WorldWideScience

Sample records for plasmid conjugal transfer

  1. Plasmid transfer by conjugation in Xylella fastidiosa.

    Science.gov (United States)

    Recombination and horizontal gene transfer have been implicated in the adaption of Xylella fastidiosa (Xf) to infect a wide variety of different plant species. There is evidence that certain strains of Xf carry native plasmids equipped with transfer and mobilization genes, suggesting conjugation as ...

  2. Conjugal plasmid transfer (pAM beta 1) in Lactobacillus plantarum.

    OpenAIRE

    Shrago, A W; Chassy, B M; Dobrogosz, W J

    1986-01-01

    The streptococcal plasmid pAM beta 1 (erythromycin resistance) was transferred via conjugation from Streptococcus faecalis to Lactobacillus plantarum and was transferred among L. plantarum strains. Streptococcus sanguis Challis was transformed with pAM beta 1 isolated from these transconjugants, and transformants harboring intact pAM beta 1 could conjugate the plasmid back to L. plantarum.

  3. Effect of TiO2 on conjugative transfer of RP4 plasmid

    International Nuclear Information System (INIS)

    Qian Di; Zhang Buchang; Yang Dong; Chen Zhaoli; Jin Min; Qiu Zhigang; Li Junwen

    2013-01-01

    Objective: To explore the effect and law of nano-titanium dioxide on the conjugative transfer of RP4 plasmid. Methods: Mating was conducted between Escherichia coli HB101 (RP4) and E. coli K12Rif in saline without stirring under certain conditions and the donor per recipient ratio was 1:1 constantly. The selective LB agar medium plates containing appropriate antibiotics were used to count the number of transconjugants and the conjugative transfer frequency. Results: Nano-titanium dioxide could promote the conjugative transfer of RP4. The nano-titanium dioxide concentration, bacterial concentration, mating temperature and mating time could affect the conjugative transfer of RP4. Conclusion: Nano-titanium dioxide can promote plasmid conjugal transfer in the liquid phase under certain conditions, which may pose a potential hazard to environmental and human health. (authors)

  4. Conjugative Plasmid Transfer in Xylella fastidiosa Is Dependent on tra and trb Operon Functions

    OpenAIRE

    Burbank, Lindsey P.; Van Horn, Christopher R.

    2017-01-01

    The insect-transmitted plant pathogen Xylella fastidiosa is capable of efficient horizontal gene transfer (HGT) and recombination. Natural transformation occurs at high rates in X. fastidiosa, but there also is evidence that certain strains of X. fastidiosa carry native plasmids equipped with transfer and mobilization genes, suggesting conjugation as an additional mechanism of HGT in some instances. Two operons, tra and trb, putatively encoding a conjugative type IV secretion system, are foun...

  5. Conjugative Plasmid Transfer in Xylella fastidiosa Is Dependent on tra and trb Operon Functions.

    Science.gov (United States)

    Burbank, Lindsey P; Van Horn, Christopher R

    2017-11-01

    The insect-transmitted plant pathogen Xylella fastidiosa is capable of efficient horizontal gene transfer (HGT) and recombination. Natural transformation occurs at high rates in X. fastidiosa , but there also is evidence that certain strains of X. fastidiosa carry native plasmids equipped with transfer and mobilization genes, suggesting conjugation as an additional mechanism of HGT in some instances. Two operons, tra and trb , putatively encoding a conjugative type IV secretion system, are found in some but not all X. fastidiosa isolates, often on native plasmids. X. fastidiosa strains that carry the conjugative transfer genes can belong to different subspecies and frequently differ in host ranges. Using X. fastidiosa strain M23 ( X. fastidiosa subsp. fastidiosa ) or Dixon ( X. fastidiosa subsp. multiplex ) as the donor strain and Temecula ( X. fastidiosa subsp. fastidiosa ) as the recipient strain, plasmid transfer was characterized using the mobilizable broad-host-range vector pBBR5pemIK. Transfer of plasmid pBBR5pemIK was observed under in vitro conditions with both donor strains and was dependent on both tra and trb operon functions. A conjugative mechanism likely contributes to gene transfer between diverse strains of X. fastidiosa , possibly facilitating adaptation to new environments or different hosts. IMPORTANCE Xylella fastidiosa is an important plant pathogen worldwide, infecting a wide range of different plant species. The emergence of new diseases caused by X. fastidiosa , or host switching of existing strains, is thought to be primarily due to the high frequency of HGT and recombination in this pathogen. Transfer of plasmids by a conjugative mechanism enables movement of larger amounts of genetic material at one time, compared with other routes of gene transfer such as natural transformation. Establishing the prevalence and functionality of this mechanism in X. fastidiosa contributes to a better understanding of HGT, adaptation, and disease emergence

  6. TOL plasmid transfer during bacterial conjugation in vitro and rhizoremediation of oil compounds in vivo

    International Nuclear Information System (INIS)

    Jussila, Minna M.; Zhao, Ji; Suominen, Leena; Lindstroem, Kristina

    2007-01-01

    Molecular profiling methods for horizontal transfer of aromatics-degrading plasmids were developed and applied during rhizoremediation in vivo and conjugations in vitro. pWW0 was conjugated from Pseudomonas to Rhizobium. The xylE gene was detected both in Rhizobium galegae bv. officinalis and bv. orientalis, but it was neither stably maintained in orientalis nor functional in officinalis. TOL plasmids were a major group of catabolic plasmids among the bacterial strains isolated from the oil-contaminated rhizosphere of Galega orientalis. A new finding was that some Pseudomonas migulae and Pseudomonas oryzihabitans strains harbored a TOL plasmid with both pWW0- and pDK1-type xylE gene. P. oryzihabitans 29 had received the archetypal TOL plasmid pWW0 from Pseudomonas putida PaW85. As an application for environmental biotechnology, the biodegradation potential of oil-polluted soil and the success of bioremediation could be estimated by monitoring changes not only in the type and amount but also in transfer of degradation plasmids. - Horizontal transfer of degradation plasmids in the oil-contaminated rhizosphere reveals the dynamic nature of the intrinsic biodegradation potential

  7. Transfer of conjugative plasmids among bacteria under environmentally relevant conditions

    DEFF Research Database (Denmark)

    Musovic, Sanin

    Mobile genetiske elementer (f.eks. plasmider), der ofte bærer ekstra funktioner såsom antibiotikaresistens, eller kataboliske- og xenobiotiske nedbrydnings gener, antages at have en meget vigtigt evolutionær rolle for bakterier. I denne PhD afhandling undersøgte jeg størrelsen af plasmid overførs...

  8. Quorum-Dependent Mannopine-Inducible Conjugative Transfer of an Agrobacterium Opine-Catabolic Plasmid

    Science.gov (United States)

    Wetzel, Margaret E.; Kim, Kun-Soo; Miller, Marilyn; Olsen, Gary J.

    2014-01-01

    The Ti plasmid in Agrobacterium tumefaciens strain 15955 carries two alleles of traR that regulate conjugative transfer. The first is a functional allele, called traR, that is transcriptionally induced by the opine octopine. The second, trlR, is a nonfunctional, dominant-negative mutant located in an operon that is inducible by the opine mannopine (MOP). Based on these findings, we predicted that there exist wild-type agrobacterial strains harboring plasmids in which MOP induces a functional traR and, hence, conjugation. We analyzed 11 MOP-utilizing field isolates and found five where MOP induced transfer of the MOP-catabolic element and increased production of the acyl-homoserine lactone (acyl-HSL) quormone. The transmissible elements in these five strains represent a set of highly related plasmids. Sequence analysis of one such plasmid, pAoF64/95, revealed that the 176-kb element is not a Ti plasmid but carries genes for catabolism of MOP, mannopinic acid (MOA), agropinic acid (AGA), and the agrocinopines. The plasmid additionally carries all of the genes required for conjugative transfer, including the regulatory genes traR, traI, and traM. The traR gene, however, is not located in the MOP catabolism region. The gene, instead, is monocistronic and located within the tra-trb-rep gene cluster. A traR mutant failed to transfer the plasmid and produced little to no quormone even when grown with MOP, indicating that TraRpAoF64/95 is the activator of the tra regulon. A traM mutant was constitutive for transfer and acyl-HSL production, indicating that the anti-activator function of TraM is conserved. PMID:24363349

  9. Treatment with Cefotaxime Affects Expression of Conjugation Associated Proteins and Conjugation Transfer Frequency of an IncI1 Plasmid in Escherichia coli

    DEFF Research Database (Denmark)

    Møller, Thea S B; Liu, Gang; Boysen, Anders

    2017-01-01

    research suggests that the effect of antibiotic treatment on plasmid conjugation frequencies, and hence the spread of resistance plasmids, may have been overestimated. We addressed the question by quantifying transfer proteins and conjugation frequencies of a blaCTX-M-1 encoding IncI1 resistance plasmid....... The frequency of plasmid conjugation, measured in an antibiotic free environment, increased significantly when the donor was pre-grown in broth containing CTX compared to growth without this drug, regardless of whether blaCTX-M-1 was located on the plasmid or in trans on the chromosome. The results shows...

  10. Interplay of a non-conjugative integrative element and a conjugative plasmid in the spread of antibiotic resistance via suicidal plasmid transfer from an aquaculture Vibrio isolate.

    Science.gov (United States)

    Nonaka, Lisa; Yamamoto, Tatsuya; Maruyama, Fumito; Hirose, Yuu; Onishi, Yuki; Kobayashi, Takeshi; Suzuki, Satoru; Nomura, Nobuhiko; Masuda, Michiaki; Yano, Hirokazu

    2018-01-01

    The capture of antimicrobial resistance genes (ARGs) by mobile genetic elements (MGEs) plays a critical role in resistance acquisition for human-associated bacteria. Although aquaculture environments are recognized as important reservoirs of ARGs, intra- and intercellular mobility of MGEs discovered in marine organisms is poorly characterized. Here, we show a new pattern of interspecies ARGs transfer involving a 'non-conjugative' integrative element. To identify active MGEs in a Vibrio ponticus isolate, we conducted whole-genome sequencing of a transconjugant obtained by mating between Escherichia coli and Vibrio ponticus. This revealed integration of a plasmid (designated pSEA1) into the chromosome, consisting of a self-transmissible plasmid backbone of the MOBH group, ARGs, and a 13.8-kb integrative element Tn6283. Molecular genetics analysis suggested a two-step gene transfer model. First, Tn6283 integrates into the recipient chromosome during suicidal plasmid transfer, followed by homologous recombination between the Tn6283 copy in the chromosome and that in the newly transferred pSEA1. Tn6283 is unusual among integrative elements in that it apparently does not encode transfer function and its excision barely generates unoccupied donor sites. Thus, its movement is analogous to the transposition of insertion sequences rather than to that of canonical integrative and conjugative elements. Overall, this study reveals the presence of a previously unrecognized type of MGE in a marine organism, highlighting diversity in the mode of interspecies gene transfer.

  11. Enterococcus faecalis Sex Pheromone cCF10 Enhances Conjugative Plasmid Transfer In Vivo.

    Science.gov (United States)

    Hirt, Helmut; Greenwood-Quaintance, Kerryl E; Karau, Melissa J; Till, Lisa M; Kashyap, Purna C; Patel, Robin; Dunny, Gary M

    2018-02-13

    Cell-cell communication mediated by peptide pheromones (cCF10 [CF]) is essential for high-frequency plasmid transfer in vitro in Enterococcus faecalis To examine the role of pheromone signaling in vivo , we established either a CF-producing (CF+) recipient or a recipient producing a biologically inactive variant of CF (CF- recipient) in a germfree mouse model 3 days before donor inoculation and determined transfer frequencies of the pheromone-inducible plasmid pCF10. Plasmid transfer was detected in the upper and middle sections of the intestinal tract 5 h after donor inoculation and was highly efficient in the absence of antibiotic selection. The transconjugant/donor ratio reached a maximum level approaching 1 on day 4 in the upper intestinal tract. Plasmid transfer was significantly lower with the CF- recipient. While rescue of the CF- mating defect by coculture with CF+ recipients is easily accomplished in vitro , no extracellular complementation occurred in vivo This suggests that most pheromone signaling in the gut occurs between recipient and donor cells in very close proximity. Plasmid-bearing cells (donors plus transconjugants) steadily increased in the population from 0.1% after donor inoculation to about 10% at the conclusion of the experiments. This suggests a selective advantage of pCF10 carriage distinct from antibiotic resistance or bacteriocin production. Our results demonstrate that pheromone signaling is required for efficient pCF10 transfer in vivo In the absence of CF+ recipients, a low level of transfer to CF- recipients occurred in the gut. This may result from low-level host-mediated induction of the donors in the gastrointestinal (GI) tract, similar to that previously observed in serum. IMPORTANCE Horizontal gene transfer is a major factor in the biology of Enterococcus faecalis , an important nosocomial pathogen. Previous studies showing efficient conjugative plasmid transfer in the gastrointestinal (GI) tracts of experimental animals did

  12. Effects of Metals on Antibiotic Resistance and Conjugal Plasmid Transfer in Soil Bacterial Communities

    DEFF Research Database (Denmark)

    Song, Jianxiao

    Antibiotic resistance currently represents one of the biggest challenges for human health and in recent years the environmental dimension of antibiotic resistance has been increasingly recognized. The soil environment serves as an important reservoir of antibiotic resistance determinants. In addi...... adaptation to metal stress did not significantly increase the permissiveness of the soil bacterial community towards conjugal plasmid transfer........ In addition to direct selection of antibiotic resistance by antibiotics, metals may co-select for antibiotic resistance via different mechanisms causing environmental selection of antibiotic resistance in metal contaminated soils. Horizontal gene transfer of mobile genetic elements (MGEs) like plasmids...... is generally considered one of the most important co-selection mechanisms as multiple resistance genes can be located on the same MGE. This PhD thesis focused on the impact of metals (Cu and Zn) on the development of antibiotic resistance in bacterial communities in soils exposed to different degrees...

  13. Conjugal transfer of plasmid pAM beta 1 in Lactobacillus reuteri and between lactobacilli and Enterococcus faecalis.

    OpenAIRE

    Tannock, G W

    1987-01-01

    The broad-host-range plasmid pAM beta 1 (erythromycin resistance) was transferred conjugally from Streptococcus lactis to Lactobacillus reuteri, L. murinus, and L. fermentum. Transfer of pAM beta 1 between two L. reuteri strains occurred, and lactobacillus transconjugants could act as donors of pAM beta 1 in crosses with Enterococcus faecalis JH2-2.

  14. Frequent conjugative transfer accelerates adaptation of a broad-host-range plasmid to an unfavorable Pseudomonas putida host.

    Science.gov (United States)

    Heuer, Holger; Fox, Randal E; Top, Eva M

    2007-03-01

    IncP-1 plasmids are known to be promiscuous, but it is not understood if they are equally well adapted to various species within their host range. Moreover, little is known about their fate in bacterial communities. We determined if the IncP-1beta plasmid pB10 was unstable in some Proteobacteria, and whether plasmid stability was enhanced after long-term carriage in a single host and when regularly switched between isogenic hosts. Plasmid pB10 was found to be very unstable in Pseudomonas putida H2, and conferred a high cost (c. 20% decrease in fitness relative to the plasmid-free host). H2(pB10) was then evolved under conditions that selected for plasmid maintenance, with or without regular plasmid transfer (host-switching). When tested in the ancestral host, the evolved plasmids were more stable and their cost was significantly reduced (9% and 16% for plasmids from host-switched and nonswitched lineages, respectively). Our findings suggest that IncP-1 plasmids can rapidly adapt to an unfavorable host by improving their overall stability, and that regular conjugative transfer accelerates this process.

  15. 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...

  16. Human Intestinal Cells Modulate Conjugational Transfer of Multidrug Resistance Plasmids between Clinical Escherichia coli Isolates

    DEFF Research Database (Denmark)

    Machado, Ana Manuel; Sommer, Morten

    2014-01-01

    Bacterial conjugation in the human gut microbiota is believed to play a major role in the dissemination of antibiotic resistance genes and virulence plasmids. However, the modulation of bacterial conjugation by the human host remains poorly understood and there is a need for controlled systems...... to study this process. We established an in vitro co-culture system to study the interaction between human intestinal cells and bacteria. We show that the conjugation efficiency of a plasmid encoding an extended spectrum beta-lactamase is reduced when clinical isolates of Escherichia coli are co...... of the intestinal cells exposed to bacteria leading to a two-fold reduction in conjugation efficiency. These results show that human gut epithelial cells can modulate bacterial conjugation and may have relevance to gene exchange in the gut....

  17. Enterococcus faecalis Sex Pheromone cCF10 Enhances Conjugative Plasmid Transfer In Vivo

    Directory of Open Access Journals (Sweden)

    Helmut Hirt

    2018-02-01

    Full Text Available Cell-cell communication mediated by peptide pheromones (cCF10 [CF] is essential for high-frequency plasmid transfer in vitro in Enterococcus faecalis. To examine the role of pheromone signaling in vivo, we established either a CF-producing (CF+ recipient or a recipient producing a biologically inactive variant of CF (CF− recipient in a germfree mouse model 3 days before donor inoculation and determined transfer frequencies of the pheromone-inducible plasmid pCF10. Plasmid transfer was detected in the upper and middle sections of the intestinal tract 5 h after donor inoculation and was highly efficient in the absence of antibiotic selection. The transconjugant/donor ratio reached a maximum level approaching 1 on day 4 in the upper intestinal tract. Plasmid transfer was significantly lower with the CF− recipient. While rescue of the CF− mating defect by coculture with CF+ recipients is easily accomplished in vitro, no extracellular complementation occurred in vivo. This suggests that most pheromone signaling in the gut occurs between recipient and donor cells in very close proximity. Plasmid-bearing cells (donors plus transconjugants steadily increased in the population from 0.1% after donor inoculation to about 10% at the conclusion of the experiments. This suggests a selective advantage of pCF10 carriage distinct from antibiotic resistance or bacteriocin production. Our results demonstrate that pheromone signaling is required for efficient pCF10 transfer in vivo. In the absence of CF+ recipients, a low level of transfer to CF− recipients occurred in the gut. This may result from low-level host-mediated induction of the donors in the gastrointestinal (GI tract, similar to that previously observed in serum.

  18. Persistence Mechanisms of Conjugative Plasmids

    DEFF Research Database (Denmark)

    Bahl, Martin Iain; Hansen, Lars H.; Sørensen, Søren Johannes

    2009-01-01

    Are plasmids selfish parasitic DNA molecules or an integrated part of the bacterial genome? This chapter reviews the current understanding of the persistence mechanisms of conjugative plasmids harbored by bacterial cells and populations. The diversity and intricacy of mechanisms affecting the suc...

  19. A study on the role that quorum sensing play in antibiotic-resistant plasmid conjugative transfer in Escherichia coli.

    Science.gov (United States)

    Zhang, Yueheng; Ma, Qingping; Su, Bingmei; Chen, Rui; Lin, Juan; Lin, Zhifen; Wang, Dali; Yu, Yang

    2018-03-01

    Horizontal genes transfer (HGT) plays an important role in the dissemination of antibiotic resistance genes (ARGs) in the environment. However, the mechanisms of HGT of ARGs under the influence of antibiotics in sub-MIC remain rarely explored. Moreover, given its collective nature, HGT was considered to be relative to quorum sensing (QS) system. To investigate whether QS has any impact on horizontal gene transfer of ARGs, experiments were conducted to determine the conjugative efficiency of plasmid RP4 on Escherichia coli (E.coli) under the influences of tetracyclines (TCs), quorum sensing autoinducers (AIs) and quorum sensing inhibitors (QSIs). The results indicated that the sub-MIC TCs could facilitate the conjugative transfer of RP4, a process which could be enhanced by AIs but inhibited by QSIs. This study demonstrated the roles that QS played in the dissemination of ARGs, and provided theoretical insights into the mechanism of HGT of ARGs in the environment.

  20. Evidence for the role of horizontal transfer in generating pVT1, a large mosaic conjugative plasmid from the clam pathogen, Vibrio tapetis.

    Directory of Open Access Journals (Sweden)

    Gaël Erauso

    Full Text Available The marine bacterium Vibrio tapetis is the causative agent of the brown ring disease, which affects the clam Ruditapes philippinarum and causes heavy economic losses in North of Europe and in Eastern Asia. Further characterization of V. tapetis isolates showed that all the investigated strains harbored at least one large plasmid. We determined the sequence of the 82,266 bp plasmid pVT1 from the CECT4600(T reference strain and analyzed its genetic content. pVT1 is a mosaic plasmid closely related to several conjugative plasmids isolated from Vibrio vulnificus strains and was shown to be itself conjugative in Vibrios. In addition, it contains DNA regions that have similarity with several other plasmids from marine bacteria (Vibrio sp., Shewanella sp., Listonella anguillarum and Photobacterium profundum. pVT1 contains a number of mobile elements, including twelve Insertion Sequences or inactivated IS genes and an RS1 phage element related to the CTXphi phage of V. cholerae. The genetic organization of pVT1 underscores an important role of horizontal gene transfer through conjugative plasmid shuffling and transposition events in the acquisition of new genetic resources and in generating the pVT1 modular organization. In addition, pVT1 presents a copy number of 9, relatively high for a conjugative plasmid, and appears to belong to a new type of replicon, which may be specific to Vibrionaceae and Shewanelleacae.

  1. Deciphering conjugative plasmid permissiveness in wastewater microbiomes

    DEFF Research Database (Denmark)

    Jacquiod, Samuel Jehan Auguste; Brejnrod, Asker Daniel; Milani, Stefan Morberg

    2017-01-01

    Wastewater treatment plants (WWTPs) are designed to robustly treat polluted water. They are characterized by ceaseless flows of organic, chemical and microbial matter, followed by treatment steps before environmental release. WWTPs are hotspots of horizontal gene transfer between bacteria via...... still remains largely uncharted. Furthermore, current in vitro methods used to assess conjugation in complex microbiomes do not include in situ behaviours of recipient cells, resulting in partial understanding of transfers. We investigated the in vitro conjugation capacities of WWTP microbiomes from...... inlet sewage and outlet treated water using the broad-host range IncP-1 conjugative plasmid, pKJK5. A thorough molecular approach coupling metagenomes to 16S rRNA DNA/cDNA amplicon sequencing was established to characterize microbiomes using the ecological concept of functional response groups. A broad...

  2. The master regulator of IncA/C plasmids is recognized by the Salmonella Genomic island SGI1 as a signal for excision and conjugal transfer.

    Science.gov (United States)

    Kiss, János; Papp, Péter Pál; Szabó, Mónika; Farkas, Tibor; Murányi, Gábor; Szakállas, Erik; Olasz, Ferenc

    2015-10-15

    The genomic island SGI1 and its variants, the important vehicles of multi-resistance in Salmonella strains, are integrative elements mobilized exclusively by the conjugative IncA/C plasmids. Integration and excision of the island are carried out by the SGI1-encoded site-specific recombinase Int and the recombination directionality factor Xis. Chromosomal integration ensures the stable maintenance and vertical transmission of SGI1, while excision is the initial step of horizontal transfer, followed by conjugation and integration into the recipient. We report here that SGI1 not only exploits the conjugal apparatus of the IncA/C plasmids but also utilizes the regulatory mechanisms of the conjugation system for the exact timing and activation of excision to ensure efficient horizontal transfer. This study demonstrates that the FlhDC-family activator AcaCD, which regulates the conjugation machinery of the IncA/C plasmids, serves as a signal of helper entry through binding to SGI1 xis promoter and activating SGI1 excision. Promoters of int and xis genes have been identified and the binding site of the activator has been located by footprinting and deletion analyses. We prove that expression of xis is activator-dependent while int is constitutively expressed, and this regulatory mechanism is presumably responsible for the efficient transfer and stable maintenance of SGI1. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  3. Plasmid Conjugation in E. coli and Drug Resistance | Igwe ...

    African Journals Online (AJOL)

    This study aimed at determining the antibiotics susceptibility pattern of E. coli isolates claimed to be multidrug resistance using disc diffusion method. It also determined the presence of transferable resistance plasmids through conjugation and evaluated the medical significance of plasmid encoding E. coli and drug ...

  4. Two novel conjugative plasmids from a single strain of Sulfolobus

    NARCIS (Netherlands)

    Erauso, G.; Stedman, K.M.; Werken, van de H.J.G.; Zillig, W.; Oost, van der J.

    2006-01-01

    Two conjugative plasmids (CPs) were isolated and characterized from the same 'Sulfolobus islandicus' strain, SOG2/4, The plasmids were separated from each other and transferred into Sulfolobus soltataricus. One has a high copy number and is not stable (pSOG1) whereas the other has a low copy number

  5. CRISPR-Cas and Restriction-Modification Act Additively against Conjugative Antibiotic Resistance Plasmid Transfer in Enterococcus faecalis

    OpenAIRE

    Price, Valerie J.; Huo, Wenwen; Sharifi, Ardalan; Palmer, Kelli L.

    2016-01-01

    ABSTRACT Enterococcus faecalis is an opportunistic pathogen and a leading cause of nosocomial infections. Conjugative pheromone-responsive plasmids are narrow-host-range mobile genetic elements (MGEs) that are rapid disseminators of antibiotic resistance in the faecalis species. Clustered regularly interspaced short palindromic repeat (CRISPR)-Cas and restriction-modification confer acquired and innate immunity, respectively, against MGE acquisition in bacteria. Most multidrug-resistant E.?fa...

  6. Conjugative plasmid transfer in Xylella fastidiosa is dependent on tra and trb operon functions

    Science.gov (United States)

    The insect-transmitted plant pathogen Xylella fastidiosa is capable of efficient horizontal gene transfer and recombination, leading to diversity between strains and the categorization of X. fastidiosa into multiple subspecies. Although natural transformation is shown to occur at high rates in X. fa...

  7. Conjugal properties of the Sinorhizobium meliloti plasmid mobilome.

    Science.gov (United States)

    Pistorio, Mariano; Giusti, María A; Del Papa, María F; Draghi, Walter O; Lozano, Mauricio J; Tejerizo, Gonzalo Torres; Lagares, Antonio

    2008-09-01

    The biology and biochemistry of plasmid transfer in soil bacteria is currently under active investigation because of its central role in prokaryote adaptation and evolution. In this work, we examined the conjugal properties of the cryptic plasmids present in a collection of the N(2)-fixing legume-symbiont Sinorhizobium meliloti. The study was performed on 65 S. meliloti isolates recovered from 25 humic soils of Argentina, which were grouped into 22 plasmid-profile types [i.e. plasmid operational taxonomic units (OTUs)]. The cumulative Shannon index calculated for the observed plasmid profiles showed a clear saturation plateau, thus indicating an adequate representation of the S. meliloti plasmid-profile types in the isolates studied. The results show that isolates of nearly 14% of the plasmid OTUs hosted transmissible plasmids and that isolates of 29% of the plasmid OTUs were able to retransfer the previously characterized mobilizable-cryptic plasmid pSmeLPU88b to a third recipient strain. It is noteworthy that isolates belonging to 14% of the plasmid OTUs proved to be refractory to the entrance of the model plasmid pSmeLPU88b, suggesting either the presence of surface exclusion phenomena or the occurrence of restriction incompatibility with the incoming replicon. Incompatibility for replication between resident plasmids and plasmid pSmeLPU88b was observed in c. 20% of the OTUs. The results reported here reveal a widespread compatibility among the conjugal functions of the cryptic plasmids in S. meliloti, and this fact, together with the observed high proportion of existing donor genotypes, points to the extrachromosomal compartment of the species as being an extremely active plasmid mobilome.

  8. CRISPR-Cas and Restriction-Modification Act Additively against Conjugative Antibiotic Resistance Plasmid Transfer in Enterococcus faecalis.

    Science.gov (United States)

    Price, Valerie J; Huo, Wenwen; Sharifi, Ardalan; Palmer, Kelli L

    2016-01-01

    Enterococcus faecalis is an opportunistic pathogen and a leading cause of nosocomial infections. Conjugative pheromone-responsive plasmids are narrow-host-range mobile genetic elements (MGEs) that are rapid disseminators of antibiotic resistance in the faecalis species. Clustered regularly interspaced short palindromic repeat (CRISPR)-Cas and restriction-modification confer acquired and innate immunity, respectively, against MGE acquisition in bacteria. Most multidrug-resistant E. faecalis isolates lack CRISPR-Cas and possess an orphan locus lacking cas genes, CRISPR2, that is of unknown function. Little is known about restriction-modification defense in E. faecalis. Here, we explore the hypothesis that multidrug-resistant E. faecalis strains are immunocompromised. We assessed MGE acquisition by E. faecalis T11, a strain closely related to the multidrug-resistant hospital isolate V583 but which lacks the ~620 kb of horizontally acquired genome content that characterizes V583. T11 possesses the E. faecalis CRISPR3-cas locus and a predicted restriction-modification system, neither of which occurs in V583. We demonstrate that CRISPR-Cas and restriction-modification together confer a 4-log reduction in acquisition of the pheromone-responsive plasmid pAM714 in biofilm matings. Additionally, we show that the orphan CRISPR2 locus is functional for genome defense against another pheromone-responsive plasmid, pCF10, only in the presence of cas9 derived from the E. faecalis CRISPR1-cas locus, which most multidrug-resistant E. faecalis isolates lack. Overall, our work demonstrated that the loss of only two loci led to a dramatic reduction in genome defense against a clinically relevant MGE, highlighting the critical importance of the E. faecalis accessory genome in modulating horizontal gene transfer. Our results rationalize the development of antimicrobial strategies that capitalize upon the immunocompromised status of multidrug-resistant E. faecalis. IMPORTANCE

  9. The BlcC (AttM) lactonase of Agrobacterium tumefaciens does not quench the quorum-sensing system that regulates Ti plasmid conjugative transfer.

    Science.gov (United States)

    Khan, Sharik R; Farrand, Stephen K

    2009-02-01

    The conjugative transfer of Agrobacterium plasmids is controlled by a quorum-sensing system consisting of TraR and its acyl-homoserine lactone (HSL) ligand. The acyl-HSL is essential for the TraR-mediated activation of the Ti plasmid Tra genes. Strains A6 and C58 of Agrobacterium tumefaciens produce a lactonase, BlcC (AttM), that can degrade the quormone, leading some to conclude that the enzyme quenches the quorum-sensing system. We tested this hypothesis by examining the effects of the mutation, induction, or mutational derepression of blcC on the accumulation of acyl-HSL and on the conjugative competence of strain C58. The induction of blc resulted in an 8- to 10-fold decrease in levels of extracellular acyl-HSL but in only a twofold decrease in intracellular quormone levels, a measure of the amount of active intracellular TraR. The induction or mutational derepression of blc as well as a null mutation in blcC had no significant effect on the induction of or continued transfer of pTiC58 from donors in any stage of growth, including stationary phase. In matings performed in developing tumors, wild-type C58 transferred the Ti plasmid to recipients, yielding transconjugants by 14 to 21 days following infection. blcC-null donors yielded transconjugants 1 week earlier, but by the following week, transconjugants were recovered at numbers indistinguishable from those of the wild type. Donors mutationally derepressed for blcC yielded transconjugants in planta at numbers 10-fold lower than those for the wild type at weeks 2 and 3, but by week 4, the two donors showed no difference in recoverable transconjugants. We conclude that BlcC has no biologically significant effect on Ti plasmid transfer or its regulatory system.

  10. Plasmid transfer by conjugation as a possible route of horizontal gene transfer and recombination in Xylella fastidiosa

    Science.gov (United States)

    Horizontal gene transfer is an important component of evolution and adaptation of bacterial species. Xylella fastidiosa has the ability to incorporate exogenous DNA into its genome by homologous recombination at relatively high rates. This genetic recombination is believed to play a role in adaptati...

  11. 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...

  12. Identification of a Novel Conjugative Plasmid in Mycobacteria That Requires Both Type IV and Type VII Secretion

    KAUST Repository

    Ummels, R.

    2014-09-23

    Conjugative plasmids have been identified in a wide variety of different bacteria, ranging from proteobacteria to firmicutes, and conjugation is one of the most efficient routes for horizontal gene transfer. The most widespread mechanism of plasmid conjugation relies on different variants of the type IV secretion pathway. Here, we describe the identification of a novel type of conjugative plasmid that seems to be unique for mycobacteria. Interestingly, while this plasmid is efficiently exchanged between different species of slow-growing mycobacteria, including Mycobacterium tuberculosis, it could not be transferred to any of the fast-growing mycobacteria tested. Genetic analysis of the conjugative plasmid showed the presence of a locus containing homologues of three type IV secretion system components and a relaxase. In addition, a new type VII secretion locus was present. Using transposon insertion mutagenesis, we show that in fact both these secretion systems are essential for conjugation, indicating that this plasmid represents a new class of conjugative plasmids requiring two secretion machineries. This plasmid could form a useful new tool to exchange or introduce DNA in slow-growing mycobacteria. IMPORTANCE: Conjugative plasmids play an important role in horizontal gene transfer between different bacteria and, as such, in their adaptation and evolution. This effect is most obvious in the spread of antibiotic resistance genes. Thus far, conjugation of natural plasmids has been described only rarely for mycobacterial species. In fact, it is generally accepted that M. tuberculosis does not show any recent sign of horizontal gene transfer. In this study, we describe the identification of a new widespread conjugative plasmid that can also be efficiently transferred to M. tuberculosis. This plasmid therefore poses both a threat and an opportunity. The threat is that, through the acquisition of antibiotic resistance markers, this plasmid could start a rapid spread of

  13. Growth dependence of conjugation explains limited plasmid invasion in biofilms: an individual‐based modelling study

    DEFF Research Database (Denmark)

    Merkey, Brian; Lardon, Laurent; Seoane, Jose Miguel

    2011-01-01

    Plasmid invasion in biofilms is often surprisingly limited in spite of the close contact of cells in a biofilm. We hypothesized that this poor plasmid spread into deeper biofilm layers is caused by a dependence of conjugation on the growth rate (relative to the maximum growth rate) of the donor......, we find that invasion of a resident biofilm is indeed limited when plasmid transfer depends on growth, but not so in the absence of growth dependence. Using sensitivity analysis we also find that parameters related to timing (i.e. a lag before the transconjugant can transfer, transfer proficiency...... and scan speed) and spatial reach (EPS yield, conjugal pilus length) are more important for successful plasmid invasion than the recipients' growth rate or the probability of segregational loss. While this study identifies one factor that can limit plasmid invasion in biofilms, the new individual...

  14. Identification of a Novel Conjugative Plasmid in Mycobacteria That Requires Both Type IV and Type VII Secretion

    KAUST Repository

    Ummels, R.; Abdallah, A. M.; Kuiper, V.; Aajoud, A.; Sparrius, M.; Naeem, R.; Spaink, H. P.; van Soolingen, D.; Pain, Arnab; Bitter, W.

    2014-01-01

    Conjugative plasmids play an important role in horizontal gene transfer between different bacteria and, as such, in their adaptation and evolution. This effect is most obvious in the spread of antibiotic resistance genes. Thus far, conjugation of natural plasmids has been described only rarely for mycobacterial species. In fact, it is generally accepted that M. tuberculosis does not show any recent sign of horizontal gene transfer. In this study, we describe the identification of a new widespread conjugative plasmid that can also be efficiently transferred to M. tuberculosis. This plasmid therefore poses both a threat and an opportunity. The threat is that, through the acquisition of antibiotic resistance markers, this plasmid could start a rapid spread of antibiotic resistance genes between pathogenic mycobacteria. The opportunity is that we could use this plasmid to generate new tools for the efficient introduction of foreign DNA in slow-growing mycobacteria.

  15. The extended regulatory networks of SXT/R391 integrative and conjugative elements and IncA/C conjugative plasmids.

    Science.gov (United States)

    Poulin-Laprade, Dominic; Carraro, Nicolas; Burrus, Vincent

    2015-01-01

    Nowadays, healthcare systems are challenged by a major worldwide drug resistance crisis caused by the massive and rapid dissemination of antibiotic resistance genes and associated emergence of multidrug resistant pathogenic bacteria, in both clinical and environmental settings. Conjugation is the main driving force of gene transfer among microorganisms. This mechanism of horizontal gene transfer mediates the translocation of large DNA fragments between two bacterial cells in direct contact. Integrative and conjugative elements (ICEs) of the SXT/R391 family (SRIs) and IncA/C conjugative plasmids (ACPs) are responsible for the dissemination of a broad spectrum of antibiotic resistance genes among diverse species of Enterobacteriaceae and Vibrionaceae. The biology, diversity, prevalence and distribution of these two families of conjugative elements have been the subject of extensive studies for the past 15 years. Recently, the transcriptional regulators that govern their dissemination through the expression of ICE- or plasmid-encoded transfer genes have been described. Unrelated repressors control the activation of conjugation by preventing the expression of two related master activator complexes in both types of elements, i.e., SetCD in SXT/R391 ICEs and AcaCD in IncA/C plasmids. Finally, in addition to activating ICE- or plasmid-borne genes, these master activators have been shown to specifically activate phylogenetically unrelated mobilizable genomic islands (MGIs) that also disseminate antibiotic resistance genes and other adaptive traits among a plethora of pathogens such as Vibrio cholerae and Salmonella enterica.

  16. The extended regulatory networks of SXT/R391 integrative and conjugative elements and IncA/C conjugative plasmids.

    Directory of Open Access Journals (Sweden)

    Dominic ePoulin-Laprade

    2015-08-01

    Full Text Available Nowadays, healthcare systems are challenged by a major worldwide drug resistance crisis caused by the massive and rapid dissemination of antibiotic resistance genes and associated emergence of multidrug resistant pathogenic bacteria, in both clinical and environmental settings. Conjugation is the main driving force of gene transfer among microorganisms. This mechanism of horizontal gene transfer mediates the translocation of large DNA fragments between two bacterial cells in direct contact. Integrative and conjugative elements (ICEs of the SXT/R391 family (SRIs and IncA/C conjugative plasmids (ACPs are responsible for the dissemination of a broad spectrum of antibiotic resistance genes among diverse species of Enterobacteriaceae and Vibrionaceae. The biology, diversity, prevalence and distribution of these two families of conjugative elements have been the subject of extensive studies for the past 15 years. Recently, the transcriptional regulators that govern their dissemination through the expression of ICE- or plasmid-encoded transfer genes have been described. Unrelated repressors control the activation of conjugation by preventing the expression of two related master activator complexes in both types of elements, i.e. SetCD in SXT/R391 ICEs and AcaCD in IncA/C plasmids. Finally, in addition to activating ICE- or plasmid-borne genes, these master activators have been shown to specifically activate phylogenetically unrelated mobilizable genomic islands (MGIs that also disseminate antibiotic resistance genes and other adaptive traits among a plethora of pathogens such as Vibrio cholerae and Salmonella enterica.

  17. Impact of co-carriage of IncA/C plasmids with additional plasmids on the transfer of antimicrobial resistance in Salmonella enterica isolates.

    Science.gov (United States)

    Han, Jing; Pendleton, Sean J; Deck, Joanna; Singh, Ruby; Gilbert, Jeffrey; Johnson, Timothy J; Sanad, Yasser M; Nayak, Rajesh; Foley, Steven L

    2018-04-20

    Antimicrobial resistance in Salmonella enterica is often plasmid encoded. A key resistance plasmid group is the incompatibility group (Inc) A/C plasmids that often carry multiple resistance determinants. Previous studies showed that IncA/C plasmids were often co-located with other plasmids. The current study was undertaken to evaluate the impact of plasmid co-carriage on antimicrobial resistance and plasmid transfer. A total of 1267 Salmonella isolates, representing multiple serotypes and sources were previously subjected to susceptibility testing and 251 isolates with resistance to at least 5 antimicrobial agents were identified for further study. Each isolate was subjected to PCR-based replicon typing, and those with IncA/C plasmids were selected for plasmid isolation, PCR-based mapping of IncA/C plasmid backbone genes, and conjugation assays to evaluate resistance plasmid transferability. Of the 87 identified IncA/C positive isolates, approximately 75% carried a plasmid with another identified replicon type, with the most common being I1 (39%), FIA, FIIA, FIB and HI2 (each 15%). PCR-based mapping indicated significant diversity in IncA/C backbone content, especially in regions encoding transfer-associated and hypothetical proteins. Conjugation experiments showed that nearly 68% of the isolates transferred resistance plasmids, with 90% containing additional identified plasmids or larger (>50 kb) non-typeable plasmids. The majority of IncA/C-positive strains were able to conjugally transfer antimicrobial resistance to the recipient, encoded by IncA/C and/or co-carried plasmids. These findings highlight the importance of co-located plasmids for resistance dissemination either by directly transferring resistance genes or by potentially providing the needed conjugation machinery for IncA/C plasmid transfer. Copyright © 2018. Published by Elsevier B.V.

  18. Effect of ionizing radition on conjugative R plasmid in Escherichia coli

    International Nuclear Information System (INIS)

    Kmetova, M.; Puzova, H.; Rexa, R.

    1986-01-01

    Five-fold cyclic gamma irradiation of E. coli strain No. 214 with conjugative R plasmid with doses of 150 Gy, with the exception of chloramphenicol, did not essentially affect the expression of the examined determinants of resistance to antimicrobial substances (tetracycline, streptomycin, chloramphenicol, canamycin, ampicillin, sulfamethoxidine). The dose of 150 Gy from the first irradiation of the strain reduced the transfer frequency of the R plasmid approximately hundred-fold. After the second up to the fourth irradiation of the strain the transfer frequency went back to approximately its original value. (author)

  19. Recombinogenic engineering of conjugative plasmids with fluorescent marker cassettes

    DEFF Research Database (Denmark)

    Reisner, A.; Molin, Søren; Zechner, E.L.

    2002-01-01

    An efficient approach for the insertion of fluorescent marker genes with sequence specificity into conjugative plasmids in Escherichia coli is described. For this purpose, homologous recombination of linear double-stranded targeting DNA was mediated by the bacteriophage lambda recombination...... resistance genes and fluorescent markers. The choice of 5' non-homologous extensions in primer pairs used for amplifying the marker cassettes determines the site specificity of the targeting DNA. This methodology is applicable to the modification of all plasmids that replicate in E coli and is not restricted...

  20. Metal stressors consistently modulate bacterial conjugal plasmid uptake potential in a phylogenetically conserved manner

    DEFF Research Database (Denmark)

    Klümper, Uli; Dechesne, Arnaud; Riber, Leise

    2017-01-01

    The environmental stimulants and inhibitors of conjugal plasmid transfer in microbial communities are poorly understood. Specifically, it is not known whether exposure to stressors may cause a community to alter its plasmid uptake ability. We assessed whether metals (Cu, Cd, Ni, Zn) and one metal...... that community permissiveness is sensitive to metal(loid) stress in a manner that is both partially consistent across stressors and phylogenetically conserved.The ISME Journal advance online publication, 2 August 2016; doi:10.1038/ismej.2016.98....

  1. Protocol for Evaluating the Permissiveness of Bacterial Communities Toward Conjugal Plasmids by Quantification and Isolation of Transconjugants

    DEFF Research Database (Denmark)

    Klümper, Uli; Dechesne, Arnaud; Smets, Barth F.

    2014-01-01

    may encode catabolic pathways, virulence factors, and antibiotic or metal resistances, it is of environmental, evolutionary, and medical relevance to track and monitor the fate of plasmids in mixed microbial community. When assessing the short-term and long-term implications of conjugal plasmid...... a gfp-tagged plasmid in a mCherry red fluorescently tagged donor strain repressing gfp expression. We take advantage of fluorescent marker genes to microscopically detect plasmid transfer events and use subsequent high-throughput fluorescence-activated cell sorting (FACS) to isolate...

  2. A conjugative 38 kB plasmid is present in multiple subspecies of Xylella fastidiosa.

    Science.gov (United States)

    Rogers, Elizabeth E; Stenger, Drake C

    2012-01-01

    A ≈ 38kB plasmid (pXF-RIV5) was present in the Riv5 strain of Xylella fastidiosa subsp. multiplex isolated from ornamental plum in southern California. The complete nucleotide sequence of pXF-RIV5 is almost identical to that of pXFAS01 from X. fastidiosa subsp. fastidiosa strain M23; the two plasmids vary at only 6 nucleotide positions. BLAST searches and phylogenetic analyses indicate pXF-RIV5 and pXFAS01 share some similarity to chromosomal and plasmid (pXF51) sequences of X. fastidiosa subsp. pauca strain 9a5c and more distant similarity to plasmids from a wide variety of bacteria. Both pXF-RIV5 and pXFAS01 encode homologues of a complete Type IV secretion system involved in conjugation and DNA transfer among bacteria. Mating pair formation proteins (Trb) from Yersinia pseudotuberculosis IP31758 are the mostly closely related non-X. fastidiosa proteins to most of the Trb proteins encoded by pXF-RIV5 and pXFAS01. Unlike many bacterial conjugative plasmids, pXF-RIV5 and pXFAS01 do not carry homologues of known accessory modules that confer selective advantage on host bacteria. However, both plasmids encode seven hypothetical proteins of unknown function and possess a small transposon-associated region encoding a putative transposase and associated factor. Vegetative replication of pXF-RIV5 and pXFAS01 appears to be under control of RepA protein and both plasmids have an origin of DNA replication (oriV) similar to that of pRP4 and pR751 from Escherichia coli. In contrast, conjugative plasmids commonly encode TrfA and have an oriV similar to those found in IncP-1 incompatibility group plasmids. The presence of nearly identical plasmids in single strains from two distinct subspecies of X. fastidiosa is indicative of recent horizontal transfer, probably subsequent to the introduction of subspecies fastidiosa to the United States in the late 19(th) century.

  3. A conjugative 38 kB plasmid is present in multiple subspecies of Xylella fastidiosa.

    Directory of Open Access Journals (Sweden)

    Elizabeth E Rogers

    Full Text Available A ≈ 38kB plasmid (pXF-RIV5 was present in the Riv5 strain of Xylella fastidiosa subsp. multiplex isolated from ornamental plum in southern California. The complete nucleotide sequence of pXF-RIV5 is almost identical to that of pXFAS01 from X. fastidiosa subsp. fastidiosa strain M23; the two plasmids vary at only 6 nucleotide positions. BLAST searches and phylogenetic analyses indicate pXF-RIV5 and pXFAS01 share some similarity to chromosomal and plasmid (pXF51 sequences of X. fastidiosa subsp. pauca strain 9a5c and more distant similarity to plasmids from a wide variety of bacteria. Both pXF-RIV5 and pXFAS01 encode homologues of a complete Type IV secretion system involved in conjugation and DNA transfer among bacteria. Mating pair formation proteins (Trb from Yersinia pseudotuberculosis IP31758 are the mostly closely related non-X. fastidiosa proteins to most of the Trb proteins encoded by pXF-RIV5 and pXFAS01. Unlike many bacterial conjugative plasmids, pXF-RIV5 and pXFAS01 do not carry homologues of known accessory modules that confer selective advantage on host bacteria. However, both plasmids encode seven hypothetical proteins of unknown function and possess a small transposon-associated region encoding a putative transposase and associated factor. Vegetative replication of pXF-RIV5 and pXFAS01 appears to be under control of RepA protein and both plasmids have an origin of DNA replication (oriV similar to that of pRP4 and pR751 from Escherichia coli. In contrast, conjugative plasmids commonly encode TrfA and have an oriV similar to those found in IncP-1 incompatibility group plasmids. The presence of nearly identical plasmids in single strains from two distinct subspecies of X. fastidiosa is indicative of recent horizontal transfer, probably subsequent to the introduction of subspecies fastidiosa to the United States in the late 19(th century.

  4. Mobility of the native Bacillus subtilis conjugative plasmid pLS20 is regulated by intercellular signaling.

    Science.gov (United States)

    Singh, Praveen K; Ramachandran, Gayetri; Ramos-Ruiz, Ricardo; Peiró-Pastor, Ramón; Abia, David; Wu, Ling J; Meijer, Wilfried J J

    2013-10-01

    Horizontal gene transfer mediated by plasmid conjugation plays a significant role in the evolution of bacterial species, as well as in the dissemination of antibiotic resistance and pathogenicity determinants. Characterization of their regulation is important for gaining insights into these features. Relatively little is known about how conjugation of Gram-positive plasmids is regulated. We have characterized conjugation of the native Bacillus subtilis plasmid pLS20. Contrary to the enterococcal plasmids, conjugation of pLS20 is not activated by recipient-produced pheromones but by pLS20-encoded proteins that regulate expression of the conjugation genes. We show that conjugation is kept in the default "OFF" state and identified the master repressor responsible for this. Activation of the conjugation genes requires relief of repression, which is mediated by an anti-repressor that belongs to the Rap family of proteins. Using both RNA sequencing methodology and genetic approaches, we have determined the regulatory effects of the repressor and anti-repressor on expression of the pLS20 genes. We also show that the activity of the anti-repressor is in turn regulated by an intercellular signaling peptide. Ultimately, this peptide dictates the timing of conjugation. The implications of this regulatory mechanism and comparison with other mobile systems are discussed.

  5. Plasmid Transfer in the Ocean – A Case Study from the Roseobacter Group

    Directory of Open Access Journals (Sweden)

    Jörn Petersen

    2017-07-01

    Full Text Available Plasmid mediated horizontal gene transfer (HGT has been speculated to be one of the prime mechanisms for the adaptation of roseobacters (Rhodobacteraceae to their ecological niches in the marine habitat. Their plasmids contain ecologically crucial functional modules of up to ∼40-kb in size, e.g., for aerobic anoxygenic photosynthesis, flagellar formation and the biosynthesis of the antibiotic tropodithietic acid. Furthermore, the widely present type four secretion system (T4SS of roseobacters has been shown to mediate conjugation across genus barriers, albeit in the laboratory. Here we discovered that Confluentimicrobium naphthalenivorans NS6T, a tidal flat bacterium isolated in Korea, carries a 185-kb plasmid, which exhibits a long-range synteny with the conjugative 126-kb plasmid of Dinoroseobacter shibae DFL12T. Both replicons are stably maintained by RepABC operons of the same compatibility group (-2 and they harbor a homologous T4SS. Principal component analysis of the codon usage shows a large similarity between the two plasmids, while the chromosomes are very distinct, showing that neither of the two bacterial species represents the original host of those RepABC-2 type plasmids. The two species do not share a common habitat today and they are phylogenetically only distantly related. Our finding demonstrates the first clear-cut evidence for conjugational plasmid transfer across biogeographical and phylogenetic barriers in Rhodobacteraceae and documents the importance of conjugative HGT in the ocean.

  6. Processing of Nonconjugative Resistance Plasmids by Conjugation Nicking Enzyme of Staphylococci

    Energy Technology Data Exchange (ETDEWEB)

    Pollet, Rebecca M.; Ingle, James D.; Hymes, Jeff P.; Eakes, Thomas C.; Eto, Karina Yui; Kwong, Stephen M.; Ramsay, Joshua P.; Firth, Neville; Redinbo, Matthew R. (Curtin U.); (Sydney); (UNC)

    2016-01-04

    Antimicrobial resistance inStaphylococcus aureuspresents an increasing threat to human health. This resistance is often encoded on mobile plasmids, such as pSK41; however, the mechanism of transfer of these plasmids is not well understood. In this study, we first examine key protein-DNA interactions formed by the relaxase enzyme, NES, which initiates and terminates the transfer of the multidrug resistance plasmid pSK41. Two loops on the NES protein, hairpin loops 1 and 2, form extensive contacts with the DNA hairpin formed at theoriTregion of pSK41, and here we establish that these contacts are essential for proper DNA cleavage and religation by the full 665-residue NES proteinin vitro. Second, pSK156 and pCA347 are nonconjugativeStaphylococcus aureusplasmids that contain sequences similar to theoriTregion of pSK41 but differ in the sequence predicted to form a DNA hairpin. We show that pSK41-encoded NES is able to bind, cleave, and religate theoriTsequences of these nonconjugative plasmidsin vitro. Although pSK41 could mobilize a coresident plasmid harboring its cognateoriT, it was unable to mobilize plasmids containing the pSK156 and pCA347 variantoriTmimics, suggesting that an accessory protein like that previously shown to confer specificity in the pWBG749 system may also be involved in transmission of plasmids containing a pSK41-likeoriT. These data indicate that the conjugative relaxase intransmechanism recently described for the pWBG749 family of plasmids also applies to the pSK41 family of plasmids, further heightening the potential significance of this mechanism in the horizontal transfer of staphylococcal plasmids.

    IMPORTANCEUnderstanding the

  7. The master activator of IncA/C conjugative plasmids stimulates genomic islands and multidrug resistance dissemination.

    Science.gov (United States)

    Carraro, Nicolas; Matteau, Dominick; Luo, Peng; Rodrigue, Sébastien; Burrus, Vincent

    2014-10-01

    Dissemination of antibiotic resistance genes occurs mostly by conjugation, which mediates DNA transfer between cells in direct contact. Conjugative plasmids of the IncA/C incompatibility group have become a substantial threat due to their broad host-range, the extended spectrum of antimicrobial resistance they confer, their prevalence in enteric bacteria and their very efficient spread by conjugation. However, their biology remains largely unexplored. Using the IncA/C conjugative plasmid pVCR94ΔX as a prototype, we have investigated the regulatory circuitry that governs IncA/C plasmids dissemination and found that the transcriptional activator complex AcaCD is essential for the expression of plasmid transfer genes. Using chromatin immunoprecipitation coupled with exonuclease digestion (ChIP-exo) and RNA sequencing (RNA-seq) approaches, we have identified the sequences recognized by AcaCD and characterized the AcaCD regulon. Data mining using the DNA motif recognized by AcaCD revealed potential AcaCD-binding sites upstream of genes involved in the intracellular mobility functions (recombination directionality factor and mobilization genes) in two widespread classes of genomic islands (GIs) phylogenetically unrelated to IncA/C plasmids. The first class, SGI1, confers and propagates multidrug resistance in Salmonella enterica and Proteus mirabilis, whereas MGIVmi1 in Vibrio mimicus belongs to a previously uncharacterized class of GIs. We have demonstrated that through expression of AcaCD, IncA/C plasmids specifically trigger the excision and mobilization of the GIs at high frequencies. This study provides new evidence of the considerable impact of IncA/C plasmids on bacterial genome plasticity through their own mobility and the mobilization of genomic islands.

  8. Estimating the Transfer Range of Plasmids Encoding Antimicrobial Resistance in a Wastewater Treatment Plant Microbial Community

    DEFF Research Database (Denmark)

    Li, Liguan; Dechesne, Arnaud; He, Zhiming

    2018-01-01

    sludge microbial community was challenged in standardized filter matings with one of three multidrug resistance plasmids (pKJK5, pB10, and RP4) harbored by Escherichia coli or Pseudomonas putida. Different donor–plasmid combinations had distinct transfer frequencies, ranging from 3 to 50 conjugation...... events per 100000 cells of the WWTP microbial community. In addition, transfer was observed to a broad phylogenetic range of 13 bacterial phyla with several taxa containing potentially pathogenic species. Preferential transfer to taxa belonging to the predicted evolutionary host range of the plasmids...... ARG transmission. However, the contribution of microbial communities in WWTPs to ARG dissemination remains poorly understood. Here, we examined for the first time plasmid permissiveness of an activated sludge microbial community by utilizing an established fluorescent bioreporter system. The activated...

  9. Cholesterol-conjugated supramolecular assemblies of low generations polyamidoamine dendrimers for enhanced EGFP plasmid DNA transfection

    Energy Technology Data Exchange (ETDEWEB)

    Golkar, Nasim; Samani, Soliman Mohammadi; Tamaddon, Ali Mohammad, E-mail: amtamadon@gmail.com [Shiraz University of Medical Sciences, Department of Pharmaceutics, School of Pharmacy (Iran, Islamic Republic of)

    2016-05-15

    Aimed to prepare an enhanced gene delivery system with low cytotoxicity and high transfection efficiency, various cholesterol-conjugated derivates of low generation polyamidoamine (PAMAM) dendrimers were prepared. The conjugates were characterized by TNBS assay, FTIR, and {sup 1}H-NMR spectroscopy. Self-assembly of the dendrimer conjugates (G1-Chol, G2-Chol, and G3-Chol) was investigated by pyrene assay. Following formation of the complexes between enhanced green fluorescence protein plasmid and the dendrimer conjugates at various N (primary amine)/P (phosphate) mole ratios, plasmid condensation, biologic stability, cytotoxicity, and protein expression were investigated. The conjugates self-assembled into micellar dispersions with the critical micelle concentration values (<50 µg/ml) depending on the dendrimer generation and cholesterol/amine mole ratio. Cholesterol conjugation resulted in higher resistance of the condensed plasmid DNA in a competition assay with heparin sulfate. Also, the transfection efficiency was determined higher for the cholesterol conjugates than unmodified dendrimers in HepG2 cells, showing the highest for G2-Chol at 40 % degree of cholesterol modification (G2-Chol{sub 40 %}) among various dendrimer generations. Interestingly, such conjugate showed a complete protection of plasmid against serum nucleases. Our results confirmed that the cholesterol conjugation to PAMAM dendrimers of low generations bearing little cytotoxicity improves their several physicochemical and biological characteristics required for an enhanced delivery of plasmid DNA into cells.

  10. Repression/depression of conjugative plasmids and their influence on the mutation-selection balance in static environments.

    Directory of Open Access Journals (Sweden)

    Yoav Atsmon-Raz

    Full Text Available We study the effect that conjugation-mediated Horizontal Gene Transfer (HGT has on the mutation-selection balance of a population in a static environment. We consider a model whereby a population of unicellular organisms, capable of conjugation, comes to mutation-selection balance in the presence of an antibiotic, which induces a first-order death rate constant [Formula: see text] for genomes that are not resistant. We explicitly take into consideration the repression/de-repression dynamics of the conjugative plasmid, and assume that a de-repressed plasmid remains temporarily de-repressed after copying itself into another cell. We assume that both repression and de-repression are characterized by first-order rate constants [Formula: see text]and [Formula: see text], respectively. We find that conjugation has a deleterious effect on the mean fitness of the population, suggesting that HGT does not provide a selective advantage in a static environment, but is rather only useful for adapting to new environments. This effect can be ameliorated by repression, suggesting that while HGT is not necessarily advantageous for a population in a static environment, its deleterious effect on the mean fitness can be negated via repression. Therefore, it is likely that HGT is much more advantageous in a dynamic landscape. Furthermore, in the limiting case of a vanishing spontaneous de-repression rate constant, we find that the fraction of conjugators in the population undergoes a phase transition as a function of population density. Below a critical population density, the fraction of conjugators is zero, while above this critical population density the fraction of conjugators rises continuously to one. Our model for conjugation-mediated HGT is related to models of infectious disease dynamics, where the conjugators play the role of the infected (I class, and the non-conjugators play the role of the susceptible (S class.

  11. Isolation of a conjugative F-like plasmid from a multidrug-resistant Escherichia coli strain CM6 using tandem shock wave-mediated transformation.

    Science.gov (United States)

    Soto-Alonso, G; Cruz-Medina, J A; Caballero-Pérez, J; Arvizu-Hernández, I; Ávalos-Esparza, L M; Cruz-Hernández, A; Romero-Gómez, S; Rodríguez, A L; Pastrana-Martínez, X; Fernández, F; Loske, A M; Campos-Guillén, J

    2015-07-01

    Genetic characterization of plasmids from bacterial strains provides insight about multidrug resistance. Ten wild type Escherichia coli (E. coli) strains isolated from cow fecal samples were characterized by their antibiotic resistance profile, plasmid patterns and three different identification methods. From one of the strains, a fertility factor-like plasmid was replicated using tandem shock wave-mediated transformation. Underwater shock waves with a positive pressure peak of up to approximately 40 MPa, followed by a pressure trough of approximately -19 MPa were generated using an experimental piezoelectric shock wave source. Three different shock wave energies and a fixed delay of 750 μs were used to study the relationship between energy and transformation efficiency (TE), as well as the influence of shock wave energy on the integrity of the plasmid. Our results showed that the mean shock wave-mediated TE and the integrity of the large plasmid (~70 kb) were reduced significantly at the energy levels tested. The sequencing analysis of the plasmid revealed a high identity to the pHK17a plasmid, including the replication system, which was similar to the plasmid incompatibility group FII. It also showed that it carried an extended spectrum beta-lactamase gene, ctx-m-14. Furthermore, diverse genes for the conjugative mechanism were identified. Our results may be helpful in improving methodologies for conjugative plasmid transfer and directly selecting the most interesting plasmids from environmental samples. Copyright © 2015 Elsevier B.V. All rights reserved.

  12. Characterization of replication and conjugation of plasmid pWTY27 from a widely distributed Streptomyces species

    Directory of Open Access Journals (Sweden)

    Wang Tao

    2012-11-01

    Full Text Available Abstract Background Streptomyces species are widely distributed in natural habitats, such as soils, lakes, plants and some extreme environments. Replication loci of several Streptomyces theta-type plasmids have been reported, but are not characterized in details. Conjugation loci of some Streptomyces rolling-circle-type plasmids are identified and mechanism of conjugal transferring are described. Results We report the detection of a widely distributed Streptomyces strain Y27 and its indigenous plasmid pWTY27 from fourteen plants and four soil samples cross China by both culturing and nonculturing methods. The complete nucleotide sequence of pWTY27 consisted of 14,288 bp. A basic locus for plasmid replication comprised repAB genes and an adjacent iteron sequence, to a long inverted-repeat (ca. 105 bp of which the RepA protein bound specifically in vitro, suggesting that RepA may recognize a second structure (e.g. a long stem-loop of the iteron DNA. A plasmid containing the locus propagated in linear mode when the telomeres of a linear plasmid were attached, indicating a bi-directional replication mode for pWTY27. As for rolling-circle plasmids, a single traA gene and a clt sequence (covering 16 bp within traA and its adjacent 159 bp on pWTY27 were required for plasmid transfer. TraA recognized and bound specifically to the two regions of the clt sequence, one containing all the four DC1 of 7 bp (TGACACC and one DC2 (CCCGCCC and most of IC1, and another covering two DC2 and part of IC1, suggesting formation of a high-ordered DNA-protein complex. Conclusions This work (i isolates a widespread Streptomyces strain Y27 and sequences its indigenous theta-type plasmid pWTY27; (ii identifies the replication and conjugation loci of pWTY27 and; (iii characterizes the binding sequences of the RepA and TraA proteins.

  13. Characterization of replication and conjugation of plasmid pWTY27 from a widely distributed Streptomyces species

    Science.gov (United States)

    2012-01-01

    Background Streptomyces species are widely distributed in natural habitats, such as soils, lakes, plants and some extreme environments. Replication loci of several Streptomyces theta-type plasmids have been reported, but are not characterized in details. Conjugation loci of some Streptomyces rolling-circle-type plasmids are identified and mechanism of conjugal transferring are described. Results We report the detection of a widely distributed Streptomyces strain Y27 and its indigenous plasmid pWTY27 from fourteen plants and four soil samples cross China by both culturing and nonculturing methods. The complete nucleotide sequence of pWTY27 consisted of 14,288 bp. A basic locus for plasmid replication comprised repAB genes and an adjacent iteron sequence, to a long inverted-repeat (ca. 105 bp) of which the RepA protein bound specifically in vitro, suggesting that RepA may recognize a second structure (e.g. a long stem-loop) of the iteron DNA. A plasmid containing the locus propagated in linear mode when the telomeres of a linear plasmid were attached, indicating a bi-directional replication mode for pWTY27. As for rolling-circle plasmids, a single traA gene and a clt sequence (covering 16 bp within traA and its adjacent 159 bp) on pWTY27 were required for plasmid transfer. TraA recognized and bound specifically to the two regions of the clt sequence, one containing all the four DC1 of 7 bp (TGACACC) and one DC2 (CCCGCCC) and most of IC1, and another covering two DC2 and part of IC1, suggesting formation of a high-ordered DNA-protein complex. Conclusions This work (i) isolates a widespread Streptomyces strain Y27 and sequences its indigenous theta-type plasmid pWTY27; (ii) identifies the replication and conjugation loci of pWTY27 and; (iii) characterizes the binding sequences of the RepA and TraA proteins. PMID:23134842

  14. Unique Helicase Determinants in the Essential Conjugative TraI Factor from Salmonella enterica Serovar Typhimurium Plasmid pCU1

    Energy Technology Data Exchange (ETDEWEB)

    McLaughlin, Krystle J.; Nash, Rebekah P.; Redinbo, Mathew R. (UNC)

    2014-06-16

    The widespread development of multidrug-resistant bacteria is a major health emergency. Conjugative DNA plasmids, which harbor a wide range of antibiotic resistance genes, also encode the protein factors necessary to orchestrate the propagation of plasmid DNA between bacterial cells through conjugative transfer. Successful conjugative DNA transfer depends on key catalytic components to nick one strand of the duplex DNA plasmid and separate the DNA strands while cell-to-cell transfer occurs. The TraI protein from the conjugative Salmonella plasmid pCU1 fulfills these key catalytic roles, as it contains both single-stranded DNA-nicking relaxase and ATP-dependent helicase domains within a single, 1,078-residue polypeptide. In this work, we unraveled the helicase determinants of Salmonella pCU1 TraI through DNA binding, ATPase, and DNA strand separation assays. TraI binds DNA substrates with high affinity in a manner influenced by nucleic acid length and the presence of a DNA hairpin structure adjacent to the nick site. TraI selectively hydrolyzes ATP, and mutations in conserved helicase motifs eliminate ATPase activity. Surprisingly, the absence of a relatively short (144-residue) domain at the extreme C terminus of the protein severely diminishes ATP-dependent strand separation. Collectively, these data define the helicase motifs of the conjugative factor TraI from Salmonella pCU1 and reveal a previously uncharacterized C-terminal functional domain that uncouples ATP hydrolysis from strand separation activity.

  15. Development of pVCR94ΔX from Vibrio cholerae, a prototype for studying multidrug resistant IncA/C conjugative plasmids.

    Science.gov (United States)

    Carraro, Nicolas; Sauvé, Maxime; Matteau, Dominick; Lauzon, Guillaume; Rodrigue, Sébastien; Burrus, Vincent

    2014-01-01

    Antibiotic resistance has grown steadily in Vibrio cholerae over the last few decades to become a major threat in countries affected by cholera. Multi-drug resistance (MDR) spreads among clinical and environmental V. cholerae strains by lateral gene transfer often mediated by integrative and conjugative elements (ICEs) of the SXT/R391 family. However, in a few reported but seemingly isolated cases, MDR in V. cholerae was shown to be associated with other self-transmissible genetic elements such as conjugative plasmids. IncA/C conjugative plasmids are often found associated with MDR in isolates of Enterobacteriaceae. To date, IncA/C plasmids have not been commonly found in V. cholerae or other species of Vibrio. Here we present a detailed analysis of pVCR94ΔX derived from pVCR94, a novel IncA/C conjugative plasmid identified in a V. cholerae clinical strain isolated during the 1994 Rwandan cholera outbreak. pVCR94 was found to confer resistance to sulfamethoxazole, trimethoprim, ampicillin, streptomycin, tetracycline, and chloramphenicol and to transfer at very high frequency. Sequence analysis revealed its mosaic nature as well as high similarity of the core genes responsible for transfer and maintenance with other IncA/C plasmids and ICEs of the SXT/R391 family. Although IncA/C plasmids are considered a major threat in antibiotics resistance, their basic biology has received little attention, mostly because of the difficulty to genetically manipulate these MDR conferring elements. Therefore, we developed a convenient derivative from pVCR94, pVCR94Δ X, a 120.5-kb conjugative plasmid which only codes for sulfamethoxazole resistance. Using pVCR94Δ X, we identified the origin of transfer (oriT) and discovered an essential gene for transfer, both located within the shared backbone, allowing for an annotation update of all IncA/C plasmids. pVCR94Δ X may be a useful model that will provide new insights on the basic biology of IncA/C conjugative plasmids.

  16. Conjugative plasmids: Vessels of the communal gene pool

    DEFF Research Database (Denmark)

    Norman, Anders; Hansen, Lars H.; Sørensen, Søren Johannes

    2009-01-01

    to the hosts and, potentially, other resident prokaryotes within specific environmental niches. Insight into the evolution of plasmid modules therefore contributes to our knowledge of gene dissemination and evolution within prokaryotic communities. This communal pool provides the prokaryotes with an important...... mechanistic framework for obtaining adaptability and functional diversity that alleviates the need for large genomes of specialized ‘private genes'....

  17. 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...

  18. Effect of bacterial distribution and activity on conjugal transfer on the phylloplane of the bush bean (Phaseolus vulgaris)

    DEFF Research Database (Denmark)

    Normander, Bo; Christensen, Bjarke Bak; Molin, Søren

    1998-01-01

    Conjugal plasmid transfer was examined on the phylloplane of bean (Phaseolus vulgaris) and related to the spatial distribution pattern and metabolic activity of the bacteria. The donor (Pseudomonas putida KT2442) harbored a derivative of the TOL plasmid, which conferred kanamycin resistance and had...

  19. Diversity of Clostridium perfringens isolates from various sources and prevalence of conjugative plasmids.

    Science.gov (United States)

    Park, Miseon; Deck, Joanna; Foley, Steven L; Nayak, Rajesh; Songer, J Glenn; Seibel, Janice R; Khan, Saeed A; Rooney, Alejandro P; Hecht, David W; Rafii, Fatemeh

    2016-04-01

    Clostridium perfringens is an important pathogen, causing food poisoning and other mild to severe infections in humans and animals. Some strains of C. perfringens contain conjugative plasmids, which may carry antimicrobial resistance and toxin genes. We studied genomic and plasmid diversity of 145 C. perfringens type A strains isolated from soils, foods, chickens, clinical samples, and domestic animals (porcine, bovine and canine), from different geographic areas in the United States between 1994 and 2006, using multiple-locus variable-number tandem repeat analysis (MLVA) and/or pulsed-field gel electrophoresis (PFGE). MLVA detected the genetic diversity in a majority of the isolates. PFGE, using SmaI and KspI, confirmed the MLVA results but also detected differences among the strains that could not be differentiated by MLVA. All of the PFGE profiles of the strains were different, except for a few of the epidemiologically related strains, which were identical. The PFGE profiles of strains isolated from the same domestic animal species were clustered more closely with each other than with other strains. However, a variety of C. perfringens strains with distinct genetic backgrounds were found among the clinical isolates. Variation was also observed in the size and number of plasmids in the strains. Primers for the internal fragment of a conjugative tcpH gene of C. perfringens plasmid pCPF4969 amplified identical size fragments from a majority of strains tested; and this gene hybridized to the various-sized plasmids of these strains. The sequences of the PCR-amplified tcpH genes from 12 strains showed diversity among the tcpH genes. Regardless of the sources of the isolates, the genetic diversity of C. perfringens extended to the plasmids carrying conjugative genes. Published by Elsevier Ltd.

  20. Transfer of the lambdadv plasmid to new bacterial hosts

    International Nuclear Information System (INIS)

    Kellenberger-Gujer, G.; Boy de la Tour, E.; Berg, D.E.

    1974-01-01

    Lambda dv, which was derived from bacteriophage lambda, replicates autonomously as a plasmid in Escherichia coli and consists of only the immunity region (imm/sup lambda/) and DNA replication genes (O, P) of the ancestral phage. Addition phages (lambda imm 21 --lambda dv) carry the lambda dv fragment inserted as a tandem duplication in their genome (sequence A imm 21 O P imm/sup lambda/ O P R) are formed as recombinants after lambda imm 21 infection of strains carrying lambda dv. Addition phages were used to transfer lambda dv to new bacterial hosts. Lambda dv transfer by excision of the lambda dv segment from the addition phage genome requires a bacterial Rec or a phage Red recombination system. Successful transfer is stimulated by uv irradiation of the addition phage before infection. Some properties of the newly transferred lambda dv plasmids are described. (U.S.)

  1. Conjugative IncFI plasmids carrying CTX-M-15 among Escherichia coli ESBL producing isolates at a University hospital in Germany

    Directory of Open Access Journals (Sweden)

    Hain Torsten

    2009-06-01

    Full Text Available Abstract Background Multi-drug-resistant, extended-spectrum β-lactamase (ESBL-producing Enterobacteriaceae, constitute an emerging public-health concern. Little data on the molecular epidemiology of ESBL producing Escherichia coli is available in Germany. Here we describe the prevalence and molecular epidemiology of ESBL producing-Escherichia coli isolates at a German University hospital. Methods We analysed 63 non-duplicate clinical ESBL isolates obtained over an 8-month period using PCR and sequence-based ESBL allele typing, plasmid replicon typing, phylogenetic group typing. Pulsed-field gel electrophoresis (PFGE based genotyping and plasmid profiling was performed, as well as confirmatory DNA-based hybridization assays. Results Examination of the 63 Escherichia coli isolates revealed an almost equal distribution among the E. coli phylogenetic groups A, B1, B2 and D. High prevalence (36/63 of the CTX-M-15 gene was observed and an analysis of PFGE-based patterns revealed the presence of this CTX-M allele in multiple clones. Resistance to cefotaxime was a transferable trait and a commonly occurring 145.5 kb conjugative IncFI plasmid was detected in 65% of E. coli carrying the CTX-M-15 allele. The rate of transferable antibiotic resistances for GM, SXT, TET, GM-SXT-TET, SXT-TET and GM-TET was 33%, 61%, 61%, 27%, 44% and 11%, respectively. The remaining strains did not have a common IncFI plasmid but harboured transferable IncFI plasmids with sizes that ranged from 97 to 242.5 kb. Conclusion Our data demonstrate the presence of IncFI plasmids within the prevailing E. coli population in a hospital setting and suggest that the dissemination of CTX-M-15 allele is associated to lateral transfer of these well-adapted, conjugative IncFI plasmids among various E. coli genotypes.

  2. The Bacillus subtilis Conjugative Plasmid pLS20 Encodes Two Ribbon-Helix-Helix Type Auxiliary Relaxosome Proteins That Are Essential for Conjugation.

    Science.gov (United States)

    Miguel-Arribas, Andrés; Hao, Jian-An; Luque-Ortega, Juan R; Ramachandran, Gayetri; Val-Calvo, Jorge; Gago-Córdoba, César; González-Álvarez, Daniel; Abia, David; Alfonso, Carlos; Wu, Ling J; Meijer, Wilfried J J

    2017-01-01

    Bacterial conjugation is the process by which a conjugative element (CE) is transferred horizontally from a donor to a recipient cell via a connecting pore. One of the first steps in the conjugation process is the formation of a nucleoprotein complex at the origin of transfer ( oriT ), where one of the components of the nucleoprotein complex, the relaxase, introduces a site- and strand specific nick to initiate the transfer of a single DNA strand into the recipient cell. In most cases, the nucleoprotein complex involves, besides the relaxase, one or more additional proteins, named auxiliary proteins, which are encoded by the CE and/or the host. The conjugative plasmid pLS20 replicates in the Gram-positive Firmicute bacterium Bacillus subtilis . We have recently identified the relaxase gene and the oriT of pLS20, which are separated by a region of almost 1 kb. Here we show that this region contains two auxiliary genes that we name aux1 LS20 and aux2 LS20 , and which we show are essential for conjugation. Both Aux1 LS20 and Aux2 LS20 are predicted to contain a Ribbon-Helix-Helix DNA binding motif near their N-terminus. Analyses of the purified proteins show that Aux1 LS20 and Aux2 LS20 form tetramers and hexamers in solution, respectively, and that they both bind preferentially to oriT LS20 , although with different characteristics and specificities. In silico analyses revealed that genes encoding homologs of Aux1 LS20 and/or Aux2 LS20 are located upstream of almost 400 relaxase genes of the Rel LS20 family (MOB L ) of relaxases. Thus, Aux1 LS20 and Aux2 LS20 of pLS20 constitute the founding member of the first two families of auxiliary proteins described for CEs of Gram-positive origin.

  3. The Bacillus subtilis Conjugative Plasmid pLS20 Encodes Two Ribbon-Helix-Helix Type Auxiliary Relaxosome Proteins That Are Essential for Conjugation

    Directory of Open Access Journals (Sweden)

    Andrés Miguel-Arribas

    2017-11-01

    Full Text Available Bacterial conjugation is the process by which a conjugative element (CE is transferred horizontally from a donor to a recipient cell via a connecting pore. One of the first steps in the conjugation process is the formation of a nucleoprotein complex at the origin of transfer (oriT, where one of the components of the nucleoprotein complex, the relaxase, introduces a site- and strand specific nick to initiate the transfer of a single DNA strand into the recipient cell. In most cases, the nucleoprotein complex involves, besides the relaxase, one or more additional proteins, named auxiliary proteins, which are encoded by the CE and/or the host. The conjugative plasmid pLS20 replicates in the Gram-positive Firmicute bacterium Bacillus subtilis. We have recently identified the relaxase gene and the oriT of pLS20, which are separated by a region of almost 1 kb. Here we show that this region contains two auxiliary genes that we name aux1LS20 and aux2LS20, and which we show are essential for conjugation. Both Aux1LS20 and Aux2LS20 are predicted to contain a Ribbon-Helix-Helix DNA binding motif near their N-terminus. Analyses of the purified proteins show that Aux1LS20 and Aux2LS20 form tetramers and hexamers in solution, respectively, and that they both bind preferentially to oriTLS20, although with different characteristics and specificities. In silico analyses revealed that genes encoding homologs of Aux1LS20 and/or Aux2LS20 are located upstream of almost 400 relaxase genes of the RelLS20 family (MOBL of relaxases. Thus, Aux1LS20 and Aux2LS20 of pLS20 constitute the founding member of the first two families of auxiliary proteins described for CEs of Gram-positive origin.

  4. Protein-Nanocrystal Conjugates Support a Single Filament Polymerization Model in R1 Plasmid Segregation

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Charina L.; Claridge, Shelley A.; Garner, Ethan C.; Alivisatos, A. Paul; Mullins, R. Dyche

    2008-07-15

    To ensure inheritance by daughter cells, many low-copy number bacterial plasmids, including the R1 drug-resistance plasmid, encode their own DNA segregation systems. The par operon of plasmid R1 directs construction of a simple spindle structure that converts free energy of polymerization of an actin-like protein, ParM, into work required to move sister plasmids to opposite poles of rod-shaped cells. The structures of individual components have been solved, but little is known about the ultrastructure of the R1 spindle. To determine the number of ParM filaments in a minimal R1 spindle, we used DNA-gold nanocrystal conjugates as mimics of the R1 plasmid. Wefound that each end of a single polar ParM filament binds to a single ParR/parC-gold complex, consistent with the idea that ParM filaments bind in the hollow core of the ParR/parC ring complex. Our results further suggest that multifilament spindles observed in vivo are associated with clusters of plasmidssegregating as a unit.

  5. Novel assay to measure the plasmid mobilizing potential of mixed microbial communities

    DEFF Research Database (Denmark)

    Klümper, Uli; Droumpali, Ariadni; Dechesne, Arnaud

    2014-01-01

    Mobilizable plasmids lack necessary genes for complete conjugation and are therefore non-self-transmissible. Instead, they rely on the conjugation system of conjugal plasmids to be horizontally transferred to new recipients. While community permissiveness, the fraction of a mixed microbial...... community that can receive self-transmissible conjugal plasmids, has been studied, the intrinsic ability of a community to mobilize plasmids that lack conjugation systems is unexplored. Here, we present a novel framework and experimental method to estimate the mobilization potential of mixed communities. We...... of the donors receiving the conjugal plasmid in the first step. Further work is needed to establish how plasmid mobilization potential varies within and across microbial communities....

  6. Theory of periodic conjugate heat transfer

    CERN Document Server

    Zudin, Yuri B

    2016-01-01

    This book presents the theory of periodic conjugate heat transfer in detail. It offers a simplified description of the interaction between a solid body and a fluid as a boundary value problem of the heat conduction equation for the solid body.

  7. Conjugate problems in convective heat transfer

    CERN Document Server

    Dorfman, Abram S

    2009-01-01

    The conjugate heat transfer (CHT) problem takes into account the thermal interaction between a body and fluid flowing over or through it, a key consideration in both mechanical and aerospace engineering. Presenting more than 100 solutions of non-isothermal and CHT problems, this title considers the approximate solutions of CHT problems.

  8. The detection and sequencing of a broad-host-range conjugative IncP-1β plasmid in an epidemic strain of Mycobacterium abscessus subsp. bolletii.

    Directory of Open Access Journals (Sweden)

    Sylvia Cardoso Leão

    Full Text Available BACKGROUND: An extended outbreak of mycobacterial surgical infections occurred in Brazil during 2004-2008. Most infections were caused by a single strain of Mycobacterium abscessus subsp. bolletii, which was characterized by a specific rpoB sequevar and two highly similar pulsed-field gel electrophoresis (PFGE patterns differentiated by the presence of a ∼50 kb band. The nature of this band was investigated. METHODOLOGY/PRINCIPAL FINDINGS: Genomic sequencing of the prototype outbreak isolate INCQS 00594 using the SOLiD platform demonstrated the presence of a 56,267-bp [corrected] circular plasmid, designated pMAB01. Identity matrices, genetic distances and phylogeny analyses indicated that pMAB01 belongs to the broad-host-range plasmid subgroup IncP-1β and is highly related to BRA100, pJP4, pAKD33 and pB10. The presence of pMAB01-derived sequences in 41 M. abscessus subsp. bolletii isolates was evaluated using PCR, PFGE and Southern blot hybridization. Sixteen of the 41 isolates showed the presence of the plasmid. The plasmid was visualized as a ∼50-kb band using PFGE and Southern blot hybridization in 12 isolates. The remaining 25 isolates did not exhibit any evidence of this plasmid. The plasmid was successfully transferred to Escherichia coli by conjugation and transformation. Lateral transfer of pMAB01 to the high efficient plasmid transformation strain Mycobacterium smegmatis mc(2155 could not be demonstrated. CONCLUSIONS/SIGNIFICANCE: The occurrence of a broad-host-range IncP-1β plasmid in mycobacteria is reported for the first time. Thus, genetic exchange could result in the emergence of specific strains that might be better adapted to cause human disease.

  9. Plasmid transferability of KPC into a virulent K2 serotype Klebsiella pneumoniae.

    Science.gov (United States)

    Siu, Leung-Kei Kristopher; Huang, David B; Chiang, Tom

    2014-03-31

    KPC-producing carbapenem-resistant Klebsiella pneumoniae (CRKP) infections are associated with high mortality; however, their virulence determinants are not well defined. We investigated the virulence and plasmid transferability among KPC-containing K. pneumoniae isolates. KPC-2 and -3 were successfully conjugated and retained by a virulent K2 K. pneumoniae recipient isolate. Antimicrobial susceptibility testing showed KPC-2 and -3 donor strains were resistant to more than four classes of antibiotics while the K2 isolate was only initially resistant to ampicillin. After conjugation of KPC-2 and -3, the K2 K. pneumoniae transconjugants became resistant to all beta-lactams. Additionally, the KPC K2 K. pneumoniae transconjugants continued to retain its high serum resistance and murine lethality. Conjugation and retainment of KPC by virulent K2 K. pneumoniae and the ability of the tranconjugants to maintain its high serum resistance and murine lethality after conjugation was demonstrated in this study. These findings are concerning for the potential of KPC-like genes to disseminate among virulent K. pneumoniae isolates.

  10. Conjugate Problems in Convective Heat Transfer: Review

    Directory of Open Access Journals (Sweden)

    Abram Dorfman

    2009-01-01

    Full Text Available A review of conjugate convective heat transfer problems solved during the early and current time of development of this modern approach is presented. The discussion is based on analytical solutions of selected typical relatively simple conjugate problems including steady-state and transient processes, thermal material treatment, and heat and mass transfer in drying. This brief survey is accompanied by the list of almost two hundred publications considering application of different more and less complex analytical and numerical conjugate models for simulating technology processes and industrial devices from aerospace systems to food production. The references are combined in the groups of works studying similar problems so that each of the groups corresponds to one of selected analytical solutions considered in detail. Such structure of review gives the reader the understanding of early and current situation in conjugate convective heat transfer modeling and makes possible to use the information presented as an introduction to this area on the one hand, and to find more complicated publications of interest on the other hand.

  11. Unraveling the regulatory network of IncA/C plasmid mobilization: When genomic islands hijack conjugative elements.

    Science.gov (United States)

    Carraro, Nicolas; Matteau, Dominick; Burrus, Vincent; Rodrigue, Sébastien

    2015-01-01

    Conjugative plasmids of the A/C incompatibility group (IncA/C) have become substantial players in the dissemination of multidrug resistance. These large conjugative plasmids are characterized by their broad host-range, extended spectrum of antimicrobials resistance, and prevalence in enteric bacteria recovered from both environmental and clinical settings. Until recently, relatively little was known about the basic biology of IncA/C plasmids, mostly because of the hindrance of multidrug resistance for molecular biology experiments. To circumvent this issue, we previously developed pVCR94ΔX, a convenient prototype that codes for a reduced set of antibiotic resistances. Using pVCR94ΔX, we then characterized the regulatory pathway governing IncA/C plasmid dissemination. We found that the expression of roughly 2 thirds of the genes encoded by this plasmid, including large operons involved in the conjugation process, depends on an FlhCD-like master activator called AcaCD. Beyond the mobility of IncA/C plasmids, AcaCD was also shown to play a key role in the mobilization of different classes of genomic islands (GIs) identified in various pathogenic bacteria. By doing so, IncA/C plasmids can have a considerable impact on bacterial genomes plasticity and evolution.

  12. An In Vitro Chicken Gut Model Demonstrates Transfer of a Multidrug Resistance Plasmid from Salmonella to Commensal Escherichia coli.

    Science.gov (United States)

    Card, Roderick M; Cawthraw, Shaun A; Nunez-Garcia, Javier; Ellis, Richard J; Kay, Gemma; Pallen, Mark J; Woodward, Martin J; Anjum, Muna F

    2017-07-18

    The chicken gastrointestinal tract is richly populated by commensal bacteria that fulfill various beneficial roles for the host, including helping to resist colonization by pathogens. It can also facilitate the conjugative transfer of multidrug resistance (MDR) plasmids between commensal and pathogenic bacteria which is a significant public and animal health concern as it may affect our ability to treat bacterial infections. We used an in vitro chemostat system to approximate the chicken cecal microbiota, simulate colonization by an MDR Salmonella pathogen, and examine the dynamics of transfer of its MDR plasmid harboring several genes, including the extended-spectrum beta-lactamase bla CTX-M1 We also evaluated the impact of cefotaxime administration on plasmid transfer and microbial diversity. Bacterial community profiles obtained by culture-independent methods showed that Salmonella inoculation resulted in no significant changes to bacterial community alpha diversity and beta diversity, whereas administration of cefotaxime caused significant alterations to both measures of diversity, which largely recovered. MDR plasmid transfer from Salmonella to commensal Escherichia coli was demonstrated by PCR and whole-genome sequencing of isolates purified from agar plates containing cefotaxime. Transfer occurred to seven E. coli sequence types at high rates, even in the absence of cefotaxime, with resistant strains isolated within 3 days. Our chemostat system provides a good representation of bacterial interactions, including antibiotic resistance transfer in vivo It can be used as an ethical and relatively inexpensive approach to model dissemination of antibiotic resistance within the gut of any animal or human and refine interventions that mitigate its spread before employing in vivo studies. IMPORTANCE The spread of antimicrobial resistance presents a grave threat to public health and animal health and is affecting our ability to respond to bacterial infections

  13. Blending genomes: distributive conjugal transfer in mycobacteria, a sexier form of HGT.

    Science.gov (United States)

    Gray, Todd A; Derbyshire, Keith M

    2018-04-18

    This review discusses a novel form of horizontal gene transfer (HGT) found in mycobacteria called Distributive Conjugal Transfer (DCT). While satisfying the criteria for conjugation, DCT occurs by a mechanism so distinct from oriT-mediated conjugation that it could be considered a fourth category of HGT. DCT involves the transfer of chromosomal DNA between mycobacteria and, most significantly, generates transconjugants with mosaic genomes of the parental strains. Multiple segments of donor chromosomal DNA can be co-transferred regardless of their location or the genetic selection and, as a result, the transconjugant genome contains many donor-derived segments; hence the name DCT. This distinguishing feature of DCT separates it from the other known mechanisms of HGT, which generally result in the introduction of a single, defined segment of DNA into the recipient chromosome (Fig. ). Moreover, these mosaic progeny are generated from a single conjugal event, which provides enormous capacity for rapid adaptation and evolution, again distinguishing it from the three classical modes of HGT. Unsurprisingly, the unusual mosaic products of DCT are generated by a conjugal mechanism that is also unusual. Here, we will describe the unique features of DCT and contrast those to other mechanisms of HGT, both from a mechanistic and an evolutionary perspective. Our focus will be on transfer of chromosomal DNA, as opposed to plasmid mobilization, because DCT mediates transfer of chromosomal DNA and is a chromosomally encoded process. © 2018 John Wiley & Sons Ltd.

  14. Effect of thiol pendant conjugates on plasmid DNA binding, release, and stability of polymeric delivery vectors.

    Science.gov (United States)

    Bacalocostantis, Irene; Mane, Viraj P; Kang, Michael S; Goodley, Addison S; Muro, Silvia; Kofinas, Peter

    2012-05-14

    Polymers have attracted much attention as potential gene delivery vectors due to their chemical and structural versatility. However, several challenges associated with polymeric carriers, including low transfection efficiencies, insufficient cargo release, and high cytotoxicity levels have prevented clinical implementation. Strong electrostatic interactions between polymeric carriers and DNA cargo can prohibit complete cargo release within the cell. As a result, cargo DNA never reaches the cell's nucleus where gene expression takes place. In addition, highly charged cationic polymers have been correlated with high cytotoxicity levels, making them unsuitable carriers in vivo. Using poly(allylamine) (PAA) as a model, we investigated how pH-sensitive disulfide cross-linked polymer networks can improve the delivery potential of cationic polymer carriers. To accomplish this, we conjugated thiol-terminated pendant chains onto the primary amines of PAA using 2-iminothiolane, developing three new polymer vectors with 5, 13, or 20% thiol modification. Unmodified PAA and thiol-conjugated polymers were tested for their ability to bind and release plasmid DNA, their capacity to protect genetic cargo from enzymatic degradation, and their potential for endolysosomal escape. Our results demonstrate that polymer-plasmid complexes (polyplexes) formed by the 13% thiolated polymer demonstrate the greatest delivery potential. At high N/P ratios, all thiolated polymers (but not unmodified counterparts) were able to resist decomplexation in the presence of heparin, a negatively charged polysaccharide used to mimic in vivo polyplex-protein interactions. Further, all thiolated polymers exhibited higher buffering capacities than unmodified PAA and, therefore, have a greater potential for endolysosomal escape. However, 5 and 20% thiolated polymers exhibited poor DNA binding-release kinetics, making them unsuitable carriers for gene delivery. The 13% thiolated polymers, on the other hand

  15. Conjugate Heat Transfer Study in Hypersonic Flows

    Science.gov (United States)

    Sahoo, Niranjan; Kulkarni, Vinayak; Peetala, Ravi Kumar

    2018-04-01

    Coupled and decoupled conjugate heat transfer (CHT) studies are carried out to imitate experimental studies for heat transfer measurement in hypersonic flow regime. The finite volume based solvers are used for analyzing the heat interaction between fluid and solid domains. Temperature and surface heat flux signals are predicted by both coupled and decoupled CHT analysis techniques for hypersonic Mach numbers. These two methodologies are also used to study the effect of different wall materials on surface parameters. Effectiveness of these CHT solvers has been verified for the inverse problem of wall heat flux recovery using various techniques reported in the literature. Both coupled and decoupled CHT techniques are seen to be equally useful for prediction of local temperature and heat flux signals prior to the experiments in hypersonic flows.

  16. Double diffusive conjugate heat transfer: Part I

    Science.gov (United States)

    Azeem, Soudagar, Manzoor Elahi M.

    2018-05-01

    The present work is undertaken to investigate the effect of solid wall being placed at left of square cavity filled with porous medium. The presence of a solid wall in the porous medium turns the situation into a conjugate heat transfer problem. The boundary conditions are such that the left vertical surface is maintained at highest temperature and concentration whereas right vertical surface at lowest temperature and concentration in the medium. The top and bottom surfaces are adiabatic. The additional conduction equation along with the regular momentum and energy equations of porous medium are solved in an iterative manner with the help of finite element method. It is seen that the heat and mass transfer rate is lesser due to smaller thermal and concentration gradients.

  17. Theory of Periodic Conjugate Heat Transfer

    CERN Document Server

    Zudin, Yuri B

    2012-01-01

    This book presents the theory of periodic conjugate heat transfer in a detailed way. The effects of thermophysical properties and geometry of a solid body on the commonly used and experimentally determined heat transfer coefficient are analytically presented from a general point of view. The main objective of the book is a simplified description of the interaction between a solid body and a fluid as a boundary value problem of the heat conduction equation for the solid body. At the body surface, the true heat transfer coefficient is composed of two parts: the true mean value resulting from the solution of the steady state heat transfer problem and a periodically variable part, the periodic time and length to describe the oscillatory hydrodynamic effects. The second edition is extended by (i) the analysis of stability boundaries in helium flow at supercritical conditions in a heated channel with respect to the interaction between a solid body and a fluid; (ii) a periodic model and a method of heat transfer sim...

  18. Effects of different replicons in conjugative plasmids on transformation efficiency, plasmid stability, gene expression and n-butanol biosynthesis in Clostridium tyrobutyricum

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Mingrui; Du, Yinming; Jiang, Wenyan; Chang, Wei-Lun; Yang, Shang-Tian [Ohio State Univ., Columbus, OH (United States). William G. Lowrie Dept. of Chemical and Biomolecular Engineering; Tang, I-Ching [Bioprocessing Innovative Company, Dublin, OH (United States)

    2012-01-15

    Clostridium tyrobutyricum ATCC 25755 can produce butyric acid, acetic acid, and hydrogen as the main products from various carbon sources. In this study, C. tyrobutyricum was used as a host to produce n-butanol by expressing adhE2 gene under the control of a native thiolase promoter using four different conjugative plasmids (pMTL82151, 83151, 84151, and 85151) each with a different replicon (pBP1 from C. botulinum NCTC2916, pCB102 from C. butyricum, pCD6 from Clostridium difficile, and pIM13 from Bacillus subtilis). The effects of different replicons on transformation efficiency, plasmid stability, adhE2 expression and aldehyde/alcohol dehydrogenase activities, and butanol production by different mutants of C. tyrobutyricum were investigated. Among the four plasmids and replicons studied, pMTL82151 with pBP1 gave the highest transformation efficiency, plasmid stability, gene expression, and butanol biosynthesis. Butanol production from various substrates, including glucose, xylose, mannose, and mannitol were then investigated with the best mutant strain harboring adhE2 in pMTL82151. A high butanol titer of 20.5 g/L with 0.33 g/g yield and 0.32 g/L h productivity was obtained with mannitol as the substrate in batch fermentation with pH controlled at {proportional_to}6.0. (orig.)

  19. Circulation of a multiresistant, conjugative, IncA/C plasmid within the nosocomial Providencia stuartii population in the Athens area.

    Science.gov (United States)

    Giakkoupi, Panagiota; Tryfinopoulou, Kyriaki; Polemis, Michalis; Pappa, Olga; Miriagou, Vivi; Vatopoulos, Alkiviadis

    2015-05-01

    The objective of the study is to report a multidrug-resistant outbreak of Providencia stuartii that occurred in inpatients in the Athens area in 2012 resulting from a very successful transmissible A/C multidrug-resistant plasmid. Thirteen multidrug-resistant P. stuartii clinical isolates from 5 hospitals were studied. Molecular typing was performed by pulsed-field gel electrophoresis. Antibiotic resistance genes and their genetic surround were detected by PCR and sequencing. Plasmid analysis included conjugation experiments using liquid cultures, sizing by S1 digestion, and incompatibility replicon typing by PCR. Isolates were grouped into 2 distinct clonal types A and B, exhibiting similarity less than 70%. Isolates of type A were recovered from patients hospitalized in 4 different hospitals with no obvious epidemiological linkage, while isolates of type B were recovered from patients treated in a single hospital. Both clonal types harbored a conjugative plasmid of 130 bp and IncA/C replicon type carrying 5 β-lactamase genes bla(SHV-5), bla(VEB-1), bla(VIM-1), bla(OXA-10), and bla(TEM-1) and aminoglycosides resistant determinants. All β-lactamase genes were included in stable structures as IS26, IS1999, and In-e541. The current plasmid seemed to have many common determinants with previously reported plasmids derived from P. stuartii and Proteus mirabilis clinical isolates and exhibited the ability to circulate in nosocomial bacterial populations. Copyright © 2015 Elsevier Inc. All rights reserved.

  20. Identification of the rctA Gene, Which Is Required for Repression of Conjugative Transfer of Rhizobial Symbiotic Megaplasmids†

    Science.gov (United States)

    Pérez-Mendoza, Daniel; Sepúlveda, Edgardo; Pando, Victoria; Muñoz, Socorro; Nogales, Joaquina; Olivares, José; Soto, Maria J.; Herrera-Cervera, José A.; Romero, David; Brom, Susana; Sanjuán, Juan

    2005-01-01

    An analysis of the conjugative transfer of pRetCFN42d, the symbiotic plasmid (pSym) of Rhizobium etli, has revealed a novel gene, rctA, as an essential element of a regulatory system for silencing the conjugative transfer of R. etli pSym by repressing the transcription of conjugal transfer genes in standard laboratory media. The rctA gene product lacks sequence conservation with other proteins of known function but may belong to the winged-helix DNA-binding subfamily of transcriptional regulators. Similar to that of many transcriptional repressors, rctA transcription seems to be positively autoregulated. rctA expression is greatly reduced upon overexpression of another gene, rctB, previously identified as a putative activator of R. etli pSym conjugal transfer. Thus, rctB seems to counteract the repressive action of rctA. rctA homologs are present in at least three other bacterial genomes within the order Rhizobiales, where they are invariably located adjacent to and divergently transcribed from putative virB-like operons. We show that similar to that of R. etli pSym, conjugative transfer of the 1.35-Mb symbiotic megaplasmid A of Sinorhizobium meliloti is also subjected to the inhibitory action of rctA. Our data provide strong evidence that the R. etli and S. meliloti pSym plasmids are indeed self-conjugative plasmids and that this property would only be expressed under optimal, as yet unknown conditions that entail inactivation of the rctA function. The rctA gene seems to represent novel but probably widespread regulatory systems controlling the transfer of conjugative elements within the order Rhizobiales. PMID:16237017

  1. The multidrug resistance IncA/C transferable plasmid encodes a novel domain-swapped dimeric protein-disulfide isomerase.

    Science.gov (United States)

    Premkumar, Lakshmanane; Kurth, Fabian; Neyer, Simon; Schembri, Mark A; Martin, Jennifer L

    2014-01-31

    The multidrug resistance-encoding IncA/C conjugative plasmids disseminate antibiotic resistance genes among clinically relevant enteric bacteria. A plasmid-encoded disulfide isomerase is associated with conjugation. Sequence analysis of several IncA/C plasmids and IncA/C-related integrative and conjugative elements (ICE) from commensal and pathogenic bacteria identified a conserved DsbC/DsbG homolog (DsbP). The crystal structure of DsbP reveals an N-terminal domain, a linker region, and a C-terminal catalytic domain. A DsbP homodimer is formed through domain swapping of two DsbP N-terminal domains. The catalytic domain incorporates a thioredoxin-fold with characteristic CXXC and cis-Pro motifs. Overall, the structure and redox properties of DsbP diverge from the Escherichia coli DsbC and DsbG disulfide isomerases. Specifically, the V-shaped dimer of DsbP is inverted compared with EcDsbC and EcDsbG. In addition, the redox potential of DsbP (-161 mV) is more reducing than EcDsbC (-130 mV) and EcDsbG (-126 mV). Other catalytic properties of DsbP more closely resemble those of EcDsbG than EcDsbC. These catalytic differences are in part a consequence of the unusual active site motif of DsbP (CAVC); substitution to the EcDsbC-like (CGYC) motif converts the catalytic properties to those of EcDsbC. Structural comparison of the 12 independent subunit structures of DsbP that we determined revealed that conformational changes in the linker region contribute to mobility of the catalytic domain, providing mechanistic insight into DsbP function. In summary, our data reveal that the conserved plasmid-encoded DsbP protein is a bona fide disulfide isomerase and suggest that a dedicated oxidative folding enzyme is important for conjugative plasmid transfer.

  2. Conjugative plasmid pAW63 brings new insights into the genesis of the Bacillus anthracis virulence plasmid pXO2 and of the Bacillus thuringiensis plasmid pBT9727

    Directory of Open Access Journals (Sweden)

    Mahillon Jacques

    2005-07-01

    Full Text Available Abstract Background Bacillus cereus, Bacillus anthracis and Bacillus thuringiensis belong to the genetically close-knit Bacillus cereus sensu lato group, a family of rod-shaped Gram-positive bacteria. pAW63 is the first conjugative plasmid from the B. cereus group to be completely sequenced. Results The 71,777 bp nucleotide sequence of pAW63 reveals a modular structure, including a 42 kb tra region encoding homologs of the Type IV secretion systems components VirB11, VirB4 and VirD4, as well as homologs of Gram-positive conjugation genes from Enterococcus, Lactococcus, Listeria, Streptococcus and Staphylococcus species. It also firmly establishes the existence of a common backbone between pAW63, pXO2 from Bacillus anthracis and pBT9727 from the pathogenic Bacillus thuringiensis serovar konkukian strain 97-27. The alignment of these three plasmids highlights the presence of well conserved segments, in contrast to distinct regions of high sequence plasticity. The study of their specific differences has provided a three-point reference framework that can be exploited to formulate solid hypotheses concerning the functionalities and the molecular evolution of these three closely related plasmids. This has provided insight into the chronology of their divergence, and led to the discovery of two Type II introns on pAW63, matching copies of the mobile element IS231L in different loci of pXO2 and pBT9727, and the identification on pXO2 of a 37 kb pathogenicity island (PAI containing the anthrax capsule genes. Conclusion The complete sequence determination of pAW63 has led to a functional map of the plasmid yielding insights into its conjugative apparatus, which includes T4SS-like components, as well as its resemblance to other large plasmids of Gram-positive bacteria. Of particular interest is the extensive homology shared between pAW63 and pXO2, the second virulence plasmid of B. anthracis, as well as pBT9727 from the pathogenic strain B. thuringiensis

  3. Nucleotide Sequences and Comparison of Two Large Conjugative Plasmids from Different Campylobacter species

    National Research Council Canada - National Science Library

    Batchelor, Roger A; Pearson, Bruce M; Friis, Lorna M; Guerry, Patricia; Wells, Jerry M

    2004-01-01

    .... Both plasmids are mosaic in structure, having homologues of genes found in a variety of different commensal and pathogenic bacteria, but nevertheless, showed striking similarities in DNA sequence...

  4. Intergenic and intragenic conjugal transfer of multiple antibiotic ...

    African Journals Online (AJOL)

    Conjugation process was conducted to determine the means of transferring ... In this study, it was surprisingly observed that tetracycline resistant gene was ... among pathogenic bacteria, particularly since antibiotics are indiscriminately used in ...

  5. Bacteriophytochromes control conjugation in Agrobacterium fabrum.

    Science.gov (United States)

    Bai, Yingnan; Rottwinkel, Gregor; Feng, Juan; Liu, Yiyao; Lamparter, Tilman

    2016-08-01

    Bacterial conjugation, the transfer of single stranded plasmid DNA from donor to recipient cell, is mediated through the type IV secretion system. We performed conjugation assays using a transmissible artificial plasmid as reporter. With this assay, conjugation in Agrobacterium fabrum was modulated by the phytochromes Agp1 and Agp2, photoreceptors that are most sensitive in the red region of visible light. In conjugation studies with wild-type donor cells carrying a pBIN-GUSINT plasmid as reporter that lacked the Ti (tumor inducing) plasmid, no conjugation was observed. When either agp1(-) or agp2(-) knockout donor strains were used, plasmid DNA was delivered to the recipient, indicating that both phytochromes suppress conjugation in the wild type donor. In the recipient strains, the loss of Agp1 or Agp2 led to diminished conjugation. When wild type cells with Ti plasmid and pBIN-GUS reporter plasmid were used as donor, a high rate of conjugation was observed. The DNA transfer was down regulated by red or far-red light by a factor of 3.5. With agp1(-) or agp2(-) knockout donor cells, conjugation in the dark was about 10 times lower than with the wild type donor, and with the double knockout donor no conjugation was observed. These results imply that the phytochrome system has evolved to inhibit conjugation in the light. The decrease of conjugation under different temperature correlated with the decrease of phytochrome autophosphorylation. Copyright © 2015 Elsevier B.V. All rights reserved.

  6. Requirement for Vibrio cholerae integration host factor in conjugative DNA transfer.

    Science.gov (United States)

    McLeod, Sarah M; Burrus, Vincent; Waldor, Matthew K

    2006-08-01

    The requirement for host factors in the transmission of integrative and conjugative elements (ICEs) has not been extensively explored. Here we tested whether integration host factor (IHF) or Fis, two host-encoded nucleoid proteins, are required for transfer of SXT, a Vibrio cholerae-derived ICE that can be transmitted to many gram-negative species. Fis did not influence the transfer of SXT to or from V. cholerae. In contrast, IHF proved to be required for V. cholerae to act as an SXT donor. In the absence of IHF, V. cholerae displayed a modest defect for serving as an SXT recipient. Surprisingly, SXT integration into or excision from the V. cholerae chromosome, which requires an SXT-encoded integrase related to lambda integrase, did not require IHF. Therefore, the defect in SXT transmission in the V. cholerae IHF mutant is probably not related to IHF's ability to promote DNA recombination. The V. cholerae IHF mutant was also highly impaired as a donor of RP4, a broad-host-range conjugative plasmid. Thus, the V. cholerae IHF mutant appears to have a general defect in conjugation. Escherichia coli IHF mutants were not impaired as donors or recipients of SXT or RP4, indicating that IHF is a V. cholerae-specific conjugation factor.

  7. A genetic study of a Staphylococus aureus plasmid involving cure and transference

    Directory of Open Access Journals (Sweden)

    Ana Lúcia Costa Darini

    Full Text Available High frequency transfer and elimination of drug resistance may indicate an extrachromosomal inheritance of genetic determinants. This study shows the cure and transfer of a small plasmid and tetracycline resistance in Staphylococcus aureus 1030 (55TetR strains. Several methods are available for plasmid elimination. We used ethidium bromide, an agent that binds to DNA, and thus inhibits DNA polymerase. This caused a high frequency of loss of the small plasmid and resistance to tetracycline. Transfer of tetracycline resistance was done in a mixed culture at a frequency of 10-6. This type of study is very important to physicians and epidemiology investigators and provides better knowledge on antibiotic-resistance mechanisms that may occur in vivo in a hospital environment.

  8. Transfer and Persistence of a Multi-Drug Resistance Plasmid in situ of the Infant Gut Microbiota in the Absence of Antibiotic Treatment

    Directory of Open Access Journals (Sweden)

    Heidi Gumpert

    2017-09-01

    Full Text Available The microbial ecosystem residing in the human gut is believed to play an important role in horizontal exchange of virulence and antibiotic resistance genes that threatens human health. While the diversity of gut-microorganisms and their genetic content has been studied extensively, high-resolution insight into the plasticity, and selective forces shaping individual genomes is scarce. In a longitudinal study, we followed the dynamics of co-existing Escherichia coli lineages in an infant not receiving antibiotics. Using whole genome sequencing, we observed large genomic deletions, bacteriophage infections, as well as the loss and acquisition of plasmids in these lineages during their colonization of the human gut. In particular, we captured the exchange of multidrug resistance genes, and identified a clinically relevant conjugative plasmid mediating the transfer. This resistant transconjugant lineage was maintained for months, demonstrating that antibiotic resistance genes can disseminate and persist in the gut microbiome; even in absence of antibiotic selection. Furthermore, through in vivo competition assays, we suggest that the resistant transconjugant can persist through a fitness advantage in the mouse gut in spite of a fitness cost in vitro. Our findings highlight the dynamic nature of the human gut microbiota and provide the first genomic description of antibiotic resistance gene transfer between bacteria in the unperturbed human gut. These results exemplify that conjugative plasmids, harboring resistance determinants, can transfer and persists in the gut in the absence of antibiotic treatment.

  9. Conjugal gene transfer between bacteria in soil and rhizosphere

    NARCIS (Netherlands)

    Smit, E.

    1994-01-01

    The extent of possible conjugal transfer of recombinant DNA present in genetically engineered microorganisms (GEMs) was studied. Occurrence of transfer of recombinant DNA is only one of the concerns regarding the use of GEMs (Chapter 2). Other potential hazards preventing the application of

  10. Selective pressure affects transfer and establishment of a Lactobacillus plantarum resistance plasmid in the gastrointestinal environment

    DEFF Research Database (Denmark)

    Feld, Louise; Schjorring, S.; Hammer, Karin

    2008-01-01

    Objectives and methods: A Lactobacillus plantarum strain recently isolated from French raw-milk cheese was tested for its ability to transfer a small plasmid pLFE1 harbouring the erythromycin resistance gene erm(B) to Enterococcus faecalis. Mating was studied in vitro and in different gastrointes......Objectives and methods: A Lactobacillus plantarum strain recently isolated from French raw-milk cheese was tested for its ability to transfer a small plasmid pLFE1 harbouring the erythromycin resistance gene erm(B) to Enterococcus faecalis. Mating was studied in vitro and in different...

  11. High-level fluoroquinolone resistant Salmonella enterica serovar Kentucky ST198 epidemic clone with IncA/C conjugative plasmid carrying bla(CTX-M-25) gene.

    Science.gov (United States)

    Wasyl, Dariusz; Kern-Zdanowicz, Izabela; Domańska-Blicharz, Katarzyna; Zając, Magdalena; Hoszowski, Andrzej

    2015-01-30

    Multidrug resistant Salmonella Kentucky strains have been isolated from turkeys in Poland since 2009. Multiple mutations within chromosomal genes gyrA and parC were responsible for high-level ciprofloxacin resistance. One of the isolates was extended spectrum β-lactamase- (ESBL) positive: the strain 1643/2010 carried a conjugative 167,779 bps plasmid of IncA/C family. The sequence analysis revealed that it carried a blaCTX-M-25 gene and an integron with another β-lactamase encoding gene-blaOXA-21. This is the first known report of a CTX-M-25 encoding gene both in Poland and in Salmonella Kentucky world-wide, as well as in the IncA/C plasmid. Analysis of the integron showed a novel arrangement of gene cassettes-aacA4, aacC-A1 and blaOXA-21 where the latter might result from an intergeneric gene transfer. The study confirmed Salmonella Kentucky population isolated in Poland belongs to global epidemics of high level fluoroquinolone resistant clone ST198 that can carry rare β-lactamase genes. Copyright © 2014 Elsevier B.V. All rights reserved.

  12. Conjugate Image Theory Applied on Capacitive Wireless Power Transfer

    OpenAIRE

    Ben Minnaert; Nobby Stevens

    2017-01-01

    Wireless power transfer using a magnetic field through inductive coupling is steadily entering the market in a broad range of applications. However, for certain applications, capacitive wireless power transfer using electric coupling might be preferable. In order to obtain a maximum power transfer efficiency, an optimal compensation network must be designed at the input and output ports of the capacitive wireless link. In this work, the conjugate image theory is applied to determine this opti...

  13. Conjugate heat and mass transfer in heat mass exchanger ducts

    CERN Document Server

    Zhang, Li-Zhi

    2013-01-01

    Conjugate Heat and Mass Transfer in Heat Mass Exchanger Ducts bridges the gap between fundamentals and recent discoveries, making it a valuable tool for anyone looking to expand their knowledge of heat exchangers. The first book on the market to cover conjugate heat and mass transfer in heat exchangers, author Li-Zhi Zhang goes beyond the basics to cover recent advancements in equipment for energy use and environmental control (such as heat and moisture recovery ventilators, hollow fiber membrane modules for humidification/dehumidification, membrane modules for air purification, desi

  14. Quantifying and visualizing the transfer of exogenous plasmids to environmental microbial communities

    DEFF Research Database (Denmark)

    Dechesne, Arnaud

    2015-01-01

    range plasmids, with common transfer across the Gram ‘barrier’. We next looked for factors that modulate permissiveness and, in particular, identified a taxon-specific effect imposed by metals when supplemented in concentrations that cause partial inhibition of the community metabolic activity. Overall...

  15. Minimal and contributing sequence determinants of the cis-acting locus of transfer (clt) of streptomycete plasmid pIJ101 occur within an intrinsically curved plasmid region.

    Science.gov (United States)

    Ducote, M J; Prakash, S; Pettis, G S

    2000-12-01

    Efficient interbacterial transfer of streptomycete plasmid pIJ101 requires the pIJ101 tra gene, as well as a cis-acting plasmid function known as clt. Here we show that the minimal pIJ101 clt locus consists of a sequence no greater than 54 bp in size that includes essential inverted-repeat and direct-repeat sequences and is located in close proximity to the 3' end of the korB regulatory gene. Evidence that sequences extending beyond the minimal locus and into the korB open reading frame influence clt transfer function and demonstration that clt-korB sequences are intrinsically curved raise the possibility that higher-order structuring of DNA and protein within this plasmid region may be an inherent feature of efficient pIJ101 transfer.

  16. Large IncHI2-plasmids encode extended-spectrum β-lactamases (ESBLs) in Enterobacter spp. bloodstream isolates, and support ESBL-transfer to Escherichia coli.

    Science.gov (United States)

    Nilsen, E; Haldorsen, B C; Sundsfjord, A; Simonsen, G S; Ingebretsen, A; Naseer, U; Samuelsen, O

    2013-11-01

    We investigated the prevalence of extended-spectrum β-lactamases (ESBLs) in Enterobacter spp. bloodstream isolates from 19 hospital laboratories in Norway during 2011. A total of 62/230 (27%) isolates were resistant to third-generation cephalosporins and four (1.7%) were ESBL-positive; blaCTX -M-15 (n = 3) and blaSHV -12 (n = 1). This is comparable to the prevalence of ESBLs in clinical isolates of Escherichia coli and Klebsiella pneumoniae in Norway during the same period. All ESBL-positive isolates were multidrug resistant (MDR) and harboured plasmid-mediated quinolone resistance. Three isolates supported transfer of large IncHI2-plasmids harbouring ESBL- and MDR-encoding genes to E. coli recipients by in vitro conjugation. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  17. Transfer and persistence of a multi-drug resistance plasmid in situ of the infant gut microbiota in the absence of antibiotic treatment

    DEFF Research Database (Denmark)

    Gumpert, Heidi; Kubicek-Sutherland, Jessica Z.; Porse, Andreas

    2017-01-01

    lineage was maintained for months, demonstrating that antibiotic resistance genes can disseminate and persist in the gut microbiome; even in absence of antibiotic selection. Furthermore, through in vivo competition assays, we suggest that the resistant transconjugant can persist through a fitness......The microbial ecosystem residing in the human gut is believed to play an important role in horizontal exchange of virulence and antibiotic resistance genes that threatens human health. While the diversity of gut-microorganisms and their genetic content has been studied extensively, high...... infections, as well as the loss and acquisition of plasmids in these lineages during their colonization of the human gut. In particular, we captured the exchange of multidrug resistance genes, and identified a clinically relevant conjugative plasmid mediating the transfer. This resistant transconjugant...

  18. Numerical Modeling of Conjugate Heat Transfer in Fluid Network

    Science.gov (United States)

    Majumdar, Alok

    2004-01-01

    Fluid network modeling with conjugate heat transfer has many applications in Aerospace engineering. In modeling unsteady flow with heat transfer, it is important to know the variation of wall temperature in time and space to calculate heat transfer between solid to fluid. Since wall temperature is a function of flow, a coupled analysis of temperature of solid and fluid is necessary. In cryogenic applications, modeling of conjugate heat transfer is of great importance to correctly predict boil-off rate in propellant tanks and chill down of transfer lines. In TFAWS 2003, the present author delivered a paper to describe a general-purpose computer program, GFSSP (Generalized Fluid System Simulation Program). GFSSP calculates flow distribution in complex flow circuit for compressible/incompressible, with or without heat transfer or phase change in all real fluids or mixtures. The flow circuit constitutes of fluid nodes and branches. The mass, energy and specie conservation equations are solved at the nodes where as momentum conservation equations are solved at the branches. The proposed paper describes the extension of GFSSP to model conjugate heat transfer. The network also includes solid nodes and conductors in addition to fluid nodes and branches. The energy conservation equations for solid nodes solves to determine the temperatures of the solid nodes simultaneously with all conservation equations governing fluid flow. The numerical scheme accounts for conduction, convection and radiation heat transfer. The paper will also describe the applications of the code to predict chill down of cryogenic transfer line and boil-off rate of cryogenic propellant storage tank.

  19. Conjugate Compressible Fluid Flow and Heat Transfer in Ducts

    Science.gov (United States)

    Cross, M. F.

    2011-01-01

    A computational approach to modeling transient, compressible fluid flow with heat transfer in long, narrow ducts is presented. The primary application of the model is for analyzing fluid flow and heat transfer in solid propellant rocket motor nozzle joints during motor start-up, but the approach is relevant to a wide range of analyses involving rapid pressurization and filling of ducts. Fluid flow is modeled through solution of the spatially one-dimensional, transient Euler equations. Source terms are included in the governing equations to account for the effects of wall friction and heat transfer. The equation solver is fully-implicit, thus providing greater flexibility than an explicit solver. This approach allows for resolution of pressure wave effects on the flow as well as for fast calculation of the steady-state solution when a quasi-steady approach is sufficient. Solution of the one-dimensional Euler equations with source terms significantly reduces computational run times compared to general purpose computational fluid dynamics packages solving the Navier-Stokes equations with resolved boundary layers. In addition, conjugate heat transfer is more readily implemented using the approach described in this paper than with most general purpose computational fluid dynamics packages. The compressible flow code has been integrated with a transient heat transfer solver to analyze heat transfer between the fluid and surrounding structure. Conjugate fluid flow and heat transfer solutions are presented. The author is unaware of any previous work available in the open literature which uses the same approach described in this paper.

  20. Nucleotide Sequences and Comparison of Two Large Conjugative Plasmids from Different Campylobacter species

    Science.gov (United States)

    2004-01-01

    predicted protein of 24 kDa that shows 33% identity and 56% similarity to an invertase from Shewanella oneidensis. Invertases and resolvases have been...genes encoding the putative enzymes involved in DNA processing and transfer such as the nickase, helicase, primase, invertase and single-stand-binding...139–153. Edited by I. Nachamkin & M. J. Blaser. Washington, DC: American Society for Microbiology. Pansegrau, W. & Lanka, E. (1996). Enzymology of

  1. A standardized conjugation protocol to asses antibiotic resistance transfer between lactococcal species

    DEFF Research Database (Denmark)

    Lampkowska, Joanna; Feld, Louise; Monaghan, Aine

    2008-01-01

    Optimal conditions and a standardized method for conjugation between two model lactococcal strains, Lactococcus lactis SH4174 (pAM beta 1-containing, erythromycin resistant donor) and L. lactis Bu2-60 (plasmid-free, erythromycin sensitive recipient), were developed and tested in a inter-laborator...

  2. A standardized conjugation protocol to assess antibiotic resistance transfer between lactococcal species

    NARCIS (Netherlands)

    Lampkowska, J.; Feld, L.; Monaghan, A.; Toomey, N.; Schjørring, S.; Jacobsen, B.; Voet, van der H.; Andersen, S.R.; Bolton, D.; Aarts, H.J.M.; Krogfelt, K.A.; Wilcks, A.; Bardowski, J.K.

    2008-01-01

    Optimal conditions and a standardized method for conjugation between two model lactococcal strains, Lactococcus lactis SH4174 (pAMbeta1-containing, erythromycin resistant donor) and L. lactis Bu2-60 (plasmid-free, erythromycin sensitive recipient), were developed and tested in a inter-laboratory

  3. Material Selection for Microchannel Heatsink: Conjugate Heat Transfer Simulation

    Science.gov (United States)

    Uday Kumar, A.; Javed, Arshad; Dubey, Satish K.

    2018-04-01

    Heat dissipation during the operation of electronic devices causes rise in temperature, which demands an effective thermal management for their performance, life and reliability. Single phase liquid cooling in microchannels is an effective and proven technology for electronics cooling. However, due to the ongoing trends of miniaturization and developments in the microelectronics technology, the future needs of heat flux dissipation rate are expected to rise to 1 kW/cm2. Air cooled systems are unable to meet this demand. Hence, liquid cooled heatsinks are preferred. This paper presents conjugate heat transfer simulation of single phase flow in microchannels with application to electronic cooling. The numerical model is simulated for different materials: copper, aluminium and silicon as solid and water as liquid coolant. The performances of microchannel heatsink are analysed for mass flow rate range of 20-40 ml/min. The investigation has been carried out on same size of electronic chip and heat flux in order to have comparative study of different materials. This paper is divided into two sections: fabrication techniques and numerical simulation for different materials. In the first part, a brief discussion of fabrication techniques of microchannel heatsink have been presented. The second section presents conjugate heat transfer simulation and parametric investigation for different material microchannel heatsink. The presented study and findings are useful for selection of materials for microchannel heatsink.

  4. Conjugate Image Theory Applied on Capacitive Wireless Power Transfer

    Directory of Open Access Journals (Sweden)

    Ben Minnaert

    2017-01-01

    Full Text Available Wireless power transfer using a magnetic field through inductive coupling is steadily entering the market in a broad range of applications. However, for certain applications, capacitive wireless power transfer using electric coupling might be preferable. In order to obtain a maximum power transfer efficiency, an optimal compensation network must be designed at the input and output ports of the capacitive wireless link. In this work, the conjugate image theory is applied to determine this optimal network as a function of the characteristics of the capacitive wireless link, as well for the series as for the parallel topology. The results are compared with the inductive power transfer system. Introduction of a new concept, the coupling function, enables the description of the compensation network of both an inductive and a capacitive system in two elegant equations, valid for the series and the parallel topology. This approach allows better understanding of the fundamentals of the wireless power transfer link, necessary for the design of an efficient system.

  5. Dynamic impedance compensation for wireless power transfer using conjugate power

    Science.gov (United States)

    Liu, Suqi; Tan, Jianping; Wen, Xue

    2018-02-01

    Wireless power transfer (WPT) via coupled magnetic resonances has been in development for over a decade. However, the frequency splitting phenomenon occurs in the over-coupled region. Thus, the output power of the two-coil system achieves the maximum output power at the two splitting angular frequencies, and not at the natural resonant angular frequency. According to the maximum power transfer theorem, the impedance compensation method was adopted in many WPT projects. However, it remains a challenge to achieve the maximum output power and transmission efficiency in a fixed-frequency mode. In this study, dynamic impedance compensation for WPT was presented by utilizing the compensator within a virtual three-coil WPT system. First, the circuit model was established and transfer characteristics of a system were studied by utilizing circuit theories. Second, the power superposition of the WPT system was carefully researched. When a pair of compensating coils was inserted into the transmitter loop, the conjugate power of the compensator loop was created via magnetic coupling of the two compensating coils that insert into the transmitter loop. The mechanism for dynamic impedance compensation for wireless power transfer was then provided by investigating a virtual three-coil WPT system. Finally, the experimental circuit of a virtual three-coil WPT system was designed, and experimental results are consistent with the theoretical analysis, which achieves the maximum output power and transmission efficiency.

  6. Dynamic impedance compensation for wireless power transfer using conjugate power

    Directory of Open Access Journals (Sweden)

    Suqi Liu

    2018-02-01

    Full Text Available Wireless power transfer (WPT via coupled magnetic resonances has been in development for over a decade. However, the frequency splitting phenomenon occurs in the over-coupled region. Thus, the output power of the two-coil system achieves the maximum output power at the two splitting angular frequencies, and not at the natural resonant angular frequency. According to the maximum power transfer theorem, the impedance compensation method was adopted in many WPT projects. However, it remains a challenge to achieve the maximum output power and transmission efficiency in a fixed-frequency mode. In this study, dynamic impedance compensation for WPT was presented by utilizing the compensator within a virtual three-coil WPT system. First, the circuit model was established and transfer characteristics of a system were studied by utilizing circuit theories. Second, the power superposition of the WPT system was carefully researched. When a pair of compensating coils was inserted into the transmitter loop, the conjugate power of the compensator loop was created via magnetic coupling of the two compensating coils that insert into the transmitter loop. The mechanism for dynamic impedance compensation for wireless power transfer was then provided by investigating a virtual three-coil WPT system. Finally, the experimental circuit of a virtual three-coil WPT system was designed, and experimental results are consistent with the theoretical analysis, which achieves the maximum output power and transmission efficiency.

  7. Virus-sized self-assembling lamellar complexes between plasmid DNA and cationic micelles promote gene transfer

    Science.gov (United States)

    Pitard, Bruno; Aguerre, Olivier; Airiau, Marc; Lachagès, Anne-Marie; Boukhnikachvili, Tsiala; Byk, Gérardo; Dubertret, Catherine; Herviou, Christian; Scherman, Daniel; Mayaux, Jean-François; Crouzet, Joël

    1997-01-01

    Gene therapy is based on the vectorization of genes to target cells and their subsequent expression. Cationic amphiphile-mediated delivery of plasmid DNA is the nonviral gene transfer method most often used. We examined the supramolecular structure of lipopolyamine/plasmid DNA complexes under various condensing conditions. Plasmid DNA complexation with lipopolyamine micelles whose mean diameter was 5 nm revealed three domains, depending on the lipopolyamine/plasmid DNA ratio. These domains respectively corresponded to negatively, neutrally, and positively charged complexes. Transmission electron microscopy and x-ray scattering experiments on complexes originating from these three domains showed that although their morphology depends on the lipopolyamine/plasmid DNA ratio, their particle structure consists of ordered domains characterized by even spacing of 80 Å, irrespective of the lipid/DNA ratio. The most active lipopolyamine/DNA complexes for gene transfer were positively charged. They were characterized by fully condensed DNA inside spherical particles (diameter: 50 nm) sandwiched between lipid bilayers. These results show that supercoiled plasmid DNA is able to transform lipopolyamine micelles into a supramolecular organization characterized by ordered lamellar domains. PMID:9405626

  8. Conjugative transfer of clostridial shuttle vectors from Escherichia coli to Clostridium difficile through circumvention of the restriction barrier.

    Science.gov (United States)

    Purdy, Des; O'Keeffe, Triona A T; Elmore, Michael; Herbert, Mike; McLeod, Anne; Bokori-Brown, Monika; Ostrowski, Anna; Minton, Nigel P

    2002-10-01

    Progress towards understanding the molecular basis of virulence in Clostridium difficile has been hindered by the lack of effective gene transfer systems. We have now, for the first time, developed procedures that may be used to introduce autonomously replicating vectors into this organism through their conjugative, oriT-based mobilization from Escherichia coli donors. Successful transfer was achieved through the use of a plasmid replicon isolated from an indigenous C. difficile plasmid, pCD6, and through the characterization and subsequent circumvention of host restriction/modification (RM) systems. The characterized replicon is the first C. difficile plasmid replicon to be sequenced and encodes a large replication protein (RepA) and a repetitive region composed of a 35 bp iteron sequence repeated seven times. Strain CD6 has two RM systems, CdiCD6I/M.CdiCD6I and CdiCD6II/M. CdiCD6II, with equivalent specificities to Sau96I/M. Sau96I (5'-GGNMCC-3') and MboI/M. MboI (5'-GMATC-3') respectively. A second strain (CD3) possesses a type IIs restriction enzyme, Cdi I, which cleaves the sequence 5'-CATCG-3' between the fourth and fifth nucleotide to give a blunt-ended fragment. This is the first time that an enzyme with this specificity has been reported. The sequential addition of this site to vectors showed that each site caused between a five- and 16-fold reduction in transfer efficiency. The transfer efficiencies achieved with both strains equated to between 1.0 x 10-6 and 5.5 x 10-5 transconjugants per donor.

  9. Polyhedral meshing in numerical analysis of conjugate heat transfer

    Science.gov (United States)

    Sosnowski, Marcin; Krzywanski, Jaroslaw; Grabowska, Karolina; Gnatowska, Renata

    2018-06-01

    Computational methods have been widely applied in conjugate heat transfer analysis. The very first and crucial step in such research is the meshing process which consists in dividing the analysed geometry into numerous small control volumes (cells). In Computational Fluid Dynamics (CFD) applications it is desirable to use the hexahedral cells as the resulting mesh is characterized by low numerical diffusion. Unfortunately generating such mesh can be a very time-consuming task and in case of complicated geometry - it may not be possible to generate cells of good quality. Therefore tetrahedral cells have been implemented into commercial pre-processors. Their advantage is the ease of its generation even in case of very complex geometry. On the other hand tetrahedrons cannot be stretched excessively without decreasing the mesh quality factor, so significantly larger number of cells has to be used in comparison to hexahedral mesh in order to achieve a reasonable accuracy. Moreover the numerical diffusion of tetrahedral elements is significantly higher. Therefore the polyhedral cells are proposed within the paper in order to combine the advantages of hexahedrons (low numerical diffusion resulting in accurate solution) and tetrahedrons (rapid semi-automatic generation) as well as to overcome the disadvantages of both the above mentioned mesh types. The major benefit of polyhedral mesh is that each individual cell has many neighbours, so gradients can be well approximated. Polyhedrons are also less sensitive to stretching than tetrahedrons which results in better mesh quality leading to improved numerical stability of the model. In addition, numerical diffusion is reduced due to mass exchange over numerous faces. This leads to a more accurate solution achieved with a lower cell count. Therefore detailed comparison of numerical modelling results concerning conjugate heat transfer using tetrahedral and polyhedral meshes is presented in the paper.

  10. Conjugate heat transfer simulations of advanced research reactor fuel

    Energy Technology Data Exchange (ETDEWEB)

    Piro, M.H.A., E-mail: pirom@aecl.ca; Leitch, B.W.

    2014-07-01

    Highlights: • Temperature predictions are enhanced by coupling heat transfer in solid and fluid zones. • Seven different cases are considered to observe trends in predicted temperature and pressure. • The seven cases consider high/medium/low power, flow, burnup, fuel material and geometry. • Simulations provide temperature predictions for performance/safety. Boiling is unlikely. • Simulations demonstrate that a candidate geometry can enhance performance/safety. - Abstract: The current work presents numerical simulations of coupled fluid flow and heat transfer of advanced U–Mo/Al and U–Mo/Mg research reactor fuels in support of performance and safety analyses. The objective of this study is to enhance predictions of the flow regime and fuel temperatures through high fidelity simulations that better capture various heat transfer pathways and with a more realistic geometric representation of the fuel assembly in comparison to previous efforts. Specifically, thermal conduction, convection and radiation mechanisms are conjugated between the solid and fluid regions. Also, a complete fuel element assembly is represented in three dimensional space, permitting fluid flow and heat transfer to be simulated across the entire domain. Seven case studies are examined that vary the coolant inlet conditions, specific power, and burnup to investigate the predicted changes in the pressure drop in the coolant and the fuel, clad and coolant temperatures. In addition, an alternate fuel geometry is considered with helical fins (replacing straight fins in the existing design) to investigate the relative changes in predicted fluid and solid temperatures. Numerical simulations predict that the clad temperature is sensitive to changes in the thermal boundary layer in the coolant, particularly in simultaneously developing flow regions, while the temperature in the fuel is anticipated to be unaffected. Finally, heat transfer between fluid and solid regions is enhanced with

  11. Transfer of the pheromone-inducible plasmid pCF10 among Enterococcus faecalis microorganisms colonizing the intestine of mini-pigs

    DEFF Research Database (Denmark)

    Licht, Tine Rask; Laugesen, D.; Jensen, Lars Bogø

    2002-01-01

    A new animal model, the streptomycin-treated mini-pig, was developed in order to allow colonization of defined strains of Enterococcus faecalis in numbers sufficient to study plasmid transfer. Transfer of the pheromone-inducible pCF10 plasmid between streptomycin-resistant strains of E. faecalis OG...

  12. Liposome-enhanced transformation of Streptococcus lactis and plasmid transfer by intergeneric protoplast fusion of Streptococcus lactis and Bacillus subtilis

    NARCIS (Netherlands)

    Vossen, Jos M.B.M. van der; Kok, Jan; Lelie, Daniel van der; Venema, Gerhardus

    An efficient protoplast transformation system and a procedure of plasmid transfer by means of protoplast fusion is described for Streptococcus lactis. Protoplasts of S. lactis IL1403 and S. lactis MG1363 were transformed by pGK12 [2.9 MDa erythromycin resistance (Emr)] with an efficiency of 3 × 10^5

  13. An immersed-boundary method for conjugate heat transfer analysis

    Energy Technology Data Exchange (ETDEWEB)

    Song, Jeong Chul; Lee, Joon Sik [Seoul National University, Seoul (Korea, Republic of); Ahn, Joon [Kookmin University, Seoul (Korea, Republic of)

    2017-05-15

    An immersed-boundary method is proposed for the analysis of conjugate problems of convective heat transfer in conducting solids. In- side the solid body, momentum forcing is applied to set the velocity to zero. A thermal conductivity ratio and a heat capacity ratio, between the solid body and the fluid, are introduced so that the energy equation is reduced to the heat diffusion equation. At the solid fluid interface, an effective conductivity is introduced to satisfy the heat flux continuity. The effective thermal conductivity is obtained by considering the heat balance at the interface or by using a harmonic mean formulation. The method is first validated against the analytic solution to the heat transfer problem in a fully developed laminar channel flow with conducting solid walls. Then it is applied to a laminar channel flow with a heated, block-shaped obstacle to show its validity for geometry with sharp edges. Finally the validation for a curvilinear solid body is accomplished with a laminar flow through arrayed cylinders.

  14. Plasmids foster diversification and adaptation of bacterial populations in soil.

    Science.gov (United States)

    Heuer, Holger; Smalla, Kornelia

    2012-11-01

    It is increasingly being recognized that the transfer of conjugative plasmids across species boundaries plays a vital role in the adaptability of bacterial populations in soil. There are specific driving forces and constraints of plasmid transfer within bacterial communities in soils. Plasmid-mediated genetic variation allows bacteria to respond rapidly with adaptive responses to challenges such as irregular antibiotic or metal concentrations, or opportunities such as the utilization of xenobiotic compounds. Cultivation-independent detection and capture of plasmids from soil bacteria, and complete sequencing have provided new insights into the role and ecology of plasmids. Broad host range plasmids such as those belonging to IncP-1 transfer a wealth of accessory functions which are carried by similar plasmid backbones. Plasmids with a narrower host range can be more specifically adapted to particular species and often transfer genes which complement chromosomally encoded functions. Plasmids seem to be an ancient and successful strategy to ensure survival of a soil population in spatial and temporal heterogeneous conditions with various environmental stresses or opportunities that occur irregularly or as a novel challenge in soil. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  15. Mutation in ESBL Plasmid from Escherichia coli O104:H4 Leads Autoagglutination and Enhanced Plasmid Dissemination

    Directory of Open Access Journals (Sweden)

    Mickaël Poidevin

    2018-02-01

    Full Text Available Conjugative plasmids are one of the main driving force of wide-spreading of multidrug resistance (MDR bacteria. They are self-transmittable via conjugation as carrying the required set of genes and cis-acting DNA locus for direct cell-to-cell transfer. IncI incompatibility plasmids are nowadays often associated with extended-spectrum beta-lactamases producing Enterobacteria in clinic and environment. pESBL-EA11 was isolated from Escherichia coli O104:H4 outbreak strain in Germany in 2011. During the previous study identifying transfer genes of pESBL-EA11, it was shown that transposon insertion at certain DNA region of the plasmid, referred to as Hft, resulted in great enhancement of transfer ability. This suggested that genetic modifications can enhance dissemination of MDR plasmids. Such ‘superspreader’ mutations have attracted little attention so far despite their high potential to worsen MDR spreading. Present study aimed to gain our understanding on regulatory elements that involved pESBL transfer. While previous studies of IncI plasmids indicated that immediate downstream gene of Hft, traA, is not essential for conjugative transfer, here we showed that overexpression of TraA in host cell elevated transfer rate of pESBL-EA11. Transposon insertion or certain nucleotide substitutions in Hft led strong TraA overexpression which resulted in activation of essential regulator TraB and likely overexpression of conjugative pili. Atmospheric Scanning Electron Microscopy observation suggested that IncI pili are distinct from other types of conjugative pili (such as long filamentous F-type pili and rather expressed throughout the cell surface. High transfer efficiency in the mutant pESBL-EA11 was involved with hyperpiliation which facilitates cell-to-cell adhesion, including autoagglutination. The capability of plasmids to evolve to highly transmissible mutant is alarming, particularly it might also have adverse effect on host pathogenicity.

  16. Direct repeat sequences are essential for function of the cis-acting locus of transfer (clt) of Streptomyces phaeochromogenes plasmid pJV1.

    Science.gov (United States)

    Franco, Bernardo; González-Cerón, Gabriela; Servín-González, Luis

    2003-11-01

    The functionality of direct and inverted repeat sequences inside the cis acting locus of transfer (clt) of the Streptomyces plasmid pJV1 was determined by testing the effect of different deletions on plasmid transfer. The results show that the single most important element for pJV1 clt function is a series of evenly spaced 9 bp long direct repeats which match the consensus CCGCACA(C/G)(C/G), since their deletion caused a dramatic reduction in plasmid transfer. The presence of these repeats in the absence of any other clt sequences allowed plasmid transfer to occur at a frequency that was at least two orders of magnitude higher than that obtained in the complete absence of clt. A database search revealed regions with a similar organization, and in the same position, in Streptomyces plasmids pSN22 and pSLS, which have transfer proteins homologous to those of pJV1.

  17. Plasmid Transfer of Plasminogen K1-5 Reduces Subcutaneous Hepatoma Growth by Affecting Inflammatory Factors

    Directory of Open Access Journals (Sweden)

    Lea A. Koch

    2014-01-01

    Full Text Available There is evidence that plasminogen K1-5 (PlgK1-5 directly affects tumour cells and inflammation. Therefore, we analysed if PlgK1-5 has immediate effects on hepatoma cells and inflammatory factors in vitro and in vivo. In vitro, effects of plasmid encoding PlgK1-5 (pK1-5 on Hepa129, Hepa1-6, and HuH7 cell viability, apoptosis, and proliferation as well as VEGF and TNF-alpha expression and STAT3-phosphorylation were investigated. In vivo, tumour growth, proliferation, vessel density, and effects on vascular endothelial growth factor (VEGF and tumour necrosis factor alpha (TNF-alpha expression were examined following treatment with pK1-5. In vivo, pK1-5 halved cell viability; cell death was increased by up to 15% compared to the corresponding controls. Proliferation was not affected. VEGF, TNF-alpha, and STAT3-phosphorylation were affected following treatment with pK1-5. In vivo, ten days after treatment initiation, pK1-5 reduced subcutaneous tumour growth by 32% and mitosis by up to 77% compared to the controls. Vessel density was reduced by 50%. TNF-alpha levels in tumour and liver tissue were increased, whereas VEGF levels in tumours and livers were reduced after pK1-5 treatment. Taken together, plasmid gene transfer of PlgK1-5 inhibits hepatoma (cell growth not only by reducing vessel density but also by inducing apoptosis, inhibiting proliferation, and triggering inflammation.

  18. Development of plasmid vector and electroporation condition for gene transfer in sporogenic lactic acid bacterium, Bacillus coagulans.

    Science.gov (United States)

    Rhee, Mun Su; Kim, Jin-Woo; Qian, Yilei; Ingram, L O; Shanmugam, K T

    2007-07-01

    Bacillus coagulans is a sporogenic lactic acid bacterium that ferments glucose and xylose, major components of plant biomass, a potential feedstock for cellulosic ethanol. The temperature and pH for optimum rate of growth of B. coagulans (50 to 55 degrees C, pH 5.0) are very similar to that of commercially developed fungal cellulases (50 degrees C; pH 4.8). Due to this match, simultaneous saccharification and fermentation (SSF) of cellulose to products by B. coagulans is expected to require less cellulase than needed if the SSF is conducted at a sub-optimal temperature, such as 30 degrees C, the optimum for yeast, the main biocatalyst used by the ethanol industry. To fully exploit B. coagulans as a platform organism, we have developed an electroporation method to transfer plasmid DNA into this genetically recalcitrant bacterium. We also constructed a B. coagulans/E. coli shuttle vector, plasmid pMSR10 that contains the rep region from a native plasmid (pMSR0) present in B. coagulans strain P4-102B. The native plasmid, pMSR0 (6823bp), has 9 ORFs, and replicates by rolling-circle mode of replication. Plasmid pNW33N, developed for Geobacillus stearothermophilus, was also transformed into this host and stably maintained while several other Bacillus/Escherichia coli shuttle vector plasmids were not transformed into B. coagulans. The transformation efficiency of B. coagulans strain P4-102B using the plasmids pNW33N or pMSR10 was about 1.5x10(16) per mole of DNA. The availability of shuttle vectors and an electroporation method is expected to aid in genetic and metabolic engineering of B. coagulans.

  19. Ecological and genetic determinants of plasmid distribution in Escherichia coli.

    Science.gov (United States)

    Medaney, Frances; Ellis, Richard J; Raymond, Ben

    2016-11-01

    Bacterial plasmids are important carriers of virulence and antibiotic resistance genes. Nevertheless, little is known of the determinants of plasmid distribution in bacterial populations. Here the factors affecting the diversity and distribution of the large plasmids of Escherichia coli were explored in cattle grazing on semi-natural grassland, a set of populations with low frequencies of antibiotic resistance genes. Critically, the population genetic structure of bacterial hosts was chararacterized. This revealed structured E. coli populations with high diversity between sites and individuals but low diversity within cattle hosts. Plasmid profiles, however, varied considerably within the same E. coli genotype. Both ecological and genetic factors affected plasmid distribution: plasmid profiles were affected by site, E. coli diversity, E. coli genotype and the presence of other large plasmids. Notably 3/26 E. coli serotypes accounted for half the observed plasmid-free isolates indicating that within species variation can substantially affect carriage of the major conjugative plasmids. The observed population structure suggest that most of the opportunities for within species plasmid transfer occur between different individuals of the same genotype and support recent experimental work indicating that plasmid-host coevolution, and epistatic interactions on fitness costs are likely to be important in determining occupancy. © 2016 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.

  20. Transposon characterization of vancomycin-resistant Enterococcus faecium (VREF) and dissemination of resistance associated with transferable plasmids

    DEFF Research Database (Denmark)

    Migura, Lourdes Garcia; Liebana, Ernesto; Jensen, Lars Bogø

    2007-01-01

    Objectives: VanA glycopeptide resistance has persisted on broiler farms in the UK despite the absence of the antimicrobial selective pressure, avoparcin. This study aimed to investigate the contribution of horizontal gene transfer of Tn 1546 versus clonal spread in the dissemination of the resist......Objectives: VanA glycopeptide resistance has persisted on broiler farms in the UK despite the absence of the antimicrobial selective pressure, avoparcin. This study aimed to investigate the contribution of horizontal gene transfer of Tn 1546 versus clonal spread in the dissemination...... plasmid replicons, associated with antimicrobial resistance on several unrelated farms. Conclusions: Horizontal transfer of vancomycin resistance may play a more important role in the persistence of antimicrobial resistance than clonal spread. The presence of different plasmid replicons, associated...... with antimicrobial resistance on several unrelated farms, illustrates the ability of these enterococci to acquire and disseminate mobile genetic elements within integrated livestock systems....

  1. Complete nucleotide sequence and analysis of two conjugative broad host range plasmids from a marine microbial biofilm.

    Directory of Open Access Journals (Sweden)

    Peter Norberg

    Full Text Available The complete nucleotide sequence of plasmids pMCBF1 and pMCBF6 was determined and analyzed. pMCBF1 and pMCBF6 form a novel clade within the IncP-1 plasmid family designated IncP-1 ς. The plasmids were exogenously isolated earlier from a marine biofilm. pMCBF1 (62 689 base pairs; bp and pMCBF6 (66 729 bp have identical backbones, but differ in their mercury resistance transposons. pMCBF1 carries Tn5053 and pMCBF6 carries Tn5058. Both are flanked by 5 bp direct repeats, typical of replicative transposition. Both insertions are in the vicinity of a resolvase gene in the backbone, supporting the idea that both transposons are "res-site hunters" that preferably insert close to and use external resolvase functions. The similarity of the backbones indicates recent insertion of the two transposons and the ongoing dynamics of plasmid evolution in marine biofilms. Both plasmids also carry the insertion sequence ISPst1, albeit without flanking repeats. ISPs1is located in an unusual site within the control region of the plasmid. In contrast to most known IncP-1 plasmids the pMCBF1/pMCBF6 backbone has no insert between the replication initiation gene (trfA and the vegetative replication origin (oriV. One pMCBF1/pMCBF6 block of about 2.5 kilo bases (kb has no similarity with known sequences in the databases. Furthermore, insertion of three genes with similarity to the multidrug efflux pump operon mexEF and a gene from the NodT family of the tripartite multi-drug resistance-nodulation-division (RND system in Pseudomonas aeruginosa was found. They do not seem to confer antibiotic resistance to the hosts of pMCBF1/pMCBF6, but the presence of RND on promiscuous plasmids may have serious implications for the spread of antibiotic multi-resistance.

  2. DNA sequence analysis of plasmids from multidrug resistant Salmonella enterica serotype Heidelberg isolates.

    Directory of Open Access Journals (Sweden)

    Jing Han

    Full Text Available Salmonella enterica serovar Heidelberg is among the most detected serovars in swine and poultry, ranks among the top five serotypes associated with human salmonellosis and is disproportionately associated with invasive infections and mortality in humans. Salmonella are known to carry plasmids associated with antimicrobial resistance and virulence. To identify plasmid-associated genes in multidrug resistant S. enterica serovar Heidelberg, antimicrobial resistance plasmids from five isolates were sequenced using the 454 LifeSciences pyrosequencing technology. Four of the isolates contained incompatibility group (Inc A/C multidrug resistance plasmids harboring at least eight antimicrobial resistance genes. Each of these strains also carried a second resistance plasmid including two IncFIB, an IncHI2 and a plasmid lacking an identified Inc group. The fifth isolate contained an IncI1 plasmid, encoding resistance to gentamicin, streptomycin and sulfonamides. Some of the IncA/C plasmids lacked the full concert of transfer genes and yet were able to be conjugally transferred, likely due to the transfer genes carried on the companion plasmids in the strains. Several non-IncA/C resistance plasmids also carried putative virulence genes. When the sequences were compared to previously sequenced plasmids, it was found that while all plasmids demonstrated some similarity to other plasmids, they were unique, often due to differences in mobile genetic elements in the plasmids. Our study suggests that Salmonella Heidelberg isolates harbor plasmids that co-select for antimicrobial resistance and virulence, along with genes that can mediate the transfer of plasmids within and among other bacterial isolates. Prevalence of such plasmids can complicate efforts to control the spread of S. enterica serovar Heidelberg in food animal and human populations.

  3. Complete Sequence of a F33:A-:B- Conjugative Plasmid Carrying the oqxAB, fosA3 and blaCTX-M-55 Elements from a Foodborne Escherichia coli Strain

    Directory of Open Access Journals (Sweden)

    Marcus Ho-yin Wong

    2016-10-01

    Full Text Available This study reports the complete sequence of pE80, a conjugative IncFII plasmid recovered from an E. coli strain isolated from chicken meat. This plasmid harbors multiple resistance determinants including oqxAB, fosA3, blaCTX-M-55 and blaTEM-1, and is a close variant of the recently reported p42-2 element, which was recovered from E. coli of veterinary source. Recovery of pE80 constitutes evidence that evolution or genetic re-arrangement of IncFII type plasmids residing in animal-borne organisms is an active event, which involves acquisition and integration of foreign resistance elements into the plasmid backbone. Dissemination of these plasmids may further compromise the effectiveness of current antimicrobial strategies.

  4. Gene Transfer into the Lung by Nanoparticle Dextran-Spermine/Plasmid DNA Complexes

    Directory of Open Access Journals (Sweden)

    Syahril Abdullah

    2010-01-01

    Full Text Available A novel cationic polymer, dextran-spermine (D-SPM, has been found to mediate gene expression in a wide variety of cell lines and in vivo through systemic delivery. Here, we extended the observations by determining the optimal conditions for gene expression of D-SPM/plasmid DNA (D-SPM/pDNA in cell lines and in the lungs of BALB/c mice via instillation delivery. In vitro studies showed that D-SPM could partially protect pDNA from degradation by nuclease and exhibited optimal gene transfer efficiency at D-SPM to pDNA weight-mixing ratio of 12. In the lungs of mice, the levels of gene expression generated by D-SPM/pDNA are highly dependent on the weight-mixing ratio of D-SPM to pDNA, amount of pDNA in the complex, and the assay time postdelivery. Readministration of the complex at day 1 following the first dosing showed no significant effect on the retention and duration of gene expression. The study also showed that there was a clear trend of increasing size of the complexes as the amount of pDNA was increased, where the sizes of the D-SPM/pDNA complexes were within the nanometer range.

  5. Comparative genomics of the IncA/C multidrug resistance plasmid family.

    Science.gov (United States)

    Fricke, W Florian; Welch, Timothy J; McDermott, Patrick F; Mammel, Mark K; LeClerc, J Eugene; White, David G; Cebula, Thomas A; Ravel, Jacques

    2009-08-01

    Multidrug resistance (MDR) plasmids belonging to the IncA/C plasmid family are widely distributed among Salmonella and other enterobacterial isolates from agricultural sources and have, at least once, also been identified in a drug-resistant Yersinia pestis isolate (IP275) from Madagascar. Here, we present the complete plasmid sequences of the IncA/C reference plasmid pRA1 (143,963 bp), isolated in 1971 from the fish pathogen Aeromonas hydrophila, and of the cryptic IncA/C plasmid pRAx (49,763 bp), isolated from Escherichia coli transconjugant D7-3, which was obtained through pRA1 transfer in 1980. Using comparative sequence analysis of pRA1 and pRAx with recent members of the IncA/C plasmid family, we show that both plasmids provide novel insights into the evolution of the IncA/C MDR plasmid family and the minimal machinery necessary for stable IncA/C plasmid maintenance. Our results indicate that recent members of the IncA/C plasmid family evolved from a common ancestor, similar in composition to pRA1, through stepwise integration of horizontally acquired resistance gene arrays into a conserved plasmid backbone. Phylogenetic comparisons predict type IV secretion-like conjugative transfer operons encoded on the shared plasmid backbones to be closely related to a group of integrating conjugative elements, which use conjugative transfer for horizontal propagation but stably integrate into the host chromosome during vegetative growth. A hipAB toxin-antitoxin gene cluster found on pRA1, which in Escherichia coli is involved in the formation of persister cell subpopulations, suggests persistence as an early broad-spectrum antimicrobial resistance mechanism in the evolution of IncA/C resistance plasmids.

  6. Simplified 3d CFD flow simulation of a turbojet disc cavity with conjugate heat transfer

    CSIR Research Space (South Africa)

    Snedden, Glen C

    2003-09-01

    Full Text Available A comprehensive computational fluid dynamics (CFD) model of an actual disc cavity, complete with rotation and conjugate heat transfer, is presented. The model uses a commercially available code with geometrical accuracy including a labyrinth seal...

  7. Characterization of new plasmids from methylotrophic bacteria.

    Science.gov (United States)

    Brenner, V; Holubová, I; Benada, O; Hubácek, J

    1991-07-01

    Several tens of methanol-utilizing bacterial strains isolated from soil were screened for the presence of plasmids. From the obligate methylotroph Methylomonas sp. strain R103a plasmid pIH36 (36 kb) was isolated and its restriction map was constructed. In pink-pigmented facultative methylotrophs (PPFM), belonging to the genus Methylobacterium four plasmids were detected: plasmids pIB200 (200 kb) and pIB14 (14 kb) in the strain R15d and plasmids pWU14 (14 kb) and pWU7 (7.8 kb) in the strain M17. Because of the small size and the presence of several unique REN sites (HindIII, EcoRI, NcoI), plasmid pWU7 was chosen for the construction of a vector for cloning in methylotrophs. Cointegrates pKWU7A and pKWU7B were formed between pWU7 and the E. coli plasmid pK19 Kmr, which were checked for conjugative transfer from E. coli into the methylotrophic host.

  8. SMV1 virus-induced CRISPR spacer acquisition from the conjugative plasmid pMGB1 in Sulfolobus solfataricus P2

    DEFF Research Database (Denmark)

    Erdmann, Susanne; Shah, Shiraz Ali; Garrett, Roger Antony

    2013-01-01

    Organisms of the crenarchaeal order Sulfolobales carry complex CRISPR (clustered regularly interspaced short palindromic repeats) adaptive immune systems. These systems are modular and show extensive structural and functional diversity, especially in their interference complexes. The primary...... targets are an exceptional range of diverse viruses, many of which propagate stably within cells and follow lytic life cycles without producing cell lysis. These properties are consistent with the difficulty of activating CRISPR spacer uptake in the laboratory, but appear to conflict with the high...... complexity and diversity of the CRISPR immune systems that are found among the Sulfolobales. In the present article, we re-examine the first successful induction of archaeal spacer acquisition in our laboratory that occurred exclusively for the conjugative plasmid pMGB1 in Sulfolobus solfataricus P2...

  9. Prevalence and characterization of plasmids carrying sulfonamide resistance genes among Escherichia coli from pigs, pig carcasses and human

    DEFF Research Database (Denmark)

    Shuyu, Wu; Dalsgaard, A.; Hammerum, A. M.

    2010-01-01

    isolates. Fifty-seven sulfonamide-resistant E. coli were selected based on presence of sul resistance genes and subjected to conjugation and/or transformation experiments. S1 nuclease digestion followed by pulsed-field gel electrophoresis was used to visualize and determine the size of plasmids. Plasmids...... and humans) were 65%, 45% and 12% for sul2, sul1, and sul3, respectively. Transfer of resistance through conjugation was observed in 42/57 isolates. Resistances to streptomycin, ampicillin and trimethoprim were co-transferred in most strains. Class 1 integrons were present in 80% of sul1-carrying plasmids...

  10. Heat loss prediction of a confined premixed jet flame using a conjugate heat transfer approach

    NARCIS (Netherlands)

    Gövert, S.; Mira, D.; Zavala-Ake, M.; Kok, J.B.W.; Vázquez, M.; Houzeaux, G.

    2017-01-01

    The presented work addresses the investigation of the heat loss of a confined turbulent jet flame in a lab-scale combustor using a conjugate-heat transfer approach and large-eddy simulation. The analysis includes the assessment of the principal mechanisms of heat transfer in this combustion chamber:

  11. The Salmonella genomic island 1 is specifically mobilized in trans by the IncA/C multidrug resistance plasmid family.

    Science.gov (United States)

    Douard, Gregory; Praud, Karine; Cloeckaert, Axel; Doublet, Benoît

    2010-12-20

    The Salmonella genomic island 1 (SGI1) is a Salmonella enterica-derived integrative mobilizable element (IME) containing various complex multiple resistance integrons identified in several S. enterica serovars and in Proteus mirabilis. Previous studies have shown that SGI1 transfers horizontally by in trans mobilization in the presence of the IncA/C conjugative helper plasmid pR55. Here, we report the ability of different prevalent multidrug resistance (MDR) plasmids including extended-spectrum β-lactamase (ESBL) gene-carrying plasmids to mobilize the multidrug resistance genomic island SGI1. Through conjugation experiments, none of the 24 conjugative plasmids tested of the IncFI, FII, HI2, I1, L/M, N, P incompatibility groups were able to mobilize SGI1 at a detectable level (transfer frequency IncA/C incompatibility group. Several conjugative IncA/C MDR plasmids as well as the sequenced IncA/C reference plasmid pRA1 of 143,963 bp were shown to mobilize in trans SGI1 from a S. enterica donor to the Escherichia coli recipient strain. Depending on the IncA/C plasmid used, the conjugative transfer of SGI1 occurred at frequencies ranging from 10(-3) to 10(-6) transconjugants per donor. Of particular concern, some large IncA/C MDR plasmids carrying the extended-spectrum cephalosporinase bla(CMY-2) gene were shown to mobilize in trans SGI1. The ability of the IncA/C MDR plasmid family to mobilize SGI1 could contribute to its spread by horizontal transfer among enteric pathogens. Moreover, the increasing prevalence of IncA/C plasmids in MDR S. enterica isolates worldwide has potential implications for the epidemic success of the antibiotic resistance genomic island SGI1 and its close derivatives.

  12. Preclinical evaluation of a Haemophilus influenzae type b conjugate vaccine process intended for technology transfer.

    Science.gov (United States)

    Hamidi, Ahd; Verdijk, Pauline; Kreeftenberg, Hans

    2014-01-01

    Introduction of Haemophilus influenzae type b (Hib) vaccine in low- and middle-income countries has been limited by cost and availability of Hib conjugate vaccines for a long time. It was previously recognized by the Institute for Translational Vaccinology (Intravacc, originating from the former Vaccinology Unit of the National Institute of Public Health [RIVM] and the Netherlands Vaccine Institute [NVI]) that local production of a Hib conjugate vaccine would increase the affordability and sustainability of the vaccine and thereby help to speed up Hib introduction in these countries. A new affordable and a non-infringing production process for a Hib conjugate vaccine was developed, including relevant quality control tests, and the technology was transferred to a number of vaccine manufacturers in India, Indonesia, and China. As part of the Hib technology transfer project managed by Intravacc, a preclinical toxicity study was conducted in the Netherlands to test the safety and immunogenicity of this new Hib conjugate vaccine. The data generated by this study were used by the technology transfer partners to accelerate the clinical development of the new Hib conjugate vaccine. A repeated dose toxicity and local tolerance study in rats was performed to assess the reactogenicity and immunogenicity of a new Hib conjugate vaccine compared to a licensed vaccine. The results showed that the vaccine was well tolerated and immunogenic in rats, no major differences in both safety and immunogenicity in rats were found between the vaccine produced according to the production process developed by Intravacc and the licensed one. Rats may be useful to verify the immunogenicity of Hib conjugate vaccines and for preclinical evaluation. In general, nonclinical evaluation of the new Hib conjugate vaccine, including this proof of concept (safety and immunogenicity study in rats), made it possible for technology transfer partners, having implemented the original process with no changes

  13. Transcriptional regulation of pWW0 transfer genes in Pseudomonas putida KT2440

    DEFF Research Database (Denmark)

    Lambertsen, L.M.; Molin, Søren; Kroer, N.

    2004-01-01

    The conjugative IncP-9 plasmid pWW0 (TOL) carries transfer genes, many of whose functions can be predicted from sequence similarities to the well-studied IncW and IncP-1 plasmids, and that are clustered with the replication and maintenance genes of the plasmid core. In this study we show that the...

  14. Improvement of in vivo transfer of plasmid DNA in muscle : Comparison of electroporation versus ultrasound

    NARCIS (Netherlands)

    Kusumanto, Yoka H.; Mulder, Nanno H.; Dam, Wendy A.; Losen, Mario H.; Meijer, Coby; Hospers, Geke A. P.

    Plasmid-based gene delivery to muscle is a treatment strategy for many diseases with potential advantages above viral-based gene delivery methods, however, with a relative low transfection efficiency. We compared two physical methods-electroporation and ultrasound-that facilitate DNA uptake into

  15. [Role of proton-motive force in the conjugative DNA transport in Staphylococci].

    Science.gov (United States)

    Gavriliuk, V G; Vinnikov, A I

    1997-01-01

    Sensitivity of the conjugative process in staphylococci to the action of uncouplers of oxidative phosphorylation and inhibitors of electron transport systems have been proved, that testifies to the energy-dependent character of conjugative transport of DNA. Proceeding of the conjugation process depends upon the generation of delta microH+ on the membrane of both the donor and recipient cells. contribution of protonmotive forces to providing for the transfer of plasmids during conjugation to staphylococci has been defined.

  16. Solving implicit multi-mesh flow and conjugate heat transfer problems with RELAP-7

    International Nuclear Information System (INIS)

    Zou, L.; Peterson, J.; Zhao, H.; Zhang, H.; Andrs, D.; Martineau, R.

    2013-01-01

    The fully implicit simulation capability of RELAP-7 to solve multi-mesh flow and conjugate heat transfer problems for reactor system safety analysis is presented. Compared to general single-mesh simulations, the reactor system safety analysis-type of code has unique challenges due to its highly simplified, interconnected, one-dimensional, and zero-dimensional flow network describing multiple physics with significantly different time and length scales. To use the Jacobian-free Newton Krylov-type of solver, preconditioning is generally required for the Krylov method. The uniqueness of the reactor safety analysis-type of code in treating the interconnected flow network and conjugate heat transfer also introduces challenges in providing preconditioning matrix. Typical flow and conjugate heat transfer problems involved in reactor safety analysis using RELAP-7, as well as the special treatment on the preconditioning matrix are presented in detail. (authors)

  17. Conjugated heat transfer in laminar flow between parallel-plates channel

    International Nuclear Information System (INIS)

    Guedes, R.O.C.; Cotta, R.M.; Brum, N.C.L.

    1989-01-01

    An analysis is made of conjugated convective-conductive heat transfer in laminar flow of a newtonian fluid between parallel-plates channel, taking into account the longitudinal conduction along the duct walls only, by neglecting the transversal temperature gradients in the solid. This extended Graetz-type problem is then analytically handled through the generalized integral transform technique, providing accurate numerical results for quantities of practical interest sucyh as bulk and wall temperatures, and Nusselt numbers. The effects of a conjugation parameter and Biot number on heat transfer behavior are then investigated. (author)

  18. PRECONDITIONED BI-CONJUGATE GRADIENT METHOD FOR RADIATIVE TRANSFER IN SPHERICAL MEDIA

    International Nuclear Information System (INIS)

    Anusha, L. S.; Nagendra, K. N.; Paletou, F.; Leger, L.

    2009-01-01

    A robust numerical method called the Preconditioned Bi-Conjugate Gradient (Pre-BiCG) method is proposed for the solution of the radiative transfer equation in spherical geometry. A variant of this method called Stabilized Preconditioned Bi-Conjugate Gradient (Pre-BiCG-STAB) is also presented. These are iterative methods based on the construction of a set of bi-orthogonal vectors. The application of the Pre-BiCG method in some benchmark tests shows that the method is quite versatile, and can handle difficult problems that may arise in astrophysical radiative transfer theory.

  19. Crystal growth of new charge-transfer salts based on π-conjugated donor molecules

    Energy Technology Data Exchange (ETDEWEB)

    Morherr, Antonia, E-mail: morherr@stud.uni-frankfurt.de [Physikalisches Institut, Goethe-Universität Frankfurt am Main, 60438 Frankfurt am Main (Germany); Witt, Sebastian [Physikalisches Institut, Goethe-Universität Frankfurt am Main, 60438 Frankfurt am Main (Germany); Chernenkaya, Alisa [Graduate School Materials Science in Mainz, 55128 Mainz (Germany); Institut für Physik, Johannes Gutenberg-Universität, 55099 Mainz (Germany); Bäcker, Jan-Peter [Physikalisches Institut, Goethe-Universität Frankfurt am Main, 60438 Frankfurt am Main (Germany); Schönhense, Gerd [Institut für Physik, Johannes Gutenberg-Universität, 55099 Mainz (Germany); Bolte, Michael [Institut für anorganische Chemie, Goethe-Universität Frankfurt am Main, 60438 Frankfurt am Main (Germany); Krellner, Cornelius [Physikalisches Institut, Goethe-Universität Frankfurt am Main, 60438 Frankfurt am Main (Germany)

    2016-09-01

    New charge transfer crystals of π-conjugated, aromatic molecules (phenanthrene and picene) as donors were obtained by physical vapor transport. The melting behavior, optimization of crystal growth and the crystal structure are reported for charge transfer salts with (fluorinated) tetracyanoquinodimethane (TCNQ-F{sub x}, x=0, 2, 4), which was used as acceptor material. The crystal structures were determined by single-crystal X-ray diffraction. Growth conditions for different vapor pressures in closed ampules were applied and the effect of these starting conditions for crystal size and quality is reported. The process of charge transfer was investigated by geometrical analysis of the crystal structure and by infrared spectroscopy on single crystals. With these three different acceptor strengths and the two sets of donor materials, it is possible to investigate the distribution of the charge transfer systematically. This helps to understand the charge transfer process in this class of materials with π-conjugated donor molecules.

  20. Conjugate heat transfer of laminar film condensation along a horizontal plate

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Euk Soo [Pusan National Univesity, Busan (Korea, Republic of)

    2006-03-15

    This paper proposes appropriate conjugate parameters and dimensionless temperatures to analysis the conjugate problem of heat conduction in solid wall coupled with laminar film condensation flow adjacent to horizontal flat plate. An efficient methods for some fluids are proposed for its solution. The momentum and energy balance equations are reduced to a nonlinear system of ordinary differential equations with four parameters: the Prandtl number, Pr, Modified Jacob number, Ja{sup *}/Pr, defined by an overall temperature difference, a property ratio {radical}{rho}{sub {iota}}{mu}{sub {iota}} {radical}{rho}{sub {upsilon}}{mu}{sub {upsilon}} and the conjugate parameter {zeta}. The obtained similarity solution reveals the effect of the conjugate parameter, and the results are compared with the simplified solution. The variations of the heat transfer rates as well as the interface temperature and frictions along the plate are shown explicitly.

  1. Quorum-dependent transfer of the opine-catabolic plasmid pAoF64/95 is regulated by a novel mechanism involving inhibition of the TraR antiactivator TraM.

    Science.gov (United States)

    Wetzel, Margaret E; Asenstorfer, Robert E; Tate, Max E; Farrand, Stephen K

    2018-04-10

    We previously described a plasmid of Agrobacterium spp., pAoF64/95, in which the quorum-sensing system that controls conjugative transfer is induced by the opine mannopine. We also showed that the quorum-sensing regulators TraR, TraM, and TraI function similarly to their counterparts in other repABC plasmids. However, traR, unlike its counterpart on Ti plasmids, is monocistronic and not located in an operon that is inducible by the conjugative opine. Here, we report that both traR and traM are expressed constitutively and not regulated by growth with mannopine. We report two additional regulatory genes, mrtR and tmsP, that are involved in a novel mechanism of control of TraR activity. Both genes are located in the distantly linked region of pAoF64/95 encoding mannopine utilization. MrtR, in the absence of mannopine, represses the four-gene mocC operon as well as tmsP, which is the distal gene of the eight-gene motA operon. As judged by a bacterial two-hybrid analysis, TmsP, which shows amino acid sequence relatedness with the TraM-binding domain of TraR, interacts with the antiactivator. We propose a model in which mannopine, acting through the repressor MrtR, induces expression of TmsP which then titrates the levels of TraM thereby freeing TraR to activate the tra regulon. © 2018 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

  2. Conjugate heat transfer effects on wall bubble nucleation in subcooled flashing flows

    International Nuclear Information System (INIS)

    Peterson, P.F.; Hijikata, K.

    1990-01-01

    A variety of models have been proposed to explain observations that large liquid superheat is required to initiate nucleation in flashing flows of subcooled liquids in nozzles, cracks and pipes. In such flows an abrupt change in the fluid temperature occurs downstream of the nucleating cavities. This paper examines the subcooling of the nucleating cavities due to conjugate heat transfer to the cold downstream fluid. This examination suggests a mechanism limiting the maximum active cavity size. Simple analysis shows that, of the total superheat required to initiate flashing, a substantial portion results from conjugate wall subcooling, which decreases the cavity vapor pressure. The specific case of flashing critical nozzle flow is examined in detail. Here boundary-layer laminarization due to the strong favorable pressure gradient aids the analysis of conjugate heat transfer

  3. Substrate analog interaction with MCR-1 offers insight into the rising threat of the plasmid-mediated transferable colistin resistance.

    Science.gov (United States)

    Wei, Pengcheng; Song, Guangji; Shi, Mengyang; Zhou, Yafei; Liu, Yang; Lei, Jun; Chen, Peng; Yin, Lei

    2018-02-01

    Colistin is considered a last-resort antibiotic against most gram-negative bacteria. Recent discoveries of a plasmid-mediated, transferable mobilized colistin-resistance gene ( mcr-1) on all continents have heralded the imminent emergence of pan-drug-resistant superbacteria. The inner-membrane protein MCR-1 can catalyze the transfer of phosphoethanolamine (PEA) to lipid A, resulting in colistin resistance. However, little is known about the mechanism, and few drugs exist to address this issue. We present crystal structures revealing the MCR-1 catalytic domain (cMCR-1) as a monozinc metalloprotein with ethanolamine (ETA) and d-glucose, respectively, thus highlighting 2 possible substrate-binding pockets in the MCR-1-catalyzed PEA transfer reaction. Mutation of the residues involved in ETA and d-glucose binding impairs colistin resistance in recombinant Escherichia coli containing full-length MCR-1. Partial analogs of the substrate are used for cocrystallization with cMCR-1, providing valuable information about the family of PEA transferases. One of the analogs, ETA, causes clear inhibition of polymyxin B resistance, highlighting its potential for drug development. These data demonstrate the crucial role of the PEA- and lipid A-binding pockets and provide novel insights into the structure-based mechanisms, important drug-target hot spots, and a drug template for further drug development to combat the urgent, rising threat of MCR-1-mediated antibiotic resistance.-Wei, P., Song, G., Shi, M., Zhou, Y., Liu, Y., Lei, J., Chen, P., Yin, L. Substrate analog interaction with MCR-1 offers insight into the rising threat of the plasmid-mediated transferable colistin resistance.

  4. Conjugate heat transfer for turbulent flow in a thick walled plain pipe

    Directory of Open Access Journals (Sweden)

    Canli Eyub

    2018-01-01

    Full Text Available Laminar and turbulent flow have their own characteristics in respect of heat transfer in pipes. While conjugate heat transfer is a major concern for a thick walled pipe with laminar flow inside it, there are limited studies about a turbulent flow in a thick walled plain pipe considering the conjugate heat transfer. In order to conduct such a work by means of in-house developed code, it was desired to make a preliminary investigation with commercially available CFD codes. ANSYS CFD was selected as the tool since it has a positive reputation in the literature for reliability. Defined heat transfer problem was solved with SIMPLE and Coupled Schemes for pressure velocity coupling and results are presented accordingly.

  5. Dynamic impedance compensation for wireless power transfer using conjugate power

    OpenAIRE

    Suqi Liu; Jianping Tan; Xue Wen

    2018-01-01

    Wireless power transfer (WPT) via coupled magnetic resonances has been in development for over a decade. However, the frequency splitting phenomenon occurs in the over-coupled region. Thus, the output power of the two-coil system achieves the maximum output power at the two splitting angular frequencies, and not at the natural resonant angular frequency. According to the maximum power transfer theorem, the impedance compensation method was adopted in many WPT projects. However, it remains a c...

  6. Plasmid-Mediated Antimicrobial Resistance in Staphylococci and Other Firmicutes.

    Science.gov (United States)

    Schwarz, Stefan; Shen, Jianzhong; Wendlandt, Sarah; Fessler, Andrea T; Wang, Yang; Kadlec, Kristina; Wu, Cong-Ming

    2014-12-01

    In staphylococci and other Firmicutes, resistance to numerous classes of antimicrobial agents, which are commonly used in human and veterinary medicine, is mediated by genes that are associated with mobile genetic elements. The gene products of some of these antimicrobial resistance genes confer resistance to only specific members of a certain class of antimicrobial agents, whereas others confer resistance to the entire class or even to members of different classes of antimicrobial agents. The resistance mechanisms specified by the resistance genes fall into any of three major categories: active efflux, enzymatic inactivation, and modification/replacement/protection of the target sites of the antimicrobial agents. Among the mobile genetic elements that carry such resistance genes, plasmids play an important role as carriers of primarily plasmid-borne resistance genes, but also as vectors for nonconjugative and conjugative transposons that harbor resistance genes. Plasmids can be exchanged by horizontal gene transfer between members of the same species but also between bacteria belonging to different species and genera. Plasmids are highly flexible elements, and various mechanisms exist by which plasmids can recombine, form cointegrates, or become integrated in part or in toto into the chromosomal DNA or into other plasmids. As such, plasmids play a key role in the dissemination of antimicrobial resistance genes within the gene pool to which staphylococci and other Firmicutes have access. This chapter is intended to provide an overview of the current knowledge of plasmid-mediated antimicrobial resistance in staphylococci and other Firmicutes.

  7. Horizontal gene transfer of a ColV plasmid has resulted in a dominant avian clonal type of Salmonella enterica serovar Kentucky.

    Directory of Open Access Journals (Sweden)

    Timothy J Johnson

    Full Text Available Salmonella enterica continues to be a significant cause of foodborne gastrointestinal illness in humans. A wide variety of Salmonella serovars have been isolated from production birds and from retail poultry meat. Recently, though, S. enterica subsp. enterica serovar Kentucky has emerged as one of the prominent Salmonella serovars isolated from broiler chickens. Recent work suggests that its emergence apparently coincides with its acquisition of a ColV virulence plasmid. In the present study, we examined 902 Salmonella isolates belonging to 59 different serovars for the presence of this plasmid. Of the serovars examined, the ColV plasmid was found only among isolates belonging to the serovars Kentucky (72.9%, Typhimurium (15.0% and Heidelberg (1.7%. We demonstrated that a single PFGE clonal type of S. Kentucky harbors this plasmid, and acquisition of this plasmid by S. Kentucky significantly increased its ability to colonize the chicken cecum and cause extraintestinal disease. Comparison of the completed sequences of three ColV plasmids from S. Kentucky isolated from different geographical locales, timepoints and sources revealed a nearly identical genetic structure with few single nucleotide changes or insertions/deletions. Overall, it appears that the ColV plasmid was recently acquired by a single clonal type S. Kentucky and confers to its host enhanced colonization and fitness capabilities. Thus, the potential for horizontal gene transfer of virulence and fitness factors to Salmonella from other enteric bacteria exists in poultry, representing a potential human health hazard.

  8. Mobilisation and remobilisation of a large archetypal pathogenicity island of uropathogenic Escherichia coli in vitro support the role of conjugation for horizontal transfer of genomic islands

    Directory of Open Access Journals (Sweden)

    Hochhut Bianca

    2011-09-01

    Full Text Available Abstract Background A substantial amount of data has been accumulated supporting the important role of genomic islands (GEIs - including pathogenicity islands (PAIs - in bacterial genome plasticity and the evolution of bacterial pathogens. Their instability and the high level sequence similarity of different (partial islands suggest an exchange of PAIs between strains of the same or even different bacterial species by horizontal gene transfer (HGT. Transfer events of archetypal large genomic islands of enterobacteria which often lack genes required for mobilisation or transfer have been rarely investigated so far. Results To study mobilisation of such large genomic regions in prototypic uropathogenic E. coli (UPEC strain 536, PAI II536 was supplemented with the mobRP4 region, an origin of replication (oriVR6K, an origin of transfer (oriTRP4 and a chloramphenicol resistance selection marker. In the presence of helper plasmid RP4, conjugative transfer of the 107-kb PAI II536 construct occured from strain 536 into an E. coli K-12 recipient. In transconjugants, PAI II536 existed either as a cytoplasmic circular intermediate (CI or integrated site-specifically into the recipient's chromosome at the leuX tRNA gene. This locus is the chromosomal integration site of PAI II536 in UPEC strain 536. From the E. coli K-12 recipient, the chromosomal PAI II536 construct as well as the CIs could be successfully remobilised and inserted into leuX in a PAI II536 deletion mutant of E. coli 536. Conclusions Our results corroborate that mobilisation and conjugal transfer may contribute to evolution of bacterial pathogens through horizontal transfer of large chromosomal regions such as PAIs. Stabilisation of these mobile genetic elements in the bacterial chromosome result from selective loss of mobilisation and transfer functions of genomic islands.

  9. Production of Fibronectin Binding Protein A at the surface of Lactococcus lactis increases plasmid transfer in vitro and in vivo.

    Directory of Open Access Journals (Sweden)

    Daniela Pontes

    Full Text Available Lactococci are noninvasive lactic acid bacteria frequently used as protein delivery vectors and, more recently, as DNA delivery vehicles. We previously showed that Lactococcus lactis (LL expressing the Fibronectin-Binding Protein A of Staphylococcus aureus (LL-FnBPA+ showed higher internalization rates in vitro in Caco-2 cells than the native (wt lactococci and were able to deliver a eukaryotic Green Fluorescent Protein (GFP expression plasmid in 1% of human Caco-2 cells. Here, using the bovine beta-lactoglobulin (BLG, one of the major cow's milk allergen, and GFP we characterized the potential of LL-FnBPA+ as an in vivo DNA vaccine delivery vehicle. We first showed that the invasive strain LL-FnBPA+ carrying the plasmid pValac:BLG (LL-FnBPA+ BLG was more invasive than LL-BLG and showed the same invasivity as LL-FnBPA+. Then we demonstrated that the Caco-2 cells, co-incubated with LL-FnBPA+ BLG produced up to 30 times more BLG than the Caco-2 cells co-incubated with the non invasive LL-BLG. Using two different gene reporters, BLG and GFP, and two different methods of detection, EIA and fluorescence microscopy, we showed in vivo that: i in order to be effective, LL-FnBPA+ required a pre-coating with Fetal Calf Serum before oral administration; ii plasmid transfer occurred in enterocytes without regard to the strains used (invasive or not; iii the use of LL-FnBPA+ increased the number of mice producing BLG, but not the level of BLG produced. We thus confirmed the good potential of invasive recombinant lactic acid bacteria as DNA delivery vector in vivo.

  10. Effect of Conjugation Length on Photoinduced Charge-Transfer in π-Conjugated Oligomer-Acceptor Dyads

    KAUST Repository

    Jiang, Junlin

    2017-05-25

    A series of -conjugated oligomer-acceptor dyads were synthesized that feature oligo(phenylene ethynylene) (OPE) conjugated backbones end-capped with a naphthalene diimide (NDI) acceptor. The OPE segments vary in length from 4 to 8 phenylene ethynene units (PEn-NDI, where n = 4, 6 and 8). Fluorescence and transient absorption spectroscopy reveals that intramolecular OPE NDI charge transfer dominates the deactivation of excited states of the PEn-NDI oligomers. Both charge separation (CS) and charge recombination (CR) are strongly exothermic (G0CS ~ -1.1 and G0CR ~ -2.0 eV), and the driving forces do not vary much across the series because the oxidation and reduction potentials and singlet energies of the OPEs do not vary much with their length. Bimolecular photoinduced charge transfer between model OPEs that do not contain the NDI acceptors with methyl viologen was studied, and the results reveal that the absorption of the cation radical state (OPE+•) remains approximately constant ( ~ 575 nm) regardless of oligomer length. This finding suggests that the cation radical (polaron) of the OPE is relatively localized, effectively occupying a confined segment of n 4 repeat units in the longer oligomers. Photoinduced intramolecular electron transfer dynamics in the PEn-NDI series was investigated by UV-visible femtosecond transient absorption spectroscopy with visible and mid-infrared probes. Charge separation occurs on the 1 – 10 ps timescale, with the rates decreasing slightly with increased oligomer length (βCS ~ 0.15 Å-1). The rate for charge-recombination decreases in the sequence PE4-NDI > PE6-NDI ~ PE8-NDI. The discontinuous distance dependence in the rate for charge recombination may be related to the spatial localization of the positive polaron state in the longer oligomers.

  11. Effect of Conjugation Length on Photoinduced Charge-Transfer in π-Conjugated Oligomer-Acceptor Dyads

    KAUST Repository

    Jiang, Junlin; Alsam, Amani Abdu; Wang, Shanshan; Aly, Shawkat Mohammede; Pan, Zhenxing; Mohammed, Omar F.; Schanze, Kirk S.

    2017-01-01

    A series of -conjugated oligomer-acceptor dyads were synthesized that feature oligo(phenylene ethynylene) (OPE) conjugated backbones end-capped with a naphthalene diimide (NDI) acceptor. The OPE segments vary in length from 4 to 8 phenylene ethynene units (PEn-NDI, where n = 4, 6 and 8). Fluorescence and transient absorption spectroscopy reveals that intramolecular OPE NDI charge transfer dominates the deactivation of excited states of the PEn-NDI oligomers. Both charge separation (CS) and charge recombination (CR) are strongly exothermic (G0CS ~ -1.1 and G0CR ~ -2.0 eV), and the driving forces do not vary much across the series because the oxidation and reduction potentials and singlet energies of the OPEs do not vary much with their length. Bimolecular photoinduced charge transfer between model OPEs that do not contain the NDI acceptors with methyl viologen was studied, and the results reveal that the absorption of the cation radical state (OPE+•) remains approximately constant ( ~ 575 nm) regardless of oligomer length. This finding suggests that the cation radical (polaron) of the OPE is relatively localized, effectively occupying a confined segment of n 4 repeat units in the longer oligomers. Photoinduced intramolecular electron transfer dynamics in the PEn-NDI series was investigated by UV-visible femtosecond transient absorption spectroscopy with visible and mid-infrared probes. Charge separation occurs on the 1 – 10 ps timescale, with the rates decreasing slightly with increased oligomer length (βCS ~ 0.15 Å-1). The rate for charge-recombination decreases in the sequence PE4-NDI > PE6-NDI ~ PE8-NDI. The discontinuous distance dependence in the rate for charge recombination may be related to the spatial localization of the positive polaron state in the longer oligomers.

  12. Photoluminescence quenching through resonant energy transfer in blends of conjugated polymer with low-molecular acceptor

    International Nuclear Information System (INIS)

    Zapunidi, S. A.; Paraschuk, D. Yu.

    2008-01-01

    A model is proposed for photoluminescence quenching due to resonant energy transfer in a blend of a conjugated polymer and a low-molecular energy acceptor. An analytical dependence of the normalized photoluminescence intensity on the acceptor concentration is derived for the case of a homogeneous blend. This dependence can be described by two fitting parameters related to the Foerster radii for energy transfer between conjugated segments of the polymer and between the conjugated polymer segment and the energy acceptor. Asymptotic approximations are obtained for the model dependence that make it possible to estimate the contribution from the spatial migration of excitons to the photoluminescence quenching. The proposed model is used to analyze experimental data on the photoluminescence quenching in a blend of the soluble derivative of poly(p-phenylene vinylene) and trinitrofluorenone [13]. The Foerster radius for resonant energy transfer between the characteristic conjugated segment of poly(p-phenylene vinylene) and the energy acceptor is determined to be r F = 2.6 ± 0.3 nm

  13. Photoinduced partial charge transfer between conjugated polymer and fullerene in solutions

    International Nuclear Information System (INIS)

    Lin Hongzhen; Weng Yufeng; Huang Hongmin; He Qingguo; Zheng Min; Bai Fenglian

    2004-01-01

    Photoinduced charge transfer between a conjugated polymer and C 60 and the related processes were investigated in dilute solutions. The substantial fluorescence quenching is correlated with the efficient exciton diffusion within the polymer chains, according to which a sphere-of-action mechanism is proposed. An emissive exciplex was found formed between the conjugated polymer and fullerene in a nonpolar solvent, indicating the occurrence of a photoinduced partial charge transfer process. The low-energy sites in the polymer are believed to play a crucial role in the partial charge transfer. The asymmetry of the exciplex provides a method for evaluating the tendency of photoinduced charge separation between the donor and the acceptor. This method allows screening candidates for photovoltaic applications

  14. Charge transfer in conjugated oligomers encapsulated into carbon nanotubes

    Energy Technology Data Exchange (ETDEWEB)

    Almadori, Y.; Alvarez, L.; Michel, T.; Le Parc, R.; Bantignies, J.L.; Hermet, P.; Sauvajol, J.L. [Laboratoire Charles Coulomb UMR 5521, Universite Montpellier 2, 34095 Montpellier (France); Laboratoire Charles Coulomb UMR 5521, CNRS, 34095 Montpellier (France); Arenal, R. [Laboratoire d' Etude des Microstructures, CNRS-ONERA, 92322 Chatillon (France); Laboratorio de Microscopias Avanzadas, Instituto de Nanociencia de Aragon, U. Zaragoza, 50018 Zaragoza (Spain); Babaa, R. [Laboratoire de Chimie des Surfaces et Interfaces, CEA, IRAMIS, SPCSI, 91191 Gif-sur-Yvette Cedex (France); Chemical Engineering Department, University of Technology PETRONAS, UTP, Ipoh-Perak (Malaysia); Jouselme, B.; Palacin, S. [Laboratoire de Chimie des Surfaces et Interfaces, CEA, IRAMIS, SPCSI, 91191 Gif-sur-Yvette Cedex (France)

    2011-11-15

    This study deals with a hybrid system consisting in quaterthiophene derivative encapsulated inside single-walled and multi-walled carbon nanotubes. Investigations of the encapsulation step are performed by transmission electron microscopy. Raman spectroscopy data point out different behaviors depending on the laser excitation energy with respect to the optical absorption of quaterthiophene. At low excitation energy (far from the oligomer resonance window) there is no significant modification of the Raman spectra before and after encapsulation. By contrast, at high excitation energy (close to the oligomer resonance window), Raman spectra exhibit a G-band shift together with an important RBM intensity loss, suggesting a significant charge transfer between the inserted molecule and the host nanotubes. Those results suggest a photo induced process leading to a significant charge transfer. (Copyright copyright 2011 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

  15. Pentanidium-catalyzed enantioselective phase-transfer conjugate addition reactions

    KAUST Repository

    Ma, Ting

    2011-03-09

    A new chiral entity, pentanidium, has been shown to be an excellent chiral phase-transfer catalyst. The enantioselective Michael addition reactions of tert-butyl glycinate-benzophenone Schiff base with various α,β- unsaturated acceptors provide adducts with high enantioselectivities. A successful gram-scale experiment at a low catalyst loading of 0.05 mol % indicates the potential for practical applications of this methodology. Phosphoglycine ester analogues can also be utilized as the Michael donor, affording enantioenriched α-aminophosphonic acid derivatives and phosphonic analogues of (S)-proline. © 2011 American Chemical Society.

  16. A computationally efficient method for full-core conjugate heat transfer modeling of sodium fast reactors

    Energy Technology Data Exchange (ETDEWEB)

    Hu, Rui, E-mail: rhu@anl.gov; Yu, Yiqi

    2016-11-15

    Highlights: • Developed a computationally efficient method for full-core conjugate heat transfer modeling of sodium fast reactors. • Applied fully-coupled JFNK solution scheme to avoid the operator-splitting errors. • The accuracy and efficiency of the method is confirmed with a 7-assembly test problem. • The effects of different spatial discretization schemes are investigated and compared to the RANS-based CFD simulations. - Abstract: For efficient and accurate temperature predictions of sodium fast reactor structures, a 3-D full-core conjugate heat transfer modeling capability is developed for an advanced system analysis tool, SAM. The hexagon lattice core is modeled with 1-D parallel channels representing the subassembly flow, and 2-D duct walls and inter-assembly gaps. The six sides of the hexagon duct wall and near-wall coolant region are modeled separately to account for different temperatures and heat transfer between coolant flow and each side of the duct wall. The Jacobian Free Newton Krylov (JFNK) solution method is applied to solve the fluid and solid field simultaneously in a fully coupled fashion. The 3-D full-core conjugate heat transfer modeling capability in SAM has been demonstrated by a verification test problem with 7 fuel assemblies in a hexagon lattice layout. Additionally, the SAM simulation results are compared with RANS-based CFD simulations. Very good agreements have been achieved between the results of the two approaches.

  17. In Vivo Transmission of an IncA/C Plasmid in Escherichia coli Depends on Tetracycline Concentration, and Acquisition of the Plasmid Results in a Variable Cost of Fitness.

    Science.gov (United States)

    Johnson, Timothy J; Singer, Randall S; Isaacson, Richard E; Danzeisen, Jessica L; Lang, Kevin; Kobluk, Kristi; Rivet, Bernadette; Borewicz, Klaudyna; Frye, Jonathan G; Englen, Mark; Anderson, Janet; Davies, Peter R

    2015-05-15

    IncA/C plasmids are broad-host-range plasmids enabling multidrug resistance that have emerged worldwide among bacterial pathogens of humans and animals. Although antibiotic usage is suspected to be a driving force in the emergence of such strains, few studies have examined the impact of different types of antibiotic administration on the selection of plasmid-containing multidrug resistant isolates. In this study, chlortetracycline treatment at different concentrations in pig feed was examined for its impact on selection and dissemination of an IncA/C plasmid introduced orally via a commensal Escherichia coli host. Continuous low-dose administration of chlortetracycline at 50 g per ton had no observable impact on the proportions of IncA/C plasmid-containing E. coli from pig feces over the course of 35 days. In contrast, high-dose administration of chlortetracycline at 350 g per ton significantly increased IncA/C plasmid-containing E. coli in pig feces (P IncA/C plasmid to other indigenous E. coli hosts. There was no evidence of conjugal transfer of the IncA/C plasmid to bacterial species other than E. coli. In vitro competition assays demonstrated that bacterial host background substantially impacted the cost of IncA/C plasmid carriage in E. coli and Salmonella. In vitro transfer and selection experiments demonstrated that tetracycline at 32 μg/ml was necessary to enhance IncA/C plasmid conjugative transfer, while subinhibitory concentrations of tetracycline in vitro strongly selected for IncA/C plasmid-containing E. coli. Together, these experiments improve our knowledge on the impact of differing concentrations of tetracycline on the selection of IncA/C-type plasmids. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  18. The qacC Gene Has Recently Spread between Rolling Circle Plasmids of Staphylococcus, Indicative of a Novel Gene Transfer Mechanism

    DEFF Research Database (Denmark)

    Wassenaar, Trudy M; Ussery, David W; Ingmer, Hanne

    2016-01-01

    and transferred to acceptor RC-plasmids without assistance of other genes, by means of its location in between the Double Strand replication Origin (DSO) and the Single-Strand replication Origin (SSO). The proposed mobilization model of this DSO-qacC-SSO element represents a novel mechanism of gene mobilization...

  19. Outbreak of NDM-1-Producing Klebsiella pneumoniae in a Dutch Hospital, with Interspecies Transfer of the Resistance Plasmid and Unexpected Occurrence in Unrelated Health Care Centers

    NARCIS (Netherlands)

    Bosch, Thijs; Lutgens, Suzanne P M; Hermans, Mirjam H A; Wever, Peter C; Schneeberger, Peter M.; Renders, Nicole H M; Leenders, Alexander C. A. P.; Kluytmans, Jan A. J. W.; Schoffelen, Annelot F; Notermans, Daan; Witteveen, Sandra; Bathoorn, Erik; Schouls, Leo M.

    In the Netherlands, the number of cases of infection with New Delhi metallo-beta-lactamase (NDM)-positive Enterobacteriaceae is low. Here, we report an outbreak of NDM-1-producing Klebsiella pneumoniae infection in a Dutch hospital with interspecies transfer of the resistance plasmid and unexpected

  20. Outbreak of NDM-1-Producing Klebsiella pneumoniae in a Dutch Hospital, with Interspecies Transfer of the Resistance Plasmid and Unexpected Occurrence in Unrelated Health Care Centers.

    NARCIS (Netherlands)

    Bosch, Thijs; Lutgens, Suzanne P M; Hermans, Mirjam H A; Wever, Peter C; Schneeberger, Peter M; Renders, Nicole H M; Leenders, Alexander C A P; Kluytmans, Jan A J W; Schoffelen, Annelot; Notermans, Daan; Witteveen, Sandra; Bathoorn, Erik; Schouls, Leo M

    In the Netherlands, the number of cases of infection with New Delhi metallo-beta-lactamase (NDM)-positive Enterobacteriaceae is low. Here, we report an outbreak of NDM-1-producing Klebsiella pneumoniae infection in a Dutch hospital with interspecies transfer of the resistance plasmid and unexpected

  1. Requirement for Vibrio cholerae Integration Host Factor in Conjugative DNA Transfer

    OpenAIRE

    McLeod, Sarah M.; Burrus, Vincent; Waldor, Matthew K.

    2006-01-01

    The requirement for host factors in the transmission of integrative and conjugative elements (ICEs) has not been extensively explored. Here we tested whether integration host factor (IHF) or Fis, two host-encoded nucleoid proteins, are required for transfer of SXT, a Vibrio cholerae-derived ICE that can be transmitted to many gram-negative species. Fis did not influence the transfer of SXT to or from V. cholerae. In contrast, IHF proved to be required for V. cholerae to act as an SXT donor. I...

  2. The qacC gene has recently spread between rolling circle plasmids of Staphylococcus, indicative of a novel gene transfer mechanism

    Directory of Open Access Journals (Sweden)

    Trudy M. Wassenaar

    2016-09-01

    Full Text Available Resistance of Staphylococcus species to quaternary ammonium compounds, frequently used as disinfectants and biocides, can be attributed to qac genes. These qac gene products belong to the Small Multidrug Resistant (SMR protein family, and are often encoded by rolling-circle (RC replicating plasmids. Four classes of SMR-type qac gene families have been described in Staphylococcus species: qacC, qacG, qacJ and qacH. Within their class, these genes are highly conserved, but qacC genes are extremely conserved, although they are found in variable plasmid backgrounds. The lower degree of sequence identity of these plasmids compared to the strict nucleotide conservation of their qacC means that this gene has recently spread. In the absence of insertion sequences or other genetic elements explaining the mobility, we sought for an explanation of mobilization by sequence comparison. Publically available sequences of qac genes, their flanking genes and the replication gene that is invariably present in RC-plasmids were compared to reconstruct the evolutionary history of these plasmids and to explain the recent spread of qacC. Here we propose a new model that explains how qacC is mobilized and transferred to acceptor RC-plasmids without assistance of other genes, by means of its location in between the Double Strand replication Origin (DSO and the Single-Strand replication Origin (SSO. The proposed mobilization model of this DSO-qacC-SSO element represents a novel mechanism of gene mobilization in RC-plasmids, which has also been employed by other genes, such as lnuA (conferring lincomycin resistance. The proposed gene mobility has aided to the wide spread of clinically relevant resistance genes in Staphylococcus populations.

  3. First detection of AmpC β-lactamase bla(CMY-2) on a conjugative IncA/C plasmid in a Vibrio parahaemolyticus isolate of food origin.

    Science.gov (United States)

    Li, Ruichao; Lin, Dachuan; Chen, Kaichao; Wong, Marcus Ho Yin; Chen, Sheng

    2015-07-01

    Vibrio parahaemolyticus is an important causative agent of gastroenteritis, with the consumption of contaminated seafood being the major transmission route. Resistance to penicillin is common among V. parahaemolyticus strains, whereas cephalosporin resistance remains rare. In an attempt to assess the current prevalence and characteristics of antibiotic resistance of this pathogen in common food samples, a total of 54 (17% of the total samples) V. parahaemolyticus strains were isolated from 318 meat and seafood samples purchased from supermarkets and wet markets in Shenzhen, China, in 2013. These isolates exhibited high-level resistance to ampicillin, yet they were mostly susceptible to other antimicrobials, except for two that were resistant to extended-spectrum cephalosporins. The β-lactamase gene blaPER-1 was detectable in one strain, V. parahaemolyticus V43, which was resistant to both third- and fourth-generation cephalosporins. Compared to other blaPER-1-positive V. parahaemolyticus strains reported in our previous studies, strain V43 was found to harbor an ∼200-kb conjugative plasmid carrying genes that were different from the antimicrobial resistance genes reported from the previous studies. The β-lactamase gene blaCMY-2 was detectable for the first time in another V. parahaemolyticus isolate, V4, which was resistant to third-generation cephalosporins. This blaCMY-2 gene was shown to be located in an ∼150-kb IncA/C-type conjugative plasmid with a genetic structure consisting of traB-traV-traA-ISEcp1-blaCMY-2-blc-sugE-encR-orf1-orf2-orf3-orf4-dsbC-traC, which is identical to that of other IncA/C conjugative plasmids in Enterobacteriaceae, albeit with a different size. These findings indicate that the transmission of extended-spectrum-β-lactamase (ESBL) and AmpC β-lactamase genes via conjugative plasmids can mediate the development of extended-spectrum cephalosporin resistance in V. parahaemolyticus, thereby posing a potential threat to public health

  4. The broad-host-range plasmid pSFA231 isolated from petroleum-contaminated sediment represents a new member of the PromA plasmid family.

    Science.gov (United States)

    Li, Xiaobin; Top, Eva M; Wang, Yafei; Brown, Celeste J; Yao, Fei; Yang, Shan; Jiang, Yong; Li, Hui

    2014-01-01

    A self-transmissible broad-host-range (BHR) plasmid pSFA231 was isolated from petroleum-contaminated sediment in Shen-fu wastewater irrigation zone, China, using the triparental mating exogenous plasmid capture method. Based on its complete sequence the plasmid has a size of 41.5 kb and codes for 50 putative open reading frames (orfs), 29 of which represent genes involved in replication, partitioning and transfer functions of the plasmid. Phylogenetic analysis grouped pSFA231 into the newly defined PromA plasmid family, which currently includes five members. Further comparative genomic analysis shows that pSFA231 shares the common backbone regions with the other PromA plasmids, i.e., genes involved in replication, maintenance and control, and conjugative transfer. Nevertheless, phylogenetic divergence was found in specific gene products. We propose to divide the PromA group into two subgroups, PromA-α (pMRAD02, pSB102) and PromA-β (pMOL98, pIPO2T, pSFA231, pTer331), based on the splits network analysis of the RepA protein. Interestingly, a cluster of hypothetical orfs located between parA and traA of pSFA231 shows high similarity with the corresponding regions on pMOL98, pIPO2T, and pTer331, suggesting these hypothetical orfs may represent "essential" plasmid backbone genes for the PromA-β subgroup. Alternatively, they may also be accessory genes that were first acquired and then stayed as the plasmid diverged. Our study increases the available collection of complete genome sequences of BHR plasmids, and since pSFA231 is the only characterized PromA plasmid from China, our findings also enhance our understanding of the genetic diversity of this plasmid group in different parts of the world.

  5. The broad-host-range plasmid pSFA231 isolated from petroleum-contaminated sediment represents a new member of the PromA plasmid family

    Directory of Open Access Journals (Sweden)

    Xiaobin eLi

    2015-01-01

    Full Text Available A self-transmissible broad-host-range (BHR plasmid pSFA231 was isolated from petroleum-contaminated sediment in Shen-fu wastewater irrigation zone, China, using the triparental mating exogenous plasmid capture method. Based on its complete sequence the plasmid has a size of 41.5 kb and codes for 50 putative open reading frames (orfs, 28 of which represent genes involved in replication, partitioning and transfer functions of the plasmid. Phylogenetic analysis grouped pSFA231 into the newly defined PromA plasmid family, which currently includes five members. Further comparative genomic analysis shows that pSFA231 shares the common backbone regions with the other PromA plasmids, i.e., genes involved in replication, maintenance and control, and conjugative transfer. Nevertheless, phylogenetic divergence was found in specific gene products. We propose to divide the PromA group into two subgroups, PromA-α (pMRAD02, pSB102 and PromA-β (pMOL98, pIPO2T, pSFA231, pTer331, based on the splits network analysis of the RepA protein. Interestingly, a cluster of hypothetical orfs located between parA and traA of pSFA231 shows high similarity with the corresponding regions on pMOL98, pIPO2T and pTer331, suggesting these hypothetical orfs may represent ‘‘essential’’ plasmid backbone genes for the PromA-β subgroup. Alternatively, they may also be accessory genes that were first acquired and then stayed as the plasmid diverged. Our study increases the available collection of complete genome sequences of BHR plasmids, and since pSFA231 is the only characterized PromA plasmid from China, our findings also enhance our understanding of the genetic diversity of this plasmid group in different parts of the world.

  6. Transferable antibiotic resistance plasmids from biogas plant digestates often belong to the IncP-1 epsilon subgroup

    Czech Academy of Sciences Publication Activity Database

    Wolters, B.; Kyselková, Martina; Krögerrecklenfort, E.; Kreuzig, R.; Smalla, K.

    2015-01-01

    Roč. 5, January (2015), Article 765 ISSN 1664-302X R&D Projects: GA MŠk(CZ) EE2.3.30.0032 Institutional support: RVO:60077344 Keywords : IncP-1 epsilon plasmid * class 1 integrons * biogas plant digestate * antibiotic resistance * exogenous plasmid isolation Subject RIV: EE - Microbiology, Virology Impact factor: 4.165, year: 2015

  7. Fluorescence resonance energy transfer between conjugated molecules infiltrated in three-dimensional opal photonic crystals

    International Nuclear Information System (INIS)

    Zou, Lu; Sui, Ning; Wang, Ying-Hui; Qian, Cheng; Ma, Yu-Guang; Zhang, Han-Zhuang

    2015-01-01

    Fluorescence resonance energy transfer (FRET) from Coumarin 6 (C-6) to Sulforhodamine B (S-B) infiltrated into opal PMMA (poly-methyl-methacrylate) photonic crystals (PCs) has been studied in detail. The intrinsic mesh micro-porous structure of opal PCs could increase the luminescent efficiency through inhibiting the intermolecular interaction. Meanwhile, its structure of periodically varying refractive indices could also modify the FRET through affecting the luminescence characteristics of energy donor or energy acceptor. The results demonstrate that the FRET efficiency between conjugated dyes was easily modified by opal PCs. - Highlights: • We investigate the fluorescence resonance energy transfer between two kinds of dyes. • These two kinds of dyes are infiltrated in PMMA opal photonic crystals. • The structure of opal PCs could improve the luminescent characteristics. • The structure of opal PCs could improve the energy transfer characteristics

  8. An individual-based approach to explain plasmid invasion in bacterial populations

    DEFF Research Database (Denmark)

    Seoane, Jose Miguel; Yankelevich, Tatiana; Dechesne, Arnaud

    2011-01-01

    We present an individual-based experimental framework to identify and estimate the main parameters governing bacterial conjugation at the individual cell scale. From this analysis, we have established that transient periods of unregulated plasmid transfer within newly formed transconjugant cells...... of the growth cycle and that nongrowing cells, even when exposed to high nutrient concentrations, do not display conjugal activity. These results do not support previous hypotheses relating conjugation decay in the deeper cell layers of bacterial biofilms to nutrient depletion and low physiological activity. We...

  9. Plasmids in Gram negatives: molecular typing of resistance plasmids.

    Science.gov (United States)

    Carattoli, Alessandra

    2011-12-01

    A plasmid is defined as a double stranded, circular DNA molecule capable of autonomous replication. By definition, plasmids do not carry genes essential for the growth of host cells under non-stressed conditions but they have systems which guarantee their autonomous replication also controlling the copy number and ensuring stable inheritance during cell division. Most of the plasmids confer positively selectable phenotypes by the presence of antimicrobial resistance genes. Plasmids evolve as an integral part of the bacterial genome, providing resistance genes that can be easily exchanged among bacteria of different origin and source by conjugation. A multidisciplinary approach is currently applied to study the acquisition and spread of antimicrobial resistance in clinically relevant bacterial pathogens and the established surveillance can be implemented by replicon typing of plasmids. Particular plasmid families are more frequently detected among Enterobacteriaceae and play a major role in the diffusion of specific resistance genes. For instance, IncFII, IncA/C, IncL/M, IncN and IncI1 plasmids carrying extended-spectrum beta-lactamase genes and acquired AmpC genes are currently considered to be "epidemic resistance plasmids", being worldwide detected in Enterobacteriaceae of different origin and sources. The recognition of successful plasmids is an essential first step to design intervention strategies preventing their spread. Copyright © 2011 Elsevier GmbH. All rights reserved.

  10. Conjugated heat transfer and temperature distributions in a gas turbine combustion liner under base-load operation

    International Nuclear Information System (INIS)

    Kim, Kyung Min; Yun, Nam Geon; Jeon, Yun Heung; Lee, Dong Hyun; Cho, Yung Hee

    2010-01-01

    Prediction of temperature distributions on hot components is important in development of a gas turbine combustion liner. The present study investigated conjugated heat transfer to obtain temperature distributions in a combustion liner with six combustion nozzles. 3D numerical simulations using FVM commercial codes, Fluent and CFX were performed to calculate combustion and heat transfer distributions. The temperature distributions in the combustor liner were calculated by conjugation of conduction and convection (heat transfer coefficients) obtained by combustion and cooling flow analysis. The wall temperature was the highest on the attachment points of the combustion gas from combustion nozzles, but the temperature gradient was high at the after shell section with low wall temperature

  11. Conjugate heat transfer analysis for in-vessel retention with external reactor vessel cooling

    International Nuclear Information System (INIS)

    Park, Jong-Woon; Bae, Jae-ho; Song, Hyuk-Jin

    2016-01-01

    Highlights: • A conjugate heat transfer analysis method is applied for in-vessel corium retention. • 3D heat diffusion has a formidable effect in alleviating focusing heat load from metallic layer. • The focusing heat load is decreased by about 2.5 times on the external surface. - Abstract: A conjugate heat transfer analysis method for the thermal integrity of a reactor vessel under external reactor vessel cooling conditions is developed to resolve light metal layer focusing effect issue for in-vessel retention. The method calculates steady-state three-dimensional temperature distribution of a reactor vessel using coupled conjugate heat transfer between in-vessel three-layered stratified corium (metallic pool, oxide pool and heavy metal and polar-angle dependent boiling heat transfer at the outer surface of a reactor vessel). The three-layer corium heat transfer model is utilizing lumped-parameter thermal-resistance circuit method. For the ex-vessel boiling boundary conditions, nucleate, transition and film boiling are considered. The thermal integrity of a reactor vessel is addressed in terms of heat flux at the outer-most nodes of the vessel and remaining thickness profile. The vessel three-dimensional heat conduction is validated against a commercial code. It is found that even though the internal heat flux from the metal layer goes far beyond critical heat flux (CHF) the heat flux from the outermost nodes of the vessel may be maintained below CHF due to massive vessel heat diffusion. The heat diffusion throughout the vessel is more pronounced for relatively low heat generation rate in an oxide pool. Parametric calculations are performed considering thermal conditions such as peak heat flux from a light metal layer, heat generation in an oxide pool and external boiling conditions. The major finding is that the most crucial factor for success of in-vessel retention is not the mass of the molten light metal above the oxide pool but the heat generation rate

  12. Proposed model for the high rate of rearrangement and rapid migration observed in some IncA/C plasmid lineages.

    Science.gov (United States)

    Meinersmann, R J; Lindsey, R L; Bono, J L; Smith, T P; Oakley, B B

    2013-08-01

    IncA/C plasmids are a class of plasmids from the Enterobacteriaceae that are relatively large (49 to >180 kbp), that are readily transferred by conjugation, and that carry multiple antimicrobial resistance genes. Reconstruction of the phylogeny of these plasmids has been difficult because of the high rate of remodeling by recombination-mediated horizontal gene transfer (HGT). We hypothesized that evaluation of nucleotide polymorphisms relative to the rate of HGT would help to develop a clock to show whether anthropic practices have had significant influences on the lineages of the plasmid. A system was developed to rapidly sequence up to 191 known open reading frames from each of 39 recently isolated IncA/C plasmids from a diverse panel of Salmonella enterica and Escherichia coli strains. With these data plus sequences from GenBank, we were able to distinguish six distinct lineages that had extremely low numbers of polymorphisms within each lineage, especially among the largest group designated as group 1. Two regions, each about half the plasmid in size, could be distinguished with a separate lineal pattern. The distribution of group 1 showed that it has migrated extremely rapidly with fewer polymorphisms than can be expected in 2,000 years. Remodeling by frequent HGT was evident, with a pattern that appeared to have the highest rate just upstream of the putative conjugation origin of transfer (oriT). It seems likely that when an IncA/C plasmid is transferred by conjugation there is an opportunity for plasmid remodeling adjacent to the oriT, which was also adjacent to a multiple antimicrobial resistance gene cassette.

  13. Prevalence and characterization of plasmids carrying sulfonamide resistance genes among Escherichia coli from pigs, pig carcasses and human.

    Science.gov (United States)

    Wu, Shuyu; Dalsgaard, Anders; Hammerum, Anette M; Porsbo, Lone J; Jensen, Lars B

    2010-07-30

    Sulfonamide resistance is very common in Escherichia coli. The aim of this study was to characterize plasmids carrying sulfonamide resistance genes (sul1, sul2 and sul3) in E. coli isolated from pigs and humans with a specific objective to assess the genetic diversity of plasmids involved in the mobility of sul genes. A total of 501 E. coli isolates from pig feces, pig carcasses and human stools were tested for their susceptibility to selected antimicrobial. Multiplex PCR was conducted to detect the presence of three sul genes among the sulfonamide-resistant E. coli isolates. Fifty-seven sulfonamide-resistant E. coli were selected based on presence of sul resistance genes and subjected to conjugation and/or transformation experiments. S1 nuclease digestion followed by pulsed-field gel electrophoresis was used to visualize and determine the size of plasmids. Plasmids carrying sul genes were characterized by PCR-based replicon typing to allow a comparison of the types of sul genes, the reservoir and plasmid present. A total of 109/501 isolates exhibited sulfonamide resistance. The relative prevalences of sul genes from the three reservoirs (pigs, pig carcasses and humans) were 65%, 45% and 12% for sul2, sul1, and sul3, respectively. Transfer of resistance through conjugation was observed in 42/57 isolates. Resistances to streptomycin, ampicillin and trimethoprim were co-transferred in most strains. Class 1 integrons were present in 80% of sul1-carrying plasmids and 100% of sul3-carrying plasmids, but only in 5% of sul2-carrying plasmids. The sul plasmids ranged from 33 to 160-kb in size and belonged to nine different incompatibility (Inc) groups: FII, FIB, I1, FIA, B/O, FIC, N, HI1 and X1. IncFII was the dominant type in sul2-carrying plasmids (52%), while IncI1 was the most common type in sul1 and sul3-carrying plasmids (33% and 45%, respectively). Multireplicons were found associated with all three sul genes. Sul genes were distributed widely in E. coli isolated

  14. Prevalence and characterization of plasmids carrying sulfonamide resistance genes among Escherichia coli from pigs, pig carcasses and human

    Directory of Open Access Journals (Sweden)

    Hammerum Anette M

    2010-07-01

    Full Text Available Abstract Background Sulfonamide resistance is very common in Escherichia coli. The aim of this study was to characterize plasmids carrying sulfonamide resistance genes (sul1, sul2 and sul3 in E. coli isolated from pigs and humans with a specific objective to assess the genetic diversity of plasmids involved in the mobility of sul genes. Methods A total of 501 E. coli isolates from pig feces, pig carcasses and human stools were tested for their susceptibility to selected antimicrobial. Multiplex PCR was conducted to detect the presence of three sul genes among the sulfonamide-resistant E. coli isolates. Fifty-seven sulfonamide-resistant E. coli were selected based on presence of sul resistance genes and subjected to conjugation and/or transformation experiments. S1 nuclease digestion followed by pulsed-field gel electrophoresis was used to visualize and determine the size of plasmids. Plasmids carrying sul genes were characterized by PCR-based replicon typing to allow a comparison of the types of sul genes, the reservoir and plasmid present. Results A total of 109/501 isolates exhibited sulfonamide resistance. The relative prevalences of sul genes from the three reservoirs (pigs, pig carcasses and humans were 65%, 45% and 12% for sul2, sul1, and sul3, respectively. Transfer of resistance through conjugation was observed in 42/57 isolates. Resistances to streptomycin, ampicillin and trimethoprim were co-transferred in most strains. Class 1 integrons were present in 80% of sul1-carrying plasmids and 100% of sul3-carrying plasmids, but only in 5% of sul2-carrying plasmids. The sul plasmids ranged from 33 to 160-kb in size and belonged to nine different incompatibility (Inc groups: FII, FIB, I1, FIA, B/O, FIC, N, HI1 and X1. IncFII was the dominant type in sul2-carrying plasmids (52%, while IncI1 was the most common type in sul1 and sul3-carrying plasmids (33% and 45%, respectively. Multireplicons were found associated with all three sul genes

  15. Stochastic modelling of conjugate heat transfer in near-wall turbulence

    International Nuclear Information System (INIS)

    Pozorski, Jacek; Minier, Jean-Pierre

    2006-01-01

    The paper addresses the conjugate heat transfer in turbulent flows with temperature assumed to be a passive scalar. The Lagrangian approach is applied and the heat transfer is modelled with the use of stochastic particles. The intensity of thermal fluctuations in near-wall turbulence is determined from the scalar probability density function (PDF) with externally provided dynamical statistics. A stochastic model for the temperature field in the wall material is proposed and boundary conditions for stochastic particles at the solid-fluid interface are formulated. The heated channel flow with finite-thickness walls is considered as a validation case. Computation results for the mean temperature profiles and the variance of thermal fluctuations are presented and compared with available DNS data

  16. Stochastic modelling of conjugate heat transfer in near-wall turbulence

    Energy Technology Data Exchange (ETDEWEB)

    Pozorski, Jacek [Institute of Fluid-Flow Machinery, Polish Academy of Sciences, Fiszera 14, 80952 Gdansk (Poland)]. E-mail: jp@imp.gda.pl; Minier, Jean-Pierre [Research and Development Division, Electricite de France, 6 quai Watier, 78400 Chatou (France)

    2006-10-15

    The paper addresses the conjugate heat transfer in turbulent flows with temperature assumed to be a passive scalar. The Lagrangian approach is applied and the heat transfer is modelled with the use of stochastic particles. The intensity of thermal fluctuations in near-wall turbulence is determined from the scalar probability density function (PDF) with externally provided dynamical statistics. A stochastic model for the temperature field in the wall material is proposed and boundary conditions for stochastic particles at the solid-fluid interface are formulated. The heated channel flow with finite-thickness walls is considered as a validation case. Computation results for the mean temperature profiles and the variance of thermal fluctuations are presented and compared with available DNS data.

  17. Theory of interfacial charge-transfer complex photophysics in π-conjugated polymer-fullerene blends

    Science.gov (United States)

    Aryanpour, K.; Psiachos, D.; Mazumdar, S.

    2010-03-01

    We present a theory of the electronic structure and photophysics of 1:1 blends of derivatives of polyparaphenylenevinylene and fullerenes [1]. Within the same Coulomb-correlated Hamiltonian applied previously to interacting chains of single-component π-conjugated polymers [2], we find an exciplex state that occurs below the polymer's optical exciton. Weak absorption from the ground state occurs to the exciplex. We explain transient photoinduced absorptions in the blend [3], observed for both above-gap and below-gap photoexcitations, within our theory. Photoinduced absorptions for above-gap photoexcitation are from the optical exciton as well as the exciplex, while for below-gap photoexcitation induced absorptions are from the exciplex alone. In neither case are free polarons generated in the time scale of the experiment. Importantly, the photophysics of films of single-component π-conjugated polymers and blends can both be understood by extending Mulliken's theory of ground state charge-transfer to the case of excited state charge-transfer. [1] K. Aryanpour, D. Psiachos, and S. Mazumdar, arXiv:0908.0366 [2] D. Psiachos and S. Mazumdar, Phys. Rev. B. 79 155106 (2009) [3] T. Drori et al., Phys. Rev. Lett. 101, 037402 (2008)

  18. DNS of turbulent channel flow with conjugate heat transfer at Prandtl number 0.01

    Energy Technology Data Exchange (ETDEWEB)

    Tiselj, Iztok, E-mail: iztok.tiselj@ijs.si [' Jozef Stefan' Institute, Jamova 39, SI-1000 Ljubljana (Slovenia); Cizelj, Leon, E-mail: leon.cizelj@ijs.si [' Jozef Stefan' Institute, Jamova 39, SI-1000 Ljubljana (Slovenia)

    2012-12-15

    Highlights: Black-Right-Pointing-Pointer DNS database for turbulent channel flow at Prandtl number 0.01 and various Re{sub {tau}}. Black-Right-Pointing-Pointer Two ideal boundary condition analyzed: non-fluctuating and fluctuating temperature. Black-Right-Pointing-Pointer DNS database with conjugate heat transfer for liquid sodium-steel contact. Black-Right-Pointing-Pointer Penetration of the turbulent temperature fluctuations into the solid wall analyzed. - Abstract: Direct Numerical Simulation (DNS) of the fully developed velocity and temperature fields in a turbulent channel flow coupled with the unsteady conduction in the heated walls was carried out. Simulations were performed with passive scalar approximation at Prandtl number 0.01, which roughly corresponds to the Prandtl number of liquid sodium. DNSs were performed at friction Reynolds numbers 180, 395 and 590. The obtained statistical quantities like mean temperatures, profiles of the root-mean-square (RMS) temperature fluctuations for various thermal properties of wall and fluid, and various wall thicknesses were obtained from a pseudo-spectral channel-flow code. Even for the highest implemented Reynolds number the temperature profile in the fluid does not exhibit log-law region and the near-wall RMS temperature fluctuations show Reynolds number dependence. Conjugate heat transfer simulations of liquid sodium-steel system point to a relatively intensive penetration of turbulent temperature fluctuations into the heated wall. Database containing the results is available in a digital form.

  19. DNS of turbulent channel flow with conjugate heat transfer at Prandtl number 0.01

    International Nuclear Information System (INIS)

    Tiselj, Iztok; Cizelj, Leon

    2012-01-01

    Highlights: ► DNS database for turbulent channel flow at Prandtl number 0.01 and various Re τ . ► Two ideal boundary condition analyzed: non-fluctuating and fluctuating temperature. ► DNS database with conjugate heat transfer for liquid sodium–steel contact. ► Penetration of the turbulent temperature fluctuations into the solid wall analyzed. - Abstract: Direct Numerical Simulation (DNS) of the fully developed velocity and temperature fields in a turbulent channel flow coupled with the unsteady conduction in the heated walls was carried out. Simulations were performed with passive scalar approximation at Prandtl number 0.01, which roughly corresponds to the Prandtl number of liquid sodium. DNSs were performed at friction Reynolds numbers 180, 395 and 590. The obtained statistical quantities like mean temperatures, profiles of the root-mean-square (RMS) temperature fluctuations for various thermal properties of wall and fluid, and various wall thicknesses were obtained from a pseudo-spectral channel-flow code. Even for the highest implemented Reynolds number the temperature profile in the fluid does not exhibit log-law region and the near-wall RMS temperature fluctuations show Reynolds number dependence. Conjugate heat transfer simulations of liquid sodium–steel system point to a relatively intensive penetration of turbulent temperature fluctuations into the heated wall. Database containing the results is available in a digital form.

  20. Conjugate Heat Transfer of Mixed Convection for Viscoelastic Fluid Past a Stretching Sheet

    Directory of Open Access Journals (Sweden)

    Kai-Long Hsiao

    2007-01-01

    Full Text Available A conjugate heat transfer problem of a second-grade viscoelastic fluid past a stretching sheet has been studied. Governing equations include heat conduction equation of a stretching sheet, continuity equation, momentum equation, and energy equation of a second-grade fluid, analyzed by a combination of a series expansion method, the similarity transformation, and a second-order accurate finite-difference method. These solutions are used to iterate with the heat conduction equation of the stretching sheet to obtain distributions of the local convective heat transfer coefficient and the stretching sheet temperature. Ranges of dimensionless parameters, the Prandtl number Pr, the elastic number E and the conduction-convection coefficient Ncc are from 0.001 to 10, 0.0001 to 0.01, and 0.5 to 2.0, respectively. A parameter G, which is used to represent the dominance of the buoyant effect, is present in governing equations. Results indicated that elastic effect in the flow could increase the local heat transfer coefficient and enhance the heat transfer of a stretching sheet. In addition, same as the results from Newtonian fluid flow and conduction analysis of a stretching sheet, a better heat transfer is obtained with a larger Ncc, G, and E.

  1. Characterization of a Large Antibiotic Resistance Plasmid Found in Enteropathogenic Escherichia coli Strain B171 and Its Relatedness to Plasmids of Diverse E. coli and Shigella Strains.

    Science.gov (United States)

    Hazen, Tracy H; Michalski, Jane; Nagaraj, Sushma; Okeke, Iruka N; Rasko, David A

    2017-09-01

    Enteropathogenic Escherichia coli (EPEC) is a leading cause of severe infantile diarrhea in developing countries. Previous research has focused on the diversity of the EPEC virulence plasmid, whereas less is known regarding the genetic content and distribution of antibiotic resistance plasmids carried by EPEC. A previous study demonstrated that in addition to the virulence plasmid, reference EPEC strain B171 harbors a second, larger plasmid that confers antibiotic resistance. To further understand the genetic diversity and dissemination of antibiotic resistance plasmids among EPEC strains, we describe the complete sequence of an antibiotic resistance plasmid from EPEC strain B171. The resistance plasmid, pB171_90, has a completed sequence length of 90,229 bp, a GC content of 54.55%, and carries protein-encoding genes involved in conjugative transfer, resistance to tetracycline ( tetA ), sulfonamides ( sulI ), and mercury, as well as several virulence-associated genes, including the transcriptional regulator hha and the putative calcium sequestration inhibitor ( csi ). In silico detection of the pB171_90 genes among 4,798 publicly available E. coli genome assemblies indicates that the unique genes of pB171_90 ( csi and traI ) are primarily restricted to genomes identified as EPEC or enterotoxigenic E. coli However, conserved regions of the pB171_90 plasmid containing genes involved in replication, stability, and antibiotic resistance were identified among diverse E. coli pathotypes. Interestingly, pB171_90 also exhibited significant similarity with a sequenced plasmid from Shigella dysenteriae type I. Our findings demonstrate the mosaic nature of EPEC antibiotic resistance plasmids and highlight the need for additional sequence-based characterization of antibiotic resistance plasmids harbored by pathogenic E. coli . Copyright © 2017 American Society for Microbiology.

  2. Numerical study of the conjugate heat transfer in a horizontal pipe heated by Joulean effect

    Directory of Open Access Journals (Sweden)

    Touahri Sofiane

    2012-01-01

    Full Text Available The three dimensional mixed convection heat transfer in a electrically heated horizontal pipe conjugated to a thermal conduction through the entire solid thickness is investigated by taking into account the thermal dependence of the physical properties of the fluid and the outer heat losses. The model equations of continuity, momentum and energy are numerically solved by the finite volume method. The pipe thickness, the Prandtl and the Reynolds numbers are fixed while the Grashof number is varied from 104to107. The results obtained show that the dynamic and thermal fields for mixed convection are qualitatively and quantitatively different from those of forced convection, and the local Nusselt number at the interface solid-fluid is not uniform: it has considerable axial and azimuthally variations. The effect of physical variables of the fluid depending on temperature is significant, which justifies its inclusion. The heat transfer is quantified by the local and average Nusselt numbers. We found that the average Nusselt number of solid-fluid interface of the duct increases with the increase of Grashof number. We have equally found out that the heat transfer is improved thanks to the consideration of the thermo dependence of the physical properties. We have tried modelling the average Nusselt number as a function of Richardson number. With the parameters used, the heat transfer is quantified by the correlation: NuA=12.0753 Ri0.156

  3. Compatibility and entry exclusion of IncA and IncC plasmids revisited: IncA and IncC plasmids are compatible.

    Science.gov (United States)

    Ambrose, Stephanie J; Harmer, Christopher J; Hall, Ruth M

    2018-02-24

    In an early study, IncA and IncC plasmids that were reported to be compatible were grouped as the "A-C complex" based on similarities and on strong entry exclusion. However, recently, the term IncA/C has been used frequently to describe plasmids belonging to both of these two groups. Granted that the supporting data was not included in the original reports and that the consensus iteron sequences have since been shown to be essentially identical, we have addressed the question again. The original IncA plasmid, RA1, and the IncC plasmid pRMH760, were introduced into the same cell by transformation, and were found to be maintained stably for over 100 generations in the absence of selection for either plasmid, i.e. they were compatible. We conclude that use of the term IncA/C for this important plasmid group is indeed incorrect and it causes unnecessary confusion. Granted the importance of IncC plasmids in the spread of antibiotic resistance genes, we recommend that use of the misleading terms IncA/C, IncA/C 1 and IncA/C 2 should cease. In addition, RA1 and pRMH760 were shown to each completely prevent entry of the other via conjugative transfer into the cell they reside in. Copyright © 2018 Elsevier Inc. All rights reserved.

  4. IncA/C Conjugative Plasmids Mobilize a New Family of Multidrug Resistance Islands in Clinical Vibrio cholerae Non-O1/Non-O139 Isolates from Haiti.

    Science.gov (United States)

    Carraro, Nicolas; Rivard, Nicolas; Ceccarelli, Daniela; Colwell, Rita R; Burrus, Vincent

    2016-07-19

    Mobile genetic elements play a pivotal role in the adaptation of bacterial populations, allowing them to rapidly cope with hostile conditions, including the presence of antimicrobial compounds. IncA/C conjugative plasmids (ACPs) are efficient vehicles for dissemination of multidrug resistance genes in a broad range of pathogenic species of Enterobacteriaceae ACPs have sporadically been reported in Vibrio cholerae, the infectious agent of the diarrheal disease cholera. The regulatory network that controls ACP mobility ultimately depends on the transcriptional activation of multiple ACP-borne operons by the master activator AcaCD. Beyond ACP conjugation, AcaCD has also recently been shown to activate the expression of genes located in the Salmonella genomic island 1 (SGI1). Here, we describe MGIVchHai6, a novel and unrelated mobilizable genomic island (MGI) integrated into the 3' end of trmE in chromosome I of V. cholerae HC-36A1, a non-O1/non-O139 multidrug-resistant clinical isolate recovered from Haiti in 2010. MGIVchHai6 contains a mercury resistance transposon and an integron In104-like multidrug resistance element similar to the one of SGI1. We show that MGIVchHai6 excises from the chromosome in an AcaCD-dependent manner and is mobilized by ACPs. Acquisition of MGIVchHai6 confers resistance to β-lactams, sulfamethoxazole, tetracycline, chloramphenicol, trimethoprim, and streptomycin/spectinomycin. In silico analyses revealed that MGIVchHai6-like elements are carried by several environmental and clinical V. cholerae strains recovered from the Indian subcontinent, as well as from North and South America, including all non-O1/non-O139 clinical isolates from Haiti. Vibrio cholerae, the causative agent of cholera, remains a global public health threat. Seventh-pandemic V. cholerae acquired multidrug resistance genes primarily through circulation of SXT/R391 integrative and conjugative elements. IncA/C conjugative plasmids have sporadically been reported to

  5. A conjugate gradient method for solving the non-LTE line radiation transfer problem

    Science.gov (United States)

    Paletou, F.; Anterrieu, E.

    2009-12-01

    This study concerns the fast and accurate solution of the line radiation transfer problem, under non-LTE conditions. We propose and evaluate an alternative iterative scheme to the classical ALI-Jacobi method, and to the more recently proposed Gauss-Seidel and successive over-relaxation (GS/SOR) schemes. Our study is indeed based on applying a preconditioned bi-conjugate gradient method (BiCG-P). Standard tests, in 1D plane parallel geometry and in the frame of the two-level atom model with monochromatic scattering are discussed. Rates of convergence between the previously mentioned iterative schemes are compared, as are their respective timing properties. The smoothing capability of the BiCG-P method is also demonstrated.

  6. Conjugate Heat Transfer Characteristics of Laminar Flows Through a Backward Facing Step Duct

    Directory of Open Access Journals (Sweden)

    Bayram CELİK

    2017-10-01

    Full Text Available Present study investigates the effects of solid to fluid conductivity ratio, Prandtl and Reynolds numbers, and solid wall thickness on conjugate heat transfer for a backward facing step duct with a conductive solid bottom wall. Although Kanna and Das performed a case study in 2006 for the same problem, the benchmark studies conducted later including the present one revealed that their results are arguable. Ramsak states in a study “Professor Kanna has confirmed in personal communication that their results are probably wrong”. The temperature and Nusselt number variations along the solid-fluid interface presented here are in excellent agreement with those obtained by Ramsak. The analyses presented here reveal that even though the decrease in Pr and the increase in solid to fluid conductivity ratio have similar global influence with the thinning wall on the interface temperature, the influence of the former parameters are limited in recirculation zone.

  7. Numerical simulation of conjugate heat transfer in electronic cooling and analysis based on field synergy principle

    International Nuclear Information System (INIS)

    Cheng, Y.P.; Lee, T.S.; Low, H.T.

    2008-01-01

    In this paper, the conjugate heat transfer in electronic cooling is numerically simulated with the newly proposed algorithm CLEARER on collocated grid. Because the solid heat source and substrate are isolated from the boundary, special attention is given to deal with the velocity and temperature in the solid region in the full field computation. The influence of openings on the substrate, heat source height and their distribution along the substrate on the maximum temperature and overall Nusselt number is investigated. The numerical results show that the openings on the substrate can enhance the heat transfer as well as increasing the heat source height, meanwhile, by arranging the heat sources coarsely in the front part and densely in the rear part of the substrate, the thermal performance can also be increased. Then the results are analyzed from the viewpoint of field synergy principle, and it is shown that the heat transfer improvement can all be attributed to the better synergy between the velocity field and temperature field, which may offer some guidance in the design of electronic devices

  8. On uncertainty and local sensitivity analysis for transient conjugate heat transfer problems

    International Nuclear Information System (INIS)

    Rauch, Christian

    2012-01-01

    The need for simulating real-world behavior of automobiles has led to more and more sophisticated models being added of various physical phenomena for being coupled together. This increases the number of parameters to be set and, consequently, the required knowledge of their relative importance for the solution and the theory behind them. Sensitivity and uncertainty analysis provides the knowledge of parameter importance. In this paper a thermal radiation solver is considered that performs conduction calculations and receives heat transfer coefficient and fluid temperate at a thermal node. The equations of local, discrete, and transient sensitivities for the conjugate heat transfer model solved by the finite difference method are being derived for some parameters. In the past, formulations for the finite element method have been published. This paper builds on the steady-state formulation published previously by the author. A numerical analysis on the stability of the solution matrix is being conducted. From those normalized sensitivity coefficients are calculated dimensionless uncertainty factors. On a simplified example the relative importance of the heat transfer modes at various locations is then investigated by those uncertainty factors and their changes over time

  9. Photoinduced charge and energy transfer in dye-doped conjugated polymers

    International Nuclear Information System (INIS)

    Veldman, Dirk; Bastiaansen, Jolanda J.A.M.; Langeveld-Voss, Bea M.W.; Sweelssen, Joergen; Koetse, Marc M.; Meskers, Stefan C.J.; Janssen, Rene A.J.

    2006-01-01

    Conjugated polymer-molecular dye blends of MDMO-PPV (poly[2-methoxy-5-(3',7'-dimethyloctyloxy)-1,4-phenylenevinylene]) and PF1CVTP (poly[9,9-dioctylfluorene-2,7-diyl-alt-2,5-bis(2-thienyl-1-cyanovinyl) -1-(3',7= '-dimethyloctyloxy)-4-methoxybenzene-5'',5''-diyl]) with three dipyrrometheneboron difluoride (bodipy) dyes were studied by (time-resolved) fluorescence and photoinduced absorption spectroscopy to determine quantitatively the relation between the electronic HOMO and LUMO levels and the occurrence of energy or charge transfer after optical excitation. We find that for MDMO-PPV photoinduced charge transfer to the dyes occurs, while photoexcitation of PF1CVTP exclusively results in energy transfer. The differences can be rationalized by assuming that the energy of the charge separated state is 0.33-0.45 eV higher than the energy determined from oxidation and reduction potentials of donor and acceptor, respectively. This provides an important design rule to identify appropriate materials for polymer solar cells that can have a high open-circuit voltage

  10. Conjugate heat transfer LES of thermal mixing in a T-junction

    Energy Technology Data Exchange (ETDEWEB)

    Timperi, Antti, E-mail: antti.timperi@vtt.fi

    2014-07-01

    Highlights: • Turbulent mixing of hot and cold water in a T-junction is studied with LES including conjugate heat transfer. • The numerical simulations are in good agreement with experimental data in the bulk of the flow. • Comparison of different inlet and wall boundary conditions shows significant differences only near walls. • Effects of mesh and flow velocity on the velocity and temperature fluctuations are also considered. - Abstract: Turbulent mixing of hot and cold fluids may lead to high-cycle thermal fatigue in piping of nuclear power plants. In this work, the mixing in a T-junction experiment is studied by large-eddy simulation (LES). Conjugate heat transfer (CHT) between fluid and pipe wall is studied by replacing the plexiglass pipe of the experiment with a steel one. Different inlet and wall boundary conditions are first considered. Steady and turbulent inlets are compared, as well as adiabatic and CHT wall conditions. The turbulent inlets are created by using the vortex method which is validated for fully developed flow. The inlets are shown to have only small effect in bulk of the flow, but non-negligible effect near walls. The adiabatic and CHT cases show practically no difference in the logarithmic layer and upward, whereas near walls the difference becomes significant due to thermal inertia of the pipe wall. In bulk of the flow, the mean and fluctuating quantities show good agreement with the experiment. CHT simulations by using different meshes and flow velocities are then considered. A coarse mesh is found to yield qualitative agreement but significant errors in e.g. Reynolds stresses near walls. Temperature fluctuation intensity at the pipe inner surface is fairly similar for both meshes. Normalized profiles and spectra for different velocities are qualitatively similar, but some differences exist e.g. in distributions of wall temperature fluctuation intensity.

  11. Impact of nuclear lattice relaxation on the excitation energy transfer along a chain of pi-conjugated molecules

    NARCIS (Netherlands)

    Schmid, S.A.; Abbel, R.J.; Schenning, A.P.H.J.; Meijer, E.W.; Herz, L.M.

    2010-01-01

    We have investigated the extent to which delocalization of the ground-state and excited-state wave functions of a p-conjugated molecule affects the excitation energy transfer (EET) between such molecules. Using femtosecond photoluminescence spectroscopy, we experimentally monitored the EET along

  12. A Novel IncA/C1 Group Conjugative Plasmid, Encoding VIM-1 Metallo-Beta-Lactamase, Mediates the Acquisition of Carbapenem Resistance in ST104 Klebsiella pneumoniae Isolates from Neonates in the Intensive Care Unit of V. Monaldi Hospital in Naples.

    Science.gov (United States)

    Esposito, Eliana P; Gaiarsa, Stefano; Del Franco, Mariateresa; Crivaro, Valeria; Bernardo, Mariano; Cuccurullo, Susanna; Pennino, Francesca; Triassi, Maria; Marone, Piero; Sassera, Davide; Zarrilli, Raffaele

    2017-01-01

    The emergence of carbapenemase producing Enterobacteriaceae has raised major public health concern. The aim of this study was to investigate the molecular epidemiology and the mechanism of carbapenem resistance acquisition of multidrug-resistant Klebsiella pneumoniae isolates from 20 neonates in the neonatal intensive care unit (NICU) of the V. Monaldi Hospital in Naples, Italy, from April 2015 to March 2016. Genotype analysis by pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST) identified PFGE type A and subtypes A1 and A2 in 17, 2, and 1 isolates, respectively, and assigned all isolates to sequence type (ST) 104. K. pneumoniae isolates were resistant to all classes of β-lactams including carbapenems, fosfomycin, gentamicin, and trimethoprim-sulfamethoxazole, but susceptible to quinolones, amikacin, and colistin. Conjugation experiments demonstrated that resistance to third-generation cephems and imipenem could be transferred along with an IncA/C plasmid containing the extended spectrum β-lactamase bla SHV -12 and carbapenem-hydrolyzing metallo-β-lactamase bla V IM-1 genes. The plasmid that we called pIncAC_KP4898 was 156,252 bp in size and included a typical IncA/C backbone, which was assigned to ST12 and core genome (cg) ST12.1 using the IncA/C plasmid MLST (PMLST) scheme. pIncAC_KP4898 showed a mosaic structure with bla V IM-1 into a class I integron, bla SHV -12 flanked by IS6 elements, a mercury resistance and a macrolide 2'-phosphotransferase clusters, ant(3″), aph(3″), aacA4, qnrA1, sul1 , and dfrA14 conferring resistance to aminoglycosides, quinolones, sulfonamides, and trimethoprim, respectively, several genes predicted to encode transfer functions and proteins involved in DNA transposition. The acquisition of pIncAC_KP4898 carrying bla V IM-1 and bla SHV -12 contributed to the spread of ST104 K. pneumoniae in the NICU of V. Monaldi Hospital in Naples.

  13. A Novel IncA/C1 Group Conjugative Plasmid, Encoding VIM-1 Metallo-Beta-Lactamase, Mediates the Acquisition of Carbapenem Resistance in ST104 Klebsiella pneumoniae Isolates from Neonates in the Intensive Care Unit of V. Monaldi Hospital in Naples

    Directory of Open Access Journals (Sweden)

    Eliana P. Esposito

    2017-11-01

    Full Text Available The emergence of carbapenemase producing Enterobacteriaceae has raised major public health concern. The aim of this study was to investigate the molecular epidemiology and the mechanism of carbapenem resistance acquisition of multidrug-resistant Klebsiella pneumoniae isolates from 20 neonates in the neonatal intensive care unit (NICU of the V. Monaldi Hospital in Naples, Italy, from April 2015 to March 2016. Genotype analysis by pulsed-field gel electrophoresis (PFGE and multi-locus sequence typing (MLST identified PFGE type A and subtypes A1 and A2 in 17, 2, and 1 isolates, respectively, and assigned all isolates to sequence type (ST 104. K. pneumoniae isolates were resistant to all classes of β-lactams including carbapenems, fosfomycin, gentamicin, and trimethoprim–sulfamethoxazole, but susceptible to quinolones, amikacin, and colistin. Conjugation experiments demonstrated that resistance to third-generation cephems and imipenem could be transferred along with an IncA/C plasmid containing the extended spectrum β-lactamase blaSHV -12 and carbapenem-hydrolyzing metallo-β-lactamase blaV IM-1 genes. The plasmid that we called pIncAC_KP4898 was 156,252 bp in size and included a typical IncA/C backbone, which was assigned to ST12 and core genome (cg ST12.1 using the IncA/C plasmid MLST (PMLST scheme. pIncAC_KP4898 showed a mosaic structure with blaV IM-1 into a class I integron, blaSHV -12 flanked by IS6 elements, a mercury resistance and a macrolide 2′-phosphotransferase clusters, ant(3″, aph(3″, aacA4, qnrA1, sul1, and dfrA14 conferring resistance to aminoglycosides, quinolones, sulfonamides, and trimethoprim, respectively, several genes predicted to encode transfer functions and proteins involved in DNA transposition. The acquisition of pIncAC_KP4898 carrying blaV IM-1 and blaSHV -12 contributed to the spread of ST104 K. pneumoniae in the NICU of V. Monaldi Hospital in Naples.

  14. Solution of the conjugated heat transfer problem for the fuel elements assemblies

    International Nuclear Information System (INIS)

    Golba, V.S.; Ivanenko, I.J.; Zinina, G.A.

    1997-01-01

    The paper presents the assemblies conjugated heat conductivity problem calculation and experimental method. The method is based on the temperature superposition modified concept and subchannel method and allows to predict the fuel elements surface temperatures with availability of fuel elements inside structure of any complication caused by technological and working defects and with availability of depositions with low heat conductivity on the fuel elements surfaces. According to the method developed the partial solutions of the heat conductivity equation at the heat removal boundaries (solid-liquid) are found separately for the fuel elements and for the liquid. The heat conductivity equation partial solutions for the fuel elements are predicted by calculations. The coolant heat conductivity equation partial solution ('influence functions') data massif is obtained in present work experimentally in the fuel assembly model consists of 7 tube bundle of fuel elements imitators placed in right grating with relative grating step equal to 1.1 and cooled by eutectic alloy Pb-Bi. It is shown that 'subchannel prediction method' decreases the crosswise heat transfer in comparison with crosswise heat transfer, when the fuel element inside structure is taken into account. Also in the paper it is shown that it is possible to realize the assembly temperature prediction method suggested without carrying out the experiments in the assembly's model in order to get the external problem influence functions'. (author)

  15. Three dimensional conjugated heat transfer analysis in sodium fast reactor wire-wrapped fuel assembly

    International Nuclear Information System (INIS)

    Peniguel, C.; Rupp, I.; Juhel, JP.; Rolfo, S.; Guillaud, M.; Gervais, N.

    2009-01-01

    Fast reactors with liquid metal coolant have recently received a renewed interest owing to a more efficient usage of the primary uranium resources, and they are one of the proposal for the next Generation IV. In order to evaluate nuclear power plant design and safety, 3D analysis of the flow and heat transfer in a wire spacer fuel assembly are ongoing at EDF. The introduction of the wire wrapped spacers, helically wound along the pin axis, enhances the mixing of the coolant between sub-channels and prevents contact between the fuel pins. The mesh generation step constitutes a challenging task if a reasonable amount of cells in conjunction with a suitable spatial discretization is wanted. Several approaches have been investigated and will be presented. Quite complex global flow patterns are found using either k-ε or preferably Reynolds Stress turbulent models. Preliminary conjugated heat transfer calculations using a coupling between the finite element thermal code SYRTHES and the finite volume CFD code Code Saturne are also shown. (author)

  16. pTC Plasmids from Sulfolobus Species in the Geothermal Area of Tengchong, China: Genomic Conservation and Naturally-Occurring Variations as a Result of Transposition by Mobile Genetic Elements.

    Science.gov (United States)

    Xiang, Xiaoyu; Huang, Xiaoxing; Wang, Haina; Huang, Li

    2015-02-12

    Plasmids occur frequently in Archaea. A novel plasmid (denoted pTC1) containing typical conjugation functions has been isolated from Sulfolobus tengchongensis RT8-4, a strain obtained from a hot spring in Tengchong, China, and characterized. The plasmid is a circular double-stranded DNA molecule of 20,417 bp. Among a total of 26 predicted pTC1 ORFs, 23 have homologues in other known Sulfolobus conjugative plasmids (CPs). pTC1 resembles other Sulfolobus CPs in genome architecture, and is most highly conserved in the genomic region encoding conjugation functions. However, attempts to demonstrate experimentally the capacity of the plasmid for conjugational transfer were unsuccessful. A survey revealed that pTC1 and its closely related plasmid variants were widespread in the geothermal area of Tengchong. Variations of the plasmids at the target sites for transposition by an insertion sequence (IS) and a miniature inverted-repeat transposable element (MITE) were readily detected. The IS was efficiently inserted into the pTC1 genome, and the inserted sequence was inactivated and degraded more frequently in an imprecise manner than in a precise manner. These results suggest that the host organism has evolved a strategy to maintain a balance between the insertion and elimination of mobile genetic elements to permit genomic plasticity while inhibiting their fast spreading.

  17. pTC Plasmids from Sulfolobus Species in the Geothermal Area of Tengchong, China: Genomic Conservation and Naturally-Occurring Variations as a Result of Transposition by Mobile Genetic Elements

    Directory of Open Access Journals (Sweden)

    Xiaoyu Xiang

    2015-02-01

    Full Text Available Plasmids occur frequently in Archaea. A novel plasmid (denoted pTC1 containing typical conjugation functions has been isolated from Sulfolobus tengchongensis RT8-4, a strain obtained from a hot spring in Tengchong, China, and characterized. The plasmid is a circular double-stranded DNA molecule of 20,417 bp. Among a total of 26 predicted pTC1 ORFs, 23 have homologues in other known Sulfolobus conjugative plasmids (CPs. pTC1 resembles other Sulfolobus CPs in genome architecture, and is most highly conserved in the genomic region encoding conjugation functions. However, attempts to demonstrate experimentally the capacity of the plasmid for conjugational transfer were unsuccessful. A survey revealed that pTC1 and its closely related plasmid variants were widespread in the geothermal area of Tengchong. Variations of the plasmids at the target sites for transposition by an insertion sequence (IS and a miniature inverted-repeat transposable element (MITE were readily detected. The IS was efficiently inserted into the pTC1 genome, and the inserted sequence was inactivated and degraded more frequently in an imprecise manner than in a precise manner. These results suggest that the host organism has evolved a strategy to maintain a balance between the insertion and elimination of mobile genetic elements to permit genomic plasticity while inhibiting their fast spreading.

  18. Germination and conjugation of Bacillus thuringiensis subsp. israelensis in the intestine of gnotobiotic rats

    DEFF Research Database (Denmark)

    Wilcks, Andrea; Ørum-Smidt, Lasse; Bahl, Martin Iain

    2008-01-01

    of growth and sporulation of vegetative cells. For the first time conjugative plasmid transfer in a mammalian intestinal tract was shown between two B. thuringiensis strains. Significance and Impact of the Study: Strains of B. thuringiensis are used worldwide to combat insect pests, and this study brings...

  19. Survival and conjugal transfer between Bacillus thuringiensis strains in aquatic environment Sobrevivência e conjugação de Bacillus thuringiensis em ambiente aquático

    Directory of Open Access Journals (Sweden)

    Luciana Furlaneto

    2000-10-01

    Full Text Available Field and laboratory studies were conducted to assess the survival of cells and spores and plasmid transfer between Bacillus thuringienis strains in aquatic environment. Results indicated that cells and spores of B. thuringiensis can survive for 10 days in water, without altering their number. The sporulation process began after 12-15 hours of inoculation of water. B. thuringiensis was able to transfer conjugative plasmids in the aquatic environment.O presente trabalho é um estudo sobre a sobrevivência e a conjugação de linhagens de Bacillus thuringiensis em água. Os experimentos conduzidos no laboratório mostram que as células e os esporos de B. thuringiensis podem persistir pelo menos 10 dias na água. A esporulação inicia-se 12-15 horas após a inoculação. O processo de conjugação foi demonstrado em diferentes ambientes aquáticos, tanto em condições de laboratório quanto no meio ambiente.

  20. Spontaneous mutability and light-induced mutagenesis in Salmonella typhimurium: effects of an R-plasmid

    International Nuclear Information System (INIS)

    Valdivia, L.

    1979-01-01

    The UV-protecting plasmid R46 was transferred by conjugation to a genetically marked mouse-virulent Salmonella typhimurium strain, not derived from LT2; in this host the plasmid conferred UV protection and enhanced UV mutagenesis just as it does in LT2 lines. Tra - derivatives of R46 encountered during transduction retained UV-protecting and mutagenesis-enhancing ability. Stored strains carrying the R46-derived plasmids with strong mutator effect but not UV-protecting had lost most of their original streptomycin resistance but were slightly resistant to spectinomycin; attempts to transfer such plasmids failed. R46 enhanced the weak mutagenic effect of visible light on several his and trp mutants of strain LT2, including some whose frequency of spontaneous reversion was not increased by the plasmid. A mutagenic effect was produced by visible-light irradiation of hisG46(R46), either growing cells or nonmultiplying (histidine-deprived cells at 10 0 C). Presence of catalase or cyanide during irradiation did not prevent mutagenesis, which excludes some hypothetical mechanisms. Visible-light irradiation of hisG46 or hisG46(R46) under strict anaerobiosis had little or no mutagenic effect (controls showed that revertants if produced would have been detected). This is as expected if visible-light irradiation in air causes photodynamic damage to DNA and mutations are produced during error-prone, plasmid-enhanced repair

  1. Photoinduced electron transfer for an eosin-tyrosine conjugate. Activity of the tyrosinate anion in long-range electron transfer in a protein-like polymer matrix

    Energy Technology Data Exchange (ETDEWEB)

    Jones, G. II; Feng, Z.; Oh, C. [Boston Univ., MA (United States)

    1995-03-23

    The Xanthene dye eosin Y has been modified via a thiohydantoin link to the amine terminus of the amino acid L-tyrosine. Photochemical electron transfer involving the singlet state of the dye and the attached phenol-containing residue led to a reduction in eosin fluorescence quantum yield and lifetime for aqueous solutions at elevated pH. The conjugate provided an electron transfer product of relatively long lifetime (1 {mu}s range) observed by flash photolysis of solutions at pH 12.0, conditions under which the tyrosine moiety is ionized. The effects of binding of the conjugate in the polymer poly(vinylpyrrolidone) (PVP) on the rates of electron transfer of species of different charge type were examined. 30 refs., 5 figs., 1 tab.

  2. Effect of pheromone induction on transfer of the Enterococcus faecalis plasmid pCF10 in intestinal mucus ex vivo

    DEFF Research Database (Denmark)

    Licht, Tine Rask; Hammerum, Anette Marie; Jensen, Lars Bogø

    2001-01-01

    The effect of synthetic sex pheromone on pheromone-inducible conjugation between the isogenic Enterococcus faecalis strains OG1RF and OG1SS was investigated in (i) Todd-Hewitt broth medium and (ii) intestinal mucus isolated from germ-free rats. In broth, the presence of synthetic pheromone cCF10...

  3. Broad-Host-Range IncP-1 plasmids and their resistance potential

    Directory of Open Access Journals (Sweden)

    Magdalena ePopowska

    2013-03-01

    Full Text Available The plasmids of the incompatibility group IncP-1, also called IncP, as extrachromosomal genetic elements can transfer and replicate virtually in all Gram-negative bacteria. They are composed of backbone genes that encode a variety of essential functions and accessory genes that have implications for human health and environmental bioremediation. Broad-host-range IncP plasmids are known to spread genes between distinct phylogenetic groups of bacteria. These genes often code for resistances to a broad spectrum of antibiotics, heavy metals and quaternary ammonium compounds used as disinfectants. The backbone of these plasmids carries modules that enable them to effectively replicate, move to a new host via conjugative transfer and to be stably maintained in bacterial cells. The adaptive, resistance and virulence genes are mainly located on mobile genetic elements integrated between the functional plasmid backbone modules. Environmental studies have demonstrated the wide distribution of IncP-like replicons in manure, soils and wastewater treatment plants. They also are present in strains of pathogenic or opportunistic bacteria, which can be a cause for concern, because they may encode multiresistance. Their broad distribution suggests that IncP plasmids play a crucial role in bacterial adaptation by utilizing horizontal gene transfer. This review summarizes the variety of genetic information and physiological functions carried by IncP plasmids, which can contribute to the spread of antibiotic and heavy metal resistance while also mediating the process of bioremediation of pollutants. Due to the location of the resistance genes on plasmids with a broad host range and the presence of transposons carrying these genes it seems that the spread of these genes would be possible and quite hazardous in infection control. Future studies are required to determine the level of risk of the spread of resistance genes located on these plasmids.

  4. The Plasmid Mobilome of the Model Plant-Symbiont Sinorhizobium meliloti: Coming up with New Questions and Answers.

    Science.gov (United States)

    Lagares, Antonio; Sanjuán, Juan; Pistorio, Mariano

    2014-10-01

    Rhizobia are Gram-negative Alpha- and Betaproteobacteria living in the underground which have the ability to associate with legumes for the establishment of nitrogen-fixing symbioses. Sinorhizobium meliloti in particular-the symbiont of Medicago, Melilotus, and Trigonella spp.-has for the past decades served as a model organism for investigating, at the molecular level, the biology, biochemistry, and genetics of a free-living and symbiotic soil bacterium of agricultural relevance. To date, the genomes of seven different S. meliloti strains have been fully sequenced and annotated, and several other draft genomic sequences are also available. The vast amount of plasmid DNA that S. meliloti frequently bears (up to 45% of its total genome), the conjugative ability of some of those plasmids, and the extent of the plasmid diversity has provided researchers with an extraordinary system to investigate functional and structural plasmid molecular biology within the evolutionary context surrounding a plant-associated model bacterium. Current evidence indicates that the plasmid mobilome in S. meliloti is composed of replicons varying greatly in size and having diverse conjugative systems and properties along with different evolutionary stabilities and biological roles. While plasmids carrying symbiotic functions (pSyms) are known to have high structural stability (approaching that of chromosomes), the remaining plasmid mobilome (referred to as the non-pSym, functionally cryptic, or accessory compartment) has been shown to possess remarkable diversity and to be highly active in conjugation. In light of the modern genomic and current biochemical data on the plasmids of S. meliloti, the current article revises their main structural components, their transfer and regulatory mechanisms, and their potential as vehicles in shaping the evolution of the rhizobial genome.

  5. The SXT conjugative element and linear prophage N15 encode toxin-antitoxin-stabilizing systems homologous to the tad-ata module of the Paracoccus aminophilus plasmid pAMI2.

    Science.gov (United States)

    Dziewit, Lukasz; Jazurek, Magdalena; Drewniak, Lukasz; Baj, Jadwiga; Bartosik, Dariusz

    2007-03-01

    A group of proteic toxin-antitoxin (TA) cassettes whose representatives are widely distributed among bacterial genomes has been identified. These cassettes occur in chromosomes, plasmids, bacteriophages, and noncomposite transposons, as well as in the SXT conjugative element of Vibrio cholerae. The following four homologous loci were subjected to detailed comparative studies: (i) tad-ata from plasmid pAMI2 of Paracoccus aminophilus (the prototype of this group), (ii) gp49-gp48 from the linear bacteriophage N15 of Escherichia coli, (iii) s045-s044 from SXT, and (iv) Z3230-Z3231 from the genomic island of enterohemorrhagic Escherichia coli O157:H7 strain EDL933. Functional analysis revealed that all but one of these loci (Z3230-Z3231) are able to stabilize heterologous replicons, although the host ranges varied. The TA cassettes analyzed have the following common features: (i) the toxins are encoded by the first gene of each operon; (ii) the antitoxins contain a predicted helix-turn-helix motif of the XRE family; and (iii) the cassettes have two promoters that are different strengths, one which is located upstream of the toxin gene and one which is located upstream of the antitoxin gene. All four toxins tested are functional in E. coli; overexpression of the toxins (in the absence of antitoxin) results in a bacteriostatic effect manifested by elongation of bacterial cells and growth arrest. The toxins have various effects on cell viability, which suggests that they may recognize different intracellular targets. Preliminary data suggest that different cellular proteases are involved in degradation of antitoxins encoded by the loci analyzed.

  6. Analytical solutions of steady-state conjugate heat transfer in ducts with turbulent flow

    International Nuclear Information System (INIS)

    Cerqueira, Djane R.; Jian Su

    2007-01-01

    In this work, we present an approximate analytical solution of the steady-state conjugate heat transfer of turbulent forced convection in a circular pipe with wall axial heat conduction and external convective boundary conditions. Improved lumped differential approach based on two points Hermite approximation for integrals was applied to reduce the heat conduction equation in the solid into a second-order ordinary differential equation for the radially averaged solid temperature. The energy equation in the fluid was solved by applying the generalized integral transform technique (GITT). The Sturm-Lioville eigenproblem for fluid energy equation in the cylindrical coordinate system was solved by the sign-count method. The truncated system of N ordinary differential equations for transformed potentials of the fluid temperature and the second-order ordinary differential equation for radially averaged solid temperature formed a homogeneous system of N+2 ordinary differential equations, which was solved analytically. The effects of the fluid-solid thermal conductivity ratio on the Nusselt number, the average fluid and solid temperatures, and the fluid-solid interface temperature were investigated. (author)

  7. Numerical analysis for the conjugate heat transfer of skin under various temperature conditions of contrast therapy

    Energy Technology Data Exchange (ETDEWEB)

    Park, Da Ae; Oh, Han Nah; Choi, Hyoung Gwon [Dept. of Mechanical and Automotive Engineering, Seoul National University of Science and Technology, Seoul (Korea, Republic of); Jeon, Byoung Jin; Kim, Eun Jeoung; Lee, Seung Deok [Yonsei University, Seoul (Korea, Republic of)

    2015-11-15

    In this paper, the contrast therapy of skin was numerically investigated by solving the conjugate heat transfer problem. A finite volume method based on the SIMPLE algorithm was adopted to solve the axisymmetric incompressible Navier-Stokes equations, coupled with an energy equation. These equations are strongly coupled with the Pennes bio-heat equation in order to consider the effect of blood perfusion rate. We investigated the thermal response of skin at some selected depths for various input temperature profiles of a stimulator for contrast therapy. From the numerical simulations, the regions with cold/hot threshold temperatures were found for five input temperature profiles. It was shown that the temperature varies mildly for different input profiles as the depth increases, owing to the Pennes effect. The input temperatures for effective hot/cold stimulation of dermis layer were found to be 47 degrees C and 7 degrees C, respectively. The present numerical results will be used for finding an optimal temperature profile of a stimulator for contrast therapy.

  8. Transfer of copper from a chelated 67Cu-antibody conjugate to ceruloplasmin in lymphoma patients

    International Nuclear Information System (INIS)

    Mirick, Gary R.; O'Donnell, Robert T.; DeNardo, Sally J.; Shen Sui; Meares, Claude F.; DeNardo, Gerald L.

    1999-01-01

    The Lym-1 monoclonal antibody was conjugated with the bifunctional chelating agent 6-[p-(bromoacetamido)benzyl]-1,4,8,11-tetraazacyclotetradecane-N,N',N'',N' -tetraacetic acid (BAT), using 2IT as a linker, and radiolabeled with 67 Cu to make the radiopharmaceutical, 67 Cu-2IT-BAT-Lym-1. Ten patients received a total of 18 doses of 67 Cu-2IT-BAT-Lym-1 as targeted, systemic radiotherapy. The beta phase of blood clearance, when corrected for 67 Cu decay, was positive or flat, a phenomenon not observed in similar patients treated with 131 I-Lym-1. The flat beta phase of blood clearance suggested recycling of 67 Cu from 67 Cu-2IT-BAT-Lym-1 to another plasma protein. Therefore, the amount of 67 Cu transferred from the radiopharmaceutical to CP, Alb, and TF was measured using affinity-purified polyclonal antibodies. The fraction of plasma 67 Cu precipitated by anti-human CP increased daily; most blood radioactivity was 67 Cu-CP after a median of 4 days (range 2-7 days). The transfer of 67 Cu to CP was observed in all patients and was consistent from dose to dose within the same patient. An average of 2.8±1.5% (range 0.8-7.8%) of the 67 Cu dose (%ID) was transferred to CP. The release rate of 67 Cu-CP from the liver into the blood was 0.9±0.4 %ID/day for the first 3 days. The 67 Cu-CP effective clearance half-life was 3.7 ± 0.7 days. Subtraction of the 67 Cu-CP activity from the total blood radioactivity yielded a biphasic blood clearance similar to that obtained for patients given 131 I-Lym-1. Cu-67-CP increased the AUC for whole blood by 24 ± 10%. The %ID of 67 Cu recycled correlated with GGT, ALT, and alkaline phosphatase levels; r=0.958 (p 67 Cu-2IT-BAT-Lym-1 and recycles a small fraction of the 67 Cu, transferring it to CP

  9. The Application of Discontinuous Galerkin Methods in Conjugate Heat Transfer Simulations of Gas Turbines

    Directory of Open Access Journals (Sweden)

    Zeng-Rong Hao

    2014-11-01

    Full Text Available The performance of modern heavy-duty gas turbines is greatly determined by the accurate numerical predictions of thermal loading on the hot-end components. The purpose of this paper is: (1 to present an approach applying a novel numerical technique—the discontinuous Galerkin (DG method—to conjugate heat transfer (CHT simulations, develop the engineering-oriented numerical platform, and validate the feasibility of the methodology and tool preliminarily; and (2 to utilize the constructed platform to investigate the aerothermodynamic features of a typical transonic turbine vane with convection cooling. Fluid dynamic and solid heat conductive equations are discretized into explicit DG formulations. A centroid-expanded Taylor basis is adopted for various types of elements. The Bassi-Rebay method is used in the computation of gradients. A coupled strategy based on a data exchange process via numerical flux on interface quadrature points is simply devised. Additionally, various turbulence Reynolds-Averaged-Navier-Stokes (RANS models and the local-variable-based transition model γ-Reθ are assimilated into the integral framework, combining sophisticated modelling with the innovative algorithm. Numerical tests exhibit good consistency between computational and analytical or experimental results, demonstrating that the presented approach and tool can handle well general CHT simulations. Application and analysis in the turbine vane, focusing on features around where there in cluster exist shock, separation and transition, illustrate the effects of Bradshaw’s shear stress limitation and separation-induced-transition modelling. The general overestimation of heat transfer intensity behind shock is conjectured to be associated with compressibility effects on transition modeling. This work presents an unconventional formulation in CHT problems and achieves its engineering applications in gas turbines.

  10. Characterization and plasmid elimination of NDM-1-producing Acinetobacter calcoaceticus from China.

    Directory of Open Access Journals (Sweden)

    Yang Sun

    Full Text Available The presence of multidrug-resistant bacterial pathogens in the environment poses a serious threat to public health. The opportunistic Acinetobacter spp. are among the most prevalent causes of nosocomial infections. Here, we performed complete genome sequencing of the Acinetobacter calcoaceticus strain XM1570, which was originally cultivated from the sputum of a patient diagnosed with pneumonia in Xiamen in 2010. We identified carbapenem resistance associated gene bla(NDM-1 located on a 47.3-kb plasmid. Three methods--natural reproduction, sodium dodecyl sulfate treatment and nalidixic acid treatment--were used to eliminate the bla(NDM-1-encoding plasmid, which achieved elimination rates of 3.32% (10/301, 83.78% (278/332, and 84.17% (298/354, respectively. Plasmid elimination dramatically increased antibiotic sensitivity, reducing the minimum bacteriostatic concentration of meropenem from 256 µg/ml in the clinical strain to 0.125 µg/ml in the plasmid-eliminated strain. Conjugation transfer assays showed that the bla(NDM-1-containing plasmid could be transferred into Escherichia coli DH5α:pBR322 in vitro as well as in vivo in mice. The bla(NDM-1 genetic environment was in accordance with that of other bla(NDM-1 genes identified from India, Japan, and Hong-Kong. The multilocus sequence type of the isolate was identified as ST-70. Two novel genes encoding intrinsic OXA and ADC were identified and named as OXA-417 and ADC-72. The finding of bla(NDM-1 in species like A. calcoaceticus demonstrates the wide spread of this gene in gram-negative bacteria which is possible by conjugative plasmid transfer. The results of this study may help in the development of a treatment strategy for controlling NDM-1 bacterial infection and transmission.

  11. Horizontal transfer of tet(M) and erm(B) resistance plasmids from food strains of Lactobacillus plantarum to Enterococcus faecalis JH2-2 in the gastrointestinal tract of gnotobiotic rats

    DEFF Research Database (Denmark)

    Jacobsen, Louise; Wilcks, Andrea; Hammer, Karin

    2007-01-01

    Two wild-type strains of Lactobacillus plantarum previously isolated from fermented dry sausages were analysed for their ability to transfer antibiotic resistance plasmids in the gastrointestinal tract. For this purpose, we used gnotobiotic rats as an in vivo model. Rats were initially inoculated......(2) CFU g(-1) faeces towards the end of the experiment. For erm(B)-TCs, the number was significantly higher and increased to c. 10(3) CFU g(-1) faeces. To our knowledge, this is the first study showing in vivo transfer of wild-type antibiotic resistance plasmids from L. plantarum to E. faecalis....

  12. Origin and Evolution of Rickettsial Plasmids.

    Directory of Open Access Journals (Sweden)

    Khalid El Karkouri

    Full Text Available Rickettsia species are strictly intracellular bacteria that have undergone a reductive genomic evolution. Despite their allopatric lifestyle, almost half of the 26 currently validated Rickettsia species have plasmids. In order to study the origin, evolutionary history and putative roles of rickettsial plasmids, we investigated the evolutionary processes that have shaped 20 plasmids belonging to 11 species, using comparative genomics and phylogenetic analysis between rickettsial, microbial and non-microbial genomes.Plasmids were differentially present among Rickettsia species. The 11 species had 1 to 4 plasmid (s with a size ranging from 12 kb to 83 kb. We reconstructed pRICO, the last common ancestor of the current rickettsial plasmids. pRICO was vertically inherited mainly from Rickettsia/Orientia chromosomes and diverged vertically into a single or multiple plasmid(s in each species. These plasmids also underwent a reductive evolution by progressive gene loss, similar to that observed in rickettsial chromosomes, possibly leading to cryptic plasmids or complete plasmid loss. Moreover, rickettsial plasmids exhibited ORFans, recent gene duplications and evidence of horizontal gene transfer events with rickettsial and non-rickettsial genomes mainly from the α/γ-proteobacteria lineages. Genes related to maintenance and plasticity of plasmids, and to adaptation and resistance to stress mostly evolved under vertical and/or horizontal processes. Those involved in nucleotide/carbohydrate transport and metabolism were under the influence of vertical evolution only, whereas genes involved in cell wall/membrane/envelope biogenesis, cycle control, amino acid/lipid/coenzyme and secondary metabolites biosynthesis, transport and metabolism underwent mainly horizontal transfer events.Rickettsial plasmids had a complex evolution, starting with a vertical inheritance followed by a reductive evolution associated with increased complexity via horizontal gene

  13. Numerical analysis of unsteady conjugate heat transfer for initial evolution of thermal stratification in a curved pipe

    International Nuclear Information System (INIS)

    Jo, Jong Chull; Kim, Wee Kyung; Kim, Yun Il; Cho, Sang Jin; Choi, Seok Ki

    2000-01-01

    A detailed numerical analysis of initial evolution of thermal stratification in a curved pipe with a finite wall thickness is performed. A primary emphasis of the present study is placed on the investigation of the effect of existence of pipe wall thickness on the evolution of thermal stratification. A simple and convenient numerical method of treating the unsteady conjugate heat transfer in Cartesian as well as non-orthogonal coordinate systems is presented. The proposed unsteady conjugate heat transfer analysis method is implemented in a finite volume thermal-hydraulic computer code based on a cell-centered, non-staggered grid arrangement, the SIMPLEC algorithm and a higher-order bounded convection scheme. Calculations are performed for initial evolution of thermal stratification with high Richardson number in a curved pipe. The predicted results show that the thermally stratified flow and transient conjugate heat transfer in a curved pipe with a specified wall thickness can be satisfactorily analyzed by using the numerical method presented in this paper. As the result, the present analysis method is considered to be effective for the determination of transient temperature distributions in the wall of curved piping system subjected to internally thermal stratification. In addition, the method can be extended to be applicable for the simulation of turbulent flow of thermally stratified fluid

  14. Comparative Genomics of Rhodococcus equi Virulence Plasmids Indicates Host-Driven Evolution of the vap Pathogenicity Island.

    Science.gov (United States)

    MacArthur, Iain; Anastasi, Elisa; Alvarez, Sonsiray; Scortti, Mariela; Vázquez-Boland, José A

    2017-05-01

    The conjugative virulence plasmid is a key component of the Rhodococcus equi accessory genome essential for pathogenesis. Three host-associated virulence plasmid types have been identified the equine pVAPA and porcine pVAPB circular variants, and the linear pVAPN found in bovine (ruminant) isolates. We recently characterized the R. equi pangenome (Anastasi E, et al. 2016. Pangenome and phylogenomic analysis of the pathogenic actinobacterium Rhodococcus equi. Genome Biol Evol. 8:3140-3148.) and we report here the comparative analysis of the virulence plasmid genomes. Plasmids within each host-associated type were highly similar despite their diverse origins. Variation was accounted for by scattered single nucleotide polymorphisms and short nucleotide indels, while larger indels-mostly in the plasticity region near the vap pathogencity island (PAI)-defined plasmid genomic subtypes. Only one of the plasmids analyzed, of pVAPN type, was exceptionally divergent due to accumulation of indels in the housekeeping backbone. Each host-associated plasmid type carried a unique PAI differing in vap gene complement, suggesting animal host-specific evolution of the vap multigene family. Complete conservation of the vap PAI was observed within each host-associated plasmid type. Both diversity of host-associated plasmid types and clonality of specific chromosomal-plasmid genomic type combinations were observed within the same R. equi phylogenomic subclade. Our data indicate that the overall strong conservation of the R. equi host-associated virulence plasmids is the combined result of host-driven selection, lateral transfer between strains, and geographical spread due to international livestock exchanges. © The Author(s) 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  15. Survival and evolution of a large multidrug resistance plasmid in new clinical bacterial hosts

    DEFF Research Database (Denmark)

    Porse, Andreas; Schønning, Kristian; Munck, Christian

    2016-01-01

    Large conjugative plasmids are important drivers of bacterial evolution and contribute significantly to the dissemination of antibiotic resistance. Although plasmid borne multidrug resistance is recognized as one of the main challenges in modern medicine, the adaptive forces shaping the evolution...

  16. Social behaviour and decision making in bacterial conjugation

    Directory of Open Access Journals (Sweden)

    Günther eKoraimann

    2014-04-01

    Full Text Available Bacteria frequently acquire novel genes by HGT (horizontal gene transfer. HGT through the process of bacterial conjugation is highly efficient and depends on the presence of conjugative plasmids (CPs or integrated conjugative elements (ICEs that provide the necessary genes for DNA transmission. This review focuses on recent advancements in our understanding of ssDNA transfer systems and regulatory networks ensuring timely and spatially controlled DNA transfer (tra gene expression. As will become obvious by comparing different systems, by default, tra genes are shut off in cells in which conjugative elements are present. Only when conditions are optimal, donor cells – through epigenetic alleviation of negatively acting roadblocks and direct stimulation of DNA transfer genes – become transfer competent. These transfer competent cells have developmentally transformed into specialized cells capable of secreting ssDNA via a T4S (type IV secretion complex directly into recipient cells. Intriguingly, even under optimal conditions, only a fraction of the population undergoes this transition, a finding that indicates specialization and cooperative, social behavior. Thereby, at the population level, the metabolic burden and other negative consequences of tra gene expression are greatly reduced without compromising the ability to horizontally transfer genes to novel bacterial hosts. This undoubtedly intelligent strategy may explain why conjugative elements – CPs and ICEs – have been successfully kept in and evolved with bacteria to constitute a major driving force of bacterial evolution.

  17. Transfer of herpes simplex virus thymidine kinase synthesized in bacteria by a high-expression plasmid to tissue culture cells by protoplast fusion

    International Nuclear Information System (INIS)

    Waldman, A.S.; Milman, G.

    1984-01-01

    The introduction of a protein into living tissue culture cells may permit the in vivo study of functions of the protein. The authors have previously described a high-efficiency-expression plasmid, pHETK2, containing the herpes simplex virus type 1 thymidine kinase (TK) gene which, upon temperature induction, causes TK to be synthesized as greater than 4% of the bacterial protein. In this report it is shown that enzymatically active TK was transferred to mouse Ltk- cells by polyethylene glycol-mediated fusion with protoplasts prepared from bacteria containing induced levels of TK. The presence of TK in the Ltk- cells was detected by the incorporation of [ 3 H]thymidine into cell nuclei as measured by autoradiography

  18. Conjugated heat transfer of natural convection in pool with internal heat sources and convection in the tube

    International Nuclear Information System (INIS)

    Li Longjian; Liu Hongtao; Cui Wenzhi

    2007-01-01

    The conjugated heat transfer of natural convection in pool with internal heat source and the forced convection in the tube was analyzed, and the corresponding three-dimensional physical and mathematical model was proposed. A control volume based finite element method was employed to solve numerically the problem. The computations were performed for different internal heat source intensity of the pool and the different flow velocity in the tube. The computed heat transfer coefficients on the inner and outer wall showed well consistency of those calculated with the empirical correlations. Compared with the measured total heat transfer coefficients between the fluids in and out of the tube, the computed ones showed also the well consistency, which implied that the numerical model proposed in this paper was reliable. The research results revealed that the total heat transfer coefficients between the fluids were strongly affected by the internal heat source intensity of the pool liquid and the flow velocity in the tube. (authors)

  19. Isolation and characterization of kikA, a region on IncN group plasmids that determines killing of Klebsiella oxytoca.

    OpenAIRE

    Hengen, P N; Denicourt, D; Iyer, V N

    1992-01-01

    Transfer of the IncN group plasmid pCU1 from Escherichia coli to Klebsiella oxytoca by conjugation kills a large proportion (90 to 95%) of the recipients of plasmid DNA, whereas transfer to E. coli or even to the closely related Enterobacter aerogenes does not. Two regions, kikA and kikB, have been identified on pCU1 that contribute to the Kik (killing in klebsiellas) phenotype. We have localized the kikA region to 500 bp by deletion analysis and show by DNA-DNA hybridization that kikA is hig...

  20. Sub-inhibitory concentrations of heavy metals facilitate the horizontal transfer of plasmid-mediated antibiotic resistance genes in water environment.

    Science.gov (United States)

    Zhang, Ye; Gu, April Z; Cen, Tianyu; Li, Xiangyang; He, Miao; Li, Dan; Chen, Jianmin

    2018-06-01

    Although widespread antibiotic resistance has been mostly attributed to the selective pressure generated by overuse and misuse of antibiotics, recent growing evidence suggests that chemicals other than antibiotics, such as certain metals, can also select and stimulate antibiotic resistance via both co-resistance and cross-resistance mechanisms. For instance, tetL, merE, and oprD genes are resistant to both antibiotics and metals. However, the potential de novo resistance induced by heavy metals at environmentally-relevant low concentrations (much below theminimum inhibitory concentrations [MICs], also referred as sub-inhibitory) has hardly been explored. This study investigated and revealed that heavy metals, namely Cu(II), Ag(I), Cr(VI), and Zn(II), at environmentally-relevant and sub-inhibitory concentrations, promoted conjugative transfer of antibiotic resistance genes (ARGs) between E. coli strains. The mechanisms of this phenomenon were further explored, which involved intracellular reactive oxygen species (ROS) formation, SOS response, increased cell membrane permeability, and altered expression of conjugation-relevant genes. These findings suggest that sub-inhibitory levels of heavy metals that widely present in various environments contribute to the resistance phenomena via facilitating horizontal transfer of ARGs. This study provides evidence from multiple aspects implicating the ecological effect of low levels of heavy metals on antibiotic resistance dissemination and highlights the urgency of strengthening efficacious policy and technology to control metal pollutants in the environments. Copyright © 2018 Elsevier Ltd. All rights reserved.

  1. Sequential acquisition of R-plasmids in vivo by Salmonella typhimurium.

    Science.gov (United States)

    Platt, D J; Sommerville, J S; Gribben, J

    1984-01-01

    Salmonella typhimurium, resistant only to trimethoprim and sulphamethoxazole, was isolated from the faeces and blood of a chronic alcoholic patient in acute renal failure. The isolates harboured an 18 Md non-conjugative plasmid. He was dialysed peritoneally and treated with ampicillin; four days later there was no clinical improvement and his peritoneal dialysis fluid (PDF) had become infected. Salm. typhimurium was isolated from faeces and PDF. Both isolates were additionally resistant to ampicillin and contained two plasmids (55 Md and 18 Md). Therapy was changed to chloramphenicol and gentamicin was added to the PDF. Two weeks later Salm. typhimurium was again isolated from PDF and faeces. The PDF isolate was unchanged but 4% of the colonies isolated from this faecal specimen were resistant to chloramphenicol and had acquired an additional 62 Md plasmid. From all PDF and faecal specimens two different strains of Escherichia coli and one strain of Klebsiella pneumoniae were isolated which contained plasmids indistinguishable, on the basis of molecular weight and transferable resistance markers, from those acquired by Salm. typhimurium. The transferability of these plasmids in vitro to E. coli K12 and to the patient's initial Salm. typhimurium was studied and the results discussed.

  2. Dissemination of plasmid-encoded AmpC β-lactamases in antimicrobial resistant Salmonella serotypes originating from humans, pigs and the swine environment.

    Science.gov (United States)

    Keelara, Shivaramu; Thakur, Siddhartha

    2014-09-17

    The aim of this study was to characterize and determine the inter-serovar exchange of AmpC β-lactamase conferring plasmids isolated from humans, pigs and the swine environment. Plasmids isolated from a total of 21 antimicrobial resistant (AMR) Salmonella isolates representing human clinical cases (n=6), pigs (n=6) and the swine farm environment (n=9) were characterized by replicon typing and restriction digestion, inter-serovar transferability by conjugation, and presence of AmpC β-lactamase enzyme encoding gene blaCMY-2 by southern hybridization. Based on replicon typing, the majority (17/21, 81%) of the plasmids belonged to the I1-Iγ Inc group and were between 70 and 103kb. The potential for inter-serovar plasmid transfer was further confirmed by the PCR detection of AMR genes on the plasmids isolated from trans-conjugants. Plasmids from Salmonella serovars Anatum, Ouakam, Johannesburg and Typhimurium isolated from the same cohort of pigs and their environment and S. Heidelberg from a single human clinical isolate had identical plasmids based on digestion with multiple restriction enzymes (EcoRI, HindIII and PstI) and southern blotting. We demonstrated likely horizontal inter-serovar exchange of plasmid-encoding AmpC β-lactamases resistance among MDR Salmonella serotypes isolated from pigs, swine farm environment and clinical human cases. This study provides valuable information on the role of the swine farm environment and by extension other livestock farm environments, as a potential reservoir of resistant bacterial strains that potentially transmit resistance determinants to livestock, in this case, swine, humans and possibly other hosts by horizontal exchange of plasmids. Copyright © 2014 Elsevier B.V. All rights reserved.

  3. Sequence Analysis of IncA/C and IncI1 Plasmids Isolated from Multidrug-Resistant Salmonella Newport Using Single-Molecule Real-Time Sequencing.

    Science.gov (United States)

    Cao, Guojie; Allard, Marc; Hoffmann, Maria; Muruvanda, Tim; Luo, Yan; Payne, Justin; Meng, Kevin; Zhao, Shaohua; McDermott, Patrick; Brown, Eric; Meng, Jianghong

    2018-04-05

    Multidrug-resistant (MDR) plasmids play an important role in disseminating antimicrobial resistance genes. To elucidate the antimicrobial resistance gene compositions in A/C incompatibility complex (IncA/C) plasmids carried by animal-derived MDR Salmonella Newport, and to investigate the spread mechanism of IncA/C plasmids, this study characterizes the complete nucleotide sequences of IncA/C plasmids by comparative analysis. Complete nucleotide sequencing of plasmids and chromosomes of six MDR Salmonella Newport strains was performed using PacBio RSII. Open reading frames were assigned using prokaryotic genome annotation pipeline (PGAP). To understand genomic diversity and evolutionary relationships among Salmonella Newport IncA/C plasmids, we included three complete IncA/C plasmid sequences with similar backbones from Salmonella Newport and Escherichia coli: pSN254, pAM04528, and peH4H, and additional 200 draft chromosomes. With the exception of canine isolate CVM22462, which contained an additional IncI1 plasmid, each of the six MDR Salmonella Newport strains contained only the IncA/C plasmid. These IncA/C plasmids (including references) ranged in size from 80.1 (pCVM21538) to 176.5 kb (pSN254) and carried various resistance genes. Resistance genes floR, tetA, tetR, strA, strB, sul, and mer were identified in all IncA/C plasmids. Additionally, bla CMY-2 and sugE were present in all IncA/C plasmids, excepting pCVM21538. Plasmid pCVM22462 was capable of being transferred by conjugation. The IncI1 plasmid pCVM22462b in CVM22462 carried bla CMY-2 and sugE. Our data showed that MDR Salmonella Newport strains carrying similar IncA/C plasmids clustered together in the phylogenetic tree using chromosome sequences and the IncA/C plasmids from animal-derived Salmonella Newport contained diverse resistance genes. In the current study, we analyzed genomic diversities and phylogenetic relationships among MDR Salmonella Newport using complete plasmids and chromosome

  4. Smooth and robust solutions for Dirichlet boundary control of fluid-solid conjugate heat transfer problems

    KAUST Repository

    Yan, Yan

    2015-01-01

    We study a new optimization scheme that generates smooth and robust solutions for Dirichlet velocity boundary control (DVBC) of conjugate heat transfer (CHT) processes. The solutions to the DVBC of the incompressible Navier-Stokes equations are typically nonsmooth, due to the regularity degradation of the boundary stress in the adjoint Navier-Stokes equations. This nonsmoothness is inherited by the solutions to the DVBC of CHT processes, since the CHT process couples the Navier-Stokes equations of fluid motion with the convection-diffusion equations of fluid-solid thermal interaction. Our objective in the CHT boundary control problem is to select optimally the fluid inflow profile that minimizes an objective function that involves the sum of the mismatch between the temperature distribution in the fluid system and a prescribed temperature profile and the cost of the control.Our strategy to resolve the nonsmoothness of the boundary control solution is based on two features, namely, the objective function with a regularization term on the gradient of the control profile on both the continuous and the discrete levels, and the optimization scheme with either explicit or implicit smoothing effects, such as the smoothed Steepest Descent and the Limited-memory Broyden-Fletcher-Goldfarb-Shanno (L-BFGS) methods. Our strategy to achieve the robustness of the solution process is based on combining the smoothed optimization scheme with the numerical continuation technique on the regularization parameters in the objective function. In the section of numerical studies, we present two suites of experiments. In the first one, we demonstrate the feasibility and effectiveness of our numerical schemes in recovering the boundary control profile of the standard case of a Poiseuille flow. In the second one, we illustrate the robustness of our optimization schemes via solving more challenging DVBC problems for both the channel flow and the flow past a square cylinder, which use initial

  5. Identification of oriT and a recombination hot spot in the IncA/C plasmid backbone.

    Science.gov (United States)

    Hegyi, Anna; Szabó, Mónika; Olasz, Ferenc; Kiss, János

    2017-09-06

    Dissemination of multiresistance has been accelerating among pathogenic bacteria in recent decades. The broad host-range conjugative plasmids of the IncA/C family are effective vehicles of resistance determinants in Gram-negative bacteria. Although more than 150 family members have been sequenced to date, their conjugation system and other functions encoded by the conserved plasmid backbone have been poorly characterized. The key cis-acting locus, the origin of transfer (oriT), has not yet been unambiguously identified. We present evidence that IncA/C plasmids have a single oriT locus immediately upstream of the mobI gene encoding an indispensable transfer factor. The fully active oriT spans ca. 150-bp AT-rich region overlapping the promoters of mobI and contains multiple inverted and direct repeats. Within this region, the core domain of oriT with reduced but detectable transfer activity was confined to a 70-bp segment containing two inverted repeats and one copy of a 14-bp direct repeat. In addition to oriT, a second locus consisting of a 14-bp imperfect inverted repeat was also identified, which mimicked the function of oriT but which was found to be a recombination site. Recombination between two identical copies of these sites is RecA-independent, requires a plasmid-encoded recombinase and resembles the functioning of dimer-resolution systems.

  6. Validation of ANSYS CFX for gas and liquid metal flows with conjugate heat transfer within the European project THINS

    Energy Technology Data Exchange (ETDEWEB)

    Papukchiev, A., E-mail: angel.papukchiev@grs.de; Buchholz, S.

    2017-02-15

    Highlights: • ANSYS CFX is validated for gas and liquid metal flows. • L-STAR and TALL-3D experiments are simulated. • Complex flow and heat transfer phenomena are modelled. • Conjugate heat transfer has to be considered in CFD analyses. - Abstract: Within the FP7 European project THINS (Thermal Hydraulics of Innovative Nuclear Systems), numerical tools for the simulation of the thermal-hydraulics of next generation rector systems were developed, applied and validated for innovative coolants. The Gesellschaft fuer Anlagen- und Reaktorsicherheit (GRS) gGmbH participated in THINS with activities related to the development and validation of computational fluid dynamics (CFD) and coupled System Thermal Hydraulics (STH) – CFD codes. High quality measurements from the L-STAR and TALL-3D experiments were used to assess the numerical results. Two-equation eddy viscosity and scale resolving turbulence models were used in the validation process of ANSYS CFX for gas and liquid metal flows with conjugate heat transfer. This paper provides a brief overview on the main results achieved at GRS within the project.

  7. Broad host range plasmids can invade an unexpectedly diverse fraction of a soil bacterial community

    DEFF Research Database (Denmark)

    Klümper, Uli; Riber, Leise; Dechesne, Arnaud

    2014-01-01

    and Actinobacteria suggests that inter-Gram plasmid transfer of IncP-1 and IncPromA-type plasmids is a frequent phenomenon. While the plasmid receiving fractions of the community were both plasmid- and donor- dependent, we identified a core super-permissive fraction that could take up different plasmids from diverse...

  8. Interface model conditions for a non-equilibrium heat transfer model for conjugate fluid/porous/solid domains

    International Nuclear Information System (INIS)

    Betchen, L.J.; Straatman, A.G.

    2005-01-01

    A mathematical and numerical model for the treatment of conjugate fluid flow and heat transfer problems in domains containing pure fluid, porous, and pure solid regions has been developed. The model is general and physically reasoned, and allows for local thermal non-equilibrium in the porous region. The model is developed for implementation on a simple collocated finite volume grid. Of particular novelty are the conditions implemented at the interfaces between porous regions, and those containing a pure solid or pure fluid. The model is validated by simulation of a three-dimensional porous plug problem for which experimental results are available. (author)

  9. Conjugate gradient determination of optimal plane changes for a class of three-impulse transfers between noncoplanar circular orbits

    Science.gov (United States)

    Burrows, R. R.

    1972-01-01

    A particular type of three-impulse transfer between two circular orbits is analyzed. The possibility of three plane changes is recognized, and the problem is to optimally distribute these plane changes to minimize the sum of the individual impulses. Numerical difficulties and their solution are discussed. Numerical results obtained from a conjugate gradient technique are presented for both the case where the individual plane changes are unconstrained and for the case where they are constrained. Possibly not unexpectedly, multiple minima are found. The techniques presented could be extended to the finite burn case, but primarily the contents are addressed to preliminary mission design and vehicle sizing.

  10. Integrated conjugate heat transfer analysis method for in-vessel retention with external reactor vessel cooling - 15477

    International Nuclear Information System (INIS)

    Park, J.W.; Bae, J.H.; Seol, W.C.

    2015-01-01

    An integrated conjugate heat transfer analysis method for the thermal integrity of a reactor vessel under external reactor vessel cooling conditions is developed to resolve light metal layer focusing effect issue. The method calculates steady-state 3-dimensional temperature distribution of a reactor vessel using coupled conjugate heat transfer between in-vessel 3-layered stratified corium (metallic pool, oxide pool and heavy metal) and polar-angle dependent boiling heat transfer at the outer surface of a reactor vessel. The 3-layer corium heat transfer model is utilizing lumped-parameter thermal-resistance circuit method and ex-vessel boiling regimes are parametrically considered. The thermal integrity of a reactor vessel is addressed in terms of un-molten thickness profile. The vessel 3-dimensional heat conduction is validated against a commercial code. It is found that even though the internal heat flux from the metal layer goes far beyond critical heat flux (CHF) the heat flux from the outermost nodes of the vessel may be maintained below CHF due to massive vessel heat diffusion. The heat diffusion throughout the vessel is more pronounced for relatively low heat generation rate in an oxide pool. Parametric calculations are performed considering thermal conditions such as peak heat flux from a light metal layer, heat generation in an oxide pool and external boiling conditions. The major finding is that the most crucial factor for success of in-vessel retention is not the mass of the molten light metal above the oxide pool but the heat generation rate inside the oxide pool and the 3-dimensional vessel heat transfer provides a much larger minimum vessel wall thickness. (authors)

  11. Destabilization of IncA and IncC plasmids by SGI1 and SGI2 type Salmonella genomic islands.

    Science.gov (United States)

    Harmer, Christopher J; Hamidian, Mohammad; Ambrose, Stephanie J; Hall, Ruth M

    Both the Salmonella genomic islands (SGI) and the conjugative IncC plasmids are known to contribute substantially to the acquisition of resistance to multiple antibiotics, and plasmids in the A/C group are known to mobilize the Salmonella genomic island SGI1, which also carries multiple antibiotic resistance genes. Plasmid pRMH760 (IncC; A/C 2 ) was shown to mobilize SGI1 variants SGI1-I, SGI1-F, SGI1-K and SGI2 from Salmonella enterica to Escherichia coli where it was integrated at the preferred location, at the end of the trmE (thdF) gene. The plasmid was transferred at a similar frequency. However, we observed that co-transfer of the SGI and the plasmid was rarer. In E. coli to E. coli transfer, the frequency of transfer of the IncC plasmid pRMH760 was at least 1000-fold lower when the donor carried SGI1-I or SGI1-K, indicating that the SGI suppresses transfer of the plasmid. In addition, pRMH760 was rapidly lost from both E. coli and S. enterica strains that also carried SGI1-I, SGI1-F or SGI2. However, plasmid loss was not seen when the SGI1 variant was SGI1-K, which lacks two segments of the SGI1 backbone. The complete sequence of the SGI1-I and SGI1-F were determined and SGI1-K also carries two single base substitutions relative to SGI1-I. The IncA (A/C 1 ) plasmid RA1 was also shown to mobilize SGI2-A and though there are significant differences between the backbones of IncA and IncC plasmids, RA1 was also rapidly lost when SGI2-A was present in the same cell. We conclude that there are multiple interactions, both cooperative and antagonistic, between an IncA or IncC plasmid and the SGI1 and SGI2 family genomic islands. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Horizontal Transfer of the Salmonella enterica Serovar Infantis Resistance and Virulence Plasmid pESI to the Gut Microbiota of Warm-Blooded Hosts

    Directory of Open Access Journals (Sweden)

    Gili Aviv

    2016-09-01

    Full Text Available Salmonella enterica serovar Infantis is one of the prevalent salmonellae worldwide. Recently, we showed that the emergence of S. Infantis in Israel was facilitated by the acquisition of a unique megaplasmid (pESI conferring multidrug resistance and increased virulence phenotypes. Here we elucidate the ecology, transmission properties, and regulation of pESI. We show that despite its large size (~280 kb, pESI does not impose a significant metabolic burden in vitro and that it has been recently fixed in the domestic S. Infantis population. pESI conjugation and the transcription of its pilus (pil genes are inhibited at the ambient temperature (27°C and by ≥1% bile but increased under temperatures of 37 to 41°C, oxidative stress, moderate osmolarity, and the microaerobic conditions characterizing the intestinal environment of warm-blooded animals. The pESI-encoded protein TraB and the oxygen homeostasis regulator Fnr were identified as transcriptional regulators of pESI conjugation. Using the mouse model, we show that following S. Infantis infection, pESI can be horizontally transferred to the gut microbiota, including to commensal Escherichia coli strains. Possible transfer, but not persistence, of pESI was also observed into Gram-positive mouse microbiota species, especially Lactobacillus reuteri. Moreover, pESI was demonstrated to further disseminate from gut microbiota to S. enterica serovar Typhimurium, in the context of gastrointestinal infection. These findings exhibit the ability of a selfish clinically relevant megaplasmid to distribute to and from the microbiota and suggest an overlooked role of the microbiota as a reservoir of mobile genetic elements and intermediator in the spread of resistance and virulence genes between commensals and pathogenic bacteria.

  13. Effect of genomic location on horizontal transfer of a recombinant gene cassette between Pseudomonas strains in the rhizosphere and spermosphere of barley seedlings

    DEFF Research Database (Denmark)

    Sengelov, G.; Kristensen, K. J.; Sørensen, Anders Morten Hay

    2001-01-01

    , horizontal transfer of a recombinant gene cassette inserted into the chromosome of a Pseudomonas strutzeri strain, into a mobilizable plasmid (pAGM42), and into a conjugative plasmid (pKJK5) isolated from barley rhizosphere was investigated. Horizontal transfer efficiencies of the gene cassette inserted...... efficiencies were up to 4.36 x 10(-3) transconjugants/(donors x recipients)(1/2). Transfer of chromosomal encoded genes could not be detected in the microcosms by conjugation or transformation. However, transformation did occur by using the same bacterial strains under laboratory conditions. The rhizosphere...

  14. In Vivo Transfer and Microevolution of Avian Native IncA/C2blaNDM-1-Carrying Plasmid pRH-1238 during a Broiler Chicken Infection Study.

    Science.gov (United States)

    Hadziabdic, Sead; Fischer, Jennie; Malorny, Burkhard; Borowiak, Maria; Guerra, Beatriz; Kaesbohrer, Annemarie; Gonzalez-Zorn, Bruno; Szabo, Istvan

    2018-04-01

    The emergence and spread of carbapenemase-producing Enterobacteriaceae (CPE) in wildlife and livestock animals pose an important safety concern for public health. With our in vivo broiler chicken infection study, we investigated the transfer and experimental microevolution of the bla NDM-1 -carrying IncA/C 2 plasmid (pRH-1238) introduced by avian native Salmonella enterica subsp. enterica serovar Corvallis without inducing antibiotic selection pressure. We evaluated the dependency of the time point of inoculation on donor ( S Corvallis [12-SA01738]) and plasmid-free Salmonella recipient [d-tartrate-fermenting (d-Ta + ) S Paratyphi B (13-SA01617), referred to here as S Paratyphi B (d-Ta + )] excretion by quantifying their excretion dynamics. Using plasmid profiling by S1 nuclease-restricted pulsed-field gel electrophoresis, we gained insight into the variability of the native plasmid content among S Corvallis reisolates as well as plasmid acquisition in S Paratyphi B (d-Ta + ) and the enterobacterial gut microflora. Whole-genome sequencing enabled us to gain an in-depth insight into the microevolution of plasmid pRH-1238 in S Corvallis and enterobacterial recipient isolates. Our study revealed that the fecal excretion of avian native carbapenemase-producing S Corvallis is significantly higher than that of S Paratyphi (d-Ta + ) and is not hampered by S Paratyphi (d-Ta + ). Acquisition of pRH-1238 in other Enterobacteriaceae and several events of plasmid pRH-1238 transfer to different Escherichia coli sequence types and Klebsiella pneumoniae demonstrated an interspecies broad host range. Regardless of the microevolutionary structural deletions in pRH-1238, the single carbapenem resistance marker bla NDM-1 was maintained on pRH-1238 throughout the trial. Furthermore, we showed the importance of the gut E. coli population as a vector of pRH-1238. In a potential scenario of the introduction of NDM-1-producing S Corvallis into a broiler flock, the pRH-1238 plasmid could

  15. Characterization of class 1 integrons associated with R-plasmids in clinical Aeromonas salmonicida isolates from various geographical areas

    DEFF Research Database (Denmark)

    Schmidt, A.S.; Bruun, Morten Sichlau; Larsen, J.L.

    2001-01-01

    Class 1 integrons were found in 26 of 40 antibiotic-resistant isolates of the fish pathogen Aeromonas salmonicida from Northern Europe and North America. Three different dhfr genes, conferring trimethoprim resistance, and one ant(3 " )1a aminoglycoside resistance gene were identified as gene...... inserts. The gene cassettes tended to be conserved among isolates from a particular geographical area. Nineteen isolates transferred R- plasmids carrying different tet determinants to Escherichia coli in filter mating assays, and in 15 cases, the class 1 integrons were co-transferred. Transferable...... sulphadiazine, trimethoprim and streptomycin resistances were invariably encoded by integrons. It thus appears that integron-encoded antibiotic resistance genes contribute substantially to the horizontal spread of antimicrobial resistance within this species, being associated with conjugative plasmids....

  16. Evidence for the horizontal transfer of an integrase gene from a fusellovirus to a pRN-like plasmid within a single strain of Sulfolobus and the implications for plasmid survival

    DEFF Research Database (Denmark)

    Peng, Xu

    2008-01-01

    of the integrase gene occurs in the viral attachment site (attP), which corresponds to the anticodon region of the targeted tRNA gene in the host chromosome. This point mutation confers on pXZ1 the ability to integrate into the tRNA(Glu)[CUC] gene, which differs from the integration site of SSV4, t......RNA(Glu)[UUC]. SSV4 and pXZ1 were also shown experimentally to integrate into separate sites on the host chromosome. This is believed to be the first report of a pRN plasmid sharing its natural host with a fusellovirus and carrying a highly similar integrase gene....

  17. Numerical analysis of the transient conjugated heat transfer in a circular duct with a power-law fluid

    Energy Technology Data Exchange (ETDEWEB)

    Luna, N. [Secretaria de Energia, Direccion de Operacion Petrolera, Mexico DF (Mexico); Mendez, F. [UNAM, Facultad de Ingenieria, Mexico DF (Mexico); Bautista, O. [ITESM, Division de Ingenieria y Arquitectura, Mexico DF (Mexico)

    2005-05-01

    We treat numerically in this paper, the transient analysis of a conjugated heat transfer process in the thermal entrance region of a circular tube with a fully developed laminar power-law fluid flow. We apply the quasi-steady approximation for the power-law fluid, identifying the suitable time scales of the process. Thus, the energy equation in the fluids is solved analytically using the well-known integral boundary layer technique. This solution is coupled to the transient energy equation for the solid where the transverse and longitudinal heat conduction effects are taken into account. The numerical results for the temporal evolution of the average temperature of the tube wall, {theta}{sub av,} is plotted for different nondimensional parameters such as conduction parameter, {alpha}, the aspect ratios of the tube, {epsilon} and {epsilon}{sub 0} and the index of power-law fluid, n. (orig.)

  18. NUMERICAL MODELING OF CONJUGATE HEAT TRANSFER IN AN INSULATED GLASS UNIT (IGU WITH ACCOUNT FOR ITS DEFORMATION

    Directory of Open Access Journals (Sweden)

    Golubev Stanislav Sergeevich

    2012-12-01

    The effects of different climatic impacts lead to the deformation of glasses within an IGU (and its vertical cavity, respectively. Deformation of glasses and vertical cavities reduces the thermal resistance of an IGU. A numerical simulation of conjugate heat transfer within an IGU was implemented as part of the research into this phenomenon. Calculations were performed in ANSYS FLUENT CFD package. Basic equations describing the conservation of mass, conservation of momentum (in the Boussinesq approximation, conservation of energy were solved. Also, the radiation of the cavity wall was taken into account. Vertical walls were considered as non-isothermal, while horizontal walls were adiabatic. Calculations were made for several patterns of glass deformations. Calculation results demonstrate that the heat flow over vertical walls intensifies as the distance between centres of IGU glasses is reduced. The temperature in the central area of the hot glass drops.

  19. Excitation energy transfer in ruthenium (II)-porphyrin conjugates led to enhanced emission quantum yield and 1O2 generation

    International Nuclear Information System (INIS)

    Pan, Jie; Jiang, Lijun; Chan, Chi-Fai; Tsoi, Tik-Hung; Shiu, Kwok-Keung; Kwong, Daniel W.J.; Wong, Wing-Tak; Wong, Wai-Kwok; Wong, Ka-Leung

    2017-01-01

    Porphyrins are good photodynamic therapy (PDT) agents due to its flexibility for modifications to achieve tumor localization and photo-cytotoxicity against cancer. Yet they are not perfect. In a Ru(polypyridyl)-porphyrin system, the Ru(polypyridyl) moiety improves the water solubility and cell permeability. Consider the similar excited state energies between Ru(polypyridyl) and porphyrin moieties; a small perturbation (e.g. Zn(II) metalation) would lead to a marked change in the energy migration process. In this work, we have synthesized a series of porphyrins conjugated with Ru(polypyridyl) complexes using different linkers and investigated their photophysical properties, which included singlet oxygen quantum yield and their in vitro biological properties, resulting from linker variation and porphyrin modification by Zn(II) metalation. - Graphical abstract: Four amphiphilic ruthenium(II)-porphyrin complexes were prepared that display energy transfer conversion with zinc coordination, lysosome specific target, low dark toxicity and efficient photodynamic therapy.

  20. Tetrapeptide-coumarin conjugate 3D networks based on hydrogen-bonded charge transfer complexes: gel formation and dye release.

    Science.gov (United States)

    Guo, Zongxia; Gong, Ruiying; Jiang, Yi; Wan, Xiaobo

    2015-08-14

    Oligopeptide-based derivatives are important synthons for bio-based functional materials. In this article, a Gly-(L-Val)-Gly-(L-Val)-coumarin (GVGV-Cou) conjugate was synthesized, which forms 3D networks in ethanol. The gel nanostructures were characterized by UV-vis spectroscopy, FT-IR spectroscopy, X-ray diffraction (XRD), SEM and TEM. It is suggested that the formation of charge transfer (CT) complexes between the coumarin moieties is the main driving force for the gel formation. The capability of the gel to encapsulate and release dyes was explored. Both Congo Red (CR) and Methylene Blue (MB) can be trapped in the CT gel matrix and released over time. The present gel might be used as a functional soft material for guest encapsulation and release.

  1. Solution to Two-Dimensional Steady Inverse Heat Transfer Problems with Interior Heat Source Based on the Conjugate Gradient Method

    Directory of Open Access Journals (Sweden)

    Shoubin Wang

    2017-01-01

    Full Text Available The compound variable inverse problem which comprises boundary temperature distribution and surface convective heat conduction coefficient of two-dimensional steady heat transfer system with inner heat source is studied in this paper applying the conjugate gradient method. The introduction of complex variable to solve the gradient matrix of the objective function obtains more precise inversion results. This paper applies boundary element method to solve the temperature calculation of discrete points in forward problems. The factors of measuring error and the number of measuring points zero error which impact the measurement result are discussed and compared with L-MM method in inverse problems. Instance calculation and analysis prove that the method applied in this paper still has good effectiveness and accuracy even if measurement error exists and the boundary measurement points’ number is reduced. The comparison indicates that the influence of error on the inversion solution can be minimized effectively using this method.

  2. Ultrafast Photoinduced Electron Transfer in a π-Conjugated Oligomer/Porphyrin Complex

    KAUST Repository

    Aly, Shawkat Mohammede; Goswami, Subhadip; Alsulami, Qana; Schanze, Kirk S.; Mohammed, Omar F.

    2014-01-01

    Controlling charge transfer (CT), charge separation (CS), and charge recombination (CR) at the donor-acceptor interface is extremely important to optimize the conversion efficiency in solar cell devices. In general, ultrafast CT and slow CR

  3. Coupled sensitizer-catalyst dyads: electron-transfer reactions in a perylene-polyoxometalate conjugate.

    Science.gov (United States)

    Odobel, Fabrice; Séverac, Marjorie; Pellegrin, Yann; Blart, Errol; Fosse, Céline; Cannizzo, Caroline; Mayer, Cédric R; Elliott, Kristopher J; Harriman, Anthony

    2009-01-01

    Ultrafast discharge of a single-electron capacitor: A variety of intramolecular electron-transfer reactions are apparent for polyoxometalates functionalized with covalently attached perylene monoimide chromophores, but these are restricted to single-electron events. (et=electron transfer, cr=charge recombination, csr=charge-shift reaction, PER=perylene, POM=polyoxometalate).A new strategy is introduced that permits covalent attachment of an organic chromophore to a polyoxometalate (POM) cluster. Two examples are reported that differ according to the nature of the anchoring group and the flexibility of the linker. Both POMs are functionalized with perylene monoimide units, which function as photon collectors and form a relatively long-lived charge-transfer state under illumination. They are reduced to a stable pi-radical anion by electrolysis or to a protonated dianion under photolysis in the presence of aqueous triethanolamine. The presence of the POM opens up an intramolecular electron-transfer route by which the charge-transfer state reduces the POM. The rate of this process depends on the molecular conformation and appears to involve through-space interactions. Prior reduction of the POM leads to efficient fluorescence quenching, again due to intramolecular electron transfer. In most cases, it is difficult to resolve the electron-transfer products because of relatively fast reverse charge shift that occurs within a closed conformer. Although the POM can store multiple electrons, it has not proved possible to use these systems as molecular-scale capacitors because of efficient electron transfer from the one-electron-reduced POM to the excited singlet state of the perylene monoimide.

  4. Identification of novel Clostridium perfringens type E strains that carry an iota toxin plasmid with a functional enterotoxin gene.

    Directory of Open Access Journals (Sweden)

    Kazuaki Miyamoto

    Full Text Available Clostridium perfringens enterotoxin (CPE is a major virulence factor for human gastrointestinal diseases, such as food poisoning and antibiotic associated diarrhea. The CPE-encoding gene (cpe can be chromosomal or plasmid-borne. Recent development of conventional PCR cpe-genotyping assays makes it possible to identify cpe location (chromosomal or plasmid in type A isolates. Initial studies for developing cpe genotyping assays indicated that all cpe-positive strains isolated from sickened patients were typable by cpe-genotypes, but surveys of C. perfringens environmental strains or strains from feces of healthy people suggested that this assay might not be useful for some cpe-carrying type A isolates. In the current study, a pulsed-field gel electrophoresis Southern blot assay showed that four cpe-genotype untypable isolates carried their cpe gene on a plasmid of ∼65 kb. Complete sequence analysis of the ∼65 kb variant cpe-carrying plasmid revealed no intact IS elements and a disrupted cytosine methyltransferase (dcm gene. More importantly, this plasmid contains a conjugative transfer region, a variant cpe gene and variant iota toxin genes. The toxin genes encoded by this plasmid are expressed based upon the results of RT-PCR assays. The ∼65 kb plasmid is closely related to the pCPF4969 cpe plasmid of type A isolates. MLST analyses indicated these isolates belong to a unique cluster of C. perfringens. Overall, these isolates carrying a variant functional cpe gene and iota toxin genes represent unique type E strains.

  5. Dealing with the evolutionary downside of CRISPR immunity: bacteria and beneficial plasmids.

    Directory of Open Access Journals (Sweden)

    Wenyan Jiang

    Full Text Available The immune systems that protect organisms from infectious agents invariably have a cost for the host. In bacteria and archaea CRISPR-Cas loci can serve as adaptive immune systems that protect these microbes from infectiously transmitted DNAs. When those DNAs are borne by lytic viruses (phages, this protection can provide a considerable advantage. CRISPR-Cas immunity can also prevent cells from acquiring plasmids and free DNA bearing genes that increase their fitness. Here, we use a combination of experiments and mathematical-computer simulation models to explore this downside of CRISPR-Cas immunity and its implications for the maintenance of CRISPR-Cas loci in microbial populations. We analyzed the conjugational transfer of the staphylococcal plasmid pG0400 into Staphylococcus epidermidis RP62a recipients that bear a CRISPR-Cas locus targeting this plasmid. Contrary to what is anticipated for lytic phages, which evade CRISPR by mutations in the target region, the evasion of CRISPR immunity by plasmids occurs at the level of the host through loss of functional CRISPR-Cas immunity. The results of our experiments and models indicate that more than 10(-4 of the cells in CRISPR-Cas positive populations are defective or deleted for the CRISPR-Cas region and thereby able to receive and carry the plasmid. Most intriguingly, the loss of CRISPR function even by large deletions can have little or no fitness cost in vitro. These theoretical and experimental results can account for the considerable variation in the existence, number and function of CRISPR-Cas loci within and between bacterial species. We postulate that as a consequence of the opposing positive and negative selection for immunity, CRISPR-Cas systems are in a continuous state of flux. They are lost when they bear immunity to laterally transferred beneficial genes, re-acquired by horizontal gene transfer, and ascend in environments where phage are a major source of mortality.

  6. Small RNA-based feedforward loop with AND-gate logic regulates extrachromosomal DNA transfer in Salmonella.

    Science.gov (United States)

    Papenfort, Kai; Espinosa, Elena; Casadesús, Josep; Vogel, Jörg

    2015-08-25

    Horizontal gene transfer via plasmid conjugation is a major driving force in microbial evolution but constitutes a complex process that requires synchronization with the physiological state of the host bacteria. Although several host transcription factors are known to regulate plasmid-borne transfer genes, RNA-based regulatory circuits for host-plasmid communication remain unknown. We describe a posttranscriptional mechanism whereby the Hfq-dependent small RNA, RprA, inhibits transfer of pSLT, the virulence plasmid of Salmonella enterica. RprA employs two separate seed-pairing domains to activate the mRNAs of both the sigma-factor σ(S) and the RicI protein, a previously uncharacterized membrane protein here shown to inhibit conjugation. Transcription of ricI requires σ(S) and, together, RprA and σ(S) orchestrate a coherent feedforward loop with AND-gate logic to tightly control the activation of RicI synthesis. RicI interacts with the conjugation apparatus protein TraV and limits plasmid transfer under membrane-damaging conditions. To our knowledge, this study reports the first small RNA-controlled feedforward loop relying on posttranscriptional activation of two independent targets and an unexpected role of the conserved RprA small RNA in controlling extrachromosomal DNA transfer.

  7. Conjugate heat transfer in a porous cavity filled with nano-fluids and heated by a triangular thick wall

    International Nuclear Information System (INIS)

    Chamkha, Ali J.; Ismael, Muneer A.

    2013-01-01

    The conjugate natural convection-conduction heat transfer in a square domain composed of nano-fluids filled porous cavity heated by a triangular solid wall is studied under steady-state conditions. The vertical and horizontal walls of the triangular solid wall are kept isothermal and at the same hot temperature Th. The other boundaries surrounding the porous cavity are kept adiabatic except the right vertical wall where it is kept isothermally at the lower temperature T c . Equations governing the heat transfer in the triangular wall and heat and nano-fluid flow, based on the Darcy model, in the nano-fluid-saturated porous medium together with the derived relation of the interface temperature are solved numerically using the over-successive relaxation finite-difference method. A temperature independent nano-fluids properties model is adopted. Three nano-particle types dispersed in one base fluid (water) are investigated. The investigated parameters are the nano-particles volume fraction φ (0-0.2), Rayleigh number Ra (10-1000), solid wall to base-fluid saturated porous medium thermal conductivity ratio K ro (0.44, 1, 23.8), and the triangular wall thickness D (0.1-1). The results are presented in the conventional form; contours of streamlines and isotherms and the local and average Nusselt numbers. At a very low Rayleigh number Ra = 10, a significant enhancement in heat transfer within the porous cavity with φ is observed. Otherwise, the heat transfer may be enhanced or deteriorated with φ depending on the wall thickness D and the Rayleigh number Ra. At high Rayleigh numbers and low conductivity ratios, critical values of D, regardless of 4, are observed and accounted. (authors)

  8. Conjugal transfer of aac(6')Ie-aph(2″)Ia gene from native species and mechanism of regulation and cross resistance in Enterococcus faecalis MCC3063 by real time-PCR.

    Science.gov (United States)

    Jaimee, G; Halami, P M

    2017-09-01

    High level aminoglycoside resistance (HLAR) in the lactic acid bacteria (LAB) derived from food animals is detrimental. The aim of this study was to investigate the localization and conjugal transfer of aminoglycoside resistance genes, aac(6')Ie-aph(2″)Ia and aph(3')IIIa in different Enterococcus species. The cross resistance patterns in Enterococcus faecalis MCC3063 to clinically important aminoglycosides by real time PCR were also studied. Southern hybridization experiments revealed the presence of aac(6')Ie-aph(2 ″ )Ia and aph(3')IIIa genes conferring HLAR in high molecular weight plasmids except in Lactobacillus plantarum. The plasmid encoded bifunctional aac(6')Ie-aph(2″)Ia gene was transferable from Enterococcus avium (n = 2), E. cecorum (n = 1), E. faecalis (n = 1) and Pediococcus lolii (n = 1) species into the recipient strain; E. faecalis JH2-2 by filter mating experiments thus indicating the possible risks of gene transfer into pathogenic strains. Molecular analysis of cross resistance patterns in native isolate of E. faecalis MCC3063 carrying aac(6')Ie-aph(2″)Ia and aph(3')IIIa gene was displayed by quantification of the mRNA levels in this study. For this, the culture was induced with increasing concentrations of gentamicin, kanamycin and streptomycin (2048, 4096, 8192, 16384 μg/mL) individually. The increasing concentrations of gentamicin and kanamycin induced the expression of the aac(6')Ie-aph(2″)Ia and aph(3')IIIa resistance genes, respectively. Interestingly, it was observed that induction with streptomycin triggered a significant fold increase in the expression of the aph(3')IIIa gene which otherwise was not known to modify the aminoglycoside. This is noteworthy as streptomycin was found to confer cross resistance to structurally unrelated kanamycin. Also, expression of the aph(3')IIIa gene when induced with streptomycin, revealed that bacteria harbouring this gene will be able to overcome streptomycin bactericidal action at

  9. An approximate block Newton method for coupled iterations of nonlinear solvers: Theory and conjugate heat transfer applications

    Science.gov (United States)

    Yeckel, Andrew; Lun, Lisa; Derby, Jeffrey J.

    2009-12-01

    A new, approximate block Newton (ABN) method is derived and tested for the coupled solution of nonlinear models, each of which is treated as a modular, black box. Such an approach is motivated by a desire to maintain software flexibility without sacrificing solution efficiency or robustness. Though block Newton methods of similar type have been proposed and studied, we present a unique derivation and use it to sort out some of the more confusing points in the literature. In particular, we show that our ABN method behaves like a Newton iteration preconditioned by an inexact Newton solver derived from subproblem Jacobians. The method is demonstrated on several conjugate heat transfer problems modeled after melt crystal growth processes. These problems are represented by partitioned spatial regions, each modeled by independent heat transfer codes and linked by temperature and flux matching conditions at the boundaries common to the partitions. Whereas a typical block Gauss-Seidel iteration fails about half the time for the model problem, quadratic convergence is achieved by the ABN method under all conditions studied here. Additional performance advantages over existing methods are demonstrated and discussed.

  10. Color optimization of conjugated-polymer/InGaN hybrid white light emitting diodes by incomplete energy transfer

    International Nuclear Information System (INIS)

    Chang, Chi-Jung; Lai, Chun-Feng; Madhusudhana Reddy, P.; Chen, Yung-Lin; Chiou, Wei-Yung; Chang, Shinn-Jen

    2015-01-01

    By using the wavelength conversion method, white light emitting diodes (WLEDs) were produced by applying mixtures of polysiloxane and fluorescent polymers on InGaN based light emitting diodes. UV curable organic–inorganic hybrid materials with high refractive index (1.561), compromised optical, thermal and mechanical properties was used as encapsulants. Red light emitting fluorescent FABD polymer (with 9,9-dioctylfluorene (F), anthracene (A) and 2,1,3-benzothiadiazole (B), and 4,7-bis(2-thienyl)-2,1,3-benzothiadiazole (D) repeating units) and green light emitting fluorescent FAB polymer were used as wavelength converters. The encapsulant/fluorescent polymer mixture and InGaN produce the white light by incomplete energy transfer mechanism. WLEDs with high color rendering index (CRI, about 93), and tunable correlated color temperature (CCT) properties can be produced by controlling the composition and chemical structures of encapsulating polymer and fluorescent polymer in hybrid materials, offering cool-white and neutral-white LEDs. - Highlights: • Highly efficient white light-emitting diodes (WLEDs) were produced. • Conjugated-polymer/InGaN hybrid WLEDs by incomplete energy transfer mechanism. • WLEDs with high color-rendering index and tunable correlated color temperature. • Polysiloxane encapsulant with superior optical, mechanical and thermal properties

  11. Photoinduced electron-transfer from pi-conjugated polymers onto buckminsterfullerene, fulleroids, and methanofullerenes

    NARCIS (Netherlands)

    Janssen, R. A. J.; Hummelen, J. C.; Lee, Kwanghee; Pakbaz, K.; Sariciftci, N. S.; Heeger, A. J.; Wudl, F

    1995-01-01

    We present near-steady-state photoinduced absorption (PIA), photoluminescence, and light-induced electron spin resonance (LESR) studies on photoinduced electron transfer reactions from poly(bis-2,5-epi-cholestanoxy-1,4-phenylene vinylene) (BeCHA-PPV) as a donor to Buckminsterfullerene (C60) and a

  12. Fully-developed conjugate heat transfer in porous media with uniform heating

    NARCIS (Netherlands)

    Lopez Penha, D.J.; Stolz, S.; Kuerten, Johannes G.M.; Nordlund, M.; Kuczaj, Arkadiusz K.; Geurts, Bernardus J.

    2012-01-01

    We propose a computational method for approximating the heat transfer coefficient of fully-developed flow in porous media. For a representative elementary volume of the porous medium we develop a transport model subject to periodic boundary conditions that describes incompressible fluid flow through

  13. Charge-transfer mobility and electrical conductivity of PANI as conjugated organic semiconductors.

    Science.gov (United States)

    Zhang, Yahong; Duan, Yuping; Song, Lulu; Zheng, Daoyuan; Zhang, Mingxing; Zhao, Guangjiu

    2017-09-21

    The intramolecular charge transfer properties of a phenyl-end-capped aniline tetramer (ANIH) and a chloro-substituted derivative (ANICl) as organic semiconductors were theoretically studied through the first-principles calculation based on the Marcus-Hush theory. The reorganization energies, intermolecular electronic couplings, angular resolution anisotropic mobilities, and density of states of the two crystals were evaluated. The calculated results demonstrate that both ANIH and ANICl crystals show the higher electron transfer mobilities than the hole-transfer mobilities, which means that the two crystals should prefer to function as n-type organic semiconductors. Furthermore, the angle dependence mobilities of the two crystals show remarkable anisotropic character. The maximum mobility μ max of ANIH and ANICl crystals is 1.3893 and 0.0272 cm 2 V -1 s -1 , which appear at the orientation angles near 176°/356° and 119°/299° of a conducting channel on the a-b reference plane. It is synthetically evaluated that the ANIH crystal possesses relatively lower reorganization energy, higher electronic coupling, and electron transfer mobility, which means that the ANIH crystal may be the more ideal candidate as a high performance n-type organic semiconductor material. The systematic theoretical studies on organic crystals should be conducive to evaluating the charge-transport properties and designing higher performance organic semiconductor materials.

  14. Evaluation of conjugate, radial heat transfer in an internally insulated composite pipe

    NARCIS (Netherlands)

    Reurings, C.; Koussios, S.; Bergsma, O.K.; Vergote, K.

    2015-01-01

    In order to compete with steel, a fibre-reinforced composite exhaust wall with a general-purpose resin system requires an effective but lightweight insulation layer. However a lack of experimental methods for heat transfer from turbulent gas flow to pipe walls lined with a porous insulation layer

  15. Permissiveness of soil microbial communities towards broad host range plasmids

    DEFF Research Database (Denmark)

    Klümper, Uli

    . Plasmids are implicated in the rapid spread of antibiotic resistance and the emergence of multi-resistant pathogenic bacteria, making it crucial to be able to quantify, understand, and, ideally, control plasmid transfer in mixed microbial communities. The fate of plasmids in microbial communities...... of microbial communities may be directly interconnected through transfer of BHR plasmids at a so far unrecognized level. The developed method furthermore enabled me to explore how agronomic practices may affect gene transfer in soil microbial communities. I compared bacterial communities extracted from plots...

  16. On the application of Chimera/unstructured hybrid grids for conjugate heat transfer

    Science.gov (United States)

    Kao, Kai-Hsiung; Liou, Meng-Sing

    1995-01-01

    A hybrid grid system that combines the Chimera overset grid scheme and an unstructured grid method is developed to study fluid flow and heat transfer problems. With the proposed method, the solid structural region, in which only the heat conduction is considered, can be easily represented using an unstructured grid method. As for the fluid flow region external to the solid material, the Chimera overset grid scheme has been shown to be very flexible and efficient in resolving complex configurations. The numerical analyses require the flow field solution and material thermal response to be obtained simultaneously. A continuous transfer of temperature and heat flux is specified at the interface, which connects the solid structure and the fluid flow as an integral system. Numerical results are compared with analytical and experimental data for a flat plate and a C3X cooled turbine cascade. A simplified drum-disk system is also simulated to show the effectiveness of this hybrid grid system.

  17. Two-Dimensional Variable Property Conjugate Heat Transfer Simulation of Nanofluids in Microchannels

    International Nuclear Information System (INIS)

    Ramiar, A.; Ranjbar, A.A.

    2013-01-01

    Laminar two-dimensional forced convective heat transfer of CuO-water and Al 2 O 3 -water nanofluids in a horizontal microchannel has been studied numerically, considering axial conduction effects in both solid and liquid regions and variable thermal conductivity and dynamic viscosity. The results show that using nanoparticles with higher thermal conductivities will intensify enhancement of heat transfer characteristics and slightly increases shear stress on the wall. The obtained results show more steep changes in Nusselt number for lower diameters and also higher values of Nusselt number by decreasing the diameter of nanoparticles. Also, by utilizing conduction number as the criterion, it was concluded from the results that adding nanoparticles will intensify the axial conduction effect in the geometry considered.

  18. Imidazole as a parent π-conjugated backbone in charge-transfer chromophores

    Directory of Open Access Journals (Sweden)

    Jiří Kulhánek

    2012-01-01

    Full Text Available Research activities in the field of imidazole-derived push–pull systems featuring intramolecular charge transfer (ICT are reviewed. Design, synthetic pathways, linear and nonlinear optical properties, electrochemistry, structure–property relationships, and the prospective application of such D-π-A organic materials are described. This review focuses on Y-shaped imidazoles, bi- and diimidazoles, benzimidazoles, bis(benzimidazoles, imidazole-4,5-dicarbonitriles, and imidazole-derived chromophores chemically bound to a polymer chain.

  19. Conjugate heat transfer analysis of an energy conversion device with an updated numerical model obtained through inverse identification

    International Nuclear Information System (INIS)

    Hey, Jonathan; Malloy, Adam C.; Martinez-Botas, Ricardo; Lamperth, Michael

    2015-01-01

    Highlights: • Conjugate heat transfer analysis of an electric machine. • Inverse identification method for estimating the model parameters. • Experimentally determined thermal properties and electromagnetic losses. • Coupling of inverse identification method with a numerical model. • Improved modeling accuracy through introduction of interface material. - Abstract: Energy conversion devices undergo thermal loading during their operation as a result of inefficiencies in the energy conversion process. This will eventually lead to degradation and possible failure of the device if the heat generated is not properly managed. The ability to accurately predict the thermal behavior of such a device during the initial developmental stage is an important requirement. However, accurate predictions of critical temperature is challenging due to the variation of heat transfer parameters from one device to another. The ability to determine the model parameters is key to accurately representing the heat transfer in such a device. This paper presents the use of an inverse identification technique to estimate the model parameters of an energy conversion device designed for vehicular applications. To simulate the imperfect contact and the presence of insulating materials in the permanent magnet electric machine, thin material are introduced at the component interface of the numerical model. The proposed inverse identification method is used to estimate the equivalent thermal conductance of the thin material. In addition, the electromagnetic losses generated in the permanent magnet is also derived indirectly from the temperature measurement using the same method. With the thermal properties and input parameters of the numerical model obtained from the inverse identification method, the critical temperature of the device can be predicted more accurately. The deviation between the maximum measured and predicted winding temperature is less than 2.4%

  20. Experimental study of conjugate heat transfer from liquid metal layer cooled by overlying freon

    International Nuclear Information System (INIS)

    Cho, J.S.; Suh, K.Y.; Chung, C.H.; Park, R.J.; Kim, S.B.

    2001-01-01

    Steady-state and transient experiments were performed for the heat transfer from the liquid metal pool with overlying Freon (R113) coolant in the process of boiling. The simulant molten pool material is tin (Sn) with the melting temperature of 232 Celsius degrees. The metal pool is heated from the bottom surface and the coolant is injected onto the molten metal pool. Tests were conducted under the condition of the bottom surface heating in the test section and the forced convection of the R113 coolant being injected onto the molten metal pool. The bottom heating condition was varied from 8 kW to 14 kW. The temperature distributions of the metal layer and coolant were obtained in the steady-state experiment. The boiling mechanism of the R113 coolant was changed from the nucleate boiling to film boiling in the transient experiment. The critical heat flux (CHF) phenomenon was observed during the transition from the nucleate boiling to the film boiling. Also, the Nusselt (Nu) number and the Rayleigh (Ra) number in the molten metal pool region were obtained as functions of time. Analysis was done for the relationship between the heat flux and the temperature difference between the metal layer surface and the boiling coolant. In this experiment, the heat transfer is achieved with accompanying solidification in the molten metal pool by the boiling R113 coolant there above. The present test results of the natural convection heat transfer on the molten metal pool are higher than those of the liquid metal natural convection heat transfer without coolant boiling. It can be interpreted that the heat transfer rate is enhanced by the overlying boiling coolant having the high heat removal rate. Analysis of the relationship between the heat flux and the difference between the metal layer surface temperature and the coolant bulk boiling temperature revealed that the CHF occurs when the temperature difference reaches a neighborhood of 50 Celsius degrees. Also, if the temperature

  1. Isolation of novel IncA/C and IncN fluoroquinolone resistance plasmids from an antibiotic-polluted lake.

    Science.gov (United States)

    Flach, Carl-Fredrik; Johnning, Anna; Nilsson, Ida; Smalla, Kornelia; Kristiansson, Erik; Larsson, D G Joakim

    2015-10-01

    Antibiotic-polluted environments may function as reservoirs for novel resistance plasmids not yet encountered in pathogens. The aims of this study were to assess the potential of resistance transfer between bacteria from such environments and Escherichia coli, and to characterize the conjugative elements involved. Sediment samples from Kazipally lake and Asanikunta tank, two Indian lakes with a history of severe pollution with fluoroquinolones, were investigated. Proportions of resistant bacteria were determined by selective cultivation, while horizontal gene transfer was studied using a GFP-tagged E. coli as recipient. Retrieved transconjugants were tested for susceptibility by Etest(®) and captured conjugative resistance elements were characterized by WGS. The polluted lakes harboured considerably higher proportions of ciprofloxacin-resistant and sulfamethoxazole-resistant bacteria than did other Indian and Swedish lakes included for comparison (52% versus 2% and 60% versus 7%, respectively). Resistance plasmids were captured from Kazipally lake, but not from any of the other lakes; in the case of Asanikunta tank because of high sediment toxicity. Eight unique IncA/C and IncN resistance plasmids were identified among 11 sequenced transconjugants. Five plasmids were fully assembled, and four of these carried the quinolone resistance gene qnrVC1, which has previously only been found on chromosomes. Acquired resistance genes, in the majority of cases associated with class 1 integrons, could be linked to decreased susceptibility to several different classes of antibiotics. Our study shows that environments heavily polluted with antibiotics contain novel multiresistance plasmids transferrable to E. coli. © The Author 2015. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  2. Incidence of plasmid-linked antibiotic-heavy metal resistant enterics in water-sediment from agricultural and harbor sites

    Energy Technology Data Exchange (ETDEWEB)

    Mietz, J.A.; Sjorgren, R.E.

    1983-01-01

    This study assesses the impact of antibiotics used in clinical and veterinary practices on the incidence of antibiotic-heavy metal resistant enterics in fresh water and sediment from agricultural and harbor sample sites. A total of 848 bacterial strains of the family Enterobacteriaceae was isolated from agricultural and lake harbor samples. These were examined for anitbiotic-heavy metal resistance. A select smaller number of these isolates were also examined for the presence of plasmids and ability to transfer antibiotic resistance via conjugation or transformation. More than 85% of the 848 isolates from all four sites were resistant to Pb, Zn, and Co while 5.6% to 16% were resistant to Te and 2.4% to 5.7% to Hg. Of the total isolates tested, 87% were resistant to six or more antibiotics and 74% were also simultaneously resistant to Co, Zn, and Pb. Testing the resistance of the water isolates to antibiotics used solely in animal husbandry-veterinary medicine indicated that 55.6% of the agricultural isolates possessed resistance to these antibiotics while only 31.9% of the isolates from harbor water showed resistance to the same antibiotics. Of 41 ampicillin resistant isolates examined, 16 (39%) were capable of transferring antibiotic-heavy resistance markers via conjugation. From this same group, plasmid DNA preparations were made. Of these latter preparations, 67% transformed recipient E. coli cells while 58% possessed discernible, often multiple plasmids when examined by gel electrophoresis.

  3. Conjugate heat transfer investigation on the cooling performance of air cooled turbine blade with thermal barrier coating

    Science.gov (United States)

    Ji, Yongbin; Ma, Chao; Ge, Bing; Zang, Shusheng

    2016-08-01

    A hot wind tunnel of annular cascade test rig is established for measuring temperature distribution on a real gas turbine blade surface with infrared camera. Besides, conjugate heat transfer numerical simulation is performed to obtain cooling efficiency distribution on both blade substrate surface and coating surface for comparison. The effect of thermal barrier coating on the overall cooling performance for blades is compared under varied mass flow rate of coolant, and spatial difference is also discussed. Results indicate that the cooling efficiency in the leading edge and trailing edge areas of the blade is the lowest. The cooling performance is not only influenced by the internal cooling structures layout inside the blade but also by the flow condition of the mainstream in the external cascade path. Thermal barrier effects of the coating vary at different regions of the blade surface, where higher internal cooling performance exists, more effective the thermal barrier will be, which means the thermal protection effect of coatings is remarkable in these regions. At the designed mass flow ratio condition, the cooling efficiency on the pressure side varies by 0.13 for the coating surface and substrate surface, while this value is 0.09 on the suction side.

  4. A sensitivity function-based conjugate gradient method for optical tomography with the frequency-domain equation of radiative transfer

    International Nuclear Information System (INIS)

    Kim, Hyun Keol; Charette, Andre

    2007-01-01

    The Sensitivity Function-based Conjugate Gradient Method (SFCGM) is described. This method is used to solve the inverse problems of function estimation, such as the local maps of absorption and scattering coefficients, as applied to optical tomography for biomedical imaging. A highly scattering, absorbing, non-reflecting, non-emitting medium is considered here and simultaneous reconstructions of absorption and scattering coefficients inside the test medium are achieved with the proposed optimization technique, by using the exit intensity measured at boundary surfaces. The forward problem is solved with a discrete-ordinates finite-difference method on the framework of the frequency-domain full equation of radiative transfer. The modulation frequency is set to 600 MHz and the frequency data, obtained with the source modulation, is used as the input data. The inversion results demonstrate that the SFCGM can retrieve simultaneously the spatial distributions of optical properties inside the medium within a reasonable accuracy, by significantly reducing a cross-talk between inter-parameters. It is also observed that the closer-to-detector objects are better retrieved

  5. High Prevalence of Plasmid-Mediated Quinolone Resistance and IncQ Plasmids Carrying qnrS2 Gene in Bacteria from Rivers near Hospitals and Aquaculture in China.

    Directory of Open Access Journals (Sweden)

    Yanping Wen

    Full Text Available Effluents from hospital and aquaculture are considered important sources of quinolone resistance. However, little information is available on the impact of this effluent on nearby rivers. In this study, 188 ciprofloxacin-resistant bacterial isolates obtained from rivers near hospitals and aquaculture were screened for plasmid-mediated quinolone resistance (PMQR genes. Species identification, antibiotic susceptibility testing, and PMQR gene transferability assessment were conducted for PMQR-positive bacteria. Representative qnrS2-encoding plasmids were subsequently sequenced using a primer-walking approach. In total, 44 isolates (23.4% were positive for qnr genes (16 qnrB2, 3 qnrS1, and 25 qnrS2 and 32 isolates (17.0% were positive for aac(6'-Ib-cr. Other PMQR genes were not detected. The qnrB2 and aac(6'-Ib-cr genes had a higher prevalence in aquaculture samples than in hospital samples, and were significantly associated with Enterobacteriaceae (p < 0.05. In contrast, the prevalence of qnrS2 was not site-related, but was significantly associated with Aeromonas spp. (p < 0.05. All PMQR isolates were resistant to three or more classes of antibiotics. Eleven qnrS2-harboring plasmids from Aeromonas spp., including a novel conjugative plasmid pHP18, were selected for sequencing. These plasmids were small in size (6,388-16,197 bp and belonged to the IncQ or IncU plasmid family, with qnrS2 being part of a mobile insertion cassette. Taken together, our findings suggest that aquaculture is a possible source for aac(6'-Ib-cr and qnrB2 dissemination, and demonstrate the ubiquity of qnrS2 in aquatic environments. Finally, Aeromonas spp. served as vectors for qnrS2 with the help of IncQ-type plasmids.

  6. Plasmid segregation mechanisms

    DEFF Research Database (Denmark)

    Ebersbach, G.; Gerdes, Kenn

    2005-01-01

    Bacterial plasmids encode partitioning (par) loci that ensure ordered plasmid segregation prior to cell division. par loci come in two types: those that encode actin-like ATPases and those that encode deviant Walker-type ATPases. ParM, the actin-like ATPase of plasmid R1, forms dynamic filaments...... that segregate plasmids paired at mid-cell to daughter cells. Like microtubules, ParM filaments exhibit dynamic instability (i.e., catastrophic decay) whose regulation is an important component of the DNA segregation process. The Walker box ParA ATPases are related to MinD and form highly dynamic, oscillating...... filaments that are required for the subcellular movement and positioning of plasmids. The role of the observed ATPase oscillation is not yet understood. However, we propose a simple model that couples plasmid segregation to ParA oscillation. The model is consistent with the observed movement...

  7. Ultrafast Photoinduced Electron Transfer in a π-Conjugated Oligomer/Porphyrin Complex

    KAUST Repository

    Aly, Shawkat Mohammede

    2014-10-02

    Controlling charge transfer (CT), charge separation (CS), and charge recombination (CR) at the donor-acceptor interface is extremely important to optimize the conversion efficiency in solar cell devices. In general, ultrafast CT and slow CR are desirable for optimal device performance. In this Letter, the ultrafast excited-state CT between platinum oligomer (DPP-Pt(acac)) as a new electron donor and porphyrin as an electron acceptor is monitored for the first time using femtosecond (fs) transient absorption (TA) spectroscopy with broad-band capability and 120 fs temporal resolution. Turning the CT on/off has been shown to be possible either by switching from an organometallic oligomer to a metal-free oligomer or by controlling the charge density on the nitrogen atom of the porphyrin meso unit. Our time-resolved data show that the CT and CS between DPP-Pt(acac) and cationic porphyrin are ultrafast (approximately 1.5 ps), and the CR is slow (ns time scale), as inferred from the formation and the decay of the cationic and anionic species. We also found that the metallic center in the DPP-Pt(acac) oligomer and the positive charge on the porphyrin are the keys to switching on/off the ultrafast CT process.

  8. Why close a bacterial genome? The plasmid of Alteromonas macleodii HOT1A3 is a vector for inter-specific transfer of a flexible genomic island

    Directory of Open Access Journals (Sweden)

    Eduard eFadeev

    2016-03-01

    Full Text Available Genome sequencing is rapidly becoming a staple technique in environmental and clinical microbiology, yet computational challenges still remain, leading to many draft genomes which are typically fragmented into many contigs. We sequenced and completely assembled the genome of a marine heterotrophic bacterium, Alteromonas macleodii HOT1A3, and compared its full genome to several draft genomes obtained using different reference-based and de-novo methods. In general, the de-novo assemblies clearly outperformed the reference-based or hybrid ones, covering>99% of the genes and representing essentially all of the gene functions. However, only the fully closed genome (~4.5Mbp allowed us to identify the presence of a large, 148 kbp plasmid, pAM1A3. While HOT1A3 belongs to Alteromonas macleodii, typically found in surface waters (surface ecotype, this plasmid consists of an almost complete flexible genomic island, containing many genes involved in metal resistance previously identified in the genomes of Alteromonas mediterranea (deep ecotype. Indeed, similar to A. mediterranea, A. macleodii HOT1A3 grows at concentrations of zinc, mercury and copper that are inhibitory for other A. macleodii strains. The presence of a plasmid encoding almost an entire flexible genomic island suggests that wholesale genomic exchange between heterotrophic marine bacteria belonging to related but ecologically different populations is not uncommon.

  9. Microarray-based analysis of IncA/C plasmid-associated genes from multidrug-resistant Salmonella enterica.

    Science.gov (United States)

    Lindsey, Rebecca L; Frye, Jonathan G; Fedorka-Cray, Paula J; Meinersmann, Richard J

    2011-10-01

    In the family Enterobacteriaceae, plasmids have been classified according to 27 incompatibility (Inc) or replicon types that are based on the inability of different plasmids with the same replication mechanism to coexist in the same cell. Certain replicon types such as IncA/C are associated with multidrug resistance (MDR). We developed a microarray that contains 286 unique 70-mer oligonucleotide probes based on sequences from five IncA/C plasmids: pYR1 (Yersinia ruckeri), pPIP1202 (Yersinia pestis), pP99-018 (Photobacterium damselae), pSN254 (Salmonella enterica serovar Newport), and pP91278 (Photobacterium damselae). DNA from 59 Salmonella enterica isolates was hybridized to the microarray and analyzed for the presence or absence of genes. These isolates represented 17 serovars from 14 different animal hosts and from different geographical regions in the United States. Qualitative cluster analysis was performed using CLUSTER 3.0 to group microarray hybridization results. We found that IncA/C plasmids occurred in two lineages distinguished by a major insertion-deletion (indel) region that contains genes encoding mostly hypothetical proteins. The most variable genes were represented by transposon-associated genes as well as four antimicrobial resistance genes (aphA, merP, merA, and aadA). Sixteen mercury resistance genes were identified and highly conserved, suggesting that mercury ion-related exposure is a stronger pressure than anticipated. We used these data to construct a core IncA/C genome and an accessory genome. The results of our studies suggest that the transfer of antimicrobial resistance determinants by transfer of IncA/C plasmids is somewhat less common than exchange within the plasmids orchestrated by transposable elements, such as transposons, integrating and conjugative elements (ICEs), and insertion sequence common regions (ISCRs), and thus pose less opportunity for exchange of antimicrobial resistance.

  10. Integration of replication-defective R68.45-like plasmids into the Pseudomonas aeruginosa chromosome.

    Science.gov (United States)

    Reimmann, C; Rella, M; Haas, D

    1988-06-01

    R68.45 and other similar broad-host-range (IncP) plasmids carrying a tandem repeat of the 2.1 kb insertion element IS21 mobilize the chromosome of many different Gram-negative bacteria. To analyse the structure of R68.45-chromosome cointegrates, whose involvement in the mobilization process had been postulated previously, we selected for the stable integration of R68.45-like plasmids into the Pseudomonas aeruginosa chromosome. Two plasmids were chosen: pME28, a transfer-deficient, mobilizable RP1 derivative with an inactive replication control (trfA) gene, and pME487, an R68.45 derivative with a trfA(ts) mutation causing temperature-sensitive replication. Chromosomally integrated pME28 and pME487 were found to be flanked by single IS21 elements. This structure is in agreement with a 'cut-and-paste' mode of R68.45 transposition. pME28 and pME487 showed a low specificity of insertion but rarely (less than 0.1%) induced auxotrophic mutations. Hfr (high-frequency-of-recombination) donors of P. aeruginosa could be obtained by chromosomal integration of pME487 or pME28; in the latter case, the transfer functions lacking from pME28 had to be provided in trans on an autonomous plasmid. Hfr donors gave higher conjugational linkage and transferred longer stretches of the P. aeruginosa chromosome than did R68.45 donors. This suggests that the integration of R68.45 into the donor chromosome is short-lived in P. aeruginosa.

  11. Photoinduced electron transfer between Fe(III) and adenosine triphosphate-BODIPY conjugates: Application to alkaline-phosphatase-linked immunoassay.

    Science.gov (United States)

    Lin, Jia-Hui; Yang, Ya-Chun; Shih, Ya-Chen; Hung, Szu-Ying; Lu, Chi-Yu; Tseng, Wei-Lung

    2016-03-15

    Fluorescent boron dipyrromethene (BODIPY) analogs are often used as sensors for detecting various species because of their relatively high extinction coefficients, outstanding fluorescence quantum yields, photostability, and pH-independent fluorescence. However, there is little-to-no information in the literature that describes the use of BODIPY analogs for detecting alkaline phosphatase (ALP) activity and inhibition. This study discovered that the fluorescence of BODIPY-conjugated adenosine triphosphate (BODIPY-ATP) was quenched by Fe(III) ions through photoinduced electron transfer. The ALP-catalyzed hydrolysis of BODIPY-ATP resulted in the formation of BODIPY-adenosine and phosphate ions. The fluorescence of the generated BODIPY-adenosine was insensitive to the change in the concentration of Fe(III) ions. Thus, the Fe(III)-induced fluorescence quenching of BODIPY-ATP can be paired with its ALP-mediated dephosphorylation to design a turn-on fluorescence probe for ALP sensing. A method detection limit at a signal-to-noise ratio of 3 for ALP was estimated to be 0.02 units/L (~6 pM; 1 ng/mL). This probe was used for the screening of ALP inhibitors, including Na3VO4, imidazole, and arginine. Because ALP is widely used in enzyme-linked immunosorbent assays, the probe was coupled to an ALP-linked immunosorbent assay for the sensitive and selective detection of immunoglobulin G (IgG). The lowest detectable concentration for IgG in this system was 5 ng/mL. Compared with the use of 3,6-fluorescein diphosphate as a signal reporter in an ALP-linked immunosorbent assay, the proposed system provided comparable sensitivity, large linear range, and high stability over temperature and pH changes. Copyright © 2015 Elsevier B.V. All rights reserved.

  12. Acquisition through Horizontal Gene Transfer of Plasmid pSMA198 by Streptococcus macedonicus ACA-DC 198 Points towards the Dairy Origin of the Species

    Science.gov (United States)

    Papadimitriou, Konstantinos; Anastasiou, Rania; Maistrou, Eleni; Plakas, Thomas; Papandreou, Nikos C.; Hamodrakas, Stavros J.; Ferreira, Stéphanie; Supply, Philip; Renault, Pierre; Pot, Bruno; Tsakalidou, Effie

    2015-01-01

    Background Streptococcus macedonicus is an intriguing streptococcal species whose most frequent source of isolation is fermented foods similarly to Streptococcus thermophilus. However, S. macedonicus is closely related to commensal opportunistic pathogens of the Streptococcus bovis/Streptococcus equinus complex. Methodology/Principal Findings We analyzed the pSMA198 plasmid isolated from the dairy strain Streptococcus macedonicus ACA-DC 198 in order to provide novel clues about the main ecological niche of this bacterium. pSMA198 belongs to the narrow host range pCI305/pWV02 family found primarily in lactococci and to the best of our knowledge it is the first such plasmid to be reported in streptococci. Comparative analysis of the pSMA198 sequence revealed a high degree of similarity with plasmids isolated from Lactococcus lactis strains deriving from milk or its products. Phylogenetic analysis of the pSMA198 Rep showed that the vast majority of closely related proteins derive from lactococcal dairy isolates. Additionally, cloning of the pSMA198 ori in L. lactis revealed a 100% stability of replication over 100 generations. Both pSMA198 and the chromosome of S. macedonicus exhibit a high percentage of potential pseudogenes, indicating that they have co-evolved under the same gene decay processes. We identified chromosomal regions in S. macedonicus that may have originated from pSMA198, also supporting a long co-existence of the two replicons. pSMA198 was also found in divergent biotypes of S. macedonicus and in strains isolated from dispersed geographic locations (e.g. Greece and Switzerland) showing that pSMA198’s acquisition is not a recent event. Conclusions/Significance Here we propose that S. macedonicus acquired plasmid pSMA198 from L. lactis via an ancestral genetic exchange event that took place most probably in milk or dairy products. We provide important evidence that point towards the dairy origin of this species. PMID:25584532

  13. Energy Transfer between Conjugated Colloidal Ga2O3 and CdSe/CdS Core/Shell Nanocrystals for White Light Emitting Applications

    Directory of Open Access Journals (Sweden)

    Paul C. Stanish

    2016-02-01

    Full Text Available Developing solid state materials capable of generating homogeneous white light in an energy efficient and resource-sustainable way is central to the design of new and improved devices for various lighting applications. Most currently-used phosphors depend on strategically important rare earth elements, and rely on a multicomponent approach, which produces sub-optimal quality white light. Here, we report the design and preparation of a colloidal white-light emitting nanocrystal conjugate. This conjugate is obtained by linking colloidal Ga2O3 and II–VI nanocrystals in the solution phase with a short bifunctional organic molecule (thioglycolic acid. The two types of nanocrystals are electronically coupled by Förster resonance energy transfer owing to the short separation between Ga2O3 (energy donor and core/shell CdSe/CdS (energy acceptor nanocrystals, and the spectral overlap between the photoluminescence of the donor and the absorption of the acceptor. Using steady state and time-resolved photoluminescence spectroscopies, we quantified the contribution of the energy transfer to the photoluminescence spectral power distribution and the corresponding chromaticity of this nanocrystal conjugate. Quantitative understanding of this new system allows for tuning of the emission color and the design of quasi-single white light emitting inorganic phosphors without the use of rare-earth elements.

  14. Energy Transfer between Conjugated Colloidal Ga2O3 and CdSe/CdS Core/Shell Nanocrystals for White Light Emitting Applications

    Science.gov (United States)

    Stanish, Paul C.; Radovanovic, Pavle V.

    2016-01-01

    Developing solid state materials capable of generating homogeneous white light in an energy efficient and resource-sustainable way is central to the design of new and improved devices for various lighting applications. Most currently-used phosphors depend on strategically important rare earth elements, and rely on a multicomponent approach, which produces sub-optimal quality white light. Here, we report the design and preparation of a colloidal white-light emitting nanocrystal conjugate. This conjugate is obtained by linking colloidal Ga2O3 and II–VI nanocrystals in the solution phase with a short bifunctional organic molecule (thioglycolic acid). The two types of nanocrystals are electronically coupled by Förster resonance energy transfer owing to the short separation between Ga2O3 (energy donor) and core/shell CdSe/CdS (energy acceptor) nanocrystals, and the spectral overlap between the photoluminescence of the donor and the absorption of the acceptor. Using steady state and time-resolved photoluminescence spectroscopies, we quantified the contribution of the energy transfer to the photoluminescence spectral power distribution and the corresponding chromaticity of this nanocrystal conjugate. Quantitative understanding of this new system allows for tuning of the emission color and the design of quasi-single white light emitting inorganic phosphors without the use of rare-earth elements. PMID:28344289

  15. Novel archaeal plasmid pAH1 and its interactions with the lipothrixvirus AFV1

    DEFF Research Database (Denmark)

    Basta, Tamara; Smyth, John; Forterre, Patrick

    2009-01-01

    . Although nucleotide sequence comparisons revealed extensive intergenomic exchange during the evolution of archaeal conjugative plasmids, pAH1 was shown to be stably maintained suggesting that the host system is suitable for studying plasmid-virus interactions. AFV1 infection and propagation leads to a loss...... of the circular form of pAH1 and this effect correlates positively with the increase in the intracellular quantity of AFV1 DNA. We infer that the virus inhibits plasmid replication since no pAH1 degradation was observed. This mechanism of archaeal viral inhibition of plasmid propagation is not observed...... in bacteria where relevant bacteriophages either are dependent on a conjugative plasmid for successful infection or are excluded by a resident plasmid....

  16. Size unlimited markerless deletions by a transconjugative plasmid-system in Bacillus licheniformis.

    Science.gov (United States)

    Rachinger, Michael; Bauch, Melanie; Strittmatter, Axel; Bongaerts, Johannes; Evers, Stefan; Maurer, Karl-Heinz; Daniel, Rolf; Liebl, Wolfgang; Liesegang, Heiko; Ehrenreich, Armin

    2013-09-20

    Conjugative shuttle vectors of the pKVM series, based on an IncP transfer origin and the pMAD vector with a temperature sensitive replication were constructed to establish a markerless gene deletion protocol for Bacilli without natural competence such as the exoenzyme producer Bacillus licheniformis. The pKVM plasmids can be conjugated to strains of B. licheniformis and B. subtilis. For chromosomal gene deletion, regions flanking the target gene are fused and cloned in a pKVM vector prior to conjugative transfer from Escherichia coli to B. licheniformis. Appropriate markers on the vector backbone allow for the identification of the integration at the target locus and thereafter the vector excision, both events taking place via homologous recombination. The functionality of the deletion system was demonstrated with B. licheniformis by a markerless 939 bp in-frame deletion of the yqfD gene and the deletion of a 31 kbp genomic segment carrying a PBSX-like prophage. Copyright © 2013 Elsevier B.V. All rights reserved.

  17. The role of the achiral template in enantioselective transformations. Radical conjugate additions to alpha-methacrylates followed by hydrogen atom transfer.

    Science.gov (United States)

    Sibi, Mukund P; Sausker, Justin B

    2002-02-13

    We have evaluated various achiral templates (1a-g, 10, and 16) in conjunction with chiral Lewis acids in the conjugate addition of nucleophilic radicals to alpha-methacrylates followed by enantioselective H-atom transfer. Of these, a novel naphthosultam template (10) gave high enantioselectivity in the H-atom-transfer reactions with ee's up to 90%. A chiral Lewis acid derived from MgBr(2) and bisoxazoline (2) gave the highest selectivity in the enantioselective hydrogen-atom-transfer reactions. Non-C(2) symmetric oxazolines (20-25) have also been examined as ligands, and of these, compound 25 gave optimal results (87% yield and 80% ee). Insights into rotamer control in alpha-substituted acrylates and the critical role of the tetrahedral sulfone moiety in realizing high selectivity are discussed.

  18. Comparisons of Salmonella conjugation and virulence gene hyperexpression mediated by rumen protozoa from domestic and exotic ruminants.

    Science.gov (United States)

    Brewer, Matt T; Xiong, Nalee; Dier, Jeffery D; Anderson, Kristi L; Rasmussen, Mark A; Franklin, Sharon K; Carlson, Steve A

    2011-08-05

    Recent studies have identified a phenomenon in which ciliated protozoa engulf Salmonella and the intra-protozoal environment hyperactivates virulence gene expression and provides a venue for conjugal transfer of antibiotic resistance plasmids. The former observation is relegated to Salmonella bearing the SGI1 multiresistance integron while the latter phenomenon appears to be a more generalized event for recipient Salmonella. Our previous studies have assessed virulence gene hyperexpression only with protozoa from the bovine rumen while conjugal transfer has been demonstrated in rumen protozoa from cattle and goats. The present study examined virulence gene hyperexpression for Salmonella exposed to rumen protozoa obtained from cattle, sheep, goats, or two African ruminants (giraffe and bongo). Conjugal transfer was also assessed in these protozoa using Salmonella as the recipient. Virulence gene hyperexpression was only observed following exposure to the rumen protozoa from cattle and sheep while elevated virulence was also observed in these animals. Conjugal transfer events were, however, observed in all protozoa evaluated. It therefore appears that the protozoa-based hypervirulence is not universal to all ruminants while conjugal transfer is more ubiquitous. Copyright © 2011 Elsevier B.V. All rights reserved.

  19. Numerical analysis of natural convection and radiation heat transfer from various shaped thin fin-arrays placed on a horizontal plate-a conjugate analysis

    International Nuclear Information System (INIS)

    Dogan, M.; Sivrioglu, Mecit; Yılmaz, Onder

    2014-01-01

    Highlights: • Optimum fin shape is determined for natural convection and radiation heat transfer. • Fin array with the optimum shape has a much greater average heat transfer coefficient. • The most important factors affecting the heat transfer coefficient are determined. - Abstract: Steady state natural convection and radiation heat transfer from various shaped thin fin-arrays on a horizontal base plate has been numerically investigated. A conjugate analysis has been carried out in which the conservation equations of mass, momentum and energy for the fluid in the two fin enclosure are solved together with the heat conduction equation in the fin and the base plate. Heat transfer by radiation is also considered in analysis. The heat transfer coefficient has been determined for each of the fin array considered in the present study at the same base and the same total area. The results of the analysis show that there are some important geometrical factors affecting the design of fin arrays. Taking into consideration these factors, an optimum fin shape that yields the highest average heat transfer coefficient has been determined

  20. Stimulation of chondrocytes in vitro by gene transfer with plasmids coding for epidermal growth factor (hEGF) and basic fibroblast growth factor (bFGF)

    DEFF Research Database (Denmark)

    Schmal, H; Mehlhorn, A T; Zwingmann, J

    2005-01-01

    Human epidermal growth factor (hEGF) and basic fibroblast growth factor (bFGF) influence critical characteristics of chondrocytes. The effects on metabolism and differentiation were evaluated following transfection using specific plasmids coding for both cytokines. Chondrocytes were isolated from...... of recombinant hEGF and bFGF resulted in a significant increase in cell proliferation and glucosaminoglycan production. Chondrocytes were transfected with vectors coding for either hEGF or bFGF and the production of these proteins was measured in supernatants by ELISA. Expression kinetics showed different...... patterns: hEGF was detectable 2.5 days following transfection and peaked at day 5.5, whereas bFGF-production reached its maximum 1.5 days after transfection, declining thereafter. Chondrocytes endogenously produced significant amounts of bFGF within 5 days following isolation. Proliferation of h...

  1. Chlamydial plasmids and bacteriophages.

    Science.gov (United States)

    Pawlikowska-Warych, Małgorzata; Śliwa-Dominiak, Joanna; Deptuła, Wiesław

    2015-01-01

    Chlamydia are absolute pathogens of humans and animals; despite being rather well recognised, they are still open for discovery. One such discovery is the occurrence of extrachromosomal carriers of genetic information. In prokaryotes, such carriers include plasmids and bacteriophages, which are present only among some Chlamydia species. Plasmids were found exclusively in Chlamydia (C.) trachomatis, C. psittaci, C. pneumoniae, C. suis, C. felis, C. muridarum and C. caviae. In prokaryotic organisms, plasmids usually code for genes that facilitate survival of the bacteria in the environment (although they are not essential). In chlamydia, their role has not been definitely recognised, apart from the fact that they participate in the synthesis of glycogen and encode proteins responsible for their virulence. Furthermore, in C. suis it was evidenced that the plasmid is integrated in a genomic island and contains the tetracycline-resistance gene. Bacteriophages specific for chlamydia (chlamydiaphages) were detected only in six species: C. psittaci, C. abortus, C. felis, C. caviae C. pecorum and C. pneumoniae. These chlamydiaphages cause inhibition of the developmental cycle, and delay transformation of reticulate bodies (RBs) into elementary bodies (EBs), thus reducing the possibility of infecting other cells in time. Plasmids and bacteriophages can be used in the diagnostics of chlamydioses; although especially in the case of plasmids, they are already used for detection of chlamydial infections. In addition, bacteriophages could be used as therapeutic agents to replace antibiotics, potentially addressing the problem of increasing antibiotic-resistance among chlamydia.

  2. Identification of a Single Strand Origin of Replication in the Integrative and Conjugative Element ICEBs1 of Bacillus subtilis.

    Directory of Open Access Journals (Sweden)

    Laurel D Wright

    2015-10-01

    Full Text Available We identified a functional single strand origin of replication (sso in the integrative and conjugative element ICEBs1 of Bacillus subtilis. Integrative and conjugative elements (ICEs, also known as conjugative transposons are DNA elements typically found integrated into a bacterial chromosome where they are transmitted to daughter cells by chromosomal replication and cell division. Under certain conditions, ICEs become activated and excise from the host chromosome and can transfer to neighboring cells via the element-encoded conjugation machinery. Activated ICEBs1 undergoes autonomous rolling circle replication that is needed for the maintenance of the excised element in growing and dividing cells. Rolling circle replication, used by many plasmids and phages, generates single-stranded DNA (ssDNA. In many cases, the presence of an sso enhances the conversion of the ssDNA to double-stranded DNA (dsDNA by enabling priming of synthesis of the second DNA strand. We initially identified sso1 in ICEBs1 based on sequence similarity to the sso of an RCR plasmid. Several functional assays confirmed Sso activity. Genetic analyses indicated that ICEBs1 uses sso1 and at least one other region for second strand DNA synthesis. We found that Sso activity was important for two key aspects of the ICEBs1 lifecycle: 1 maintenance of the plasmid form of ICEBs1 in cells after excision from the chromosome, and 2 stable acquisition of ICEBs1 following transfer to a new host. We identified sequences similar to known plasmid sso's in several other ICEs. Together, our results indicate that many other ICEs contain at least one single strand origin of replication, that these ICEs likely undergo autonomous replication, and that replication contributes to the stability and spread of these elements.

  3. High-yield nontoxic gene transfer through conjugation of the CM₁₈-Tat₁₁ chimeric peptide with nanosecond electric pulses.

    Science.gov (United States)

    Salomone, Fabrizio; Breton, Marie; Leray, Isabelle; Cardarelli, Francesco; Boccardi, Claudia; Bonhenry, Daniel; Tarek, Mounir; Mir, Lluis M; Beltram, Fabio

    2014-07-07

    We report a novel nontoxic, high-yield, gene delivery system based on the synergistic use of nanosecond electric pulses (NPs) and nanomolar doses of the recently introduced CM18-Tat11 chimeric peptide (sequence of KWKLFKKIGAVLKVLTTGYGRKKRRQRRR, residues 1-7 of cecropin-A, 2-12 of melittin, and 47-57 of HIV-1 Tat protein). This combined use makes it possible to drastically reduce the required CM18-Tat11 concentration and confines stable nanopore formation to vesicle membranes followed by DNA release, while no detectable perturbation of the plasma membrane is observed. Two different experimental assays are exploited to quantitatively evaluate the details of NPs and CM18-Tat11 cooperation: (i) cytofluorimetric analysis of the integrity of synthetic 1,2-dioleoyl-sn-glycero-3-phosphocholine giant unilamellar vesicles exposed to CM18-Tat11 and NPs and (ii) the in vitro transfection efficiency of a green fluorescent protein-encoding plasmid conjugated to CM18-Tat11 in the presence of NPs. Data support a model in which NPs induce membrane perturbation in the form of transient pores on all cellular membranes, while the peptide stabilizes membrane defects selectively within endosomes. Interestingly, atomistic molecular dynamics simulations show that the latter activity can be specifically attributed to the CM18 module, while Tat11 remains essential for cargo binding and vector subcellular localization. We argue that this result represents a paradigmatic example that can open the way to other targeted delivery protocols.

  4. Effect of tif expression, irradiation of recipient and presence of plasmid pKM101 on recovery of a marker from a donor exposed to ultraviolet light prior to conjugation

    International Nuclear Information System (INIS)

    Wright, A. von; Bridges, B.A.

    1980-01-01

    To detect the effect of the postulated inducible error-prone repair system ('SOS repair') on the bacterial chromosome an Hfr Escherichia coli strain JC5088 rec A was u.v.-irradiated immediately before mating it with recipients in which SOS repair was supposed to be functioning either through tif expression, u.v. irradiation or the presence of plasmid pKM101. The recombinant yields of these crosses were compared with those obtained in corresponding crosses with recipients in which SOS repair either was not induced or was totally eliminated by the lexA mutation. No difference in marker recovery efficiency could be detected between these two sets of recipients and thus no induced repair process acting on donor DNA could be demonstrated. The possible reasons for this finding are discussed. (author)

  5. Evolution of Regions Containing Antibiotic Resistance Genes in FII-2-FIB-1 ColV-Colla Virulence Plasmids.

    Science.gov (United States)

    Moran, Robert A; Hall, Ruth M

    2018-05-01

    Three ColV virulence plasmids carrying antibiotic resistance genes were assembled from draft genome sequences of commensal ST95, ST131, and ST2705 Escherichia coli isolates from healthy Australians. Plasmids pCERC4, pCERC5, and pCERC9 include almost identical backbones containing FII-2 and FIB-1 replicons and the conserved ColV virulence region with an additional ColIa determinant. Only pCERC5 includes a complete, uninterrupted F-like transfer region and was able to conjugate. pCERC5 and pCERC9 contain Tn1721, carrying the tet(A) tetracycline resistance determinant in the same location, with Tn2 (bla TEM ; ampicillin resistance) interrupting the Tn1721 in pCERC5. pCERC4 has a Tn1721/Tn21 hybrid transposon carrying dfrA5 (trimethoprim resistance) and sul1 (sulfamethoxazole resistance) in a class 1 integron. Four FII-2:FIB-1 ColV-ColIa plasmids in the GenBank nucleotide database have a related transposon in the same position, but an IS26 has reshaped the resistance gene region, deleting 2,069 bp of the integron 3'-CS, including sul1, and serving as a target for IS26 translocatable units containing bla TEM , sul2 and strAB (streptomycin resistance), or aphA1 (kanamycin/neomycin resistance). Another ColV-ColIa plasmid containing a related resistance gene region has lost the FII replicon and acquired a unique transfer region via recombination within the resistance region and at oriT. Eighteen further complete ColV plasmid sequences in GenBank contained FIB-1, but the FII replicons were of three types, FII-24, FII-18, and a variant of FII-36.

  6. Plasmids and rickettsial evolution: insight from Rickettsia felis.

    Directory of Open Access Journals (Sweden)

    Joseph J Gillespie

    2007-03-01

    Full Text Available The genome sequence of Rickettsia felis revealed a number of rickettsial genetic anomalies that likely contribute not only to a large genome size relative to other rickettsiae, but also to phenotypic oddities that have confounded the categorization of R. felis as either typhus group (TG or spotted fever group (SFG rickettsiae. Most intriguing was the first report from rickettsiae of a conjugative plasmid (pRF that contains 68 putative open reading frames, several of which are predicted to encode proteins with high similarity to conjugative machinery in other plasmid-containing bacteria.Using phylogeny estimation, we determined the mode of inheritance of pRF genes relative to conserved rickettsial chromosomal genes. Phylogenies of chromosomal genes were in agreement with other published rickettsial trees. However, phylogenies including pRF genes yielded different topologies and suggest a close relationship between pRF and ancestral group (AG rickettsiae, including the recently completed genome of R. bellii str. RML369-C. This relatedness is further supported by the distribution of pRF genes across other rickettsiae, as 10 pRF genes (or inactive derivatives also occur in AG (but not SFG rickettsiae, with five of these genes characteristic of typical plasmids. Detailed characterization of pRF genes resulted in two novel findings: the identification of oriV and replication termination regions, and the likelihood that a second proposed plasmid, pRFdelta, is an artifact of the original genome assembly.Altogether, we propose a new rickettsial classification scheme with the addition of a fourth lineage, transitional group (TRG rickettsiae, that is unique from TG and SFG rickettsiae and harbors genes from possible exchanges with AG rickettsiae via conjugation. We offer insight into the evolution of a plastic plasmid system in rickettsiae, including the role plasmids may have played in the acquirement of virulence traits in pathogenic strains, and the

  7. Photoinduced electron transfer in a Watson-Crick base-paired, 2-aminopurine:uracil-C60 hydrogen bonding conjugate.

    Science.gov (United States)

    D'Souza, Francis; Gadde, Suresh; Islam, D-M Shafiqul; Pang, Siew-Cheng; Schumacher, Amy Lea; Zandler, Melvin E; Horie, Rumiko; Araki, Yasuyaki; Ito, Osamu

    2007-02-07

    A fluorescent reporter molecule, 2-aminopurine was self-assembled via Watson-Crick base-pairing to a uracil appended fullerene to form a donor-acceptor conjugate; efficient photoinduced charge separation was confirmed by time-resolved emission and transient absorption spectral studies.

  8. Time-accurate CFD conjugate analysis of transient measurements of the heat-transfer coefficient in a channel with pin fins

    Directory of Open Access Journals (Sweden)

    Tom I-P. Shih

    2013-03-01

    Full Text Available Heat-transfer coefficients (HTC on surfaces exposed to convection environments are often measured by transient techniques such as thermochromic liquid crystal (TLC or infrared thermography. In these techniques, the surface temperature is measured as a function of time, and that measurement is used with the exact solution for unsteady, zero-dimensional (0-D or one-dimensional (1-D heat conduction into a solid to calculate the local HTC. When using the 0-D or 1-D exact solutions, the transient techniques assume the HTC and the free-stream or bulk temperature characterizing the convection environment to be constants in addition to assuming the conduction into the solid to be 0-D or 1-D. In this study, computational fluid dynamics (CFD conjugate analyses were performed to examine the errors that might be invoked by these assumptions for a problem, where the free-stream/bulk temperature and the heat-transfer coefficient vary appreciably along the surface and where conduction into the solid may not be 0-D or 1-D. The problem selected to assess these errors is flow and heat transfer in a channel lined with a staggered array of pin fins. This conjugate study uses three-dimensional (3-D unsteady Reynolds-averaged Navier–Stokes (RANS closed by the shear-stress transport (SST turbulence model for the gas phase (wall functions not used and the Fourier law for the solid phase. The errors in the transient techniques are assessed by comparing the HTC predicted by the time-accurate conjugate CFD with those predicted by the 0-D and 1-D exact solutions, where the surface temperatures needed by the exact solutions are taken from the time-accurate conjugate CFD solution. Results obtained show that the use of the 1-D exact solution for the semi-infinite wall to give reasonably accurate “transient” HTC (less than 5% relative error. Transient techniques that use the 0-D exact solution for the pin fins were found to produce large errors (up to 160% relative error

  9. A combined ANN-GA and experimental based technique for the estimation of the unknown heat flux for a conjugate heat transfer problem

    Science.gov (United States)

    M K, Harsha Kumar; P S, Vishweshwara; N, Gnanasekaran; C, Balaji

    2018-05-01

    The major objectives in the design of thermal systems are obtaining the information about thermophysical, transport and boundary properties. The main purpose of this paper is to estimate the unknown heat flux at the surface of a solid body. A constant area mild steel fin is considered and the base is subjected to constant heat flux. During heating, natural convection heat transfer occurs from the fin to ambient. The direct solution, which is the forward problem, is developed as a conjugate heat transfer problem from the fin and the steady state temperature distribution is recorded for any assumed heat flux. In order to model the natural convection heat transfer from the fin, an extended domain is created near the fin geometry and air is specified as a fluid medium and Navier Stokes equation is solved by incorporating the Boussinesq approximation. The computational time involved in executing the forward model is then reduced by developing a neural network (NN) between heat flux values and temperatures based on back propagation algorithm. The conjugate heat transfer NN model is now coupled with Genetic algorithm (GA) for the solution of the inverse problem. Initially, GA is applied to the pure surrogate data, the results are then used as input to the Levenberg- Marquardt method and such hybridization is proven to result in accurate estimation of the unknown heat flux. The hybrid method is then applied for the experimental temperature to estimate the unknown heat flux. A satisfactory agreement between the estimated and actual heat flux is achieved by incorporating the hybrid method.

  10. Investigation of diversity of plasmids carrying the blaTEM-52 gene

    DEFF Research Database (Denmark)

    Bielak, Eliza Maria; Bergenholtz, Rikke D.; Jørgensen, Mikael Skaanning

    2011-01-01

    OBJECTIVES: To investigate the diversity of plasmids that carry blaTEM-52 genes among Escherichia coli and Salmonella enterica originating from animals, meat products and humans. METHODS: A collection of 22 blaTEM-52-encoding plasmids was characterized by restriction fragment length polymorphism...... of self-transfer to a plasmid-free E. coli recipient. CONCLUSIONS: The blaTEM-52 gene found in humans could have been transmitted on transferable plasmids originating from animal sources. Some of the blaTEM-52 plasmids carry replicons that differ from the classical ones. Two novel replicons were detected...

  11. Yeast transformation mediated by Agrobacterium strains harboring an Ri plasmid: comparative study between GALLS of an Ri plasmid and virE of a Ti plasmid.

    Science.gov (United States)

    Kiyokawa, Kazuya; Yamamoto, Shinji; Sato, Yukari; Momota, Naoto; Tanaka, Katsuyuki; Moriguchi, Kazuki; Suzuki, Katsunori

    2012-07-01

    Agrobacterium strains containing a Ti plasmid can transfer T-DNA not only to plants but also to fungi, including the yeast Saccharomyces cerevisiae. However, no Agrobacterium strain harboring an Ri plasmid has been evaluated in fungal transformation. Some Ri plasmids have GALLS , instead of virE1 and virE2. GALLS protein can functionally substitute in plant transformation for a structurally different protein VirE2. In this study, we compared the yeast transformation ability among Agrobacterium donors: a strain containing a Ti plasmid, strains harboring either an agropine-type or a mikimopine-type Ri plasmid, and a strain having a modified Ri plasmid supplemented with a Ti plasmid type virE operon. Agrobacterium strains possessing GALLS transformed yeast cells far less efficiently than the strain containing virE operon. Production of GALLS in recipient yeast cells improved the yeast transformation mediated by an Agrobacterium strain lacking neither GALLS nor virE operon. A reporter assay to detect mobilization of the proteins fused with Cre recombinase revealed that VirE2 protein is much more abundant in yeast cells than GALLS. Based on these results, we concluded that the low yeast transformability mediated by Agrobacterium strains having the Ri plasmid is because of low amount of mobilized GALLS in yeast cells. © 2012 The Authors Journal compilation © 2012 by the Molecular Biology Society of Japan/Blackwell Publishing Ltd.

  12. Inhibition of bacterial conjugation by phage M13 and its protein g3p: quantitative analysis and model.

    Directory of Open Access Journals (Sweden)

    Abraham Lin

    Full Text Available Conjugation is the main mode of horizontal gene transfer that spreads antibiotic resistance among bacteria. Strategies for inhibiting conjugation may be useful for preserving the effectiveness of antibiotics and preventing the emergence of bacterial strains with multiple resistances. Filamentous bacteriophages were first observed to inhibit conjugation several decades ago. Here we investigate the mechanism of inhibition and find that the primary effect on conjugation is occlusion of the conjugative pilus by phage particles. This interaction is mediated primarily by phage coat protein g3p, and exogenous addition of the soluble fragment of g3p inhibited conjugation at low nanomolar concentrations. Our data are quantitatively consistent with a simple model in which association between the pili and phage particles or g3p prevents transmission of an F plasmid encoding tetracycline resistance. We also observe a decrease in the donor ability of infected cells, which is quantitatively consistent with a reduction in pili elaboration. Since many antibiotic-resistance factors confer susceptibility to phage infection through expression of conjugative pili (the receptor for filamentous phage, these results suggest that phage may be a source of soluble proteins that slow the spread of antibiotic resistance genes.

  13. Effects of phosphorylatable short peptide-conjugated chitosan-mediated IL-1Ra and igf-1 gene transfer on articular cartilage defects in rabbits.

    Directory of Open Access Journals (Sweden)

    Ronglan Zhao

    Full Text Available Previously, we reported an improvement in the transfection efficiency of the plasmid DNA-chitosan (pDNA/CS complex by the utilization of phosphorylatable short peptide-conjugated chitosan (pSP-CS. In this study, we investigated the effects of pSP-CS-mediated gene transfection of interleukin-1 receptor antagonist protein (IL-1Ra combined with insulin-like growth factor-1 (IGF-1 in rabbit chondrocytes and in a rabbit model of cartilage defects. pBudCE4.1-IL-1Ra+igf-1, pBudCE4.1-IL-1Ra and pBudCE4.1-igf-1 were constructed and combined with pSP-CS to form pDNA/pSP-CS complexes. These complexes were transfected into rabbit primary chondrocytes or injected into the joint cavity. Seven weeks after treatment, all rabbits were sacrificed and analyzed. High levels of IL-1Ra and igf-1 expression were detected both in the cell culture supernatant and in the synovial fluid. In vitro, the transgenic complexes caused significant proliferation of chondrocytes, promotion of glycosaminoglycan (GAG and collagen II synthesis, and inhibition of chondrocyte apoptosis and nitric oxide (NO synthesis. In vivo, the exogenous genes resulted in increased collagen II synthesis and reduced NO and GAG concentrations in the synovial fluid; histological studies revealed that pDNA/pSP-CS treatment resulted in varying degrees of hyaline-like cartilage repair and Mankin score decrease. The co-expression of both genes produced greater effects than each single gene alone both in vitro and in vivo. The results suggest that pSP-CS is a good candidate for use in gene therapy for the treatment of cartilage defects and that igf-1 and IL-1Ra co-expression produces promising biologic effects on cartilage defects.

  14. Characterization of a novel plasmid type and various genetic contexts of bla OXA-58 in Acinetobacter spp. from multiple cities in China.

    Directory of Open Access Journals (Sweden)

    Yiqi Fu

    Full Text Available BACKGROUND/OBJECTIVE: Several studies have described the epidemiological distribution of blaOXA-58-harboring Acinetobacter baumannii in China. However, there is limited data concerning the replicon types of blaOXA-58-carrying plasmids and the genetic context surrounding blaOXA-58 in Acinetobacter spp. in China. METHODOLOGY/PRINCIPAL FINDINGS: Twelve non-duplicated blaOXA-58-harboring Acinetobacter spp. isolates were collected from six hospitals in five different cities between 2005 and 2010. The molecular epidemiology of the isolates was carried out using PFGE and multilocus sequence typing. Carbapenemase-encoding genes and plasmid replicase genes were identified by PCR. The genetic location of blaOXA-58 was analyzed using S1-nuclease method. Plasmid conjugation and electrotransformation were performed to evaluate the transferability of blaOXA-58-harboring plasmids. The genetic structure surrounding blaOXA-58 was determined by cloning experiments. The twelve isolates included two Acinetobacter pittii isolates (belong to one pulsotype, three Acinetobacter nosocomialis isolates (belong to two pulsotypes and seven Acinetobacter baumannii isolates (belong to two pulsotypes/sequence types. A. baumannii ST91 was found to be a potential multidrug resistant risk clone carrying both blaOXA-58 and blaOXA-23. blaOXA-58 located on plasmids varied from ca. 52 kb to ca. 143 kb. All plasmids can be electrotransformed to A. baumannii recipient, but were untypeable by the current replicon typing scheme. A novel plasmid replicase named repAci10 was identified in blaOXA-58-harboring plasmids of two A. pittii isolates, three A. nosocomialis isolates and two A. baumannii isolates. Four kinds of genetic contexts of blaOXA-58 were identified. The transformants of plasmids with structure of IS6 family insertion sequence (ISOur1, IS1008 or IS15-ΔISAba3-like element-blaOXA-58 displayed carbapenem nonsusceptible, while others with structure of intact ISAba3-like element

  15. Dissemination of NDM-1-Producing Enterobacteriaceae Mediated by the IncX3-Type Plasmid

    Science.gov (United States)

    Fu, Ying; Du, Xiaoxing; Shen, Yuqin; Yu, Yunsong

    2015-01-01

    The emergence and spread of NDM-1-producing Enterobacteriaceae have resulted in a worldwide public health risk that has affected some provinces of China. China is an exceptionally large country, and there is a crucial need to investigate the epidemic of bla NDM-1-positive Enterobacteriaceae in our province. A total of 186 carbapenem-resistant Enterobacteriaceae isolates (CRE) were collected in a grade-3 hospital in Zhejiang province. Carbapenem-resistant genes, including bla KPC, bla IMP, bla VIM, bla OXA-48 and bla NDM-1 were screened and sequenced. Ninety isolates were identified as harboring the bla KPC-2 genes, and five bla NDM-1-positive isolates were uncovered. XbaI-PFGE revealed that three bla NDM-1-positive K. pneumoniae isolates belonged to two different clones. S1-PFGE and southern blot suggested that the bla NDM-1 genes were located on IncX3-type plasmids with two different sizes ranging from 33.3 to 54.7 kb (n=4) and 104.5 to 138.9 kb (n=1), respectively, all of which could easily transfer to Escherichia coli by conjugation and electrotransformation. The high-throughput sequencing of two plasmids was performed leading to the identification of a smaller 54-kb plasmid, which had high sequence similarity with a previously reported pCFNDM-CN, and a larger plasmid in which only a 7.8-kb sequence of a common gene environment around bla NDM-1 (bla NDM-1-trpF- dsbC-cutA1-groEL-ΔInsE,) was detected. PCR mapping and sequencing demonstrated that four smaller bla NDM-1 plasmids contained a common gene environment around bla NDM-1 (IS5-bla NDM-1-trpF- dsbC-cutA1-groEL). We monitored the CRE epidemic in our hospital and determined that KPC-2 carbapenemase was a major risk to patient health and the IncX3-type plasmid played a vital role in the spread of the bla NDM-1 gene among the CRE. PMID:26047502

  16. Plasmid-mediated UV-protection in Streptococcus lactis

    Energy Technology Data Exchange (ETDEWEB)

    Chopin, M.C.; Rouault, A. (Institut National de la Recherche Agronomique, Rennes (France). Lab. de Recherches de Technologie Laitiere); Moillo-Batt, A. (Institut National de la Sante et de la Recherche Medicale (INSERM), Hopital de Pontchaillon, 35 - Rennes (France))

    1985-02-01

    Streptococcus lactis strain IL594 contains 9 plasmids, designated pIL1 to pIL9. On the basis of protoplast-induced curing experiments the authors showed that derivatives containing pIL7 were resistant to UV-irradiation while derivatives lacking pIL7 were sensitive. The pIL7-determined UV-protection was confirmed by co-transfer of the plasmid and of the character into a plasmid-free derivative of S. lactis IL594. Moreover, prophage induction required higher UV-fluence in this derivative carrying pIL7 than in the plasmid-free strain. This is the first report of a plasmid-mediated UV-protection in group N streptococci.

  17. Full sequence and comparative analysis of the plasmid pAPEC-1 of avian pathogenic E. coli chi7122 (O78:K80:H9.

    Directory of Open Access Journals (Sweden)

    Melha Mellata

    Full Text Available Extra-intestinal pathogenic E. coli (ExPEC, including Avian Pathogenic E. coli (APEC, are very diverse. They cause a complex of diseases in Human, animals, and birds. Even though large plasmids are often associated with the virulence of ExPEC, their characterization is still in its infancy.We fully sequenced and analyzed the large plasmid pAPEC-1 (103,275-bp associated with the APEC strain chi7122, from worldwide serogroup O78ratioK80ratioH9. A putative virulence region spanning an 80-kb region of pAPEC-1 possesses four iron acquisition systems (iutA iucABCD, sitABCD, iroBCDN, and temperature-sensitive hemagglutinin tsh, a colicin V operon, increasing serum sensitivity iss, ompT, hlyF, and etsABC. Thirty three ORFs in pAPEC-1 are identified as insertion sequences (ISs that belong to nine families with diverse origins. The full length of the transfer region in pAPEC-1 (11 kb is shorter compared to the tra region of other sequenced F plasmids; the absence of some tra genes in pAPEC-1 affects its self-transferability, and the conjugative function of the plasmid was effective only in the presence of other plasmids. Two-replicon systems, repFIIA-repFIC and repFIB, and two post-segregational systems, srnB and hok/sok, are also present in the sequence of pAPEC-1. The comparison of the pAPEC-1 sequence with the two available plasmid sequences reveals more gene loss and reorganization than previously appreciated. The presence of pAPEC-1-associated genes is assessed in human ExPEC by PCR. Many patterns of association between genes are found.The pathotype typical of pAPEC-1 was present in some human strains, which indicates a horizontal transfer between strains and the zoonotic risk of APEC strains. ColV plasmids could have common virulence genes that could be acquired by transposition, without sharing genes of plasmid function.

  18. IncF Plasmids Are Commonly Carried by Antibiotic Resistant Escherichia coli Isolated from Drinking Water Sources in Northern Tanzania

    OpenAIRE

    Lyimo, Beatus; Buza, Joram; Subbiah, Murugan; Temba, Sylivester; Kipasika, Honest; Smith, Woutrina; Call, Douglas R.

    2016-01-01

    The aim of this study was to identify the replicon types of plasmids, conjugation efficiencies, and the complement of antibiotic resistance genes for a panel of multidrug resistant E. coli isolates from surface waters in northern Tanzania. Standard membrane filtration was used to isolate and uidA PCR was used to confirm the identity of strains as E. coli. Antibiotic susceptibility was determined by breakpoint assay and plasmid conjugation was determined by filter-mating experiments. PCR and s...

  19. Expansion of plasmid mediated blaACT-2 among Pseudomonas aeruginosa associated with postoperative infection and its transcriptional response under cephalosporin stress.

    Directory of Open Access Journals (Sweden)

    Birson Ingti, Deepjyoti Paul, Anand Prakash Maurya

    2017-06-01

    Full Text Available Objectives: Organisms harboring multiple plasmid mediated β-lactamases are major concerns in nosocomial infections. Among these plasmid mediated β-lactamases, ACT (EBC family is a clinically important enzyme capable of hydrolyzing broad spectrum cephalosporins. Therefore, the present study was undertaken to determine the prevalence of ACT determinant along with other co-existing β-lactamase genes in P. aeruginosa strains. Methods: A total of 176 Pseudomonas isolates were phenotypically screened for the presence of AmpC β-lactamase by M3DET Method followed by Molecular detection using PCR assay. Transcriptional evaluation of blaACT-2 gene was analyzed by RT-PCR and its transferability was performed by transformation and conjugation. Results: Present study demonstrates the presence of ACT-2 allele among 12 strains of P. aeruginosa. Co-existence of other β-lactamase genes were encountered among ACT-2 harboring strains which includes CTX-M (n=2, SHV (n=3, TEM (n=2, VEB (n=2, OXA-10 (n=1, CIT (n=2 and DHA (n=3. Fingerprinting by REP PCR revealed the isolates harboring ACT-2 to be distinct and these isolates showed high resistance to expanded-spectrum cephalosporins and even to carbapenem group of drugs. This ACT-2 allele was encoded in the plasmid (L/M, FIA, FIB Inc. Group and conjugatively transferable. Transcriptional analysis revealed a significant increase in ACT-2 expression (483 fold when induced by ceftriaxone at 4 µg/ml followed by ceftazidime at 8 µg/ml (31 fold and cefotaxime 4 µg/ml (8 fold. Conclusion: In this study detection of ACT-2 plasmid mediated AmpC β-lactamase along with other β-lactamase genes in clinical isolates of P. aeruginosa represents a serious therapeutic challenge. Therefore, revision in antimicrobial policy is required for effective treatment of patients infected with pathogen expressing this mechanism. J Microbiol Infect Dis 2017; 7(2: 75-82

  20. Fundamental study on gene transfer utilizing magnetic force and jet injector

    Energy Technology Data Exchange (ETDEWEB)

    Hasegawa, T.; Nakagami, H.; Akiyama, Y.; Nishjima, S. [Osaka University, Osaka (Japan)

    2017-03-15

    Recently, DNA vaccination is attracting attentions as a new therapeutic method for lifestyle diseases and autoimmune diseases. However, its clinical applications are limited because a safe and efficient gene transfer method has not been established yet. In this study, a new method of gene transfer was proposed which utilizes the jet injection and the magnetic transfection. The jet injection is a method to inject medical liquid by momentary high pressure without needle. The injected liquid diffuses in the bio tissue and the endocytosis is considered to be improved by the diffusion. The magnetic transfection is a method to deliver the conjugates of plasmid DNA and magnetic particles to the desired site by external magnetic field. It is expected that jet injection of the conjugates causes slight membrane disruptions and the traction of the conjugates by magnetic field induces the efficient gene transfer. In conclusion, the possibility of improvement of the gene expression by the combination of jet injection and magnetic transfection was confirmed.

  1. Fundamental study on gene transfer utilizing magnetic force and jet injector

    International Nuclear Information System (INIS)

    Hasegawa, T.; Nakagami, H.; Akiyama, Y.; Nishjima, S.

    2017-01-01

    Recently, DNA vaccination is attracting attentions as a new therapeutic method for lifestyle diseases and autoimmune diseases. However, its clinical applications are limited because a safe and efficient gene transfer method has not been established yet. In this study, a new method of gene transfer was proposed which utilizes the jet injection and the magnetic transfection. The jet injection is a method to inject medical liquid by momentary high pressure without needle. The injected liquid diffuses in the bio tissue and the endocytosis is considered to be improved by the diffusion. The magnetic transfection is a method to deliver the conjugates of plasmid DNA and magnetic particles to the desired site by external magnetic field. It is expected that jet injection of the conjugates causes slight membrane disruptions and the traction of the conjugates by magnetic field induces the efficient gene transfer. In conclusion, the possibility of improvement of the gene expression by the combination of jet injection and magnetic transfection was confirmed

  2. Excitation energy transfer in ruthenium (II)-porphyrin conjugates led to enhanced emission quantum yield and {sup 1}O{sub 2} generation

    Energy Technology Data Exchange (ETDEWEB)

    Pan, Jie; Jiang, Lijun; Chan, Chi-Fai [Department of Chemistry, Hong Kong Baptist University, Kowloon Tong, Hong Kong Special Administrative Region (Hong Kong); Tsoi, Tik-Hung [Department of Applied Biology and Chemical Technology, Hung Hom, Hong Kong Special Administrative Region (Hong Kong); Shiu, Kwok-Keung; Kwong, Daniel W.J. [Department of Chemistry, Hong Kong Baptist University, Kowloon Tong, Hong Kong Special Administrative Region (Hong Kong); Wong, Wing-Tak [Department of Applied Biology and Chemical Technology, Hung Hom, Hong Kong Special Administrative Region (Hong Kong); Wong, Wai-Kwok, E-mail: wkwong@hkbu.edu.hk [Department of Chemistry, Hong Kong Baptist University, Kowloon Tong, Hong Kong Special Administrative Region (Hong Kong); Wong, Ka-Leung, E-mail: klwong@hkbu.edu.hk [Department of Chemistry, Hong Kong Baptist University, Kowloon Tong, Hong Kong Special Administrative Region (Hong Kong)

    2017-04-15

    Porphyrins are good photodynamic therapy (PDT) agents due to its flexibility for modifications to achieve tumor localization and photo-cytotoxicity against cancer. Yet they are not perfect. In a Ru(polypyridyl)-porphyrin system, the Ru(polypyridyl) moiety improves the water solubility and cell permeability. Consider the similar excited state energies between Ru(polypyridyl) and porphyrin moieties; a small perturbation (e.g. Zn(II) metalation) would lead to a marked change in the energy migration process. In this work, we have synthesized a series of porphyrins conjugated with Ru(polypyridyl) complexes using different linkers and investigated their photophysical properties, which included singlet oxygen quantum yield and their in vitro biological properties, resulting from linker variation and porphyrin modification by Zn(II) metalation. - Graphical abstract: Four amphiphilic ruthenium(II)-porphyrin complexes were prepared that display energy transfer conversion with zinc coordination, lysosome specific target, low dark toxicity and efficient photodynamic therapy.

  3. Mortalin antibody-conjugated quantum dot transfer from human mesenchymal stromal cells to breast cancer cells requires cell–cell interaction

    Energy Technology Data Exchange (ETDEWEB)

    Pietilä, Mika [National Institute of Advanced industrial Sciences and Technology, Tsukuba, Ibaraki 305 8562 (Japan); Lehenkari, Petri [Institute of Biomedicine, Department of Anatomy and Cell Biology, University of Oulu, Aapistie 7, P.O. Box 5000, FIN-90014 (Finland); Institute of Clinical Medicine, Division of Surgery, University of Oulu and Clinical Research Centre, Department of Surgery and Intensive Care, Oulu University Hospital, Aapistie 5a, P.O. Box 5000, FIN-90014 (Finland); Kuvaja, Paula [Institute of Biomedicine, Department of Anatomy and Cell Biology, University of Oulu, Aapistie 7, P.O. Box 5000, FIN-90014 (Finland); Department of Pathology, Oulu University Hospital, P.O. Box 50, FIN-90029 OYS, Oulu (Finland); Kaakinen, Mika [Biocenter Oulu, University of Oulu, P.O. Box 5000, FI-90014 (Finland); Kaul, Sunil C.; Wadhwa, Renu [National Institute of Advanced industrial Sciences and Technology, Tsukuba, Ibaraki 305 8562 (Japan); Uemura, Toshimasa, E-mail: t.uemura@aist.go.jp [National Institute of Advanced industrial Sciences and Technology, Tsukuba, Ibaraki 305 8562 (Japan)

    2013-11-01

    The role of tumor stroma in regulation of breast cancer growth has been widely studied. However, the details on the type of heterocellular cross-talk between stromal and breast cancer cells (BCCs) are still poorly known. In the present study, in order to investigate the intercellular communication between human mesenchymal stromal cells (hMSCs) and breast cancer cells (BCCs, MDA-MB-231), we recruited cell-internalizing quantum dots (i-QD) generated by conjugation of cell-internalizing anti-mortalin antibody and quantum dots (QD). Co-culture of illuminated and color-coded hMSCs (QD655) and BCCs (QD585) revealed the intercellular transfer of QD655 signal from hMSCs to BCCs. The amount of QD double positive BCCs increased gradually within 48 h of co-culture. We found prominent intercellular transfer of QD655 in hanging drop co-culture system and it was non-existent when hMSCs and BBCs cells were co-cultured in trans-well system lacking imminent cell–cell contact. Fluorescent and electron microscope analyses also supported that the direct cell-to-cell interactions may be required for the intercellular transfer of QD655 from hMSCs to BCCs. To the best of our knowledge, the study provides a first demonstration of transcellular crosstalk between stromal cells and BCCs that involve direct contact and may also include a transfer of mortalin, an anti-apoptotic and growth-promoting factor enriched in cancer cells.

  4. New plasmid-mediated aminoglycoside 6'-N-acetyltransferase, AAC(6')-Ian, and ESBL, TLA-3, from a Serratia marcescens clinical isolate.

    Science.gov (United States)

    Jin, Wanchun; Wachino, Jun-Ichi; Kimura, Kouji; Yamada, Keiko; Arakawa, Yoshichika

    2015-05-01

    Enterobacteriaceae clinical isolates showing amikacin resistance (MIC 64 to >256 mg/L) in the absence of 16S rRNA methyltransferase (MTase) genes were found. The aim of this study was to clarify the molecular mechanisms underlying amikacin resistance in Enterobacteriaceae clinical isolates that do not produce 16S rRNA MTases. PCR was performed to detect already-known amikacin resistance determinants. Cloning experiments and sequence analyses were performed to characterize unknown amikacin resistance determinants. Transfer of amikacin resistance determinants was performed by conjugation and transformation. The complete nucleotide sequence of the plasmids was determined by next-generation sequencing technology. Amikacin resistance enzymes were purified with a column chromatography system. The enzymatic function of the purified protein was investigated by thin-layer chromatography (TLC) and HPLC. Among the 14 isolates, 9 were found to carry already-known amikacin resistance determinants such as aac(6')-Ia and aac(6')-Ib. Genetic analyses revealed the presence of a new amikacin acetyltransferase gene, named aac(6')-Ian, located on a 169 829 bp transferable plasmid (p11663) of the Serratia marcescens strain NUBL-11663, one of the five strains negative for known aac(6') genes by PCR. Plasmid p11663 also carried a novel ESBL gene, named blaTLA-3. HPLC and TLC analyses demonstrated that AAC(6')-Ian catalysed the transfer of an acetyl group from acetyl coenzyme A onto an amine at the 6'-position of various aminoglycosides. We identified aac(6')-Ian as a novel amikacin resistance determinant together with a new ESBL gene, blaTLA-3, on a transferable plasmid of a S. marcescens clinical isolate. © The Author 2015. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  5. Transfer of Antibiotic Resistance in Staphylococcus aureus

    DEFF Research Database (Denmark)

    Haaber, Jakob; Penadés, José R; Ingmer, Hanne

    2017-01-01

    Staphylococcus aureus is a serious human pathogen with remarkable adaptive powers. Antibiotic-resistant clones rapidly emerge mainly by acquisition of antibiotic-resistance genes from other S. aureus strains or even from other genera. Transfer is mediated by a diverse complement of mobile genetic...... of plasmids that can be transferred by conjugation and the efficiency with which transduction occurs. Here, we review the main routes of antibiotic resistance gene transfer in S. aureus in the context of its biology as a human commensal and a life-threatening pathogen. Staphylococcus aureus cells...... are effective in exchanging mobile genetic elements, including antibiotic-resistance genes.During colonization or infection of host organisms, the exchange appears to be particularly effective.Bacteriophage-mediated transfer involves both transduction and autotransduction, which may enable lysogenic S. aureus...

  6. A classification system for plasmids from Enterococci and other Gram-positive bacteria

    DEFF Research Database (Denmark)

    Jensen, Lars Bogø; Garcia-Migura, Lourdes; Valenzuela, Antonio Jesus Sanchez

    2010-01-01

    A classification system for plasmids isolated from enterococci and other Gram-positive bacteria was developed based on 111 published plasmid sequences from enterococci and other Gram-positive bacteria; mostly staphylococci. Based on PCR amplification of conserved areas of the replication initiating....... Furthermore, conjugation experiments were performed obtaining 30 transconjugants when selecting for antimicrobial resistance. Among them 19 gave no positive amplicons indicating presence of rep-families not tested for in this experimental setup....

  7. Antibiotic resistance plasmids of Staphylococcus aureus and their clinical importance

    International Nuclear Information System (INIS)

    Lacey, R.W.

    1975-01-01

    A variety of plasmids were isolated physically, and most antibiotic resistance is thought to be plasmid mediated. A number of characters (e.g., resistance to erythromycin or methicillin, and production of pigment) are determined by genes that do not give clear indications of either plasmid or chromosomal location. Although the formation of a particular plasmid is probably, even in bacterial terms, a very rare event, once formed such an element can spread rapidly among the bacterial population. The spectacular increase in the incidence of penicillinase-producing hospital strains in the late 1940's could have been due in part to this process. Evidence is stronger, however, for the intercell transfer of recently isolated plasmids coding for resistance to fusidic acid (and penicillinase production), or for neomycin, or for tetracycline resistance. Study of bacterial plasmids can resolve fundamental biochemical problems, and give some insight into the life of the cell at the molecular level. But the immediate application of the study of staphylococcal plasmids may be directed towards improving the effectiveness of antibiotic therapy. The most important aspect of future anti-staphylococcal chemotherapy should thus be the limitation of the use of antibiotics, particularly for application to the skin and nose. (U.S.)

  8. Degenerate primer MOB typing of multiresistant clinical isolates of E. coli uncovers new plasmid backbones.

    Science.gov (United States)

    Garcillán-Barcia, M Pilar; Ruiz del Castillo, Belén; Alvarado, Andrés; de la Cruz, Fernando; Martínez-Martínez, Luis

    2015-01-01

    Degenerate Primer MOB Typing is a PCR-based protocol for the classification of γ-proteobacterial transmissible plasmids in five phylogenetic relaxase MOB families. It was applied to a multiresistant E. coli collection, previously characterized by PCR-based replicon-typing, in order to compare both methods. Plasmids from 32 clinical isolates of multiresistant E. coli (19 extended spectrum beta-lactamase producers and 13 non producers) and their transconjugants were analyzed. A total of 95 relaxases were detected, at least one per isolate, underscoring the high potential of these strains for antibiotic-resistance transmission. MOBP12 and MOBF12 plasmids were the most abundant. Most MOB subfamilies detected were present in both subsets of the collection, indicating a shared mobilome among multiresistant E. coli. The plasmid profile obtained by both methods was compared, which provided useful data upon which decisions related to the implementation of detection methods in the clinic could be based. The phylogenetic depth at which replicon and MOB-typing classify plasmids is different. While replicon-typing aims at plasmid replication regions with non-degenerate primers, MOB-typing classifies plasmids into relaxase subfamilies using degenerate primers. As a result, MOB-typing provides a deeper phylogenetic depth than replicon-typing and new plasmid groups are uncovered. Significantly, MOB typing identified 17 plasmids and an integrative and conjugative element, which were not detected by replicon-typing. Four of these backbones were different from previously reported elements. Copyright © 2014 Elsevier Inc. All rights reserved.

  9. Characterization of Endogenous Plasmids from Lactobacillus salivarius UCC118▿ †

    OpenAIRE

    Fang, Fang; Flynn, Sarah; Li, Yin; Claesson, Marcus J.; van Pijkeren, Jan-Peter; Collins, J. Kevin; van Sinderen, Douwe; O'Toole, Paul W.

    2008-01-01

    The genome of Lactobacillus salivarius UCC118 comprises a 1.83-Mb chromosome, a 242-kb megaplasmid (pMP118), and two smaller plasmids of 20 kb (pSF118-20) and 44 kb (pSF118-44). Annotation and bioinformatic analyses suggest that both of the smaller plasmids replicate by a theta replication mechanism. Furthermore, it appears that they are transmissible, although neither possesses a complete set of conjugation genes. Plasmid pSF118-20 encodes a toxin-antitoxin system composed of pemI and pemK h...

  10. TOL Plasmid Carriage Enhances Biofilm Formation and Increases Extracellular DNA Content in Pseudomonas Putida KT2440

    DEFF Research Database (Denmark)

    Smets, Barth F.; D'Alvise, Paul; Yankelovich, T.

    laser scanning microscopy. The TOL-carrying strains formed pellicles and thick biofilms, whereas the same strains without the plasmid displayed little adherent growth. Microscopy using fluorescent nucleic acid- specific stains (cytox orange, propidium iodide) revealed differences in production...... combined with specific cytostains; release of cytoplasmic material was assayed by a β-glucosidase assay. Enhanced cell lysis due to plasmid carriage was ruled out as the mechanism for eDNA release. We report, for the first time, that carriage of a conjugative plasmid leads to increased biofilm formation...

  11. TOL plasmid carriage enhances biofilm formation and increases extracellular DNA content in Pseudomonas putida KT2440

    DEFF Research Database (Denmark)

    D'Alvise, Paul; Sjoholm, O.R.; Yankelevich, T.

    2010-01-01

    laser scanning microscopy. The TOL-carrying strains formed pellicles and thick biofilms, whereas the same strains without the plasmid displayed little adherent growth. Microscopy using fluorescent nucleic acid-specific stains revealed differences in the production of extracellular polymeric substances......: TOL carriage leads to more extracellular DNA (eDNA) in pellicles and biofilms. Pellicles were dissolved by DNase I treatment. Enhanced cell lysis due to plasmid carriage was ruled out as the mechanism for eDNA release. We report, for the first time, that carriage of a conjugative plasmid leads...

  12. Global Transcriptional Regulation of Backbone Genes in Broad-Host-Range Plasmid RA3 from the IncU Group Involves Segregation Protein KorB (ParB Family).

    Science.gov (United States)

    Kulinska, Anna; Godziszewska, Jolanta; Wojciechowska, Anna; Ludwiczak, Marta; Jagura-Burdzy, Grazyna

    2016-04-01

    The KorB protein of the broad-host-range conjugative plasmid RA3 from the IncU group belongs to the ParB family of plasmid and chromosomal segregation proteins. As a partitioning DNA-binding factor, KorB specifically recognizes a 16-bp palindrome which is an essential motif in the centromere-like sequence parSRA3, forms a segrosome, and together with its partner IncC (ParA family) participates in active DNA segregation ensuring stable plasmid maintenance. Here we show that by binding to this palindromic sequence, KorB also acts as a repressor for the adjacent mobC promoter driving expression of the mobC-nicoperon, which is involved in DNA processing during conjugation. Three other promoters, one buried in the conjugative transfer module and two divergent promoters located at the border between the replication and stability regions, are regulated by KorB binding to additional KorB operators (OBs). KorB acts as a repressor at a distance, binding to OBs separated from their cognate promoters by between 46 and 1,317 nucleotides. This repressor activity is facilitated by KorB spreading along DNA, since a polymerization-deficient KorB variant with its dimerization and DNA-binding abilities intact is inactive in transcriptional repression. KorB may act as a global regulator of RA3 plasmid functions in Escherichia coli, since its overexpression in transnegatively interferes with mini-RA3 replication and stable maintenance of RA3. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  13. Modeling 3D conjugate heat and mass transfer for turbulent air drying of Chilean papaya in a direct contact dryer

    Science.gov (United States)

    Lemus-Mondaca, Roberto A.; Vega-Gálvez, Antonio; Zambra, Carlos E.; Moraga, Nelson O.

    2017-01-01

    A 3D model considering heat and mass transfer for food dehydration inside a direct contact dryer is studied. The k- ɛ model is used to describe turbulent air flow. The samples thermophysical properties as density, specific heat, and thermal conductivity are assumed to vary non-linearly with temperature. FVM, SIMPLE algorithm based on a FORTRAN code are used. Results unsteady velocity, temperature, moisture, kinetic energy and dissipation rate for the air flow are presented, whilst temperature and moisture values for the food also are presented. The validation procedure includes a comparison with experimental and numerical temperature and moisture content results obtained from experimental data, reaching a deviation 7-10 %. In addition, this turbulent k- ɛ model provided a better understanding of the transport phenomenon inside the dryer and sample.

  14. Explanatory chapter: how plasmid preparation kits work.

    Science.gov (United States)

    Koontz, Laura

    2013-01-01

    To isolate plasmid DNA from bacteria using a commercial plasmid miniprep kit (if interested, compare this protocol with Isolation of plasmid DNA from bacteria). Copyright © 2013 Elsevier Inc. All rights reserved.

  15. An adaptive simplex cut-cell method for high-order discontinuous Galerkin discretizations of elliptic interface problems and conjugate heat transfer problems

    Science.gov (United States)

    Sun, Huafei; Darmofal, David L.

    2014-12-01

    In this paper we propose a new high-order solution framework for interface problems on non-interface-conforming meshes. The framework consists of a discontinuous Galerkin (DG) discretization, a simplex cut-cell technique, and an output-based adaptive scheme. We first present a DG discretization with a dual-consistent output evaluation for elliptic interface problems on interface-conforming meshes, and then extend the method to handle multi-physics interface problems, in particular conjugate heat transfer (CHT) problems. The method is then applied to non-interface-conforming meshes using a cut-cell technique, where the interface definition is completely separate from the mesh generation process. No assumption is made on the interface shape (other than Lipschitz continuity). We then equip our strategy with an output-based adaptive scheme for an accurate output prediction. Through numerical examples, we demonstrate high-order convergence for elliptic interface problems and CHT problems with both smooth and non-smooth interface shapes.

  16. Cloning in Streptococcus lactis of plasmid-mediated UV resistance and effect on prophage stability

    International Nuclear Information System (INIS)

    Chopin, M.C.; Chopin, A.; Rouault, A.; Simon, D.

    1986-01-01

    Plasmid pIL7 (33 kilobases) from Streptococcus lactis enhances UV resistance and prophage stability. A 5.4-kilobase pIL7 fragment carrying genes coding for both characters was cloned into S. lactis, using plasmid pHV1301 as the cloning vector. The recombinant plasmid was subsequently transferred to three other S. lactis strains by transformation or protoplast fusion. Cloned genes were expressed in all tested strains

  17. Plasmid Conjugation in E. coli and Drug Resistance

    African Journals Online (AJOL)

    Prof. Ogunji

    2Department of Pharmaceutics and Pharmaceutical Microbiology, Ahmadu Bello University, Zaria ... decreasing the emergence of resistant bacteria strains within our environment. ..... Cowan and Steel's Manual for the Identification of Medical.

  18. In Silico Detection and Typing of Plasmids using PlasmidFinder and Plasmid Multilocus Sequence Typing

    DEFF Research Database (Denmark)

    Carattoli, Alessandra; Zankari, Ea; García-Fernández, Aurora

    2014-01-01

    In the work presented here, we designed and developed two easy-to-use Web tools for in silico detection and characterization of whole-genome sequence (WGS) and whole-plasmid sequence data from members of the family Enterobacteriaceae. These tools will facilitate bacterial typing based on draft...... genomes of multidrug-resistant Enterobacteriaceae species by the rapid detection of known plasmid types. Replicon sequences from 559 fully sequenced plasmids associated with the family Enterobacteriaceae in the NCBI nucleotide database were collected to build a consensus database for integration...... sequences identified in the 559 fully sequenced plasmids. For plasmid multilocus sequence typing (pMLST) analysis, a database that is updated weekly was generated from www.pubmlst.org and integrated into a Web tool called pMLST. Both databases were evaluated using draft genomes from a collection...

  19. The Conjugative Relaxase TrwC Promotes Integration of Foreign DNA in the Human Genome.

    Science.gov (United States)

    González-Prieto, Coral; Gabriel, Richard; Dehio, Christoph; Schmidt, Manfred; Llosa, Matxalen

    2017-06-15

    Bacterial conjugation is a mechanism of horizontal DNA transfer. The relaxase TrwC of the conjugative plasmid R388 cleaves one strand of the transferred DNA at the oriT gene, covalently attaches to it, and leads the single-stranded DNA (ssDNA) into the recipient cell. In addition, TrwC catalyzes site-specific integration of the transferred DNA into its target sequence present in the genome of the recipient bacterium. Here, we report the analysis of the efficiency and specificity of the integrase activity of TrwC in human cells, using the type IV secretion system of the human pathogen Bartonella henselae to introduce relaxase-DNA complexes. Compared to Mob relaxase from plasmid pBGR1, we found that TrwC mediated a 10-fold increase in the rate of plasmid DNA transfer to human cells and a 100-fold increase in the rate of chromosomal integration of the transferred DNA. We used linear amplification-mediated PCR and plasmid rescue to characterize the integration pattern in the human genome. DNA sequence analysis revealed mostly reconstituted oriT sequences, indicating that TrwC is active and recircularizes transferred DNA in human cells. One TrwC-mediated site-specific integration event was detected, proving that TrwC is capable of mediating site-specific integration in the human genome, albeit with very low efficiency compared to the rate of random integration. Our results suggest that TrwC may stabilize the plasmid DNA molecules in the nucleus of the human cell, probably by recircularization of the transferred DNA strand. This stabilization would increase the opportunities for integration of the DNA by the host machinery. IMPORTANCE Different biotechnological applications, including gene therapy strategies, require permanent modification of target cells. Long-term expression is achieved either by extrachromosomal persistence or by integration of the introduced DNA. Here, we studied the utility of conjugative relaxase TrwC, a bacterial protein with site

  20. A degenerate primer MOB typing (DPMT method to classify gamma-proteobacterial plasmids in clinical and environmental settings.

    Directory of Open Access Journals (Sweden)

    Andrés Alvarado

    Full Text Available Transmissible plasmids are responsible for the spread of genetic determinants, such as antibiotic resistance or virulence traits, causing a large ecological and epidemiological impact. Transmissible plasmids, either conjugative or mobilizable, have in common the presence of a relaxase gene. Relaxases were previously classified in six protein families according to their phylogeny. Degenerate primers hybridizing to coding sequences of conserved amino acid motifs were designed to amplify related relaxase genes from γ-Proteobacterial plasmids. Specificity and sensitivity of a selected set of 19 primer pairs were first tested using a collection of 33 reference relaxases, representing the diversity of γ-Proteobacterial plasmids. The validated set was then applied to the analysis of two plasmid collections obtained from clinical isolates. The relaxase screening method, which we call "Degenerate Primer MOB Typing" or DPMT, detected not only most known Inc/Rep groups, but also a plethora of plasmids not previously assigned to any Inc group or Rep-type.

  1. Novel plasmids and resistance phenotypes in Yersinia pestis: unique plasmid inventory of strain Java 9 mediates high levels of arsenic resistance.

    Science.gov (United States)

    Eppinger, Mark; Radnedge, Lyndsay; Andersen, Gary; Vietri, Nicholas; Severson, Grant; Mou, Sherry; Ravel, Jacques; Worsham, Patricia L

    2012-01-01

    Growing evidence suggests that the plasmid repertoire of Yersinia pestis is not restricted to the three classical virulence plasmids. The Java 9 strain of Y. pestis is a biovar Orientalis isolate obtained from a rat in Indonesia. Although it lacks the Y. pestis-specific plasmid pMT, which encodes the F1 capsule, it retains virulence in mouse and non-human primate animal models. While comparing diverse Y. pestis strains using subtractive hybridization, we identified sequences in Java 9 that were homologous to a Y. enterocolitica strain carrying the transposon Tn2502, which is known to encode arsenic resistance. Here we demonstrate that Java 9 exhibits high levels of arsenic and arsenite resistance mediated by a novel promiscuous class II transposon, named Tn2503. Arsenic resistance was self-transmissible from Java 9 to other Y. pestis strains via conjugation. Genomic analysis of the atypical plasmid inventory of Java 9 identified pCD and pPCP plasmids of atypical size and two previously uncharacterized cryptic plasmids. Unlike the Tn2502-mediated arsenic resistance encoded on the Y. enterocolitica virulence plasmid; the resistance loci in Java 9 are found on all four indigenous plasmids, including the two novel cryptic plasmids. This unique mobilome introduces more than 105 genes into the species gene pool. The majority of these are encoded by the two entirely novel self-transmissible plasmids, which show partial homology and synteny to other enterics. In contrast to the reductive evolution in Y. pestis, this study underlines the major impact of a dynamic mobilome and lateral acquisition in the genome evolution of the plague bacterium.

  2. Novel plasmids and resistance phenotypes in Yersinia pestis: unique plasmid inventory of strain Java 9 mediates high levels of arsenic resistance.

    Directory of Open Access Journals (Sweden)

    Mark Eppinger

    Full Text Available Growing evidence suggests that the plasmid repertoire of Yersinia pestis is not restricted to the three classical virulence plasmids. The Java 9 strain of Y. pestis is a biovar Orientalis isolate obtained from a rat in Indonesia. Although it lacks the Y. pestis-specific plasmid pMT, which encodes the F1 capsule, it retains virulence in mouse and non-human primate animal models. While comparing diverse Y. pestis strains using subtractive hybridization, we identified sequences in Java 9 that were homologous to a Y. enterocolitica strain carrying the transposon Tn2502, which is known to encode arsenic resistance. Here we demonstrate that Java 9 exhibits high levels of arsenic and arsenite resistance mediated by a novel promiscuous class II transposon, named Tn2503. Arsenic resistance was self-transmissible from Java 9 to other Y. pestis strains via conjugation. Genomic analysis of the atypical plasmid inventory of Java 9 identified pCD and pPCP plasmids of atypical size and two previously uncharacterized cryptic plasmids. Unlike the Tn2502-mediated arsenic resistance encoded on the Y. enterocolitica virulence plasmid; the resistance loci in Java 9 are found on all four indigenous plasmids, including the two novel cryptic plasmids. This unique mobilome introduces more than 105 genes into the species gene pool. The majority of these are encoded by the two entirely novel self-transmissible plasmids, which show partial homology and synteny to other enterics. In contrast to the reductive evolution in Y. pestis, this study underlines the major impact of a dynamic mobilome and lateral acquisition in the genome evolution of the plague bacterium.

  3. Functional properties and structural requirements of the plasmid pMV158-encoded MobM relaxase domain.

    Science.gov (United States)

    Fernández-López, Cris; Pluta, Radoslaw; Pérez-Luque, Rosa; Rodríguez-González, Lorena; Espinosa, Manuel; Coll, Miquel; Lorenzo-Díaz, Fabián; Boer, D Roeland

    2013-07-01

    A crucial element in the horizontal transfer of mobilizable and conjugative plasmids is the relaxase, a single-stranded endonuclease that nicks the origin of transfer (oriT) of the plasmid DNA. The relaxase of the pMV158 mobilizable plasmid is MobM (494 residues). In solution, MobM forms a dimer through its C-terminal domain, which is proposed to anchor the protein to the cell membrane and to participate in type 4 secretion system (T4SS) protein-protein interactions. In order to gain a deeper insight into the structural MobM requirements for efficient DNA catalysis, we studied two endonuclease domain variants that include the first 199 or 243 amino acid residues (MobMN199 and MobMN243, respectively). Our results confirmed that the two proteins behaved as monomers in solution. Interestingly, MobMN243 relaxed supercoiled DNA and cleaved single-stranded oligonucleotides harboring oriTpMV158, whereas MobMN199 was active only on supercoiled DNA. Protein stability studies using gel electrophoresis and mass spectrometry showed increased susceptibility to degradation at the domain boundary between the N- and C-terminal domains, suggesting that the domains change their relative orientation upon DNA binding. Overall, these results demonstrate that MobMN243 is capable of nicking the DNA substrate independently of its topology and that the amino acids 200 to 243 modulate substrate specificity but not the nicking activity per se. These findings suggest that these amino acids are involved in positioning the DNA for the nuclease reaction rather than in the nicking mechanism itself.

  4. Salmonella Typhimurium ST213 is associated with two types of IncA/C plasmids carrying multiple resistance determinants.

    Science.gov (United States)

    Wiesner, Magdalena; Calva, Edmundo; Fernández-Mora, Marcos; Cevallos, Miguel A; Campos, Freddy; Zaidi, Mussaret B; Silva, Claudia

    2011-01-11

    Salmonella Typhimurium ST213 was first detected in the Mexican Typhimurium population in 2001. It is associated with a multi-drug resistance phenotype and a plasmid-borne blaCMY-2 gene conferring resistance to extended-spectrum cephalosporins. The objective of the current study was to examine the association between the ST213 genotype and blaCMY-2 plasmids. The blaCMY-2 gene was carried by an IncA/C plasmid. ST213 strains lacking the blaCMY-2 gene carried a different IncA/C plasmid. PCR analysis of seven DNA regions distributed throughout the plasmids showed that these IncA/C plasmids were related, but the presence and absence of DNA stretches produced two divergent types I and II. A class 1 integron (dfrA12, orfF and aadA2) was detected in most of the type I plasmids. Type I contained all the plasmids carrying the blaCMY-2 gene and a subset of plasmids lacking blaCMY-2. Type II included all of the remaining blaCMY-2-negative plasmids. A sequence comparison of the seven DNA regions showed that both types were closely related to IncA/C plasmids found in Escherichia, Salmonella, Yersinia, Photobacterium, Vibrio and Aeromonas. Analysis of our Typhimurium strains showed that the region containing the blaCMY-2 gene is inserted between traA and traC as a single copy, like in the E. coli plasmid pAR060302. The floR allele was identical to that of Newport pSN254, suggesting a mosaic pattern of ancestry with plasmids from other Salmonella serovars and E. coli. Only one of the tested strains was able to conjugate the IncA/C plasmid at very low frequencies (10-7 to 10-9). The lack of conjugation ability of our IncA/C plasmids agrees with the clonal dissemination trend suggested by the chromosomal backgrounds and plasmid pattern associations. The ecological success of the newly emerging Typhimurium ST213 genotype in Mexico may be related to the carriage of IncA/C plasmids. We conclude that types I and II of IncA/C plasmids originated from a common ancestor and that the

  5. Plasmid-mediated AmpC-type beta-lactamase isolated from Klebsiella pneumoniae confers resistance to broad-spectrum beta-lactams, including moxalactam.

    Science.gov (United States)

    Horii, T; Arakawa, Y; Ohta, M; Ichiyama, S; Wacharotayankun, R; Kato, N

    1993-01-01

    Klebsiella pneumoniae NU2936 was isolated from a patient and was found to produce a plasmid-encoded beta-lactamase (MOX-1) which conferred resistance to broad spectrum beta-lactams, including moxalactam, flomoxef, ceftizoxime, cefotaxime, and ceftazidime. Resistance could be transferred from K. pneumoniae NU2936 to Escherichia coli CSH2 by conjugation with a transfer frequency of 5 x 10(-7). The structural gene of MOX-1 (blaMOX-1) was cloned and expressed in E. coli HB101. The MIC of moxalactam for E. coli HB101 producing MOX-1 was > 512 micrograms/ml. The apparent molecular mass and pI of this enzyme were calculated to be 38 kDa and 8.9, respectively. Hg2+ and Cu2+ failed to block enzyme activity, and the presence of EDTA in the reaction buffer did not reduce the enzyme activity. However, clavulanate and cloxacillin, serine beta-lactamase inhibitors, inhibited the enzyme activity competitively (Kis = 5.60 and 0.35 microM, respectively). The kinetic study of MOX-1 suggested that it effectively hydrolyzed broad-spectrum beta-lactams. A hybridization study confirmed that blaMOX-1 is encoded on a large resident plasmid (pRMOX1; 180 kb) of strain NU2936. By deletion analysis, the functional region was localized within a 1.2-kb region of the plasmid. By amino acid sequencing, 18 of 33 amino acid residues at the N terminus of MOX-1 were found to be identical to those of Pseudomonas aeruginosa AmpC. These findings suggest that MOX-1 is a plasmid-mediated AmpC-type beta-lactamase that provides enteric bacteria resistance to broad-spectrum beta-lactams, including moxalactam. Images PMID:8517725

  6. A method of genetically engineering acidophilic, heterotrophic, bacteria by electroporation and conjugation

    Energy Technology Data Exchange (ETDEWEB)

    Roberto, F.F.; Glenn, A.W.; Ward, T.E.

    1990-08-07

    A method of genetically manipulating an acidophilic bacteria is provided by two different procedures. Using electroporation, chimeric and broad-host range plasmids are introduced into Acidiphilium. Conjugation is also employed to introduce broad-host range plasmids into Acidiphilium at neutral pH.

  7. Isolation and characterization of the pesticide-degrading plasmid pJP1 from Alcaligenes paradoxus

    International Nuclear Information System (INIS)

    Fisher, P.R.; Appleton, J.; Pemberton, J.M.

    1978-01-01

    A strain of Alcaligenes paradoxus, unable to degrade phenoxyacetic acid, was shown to degrade two synthetic derivatives of this molecule, the herbicides 2,4-dichlorophenoxyacetic acid and 2-methyl-4-chlorophenoxyacetic acid. The ability to degrade these pesticides is encoded by a 58-megadalton conjugal plasmid, pJP1

  8. Fluorescent strategy based on cationic conjugated polymer fluorescence resonance energy transfer for the quantification of 5-(hydroxymethyl)cytosine in genomic DNA.

    Science.gov (United States)

    Hong, Tingting; Wang, Tianlu; Guo, Pu; Xing, Xiwen; Ding, Fei; Chen, Yuqi; Wu, Jinjun; Ma, Jingwei; Wu, Fan; Zhou, Xiang

    2013-11-19

    DNA methylation is dynamically reprogrammed during early embryonic development in mammals. It can be explained partially by the discovery of 5-(hydroxymethyl)cytosine (5-hmC), 5-formylcytosine (5-fC), and 5-carboxylcytosine (5-caC), which are identified as key players involved in both active and passive demethylation pathways. As one of the ten-eleven translocation oxidation products, 5-hmC was found relatively abundant in neuron cells and embryonic stem cells. Herein we report a new method for 5-hmC quantification in genomic DNA based on CCP-FRET (cationic conjugated polymers act as the energy donor and induce fluorescence resonance energy transfer) assay combined with KRuO4 oxidation. 5-hmC in genomic DNA can be selectively transformed into 5-fC by the oxidation of KRuO4 and then labeled with hydroxylamine-BODIPY (BODIPY = 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene) fluorophore through the reaction between 5-fC and hydroxylamine-BODIPY. After the fluorescently labeled DNA was captured by CCP through electrostatic interactions, a significant FRET between CCP and hydroxylamine-BODIPY fluorophore was observed. This CCP-FRET-based assay benefits from light-harvesting, large Stokes shift, and optical signal amplification properties of the CCP. Furthermore, this CCP-FRET-based assay was quite successfully demonstrated for the 5-hmC quantification in three types of cells (mESc, HeLa, HEK 293T), providing a much more convenient choice for 5-hmC quantification in genomic DNA.

  9. Non-additive costs and interactions alter the competitive dynamics of co-occurring ecologically distinct plasmids.

    Science.gov (United States)

    Morton, Elise R; Platt, Thomas G; Fuqua, Clay; Bever, James D

    2014-03-22

    Plasmids play an important role in shaping bacterial evolution and adaptation to heterogeneous environments. As modular genetic elements that are often conjugative, the selective pressures that act on plasmid-borne genes are distinct from those that act on the chromosome. Many bacteria are co-infected by multiple plasmids that impart niche-specific phenotypes. Thus, in addition to host-plasmid dynamics, interactions between co-infecting plasmids are likely to be important drivers of plasmid population dynamics, evolution and ecology. Agrobacterium tumefaciens is a facultative plant pathogen that commonly harbours two distinct megaplasmids. Virulence depends on the presence of the tumour-inducing (Ti) plasmid, with benefits that are primarily restricted to the disease environment. Here, we demonstrate that a second megaplasmid, the At plasmid, confers a competitive advantage in the rhizosphere. To assess the individual and interactive costs of these plasmids, we generated four isogenic derivatives: plasmidless, pAt only, pTi only and pAtpTi, and performed pairwise competitions under carbon-limiting conditions. These studies reveal a low cost to the virulence plasmid when outside of the disease environment, and a strikingly high cost to the At plasmid. In addition, the costs of pAt and pTi in the same host were significantly lower than predicted based on single plasmid costs, signifying the first demonstration of non-additivity between naturally occurring co-resident plasmids. Based on these empirically demonstrated costs and benefits, we developed a resource-consumer model to generate predictions about the frequencies of these genotypes in relevant environments, showing that non-additivity between co-residing plasmids allows for their stable coexistence across environments.

  10. Plasmid and chromosome partitioning: surprises from phylogeny

    DEFF Research Database (Denmark)

    Gerdes, Kenn; Møller-Jensen, Jakob; Bugge Jensen, Rasmus

    2000-01-01

    Plasmids encode partitioning genes (par) that are required for faithful plasmid segregation at cell division. Initially, par loci were identified on plasmids, but more recently they were also found on bacterial chromosomes. We present here a phylogenetic analysis of par loci from plasmids and chr...

  11. Insights into dynamics of mobile genetic elements in hyperthermophilic environments from five new Thermococcus plasmids.

    Directory of Open Access Journals (Sweden)

    Mart Krupovic

    Full Text Available Mobilome of hyperthermophilic archaea dwelling in deep-sea hydrothermal vents is poorly characterized. To gain insight into genetic diversity and dynamics of mobile genetic elements in these environments we have sequenced five new plasmids from different Thermococcus strains that have been isolated from geographically remote hydrothermal vents. The plasmids were ascribed to two subfamilies, pTN2-like and pEXT9a-like. Gene content and phylogenetic analyses illuminated a robust connection between pTN2-like plasmids and Pyrococcus abyssi virus 1 (PAV1, with roughly half of the viral genome being composed of genes that have homologues in plasmids. Unexpectedly, pEXT9a-like plasmids were found to be closely related to the previously sequenced plasmid pMETVU01 from Methanocaldococcus vulcanius M7. Our data suggests that the latter observation is most compatible with an unprecedented horizontal transfer of a pEXT9a-like plasmid from Thermococcales to Methanococcales. Gene content analysis revealed that thermococcal plasmids encode Hfq-like proteins and toxin-antitoxin (TA systems of two different families, VapBC and RelBE. Notably, although abundant in archaeal genomes, to our knowledge, TA and hfq-like genes have not been previously found in archaeal plasmids or viruses. Finally, the plasmids described here might prove to be useful in developing new genetic tools for hyperthermophiles.

  12. Long- term manure exposure increases soil bacterial community potential for plasmid uptake

    DEFF Research Database (Denmark)

    Musovic, Sanin; Klümper, Uli; Dechesne, Arnaud

    2014-01-01

    Microbial communities derived from soils subject to different agronomic treatments were challenged with three broad host range plasmids, RP4, pIPO2tet and pRO101, via solid surface filter matings to assess their permissiveness. Approximately 1 in 10 000 soil bacterial cells could receive and main......Microbial communities derived from soils subject to different agronomic treatments were challenged with three broad host range plasmids, RP4, pIPO2tet and pRO101, via solid surface filter matings to assess their permissiveness. Approximately 1 in 10 000 soil bacterial cells could receive...... and maintain the plasmids. The community permissiveness increased up to 100% in communities derived from manured soil. While the plasmid transfer frequency was significantly influenced by both the type of plasmid and the agronomic treatment, the diversity of the transconjugal pools was purely plasmid dependent...

  13. Guanidinylated polyethyleneimine-polyoxypropylene-polyoxyethylene conjugates as gene transfection agents.

    Science.gov (United States)

    Bromberg, Lev; Raduyk, Svetlana; Hatton, T Alan; Concheiro, Angel; Rodriguez-Valencia, Cosme; Silva, Maite; Alvarez-Lorenzo, Carmen

    2009-05-20

    Conjugates of linear and branched polyethyleneimine (PEI) and monoamine polyether Jeffamine M-2070 (PO/EO mol ratio 10/31, 2000 Da) were synthesized through polyether activation by cyanuric chloride followed by attachment to PEI and guanidinylation by 1H-pyrazole-carboxamidine hydrochloride. The resulting guanidinylated PEI-polyether conjugates (termed gPEI-Jeffamine) efficiently complexed plasmid DNA, and their polyplexes possessed enhanced colloidal stability in the presence of serum proteins. In vitro studies with mammalian CHO-1, 3T3, and Cos-7 cell lines demonstrated improved transfection efficiency of the pCMVbeta-gal plasmid/gPEI-Jeffamine polyplexes. The guanidinylation of the amino groups of PEI and the conjugation of PEI with the Jeffamine polyether enhanced the conjugates' interaction with genetic material and reduced the cytotoxicity of the polyplexes in experiments with the L929 cell line.

  14. Bacteria isolated from pristine high altitude environments in the Argentinean Andean wetlands: plasmid profile and multiple antibiotic resistance

    International Nuclear Information System (INIS)

    Dib, J.R.; Martinez, M.A.; Sineriz, F.; Farias, M.E.

    2005-01-01

    Full text: Andean wetlands, placed in the North-Western Argentine at 4,600 m altitude, are attractive for both, environmental and biotechnology studies. Most of these wetlands are completely remote and inaccessible, having a high salinity and metal contents, a wide range of daily temperature changes, and an important intensity of solar UV-B radiation. Bacteria isolated from these environments were identified by 16SrDNA sequence and resulted in Gram-positive colored bacteria. Interesting features, to our knowledge never reported so far from bacteria isolates from these pristine high altitude lake-environments, such as similar plasmids profiles and multiple antibiotic resistances are the focus of this work. At least two plasmids were found in all isolates studied by using modifications of the alkaline Iysis method. Their preliminary characterization in this work includes size, incompatibility group through PCR, genetic transference to suitable hosts by transformation and conjugation, and studies of possible relationships of them with antibiotic resistances. (author)

  15. Imipenem-resistance in Serratia marcescens is mediated by plasmid expression of KPC-2.

    Science.gov (United States)

    Su, W-Q; Zhu, Y-Q; Deng, N-M; Li, L

    2017-04-01

    Imipenem is a broad-spectrum carbapenem antibiotic with applications against severe bacterial infections. Here, we describe the identification of imipenem-resistant Serratia marcescens in our hospital and the role of plasmid-mediated KPC-2 expression in imipenem resistance. We used the modified Hodge test to detect carbapenemase produced in imipenem-resistant strains. His resistance can be transferred to E. coli in co-culture tests, which implicates the plasmid in imipenem resistance. PCR amplification from the plasmid identified two products consistent with KPC-2 of 583 and 1050 bp that were also present in E. coli after co-culture. The restriction pattern for both plasmids was identical, supporting the transfer from the S. marcescens isolate to E. coli. Finally, gene sequencing confirmed KPC-2 in the plasmid. Due to the presence of KPC-2 in the imipenem-resistant S. marcescens, we propose that KPC-2 mediates antibiotic resistance in the S. marcescens isolate.

  16. Technology transfer

    International Nuclear Information System (INIS)

    1998-01-01

    On the base of technological opportunities and of the environmental target of the various sectors of energy system this paper intend to conjugate the opportunity/objective with economic and social development through technology transfer and information dissemination [it

  17. Functionalized tetrapod-like ZnO nanostructures for plasmid DNA purification, polymerase chain reaction and delivery

    International Nuclear Information System (INIS)

    Nie Leng; Gao Lizeng; Yan Xiyun; Wang Taihong

    2007-01-01

    Functionalized tetrapodal ZnO nanostructures are tested in plasmid DNA experiments (1) as a solid-phase adsorbent for plasmid DNA purification (2) as improving reagents in a polymerase chain reaction (PCR) and (3) as novel carriers for gene delivery. The amino-modification, the tetrapod-like shape of the nanostructure and its high biocompatibility all contribute to measurements showing promise for applications. A sol-gel method is used for silica coating and amino-modification. Plasmid DNA is purified through reversible conjugations of amino-modified ZnO tetrapods with DNA. Also, as additional reagents, functionalized tetrapods are shown to improve the amount of PCR product. For transfection, ZnO tetrapods provide some protection against deoxyribonuclease cleavage of plasmid DNA and deliver plasmid DNA into cells with little cytotoxicity

  18. Antimicrobial susceptibility pattern and plasmid-mediated ...

    African Journals Online (AJOL)

    negative Staphylococci (CoNS) were isolated from clinical samples and isolates subjected to antibiotic susceptibility testing, plasmid curing and plasmid DNA isolation. Result: The highest percentages isolates were recovered from urine samples and ...

  19. Plasmid-Mediated Quinolone Resistance in Shigella flexneri Isolated From Macaques

    Directory of Open Access Journals (Sweden)

    Anthony J. Mannion

    2018-03-01

    Full Text Available Non-human primates (NHPs for biomedical research are commonly infected with Shigella spp. that can cause acute dysentery or chronic episodic diarrhea. These animals are often prophylactically and clinically treated with quinolone antibiotics to eradicate these possible infections. However, chromosomally- and plasmid-mediated antibiotic resistance has become an emerging concern for species in the family Enterobacteriaceae. In this study, five individual isolates of multi-drug resistant Shigella flexneri were isolated from the feces of three macaques. Antibiotic susceptibility testing confirmed resistance or decreased susceptibility to ampicillin, amoxicillin-clavulanic acid, cephalosporins, gentamicin, tetracycline, ciprofloxacin, enrofloxacin, levofloxacin, and nalidixic acid. S. flexneri isolates were susceptible to trimethoprim-sulfamethoxazole, and this drug was used to eradicate infection in two of the macaques. Plasmid DNA from all isolates was positive for the plasmid-encoded quinolone resistance gene qnrS, but not qnrA and qnrB. Conjugation and transformation of plasmid DNA from several S. flexneri isolates into antibiotic-susceptible Escherichia coli strains conferred the recipients with resistance or decreased susceptibility to quinolones and beta-lactams. Genome sequencing of two representative S. flexneri isolates identified the qnrS gene on a plasmid-like contig. These contigs showed >99% homology to plasmid sequences previously characterized from quinolone-resistant Shigella flexneri 2a and Salmonella enterica strains. Other antibiotic resistance genes and virulence factor genes were also identified in chromosome and plasmid sequences in these genomes. The findings from this study indicate macaques harbor pathogenic S. flexneri strains with chromosomally- and plasmid-encoded antibiotic resistance genes. To our knowledge, this is the first report of plasmid-mediated quinolone resistance in S. flexneri isolated from NHPs and warrants

  20. Host-Specific Patterns of Genetic Diversity among IncI1-I gamma and IncK Plasmids Encoding CMY-2 beta-Lactamase in Escherichia coli Isolates from Humans, Poultry Meat, Poultry, and Dogs in Denmark

    DEFF Research Database (Denmark)

    Hansen, Katrine Hartung; Bortolaia, Valeria; Nielsen, Christine Ahl

    2016-01-01

    and commensal E. coli isolates collected from 2006 to 2012 from humans, retail poultry meat, broilers, and dogs. Multilocus sequence typing (MLST), antimicrobial susceptibility testing, and conjugation were performed in conjunction with plasmid replicon typing, plasmid multilocus sequence typing (p......MLST), restriction fragment length polymorphism (RFLP), and sequencing of selected bla(CMY-2)-harboring plasmids. MLST revealed high strain diversity, with few E. coli lineages occurring in multiple host species and sample types. bla(CMY-2) was detected on plasmids in 83 (89%) isolates. Most (75%) of the plasmids...... were conjugative and did not (96%) cotransfer resistance to antimicrobials other than cephalosporins. The main replicon types identified were IncI1-I gamma (55%) and IncK (39%). Isolates from different host species mainly carried distinct plasmid subtypes. Seven of the 18 human isolates harbored IncI1...

  1. Transfer

    DEFF Research Database (Denmark)

    Wahlgren, Bjarne; Aarkrog, Vibe

    Bogen er den første samlede indføring i transfer på dansk. Transfer kan anvendes som praksis-filosofikum. Den giver en systematisk indsigt til den studerende, der spørger: Hvordan kan teoretisk viden bruges til at reflektere over handlinger i situationer, der passer til min fremtidige arbejdsplads?...

  2. Two highly divergent lineages of exfoliative toxin B-encoding plasmids revealed in impetigo strains of Staphylococcus aureus.

    Science.gov (United States)

    Botka, Tibor; Růžičková, Vladislava; Svobodová, Karla; Pantůček, Roman; Petráš, Petr; Čejková, Darina; Doškař, Jiří

    2017-09-01

    Exfoliative toxin B (ETB) encoded by some large plasmids plays a crucial role in epidermolytic diseases caused by Staphylococcus aureus. We have found as yet unknown types of etb gene-positive plasmids isolated from a set of impetigo strains implicated in outbreaks of pemphigus neonatorum in Czech maternity hospitals. Plasmids from the strains of clonal complex CC121 were related to archetypal plasmid pETB TY4 . Sharing a 33-kb core sequence including virulence genes for ETB, EDIN C, and lantibiotics, they were assigned to a stand-alone lineage, named pETB TY4 -based plasmids. Differing from each other in the content of variable DNA regions, they formed four sequence types. In addition to them, a novel unique plasmid pETB608 isolated from a strain of ST130 was described. Carrying conjugative cluster genes, as well as new variants of etb and edinA genes, pETB608 could be regarded as a source of a new lineage of ETB plasmids. We have designed a helpful detection assay, which facilitates the precise identification of the all described types of ETB plasmids. Copyright © 2017 Elsevier GmbH. All rights reserved.

  3. Plasmid profiling of bacterial isolates from confined environments

    Science.gov (United States)

    van Houdt, Rob; Provoost, Ann; Coninx, Ilse; Leys, Natalie; Mergeay, Max

    Plasmid profiling of bacterial isolates from confined environments R. Van Houdt, I. Coninx, A. Provoost, N. Leys, and M. Mergeay Expertise group for Molecular and Cellular Biology, Institute for Environment, Health and Safety, Belgian Nuclear Research Centre (SCK•CEN), Boeretang 200, B-2400 Mol, Belgium. Human exploration of extreme and isolated hostile environments such as space requires special confined small volume habitats to protect and house the crew. However, human confinement in such small volume habitats has restrictions on waste disposal and personal hygiene and inevitably generates a particular community of microorganisms within the habitat. These microorganisms are mainly originating from the crew (skin, mucous membranes, upper respiratory tract, mouth, and gastrointestinal tract) but also include the residing environmental microorganisms. Earth-based confined habitats such as the Antarctic Research Station Concordia are used as test beds for long-duration spaceflights to study the physiologic and psychological adaptation to isolated environments. The dynamics of the environmental microbial population in such a test bed could render additional insights in assessing the potential health risks in long-duration space missions. Not only total bacterial contamination levels are important, but it is essential to identify also the predominant microbial taxa and their mobile genetic elements (MGE). These MGEs could be exchanged between bacteria by horizontal gene transfer and may alter the pathogenic potential since they often carry antibiotic resistance or more in general adaptation-enhancing traits. In this study several bacterial strains isolated in the Concordia research station were examined for their plasmid content. An optimized protocol for extraction of large plasmids showed the present of at least one plasmid in 50% of the strains. For all strains the minimal inhibitory concentration of a range of antibiotics was determined indicating resistance to

  4. Plasmid DNA Delivery: Nanotopography Matters.

    Science.gov (United States)

    Song, Hao; Yu, Meihua; Lu, Yao; Gu, Zhengying; Yang, Yannan; Zhang, Min; Fu, Jianye; Yu, Chengzhong

    2017-12-20

    Plasmid DNA molecules with unique loop structures have widespread bioapplications, in many cases relying heavily on delivery vehicles to introduce them into cells and achieve their functions. Herein, we demonstrate that control over delicate nanotopography of silica nanoparticles as plasmid DNA vectors has significant impact on the transfection efficacy. For silica nanoparticles with rambutan-, raspberry-, and flower-like morphologies composed of spike-, hemisphere-, and bowl-type subunit nanotopographies, respectively, the rambutan-like nanoparticles with spiky surfaces demonstrate the highest plasmid DNA binding capability and transfection efficacy of 88%, higher than those reported for silica-based nanovectors. Moreover, it is shown that the surface spikes of rambutan nanoparticles provide a continuous open space to bind DNA chains via multivalent interactions and protect the gene molecules sheltered in the spiky layer against nuclease degradation, exhibiting no significant transfection decay. This unique protection feature is in great contrast to a commercial transfection agent with similar transfection performance but poor protection capability against enzymatic cleavage. Our study provides new understandings in the rational design of nonviral vectors for efficient gene delivery.

  5. Giant linear plasmids in Streptomyces: a treasure trove of antibiotic biosynthetic clusters.

    Science.gov (United States)

    Kinashi, Haruyasu

    2011-01-01

    Many giant linear plasmids have been isolated from Streptomyces by using pulsed-field gel electrophoresis and some of them were found to carry an antibiotic biosynthetic cluster(s); SCP1 carries biosynthetic genes for methylenomycin, pSLA2-L for lankacidin and lankamycin, and pKSL for lasalocid and echinomycin. Accumulated data suggest that giant linear plasmids have played critical roles in genome evolution and horizontal transfer of secondary metabolism. In this review, I summarize typical examples of giant linear plasmids whose involvement in antibiotic production has been studied in some detail, emphasizing their finding processes and interaction with the host chromosomes. A hypothesis on horizontal transfer of secondary metabolism involving giant linear plasmids is proposed at the end.

  6. Genomic analysis of Acidianus hospitalis W1 a host for studying crenarchaeal virus and plasmid life cycles

    DEFF Research Database (Denmark)

    You, X. Y.; Liu, Chao; Wang, S. Y.

    2011-01-01

    The Acidianus hospitalis W1 genome consists of a minimally sized chromosome of about 2.13 Mb and a conjugative plasmid pAH1 and it is a host for the model filamentous lipothrixvirus AFV1. The chromosome carries three putative replication origins in conserved genomic regions and two large regions ...

  7. High prevalence of plasmid-mediated 16S rRNA methylase gene rmtB among Escherichia coli clinical isolates from a Chinese teaching hospital

    Directory of Open Access Journals (Sweden)

    Zhang Xue-qing

    2010-06-01

    Full Text Available Abstract Background Recently, production of 16S rRNA methylases by Gram-negative bacilli has emerged as a novel mechanism for high-level resistance to aminoglycosides by these organisms in a variety of geographic locations. Therefore, the spread of high-level aminoglycoside resistance determinants has become a great concern. Methods Between January 2006 and July 2008, 680 distinct Escherichia coli clinical isolates were collected from a teaching hospital in Wenzhou, China. PCR and DNA sequencing were used to identify 16S rRNA methylase and extended-spectrum β-lactamase (ESBL genes, including armA and rmtB, and in situ hybridization was performed to determine the location of 16S rRNA methylase genes. Conjugation experiments were subsequently performed to determine whether aminoglycoside resistance was transferable from the E. coli isolates via 16S rRNA methylase-bearing plasmids. Homology of the isolates harboring 16S rRNA methylase genes was determined using pulse-field gel electrophoresis (PFGE. Results Among the 680 E. coli isolates, 357 (52.5%, 346 (50.9% and 44 (6.5% isolates were resistant to gentamicin, tobramycin and amikacin, respectively. Thirty-seven of 44 amikacin-resistant isolates harbored 16S rRNA methylase genes, with 36 of 37 harboring the rmtB gene and only one harboring armA. The positive rates of 16S rRNA methylase genes among all isolates and amikacin-resistant isolates were 5.4% (37/680 and 84.1% (37/44, respectively. Thirty-one isolates harboring 16S rRNA methylase genes also produced ESBLs. In addition, high-level aminoglycoside resistance could be transferred by conjugation from four rmtB-positive donors. The plasmids of incompatibility groups IncF, IncK and IncN were detected in 34, 3 and 3 isolates, respectively. Upstream regions of the armA gene contained ISCR1 and tnpU, the latter a putative transposase gene,. Another putative transposase gene, tnpD, was located within a region downstream of armA. Moreover, a

  8. Transfer of DNA from Bacteria to Eukaryotes

    Directory of Open Access Journals (Sweden)

    Benoît Lacroix

    2016-07-01

    Full Text Available Historically, the members of the Agrobacterium genus have been considered the only bacterial species naturally able to transfer and integrate DNA into the genomes of their eukaryotic hosts. Yet, increasing evidence suggests that this ability to genetically transform eukaryotic host cells might be more widespread in the bacterial world. Indeed, analyses of accumulating genomic data reveal cases of horizontal gene transfer from bacteria to eukaryotes and suggest that it represents a significant force in adaptive evolution of eukaryotic species. Specifically, recent reports indicate that bacteria other than Agrobacterium, such as Bartonella henselae (a zoonotic pathogen, Rhizobium etli (a plant-symbiotic bacterium related to Agrobacterium, or even Escherichia coli, have the ability to genetically transform their host cells under laboratory conditions. This DNA transfer relies on type IV secretion systems (T4SSs, the molecular machines that transport macromolecules during conjugative plasmid transfer and also during transport of proteins and/or DNA to the eukaryotic recipient cells. In this review article, we explore the extent of possible transfer of genetic information from bacteria to eukaryotic cells as well as the evolutionary implications and potential applications of this transfer.

  9. Protein Structure Initiative Material Repository: an open shared public resource of structural genomics plasmids for the biological community

    Science.gov (United States)

    Cormier, Catherine Y.; Mohr, Stephanie E.; Zuo, Dongmei; Hu, Yanhui; Rolfs, Andreas; Kramer, Jason; Taycher, Elena; Kelley, Fontina; Fiacco, Michael; Turnbull, Greggory; LaBaer, Joshua

    2010-01-01

    The Protein Structure Initiative Material Repository (PSI-MR; http://psimr.asu.edu) provides centralized storage and distribution for the protein expression plasmids created by PSI researchers. These plasmids are a resource that allows the research community to dissect the biological function of proteins whose structures have been identified by the PSI. The plasmid annotation, which includes the full length sequence, vector information and associated publications, is stored in a freely available, searchable database called DNASU (http://dnasu.asu.edu). Each PSI plasmid is also linked to a variety of additional resources, which facilitates cross-referencing of a particular plasmid to protein annotations and experimental data. Plasmid samples can be requested directly through the website. We have also developed a novel strategy to avoid the most common concern encountered when distributing plasmids namely, the complexity of material transfer agreement (MTA) processing and the resulting delays this causes. The Expedited Process MTA, in which we created a network of institutions that agree to the terms of transfer in advance of a material request, eliminates these delays. Our hope is that by creating a repository of expression-ready plasmids and expediting the process for receiving these plasmids, we will help accelerate the accessibility and pace of scientific discovery. PMID:19906724

  10. Comparative Sequence Analysis of Multidrug-Resistant IncA/C Plasmids from Salmonella enterica.

    Science.gov (United States)

    Hoffmann, Maria; Pettengill, James B; Gonzalez-Escalona, Narjol; Miller, John; Ayers, Sherry L; Zhao, Shaohua; Allard, Marc W; McDermott, Patrick F; Brown, Eric W; Monday, Steven R

    2017-01-01

    Determinants of multidrug resistance (MDR) are often encoded on mobile elements, such as plasmids, transposons, and integrons, which have the potential to transfer among foodborne pathogens, as well as to other virulent pathogens, increasing the threats these traits pose to human and veterinary health. Our understanding of MDR among Salmonella has been limited by the lack of closed plasmid genomes for comparisons across resistance phenotypes, due to difficulties in effectively separating the DNA of these high-molecular weight, low-copy-number plasmids from chromosomal DNA. To resolve this problem, we demonstrate an efficient protocol for isolating, sequencing and closing IncA/C plasmids from Salmonella sp. using single molecule real-time sequencing on a Pacific Biosciences (Pacbio) RS II Sequencer. We obtained six Salmonella enterica isolates from poultry, representing six different serovars, each exhibiting the MDR-Ampc resistance profile. Salmonella plasmids were obtained using a modified mini preparation and transformed with Escherichia coli DH10Br. A Qiagen Large-Construct kit™ was used to recover highly concentrated and purified plasmid DNA that was sequenced using PacBio technology. These six closed IncA/C plasmids ranged in size from 104 to 191 kb and shared a stable, conserved backbone containing 98 core genes, with only six differences among those core genes. The plasmids encoded a number of antimicrobial resistance genes, including those for quaternary ammonium compounds and mercury. We then compared our six IncA/C plasmid sequences: first with 14 IncA/C plasmids derived from S. enterica available at the National Center for Biotechnology Information (NCBI), and then with an additional 38 IncA/C plasmids derived from different taxa. These comparisons allowed us to build an evolutionary picture of how antimicrobial resistance may be mediated by this common plasmid backbone. Our project provides detailed genetic information about resistance genes in

  11. Comparative Sequence Analysis of Multidrug-Resistant IncA/C Plasmids from Salmonella enterica

    Directory of Open Access Journals (Sweden)

    Maria Hoffmann

    2017-08-01

    Full Text Available Determinants of multidrug resistance (MDR are often encoded on mobile elements, such as plasmids, transposons, and integrons, which have the potential to transfer among foodborne pathogens, as well as to other virulent pathogens, increasing the threats these traits pose to human and veterinary health. Our understanding of MDR among Salmonella has been limited by the lack of closed plasmid genomes for comparisons across resistance phenotypes, due to difficulties in effectively separating the DNA of these high-molecular weight, low-copy-number plasmids from chromosomal DNA. To resolve this problem, we demonstrate an efficient protocol for isolating, sequencing and closing IncA/C plasmids from Salmonella sp. using single molecule real-time sequencing on a Pacific Biosciences (Pacbio RS II Sequencer. We obtained six Salmonella enterica isolates from poultry, representing six different serovars, each exhibiting the MDR-Ampc resistance profile. Salmonella plasmids were obtained using a modified mini preparation and transformed with Escherichia coli DH10Br. A Qiagen Large-Construct kit™ was used to recover highly concentrated and purified plasmid DNA that was sequenced using PacBio technology. These six closed IncA/C plasmids ranged in size from 104 to 191 kb and shared a stable, conserved backbone containing 98 core genes, with only six differences among those core genes. The plasmids encoded a number of antimicrobial resistance genes, including those for quaternary ammonium compounds and mercury. We then compared our six IncA/C plasmid sequences: first with 14 IncA/C plasmids derived from S. enterica available at the National Center for Biotechnology Information (NCBI, and then with an additional 38 IncA/C plasmids derived from different taxa. These comparisons allowed us to build an evolutionary picture of how antimicrobial resistance may be mediated by this common plasmid backbone. Our project provides detailed genetic information about

  12. Frequency and diversity of small cryptic plasmids in the genus Rahnella

    Directory of Open Access Journals (Sweden)

    Summers David K

    2010-02-01

    importance of plasmids for lateral gene transfer (including chromosomal sequences to distinct genera.

  13. Comparative genomics of the pIPO2/pSB102 family of environmental plasmids : sequence, evolution, and ecology of pTer331 isolated from Collimonas fungivorans Ter331

    NARCIS (Netherlands)

    Mela, Francesca; Fritsche, Kathrin; Boersma, Hidde; van Elsas, Jan D.; Bartels, Daniela; Meyer, Folker; de Boer, Wietse; van Veen, Johannes A.; Leveau, Johan H. J.

    2008-01-01

    Plasmid pTer331 from the bacterium Collimonas fungivorans Ter331 is a new member of the pIPO2/pSB102 family of environmental plasmids. The 40 457-bp sequence of pTer331 codes for 44 putative ORFs, most of which represent genes involved in replication, partitioning and transfer of the plasmid. We

  14. TRANSFER

    African Journals Online (AJOL)

    This paper reports on further studies on long range energy transfer between curcumine as donor and another thiazine dye, thionine, which is closely related to methylene blue as energy harvester (Figure 1). Since thionine is known to have a higher quantum yield of singlet oxygen sensitization than methylene blue [8], it is ...

  15. The sudden dominance of blaCTX-M harbouring plasmids in Shigella spp. Circulating in Southern Vietnam.

    Directory of Open Access Journals (Sweden)

    Nhu Thi Khanh Nguyen

    2010-06-01

    Full Text Available Plasmid mediated antimicrobial resistance in the Enterobacteriaceae is a global problem. The rise of CTX-M class extended spectrum beta lactamases (ESBLs has been well documented in industrialized countries. Vietnam is representative of a typical transitional middle income country where the spectrum of infectious diseases combined with the spread of drug resistance is shifting and bringing new healthcare challenges.We collected hospital admission data from the pediatric population attending the hospital for tropical diseases in Ho Chi Minh City with Shigella infections. Organisms were cultured from all enrolled patients and subjected to antimicrobial susceptibility testing. Those that were ESBL positive were subjected to further investigation. These investigations included PCR amplification for common ESBL genes, plasmid investigation, conjugation, microarray hybridization and DNA sequencing of a bla(CTX-M encoding plasmid.We show that two different bla(CTX-M genes are circulating in this bacterial population in this location. Sequence of one of the ESBL plasmids shows that rather than the gene being integrated into a preexisting MDR plasmid, the bla(CTX-M gene is located on relatively simple conjugative plasmid. The sequenced plasmid (pEG356 carried the bla(CTX-M-24 gene on an ISEcp1 element and demonstrated considerable sequence homology with other IncFI plasmids.The rapid dissemination, spread of antimicrobial resistance and changing population of Shigella spp. concurrent with economic growth are pertinent to many other countries undergoing similar development. Third generation cephalosporins are commonly used empiric antibiotics in Ho Chi Minh City. We recommend that these agents should not be considered for therapy of dysentery in this setting.

  16. Modelling conjugation with stochastic differential equations

    DEFF Research Database (Denmark)

    Philipsen, Kirsten Riber; Christiansen, Lasse Engbo; Hasman, Henrik

    2010-01-01

    Enterococcus faecium strains in a rich exhaustible media. The model contains a new expression for a substrate dependent conjugation rate. A maximum likelihood based method is used to estimate the model parameters. Different models including different noise structure for the system and observations are compared......Conjugation is an important mechanism involved in the transfer of resistance between bacteria. In this article a stochastic differential equation based model consisting of a continuous time state equation and a discrete time measurement equation is introduced to model growth and conjugation of two...... using a likelihood-ratio test and Akaike's information criterion. Experiments indicating conjugation on the agar plates selecting for transconjugants motivates the introduction of an extended model, for which conjugation on the agar plate is described in the measurement equation. This model is compared...

  17. Quantitative assessment of the role played by plasmid R446b in the overall U. V. response in EscHer-ICHIA COLI ymel

    Energy Technology Data Exchange (ETDEWEB)

    Farhan, A A

    1987-01-01

    The present study concerned with the quantitative assessment of the role of plasmid R446b in the overall U.V. response in E.Coli ymel. The plasmid was, therefore, transferred to the strain by mating. The transfer of the plasmid has no influence on the microbioloicaLl characteristics evaluated from growth profiles under the rigorously controlled experimental conditions. On the other hand, the plasmid reduced the typical U.V. response of mor than 40 as measured by inactivation constant values to the dose in survival curves. (19 figs., 4 tabs., 99 refs.).

  18. 蛋白涂层支架携带一氧化氮合酶基因转染%The Feasibility of Local Plasmid-Mediated Human Inducible Nitri c Oxide Synthase Gene Transfer to Coronary Artery with Protein-Coated Metallic Stents in Mini-Swine Model

    Institute of Scientific and Technical Information of China (English)

    戴军; 高润霖; 汤健; 宋来凤; 魏英杰; 宋莉; 李永利; 唐承君; 史瑞文

    2001-01-01

    Objective:To assess the feasibility of plasmid-medicated local transfer of inducible nitric oxide synthase (iNOS) gene to coronary artery using protein-co ated metallic stents in mini-swine medel.   Methods:The metallic stent was coated by cross-linked gelatin and mounted o n 3.0 mm Percutaneous transluminal coronary angioplasty (PTCA) balloon,then End otoxin-free ultrapure Endotoxin-free ultrapure Plasmid pcDNA3hepiNOS under the control of the comegalovirus (CMV) promoter/enhancer was absorbed on the stent. Protein-coated stainless steel stents were used as controls.All stents were imp lanted into the middle segment of left anterior descending artery through 7F lar ge luman gui ding catheter.(The ratio of balloon to vessel diameter was 1.1-1.3∶1).Total RNA of stented segment of coronary artery was extracted,its reverse transeriptio n polymerase chain reaction (RT-PCR) and immunohistochemical staining were perf ormed through routine methods.   Results:At the 7th day after stenting,RT-PCR and immunohistochemical staining confirmed the expression of plasmid pcDNA3hepiNOS mRNA and presence of its protein at gene transered vessels (n=2) but there was no expression in remote organs.Endothelialization of the vessel was observed in all animals throu gh scanning electromicroscopy.   Conclusions:Local plasmid-mediated human inducible nitric oxide synthase ge ne transfer to coronary artery with protein-coated metallic stents is feasible in mini-swine model.%目的:探索蛋白涂层支架携带质粒介导人肝脏诱导型一氧化氮合酶(iNOS)基因转染小型猪冠状动脉可行性。 方法:使用蛋白支架吸附去内毒素纯化质粒,以常规支架置入技术置入小型猪冠状动脉前降支中段。置入后第7天取出前降支置入段,分别提取总核糖核酸(RAN)并进行逆向多聚酶链反应(RT-PCR),免疫组化染色检测导入人肝脏iNOS蛋白的表达。 结果:小型猪前降支置入支架处显示人iNOS基

  19. Reconstructing the complex evolutionary history of mobile plasmids in red algal genomes

    Science.gov (United States)

    Lee, JunMo; Kim, Kyeong Mi; Yang, Eun Chan; Miller, Kathy Ann; Boo, Sung Min; Bhattacharya, Debashish; Yoon, Hwan Su

    2016-01-01

    The integration of foreign DNA into algal and plant plastid genomes is a rare event, with only a few known examples of horizontal gene transfer (HGT). Plasmids, which are well-studied drivers of HGT in prokaryotes, have been reported previously in red algae (Rhodophyta). However, the distribution of these mobile DNA elements and their sites of integration into the plastid (ptDNA), mitochondrial (mtDNA), and nuclear genomes of Rhodophyta remain unknown. Here we reconstructed the complex evolutionary history of plasmid-derived DNAs in red algae. Comparative analysis of 21 rhodophyte ptDNAs, including new genome data for 5 species, turned up 22 plasmid-derived open reading frames (ORFs) that showed syntenic and copy number variation among species, but were conserved within different individuals in three lineages. Several plasmid-derived homologs were found not only in ptDNA but also in mtDNA and in the nuclear genome of green plants, stramenopiles, and rhizarians. Phylogenetic and plasmid-derived ORF analyses showed that the majority of plasmid DNAs originated within red algae, whereas others were derived from cyanobacteria, other bacteria, and viruses. Our results elucidate the evolution of plasmid DNAs in red algae and suggest that they spread as parasitic genetic elements. This hypothesis is consistent with their sporadic distribution within Rhodophyta. PMID:27030297

  20. Characterization of Endogenous Plasmids from Lactobacillus salivarius UCC118▿ †

    Science.gov (United States)

    Fang, Fang; Flynn, Sarah; Li, Yin; Claesson, Marcus J.; van Pijkeren, Jan-Peter; Collins, J. Kevin; van Sinderen, Douwe; O'Toole, Paul W.

    2008-01-01

    The genome of Lactobacillus salivarius UCC118 comprises a 1.83-Mb chromosome, a 242-kb megaplasmid (pMP118), and two smaller plasmids of 20 kb (pSF118-20) and 44 kb (pSF118-44). Annotation and bioinformatic analyses suggest that both of the smaller plasmids replicate by a theta replication mechanism. Furthermore, it appears that they are transmissible, although neither possesses a complete set of conjugation genes. Plasmid pSF118-20 encodes a toxin-antitoxin system composed of pemI and pemK homologs, and this plasmid could be cured when PemI was produced in trans. The minimal replicon of pSF118-20 was determined by deletion analysis. Shuttle vector derivatives of pSF118-20 were generated that included the replication region (pLS203) and the replication region plus mobilization genes (pLS208). The plasmid pLS203 was stably maintained without selection in Lactobacillus plantarum, Lactobacillus fermentum, and the pSF118-20-cured derivative strain of L. salivarius UCC118 (strain LS201). Cloning in pLS203 of genes encoding luciferase and green fluorescent protein, and expression from a constitutive L. salivarius promoter, demonstrated the utility of this vector for the expression of heterologous genes in Lactobacillus. This study thus expands the knowledge base and vector repertoire of probiotic lactobacilli. PMID:18390685

  1. Photoinduced electron transfer involving eosin-tryptophan conjugates. Long-lived radical pair states for systems incorporating aromatic amino acid side chains

    Energy Technology Data Exchange (ETDEWEB)

    Jones, G. II; Farahat, C.W.; Oh, C. (Boston Univ., MA (United States))

    1994-07-14

    The electron-transfer photochemistry of the covalent derivatives of the dye eosin, in which the xanthene dye is covalently attached to the amino acid L-tryptophan via the thiohydantoin derivative, the tryptophan dipeptide, and an ethyl ester derivative, has been investigated. The singlet excited state of the dye is significantly quenched on attachment of the aromatic amino acid residue. Dye triplet states are also intercepted through intramolecular interaction of excited dye and amino acid pendants. Flash photolysis experiments verify that this interaction involves electron transfer from the indole side chains of tryptophan. Rate constants for electron transfer are discussed in terms of the distance relationships for the eosin chromophore and aromatic redox sites on peptide derivatives, the pathway for [sigma]-[pi] through-bond interaction between redox sites, and the multiplicity and state of protonation for electron-transfer intermediates. Selected electron-transfer photoreactions were studied under conditions of binding of the peptide derivatives in a high molecular weight, water-soluble, globular polymer, poly(vinyl-2-pyrrolidinone). 28 refs., 4 figs., 1 tab.

  2. Influence of tra genes of IncP and F plasmids on the mobilization of small Kanamycin resistance ColE1-Like plasmids in bacterial biofilms

    Science.gov (United States)

    Background: Horizontal gene transfer is a mechanism for movement of antibiotic resistance genes among bacteria. Some small kanamycin resistance (KanR) ColE1-like plasmids isolated from different serotypes of Salmonella enterica were shown to carry mobilization genes; although not self-transmissibl...

  3. Realization of large area flexible fullerene - conjugated polymer photocells: a route to plastic solar cells

    NARCIS (Netherlands)

    Brabec, C.J.; Padinger, F.; Hummelen, J.C.; Janssen, R.A.J.; Sariciftci, N.S.

    1999-01-01

    Bulk donor — acceptor heterojunctions between conjugated polymers and fullerenes have been utilized for photovoltaic devices with quantum efficiencies of around 1%. These devices are based on the photoinduced, ultrafast electron transfer between non degenerate ground state conjugated polymers and

  4. Large-scale preparation of plasmid DNA.

    Science.gov (United States)

    Heilig, J S; Elbing, K L; Brent, R

    2001-05-01

    Although the need for large quantities of plasmid DNA has diminished as techniques for manipulating small quantities of DNA have improved, occasionally large amounts of high-quality plasmid DNA are desired. This unit describes the preparation of milligram quantities of highly purified plasmid DNA. The first part of the unit describes three methods for preparing crude lysates enriched in plasmid DNA from bacterial cells grown in liquid culture: alkaline lysis, boiling, and Triton lysis. The second part describes four methods for purifying plasmid DNA in such lysates away from contaminating RNA and protein: CsCl/ethidium bromide density gradient centrifugation, polyethylene glycol (PEG) precipitation, anion-exchange chromatography, and size-exclusion chromatography.

  5. Group II intron inhibits conjugative relaxase expression in bacteria by mRNA targeting

    Science.gov (United States)

    Piazza, Carol Lyn; Smith, Dorie

    2018-01-01

    Group II introns are mobile ribozymes that are rare in bacterial genomes, often cohabiting with various mobile elements, and seldom interrupting housekeeping genes. What accounts for this distribution has not been well understood. Here, we demonstrate that Ll.LtrB, the group II intron residing in a relaxase gene on a conjugative plasmid from Lactococcus lactis, inhibits its host gene expression and restrains the naturally cohabiting mobile element from conjugative horizontal transfer. We show that reduction in gene expression is mainly at the mRNA level, and results from the interaction between exon-binding sequences (EBSs) in the intron and intron-binding sequences (IBSs) in the mRNA. The spliced intron targets the relaxase mRNA and reopens ligated exons, causing major mRNA loss. Taken together, this study provides an explanation for the distribution and paucity of group II introns in bacteria, and suggests a potential force for those introns to evolve into spliceosomal introns. PMID:29905149

  6. Plasmid Complement of Lactococcus lactis NCDO712 Reveals a Novel Pilus Gene Cluster.

    Science.gov (United States)

    Tarazanova, Mariya; Beerthuyzen, Marke; Siezen, Roland; Fernandez-Gutierrez, Marcela M; de Jong, Anne; van der Meulen, Sjoerd; Kok, Jan; Bachmann, Herwig

    2016-01-01

    Lactococcus lactis MG1363 is an important gram-positive model organism. It is a plasmid-free and phage-cured derivative of strain NCDO712. Plasmid-cured strains facilitate studies on molecular biological aspects, but many properties which make L. lactis an important organism in the dairy industry are plasmid encoded. We sequenced the total DNA of strain NCDO712 and, contrary to earlier reports, revealed that the strain carries 6 rather than 5 plasmids. A new 50-kb plasmid, designated pNZ712, encodes functional nisin immunity (nisCIP) and copper resistance (lcoRSABC). The copper resistance could be used as a marker for the conjugation of pNZ712 to L. lactis MG1614. A genome comparison with the plasmid cured daughter strain MG1363 showed that the number of single nucleotide polymorphisms that accumulated in the laboratory since the strains diverted more than 30 years ago is limited to 11 of which only 5 lead to amino acid changes. The 16-kb plasmid pSH74 was found to contain a novel 8-kb pilus gene cluster spaCB-spaA-srtC1-srtC2, which is predicted to encode a pilin tip protein SpaC, a pilus basal subunit SpaB, and a pilus backbone protein SpaA. The sortases SrtC1/SrtC2 are most likely involved in pilus polymerization while the chromosomally encoded SrtA could act to anchor the pilus to peptidoglycan in the cell wall. Overexpression of the pilus gene cluster from a multi-copy plasmid in L. lactis MG1363 resulted in cell chaining, aggregation, rapid sedimentation and increased conjugation efficiency of the cells. Electron microscopy showed that the over-expression of the pilus gene cluster leads to appendices on the cell surfaces. A deletion of the gene encoding the putative basal protein spaB, by truncating spaCB, led to more pilus-like structures on the cell surface, but cell aggregation and cell chaining were no longer observed. This is consistent with the prediction that spaB is involved in the anchoring of the pili to the cell.

  7. [Replication of Streptomyces plasmids: the DNA nucleotide sequence of plasmid pSB 24.2].

    Science.gov (United States)

    Bolotin, A P; Sorokin, A V; Aleksandrov, N N; Danilenko, V N; Kozlov, Iu I

    1985-11-01

    The nucleotide sequence of DNA in plasmid pSB 24.2, a natural deletion derivative of plasmid pSB 24.1 isolated from S. cyanogenus was studied. The plasmid amounted by its size to 3706 nucleotide pairs. The G-C composition was equal to 73 per cent. The analysis of the DNA structure in plasmid pSB 24.2 revealed the protein-encoding sequence of DNA, the continuity of which was significant for replication of the plasmid containing more than 1300 nucleotide pairs. The analysis also revealed two A-T-rich areas of DNA, the G-C composition of which was less than 55 per cent and a DNA area with a branched pin structure. The results may be of value in investigation of plasmid replication in actinomycetes and experimental cloning of DNA with this plasmid as a vector.

  8. DNA-binding proteins regulating pIP501 transfer and replication

    Directory of Open Access Journals (Sweden)

    Elisabeth Grohmann

    2016-08-01

    Full Text Available pIP501 is a Gram-positive broad-host-range model plasmid intensively used for studying plasmid replication and conjugative transfer. It is a multiple antibiotic resistance plasmid frequently found in clinical Enterococcus faecalis and Enterococcus faecium isolates. Replication of pIP501 proceeds unidirectionally by a theta mechanism. The minimal replicon of pIP501 is composed of the repR gene encoding the essential rate-limiting replication initiator protein RepR and the origin of replication, oriR, located downstream of repR. RepR is similar to RepE of related streptococcal plasmid pAMβ1, which has been shown to possess RNase activity cleaving free RNA molecules in close proximity of the initiation site of DNA synthesis. Replication of pIP501 is controlled by the concerted action of a small protein, CopR, and an antisense RNA, RNAIII. CopR has a dual role: It acts as transcriptional repressor at the repR promoter and prevents convergent transcription of RNAIII and repR mRNA (RNAII, thereby indirectly increasing RNAIII synthesis. CopR binds asymmetrically as a dimer at two consecutive binding sites upstream of and overlapping with the repR promoter. RNAIII induces transcriptional attenuation within the leader region of the repR mRNA (RNAII. Deletion of either control component causes a 10- to 20-fold increase of plasmid copy number, while simultaneous deletions have no additional effect. Conjugative transfer of pIP501 depends on a type IV secretion system (T4SS encoded in a single operon. Its transfer host-range is considerably broad, as it has been transferred to virtually all Gram-positive bacteria including filamentous streptomycetes and even the Gram-negative Escherichia coli. Expression of the 15 genes encoding the T4SS is tightly controlled by binding of the relaxase TraA, the transfer initiator protein, to the operon promoter, which overlaps with the origin of transfer (oriT. The T4SS operon encodes the DNA-binding proteins TraJ (VirD4

  9. Damage of plasmid DNA by high energy ions

    International Nuclear Information System (INIS)

    Michaelidesova, A.; Pachnerova Brabcova, K.; Davidkova, M.

    2018-01-01

    The aim of the study was to determine the degree of direct DNA damage by high-energy ions, which are one of the components of cosmic rays, and therefore the knowledge of the biological effects of these ions is key to long-term space missions with human crew. The pBR322 plasmid containing 4361 base pairs was used in this study. The aqueous solution of plasmid pBR322 was transferred on ice to Japan to the Heavy Ion Medical Accelerator in Chiba, the Research Center for Charged Particle Therapy. Just before the experiment, the droplets of solution of known concentration were applied to the slides and the water was allowed to evaporate to produce dry DNA samples. Half of the slides were irradiated with 290 MeV/u of carbon ions and a dose rate of 20 Gy/min. The other half of the slides were irradiated with helium nuclei of 150 MeV/hr and a dose rate of 12.6 Gy/min. Both sets of slides were irradiated with doses of 0-1,400 Gy with a 200 Gy step. After irradiation, the samples were re-dissolved in distilled water, frozen and transported on ice to the Czech Republic for processing. Samples were analyzed by agarose gel electrophoresis. The plasmid was evaluated separately to determine the degree of radiation induced lesions and further to incubation with enzymes recognizing basal damage. (authors)

  10. 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.

  11. 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.

  12. Plasmid fingerprinting and virulence gene detection among indigenous strains of salmonella enterica serovar enteritidis

    International Nuclear Information System (INIS)

    Sajid, S.U.; Schwarz, S.

    2009-01-01

    Salmonella enterica serovar Enteritidis is an important frequently reported zoonotic pathogen and a common cause of human gastroenteritis worldwide. The highly conserved Serospecific plasmids (SSPs) and Salmonella plasmid virulence (Spv) genes have been shown to mediate extra-intestinal colonization and systemic infection. The objective of current study was to document the presence of SSPs and SpvB/SpvC genes prevailing in the indigenous population of serovar Enteritidis. A total of 48 epidemiologically unrelated strains of Salmonella enteritidis were included in the study. Preparation of plasmids DNA suitable for endonuclease digestion and separation of respective fragments by agarose gel electrophoresis followed previously described protocols. The plasmids of Escherichia coli V517, 1-kbp ladder, and lambda DNA HindIII fragments served as DNA size standards. Transfer of DNA fragments from agarose gels to nitrocellulose membranes was achieved by capillary blot procedure. An ECL labeled 3.6 kbp HindIII fragment of plasmid PRQ 51 was used as probe for SpvB/SpvC gene detection. Plasmid DNA fingerprinting revealed the presence of two different profiles of approximately 55 kbp and 90 kbp and were identified as virulence plasmids by DNA hybridization. The SpvB/SpvC genes were located on HindIII fragments of 3.6 kbp in each of the two types of virulence plasmids. The study confirms the presence of SSPs and SpvB/SpvC genes in indigenous strains of S. enteritidis isolated from Northern Punjab area of Pakistan and substantiate the previous data on such findings from other parts of the world. (author)

  13. Interaction and energy transfer studies between bovine serum albumin and CdTe quantum dots conjugates: CdTe QDs as energy acceptor probes.

    Science.gov (United States)

    Kotresh, M G; Inamdar, L S; Shivkumar, M A; Adarsh, K S; Jagatap, B N; Mulimani, B G; Advirao, G M; Inamdar, S R

    2017-06-01

    In this paper, a systematic investigation of the interaction of bovine serum albumin (BSA) with water-soluble CdTe quantum dots (QDs) of two different sizes capped with carboxylic thiols is presented based on steady-state and time-resolved fluorescence measurements. Efficient Förster resonance energy transfer (FRET) was observed to occur from BSA donor to CdTe acceptor as noted from reduction in the fluorescence of BSA and enhanced fluorescence from CdTe QDs. FRET parameters such as Förster distance, spectral overlap integral, FRET rate constant and efficiency were determined. The quenching of BSA fluorescence in aqueous solution observed in the presence of CdTe QDs infers that fluorescence resonance energy transfer is primarily responsible for the quenching phenomenon. Bimolecular quenching constant (k q ) determined at different temperatures and the time-resolved fluorescence data provide additional evidence for this. The binding stoichiometry and various thermodynamic parameters are evaluated by using the van 't Hoff equation. The analysis of the results suggests that the interaction between BSA and CdTe QDs is entropy driven and hydrophobic forces play a key role in the interaction. Binding of QDs significantly shortened the fluorescence lifetime of BSA which is one of the hallmarks of FRET. The effect of size of the QDs on the FRET parameters are discussed in the light of FRET parameters obtained. Copyright © 2016 John Wiley & Sons, Ltd.

  14. Comparative symbiotic plasmid analysis indicates that symbiosis gene ancestor type affects plasmid genetic evolution.

    Science.gov (United States)

    Wang, X; Zhao, L; Zhang, L; Wu, Y; Chou, M; Wei, G

    2018-07-01

    Rhizobial symbiotic plasmids play vital roles in mutualistic symbiosis with legume plants by executing the functions of nodulation and nitrogen fixation. To explore the gene composition and genetic constitution of rhizobial symbiotic plasmids, comparison analyses of 24 rhizobial symbiotic plasmids derived from four rhizobial genera was carried out. Results illustrated that rhizobial symbiotic plasmids had higher proportion of functional genes participating in amino acid transport and metabolism, replication; recombination and repair; carbohydrate transport and metabolism; energy production and conversion and transcription. Mesorhizobium amorphae CCNWGS0123 symbiotic plasmid - pM0123d had similar gene composition with pR899b and pSNGR234a. All symbiotic plasmids shared 13 orthologous genes, including five nod and eight nif/fix genes which participate in the rhizobia-legume symbiosis process. These plasmids contained nod genes from four ancestors and fix genes from six ancestors. The ancestral type of pM0123d nod genes was similar with that of Rhizobium etli plasmids, while the ancestral type of pM0123d fix genes was same as that of pM7653Rb. The phylogenetic trees constructed based on nodCIJ and fixABC displayed different topological structures mainly due to nodCIJ and fixABC ancestral type discordance. The study presents valuable insights into mosaic structures and the evolution of rhizobial symbiotic plasmids. This study compared 24 rhizobial symbiotic plasmids that included four genera and 11 species, illuminating the functional gene composition and symbiosis gene ancestor types of symbiotic plasmids from higher taxonomy. It provides valuable insights into mosaic structures and the evolution of symbiotic plasmids. © 2018 The Society for Applied Microbiology.

  15. Co-evolution of genomes and plasmids within Chlamydia trachomatis and the emergence in Sweden of a new variant strain

    Directory of Open Access Journals (Sweden)

    Skilton Rachel J

    2009-05-01

    suggest that the plasmid is not a highly mobile genetic element and does not transfer readily between isolates. Comparative analysis of the plasmid sequences has revealed the most conserved regions that should be used to design future plasmid based nucleic acid amplification tests, to avoid diagnostic failures.

  16. Analysis of plasmid genes by phylogenetic profiling and visualization of homology relationships using Blast2Network

    Directory of Open Access Journals (Sweden)

    Bazzicalupo Marco

    2008-12-01

    Full Text Available Abstract Background Phylogenetic methods are well-established bioinformatic tools for sequence analysis, allowing to describe the non-independencies of sequences because of their common ancestor. However, the evolutionary profiles of bacterial genes are often complicated by hidden paralogy and extensive and/or (multiple horizontal gene transfer (HGT events which make bifurcating trees often inappropriate. In this context, plasmid sequences are paradigms of network-like relationships characterizing the evolution of prokaryotes. Actually, they can be transferred among different organisms allowing the dissemination of novel functions, thus playing a pivotal role in prokaryotic evolution. However, the study of their evolutionary dynamics is complicated by the absence of universally shared genes, a prerequisite for phylogenetic analyses. Results To overcome such limitations we developed a bioinformatic package, named Blast2Network (B2N, allowing the automatic phylogenetic profiling and the visualization of homology relationships in a large number of plasmid sequences. The software was applied to the study of 47 completely sequenced plasmids coming from Escherichia, Salmonella and Shigella spps. Conclusion The tools implemented by B2N allow to describe and visualize in a new way some of the evolutionary features of plasmid molecules of Enterobacteriaceae; in particular it helped to shed some light on the complex history of Escherichia, Salmonella and Shigella plasmids and to focus on possible roles of unannotated proteins. The proposed methodology is general enough to be used for comparative genomic analyses of bacteria.

  17. Plasmid fermentation process for DNA immunization applications.

    Science.gov (United States)

    Carnes, Aaron E; Williams, James A

    2014-01-01

    Plasmid DNA for immunization applications must be of the highest purity and quality. The ability of downstream purification to efficiently produce a pure final product is directly influenced by the performance of the upstream fermentation process. While several clinical manufacturing facilities already have validated fermentation processes in place to manufacture plasmid DNA for use in humans, a simple and inexpensive laboratory-scale fermentation process can be valuable for in-house production of plasmid DNA for use in animal efficacy studies. This chapter describes a simple fed-batch fermentation process for producing bacterial cell paste enriched with high-quality plasmid DNA. A constant feeding strategy results in a medium cell density culture with continuously increasing plasmid amplification towards the end of the process. Cell banking and seed culture preparation protocols, which can dramatically influence final product yield and quality, are also described. These protocols are suitable for production of research-grade plasmid DNA at the 100 mg-to-1.5 g scale from a typical 10 L laboratory benchtop fermentor.

  18. Plasmid-mediated mineralization of 4-chlorobiphenyl

    International Nuclear Information System (INIS)

    Shields, M.S.; Hooper, S.W.; Sayler, G.S.

    1985-01-01

    Strains of Alcaligenes and Acinetobacter spp. were isolated from a mixed culture already proven to be proficient at complete mineralization of monohalogenated biphenyls. These strains were shown to harbor a 35 x 10(6)-dalton plasmid mediating a complete pathway for 4-chlorobiphenyl (4CB) oxidation. Subsequent plasmid curing of these bacteria resulted in the abolishment of the 4CB mineralization phenotype and loss of even early 4CB metabolism by Acinetobacter spp. Reestablishment of the Alcaligenes plasmid, denoted pSS50, in the cured Acinetobacter spp. via filter surface mating resulted in the restoration of 4CB mineralization abilities. 4CB mineralization, however, proved to be an unstable characteristic in some subcultured strains. Such loss was not found to coincide with any detectable alteration in plasmid size. Cultures capable of complete mineralization, as well as those limited to partial metabolism of 4CB, produced 4-chlorobenzoate as a metabolite. Demonstration of mineralization of a purified 14 C-labeled chlorobenzoate showed it to be a true intermediate in 4CB mineralization. Unlike the mineralization capability, the ability to produce a metabolite has proven to be stable on subculture. These results indicate the occurrence of a novel plasmid, or evolved catabolic plasmid, that mediates the complete mineralization of 4CB

  19. Heavy ion induced damage to plasmid DNA : plateau region vs. spread out Bragg-peak

    NARCIS (Netherlands)

    Dang, H.M.; van Goethem, M.J.; van der Graaf, E.R.; Brandenburg, S.; Hoekstra, R.A.; Schlathölter, T.A.

    We have investigated the damage of synthetic plasmid pBR322 DNA in dilute aqueous solutions induced by fast carbon ions. The relative contribution of indirect damage and direct damage to the DNA itself is expected to vary with linear energy transfer along the ion track, with the direct damage

  20. Host range of enterococcal vanA plasmids among Gram-positive intestinal bacteria

    DEFF Research Database (Denmark)

    Werner, Guido; Freitas, Ana R.; Coque, Teresa M.

    2010-01-01

    recipients. Transfer rates were calculated per donor and recipient. Transconjugants were confirmed by determining their phenotypic and genotypic properties. Stability of plasmids in the new host was assessed in long-term growth experiments. RESULTS: In total, 282 enterococcal matings and 73 inter...

  1. Behavior of IncQ Plasmids in Agrobacterium tumefaciens

    NARCIS (Netherlands)

    Hille, Jacques; Schilperoort, Rob

    1981-01-01

    Inc-Q plasmids were introduced into Agrobacterium tumefuciens, by mobilization from Escherichia coli with an Inc-P plasmid, or by transformation with purified plasmid DNA. It was found that they were stably maintained. The presence of an Inc-Q plasmid did not influence tumorigenicity. These results

  2. Comparative metagenomic analysis of plasmid encoded functions in the human gut microbiome

    Directory of Open Access Journals (Sweden)

    Marchesi Julian R

    2010-01-01

    Full Text Available Abstract Background Little is known regarding the pool of mobile genetic elements associated with the human gut microbiome. In this study we employed the culture independent TRACA system to isolate novel plasmids from the human gut microbiota, and a comparative metagenomic analysis to investigate the distribution and relative abundance of functions encoded by these plasmids in the human gut microbiome. Results Novel plasmids were acquired from the human gut microbiome, and homologous nucleotide sequences with high identity (>90% to two plasmids (pTRACA10 and pTRACA22 were identified in the multiple human gut microbiomes analysed here. However, no homologous nucleotide sequences to these plasmids were identified in the murine gut or environmental metagenomes. Functions encoded by the plasmids pTRACA10 and pTRACA22 were found to be more prevalent in the human gut microbiome when compared to microbial communities from other environments. Among the most prevalent functions identified was a putative RelBE toxin-antitoxin (TA addiction module, and subsequent analysis revealed that this was most closely related to putative TA modules from gut associated bacteria belonging to the Firmicutes. A broad phylogenetic distribution of RelE toxin genes was observed in gut associated bacterial species (Firmicutes, Bacteroidetes, Actinobacteria and Proteobacteria, but no RelE homologues were identified in gut associated archaeal species. We also provide indirect evidence for the horizontal transfer of these genes between bacterial species belonging to disparate phylogenetic divisions, namely Gram negative Proteobacteria and Gram positive species from the Firmicutes division. Conclusions The application of a culture independent system to capture novel plasmids from the human gut mobile metagenome, coupled with subsequent comparative metagenomic analysis, highlighted the unexpected prevalence of plasmid encoded functions in the gut microbial ecosystem. In

  3. Analysis of conjugated heat transfer, in transient state of the first stage of a gas turbine; Analisis de transferencia de calor conjugada, en estado transitorio, de la primera etapa de una turbina de gas

    Energy Technology Data Exchange (ETDEWEB)

    Campos Amezcua, Alfonso; Mazur C, Zdzislaw [Instituto de Investigaciones Electricas, Cuernavaca, Morelos (Mexico); Gallegos Munoz, Armando [Facultad de Ingenieria Mecanica, Electrica y Electronica (FIMEE), Universidad de Guanajuato, Guanajuato (Mexico)

    2007-11-15

    This article presents an analysis of conjugated heat transfer in the first stage of movable blades during the starting of a gas turbine, covering a period of 1,012 seconds. The developed computer model is in 3D and uses as initial and border conditions typical starting curves for stack gases, the cooling air and the angular velocity of the blades. As a result of the numerical predictions, the temperature distributions in stack gases, the trowel of the blade and the cooling air are included, doing emphasis in the results obtained in the solid (body of the blade), since these are used for thermo-mechanical stress analysis and later estimation of the blade residual life. [Spanish] Este articulo presenta un analisis de transferencia de calor conjugada en la primera etapa de alabes moviles, durante el arranque de una turbina de gas, cubriendo un periodo de 1.012 segundos. El modelo computacional desarrollado es en tres dimensiones y utiliza como condiciones iniciales y de frontera curvas de arranque tipicas para los gases de combustion, el aire de enfriamiento y la velocidad angular de los alabes. Como resultado de las predicciones numericas, se incluyen las distribuciones de temperatura en los gases de combustion, la paleta del alabe y el aire de enfriamiento, haciendo enfasis en los resultados obtenidos en el solido (cuerpo del alabe), ya que estos se utilizan para analisis de esfuerzos termomecanicos y posterior estimacion de vida residual del alabe.

  4. A finite element solution to conjugated heat transfer in tissue using magnetic resonance angiography to measure the in vitro velocity field

    Science.gov (United States)

    Dutton, Andrew William

    1993-12-01

    A combined numerical and experimental system for tissue heat transfer analysis was developed. The goal was to develop an integrated set of tools for studying the problem of providing accurate temperature estimation for use in hyperthermia treatment planning in a clinical environment. The completed system combines (1) Magnetic Resonance Angiography (MRA) to non-destructively measure the velocity field in situ, (2) the Streamwise Upwind Petrov-Galerkin finite element solution to the 3D steady state convective energy equation (CEE), (3) a medical image based automatic 3D mesh generator, and (4) a Gaussian type estimator to determine unknown thermal model parameters such as thermal conductivity, blood perfusion, and blood velocities from measured temperature data. The system was capable of using any combination of three thermal models (1) the Convective Energy Equation (CEE), (2) the Bioheat Transfer Equation (BHTE), and (3) the Effective Thermal Conductivity Equation (ETCE) Incorporation of the theoretically correct CEE was a significant theoretical advance over approximate models made possible by the use of MRA to directly measure the 3D velocity field in situ. Experiments were carried out in a perfused alcohol fixed canine liver with hyperthermia induced through scanned focused ultrasound Velocity fields were measured using Phase Contrast Angiography. The complete system was then used to (1) develop a 3D finite element model based upon user traced outlines over a series of MR images of the liver and (2) simulate temperatures at steady state using the CEE, BHTE, and ETCE thermal models in conjunction with the gauss estimator. Results of using the system on an in vitro liver preparation indicate the need for improved accuracy in the MRA scans and accurate spatial registration between the thermocouple junctions, the measured velocity field, and the scanned ultrasound power No individual thermal model was able to meet the desired accuracy of 0.5 deg C, the resolution

  5. Protection from ischemic heart injury by a vigilant heme oxygenase-1 plasmid system.

    Science.gov (United States)

    Tang, Yao Liang; Tang, Yi; Zhang, Y Clare; Qian, Keping; Shen, Leping; Phillips, M Ian

    2004-04-01

    Although human heme oxygenase-1 (hHO-1) could provide a useful approach for cellular protection in the ischemic heart, constitutive overexpression of hHO-1 may lead to unwanted side effects. To avoid this, we designed a hypoxia-regulated hHO-1 gene therapy system that can be switched on and off. This vigilant plasmid system is composed of myosin light chain-2v promoter and a gene switch that is based on an oxygen-dependent degradation domain from the hypoxia inducible factor-1-alpha. The vector can sense ischemia and switch on the hHO-1 gene system, specifically in the heart. In an in vivo experiment, the vigilant hHO-1 plasmid or saline was injected intramyocardially into myocardial infarction mice or sham operation mice. After gene transfer, expression of hHO-1 was only detected in the ischemic heart treated with vigilant hHO-1 plasmids. Masson trichrome staining showed significantly fewer fibrotic areas in vigilant hHO-1 plasmids-treated mice compared with saline control (43.0%+/-4.8% versus 62.5%+/-3.3%, PhHO-1 expression in peri-infarct border areas, concomitant with higher Bcl-2 levels and lower Bax, Bak, and caspase 3 levels in the ischemic myocardium compared with saline control. By use of a cardiac catheter, heart from vigilant hHO-1 plasmids-treated mice showed improved recovery of contractile and diastolic performance after myocardial infarction compared with saline control. This study documents the beneficial regulation and therapeutic potential of vigilant plasmid-mediated hHO-1 gene transfer. This novel gene transfer strategy can provide cardiac-specific protection from future repeated bouts of ischemic injury.

  6. The complete sequence and comparative analysis of a multidrug- resistance and virulence multireplicon IncFII plasmid pEC302/04 from an extraintestinal pathogenic Escherichia coli EC302/04 indicate extensive diversity of IncFII plasmids

    Directory of Open Access Journals (Sweden)

    Wing Sze eHo

    2016-01-01

    Full Text Available Extraintestinal pathogenic Escherichia coli (ExPEC that causes extraintestinal infections often harbor plasmids encoding fitness traits such as resistance and virulence determinants that are of clinical importance. We determined the complete nucleotide sequence of plasmid pEC302/04 from a multidrug-resistant E. coli EC302/04 which was isolated from the tracheal aspirate of a patient in Malaysia. In addition, we also performed comparative sequence analyses of 18 related IncFIIA plasmids to determine the phylogenetic relationship and diversity of these plasmids. The 140,232 bp pEC302/04 is a multireplicon plasmid that bears three replication systems (FII, FIA and FIB with subtype of F2:A1:B1. The plasmid is self-transmissible with a complete transfer region. pEC302/04 also carries antibiotic resistance genes such as blaTEM-1 and a class I integron containing sul1, cml and aadA resistance genes, conferring multidrug resistance (MDR to its host, E. coli EC302/04. Besides, two iron acquisition systems (SitABCD and IutA-IucABCD which are the conserved virulence determinants of ExPEC-colicin V or B and M (ColV/ColBM-producing plasmids were identified in pEC302/04. Multiple toxin-antitoxin (TA-based addiction systems (i.e., PemI/PemK, VagC/VagD, CcdA/CcdB, and Hok/Sok and a plasmid partitioning system, ParAB and PsiAB, which are important for plasmid maintenance were also found.Comparative plasmid analysis revealed only one conserved gene, the repA1 as the core genome, showing that there is an extensive diversity among the IncFIIA plasmids. The phylogenetic relationship of 18 IncF plasmids based on the core regions revealed that ColV/ColBM-plasmids and non-ColV/ColBM plasmids were separated into two distinct groups. These plasmids, which carry highly diverse genetic contents, are also mosaic in nature. The atypical combination of genetic materials, i.e., the MDR- and ColV/ColBM-plasmid-virulence encoding regions in a single ExPEC plasmid is rare but of

  7. The Complete Sequence and Comparative Analysis of a Multidrug-Resistance and Virulence Multireplicon IncFII Plasmid pEC302/04 from an Extraintestinal Pathogenic Escherichia coli EC302/04 Indicate Extensive Diversity of IncFII Plasmids.

    Science.gov (United States)

    Ho, Wing Sze; Yap, Kien-Pong; Yeo, Chew Chieng; Rajasekaram, Ganeswrie; Thong, Kwai Lin

    2015-01-01

    Extraintestinal pathogenic Escherichia coli (ExPEC) that causes extraintestinal infections often harbor plasmids encoding fitness traits such as resistance and virulence determinants that are of clinical importance. We determined the complete nucleotide sequence of plasmid pEC302/04 from a multidrug-resistant E. coli EC302/04 which was isolated from the tracheal aspirate of a patient in Malaysia. In addition, we also performed comparative sequence analyses of 18 related IncFIIA plasmids to determine the phylogenetic relationship and diversity of these plasmids. The 140,232 bp pEC302/04 is a multireplicon plasmid that bears three replication systems (FII, FIA, and FIB) with subtype of F2:A1:B1. The plasmid is self-transmissible with a complete transfer region. pEC302/04 also carries antibiotic resistance genes such as bla TEM-1 and a class I integron containing sul1, cml and aadA resistance genes, conferring multidrug resistance (MDR) to its host, E. coli EC302/04. Besides, two iron acquisition systems (SitABCD and IutA-IucABCD) which are the conserved virulence determinants of ExPEC-colicin V or B and M (ColV/ColBM)-producing plasmids were identified in pEC302/04. Multiple toxin-antitoxin (TA)-based addiction systems (i.e., PemI/PemK, VagC/VagD, CcdA/CcdB, and Hok/Sok) and a plasmid partitioning system, ParAB, and PsiAB, which are important for plasmid maintenance were also found. Comparative plasmid analysis revealed only one conserved gene, the repA1 as the core genome, showing that there is an extensive diversity among the IncFIIA plasmids. The phylogenetic relationship of 18 IncF plasmids based on the core regions revealed that ColV/ColBM-plasmids and non-ColV/ColBM plasmids were separated into two distinct groups. These plasmids, which carry highly diverse genetic contents, are also mosaic in nature. The atypical combination of genetic materials, i.e., the MDR- and ColV/ColBM-plasmid-virulence encoding regions in a single ExPEC plasmid is rare but of clinical

  8. Stochastic differential equations used to model conjugation

    DEFF Research Database (Denmark)

    Philipsen, Kirsten Riber; Christiansen, Lasse Engbo

    Stochastic differential equations (SDEs) are used to model horizontal transfer of antibiotic resis- tance by conjugation. The model describes the concentration of donor, recipient, transconjugants and substrate. The strength of the SDE model over the traditional ODE models is that the noise can...

  9. Molecular Characterization of Plasmids Encoding CTX-M β-Lactamases and their Associated Addiction Systems Circulating Among Escherichia coli from Retail Chickens, Chicken Farms, and Slaughterhouses in Korea.

    Science.gov (United States)

    Jo, Su-Jin; Woo, Gun-Jo

    2016-02-01

    Extended-spectrum β-lactamases (ESBLs), particularly those of the CTX-M types, are the predominant resistance determinants of Escherichia coli that are rapidly spreading worldwide. To determine CTX-M types, E. coli isolates were collected from retail chickens (n = 390) and environmental samples from chicken farms (n = 32) and slaughterhouses (n = 67) in Korea. Fifteen strains harboring blaCTX-M genes were isolated from 358 E. coli isolates. The most common CTX-M type was eight of CTX-M-15, followed by six of CTX-M-1 and one of CTX-M- 14. The blaCTX-M genes were identified in the isolates from retail chickens (n = 9), followed by feces, water pipes, floors, and walls. Conjugations confirmed the transferability of the plasmids carrying blaCTX-M genes to the recipient E. coli J53 strain. Furthermore, eight addiction systems carried by the replicons in CTX-M types were confirmed. The dominant system was identified as ccdAB, vagCD, and pndAC in donor strains and transconjugants. The clonal relationship between the two strains carrying blaCTX-M genes indicates that E. coli may transmit from the farm to retail chickens, suggesting a possible public health risk. Our findings demonstrate that the detection of CTX-M types in E. coli isolates is important for tracking ESBL production in animals, and suggest linkage of multiple addiction systems in plasmids bearing blaCTX-M genes.

  10. Transformation of Cowpea Vigna unguiculata Cells with an Antibiotic Resistance Gene Using a Ti-Plasmid-Derived Vector

    NARCIS (Netherlands)

    Hille, Jacques; Goldbach, Rob

    1986-01-01

    A chimaeric antibiotic resistance gene was transferred to cowpea (Vigna unguiculata), a member of the legume family. This transfer was established by inoculating cowpea leaf discs with an Agrobacterium tumefaciens strain harboring a Ti-plasmid-derived vector that contained two copies of a chimaeric

  11. An abyssal mobilome: viruses, plasmids and vesicles from deep-sea hydrothermal vents.

    Science.gov (United States)

    Lossouarn, Julien; Dupont, Samuel; Gorlas, Aurore; Mercier, Coraline; Bienvenu, Nadege; Marguet, Evelyne; Forterre, Patrick; Geslin, Claire

    2015-12-01

    Mobile genetic elements (MGEs) such as viruses, plasmids, vesicles, gene transfer agents (GTAs), transposons and transpovirions, which collectively represent the mobilome, interact with cellular organisms from all three domains of life, including those thriving in the most extreme environments. While efforts have been made to better understand deep-sea vent microbial ecology, our knowledge of the mobilome associated with prokaryotes inhabiting deep-sea hydrothermal vents remains limited. Here we focus on the abyssal mobilome by reviewing accumulating data on viruses, plasmids and vesicles associated with thermophilic and hyperthermophilic Bacteria and Archaea present in deep-sea hydrothermal vents. Copyright © 2015 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  12. Formation of Escherichia coli Hfr strains by integrative suppression with the P group plasmid RP1.

    OpenAIRE

    Martin, R R; Thorlton, C L; Unger, L

    1981-01-01

    Hfr strains of Escherichia coli were obtained by integrative suppression of a dnaA(Ts) mutation by the Inc P-1 plasmid RP1 without prior creation of an unnatural homology between the plasmid and the E. coli chromosome. Unmodified RP1 mobilized the polarized transfer of the chromosome in a counterclock-wise direction from a distinct origin between 81 min (pyrE) and 82 min (dnaA) with pyrE as a leading marker. Inheritance of RP1-Hfr chromosomal and antibiotic resistance genes was due to recombi...

  13. Translocation of integron-associated resistance in a natural system: Acquisition of resistance determinants by Inc P and Inc W Plasmids from Salmonella enterica Typhimurium DT104

    DEFF Research Database (Denmark)

    Sandvang, Dorthe; Diggle, M.; Platt, D.J.

    2002-01-01

    to determinate the genetic content. Translocation to R751 and R388 was associated with the loss of the indigenous trimethoprim cassette to both plasmids and also acquisition of sulfonamide resistance by R751 and RP4::Tn7, which indicated movement of the 3' terminus of one or both of the DT104 integrons......Salmonella enterica Typhimurium DT104, 961368, a veterinary field isolate that encodes a chromosomal cluster of resistance genes as well as two integrons, was used to study the mobility of resistance cassettes (aadA2 and pse-1) and nonintegron-associated resistance determinants (chloramphenicol...... and tetracycline). A range of natural plasmids was used as targets for the translocation of resistance. Plasmids that acquired resistance from the DT104 chromosome were segregated by conjugation into Escherichia coli K12. Plasmids R751, R388, and RP4::Tn7 acquired several combinations of resistance determinant...

  14. An Enterobacter plasmid as a new genetic background for the transposon Tn1331

    Directory of Open Access Journals (Sweden)

    Alavi MR

    2011-11-01

    the Enterobacter plasmid pPIGDM1 enables this transposon to propagate in this Enterobacter. Since Tn1331 was previously isolated only from Klebsiella, this report suggests horizontal transfer of this transposon between the two bacterial genera.Keywords: transposon Tn1331, Enterobacter, wound infection

  15. Horizontol dissemination of TEM- and SHV-typr beta-lactamase genes-carrying resistance plasmids amongst clonical isolates of Enterobacteriaceae Disseminação horizontal de plasmídios de resistência contendo genes de beta-lactamase dos tipos TEM e SHV entre isolados clínicos de Enterobacteriaceae

    Directory of Open Access Journals (Sweden)

    Osman Birol Ozgumus

    2008-12-01

    Full Text Available The extended-spectrum β-lactamase (ESBL-producing bacteria have been isolated at increasing frequency worldwide. Expression of ESBL is often associated with multidrug resistance and dissemination by resistance plasmids. During a two-month period in 2000, 133 clinical isolates of enterobacterial strains were randomly collected from outpatients and inpatients at a university hospital in Turkey. The ESBL producing strains were determined by double-disk synergy (DDS testing. Twenty ESBL producing strains (15% including Escherichia coli (n = 9, Klebsiella pneumoniae (n = 7, Klebsiella oxytoca (n = 2 and Enterobacter aerogenes (n = 2 were detected and further analyzed for their resistance transfer features, plasmid profile and nature of the resistance genes. Plasmid transfer assays were performed using broth mating techniques. TEM- and SHV- genes were analyzed by polymerase chain reaction (PCR and hybridization using specific probes. EcoRI restriction enzyme analyses of R plasmids were used in the detection of epidemic plasmids. Fourteen plasmid profiles (A, B1, B2, C1, and C2 to L were obtained with EcoRI restriction enzyme analysis. Most of these plasmids were detected to carry both TEM- and SHV-derived genes by PCR, and confirmed by localizing each gene by hybridization assay. Epidemiological evidence indicated that there was an apparent horizontal dissemination of conjugative R plasmids among multidrug-resistant enterobacterial genera and species in this hospitalO isolamento de bactérias produtoras de beta-lactamases de espectro expandido (ESBL está aumentando no mundo todo. Freqüentemente, a expressão de ESBL está associada com resistência a múltiplas drogas e disseminação por plasmídios de resistência. Durante um período de dois meses em 2000, 133 isolados clínicos de cepas de enterobactérias foram obtidos aleatoriamente de pacientes internos e externos de um hospital universitário na Turquia. As cepas produtoras de ESBL foram

  16. SHV-type extended-spectrum beta-lactamase (ESBL are encoded in related plasmids from enterobacteria clinical isolates from Mexico beta-Lactamasas de espectro extendido (BLEE tipo SHV están codificadas en plásmidos relacionados en aislamientos clínicos de enterobacterias en México

    Directory of Open Access Journals (Sweden)

    Ulises Garza-Ramos

    2007-12-01

    Full Text Available OBJECTIVE: In this work we report the molecular characterization of beta-lactam antibiotics resistance conferred by genes contained in plasmids from enterobacteria producing extended-spectrum beta-lactamases (ESBL. MATERIAL AND METHODS: Fourteen enterobacterial clinical isolates selected from a group of strains obtained from seven different hospitals in Mexico during 1990-1992 and 1996-1998 were analyzed at the Bacterial Resistance Laboratory (National Institute Public Health, Cuernavaca. Molecular characterization included PFGE, IEF of beta-lactamases, bacterial conjugation, PCR amplification and DNA sequencing, plasmid extraction and restriction. RESULTS: Isolates were genetically unrelated. ESBL identified were SHV-2 (5/14 and SHV-5 (9/14 type. Cephalosporin-resistance was transferable in 9 of 14 (64% clinical isolates with only one conjugative plasmid, DNA finger printing showed a similar band pattern in plasmids. CONCLUSIONS: The dissemination of cephalosporin resistance was due to related plasmids carrying the ESBL genes.OBJETIVO: En este trabajo se reporta la caracterización molecular de la resistencia a antibiótico beta-lactámicos conferida por genes contenidos en plásmidos de enterobacterias productoras de beta-lactamasas de espectro extendido (BLEEs. MATERIAL Y MÉTODOS: Catorce aislamientos clínicos de enterobacterias fueron seleccionados por conveniencia de un banco de cepas obtenidas de siete diferentes hospitales de México durante los periodos 1990-1992 y 1996-1998 y fueron procesados en el Laboratorio de Resistencia Bacteriana (Instituto Nacional de Salud Pública, Cuernavaca. En la caracterización se empleó PFGE, IEF para beta-lactamasas, conjugación bacteriana, amplificación por PCR y secuenciación de DNA, extracción y restricción de plásmidos. RESULTADOS: Las 14 cepas fueron no relacionadas genéticamente. Se identificaron BLEEs tipo SHV-2 (5/14 y SHV-5 (9/14. La resistencia a cefalosporinas fue transferida por

  17. Optical phase conjugation

    CERN Document Server

    Fisher, Robert A

    1983-01-01

    This book appears at a time of intense activity in optical phase conjugation. We chose not to await the maturation of the field, but instead to provide this material in time to be useful in its development. We have tried very hard to elucidate and interrelate the various nonlinear phenomena which can be used for optical phase conjugation.

  18. Development of Tat-Conjugated Dendrimer for Transdermal DNA Vaccine Delivery.

    Science.gov (United States)

    Bahadoran, Azadeh; Moeini, Hassan; Bejo, Mohd Hair; Hussein, Mohd Zobir; Omar, Abdul Rahman

    In order to enhance cellular uptake and to facilitate transdermal delivery of DNA vaccine, polyamidoamine (PAMAM) dendrimers conjugated with HIV transactivator of transcription (TAT) was developed. First, the plasmid DNA (pIRES-H5/GFP) nanoparticle was formulated using PAMAM dendrimer and TAT peptide and then characterized for surface charge, particle size, DNA encapsulation and protection of the pIRES-H5/GFP DNA plasmid to enzymatic digestion. Subsequently, the potency of the TAT-conjugated dendrimer for gene delivery was evaluated through in vitro transfection into Vero cells followed by gene expression analysis including western blotting, fluorescent microscopy and PCR. The effect of the TAT peptide on cellular uptake of DNA vaccine was studied by qRT-PCR and flow cytometry. Finally, the ability of TAT-conjugated PAMAM dendrimer for transdermal delivery of the DNA plasmid was assessed through artificial membranes followed by qRT-PCR and flow cytometry. TAT-conjugated PAMAM dendrimer showed the ability to form a compact and nanometre-sized polyplexes with the plasmid DNA, having the size range of 105 to 115 nm and a positive charge of +42 to +45 mV over the N/P ratio of 6:1(+/-).  In vitro transfection analysis into Vero cells confirms the high potency of TAT-conjugated PAMAM dendrimer to enhance the cellular uptake of DNA vaccine.  The permeability value assay through artificial membranes reveals that TAT-conjugated PAMAM has more capacity for transdermal delivery of the DNA compared to unmodified PAMAM dendrimer (Pdendrimer is a promising non-viral vector for transdermal use.This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

  19. Characteristics of plasmids in multi-drug-resistant Enterobacteriaceae isolated during prospective surveillance of a newly opened hospital in Iraq.

    Directory of Open Access Journals (Sweden)

    Xiao-Zhe Huang

    Full Text Available BACKGROUND: Gram-negative multidrug-resistant (MDR bacteria are major causes of nosocomial infections, and antibiotic resistance in these organisms is often plasmid mediated. Data are scarce pertaining to molecular mechanisms of antibiotic resistance in resource constrained areas such as Iraq. METHODOLOGY/PRINCIPAL FINDINGS: In this study, all MDR Enterobacteriaceae (n = 38 and randomly selected non-MDR counterparts (n = 41 isolated from patients, healthcare workers and environmental surfaces in a newly opened hospital in Iraq were investigated to characterize plasmids found in these isolates and determine their contribution to antibiotic resistance. Our results demonstrated that MDR E. coli and K. pneumoniae isolates harbored significantly more (≥ 3 plasmids compared to their non-MDR counterparts, which carried ≤ 2 plasmids (p<0.01. Various large plasmids (~52 to 100 kb from representative isolates were confirmed to contain multiple resistance genes by DNA microarray analysis. Aminoglycoside (acc, aadA, aph, strA/B, and ksgA, β-lactam (bla(TEM1, bla(AMPC, bla(CTX-M-15, bla(OXA-1, bla(VIM-2 and bla(SHV, sulfamethoxazole/trimethoprim (sul/dfr, tetracycline (tet and chloramphenicol (cat resistance genes were detected on these plasmids. Additionally, multiple plasmids carrying multiple antibiotic resistance genes were found in the same host strain. Genetic transfer-associated genes were identified on the plasmids from both MDR and non-MDR isolates. Seven plasmid replicon types (FII, FIA, FIB, B/O, K, I1 and N were detected in the isolates, while globally disseminated IncA/C and IncHI1 plasmids were not detected in these isolates. CONCLUSIONS/SIGNIFICANCE: This is the first report of the characteristics of the plasmids found in Enterobacteriaceae isolated following the opening of a new hospital in Iraq. The information provided here furthers our understanding of the mechanisms of drug resistance in this specific region and their evolutionary

  20. Comparative genomics of multidrug resistance-encoding IncA/C plasmids from commensal and pathogenic Escherichia coli from multiple animal sources.

    Science.gov (United States)

    Fernández-Alarcón, Claudia; Singer, Randall S; Johnson, Timothy J

    2011-01-01

    Incompatibility group A/C (IncA/C) plasmids have received recent attention for their broad host range and ability to confer resistance to multiple antimicrobial agents. Due to the potential spread of multidrug resistance (MDR) phenotypes from foodborne pathogens to human pathogens, the dissemination of these plasmids represents a public health risk. In this study, four animal-source IncA/C plasmids isolated from Escherichia coli were sequenced and analyzed, including isolates from commercial dairy cows, pigs and turkeys in the U.S. and Chile. These plasmids were initially selected because they either contained the floR and tetA genes encoding for florfenicol and tetracycline resistance, respectively, and/or the bla(CMY-2) gene encoding for extended spectrum β-lactamase resistance. Overall, sequence analysis revealed that each of the four plasmids retained a remarkably stable and conserved backbone sequence, with differences observed primarily within their accessory regions, which presumably have evolved via horizontal gene transfer events involving multiple modules. Comparison of these plasmids with other available IncA/C plasmid sequences further defined the core and accessory elements of these plasmids in E. coli and Salmonella. Our results suggest that the bla(CMY-2) plasmid lineage appears to have derived from an ancestral IncA/C plasmid type harboring floR-tetAR-strAB and Tn21-like accessory modules. Evidence is mounting that IncA/C plasmids are widespread among enteric bacteria of production animals and these emergent plasmids have flexibility in their acquisition of MDR-encoding modules, necessitating further study to understand the evolutionary mechanisms involved in their dissemination and stability in bacterial populations.

  1. Comparative genomics of multidrug resistance-encoding IncA/C plasmids from commensal and pathogenic Escherichia coli from multiple animal sources.

    Directory of Open Access Journals (Sweden)

    Claudia Fernández-Alarcón

    Full Text Available Incompatibility group A/C (IncA/C plasmids have received recent attention for their broad host range and ability to confer resistance to multiple antimicrobial agents. Due to the potential spread of multidrug resistance (MDR phenotypes from foodborne pathogens to human pathogens, the dissemination of these plasmids represents a public health risk. In this study, four animal-source IncA/C plasmids isolated from Escherichia coli were sequenced and analyzed, including isolates from commercial dairy cows, pigs and turkeys in the U.S. and Chile. These plasmids were initially selected because they either contained the floR and tetA genes encoding for florfenicol and tetracycline resistance, respectively, and/or the bla(CMY-2 gene encoding for extended spectrum β-lactamase resistance. Overall, sequence analysis revealed that each of the four plasmids retained a remarkably stable and conserved backbone sequence, with differences observed primarily within their accessory regions, which presumably have evolved via horizontal gene transfer events involving multiple modules. Comparison of these plasmids with other available IncA/C plasmid sequences further defined the core and accessory elements of these plasmids in E. coli and Salmonella. Our results suggest that the bla(CMY-2 plasmid lineage appears to have derived from an ancestral IncA/C plasmid type harboring floR-tetAR-strAB and Tn21-like accessory modules. Evidence is mounting that IncA/C plasmids are widespread among enteric bacteria of production animals and these emergent plasmids have flexibility in their acquisition of MDR-encoding modules, necessitating further study to understand the evolutionary mechanisms involved in their dissemination and stability in bacterial populations.

  2. Mechanisms Involved in Acquisition of blaNDM Genes by IncA/C2 and IncFIIY Plasmids.

    Science.gov (United States)

    Wailan, Alexander M; Sidjabat, Hanna E; Yam, Wan Keat; Alikhan, Nabil-Fareed; Petty, Nicola K; Sartor, Anna L; Williamson, Deborah A; Forde, Brian M; Schembri, Mark A; Beatson, Scott A; Paterson, David L; Walsh, Timothy R; Partridge, Sally R

    2016-07-01

    blaNDM genes confer carbapenem resistance and have been identified on transferable plasmids belonging to different incompatibility (Inc) groups. Here we present the complete sequences of four plasmids carrying a blaNDM gene, pKP1-NDM-1, pEC2-NDM-3, pECL3-NDM-1, and pEC4-NDM-6, from four clinical samples originating from four different patients. Different plasmids carry segments that align to different parts of the blaNDM region found on Acinetobacter plasmids. pKP1-NDM-1 and pEC2-NDM-3, from Klebsiella pneumoniae and Escherichia coli, respectively, were identified as type 1 IncA/C2 plasmids with almost identical backbones. Different regions carrying blaNDM are inserted in different locations in the antibiotic resistance island known as ARI-A, and ISCR1 may have been involved in the acquisition of blaNDM-3 by pEC2-NDM-3. pECL3-NDM-1 and pEC4-NDM-6, from Enterobacter cloacae and E. coli, respectively, have similar IncFIIY backbones, but different regions carrying blaNDM are found in different locations. Tn3-derived inverted-repeat transposable elements (TIME) appear to have been involved in the acquisition of blaNDM-6 by pEC4-NDM-6 and the rmtC 16S rRNA methylase gene by IncFIIY plasmids. Characterization of these plasmids further demonstrates that even very closely related plasmids may have acquired blaNDM genes by different mechanisms. These findings also illustrate the complex relationships between antimicrobial resistance genes, transposable elements, and plasmids and provide insights into the possible routes for transmission of blaNDM genes among species of the Enterobacteriaceae family. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  3. High diversity of genes and plasmids encoding resistance to third-generation cephalosporins and quinolones in clinical Escherichia coli from commercial poultry flocks in Italy

    DEFF Research Database (Denmark)

    Niero, Giulia; Bortolaia, Valeria; Vanni, Michele

    2018-01-01

    = 98) and layers (n = 22) between 2008 and 2012. 3GC-resistant isolates were screened for extended-spectrum and AmpC β-lactamase (ESBL/AmpC), while all isolates were tested for plasmid-mediated quinolone resistance (PMQR) genes. ESBL/AmpC- and PMQR-positive isolates were typed by pulsed-field gel......% of isolates from turkeys, broilers and layers, respectively. We identified seven ESBL/AmpC-encoding plasmid types, usually conjugative (78%), with a marked prevalence of IncI1/pST3 plasmids carrying blaCTX-M-1. PMQR occurred less frequently among isolates from turkeys (0.9%) compared to those from broilers (5......%) and layers (4%). The PMQR genes qnrS, qnrB19 and oqxA/B were located on three plasmid types and two non-typeable plasmids, mostly (85%) conjugative. ESBL/AmpC- and PMQR-positive isolates were genetically unrelated and 64% of them were additionally resistant to aminoglycosides, sulfonamides and tetracyclines...

  4. Standardized Cloning and Curing of Plasmids

    DEFF Research Database (Denmark)

    Lauritsen, Ida; Kim, Se Hyeuk; Porse, Andreas

    2018-01-01

    and exchange of genetic parts in the Standard European Vectors Architecture (SEVA) vector system. Additionally, to facilitate rapid testing and iterative bioengineering using different vector designs, we provide a one-step protocol for a universal CRISPR-Cas9-based plasmid curing system (pFREE) and demonstrate...

  5. Optimization of plasmid electrotransformation into Escherichia coli ...

    African Journals Online (AJOL)

    In order to improve electroporation, optical density of bacteria, recovery time and electrical parameter (field strength and capacitance) were optimized using the Taguchi statistical method. ANOVA of obtained data indicated that the optimal conditions of electrotransformation of pET-28a (+) plasmid into Escherichia coli ...

  6. Plasmid mediated quinolone resistance in Enterobacteriaceae

    NARCIS (Netherlands)

    Veldman, K.T.; LS Klinisch Onderzoek Wagenaar

    2014-01-01

    This thesis describes the occurrence of Plasmid Mediated Quinolone Resistance (PMQR) in Salmonella and E. coli from The Netherlands and other European countries. Furthermore, the genetic background of these genes was characterized. Fluoroquinolones are widely used antibiotics in both human and

  7. Plasmid and chromosome segregation in prokaryotes

    DEFF Research Database (Denmark)

    Møller-Jensen, Jakob; Bugge Jensen, Rasmus; Gerdes, Kenn

    2000-01-01

    Recent major advances in the understanding of prokaryotic DNA segregation have been achieved by using fluorescence microscopy to visualize the localization of cellular components. Plasmids and bacterial chromosomes are partitioned in a highly dynamic fashion, suggesting the presence of a mitotic...

  8. Antimicrobial resistance and plasmid profiles of Aeromonas ...

    African Journals Online (AJOL)

    The purpose of this study was to investigate the presence of Aeromonas hydrophila at commonly used water collection points on the River Njoro and to determine the in-vitro antimicrobial susceptibility and plasmid profiles of isolates. In total, 126 samples were collected and 36.5% of them were positive for A. hydrophila.

  9. Antimicrobial resistance patterns and plasmid profiles of ...

    African Journals Online (AJOL)

    Objectives: To determine the frequency of resistance of Staphylococcus aureus to various antimicrobial agents, and the relationship between antimicrobial resistance of the isolates and carriage of plasmids. Design: A random sampling of milk and meat samples was carried out. Setting: Milk was collected from various dairy ...

  10. Simple method for identification of plasmid-coded proteins

    International Nuclear Information System (INIS)

    Sancar, A.; Hack, A.M.; Rupp, W.D.

    1979-01-01

    Proteins encoded by plasmid DNA are specifically labeled in uv-irradiated cells of Escherichia coli carrying recA and uvrA mutations because extensive degradation of the chromosome DNA occurs concurrently with amplification of plasmid DNA

  11. Solid lipid nanoparticles mediate non-viral delivery of plasmid DNA to dendritic cells

    Science.gov (United States)

    Penumarthi, Alekhya; Parashar, Deepti; Abraham, Amanda N.; Dekiwadia, Chaitali; Macreadie, Ian; Shukla, Ravi; Smooker, Peter M.

    2017-06-01

    There is an increasing demand for novel DNA vaccine delivery systems, mainly for the non-viral type as they are considered relatively safe. Therefore, solid lipid nanoparticles (SLNs) were investigated for their suitability as a non-viral DNA vaccine delivery system. SLNs were synthesised by a modified solvent-emulsification method in order to study their potential to conjugate with plasmid DNA and deliver them in vitro to dendritic cells using eGFP as the reporter plasmid. The DNA-SLN complexes were characterised by electron microscopy, gel retardation assays and dynamic light scattering. The cytotoxicity assay data supported their biocompatibility and was used to estimate safe threshold concentration resulting in high transfection rate. The transfection efficiency of these complexes in a dendritic cell line was shown to increase significantly compared to plasmid alone, and was comparable to that mediated by lipofectamine. Transmission electron microscopy studies delineated the pathway of cellular uptake. Endosomal escape was observed supporting the mechanism of transfection.

  12. Plasmid mediated enhancement of uv resistance in Streptococcus faecalis

    International Nuclear Information System (INIS)

    Miehl, R.; Miller, M.; Yasbin, R.E.

    1980-01-01

    A 38.5-Mdal plasmid of Streptococcus faecalis subdp. zymogenes has been shown to enhance survival following uv irradiation. In addition, the presence of this plasmid increases the mutation frequencies following uv irradiation and enhanced W-reactivation. The data presented indicate that S. faecalis has an inducible error-prone repair system and that the plasmid enhances these repair functions

  13. Qualidade conjugal: mapeando conceitos

    Directory of Open Access Journals (Sweden)

    Clarisse Mosmann

    2006-12-01

    Full Text Available Apesar da ampla utilização do conceito de qualidade conjugal, identifica-se falta de clareza conceitual acerca das variáveis que o compõem. Esse artigo apresenta revisão da literatura na área com o objetivo de mapear o conceito de qualidade conjugal. Foram analisadas sete principais teorias sobre o tema: Troca Social, Comportamental, Apego, Teoria da Crise, Interacionismo Simbólico. Pelos postulados propostos nas diferentes teorias, podem-se identificar três grupos de variáveis fundamentais na definição da qualidade conjugal: recursos pessoais dos cônjuges, contexto de inserção do casal e processos adaptativos. Neste sentido, a qualidade conjugal é resultado do processo dinâmico e interativo do casal, razão deste caráter multidimensional.

  14. Conjugate Gaze Palsies

    Science.gov (United States)

    ... version Home Brain, Spinal Cord, and Nerve Disorders Cranial Nerve Disorders Conjugate Gaze Palsies Horizontal gaze palsy Vertical ... Version. DOCTORS: Click here for the Professional Version Cranial Nerve Disorders Overview of the Cranial Nerves Internuclear Ophthalmoplegia ...

  15. Conjugated Polymer Solar Cells

    National Research Council Canada - National Science Library

    Paraschuk, Dmitry Y

    2006-01-01

    This report results from a contract tasking Moscow State University as follows: Conjugated polymers are promising materials for many photonics applications, in particular, for photovoltaic and solar cell devices...

  16. Polymers for Protein Conjugation

    Directory of Open Access Journals (Sweden)

    Gianfranco Pasut

    2014-01-01

    Full Text Available Polyethylene glycol (PEG at the moment is considered the leading polymer for protein conjugation in view of its unique properties, as well as to its low toxicity in humans, qualities which have been confirmed by its extensive use in clinical practice. Other polymers that are safe, biodegradable and custom-designed have, nevertheless, also been investigated as potential candidates for protein conjugation. This review will focus on natural polymers and synthetic linear polymers that have been used for protein delivery and the results associated with their use. Genetic fusion approaches for the preparation of protein-polypeptide conjugates will be also reviewed and compared with the best known chemical conjugation ones.

  17. Construction of Biologically Functional Bacterial Plasmids In Vitro

    Science.gov (United States)

    Cohen, Stanley N.; Chang, Annie C. Y.; Boyer, Herbert W.; Helling, Robert B.

    1973-01-01

    The construction of new plasmid DNA species by in vitro joining of restriction endonuclease-generated fragments of separate plasmids is described. Newly constructed plasmids that are inserted into Escherichia coli by transformation are shown to be biologically functional replicons that possess genetic properties and nucleotide base sequences from both of the parent DNA molecules. Functional plasmids can be obtained by reassociation of endonuclease-generated fragments of larger replicons, as well as by joining of plasmid DNA molecules of entirely different origins. Images PMID:4594039

  18. Plasmid Flux in Escherichia coli ST131 Sublineages, Analyzed by Plasmid Constellation Network (PLACNET), a New Method for Plasmid Reconstruction from Whole Genome Sequences

    Science.gov (United States)

    Garcillán-Barcia, M. Pilar; Mora, Azucena; Blanco, Jorge; Coque, Teresa M.; de la Cruz, Fernando

    2014-01-01

    Bacterial whole genome sequence (WGS) methods are rapidly overtaking classical sequence analysis. Many bacterial sequencing projects focus on mobilome changes, since macroevolutionary events, such as the acquisition or loss of mobile genetic elements, mainly plasmids, play essential roles in adaptive evolution. Existing WGS analysis protocols do not assort contigs between plasmids and the main chromosome, thus hampering full analysis of plasmid sequences. We developed a method (called plasmid constellation networks or PLACNET) that identifies, visualizes and analyzes plasmids in WGS projects by creating a network of contig interactions, thus allowing comprehensive plasmid analysis within WGS datasets. The workflow of the method is based on three types of data: assembly information (including scaffold links and coverage), comparison to reference sequences and plasmid-diagnostic sequence features. The resulting network is pruned by expert analysis, to eliminate confounding data, and implemented in a Cytoscape-based graphic representation. To demonstrate PLACNET sensitivity and efficacy, the plasmidome of the Escherichia coli lineage ST131 was analyzed. ST131 is a globally spread clonal group of extraintestinal pathogenic E. coli (ExPEC), comprising different sublineages with ability to acquire and spread antibiotic resistance and virulence genes via plasmids. Results show that plasmids flux in the evolution of this lineage, which is wide open for plasmid exchange. MOBF12/IncF plasmids were pervasive, adding just by themselves more than 350 protein families to the ST131 pangenome. Nearly 50% of the most frequent γ–proteobacterial plasmid groups were found to be present in our limited sample of ten analyzed ST131 genomes, which represent the main ST131 sublineages. PMID:25522143

  19. Plasmid flux in Escherichia coli ST131 sublineages, analyzed by plasmid constellation network (PLACNET), a new method for plasmid reconstruction from whole genome sequences.

    Science.gov (United States)

    Lanza, Val F; de Toro, María; Garcillán-Barcia, M Pilar; Mora, Azucena; Blanco, Jorge; Coque, Teresa M; de la Cruz, Fernando

    2014-12-01

    Bacterial whole genome sequence (WGS) methods are rapidly overtaking classical sequence analysis. Many bacterial sequencing projects focus on mobilome changes, since macroevolutionary events, such as the acquisition or loss of mobile genetic elements, mainly plasmids, play essential roles in adaptive evolution. Existing WGS analysis protocols do not assort contigs between plasmids and the main chromosome, thus hampering full analysis of plasmid sequences. We developed a method (called plasmid constellation networks or PLACNET) that identifies, visualizes and analyzes plasmids in WGS projects by creating a network of contig interactions, thus allowing comprehensive plasmid analysis within WGS datasets. The workflow of the method is based on three types of data: assembly information (including scaffold links and coverage), comparison to reference sequences and plasmid-diagnostic sequence features. The resulting network is pruned by expert analysis, to eliminate confounding data, and implemented in a Cytoscape-based graphic representation. To demonstrate PLACNET sensitivity and efficacy, the plasmidome of the Escherichia coli lineage ST131 was analyzed. ST131 is a globally spread clonal group of extraintestinal pathogenic E. coli (ExPEC), comprising different sublineages with ability to acquire and spread antibiotic resistance and virulence genes via plasmids. Results show that plasmids flux in the evolution of this lineage, which is wide open for plasmid exchange. MOBF12/IncF plasmids were pervasive, adding just by themselves more than 350 protein families to the ST131 pangenome. Nearly 50% of the most frequent γ-proteobacterial plasmid groups were found to be present in our limited sample of ten analyzed ST131 genomes, which represent the main ST131 sublineages.

  20. Plasmid flux in Escherichia coli ST131 sublineages, analyzed by plasmid constellation network (PLACNET, a new method for plasmid reconstruction from whole genome sequences.

    Directory of Open Access Journals (Sweden)

    Val F Lanza

    2014-12-01

    Full Text Available Bacterial whole genome sequence (WGS methods are rapidly overtaking classical sequence analysis. Many bacterial sequencing projects focus on mobilome changes, since macroevolutionary events, such as the acquisition or loss of mobile genetic elements, mainly plasmids, play essential roles in adaptive evolution. Existing WGS analysis protocols do not assort contigs between plasmids and the main chromosome, thus hampering full analysis of plasmid sequences. We developed a method (called plasmid constellation networks or PLACNET that identifies, visualizes and analyzes plasmids in WGS projects by creating a network of contig interactions, thus allowing comprehensive plasmid analysis within WGS datasets. The workflow of the method is based on three types of data: assembly information (including scaffold links and coverage, comparison to reference sequences and plasmid-diagnostic sequence features. The resulting network is pruned by expert analysis, to eliminate confounding data, and implemented in a Cytoscape-based graphic representation. To demonstrate PLACNET sensitivity and efficacy, the plasmidome of the Escherichia coli lineage ST131 was analyzed. ST131 is a globally spread clonal group of extraintestinal pathogenic E. coli (ExPEC, comprising different sublineages with ability to acquire and spread antibiotic resistance and virulence genes via plasmids. Results show that plasmids flux in the evolution of this lineage, which is wide open for plasmid exchange. MOBF12/IncF plasmids were pervasive, adding just by themselves more than 350 protein families to the ST131 pangenome. Nearly 50% of the most frequent γ-proteobacterial plasmid groups were found to be present in our limited sample of ten analyzed ST131 genomes, which represent the main ST131 sublineages.

  1. IncA/C plasmids harboured in serious multidrug-resistant Vibrio cholerae serogroup O139 strains in China.

    Science.gov (United States)

    Wang, Ruibai; Yu, Dong; Zhu, Lianhui; Li, Jie; Yue, Junjie; Kan, Biao

    2015-03-01

    Vibrio cholerae serogroup O139 emerged in 1992 and is one of two major serogroups to have caused cholera epidemics. After 1998, serious multidrug-resistant (MDR) O139 strains quickly became common in China, showing a multidrug resistance profile to eight antibiotics. It is a great threat to public health, and elucidation of its mechanisms of resistance will provide a helpful guide for the clinical treatment and prevention of cholera. In this study, mega-plasmids from MDR V. cholerae O139 strains were identified by pulsed-field gel electrophoresis (PFGE) without enzyme digestion. One plasmid was isolated and sequenced, belonging to the IncA/C family. Ten antibiotic resistance genes were found in the MDR regions, including a blaTEM-20 gene, and these genes endowed the host with resistance to seven antibiotics. This kind of plasmid was positive in 71.2% (198/278) of toxigenic O139 strains, and the rate of plasmid positivity was consistent with the yearly change in MDR rates of these strains. This study reveals an important role of the IncA/C family plasmid in the spread of multiple antibiotic resistance of epidemic V. cholerae serogroup O139 strains, which has recombined with plasmids from different bacterial species and transferred among V. cholerae strains. Copyright © 2014 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

  2. Detection of an IncA/C plasmid encoding VIM-4 and CMY-4 β-lactamases in Klebsiella oxytoca and Citrobacter koseri from an inpatient in a cardiac rehabilitation unit.

    Science.gov (United States)

    Caltagirone, Mariasofia; Bitar, Ibrahim; Piazza, Aurora; Spalla, Melissa; Nucleo, Elisabetta; Navarra, Antonella; Migliavacca, Roberta

    2015-07-01

    A 62-year-old patient was transferred to the cardiac rehabilitation unit of the I.R.C.C.S. Fondazione S. Maugeri after undergoing a heart transplantation at the Acute Care Hospital I.R.C.C.S. S. Matteo of Pavia. On 1 August 2013 and during hospitalization in the rehabilitation unit, Klebsiella oxytoca and Citrobacter koseri clinical isolates were simultaneously recovered from the patient's preputial swab. Both the K. oxytoca and C. koseri strains were carbapenem- resistant by MicroScan System (Beckman Coulter). Carbapenem-resistant K. pneumoniae had previously been reported in the same rehabilitation facility. The aim of the study was to identify the carbapenem resistance mechanisms among the enterobacterial species recovered. Phenotypic screening tests useful to detect the β-lactamases/carbapenemases were performed. Carbapenem MICs were obtained by Etest. AmpC and MBL encoding genes were identified by PCR and sequencing. Conjugation assays and plasmid characterization were performed. Both of the K. oxytoca and C. koseri isolates were multi drug resistant, showing resistance to amoxicillin-clavulanic acid, three generation cephalosporins, ertapenem (K. oxytoca MIC, >32 mg/L; C. koseri MIC, 4 mg/L), imipenem (K. oxytoca MIC, 4 mg/L; C. koseri MIC, 12 mg/L), thrimethoprim sulphamethoxazole and gentamicin. Susceptibility was retained to fluoroquinolones, colistin and tigecycline. Molecular characterization confirmed the co-presence of blaCMY-4 and blaVIM-4 determinants in a 150 Kb transferable plasmid of IncA/C group. This case is the first detection in Italy of the K. oxytoca and C. koseri clinical isolates co-producing the CMY-4 and VIM-4 enzymes.

  3. An abyssal mobilome: Viruses, plasmids and vesicles from deep-sea hydrothermal vents

    OpenAIRE

    Lossouarn, Julien; Dupont, Samuel; Gorlas, Aurore; Mercier, Coraline; Bienvenu, Nadege; Marguet, Evelyne; Forterre, Patrick; Geslin, Claire

    2015-01-01

    Mobile genetic elements (MGEs) such as viruses, plasmids, vesicles, gene transfer agents (GTAs), transposons and transpovirions, which collectively represent the mobilome, interact with cellular organisms from all three domains of life, including those thriving in the most extreme environments. While efforts have been made to better understand deep-sea vent microbial ecology, our knowledge of the mobilome associated with prokaryotes inhabiting deep-sea hydrothermal vents remains limited. Here...

  4. Molecular analysis of an integrative conjugative element, ICEH, present in the chromosome of different strains of Mycoplasma hyopneumoniae

    Directory of Open Access Journals (Sweden)

    Paulo Marcos Pinto

    2007-01-01

    Full Text Available Diversification of bacterial species and pathotypes is largely caused by lateral gene transfer (LGT of diverse mobile DNA elements such as plasmids, phages, transposons and genomic islands. Thus, acquisition of new phenotypes by LGT is very important for bacterial evolution and relationship with hosts. This paper reports a 23 kb region containing fourteen CDSs with similarity to the previous described Integrative Conjugal Element of Mycoplasma fermentans (ICEF. This element, named ICEH, is present as one copy at distinct integration sites in the chromosome of 7448 and 232 pathogenic strains and is absent in the type strain J (non-pathogenic. Notable differences in the nucleotide composition of the insertion sites were detected, and could be correlated to a lack of specificity of the ICEH integrase. Although present in strains of the same organism, the ICEH elements are more divergent than the typical similarity between other chromosomal locus of Mycoplasma hyopneunomiae, suggesting an accelerated evolution of these constins or an ongoing process of degeneration, while maintaining conservation of the tra genes. An extrachromosomal form of this element had been detected in the 7448 strain, suggesting a possible involvement in its mobilization and transference of CDSs to new hosts.

  5. Abortive phage-infection and UV-protection markers on ColI plasmids from epidemic strains of Salmonella

    International Nuclear Information System (INIS)

    Barker, R.M.

    1988-01-01

    Cultures of Escherichia coli carrying ColI plasmids received in conjugation from strains of Salmonella typhimurium and S. agona were examined for abortive infection (Abi) of phage BF23 and for enhanced resistance to the lethal action of UV-irradiation (Uvr). The Abi character of stored cultures of E. coli was also compared with the reaction of the same stock culture tested 5 years before. Seven of the eight potential types differentiated by three characters were represented among 160 ColI plasmids: ColIa Abi + Uvr + (3 plasmids), ColIa Abi - Uvr + (1), ColIa Abi - Uvr-> (2), ColIb Abi + Uvr + (85), ColIb Abi + Uvr - (5), ColIb Abi - Uvr + (4), ColIb Abi-? Uvr - (60). Recognition that different plasmid types could be carried by strains of a clone proved useful in the interpretation of the epidemic spread of strains of S. typhimurium of phage type/biotype 141/9f in Scotland and in tracing the ancestry of a recently emerged rhamnose non-fermenting mutant strain of S. agona. (author)

  6. [Isolation and characterization of petroleum catabolic broad-host-range plasmids from Shen-Fu wastewater irrigation zone].

    Science.gov (United States)

    Wang, Ya-Fei; Wang, Ya-Fei; Li, Hui; Li, Xiao-Bin

    2013-11-01

    Based on triparental mating, we isolated a total of eight broad host range (BHR) petroleum hydrocarbon catabolic plasmids from the soils, sediments, and wastewater samples in the Shen-Fu irrigation zone. The antibiotic resistance of the plasmids was tested, and then, the plasmids were transferred to Escherichia coli EC100. The plasmids carrying no antibiotic resistance were tagged by miniTn5 transposon consisting of antibiotic resistant genes. The PCR-based incompatibility test revealed that the pS3-2C and pS4-6G belonged to Inc P group, the pS3-2G, pW22-3G, and pA15-7G belonged to Inc N group, the pS7-2G was identified as Inc W plasmid, and the pA23-1G and pA10-1C were placed into Inc Q group. By adopting the reported PCR amplification methods of petroleum hydrocarbon-degrading catabolic genes, the petroleum-degrading capability of these BHR plasmids were preliminarily analyzed. The plasmids pS3-2G, pS7-2G, pA23-1G, pW22-3G, and pA10-1C carried aromatic ring- hydroxylating dioxygenase gene phdA and toluene monooxygenase gene touA; the plasmid pA15-7G carried touA and toluene dioxygenase gene tod; the plasmid pS3-2C carried ben, phdA, and tod; whereas the pS4-6G only carried ben. The host range test showed that all the isolated plasmids except pS3-2C could be transferred and maintained stably in the representative strains Agrobacterium tumefaciens C58, Cupriavidus necator JMP228, and E. coli EC100 of the alpha-, beta-, and gamma-Proteobacteria, respectively.

  7. CARTOGRAPHIE DU PLASMIDE pSU100, PLASMIDE CRYPTIQUE DE LACTOBACILLUS CASEI

    Directory of Open Access Journals (Sweden)

    F BENSALAH

    2003-06-01

    Ce plasmide appelé pSU100 a été cloné dans le vecteur de transformation pUC18 au site EcoRI chez E. coli JM103. Les profils électrophorétiques de restriction obtenus par des digestions simples, doubles et triples sous l’action de 33 endonucléases, ont contribué à l’élaboration d’une carte de restriction de ce plasmide. Cinq sites uniques ont été identifiés, ainsi que d’autres sites doubles et multiples. Une étude préliminaire du rôle physiologique de ce plasmide a permis de déceler une résistance à la kanamycine.

  8. A new approach to produce amino-carbon nanotubes as plasmid transfection vector by [2 + 1] cycloaddition of nitrenes

    International Nuclear Information System (INIS)

    Jiang Yongjian; Jin Chen; Yang Feng; Yu Xianjun; Wang Guojian; Cheng Si; Di, Yang; Li, Ji; Fu, Deliang; N, Quanxing

    2011-01-01

    Amino-carbon nanotubes (amino-CNTs) can conjugate with the DNA by electrostatic interactions and shuttle the DNA to the cell cytoplasm or even the nucleus. Here we report a new approach to produce amino-CNTs by cycloaddition of nitrenes. Fourier transform infrared spectroscopy was used to verify the success of the functionalization, and the functionalization degree was calculated by thermal gravity analysis. Transmission electron microscope (TEM) was used to observe the solubility of the CNTs and the interactions of the amino-CNTs with the plasmids. Cell ultrathin sections were made and observed under the TEM to confirm the amino-CNTs enter the cells. Transfection experiments ultimately verify the amino-CNTs produced through cycloadditions of nitrenes can serve as plasmid vector.

  9. Homology among tet determinants in conjugative elements of streptococci

    Energy Technology Data Exchange (ETDEWEB)

    Smith, M.D.; Hazum, S.; Guild, W.R.

    1981-10-01

    A mutation to tetracycline sensitivity in a resistant strain of Streptococcus pneumoniae was shown by several criteria to be due to a point mutation in the conjugative o(cat-tet) element found in the chromosomes of strains derived from BM6001, a clinical strain resistant to tetracycline and chloramphenicol. Strains carrying the mutation were transformed back to tetracycline resistance with the high efficiency of a point marker by donor deoxyribonucleic acids from its ancestral strain and from nine other clinical isolates of pneumococcus and by deoxyribonucleic acids from Group D Streptococcus faecalis and Group B Streptococcus agalactiae strains that also carry conjugative tet elements in their chromosomes. It was not transformed to resistance by tet plasmid deoxyribonucleic acids from either gram-negative or gram-positive species, except for one that carried transposon TN916, the conjugative tet element present in the chromosomes of some S. faecalis strains. The results showed that the tet determinants in conjugative elements of several streptococcal species share a high degree of deoxyribonucleic acid sequence homology and suggested that they differ from other tet genes.

  10. Mosaic Structure of a Multiple-Drug-Resistant, Conjugative Plasmid from Campylobacter jejuni

    National Research Council Canada - National Science Library

    Nirdnoy, Warawadee; Mason, Carl J; Guerry, Patricia

    2005-01-01

    ..., where it apparently integrated into the chromosome and expressed high-level resistance to multiple aminoglycoside antibiotics. This work provides new information about both the nature of drug resistance in C...

  11. Folic acid conjugated mPEG-PEI600 as an efficient non-viral vector for targeted nucleic acid delivery.

    Science.gov (United States)

    Xu, Zhenhua; Jin, Jiefu; Siu, Leo K S; Yao, Hong; Sze, Johnny; Sun, Hongzhe; Kung, Hsiang-Fu; Poon, Wai Sang; Ng, Samuel S M; Lin, Marie C

    2012-04-15

    In this study we describe a novel polymer, mPPS-FA, synthesized as a potential gene transfer vector. To complete mPPS-FA, folic acid was conjugated to a backbone (named mPPS) consisting of a copolymer of methyl PEG-2000, PEI-600, and sebacoyl chloride. (1)H NMR, FT-IR, and UV spectroscopy were used to characterize the structure of mPPS-FA. It was revealed that mPPS-FA holds the ability to bind plasmid DNA yielding positively charged particles (polyplexes). Dynamic light scattering (DLS) and TEM techniques were used to study the size and morphology of the formed mPPS-FA/DNA nanocomplexes. The mPPS-FA/DNA nanoparticles exhibited low cytotoxicity as transfection of B16-F0, U87MG, CHO-1, and Ho-8910 cells produced >80% viability indicating low cytotoxicity of the polymer. The ability of mPPS-FA to deliver EGFP plasmid to melanoma B16-F0, U87, CHO-1, Ho-8910, and A549 cells was investigated in vitro as compared to the lipid-based transfection agent Lipofectamine2000 and Linear PEI 22 kDa (L-PEI 22 kDa). We found that mPPS-FA/DNA complexes yielded the highest GFP transfection efficiency in B16-F0, U87, CHO-1, and Ho-8910 cells, which all highly express folate receptors (FR), at an mPPS-FA/DNA ratio (w/w) of 15. Furthermore, the transfection of mPPS-FA/DNA complexes in CHO-1 cells could be competitively blocked by free folic acid molecules. In contrast, in low FR expressing A549 cells, mPPS-FA showed similar low transfection efficiency as mPPS. Taken together, mPPS-FA showed the highest efficiency in vitro and the potential to be developed as a nonviral gene carrier. Copyright © 2012 Elsevier B.V. All rights reserved.

  12. Peptide-Carrier Conjugation

    DEFF Research Database (Denmark)

    Hansen, Paul Robert

    2015-01-01

    To produce antibodies against synthetic peptides it is necessary to couple them to a protein carrier. This chapter provides a nonspecialist overview of peptide-carrier conjugation. Furthermore, a protocol for coupling cysteine-containing peptides to bovine serum albumin is outlined....

  13. Photoluminescence in conjugated polymers

    International Nuclear Information System (INIS)

    Furst, J.E.; Laugesen, R.; Dastoor, P.; McNeill, C.

    2002-01-01

    Full text: Conjugated polymers combine the electronic and optical properties of semiconductors with the processability of polymers. They contain a sequence of alternate single and double carbon bonds so that the overlap of unhybridised p z orbitals creates a delocalised ρ system which gives semiconducting properties with p-bonding (valence) and p* -antibonding (conduction) bands. Photoluminesence (PL) in conjugated polymers results from the radiative decay of singlet excitons confined to a single chain. The present work is the first in a series of studies in our laboratory that will characterize the optical properties of conjugated polymers. The experiment involves the illumination of thin films of conjugated polymer with UV light (I=360 nm) and observing the subsequent fluorescence using a custom-built, fluorescence spectrometer. Photoluminesence spectra provide basic information about the structure of the polymer film. A typical spectrum is shown in the accompanying figure. The position of the first peak is related to the polymer chain length and resolved multiple vibronic peaks are an indication of film structure and morphology. We will also present results related to the optical degradation of these materials when exposed to air and UV light

  14. DNA-cell conjugates

    Science.gov (United States)

    Hsiao, Shih-Chia; Francis, Matthew B.; Bertozzi, Carolyn; Mathies, Richard; Chandra, Ravi; Douglas, Erik; Twite, Amy; Toriello, Nicholas; Onoe, Hiroaki

    2018-05-15

    The present invention provides conjugates of DNA and cells by linking the DNA to a native functional group on the cell surface. The cells can be without cell walls or can have cell walls. The modified cells can be linked to a substrate surface and used in assay or bioreactors.

  15. DNA-cell conjugates

    Science.gov (United States)

    Hsiao, Shih-Chia; Francis, Matthew B.; Bertozzi, Carolyn; Mathies, Richard; Chandra, Ravi; Douglas, Erik; Twite, Amy; Toriello, Nicholas; Onoe, Hiroaki

    2016-05-03

    The present invention provides conjugates of DNA and cells by linking the DNA to a native functional group on the cell surface. The cells can be without cell walls or can have cell walls. The modified cells can be linked to a substrate surface and used in assay or bioreactors.

  16. Drug resistance plasmids in Lactobacillus acidophilus and Lactobacillus reuteri.

    OpenAIRE

    Vescovo, M; Morelli, L; Bottazzi, V

    1982-01-01

    Sixteen strains of Lactobacillus reuteri and 20 strains of Lactobacillus acidophilus were tested for resistance to 22 antibiotics by using commercially available sensitivity disks. Evidence suggesting linkage of these resistances to plasmids was obtained by "curing" experiments with acridine dyes and high growth temperatures. Examination of plasmid patterns of agarose gel electrophoresis provided further evidence of loss in plasmid DNA under curing conditions in some of the strains examined.

  17. ICESag37, a Novel Integrative and Conjugative Element Carrying Antimicrobial Resistance Genes and Potential Virulence Factors in Streptococcus agalactiae.

    Science.gov (United States)

    Zhou, Kaixin; Xie, Lianyan; Han, Lizhong; Guo, Xiaokui; Wang, Yong; Sun, Jingyong

    2017-01-01

    ICE Sag37 , a novel integrative and conjugative element carrying multidrug resistance and potential virulence factors, was characterized in a clinical isolate of Streptococcus agalactiae . Two clinical strains of S. agalactiae , Sag37 and Sag158, were isolated from blood samples of new-borns with bacteremia. Sag37 was highly resistant to erythromycin and tetracycline, and susceptible to levofloxacin and penicillin, while Sag158 was resistant to tetracycline and levofloxacin, and susceptible to erythromycin. Transfer experiments were performed and selection was carried out with suitable antibiotic concentrations. Through mating experiments, the erythromycin resistance gene was found to be transferable from Sag37 to Sag158. Sma I-PFGE revealed a new Sma I fragment, confirming the transfer of the fragment containing the erythromycin resistance gene. Whole genome sequencing and sequence analysis revealed a mobile element, ICE Sag37 , which was characterized using several molecular methods and in silico analyses. ICE Sag37 was excised to generate a covalent circular intermediate, which was transferable to S. agalactiae . Inverse PCR was performed to detect the circular form. A serine family integrase mediated its chromosomal integration into rumA , which is a known hotspot for the integration of streptococcal ICEs. The integration site was confirmed using PCR. ICE Sag37 carried genes for resistance to multiple antibiotics, including erythromycin [ erm(B) ], tetracycline [ tet(O) ], and aminoglycosides [ aadE, aphA , and ant(6) ]. Potential virulence factors, including a two-component signal transduction system ( nisK/nisR ), were also observed in ICE Sag37 . S1-PFGE analysis ruled out the existence of plasmids. ICE Sag37 is the first ICE Sa2603 family-like element identified in S. agalactiae carrying both resistance and potential virulence determinants. It might act as a vehicle for the dissemination of multidrug resistance and pathogenicity among S. agalactiae .

  18. Plasmid P1 replication: negative control by repeated DNA sequences.

    OpenAIRE

    Chattoraj, D; Cordes, K; Abeles, A

    1984-01-01

    The incompatibility locus, incA, of the unit-copy plasmid P1 is contained within a fragment that is essentially a set of nine 19-base-pair repeats. One or more copies of the fragment destabilizes the plasmid when present in trans. Here we show that extra copies of incA interfere with plasmid DNA replication and that a deletion of most of incA increases plasmid copy number. Thus, incA is not essential for replication but is required for its control. When cloned in a high-copy-number vector, pi...

  19. Generation of a CRISPR database for Yersinia pseudotuberculosis complex and role of CRISPR-based immunity in conjugation.

    Science.gov (United States)

    Koskela, Katja A; Mattinen, Laura; Kalin-Mänttäri, Laura; Vergnaud, Gilles; Gorgé, Olivier; Nikkari, Simo; Skurnik, Mikael

    2015-11-01

    The clustered regularly interspaced short palindromic repeat - CRISPR-associated genes (CRISPR-Cas) system is used by bacteria and archaea against invading conjugative plasmids or bacteriophages. Central to this immunity system are genomic CRISPR loci that contain fragments of invading DNA. These are maintained as spacers in the CRISPR loci between direct repeats and the spacer composition in any bacterium reflects its evolutionary history. We analysed the CRISPR locus sequences of 335 Yersinia pseudotuberculosis complex strains. Altogether 1902 different spacer sequences were identified and these were used to generate a database for the spacer sequences. Only ∼10% of the spacer sequences found matching sequences. In addition, surprisingly few spacers were shared by Yersinia pestis and Y. pseudotuberculosis strains. Interestingly, 32 different protospacers were present in the conjugative plasmid pYptb32953. The corresponding spacers were identified from 35 different Y. pseudotuberculosis strains indicating that these strains had encountered pYptb32953 earlier. In conjugation experiments, pYptb32953-specific spacers generally prevented conjugation with spacer-positive and spacer-free strains. However, some strains with one to four spacers were invaded by pYptb32953 and some spacer-free strains were fully resistant. Also some spacer-positive strains were intermediate resistant to conjugation. This suggests that one or more other defence systems are determining conjugation efficiency independent of the CRISPR-Cas system. © 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.

  20. Resistant plasmid profile analysis of multidrug resistant Escherichia ...

    African Journals Online (AJOL)

    Multiple drug resistance isolates causing UTI has seri- ous implications for the empiric therapy against patho- genic isolates and for the possible co-selection of antimicrobial resistant mediated by multi drug resistant plasmids21,22. E. coli from clinical isolates are known to harbour plasmids of different molecular sizes23.

  1. Application of methylation in improving plasmid transformation into Helicobacter pylori.

    Science.gov (United States)

    Zhao, Huilin; Xu, Linlin; Rong, Qianyu; Xu, Zheng; Ding, Yunfei; Zhang, Ying; Wu, Yulong; Li, Boqing; Ji, Xiaofei

    2018-05-23

    Helicobacter pylori is an important gastrointestinal pathogen. Its strains possess different levels of powerful restriction modification systems, which are significant barriers to genetic tools used for studying the role of functional genes in its pathogenesis. Methylating vectors in vitro was reported as an alternative to overcome this barrier in several bacteria. In this study we used two H. pylori-E. coli shuttle plasmids and several single/double-crossover homologous recombination gene-targeting plasmids, to test the role of methylation in H. pylori transformation. According to our results, transformants could be obtained only after shuttle plasmids were methylated before transformation. It is helpful in gene complementation and over-expression although at a low frequency. The frequency of gene-targeting transformation was also increased after methylation, especially for the single-crossover recombination plasmids, the transformants of which could only be obtained after methylation. For the double-crossover recombination targeting plasmids, the initial yield of transformants was 0.3-0.8 × 10 2 CFUs per microgram plasmid DNA. With the help of methylation, the yield was increased to 0.4-1.3 × 10 2 CFUs per microgram plasmid DNA. These results suggest that in vitro methylation can improve H. pylori transformation by different plasmids, which will benefit the pathogenic mechanism research. Copyright © 2018. Published by Elsevier B.V.

  2. Functional analysis of three plasmids from Lactobacillus plantarum

    NARCIS (Netherlands)

    Kranenburg, R. van; Golic, N.; Bongers, R.; Leer, R.J.; Vos, W.M. de; Siezen, R.J.; Kleerebezem, M.

    2005-01-01

    Lactobacillus plantarum WCFS1 harbors three plasmids, pWCFS101, pWCFS102, and pWCFS103, with sizes of 1,917, 2,365, and 36,069 bp, respectively. The two smaller plasmids are of unknown function and contain replication genes that are likely to function via the rolling-circle replication mechanism.

  3. Production and pharmaceutical formulation of plasmid DNA vaccines

    NARCIS (Netherlands)

    van der Heijden, I.

    2013-01-01

    Research leading to the thesis ‘Production and pharmaceutical formulation of plasmid DNA vaccines‘ can be divided into two parts. The first part describes the development of a Good Manufacturing Practice (GMP) compliant plasmid DNA production process of pDNA vaccines for the treatment of Human

  4. The technology of large-scale pharmaceutical plasmid purification ...

    African Journals Online (AJOL)

    Further test demonstrated that the pcDNAlacZ purified with CTAB and authoritative endotoxin-free plasmid Kit had the similar transfection efficiency in vivo and in vitro. CTAB can be used for plasmid purification; the main advantages of the DNAs purified with CTAB include the avoidance of animal-derived enzymes, toxic ...

  5. Identification of IncA/C Plasmid Replication and Maintenance Genes and Development of a Plasmid Multilocus Sequence Typing Scheme.

    Science.gov (United States)

    Hancock, Steven J; Phan, Minh-Duy; Peters, Kate M; Forde, Brian M; Chong, Teik Min; Yin, Wai-Fong; Chan, Kok-Gan; Paterson, David L; Walsh, Timothy R; Beatson, Scott A; Schembri, Mark A

    2017-02-01

    Plasmids of incompatibility group A/C (IncA/C) are becoming increasingly prevalent within pathogenic Enterobacteriaceae They are associated with the dissemination of multiple clinically relevant resistance genes, including bla CMY and bla NDM Current typing methods for IncA/C plasmids offer limited resolution. In this study, we present the complete sequence of a bla NDM-1 -positive IncA/C plasmid, pMS6198A, isolated from a multidrug-resistant uropathogenic Escherichia coli strain. Hypersaturated transposon mutagenesis, coupled with transposon-directed insertion site sequencing (TraDIS), was employed to identify conserved genetic elements required for replication and maintenance of pMS6198A. Our analysis of TraDIS data identified roles for the replicon, including repA, a toxin-antitoxin system; two putative partitioning genes, parAB; and a putative gene, 053 Construction of mini-IncA/C plasmids and examination of their stability within E. coli confirmed that the region encompassing 053 contributes to the stable maintenance of IncA/C plasmids. Subsequently, the four major maintenance genes (repA, parAB, and 053) were used to construct a new plasmid multilocus sequence typing (PMLST) scheme for IncA/C plasmids. Application of this scheme to a database of 82 IncA/C plasmids identified 11 unique sequence types (STs), with two dominant STs. The majority of bla NDM -positive plasmids examined (15/17; 88%) fall into ST1, suggesting acquisition and subsequent expansion of this bla NDM -containing plasmid lineage. The IncA/C PMLST scheme represents a standardized tool to identify, track, and analyze the dissemination of important IncA/C plasmid lineages, particularly in the context of epidemiological studies. Copyright © 2017 American Society for Microbiology.

  6. Expansion of the IncX plasmid family for improved identification and typing of novel plasmids in drug-resistant Enterobacteriaceae

    DEFF Research Database (Denmark)

    Johnson, Timothy J.; Bielak, Eliza Maria; Fortini, Daniela

    2012-01-01

    and biofilm formation. Previous plasmid-based replicon typing procedures have indicated that the prevalence of IncX plasmids is low among members of the Enterobacteriaceae. However, examination of a number of IncX-like plasmid sequences and their occurrence in various organisms suggests that IncX plasmid...

  7. Plasmid-mediated UV-protection in Streptococcus lactis

    International Nuclear Information System (INIS)

    Chopin, M.-C.; Rouault, A.

    1985-01-01

    Streptococcus lactis strain IL594 contains 9 plasmids, designated pIL1 to pIL9. On the basis of protoplast-induced curing experiments the authors showed that derivatives containing pIL7 were resistant to UV-irradiation while derivatives lacking pIL7 were sensitive. The pIL7-determined UV-protection was confirmed by cotransfer of the plasmid and of the character into a plasmid-free derivative of S. lactis IL594. Moreover, prophage induction required higher UV-fluence in this derivative carrying pIL7 than in the plasmid-free strain. This is the first report of a plasmid-mediated UV-protection in group N streptococci. (orig.)

  8. Characterization of the IncA/C plasmid pSCEC2 from Escherichia coli of swine origin that harbours the multiresistance gene cfr.

    Science.gov (United States)

    Zhang, Wan-Jiang; Xu, Xing-Ran; Schwarz, Stefan; Wang, Xiu-Mei; Dai, Lei; Zheng, Hua-Jun; Liu, Siguo

    2014-02-01

    To determine the complete nucleotide sequence of the multidrug resistance plasmid pSCEC2, isolated from a porcine Escherichia coli strain, and to analyse it with particular reference to the cfr gene region. Plasmid pSCEC2 was purified from its E. coli J53 transconjugant and then sequenced using the 454 GS-FLX System. After draft assembly, predicted gaps were closed by PCR with subsequent sequencing of the amplicons. Plasmid pSCEC2 is 135 615 bp in size and contains 200 open reading frames for proteins of ≥100 amino acids. Analysis of the sequence of pSCEC2 revealed two resistance gene segments. The 4.4 kb cfr-containing segment is flanked by two IS256 elements in the same orientation, which are believed to be involved in the dissemination of the rRNA methylase gene cfr. The other segment harbours the resistance genes floR, tet(A)-tetR, strA/strB and sul2, which have previously been found on other IncA/C plasmids. Except for these two resistance gene regions, the pSCEC2 backbone displayed >99% nucleotide sequence identity to that of other IncA/C family plasmids isolated in France, Chile and the USA. The cfr gene was identified on an IncA/C plasmid, which is well known for its broad host range and transfer and maintenance properties. The location on such a plasmid will further accelerate the dissemination of cfr and co-located resistance genes among different Gram-negative bacteria. The genetic context of cfr on plasmid pSCEC2 underlines the complexity of cfr transfer events and confirms the role that insertion sequences play in the spread of cfr.

  9. Physical structure and genetic expression of the sulfonamide-resistance plasmid pLS80 and its derivatives in Streptococcus pneumoniae and Bacillus subtilis

    Energy Technology Data Exchange (ETDEWEB)

    Lopez, P.; Espinosa, M.; Lacks, S.A.

    1984-01-01

    The 10-kb chromosomal fragment of Streptococcus pneumoniae cloned in pLS80 contains the sul-d allele of the pneumococcal gene for dihydropteroate synthase. As a single copy in the chromosome this allele confers resistance to sulfanilamide at 0.2 mg/ml; in the multicopy plasmid it confers resistance to 2.0 mg/ml. The sul-d mutation was mapped by restriction analysis to a 0.4-kb region. A spontaneous deletion beginning approx. 1.5 kb to the right of the sul-d mutation prevented gene function, possibly by removing a promoter. This region could be restored by chromosomal facilitation and be demonstrated in the plasmid by selection for sulfonamide resistance. Under selection for a vector marker, tetracycline resistance, only the deleted plasmid was detectable, apparently as a result of plasmid segregation and the advantageous growth rates of cells with smaller plasmids. When such cells were selected for sulfonamide resistance, the deleted region returned to the plasmid, presumably by equilibration between the chromosome and the plasmid pool, to give a low frequency (approx. 10/sup -3/) of cells resistant to sulfanilamide at 2.0 mg/ml. Models for the mechanisms of chromosomal facilitation and equilibration are proposed. Several derivatives of pLS80 could be transferred to Bacillus subtilis, where they conferred resistance to sulfanilamide at 2 mg/ml, thereby demonstrating cross-species expression of the pneumococcal gene. Transfer of the plasmids to B. subtilis gave rise to large deletions to the left of the sul-d marker, but these deletions did not interfere with the sul-d gene function. Restriction maps of pLS80 and its variously deleted derivatives are presented.

  10. plasmid in Saccharomyces species and in Saccharomyces cerevisiae.

    Science.gov (United States)

    Strope, Pooja K; Kozmin, Stanislav G; Skelly, Daniel A; Magwene, Paul M; Dietrich, Fred S; McCusker, John H

    2015-12-01

    We determined that extrachromosomal 2μ plasmid was present in 67 of the Saccharomyces cerevisiae 100-genome strains; in addition to variation in the size and copy number of 2μ, we identified three distinct classes of 2μ. We identified 2μ presence/absence and class associations with populations, clinical origin and nuclear genotypes. We also screened genome sequences of S. paradoxus, S. kudriavzevii, S. uvarum, S. eubayanus, S. mikatae, S. arboricolus and S. bayanus strains for both integrated and extrachromosomal 2μ. Similar to S. cerevisiae, we found no integrated 2μ sequences in any S. paradoxus strains. However, we identified part of 2μ integrated into the genomes of some S. uvarum, S. kudriavzevii, S. mikatae and S. bayanus strains, which were distinct from each other and from all extrachromosomal 2μ. We identified extrachromosomal 2μ in one S. paradoxus, one S. eubayanus, two S. bayanus and 13 S. uvarum strains. The extrachromosomal 2μ in S. paradoxus, S. eubayanus and S. cerevisiae were distinct from each other. In contrast, the extrachromosomal 2μ in S. bayanus and S. uvarum strains were identical with each other and with one of the three classes of S. cerevisiae 2μ, consistent with interspecific transfer. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  11. Plasmid borne Carbapenem-Hydrolyzing Class D β-Lactamases (CHDLs) and AdeABC efflux pump conferring carbapenem-tigecycline resistance among Acinetobacter baumannii isolates harboring TnAbaRs.

    Science.gov (United States)

    Savari, Mohammad; Ekrami, Alireza; Shoja, Saeed; Bahador, Abbas

    2017-03-01

    Here we studied the prevalence and mechanisms of simultaneous resistance to carbapenem and tigecycline and accumulation of resistance determinants reservoirs in genome of Acinetobacter baumannii (A. baumannii) clinical isolates. Susceptibility of the isolates were measured to 18 antimicrobial agents. Genetic diversity of the microbial population was determined using the International Clonal lineage typing (IC typing), multiple locus VNTR analysis (MLVA) and plasmid profiling methods. To detect the AbaRs, Carbapenem-Hydrolyzing Class D β-Lactamases (CHDLs) genes, AdeABC efflux pump genes and resistance determinants, PCR was used. Filter mating experiments were used to prove that if carbapenem resistance genes are located on conjugative plasmids or not. Among the A. baumannii clinical isolates, 40.8% were carbapenem-tigecycline resistant and in this population, 46.9% were belonging to IC I, IC II or IC III and 53.1% were IC variants. These isolates had fallen in 40 MLVA types and were harboring plasmids in multiple numbers and sizes. In this study, bla OXA-23-like was the most prevalent CHDL and conjugation analysis proved that the carbapenem resistance genes are located on conjugative plasmids. All efflux pump genes, except for adeC, were detected in all carbapenem-tigecycline resistant A. baumannii (CTRAb) isolates. Resistance determinants were distributed in both TnAbaRs and R plasmids with a shift toward the R plasmids. Emerging of carbapenem resistant A. baumannii (CRAB) with simultaneous resistance to the last line therapy including tigecycline represent emerging of extensively drug resistance (XDR) and pandrug resistance (PDR) phenotypes that would be a great threat to our public health system. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Restriction Fragment Length Polymorphisms of Virulence Plasmids in Rhodococcus equi

    Science.gov (United States)

    Takai, Shinji; Shoda, Masato; Sasaki, Yukako; Tsubaki, Shiro; Fortier, Guillaume; Pronost, Stephane; Rahal, Karim; Becu, Teotimo; Begg, Angela; Browning, Glenn; Nicholson, Vivian M.; Prescott, John F.

    1999-01-01

    Virulent Rhodococcus equi, which is a well-known cause of pyogranulomatous pneumonia in foals, possesses a large plasmid encoding virulence-associated 15- to 17-kDa antigens. Foal and soil isolates from five countries—Argentina, Australia, Canada, France, and Japan—were investigated for the presence of 15- to 17-kDa antigens by colony blotting, using the monoclonal antibody 10G5, and the gene coding for 15- to 17-kDa antigens by PCR. Plasmid DNAs extracted from positive isolates were digested with restriction endonucleases BamHI, EcoRI, EcoT22I, and HindIII, and the digestion patterns that resulted divided the plasmids of virulent isolates into five closely related types. Three of the five types had already been reported in Canadian and Japanese isolates, and the two new types had been found in French and Japanese isolates. Therefore, we tentatively designated these five types 85-kb type I (pREAT701), 85-kb type II (a new type), 87-kb type I (EcoRI and BamHI type 2 [V. M. Nicholson and J. F. Prescott, J. Clin. Microbiol. 35:738–740, 1997]), 87-kb type II (a new type), and 90-kb (pREL1) plasmids. The 85-kb type I plasmid was found in isolates from Argentina, Australia, Canada, and France. Plasmid 87-kb type I was isolated in specimens from Argentina, Canada, and France. The 85-kb type II plasmid appeared in isolates from France. On the other hand, plasmids 87-kb type II and 90-kb were found only in isolates from Japan. These results revealed geographic differences in the distribution of the virulence plasmids found in the five countries and suggested that the restriction fragment length polymorphism of virulence plasmids might be useful to elucidate the molecular epidemiology of virulent R. equi in the world. PMID:10488224

  13. Development of a chromosome-plasmid balanced lethal system for Lactobacillus acidophilus with thyA gene as selective marker.

    Science.gov (United States)

    Fu, X; Xu, J G

    2000-01-01

    A chromosome-plasmid balanced lethal gene delivery system for Lactobacillus acidophilus based on the thyA gene was developed. The selected L. acidophilus DOM La strain carries a mutated thyA gene and has an obligate requirement for thymidine. This strain can be used as a host for the constructed shuttle vector pFXL03, lacking antibiotic-resistant markers but having the wild-type thyA gene from L. casei which complements the thyA chromosomal mutation. The vector also contains the replicon region from plasmid pUC19 and that of the Lactococcus plasmid pWV01, which allows the transfer between Escherichia coli, L. casei and L. acidophilus. Eight unique restriction sites (i.e., PstI, HindIII, SphI, SalI, AccI, XbaI, KpnI and SacI) are available for cloning. After 40-time transfers in modified MRS medium, no plasmid loss was observed. The vector pFXL03 is potentially useful as a food-grade vaccine delivery system for L. acidophilus.

  14. Rapid, facile synthesis of conjugated polymer zwitterions in ionic liquids

    Energy Technology Data Exchange (ETDEWEB)

    Page, Zachariah A. [Polymer Science & Engineering Department; Conte Center for Polymer Research; University of Massachusetts; Amherst, USA; Liu, Feng [Polymer Science & Engineering Department; Conte Center for Polymer Research; University of Massachusetts; Amherst, USA; Russell, Thomas P. [Polymer Science & Engineering Department; Conte Center for Polymer Research; University of Massachusetts; Amherst, USA; Emrick, Todd [Polymer Science & Engineering Department; Conte Center for Polymer Research; University of Massachusetts; Amherst, USA

    2014-01-01

    Ionic liquids (ILs) were utilized for the rapid air-stable Suzuki polymerization of polar zwitterionic thiophene monomers, precluding the need for volatile organic solvents, phosphine ligands and phase transfer catalysts typically used in conjugated polymer synthesis.

  15. The local transfer of plasmid-mediated human inducible nitric oxide synthase gene with protein-coated metallic stents inhibits intimal hyperplasia following coronary angioplasty in mini-swine model%蛋白涂层支架携带质粒介导的一氧化氮 合酶基因预防冠状动脉球囊扩张后 再狭窄的实验研究

    Institute of Scientific and Technical Information of China (English)

    戴军; 高润霖; 宋来凤; 阮英茆; 陈纪林; 李永力; 孟亮; 唐承君; 史瑞文; 汤键

    2001-01-01

    Objective To assess the effect of local transfer ofplasmid-mediated inducible Nitric Oxide synthase gene using protein-coated metallic stents of inhibiting neointimal hyperplasia after coronary angioplasty. Methods The metallic stent was coated by cross-linked gelatin and mounted on 3.0 mm PTCA balloon,then endotoxin-free ultrapure plasmid pcDNA3hepiNOS was absorbed on the stent. Protein-coated stainless steel stents were used as controls. All stents were implanted into the middle segment of LAD. The ratio of balloon to vessel diameter was 1.1-1.3:1. Results At the 7th day after stenting, RT-PCR and immunohistochemical staining confirmed the expression of iNOS mRNA and presence of its protein at gene transferred vessels (n=2), but there was no expression in remote organs. After 3 months of stenting coronary angiograms showed that there was no restenosis in all animals transferred plasmid pcDNA3hepiNOS(n=5) while restenosis developed in all animals of the control group (n=5). Morphometric analysis showed that lumen diameter loss (0.61±0.30) mm vs (1.58±0.31) mm (P<0.001), residual lumen diameter (1.00±0.51) mm vs (0.36±0.32) mm (P<0.05), neointimal area (1.65±0.83) mm2 vs (2.83±0.83) mm2 (P<0.05), mean percentage of area stenosis (26.45±7.45) mm2 vs (94.2±14.3) mm2 (P<0.001) were significantly less than those in control group. The proportion of the neointimal area to media area(I/M) reduced to 59.84% in pcDNA3hepiNOS group. Conclusions Local plasmid-mediated human iNOS gene transferred with protein-coated metallic stents significantly inhibited intimal hyperplasia,which was usually a causative factor of retenosis after coronary angioplasty, in mini-swine model.%目的 为评价蛋白涂层金属支架携带质粒介导的诱导性一氧化氮合酶(iNOS)基因局部转染血管壁,预防冠状动脉内血管成形术后再狭窄的效果。方法 金属支架涂层为胶联明胶制成。载体为去内毒素纯化pcDNA3。采用标准球囊导管技

  16. Binding mechanisms for histamine and agmatine ligands in plasmid deoxyribonucleic acid purifications.

    Science.gov (United States)

    Sousa, Ângela; Pereira, Patrícia; Sousa, Fani; Queiroz, João A

    2014-10-31

    Histamine and agmatine amino acid derivatives were immobilized into monolithic disks, in order to combine the specificity and selectivity of the ligand with the high mass transfer and binding capacity offered by monolithic supports, to purify potential plasmid DNA biopharmaceuticals. Different elution strategies were explored by changing the type and salt concentration, as well as the pH, in order to understand the retention pattern of different plasmids isoforms The pVAX1-LacZ supercoiled isoform was isolated from a mixture of pDNA isoforms by using NaCl increasing stepwise gradient and also by ammonium sulfate decreasing stepwise gradient, in both histamine and agmatine monoliths. Acidic pH in the binding buffer mainly strengthened ionic interactions with both ligands in the presence of sodium chloride. Otherwise, for histamine ligand, pH values higher than 7 intensified hydrophobic interactions in the presence of ammonium sulfate. In addition, circular dichroism spectroscopy studies revealed that the binding and elution chromatographic conditions, such as the combination of high ionic strength with extreme pH values can reversibly influence the structural stability of the target nucleic acid. Therefore, ascending sodium chloride gradients with pH manipulation can be preferable chromatographic conditions to be explored in the purification of plasmid DNA biopharmaceuticals, in order to avoid the environmental impact of ammonium sulfate. Copyright © 2014. Published by Elsevier B.V.

  17. Characterization of Plasmid-Mediated Quinolone Resistance Determinants in High-Level Quinolone-Resistant Enterobacteriaceae Isolates from the Community: First Report of qnrD Gene in Algeria.

    Science.gov (United States)

    Yanat, Betitera; Machuca, Jesús; Díaz-De-Alba, Paula; Mezhoud, Halima; Touati, Abdelaziz; Pascual, Álvaro; Rodríguez-Martínez, José-Manuel

    2017-01-01

    The objective was to assess the prevalence of plasmid-mediated quinolone resistance (PMQR)-producing isolates in a collection of quinolone-resistant Enterobacteriaceae of community origin isolated in Bejaia, Algeria. A total of 141 nalidixic acid-resistant Enterobacteriaceae community isolates were collected in Bejaia (Northern Algeria) and screened for PMQR genes using polymerase chain reaction (PCR). For PMQR-positive strains, antimicrobial susceptibility testing was performed by broth microdilution and disk diffusion. Mutations in the quinolone resistance-determining regions of the target genes, gyrA and parC, were detected with a PCR-based method and sequencing. Southern blotting, conjugation and transformation assays and molecular typing by pulsed-field gel electrophoresis (PFGE), and multilocus sequence typing were also performed. The prevalence of PMQR-producing Enterobacteriaceae isolates was 13.5% (19/141); 11 of these isolates produced Aac(6')-Ib-cr and 8 were qnr-positive (4 qnrB1-like, 2 qnrS1-like, and 2 qnrD1-like), including the association with aac(6')-Ib-cr gene in three cases. PMQR gene transfer by conjugation was successful in 6 of 19 isolates tested. PFGE revealed that most of the PMQR-positive Escherichia coli isolates were unrelated, except for two groups comprising two and four isolates, respectively, including the virulent multidrug-resistant clone E. coli ST131 that were clonally related. Our findings indicate that PMQR determinants are prevalent in Enterobacteriaceae isolates from the community studied. We describe the first report of the qnrD gene in Algeria.

  18. Emergence of carbapenem resistant Escherichia coli isolates producing blaNDM and blaOXA-48-like carried on IncA/C and IncL/M plasmids at two Iranian university hospitals.

    Science.gov (United States)

    Solgi, Hamid; Giske, Christian G; Badmasti, Farzad; Aghamohammad, Shadi; Havaei, Seyed Asghar; Sabeti, Shahram; Mostafavizadeh, Kamyar; Shahcheraghi, Fereshteh

    2017-11-01

    The emergence of carbapenem resistance among Escherichia coli is a serious threat to public health. The objective of this study was to investigate resistance genes and clonality of carbapenem resistant E. coli in Iran. Between February 2015 and July 2016, a total of 32 non-duplicate E. coli isolates that were ertapenem resistant or intermediate (R/I-ETP) were collected from patient clinical or surveillance cultures (rectal swabs) at two university hospitals. Resistance genes were identified by PCR and sequencing. Conjugation experiments, PCR-based replicon typing, PFGE and multilocus sequence typing (MLST) were performed. PCR assays showed, among the 32 isolates, twenty-nine strains produced carbapenemase genes. The predominant carbapenemase was bla OXA-48 (82.8%), followed by bla NDM-1 (31%), bla NDM-7 (6.9%) and bla OXA-181 (3.4%). Seven of the bla NDM positive isolates co-harbored bla OXA-48 carbapenemases. The bla NDM and bla OXA-48 were found in IncA/C and IncL/M conjugative plasmids, respectively. The bla CTX-M-15 , qnrA and intI1 genes were also present in most isolates. The PFGE revealed genetic diversity among the 28 E. coli isolates, which belonged to six minor PFGE clusters and 14 isolates were singletons. The 26 isolates were distributed into 18 STs, of which two were dominant (ST648 and ST167). We identified one bla NDM-1 -positive ST131 E. coli isolates that harbor the bla CTX-M-15 and bla TEM genes. Horizontal transfer of IncA/C and IncL/M plasmids has likely facilitated the spread of the bla OXA-48 and bla NDM genes among E. coli. Their clonal diversity and the presence of faecal carriers in isolates suggest an endemic spread of OXA-48 and NDM. Therefore, it emphasizes the critical importance of monitoring and controlling the spread of carbapenem resistant E. coli. Copyright © 2017. Published by Elsevier B.V.

  19. Plasmid-associated sensitivity of Bacillus thuringiensis to UV light

    International Nuclear Information System (INIS)

    Benoit, T.G.; Wilson, G.R.; Bull, D.L.; Aronson, A.I.

    1990-01-01

    Spores and vegetative cells of Bacillus thuringiensis were more sensitive to UV light than were spores or cells of plasmid-cured B. thuringiensis strains or of the closely related Bacillus cereus. Introduction of B. thuringiensis plasmids into B. cereus by cell mating increased the UV sensitivity of the cells and spores. Protoxins encoded by one or more B. thuringiensis plasmids were not involved in spore sensitivity, since a B. thuringiensis strain conditional for protoxin accumulation was equally sensitive at the permissive and nonpermissive temperatures. In addition, introduction of either a cloned protoxin gene, the cloning vector, or another plasmid not containing a protoxin gene into a plasmid-cured strain of B. thuringiensis all increased the UV sensitivity of the spores. Although the variety of small, acid-soluble proteins was the same in the spores of all strains examined, the quantity of dipicolinic acid was about twice as high in the plasmid-containing strains, and this may account for the differences in UV sensitivity of the spores. The cells of some strains harboring only B. thuringiensis plasmids were much more sensitive than cells of any of the other strains, and the differences were much greater than observed with spores

  20. Biofilm plasmids with a rhamnose operon are widely distributed determinants of the 'swim-or-stick' lifestyle in roseobacters.

    Science.gov (United States)

    Michael, Victoria; Frank, Oliver; Bartling, Pascal; Scheuner, Carmen; Göker, Markus; Brinkmann, Henner; Petersen, Jörn

    2016-10-01

    Alphaproteobacteria of the metabolically versatile Roseobacter group (Rhodobacteraceae) are abundant in marine ecosystems and represent dominant primary colonizers of submerged surfaces. Motility and attachment are the prerequisite for the characteristic 'swim-or-stick' lifestyle of many representatives such as Phaeobacter inhibens DSM 17395. It has recently been shown that plasmid curing of its 65-kb RepA-I-type replicon with >20 genes for exopolysaccharide biosynthesis including a rhamnose operon results in nearly complete loss of motility and biofilm formation. The current study is based on the assumption that homologous biofilm plasmids are widely distributed. We analyzed 33 roseobacters that represent the phylogenetic diversity of this lineage and documented attachment as well as swimming motility for 60% of the strains. All strong biofilm formers were also motile, which is in agreement with the proposed mechanism of surface attachment. We established transposon mutants for the four genes of the rhamnose operon from P. inhibens and proved its crucial role in biofilm formation. In the Roseobacter group, two-thirds of the predicted biofilm plasmids represent the RepA-I type and their physiological role was experimentally validated via plasmid curing for four additional strains. Horizontal transfer of these replicons was documented by a comparison of the RepA-I phylogeny with the species tree. A gene content analysis of 35 RepA-I plasmids revealed a core set of genes, including the rhamnose operon and a specific ABC transporter for polysaccharide export. Taken together, our data show that RepA-I-type biofilm plasmids are essential for the sessile mode of life in the majority of cultivated roseobacters.

  1. AAVS1-Targeted Plasmid Integration in AAV Producer Cell Lines.

    Science.gov (United States)

    Luo, Yuxia; Frederick, Amy; Martin, John M; Scaria, Abraham; Cheng, Seng H; Armentano, Donna; Wadsworth, Samuel C; Vincent, Karen A

    2017-06-01

    Adeno-associated virus (AAV) producer cell lines are created via transfection of HeLaS3 cells with a single plasmid containing three components (the vector sequence, the AAV rep and cap genes, and a selectable marker gene). As this plasmid contains both the cis (Rep binding sites) and trans (Rep protein encoded by the rep gene) elements required for site-specific integration, it was predicted that plasmid integration might occur within the AAVS1 locus on human chromosome 19 (chr19). The objective of this study was to investigate whether integration in AAVS1 might be correlated with vector yield. Plasmid integration sites within several independent cell lines were assessed via Southern, fluorescence in situ hybridization (FISH) and PCR analyses. In the Southern analyses, the presence of fragments detected by both rep- and AAVS1-specific probes suggested that for several mid- and high-producing lines, plasmid DNA had integrated into the AAVS1 locus. Analysis with puroR and AAVS1-specific probes suggested that integration in AAVS1 was a more widespread phenomenon. High-producing AAV2-secreted alkaline phosphatase (SEAP) lines (masterwell 82 [MW82] and MW278) were evaluated via FISH using probes specific for the plasmid, AAVS1, and a chr19 marker. FISH analysis detected two plasmid integration sites in MW278 (neither in AAVS1), while a total of three sites were identified in MW82 (two in AAVS1). An inverse PCR assay confirmed integration within AAVS1 for several mid- and high-producing lines. In summary, the FISH, Southern, and PCR data provide evidence of site-specific integration of the plasmid within AAVS1 in several AAV producer cell lines. The data also suggest that integration in AAVS1 is a general phenomenon that is not necessarily restricted to high producers. The results also suggest that plasmid integration within the AAVS1 locus is not an absolute requirement for a high vector yield.

  2. Nanoparticles as conjugated delivery agents for therapeutic applications

    Science.gov (United States)

    Muroski, Megan Elizabeth

    This dissertation explores the use of nanoparticles as conjugated delivery agents. Chapter 1 is a general introduction. Chapter 2 discusses the delivery by a nanoparticle platform provides a method to manipulate gene activation, by taking advantage of the high surface area of a nanoparticle and the ability to selectively couple a desired biological moiety to the NP surface. The nanoparticle based transfection approach functions by controlled release of gene regulatory elements from a 6 nm AuNP (gold nanoparticle) surface. The endosomal release of the regulatory elements from the nanoparticle surface results in endogenous protein knockdown simultaneously with exogenous protein expression for the first 48 h. The use of fluorescent proteins as the endogenous and exogenous signals for protein expression enables the efficiency of co-delivery of siRNA (small interfering RNA) for GFP (green fluorescent protein) knockdown and a dsRed-express linearized plasmid for induction to be optically analyzed in CRL-2794, a human kidney cell line expressing an unstable green fluorescent protein. Delivery of the bimodal nanoparticle in cationic liposomes results in 20% GFP knockdown within 24 h of delivery and continues exhibiting knockdown for up to 48 h for the bimodal agent. Simultaneous dsRed expression is observed to initiate within the same time frame with expression levels reaching 34% after 25 days although cells have divided approximately 20 times, implying daughter cell transfection has occurred. Fluorescence cell sorting results in a stable colony, as demonstrated by Western blot analysis. The simultaneous delivery of siRNA and linearized plasmid DNA on the surface of a single nanocrystal provides a unique method for definitive genetic control within a single cell and leads to a very efficient cell transfection protocol. In Chapter 3, we wanted to understand the NP complex within the cell, and to look at the dynamics of release utilizing nanometal surface energy transfer as

  3. Genetic islands in pome fruit pathogenic and nonpathogenic Erwinia species and related plasmids

    Directory of Open Access Journals (Sweden)

    Pablo eLlop

    2015-08-01

    Full Text Available New pathogenic bacteria species belonging to the genus Erwinia associated with pome fruit trees (Erwinia pyrifoliae, E. piriflorinigrans, E. uzenensis have been increasingly described in the last years, and comparative analyses have found that all these species share several genetic characteristics. Studies at different level (whole genome comparison, virulence genes, plasmid content, etc. show a high intraspecies homogeneity (i.e. among E. amylovora strains and also abundant similarities appear between the different Erwinia species: presence of plasmids of similar size in the pathogenic species; high similarity in several genes associated with exopolysaccharide production and hence, with virulence, as well as in some other genes, in the chromosomes. Many genetic similarities have been observed also among some of the plasmids (and genomes from the pathogenic species and E. tasmaniensis or E. billingiae, two epiphytic species on the same hosts. The amount of genetic material shared in this genus varies from individual genes to clusters, genomic islands and genetic material that even may constitute a whole plasmid. Recent research on evolution of erwinias point out the horizontal transfer acquisition of some genomic islands that were subsequently lost in some species and several pathogenic traits that are still present. How this common material has been obtained and is efficiently maintained in different species belonging to the same genus sharing a common ecological niche provides an idea of the origin and evolution of the pathogenic Erwinia and the interaction with nonpathogenic species present in the same niche, and the role of the genes that are conserved in all of them.

  4. Origin and Evolution of Nitrogen Fixation Genes on Symbiosis Islands and Plasmid in Bradyrhizobium

    Science.gov (United States)

    Okubo, Takashi; Piromyou, Pongdet; Tittabutr, Panlada; Teaumroong, Neung; Minamisawa, Kiwamu

    2016-01-01

    The nitrogen fixation (nif) genes of nodule-forming Bradyrhizobium strains are generally located on symbiosis islands or symbiosis plasmids, suggesting that these genes have been transferred laterally. The nif genes of rhizobial and non-rhizobial Bradyrhizobium strains were compared in order to infer the evolutionary histories of nif genes. Based on all codon positions, the phylogenetic tree of concatenated nifD and nifK sequences showed that nifDK on symbiosis islands formed a different clade from nifDK on non-symbiotic loci (located outside of symbiosis islands and plasmids) with elongated branches; however, these genes were located in close proximity, when only the 1st and 2nd codon positions were analyzed. The guanine (G) and cytosine (C) content of the 3rd codon position of nifDK on symbiosis islands was lower than that on non-symbiotic loci. These results suggest that nif genes on symbiosis islands were derived from the non-symbiotic loci of Bradyrhizobium or closely related strains and have evolved toward a lower GC content with a higher substitution rate than the ancestral state. Meanwhile, nifDK on symbiosis plasmids clustered with nifDK on non-symbiotic loci in the tree representing all codon positions, and the GC content of symbiotic and non-symbiotic loci were similar. These results suggest that nif genes on symbiosis plasmids were derived from the non-symbiotic loci of Bradyrhizobium and have evolved with a similar evolutionary pattern and rate as the ancestral state. PMID:27431195

  5. High Prevalence of Plasmid-Mediated Quinolone Resistance and IncQ Plasmids Carrying qnrS2 Gene in Bacteria from Rivers near Hospitals and Aquaculture in China

    OpenAIRE

    Wen, Yanping; Pu, Xiaoying; Zheng, Wei; Hu, Guang

    2016-01-01

    Effluents from hospital and aquaculture are considered important sources of quinolone resistance. However, little information is available on the impact of this effluent on nearby rivers. In this study, 188 ciprofloxacin-resistant bacterial isolates obtained from rivers near hospitals and aquaculture were screened for plasmid-mediated quinolone resistance (PMQR) genes. Species identification, antibiotic susceptibility testing, and PMQR gene transferability assessment were conducted for PMQR-p...

  6. Crystal structures of the F and pSLT plasmid TraJ N-terminal regions reveal similar homodimeric PAS folds with functional interchangeability

    Energy Technology Data Exchange (ETDEWEB)

    Lu, Jun; Wu, Ruiying; Adkins, Joshua N.; Joachimiak, Andrzej; Glover, Mark

    2014-09-16

    In the F-family of conjugative plasmids, TraJ is an essential transcriptional activator of the tra operon that encodes most of the proteins required for conjugation. Here we report for the first time the X-ray crystal structures of the TraJ N-terminal regions from the prototypic F plasmid (TraJF11-130) and from the Salmonella virulence plasmid pSLT (TraJpSLT 1-128). Both proteins form similar homodimeric Per-ARNT-Sim (PAS) fold structures. Mutational analysis reveals that the observed dimeric interface is critical for TraJF transcriptional activation, indicating that dimerization of TraJ is required for its in vivo function. An artificial ligand (oxidized dithiothreitol) occupies a cavity in the TraJF dimer interface, while a smaller cavity in corresponding region of the TraJpSLT structure lacks a ligand. Gas chromatography/mass spectrometry-electron ionization analysis of dithiothreitol-free TraJF suggests indole may be the natural TraJ ligand; however, disruption of the indole biosynthetic pathway does not affect TraJF function. Heterologous PAS domains from pSLT and R100 TraJ can functionally replace the TraJF PAS domain, suggesting that TraJ allelic specificity is mediated by the region C-terminal to the PAS domain.

  7. Organometallic B12-DNA conjugate

    DEFF Research Database (Denmark)

    Hunger, Miriam; Mutti, Elena; Rieder, Alexander

    2014-01-01

    Design, synthesis, and structural characterization of a B12-octadecanucleotide are presented herein, a new organometallic B12-DNA conjugate. In such covalent conjugates, the natural B12 moiety may be a versatile vector for controlled in vivo delivery of oligonucleotides to cellular targets in hum...

  8. Characterization of tet(Y)-carrying LowGC plasmids exogenously captured from cow manure at a conventional dairy farm

    Czech Academy of Sciences Publication Activity Database

    Kyselková, Martina; Chrudimský, Tomáš; Husník, Filip; Chroňáková, Alica; Heuer, H.; Smalla, K.; Elhottová, Dana

    2016-01-01

    Roč. 92, č. 6 (2016), č. článku fiw075. ISSN 0168-6496 R&D Projects: GA ČR GAP504/10/2077; GA MŠk(CZ) EE2.3.30.0032; GA MŠk(CZ) LD13046 Institutional support: RVO:60077344 Keywords : cattle manure * LowGC plasmids * tetracycline resistance * tet(Y) * Acinetobacter * horizontal gene transfer Subject RIV: EH - Ecology, Behaviour Impact factor: 3.720, year: 2016

  9. The interconnection between biofilm formation and horizontal gene transfer

    DEFF Research Database (Denmark)

    Madsen, Jonas Stenløkke; Burmølle, Mette; Hansen, Lars H.

    2012-01-01

    Recent research has revealed that horizontal gene transfer and biofilm formation are connected processes. Although published research investigating this interconnectedness is still limited, we will review this subject in order to highlight the potential of these observations because....... Biofilms, furthermore, promote plasmid stability and may enhance the host range of mobile genetic elements that are transferred horizontally. Plasmids, on the other hand, are very well suited to promote the evolution of social traits such as biofilm formation. This, essentially, transpires because plasmids...... of their believed importance in the understanding of the adaptation and subsequent evolution of social traits in bacteria. Here, we discuss current evidence for such interconnectedness centred on plasmids. Horizontal transfer rates are typically higher in biofilm communities compared with those in planktonic states...

  10. Hydrophobic and electrostatic interactions between cell penetrating peptides and plasmid DNA are important for stable non-covalent complexation and intracellular delivery.

    Science.gov (United States)

    Upadhya, Archana; Sangave, Preeti C

    2016-10-01

    Cell penetrating peptides are useful tools for intracellular delivery of nucleic acids. Delivery of plasmid DNA, a large nucleic acid, poses a challenge for peptide mediated transport. The paper investigates and compares efficacy of five novel peptide designs for complexation of plasmid DNA and subsequent delivery into cells. The peptides were designed to contain reported DNA condensing agents and basic cell penetrating sequences, octa-arginine (R 8 ) and CHK 6 HC coupled to cell penetration accelerating peptides such as Bax inhibitory mutant peptide (KLPVM) and a peptide derived from the Kaposi fibroblast growth factor (kFGF) membrane translocating sequence. A tryptophan rich peptide, an analogue of Pep-3, flanked with CH 3 on either ends was also a part of the study. The peptides were analysed for plasmid DNA complexation, protection of peptide-plasmid DNA complexes against DNase I, serum components and competitive ligands by simple agarose gel electrophoresis techniques. Hemolysis of rat red blood corpuscles (RBCs) in the presence of the peptides was used as a measure of peptide cytotoxicity. Plasmid DNA delivery through the designed peptides was evaluated in two cell lines, human cervical cancer cell line (HeLa) and (NIH/3 T3) mouse embryonic fibroblasts via expression of the secreted alkaline phosphatase (SEAP) reporter gene. The importance of hydrophobic sequences in addition to cationic sequences in peptides for non-covalent plasmid DNA complexation and delivery has been illustrated. An alternative to the employment of fatty acid moieties for enhanced gene transfer has been proposed. Comparison of peptides for plasmid DNA complexation and delivery of peptide-plasmid DNA complexes to cells estimated by expression of a reporter gene, SEAP. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.

  11. Comparative assessment of plasmid DNA delivery by encapsulation ...

    African Journals Online (AJOL)

    Tropical Journal of Pharmaceutical Research January 2018; 17 (1): 1-10 ... Purpose: To compare the gene delivery effectiveness of plasmid DNA (pDNA) ..... Intramuscular delivery of DNA ... copolymeric system for gene delivery in complete.

  12. The technology of large-scale pharmaceutical plasmid purification ...

    African Journals Online (AJOL)

    STORAGESEVER

    2010-01-04

    Jan 4, 2010 ... DNA vaccine, the cost of purification must be decreased. Although commonly .... Three mice were killed every 4 days interval. Tissues of heart, liver, .... Now, methods such as chromatography had good prospects in plasmid ...

  13. Mechanisms of Evolution in High-Consequence Drug Resistance Plasmids

    Directory of Open Access Journals (Sweden)

    Susu He

    2016-12-01

    Full Text Available The dissemination of resistance among bacteria has been facilitated by the fact that resistance genes are usually located on a diverse and evolving set of transmissible plasmids. However, the mechanisms generating diversity and enabling adaptation within highly successful resistance plasmids have remained obscure, despite their profound clinical significance. To understand these mechanisms, we have performed a detailed analysis of the mobilome (the entire mobile genetic element content of a set of previously sequenced carbapenemase-producing Enterobacteriaceae (CPE from the National Institutes of Health Clinical Center. This analysis revealed that plasmid reorganizations occurring in the natural context of colonization of human hosts were overwhelmingly driven by genetic rearrangements carried out by replicative transposons working in concert with the process of homologous recombination. A more complete understanding of the molecular mechanisms and evolutionary forces driving rearrangements in resistance plasmids may lead to fundamentally new strategies to address the problem of antibiotic resistance.

  14. Photoinduced silver nanoparticles/nanorings on plasmid DNA scaffolds.

    Science.gov (United States)

    Liu, Jianhua; Zhang, Xiaoliang; Yu, Mei; Li, Songmei; Zhang, Jindan

    2012-01-23

    Biological scaffolds are being actively explored for the synthesis of nanomaterials with novel structures and unexpected properties. Toroidal plasmid DNA separated from the Bacillus host is applied as a sacrificial mold for the synthesis of silver nanoparticles and nanorings. The photoirradiation method is applied to reduce Ag(I) on the plasmid. The nanoparticles are obtained by varying the concentration of the Ag(I) ion solution and the exposure time of the plasmid-Ag(I) complex under UV light at 254 nm and room temperature. It is found that the plasmid serves not only as a template but also as a reductant to drive the silver nucleation and deposition. The resulting nanoparticles have a face-centered cubic (fcc) crystal structure and 20-30 nm average diameter. The detailed mechanism is discussed, and other metals or alloys could also be synthesized with this method. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Isolation and properties of plasmids from Deinococcus radiodurans Sark

    International Nuclear Information System (INIS)

    Sjarief, S.H.; Kikuchi, Masahiro; Kurita, Hiromi; Kitayama, Shigeru; Watanabe, Hiroshi.

    1990-05-01

    Radioresistant bacterium, Deinococcus radiodurans, can repair completely almost all of DNA damages including double strand breaks induced by gamma-rays up to about 5 kGy. In order to reveal the repair mechanism, it is necessary to develop a cloning vector available for the genetic analysis. We tried to isolate plasmids from D.radiodurans Sark strain. In the present paper the isolation and properties of plasmids were described. (author)

  16. Mechanisms of Evolution in High-Consequence Drug Resistance Plasmids.

    Science.gov (United States)

    He, Susu; Chandler, Michael; Varani, Alessandro M; Hickman, Alison B; Dekker, John P; Dyda, Fred

    2016-12-06

    The dissemination of resistance among bacteria has been facilitated by the fact that resistance genes are usually located on a diverse and evolving set of transmissible plasmids. However, the mechanisms generating diversity and enabling adaptation within highly successful resistance plasmids have remained obscure, despite their profound clinical significance. To understand these mechanisms, we have performed a detailed analysis of the mobilome (the entire mobile genetic element content) of a set of previously sequenced carbapenemase-producing Enterobacteriaceae (CPE) from the National Institutes of Health Clinical Center. This analysis revealed that plasmid reorganizations occurring in the natural context of colonization of human hosts were overwhelmingly driven by genetic rearrangements carried out by replicative transposons working in concert with the process of homologous recombination. A more complete understanding of the molecular mechanisms and evolutionary forces driving rearrangements in resistance plasmids may lead to fundamentally new strategies to address the problem of antibiotic resistance. The spread of antibiotic resistance among Gram-negative bacteria is a serious public health threat, as it can critically limit the types of drugs that can be used to treat infected patients. In particular, carbapenem-resistant members of the Enterobacteriaceae family are responsible for a significant and growing burden of morbidity and mortality. Here, we report on the mechanisms underlying the evolution of several plasmids carried by previously sequenced clinical Enterobacteriaceae isolates from the National Institutes of Health Clinical Center (NIH CC). Our ability to track genetic rearrangements that occurred within resistance plasmids was dependent on accurate annotation of the mobile genetic elements within the plasmids, which was greatly aided by access to long-read DNA sequencing data and knowledge of their mechanisms. Mobile genetic elements such as

  17. Conjugation in Escherichia coli

    Science.gov (United States)

    Boyer, Herbert

    1966-01-01

    Boyer, Herbert (Yale University, New Haven, Conn.). Conjugation in Escherichia coli. J. Bacteriol. 91:1767–1772. 1966.—The sex factor of Escherichia coli K-12 was introduced into an E. coli B/r strain by circumventing the host-controlled modification and restriction incompatibilities known to exist between these closely related strains. The sexual properties of the constructed F+ B strain and its Hfr derivatives were examined. These studies showed that the E. coli strain B/r F+ and Hfr derivatives are similar to the E. coli strain K-12 F+ and Hfr derivatives. However, the site of sex factor integration was found to be dependent on the host genome. PMID:5327905

  18. Electrochromic in conjugated polymers

    International Nuclear Information System (INIS)

    Picado Valenzuela, Alfredo

    2007-01-01

    This revision considered object the description of one of the materials with the greatest potential in the field of electrochromic (mainly in the visible region): the conjugated polymers (CP), area of enormous potential both now and in a short time ahead. The CP are insulating materials and organic semiconductors in a state not doped. They can be doped positively or negatively being observed a significant increase in the conductivity and being generated a color change in these materials. The understanding of how optical properties vary based on the chemical structure of the polymer or its mixtures and more precisely of the alternatives that can be entered into the conjugated system or π system to obtain a material that besides to be flexible, environmentally stable, presents the colored states. The revision was centred chiefly in the polypyrrole (Ppy), the polythiophene (PTh) and their derivatives such as poly (3.4-ethylenedioxythiophene) (PEDOT). The advantage of using monomers with variable structure, to adjust the composition of the copolymer, or to blend with the PC, allows to obtain a variety of colored states that can be modulated through the visible spectrum and even with applications to wavelengths outside of this region. Because the PC presented at least two different colored states can be varied continuously as a function of the voltage applied. In some cases, they may submit multicoloured statements, which offers a range of possibilities for their application in flexible electronic devices type screens and windows. Applications include smart windows, camouflage clothing and data screens. This type of material is emerging as one of the substitutes of the traditional inorganic semiconductor, with the advantage of its low cost, high flexibility and the possibility to generate multiple colors through the handling of the monomers in the structure and control of energy of his band gap. (author) [es

  19. The characteristics of micrococcus (deinococcus) radiodurans sark plasmids

    International Nuclear Information System (INIS)

    Sjarief, Sri Hariani; Kikuchi, Masahiro; Watanabe, Hiroshi.

    1994-01-01

    The characterization of micrococcus (deinococcus) radiodurans sark plasmids. This bacterium has been classified as a new genus deinococcus radiodurans which is resistant to gamma-rays. It can repair itself completely almost all of DNA damages including double strand breaks induced by gamma-rays up to about 5 KGy. To reveal the repair mechanism, several investigations had been done to develop a cloning vector available for the genetic analysis. For this purpose D. radiodurans Sark are to be prepared as a vector by studying the characteristics of its plasmid. Plasmids were isolated by electrophoresis using 0.6% low-melting-temperature agarose in TAE and run for 5.5 hours, followed by the identification. An antibiotic marker was also carried out in this experiment to identify its location in the genetic materials of the cell, beside making a restriction map of the plasmid. Results have shown that D. radiodurans Sark has 4 plasmids (P1, P2, P3, and P4) and the refampicin resistant genes were not found in the plasmid. (authors). 14 refs; 4 figs

  20. Multiplex PCR for detection of plasmid-mediated colistin resistance determinants, mcr-1, mcr-2, mcr-3, mcr-4 and mcr-5 for surveillance purposes

    DEFF Research Database (Denmark)

    Rebelo, Ana Rita; Bortolaia, Valeria; Kjeldgaard, Jette S.

    2018-01-01

    Background and aim: Plasmid-mediated colistin resistance mechanisms have been identified worldwide in the past years. A multiplex polymerase chain reaction (PCR) protocol for detection of all currently known transferable colistin resistance genes (mcr-1 to mcr-5, and variants) in Enterobacteriace...

  1. Construction of recombinant plasmid pIRESEgr-IFN γ and its expression in Lewis lung carcinoma induced by irradiation

    International Nuclear Information System (INIS)

    Yang Wei; Li Xiuyi; Gong Shouliang; Sun Ting; Gong Pingsheng

    2007-01-01

    Objective: To construct the recombinant plasmid pIRESEgr-IFN γ and detect its expression in Lewis lung carcinoma induced by irradiation in vitro. Methods: The recombinant plasmid pIRESEgr-IFN γ containing Egr-1 promoter and IFN γ gene was constructed with gene recombinant technique. The plasmid was transferred into Lewis lung carcinoma by liposome in vitro. The correlations of dose- and time-effects in the expression of IFN γ gene induced by X-ray were detected by ELISA. Results: The identification with enzymes proved that Egr-1 promoter and IFN γ gene were inserted into vector pIRESlneo correctly. After X-ray irradiation with different doses, the expression of IFN γ in the supernatant of Lewis lung carcinoma transfected by pIRESEgr-IFN γ was significantly higher than that in 0 Gy group (P<0.001). After 5 Gy X-ray irradiation, the expression of IFN γ was the highest, being 4.39 times as much as that in 0 Gy group. The expression of IFN γ in the supernatant increased after 5 Gy X-ray irradiation, being 6.27 times as much as that in 0 h group 36 h after irradiation. Conclusion: The recombinant plasmid pIRESEgr-IFN γ is constructed successfully, and it has the property of enhancing the expression of IFN γ gene induced by irradiation. (authors)

  2. Comparative genomic and plasmid analysis of beer-spoiling and non-beer-spoiling Lactobacillus brevis isolates.

    Science.gov (United States)

    Bergsveinson, Jordyn; Ziola, Barry

    2017-12-01

    Beer-spoilage-related lactic acid bacteria (BSR LAB) belong to multiple genera and species; however, beer-spoilage capacity is isolate-specific and partially acquired via horizontal gene transfer within the brewing environment. Thus, the extent to which genus-, species-, or environment- (i.e., brewery-) level genetic variability influences beer-spoilage phenotype is unknown. Publicly available Lactobacillus brevis genomes were analyzed via BlAst Diagnostic Gene findEr (BADGE) for BSR genes and assessed for pangenomic relationships. Also analyzed were functional coding capacities of plasmids of LAB inhabiting extreme niche environments. Considerable genetic variation was observed in L. brevis isolated from clinical samples, whereas 16 candidate genes distinguish BSR and non-BSR L. brevis genomes. These genes are related to nutrient scavenging of gluconate or pentoses, mannose, and metabolism of pectin. BSR L. brevis isolates also have higher average nucleotide identity and stronger pangenome association with one another, though isolation source (i.e., specific brewery) also appears to influence the plasmid coding capacity of BSR LAB. Finally, it is shown that niche-specific adaptation and phenotype are plasmid-encoded for both BSR and non-BSR LAB. The ultimate combination of plasmid-encoded genes dictates the ability of L. brevis to survive in the most extreme beer environment, namely, gassed (i.e., pressurized) beer.

  3. SXT/R391 Integrative and Conjugative Elements (ICEs) Encode a Novel 'Trap-Door' Strategy for Mobile Element Escape.

    Science.gov (United States)

    Ryan, Michael P; Armshaw, Patricia; Pembroke, J Tony

    2016-01-01

    Integrative conjugative elements (ICEs) are a class of bacterial mobile elements that have the ability to mediate their own integration, excision, and transfer from one host genome to another by a mechanism of site-specific recombination, self-circularisation, and conjugative transfer. Members of the SXT/R391 ICE family of enterobacterial mobile genetic elements display an unusual UV-inducible sensitization function which results in stress induced killing of bacterial cells harboring the ICE. This sensitization has been shown to be associated with a stress induced overexpression of a mobile element encoded conjugative transfer gene, orf43, a traV homolog. This results in cell lysis and release of a circular form of the ICE. Induction of this novel system may allow transfer of an ICE, enhancing its survival potential under conditions not conducive to conjugative transfer.

  4. Phytotracker, an information management system for easy recording and tracking of plants, seeds and plasmids.

    Science.gov (United States)

    Nieuwland, Jeroen; Sornay, Emily; Marchbank, Angela; de Graaf, Barend Hj; Murray, James Ah

    2012-10-13

    A large number of different plant lines are produced and maintained in a typical plant research laboratory, both as seed stocks and in active growth. These collections need careful and consistent management to track and maintain them properly, and this is a particularly pressing issue in laboratories undertaking research involving genetic manipulation due to regulatory requirements. Researchers and PIs need to access these data and collections, and therefore an easy-to-use plant-oriented laboratory information management system that implements, maintains and displays the information in a simple and visual format would be of great help in both the daily work in the lab and in ensuring regulatory compliance. Here, we introduce 'Phytotracker', a laboratory management system designed specifically to organise and track plasmids, seeds and growing plants that can be used in mixed platform environments. Phytotracker is designed with simplicity of user operation and ease of installation and management as the major factor, whilst providing tracking tools that cover the full range of activities in molecular genetics labs. It utilises the cross-platform Filemaker relational database, which allows it to be run as a stand-alone or as a server-based networked solution available across all workstations in a lab that can be internet accessible if desired. It can also be readily modified or customised further. Phytotracker provides cataloguing and search functions for plasmids, seed batches, seed stocks and plants growing in pots or trays, and allows tracking of each plant from seed sowing, through harvest to the new seed batch and can print appropriate labels at each stage. The system enters seed information as it is transferred from the previous harvest data, and allows both selfing and hybridization (crossing) to be defined and tracked. Transgenic lines can be linked to their plasmid DNA source. This ease of use and flexibility helps users to reduce their time needed to organise

  5. Phytotracker, an information management system for easy recording and tracking of plants, seeds and plasmids

    Directory of Open Access Journals (Sweden)

    Nieuwland Jeroen

    2012-10-01

    Full Text Available Abstract Background A large number of different plant lines are produced and maintained in a typical plant research laboratory, both as seed stocks and in active growth. These collections need careful and consistent management to track and maintain them properly, and this is a particularly pressing issue in laboratories undertaking research involving genetic manipulation due to regulatory requirements. Researchers and PIs need to access these data and collections, and therefore an easy-to-use plant-oriented laboratory information management system that implements, maintains and displays the information in a simple and visual format would be of great help in both the daily work in the lab and in ensuring regulatory compliance. Results Here, we introduce ‘Phytotracker’, a laboratory management system designed specifically to organise and track plasmids, seeds and growing plants that can be used in mixed platform environment