenous level of glutamate by 33%. The high affinity uptake of glutamate was concentrated in the synaptosomal fraction and the decrease after the lesion was most pronounced in this fraction. Restricted lesions indicated that fibers in the fimbria/fornix coming from the subiculum were responsible for this part of the glutamate uptake in the nucleus. Local injection of kainic acid into the nucleus was accompanied by a 75% decrease in choline acetyltransferase and a 35% decrease in acetylcholineserase activities, a 70% decrease in glutamate decarboxylase activity and a 60% decrease in the high affinity uptake of ..gamma..-aminobutyrate, a 45% decrease in high affinity glutamate uptake, and no change in aromatic amino acid decarboxylase activity. Performing a lesion of the fornix after kainic acid injection led to an 85% decrease in high affinity glutamate uptake, without further affecting the other neuronal markers. The results indicate that all aminergic fibers to the nucleus accumbens are ascending in the medial forebrain bundle, that the subiculum-accumbens fibers are glutamergic and the nucleus also contains intrinsic glutamergic or aspartergic cells. Cholinergic and ..gamma..-aminobutyrate-containing cells are wholly intrinsic to the nucleus.

  20. Effect of elevated potassium ion concentrations on electrically evoked release of (/sup 3/H)acetylcholine in slices of rat hippocampus

    Energy Technology Data Exchange (ETDEWEB)

    Szerb, J C; Hadhazy, P; Dudar, J D [Dalhousie Univ., Halifax, Nova Scotia (Canada). Dept. of Physiology and Biophysics

    1978-01-01

    To establish the effect of raising the concentration of extracellular potassium ions on axonal conduction and transmitter release in a mammalian central pathway, the septohippocampal cholinergic tract, the rate of (/sup 3/H) acetylcholine release evoked by electrical stimulation was measured in rat hippocampal slices superfused with Krebs' solution containing 3-15 mM K/sup +/. The evoked release of (/sup 3/H) acetylcholine was abolished by the presence of tetrodotoxin or by the omission of Ca/sup 2 +/ in the superfusion medium, indicating that it originated from terminals depolarized by conducted action potentials. Potassium concentrations between 3 and 8 mM had no effect but 10-15 mM K/sup +/ reduced the rate of evoked release and decreased the size of the releasable pool of (/sup 3/H) acetylcholine. Decreasing the sodium content of the Krebs' solution to 97 mM or less had effects similar to those of elevated (K/sup +/). Elevated K/sup +/ (10-15 mM) reversibly reduced the size of compound action potentials in the fimbria and the alveus. The results suggest that extracellular potassium concentrations occurring under physiological conditions do not affect axonal conduction and transmitter release but that both are reduced in pathological states when extracellular (K/sup +/) above 8 mM occur.

  1. Vasopressin facilitates excitatory transmission in slices of the rat dorso-lateral septum.

    Science.gov (United States)

    Van den Hooff, P; Urban, I J

    1990-01-01

    The effect of vasopressin on neurons of the rat dorso-lateral septum (DLS) was studied in brain slices with intracellular microelectrodes. Two out of 13 neurons showed a small depolarization, spontaneous activity, and increased input resistances following a 15 min exposure to 10(-6) to 10(-8) M vasopressin (VP). These membrane effects disappeared completely within 3-5 min after the application. The remaining DLS neurons treated with these vasopressin concentrations showed an increase in glutamate-mediated excitatory postsynaptic potentials (EPSPs), evoked by stimulation of the fimbria fibers. As little as 10(-12) MVP increased these EPSPs markedly in nearly 80% of the cells studied. This increase in most of the cells disappeared within 15 min after the application period, whereas the increase in EPSPs induced by 10(-10) M VP outlasted the peptide application period for more than 30 min. Neither the blockade of GABA-ergic synaptic inhibition nor the pre-treatment of the neurons with d(CH2)5-Tyr(Me)-arginine vasopressin or 2-amino-5-phosphonovaleric acid (2-APV), antagonists for the V1 type of vasopressin receptor and NMDA receptors, respectively, interfered with the EPSPs potentiating effect of the peptide. It is concluded that a type of vasopressin receptor other then the V1 type is involved in the long-lasting potentiation of the primarily non-NMDA receptor mediated transmission in DLS neurons.

  2. Brain development in mice after prenatal irradiation: Modes of effect manifestation; dose-response-relationships and the RBE of neutrons

    International Nuclear Information System (INIS)

    Konermann, G.

    1986-01-01

    Postnatal effect manifestation in the CNS after exposures during advanced prenatal stages of development is due to both the prolonged period of neurogenesis and its complexity. Apart from acute proliferative effects, examples of two types of long-term effects in the brains of prenatally exposed mice are represented. Namely, persistent structural damage, and, fluctuating responses during the histochemical and biochemical brain maturation. Structural effects following X-ray exposure are quantified on the basis of data for diameter diminution of the cortical plate, corpus callosum and fimbria hippocampi. The studies include computerized micro-videoanalysis of neuronal branching defects. Continuous extension of exposure levels to doses as low as 0.05 Gy give evidence for the existence of thresholds for these types of structural damage in the vicinity of exposures to 0.1 Gy. The effects following X-ray exposures are partly compared with corresponding effects after neutron exposures. Our studies on postnatal maturation disturbances include proliferative responses, myelin formation, as well as the determination of different biochemical parameters (ATP, myelin-proteins, Na + /K + -balance). From our experimental findings we are able to stress the special significance of neurogenetical long-term effects for risk estimates in man. (orig.)

  3. Methylobacterium marchantiae sp. nov., a pink-pigmented, facultatively methylotrophic bacterium isolated from the thallus of a liverwort.

    Science.gov (United States)

    Schauer, S; Kämpfer, P; Wellner, S; Spröer, C; Kutschera, U

    2011-04-01

    A pink-pigmented, facultatively methylotrophic bacterium, designated strain JT1(T), was isolated from a thallus of the liverwort Marchantia polymorpha L. and was analysed by using a polyphasic approach. Comparative 16S rRNA gene sequence analysis placed the strain in a clade with Methylobacterium adhaesivum AR27(T), Methylobacterium fujisawaense DSM 5686(T), Methylobacterium radiotolerans JCM 2831(T) and Methylobacterium jeotgali S2R03-9(T), with which it showed sequence similarities of 97.8, 97.7, 97.2 and 97.4 %, respectively. However, levels of DNA-DNA relatedness between strain JT1(T) and these and the type strains of other closely related species were lower than 70 %. Cells of JT1(T) stained Gram-negative and were motile, rod-shaped and characterized by numerous fimbriae-like appendages on the outer surface of their wall (density up to 200 µm(-2)). Major fatty acids were C(18 : 1)ω7c and C(16 : 0). Based on the morphological, physiological and biochemical data presented, strain JT1(T) is considered to represent a novel species of the genus Methylobacterium, for which the name Methylobacterium marchantiae sp. nov. is proposed. The type strain is JT1(T) ( = DSM 21328(T)  = CCUG 56108(T)).

  4. Processing of fallopian tube, ovary, and endometrial surgical pathology specimens: A survey of U.S. laboratory practices.

    Science.gov (United States)

    Samimi, Goli; Trabert, Britton; Duggan, Máire A; Robinson, Jennifer L; Coa, Kisha I; Waibel, Elizabeth; Garcia, Edna; Minasian, Lori M; Sherman, Mark E

    2018-03-01

    Many high-grade serous carcinomas initiate in fallopian tubes as serous tubal intraepithelial carcinoma (STIC), a microscopic lesion identified with specimen processing according to the Sectioning and Extensive Examination of the Fimbria protocol (SEE-Fim). Given that the tubal origin of these cancers was recently recognized, we conducted a survey of pathology practices to assess processing protocols that are applied to gynecologic surgical pathology specimens in clinical contexts in which finding STIC might have different implications. We distributed a survey electronically to the American Society for Clinical Pathology list-serve to determine practice patterns and compared results between practice types by chi-square (χ2) tests for categorical variables. Free text comments were qualitatively reviewed. Survey responses were received from 159 laboratories (72 academic, 87 non-academic), which reported diverse specimen volumes and percentage of gynecologic samples. Overall, 74.1% of laboratories reported performing SEE-Fim for risk-reducing surgical specimens (82.5% academic versus 65.7% non-academic, p STIC or early cancer precursors. Published by Elsevier Inc.

  5. Serous tubal intraepithelial carcinoma, chronic fallopian tube injury, and serous carcinoma development.

    Science.gov (United States)

    Malmberg, Karin; Klynning, Charlotta; Flöter-Rådestad, Angelique; Carlson, Joseph W

    2016-06-01

    Ovarian carcinoma is the deadliest gynecological malignancy. Previous studies have suggested that the fallopian tube may be the primary site for high-grade serous carcinoma. In prophylactic salpingo-oophorectomies from women with hereditary high risk for ovarian cancer, precursors can be assessed prior to onset and studied as a model for serous cancer precursor lesions. Epidemiologic studies indicate that carcinogenesis may be a result of chronic fallopian tube injury. The aims of this study were to (1) to examine the incidence of serous tubal intraepithelial carcinoma (STIC) in relation to other clinical parameters and (2) to evaluate whether chronic fallopian tube injury was related to cancer development. This study enrolled 101 women, comprising the following three groups: hereditary (n = 60), sporadic serous cancer (n = 18; endometrial cancers were excluded), and control (n = 23). The cases were histologically examined and clinical risk factors were collected. The histological changes were compared between different patients and correlated to clinical risk factors. STICs were identified primarily on the fallopian tube fimbria. The incidence of STIC was 3 % in the hereditary patients. In sporadic serous cancer cases, 61 % were associated with STIC and tubal carcinoma (p STIC and invasive cancer were seen more often in the older patients than in the younger patients (p = 0.528). This small study, no correlation with chronic tubal injury or inflammation was identified.

  6. Progesterone Prevents High-Grade Serous Ovarian Cancer by Inducing Necroptosis of p53-Defective Fallopian Tube Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Na-Yiyuan Wu

    2017-03-01

    Full Text Available High-grade serous ovarian carcinoma (HGSOC originates mainly from the fallopian tube (FT epithelium and always carries early TP53 mutations. We previously reported that tumors initiate in the FT fimbria epithelium because of apoptotic failure and the expansion of cells with DNA double-strand breaks (DSB caused by bathing of the FT epithelial cells in reactive oxygen species (ROSs and hemoglobin-rich follicular fluid (FF after ovulation. Because ovulation is frequent and HGSOC is rare, we hypothesized that luteal-phase progesterone (P4 could eliminate p53-defective FT cells. Here we show that P4, via P4 receptors (PRs, induces necroptosis in Trp53−/− mouse oviduct epithelium and in immortalized human p53-defective fimbrial epithelium through the TNF-α/RIPK1/RIPK3/MLKL pathway. Necroptosis occurs specifically at diestrus, recovers at the proestrus phase of the estrus cycle, and can be augmented with P4 supplementation. These results reveal the mechanism of the well-known ability of progesterone to prevent ovarian cancer.

  7. Characterization of five ECF sigma factors in the genome of Pseudomonas syringae pv. syringae B728a.

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    Poulami Basu Thakur

    Full Text Available Pseudomonas syringae pv. syringae B728a, a bacterial pathogen of bean, utilizes large surface populations and extracellular signaling to initiate a fundamental change from an epiphytic to a pathogenic lifestyle. Extracytoplasmic function (ECF sigma (σ factors serve as important regulatory factors in responding to various environmental signals. Bioinformatic analysis of the B728a genome revealed 10 ECF sigma factors. This study analyzed deletion mutants of five previously uncharacterized ECF sigma factor genes in B728a, including three FecI-type ECF sigma factors (ECF5, ECF6, and ECF7 and two ECF sigma factors placed in groups ECF11 and ECF18. Transcriptional profiling by qRT-PCR analysis of ECF sigma factor mutants was used to measure expression of their associated anti-sigma and outer membrane receptor proteins, and expression of genes associated with production of extracellular polysaccharides, fimbriae, glycine betaine and syringomycin. Notably, the B728aΔecf7 mutant displayed reduced swarming and had decreased expression of CupC fimbrial genes. Growth and pathogenicity assays, using a susceptible bean host, revealed that none of the tested sigma factor genes are required for in planta growth and lesion formation.

  8. Towards a better understanding of Lactobacillus rhamnosus GG - host interactions

    Science.gov (United States)

    2014-01-01

    Lactobacillus rhamnosus GG (LGG) is one of the most widely used probiotic strains. Various health effects are well documented including the prevention and treatment of gastro-intestinal infections and diarrhea, and stimulation of immune responses that promote vaccination or even prevent certain allergic symptoms. However, not all intervention studies could show a clinical benefit and even for the same conditions, the results are not univocal. Clearly, the host phenotype governed by age, genetics and environmental factors such as the endogenous microbiota, plays a role in whether individuals are responders or non-responders. However, we believe that a detailed knowledge of the bacterial physiology and the LGG molecules that play a key role in its host-interaction capacity is crucial for a better understanding of its potential health benefits. Molecules that were yet identified as important factors governing host interactions include its adhesive pili or fimbriae, its lipoteichoic acid molecules, its major secreted proteins and its galactose-rich exopolysaccharides, as well as specific DNA motifs. Nevertheless, future studies are needed to correlate specific health effects to these molecular effectors in LGG, and also in other probiotic strains. PMID:25186587

  9. Proteus mirabilis and Urinary Tract Infections

    Science.gov (United States)

    Schaffer, Jessica N.; Pearson, Melanie M.

    2015-01-01

    Proteus mirabilis is a Gram-negative bacterium which is well-known for its ability to robustly swarm across surfaces in a striking bulls’-eye pattern. Clinically, this organism is most frequently a pathogen of the urinary tract, particularly in patients undergoing long-term catheterization. This review covers P. mirabilis with a focus on urinary tract infections (UTI), including disease models, vaccine development efforts, and clinical perspectives. Flagella-mediated motility, both swimming and swarming, is a central facet of this organism. The regulation of this complex process and its contribution to virulence is discussed, along with the type VI-secretion system-dependent intra-strain competition which occurs during swarming. P. mirabilis uses a diverse set of virulence factors to access and colonize the host urinary tract, including urease and stone formation, fimbriae and other adhesins, iron and zinc acquisition, proteases and toxins, biofilm formation, and regulation of pathogenesis. While significant advances in this field have been made, challenges remain to combatting complicated UTI and deciphering P. mirabilis pathogenesis. PMID:26542036

  10. Microarray Evaluation of Antimicrobial Resistance and Virulence of Escherichia coli Isolates from Portuguese Poultry

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    Nuno Mendonça

    2016-01-01

    Full Text Available The presence of antimicrobial resistance and virulence factors of 174 Escherichia coli strains isolated from healthy Portuguese Gallus gallus was evaluated. Resistance profiles were determined against 33 antimicrobials by microbroth dilution. Resistance was prevalent for tetracycline (70% and ampicillin (63%. Extended-spectrum beta-lactamase (ESBL phenotype was observed in 18% of the isolates. Multidrug resistance was found in 56% of isolates. A subset of 74 isolates were screened by DNA microarrays for the carriage of 88 antibiotic resistance genes and 62 virulence genes. Overall, 37 different resistance genes were detected. The most common were tet(A (72%, blaTEM (68%, and sul1 (47%, while 21% isolates harbored an ESBL gene (blaCTX-M group 1, group 2, or group 9. Of these, 96% carried the increased serum survival (iss virulence gene, while 89% presented the enterobactin siderophore receptor protein (iroN, 70% the temperature-sensitive hemagglutinin (tsh, and 68% the long polar fimbriae (lpfA virulence genes associated with extraintestinal pathogenic E. coli. In conclusion, prevalence of antibiotic resistant E. coli from the microbiota of Portuguese chickens was high, including to extended spectrum cephalosporins. The majority of isolates seems to have the potential to trigger extraintestinal human infection due to the presence of some virulence genes. However, the absence of genes specific for enteropathogenic E. coli reduces the risk for human intestinal infection.

  11. Impact of nanoscale topography on genomics and proteomics of adherent bacteria.

    Science.gov (United States)

    Rizzello, Loris; Sorce, Barbara; Sabella, Stefania; Vecchio, Giuseppe; Galeone, Antonio; Brunetti, Virgilio; Cingolani, Roberto; Pompa, Pier Paolo

    2011-03-22

    Bacterial adhesion onto inorganic/nanoengineered surfaces is a key issue in biotechnology and medicine, because it is one of the first necessary steps to determine a general pathogenic event. Understanding the molecular mechanisms of bacteria-surface interaction represents a milestone for planning a new generation of devices with unanimously certified antibacterial characteristics. Here, we show how highly controlled nanostructured substrates impact the bacterial behavior in terms of morphological, genomic, and proteomic response. We observed by atomic force microscopy (AFM) and scanning electron microscopy (SEM) that type-1 fimbriae typically disappear in Escherichia coli adherent onto nanostructured substrates, as opposed to bacteria onto reference glass or flat gold surfaces. A genetic variation of the fimbrial operon regulation was consistently identified by real time qPCR in bacteria interacting with the nanorough substrates. To gain a deeper insight into the molecular basis of the interaction mechanisms, we explored the entire proteomic profile of E. coli by 2D-DIGE, finding significant changes in the bacteria adherent onto the nanorough substrates, such as regulations of proteins involved in stress processes and defense mechanisms. We thus demonstrated that a pure physical stimulus, that is, a nanoscale variation of surface topography, may play per se a significant role in determining the morphological, genetic, and proteomic profile of bacteria. These data suggest that in depth investigations of the molecular processes of microorganisms adhering to surfaces are of great importance for the design of innovative biomaterials with active biological functionalities.

  12. Protective effect of oral administration of transgenic tobacco seeds against verocytotoxic Escherichia coli strain in piglets.

    Science.gov (United States)

    Rossi, Luciana; Dell'Orto, Vittorio; Vagni, Simona; Sala, Vittorio; Reggi, Serena; Baldi, Antonella

    2014-03-01

    The use of transgenic plants as delivery system for antigenic proteins is attractive for its simplicity and increases likelihood for local immune response at sites of infection. The aim of this study was to evaluate the protective effect of oral administration of tobacco seeds, expressing the FedA, the major protein of the F18 adhesive fimbriae, and B subunit of verocytotoxin, against verocytotoxin-producing E. coli (VTEC) strain in piglets. Forty-three early weaned piglets, were randomly divided into 4 experimental groups: 3 test groups and a control. Treatment groups orally received a bolus, with different dose of tobacco seeds on 0, 1, 2, 14 days post primary administration. After challenge, with 1*10(10) CFU of O138 Escherichia coli strain, piglets showed clinical scores significantly higher in the control group compared to orally immunized groups (P administration of recombinant tobacco seeds expressing antigenic proteins against VTEC strains can induce a protective effect against challenger strain in piglets.

  13. N-Acetylglucosamine Functions in Cell Signaling

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    James B. Konopka

    2012-01-01

    Full Text Available The amino sugar N-acetylglucosamine (GlcNAc is well known for the important structural roles that it plays at the cell surface. It is a key component of bacterial cell wall peptidoglycan, fungal cell wall chitin, and the extracellular matrix of animal cells. Interestingly, recent studies have also identified new roles for GlcNAc in cell signaling. For example, GlcNAc stimulates the human fungal pathogen Candida albicans to undergo changes in morphogenesis and expression of virulence genes. Pathogenic E. coli responds to GlcNAc by altering the expression of fimbriae and CURLI fibers that promote biofilm formation and GlcNAc stimulates soil bacteria to undergo changes in morphogenesis and production of antibiotics. Studies with animal cells have revealed that GlcNAc influences cell signaling through the posttranslational modification of proteins by glycosylation. O-linked attachment of GlcNAc to Ser and Thr residues regulates a variety of intracellular proteins, including transcription factors such as NFκB, c-myc, and p53. In addition, the specificity of Notch family receptors for different ligands is altered by GlcNAc attachment to fucose residues in the extracellular domain. GlcNAc also impacts signal transduction by altering the degree of branching of N-linked glycans, which influences cell surface signaling proteins. These emerging roles of GlcNAc as an activator and mediator of cellular signaling in fungi, animals, and bacteria will be the focus of this paper.

  14. Comparison of Inhibitory Activities of meta and para Substituted N-aryl 3-Hydroxypyridin-4-one Mannosides Towards Type 1 Fimbriated E. coli

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    Vesna Petrović Peroković

    2016-06-01

    Full Text Available In uropathogenic Escherichia coli, mannose-specific adhesion is mediated by the FimH adhesin located at the tip of type 1 fimbriae. Novel mannosylated N-aryl substituted 3-hydroxypyridin-4ones with meta substituents on the aryl part of the molecule were prepared, and their inhibitory properties towards the adhesion of E. coli to guinea pig erythrocytes explored using the hemagglutination assay. These results were compared with inhibitory potencies of analogous para derivatives. The assays revealed greater preference of FimH towards para substituted compounds in general, with p-nitro and p-methoxy substituted substrates being much more effective then the hydrophobic p-methyl compound. When substituents are in meta position the positive affect on the binding of compounds in the FimH binding site was observed with all compounds tested but the structure with an alkyl group was shown to be the most effective one. This study provides guidelines for the rational design of novel, more effective series of FimH antagonists. This work is licensed under a Creative Commons Attribution 4.0 International License.

  15. White Matter Tracts Connected to the Medial Temporal Lobe Support the Development of Mnemonic Control.

    Science.gov (United States)

    Wendelken, Carter; Lee, Joshua K; Pospisil, Jacqueline; Sastre, Marcos; Ross, Julia M; Bunge, Silvia A; Ghetti, Simona

    2015-09-01

    One of the most important factors driving the development of memory during childhood is mnemonic control, or the capacity to initiate and maintain the processes that guide encoding and retrieval operations. The ability to selectively attend to and encode relevant stimuli is a particularly useful form of mnemonic control, and is one that undergoes marked improvement over childhood. We hypothesized that structural integrity of white matter tracts, in particular those connecting medial temporal lobe memory regions to other cortical areas, and/or those connecting frontal and parietal control regions, should contribute to successful mnemonic control. To test this hypothesis, we examined the relationship between structural integrity of selected white matter tracts and an experimental measure of mnemonic control, involving enhancement of memory by attention at encoding, in 116 children aged 7-11 and 25 young adults. We observed a positive relationship between integrity of uncinate fasciculus and mnemonic enhancement across age groups. In adults, but not in children, we also observed an association between mnemonic enhancement and integrity of ventral cingulum bundle and ventral fornix/fimbria. Integrity of fronto-parietal tracts, including dorsal cingulum and superior longitudinal fasciculus, was unrelated to mnemonic enhancement. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  16. [Microbiological characterization of non-O1 Vibrio cholerae isolated in Cuba].

    Science.gov (United States)

    Bravo Fariñas, Laura; Fernández, Anabel; Ramírez, María M; Llop, Alina; Martínez, Gerardo; Hernández, Raquel I; Cabrera, Luis E; Morier, Luis; Fraga, Jorge; Núñez, Fidel A; Aguila, Adalberto

    2007-01-01

    The study of 422 non-01 Vibrio cholerae strains from nine provinces, 9 of them isolated from a water-borne disease outbreak, was performed. All the strains exhibited antimicrobial susceptibility and virulence factors. The nine strains from the outbreak were subjected to a DNA macrorestriction study based on the pulsed field electrophoresis technique. For the first time in Cuba and the Caribbean. The circulation of atypical non-01 V cholerae strains (resistent to vibriostatic compound 0129 and trimethoprim/sulfamethoxazole). The behavior of antimicrobial susceptibility evinced for the first time the circulation of two different resistence patterns in Cuba (ampicilline, trimethoprim/ sulfamethoxazole, sulfonamide and tetracycline, trimethoprim/ sulfamethoxazole, sulfonamide). The frequency of trimethoprim/ sulfamethoxazole-resistent strains was similar during the whole period of study. However, resistance to ampicilline decreased whereas resistance to tetracycline increased. The main found virulence factors were gelatinase, hemolysine, elastase and adherence to Hep-2 cells. On the other hand, the outbreak strains showed higher percentages than the others due to the presence of heat-liable toxin and fimbriae. The results of the molecular and epidemiological studies allowed giving a speedy and accurate response that explained the etiology of the first food-borne disease outbreak.

  17. Short communication: Distribution of Porphyromonas gulae fimA genotypes in oral specimens from dogs with mitral regurgitation.

    Science.gov (United States)

    Shirai, Mitsuyuki; Nomura, Ryota; Kato, Yukio; Murakami, Masaru; Kondo, Chihiro; Takahashi, Soraaki; Yamasaki, Yoshie; Matsumoto-Nakano, Michiyo; Arai, Nobuaki; Yasuda, Hidemi; Nakano, Kazuhiko; Asai, Fumitoshi

    2015-10-01

    Porphyromonas gulae, a suspected pathogen for periodontal disease in dogs, possesses approximately 41-kDa fimbriae (FimA) that are encoded by the fimA gene. In the present study, the association of fimA genotypes with mitral regurgitation (MR) was investigated. Twenty-five dogs diagnosed with MR (age range 6-13 years old, average 10.8 years) and 32 healthy dogs (8-15 years old, average 10.8 years) were selected at the participating clinics in a consecutive manner during the same time period. Oral swab specimens were collected from the dogs and bacterial DNA was extracted, then polymerase chain reaction analysis was performed using primers specific for each fimA genotype, with the dominant genotype determined. The rate for genotype C dominant specimens was 48.0% in the MR group, which was significantly higher than that in the control group (18.8%) (P <0.05). These results suggest that P. gulae fimA genotype C is associated with MR. Copyright © 2015 Elsevier Ltd. All rights reserved.

  18. Virulence factors of Escherichia coli in relation to the importance of vaccination in pigs

    Directory of Open Access Journals (Sweden)

    Daniele Araujo Pereira

    2016-08-01

    Full Text Available ABSTRACT: Enterotoxigenic Escherichia coli (ETEC is the major cause of diarrhea in newborn and weaned pigs. Bacteria adhesion to the host cell is considered a specific phenomenon among fimbrial and non-fimbrial adhesins with their respective receptors on enterocytes. Enteric disorders are related with the fimbriae F4 (K88, F5 (K99, F6 (987P, F41, and F18. In addition to ETEC, another category of E. coli , porcine pathogenic E. coli (PEPEC,can cause diarrhea in pigs; it produces the porcine attaching and effacing-associated (Paa adhesin in, which is capable to cause a typical lesion known as an attaching and effacing (A/E lesion. Immunization of sows with adhesin is important to stimulate the production of antibodies and their subsequent transfer to piglets through colostrum. The aim of this paper is to illustrate the main impacts of enteric diseases caused by E. coli in swine production and to highlight the importance of continuing research on this bacterium to improve disease prevention through vaccination.

  19. The Rcs regulon in Proteus mirabilis: implications for motility, biofilm formation, and virulence.

    Science.gov (United States)

    Howery, Kristen E; Clemmer, Katy M; Rather, Philip N

    2016-11-01

    The overall role of the Rcs phosphorelay in Proteus mirabilis is largely unknown. Previous work had demonstrated that the Rcs phosphorelay represses the flhDC operon and activates the minCDE cell division inhibition system. To identify additional cellular functions regulated by the Rcs phosphorelay, an analysis of RNA-seq data was undertaken. In this report, the results of the RNA-sequencing are discussed with an emphasis on the predicted roles of the Rcs phosphorelay in swarmer cell differentiation, motility, biofilm formation, and virulence. RcsB is shown to activate genes important for differentiation and fimbriae formation, while repressing the expression of genes important for motility and virulence. Additionally, to follow up on the RNA-Seq data, we demonstrate that an rcsB mutant is deficient in its ability to form biofilm and exhibits enhanced virulence in a Galleria mellonella waxworm model. Overall, these results indicate the Rcs regulon in P. mirabilis extends beyond flagellar genes to include those involved in biofilm formation and virulence. Furthermore, the information presented in this study may provide clues to additional roles of the Rcs phosphorelay in other members of the Enterobacteriaceae.

  20. Transcriptional Analysis of the MrpJ Network: Modulation of Diverse Virulence-Associated Genes and Direct Regulation of mrp Fimbrial and flhDC Flagellar Operons in Proteus mirabilis

    Science.gov (United States)

    Bode, Nadine J.; Debnath, Irina; Kuan, Lisa; Schulfer, Anjelique; Ty, Maureen

    2015-01-01

    The enteric bacterium Proteus mirabilis is associated with a significant number of catheter-associated urinary tract infections (UTIs). Strict regulation of the antagonistic processes of adhesion and motility, mediated by fimbriae and flagella, respectively, is essential for disease progression. Previously, the transcriptional regulator MrpJ, which is encoded by the mrp fimbrial operon, has been shown to repress both swimming and swarming motility. Here we show that MrpJ affects an array of cellular processes beyond adherence and motility. Microarray analysis found that expression of mrpJ mimicking levels observed during UTIs leads to differential expression of 217 genes related to, among other functions, bacterial virulence, type VI secretion, and metabolism. We probed the molecular mechanism of transcriptional regulation by MrpJ using transcriptional reporters and chromatin immunoprecipitation (ChIP). Binding of MrpJ to two virulence-associated target gene promoters, the promoters of the flagellar master regulator flhDC and mrp itself, appears to be affected by the condensation state of the native chromosome, although both targets share a direct MrpJ binding site proximal to the transcriptional start. Furthermore, an mrpJ deletion mutant colonized the bladders of mice at significantly lower levels in a transurethral model of infection. Additionally, we observed that mrpJ is widely conserved in a collection of recent clinical isolates. Altogether, these findings support a role of MrpJ as a global regulator of P. mirabilis virulence. PMID:25847961

  1. Proteus mirabilis and Urinary Tract Infections.

    Science.gov (United States)

    Schaffer, Jessica N; Pearson, Melanie M

    2015-10-01

    Proteus mirabilis is a Gram-negative bacterium and is well known for its ability to robustly swarm across surfaces in a striking bulls'-eye pattern. Clinically, this organism is most frequently a pathogen of the urinary tract, particularly in patients undergoing long-term catheterization. This review covers P. mirabilis with a focus on urinary tract infections (UTI), including disease models, vaccine development efforts, and clinical perspectives. Flagella-mediated motility, both swimming and swarming, is a central facet of this organism. The regulation of this complex process and its contribution to virulence is discussed, along with the type VI-secretion system-dependent intra-strain competition, which occurs during swarming. P. mirabilis uses a diverse set of virulence factors to access and colonize the host urinary tract, including urease and stone formation, fimbriae and other adhesins, iron and zinc acquisition, proteases and toxins, biofilm formation, and regulation of pathogenesis. While significant advances in this field have been made, challenges remain to combatting complicated UTI and deciphering P. mirabilis pathogenesis.

  2. Efficacy of thiolated eudragit microspheres as an oral vaccine delivery system to induce mucosal immunity against enterotoxigenic Escherichia coli in mice.

    Science.gov (United States)

    Lee, Won-Jung; Cha, Seungbin; Shin, Minkyoung; Jung, Myunghwan; Islam, Mohammad Ariful; Cho, Chong-su; Yoo, Han Sang

    2012-05-01

    A vaccine delivery system based on thiolated eudragit microsphere (TEMS) was studied in vivo for its ability to elicit mucosal immunity against enterotoxigenic Escherichia coli (ETEC). Groups of mice were orally immunized with F4 or F18 fimbriae of ETEC and F4 or F18 loaded in TEMS. Mice that were orally administered with F4 or F18 loaded TEMS showed higher antigen-specific IgG antibody responses in serum and antigen-specific IgA in saliva and feces than mice that were immunized with antigens only. In addition, oral vaccination of F4 or F18 loaded TEMS resulted in higher numbers of IgG and IgA antigen-specific antibody secreting cells in the spleen, lamina propria, and Peyer's patches of immunized mice than other groups. Moreover, TEMS administration loaded with F4 or F18 induced mixed Th1 and Th2 type responses based on similarly increased levels of IgG1 and IgG2a. These results suggest that F4 or F18 loaded TEMS may be a promising candidate for an oral vaccine delivery system to elicit systemic and mucosal immunity against ETEC. Copyright © 2012 Elsevier B.V. All rights reserved.

  3. N-terminal truncation enables crystallization of the receptor-binding domain of the FedF bacterial adhesin

    Energy Technology Data Exchange (ETDEWEB)

    De Kerpel, Maia; Van Molle, Inge [Department of Ultrastructure, Vrije Universiteit Brussel (VUB), Flanders Interuniversity Institute for Biotechnology (VIB), Pleinlaan 2, 1050 Brussels (Belgium); Brys, Lea [Department of Cellular and Molecular Immunology, Vrije Universiteit Brussel (VUB), Flanders Interuniversity Institute for Biotechnology (VIB), Pleinlaan 2, 1050 Brussels (Belgium); Wyns, Lode; De Greve, Henri; Bouckaert, Julie, E-mail: bouckaej@vub.ac.be [Department of Ultrastructure, Vrije Universiteit Brussel (VUB), Flanders Interuniversity Institute for Biotechnology (VIB), Pleinlaan 2, 1050 Brussels (Belgium)

    2006-12-01

    The N-terminal receptor-binding domain of the FedF adhesin from enterotoxigenic E. coli has been crystallized. This required the deletion of its first 14 residues, which are also cleaved off naturally. FedF is the two-domain tip adhesin of F18 fimbriae from enterotoxigenic Escherichia coli. Bacterial adherence, mediated by the N-terminal receptor-binding domain of FedF to carbohydrate receptors on intestinal microvilli, causes diarrhoea and oedema disease in newly weaned piglets and induces the secretion of Shiga toxins. A truncate containing only the receptor-binding domain of FedF was found to be further cleaved at its N-terminus. Reconstruction of this N-terminal truncate rendered FedF amenable to crystallization, resulting in crystals with space group P2{sub 1}2{sub 1}2{sub 1} and unit-cell parameters a = 36.20, b = 74.64, c = 99.03 Å that diffracted to beyond 2 Å resolution. The binding specificity of FedF was screened for on a glycan array, exposing 264 glycoconjugates, to identify specific receptors for cocrystallization with FedF.

  4. General N-and O-Linked Glycosylation of Lipoproteins in Mycoplasmas and Role of Exogenous Oligosaccharide.

    Science.gov (United States)

    Daubenspeck, James M; Jordan, David S; Simmons, Warren; Renfrow, Matthew B; Dybvig, Kevin

    2015-01-01

    The lack of a cell wall, flagella, fimbria, and other extracellular appendages and the possession of only a single membrane render the mycoplasmas structurally simplistic and ideal model organisms for the study of glycoconjugates. Most species have genomes of about 800 kb and code for few proteins predicted to have a role in glycobiology. The murine pathogens Mycoplasma arthritidis and Mycoplasma pulmonis have only a single gene annotated as coding for a glycosyltransferase but synthesize glycolipid, polysaccharide and glycoproteins. Previously, it was shown that M. arthritidis glycosylated surface lipoproteins through O-linkage. In the current study, O-linked glycoproteins were similarly found in M. pulmonis and both species of mycoplasma were found to also possess N-linked glycans at residues of asparagine and glutamine. Protein glycosylation occurred at numerous sites on surface-exposed lipoproteins with no apparent amino acid sequence specificity. The lipoproteins of Mycoplasma pneumoniae also are glycosylated. Glycosylation was dependent on the glycosidic linkages from host oligosaccharides. As far as we are aware, N-linked glycoproteins have not been previously described in Gram-positive bacteria, the organisms to which the mycoplasmas are phylogenetically related. The findings indicate that the mycoplasma cell surface is heavily glycosylated with implications for the modulation of mycoplasma-host interactions.

  5. The importance of Bordetella pertussis strains which do not produce virulence factors in the epidemiology of pertussis

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    Maciej Polak

    2017-05-01

    Full Text Available Bordetella pertussis strains, which have lost the ability to produce antigens, such as pertactin - Prn, pertussis toxin - Ptx, filamentous haemagglutinin - FHA, fimbriae type 2 and 3 - Fim 2 and 3, tracheal colonization factor - TcfA, have recently been isolated in countries with a high vaccination coverage. The emergence of such isolates might have resulted from B. pertussis natural evolution course or adaptive mechanisms, allowing increased circulation of the pathogen in vaccinated populations. So far, the majority of described mutants were deficient in the Prn production. Prn deficient isolates were found in countries which use acellular pertussis vaccines (aP in routine immunization programmes. The increase of frequency of Prn¯ strains isolation was correlated with the period of routine vaccination with aP vaccines. In most countries, the spread of these isolates has resulted from independent mutations rather than from the expansion of a single clone. Prn¯ isolates were collected from patients showing typical clinical symptoms of pertussis found for Prn+ strains. Results of in vitro and in vivo studies have shown that Prn¯, Ptx¯ and FHA¯ isolates retain cytotoxic properties, and besides Ptx¯ isolates, were lethal in intranasally infected mice. Further explanation of the impact of antigen deficiencies on virulence and transmission of B. pertussis in the context of the continuous increase of pertussis incidence is necessary to develop a new, optimized strategy of pertussis prevention.

  6. Urinary tract infections of Escherichia coli strains of chaperone-usher system.

    Science.gov (United States)

    Zalewska-Piatek, Beata M

    2011-01-01

    Urinary tract infections are a very serious health and economic problem affecting millions of people each year worldwide. The most common etiologic agent of this type of bacterial infections, involving the upper and lower urinary tract, are E. coli strains representing approximately 80% of cases. Uropathogenic E. coli strains produce several urovirulence factors which can be divided into two main types, surface virulence factors and exported virulence factors. Surface-exposed structures include mainly extracellular adhesive organelles such as fimbriae/pili necessary in adhesion, invasion, biofilm formation and cytokine induction. Among the surface-exposed polymeric adhesive structures there are three most invasive groups, type 1 pili, type P pili and Dr family of adhesins which are bioassembled via the conserved, among Gram-negative bacteria, chaperone-usher secretion system. Type 1 and P-piliated E. coli cause cystitis and pyelonephritis. The Dr family of adhesins recognizing DAF receptor is responsible for cystitis, pyelonephritis (especially in pregnant women) and diarrhoea (in infants). In addition, Dr-positive E. coli strains carry the risk of recurrent urinary tract infections. Pyelonephritis in pregnant women leads to a series of complications such as bacteremia, urosepsis, acute respiratory distress syndrome and even death. In the era of increasing drug resistance of bacteria, the development of vaccines, drugs termed pilicides and inhibitors of adhesion may be a promising tool in the fight against urogenital infections.

  7. Virulence factors, antimicrobial resistance, and plasmid content of Escherichia coli isolated in swine commercial farms Fatores de virulência, resistência aos antimicrobianos, presença de plasmídeos em Escherichia coli isoladas de amostras clínicas e ambientais de suínos

    Directory of Open Access Journals (Sweden)

    M.M. Costa

    2010-02-01

    Full Text Available Virulence factors and antimicrobial resistance patterns of Escherichia coli isolates were evaluated. A total of 80 E. coli isolates were evaluated, being 64 from clinical samples (intestinal content and fragments of organs from diarrheic piglets, seven from feces of clinically healthy piglets and sows, and nine environmental samples (five from facilities, two from feed, one from insect, and one from waste. Molecular characterization was performed by PCR detection of fimbriae and toxin genes and plasmid content determination. The isolates were also characterized according to their resistance or sensitivity to the following drugs: ampicillin, trimethoprim:sulfamethoxazole, tetracycline, amikacine, colistin, norfloxacin, florfenicol, enrofloxacin, cefalexin, trimethoprim, neomycin, chloramphenicol, and gentamicin. From 80 E. coli isolates, 53.8% were classified as enterotoxigenic E. coli (ETEC, 2.5% were shiga toxin-producing E. coli (STEC, and 43.8% showed a non specific pattern and were unclassified. One fecal isolate from non-diarrheic piglet was classified as ETEC by PCR. Clinical isolates showed resistance mainly for tetracycline and trimethoprim:sulfamethoxazole. Plasmidial DNA was observed in 70 isolates, being 78.5% of clinical isolates, 8.57% of non-diarrheic feces, and 12.8% of environment.Os fatores de virulência e a resistência aos antimicrobianos foram avaliados em Escherichia coli. Um total de 80 isolados de E. coli, sendo 64 de amostras clínicas (conteúdo intestinal e fragmentos de órgãos de leitões diarreicos, sete das fezes de porcas e leitões saudáveis e nove de amostras ambientais (cinco de instalações, dois de alimentos, um de inseto e um de esterqueira. A caracterização molecular feita pela PCR objetivou detectar fimbrias e toxinas, bem como a determinação do conteúdo de plasmídeos. Os isolados foram caracterizados quanto à resistência ou sensibilidade às seguintes drogas: ampicilina, sulfazotrim

  8. Oxygen limitation modulates pH regulation of catabolism and hydrogenases, multidrug transporters, and envelope composition in Escherichia coli K-12

    Directory of Open Access Journals (Sweden)

    Radmacher Michael D

    2006-10-01

    Full Text Available Abstract Background In Escherichia coli, pH regulates genes for amino-acid and sugar catabolism, electron transport, oxidative stress, periplasmic and envelope proteins. Many pH-dependent genes are co-regulated by anaerobiosis, but the overall intersection of pH stress and oxygen limitation has not been investigated. Results The pH dependence of gene expression was analyzed in oxygen-limited cultures of E. coli K-12 strain W3110. E. coli K-12 strain W3110 was cultured in closed tubes containing LBK broth buffered at pH 5.7, pH 7.0, and pH 8.5. Affymetrix array hybridization revealed pH-dependent expression of 1,384 genes and 610 intergenic regions. A core group of 251 genes showed pH responses similar to those in a previous study of cultures grown with aeration. The highly acid-induced gene yagU was shown to be required for extreme-acid resistance (survival at pH 2. Acid also up-regulated fimbriae (fimAC, periplasmic chaperones (hdeAB, cyclopropane fatty acid synthase (cfa, and the "constitutive" Na+/H+ antiporter (nhaB. Base up-regulated core genes for maltodextrin transport (lamB, mal, ATP synthase (atp, and DNA repair (recA, mutL. Other genes showed opposite pH responses with or without aeration, for example ETS components (cyo,nuo, sdh and hydrogenases (hya, hyb, hyc, hyf, hyp. A hypF strain lacking all hydrogenase activity showed loss of extreme-acid resistance. Under oxygen limitation only, acid down-regulated ribosome synthesis (rpl,rpm, rps. Acid up-regulated the catabolism of sugar derivatives whose fermentation minimized acid production (gnd, gnt, srl, and also a cluster of 13 genes in the gadA region. Acid up-regulated drug transporters (mdtEF, mdtL, but down-regulated penicillin-binding proteins (dacACD, mreBC. Intergenic regions containing regulatory sRNAs were up-regulated by acid (ryeA, csrB, gadY, rybC. Conclusion pH regulates a core set of genes independently of oxygen, including yagU, fimbriae, periplasmic chaperones, and nha

  9. Serous tubal intraepithelial carcinoma localizes to the tubal-peritoneal junction: a pivotal clue to the site of origin of extrauterine high-grade serous carcinoma (ovarian cancer).

    Science.gov (United States)

    Seidman, Jeffrey D

    2015-03-01

    Recent data suggest that intraepithelial carcinoma of the fallopian tube [serous tubal intraepithelial carcinoma (STIC)] is the precursor of high-grade extrauterine serous carcinoma. A more specific location for the origin of this lesion is suggested by the recently described junction between the fallopian tubal epithelium and the peritoneum [tubal-peritoneal junction (TPJ)]. Fallopian tubes from 202 patients with advanced-stage high-grade extrauterine serous carcinoma or carcinosarcoma were evaluated histologically as were 124 prophylactic salpingo-oophorectomy specimens. These included 54 patients with BRCA or other high-risk mutation or a family history of BRCA mutation and 70 with a personal or family history of breast carcinoma. STIC was found in 81 of 202 patients with serous carcinoma (40.1%). STIC was present in 73 of 141 (52%) cases in which the fimbriae were present and in 62 of 100 (62%) cases in which the TPJ was present (P not significant). In comparison with these groups, when fimbriae and TPJ were absent, STIC was found in 8 of 61 (13%) cases (PSTIC. The mean size of STIC was 1.7 mm. In 32 cases (39.5%), the lesion was flat and in 49 (60.5%), papillary. The mean size of flat STICs was 0.8 mm as compared with 2.3 mm for papillary STICs (P=0.00005). STIC was identified in the same tissue fragment as the junction in 48 cases. The mean distance of STIC to the junction was 1.8 mm. In 11 cases, STIC was flanked by peritoneal mesothelium on one side and tubal epithelium on the opposite side. In 51 patients, the mean distance of invasive carcinoma from the TPJ was 1.8 mm. This distance was 1.9 mm when STIC was present (37 cases) in comparison with 1.5 mm when STIC was absent (14 cases) (P not significant). In 27 of 42 cases (64%), STIC was contiguous with invasive carcinoma. Lamina propria invasion was present in 71% of cases in which STIC was present as compared with 26% of cases in which STIC was absent (PSTIC was present as compared with 26% of cases in

  10. A short-time scale colloidal system reveals early bacterial adhesion dynamics.

    Directory of Open Access Journals (Sweden)

    Christophe Beloin

    2008-07-01

    Full Text Available The development of bacteria on abiotic surfaces has important public health and sanitary consequences. However, despite several decades of study of bacterial adhesion to inert surfaces, the biophysical mechanisms governing this process remain poorly understood, due, in particular, to the lack of methodologies covering the appropriate time scale. Using micrometric colloidal surface particles and flow cytometry analysis, we developed a rapid multiparametric approach to studying early events in adhesion of the bacterium Escherichia coli. This approach simultaneously describes the kinetics and amplitude of early steps in adhesion, changes in physicochemical surface properties within the first few seconds of adhesion, and the self-association state of attached and free-floating cells. Examination of the role of three well-characterized E. coli surface adhesion factors upon attachment to colloidal surfaces--curli fimbriae, F-conjugative pilus, and Ag43 adhesin--showed clear-cut differences in the very initial phases of surface colonization for cell-bearing surface structures, all known to promote biofilm development. Our multiparametric analysis revealed a correlation in the adhesion phase with cell-to-cell aggregation properties and demonstrated that this phenomenon amplified surface colonization once initial cell-surface attachment was achieved. Monitoring of real-time physico-chemical particle surface properties showed that surface-active molecules of bacterial origin quickly modified surface properties, providing new insight into the intricate relations connecting abiotic surface physicochemical properties and bacterial adhesion. Hence, the biophysical analytical method described here provides a new and relevant approach to quantitatively and kinetically investigating bacterial adhesion and biofilm development.

  11. Transcriptional analysis of the MrpJ network: modulation of diverse virulence-associated genes and direct regulation of mrp fimbrial and flhDC flagellar operons in Proteus mirabilis.

    Science.gov (United States)

    Bode, Nadine J; Debnath, Irina; Kuan, Lisa; Schulfer, Anjelique; Ty, Maureen; Pearson, Melanie M

    2015-06-01

    The enteric bacterium Proteus mirabilis is associated with a significant number of catheter-associated urinary tract infections (UTIs). Strict regulation of the antagonistic processes of adhesion and motility, mediated by fimbriae and flagella, respectively, is essential for disease progression. Previously, the transcriptional regulator MrpJ, which is encoded by the mrp fimbrial operon, has been shown to repress both swimming and swarming motility. Here we show that MrpJ affects an array of cellular processes beyond adherence and motility. Microarray analysis found that expression of mrpJ mimicking levels observed during UTIs leads to differential expression of 217 genes related to, among other functions, bacterial virulence, type VI secretion, and metabolism. We probed the molecular mechanism of transcriptional regulation by MrpJ using transcriptional reporters and chromatin immunoprecipitation (ChIP). Binding of MrpJ to two virulence-associated target gene promoters, the promoters of the flagellar master regulator flhDC and mrp itself, appears to be affected by the condensation state of the native chromosome, although both targets share a direct MrpJ binding site proximal to the transcriptional start. Furthermore, an mrpJ deletion mutant colonized the bladders of mice at significantly lower levels in a transurethral model of infection. Additionally, we observed that mrpJ is widely conserved in a collection of recent clinical isolates. Altogether, these findings support a role of MrpJ as a global regulator of P. mirabilis virulence. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  12. A trial with IgY chicken antibodies to eradicate faecal carriage of Klebsiella pneumoniae and Escherichia coli producing extended-spectrum beta-lactamases

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    Anna-Karin Jonsson

    2015-11-01

    Full Text Available Background: Extended-spectrum beta-lactamase (ESBL-producing Enterobacteriaceae is an emerging therapeutic challenge, especially in the treatment of urinary tract infections. Following an outbreak of CTX-M-15 Klebsiella pneumoniae in Uppsala, Sweden, an orphan drug trial on IgY chicken antibodies was undertaken in an attempt to eradicate faecal carriage of ESBL-producing K. pneumoniae and Escherichia coli. Methods: Hens were immunised with epitopes from freeze-dried, whole-cell bacteria (ESBL-producing K. pneumoniae and E. coli and recombinant proteins of two K. pneumoniae fimbriae subunits (fimH and mrkD. The egg yolks were processed according to good manufacturing practice and the product was stored at−20°C until used. Using an internal database from the outbreak and the regular laboratory database, faecal carriers were identified and recruited from May 2005 to December 2013. The participants were randomised in a placebo-controlled 1:1 manner. Results: From 749 eligible patients, 327 (44% had deceased, and only 91 (12% were recruited and signed the informed consent. In the initial screening performed using the polymerase chain reaction, 24 participants were ESBL positive and subsequently randomised and treated with either the study drug or a placebo. The study was powered for 124 participants. Because of a very high dropout rate, the study was prematurely terminated. From the outbreak cohort (n=247, only eight patients were screened, and only one was positive with the outbreak strain in faeces. Conclusions: The present study design, using IgY chicken antibodies for the eradication of ESBL-producing K. pneumonia and E. coli, was ineffective in reaching its goal due to high mortality and other factors resulting in a low inclusion rate. Spontaneous eradication of ESBL-producing bacteria was frequently observed in recruited participants, which is consistent with previous reports.

  13. Phenotypic changes contributing to Enterobacter gergoviae biocide resistance.

    Science.gov (United States)

    Périamé, M; Philippe, N; Condell, O; Fanning, S; Pagès, J-M; Davin-Regli, A

    2015-08-01

    Enterobacter gergoviae is a recurrent contaminant of cosmetic and hygiene products. To understand how this bacterium adapts to biocides, we studied Ent. gergoviae CIP 76.01 and its triclosan and Methylisothiazolinone-chloromethylisothiazolinone (MIT-CMIT) tolerant isogenic mutants. They were compared with others also isolated from contaminated cosmetics. Phenotypic differences were noted and these included changes in the bacterial envelope and flagella along with differences in motility, and biofilm growth rates. Triclosan and MIT-CMIT derivatives expressed flagella and other MIT-CMIT derivatives exhibited some external appendages. Those bacteria expressing a high-level minimal inhibitory concentration to MIT-CMIT, expressed a strong biofilm formation. No differential phenotypes were noted for carbon source utilisation. Enterobacter gergoviae demonstrated a diverse response to both of these preservatives contained in cosmetic preparations, depending on their concentrations. Interestingly, this adaptive response is associated with modifications of filament structure-related proteins contributing to increase the organism motility and the production of biofilm. Recurrent contaminations of cosmetics products by Ent. gergoviae, needed a better understanding concerning the bacterial adaptation to preservative agents, with particular concern to triclosan and MIT-CMIT. We demonstrated that bacteria response is associated to various mechanisms represented by expression of external appendages (pili or fimbriae) that control cell motility and biofilm formation and evolving as the concentration of biocides adaptation increased. Such mechanisms which are not chemical specific can also promote a cross-resistance to other biocidal agents. The characterization of Ent. gergoviae adaptability to biocides allows industry to adjust the ranges of concentrations and composition of preservatives in formula. © 2015 The Society for Applied Microbiology.

  14. In silico analysis of usher encoding genes in Klebsiella pneumoniae and characterization of their role in adhesion and colonization.

    Directory of Open Access Journals (Sweden)

    Fida Khater

    Full Text Available Chaperone/usher (CU assembly pathway is used by a wide range of Enterobacteriaceae to assemble adhesive surface structures called pili or fimbriae that play a role in bacteria-host cell interactions. In silico analysis revealed that the genome of Klebsiella pneumoniae LM21 harbors eight chromosomal CU loci belonging to γκп and ϭ clusters. Of these, only two correspond to previously described operons, namely type 1 and type 3-encoding operons. Isogenic usher deletion mutants of K. pneumoniae LM21 were constructed for each locus and their role in adhesion to animal (Intestine 407 and plant (Arabidopsis thaliana cells, biofilm formation and murine intestinal colonization was investigated. Type 3 pili usher deleted mutant was impaired in all assays, whereas type 1 pili usher deleted mutant only showed attenuation in adhesion to plant cells and in intestinal colonization. The LM21ΔkpjC mutant was impaired in its capacity to adhere to Arabidopsis cells and to colonize the murine intestine, either alone or in co-inoculation experiments. Deletion of LM21kpgC induced a significant decrease in biofilm formation, in adhesion to animal cells and in colonization of the mice intestine. The LM21∆kpaC and LM21∆kpeC mutants were only attenuated in biofilm formation and the adhesion abilities to Arabidopsis cells, respectively. No clear in vitro or in vivo effect was observed for LM21∆kpbC and LM21∆kpdC mutants. The multiplicity of CU loci in K. pneumoniae genome and their specific adhesion pattern probably reflect the ability of the bacteria to adhere to different substrates in its diverse ecological niches.

  15. Roles of Chaperone/Usher Pathways of Yersinia pestis in a Murine Model of Plague and Adhesion to Host Cells

    Science.gov (United States)

    Hatkoff, Matthew; Runco, Lisa M.; Pujol, Celine; Jayatilaka, Indralatha; Furie, Martha B.; Bliska, James B.

    2012-01-01

    Yersinia pestis and many other Gram-negative pathogenic bacteria use the chaperone/usher (CU) pathway to assemble virulence-associated surface fibers termed pili or fimbriae. Y. pestis has two well-characterized CU pathways: the caf genes coding for the F1 capsule and the psa genes coding for the pH 6 antigen. The Y. pestis genome contains additional CU pathways that are capable of assembling pilus fibers, but the roles of these pathways in the pathogenesis of plague are not understood. We constructed deletion mutations in the usher genes for six of the additional Y. pestis CU pathways. The wild-type (WT) and usher deletion strains were compared in the murine bubonic (subcutaneous) and pneumonic (intranasal) plague infection models. Y. pestis strains containing deletions in CU pathways y0348-0352, y1858-1862, and y1869-1873 were attenuated for virulence compared to the WT strain by the intranasal, but not subcutaneous, routes of infection, suggesting specific roles for these pathways during pneumonic plague. We examined binding of the Y. pestis WT and usher deletion strains to A549 human lung epithelial cells, HEp-2 human cervical epithelial cells, and primary human and murine macrophages. Y. pestis CU pathways y0348-0352 and y1858-1862 were found to contribute to adhesion to all host cells tested, whereas pathway y1869-1873 was specific for binding to macrophages. The correlation between the virulence attenuation and host cell binding phenotypes of the usher deletion mutants identifies three of the additional CU pathways of Y. pestis as mediating interactions with host cells that are important for the pathogenesis of plague. PMID:22851745

  16. In Silico Analysis of Usher Encoding Genes in Klebsiella pneumoniae and Characterization of Their Role in Adhesion and Colonization

    Science.gov (United States)

    Khater, Fida; Balestrino, Damien; Charbonnel, Nicolas; Dufayard, Jean François; Brisse, Sylvain; Forestier, Christiane

    2015-01-01

    Chaperone/usher (CU) assembly pathway is used by a wide range of Enterobacteriaceae to assemble adhesive surface structures called pili or fimbriae that play a role in bacteria-host cell interactions. In silico analysis revealed that the genome of Klebsiella pneumoniae LM21 harbors eight chromosomal CU loci belonging to γκп and ϭ clusters. Of these, only two correspond to previously described operons, namely type 1 and type 3-encoding operons. Isogenic usher deletion mutants of K. pneumoniae LM21 were constructed for each locus and their role in adhesion to animal (Intestine 407) and plant (Arabidopsis thaliana) cells, biofilm formation and murine intestinal colonization was investigated. Type 3 pili usher deleted mutant was impaired in all assays, whereas type 1 pili usher deleted mutant only showed attenuation in adhesion to plant cells and in intestinal colonization. The LM21ΔkpjC mutant was impaired in its capacity to adhere to Arabidopsis cells and to colonize the murine intestine, either alone or in co-inoculation experiments. Deletion of LM21kpgC induced a significant decrease in biofilm formation, in adhesion to animal cells and in colonization of the mice intestine. The LM21∆kpaC and LM21∆kpeC mutants were only attenuated in biofilm formation and the adhesion abilities to Arabidopsis cells, respectively. No clear in vitro or in vivo effect was observed for LM21∆kpbC and LM21∆kpdC mutants. The multiplicity of CU loci in K. pneumoniae genome and their specific adhesion pattern probably reflect the ability of the bacteria to adhere to different substrates in its diverse ecological niches. PMID:25751658

  17. The isotopic biosignatures of photo- vs. thiotrophic bivalves: are they preserved in fossil shells?

    Science.gov (United States)

    Dreier, A; Loh, W; Blumenberg, M; Thiel, V; Hause-Reitner, D; Hoppert, M

    2014-09-01

    Symbiont-bearing and non-symbiotic marine bivalves were used as model organisms to establish biosignatures for the detection of distinctive symbioses in ancient bivalves. For this purpose, the isotopic composition of lipids (δ13C) and bulk organic shell matrix (δ13C, δ34S, δ15N) from shells of several thiotrophic, phototrophic, or non-symbiotic bivalves were compared (phototrophic: Fragum fragum, Fragum unedo, Tridacna maxima; thiotrophic: Codakia tigerina, Fimbria fimbriata, Anodontia sp.; non-symbiotic: Tapes dorsatus, Vasticardium vertebratum, Scutarcopagia sp.). ∆13C values of bulk organic shell matrices, most likely representing mainly original shell protein/chitin biomass, were depleted in thio- and phototrophic bivalves compared to non-symbiotic bivalves. As the bulk organic shell matrix also showed a major depletion of δ15N (down to -2.2 ‰) for thiotrophic bivalves, combined δ13C and δ15N values are useful to differentiate between thio-, phototrophic, and non-symbiotic lifestyles. However, the use of these isotopic signatures for the study of ancient bivalves is limited by the preservation of the bulk organic shell matrix in fossils. Substantial alteration was clearly shown by detailed microscopic analyses of fossil (late Pleistocene) T. maxima and Trachycardium lacunosum shell, demonstrating a severe loss of quantity and quality of bulk organic shell matrix with time. Likewise, the composition and δ13C-values of lipids from empty shells indicated that a large part of these compounds derived from prokaryotic decomposers. The use of lipids from ancient shells for the reconstruction of the bivalve's life style therefore appears to be restricted. © 2014 John Wiley & Sons Ltd.

  18. Studies on the characterization and regulation of alpha-1 adrenergic receptors and [3H]WB4101 binding sites in the central nervous system

    International Nuclear Information System (INIS)

    Morrow, A.L.

    1985-01-01

    The purpose of these studies has been to resolve the anomalous binding characteristics of two alpha adrenergic receptor ligands, [ 3 H]WB4101 and [ 3 H]prazosin and to study the regulation of the receptors labeled by these compounds after surgical denervation and chronic drug treatments. Preliminary studies indicated that [ 3 H]WB4101 binding sites, which were believed to represent alpha-1 adrenergic receptors, were increased in number following removal of the fimbrial afferents to the hippocampus. This increase was not due to removal of the adrenergic input into this structure since destruction of the locus coeruleus or the dorsal noradrenergic bundle did not produce the up-regulation. Characterization of alpha-1 adrenergic receptors using [ 3 H]prazosin and [ 3 H]WB4101 revealed evidence for subtypes of alpha-1 receptors designated alpha-1A and alpha-1B. The nanomolar affinity component of [ 3 H]WB4101 binding is not adrenergic but serotonergic. The serotonergic agonists, serotonin and 8-hydroxy-dipropylaminotetraline have affinities of 1.5 and 3.0 nM for this site, when studied in the presence of a 30 nM prazosin mask of the alpha-1 component of [ 3 H]WB4101 binding. Fimbria transection or 5,7 dihydroxytryptamine injections produced increases in the Bmax of the nanomolar affinity component of [ 3 H]WB4101 binding in the presence of a prazosin mask. The up-regulated site showed identical serotonergic pharmacology compared to control tissue. Thus, the author concluded that serotonergic denervation of the hippocampus produces the increase in serotonergic binding sites labeled by [ 3 H]WB4101

  19. Cranberry extract inhibits in vitro adhesion of F4 and F18+Escherichia coli to pig intestinal epithelium and reduces in vivo excretion of pigs orally challenged with F18+ verotoxigenic E. coli.

    Science.gov (United States)

    Coddens, Annelies; Loos, Michaela; Vanrompay, Daisy; Remon, Jean Paul; Cox, Eric

    2017-04-01

    F4 + E. coli and F18 + E. coli infections are an important threat for pig industry worldwide. Antibiotics are commonly used to treat infected piglets, but the emerging development of resistance against antibiotics raises major concerns. Hence, alternative therapies to prevent pigs from F4 + E. coli and F18 + E. coli infections need to be developed. Since cranberry previously showed anti-adhesive activity against uropathogenic E. coli, we aimed to investigate whether cranberry extract could also inhibit binding of F4 + E. coli and F18 + E. coli to pig intestinal epithelium. Using the in vitro villus adhesion assay, we found that low concentrations of cranberry extract (20μg or 100μg/ml) have strong inhibitory activity on F4 + E. coli (75.3%, S.D.=9.31 or 95.8%, S.D.=2.56, respectively) and F18 + E. coli adherence (100% inhibition). This effect was not due to antimicrobial activity. Moreover, cranberry extract (10mg or 100mg) could also abolish in vivo binding of F4 and F18 fimbriae to the pig intestinal epithelium in ligated loop experiments. Finally, two challenge experiments with F18 + E. coli were performed to address the efficacy of in-feed or water supplemented cranberry extract. No effect could be observed in piglets that received cranberry extract only in feed (1g/kg or 10g/kg). However, supplementation of feed (10g/kg) and drinking water (1g/L) significantly decreased excretion and diarrhea. The decreased infection resulted in a decreased serum antibody response indicating reduced exposure to F18 + E. coli. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Characterization of the biomechanical properties of T4 pili expressed by Streptococcus pneumoniae--a comparison between helix-like and open coil-like pili.

    Science.gov (United States)

    Castelain, Mickaël; Koutris, Efstratios; Andersson, Magnus; Wiklund, Krister; Björnham, Oscar; Schedin, Staffan; Axner, Ove

    2009-07-13

    Bacterial adhesion organelles, known as fimbria or pili, are expressed by gram-positive as well as gram-negative bacteria families. These appendages play a key role in the first steps of the invasion and infection processes, and they therefore provide bacteria with pathogenic abilities. To improve the knowledge of pili-mediated bacterial adhesion to host cells and how these pili behave under the presence of an external force, we first characterize, using force measuring optical tweezers, open coil-like T4 pili expressed by gram-positive Streptococcus pneumoniae with respect to their biomechanical properties. It is shown that their elongation behavior can be well described by the worm-like chain model and that they possess a large degree of flexibility. Their properties are then compared with those of helix-like pili expressed by gram-negative uropathogenic Escherichia coli (UPEC), which have different pili architecture. The differences suggest that these two types of pili have distinctly dissimilar mechanisms to adhere and sustain external forces. Helix-like pili expressed by UPEC bacteria adhere to host cells by single adhesins located at the distal end of the pili while their helix-like structures act as shock absorbers to dampen the irregularly shear forces induced by urine flow and to increase the cooperativity of the pili ensemble, whereas open coil-like pili expressed by S. pneumoniae adhere to cells by a multitude of adhesins distributed along the pili. It is hypothesized that these two types of pili represent different strategies of adhering to host cells in the presence of external forces. When exposed to significant forces, bacteria expressing helix-like pili remain attached by distributing the external force among a multitude of pili, whereas bacteria expressing open coil-like pili sustain large forces primarily by their multitude of binding adhesins which presumably detach sequentially.

  1. Carriage of extended-spectrum beta-lactamase-plasmids does not reduce fitness but enhances virulence in some strains of pandemic E. coli lineages

    Directory of Open Access Journals (Sweden)

    Katharina eSchaufler

    2016-03-01

    Full Text Available Pathogenic ESBL-producing E. coli lineages occur frequently worldwide, not only in a human health context but in animals and the environment, also in settings with low antimicrobial pressures. This study investigated the fitness costs of ESBL-plasmids and their influence on chromosomally encoded features associated with virulence, such as those involved in the planktonic and sessile behaviors of ST131 and ST648 E. coli. ESBL-plasmid-carrying wild-type E. coli strains, their corresponding ESBL-plasmid-cured variants (PCV, and complementary ESBL-carrying transformants were comparatively analyzed using growth curves, Omnilog® phenotype microarray (PM assays, macrocolony and biofilm formation, swimming motility, and RNA sequence analysis. Growth curves and PM results pointed towards similar growth and metabolic behaviors among the strains. Phenotypic differences in some strains were detected, including enhanced curli fimbriae and/or cellulose production as well as a reduced swimming capacity of some ESBL-carrying strains, as compared to their respective PCVs. RNA sequencing mostly confirmed the phenotypic results, suggesting that the chromosomally encoded csgD pathway is a key factor involved. These results contradict the hypothesis that ESBL-plasmid-carriage leads to a fitness loss in ESBL-carrying strains. Instead, the results indicate an influence of some ESBL-plasmids on chromosomally encoded features associated with virulence in some E. coli strains. In conclusion, apart from antibiotic resistance selective advantages, ESBL-plasmid-carriage may also lead to enhanced virulence or adaption to specific habitats in some strains of pandemic ESBL-producing E. coli lineages.

  2. Virulence Genes and Antibiotic Susceptibilities of Uropathogenic E. coli Strains.

    Science.gov (United States)

    Uzun, Cengiz; Oncül, Oral; Gümüş, Defne; Alan, Servet; Dayioğlu, Nurten; Küçüker, Mine Anğ

    2015-01-01

    The aim of this study is to detect the presence of and possible relation between virulence genes and antibiotic resistance in E. coli strains isolated from patients with acute, uncomplicated urinary tract infections (UTI). 62 E. coli strains isolated from patients with acute, uncomplicated urinary tract infections (50 strains isolated from acute uncomplicated cystitis cases (AUC); 12 strains from acute uncomplicated pyelonephritis cases (AUP)) were screened for virulence genes [pap (pyelonephritis-associated pili), sfa/foc (S and F1C fimbriae), afa (afimbrial adhesins), hly (hemolysin), cnf1 (cytotoxic necrotizing factor), aer (aerobactin), PAI (pathogenicity island marker), iroN (catecholate siderophore receptor), ompT (outer membrane protein T), usp (uropathogenic specific protein)] by PCR and for antimicrobial resistance by disk diffusion method according to CLSI criteria. It was found that 56 strains (90.3%) carried at least one virulence gene. The most common virulence genes were ompT (79%), aer (51.6%), PAI (51.6%) and usp (56.5%). 60% of the strains were resistant to at least one antibiotic. The highest resistance rates were against ampicillin (79%) and co-trimoxazole (41.9%). Fifty percent of the E. coli strains (31 strains) were found to be multiple resistant. Eight (12.9%) out of 62 strains were found to be ESBL positive. Statistically significant relationships were found between the absence of usp and AMP - SXT resistance, iroN and OFX - CIP resistance, PAI and SXT resistance, cnf1 and AMP resistance, and a significant relationship was also found between the presence of the afa and OFX resistance. No difference between E. coli strains isolated from two different clinical presentations was found in terms of virulence genes and antibiotic susceptibility.

  3. Administration of probiotics influences F4 (K88)-positive enterotoxigenic Escherichia coli attachment and intestinal cytokine expression in weaned pigs.

    Science.gov (United States)

    Daudelin, Jean-François; Lessard, Martin; Beaudoin, Frédéric; Nadeau, Eric; Bissonnette, Nathalie; Boutin, Yvan; Brousseau, Jean-Philippe; Lauzon, Karoline; Fairbrother, John Morris

    2011-05-23

    This study evaluated the effect of the probiotics Pediococcus acidilactici and Saccharomyces cerevisiae boulardii on the intestinal colonization of O149 enterotoxigenic Escherichia coli harbouring the F4 (K88) fimbriae (ETEC F4) and on the expression of ileal cytokines in weaned pigs. At birth, different litters of pigs were randomly assigned to one of the following treatments: 1) control without antibiotics or probiotics (CTRL); 2) reference group in which chlortetracycline and tiamulin were added to weanling feed (ATB); 3) P. acidilactici; 4) S. cerevisiae boulardii; or 5) P. acidilactici + S. cerevisiae boulardii. Probiotics were administered daily (1 × 10(9) CFU per pig) during the lactation period and after weaning (day 21). At 28 days of age, all pigs were orally challenged with an ETEC F4 strain, and a necropsy was performed 24 h later. Intestinal segments were collected to evaluate bacterial colonization in the small intestine and ileal cytokine expressions. Attachment of ETEC F4 to the intestinal mucosa was significantly reduced in pigs treated with P. acidilactici or S. cerevisiae boulardii in comparison with the ATB group (P = 0.01 and P = 0.03, respectively). In addition, proinflammatory cytokines, such as IL-6, were upregulated in ETEC F4 challenged pigs treated with P. acidilactici alone or in combination with S. cerevisiae boulardii compared with the CTRL group. In conclusion, the administration of P. acidilactici or S. cerevisiae boulardii was effective in reducing ETEC F4 attachment to the ileal mucosa, whereas the presence of P. acidilactici was required to modulate the expression of intestinal inflammatory cytokines in pigs challenged with ETEC F4.

  4. A novel growth-promoting microbe, Methylobacterium funariae sp. nov., isolated from the leaf surface of a common moss.

    Science.gov (United States)

    Schauer, S; Kutschera, U

    2011-04-01

    Land plants (embryophytes) evolved in the presence of prokaryotic microbes. As a result, numerous mutually beneficial associations (symbioses) developed that can be analyzed using a variety of methods. Here we describe the isolation and characterization of a new pink-pigmented facultatively methylotrophic symbiotic bacterium of the genus Methylobacterium (laboratory strain F3.2) that was isolated from the gametophytic phylloids of the common cord moss Funaria hygrometrica Hedw. Plantlets were collected in the field and analyzed in the laboratory. Colonies of methylobacteria were obtained by the agar-impression-method. Based on its unique phenotype (the bacterial cells are characterized by fimbriae-like appendages), a comparative 16S rRNA gene (DNA) sequence analysis, and an average DNA-DNA hybridization value of 8,4 %, compared with its most closely related sister taxon, this isolate is described as a new species, Methylobacterium funariae sp. nov. (type strain F3.2). This new epiphytic bacterium inhabits the leaf surface of "primitive" land plants such as mosses and interacts with its host organism via the secretion of phytohormones (cytokinines, auxins). These external signals are perceived by the plant cells that divide and grow more rapidly than in the absence of their prokaryotic phytosymbionts. We suggest that M. funariae sp. nov. uses methanol emitted from the stomatal pores as principal carbon source for cell metabolism. However, our novel data indicate that, in this unique symbiotic plant-microbe interaction, the uptake of amino acids leached from the surface of the epidermal cells of the green host organism may be of importance as microbial carbon- and nitrogen-source.

  5. Hypothyroidism affects differentially the cell size of epithelial cells among oviductal regions of rabbits.

    Science.gov (United States)

    Anaya-Hernández, A; Rodríguez-Castelán, J; Nicolás, L; Martínez-Gómez, M; Jiménez-Estrada, I; Castelán, F; Cuevas, E

    2015-02-01

    Oviductal regions show particular histological characteristics and functions. Tubal pathologies and hypothyroidism are related to primary and secondary infertility. The impact of hypothyroidism on the histological characteristics of oviductal regions has been scarcely studied. Our aim was to analyse the histological characteristics of oviductal regions in control and hypothyroid rabbits. Hypothyroidism was induced by oral administration of methimazole (MMI) for 30 days. For both groups, serum concentrations of thyroid and gonadal hormones were determined. Sections of oviductal regions were stained with the Masson's trichrome technique to analyse both epithelial and smooth muscle layers. The percentage of proliferative epithelial cells (anti-Ki67) in diverse oviductal regions was also quantified. Data were compared with Student t-test, Mann-Whitney U-test, or Fischer's test. In comparison with the control group, the hypothyroid group showed: (i) a low concentration of T3 and T4, but a high level of TSH; (ii) similar values of serum estradiol, progesterone and testosterone; (iii) a large size of ciliated cells in the ampulla (AMP), isthmus (IST) and utero-tubal junction (UTJ); (iv) a large size of secretory cells in the IST region; (v) a low percentage of proliferative secretory cells in the fimbria-infundibulum (FIM-INF) region; and (vi) a similar thickness of the smooth muscle layer and the cross-sectional area in the AMP and IST regions. Modifications in the size of the oviductal epithelium in hypothyroid rabbits could be related to changes in the cell metabolism that may impact on the reproductive functions achieved by oviduct. © 2014 Blackwell Verlag GmbH.

  6. Characterization of Novel Factors Involved in Swimming and Swarming Motility in Salmonella enterica Serovar Typhimurium.

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    Julia Andrea Deditius

    Full Text Available Salmonella enterica utilizes flagellar motility to swim through liquid environments and on surfaces. The biosynthesis of the flagellum is regulated on various levels, including transcriptional and posttranscriptional mechanisms. Here, we investigated the motility phenotype of 24 selected single gene deletions that were previously described to display swimming and swarming motility effects. Mutations in flgE, fliH, ydiV, rfaG, yjcC, STM1267 and STM3363 showed an altered motility phenotype. Deletions of flgE and fliH displayed a non-motile phenotype in both swimming and swarming motility assays as expected. The deletions of STM1267, STM3363, ydiV, rfaG and yjcC were further analyzed in detail for flagellar and fimbrial gene expression and filament formation. A ΔydiV mutant showed increased swimming motility, but a decrease in swarming motility, which coincided with derepression of curli fimbriae. A deletion of yjcC, encoding for an EAL domain-containing protein, increased swimming motility independent on flagellar gene expression. A ΔSTM1267 mutant displayed a hypermotile phenotype on swarm agar plates and was found to have increased numbers of flagella. In contrast, a knockout of STM3363 did also display an increase in swarming motility, but did not alter flagella numbers. Finally, a deletion of the LPS biosynthesis-related protein RfaG reduced swimming and swarming motility, associated with a decrease in transcription from flagellar class II and class III promoters and a lack of flagellar filaments.

  7. [Mechanisms of urinary tract sterility maintenance].

    Science.gov (United States)

    Okrągła, Emilia; Szychowska, Katarzyna; Wolska, Lidia

    2014-06-02

    Physiologically, urine and the urinary tract are maintained sterile because of physical and chemical properties of urine and the innate immune system's action. The urinary tract is constantly exposed to the invasion of microorganisms from the exterior environment, also because of the anatomical placement of the urethra, in the vicinity of the rectum. Particularly vulnerable to urinary tract infections (UTI) are women (an additional risk factor is pregnancy), but also the elderly and children. The main pathogens causing UTI are bacteria; in 70-95% of cases it is the bacterium Escherichia coli. Infections caused by viruses and fungi are less common and are associated with decreased immunity, pharmacotherapy, or some diseases. Bacteria have evolved a number of factors that facilitate the colonization of the urinary tract: the cover and cell membrane antigens O and K1, lipopolysaccharide (LPS), fimbriae, pile and cilia. On the other hand, the human organism has evolved mechanisms to hinder colonization of the urinary tract: mechanisms arising from the anatomical structure of the urinary tract, the physicochemical properties of the urine and the activity of the innate immune system, also known as non-specific, which isolates and destroys pathogens using immunological processes, and the mechanisms for release of antimicrobial substances such as Tamm-Horsfall protein, mucopolysaccharides, immunoglobulins IgA and IgG, lactoferrin, lipocalin, neutrophils, cytokines and antimicrobial peptides. This review aims to analyze the state of knowledge on the mechanisms to maintain the sterility of the urinary tract used by the human organism and bacterial virulence factors to facilitate the colonization of the urinary tract.

  8. Detection of virulence-associated genes in pathogenic and commensal avian Escherichia coli isolates.

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    Paixão, A C; Ferreira, A C; Fontes, M; Themudo, P; Albuquerque, T; Soares, M C; Fevereiro, M; Martins, L; Corrêa de Sá, M I

    2016-07-01

    Poultry colibacillosis due to Avian Pathogenic Escherichia coli (APEC) is responsible for several extra-intestinal pathological conditions, leading to serious economic damage in poultry production. The most commonly associated pathologies are airsacculitis, colisepticemia, and cellulitis in broiler chickens, and salpingitis and peritonitis in broiler breeders. In this work a total of 66 strains isolated from dead broiler breeders affected with colibacillosis and 61 strains from healthy broilers were studied. Strains from broiler breeders were typified with serogroups O2, O18, and O78, which are mainly associated with disease. The serogroup O78 was the most prevalent (58%). All the strains were checked for the presence of 11 virulence genes: 1) arginine succinyltransferase A (astA); ii) E.coli hemeutilization protein A (chuA); iii) colicin V A/B (cvaA/B); iv) fimbriae mannose-binding type 1 (fimC); v) ferric yersiniabactin uptake A (fyuA); vi) iron-repressible high-molecular-weight proteins 2 (irp2); vii) increased serum survival (iss); viii) iron-uptake systems of E.coli D (iucD); ix) pielonefritis associated to pili C (papC); x) temperature sensitive haemaglutinin (tsh), and xi) vacuolating autotransporter toxin (vat), by Multiplex-PCR. The results showed that all genes are present in both commensal and pathogenic E. coli strains. The iron uptake-related genes and the serum survival gene were more prevalent among APEC. The adhesin genes, except tsh, and the toxin genes, except astA, were also more prevalent among APEC isolates. Except for astA and tsh, APEC strains harbored the majority of the virulence-associated genes studied and fimC was the most prevalent gene, detected in 96.97 and 88.52% of APEC and AFEC strains, respectively. Possession of more than one iron transport system seems to play an important role on APEC survival. © 2016 Poultry Science Association Inc.

  9. Survey of protocols for the manual segmentation of the hippocampus: preparatory steps towards a joint EADC-ADNI harmonized protocol.

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    Boccardi, Marina; Ganzola, Rossana; Bocchetta, Martina; Pievani, Michela; Redolfi, Alberto; Bartzokis, George; Camicioli, Richard; Csernansky, John G; de Leon, Mony J; deToledo-Morrell, Leyla; Killiany, Ronald J; Lehéricy, Stéphane; Pantel, Johannes; Pruessner, Jens C; Soininen, H; Watson, Craig; Duchesne, Simon; Jack, Clifford R; Frisoni, Giovanni B

    2011-01-01

    Manual segmentation from magnetic resonance imaging (MR) is the gold standard for evaluating hippocampal atrophy in Alzheimer's disease (AD). Nonetheless, different segmentation protocols provide up to 2.5-fold volume differences. Here we surveyed the most frequently used segmentation protocols in the AD literature as a preliminary step for international harmonization. The anatomical landmarks (anteriormost and posteriormost slices, superior, inferior, medial, and lateral borders) were identified from 12 published protocols for hippocampal manual segmentation ([Abbreviation] first author, publication year: [B] Bartzokis, 1998; [C] Convit, 1997; [dTM] deToledo-Morrell, 2004; [H] Haller, 1997; [J] Jack, 1994; [K] Killiany, 1993; [L] Lehericy, 1994; [M] Malykhin, 2007; [Pa] Pantel, 2000; [Pr] Pruessner, 2000; [S] Soininen, 1994; [W] Watson, 1992). The hippocampi of one healthy control and one AD patient taken from the 1.5T MR ADNI database were segmented by a single rater according to each protocol. The accuracy of the protocols' interpretation and translation into practice was checked with lead authors of protocols through individual interactive web conferences. Semantically harmonized landmarks and differences were then extracted, regarding: (a) the posteriormost slice, protocol [B] being the most restrictive, and [H, M, Pa, Pr, S] the most inclusive; (b) inclusion [C, dTM, J, L, M, Pr, W] or exclusion [B, H, K, Pa, S] of alveus/fimbria; (c) separation from the parahippocampal gyrus, [C] being the most restrictive, [B, dTM, H, J, Pa, S] the most inclusive. There were no substantial differences in the definition of the anteriormost slice. This survey will allow us to operationalize differences among protocols into tracing units, measure their impact on the repeatability and diagnostic accuracy of manual hippocampal segmentation, and finally develop a harmonized protocol.

  10. Immunogenicity and safety of a CRM-conjugated meningococcal ACWY vaccine administered concomitantly with routine vaccines starting at 2 months of age.

    Science.gov (United States)

    Nolan, Terry M; Nissen, Michael D; Naz, Aftab; Shepard, Julie; Bedell, Lisa; Hohenboken, Matthew; Odrljin, Tatjana; Dull, Peter M

    2014-01-01

    Infants are at the highest risk for meningococcal disease and a broadly protective and safe vaccine is an unmet need in this youngest population. We evaluated the immunogenicity and safety of a 4-dose infant/toddler regimen of MenACWY-CRM given at 2, 4, 6, and 12 months of age concomitantly with pentavalent diphtheria-tetanus-acellular pertussis-Hemophilus influenzae type b-inactivated poliovirus-combination vaccine (DTaP-IPV/Hib), hepatitis B vaccine (HBV), 7- or 13-valent conjugate pneumococcal vaccine (PCV), and measles, mumps, and rubella vaccine (MMR). Four doses of MenACWY-CRM induced hSBA titers ≥8 in 89%, 95%, 97%, and 96% of participants against serogroups A, C, W-135, and Y, respectively. hSBA titers ≥8 were present in 76-98% of participants after the first 3 doses. A categorical linear analysis incorporating vaccine group and study center showed responses to routine vaccines administered with MenACWY-CRM were non-inferior to routine vaccines alone, except for seroresponse to the pertussis antigen fimbriae. The reactogenicity profile was not affected when MenACWY-CRM was administered concomitantly with routine vaccines. MenACWY-CRM administered with routine concomitant vaccinations in young infants was well tolerated and induced highly immunogenic responses against each of the serogroups without significant interference with the immune responses to routine infant vaccinations. Healthy 2 month old infants were randomized to receive MenACWY-CRM with routine vaccines (n = 258) or routine vaccines alone (n = 271). Immunogenicity was assessed by serum bactericidal assay using human complement (hSBA). Medically attended adverse events (AEs), serious AEs (SAEs) and AEs leading to study withdrawal were collected throughout the study period.

  11. Biofunctionalization on alkylated silicon substrate surfaces via "click" chemistry.

    Science.gov (United States)

    Qin, Guoting; Santos, Catherine; Zhang, Wen; Li, Yan; Kumar, Amit; Erasquin, Uriel J; Liu, Kai; Muradov, Pavel; Trautner, Barbara Wells; Cai, Chengzhi

    2010-11-24

    Biofunctionalization of silicon substrates is important to the development of silicon-based biosensors and devices. Compared to conventional organosiloxane films on silicon oxide intermediate layers, organic monolayers directly bound to the nonoxidized silicon substrates via Si-C bonds enhance the sensitivity of detection and the stability against hydrolytic cleavage. Such monolayers presenting a high density of terminal alkynyl groups for bioconjugation via copper-catalyzed azide-alkyne 1,3-dipolar cycloaddition (CuAAC, a "click" reaction) were reported. However, yields of the CuAAC reactions on these monolayer platforms were low. Also, the nonspecific adsorption of proteins on the resultant surfaces remained a major obstacle for many potential biological applications. Herein, we report a new type of "clickable" monolayers grown by selective, photoactivated surface hydrosilylation of α,ω-alkenynes, where the alkynyl terminal is protected with a trimethylgermanyl (TMG) group, on hydrogen-terminated silicon substrates. The TMG groups on the film are readily removed in aqueous solutions in the presence of Cu(I). Significantly, the degermanylation and the subsequent CuAAC reaction with various azides could be combined into a single step in good yields. Thus, oligo(ethylene glycol) (OEG) with an azido tag was attached to the TMG-alkyne surfaces, leading to OEG-terminated surfaces that reduced the nonspecific adsorption of protein (fibrinogen) by >98%. The CuAAC reaction could be performed in microarray format to generate arrays of mannose and biotin with varied densities on the protein-resistant OEG background. We also demonstrated that the monolayer platform could be functionalized with mannose for highly specific capturing of living targets (Escherichia coli expressing fimbriae) onto the silicon substrates.

  12. Porcine E. coli: virulence-associated genes, resistance genes and adhesion and probiotic activity tested by a new screening method.

    Science.gov (United States)

    Schierack, Peter; Rödiger, Stefan; Kuhl, Christoph; Hiemann, Rico; Roggenbuck, Dirk; Li, Ganwu; Weinreich, Jörg; Berger, Enrico; Nolan, Lisa K; Nicholson, Bryon; Römer, Antje; Frömmel, Ulrike; Wieler, Lothar H; Schröder, Christian

    2013-01-01

    We established an automated screening method to characterize adhesion of Escherichia coli to intestinal porcine epithelial cells (IPEC-J2) and their probiotic activity against infection by enteropathogenic E. coli (EPEC). 104 intestinal E. coli isolates from domestic pigs were tested by PCR for the occurrence of virulence-associated genes, genes coding for resistances to antimicrobial agents and metals, and for phylogenetic origin by PCR. Adhesion rates and probiotic activity were examined for correlation with the presence of these genes. Finally, data were compared with those from 93 E. coli isolates from wild boars. Isolates from domestic pigs carried a broad variety of all tested genes and showed great diversity in gene patterns. Adhesions varied with a maximum of 18.3 or 24.2 mean bacteria adherence per epithelial cell after 2 or 6 hours respectively. Most isolates from domestic pigs and wild boars showed low adherence, with no correlation between adhesion/probiotic activity and E. coli genes or gene clusters. The gene sfa/foc, encoding for a subunit of F1C fimbriae did show a positive correlative association with adherence and probiotic activity; however E. coli isolates from wild boars with the sfa/foc gene showed less adhesion and probiotic activity than E. coli with the sfa/foc gene isolated from domestic pigs after 6 hour incubation. In conclusion, screening porcine E. coli for virulence associated genes genes, adhesion to intestinal epithelial cells, and probiotic activity revealed a single important adhesion factor, several probiotic candidates, and showed important differences between E. coli of domestic pigs and wild boars.

  13. Porcine E. coli: virulence-associated genes, resistance genes and adhesion and probiotic activity tested by a new screening method.

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    Peter Schierack

    Full Text Available We established an automated screening method to characterize adhesion of Escherichia coli to intestinal porcine epithelial cells (IPEC-J2 and their probiotic activity against infection by enteropathogenic E. coli (EPEC. 104 intestinal E. coli isolates from domestic pigs were tested by PCR for the occurrence of virulence-associated genes, genes coding for resistances to antimicrobial agents and metals, and for phylogenetic origin by PCR. Adhesion rates and probiotic activity were examined for correlation with the presence of these genes. Finally, data were compared with those from 93 E. coli isolates from wild boars. Isolates from domestic pigs carried a broad variety of all tested genes and showed great diversity in gene patterns. Adhesions varied with a maximum of 18.3 or 24.2 mean bacteria adherence per epithelial cell after 2 or 6 hours respectively. Most isolates from domestic pigs and wild boars showed low adherence, with no correlation between adhesion/probiotic activity and E. coli genes or gene clusters. The gene sfa/foc, encoding for a subunit of F1C fimbriae did show a positive correlative association with adherence and probiotic activity; however E. coli isolates from wild boars with the sfa/foc gene showed less adhesion and probiotic activity than E. coli with the sfa/foc gene isolated from domestic pigs after 6 hour incubation. In conclusion, screening porcine E. coli for virulence associated genes genes, adhesion to intestinal epithelial cells, and probiotic activity revealed a single important adhesion factor, several probiotic candidates, and showed important differences between E. coli of domestic pigs and wild boars.

  14. Phenotypic and molecular characterization of Klebsiella spp. isolates causing community-acquired infections

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    U. Garza-Ramos

    2018-05-01

    Full Text Available Klebsiella spp. isolates from community-acquired infections were characterized. A total of 39 Klebsiella spp. isolates were obtained from outpatients at four rural hospitals in Mexico (2013–2014. The biochemical tests identified all as being K. pneumoniae. The molecular multiplex-PCR test identified 36 (92.4% K. pneumoniae isolates and one (2.5% K. variicola isolate, and phylogenetic analysis of the rpoB gene identified two isolates (5.1% belonging to K. quasipneumoniae subsp. quasipneumoniae and K. quasivariicola. The last one was confirmed by phylogenetic analysis of six-loci concatenated genes. Mostly the isolates were multidrug resistant; however, a minority were extended-spectrum β-lactamase producing (10.2%. The extended-spectrum β-lactamase CTX-M-15 gene was identified in these isolates. Analysis of biofilm production and the hypermucoviscosity phenotype showed a total of 35 (92.3% and seven (17.9% of the isolates were positive for these phenotypes respectively. The K2 (4/39, 10.2%, K5 (2/39, 5.1% and K54 (1/39, 2.5% serotypes were identified in seven (17.9% of the isolates, and only 28.5% (2/7 hypermucoviscous isolates were positive for the K2 and K5 serotypes. In general, the sequence type (ST analysis and phylogenetic analysis of seven multilocus sequence typing loci were heterogeneous; however, ST29 was the most prevalent ST in the analysed isolates, accounting for 19% (4/21 of the total isolates. Two of the four ST29 isolates had the hypermucoviscosity phenotype. The virulence factors for fimbriae were the most prevalent, followed by siderophores. Community-acquired infections are caused by various species from Klebsiella genus, with different profiles of antibiotic resistance and heterogeneous virulence factors. Keywords: Antimicrobial susceptibility, Bacterial resistance, Cephalosporin resistance, Community infection, ESBL, Hypermucoviscosity

  15. Genomic characterization of Haemophilus parasuis SH0165, a highly virulent strain of serovar 5 prevalent in China.

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    Zhuofei Xu

    Full Text Available Haemophilus parasuis can be either a commensal bacterium of the porcine respiratory tract or an opportunistic pathogen causing Glässer's disease, a severe systemic disease that has led to significant economical losses in the pig industry worldwide. We determined the complete genomic sequence of H. parasuis SH0165, a highly virulent strain of serovar 5, which was isolated from a hog pen in North China. The single circular chromosome was 2,269,156 base pairs in length and contained 2,031 protein-coding genes. Together with the full spectrum of genes detected by the analysis of metabolic pathways, we confirmed that H. parasuis generates ATP via both fermentation and respiration, and possesses an intact TCA cycle for anabolism. In addition to possessing the complete pathway essential for the biosynthesis of heme, this pathogen was also found to be well-equipped with different iron acquisition systems, such as the TonB system and ABC-type transport complexes, to overcome iron limitation during infection and persistence. We identified a number of genes encoding potential virulence factors, such as type IV fimbriae and surface polysaccharides. Analysis of the genome confirmed that H. parasuis is naturally competent, as genes related to DNA uptake are present. A nine-mer DNA uptake signal sequence (ACAAGCGGT, identical to that found in Actinobacillus pleuropneumoniae and Mannheimia haemolytica, followed by similar downstream motifs, was identified in the SH0165 genome. Genomic and phylogenetic comparisons with other Pasteurellaceae species further indicated that H. parasuis was closely related to another swine pathogenic bacteria A. pleuropneumoniae. The comprehensive genetic analysis presented here provides a foundation for future research on the metabolism, natural competence and virulence of H. parasuis.

  16. papA gene of avian pathogenic Escherichia coli.

    Science.gov (United States)

    Kariyawasam, Subhashinie; Nolan, Lisa K

    2011-12-01

    P fimbrial adhesins may be associated with the virulence of avian pathogenic Escherichia coli (APEC). However, most APECs are unable to express P fimbriae even when they are grown under conditions that favor P fimbrial expression. This failure can be explained by the complete absence of the pap operon or the presence of an incomplete pap operon in Pap-negative APEC strains. In the present study, we analyzed the pap operon, specifically the papA gene that encodes the major fimbrial shaft, to better understand the pap gene cluster at the genetic level. First, by PCR, we examined a collection of 500 APEC strains for the presence of 11 genes comprising the pap operon. Except for papA, all the other genes of the operon were present in 38% to 41.2% of APEC, whereas the papA was present only in 10.4% of the APEC tested. Using multiplex PCR to probe for allelic variants of papA, we sought to determine if the low prevalence of papA among APEC was related to genetic heterogeneity of the gene itself. It was determined that the papA of APEC always belongs to the F11 allelic variant. Finally, we sequenced the 'papA region' from two papA-negative strains, both of which contain all the other genes of the pap operon. Interestingly, both strains had an 11,104-bp contig interruptingpapA at the 281-bp position. This contig harbored a streptomycin resistance gene and a classic Tn10 transposon containing the genes that confer tetracycline resistance. However, we noted that the papA gene of every papA-negative APEC strain was not interrupted by an 11,104-bp contig. It is likely that transposons bearing antibiotic resistance genes have inserted within pap gene cluster of some APEC strains, and such genetic events may have been selected for by antibiotic use.

  17. Porphyromonas gingivalis Uses Specific Domain Rearrangements and Allelic Exchange to Generate Diversity in Surface Virulence Factors.

    Science.gov (United States)

    Dashper, Stuart G; Mitchell, Helen L; Seers, Christine A; Gladman, Simon L; Seemann, Torsten; Bulach, Dieter M; Chandry, P Scott; Cross, Keith J; Cleal, Steven M; Reynolds, Eric C

    2017-01-01

    Porphyromonas gingivalis is a keystone pathogen of chronic periodontitis. The virulence of P. gingivalis is reported to be strain related and there are currently a number of strain typing schemes based on variation in capsular polysaccharide, the major and minor fimbriae and adhesin domains of Lys-gingipain (Kgp), amongst other surface proteins. P. gingivalis can exchange chromosomal DNA between strains by natural competence and conjugation. The aim of this study was to determine the genetic variability of P. gingivalis strains sourced from international locations over a 25-year period and to determine if variability in surface virulence factors has a phylogenetic basis. Whole genome sequencing was performed on 13 strains and comparison made to 10 previously sequenced strains. A single nucleotide polymorphism-based phylogenetic analysis demonstrated a shallow tri-lobed phylogeny. There was a high level of reticulation in the phylogenetic network, demonstrating extensive horizontal gene transfer between the strains. Two highly conserved variants of the catalytic domain of the major virulence factor the Kgp proteinase (Kgp cat I and Kgp cat II) were found. There were three variants of the fourth Kgp C-terminal cleaved adhesin domain. Specific variants of the cell surface proteins FimA, FimCDE, MfaI, RagAB, Tpr, and PrtT were also identified. The occurrence of all these variants in the P. gingivalis strains formed a mosaic that was not related to the SNP-based phylogeny. In conclusion P. gingivalis uses domain rearrangements and genetic exchange to generate diversity in specific surface virulence factors.

  18. Klebsiella pneumoniae: Going on the Offense with a Strong Defense

    Science.gov (United States)

    Paczosa, Michelle K.

    2016-01-01

    SUMMARY Klebsiella pneumoniae causes a wide range of infections, including pneumonias, urinary tract infections, bacteremias, and liver abscesses. Historically, K. pneumoniae has caused serious infection primarily in immunocompromised individuals, but the recent emergence and spread of hypervirulent strains have broadened the number of people susceptible to infections to include those who are healthy and immunosufficient. Furthermore, K. pneumoniae strains have become increasingly resistant to antibiotics, rendering infection by these strains very challenging to treat. The emergence of hypervirulent and antibiotic-resistant strains has driven a number of recent studies. Work has described the worldwide spread of one drug-resistant strain and a host defense axis, interleukin-17 (IL-17), that is important for controlling infection. Four factors, capsule, lipopolysaccharide, fimbriae, and siderophores, have been well studied and are important for virulence in at least one infection model. Several other factors have been less well characterized but are also important in at least one infection model. However, there is a significant amount of heterogeneity in K. pneumoniae strains, and not every factor plays the same critical role in all virulent Klebsiella strains. Recent studies have identified additional K. pneumoniae virulence factors and led to more insights about factors important for the growth of this pathogen at a variety of tissue sites. Many of these genes encode proteins that function in metabolism and the regulation of transcription. However, much work is left to be done in characterizing these newly discovered factors, understanding how infections differ between healthy and immunocompromised patients, and identifying attractive bacterial or host targets for treating these infections. PMID:27307579

  19. Prevalence of fimA genotypes of Porphyromonas gingivalis in adolescent orthodontic patients.

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    Shuang Pan

    Full Text Available The placement of fixed orthodontic appliances may alter the composition of oral microbiota and has the potential risk of periodontal complication. Porphyromonas gingivalis fimbriae play a critical role in colonization of P. gingivalis in subgingival regions. In this study, we investigated the association between the prevalence of P. gingivalis-specific fimA genotypes and periodontal health status in adolescent orthodontic patients, to identify the pathogencity of P. gingivalis during orthodontic therapy.Sixty-one adolescent orthodontic patients were enrolled in the case group, while the control group consisted of 56 periodontally healthy adolescents. At baseline (T0, clinical parameter (gingival index was tested, and subgingival plaque samples were obtained from the lower incisors. The incidences of P. gingivalis and fimA genotypes were detected by polymerase chain reaction. All parameters were reassessed after 1 month (T1, 2 months (T2, 3 months (T3, and 6 months (T4 in the case group and then compared with those of the controls.Both microbiological and clinical parameters from orthodontic patients started to increase after placement of fixed appliances. Maximum values were reached at 3 months after placement and followed by their decreases at six months. However, the microbiological and clinical parameters in the case group were significantly higher than those of the control group. The GI of fimA II, IV-positive samples was significantly higher than that of negative samples.P. gingivalis carrying fimA II or IV was closely related to orthodontic gingivitis. In addition, proper oral hygiene control could lead to little increase in dental plaque accumulation, and exert a beneficial effect to periodontal tissues.

  20. In vivo quantitative whole-brain diffusion tensor imaging analysis of APP/PS1 transgenic mice using voxel-based and atlas-based methods

    International Nuclear Information System (INIS)

    Qin, Yuan-Yuan; Li, Mu-Wei; Oishi, Kenichi; Zhang, Shun; Zhang, Yan; Zhao, Ling-Yun; Zhu, Wen-Zhen; Lei, Hao

    2013-01-01

    Diffusion tensor imaging (DTI) has been applied to characterize the pathological features of Alzheimer's disease (AD) in a mouse model, although little is known about whether these features are structure specific. Voxel-based analysis (VBA) and atlas-based analysis (ABA) are good complementary tools for whole-brain DTI analysis. The purpose of this study was to identify the spatial localization of disease-related pathology in an AD mouse model. VBA and ABA quantification were used for the whole-brain DTI analysis of nine APP/PS1 mice and wild-type (WT) controls. Multiple scalar measurements, including fractional anisotropy (FA), trace, axial diffusivity (DA), and radial diffusivity (DR), were investigated to capture the various types of pathology. The accuracy of the image transformation applied for VBA and ABA was evaluated by comparing manual and atlas-based structure delineation using kappa statistics. Following the MR examination, the brains of the animals were analyzed for microscopy. Extensive anatomical alterations were identified in APP/PS1 mice, in both the gray matter areas (neocortex, hippocampus, caudate putamen, thalamus, hypothalamus, claustrum, amygdala, and piriform cortex) and the white matter areas (corpus callosum/external capsule, cingulum, septum, internal capsule, fimbria, and optic tract), evidenced by an increase in FA or DA, or both, compared to WT mice (p 0.05). The histopathological changes in the gray matter areas were confirmed by microscopy studies. DTI did, however, demonstrate significant changes in white matter areas, where the difference was not apparent by qualitative observation of a single-slice histological specimen. This study demonstrated the structure-specific nature of pathological changes in APP/PS1 mouse, and also showed the feasibility of applying whole-brain analysis methods to the investigation of an AD mouse model. (orig.)

  1. Systematic revision of the marbled velvet geckos (Oedura marmorata species complex, Diplodactylidae) from the Australian arid and semi-arid zones.

    Science.gov (United States)

    Oliver, Paul M; Doughty, Paul

    2016-03-08

    Lizards restricted to rocky habitats often comprise numerous deeply divergent lineages, reflecting the disjunct nature of their preferred habitat and the capacity of rocky habitats to function as evolutionary refugia. Here we review the systematics and diversity of the predominantly saxicoline Australian marbled velvet geckos (genus Oedura) in the Australian arid and semi-arid zones using newly-gathered morphological data and previously published genetic data. Earlier work showed that four largely allopatric and genetically divergent lineages are present: Western (Pilbara and Gascoyne regions), Gulf (west and south of the Gulf of Carpentaria), Central (central ranges) and Eastern (Cooper and Darling Basins). None of these four populations are conspecific with true O. marmorata, a seperate species complex that is restricted to the Top End region of the Northern Territory. Top End forms share a short, bulbous tail whereas the other four lineages treated here possess a long, tapering tail. Morphological differences among the arid and semi-arid lineages include smaller body size, tapering lamellae and a shorter tail for the Gulf population, and a partially divided rostral scale in the Western population compared to the Central and Eastern populations. Accordingly, we resurrect O. cincta de Vis from synonymy for the Central and Eastern lineages, and regard this species as being comprised of two evolutionary significant units. We also describe the Gulf and Western lineages as new species: Oedura bella sp. nov. and O. fimbria sp. nov., respectively. We note that a predominantly arboreal lineage (the Eastern lineage of O. cincta) is more widely distributed than the other lineages and is phylogenetically nested within a saxicoline clade, but tends to have a deeper head and shorter limbs, consistent with morphological variation observed in other lizard radiations including both saxicoline and arboreal taxa.

  2. Glicosilación intestinal y su relación con la interacción hospedero-patógeno.

    Directory of Open Access Journals (Sweden)

    Gregory Ramón Valdés Paneca

    2015-03-01

    Full Text Available Se evaluó el efecto bloqueador de diferentes lectinas vegetales en la adhesión de E. coli enterotoxigénicas que expresan fimbrias F4. Se realizaron dos experimentos. Primero – las vellosidades fueron incubadas con lectinas de Glycine max, Ulex europaeus, Lens culinaris y Arachis hypogaea a diferentes concentraciones y después fueron incubadas con las cepa de E. coli GIS26 (F4ac. Segundo – veinte y seis lectinas vegetales biotiniladas se enfrentaron a las vellosidades bajo 3 métodos de fijación diferentes. La unión se verificó mediante la técnica de inmunofluorescencia. Para el primer caso hubo bloqueo total de la adhesión de la E. coli GIS26 al borde en cepillo por parte de las lectinas; para el segundo caso se mostró que existe una adhesión moderada y fuerte a las células epiteliales intestinales en el 69,2% (18/26 de las lectinas. Al mismo tiempo se probaron la adhesión de lectinas biotiniladas a diferentes regiones del intestino delgado y la unión de los antígenos fimbriales F4ab, F4ac y F4ad al borde en cepillo. Se encontraron variaciones en el perfil de unión de las lectinas a lo largo de las diferentes regiones del intestino delgado y no homogeneidad en la adhesión de los antígenos fimbriales.

  3. A structural basis for sustained bacterial adhesion: biomechanical properties of CFA/I pili.

    Science.gov (United States)

    Andersson, Magnus; Björnham, Oscar; Svantesson, Mats; Badahdah, Arwa; Uhlin, Bernt Eric; Bullitt, Esther

    2012-02-03

    Enterotoxigenic Escherichia coli (ETEC) are a major cause of diarrheal disease worldwide. Adhesion pili (or fimbriae), such as the CFA/I (colonization factor antigen I) organelles that enable ETEC to attach efficiently to the host intestinal tract epithelium, are critical virulence factors for initiation of infection. We characterized the intrinsic biomechanical properties and kinetics of individual CFA/I pili at the single-organelle level, demonstrating that weak external forces (7.5 pN) are sufficient to unwind the intact helical filament of this prototypical ETEC pilus and that it quickly regains its original structure when the force is removed. While the general relationship between exertion of force and an increase in the filament length for CFA/I pili associated with diarrheal disease is analogous to that of P pili and type 1 pili, associated with urinary tract and other infections, the biomechanical properties of these different pili differ in key quantitative details. Unique features of CFA/I pili, including the significantly lower force required for unwinding, the higher extension speed at which the pili enter a dynamic range of unwinding, and the appearance of sudden force drops during unwinding, can be attributed to morphological features of CFA/I pili including weak layer-to-layer interactions between subunits on adjacent turns of the helix and the approximately horizontal orientation of pilin subunits with respect to the filament axis. Our results indicate that ETEC CFA/I pili are flexible organelles optimized to withstand harsh motion without breaking, resulting in continued attachment to the intestinal epithelium by the pathogenic bacteria that express these pili. Copyright © 2011 Elsevier Ltd. All rights reserved.

  4. Mechanisms of urinary tract sterility maintenance

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    Emilia Okrągła

    2014-06-01

    Full Text Available Physiologically, urine and the urinary tract are maintained sterile because of physical and chemical properties of urine and the innate immune system’s action. The urinary tract is constantly exposed to the invasion of microorganisms from the exterior environment, also because of the anatomical placement of the urethra, in the vicinity of the rectum. Particularly vulnerable to urinary tract infections (UTI are women (an additional risk factor is pregnancy, but also the elderly and children. The main pathogens causing UTI are bacteria; in 70-95% of cases it is the bacterium Escherichia coli. Infections caused by viruses and fungi are less common and are associated with decreased immunity, pharmacotherapy, or some diseases. Bacteria have evolved a number of factors that facilitate the colonization of the urinary tract: the cover and cell membrane antigens O and K1, lipopolysaccharide (LPS, fimbriae, pile and cilia. On the other hand, the human organism has evolved mechanisms to hinder colonization of the urinary tract: mechanisms arising from the anatomical structure of the urinary tract, the physicochemical properties of the urine and the activity of the innate immune system, also known as non-specific, which isolates and destroys pathogens using immunological processes, and the mechanisms for release of antimicrobial substances such as Tamm-Horsfall protein, mucopolysaccharides, immunoglobulins IgA and IgG, lactoferrin, lipocalin, neutrophils, cytokines and antimicrobial peptides. This review aims to analyze the state of knowledge on the mechanisms to maintain the sterility of the urinary tract used by the human organism and bacterial virulence factors to facilitate the colonization of the urinary tract.

  5. Killer whale depredation and associated costs to Alaskan sablefish, Pacific halibut and Greenland turbot longliners.

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    Megan J Peterson

    Full Text Available Killer whale (Orcinus orca depredation (whales stealing or damaging fish caught on fishing gear adversely impacts demersal longline fisheries for sablefish (Anoplopoma fimbria, Pacific halibut (Hippoglossus stenolepis and Greenland turbot (Reinhardtius hippoglossoides in the Bering Sea, Aleutian Islands and Western Gulf of Alaska. These interactions increase direct costs and opportunity costs associated with catching fish and reduce the profitability of longline fishing in western Alaska. This study synthesizes National Marine Fisheries Service observer data, National Marine Fisheries Service sablefish longline survey and fishermen-collected depredation data to: 1 estimate the frequency of killer whale depredation on longline fisheries in Alaska; 2 estimate depredation-related catch per unit effort reductions; and 3 assess direct costs and opportunity costs incurred by longliners in western Alaska as a result of killer whale interactions. The percentage of commercial fishery sets affected by killer whales was highest in the Bering Sea fisheries for: sablefish (21.4%, Greenland turbot (9.9%, and Pacific halibut (6.9%. Average catch per unit effort reductions on depredated sets ranged from 35.1-69.3% for the observed longline fleet in all three management areas from 1998-2012 (p<0.001. To compensate for depredation, fishermen set additional gear to catch the same amount of fish, and this increased fuel costs by an additional 82% per depredated set (average $433 additional fuel per depredated set. In a separate analysis with six longline vessels in 2011 and 2012, killer whale depredation avoidance measures resulted in an average additional cost of $494 per depredated vessel-day for fuel and crew food. Opportunity costs of time lost by fishermen averaged $522 per additional vessel-day on the grounds. This assessment of killer whale depredation costs represents the most extensive economic evaluation of this issue in Alaska to date and will help

  6. Microbial carriage state of peripheral blood dendritic cells (DCs) in chronic periodontitis influences DC differentiation, atherogenic potential.

    Science.gov (United States)

    Carrion, Julio; Scisci, Elizabeth; Miles, Brodie; Sabino, Gregory J; Zeituni, Amir E; Gu, Ying; Bear, Adam; Genco, Caroline A; Brown, David L; Cutler, Christopher W

    2012-09-15

    The low-grade oral infection chronic periodontitis (CP) has been implicated in coronary artery disease risk, but the mechanisms are unclear. In this study, a pathophysiological role for blood dendritic cells (DCs) in systemic dissemination of oral mucosal pathogens to atherosclerotic plaques was investigated in humans. The frequency and microbiome of CD19(-)BDCA-1(+)DC-SIGN(+) blood myeloid DCs (mDCs) were analyzed in CP subjects with or without existing acute coronary syndrome and in healthy controls. FACS analysis revealed a significant increase in blood mDCs in the following order: healthy controls < CP < acute coronary syndrome/CP. Analysis of the blood mDC microbiome by 16S rDNA sequencing showed Porphyromonas gingivalis and other species, including (cultivable) Burkholderia cepacia. The mDC carriage rate with P. gingivalis correlated with oral carriage rate and with serologic exposure to P. gingivalis in CP subjects. Intervention (local debridement) to elicit a bacteremia increased the mDC carriage rate and frequency in vivo. In vitro studies established that P. gingivalis enhanced by 28% the differentiation of monocytes into immature mDCs; moreover, mDCs secreted high levels of matrix metalloproteinase-9 and upregulated C1q, heat shock protein 60, heat shock protein 70, CCR2, and CXCL16 transcripts in response to P. gingivalis in a fimbriae-dependent manner. Moreover, the survival of the anaerobe P. gingivalis under aerobic conditions was enhanced when within mDCs. Immunofluorescence analysis of oral mucosa and atherosclerotic plaques demonstrate infiltration with mDCs, colocalized with P. gingivalis. Our results suggest a role for blood mDCs in harboring and disseminating pathogens from oral mucosa to atherosclerosis plaques, which may provide key signals for mDC differentiation and atherogenic conversion.

  7. Miniature wireless recording and stimulation system for rodent behavioural testing

    Science.gov (United States)

    Pinnell, R. C.; Dempster, J.; Pratt, J.

    2015-12-01

    Objective. Elucidation of neural activity underpinning rodent behaviour has traditionally been hampered by the use of tethered systems and human involvement. Furthermore the combination of deep-brain stimulation (DBS) and various neural recording modalities can lead to complex and time-consuming laboratory setups. For studies of this type, novel tools are required to drive forward this research. Approach. A miniature wireless system weighing 8.5 g (including battery) was developed for rodent use that combined multichannel DBS and local-field potential (LFP) recordings. Its performance was verified in a working memory task that involved 4-channel fronto-hippocampal LFP recording and bilateral constant-current fimbria-fornix DBS. The system was synchronised with video-tracking for extraction of LFP at discrete task phases, and DBS was activated intermittently at discrete phases of the task. Main results. In addition to having a fast set-up time, the system could reliably transmit continuous LFP at over 8 hours across 3-5 m distances. During the working memory task, LFP pertaining to discrete task phases was extracted and compared with well-known neural correlates of active exploratory behaviour in rodents. DBS could be wirelessly activated/deactivated at any part of the experiment during EEG recording and transmission, allowing for a seamless integration of this modality. Significance. The wireless system combines a small size with a level of robustness and versatility that can greatly simplify rodent behavioural experiments involving EEG recording and DBS. Designed for versatility and simplicity, the small size and low-cost of the system and its receiver allow for enhanced portability, fast experimental setup times, and pave the way for integration with more complex behaviour.

  8. Evaluation of antigens stability of tobacco seeds as edible vaccine against VTEC strains

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    Luciana Rossi

    2015-11-01

    Full Text Available Plants have represent a promising alternative for biopharmaceutical proteins (Ma et al., 2003; Rossi et al., 2014. Many plant based edible vaccines have been shown to be effective in inducing local immune responses (Rossi et al., 2013. Edible vaccines can activate both mucosal and systemic immunity, as they come in contact with the digestive tract lining. This dual effect would provide first-line defense against pathogens invading through the mucosa. The antigens are released in the intestines are taken up by M cells that are present over the Payer’s patches (in the ileum and the gut associated lymphoid tissue (GALT. Edible vaccines represent an important worldwide goal for the prevention of the enteric diseases, also in livestock. In particular, the enteric infections are a significant clinical problem in pigs. Verocytotoxic Escherichia (E. coli strains are responsible for serious enterotoxaemia that causes important economic losses in the pig industry. The production of a vaccine for oral administration of transgenic seeds could be a practical and efficient system to prevent the infection and to reduce the antibiotic use. This study was focused on tobacco plants, previously transformed by agroinfection for the seed-specific expression of antigenic proteins (F18 adhesive fimbriae and the B subunit of the Vt2e toxin as model of edible vaccines against verocytotoxic E. coli strains. The dietary administration of transgenic tobacco seeds promotes a significant increase in the number of mucosal IgA-producing cells of the tunica propria in both small and large intestine in mice (Rossi et al., 2013. A protective effect of oral administration of transgenic tobacco seeds was also observed against verocytotoxic Escherichia coli infection in piglets (Rossi et al., 2014. The aim of this study was to assess the seed-expression stability, that is a important requirement in the vaccine production, of F 18 and Vt2e-B heterologous genes into the progeny of

  9. Comparison of hysterosalpingography and laparoscopy in the evaluation of infertile women

    International Nuclear Information System (INIS)

    Sakar, Mehmet N.; Atay, Ahmet E.; Gul, T.; Celik, Y.

    2008-01-01

    Objective was to compare tuboperitoneal factors of infertile women hysterosalpingography (HSG) and laparoscopy. In this cohort study, 82 infertile cases were evaluated retrospectively by laparoscopy, 3 months subsequent to HSG in the Department of Gynecology and Obstetrics, Medical School of Dicle University, Diyarbakir, Turkey between March 2004 and April 2006. The findings of HSG and laparoscopy were compared. Out of 82 infertile women, pathological findings were observed in 45.1% by HSG and 54.9% had no pathological findings. On laparoscopic evaluation, however, pathological findings were observed in 65.85% and 34.15% had no pathological findings. The pathological findings were detected by laparoscopy in 20 of the 45(44.4%) patients who had had no pathological findings by HSG and no pathological findings were detected by laparoscopy in 3 of the 37 (8.1%) patients who had pathological findings by HSG. Laparoscopy revealed no pathological findings in 6 of 35 patients who had tubal pathology by HSG. The sensitivity of HSG was 63%, specificity was 89.3% and positive predictive value was 92% with a 55% predictive value and the accuracy ratio was 72%. Laparoscopy is a superior method for the result of tubal and pelvic pathologies in the evaluation of infertility. However, HSG is a more economical and elementary method suitable for evaluation of endometrial and tubal pathologies and laparoscopy is an appropriate method for examining the external part of tubae, fimbriae, the relation of tuba and ovary, endometriosis, adhesions, tuberculosis and other pathologies. Therefore, these 2 methods are not alternative but complementary. (author)

  10. Bacterial surface appendages strongly impact nanomechanical and electrokinetic properties of Escherichia coli cells subjected to osmotic stress.

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    Grégory Francius

    Full Text Available The physicochemical properties and dynamics of bacterial envelope, play a major role in bacterial activity. In this study, the morphological, nanomechanical and electrohydrodynamic properties of Escherichia coli K-12 mutant cells were thoroughly investigated as a function of bulk medium ionic strength using atomic force microscopy (AFM and electrokinetics (electrophoresis. Bacteria were differing according to genetic alterations controlling the production of different surface appendages (short and rigid Ag43 adhesins, longer and more flexible type 1 fimbriae and F pilus. From the analysis of the spatially resolved force curves, it is shown that cells elasticity and turgor pressure are not only depending on bulk salt concentration but also on the presence/absence and nature of surface appendage. In 1 mM KNO(3, cells without appendages or cells surrounded by Ag43 exhibit large Young moduli and turgor pressures (∼700-900 kPa and ∼100-300 kPa respectively. Under similar ionic strength condition, a dramatic ∼50% to ∼70% decrease of these nanomechanical parameters was evidenced for cells with appendages. Qualitatively, such dependence of nanomechanical behavior on surface organization remains when increasing medium salt content to 100 mM, even though, quantitatively, differences are marked to a much smaller extent. Additionally, for a given surface appendage, the magnitude of the nanomechanical parameters decreases significantly when increasing bulk salt concentration. This effect is ascribed to a bacterial exoosmotic water loss resulting in a combined contraction of bacterial cytoplasm together with an electrostatically-driven shrinkage of the surface appendages. The former process is demonstrated upon AFM analysis, while the latter, inaccessible upon AFM imaging, is inferred from electrophoretic data interpreted according to advanced soft particle electrokinetic theory. Altogether, AFM and electrokinetic results clearly demonstrate the

  11. Primary Papillary Serous Carcinoma of the Fallopian Tube Presenting as a Vaginal Mass: A Case Report and Review of the Literature.

    Science.gov (United States)

    Kadour-Peero, Einav; Sagi-Dain, Lena; Cohen, Gil; Korobochka, Roman; Agbarya, Abed; Bejar, Jacob; Sagi, Shlomi

    2018-05-07

    BACKGROUND There is now evidence to support that some cases of high-grade serous papillary carcinoma arise from the fallopian tubes rather than the ovaries. Common symptoms at presentation include abdominal pain and swelling, vomiting, altered bowel habit and urinary symptoms. To our knowledge, this is the first case of serous papillary carcinoma presenting as a vaginal mass lesion. CASE REPORT A 41-year-old woman was referred to the Bnai-Zion Medical Center with the main complaint of irregular vaginal bleeding, vaginal mucous discharge, and suspected pelvic mass. Physical examination showed a soft, painless mass, measuring about 10 cm in diameter located mainly in the recto-vaginal septum, but not involving the uterus. Ultrasound examination showed no abnormal ovarian or uterine findings. Transvaginal biopsies of the mass showed a poorly differentiated serous papillary carcinoma of ovarian, tubal, or peritoneal origin. The physical examination and imaging findings strongly indicated an inoperable tumor, and the patient was treated with neoadjuvant (pre-surgical) chemotherapy. Pre-operative computed tomography (CT) imaging showed the partial involvement of the colon, and so surgical treatment included total abdominal hysterectomy, bilateral salpingo-oophorectomy, omentectomy, partial vaginectomy, anterior rectal resection, and lymph node dissection. Histopathology of the surgical specimens showed a poorly differentiated serous carcinoma originating from the fimbria of the right fallopian tube. CONCLUSIONS To the best of our knowledge, this is the first report to describe primary fallopian tube papillary serous carcinoma presenting as a vaginal mass. Therefore, physicians should be aware of this possible diagnosis.

  12. Utilización de lectinas en la inhibición de la adhesión de Pasteurella multocida

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    Magda Patricia Carrillo Lamus

    2013-06-01

    Full Text Available El primer paso de la infección es la adhesión de los organismos patógenos a las células blanco. Esta característica les permite no solo penetrar y desplegar las estrategias ofensivas para iniciar la colonización, sino contribuir a la protección y el resguardo de estos a los mecanismos de defensa tanto inmunológicos como mecánicos del hospedero. Este proceso se lleva a cabo en gran medida por la interacción de lectinas, que son proteínas de origen no inmune con la capacidad de reconocer y aglutinar carbohidratos. Este mecanismo ha sido reportado para muchos microorganismos como virus y bacterias. En el caso particular de la Pasteurella multocida, que es una bacteria gramnegativa, patógena oportunista, que inicia la infección en el epitelio respiratorio de muchos animales, se han descrito sobre su superficie sustancias lectinas como la fimbria tipo IV y carbohidratos como el lipopolisacárido o la cápsula que reconocen carbohidratos y lectinas, respectivamente, sobre la superficie de las células epiteliales del tubo respiratorio, circunstancia que le permite adherirse y resguardarse del efecto mucociliar. Debido a que para la gran mayoría de estos microorganismos —incluida P. multocida— es evidente la disminución de la susceptibilidad a los antibacterianos y a la efectividad de las vacunas, se han buscado nuevas tácticas terapéuticas y profilácticas con el fin de interrumpir la infección de los patógenos por medio del bloqueo por competencia de la unión carbohidratos-lectina, donde se ve disminuida la unión del microorganismo al tejido y, por consiguiente, la infección.

  13. Anatomic guidelines defined by reformatting images on MRI for volume measurement of amygdala and hippocampus

    International Nuclear Information System (INIS)

    Hoshida, Tohru; Sakaki, Toshisuke; Uematsu, Sumio.

    1995-01-01

    Twelve patients with intractable partial epilepsy underwent MR scans at the Epilepsy Center of the Johns Hopkins Hospital. There were five women and seven men, ranging in age from five to 51 years (mean age: 26 years). Coronal images were obtained using a 3-D SPGR. The coronal images were transferred to an Allegro 5.1 workstation, and reformatted along the cardinal axes (axial and sagittal) in multiple view points. The anterior end of the amygdala was measured at the level just posterior to the disappearance of the temporal stem. The semilunar gyrus of the amygdala was separated from the ambient gyrus by the semianular sulcus that forms the boundary between the amygdala and the entorhinal cortex. The delineation of the hippocampal formation included the subicular complex, hippocampal proper, dentate gyrus, alveus, and fimbria. The uncal cleft separated the uncus above from the parahippocampal gyrus below. The roof of this cleft was formed by the hippocampus and the dentate gyrus, and the floor, by the presubiculum and subiculum. Although using some guidelines, strictly separating the hippocampal head from the posterior part of the amygdala was not feasible as was previously reported, because of the isointensity on MRI between the cortex of the amygdala and the hippocampus. The most posterior portion of the hippocampus was measured at the level of the subsplenial gyri, just below the splenium of the corpus callosum, to measure the hippocampal volume in its near totality. Therefore, it is reliable, and clinically useful, to measure the combined total volume of the amygdala and the hippocampus when comparing results with those of other centers. (S.Y.)

  14. Bio-mineralisation on the composites of silicon-based polymer and nanodiamond particles by a species of Serratia Bacteria

    International Nuclear Information System (INIS)

    Sammon, R.; Mitev, D.; Pramatarova, L.; Hikov, T.; Radeva, E.; Presker, R.

    2014-01-01

    Serratia sp. NCIMB 40259 is a non-pathogenic Gram-negative bacterium that is able to produce hydroxyapatite by a mechanism involving enzymic cleavage of organic phosphates. Serratia bacteria can attach and form a biofilm on glass, plastics, ceramics and metals and the method can be used to form three dimensional porous scaffolds and for coating 3D structures with hydroxyapatite. The production of calcium phosphate is driven by an acid phosphatase enzyme located in the bacterial cell wall, on fimbriae and within the bacterial extracellular polymeric matrix. Calcium phosphate ceramic may be obtained by two methods: In the first, crystals of calcium phosphate are formed extracellularly within the pre-formed bacterial biofilm grown on the substrata. In the second method, planktonic bacteria catalyse the formation of CaP in suspension and on solid substrata placed in the same container. Composite thin layer of silicon-based polymer and detonated nanodiamond (DND) particles was used as substrate for the process of biomineralization by a species of Serratia. The plasma polymerization (PP) method was chosen to obtain composites of silicon-based polymer, in which DND particles were incorporated. Over the past decades carbon-based nanostructures have been the focus of intense research due to their unique chemical and physical properties. Recently it was shown that the incorporation of the DND particles in a polymer matrix (an organosilicon polymer) changes their physico-chemical properties. The composite films are homogeneous, chemically resistant, thermally and mechanically stable, thus allowing a large amount of biological components to be loaded onto their surface and to be used in tissue engineering, regenerative medicine, implants, stents, biosensors and other medical and biological devices. The aim of this study was to investigate the process of biomineralisation by Serratia bacteria on various composites of silicon-based polymer and detonated nanodiamond particles

  15. Safety and immunogenicity of tetanus-diphtheria-acellular pertussis vaccine administered to children 10 or 11 years of age.

    Science.gov (United States)

    Marshall, Gary S; Pool, Vitali; Greenberg, David P; Johnson, David R; Sheng, Xiaohua; Decker, Michael D

    2014-11-01

    Boosting immunity to tetanus, diphtheria, and pertussis through the use of Tdap vaccines is routinely recommended at 11 to 12 years of age; some states, however, require Tdap for entry into middle school, which may begin at 10 years of age. This study was conducted to determine whether Tdap5 (Adacel), which is licensed for use in children beginning at 11 years of age, is as safe and immunogenic in 10-year-olds as it is in 11-year-olds. Children who had received 5 previous doses of any diphtheria-tetanus-acellular pertussis (DTaP) vaccine were enrolled in a phase IV clinical trial; 646 10-year-olds and 645 11-year-olds completed the study, which involved a single intramuscular dose of Tdap5 along with pre- and postvaccination serologies. Postvaccination geometric mean concentrations (GMCs) of antibody to pertussis antigens (pertussis toxoid, filamentous hemagglutinin, pertactin, and fimbria types 2 and 3) of 10-year-olds were noninferior to those of 11-year-olds, as were booster response rates for all pertussis antibodies, except for those to fimbrial antigens (94% and 97%, respectively). Seroprotection rates among 10-year-olds for tetanus and diphtheria were noninferior to those in 11-year-olds. Rates of injection site reactions, solicited systemic reactions, and unsolicited adverse events, adverse reactions, and serious adverse events were similar in the two groups. These data support the conclusion that Tdap5 is safe and immunogenic in 10-year-olds. (This study has been registered at ClinicalTrials.gov under registration no. NCT01311557.). Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  16. Mosaic organization of the hippocampal neuroepithelium and the multiple germinal sources of dentate granule cells

    International Nuclear Information System (INIS)

    Altman, J.; Bayer, S.A.

    1990-01-01

    This study deals with the site of origin, migration, and settling of the principal cell constituents of the rat hippocampus during the embryonic period. The results indicate that the hippocampal neuroepithelium consists of three morphogenetically discrete components--the Ammonic neuroepithelium, the primary dentate neuroepithelium, and the fimbrial glioepithelium--and that these are discrete sources of the large neurons of Ammon's horn, the smaller granular neurons of the dentate gyrus, and the glial cells of the fimbria. The putative Ammonic neuroepithelium is marked in short-survival thymidine radiograms by a high level of proliferative activity and evidence of interkinetic nuclear migration from day E16 until day E19. On days E16 and E17 a diffuse band of unlabeled cells forms outside the Ammonic neuroepithelium. These postmitotic cells are considered to be stratum radiatum and stratum oriens neurons, which are produced in large numbers as early as day E15. A cell-dense layer, the incipient stratum pyramidale, begins to form on day E18 and spindle-shaped cells can be traced to it from the Ammonic neuroepithelium. This migratory band increases in size for several days, then declines, and finally disappears by day E22. It is inferred that this migration contains the pyramidal cells of Ammon's horn that are produced mostly on days E17 through E20. The putative primary dentate neuroepithelium is distinguished from the Ammonic neuroepithelium during the early phases of embryonic development by its location, shape, and cellular dynamics. It is located around a ventricular indentation, the dentate notch, contains fewer mitotic cells near the lumen of the ventricle than the Ammonic neuroepithelium, and shows a different labeling pattern both in short-survival and sequential-survival thymidine radiograms

  17. A novel growth-promoting microbe, Methylobacterium funariae sp. nov., isolated from the leaf surface of a common moss

    Science.gov (United States)

    Schauer, S

    2011-01-01

    Land plants (embryophytes) evolved in the presence of prokaryotic microbes. As a result, numerous mutually beneficial associations (symbioses) developed that can be analyzed using a variety of methods. Here we describe the isolation and characterization of a new pink-pigmented facultatively methylotrophic symbiotic bacterium of the genus Methylobacterium (laboratory strain F3.2) that was isolated from the gametophytic phylloids of the common cord moss Funaria hygrometrica Hedw. Plantlets were collected in the field and analyzed in the laboratory. Colonies of methylobacteria were obtained by the agar-impression-method. Based on its unique phenotype (the bacterial cells are characterized by fimbriae-like appendages), a comparative 16S rRNA gene (DNA) sequence analysis and an average DNA-DNA hybridization value of 8.4%, compared with its most closely related sister taxon, this isolate is described as a new species, Methylobacterium funariae sp. nov. (type strain F3.2). This new epiphytic bacterium inhabits the leaf surface of “primitive” land plants such as mosses and interacts with its host organism via the secretion of phytohormones (cytokinines, auxins). These external signals are perceived by the plant cells that divide and grow more rapidly than in the absence of their prokaryotic phytosymbionts. We suggest that M. funariae sp. nov. uses methanol emitted from the stomatal pores as principal carbon source for cell metabolism. However, our novel data indicate that, in this unique symbiotic plant-microbe interaction, the uptake of amino acids leached from the surface of the epidermal cells of the green host organism may be of importance as microbial carbon- and nitrogen-source. PMID:21673511

  18. Seasonal monitoring of deep-sea megabenthos in Barkley Canyon cold seep by internet operated vehicle (IOV.

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    Carolina Doya

    Full Text Available Knowledge of the processes shaping deep-sea benthic communities at seasonal scales in cold-seep environments is incomplete. Cold seeps within highly dynamic regions, such as submarine canyons, where variable current regimes may occur, are particularly understudied. Novel Internet Operated Vehicles (IOVs, such as tracked crawlers, provide new techniques for investigating these ecosystems over prolonged periods. In this study a benthic crawler connected to the NEPTUNE cabled infrastructure operated by Ocean Networks Canada was used to monitor community changes across 60 m2 of a cold-seep area of the Barkley Canyon, North East Pacific, at ~890 m depth within an Oxygen Minimum Zone (OMZ. Short video-transects were run at 4-h intervals during the first week of successive calendar months, over a 14 month period (February 14th 2013 to April 14th 2014. Within each recorded transect video megafauna abundances were computed and changes in environmental conditions concurrently measured. The responses of fauna to environmental conditions as a proxy of seasonality were assessed through analysis of abundances in a total of 438 video-transects (over 92 h of total footage. 7698 fauna individuals from 6 phyla (Cnidaria, Ctenophora, Arthropoda, Echinodermata, Mollusca, and Chordata were logged and patterns in abundances of the 7 most abundant taxa (i.e. rockfish Sebastidae, sablefish Anoplopoma fimbria, hagfish Eptatretus stoutii, buccinids (Buccinoidea, undefined small crabs, ctenophores Bolinopsis infundibulum, and Scyphomedusa Poralia rufescens were identified. Patterns in the reproductive behaviour of the grooved tanner crab (Chionnecetes tanneri were also indicated. Temporal variations in biodiversity and abundance in megabenthic fauna was significantly influenced by variabilities in flow velocity flow direction (up or down canyon, dissolved oxygen concentration and month of study. Also reported here for the first time are transient mass aggregations of

  19. Maternal pravastatin prevents altered fetal brain development in a preeclamptic CD-1 mouse model.

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    Alissa R Carver

    Full Text Available Using an animal model, we have previously shown that preeclampsia results in long-term adverse neuromotor outcomes in the offspring, and this phenotype was prevented by antenatal treatment with pravastatin. This study aims to localize the altered neuromotor programming in this animal model and to evaluate the role of pravastatin in its prevention.For the preeclampsia model, pregnant CD-1 mice were randomly allocated to injection of adenovirus carrying sFlt-1 or its control virus carrying mFc into the tail vein. Thereafter they received pravastatin (sFlt-1-pra "experimental group" or water (sFlt-1 "positive control" until weaning. The mFc group ("negative control" received water. Offspring at 6 months of age were sacrificed, and whole brains underwent magnetic resonance imaging (MRI. MRIs were performed using an 11.7 Tesla vertical bore MRI scanner. T2 weighted images were acquired to evaluate the volumes of 28 regions of interest, including areas involved in adaptation and motor, spatial and sensory function. Cytochemistry and cell quantification was performed using neuron-specific Nissl stain. One-way ANOVA with multiple comparison testing was used for statistical analysis.Compared with control offspring, male sFlt-1 offspring have decreased volumes in the fimbria, periaquaductal gray, stria medullaris, and ventricles and increased volumes in the lateral globus pallidus and neocortex; however, female sFlt-1 offspring showed increased volumes in the ventricles, stria medullaris, and fasciculus retroflexus and decreased volumes in the inferior colliculus, thalamus, and lateral globus pallidus. Neuronal quantification via Nissl staining exhibited decreased cell counts in sFlt-1 offspring neocortex, more pronounced in males. Prenatal pravastatin treatment prevented these changes.Preeclampsia alters brain development in sex-specific patterns, and prenatal pravastatin therapy prevents altered neuroanatomic programming in this animal model.

  20. Antimicrobial activity of intraurethrally administered probiotic Lactobacillus casei in a murine model of Escherichia coli urinary tract infection.

    Science.gov (United States)

    Asahara, T; Nomoto, K; Watanuki, M; Yokokura, T

    2001-06-01

    The antimicrobial activity of the intraurethrally administered probiotic Lactobacillus casei strain Shirota against Escherichia coli in a murine urinary tract infection (UTI) model was examined. UTI was induced by intraurethral administration of Escherichia coli strain HU-1 (a clinical isolate from a UTI patient, positive for type 1 and P fimbriae), at a dose of 1 x 10(6) to 2 x 10(6) CFU in 20 microl of saline, into a C3H/HeN mouse bladder which had been traumatized with 0.1 N HCl followed immediately by neutralization with 0.1 N NaOH 24 h before the challenge infection. Chronic infection with the pathogen at 10(6) CFU in the urinary tract (bladder and kidneys) was maintained for more than 3 weeks after the challenge, and the number of polymorphonuclear leukocytes and myeloperoxidase activity in the urine were markedly elevated during the infection period. A single administration of L. casei Shirota at a dose of 10(8) CFU 24 h before the challenge infection dramatically inhibited E. coli growth and inflammatory responses in the urinary tract. Multiple daily treatments with L. casei Shirota during the postinfection period also showed antimicrobial activity in this UTI model. A heat-killed preparation of L. casei Shirota exerted significant antimicrobial effects not only with a single pretreatment (100 microg/mouse) but also with multiple daily treatments during the postinfection period. The other Lactobacillus strains tested, i.e., L. fermentum ATCC 14931(T), L. jensenii ATCC 25258(T), L. plantarum ATCC 14917(T), and L. reuteri JCM 1112(T), had no significant antimicrobial activity. Taken together, these results suggest that the probiotic L. casei strain Shirota is a potent therapeutic agent for UTI.

  1. Structural insight into the role of Streptococcus parasanguinis Fap1 within oral biofilm formation

    Energy Technology Data Exchange (ETDEWEB)

    Garnett, James A.; Simpson, Peter J.; Taylor, Jonathan; Benjamin, Stefi V.; Tagliaferri, Camille; Cota, Ernesto [Department of Biological Sciences, Centre for Structural Biology, Imperial College London, South Kensington, London SW7 2AZ (United Kingdom); Chen, Yi-Ywan M. [Department of Microbiology and Immunology, and Research Center for Pathogenic Bacteria, Chang Gung University, Tao-Yuan, Taiwan (China); Wu, Hui [Department of Pediatric Dentistry, University of Alabama at Birmingham, School of Dentistry, Birmingham, AL 35294 (United States); Matthews, Stephen, E-mail: s.j.matthews@imperial.ac.uk [Department of Biological Sciences, Centre for Structural Biology, Imperial College London, South Kensington, London SW7 2AZ (United Kingdom)

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer Crystal structure of Streptococcus parasanguinis Fap1-NR{sub {alpha}} at pH 5.0. Black-Right-Pointing-Pointer pH-dependent conformational changes mediated through electrostatic potential of Fap1-NR{sub {alpha}}. Black-Right-Pointing-Pointer Fap1 facilitates pH-dependent biofilms. Black-Right-Pointing-Pointer We model inter-Fap1 biofilm interactions. -- Abstract: The fimbriae-associated protein 1 (Fap1) is a major adhesin of Streptococcus parasanguinis, a primary colonizer of the oral cavity that plays an important role in the formation of dental plaque. Fap1 is an extracellular adhesive surface fibre belonging to the serine-rich repeat protein (SRRP) family, which plays a central role in the pathogenesis of streptococci and staphylococci. The N-terminal adhesive region of Fap1 (Fap1-NR) is composed of two domains (Fap1-NR{sub {alpha}} and Fap1-NR{sub {beta}}) and is projected away from the bacterial surface via the extensive serine-rich repeat region, for adhesion to the salivary pellicle. The adhesive properties of Fap1 are modulated through a pH switch in which a reduction in pH results in a rearrangement between the Fap1-NR{sub {alpha}} and Fap1-NR{sub {beta}} domains, which assists in the survival of S. parasanguinis in acidic environments. We have solved the structure of Fap1-NR{sub {alpha}} at pH 5.0 at 3.0 A resolution and reveal how subtle rearrangements of the 3-helix bundle combined with a change in electrostatic potential mediates 'opening' and activation of the adhesive region. Further, we show that pH-dependent changes are critical for biofilm formation and present an atomic model for the inter-Fap1-NR interactions which have been assigned an important role in the biofilm formation.

  2. Structural insight into the role of Streptococcus parasanguinis Fap1 within oral biofilm formation

    International Nuclear Information System (INIS)

    Garnett, James A.; Simpson, Peter J.; Taylor, Jonathan; Benjamin, Stefi V.; Tagliaferri, Camille; Cota, Ernesto; Chen, Yi-Ywan M.; Wu, Hui; Matthews, Stephen

    2012-01-01

    Highlights: ► Crystal structure of Streptococcus parasanguinis Fap1-NR α at pH 5.0. ► pH-dependent conformational changes mediated through electrostatic potential of Fap1-NR α . ► Fap1 facilitates pH-dependent biofilms. ► We model inter-Fap1 biofilm interactions. -- Abstract: The fimbriae-associated protein 1 (Fap1) is a major adhesin of Streptococcus parasanguinis, a primary colonizer of the oral cavity that plays an important role in the formation of dental plaque. Fap1 is an extracellular adhesive surface fibre belonging to the serine-rich repeat protein (SRRP) family, which plays a central role in the pathogenesis of streptococci and staphylococci. The N-terminal adhesive region of Fap1 (Fap1-NR) is composed of two domains (Fap1-NR α and Fap1-NR β ) and is projected away from the bacterial surface via the extensive serine-rich repeat region, for adhesion to the salivary pellicle. The adhesive properties of Fap1 are modulated through a pH switch in which a reduction in pH results in a rearrangement between the Fap1-NR α and Fap1-NR β domains, which assists in the survival of S. parasanguinis in acidic environments. We have solved the structure of Fap1-NR α at pH 5.0 at 3.0 A resolution and reveal how subtle rearrangements of the 3-helix bundle combined with a change in electrostatic potential mediates ‘opening’ and activation of the adhesive region. Further, we show that pH-dependent changes are critical for biofilm formation and present an atomic model for the inter-Fap1-NR interactions which have been assigned an important role in the biofilm formation.

  3. Kittiwake diets and chick production signal a 2008 regime shift in the Northeast Pacific

    Science.gov (United States)

    Hatch, Scott A.

    2013-01-01

    I examined ~2700 food samples collected from adult and nestling black-legged kittiwakes Rissa tridactyla from 1978 through 2011 on Middleton Island in the Gulf of Alaska. The kittiwake diet was composed chiefly of fish, but invertebrates were taken in appreciable quantities in April and May. Upon spring arrival at the colony, adult kittiwakes foraged regularly at night on vertically migrating mesopelagic prey—lanternfishes (Myctophidae), squids, crustaceans, and polychaetes—a behavior they largely discontinued by egg-laying. During incubation and chick-rearing, food samples contained mostly (~85% by weight) Pacific sand lance Ammodytes hexapterus, capelin Mallotus villosus, Pacific herring Clupea pallasii, sablefish Anopoploma fimbria, krill (Euphausiidae), and juvenile salmon Onchorynchus gorboscha and O. keta. A salient finding over the longitudinal study was the emergence, twice, of capelin as a dominant forage species—once in 2000 to 2003, and again in 2008 through 2011. Kittiwakes responded to capelin availability by producing markedly higher numbers of fledged young. The 2000 to 2003 event corresponded to a previously documented shift to cooler conditions in the NE Pacific, which apparently was relatively limited in magnitude or duration. The more recent transition appears stronger and may be more lasting. I submit that 2008 was an important turning point, marking a substantive reversal of warm conditions that began with the well-documented regime shift of 1977. That interpretation is consistent with the existence of a ~60 yr cycle in ocean and atmospheric conditions in the North Pacific. All else being equal, it predicts the next 20 to 30 yr will be favorable for species such as kittiwakes and Steller sea lions, which seemed to respond negatively to the 1977 to 2007 warm phase of the Pacific Decadal Oscillation.

  4. Differential induction and spread of tau pathology in young PS19 tau transgenic mice following intracerebral injections of pathological tau from Alzheimer’s disease or corticobasal degeneration brains

    Science.gov (United States)

    Boluda, Susana; Iba, Michiyo; Zhang, Bin; Raible, Kevin M.; Lee, Virginia M-Y.; Trojanowski, John Q.

    2015-01-01

    Filamentous tau pathologies are hallmark lesions of several neurodegenerative tauopathies including Alzheimer’s disease (AD) and corticobasal degeneration (CBD) which show cell type-specific and topographically distinct tau inclusions. Growing evidence supports templated transmission of tauopathies through functionally interconnected neuroanatomical pathways suggesting that different self-propagating strains of pathological tau could account for the diverse manifestations of neurodegenerative tauopathies. Here, we describe the rapid and distinct cell type-specific spread of pathological tau following intracerebral injections of CBD or AD brain extracts enriched in pathological tau (designated CBD-Tau and AD-Tau, respectively) in young human mutant P301S tau transgenic (Tg) mice (line PS19) ~6–9 months before they show onset of mutant tau transgene-induced tau pathology. At 1 month post-injection of CBD-Tau, tau inclusions developed predominantly in oligodendrocytes of the fimbria and white matter near the injection sites with infrequent intraneuronal tau aggregates. In contrast, injections of AD-Tau in young PS19 mice induced tau pathology predominantly in neuronal perikarya with little or no oligodendrocyte involvement 1 month post-injection. With longer post-injection survival intervals of up to 6 months, CBD-Tau- and AD-Tau-induced tau pathology spread to different brain regions distant from the injection sites while maintaining the cell type-specific pattern noted above. Finally, CA3 neuron loss was detected 3 months post-injection of AD-Tau but not CBD-Tau. Thus, AD-Tau and CBD-Tau represent specific pathological tau strains that spread differentially and may underlie distinct clinical and pathological features of these two tauopathies. Hence, these strains could become targets to develop disease-modifying therapies for CBD and AD. PMID:25534024

  5. Distribution of Porphyromonas gingivalis fimA genotypes in chronic apical periodontitis associated with symptoms.

    Science.gov (United States)

    Wang, Qian; Zhou, Xue-dong; Zheng, Qing-hua; Wang, Yao; Tang, Lu; Huang, Ding-ming

    2010-11-01

    Porphyromonas gingivalis (P. gingivalis) is an anaerobic bacterium involved in root canal infections whose fimbriae are classified into six genotypes (types I-V and Ib) based on nucleotide sequence. Accumulated evidence suggests there is significant association between P. gingivalis and some clinical symptoms of periodontal diseases. The present study aims to determine the prevalence of P. gingivalis fimA genotypes in apical periodontitis and to investigate the correlation between P. gingivalis fimA genotypes and clinical symptoms. Samples were obtained from 158 infected root canals with apical periodontitis. DNA was extracted and analyzed with a polymerase chain reaction-based identification assay. Odds ratios, 95% confidence intervals, and contingency coefficient were calculated for associating the fimA-specific genes with clinical symptoms. P. gingivalis was detected in 39.9% of the inflected root canal samples and was found in 44.5% of P. gingivalis-positive specimens with symptoms. Types II (69.4%) were the most frequent in the symptomatic cases followed by type IV (32.7%). The occurrence of type I (64.3%) was significantly higher than any other genotypes in the asymptomatic apical periodontitis, whereas type II and type Ib were not identified. Statistical analysis revealed that the occurrences of types II, IV, and Ib fimA were associated with greater risk of clinical signs (swelling, sinus tract, or intracanal exudates) than type I. Results from this study reinforce the association between P. gingivalis-specific fimA genotypic clones and apical periodontitis, indicating that fimA genotypes (types II, IV, and Ib) were related to the etiology of symptomatic periradicular diseases. Copyright © 2010 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  6. Administration of probiotics influences F4 (K88-positive enterotoxigenic Escherichia coli attachment and intestinal cytokine expression in weaned pigs

    Directory of Open Access Journals (Sweden)

    Daudelin Jean-François

    2011-05-01

    Full Text Available Abstract This study evaluated the effect of the probiotics Pediococcus acidilactici and Saccharomyces cerevisiae boulardii on the intestinal colonization of O149 enterotoxigenic Escherichia coli harbouring the F4 (K88 fimbriae (ETEC F4 and on the expression of ileal cytokines in weaned pigs. At birth, different litters of pigs were randomly assigned to one of the following treatments: 1 control without antibiotics or probiotics (CTRL; 2 reference group in which chlortetracycline and tiamulin were added to weanling feed (ATB; 3 P. acidilactici; 4 S. cerevisiae boulardii; or 5 P. acidilactici + S. cerevisiae boulardii. Probiotics were administered daily (1 × 109 CFU per pig during the lactation period and after weaning (day 21. At 28 days of age, all pigs were orally challenged with an ETEC F4 strain, and a necropsy was performed 24 h later. Intestinal segments were collected to evaluate bacterial colonization in the small intestine and ileal cytokine expressions. Attachment of ETEC F4 to the intestinal mucosa was significantly reduced in pigs treated with P. acidilactici or S. cerevisiae boulardii in comparison with the ATB group (P = 0.01 and P = 0.03, respectively. In addition, proinflammatory cytokines, such as IL-6, were upregulated in ETEC F4 challenged pigs treated with P. acidilactici alone or in combination with S. cerevisiae boulardii compared with the CTRL group. In conclusion, the administration of P. acidilactici or S. cerevisiae boulardii was effective in reducing ETEC F4 attachment to the ileal mucosa, whereas the presence of P. acidilactici was required to modulate the expression of intestinal inflammatory cytokines in pigs challenged with ETEC F4.

  7. Transcortical selective amygdalohippocampectomy technique through the middle temporal gyrus revisited: An anatomical study laboratory investigation.

    Science.gov (United States)

    Bozkurt, Baran; da Silva Centeno, Ricardo; Chaddad-Neto, Feres; da Costa, Marcos Devanir Silva; Goiri, Marcelo Augusto Acosta; Karadag, Ali; Tugcu, Bekir; Ovalioglu, Talat Cem; Tanriover, Necmettin; Kaya, Serdar; Yagmurlu, Kaan; Grande, Andrew

    2016-12-01

    The anterior temporal lobectomy (ATL) and selective amygdalohippocampectomy (SelAH) have been used for surgical treatment of mesial temporal lobe epilepsy. We examined the comprehensive white matter tract anatomy of the temporal lobe to gain an insight into the trans-middle temporal gyrus, a lateral approach which has been commonly used. The transmiddle temporal gyrus approach was performed in a stepwise manner on cadaveric human heads to examine the traversing white matter pathways through it and the structures located in the temporal horn. We reviewed the literature to compare the trans-middle temporal gyrus approach with other SelAH techniques based on surgical outcomes. There does not appear to be a significant difference in seizure outcome between SelAH and ATL. However, the SelAH provides a better neuropsychological outcomes than the ATL in selected patients. Each SelAH approach has individual advantages and disadvantages. Based on our anatomical study, in the transcortical amygdalohippocampectomy technique through the middle temporal gyrus the white matter pathways to be encountered. In the temporal horn, the collateral eminence, hippocampus, lateral ventricular sulcus, choroidal fissure, inferior choroidal point, choroid plexus, fimbria of the fornix, and amygdala are exposed. The subpial dissection is performed along the lateral ventricular sulcus from the collateral eminence on lateral side and from the choroidal fissure on medial side by microdissector for en bloc resection of the hippocampus proper. The trans-middle temporal gyrus approach is commonly used in treatment of mesial temporal lobe epilepsy patients. A better anatomical and functional understanding of the structures of the temporal lobe is crucial for safer and more accurate surgery. Copyright © 2016 Elsevier Ltd. All rights reserved.

  8. A trial with IgY chicken antibodies to eradicate faecal carriage of Klebsiella pneumoniae and Escherichia coli producing extended-spectrum beta-lactamases

    Science.gov (United States)

    Jonsson, Anna-Karin; Larsson, Anders; Tängdén, Thomas; Melhus, Åsa; Lannergård, Anders

    2015-01-01

    Background Extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae is an emerging therapeutic challenge, especially in the treatment of urinary tract infections. Following an outbreak of CTX-M-15 Klebsiella pneumoniae in Uppsala, Sweden, an orphan drug trial on IgY chicken antibodies was undertaken in an attempt to eradicate faecal carriage of ESBL-producing K. pneumoniae and Escherichia coli. Methods Hens were immunised with epitopes from freeze-dried, whole-cell bacteria (ESBL-producing K. pneumoniae and E. coli) and recombinant proteins of two K. pneumoniae fimbriae subunits (fimH and mrkD). The egg yolks were processed according to good manufacturing practice and the product was stored at−20°C until used. Using an internal database from the outbreak and the regular laboratory database, faecal carriers were identified and recruited from May 2005 to December 2013. The participants were randomised in a placebo-controlled 1:1 manner. Results From 749 eligible patients, 327 (44%) had deceased, and only 91 (12%) were recruited and signed the informed consent. In the initial screening performed using the polymerase chain reaction, 24 participants were ESBL positive and subsequently randomised and treated with either the study drug or a placebo. The study was powered for 124 participants. Because of a very high dropout rate, the study was prematurely terminated. From the outbreak cohort (n=247), only eight patients were screened, and only one was positive with the outbreak strain in faeces. Conclusions The present study design, using IgY chicken antibodies for the eradication of ESBL-producing K. pneumonia and E. coli, was ineffective in reaching its goal due to high mortality and other factors resulting in a low inclusion rate. Spontaneous eradication of ESBL-producing bacteria was frequently observed in recruited participants, which is consistent with previous reports. PMID:26560861

  9. Limiting Size of Fish Fillets at the Center of the Plate Improves the Sustainability of Aquaculture Production

    Directory of Open Access Journals (Sweden)

    Stephen F. Cross

    2011-07-01

    Full Text Available North American dining customers like to have a singular large piece of protein in the center of the plate. When fish is the protein of choice, the portion size from many species is limited by the overall size of the fish. Therefore, for these species, the means to achieve a singular larger portion of “center of the plate” protein is to grow a larger animal. However, fish become less efficient in converting feed to protein as they age. A second option would be to provide two smaller fillets originating from younger, more efficient fish. Here, the sustainability ramifications of these two protein provisioning strategies (single large or two small fillets are considered for three species of fish produced in aquaculture. Growth data for channel catfish (Ictalurus punctatus produced in ponds, rainbow trout (Oncorhynchus mykiss in raceways, and sablefish (Anoplopoma fimbria in marine net pens, were modeled to assess the total biomass and overall food conversion ratio for the production of small, medium or large fish. The production of small fish added an additional 50% or more biomass per year for trout, catfish, and sablefish compared to the production of large fish. Feed conversion ratios were also improved by nearly 10% for the smaller compared to larger fish of each species. Thus, even though all of these species tend to be considered aquaculture species of low environmental impact (and hence “green” or sustainable options, the product form requested by retailers and served by chefs can further increase the sustainability of these species.

  10. The effect of irradiation and cis-diamminedichloroplatinum(II) in the rat brain : analysis of histopathology at 3 and 6 months after treatment

    International Nuclear Information System (INIS)

    Lee, Kyung Ja; Chang, Seung Hee; Koo, Hea Soo

    1998-01-01

    To evalute the late effect(3 and 6 months) of cis-diamminedichloroplatinum(II)(cisplatin) on the radiation brain damage when the cisplatin was intraperitoneally infused immediately after whole brain irradiation in the rats. The histolopathological findings of the brain were examined in rat brains at 3 and 6 months after the treatment. The rats were irradiated(20 or 22.5 Gy, RT) or cisplatin was injected intraperitoneally(2, 4, or 8mg/kg, CT) and in combined treatment group, cisplatin(2mg/kg) was injected immediately after irradiation (20 or 22.5 Gy). Histopathological examination was done mostly in irradiation or cisplatin alone groups, because the rats in combined group died during experimental period except 2 rats. The rats treated with cisplatin showed marked epithelial vacuolation with perivascular edema and vascular dilatation in choroid plexus at 3 months as well as multifocal necrosis involving fimbria and cerebellar hemispheres at 3 and 6 months. The changes were more prominent in rats with 2mg/kg injection compared to rats with 8mg/kg injection. The rats with RT and combined CT and RT showed characteristic delayed irradiation effects such as focal coagulation necrosis and vascular changes, which were more marked than previous reports. Prominent perivascular and leptomeningeal astrocytic proliferation was well documented by anti-GFAP antibody. Cisplatin treatment did not enhance the effect of radiation -induced changes of blood vessels and astrocytic proliferation. The focal necrosis was the most consistently noted finding in this study, it suggested the possibility to use this as an evaluation factor for combined effects of RT and cisplatin

  11. Role of transcription factor Sp1 and RNA binding protein HuR in the downregulation of Dr+ Escherichia coli receptor protein decay accelerating factor (DAF or CD55) by nitric oxide.

    Science.gov (United States)

    Banadakoppa, Manu; Liebenthal, Daniel; Nowak, David E; Urvil, Petri; Yallampalli, Uma; Wilson, Gerald M; Kishor, Aparna; Yallampalli, Chandra

    2013-02-01

    We previously reported that nitric oxide (NO) reduces the rate of bacteremia and maternal mortality in pregnant rats with uterine infection by Escherichia coli expressing the Dr Fimbria (Dr(+) ). The epithelial invasion of Dr(+) E. coli is dependent on the expression level of its cellular receptor decay accelerating factor (DAF). NO reduces the rate of bacteremia by downregulating the expression of DAF. In this study, we elucidated the role of transcription factor Sp1 and RNA binding protein HuR in the downregulation of human DAF by NO. We generated a series of deletion mutant constructs of DAF gene 5'-untranslated region and mapped the NO-response region upstream to the core promoter region of the DAF gene. One of the several Sp1 binding sites in the DAF 5'-untranslated region was located within the NO-response region. The binding of Sp1 to this site was inhibited by NO. Furthermore, NO also promoted the degradation of DAF mRNA. The 3'-untranslated region of DAF harbors an AU-rich element and this element destabilized the mRNA transcript. NO promoted the rapid degradation of DAF mRNA by inhibiting the binding of mRNA stabilizing protein HuR to this AU-rich region. The inhibition of binding of HuR to the AU-rich region was due to the S-nitrosylation of one or more cysteine residues by NO. Thus, these data reveal the molecular mediators of transcriptional and post-transcriptional regulation of DAF by NO with implications in pathophysiology related to DAF. © 2012 The Authors Journal compilation © 2012 FEBS.

  12. The global repressor FliZ antagonizes gene expression by σS-containing RNA polymerase due to overlapping DNA binding specificity.

    Science.gov (United States)

    Pesavento, Christina; Hengge, Regine

    2012-06-01

    FliZ, a global regulatory protein under the control of the flagellar master regulator FlhDC, was shown to antagonize σ(S)-dependent gene expression in Escherichia coli. Thereby it plays a pivotal role in the decision between alternative life-styles, i.e. FlhDC-controlled flagellum-based motility or σ(S)-dependent curli fimbriae-mediated adhesion and biofilm formation. Here, we show that FliZ is an abundant DNA-binding protein that inhibits gene expression mediated by σ(S) by recognizing operator sequences that resemble the -10 region of σ(S)-dependent promoters. FliZ does so with a structural element that is similar to region 3.0 of σ(S). Within this element, R108 in FliZ corresponds to K173 in σ(S), which contacts a conserved cytosine at the -13 promoter position that is specific for σ(S)-dependent promoters. R108 as well as C(-13) are also crucial for DNA binding by FliZ. However, while a number of FliZ binding sites correspond to known σ(S)-dependent promoters, promoter activity is not a prerequisite for FliZ binding and repressor function. Thus, we demonstrate that FliZ also feedback-controls flagellar gene expression by binding to a site in the flhDC control region that shows similarity only to a -10 element of a σ(S)-dependent promoter, but does not function as a promoter.

  13. Inverse regulatory coordination of motility and curli-mediated adhesion in Escherichia coli.

    Science.gov (United States)

    Pesavento, Christina; Becker, Gisela; Sommerfeldt, Nicole; Possling, Alexandra; Tschowri, Natalia; Mehlis, Anika; Hengge, Regine

    2008-09-01

    During the transition from post-exponential to stationary phase, Escherichia coli changes from the motile-planktonic to the adhesive-sedentary "lifestyle." We demonstrate this transition to be controlled by mutual inhibition of the FlhDC/motility and sigma(S)/adhesion control cascades at two distinct hierarchical levels. At the top level, motility gene expression and the general stress response are inversely coordinated by sigma(70)/sigma(FliA)/sigma(S) competition for core RNA polymerase and the FlhDC-controlled FliZ protein acting as a sigma(S) inhibitor. At a lower level, the signaling molecule bis-(3'-5')-cyclic-diguanosine monophosphate (c-di-GMP) reduces flagellar activity and stimulates transcription of csgD, which encodes an essential activator of adhesive curli fimbriae expression. This c-di-GMP is antagonistically controlled by sigma(S)-regulated GGDEF proteins (mainly YegE) and YhjH, an EAL protein and c-di-GMP phosphodiesterase under FlhDC/FliA control. The switch from motility-based foraging to the general stress response and curli expression requires sigma(S)-modulated down-regulation of expression of the flagellar regulatory cascade as well as proteolysis of the flagellar master regulator FlhDC. Control of YhjH by FlhDC and of YegE by sigma(S) produces a fine-tuned checkpoint system that "unlocks" curli expression only after down-regulation of flagellar gene expression. In summary, these data reveal the logic and sequence of molecular events underlying the motile-to-adhesive "lifestyle" switch in E. coli.

  14. Adhesive properties of Enterobacter sakazakii to human epithelial and brain microvascular endothelial cells

    Directory of Open Access Journals (Sweden)

    Pospischil Andreas

    2006-06-01

    Full Text Available Abstract Background Enterobacter sakazakii is an opportunistic pathogen that has been associated with sporadic cases and outbreaks causing meningitis, necrotizing enterocolitis and sepsis especially in neonates. However, up to now little is known about the mechanisms of pathogenicity in E. sakazakii. A necessary state in the successful colonization, establishment and ultimately production of disease by microbial pathogens is the ability to adhere to host surfaces such as mucous membranes, gastric and intestinal epithelial or endothelial tissue. This study examined for the first time the adherence ability of 50 E. sakazakii strains to the two epithelial cell lines HEp-2 and Caco-2, as well as the brain microvascular endothelial cell line HBMEC. Furthermore, the effects of bacterial culture conditions on the adherence behaviour were investigated. An attempt was made to characterize the factors involved in adherence. Results Two distinctive adherence patterns, a diffuse adhesion and the formation of localized clusters of bacteria on the cell surface could be distinguished on all three cell lines. In some strains, a mixture of both patterns was observed. Adherence was maximal during late exponential phase, and increased with higher MOI. The adhesion capacity of E. sakazakii to HBMEC cells was affected by the addition of blood to the bacteria growth medium. Mannose, hemagglutination, trypsin digestion experiments and transmission electron microscopy suggested that the adhesion of E. sakazakii to the epithelial and endothelial cells is mainly non-fimbrial based. Conclusion Adherence experiments show heterogeneity within different E. sakazakii strains. In agreement with studies on E. cloacae, we found no relationship between the adhesive capacities in E. sakazakii and the eventual production of specific fimbriae. Further studies will have to be carried out in order to determine the adhesin(s involved in the interaction of E. sakazakii with cells and to

  15. Early life history pelagic exposure profiles of selected commercially important fish species in the Gulf of Alaska

    Science.gov (United States)

    Doyle, Miriam J.; Mier, Kathryn L.

    2016-10-01

    A synthesis of nearly four decades of ichthyoplankton survey data from the Gulf of Alaska was undertaken to provide the most comprehensive information available on the early life history ecology of five focal species: Pacific Cod (Gadus macrocephalus), Walleye Pollock (Gadus chalcogrammus), Pacific Ocean Perch (Sebastes alutus), Sablefish (Anoplopoma fimbria), and Arrowtooth Flounder (Atheresthes stomias). This analysis of historical data, along with information from published studies, is presented here in the form of ecological reviews of the species during their planktonic phase. The reviews include descriptions of temporal and spatial patterns of exposure to the environment, and interpretation regarding associated sensitivities to environmental forcing. On a temporal scale, patterns in abundance of eggs and larvae are synthesized that characterize seasonal exposure to the pelagic environment, and interannual variation that is presumed to incorporate responses to long-term environmental forcing. Spatial patterns are synthesized to identify horizontal and vertical extent of egg and larval distributions, delineate areas of primary larval habitat, and illuminate egg and larval drift pathways. The observed patterns are discussed with respect to characterizing species early life history strategies, identifying long-term adaptations to the Gulf of Alaska environment, and associated resilience and vulnerability factors that may modulate early life responses to environmental forcing in this region. For each species, gaps in knowledge are identified and are concerned primarily with the period of transition between the larval and juvenile stage, and feeding habits and ecology across seasons, habitats and sub-intervals of early ontogeny. These early life history reviews advance our ecological understanding of the pelagic phase, and fine-tune our focus for the investigation of potential response mechanisms to environmental forcing at appropriate, species-specific temporal

  16. Conditional inactivation of p53 in mouse ovarian surface epithelium does not alter MIS driven Smad2-dominant negative epithelium-lined inclusion cysts or teratomas.

    Directory of Open Access Journals (Sweden)

    Suzanne M Quartuccio

    Full Text Available Epithelial ovarian cancer is the most lethal gynecological malignancy among US women. The etiology of this disease, although poorly understood, may involve the ovarian surface epithelium or the epithelium of the fallopian tube fimbriae as the progenitor cell. Disruptions in the transforming growth factor beta (TGFβ pathway and p53 are frequently found in chemotherapy-resistant serous ovarian tumors. Transgenic mice expressing a dominant negative form of Smad2 (Smad2DN, a downstream transcription factor of the TGFβ signaling pathway, targeted to tissues of the reproductive tract were created on a FVB background. These mice developed epithelium-lined inclusion cysts, a potential precursor lesion to ovarian cancer, which morphologically resembled oviductal epithelium but exhibited protein expression more closely resembling the ovarian surface epithelium. An additional genetic "hit" of p53 deletion was predicted to result in ovarian tumors. Tissue specific deletion of p53 in the ovaries and oviducts alone was attempted through intrabursal or intraoviductal injection of Cre-recombinase expressing adenovirus (AdCreGFP into p53 (flox/flox mice. Ovarian bursal cysts were detected in some mice 6 months after intrabursal injection. No pathological abnormalities were detected in mice with intraoviductal injections, which may be related to decreased infectivity of the oviductal epithelium with adenovirus as compared to the ovarian surface epithelium. Bitransgenic mice, expressing both the Smad2DN transgene and p53 (flox/flox, were then exposed to AdCreGFP in the bursa and oviductal lumen. These mice did not develop any additional phenotypes. Exposure to AdCreGFP is not an effective methodology for conditional deletion of floxed genes in oviductal epithelium and tissue specific promoters should be employed in future mouse models of the disease. In addition, a novel phenotype was observed in mice with high expression of the Smad2DN transgene as validated

  17. Hippocampal volume reduction in congenital central hypoventilation syndrome.

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    Paul M Macey

    Full Text Available Children with congenital central hypoventilation syndrome (CCHS, a genetic disorder characterized by diminished drive to breathe during sleep and impaired CO(2 sensitivity, show brain structural and functional changes on magnetic resonance imaging (MRI scans, with impaired responses in specific hippocampal regions, suggesting localized injury.We assessed total volume and regional variation in hippocampal surface morphology to identify areas affected in the syndrome. We studied 18 CCHS (mean age+/-std: 15.1+/-2.2 years; 8 female and 32 healthy control (age 15.2+/-2.4 years; 14 female children, and traced hippocampi on 1 mm(3 resolution T1-weighted scans, collected with a 3.0 Tesla MRI scanner. Regional hippocampal volume variations, adjusted for cranial volume, were compared between groups based on t-tests of surface distances to the structure midline, with correction for multiple comparisons. Significant tissue losses emerged in CCHS patients on the left side, with a trend for loss on the right; however, most areas affected on the left also showed equivalent right-sided volume reductions. Reduced regional volumes appeared in the left rostral hippocampus, bilateral areas in mid and mid-to-caudal regions, and a dorsal-caudal region, adjacent to the fimbria.The volume losses may result from hypoxic exposure following hypoventilation during sleep-disordered breathing, or from developmental or vascular consequences of genetic mutations in the syndrome. The sites of change overlap regions of abnormal functional responses to respiratory and autonomic challenges. Affected hippocampal areas have roles associated with memory, mood, and indirectly, autonomic regulation; impairments in these behavioral and physiological functions appear in CCHS.

  18. Gene-based vaccine development for improving animal production in developing countries. Possibilities and constraints

    International Nuclear Information System (INIS)

    Egerton, J.R.

    2005-01-01

    For vaccine production, recombinant antigens must be protective. Identifying protective antigens or candidate antigens is an essential precursor to vaccine development. Even when a protective antigen has been identified, cloning of its gene does not lead directly to vaccine development. The fimbrial protein of Dichelobacter nodosus, the agent of foot-rot in ruminants, was known to be protective. Recombinant vaccines against this infection are ineffective if expressed protein subunits are not assembled as mature fimbriae. Antigenic competition between different, but closely related, recombinant antigens limited the use of multivalent vaccines based on this technology. Recombinant antigens may need adjuvants to enhance response. DNA vaccines, potentiated with genes for different cytokines, may replace the need for aggressive adjuvants, and especially where cellular immunity is essential for protection. The expression of antigens from animal pathogens in plants and the demonstration of some immunity to a disease like rinderpest after ingestion of these, suggests an alternative approach to vaccination by injection. Research on disease pathogenesis and the identification of candidate antigens is specific to the disease agent. The definition of expression systems and the formulation of a vaccine for each disease must be followed by research to establish safety and efficacy. Where vaccines are based on unique gene sequences, the intellectual property is likely to be protected by patent. Organizations, licensed to produce recombinant vaccines, expect to recover their costs and to make a profit. The consequence is that genetically-derived vaccines are expensive. The capacity of vaccines to help animal owners of poorer countries depends not only on quality and cost but also on the veterinary infrastructure where they are used. Ensuring the existence of an effective animal health infrastructure in developing countries is as great a challenge for the developed world as

  19. Genetic basis of coaggregation receptor polysaccharide biosynthesis in Streptococcus sanguinis and related species.

    Science.gov (United States)

    Yang, J; Yoshida, Y; Cisar, J O

    2014-02-01

    Interbacterial adhesion between streptococci and actinomyces promotes early dental plaque biofilm development. Recognition of coaggregation receptor polysaccharides (RPS) on strains of Streptococcus sanguinis, Streptococcus gordonii and Streptococcus oralis by Actinomyces spp. type 2 fimbriae is the principal mechanism of these interactions. Previous studies of genetic loci for synthesis of RPS (rps) and RPS precursors (rml, galE1 and galE2) in S. gordonii 38 and S. oralis 34 revealed differences between these strains. To determine whether these differences are strain-specific or species-specific, we identified and compared loci for polysaccharide biosynthesis in additional strains of these species and in several strains of the previously unstudied species, S. sanguinis. Genes for synthesis of RPS precursors distinguished the rps loci of different streptococci. Hence, rml genes for synthesis of TDP-L-Rha were in rps loci of S. oralis strains but at other loci in S. gordonii and S. sanguinis. Genes for two distinct galactose epimerases were also distributed differently. Hence, galE1 for epimerization of UDP-Glc and UDP-Gal was in galactose operons of S. gordonii and S. sanguinis strains but surprisingly, this gene was not present in S. oralis. Moreover, galE2 for epimerization of both UDP-Glc and UDP-Gal and UDP-GlcNAc and UDP-GalNAc was at a different locus in each species, including rps operons of S. sanguinis. The findings provide insight into cell surface properties that distinguish different RPS-producing streptococci and open an approach for identifying these bacteria based on the arrangement of genes for synthesis of polysaccharide precursors. Published 2013. This article is a U.S. Government work and is in the public domain in the USA.

  20. Mutations reducing replication from R-loops suppress the defects of growth, chromosome segregation and DNA supercoiling in cells lacking topoisomerase I and RNase HI activity.

    Science.gov (United States)

    Usongo, Valentine; Martel, Makisha; Balleydier, Aurélien; Drolet, Marc

    2016-04-01

    R-loop formation occurs when the nascent RNA hybridizes with the template DNA strand behind the RNA polymerase. R-loops affect a wide range of cellular processes and their use as origins of replication was the first function attributed to them. In Escherichia coli, R-loop formation is promoted by the ATP-dependent negative supercoiling activity of gyrase (gyrA and gyrB) and is inhibited by topoisomerase (topo) I (topA) relaxing transcription-induced negative supercoiling. RNase HI (rnhA) degrades the RNA moiety of R-loops. The depletion of RNase HI activity in topA null mutants was previously shown to lead to extensive DNA relaxation, due to DNA gyrase inhibition, and to severe growth and chromosome segregation defects that were partially corrected by overproducing topo III (topB). Here, DNA gyrase assays in crude cell extracts showed that the ATP-dependent activity (supercoiling) of gyrase but not its ATP-independent activity (relaxation) was inhibited in topA null cells lacking RNase HI. To characterize the cellular event(s) triggered by the absence of RNase HI, we performed a genetic screen for suppressors of the growth defect of topA rnhA null cells. Suppressors affecting genes in replication (holC2::aph and dnaT18::aph) nucleotide metabolism (dcd49::aph), RNA degradation (rne59::aph) and fimbriae synthesis (fimD22::aph) were found to reduce replication from R-loops and to restore supercoiling, thus pointing to a correlation between R-loop-dependent replication in topA rnhA mutants and the inhibition of gyrase activity and growth. Interestingly, the position of fimD on the E. coli chromosome corresponds to the site of one of the five main putative origins of replication from R-loops in rnhA null cells recently identified by next-generation sequencing, thus suggesting that the fimD22::aph mutation inactivated one of these origins. Furthermore, we show that topo III overproduction is unable to complement the growth defect of topA rnhA null mutants at low

  1. Efficacy of a single oral dose of a live bivalent E. coli vaccine against post-weaning diarrhea due to F4 and F18-positive enterotoxigenic E. coli.

    Science.gov (United States)

    Nadeau, É; Fairbrother, J M; Zentek, J; Bélanger, L; Tremblay, D; Tremblay, C-L; Röhe, I; Vahjen, W; Brunelle, M; Hellmann, K; Cvejić, D; Brunner, B; Schneider, C; Bauer, K; Wolf, R; Hidalgo, Á

    2017-08-01

    F4- and F18-positive enterotoxigenic E. coli strains (F4-ETEC and F18-ETEC) are important causes of post-weaning diarrhea (PWD) in pigs. F4 (antigenic variant ac) and F18 (ab and ac) fimbriae are major antigens that play an important role in the early stages of infection. Herein, the efficacy of a live oral vaccine consisting of two non-pathogenic E. coli strains, one F4ac- and one F18ac-positive, was evaluated using F4ac-ETEC and F18ab-ETEC challenge models. A randomized, masked, placebo-controlled, block design, parallel-group confirmatory study with two different vaccination-challenge intervals (7 and 21 days) was conducted for each challenge model. The vaccine was administered in one dose, to ≥18-day-old piglets via drinking water. Efficacy was assessed by evaluating diarrhea, clinical observations, weight gain and fecal shedding of F4-ETEC or F18-ETEC. Anti-F4 and anti-F18 immunoglobulins in blood were measured. The vaccination resulted in significant reductions in clinical PWD and fecal shedding of F4-ETEC and F18-ETEC after the 7- and 21-day-post-vaccination heterologous challenges, except for after the 21-day-post-vaccination F4-ETEC challenge, when the clinical PWD was too mild to demonstrate efficacy. A significant reduction of mortality and weight loss by vaccination were observed following the F18-ETEC challenge. The 7-day protection was associated with induction of anti-F4 and anti-F18 IgM, whereas the 21-day protection was mainly associated with anti-F4 and anti-F18 IgA. The 7-day onset and 21-day duration of protection induced by this vaccine administered once in drinking water to pigs of at least 18days of age were confirmed by protection against F4-ETEC and F18-ETEC, and induction of F4 and F18-specific immunity. Cross protection of the vaccine against F18ab-E. coli was demonstrated for both the 7- and 21-day F18-ETEC challenges. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  2. Prevalence of adhesin and toxin genes in E. coli strains isolated from diarrheic and non-diarrheic pigs from smallholder herds in northern and eastern Uganda.

    Science.gov (United States)

    Ikwap, Kokas; Larsson, Jenny; Jacobson, Magdalena; Owiny, David Okello; Nasinyama, George William; Nabukenya, Immaculate; Mattsson, Sigbrit; Aspan, Anna; Erume, Joseph

    2016-08-05

    Enterotoxigenic E. coli (ETEC) significantly contribute to diarrhea in piglets and weaners. The smallholder pig producers in Uganda identified diarrhea as one of the major problems especially in piglets. The aim of this study was to; i) characterize the virulence factors of E. coli strains isolated from diarrheic and non-diarrheic suckling piglets and weaners from smallholder herds in northern and eastern Uganda and ii) identify and describe the post-mortem picture of ETEC infection in severely diarrheic piglets. Rectal swab samples were collected from 83 piglets and weaners in 20 herds and isolated E. coli were characterized by PCR, serotyping and hemolysis. The E. coli strains carried genes for the heat stable toxins STa, STb and EAST1 and adhesins F4 and AIDA-I. The genes for the heat labile toxin LT and adhesins F5, F6, F18 and F41 were not detected in any of the E. coli isolates. Where the serogroup could be identified, E. coli isolates from the same diarrheic pig belonged to the same serogroup. The prevalence of EAST1, STb, Stx2e, STa, AIDA-I, and F4 in the E. coli isolates from suckling piglets and weaners (diarrheic and non-diarrheic combined) was 29, 26.5, 2.4, 1.2, 16, and 8.4 %, respectively. However the prevalence of F4 and AIDA-I in E. coli from diarrheic suckling piglets alone was 22.2 and 20 %, respectively. There was no significant difference in the prevalence of the individual virulence factors in E. coli from the diarrheic and non-diarrheic pigs (p > 0.05). The main ETEC strains isolated from diarrheic and non-diarrheic pigs included F4/STb/EAST1 (7.2 %), F4/STb (1.2 %), AIDA/STb/EAST1 (8 %) and AIDA/STb (8 %). At post-mortem, two diarrheic suckling piglets carrying ETEC showed intact intestinal villi, enterocytes and brush border but with a layer of cells attached to the brush border, suggestive of ETEC infections. This study has shown that the F4 fimbriae is the most predominant in E. coli from diarrheic piglets in the study area and

  3. Genotypic and phenotypic characterization of multidrug resistant Salmonella Typhimurium and Salmonella Kentucky strains recovered from chicken carcasses.

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    Rizwana Tasmin

    Full Text Available Salmonella Typhimurium is the leading cause of human non-typhoidal gastroenteritis in the US. S. Kentucky is one the most commonly recovered serovars from commercially processed poultry carcasses. This study compared the genotypic and phenotypic properties of two Salmonella enterica strains Typhimurium (ST221_31B and Kentucky (SK222_32B recovered from commercially processed chicken carcasses using whole genome sequencing, phenotype characterizations and an intracellular killing assay. Illumina MiSeq platform was used for sequencing of two Salmonella genomes. Phylogenetic analysis employing homologous alignment of a 1,185 non-duplicated protein-coding gene in the Salmonella core genome demonstrated fully resolved bifurcating patterns with varying levels of diversity that separated ST221_31B and SK222_32B genomes into distinct monophyletic serovar clades. Single nucleotide polymorphism (SNP analysis identified 2,432 (ST19 SNPs within 13 Typhimurium genomes including ST221_31B representing Sequence Type ST19 and 650 (ST152 SNPs were detected within 13 Kentucky genomes including SK222_32B representing Sequence Type ST152. In addition to serovar-specific conserved coding sequences, the genomes of ST221_31B and SK222_32B harbor several genomic regions with significant genetic differences. These included phage and phage-like elements, carbon utilization or transport operons, fimbriae operons, putative membrane associated protein-encoding genes, antibiotic resistance genes, siderophore operons, and numerous hypothetical protein-encoding genes. Phenotype microarray results demonstrated that ST221_31B is capable of utilizing certain carbon compounds more efficiently as compared to SK222_3B; namely, 1,2-propanediol, M-inositol, L-threonine, α-D-lactose, D-tagatose, adonitol, formic acid, acetoacetic acid, and L-tartaric acid. ST221_31B survived for 48 h in macrophages, while SK222_32B was mostly eliminated. Further, a 3-fold growth of ST221_31B was

  4. Immunogenicity and protective efficacy of a single-dose live non-pathogenic Escherichia coli oral vaccine against F4-positive enterotoxigenic Escherichia coli challenge in pigs.

    Science.gov (United States)

    Fairbrother, John Morris; Nadeau, Éric; Bélanger, Louise; Tremblay, Cindy-Love; Tremblay, Danielle; Brunelle, Mélanie; Wolf, Regina; Hellmann, Klaus; Hidalgo, Álvaro

    2017-01-05

    Enterotoxigenic Escherichia coli strains expressing F4 (K88) fimbriae (F4-ETEC) are one of the most important causes of post-weaning diarrhea (PWD) in pigs. F4, a major antigen, plays an important role in the early steps of the infection. Herein, the efficacy of a live oral vaccine consisting of a non-pathogenic E. coli strain expressing F4 for protection of pigs against PWD was evaluated. Three blinded, placebo-controlled, block design, parallel-group confirmatory experiments were conducted, using an F4-ETEC PWD challenge model, each with a different vaccination-challenge interval (3, 7, and 21days). The pigs were vaccinated via the drinking water with a single dose of the Coliprotec® F4 vaccine one day post-weaning. Efficacy was assessed by evaluating diarrhea, clinical observations, intestinal fluid accumulation, weight gain, intestinal colonization and fecal shedding of F4-ETEC. The immune response was evaluated by measuring serum and intestinal F4-specific antibodies. The administration of the vaccine resulted in a significant reduction of the incidence of moderate to severe diarrhea, ileal colonization by F4-ETEC, and fecal shedding of F4-ETEC after the heterologous challenge at 7 and 21days post-vaccination. The 7-day onset of protection was associated with an increase of serum anti-F4 IgM whereas the 21-day duration of protection was associated with an increase of both serum anti-F4 IgM and IgA. Significant correlations between levels of serum and intestinal secretory anti-F4 antibodies were detected. Maternally derived F4-specific serum antibodies did not interfere with the vaccine efficacy. The evaluation of protection following a challenge three days after vaccination showed a reduction of the severity and the duration of diarrhea and of fecal shedding of F4-ETEC. The 7-day onset and the 21-day duration of protection induced by Coliprotec® F4 vaccine administered once in drinking water to pigs of at least 18days of age were confirmed by protection

  5. Whole genome sequencing for the molecular characterization of carbapenem-resistant Klebsiella pneumoniae strains isolated at the Italian ASST Fatebenefratelli Sacco Hospital, 2012-2014.

    Science.gov (United States)

    Rimoldi, Sara Giordana; Gentile, Bernardina; Pagani, Cristina; Di Gregorio, Annamaria; Anselmo, Anna; Palozzi, Anna Maria; Fortunato, Antonella; Pittiglio, Valentina; Ridolfo, Anna Lisa; Gismondo, Maria Rita; Rizzardini, Giuliano; Lista, Florigio

    2017-10-10

    The emergence of carbapenem-resistant Klebsiella pneumoniae strains is threatening antimicrobial treatment. Sixty-eight carbapenemase-producing K. pneumoniae strains isolated at Luigi Sacco University Hospital-ASST Fatebenefratelli Sacco (Milan, Italy) between 2012 and 2014 were characterised microbiologically and molecularly. They were tested for drug susceptibility and carbapenemase phenotypes, investigated by means of repetitive extra-genic palindromic polymerase chain reaction (REP-PCR), and fully sequenced by means of next-generation sequencing for the in silico analysis of multi-locus sequence typing (MLST), their resistome, virulome and plasmid content, and their core single nucleotide polymorphism (SNP) genotypes. All of the samples were resistant to carbapenems, other β-lactams and ciprofloxacin; many were resistant to aminoglycosides and tigecycline; and seven were resistant to colistin. Resistome analysis revealed the presence of blaKPC genes and, less frequently blaSHV, blaTEM, blaCTX-M and blaOXA, which are related to resistance to carbapenem and other β-lactams. Other genes conferring resistance to aminoglycoside, fluoroquinolone, phenicol, sulphonamide, tetracycline, trimethoprim and macrolide-lincosamide-streptogramin were also detected. Genes related to AcrAB-TolC efflux pump-dependent and pump-independent tigecycline resistance mechanisms were investigated, but it was not possible to clearly correlate the genomic features with tigecycline resistance because of the presence of a common mutation in susceptible, intermediate and resistant strains. Concerning colistin resistance, the mgrB gene was disrupted by an IS5-like element, and the mobile mcr-1 and mcr-2 genes were not detected in two cases. The virulome profile revealed type-3 fimbriae and iron uptake system genes, which are important during the colonisation stage in the mammalian host environment. The in silico detected plasmid replicons were classified as IncFIB(pQil), IncFIB(K), Col

  6. Elucidating dynamic responses of North Pacific fish populations to climatic forcing: Influence of life-history strategy

    Science.gov (United States)

    Yatsu, A.; Aydin, K. Y.; King, J. R.; McFarlane, G. A.; Chiba, S.; Tadokoro, K.; Kaeriyama, M.; Watanabe, Y.

    2008-05-01

    In order to explore mechanistic linkages between low-frequency ocean/climate variability, and fish population responses, we undertook comparative studies of time-series of recruitment-related productivity and the biomass levels of fish stocks representing five life-history strategies in the northern North Pacific between the 1950s and the present. We selected seven species: Japanese sardine ( Sardinopus melanostictus) and California sardine ( Sardinopus sagax) (opportunistic strategists), walleye pollock ( Theragra chalcogramma, intermediate strategist), pink salmon ( Oncorhynchus gorbuscha, salmonic strategist), sablefish ( Anoplopoma fimbria) and Pacific halibut ( Hippoglossus stenolepis) (periodic strategists) and spiny dogfish ( Squalus acanthias, equilibrium strategist). The responses in terms of productivity of sardine, pink salmon, sablefish and halibut to climatic regime shifts were generally immediate, delayed, or no substantial responses depending on the particular regime shift year and fish stock (population). In walleye pollock, there were some periods of high productivity and low productivity, but not coincidental to climatic regime shifts, likely due to indirect climate forcing impacts on both bottom-up and top-down processes. Biomass of zooplankton and all fish stocks examined, except for spiny dogfish whose data were limited, indicated a decadal pattern with the most gradual changes in periodic strategists and most intensive and rapid changes in opportunistic strategists. Responses of sardine productivity to regime shifts were the most intense, probably due to the absence of density-dependent effects and the availability of refuges from predators when sardine biomass was extremely low. Spiny dogfish were least affected by environmental variability. Conversely, spiny dogfish are likely to withstand only modest harvest rates due to their very low intrinsic rate of increase. Thus, each life-history strategy type had a unique response to climatic

  7. Enterotoxigenic Escherichia coli Subclinical Infection in Pigs: Bacteriological and Genotypic Characterization and Antimicrobial Resistance Profiles.

    Science.gov (United States)

    Moredo, Fabiana A; Piñeyro, Pablo E; Márquez, Gabriela C; Sanz, Marcelo; Colello, Rocío; Etcheverría, Analía; Padola, Nora L; Quiroga, María A; Perfumo, Carlos J; Galli, Lucía; Leotta, Gerardo A

    2015-08-01

    Enterotoxigenic Escherichia coli (ETEC) is the major pathogen responsible for neonatal diarrhea, postweaning diarrhea, and edema disease in pigs. Although it can be harmless, ETEC is also present in the intestines of other animal species and humans, causing occasional diarrhea outbreaks. The evaluation of this pathogen's presence in food sources is becoming an increasingly important issue in human health. In order to determine the prevalence of ETEC in nondiarrheic pigs, 990 animals from 11 pig farms were sampled. Using end-time polymerase chain reaction (PCR), eltA, estI genes, or both, were detected in 150 (15.2%) animals. From the positive samples, 40 (26.6%) ETEC strains were isolated, showing 19 antibiotic-resistance patterns; 52.5% of these strains had multiple antibiotic resistances, and 17.5% carried the intI2 gene. The most prevalent genotypes were rfb(O157)/estII/aidA (32.5%) and estI/estII (25.0%). The estII gene was identified most frequently (97.5%), followed by estI (37.5%), astA (20.0%), and eltA (12.5%). The genes coding the fimbriae F5, F6, and F18 were detected in three single isolates. The aidA gene was detected in 20 ETEC strains associated with the estII gene. Among the isolated ETEC strains, stx(2e)/estI, stx(2e)/estI/estII, and stx(2e)/estI/estII/intI2 genotypes were identified. The ETEC belonged to 12 different serogroups; 37.5% of them belonged to serotype O157:H19. Isolates were grouped by enterobacterial repetitive intergenic consensus-PCR into 5 clusters with 100.0% similarity. In this study, we demonstrated that numerous ETEC genotypes cohabit and circulate in swine populations without clinical manifestation of neonatal diarrhea, postweaning diarrhea, or edema disease in different production stages. The information generated is important not only for diagnostic and epidemiological purposes, but also for understanding the dynamics and ecology of ETEC in pigs in different production stages that can be potentially transmitted to humans

  8. Virulence characteristics and genetic affinities of multiple drug resistant uropathogenic Escherichia coli from a semi urban locality in India.

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    Savita Jadhav

    Full Text Available Extraintestinal pathogenic Escherichia coli (ExPEC are of significant health concern. The emergence of drug resistant E. coli with high virulence potential is alarming. Lack of sufficient data on transmission dynamics, virulence spectrum and antimicrobial resistance of certain pathogens such as the uropathogenic E. coli (UPEC from countries with high infection burden, such as India, hinders the infection control and management efforts. In this study, we extensively genotyped and phenotyped a collection of 150 UPEC obtained from patients belonging to a semi-urban, industrialized setting near Pune, India. The isolates representing different clinical categories were analyzed in comparison with 50 commensal E. coli isolates from India as well as 50 ExPEC strains from Germany. Virulent strains were identified based on hemolysis, haemagglutination, cell surface hydrophobicity, serum bactericidal activity as well as with the help of O serotyping. We generated antimicrobial resistance profiles for all the clinical isolates and carried out phylogenetic analysis based on repetitive extragenic palindromic (rep-PCR. E. coli from urinary tract infection cases expressed higher percentages of type I (45% and P fimbriae (40% when compared to fecal isolates (25% and 8% respectively. Hemolytic group comprised of 60% of UPEC and only 2% of E. coli from feces. Additionally, we found that serum resistance and cell surface hydrophobicity were not significantly (p = 0.16/p = 0.51 associated with UPEC from clinical cases. Moreover, clinical isolates exhibited highest resistance against amoxicillin (67.3% and least against nitrofurantoin (57.3%. We also observed that 31.3% of UPEC were extended-spectrum beta-lactamase (ESBL producers belonging to serotype O25, of which four were also positive for O25b subgroup that is linked to B2-O25b-ST131-CTX-M-15 virulent/multiresistant type. Furthermore, isolates from India and Germany (as well as global sources were found to be

  9. Phagocytosis and Epithelial Cell Invasion by Crohn’s Disease-Associated Adherent-Invasive Escherichia coli Are Inhibited by the Anti-inflammatory Drug 6-Mercaptopurine

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    Federica Migliore

    2018-05-01

    Full Text Available Adherent-invasive Escherichia coli (AIEC strains are overrepresented in the dysbiotic microbiota of Crohn’s disease (CD patients, and contribute to the onset of the chronic inflammation typical of the disease. However, the effects of anti-inflammatory drugs used for CD treatment on AIEC virulence have not yet been investigated. In this report, we show that exposure of AIEC LF82 strain to amino-6-mercaptopurine (6-MP riboside, one of the most widely used anti-inflammatory drugs in CD, impairs its ability to adhere to, and consequently to invade, human epithelial cells. Notably, phagocytosis of LF82 treated with 6-MP by human macrophages is also reduced, suggesting that 6-MP affects AIEC cell surface determinants involved both in interaction with epithelial cells and in uptake by macrophages. Since a main target of 6-MP in bacterial cells is the inhibition of the important signal molecule c-di-GMP, we also tested whether perturbations in cAMP, another major signaling pathway in E. coli, might have similar effects on interactions with human cells. To this aim, we grew LF82 in the presence of glucose, which leads to inhibition of cAMP synthesis. Growth in glucose-supplemented medium resulted in a reduction in AIEC adhesion to epithelial cells and uptake by macrophages. Consistent with these results, both 6-MP and glucose can affect expression of cell adhesion-related genes, such as the csg genes, encoding thin aggregative fimbriae (curli. In addition, glucose strongly inhibits expression of the fim operon, encoding type 1 pili, a known AIEC determinant for adhesion to human cells. To further investigate whether 6-MP can indeed inhibit c-di-GMP signaling in AIEC, we performed biofilm and motility assays and determination of extracellular polysaccharides. 6-MP clearly affected biofilm formation and cellulose production, but also, unexpectedly, reduced cell motility, itself an important virulence factor for AIEC. Our results provide strong evidence

  10. Immune response of macrophages from young and aged mice to the oral pathogenic bacterium Porphyromonas gingivalis

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    Gibson Frank C

    2010-11-01

    Full Text Available Abstract Periodontal disease is a chronic inflammatory gum disease that in severe cases leads to tooth loss. Porphyromonas gingivalis (Pg is a bacterium closely associated with generalized forms of periodontal disease. Clinical onset of generalized periodontal disease commonly presents in individuals over the age of 40. Little is known regarding the effect of aging on inflammation associated with periodontal disease. In the present study we examined the immune response of bone marrow derived macrophages (BMM from young (2-months and aged (1-year and 2-years mice to Pg strain 381. Pg induced robust expression of cytokines; tumor necrosis factor (TNF-α, interleukin (IL-6, and IL-10, chemokines; neutrophil chemoattractant protein (KC, macrophage colony stimulating factor (MCP-1, macrophage inflammatory protein (MIP-1α and regulated upon activation normal T cell expressed and secreted (RANTES, as well as nitric oxide (NO, measured as nitrite, and prostaglandin E2 (PGE2 from BMM of young mice. BMM from the 2-year age group produced significantly less TNF-α, IL-6 and NO in response to Pg as compared with BMM from 2-months and 1-year of age. We did not observe any difference in the levels of IL-1β, IL-10 and PGE2 produced by BMM in response to Pg. BMM from 2-months and 1-year of age produced similar levels of all chemokines measured with the exception of MCP-1, which was reduced in BMM from 1-year of age. BMM from the 2-year group produced significantly less MCP-1 and MIP-1α compared with 2-months and 1-year age groups. No difference in RANTES production was observed between age groups. Employing a Pg attenuated mutant, deficient in major fimbriae (Pg DPG3, we observed reduced ability of the mutant to stimulate inflammatory mediator expression from BMMs as compared to Pg 381, irrespective of age. Taken together these results support senescence as an important facet of the reduced immunological response observed by BMM of aged host to the

  11. Fimbriiglobus ruber gen. nov., sp. nov., a Gemmata-like planctomycete from Sphagnum peat bog and the proposal of Gemmataceae fam. nov.

    Science.gov (United States)

    Kulichevskaya, Irina S; Ivanova, Anastasia A; Baulina, Olga I; Rijpstra, W Irene C; Sinninghe Damsté, Jaap S; Dedysh, Svetlana N

    2017-02-01

    An aerobic, budding, dark pink to red-pigmented bacterium was isolated from an acidic boreal Sphagnum peat bog and designated strain SP5T. Cells of this strain were non-motile spheres that were uniformly covered with crateriform pits and fimbria, and tended to form aggregates during growth in liquid media. Strain SP5T was capable of growth between pH 4.0 and pH 6.8 (optimum at pH 5.5-6.0) and at temperatures between 10 and 30 °C (optimum at 20-25 °C). The preferred growth substrates were sugars and some heteropolysaccharides. The major fatty acids were C20 : 1ω9c, C16 : 1ω9c and C16 : 0, and the major polar lipid was trimethylornithine. Cells contained also significant amounts of bound (ω-1)OH-C30 : 1 fatty acid. The quinone was menaquinone-6, and the G+C content of the DNA was 60.7 mol%. Strain SP5T was a member of the order Planctomycetales and belonged to the phylogenetic lineage defined by the genus Gemmata. It displayed 88 and 89 % 16S rRNA gene sequence similarity to Gemmata obscuriglobusUQM 2246T and 'Gemmata massiliana' IIL30, 89 % to Zavarzinella formosa A10T and 86 % to Telmatocola sphagniphila SP2T. However, strain SP5T differed from members of these genera by cell morphology, substrate utilization pattern and fatty acid composition. Based on these data, the novel isolate should be considered as representing a novel species of a new genus of planctomycetes, for which the name Fimbriiglobus ruber gen. nov., sp. nov, is proposed. The type strain is SP5T (=LMG 29572T=VKM B-3045T). We also suggest the establishment of a novel family, Gemmataceaefam. nov., which includes the phylogenetically related genera Gemmata, Zavarzinella, Telmatocola and Fimbriiglobus.

  12. Transcriptional profiling of Actinobacillus pleuropneumoniae during the acute phase of a natural infection in pigs

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    Harel Josée

    2010-02-01

    Full Text Available Abstract Background Actinobacillus pleuropneumoniae is the etiological agent of porcine pleuropneumonia, a respiratory disease which causes great economic losses worldwide. Many virulence factors are involved in the pathogenesis, namely capsular polysaccharides, RTX toxins, LPS and many iron acquisition systems. In order to identify genes that are expressed in vivo during a natural infection, we undertook transcript profiling experiments with an A. pleuropneumoniae DNA microarray, after recovery of bacterial mRNAs from serotype 5b-infected porcine lungs. AppChip2 contains 2033 PCR amplicons based on the genomic sequence of App serotype 5b strain L20, representing more than 95% of ORFs greater than 160 bp in length. Results Transcriptional profiling of A. pleuropneumoniae recovered from the lung of a pig suffering from a natural infection or following growth of the bacterial isolate in BHI medium was performed. An RNA extraction protocol combining beadbeating and hot-acid-phenol was developed in order to maximize bacterial mRNA yields and quality following total RNA extraction from lung lesions. Nearly all A. pleuropneumoniae transcripts could be detected on our microarrays, and 150 genes were deemed differentially expressed in vivo during the acute phase of the infection. Our results indicate that, for example, gene apxIVA from an operon coding for RTX toxin ApxIV is highly up-regulated in vivo, and that two genes from the operon coding for type IV fimbriae (APL_0878 and APL_0879 were also up-regulated. These transcriptional profiling data, combined with previous comparative genomic hybridizations performed by our group, revealed that 66 out of the 72 up-regulated genes are conserved amongst all serotypes and that 3 of them code for products that are predicted outer membrane proteins (genes irp and APL_0959, predicted to code for a TonB-dependent receptor and a filamentous hemagglutinin/adhesin respectively or lipoproteins (gene APL_0920. Only 4

  13. DNA microarray-based assessment of virulence potential of Shiga toxin gene-carrying Escherichia coli O104:H7 isolated from feedlot cattle feces.

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    Pragathi B Shridhar

    Full Text Available Escherichia coli O104:H4, a hybrid pathotype reported in a large 2011 foodborne outbreak in Germany, has not been detected in cattle feces. However, cattle harbor and shed in the feces other O104 serotypes, particularly O104:H7, which has been associated with sporadic cases of diarrhea in humans. The objective of our study was to assess the virulence potential of Shiga toxin-producing E. coli (STEC O104:H7 isolated from feces of feedlot cattle using DNA microarray. Six strains of STEC O104:H7 isolated from cattle feces were analyzed using FDA-E. coli Identification (ECID DNA microarray to determine their virulence profiles and compare them to the human strains (clinical of O104:H7, STEC O104:H4 (German outbreak strain, and O104:H21 (milk-associated Montana outbreak strain. Scatter plots were generated from the array data to visualize the gene-level differences between bovine and human O104 strains, and Pearson correlation coefficients (r were determined. Splits tree was generated to analyze relatedness between the strains. All O104:H7 strains, both bovine and human, similar to O104:H4 and O104:H21 outbreak strains were negative for intimin (eae. The bovine strains were positive for Shiga toxin 1 subtype c (stx1c, enterohemolysin (ehxA, tellurite resistance gene (terD, IrgA homolog protein (iha, type 1 fimbriae (fimH, and negative for genes that code for effector proteins of type III secretory system. The six cattle O104 strains were closely related (r = 0.86-0.98 to each other, except for a few differences in phage related and non-annotated genes. One of the human clinical O104:H7 strains (2011C-3665 was more closely related to the bovine O104:H7 strains (r = 0.81-0.85 than the other four human clinical O104:H7 strains (r = 0.75-0.79. Montana outbreak strain (O104:H21 was more closely related to four of the human clinical O104:H7 strains than the bovine O104:H7 strains. None of the bovine E. coli O104 strains carried genes characteristic of E

  14. Test-retest reliability and longitudinal analysis of automated hippocampal subregion volumes in healthy ageing and Alzheimer's disease populations.

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    Worker, Amanda; Dima, Danai; Combes, Anna; Crum, William R; Streffer, Johannes; Einstein, Steven; Mehta, Mitul A; Barker, Gareth J; C R Williams, Steve; O'daly, Owen

    2018-04-01

    The hippocampal formation is a complex brain structure that is important in cognitive processes such as memory, mood, reward processing and other executive functions. Histological and neuroimaging studies have implicated the hippocampal region in neuropsychiatric disorders as well as in neurodegenerative diseases. This highly plastic limbic region is made up of several subregions that are believed to have different functional roles. Therefore, there is a growing interest in imaging the subregions of the hippocampal formation rather than modelling the hippocampus as a homogenous structure, driving the development of new automated analysis tools. Consequently, there is a pressing need to understand the stability of the measures derived from these new techniques. In this study, an automated hippocampal subregion segmentation pipeline, released as a developmental version of Freesurfer (v6.0), was applied to T1-weighted magnetic resonance imaging (MRI) scans of 22 healthy older participants, scanned on 3 separate occasions and a separate longitudinal dataset of 40 Alzheimer's disease (AD) patients. Test-retest reliability of hippocampal subregion volumes was assessed using the intra-class correlation coefficient (ICC), percentage volume difference and percentage volume overlap (Dice). Sensitivity of the regional estimates to longitudinal change was estimated using linear mixed effects (LME) modelling. The results show that out of the 24 hippocampal subregions, 20 had ICC scores of 0.9 or higher in both samples; these regions include the molecular layer, granule cell layer of the dentate gyrus, CA1, CA3 and the subiculum (ICC > 0.9), whilst the hippocampal fissure and fimbria had lower ICC scores (0.73-0.88). Furthermore, LME analysis of the independent AD dataset demonstrated sensitivity to group and individual differences in the rate of volume change over time in several hippocampal subregions (CA1, molecular layer, CA3, hippocampal tail, fissure and presubiculum

  15. Involvement of a periodontal pathogen, Porphyromonas gingivalis on the pathogenesis of non-alcoholic fatty liver disease

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    Yoneda Masato

    2012-02-01

    Full Text Available Abstract Background Non-alcoholic fatty liver disease (NAFLD is a hepatic manifestation of metabolic syndrome that is closely associated with multiple factors such as obesity, hyperlipidemia and type 2 diabetes mellitus. However, other risk factors for the development of NAFLD are unclear. With the association between periodontal disease and the development of systemic diseases receiving increasing attention recently, we conducted this study to investigate the relationship between NAFLD and infection with Porphyromonas gingivalis (P. gingivalis, a major causative agent of periodontitis. Methods The detection frequencies of periodontal bacteria in oral samples collected from 150 biopsy-proven NAFLD patients (102 with non-alcoholic steatohepatitis (NASH and 48 with non-alcoholic fatty liver (NAFL patients and 60 non-NAFLD control subjects were determined. Detection of P. gingivalis and other periodontopathic bacteria were detected by PCR assay. In addition, effect of P. gingivalis-infection on mouse NAFLD model was investigated. To clarify the exact contribution of P. gingivalis-induced periodontitis, non-surgical periodontal treatments were also undertaken for 3 months in 10 NAFLD patients with periodontitis. Results The detection frequency of P. gingivalis in NAFLD patients was significantly higher than that in the non-NAFLD control subjects (46.7% vs. 21.7%, odds ratio: 3.16. In addition, the detection frequency of P. gingivalis in NASH patients was markedly higher than that in the non-NAFLD subjects (52.0%, odds ratio: 3.91. Most of the P. gingivalis fimbria detected in the NAFLD patients was of invasive genotypes, especially type II (50.0%. Infection of type II P. gingivalis on NAFLD model of mice accelerated the NAFLD progression. The non-surgical periodontal treatments on NAFLD patients carried out for 3 months ameliorated the liver function parameters, such as the serum levels of AST and ALT. Conclusions Infection with high-virulence P

  16. Inactive Gingipains from P. gingivalis Selectively Skews T Cells toward a Th17 Phenotype in an IL-6 Dependent Manner

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    Jan Potempa

    2017-04-01

    Full Text Available Gingipain cysteine proteases are considered key virulence factors of Porphyromonas gingivalis. They significantly influence antibacterial and homeostatic functions of macrophages, neutrophils, the complement system, and cytokine networks. Recent data indicate the role of P. gingivalis in T cell differentiation; however, the involvement of gingipains in this process remains elusive. Therefore, the aim of this study was to investigate the contribution of danger signals triggered by the gingipains on the generation of Th17 cells, which play a key role in protection against bacterial diseases but may cause chronic inflammation and bone resorption. To this end we compared the effects of the wild-type strain of P. gingivalis (W83 with its isogenic mutant devoid of gingipain activity (ΔKΔRAB, and bacterial cells pretreated with a highly-specific inhibitor of gingipains activity (KYTs. Antigen presenting cells (APCs, both professional (dendritic cells, and non-professional (gingival keratinocytes, exposed to viable bacteria expressed high amounts of cytokines (IL-6, IL-21, IL-23. These cytokines are reported to either stimulate or balance the Th17-dependent immune response. Surprisingly, cells infected with P. gingivalis devoid of gingipain activity showed increased levels of all tested cytokines compared to bacteria with fully active enzymes. The effect was dependent on both the reduction of cytokine proteolysis and the lack of cross-talk with other bacterial virulence factors, including LPS and fimbriae that induce de novo synthesis of cytokines. The profile of lymphocyte T differentiation from naive T cells showed enhanced generation of Th17 in response to bacteria with inactive gingipains. Moreover, we found that gingipain-dependent induction of Th17 cells was highly specific, since other T cell-subsets remained unchanged. Finally, inhibition of IL-6 signaling in dendritic cells led to a significant depletion of the Th17 population. Cumulatively

  17. Detection and Serogrouping of Dichelobacter nodosus Infection by Use of Direct PCR from Lesion Swabs To Support Outbreak-Specific Vaccination for Virulent Footrot in Sheep.

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    McPherson, Andrew S; Dhungyel, Om P; Whittington, Richard J

    2018-04-01

    Virulent footrot is an economically significant disease in most sheep-rearing countries. The disease can be controlled with vaccine targeting the fimbriae of virulent strains of the essential causative agent, Dichelobacter nodosus However, the bacterium is immunologically heterogeneous, and 10 distinct fimbrial serogroups have been identified. Ideally, in each outbreak the infecting strains would be cultured and serogrouped so that the appropriate serogroup-specific mono- or bivalent vaccine could be administered, because multivalent vaccines lack efficacy due to antigenic competition. If clinical disease expression is suspected to be incomplete, culture-based virulence tests are required to confirm the diagnosis, because control of benign footrot is economically unjustifiable. Both diagnosis and vaccination are conducted at the flock level. The aims of this study were to develop a PCR-based procedure for detecting and serogrouping D. nodosus directly from foot swabs and to determine whether this could be done accurately from the same cultured swab. A total of 269 swabs from the active margins of foot lesions of 261 sheep in 12 Merino sheep flocks in southeastern Australia were evaluated. DNA extracts taken from putative pure cultures of D. nodosus and directly from the swabs were evaluated in PCR assays for the 16S rRNA and fimA genes of D. nodosus Pure cultures were tested also by the slide agglutination test. Direct PCR using extracts from swabs was more sensitive than culture for detecting and serogrouping D. nodosus strains. Using the most sensitive sample collection method of the use of swabs in lysis buffer, D. nodosus was more likely to be detected by PCR in active than in inactive lesions, and in lesions with low levels of fecal contamination, but lesion score was not a significant factor. PCR conducted on extracts from swabs in modified Stuart's transport medium that had already been used to inoculate culture plates had lower sensitivity. Therefore, if

  18. In vivo quantitative whole-brain diffusion tensor imaging analysis of APP/PS1 transgenic mice using voxel-based and atlas-based methods

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    Qin, Yuan-Yuan [Huazhong University of Science and Technology, Department of Radiology, Tongji Hospital, Tongji Medical College, Wuhan (China); The Johns Hopkins University School of Medicine, The Russell H. Morgan Department of Radiology and Radiological Science, Baltimore, MD (United States); Li, Mu-Wei; Oishi, Kenichi [The Johns Hopkins University School of Medicine, The Russell H. Morgan Department of Radiology and Radiological Science, Baltimore, MD (United States); Zhang, Shun; Zhang, Yan; Zhao, Ling-Yun; Zhu, Wen-Zhen [Huazhong University of Science and Technology, Department of Radiology, Tongji Hospital, Tongji Medical College, Wuhan (China); Lei, Hao [Chinese Academy of Sciences, Wuhan Center for Magnetic Resonance, State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics, Wuhan Institute of Physics and Mathematics, Wuhan (China)

    2013-08-15

    Diffusion tensor imaging (DTI) has been applied to characterize the pathological features of Alzheimer's disease (AD) in a mouse model, although little is known about whether these features are structure specific. Voxel-based analysis (VBA) and atlas-based analysis (ABA) are good complementary tools for whole-brain DTI analysis. The purpose of this study was to identify the spatial localization of disease-related pathology in an AD mouse model. VBA and ABA quantification were used for the whole-brain DTI analysis of nine APP/PS1 mice and wild-type (WT) controls. Multiple scalar measurements, including fractional anisotropy (FA), trace, axial diffusivity (DA), and radial diffusivity (DR), were investigated to capture the various types of pathology. The accuracy of the image transformation applied for VBA and ABA was evaluated by comparing manual and atlas-based structure delineation using kappa statistics. Following the MR examination, the brains of the animals were analyzed for microscopy. Extensive anatomical alterations were identified in APP/PS1 mice, in both the gray matter areas (neocortex, hippocampus, caudate putamen, thalamus, hypothalamus, claustrum, amygdala, and piriform cortex) and the white matter areas (corpus callosum/external capsule, cingulum, septum, internal capsule, fimbria, and optic tract), evidenced by an increase in FA or DA, or both, compared to WT mice (p < 0.05, corrected). The average kappa value between manual and atlas-based structure delineation was approximately 0.8, and there was no significant difference between APP/PS1 and WT mice (p > 0.05). The histopathological changes in the gray matter areas were confirmed by microscopy studies. DTI did, however, demonstrate significant changes in white matter areas, where the difference was not apparent by qualitative observation of a single-slice histological specimen. This study demonstrated the structure-specific nature of pathological changes in APP/PS1 mouse, and also showed the

  19. GtfA and GtfB Are Both Required for Protein O-Glycosylation in Lactobacillus plantarum

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    Lee, I-Chiao; van Swam, Iris I.; Tomita, Satoru; Morsomme, Pierre; Rolain, Thomas; Hols, Pascal; Bron, Peter A.

    2014-01-01

    Acm2, the major autolysin of Lactobacillus plantarum WCFS1, was recently found to be O-glycosylated with N-acetylhexosamine, likely N-acetylglucosamine (GlcNAc). In this study, we set out to identify the glycosylation machinery by employing a comparative genomics approach to identify Gtf1 homologues, which are involved in fimbria-associated protein 1 (Fap1) glycosylation in Streptococcus parasanguinis. This in silico approach resulted in the identification of 6 candidate L. plantarum WCFS1 genes with significant homology to Gtf1, namely, tagE1 to tagE6. These candidate genes were targeted by systematic gene deletion, followed by assessment of the consequences on glycosylation of Acm2. We observed a changed mobility of Acm2 on SDS-PAGE in the tagE5E6 deletion strain, while deletion of other tagE genes resulted in Acm2 mobility comparable to that of the wild type. Subsequent mass spectrometry analysis of excised and in-gel-digested Acm2 confirmed the loss of glycosylation on Acm2 in the tagE5E6 deletion mutant, whereas a lectin blot using GlcNAc-specific succinylated wheat germ agglutinin (sWGA) revealed that besides Acm2, tagE5E6 deletion also abolished all but one other sWGA-reactive, protease-sensitive signal. Only complementation of both tagE5 and tagE6 restored those sWGA lectin signals, establishing that TagE5 and TagE6 are both required for the glycosylation of Acm2 as well as the vast majority of other sWGA-reactive proteins. Finally, sWGA lectin blotting experiments using a panel of 8 other L. plantarum strains revealed that protein glycosylation is a common feature in L. plantarum strains. With the establishment of these enzymes as protein glycosyltransferases, we propose to rename TagE5 and TagE6 as GtfA and GtfB, respectively. PMID:24532775

  20. Parallel processing of information about location in the amygdala, entorhinal cortex and hippocampus.

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    Gaskin, Stephane; White, Norman M

    2013-11-01

    The conditioned cue preference paradigm was used to study how rats use extra-maze cues to discriminate between 2 adjacent arms on an 8-arm radial maze, a situation in which most of the same cues can be seen from both arms but only one arm contains food. Since the food-restricted rats eat while passively confined on the food-paired arm no responses are reinforced, so the discrimination is due to Pavlovian stimulus-reward (or outcome) learning. Consistent with other evidence that rats must move around in an environment to acquire a spatial map, we found that learning the adjacent arms CCP (ACCP) required a minimum amount of active exploration of the maze with no reinforcers present prior to passive pairing of the extra-maze cues with the food reinforcer, an instance of latent learning. Temporary inactivation of the hippocampus during the pre-exposure sessions had no effect on ACCP learning, confirming other evidence that the hippocampus is not involved in latent learning. A series of experiments indentified a circuit involving fimbria-fornix and dorsal entorhinal cortex as the neural basis of latent learning in this situation. In contrast, temporary inactivation of the entorhinal cortex or hippocampus during passive training or during testing blocked ACCP learning and expression, respectively, suggesting that these two structures co-operate in using spatial information to learn the location of food on the maze during passive pairing and to express this combined information during testing. In parallel with these processes we found that the amygdala processes information leading to an equal tendency to enter both adjacent arms (even though only one was paired with food) suggesting that the stimulus information available to this structure is not sufficiently precise to discriminate between the ambiguous cues visible from the adjacent arms. Expression of the ACCP in normal rats depends on hippocampus-based learning to avoid the unpaired arm which competes with the

  1. Expression of the Flp proteins by Haemophilus ducreyi is necessary for virulence in human volunteers

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    Zwickl Beth W

    2011-09-01

    Full Text Available Abstract Background Haemophilus ducreyi, the causative agent of the sexually transmitted disease chancroid, contains a flp (fimbria like protein operon that encodes proteins predicted to contribute to adherence and pathogenesis. H. ducreyi mutants that lack expression of Flp1 and Flp2 or TadA, which has homology to NTPases of type IV secretion systems, have decreased abilities to attach to and form microcolonies on human foreskin fibroblasts (HFF. A tadA mutant is attenuated in its ability to cause disease in human volunteers and in the temperature dependent rabbit model, but a flp1flp2 mutant is virulent in rabbits. Whether a flp deletion mutant would cause disease in humans is not clear. Results We constructed 35000HPΔflp1-3, a deletion mutant that lacks expression of all three Flp proteins but has an intact tad secretion system. 35000HPΔflp1-3 was impaired in its ability to form microcolonies and to attach to HFF in vitro when compared to its parent (35000HP. Complementation of the mutant with flp1-3 in trans restored the parental phenotype. To test whether expression of Flp1-3 was necessary for virulence in humans, ten healthy adult volunteers were experimentally infected with a fixed dose of 35000HP (ranging from 54 to 67 CFU on one arm and three doses of 35000HPΔflp1-3 (ranging from 63 to 961 CFU on the other arm. The overall papule formation rate for the parent was 80% (95% confidence interval, CI, 55.2%-99.9% and for the mutant was 70.0% (95% CI, 50.5%-89.5% (P = 0.52. Mutant papules were significantly smaller (mean, 11.2 mm2 than were parent papules (21.8 mm2 24 h after inoculation (P = 0.018. The overall pustule formation rates were 46.7% (95% CI 23.7-69.7% at 30 parent sites and 6.7% (95% CI, 0.1-19.1% at 30 mutant sites (P = 0.001. Conclusion These data suggest that production and secretion of the Flp proteins contributes to microcolony formation and attachment to HFF cells in vitro. Expression of flp1-3 is also necessary for H

  2. Expression of the Flp proteins by Haemophilus ducreyi is necessary for virulence in human volunteers.

    Science.gov (United States)

    Janowicz, Diane M; Cooney, Sean A; Walsh, Jessica; Baker, Beth; Katz, Barry P; Fortney, Kate R; Zwickl, Beth W; Ellinger, Sheila; Munson, Robert S

    2011-09-22

    Haemophilus ducreyi, the causative agent of the sexually transmitted disease chancroid, contains a flp (fimbria like protein) operon that encodes proteins predicted to contribute to adherence and pathogenesis. H. ducreyi mutants that lack expression of Flp1 and Flp2 or TadA, which has homology to NTPases of type IV secretion systems, have decreased abilities to attach to and form microcolonies on human foreskin fibroblasts (HFF). A tadA mutant is attenuated in its ability to cause disease in human volunteers and in the temperature dependent rabbit model, but a flp1flp2 mutant is virulent in rabbits. Whether a flp deletion mutant would cause disease in humans is not clear. We constructed 35000HPΔflp1-3, a deletion mutant that lacks expression of all three Flp proteins but has an intact tad secretion system. 35000HPΔflp1-3 was impaired in its ability to form microcolonies and to attach to HFF in vitro when compared to its parent (35000HP). Complementation of the mutant with flp1-3 in trans restored the parental phenotype. To test whether expression of Flp1-3 was necessary for virulence in humans, ten healthy adult volunteers were experimentally infected with a fixed dose of 35000HP (ranging from 54 to 67 CFU) on one arm and three doses of 35000HPΔflp1-3 (ranging from 63 to 961 CFU) on the other arm. The overall papule formation rate for the parent was 80% (95% confidence interval, CI, 55.2%-99.9%) and for the mutant was 70.0% (95% CI, 50.5%-89.5%) (P = 0.52). Mutant papules were significantly smaller (mean, 11.2 mm2) than were parent papules (21.8 mm2) 24 h after inoculation (P = 0.018). The overall pustule formation rates were 46.7% (95% CI 23.7-69.7%) at 30 parent sites and 6.7% (95% CI, 0.1-19.1%) at 30 mutant sites (P = 0.001). These data suggest that production and secretion of the Flp proteins contributes to microcolony formation and attachment to HFF cells in vitro. Expression of flp1-3 is also necessary for H. ducreyi to initiate disease and progress to

  3. High-grade serous carcinomas arise in the mouse oviduct via defects linked to the human disease.

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    Zhai, Yali; Wu, Rong; Kuick, Rork; Sessine, Michael S; Schulman, Stephanie; Green, Megan; Fearon, Eric R; Cho, Kathleen R

    2017-09-01

    Recent studies have suggested that the most common and lethal type of 'ovarian' cancer, i.e. high-grade serous carcinoma (HGSC), usually arises from epithelium on the fallopian tube fimbriae, and not from the ovarian surface epithelium. We have developed Ovgp1-iCreER T2 mice in which the Ovgp1 promoter controls expression of tamoxifen-regulated Cre recombinase in oviductal epithelium - the murine equivalent of human fallopian tube epithelium (FTE). We employed Ovgp1-iCreER T2 mice to show that FTE-specific inactivation of several different combinations of tumour suppressor genes that are recurrently mutated in human HGSCs - namely Brca1, Trp53, Rb1, and Nf1 - results in serous tubal intraepithelial carcinomas (STICs) that progress to HGSC or carcinosarcoma, and to widespread metastatic disease in a subset of mice. The cancer phenotype is highly penetrant and more rapid in mice carrying engineered alleles of all four tumour suppressor genes. Brca1, Trp53 and Pten inactivation in the oviduct also results in STICs and HGSCs, and is associated with diffuse epithelial hyperplasia and mucinous metaplasia, which are not observed in mice with intact Pten. Oviductal tumours arise earlier in these mice than in those with Brca1, Trp53, Rb1 and Nf1 inactivation. Tumour initiation and/or progression in mice lacking conditional Pten alleles probably require the acquisition of additional defects, a notion supported by our identification of loss of the wild-type Rb1 allele in the tumours of mice carrying only one floxed Rb1 allele. Collectively, the models closely recapitulate the heterogeneity and histological, genetic and biological features of human HGSC. These models should prove useful for studying the pathobiology and genetics of HGSC in vivo, and for testing new approaches for prevention, early detection, and treatment. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2017 Pathological Society of

  4. Impact of Vortioxetine on Synaptic Integration in Prefrontal-Subcortical Circuits: Comparisons with Escitalopram

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    Shreaya Chakroborty

    2017-10-01

    Full Text Available Prefrontal-subcortical circuits support executive functions which often become dysfunctional in psychiatric disorders. Vortioxetine is a multimodal antidepressant that is currently used in the clinic to treat major depressive disorder. Mechanisms of action of vortioxetine include serotonin (5-HT transporter blockade, 5-HT1A receptor agonism, 5-HT1B receptor partial agonism, and 5-HT1D, 5-HT3, and 5-HT7 receptor antagonism. Vortioxetine facilitates 5-HT transmission in the medial prefrontal cortex (mPFC, however, the impact of this compound on related prefrontal-subcortical circuits is less clear. Thus, the current study examined the impact of systemic vortioxetine administration (0.8 mg/kg, i.v. on spontaneous spiking and spikes evoked by electrical stimulation of the mPFC in the anterior cingulate cortex (ACC, medial shell of the nucleus accumbens (msNAc, and lateral septal nucleus (LSN in urethane-anesthetized rats. We also examined whether vortioxetine modulated afferent drive in the msNAc from hippocampal fimbria (HF inputs. Similar studies were performed using the selective 5-HT reuptake inhibitor [selective serotonin reuptake inhibitors (SSRI] escitalopram (1.6 mg/kg, i.v. to enable comparisons between the multimodal actions of vortioxetine and SSRI-mediated effects. No significant differences in spontaneous activity were observed in the ACC, msNAc, and LSN across treatment groups. No significant impact of treatment on mPFC-evoked responses was observed in the ACC. In contrast, vortioxetine decreased mPFC-evoked activity recorded in the msNAc as compared to parallel studies in control and escitalopram treated groups. Thus, vortioxetine may reduce mPFC-msNAc afferent drive via a mechanism that, in addition to an SSRI-like effect, requires 5-HT receptor modulation. Recordings in the LSN revealed a significant increase in mPFC-evoked activity following escitalopram administration as compared to control and vortioxetine treated groups

  5. The Escherichia coli P and Type 1 Pilus Assembly Chaperones PapD and FimC Are Monomeric in Solution

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    Sarowar, Samema; Hu, Olivia J.; Werneburg, Glenn T.; Thanassi, David G.; Li, Huilin; Christie, P. J.

    2016-06-27

    ABSTRACT

    The chaperone/usher pathway is used by Gram-negative bacteria to assemble adhesive surface structures known as pili or fimbriae. Uropathogenic strains ofEscherichia coliuse this pathway to assemble P and type 1 pili, which facilitate colonization of the kidney and bladder, respectively. Pilus assembly requires a periplasmic chaperone and outer membrane protein termed the usher. The chaperone allows folding of pilus subunits and escorts the subunits to the usher for polymerization into pili and secretion to the cell surface. Based on previous structures of mutant versions of the P pilus chaperone PapD, it was suggested that the chaperone dimerizes in the periplasm as a self-capping mechanism. Such dimerization is counterintuitive because the chaperone G1 strand, important for chaperone-subunit interaction, is buried at the dimer interface. Here, we show that the wild-type PapD chaperone also forms a dimer in the crystal lattice; however, the dimer interface is different from the previously solved structures. In contrast to the crystal structures, we found that both PapD and the type 1 pilus chaperone, FimC, are monomeric in solution. Our findings indicate that pilus chaperones do not sequester their G1 β-strand by forming a dimer. Instead, the chaperones may expose their G1 strand for facile interaction with pilus subunits. We also found that the type 1 pilus adhesin, FimH, is flexible in solution while in complex with its chaperone, whereas the P pilus adhesin, PapGII, is rigid. Our study clarifies a crucial step in pilus biogenesis and reveals pilus-specific differences that may relate to biological function.

    IMPORTANCEPili are critical virulence factors for many bacterial pathogens. UropathogenicE. colirelies on P and type 1 pili assembled by the chaperone/usher pathway to

  6. Maternal exposure to hexachlorophene targets intermediate-stage progenitor cells of the hippocampal neurogenesis in rat offspring via dysfunction of cholinergic inputs by myelin vacuolation

    International Nuclear Information System (INIS)

    Itahashi, Megu; Abe, Hajime; Tanaka, Takeshi; Mizukami, Sayaka; Kimura, Masayuki; Yoshida, Toshinori; Shibutani, Makoto

    2015-01-01

    Highlights: • The effect of maternal exposure to HCP on rat hippocampal neurogenesis was examined. • HCP induces myelin vacuolation of nerve tracts in the septal–hippocampal pathway. • Myelin changes suppress Chrnb2-mediated cholinergic inputs to the dentate gyrus. • SGZ apoptosis occurs via the mitochondrial pathway and targets type-2b cells. • Dysfunction of cholinergic inputs is related to type-2b SGZ cell apoptosis. - Abstract: Hexachlorophene (HCP) is known to induce myelin vacuolation corresponding to intramyelinic edema of nerve fibers in the central and peripheral nervous system in animals. This study investigated the effect of maternal exposure to HCP on hippocampal neurogenesis in rat offspring using pregnant rats supplemented with 0 (controls), 100, or 300 ppm HCP in the diet from gestational day 6 to day 21 after delivery. On postnatal day (PND) 21, the numbers of T box brain 2 + progenitor cells and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling + apoptotic cells in the hippocampal subgranular zone (SGZ) decreased in female offspring at 300 ppm, which was accompanied by myelin vacuolation and punctate tubulin beta-3 chain staining of nerve fibers in the hippocampal fimbria. In addition, transcript levels of the cholinergic receptor, nicotinic beta 2 (Chrnb2) and B-cell CLL/lymphoma 2 (Bcl2) decreased in the dentate gyrus. HCP-exposure did not alter the numbers of SGZ proliferating cells and reelin- or calcium-binding protein-expressing γ-aminobutyric acid (GABA)-ergic interneuron subpopulations in the dentate hilus on PND 21 and PND 77. Although some myelin vacuolation remained, all other changes observed in HCP-exposed offspring on PND 21 disappeared on PND 77. These results suggest that maternal HCP exposure reversibly decreases type-2b intermediate-stage progenitor cells via the mitochondrial apoptotic pathway in offspring hippocampal neurogenesis at 300 ppm HCP. Neurogenesis may be affected by dysfunction

  7. Quantitative autoradiography of brain binding sites for the vesicular acetylcholine transport blocker 2-(4-phenylpiperidino)cyclohexanol (AH5183)

    International Nuclear Information System (INIS)

    Marien, M.R.; Parsons, S.M.; Altar, C.A.

    1987-01-01

    2-(4-Phenylpiperidino)cyclohexanol (AH5183) is a noncompetitive and potent inhibitor of high-affinity acetylcholine transport into cholinergic vesicles. It is reported here that [ 3 H]AH5183 binds specifically and saturably to slide-mounted sections of the rat forebrain (Kd = 1.1 to 2.2 X 10(-8) M; Bmax = 286 to 399 fmol/mg of protein). The association and dissociation rate constants for [ 3 H]AH5183 binding are 8.6 X 10(6) M-1 X min-1 and 0.18 min-1, respectively. Bound [ 3 H]AH5183 can be displaced by nonradioactive AH5183 and by the structural analog (2 alpha,3 beta,4A beta,8A alpha)-decahydro-3-(4-phenyl-1-piperidinyl)-2- naphthalenol but not by 10 microM concentrations of the cholinergic drugs acetylcholine, choline, atropine, hexamethonium, eserine, or hemicholinium-3 or by the structurally related compounds 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, 1-methyl-4-phenylpyridine, (+/-)-N-allylnormetazocine (SKF 10,047), levoxadrol, or dexoxadrol. Quantitative autoradiography reveals that [ 3 H]AH5183 binding sites are distributed heterogenously throughout the rat forebrain and are highly localized to cholinergic nerve terminal regions. At the level of the caudate nucleus-putamen, the highest concentrations of saturable [ 3 H]AH5183 binding (713-751 fmol/mg of protein) are found in the vertical limb of the diagonal band and the olfactory tubercle, with lesser amounts (334-516 fmol/mg of protein) in the caudate-putamen, nucleus accumbens, superficial layers of the cerebral cortex, and the primary olfactory cortex. At day 7 after transsection of the left fimbria, [ 3 H]AH5183 binding and choline acetyltransferase activity in the left hippocampus were reduced by 33 +/- 6% and 61 +/- 7%, respectively. These findings indicate that [ 3 H]AH5183 binds to a unique recognition site in rat brain that is topographically associated with cholinergic nerve terminals

  8. Inactive Gingipains from P. gingivalis Selectively Skews T Cells toward a Th17 Phenotype in an IL-6 Dependent Manner.

    Science.gov (United States)

    Glowczyk, Izabela; Wong, Alicia; Potempa, Barbara; Babyak, Olena; Lech, Maciej; Lamont, Richard J; Potempa, Jan; Koziel, Joanna

    2017-01-01

    Gingipain cysteine proteases are considered key virulence factors of Porphyromonas gingivalis . They significantly influence antibacterial and homeostatic functions of macrophages, neutrophils, the complement system, and cytokine networks. Recent data indicate the role of P. gingivalis in T cell differentiation; however, the involvement of gingipains in this process remains elusive. Therefore, the aim of this study was to investigate the contribution of danger signals triggered by the gingipains on the generation of Th17 cells, which play a key role in protection against bacterial diseases but may cause chronic inflammation and bone resorption. To this end we compared the effects of the wild-type strain of P. gingivalis (W83) with its isogenic mutant devoid of gingipain activity (ΔKΔRAB), and bacterial cells pretreated with a highly-specific inhibitor of gingipains activity (KYTs). Antigen presenting cells (APCs), both professional (dendritic cells), and non-professional (gingival keratinocytes), exposed to viable bacteria expressed high amounts of cytokines (IL-6, IL-21, IL-23). These cytokines are reported to either stimulate or balance the Th17-dependent immune response. Surprisingly, cells infected with P. gingivalis devoid of gingipain activity showed increased levels of all tested cytokines compared to bacteria with fully active enzymes. The effect was dependent on both the reduction of cytokine proteolysis and the lack of cross-talk with other bacterial virulence factors, including LPS and fimbriae that induce de novo synthesis of cytokines. The profile of lymphocyte T differentiation from naive T cells showed enhanced generation of Th17 in response to bacteria with inactive gingipains. Moreover, we found that gingipain-dependent induction of Th17 cells was highly specific, since other T cell-subsets remained unchanged. Finally, inhibition of IL-6 signaling in dendritic cells led to a significant depletion of the Th17 population. Cumulatively, this study

  9. [Hippocampal subfield volume alteration in post-traumatic stress disorder: a magnetic resonance imaging study].

    Science.gov (United States)

    Lu, Lu; Zhang, Lianqing; Hu, Xinyu; Hu, Xiaoxiao; Li, Lingjiang; Gong, Qiyong; Huang, Xiaoqi

    2018-04-01

    In the current study, we aim to investigate whether post-traumatic stress disorder (PTSD) is associated with structural alterations in specific subfields of hippocampus comparing with trauma-exposed control (TC) in a relatively large sample. We included 67 PTSD patients who were diagnosed under Diagnostic and Statistical Manual of Mental Disorders (4th Edition) (DSM-Ⅳ) criteria and 78 age- and sex-matched non-PTSD adult survivors who experienced similar stressors. High resolution T1 weighted images were obtained via a GE 3.0 T scanner. The structural data was automatically segmented using FreeSurfer software, and volume of whole hippocampus and subfield including CA1, CA2-3, CA4-DG, fimbria, presubiculum, subiculum and fissure were extracted. Volume differences between the two groups were statistically compared with age, years of education, duration from the events and intracranial volume (ICV) as covariates. Hemisphere, sex and diagnosis were entered as fixed factors. Relationship between morphometric measurements with Clinician-Administered PTSD Scale (CAPS) score and illness duration were performed using Pearson's correlation with SPSS. Comparing to TC, PTSD patients showed no statistically significant alteration in volumes of the whole hippocampus and all the subfields ( P > 0.05). In male patients, there were significant correlations between CAPS score and volume of right CA2-3 ( R 2 = 0.197, P = 0.034), right subiculum ( R 2 = 0.245, P = 0.016), and duration statistically correlated with right fissure ( R 2 = 0.247, P = 0.016). In female patients, CAPS scores significant correlated with volume of left presubiculum ( R 2 = 0.095, P = 0.042), left subiculum ( R 2 = 0.090, P = 0.048), and left CA4-DG ( R 2 = 0.099, P = 0.037). The main findings of the current study suggest that stress event causes non-selective damage to hippocampus in both PTSD patients and TC, and gender-specific lateralization may underlie PTSD pathology.

  10. Transcriptional and Physiological Characterizations of Escherichia coli MG1655 that have been grown under Low Shear Stress Environment for 1000 Generations

    Science.gov (United States)

    Karouia, Fathi; Tirumalai, Madhan R.; Nelman-Gonzalez, Mayra A.; Sams, Clarence F.; Ott, Mark C.; Pierson, Duane L.; Fofanov, Yuriy; Willson, Richard C.; Fox, George E.

    significantly differently. Fimbriae encoding genes were significantly up-regulated whereas genes encoding the flagellar motor complex were down-regulated. Additionally, 81 significantly expressed genes have been implicated in and/or associated with biofilm formation. The remaining up-regulated genes seemed to be involved in a response that triggered expression of genes associated with the type II secretion complex. This complex has been involved in virulence factors and members of the multidrug efflux system which confer resistance to a multitude of antimicrobial agents and antibiotics. Biofilm formation and the aggregation of cells were evaluated by scanning electron microscopy (SEM). The analysis revealed that extracellular matrix and complex cellular networking were present among cells that were exposed to the long-term LSMMG environment. In addition the response to a variety of stresses and antibiotics were examined. Significant differences were seen between long-term exposure to LSMMG and the short-term control. Changes in expression may predispose the cells to more efficiently attach to surfaces and/or other cells and thereby confer resistance to antibiotics. Future studies will seek to determine the extent to which the long-term adaptation is influenced by genomic changes. These studies will contribute to the knowledge base needed to develop countermeasures that will decrease the risks associated with astronaut health and mission integrity that are presented by microorganisms.

  11. Multiplex real-time PCR assay for detection of Escherichia coli O157:H7 and screening for non-O157 Shiga toxin-producing E. coli.

    Science.gov (United States)

    Li, Baoguang; Liu, Huanli; Wang, Weimin

    2017-11-09

    Shiga toxin-producing Escherichia coli (STEC), including E. coli O157:H7, are responsible for numerous foodborne outbreaks annually worldwide. E. coli O157:H7, as well as pathogenic non-O157:H7 STECs, can cause life-threating complications, such as bloody diarrhea (hemolytic colitis) and hemolytic-uremic syndrome (HUS). Previously, we developed a real-time PCR assay to detect E. coli O157:H7 in foods by targeting a unique putative fimbriae protein Z3276. To extend the detection spectrum of the assay, we report a multiplex real-time PCR assay to specifically detect E. coli O157:H7 and screen for non-O157 STEC by targeting Z3276 and Shiga toxin genes (stx1 and stx2). Also, an internal amplification control (IAC) was incorporated into the assay to monitor the amplification efficiency. The multiplex real-time PCR assay was developed using the Life Technology ABI 7500 System platform and the standard chemistry. The optimal amplification mixture of the assay contains 12.5 μl of 2 × Universal Master Mix (Life Technology), 200 nM forward and reverse primers, appropriate concentrations of four probes [(Z3276 (80 nM), stx1 (80 nM), stx2 (20 nM), and IAC (40 nM)], 2 μl of template DNA, and water (to make up to 25 μl in total volume). The amplification conditions of the assay were set as follows: activation of TaqMan at 95 °C for 10 min, then 40 cycles of denaturation at 95 °C for 10 s and annealing/extension at 60 °C for 60 s. The multiplex assay was optimized for amplification conditions. The limit of detection (LOD) for the multiplex assay was determined to be 200 fg of bacterial DNA, which is equivalent to 40 CFU per reaction which is similar to the LOD generated in single targeted PCRs. Inclusivity and exclusivity determinants were performed with 196 bacterial strains. All E. coli O157:H7 (n = 135) were detected as positive and all STEC strains (n = 33) were positive for stx1, or stx2, or stx1 and stx2 (Table 1). No cross reactivity was detected with Salmonella

  12. Virulence markers associated with Trueperella pyogenes infections in livestock and companion animals.

    Science.gov (United States)

    Risseti, R M; Zastempowska, E; Twarużek, M; Lassa, H; Pantoja, J C F; de Vargas, A P C; Guerra, S T; Bolaños, C A D; de Paula, C L; Alves, A C; Colhado, B S; Portilho, F V R; Tasca, C; Lara, G H B; Ribeiro, M G

    2017-08-01

    Trueperella pyogenes is an opportunistic pathogen that causes diverse pyogenic infections in livestock. The genes that encode the exotoxin pyolysin (plo) and other putative factors that promote adhesion of pathogen to host cells (fimbriae fimA, fimC, fimE, fimG, neuraminidases nanH, nanP, and collagen-binding protein cbpA) have been associated with virulence, particularly in mastitis and uterus infections of dairy cows. However, the role of these virulence markers in the pathogenicity of the agent in domestic animals infections still is incompletely understood. The genes plo, fimA, fimC, fimE, fimG, nanH, nanP, and cbpA were investigated in 71 T. pyogenes strains recovered from cattle, sheep, goats, dogs, equines, and a pig, recovered from mastitis (n = 35), and non-mastitis (n = 36) cases (abscesses, reproductive tract diseases, pneumonia, lymphadenitis, encephalitis). The most common genes harboured by the isolates were: plo (71/71 = 100·0%), fimA (70/71 = 98·6%), nanP (56/71 = 78·9%), fimE (53/71 = 74·6%), fimC (46/71 = 64·8%) and nanH (45/71 = 63·4%), whereas cbpA (6/71 = 8·4%) and fimG (4/71 = 5·6%) were uncommon. The most frequent genotypes were plo/fimA/fimE/fimC/nanH/nanP (17/71 = 23·9%), plo/fimA/fimE/nanH/nanP (13/71 = 18·3%), and plo/fimA/fimE/fimC/nanP (11/71 = 15·5%). No association was observed between the presence of genes vs clinical signs or host species. To the best of our knowledge, this is the first report on aforementioned virulence factors of pathogen detected in diseased horses and dogs. The role of particular virulence factors of Trueperella pyogenes that determine different pyogenic infections among domestic animals is poorly understood. Eight putative virulence genes and genotype profiles of 71 isolates were investigated among different clinical manifestations in domestic animals. The most common genes were plo (71/71 = 100·0%), fimA (70/71 = 98·6%), nanP (56/71 = 78·9%), fimE (53/71 = 74·6

  13. Comparative Genomics of the Ubiquitous, Hydrocarbon-degrading Genus Marinobacter

    Science.gov (United States)

    Singer, E.; Webb, E.; Edwards, K. J.

    2012-12-01

    using fimbriae and pili. Formation of biofilm with biosurfactant characteristics has been observed in Marinobacter cultures and environmental strains in relation to hydrocarbon degradation. Genomic potential exists for the synthesis of biofilm-related carbon and energy storage compounds, e.g. alginate and isoprenoid wax esters, and quorum sensing encoded by the regulatory luxR gene and N-acyl-L-homoserine lactone (AHL) signals. Halotolerance is predicted to be achieved through biosynthesis and/or import of compatible solutes, including glycine betaine, choline, ectoine, sucrose, periplasmic glucans as well as membrane channel activity regulating intracellular sodium, potassium and chloride concentration balance. Gene abundances concur with those observed in sequenced halophilic Halomonas genomes. Defense mechanisms are plentiful and include arsenate, organic solvent, copper, and mercuric resistance, compounds, which frequently occur in oil refinery wastewater. The Marinobacter genomes reflect dynamic environments and diverse interactions with viruses and other bacteria with similar metabolic strategies, as reflected by the large number of integrases and transposases. This study has provided comprehensive genomic insights into the metabolic versatility and predicted environmental impact potential of one of the most ubiquitous bacterial genera.

  14. A direct morphometric comparison of five labeling protocols for multi-atlas driven automatic segmentation of the hippocampus in Alzheimer's disease.

    Science.gov (United States)

    Nestor, Sean M; Gibson, Erin; Gao, Fu-Qiang; Kiss, Alex; Black, Sandra E

    2013-02-01

    anatomy and dorsal white matter compartments furnish the best voxel-overlap accuracies (Dice Similarity Coefficient=0.87-0.89), compared to expert manual tracings, and achieve the smallest sample sizes required to power clinical trials in MCI and AD. The greatest distribution of errors was localized to the caudal hippocampus and the alveus-fimbria compartment when these regions were excluded. The definition of the medial body did not significantly alter accuracy among more comprehensive protocols. Voxel-overlap accuracies between automatic and manual labels were lower for the more pathologically heterogeneous Sunnybrook study in comparison to the ADNI-1 sample. Finally, accuracy among protocols appears to significantly differ the most in AD subjects compared to MCI and normal elders. Together, these results suggest that selection of a candidate protocol for fully automatic multi-template based segmentation in AD can influence both segmentation accuracy when compared to expert manual labels and performance as a biomarker in MCI and AD. Copyright © 2012 Elsevier Inc. All rights reserved.

  15. Occurrence of Proteus mirabilis associated with two species of venezuelan oysters Ocorrência de Proteus mirabilis associado a duas espécies de ostras venezuelanas

    Directory of Open Access Journals (Sweden)

    Milagro Fernández-Delgado

    2007-12-01

    Full Text Available The fecal contamination of raw seafood by indicators and opportunistic pathogenic microorganisms represents a public health concern. The objective of this study was to investigate the presence of enteric bacteria colonizing oysters collected from a Venezuelan touristic area. Oyster samples were collected at the northwestern coast of Venezuela and local salinity, pH, temperature, and dissolved oxygen of seawater were recorded. Total and fecal coliforms were measured for the assessment of the microbiological quality of water and oysters, using the Multiple Tube Fermentation technique. Analyses were made using cultures and 16S rRNA gene sequencing. Diverse enrichment and selective culture methods were used to isolate enteric bacteria. We obtained pure cultures of Gram-negative straight rods with fimbriae from Isognomon alatus and Crassostrea rhizophorae. Our results show that P. mirabilis was predominant under our culture conditions. We confirmed the identity of the cultures by biochemical tests, 16S rRNA gene sequencing, and data analysis. Other enterobacteria such as Escherichia coli, Morganella morganii and Klebsiella pneumoniae were also isolated from seawater and oysters. The presence of pathogenic bacteria in oysters could have serious epidemiological implications and a potential human health risk associated with consumption of raw seafood.A contaminação fecal de frutos do mar crus por microrganismos oportunistas patogênicos representa problema de saúde pública. O objetivo deste estudo é investigar a presença de bactérias entéricas que colonizam ostras coletadas em área turística da Venezuela. Amostras de ostras foram coletadas na costa noroeste da Venezuela e foram registrados a salinidade local, pH, temperatura e o oxigênio dissolvido na água do mar. O total de coliformes fecais foi medido para a avaliação da qualidade microbiológica da água e das ostras, usando a técnica de fermentação em tubos múltiplos. An

  16. Some virulence genes of Escherichia coli isolated from cloacal swabs of healthy Alagoas Curassows (Pauxi mitu in Brazil Alguns genes de virulência de Escherichia coli isoladas de mutuns-do-nordeste (Pauxi mitu sadios no Brasil

    Directory of Open Access Journals (Sweden)

    André A.B. Saidenberg

    2013-04-01

    Full Text Available Birds of the Cracidae family (curassows, guans, and chachalacas are endemic of the Neotropics and 50 species are currently classified. Brazil has 22 species, seven of which are considered threatened. The Alagoas Curassow (Pauxi mitu species is considered extinct in the wild; but about 120 birds are alive in captivity. Conservation of this species depends entirely on correct management. Health reports of both wildlife and captive curassows are rare. In this study the presence of Escherichia coli was evaluated in 23 healthy Alagoas Curassows from two private breeding centres. E. coli was isolated from cloacal swabs, and the presence of genes encoding cytotoxic necrotising factor 1 (cnf1, alpha-haemolysin (hly, aerobactin (iuc, serum resistance (iss and the following adhesions: S fimbriae (sfa, pili associated with pyelonephritis (pap and temperature-sensitive haemagglutinin (tsh were investigated. E. coli was isolated from 78.3% (18/23 of the birds, and the percentage of curassows colonized by E. coli was similar between the two facilities. From the 22 E. coli isolates, 15 (68.2% were positive for at least one virulence factor by PCR, and the most frequently found gene was iss (50%. No curassows had clinical signs of disease. Nevertheless, the presence of some E. coli strains may be a concern to the wildlife in captivity. Additional health surveillance studies are essential to guarantee successful conservation programmes for threatened cracids in Brazil.Aves da família Cracidae (mutuns, jacutingas e aracuãs são endêmicas da região Neotropical com 50 espécies atualmente classificadas. O Brasil possui 22 espécies nesta família e sete delas são consideradas ameaçadas de extinção. O mutum-do-nordeste (Pauxi mitu é considerado extinto na natureza, no entanto, aproximadamente 120 indivíduos são mantidos em cativeiro. A conservação desta espécie depende inteiramente de um manejo correto. Informações sobre o status sanitário de mutuns

  17. Radionuclides in fishes and mussels from the Farallon Islands Nuclear Waste Dump Site, California.

    Science.gov (United States)

    Suchanek, T H; Lagunas-Solar, M C; Raabe, O G; Helm, R C; Gielow, F; Peek, N; Carvacho, O

    1996-08-01

    The Farallon Islands Nuclear Waste Dump Site (FINWDS), approximately 30 miles west of San Francisco, California, received at least 500 TBq encapsulated in more than 47,500 containers from approximately 1945 to 1970. During several seasons in 1986/87 deep-sea bottom feeding fishes (Dover sole = Microstomus pacificus; sablefish = Anoplopoma fimbria; thornyheads = Sebastolobus spp.) and intertidal mussels (Mytilus californianus) were collected from the vicinity of the FINWDS and from comparable depths at a reference site near Point Arena, CA. Tissues were analyzed for several radionuclides (137Cs, 238Pu, 239+240Pu, and 241Am). Radionuclide concentrations for fish mussel tissue ranged from non-detectable to 4,340 mBq kg(-1) wet weight, with the following means for Farallon fishes: 137Cs = 1,110 mBq kg(-1); 238Pu = 390 mBq kg(-1); 239+240Pu = 130 mBq kg(-1); and 241Am = 1,350 mBq kg(-1). There were no statistically significant differences in the radionuclide concentrations observed in samples from the Farallon Islands compared to reference samples from Point Arena, CA. Concentrations of both 238Pu and 241Am in fish tissues (from both sites) were notably higher than those reported in literature from any other sites world-wide, including potentially contaminated sites. Concentrations of 239+24OPu from both sites were typical of low values found at some contaminated sites worldwide. These results show approximately 10 times higher concentrations of 239+240Pu and approximately 40-50 times higher concentrations of 238Pu than those values reported for identical fish species from 1977 collections at the FINWDS. Radionuclide concentrations were converted to a hypothetical per capita annual radionuclide intake for adults, yielding the following values of annual Committed Effective Dose Equivalent (CEDE) from ionizing radiation emitted from these radionuclides: 0.000 mSv y(-1) for 137Cs, 0.009 mSv Y(-1) for 228Pu, and 0.003 mSv y(-1) for 239+240Pu. For 241Am, projected CEDE for

  18. Sorbitol-Fermenting Enterohemorrhagic Escherichia coli O157:H- Isolates from Czech Patients with Novel Plasmid Composition Not Previously Seen in German Isolates.

    Science.gov (United States)

    Bauwens, Andreas; Marejková, Monika; Middendorf-Bauchart, Barbara; Prager, Rita; Kossow, Annelene; Zhang, Wenlan; Karch, Helge; Mellmann, Alexander; Bielaszewska, Martina

    2017-12-01

    Sorbitol-fermenting (SF) enterohemorrhagic Escherichia coli (EHEC) O157:H - strains, first identified in Germany, have emerged as important pathogens throughout Europe. Besides chromosomally encoded Shiga toxin 2a (the major virulence factor), several putative virulence loci, including the hly , etp , and sfp operons, encoding EHEC hemolysin, type II secretion system proteins, and Sfp fimbriae, respectively, are located on the 121-kb plasmid pSFO157 in German strains. Here we report novel SF EHEC O157:H - strains isolated from patients in the Czech Republic. These strains share the core genomes and chromosomal virulence loci encoding toxins ( stx 2a and the cdtV -ABC operon) and adhesins ( eae -γ, efa1 , lpfA O157OI-141 , and lpfA O157OI-154 ) with German strains but differ essentially in their plasmids. In contrast to all previously detected SF EHEC O157:H - strains, the Czech strains carry two plasmids, of 79 kb and 86 kb. The 79-kb plasmid harbors the sfp operon, but neither of the plasmids contains the hly and etp operons. Sequence analyses demonstrated that the 79-kb plasmid (pSFO157 258/98-1) evolved from pSFO157 of German strains by deletion of a 41,534-bp region via homologous recombination, resulting in loss of the hly and etp operons. The 86-kb plasmid (pSFO157 258/98-2) displays 98% sequence similarity to a 92.7-kb plasmid of an extraintestinal pathogenic E. coli bloodstream isolate. Our finding of this novel plasmid composition in SF EHEC O157:H - strains extends the evolutionary history of EHEC O157 plasmids. Moreover, the unique molecular plasmid characteristics permit the identification of such strains, thereby facilitating further investigations of their geographic distribution, clinical significance, and epidemiology. IMPORTANCE Since their first identification in Germany in 1989, sorbitol-fermenting enterohemorrhagic Escherichia coli O157:H - (nonmotile) strains have emerged as important causes of the life-threatening disease hemolytic

  19. Sorbitol-Fermenting Enterohemorrhagic Escherichia coli O157:H− Isolates from Czech Patients with Novel Plasmid Composition Not Previously Seen in German Isolates

    Science.gov (United States)

    Bauwens, Andreas; Marejková, Monika; Middendorf-Bauchart, Barbara; Prager, Rita; Kossow, Annelene; Zhang, Wenlan; Karch, Helge

    2017-01-01

    ABSTRACT Sorbitol-fermenting (SF) enterohemorrhagic Escherichia coli (EHEC) O157:H− strains, first identified in Germany, have emerged as important pathogens throughout Europe. Besides chromosomally encoded Shiga toxin 2a (the major virulence factor), several putative virulence loci, including the hly, etp, and sfp operons, encoding EHEC hemolysin, type II secretion system proteins, and Sfp fimbriae, respectively, are located on the 121-kb plasmid pSFO157 in German strains. Here we report novel SF EHEC O157:H− strains isolated from patients in the Czech Republic. These strains share the core genomes and chromosomal virulence loci encoding toxins (stx2a and the cdtV-ABC operon) and adhesins (eae-γ, efa1, lpfAO157OI-141, and lpfAO157OI-154) with German strains but differ essentially in their plasmids. In contrast to all previously detected SF EHEC O157:H− strains, the Czech strains carry two plasmids, of 79 kb and 86 kb. The 79-kb plasmid harbors the sfp operon, but neither of the plasmids contains the hly and etp operons. Sequence analyses demonstrated that the 79-kb plasmid (pSFO157 258/98-1) evolved from pSFO157 of German strains by deletion of a 41,534-bp region via homologous recombination, resulting in loss of the hly and etp operons. The 86-kb plasmid (pSFO157 258/98-2) displays 98% sequence similarity to a 92.7-kb plasmid of an extraintestinal pathogenic E. coli bloodstream isolate. Our finding of this novel plasmid composition in SF EHEC O157:H− strains extends the evolutionary history of EHEC O157 plasmids. Moreover, the unique molecular plasmid characteristics permit the identification of such strains, thereby facilitating further investigations of their geographic distribution, clinical significance, and epidemiology. IMPORTANCE Since their first identification in Germany in 1989, sorbitol-fermenting enterohemorrhagic Escherichia coli O157:H− (nonmotile) strains have emerged as important causes of the life-threatening disease hemolytic

  20. Long-Term Evolution Studies of E. Coli under Combined Effects of Simulated Microgravity and Antibiotic.

    Science.gov (United States)

    Karouia, Fathi; Tirumalai, Madhan R.; Ott, Mark C.; Pierson, Duane L.; Fox, George E.; Tran, Quyen

    2016-07-01

    Multiple spaceflight and simulated microgravity experiments have shown changes in phenotypic microbial characteristics such as microbial growth, morphology, metabolism, genetic transfer, antibiotic and stress susceptibility, and an increase in virulence factors. However, while these studies have contributed to expand our understanding of the short-term effects of spaceflight or simulated microgravity on biological systems, it remains unclear the type of responses subsequent to long-term exposure to space environment and microgravity in particular. As such, organisms exposed to the space environment for extended periods of time may evolve in unanticipated ways thereby negatively impacting long duration space missions. We report here for the first time, an experimental study of microbial evolution in which the effect of long-term exposure to Low Shear Modeled MicroGravity (LSMMG) on microbial gene expression and physiology in Escherichia coli (E. coli) MG1655 was examined using functional genomics, and molecular techniques with and without simultaneous exposure to broad spectrum antibiotic chloramphenicol. E. coli cells were grown under simulated microgravity for 1000 generations in High Aspect Ratio Vessels (HARVs) that were either heat-sterilized (115 deg C, 15 min) or by using/rinsing the HARVs with a saturated solution of the broad-spectrum antibiotic chloramphenicol. In the case of the cells evolved using the antibiotic sterilized HARVs, the expression levels of 357 genes were significantly changed. In particular, fimbriae encoding genes were significantly up-regulated whereas genes encoding the flagellar motor complex were down-regulated. Re-sequencing of the genome revealed that a number of the flagellar genes were actually deleted. The antibiotic resistance levels of the evolved strains were analyzed using VITEK analyzer. The evolved strain was consistently resistant to the antibiotics used (viz., Ampicillin, Cefalotin, Cefurox-ime, Cefuroxime Axetil

  1. THE FORMATION OF BIOFILMS BY PSEUDOMONAS AERUGINOSA STRAINS, AND METHODS OF ITS CONTROL (REVIEW

    Directory of Open Access Journals (Sweden)

    Sarkis-Ivanova VV

    2017-03-01

    investigate this problem the main source of nosocomial diseases and persistence factor of their causative agents in hospital ecosystems from air and water to inner surfaces of catheters and body systems are represented by biofilms. Bacteria of P. aeruginosa type are human opportunistic pathogens which being a part of biofilm may cause different nosological forms of pyoinflammatory diseases with severe course and high fatality in immunocompromised patients. These microorganisms are able to contaminate external and internal surfaces of catheters, probes, respiratory tubes, lenses, and form biofilm on them. The given information determines reasonability of searching the methods of control of biofilm production in P. aeruginosa cultures which can be used to increase the effectiveness of antibiotic treatment in blue pus infection and/or decontamination of medical equipment, and another objects of hospital environment. One of the methods to control biofilms produced by P. aeruginosa on abiotic surfaces from different materials is impact of antiseptics and decontaminants. Modern decontaminants are represented by a wide spectrum of chemical compounds of different classes which have a common ability to destroy microorganisms even in low concentrations. At present the impact of bacteria on separate elements of biofilm formation is considered to be one of the most perspective targets for the action of new antimicrobial medicines. Numerous investigations of recent 40 years demonstrated that the process of biofilm formation is complex and multistage. Currently the principal directions of development of new antimicrobial preparations are the following: development of antiadhesive coatings and preparations able to interrupt the function of eternal structures of cells in charge of adhesion (fimbriae, adhesins; development of preparations that block synthesis or destabilize matrix of biofilm; development of preparations that cause cell adhesion deficiency in microcolony, and block cell