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Lopez, Jaime G., Mohamed S. Donia, and Ned S. Wingreen. "Modeling the ecology of parasitic plasmids." ISME Journal 15, no. 10 (April 8, 2021): 2843–52. http://dx.doi.org/10.1038/s41396-021-00954-6.
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AbstractPlasmids are autonomous genetic elements that can be exchanged between microorganisms via horizontal gene transfer (HGT). Despite the central role they play in antibiotic resistance and modern biotechnology, our understanding of plasmids’ natural ecology is limited. Recent experiments have shown that plasmids can spread even when they are a burden to the cell, suggesting that natural plasmids may exist as parasites. Here, we use mathematical modeling to explore the ecology of such parasitic plasmids. We first develop models of single plasmids and find that a plasmid’s population dynamics and optimal infection strategy are strongly determined by the plasmid’s HGT mechanism. We then analyze models of co-infecting plasmids and show that parasitic plasmids are prone to a “tragedy of the commons” in which runaway plasmid invasion severely reduces host fitness. We propose that this tragedy of the commons is averted by selection between competing populations and demonstrate this effect in a metapopulation model. We derive predicted distributions of unique plasmid types in genomes—comparison to the distribution of plasmids in a collection of 17,725 genomes supports a model of parasitic plasmids with positive plasmid–plasmid interactions that ameliorate plasmid fitness costs or promote the invasion of new plasmids.
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Lopez-Diaz, Maria, Nicholas Ellaby, Jane Turton, Neil Woodford, Maria Tomas, and Matthew J. Ellington. "NDM-1 carbapenemase resistance gene vehicles emergent on distinct plasmid backbones from the IncL/M family." Journal of Antimicrobial Chemotherapy 77, no. 3 (January 5, 2022): 620–24. http://dx.doi.org/10.1093/jac/dkab466.
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Abstract Objectives To assess the genetic contexts surrounding blaNDM-1 genes carried on IncM plasmids harboured by six carbapenemase-producing Enterobacterales (CPE) isolates referred to the UK Health Security Agency’s Antimicrobial Resistance and Healthcare Associated Infections (AMRHAI) Reference Unit. Methods Between 2014 and 2018, the AMRHAI Reference Unit undertook WGS of CPE isolates using Illumina NGS. Nanopore sequencing was used for selected isolates and publicly available plasmid references were downloaded. Analysis of incRNA, which encodes the antisense RNA regulating plasmidic repA gene expression, was performed and bioinformatics tools were used to analyse whole plasmid sequences. Results Of 894 NDM-positive isolates of Enterobacterales, 44 NDM-1-positive isolates of five different species (Citrobacter spp., Enterobacter cloacae, Escherichia coli, Klebsiella pneumoniae and Klebsiella oxytoca) encoded the IncRNA locus of IncM2 plasmids. Long-read sequencing of six diverse isolates revealed related IncM2, NDM-1-encoding plasmids. Plasmid ‘backbone’ areas were conserved and contrasted with highly variable resistance regions. Sub-groupings of IncM2 plasmids encoding blaNDM-1 were detected; one sub-group occurred in five different health regions of England in every year. The diversity of NDM-1-encoding resistance gene integrons and transposons and their insertions sites in the plasmids indicated that NDM-1 has been acquired repeatedly by IncM2 variants. Conclusions The use of sequencing helped inform: (i) a wide geographical distribution of isolates encoding NDM-1 on emergent IncM2 plasmids; (ii) variant plasmids have acquired NDM-1 separately; and (iii) dynamic arrangements and evolution of the resistance elements in this plasmid group. The geographical and temporal distribution of IncM2 plasmids that encode NDM-1 highlights them as a public health threat that requires ongoing monitoring.
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Li, Feifeng, Jiong Wang, Ying Jiang, Yingyi Guo, Ningjing Liu, Shunian Xiao, Likang Yao, et al. "Adaptive Evolution Compensated for the Plasmid Fitness Costs Brought by Specific Genetic Conflicts." Pathogens 12, no. 1 (January 13, 2023): 137. http://dx.doi.org/10.3390/pathogens12010137.
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New Delhi metallo-β-lactamase (NDM)-carrying IncX3 plasmids is important in the transmission of carbapenem resistance in Escherichia coli. Fitness costs related to plasmid carriage are expected to limit gene exchange; however, the causes of these fitness costs are poorly understood. Compensatory mutations are believed to ameliorate plasmid fitness costs and enable the plasmid’s wide spread, suggesting that such costs are caused by specific plasmid–host genetic conflicts. By combining conjugation tests and experimental evolution with comparative genetic analysis, we showed here that the fitness costs related to ndm/IncX3 plasmids in E. coli C600 are caused by co-mutations of multiple host chromosomal genes related to sugar metabolism and cell membrane function. Adaptive evolution revealed that mutations in genes associated with oxidative stress, nucleotide and short-chain fatty acid metabolism, and cell membranes ameliorated the costs associated with plasmid carriage. Specific genetic conflicts associated with the ndm/IncX3 plasmid in E. coli C600 involve metabolism and cell-membrane-related genes, which could be ameliorated by compensatory mutations. Collectively, our findings could explain the wide spread of IncX3 plasmids in bacterial genomes, despite their potential cost.
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Bahl, Martin Iain, Lars Hestbjerg Hansen, Tine Rask Licht, and Søren J. Sørensen. "Conjugative Transfer Facilitates Stable Maintenance of IncP-1 Plasmid pKJK5 in Escherichia coli Cells Colonizing the Gastrointestinal Tract of the Germfree Rat." Applied and Environmental Microbiology 73, no. 1 (November 3, 2006): 341–43. http://dx.doi.org/10.1128/aem.01971-06.
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ABSTRACT Quantitative determination of IncP-1 plasmid loss from Escherichia coli cells colonizing the gastrointestinal tracts of germfree rats was achieved by flow cytometry. Results show that the plasmid's ability to conjugate counteracts plasmid loss and is thus an important mechanism for the stable maintenance of IncP-1 plasmids within the gastrointestinal environment.
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Longtine, M. S., S. Enomoto, S. L. Finstad, and J. Berman. "Yeast telomere repeat sequence (TRS) improves circular plasmid segregation, and TRS plasmid segregation involves the RAP1 gene product." Molecular and Cellular Biology 12, no. 5 (May 1992): 1997–2009. http://dx.doi.org/10.1128/mcb.12.5.1997-2009.1992.
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Telomere repeat sequences (TRSs) can dramatically improve the segregation of unstable circular autonomously replicating sequence (ARS) plasmids in Saccharomyces cerevisiae. Deletion analysis demonstrated that yeast TRSs, which conform to the general sequence (C(1-3)A)n, are able to stabilize circular ARS plasmids. A number of TRS clones of different primary sequence and C(1-3)A tract length confer the plasmid stabilization phenotype. TRS sequences do not appear to improve plasmid replication efficiency, as determined by plasmid copy number analysis and functional assays for ARS activity. Pedigree analysis confirms that TRS-containing plasmids are missegregated at low frequency and that missegregated TRS-containing plasmids, like ARS plasmids, are preferentially retained by the mother cell. Plasmids stabilized by TRSs have properties that distinguish them from centromere-containing plasmids and 2 microns-based recombinant plasmids. Linear ARS plasmids, which include two TRS tracts at their termini, segregate inefficiently, while circular plasmids with one or two TRS tracts segregate efficiently, suggesting that plasmid topology or TRS accessibility interferes with TRS segregation function on linear plasmids. In strains carrying the temperature-sensitive mutant alleles rap1grc4 and rap1-5, TRS plasmids are not stable at the semipermissive temperature, suggesting that RAP1 protein is involved in TRS plasmid stability. In Schizosaccharomyces pombe, an ARS plasmid was stabilized by the addition of S. pombe telomere sequence, suggesting that the ability to improve the segregation of ARS plasmids is a general property of telomere repeats.
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Longtine, M. S., S. Enomoto, S. L. Finstad, and J. Berman. "Yeast telomere repeat sequence (TRS) improves circular plasmid segregation, and TRS plasmid segregation involves the RAP1 gene product." Molecular and Cellular Biology 12, no. 5 (May 1992): 1997–2009. http://dx.doi.org/10.1128/mcb.12.5.1997.
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Telomere repeat sequences (TRSs) can dramatically improve the segregation of unstable circular autonomously replicating sequence (ARS) plasmids in Saccharomyces cerevisiae. Deletion analysis demonstrated that yeast TRSs, which conform to the general sequence (C(1-3)A)n, are able to stabilize circular ARS plasmids. A number of TRS clones of different primary sequence and C(1-3)A tract length confer the plasmid stabilization phenotype. TRS sequences do not appear to improve plasmid replication efficiency, as determined by plasmid copy number analysis and functional assays for ARS activity. Pedigree analysis confirms that TRS-containing plasmids are missegregated at low frequency and that missegregated TRS-containing plasmids, like ARS plasmids, are preferentially retained by the mother cell. Plasmids stabilized by TRSs have properties that distinguish them from centromere-containing plasmids and 2 microns-based recombinant plasmids. Linear ARS plasmids, which include two TRS tracts at their termini, segregate inefficiently, while circular plasmids with one or two TRS tracts segregate efficiently, suggesting that plasmid topology or TRS accessibility interferes with TRS segregation function on linear plasmids. In strains carrying the temperature-sensitive mutant alleles rap1grc4 and rap1-5, TRS plasmids are not stable at the semipermissive temperature, suggesting that RAP1 protein is involved in TRS plasmid stability. In Schizosaccharomyces pombe, an ARS plasmid was stabilized by the addition of S. pombe telomere sequence, suggesting that the ability to improve the segregation of ARS plasmids is a general property of telomere repeats.
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Pollet, Rebecca M., James D. Ingle, Jeff P. Hymes, Thomas C. Eakes, Karina Yui Eto, Stephen M. Kwong, Joshua P. Ramsay, Neville Firth, and Matthew R. Redinbo. "Processing of Nonconjugative Resistance Plasmids by Conjugation Nicking Enzyme of Staphylococci." Journal of Bacteriology 198, no. 6 (January 4, 2016): 888–97. http://dx.doi.org/10.1128/jb.00832-15.
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ABSTRACTAntimicrobial 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 mechanism of antimicrobial resistance transfer in bacteria such asStaphylococcus aureusis an important step toward potentially slowing the spread of antimicrobial-resistant infections. This work establishes protein-DNA interactions essential for the transfer of theStaphylococcus aureusmultiresistance plasmid pSK41 by its relaxase, NES. This enzyme also processed variantoriT-like sequences found on numerous plasmids previously considered nontransmissible, suggesting that in conjunction with an uncharacterized accessory protein, these plasmids may be transferred horizontally via a relaxase intransmechanism. These findings have important implications for our understanding of staphylococcal resistance plasmid evolution.
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Brown, Celeste J., Diya Sen, Hirokazu Yano, Matthew L. Bauer, Linda M. Rogers, Geraldine A. Van der Auwera, and Eva M. Top. "Diverse Broad-Host-Range Plasmids from Freshwater Carry Few Accessory Genes." Applied and Environmental Microbiology 79, no. 24 (October 4, 2013): 7684–95. http://dx.doi.org/10.1128/aem.02252-13.
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ABSTRACTBroad-host-range self-transferable plasmids are known to facilitate bacterial adaptation by spreading genes between phylogenetically distinct hosts. These plasmids typically have a conserved backbone region and a variable accessory region that encodes host-beneficial traits. We do not know, however, how well plasmids that do not encode accessory functions can survive in nature. The goal of this study was to characterize the backbone and accessory gene content of plasmids that were captured from freshwater sources without selecting for a particular phenotype or cultivating their host. To do this, triparental matings were used such that the only required phenotype was the plasmid's ability to mobilize a nonconjugative plasmid. Based on complete genome sequences of 10 plasmids, only 5 carried identifiable accessory gene regions, and none carried antibiotic resistance genes. The plasmids belong to four known incompatibility groups (IncN, IncP-1, IncU, and IncW) and two potentially new groups. Eight of the plasmids were shown to have a broad host range, being able to transfer into alpha-, beta-, and gammaproteobacteria. Because of the absence of antibiotic resistance genes, we resampled one of the sites and compared the proportion of captured plasmids that conferred antibiotic resistance to their hosts with the proportion of such plasmids captured from the effluent of a local wastewater treatment plant. Few of the captured plasmids from either site encoded antibiotic resistance. A high diversity of plasmids that encode no or unknown accessory functions is thus readily found in freshwater habitats. The question remains how the plasmids persist in these microbial communities.
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Wu, Shang Wei, Kathrine Dornbusch, Göran Kronvall, and Mari Norgren. "Characterization and Nucleotide Sequence of a Klebsiella oxytoca Cryptic Plasmid Encoding a CMY-Type β-Lactamase: Confirmation that the Plasmid-Mediated Cephamycinase Originated from the Citrobacter freundii AmpC β-Lactamase." Antimicrobial Agents and Chemotherapy 43, no. 6 (June 1, 1999): 1350–57. http://dx.doi.org/10.1128/aac.43.6.1350.
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ABSTRACT Plasmid pTKH11, originally obtained by electroporation of aKlebsiella oxytoca plasmid preparation intoEscherichia coli XAC, expressed a high level of an AmpC-like β-lactamase. The enzyme, designated CMY-5, conferred resistance to extended-spectrum β-lactams in E. coli; nevertheless, the phenotype was cryptic in the K. oxytocadonor. Determination of the complete nucleotide sequence of pTKH11 revealed that the 8,193-bp plasmid encoded seven open reading frames, including that for the CMY-5 β-lactamase (bla CMY-5). Thebla CMY-5 product was similar to the plasmidic CMY-2 β-lactamase of K. pneumoniae and the chromosomal AmpC of Citrobacter freundii, with 99.7 and 97.0% identities, respectively; there was a substitution of phenylalanine in CMY-5 for isoleucine 105 in CMY-2. bla CMY-5 was followed by the Blc and SugE genes of C. freundii, and this cluster exhibited a genetic organization identical to that of the ampC region on the chromosome ofC. freundii; these results confirmed that C. freundii AmpC was the evolutionary origin of the plasmidic cephamycinases. In the K. oxytoca host, the copy number of pTKH11 was very low and the plasmid coexisted with plasmid pNBL63. Analysis of the replication regions of the two plasmids revealed 97% sequence similarity in the RNA I transcripts; this result implied that the two plasmids might be incompatible. Incompatibility of the two plasmids might explain the cryptic phenotype ofbla CMY-5 in K. oxytoca through an exclusion effect on pTKH11 by resident plasmid pNBL63.
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Paganini, Julian A., Nienke L. Plantinga, Sergio Arredondo-Alonso, Rob J. L. Willems, and Anita C. Schürch. "Recovering Escherichia coli Plasmids in the Absence of Long-Read Sequencing Data." Microorganisms 9, no. 8 (July 28, 2021): 1613. http://dx.doi.org/10.3390/microorganisms9081613.
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The incidence of infections caused by multidrug-resistant E. coli strains has risen in the past years. Antibiotic resistance in E. coli is often mediated by acquisition and maintenance of plasmids. The study of E. coli plasmid epidemiology and genomics often requires long-read sequencing information, but recently a number of tools that allow plasmid prediction from short-read data have been developed. Here, we reviewed 25 available plasmid prediction tools and categorized them into binary plasmid/chromosome classification tools and plasmid reconstruction tools. We benchmarked six tools (MOB-suite, plasmidSPAdes, gplas, FishingForPlasmids, HyAsP and SCAPP) that aim to reliably reconstruct distinct plasmids, with a special focus on plasmids carrying antibiotic resistance genes (ARGs) such as extended-spectrum beta-lactamase genes. We found that two thirds (n = 425, 66.3%) of all plasmids were correctly reconstructed by at least one of the six tools, with a range of 92 (14.58%) to 317 (50.23%) correctly predicted plasmids. However, the majority of plasmids that carried antibiotic resistance genes (n = 85, 57.8%) could not be completely recovered as distinct plasmids by any of the tools. MOB-suite was the only tool that was able to correctly reconstruct the majority of plasmids (n = 317, 50.23%), and performed best at reconstructing large plasmids (n = 166, 46.37%) and ARG-plasmids (n = 41, 27.9%), but predictions frequently contained chromosome contamination (40%). In contrast, plasmidSPAdes reconstructed the highest fraction of plasmids smaller than 18 kbp (n = 168, 61.54%). Large ARG-plasmids, however, were frequently merged with sequences derived from distinct replicons. Available bioinformatic tools can provide valuable insight into E. coli plasmids, but also have important limitations. This work will serve as a guideline for selecting the most appropriate plasmid reconstruction tool for studies focusing on E. coli plasmids in the absence of long-read sequencing data.
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Carroll, Amanda C., and Alex Wong. "Plasmid persistence: costs, benefits, and the plasmid paradox." Canadian Journal of Microbiology 64, no. 5 (May 2018): 293–304. http://dx.doi.org/10.1139/cjm-2017-0609.
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Plasmids are extrachromosomal DNA elements that can be found throughout bacteria, as well as in other domains of life. Nonetheless, the evolutionary processes underlying the persistence of plasmids are incompletely understood. Bacterial plasmids may encode genes for traits that are sometimes beneficial to their hosts, such as antimicrobial resistance, virulence, heavy metal tolerance, and the catabolism of unique nutrient sources. In the absence of selection for these traits, however, plasmids generally impose a fitness cost on their hosts. As such, plasmid persistence presents a conundrum: models predict that costly plasmids will be lost over time or that beneficial plasmid genes will be integrated into the host genome. However, laboratory and comparative studies have shown that plasmids can persist for long periods, even in the absence of positive selection. Several hypotheses have been proposed to explain plasmid persistence, including host-plasmid co-adaptation, plasmid hitchhiking, cross-ecotype transfer, and high plasmid transfer rates, but there is no clear evidence that any one model adequately resolves the plasmid paradox.
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Smith, Hilde, Alex Bossers, Frank Harders, Guanghui Wu, Neil Woodford, Stefan Schwarz, Beatriz Guerra, et al. "Characterization of Epidemic IncI1-Iγ Plasmids Harboring Ambler Class A and C Genes in Escherichia coli and Salmonella enterica from Animals and Humans." Antimicrobial Agents and Chemotherapy 59, no. 9 (June 22, 2015): 5357–65. http://dx.doi.org/10.1128/aac.05006-14.
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ABSTRACTThe aim of the study was to identify the plasmid-encoded factors contributing to the emergence and spread of epidemic IncI1-Iγ plasmids obtained fromEscherichia coliandSalmonella entericaisolates from animal and human reservoirs. For this, 251 IncI1-Iγ plasmids carrying various extended-spectrum β-lactamase (ESBL) or AmpC β-lactamase genes were compared using plasmid multilocus sequence typing (pMLST). Thirty-two of these plasmids belonging to different pMLST types were sequenced using Roche 454 and Illumina platforms. Epidemic IncI1-Iγ plasmids could be assigned to various dominant clades, whereas rarely detected plasmids clustered together as a distinct clade. Similar phylogenetic trees were obtained using only the plasmid backbone sequences, showing that the differences observed between the plasmids belonging to distinct clades resulted mainly from differences between their backbone sequences. Plasmids belonging to the various clades differed particularly in the presence/absence of genes encoding partitioning and addiction systems, which contribute to stable inheritance during cell division and plasmid maintenance. Despite this, plasmids belonging to the various phylogenetic clades also showed marked resistance gene associations, indicating the circulation of successful plasmid-gene combinations. The variation intraYandexcAgenes found in IncI1-Iγ plasmids is conserved within pMLST sequence types and plays a role in incompatibility, although functional study is needed to elucidate the role of these genes in plasmid epidemiology.
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Rebelo, João S., Célia P. F. Domingues, and Francisco Dionisio. "Plasmid Costs Explain Plasmid Maintenance, Irrespective of the Nature of Compensatory Mutations." Antibiotics 12, no. 5 (May 1, 2023): 841. http://dx.doi.org/10.3390/antibiotics12050841.
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Conjugative plasmids often carry virulence and antibiotic-resistant genes. Therefore, understanding the behavior of these extra-chromosomal DNA elements gives insights into their spread. Bacteria frequently replicate slower after plasmids’ entry, an observation inconsistent with the plasmids’ ubiquity in nature. Several hypotheses explain the maintenance of plasmids among bacterial communities. However, the numerous combinations of bacterial species and strains, plasmids, and environments claim a robust elucidatory mechanism of plasmid maintenance. Previous works have shown that donor cells already adapted to the plasmid may use the plasmid as a ‘weapon’ to compete with non-adapted plasmid-free cells. Computer simulations corroborated this hypothesis with a wide range of parameters. Here we show that donor cells benefit from harboring conjugative plasmids even if compensatory mutations in transconjugant cells occur in the plasmid, not on chromosomes. The advantage’s leading causes are as follows: mutations take time to appear, many plasmids remain costly, and re-transfer of mutated plasmids usually occurs in sites distant to the original donors, implying little competition between these cells. Research in previous decades cautioned against uncritical acceptance of the hypothesis that resistance cost helps to preserve antibiotics’ effectiveness. This work gives a new twist to this conclusion by showing that costs help antibiotic-resistant bacteria to compete with plasmid-free cells even if compensatory mutations appear in plasmids.
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Dimitriu, Tatiana, Andrew C. Matthews, and Angus Buckling. "Increased copy number couples the evolution of plasmid horizontal transmission and plasmid-encoded antibiotic resistance." Proceedings of the National Academy of Sciences 118, no. 31 (July 29, 2021): e2107818118. http://dx.doi.org/10.1073/pnas.2107818118.
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Conjugative plasmids are mobile elements that spread horizontally between bacterial hosts and often confer adaptive phenotypes, including antimicrobial resistance (AMR). Theory suggests that opportunities for horizontal transmission favor plasmids with higher transfer rates, whereas selection for plasmid carriage favors less-mobile plasmids. However, little is known about the mechanisms leading to variation in transmission rates in natural plasmids or the resultant effects on their bacterial host. We investigated the evolution of AMR plasmids confronted with different immigration rates of susceptible hosts. Plasmid RP4 did not evolve in response to the manipulations, but plasmid R1 rapidly evolved up to 1,000-fold increased transfer rates in the presence of susceptible hosts. Most evolved plasmids also conferred on their hosts the ability to grow at high concentrations of antibiotics. This was because plasmids evolved greater copy numbers as a function of mutations in the copA gene controlling plasmid replication, causing both higher transfer rates and AMR. Reciprocally, plasmids with increased conjugation rates also evolved when selecting for high levels of AMR, despite the absence of susceptible hosts. Such correlated selection between plasmid transfer and AMR could increase the spread of AMR within populations and communities.
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McMillan, Elizabeth A., Ly-Huong T. Nguyen, Lari M. Hiott, Poonam Sharma, Charlene R. Jackson, Jonathan G. Frye, and Chin-Yi Chen. "Genomic Comparison of Conjugative Plasmids from Salmonella enterica and Escherichia coli Encoding Beta-Lactamases and Capable of Mobilizing Kanamycin Resistance Col-like Plasmids." Microorganisms 9, no. 11 (October 23, 2021): 2205. http://dx.doi.org/10.3390/microorganisms9112205.
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Salmonella enterica and Escherichia coli are important human pathogens that frequently contain plasmids, both large and small, carrying antibiotic resistance genes. Large conjugative plasmids are known to mobilize small Col plasmids, but less is known about the specificity of mobilization. In the current study, six S. enterica and four E. coli strains containing large plasmids were tested for their ability to mobilize three different kanamycin resistance Col plasmids (KanR plasmids). Large conjugative plasmids from five isolates, four S. enterica and one E. coli, were able to mobilize KanR plasmids of various types. Plasmids capable of mobilizing the KanR plasmids were either IncI1 or IncX, while IncI1 and IncX plasmids with no evidence of conjugation had disrupted transfer regions. Conjugative plasmids of similar types mobilized similar KanR plasmids, but not all conjugative plasmid types were capable of mobilizing all of the KanR plasmids. These data describe some of the complexities and specificities of individual small plasmid mobilization.
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Pedersen, K., T. Tiainen, and J. L. Larsen. "Plasmid profiles, restriction fragment length polymorphisms and O-serotypes amongVibrio anguillarumisolates." Epidemiology and Infection 117, no. 3 (December 1996): 471–78. http://dx.doi.org/10.1017/s0950268800059136.
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SummaryA total of 279Vibrio anguillarumstrains were serotyped and examined for plasmid content. Plasmids were subjected to digestion with restriction enzymes. Most strains belonged to serogroup O1 (39%) and O2 (16%). In total 164 strains (53%) carried plasmids. Of the O1 and O2 isolates, 92% and 30%, respectively, carried one or more plasmids. Restriction fragment length polymorphism (RFLP) analysis of plasmid DNA indicated that plasmids belonged to several groups. Each group seemed to be restricted to a single O-serovar. The largest group was the pJM1-like plasmids among most serovar O1 strains. Most of these plasmids were about 67 kb like the pJM1 plasmid, but various derivatives ranged from 26–77 kb. RFLP studies of the 67 kb plasmids revealed 17 different restriction patterns. Some patterns were dominant among European strains whereas others were dominant among North American strains. The results confirmed the applicability of O-serotyping together with plasmid profile and restriction analysis of plasmids for typing ofV. anguillarum. They also indicated that plasmids among strains which belonged to the traditional fish pathogenic serogroups, O1 and O2, showed more homology than did strains from most other serogroups, that were usually non-pathogenic, environmental bacteria.
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Rasko, David A., M. J. Rosovitz, Ole Andreas Økstad, Derrick E. Fouts, Lingxia Jiang, Regina Z. Cer, Anne-Brit Kolstø, Steven R. Gill, and Jacques Ravel. "Complete Sequence Analysis of Novel Plasmids from Emetic and Periodontal Bacillus cereus Isolates Reveals a Common Evolutionary History among the B. cereus-Group Plasmids, Including Bacillus anthracis pXO1." Journal of Bacteriology 189, no. 1 (October 13, 2006): 52–64. http://dx.doi.org/10.1128/jb.01313-06.
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ABSTRACTThe plasmids of the members of theBacillus cereussensu lato group of organisms are essential in defining the phenotypic traits associated with pathogenesis and ecology. For example,Bacillus anthraciscontains two plasmids, pXO1 and pXO2, encoding toxin production and encapsulation, respectively, that define this species pathogenic potential, whereas the presence of a Bt toxin-encoding plasmid definesBacillus thuringiensisisolates. In this study the plasmids fromB. cereusisolates that produce emetic toxin or are linked to periodontal disease were sequenced and analyzed. Two periodontal isolates examined contained almost identical ∼272-kb plasmids, named pPER272. The emetic toxin-producing isolate contained one ∼270-kb plasmid, named pCER270, encoding the cereulide biosynthesis gene cluster. Comparative sequence analyses of theseB. cereusplasmids revealed a high degree of sequence similarity to theB. anthracispXO1 plasmid, especially in a putative replication region. These plasmids form a newly defined group of pXO1-like plasmids. However, these novel plasmids do not contain the pXO1 pathogenicity island, which in each instance is replaced by plasmid specific DNA. Plasmids pCER270 and pPER272 share regions that are not found in any other pXO1-like plasmids. Evolutionary studies suggest that these plasmids are more closely related to each other than to other identifiedB. cereusplasmids. Screening of a population ofB. cereusgroup isolates revealed that pXO1-like plasmids are more often found in association with clinical isolates. This study demonstrates that the pXO1-like plasmids may define pathogenicB. cereusisolates in the same way that pXO1 and pXO2 define theB. anthracisspecies.
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Sengupta, Manjistha, and Stuart Austin. "Prevalence and Significance of Plasmid Maintenance Functions in the Virulence Plasmids of Pathogenic Bacteria." Infection and Immunity 79, no. 7 (May 9, 2011): 2502–9. http://dx.doi.org/10.1128/iai.00127-11.
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ABSTRACTVirulence functions of pathogenic bacteria are often encoded on large extrachromosomal plasmids. These plasmids are maintained at low copy number to reduce the metabolic burden on their host. Low-copy-number plasmids risk loss during cell division. This is countered by plasmid-encoded systems that ensure that each cell receives at least one plasmid copy. Plasmid replication and recombination can produce plasmid multimers that hinder plasmid segregation. These are removed by multimer resolution systems. Equitable distribution of the resulting monomers to daughter cells is ensured by plasmid partition systems that actively segregate plasmid copies to daughter cells in a process akin to mitosis in higher organisms. Any plasmid-free cells that still arise due to occasional failures of replication, multimer resolution, or partition are eliminated by plasmid-encoded postsegregational killing systems. Here we argue that all of these three systems are essential for the stable maintenance of large low-copy-number plasmids. Thus, they should be found on all large virulence plasmids. Where available, well-annotated sequences of virulence plasmids confirm this. Indeed, virulence plasmids often appear to contain more than one example conforming to each of the three system classes. Since these systems are essential for virulence, they can be regarded as ubiquitous virulence factors. As such, they should be informative in the search for new antibacterial agents and drug targets.
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van Passel, Mark W. J., Arie van der Ende, and Aldert Bart. "Plasmid Diversity in Neisseriae." Infection and Immunity 74, no. 8 (August 2006): 4892–99. http://dx.doi.org/10.1128/iai.02087-05.
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ABSTRACT Horizontal gene transfer constitutes an important force in prokaryotic genome evolution, and it is well-known that plasmids are vehicles for DNA transfer. Chromosomal DNA is frequently exchanged between pathogenic and commensal neisseriae, but relatively little is known about plasmid diversity and prevalence among these nasopharyngeal inhabitants. We investigated the plasmid contents of 18 Neisseria lactamica isolates and 20 nasopharyngeal Neisseria meningitidis isolates. Of 18 N. lactamica strains, 9 harbored one or more plasmids, whereas only one N. meningitidis isolate contained a plasmid. Twelve plasmids were completely sequenced, while five plasmid sequences from the public databases were also included in the analyses. On the basis of nucleic acid sequences, mobilization, and replicase protein alignments, we distinguish six different plasmid groups (I to VI). Three plasmids from N. lactamica appeared to be highly similar on the nucleotide level to the meningococcal plasmids pJS-A (>99%) and pJS-B (>75%). The genetic organizations of two plasmids show a striking resemblance with that of the recently identified meningococcal disease-associated (MDA) phage, while four putative proteins encoded by these plasmids show 25% to 39% protein identity to those encoded by the MDA phage. The putative promoter of the gene encoding the replicase on these plasmids contains a polycytidine tract, suggesting that replication is subjected to phase variation. In conclusion, extensive plasmid diversity is encountered among commensal neisseriae. Members of three plasmid groups are found in both pathogenic and commensal neisseriae, indicating plasmid exchange between these species. Resemblance between plasmids and MDA phage may be indicative of dissemination of phage-related sequences among pathogenic and commensal neisseriae.
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Berg, Tracey, Neville Firth, Sumalee Apisiridej, Anusha Hettiaratchi, Amornrut Leelaporn, and Ronald A. Skurray. "Complete Nucleotide Sequence of pSK41: Evolution of Staphylococcal Conjugative Multiresistance Plasmids." Journal of Bacteriology 180, no. 17 (September 1, 1998): 4350–59. http://dx.doi.org/10.1128/jb.180.17.4350-4359.1998.
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ABSTRACT The 46.4-kb nucleotide sequence of pSK41, a prototypical multiresistance plasmid from Staphylococcus aureus, has been determined, representing the first completely sequenced conjugative plasmid from a gram-positive organism. Analysis of the sequence has enabled the identification of the probable replication, maintenance, and transfer functions of the plasmid and has provided insights into the evolution of a clinically significant group of plasmids. The basis of deletions commonly associated with pSK41 family plasmids has been investigated, as has the observed insertion site specificity of Tn552-like β-lactamase transposons within them. Several of the resistance determinants carried by pSK41-like plasmids were found to be located on up to four smaller cointegrated plasmids. pSK41 and related plasmids appear to represent a consolidation of antimicrobial resistance functions, collected by a preexisting conjugative plasmid via transposon insertion and IS257-mediated cointegrative capture of other plasmids.
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Ogbolu, David Olusoga, Oluwole Adebayo Daini, Afolabi Ogunledun, Oyebode Armstrong Terry Alli, and Mark Alexander Webber. "Dissemination of IncF plasmids carrying beta-lactamase genes in Gram-negative bacteria from Nigerian hospitals." Journal of Infection in Developing Countries 7, no. 05 (May 13, 2013): 382–90. http://dx.doi.org/10.3855/jidc.2613.
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Introduction: Production of beta-lactamases is the predominant cause of resistance to beta-lactam antibiotics in Gram-negative bacteria. We investigated the diversity of plasmid-borne beta-lactamase genes and replicon type of the plasmids carrying the respective genes in Gram-negative bacteria recovered from clinical infection in Nigerian hospitals. Methodology: A total of 134 Gram-negative bacteria of 13 species were analyzed for antimicrobial susceptibility, phenotypic and genotypic detection of various beta-lactamases, and plasmid analysis, including replicon typing. Results: Of the 134 isolates, 111 (82.8%) contained beta-lactamases, while 28 (20.9%) carried extended-spectrum beta-lactamases. PCR and sequencing identified TEM-1 in 109 isolates (81.3%), SHV-1 in 33 isolates (24.6%), OXA-1 in 15 isolates (11.2%) and CTX-M enzymes (24 CTX-M-15 and 1 CTX-M-3) in 25 isolates (18.7%). Multiplex PCR showed that 6 isolates carried plasmidic AmpCs (ACT-1, DHA-1 and CMY-2); these enzymes were detected only in isolates possessing CTX-M beta-lactamases. Of 13 (76.9%) representative plasmids investigated in detail, 9 (69.2%) were self-transferable when selected by a beta-lactam and the plasmids once transferred coded for beta-lactam resistance. Replicon typing indicated IncF as the common vector encoding for beta-lactamases. Conclusions: The study showed a diversity of beta-lactamase genes disseminated by conjugative IncF plasmids in Gram-negative bacteria; TEM-1, SHV-1, OXA-1, CTX-M-15, CTX-M-3and plasmidic AmpC enzymes are in common circulation in Nigeria.
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Chen, Liang, Kalyan D. Chavda, Roberto G. Melano, Tao Hong, Albert D. Rojtman, Michael R. Jacobs, Robert A. Bonomo, and Barry N. Kreiswirth. "Molecular Survey of the Dissemination of TwoblaKPC-Harboring IncFIA Plasmids in New Jersey and New York Hospitals." Antimicrobial Agents and Chemotherapy 58, no. 4 (February 3, 2014): 2289–94. http://dx.doi.org/10.1128/aac.02749-13.
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ABSTRACTKlebsiella pneumoniaecarbapenemase (KPC)-producingK. pneumoniaestrains have spread worldwide and become a major threat in health care facilities. Transmission ofblaKPC, the plasmid-borne KPC gene, can be mediated by clonal spread and horizontal transfer. Here, we report the complete nucleotide sequences of two novelblaKPC-3-harboring IncFIA plasmids, pBK30661 and pBK30683. pBK30661 is 74 kb in length, with a mosaic plasmid structure; it exhibits homologies to several other plasmids but lacks the plasmid transfer operon (tra) and the origin of transfer (oriT) that are required for plasmid transfer. pBK30683 is a conjugative plasmid with a cointegrated plasmid structure, comprising a 72-kb element that highly resembles pBK30661 (>99.9% nucleotide identities) and an extra 68-kb element that harborstraandoriT. A PCR scheme was designed to detect the distribution ofblaKPC-harboring IncFIA (pBK30661-like and pBK30683-like) plasmids in a collection of clinicalEnterobacteriaceaeisolates from 10 hospitals in New Jersey and New York. KPC-harboring IncFIA plasmids were found in 20% of 491K. pneumoniaeisolates, and all carriedblaKPC-3. pBK30661-like plasmids were identified mainly in the epidemic sequence type 258 (ST258)K. pneumoniaeclone, while pBK30683-like plasmids were widely distributed in ST258 and otherK. pneumoniaesequence types and among non-K. pneumoniae Enterobacteriaceaespecies. This suggests that both clonal spread and horizontal plasmid transfer contributed to the dissemination ofblaKPC-harboring IncFIA plasmids in our area. Further studies are needed to understand the distribution of this plasmid group in other health care regions and to decipher the origins of pBK30661-like and pBK30683-like plasmids.
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Wang, Pengxia, Dongmei He, Baiyuan Li, Yunxue Guo, Weiquan Wang, Xiongjian Luo, Xuanyu Zhao, and Xiaoxue Wang. "Eliminating mcr-1-harbouring plasmids in clinical isolates using the CRISPR/Cas9 system." Journal of Antimicrobial Chemotherapy 74, no. 9 (June 15, 2019): 2559–65. http://dx.doi.org/10.1093/jac/dkz246.
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Abstract Objectives To eliminate mcr-1-harbouring plasmids and MDR plasmids in clinical Escherichia coli isolates. Methods Plasmid pMBLcas9 expressing Cas9 was constructed and used to clone target single-guide RNAs (sgRNAs) for plasmid curing. The recombinant plasmid pMBLcas9-sgRNA was transferred by conjugation into two clinical E. coli isolates. The curing efficiency of different sgRNAs targeting conserved genes was tested. The elimination of targeted plasmids and the generation of transposase-mediated recombination of p14EC033a variants were characterized by PCR and DNA sequencing. Results In this study, four native plasmids in isolate 14EC033 and two native plasmids in isolate 14EC007 were successfully eliminated in a step-by-step manner using pMBLcas9. Moreover, two native plasmids in 14EC007 were simultaneously eliminated by tandemly cloning multiple sgRNAs in pMBLcas9, sensitizing 14EC007 to polymyxin and carbenicillin. In 14EC033 with two mcr-1-harbouring plasmids, IncI2 plasmid p14EC033a and IncX4 plasmid p14EC033b, a single mcr-1 sgRNA mediated the loss of p14EC033b and generated a mutant p14EC033a in which the mcr-1 gene was deleted. An insertion element, IS5, located upstream of mcr-1 in p14EC033a was responsible for transposase-mediated recombination, resulting in mcr-1 gene deletion instead of plasmid curing. Conclusions CRISPR/Cas9 can be used to efficiently sensitize clinical isolates to antibiotics in vitro. For isolates with multiple plasmids, the CRISPR/Cas9 approach can either remove each plasmid in a stepwise manner or simultaneously remove multiple plasmids in one step. Moreover, this approach can be used to delete multiple gene copies by using only one sgRNA. However, caution must be exercised to avoid unwanted recombination events during genetic manipulation.
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Guillard, Thomas, Emmanuelle Cambau, Catherine Neuwirth, Thomas Nenninger, Aurore Mbadi, Lucien Brasme, Véronique Vernet-Garnier, Odile Bajolet, and Christophe de Champs. "Description of a 2,683-Base-Pair Plasmid ContainingqnrDin Two Providencia rettgeri Isolates." Antimicrobial Agents and Chemotherapy 56, no. 1 (October 10, 2011): 565–68. http://dx.doi.org/10.1128/aac.00081-11.
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ABSTRACTqnrgenes are plasmid-mediated quinolone resistance genes mainly harbored on large conjugative multiresistant plasmids. TheqnrDgene was recently observed inSalmonella entericaon a small nonconjugative plasmid (p2007057). We describe two strains ofProvidencia rettgeriharboringqnrDon nonconjugative plasmids. The plasmids were 99% identical, with 2,683 bp and four open reading frames, includingqnrD, but exhibited only 53% identity with the plasmid found inS. enterica.
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Rosenshine, Ilan, and Moshe Mevarech. "Isolation and partial characterization of plasmids found in three Halobacterium volcanii isolates." Canadian Journal of Microbiology 35, no. 1 (January 1, 1989): 92–95. http://dx.doi.org/10.1139/m89-014.
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Three new isolates of Halobacterium volcanii were screened for the presence of plasmids. Each of the different isolates was found to contain one plasmid. These plasmids do not show any homology to each other, nor to the previously isolated plasmid pHV2. Partial restriction maps of these plasmids were determined. One of the plasmids contains chromosomal repetitive sequences as judged by the existence of homologous sequences in the chromosomal DNA of the three isolates. Using the protoplast fusion technique, we showed that at least one of the newly isolated plasmids is compatible with pHV2.Key words: Halobacterium volcanii, archaebacterium, plasmids.
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Guo, Rui, Gen Li, Leilei Lu, Shan Sun, Ting Liu, Mengsha Li, Yong Zheng, Albertha J. M. Walhout, Jun Wu, and Huixin Li. "The Plasmid pEX18Gm Indirectly Increases Caenorhabditis elegans Fecundity by Accelerating Bacterial Methionine Synthesis." International Journal of Molecular Sciences 23, no. 9 (April 30, 2022): 5003. http://dx.doi.org/10.3390/ijms23095003.
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Plasmids are mostly found in bacteria as extrachromosomal genetic elements and are widely used in genetic engineering. Exploring the mechanisms of plasmid–host interaction can provide crucial information for the application of plasmids in genetic engineering. However, many studies have generally focused on the influence of plasmids on their bacterial hosts, and the effects of plasmids on bacteria-feeding animals have not been explored in detail. Here, we use a “plasmid–bacteria–Caenorhabditis elegans” model to explore the impact of plasmids on their host bacteria and bacterivorous nematodes. First, the phenotypic responses of C. elegans were observed by feeding Escherichia coli OP50 harboring different types of plasmids. We found that E. coli OP50 harboring plasmid pEX18Gm unexpectedly increases the fecundity of C. elegans. Subsequently, we found that the plasmid pEX18Gm indirectly affects C. elegans fecundity via bacterial metabolism. To explore the underlying regulatory mechanism, we performed bacterial RNA sequencing and performed in-depth analysis. We demonstrated that the plasmid pEX18Gm upregulates the transcription of methionine synthase gene metH in the bacteria, which results in an increase in methionine that supports C. elegans fecundity. Additionally, we found that a pEX18Gm-induced increase in C. elegans can occur in different bacterial species. Our findings highlight the plasmid–bacteria–C. elegans model to reveal the mechanism of plasmids’ effects on their host and provide a new pattern for systematically studying the interaction between plasmids and multi-species.
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Qin, Zhongjun, Meijuan Shen, and Stanley N. Cohen. "Identification and Characterization of a pSLA2 Plasmid Locus Required for Linear DNA Replication and Circular Plasmid Stable Inheritance in Streptomyces lividans." Journal of Bacteriology 185, no. 22 (November 15, 2003): 6575–82. http://dx.doi.org/10.1128/jb.185.22.6575-6582.2003.
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ABSTRACT Streptomyces linear plasmids and linear chromosomes can replicate also in a circular form when their telomeres are deleted. The 17-kb linear plasmid pSLA2 has been a useful model in studies of such replicons. Here we report that the minimal origin initiating replication of pSLA2-derived plasmids as circular molecules cannot propagate these plasmids in a linear mode unless they also contain a novel plasmid-encoded locus, here named rlrA (required for linear replication). In contrast with the need for rlrA to accomplish replication of telomere-containing linear plasmids, expression of rlrA, which encodes two LuxR family regulatory domains, interferes with the establishment of pSLA2 in circular form in Streptomyces lividans transformants. The additional presence of an adjacent divergently transcribed locus, rorA (rlrA override), which strongly resembles the kor (kil override) transcription control genes identified previously on Streptomyces plasmids, reversed the detrimental effects of rlrA on plasmid establishment and additionally stabilized circular plasmid inheritance by spores during the S. lividans life cycle. While the effects of the rlrA/rorA locus of pSLA2 were seen also on linear plasmids derived from the unrelated SLP2 replicon, they did not extend to plasmids whose replication was initiated at a cloned chromosomal origin. Our results establish the existence of, and provide the initial description of, a novel plasmid-borne regulatory system that differentially affects the propagation of linear and circular plasmids in Streptomyces.
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Chen, Chin-Yi, Ly-Huong T. Nguyen, and Terence P. Strobaugh. "Sequence analysis and plasmid mobilization of a 6.6-kb kanamycin resistance plasmid, pSNC3-Kan, from a Salmonella enterica serotype Newport isolate." PLOS ONE 17, no. 7 (July 14, 2022): e0268502. http://dx.doi.org/10.1371/journal.pone.0268502.
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Research on the transfer of antibiotic resistance plasmids has been mainly focused on the large multi-drug resistance conjugative plasmids, while the transmission of small mobilizable plasmids remains under-investigated. A series of diverse ColE-like kanamycin resistance plasmids (“KanR plasmids”) from Salmonella enterica were characterized previously. In this study, the 6.6-kb pSNC3-Kan from a Salmonella enterica serotype Newport isolate was investigated. It possessed highly conserved RNA I/II and Tn602 (IS903-aph-IS903) regions to two other KanR plasmids pSe-Kan and pSBardo-Kan, but carried a mobC-mobA/BD operon. The mobilization proteins encoded by the mob operon of pSNC3-Kan showed high sequence identity (~95%) to those of an E. coli plasmid pEC34B, except that MobE was not present; and were much less conserved to those of another KanR plasmid pSN11/00Kan (43% - 86% identity). Four structurally different KanR plasmids were investigated for their ability to be mobilized by the conjugal transfer (tra) genes from F and IncP plasmids. Transfer genes derived from IncP plasmids can efficiently mobilize KanR plasmids possessing the mob operons (mobC-mobA/BD), such as pSNC3-Kan and pSN11/00Kan, in bi-parental mating experiments. On the other hand, F tra genes were able to mobilize pU302S, pSNC3-Kan and pSe-Kan, but not pSN11/00Kan. A plasmid-borne mob operon was not required for mobilization of the oriT(F)-bearing pSe-Kan by the F tra genes. This study underscores the complexity of plasmid interaction and the importance of how small mobilizable plasmids may contribute to the spread of antibiotic resistance genes.
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Dahlberg, Cecilia, and Lin Chao. "Amelioration of the Cost of Conjugative Plasmid Carriage in Eschericha coli K12." Genetics 165, no. 4 (December 1, 2003): 1641–49. http://dx.doi.org/10.1093/genetics/165.4.1641.
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Abstract Although plasmids can provide beneficial functions to their host bacteria, they might confer a physiological or energetic cost. This study examines how natural selection may reduce the cost of carrying conjugative plasmids with drug-resistance markers in the absence of antibiotic selection. We studied two plasmids, R1 and RP4, both of which carry multiple drug resistance genes and were shown to impose an initial fitness cost on Escherichia coli. To determine if and how the cost could be reduced, we subjected plasmid-containing bacteria to 1100 generations of evolution in batch cultures. Analysis of the evolved populations revealed that plasmid loss never occurred, but that the cost was reduced through genetic changes in both the plasmids and the bacteria. Changes in the plasmids were inferred by the demonstration that evolved plasmids no longer imposed a cost on their hosts when transferred to a plasmid-free clone of the ancestral E. coli. Changes in the bacteria were shown by the lowered cost when the ancestral plasmids were introduced into evolved bacteria that had been cured of their (evolved) plasmids. Additionally, changes in the bacteria were inferred because conjugative transfer rates of evolved R1 plasmids were lower in the evolved host than in the ancestral host. Our results suggest that once a conjugative bacterial plasmid has invaded a bacterial population it will remain even if the original selection is discontinued.
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Galen, James E., Jay Nair, Jin Yuang Wang, Steven S. Wasserman, Michael K. Tanner, Marcelo B. Sztein, and Myron M. Levine. "Optimization of Plasmid Maintenance in the Attenuated Live Vector Vaccine Strain Salmonella typhiCVD 908-htrA." Infection and Immunity 67, no. 12 (December 1, 1999): 6424–33. http://dx.doi.org/10.1128/iai.67.12.6424-6433.1999.
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ABSTRACT The broad objective of the research presented here is to develop a noncatalytic plasmid maintenance system for the stabilization of multicopy expression plasmids encoding foreign antigens in aSalmonella typhi live-vector vaccine strain such as CVD 908-htrA. We have enhanced the maintenance of expression plasmids at two independent levels. First, we removed dependence upon balanced-lethal maintenance systems that involve catalytic enzymes expressed from multicopy plasmids; we accomplished this through incorporation into expression plasmids of a postsegregational killing system based on the noncatalytic hok-sok plasmid addiction system from the antibiotic resistance factor pR1. We also included at least one naturally occurring plasmid partition function in our expression plasmids, which eliminates random segregation of these plasmids, thereby enhancing their inheritance and stability; to accomplish this, we incorporated either the par locus from pSC101, the parA locus from pR1, or both. We monitored the stability of optimized expression plasmids within CVD 908-htrA by quantitating expression of a variant of green fluorescent protein (GFPuv) by using flow cytometry. In this report, we demonstrate the utility of this novel plasmid maintenance system in enhancing the stability of our expression plasmids and go on to show that as the copy number of stabilized plasmids increases, the toxicity of GFPuv synthesis also increases. The implications of these observations for the rational design of immunogenic and protective bacterial live vector vaccines are discussed.
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Zhang, Ran, Ana Zeng, Ping Fang, and Zhongjun Qin. "Characterization of Replication and Conjugation of Streptomyces Circular Plasmids pFP1 and pFP11 and Their Ability To Propagate in Linear Mode with Artificially Attached Telomeres." Applied and Environmental Microbiology 74, no. 11 (April 4, 2008): 3368–76. http://dx.doi.org/10.1128/aem.00402-08.
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ABSTRACT Many Streptomyces species harbor circular plasmids (8 to 31 kb) as well as linear plasmids (12 to 1,700 kb). We report the characterization of two newly detected circular plasmids, pFP11 (35,139 bp) and pFP1 (39,360 bp). As on linear plasmids, their replication loci comprise repA genes and adjacent iterons, to which RepA proteins bind specifically in vitro. Plasmids containing the minimal iterons plus the repA locus of pFP11 were inherited extremely unstably; par and additional loci were required for stable inheritance. Surprisingly, plasmids containing replication loci from pFP11 or Streptomyces circular plasmid SCP2 but not from pFP1, SLP1, or pIJ101 propagated in a stable linear mode when the telomeres of a linear plasmid were attached. These results indicate bidirectional replication for pFP11 and SCP2. Both pFP11 and pFP1 contain, for plasmid transfer, a major functional traB gene (encoding a DNA translocase typical for Streptomyces plasmids) as well as, surprisingly, a putative traA gene (encoding a DNA nickase, characteristic of single-stranded DNA transfer of gram-negative plasmids), but this did not appear to be functional, at least in isolation.
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Ishii, Hiroshi, Fujio Hayashi, Shizuko Iyobe, and Hajime Hashimoto. "Characterization and classification of Actinobacillus (Haemophilus) pleuropneumoniae plasmids." American Journal of Veterinary Research 52, no. 11 (November 1, 1991): 1816–20. http://dx.doi.org/10.2460/ajvr.1991.52.11.1816.
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SUMMARY Actinobacillus (Haemophilus) pleuropneumoniae plasmids were characterized and classified. They were isolated from A pleuropneumoniae strains different in serotype, year isolated, or location from which isolated. Six of 8 plasmids encoded streptomycin (Sm) and sulfonamide (Su) resistance (SmSu). One of the other plasmids, pVM105, encoded ampicillin (Ap) resistance and another, pHM0, encoded no drug resistance. All SmSu plasmids were transferred to Escherichia coli strains by transformation. Among them, pABO and pMS260 were 8.1 kb and incompatible with each other; they were stable in E coli. The other SmSu plasmids, pHM1, pVM104, pVM106, and pKD25, were 4.3 kb and did not replicate stably in E coli. The former SmSu plasmids were mobilized in E coli strains by a plasmid RP4, which belonged to incompatibility (Inc) group P, but the latter plasmids were not. Further, each 8.1-kb SmSu plasmid and each 4.3-kb plasmid had the same respective restriction pattern. These results indicated that there were at least 2 types of SmSu plasmids in A pleuropneumoniae. The 2 types were classified in 2 groups: H1(pMS260 and pABO) and H2(pHM1, pVM104, pVM106, and pKD25). The H1 and H2 plasmids belonged to different Inc groups, and H2 plasmids belonged to a different Inc group from that of pHM0 and pVM105.
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Gregorova, Daniela, Jitka Matiasovicova, Alena Sebkova, Marcela Faldynova, and Ivan Rychlik. "Salmonella entericasubsp.entericaserovar Enteritidis harbours ColE1, ColE2, and rolling-circle-like replicating plasmids." Canadian Journal of Microbiology 50, no. 2 (February 1, 2004): 107–12. http://dx.doi.org/10.1139/w03-113.
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Using DNA hybridization, at least three distinct groups of low molecular mass plasmids were identified in Salmonella enterica subsp. enterica serovar Enteritidis. After sequencing representative plasmids from each group, we concluded that they belonged to ColE1, ColE2, and rolling-circle-like replicating plasmids. Plasmid pK (4245 bp) is a representative of widely distributed ColE1 plasmids. Plasmid pP (4301 bp) is homologous to ColE2 plasmids and was present predominantly in single-stranded DNA form. The smallest plasmids pJ (2096 bp) and pB (1983 bp) were classified as rolling-circle-like replicating plasmids. Both encoded only a single protein essential for their own replication, and they must have existed in an unusual molecular structure, as (i) they were capable of hybridization without denaturation, (ii) their DNA could be linearized with S1 nuclease, and (iii) even after such treatment, the ability to hybridize without denaturation did not disappear.Key words:Salmonella enterica subsp. enterica serovar Enteritidis, ColE1, ColE2, RCR, plasmid, rolling-circle replication.
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Nishida, Hiromi. "Comparative Analyses of Base Compositions, DNA Sizes, and Dinucleotide Frequency Profiles in Archaeal and Bacterial Chromosomes and Plasmids." International Journal of Evolutionary Biology 2012 (March 26, 2012): 1–5. http://dx.doi.org/10.1155/2012/342482.
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In the present paper, I compared guanine-cytosine (GC) contents, DNA sizes, and dinucleotide frequency profiles in 109 archaeal chromosomes, 59 archaeal plasmids, 1379 bacterial chromosomes, and 854 bacterial plasmids. In more than 80% of archaeal and bacterial plasmids, the GC content was lower than that of the host chromosome. Furthermore, most of the differences in GC content found between a plasmid and its host chromosome were less than 10%, and the GC content in plasmids and host chromosomes was highly correlated (Pearson’s correlation coefficient in bacteria and 0.917 in archaea). These results support the hypothesis that horizontal gene transfers have occurred frequently via plasmid distribution during evolution. GC content and chromosome size were more highly correlated in bacteria () than in archaea (). Interestingly, there was a tendency for archaea with plasmids to have higher GC content in the chromosome and plasmid than those without plasmids. Thus, the dinucleotide frequency profile of the archaeal plasmids has a bias toward high GC content.
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Ishii, Hiroshi, Tsuguaki Fukuyasu, Shizuko Iyobe, and Hajime Hashimoto. "Characterization of newly isolated plasmids from Actinobacillus pleuropneumoniae." American Journal of Veterinary Research 54, no. 5 (May 1, 1993): 701–8. http://dx.doi.org/10.2460/ajvr.1993.54.05.701.
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Summary The genetic basis of drug-resistant strains of Actinobacillus pleuropneumoniae in Japan was studied. The A pleuropneumoniae strains AV277 and AV281 that belong to serotype 2 were resistant to streptomycin (sm) and sulfonamide (sa). Both strains had an 8.1-kilobase (kb) sm-sa plasmid that was previously classified in the H1 group. The AV177 (serotype 1) strain was resistant to sm, sa, ampicillin, and kanamycin (km), but did not have any plasmids. The AV319 and AV324 (serotype 1) strains were resistant to sm, sa, tetracycline (tc), and chloramphenicol (cp). The AV318 (serotype 12) strain was resistant to sm, sa, tc, minocycline, and cp. These 3 strains (AV319, AV324, and AV318) had a 4.3-kb sm-sa plasmid and a 5.2-kb cp plasmid. The 4.3-kb plasmid was classified in the H2 group. The AV263 (serotype 1) strain was resistant to sm, sa, km, tc, and cp. It had a 5.2-kb cp plasmid and a 6.6-kb sm-sa-km plasmid. Both plasmids did not replicate stably in Escherichia coli strains. The former 5.2-kb plasmid was mobilized in E coli strains by plasmid RP4, which belonged to incompatibility P with broad host range, but the latter 6.6-kb plasmid was not so mobilized. Three 5.2-kb cp plasmids isolated from strains AV319, AV324, and AV318, had the same restriction endonuclease pattern after digestion with Ava I and EcoRI. They coexisted with H1 group plasmids in the incompatibility test, and coexisted also with H2 group plasmids of the original A pleuropneumoniae strains. Results indicated that the 5.2-kb cp plasmids could be classified in a new incompatibility group, H3. In this study, 4 types of plasmids were isolated, but no plasmids encoded tc and minocycline resistance.
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Tonin, Patricia N., and Robert B. Grant. "Genetic and physical characterization of trimethoprim resistance plasmids from Shigella sonnei and Shigella flexneri." Canadian Journal of Microbiology 33, no. 10 (October 1, 1987): 905–13. http://dx.doi.org/10.1139/m87-157.
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Analysis of six Shigella flexneri and four S. sonnei isolates with trimethoprim (Tp) resistance from clinical cases in Ontario has shown that, in all isolates, the Tp resistance is mediated by gene(s) on conjugative, multiple antibiotic-resistance plasmids. The physical and genetic characterization of these plasmids revealed that there are three different Tp resistance plasmids. One group, composed of all six S. flexneri plasmids, consists of plasmids which are about 70 megadaltons (MDa) and inhibit the fertility of an Escherichia coli Hfr strain (Fi+). A representative member of this group, pPT4, demonstrates a weak incompatibility reaction with IncFI plasmid R455-2. Another group, three of the four S. sonnei plasmids, contains plasmids which are about 43 MDa, Fi−, and mediate propagation of phage PRD1. The third group, the remaining S. sonnei plasmid, is 53 MDa,fi+, mediates propagation of phages fd and MS2, and is incompatible with IncFII plasmid R100. These plasmids also have been differentiated by restriction endonuclease fragment profiles. Analysis of pPT4 has revealed that the Tp resistance of this plasmid is transposable. The transposon, Tn536, is different from previously described Tp resistance transposons; it is 16 MDa, and in addition to Tp, it encodes resistance to mercuric chloride ions, spectinomycin, streptomycin, and sulfonamides.
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Loftie-Eaton, Wesley, and Douglas E. Rawlings. "Comparative Biology of Two Natural Variants of the IncQ-2 Family Plasmids, pRAS3.1 and pRAS3.2." Journal of Bacteriology 191, no. 20 (August 14, 2009): 6436–46. http://dx.doi.org/10.1128/jb.00864-09.
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ABSTRACT Plasmids pRAS3.1 and pRAS3.2 are two closely related, natural variants of the IncQ-2 plasmid family that have identical plasmid backbones except for two differences. Plasmid pRAS3.1 has five 6-bp repeat sequences in the promoter region of the mobB gene and four 22-bp iterons in its oriV region, whereas pRAS3.2 has only four 6-bp repeats and three 22-bp iterons. Plasmid pRAS3.1 was found to have a higher copy number than pRAS3.2, and we show that the extra 6-bp repeat results in an increase in mobB and downstream mobA/repB expression. Placement of repB (primase) behind an arabinose-inducible promoter in trans resulted in an increase in repB expression and an approximately twofold increase in the copy number of plasmids with identical numbers of 22-bp iterons. The pRAS3 plasmids were shown to have a previously unrecognized toxin-antitoxin plasmid stability module within their replicons. The ability of the pRAS3 plasmids to mobilize the oriT regions of two other plasmids of the IncQ-2 family, pTF-FC2 and pTC-F14, suggested that the mobilization proteins pRAS3 are relaxed and can mobilize oriT regions with substantially different sequences. Plasmids pRAS3.1 and pRAS3.2 were highly incompatible with plasmids pTF-FC2 and pTC-F14, and this incompatibility was removed on inactivation of an open reading frame situated downstream of the mobCDE mobilization genes rather than being due to the 22-bp oriV-associated iterons. We propose that the pRAS3 plasmids represent a third, γ incompatibility group within the IncQ-2 family plasmids.
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Phan, Minh-Duy, Claire Kidgell, Satheesh Nair, Kathryn E. Holt, Arthur K. Turner, Jason Hinds, Philip Butcher, et al. "Variation in Salmonella enterica Serovar Typhi IncHI1 Plasmids during the Global Spread of Resistant Typhoid Fever." Antimicrobial Agents and Chemotherapy 53, no. 2 (November 17, 2008): 716–27. http://dx.doi.org/10.1128/aac.00645-08.
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ABSTRACT A global collection of plasmids of the IncHI1 incompatibility group from Salmonella enterica serovar Typhi were analyzed by using a combination of DNA sequencing, DNA sequence analysis, PCR, and microarrays. The IncHI1 resistance plasmids of serovar Typhi display a backbone of conserved gene content and arrangement, within which are embedded preferred acquisition sites for horizontal DNA transfer events. The variable regions appear to be preferred acquisition sites for DNA, most likely through composite transposition, which is presumably driven by the acquisition of resistance genes. Plasmid multilocus sequence typing, a molecular typing method for IncHI1 plasmids, was developed using variation in six conserved loci to trace the spread of these plasmids and to elucidate their evolutionary relationships. The application of this method to a collection of 36 IncHI1 plasmids revealed a chronological clustering of plasmids despite their difference in geographical origins. Our findings suggest that the predominant plasmid types present after 1993 have not evolved directly from the earlier predominant plasmid type but have displaced them. We propose that antibiotic selection acts to maintain resistance genes on the plasmid, but there is also competition between plasmids encoding the same resistance phenotype.
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Feizabadi, Mohammad Mehdi, Sorour Asadi, Maryam Zohari, Sara Gharavi, and Gelavizh Etemadi. "Genetic characterization of high-level gentamicin-resistant strains of Enterococcus faecalis in Iran." Canadian Journal of Microbiology 50, no. 10 (October 1, 2004): 869–72. http://dx.doi.org/10.1139/w04-069.
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The prevalence of resistance to high levels of gentamicin among 182 isolates of Enterococcus faecalis from 2 Iranian hospitals was 42%. Gentamicin resistance was associated with conjugative plasmids (>70 kb) in most strains. Fingerprinting using EcoRI and HindIII showed genetic variation among these plasmids and gave evidence of nosocomial outbreaks and persistence of infection in different wards of the study hospitals, as well as transfer of plasmids between genetically diverse isolates. Using EcoRI, hospital-based specific plasmid fingerprints were detected for the isolates that had previously proved to be unrelated by multilocus enzyme electrophoresis, suggesting the persistence of related plasmids at each hospital, though minor changes in these related plasmids could be detected with HindIII.Key words: Enterococcus faecalis, HLGR, plasmid profiling, plasmid fingerprinting.
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Taylor, Diane E., and Elisa C. Brose. "Characterization of incompatibility group HI1 plasmids from Salmonella typhi by restriction endonuclease digestion and hybridization of DNA probes for Tn3, Tn9, and Tn10." Canadian Journal of Microbiology 31, no. 8 (August 1, 1985): 721–29. http://dx.doi.org/10.1139/m85-136.
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Chloramphenicol resistance in Salmonella typhi is mediated by plasmids of the incompatibility group H, subgroup 1 (IncHI1). Eight IncHI1 plasmids from S. typhi strains originating in Mexico, Vietnam, Thailand, and India were examined by restriction enzyme digestion. The restriction enzymes, ApaI, XbaI, and PstI were found to be most useful for comparison of plasmid DNAs. Four plasmids from S. typhi isolated in Mexico, Vietnam, and Thailand between 1972 and 1974 had identical restriction patterns with all three enzymes. The other IncHI1 plasmids showed only minor differences. However, some significant differences were noted between these IncHI1 plasmids and the prototype IncHI1 plasmid R27, which was isolated from S. typhimurium in 1961 and for which a restriction map has been constructed. Southern transfer hybridization with a nick-translated HI1 plasmid as a probe confirmed that there is a great deal of sequence homology among the IncHI1 plasmids. DNA probes were used to locate DNA sequences for ampicillin resistance (Tn3), chloramphenicol resistance (Tn9), tetracycline resistance (Tn10), and the one-way incompatibility between IncHI1 plasmids and the F factor, a characteristic property of IncHI1 plasmids. The results demonstrate that IncHI1 plasmids isolated from S. typhi from widely different geographic sources are very similar. Comparisons between the S. typhi plasmids and R27 indicated that conserved regions of DNA were those involved in conjugative transfer.
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Colgan, D. J., and D. A. Willcocks. "Host–parasite genome relationships in Acridid grasshoppers. II. Patterns of variation in the plasmids of gut bacteria of Caledia captiva and Locusta migratoria." Genome 29, no. 2 (April 1, 1987): 264–71. http://dx.doi.org/10.1139/g87-046.
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Plasmid preparations were made from 110 isolates of Enterobacter cloacae taken from the guts of members of the Caledia captiva complex of grasshoppers to ascertain whether a relationship exists between these extrachromosomal elements and taxonomic variation in the grasshoppers themselves. Fifty-two plasmids, distinguishable by mobility or restriction fragment pattern differences, were identified. Thirty-seven of these were similar in size. Five plasmids were nick translated and used to probe Southern blots. Only three instances of cross homology with another plasmid were found, implying a very high level of sequence diversity in the samples. No explanation of the size uniformity and sequence diversity of the plasmids is entirely satisfactory but it appears most likely that the variation is maintained to serve a variety of adaptive functions. No plasmid was found in grasshoppers of more than one taxon of C. captiva. This may be due to geographical limitations on the distribution of plasmids. If this is so, it remains possible that there is an association of one or more plasmids with taxonomic divergence in this grasshopper complex. Plasmid preparations were also made from 68 bacterial isolates (predominantly E. aerogenes) from laboratory-reared Locusta migratoria and from 72 isolates from other acridid grasshoppers. Plasmids of the size general in C. captiva were discovered in most of these isolates. Some smaller plasmids were also found. As judged by restriction endonuclease digests and Southern blotting, plasmid diversity is much less in this sample of L. migratoria bacteria than in the field-collected C. captiva. The plasmids reported in this paper may be considered as possible vectors for use in the genetic control of locusts. Key words: host–parasite, plasmids, grasshoppers, Enterobacter.
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Sayeed, Sameera, Jihong Li, and Bruce A. McClane. "Virulence Plasmid Diversity in Clostridium perfringens Type D Isolates." Infection and Immunity 75, no. 5 (March 5, 2007): 2391–98. http://dx.doi.org/10.1128/iai.02014-06.
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ABSTRACT Clostridium perfringens type D isolates are important in biodefense and also cause natural enterotoxemias in sheep, goats, and occasionally cattle. In these isolates, the gene (etx) encoding ε-toxin is thought to reside on poorly characterized large plasmids. Type D isolates sometimes also produce other potentially plasmid-encoded toxins, including C. perfringens enterotoxin and beta2 toxin, encoded by the cpe and cbp2 genes, respectively. In the current study we demonstrated that the etx, cpe, and cpb2 genes are carried on plasmids in type D isolates and characterized the toxin-encoding plasmids to obtain insight into their genetic organization, potential transferability, and diversity. Southern blotting of pulsed-field gels showed that the etx gene of type D isolates can be present on at least five different plasmids, whose sizes range from 48 to 110 kb. The etx plasmids also typically carried IS1151 and tcp open reading frames (ORFs) known to mediate conjugative transfer of C. perfringens plasmid pCW3. PCR studies revealed that other than their tcp ORFs, etx plasmids of type D isolates do not carry substantial portions of the conserved or variable regions in the cpe plasmids of type A isolates. Southern blotting also demonstrated that in type D isolates the cpe and cpb2 genes are sometimes present on the etx plasmid. Collectively, these findings confirmed that the virulence of type D isolates is heavily plasmid dependent and indicated that (i) a single type D isolate can carry multiple virulence plasmids, (ii) a single type D virulence plasmid can carry up to three different toxin genes, and (iii) many etx plasmids should be capable of conjugative transfer.
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McMillan, Elizabeth A., Jamie L. Wasilenko, Kaitlin A. Tagg, Jessica C. Chen, Mustafa Simmons, Sushim K. Gupta, Glenn E. Tillman, Jason Folster, Charlene R. Jackson, and Jonathan G. Frye. "Carriage and Gene Content Variability of the pESI-Like Plasmid Associated with Salmonella Infantis Recently Established in United States Poultry Production." Genes 11, no. 12 (December 18, 2020): 1516. http://dx.doi.org/10.3390/genes11121516.
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Salmonella Infantis carrying extended spectrum β-lactamase blaCTX-M-65 on a pESI-like megaplasmid has recently emerged in United States poultry. In order to determine the carriage rate and gene content variability of this plasmid in U.S. Salmonella Infantis, whole genome sequences of Salmonella isolates from humans and animals in the U.S. and internationally containing the pESI-like plasmid were analyzed. The U.S. Department of Agriculture Food Safety and Inspection Service (FSIS) identified 654 product sampling isolates containing pESI-like plasmids through hazard analysis and critical control point (HACCP) verification testing in 2017 and 2018. The Centers for Disease Control and Prevention identified 55 isolates with pESI-like plasmids in 2016–2018 through the National Antimicrobial Resistance Monitoring System. Approximately 49% of pESI-like plasmids from FSIS verification isolates and 71% from CDC NARMS contained blaCTX-M-65. Pan-plasmid genome analysis was also performed. All plasmids contained traN and more than 95% contained 172 other conserved genes; 61% contained blaCTX-M-65. In a hierarchical clustering analysis, some plasmids from U.S. animal sources clustered together and some plasmids from South America clustered together, possibly indicating multiple plasmid lineages. However, most plasmids contained similar genes regardless of origin. Carriage of the pESI-like plasmid in U.S. appears to be limited to Salmonella Infantis and carriage rates increased from 2017 to 2018.
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Mayer, L. W. "Use of plasmid profiles in epidemiologic surveillance of disease outbreaks and in tracing the transmission of antibiotic resistance." Clinical Microbiology Reviews 1, no. 2 (April 1988): 228–43. http://dx.doi.org/10.1128/cmr.1.2.228.
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Plasmids are circular deoxyribonucleic acid molecules that exist in bacteria, usually independent of the chromosome. The study of plasmids is important to medical microbiology because plasmids can encode genes for antibiotic resistance or virulence factors. Plasmids can also serve as markers of various bacterial strains when a typing system referred to as plasmid profiling, or plasmid fingerprinting is used. In these methods partially purified plasma deoxyribonucleic acid species are separated according to molecular size by agarose gel electrophoresis. In a second procedure, plasmid deoxyribonucleic acid which has been cleaved by restriction endonucleases can be separated by agarose gel electrophoresis and the resulting pattern of fragments can be used to verify the identity of bacterial isolates. Because many species of bacteria contain plasmids, plasmid profile typing has been used to investigate outbreaks of many bacterial diseases and to trace inter- and intra-species spread of antibiotic resistance.
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Rajanna, Chythanya, Tamara Revazishvili, Mohammed H. Rashid, Svetlana Chubinidze, Lela Bakanidze, Shota Tsanava, Paata Imnadze, et al. "Characterization of pPCP1 Plasmids inYersinia pestisStrains Isolated from the Former Soviet Union." International Journal of Microbiology 2010 (2010): 1–9. http://dx.doi.org/10.1155/2010/760819.
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Complete sequences of 9.5-kb pPCP1 plasmids in threeYersinia pestisstrains from the former Soviet Union (FSU) were determined and compared with those of pPCP1 plasmids in three well-characterized, non-FSUY. pestisstrains (KIM, CO92, and 91001). Two of the FSU plasmids were from strains C2614 and C2944, isolated from plague foci in Russia, and one plasmid was from strain C790 from Kyrgyzstan. Sequence analyses identified four sequence types among the six plasmids. The pPCP1 plasmids in the FSU strains were most genetically related to the pPCP1 plasmid in the KIM strain and least related to the pPCP1 plasmid inY. pestis91001. The FSU strains generally had larger pPCP1 plasmid copy numbers compared to strain CO92. Expression of the plasmid'splagene was significantly (P≤.05) higher in strain C2944 than in strain CO92. Givenpla's role inY. pestisvirulence, this difference may have important implications for the strain's virulence.
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Liu, Ziyi, Zhiqiang Wang, Xiaoyu Lu, Kai Peng, Sheng Chen, Susu He, and Ruichao Li. "Structural Diversity, Fitness Cost, and Stability of a BlaNDM-1-Bearing Cointegrate Plasmid in Klebsiella pneumoniae and Escherichia coli." Microorganisms 9, no. 12 (November 25, 2021): 2435. http://dx.doi.org/10.3390/microorganisms9122435.
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Cointegrate/hybrid plasmids combine the genetic elements of two or more plasmids and generally carry abundant antimicrobial resistance determinants. Hence, the spread of cointegrate plasmids will accelerate the transmission of AMR genes. To evaluate the transmission risk caused by cointegrate plasmids, we investigated the structural diversity, fitness cost, and stability of a cointegrate plasmid in Klebsiella pneumoniae YZ6 and Escherichia coli EC600. The cointegrate plasmid pSL131_IncA/C_IncX3 was from a clinical Salmonella Lomita strain. After transferring the plasmid into E. coli EC600 by conjugation, we observed plasmids with different structures, including a full-length original plasmid and two truncated versions. By contrast, DNA fragment deletion and blaCTX-M-14 gene insertion in the plasmid were detected in a transconjugant derived from K. pneumoniae YZ6. These results suggest that the structure of the plasmid was unstable during conjugation. Furthermore, both the full-length plasmid in EC600 and the structurally reorganized plasmid in YZ6 imposed a fitness cost on the bacterial host and enhanced biofilm formation ability. Serial passaging in antibiotic-free medium resulted in a rapid decline of the plasmid in YZ6. However, the stability of the structurally reorganized plasmid in YZ6 was improved via serial passaging in antibiotic-containing medium. SNP calling revealed that mutations of the outer membrane porin may play an essential role in this process. These findings indicate that structural versatility could contribute to the dissemination of cointegrate plasmids. Although the plasmid incurred a fitness cost in other Enterobacteriaceae species, positive selection could alleviate the adverse effects.
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Hingston, Patricia, Thomas Brenner, Lisbeth Truelstrup Hansen, and Siyun Wang. "Comparative Analysis of Listeria monocytogenes Plasmids and Expression Levels of Plasmid-Encoded Genes during Growth under Salt and Acid Stress Conditions." Toxins 11, no. 7 (July 20, 2019): 426. http://dx.doi.org/10.3390/toxins11070426.
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Listeria monocytogenes strains are known to harbour plasmids that confer resistance to sanitizers, heavy metals, and antibiotics; however, very little research has been conducted into how plasmids may influence L. monocytogenes’ ability to tolerate food-related stresses. To investigate this, a library (n = 93) of L. monocytogenes plasmid sequences were compared. Plasmid sequences were divided into two groups (G1 and G2) based on a repA phylogeny. Twenty-six unique plasmid types were observed, with 13 belonging to each of the two repA-based groups. G1 plasmids were significantly (p < 0.05) smaller than G2 plasmids but contained a larger diversity of genes. The most prevalent G1 plasmid (57,083 bp) was observed in 26 strains from both Switzerland and Canada and a variety of serotypes. Quantitative PCR (qPCR) revealed a >2-fold induction of plasmid-contained genes encoding an NADH peroxidase, cadmium ATPase, multicopper oxidase, and a ClpL chaperone protein during growth under salt (6% NaCl) and acid conditions (pH 5) and ProW, an osmolyte transporter, under salt stress conditions. No differences in salt and acid tolerance were observed between plasmid-cured and wildtype strains. This work highlights the abundance of specific plasmid types among food-related L. monocytogenes strains, the unique characteristics of G1 and G2 plasmids, and the possible contributions of plasmids to L. monocytogenes tolerance to food-related stresses.
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Paganini, Julian A., Jesse J. Kerkvliet, Lisa Vader, Nienke L. Plantinga, Rodrigo Meneses, Jukka Corander, Rob J. L. Willems, Sergio Arredondo-Alonso, and Anita C. Schürch. "PlasmidEC and gplas2: an optimized short-read approach to predict and reconstruct antibiotic resistance plasmids in Escherichia coli." Microbial Genomics 10, no. 2 (February 20, 2024). http://dx.doi.org/10.1099/mgen.0.001193.
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Accurate reconstruction of Escherichia coli antibiotic resistance gene (ARG) plasmids from Illumina sequencing data has proven to be a challenge with current bioinformatic tools. In this work, we present an improved method to reconstruct E. coli plasmids using short reads. We developed plasmidEC, an ensemble classifier that identifies plasmid-derived contigs by combining the output of three different binary classification tools. We showed that plasmidEC is especially suited to classify contigs derived from ARG plasmids with a high recall of 0.941. Additionally, we optimized gplas, a graph-based tool that bins plasmid-predicted contigs into distinct plasmid predictions. Gplas2 is more effective at recovering plasmids with large sequencing coverage variations and can be combined with the output of any binary classifier. The combination of plasmidEC with gplas2 showed a high completeness (median=0.818) and F1-Score (median=0.812) when reconstructing ARG plasmids and exceeded the binning capacity of the reference-based method MOB-suite. In the absence of long-read data, our method offers an excellent alternative to reconstruct ARG plasmids in E. coli.
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Kottara, Anastasia, James P. J. Hall, and Michael A. Brockhurst. "The proficiency of the original host species determines community-level plasmid dynamics." FEMS Microbiology Ecology 97, no. 4 (February 13, 2021). http://dx.doi.org/10.1093/femsec/fiab026.
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ABSTRACTPlasmids are common in natural bacterial communities, facilitating bacterial evolution via horizontal gene transfer. Bacterial species vary in their proficiency to host plasmids: whereas plasmids are stably maintained in some species regardless of selection for plasmid-encoded genes, in other species, even beneficial plasmids are rapidly lost. It is, however, unclear how this variation in host proficiency affects plasmid persistence in communities. Here, we test this using multispecies bacterial soil communities comprising species varying in their proficiency to host a large conjugative mercury resistance plasmid, pQBR103. The plasmid reached higher community-level abundance where beneficial and when introduced to the community in a more proficient host species. Proficient plasmid host species were also better able to disseminate the plasmid to a wider diversity of host species. These findings suggest that the dynamics of plasmids in natural bacterial communities depend not only upon the plasmid's attributes and the selective environment but also upon the proficiency of their host species.
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Wang, Teng, and Lingchong You. "The persistence potential of transferable plasmids." Nature Communications 11, no. 1 (November 4, 2020). http://dx.doi.org/10.1038/s41467-020-19368-7.
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Abstract Conjugative plasmids can mediate the spread and maintenance of diverse traits and functions in microbial communities. This role depends on the plasmid’s ability to persist in a population. However, for a community consisting of multiple populations transferring multiple plasmids, the conditions underlying plasmid persistence are poorly understood. Here, we describe a plasmid-centric framework that makes it computationally feasible to analyze gene flow in complex communities. Using this framework, we derive the ‘persistence potential’: a general, heuristic metric that predicts the persistence and abundance of any plasmids. We validate the metric with engineered microbial consortia transferring mobilizable plasmids and with quantitative data available in the literature. We believe that our framework and the resulting metric will facilitate a quantitative understanding of natural microbial communities and the engineering of microbial consortia.