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306100.8610Tetracycline-resistance encoding plasmids from Paenibacillus larvae, the causal agent of American foulbrood disease, isolated from commercial honeys. Paenibacillus larvae, the causal agent of American foulbrood disease in honeybees, acquires tetracycline-resistance via native plasmids carrying known tetracycline-resistance determinants. From three P. larvae tetracycline-resistant strains isolated from honeys, 5-kb-circular plasmids with almost identical sequences, designated pPL373 in strain PL373, pPL374 in strain PL374, and pPL395 in strain PL395, were isolated. These plasmids were highly similar (99%) to small tetracycline-encoding plasmids (pMA67, pBHS24, pBSDMV46A, pDMV2, pSU1, pAST4, and pLS55) that replicate by the rolling circle mechanism. Nucleotide sequences comparisons showed that pPL373, pPL374, and pPL395 mainly differed from the previously reported P. larvae plasmid pMA67 in the oriT region and mob genes. These differences suggest alternative mobilization and/or conjugation capacities. Plasmids pPL373, pPL374, and pPL395 were individually transferred by electroporation and stably maintained in tetracycline-susceptible P. larvae NRRL B-14154, in which they autonomously replicated. The presence of nearly identical plasmids in five different genera of gram-positive bacteria, i.e., Bhargavaea, Bacillus, Lactobacillus, Paenibacillus, and Sporosarcina, inhabiting diverse ecological niches provides further evidence of the genetic transfer of tetracycline resistance among environmental bacteria from soils, food, and marine habitats and from pathogenic bacteria such as P. larvae.201425296446
300710.8537Analysis of the complete nucleotide sequence of an Actinobacillus pleuropneumoniae streptomycin-sulfonamide resistance plasmid, pMS260. pMS260 is an 8.1-kb non-conjugative but mobilizable plasmid that was isolated from Actinobacillus pleuropneumoniae and encodes streptomycin (SM) and sulfonamide (SA) resistances. The analysis of the complete nucleotide sequence of the plasmid revealed a high degree of similarity between pMS260 and the broad-host-range IncQ family plasmids. pMS260 had a single copy of an origin of vegetative replication (oriV). This sequence was identical to a functional oriV of the IncQ-like plasmid pIE1130 that had been exogenously isolated from piggery manure. However, pMS260 did not carry the second IncQ plasmid RSF1010-like oriV region present in pIE1130. A pIE1130-identical transfer origin was also found in pMS260. In addition, the deduced amino acid sequences from 10 open reading frames identified in pMS260 were entirely or nearly identical to those from genes for the replication, mobilization, and SM-SA resistance of pIE1130, indicating that pMS260 belongs to the IncQ-1 gamma subgroup. pMS260 is physically indistinguishable from pIE1130 apart from two DNA regions that contain the chloramphenicol and kanamycin resistance genes (catIII and aphI, respectively) and the second oriV-like region of pIE1130. The codon bias analysis of each gene of pIE1130 and the presence of potential recombination sites in the sulII-strA intergenic regions suggest that pIE1130 seems to have acquired the catIII and aphI genes more recently than the other genes of pIE1130. Therefore, pMS260 may be the ancestor of pIE1130. Information regarding the broad-host-range replicon of pMS260 will be useful in the development of genetic systems for a wide range of bacteria including A. pleuropneumoniae.200414711528
587120.8536Plasmid-mediated florfenicol resistance in Pasteurella trehalosi. OBJECTIVES: A florfenicol-resistant Pasteurella trehalosi isolate from a calf was investigated for the presence and the location of the gene floR. METHODS: The P. trehalosi isolate 13698 was investigated for its in vitro susceptibility to antimicrobial agents and its plasmid content. A 14.9 kb plasmid, designated pCCK13698, was identified by transformation into Pasteurella multocida to mediate resistance to florfenicol, chloramphenicol and sulphonamides. The plasmid was sequenced completely and analysed for its structure and organization. RESULTS: Plasmid pCCK13698 exhibited extended similarity to plasmid pHS-Rec from Haemophilus parasuis including the region carrying the parA, repB, rec and int genes. Moreover, it revealed similarities to plasmid RSF1010 in the parts covering the mobC and repA-repC genes and to plasmid pMVSCS1 in the parts covering the sul2-catA3-strA gene cluster. Moreover, the floR gene area corresponded to that of transposon TnfloR. In addition, two complete insertion sequences were detected that were highly similar to IS1593 from Mannheimia haemolytica and IS26 from Enterobacteriaceae. Several potential recombination sites were identified that might explain the development of plasmid pCCK13698 by recombination events. CONCLUSIONS: The results of this study showed that in the bovine pathogen P. trehalosi, floR-mediated resistance to chloramphenicol and florfenicol was associated with a plasmid, which also carried functionally active genes for resistance to sulphonamides (sul2) and chloramphenicol (catA3). This is to the best of our knowledge the first report of resistance genes in P. trehalosi and only the second report of the presence of a florfenicol-resistance gene in target bacteria of the family Pasteurellaceae.200616670108
301730.8535The ancient small mobilizable plasmid pALWED1.8 harboring a new variant of the non-cassette streptomycin/spectinomycin resistance gene aadA27. The small mobilizable plasmid pALWED1.8 containing a novel variant of the streptomycin/spectinomycin resistance gene aadA27 was isolated from the permafrost strains of Acinetobacter lwoffii. The 4135bp plasmid carries mobА and mobC genes that mediate its mobilization by conjugative plasmids. The nucleotide sequences of mobА and mobC are similar to those of mobilization genes of the modern plasmid pRAY* and its variants, which contain aadB gene, and are widespread among the pathogenic strains of Acinetobacter baumannii. Almost identical pALWED1.8 variants were detected in modern environmental Аcinetobacter strains. A highly similar plasmid was revealed in a strain of Acinetobacter parvus isolated from mouse intestine. Furthermore, we discovered six previously unidentified variants of plasmids related to pALWED1.8 and pRAY* in public databases. In contrast to most known variants of aadA which are cassette genes associated with integrons, the aadA27 variant harbored by pALWED1.8 is a non-cassette, autonomously transcribed gene. Non-cassette aadA genes with 96% sequence identity to aadA27 were detected in the chromosomes of Acinetobacter gyllenbergii and several uncharacterized strains of Аcinetobacter sp. Moreover, we revealed that the autonomous aadA-like genes are present in the chromosomes of many gram-positive and gram-negative bacteria. The phylogenetic analysis of amino acid sequences of all identified AadA proteins showed the following: (i) cassette aadA genes form a separate monophyletic group and mainly reside on plasmids and (ii) chromosomal non-cassette aadA genes are extremely diverse and can be inherited both vertical and via horizontal gene transfer.201626896789
587240.8534Characterization of the plasmids harbouring the florfenicol resistance gene floR in Glaesserella parasuis and Actinobacillus indolicus. OBJECTIVES: The aim of this study was to characterize the floR-carrying plasmids originating from Glaesserella parasuis and Actinobacillus indolicus isolated from pigs with respiratory disease in China. METHODS: A total of 125 G. parasuis and 28 A. indolicus strains collected between 2009 and 2022 were screened for florfenicol resistance. Characterization of floR-positive isolates and plasmids were determined by antimicrobial susceptibility testing, serotyping, multilocus sequence typing (MLST), conjugation and transformation assays, whole-genome sequencing (WGS), and phylogenetic analysis. RESULTS: One A. indolicus and six G. parasuis were identified as positive for floR. The six G. parasuis were assigned to four different serovars, including serovars 6, 7, 9, and unknown. In addition to strain XP11, six floR genes were located on plasmids. The six floR-bearing plasmids could be transformed into Pasteurella multocida and divided into two different types, including ∼5000 bp and ∼6000 bp plasmids. The ∼5000 bp plasmids consisting of rep, lysR, mobB, and floR genes, exhibited high similarity among Pasteurellaceae bacteria. Furthermore, the ∼6000 bp plasmids, consisting of rep, lysR, mobC, mobA/L, and floR genes, showed high similarity between G. parasuis and Actinobacillus Spp. Notably, WGS results showed that the floR modules of the two types of plasmids could be transferred and integrated into the diverse Pasteurellaceae- origined plasmids. CONCLUSION: This study firstly reported the characterization of floR-carrying plasmids from A. indolicus and a non-virulent serovar of G. parasuis in pigs in China and elucidated the transmission mechanism of the floR resistance gene among the Pasteurellaceae family.202337726088
301850.8531The large Bacillus plasmid pTB19 contains two integrated rolling-circle plasmids carrying mobilization functions. Plasmid pTB19 is a 27-kb plasmid originating from a thermophilic Bacillus species. It was shown previously that pTB19 contains an integrated copy of the rolling-circle type plasmid pTB913. Here we describe the analysis of a 4324-bp region of pTB19 conferring resistance to tetracycline. The nucleotide sequence of this region revealed all the characteristics of a second plasmid replicating via the rolling-circle mechanism. This sequence contained (i) the tetracycline resistance marker of pTB19, which is highly similar to other tetL-genes of gram-positive bacteria; (ii) a hybrid mob gene, which bears relatedness to both the mob-genes of pUB110 and pTB913; (iii) a palU type minus origin identical to those of pUB110 and pTB913; and (iv) a plus origin of replication similar to that of pTB913. A repB-type replication initiation gene sequence identical to that of pTB913 was present, which lacked the middle part (492 bp), thus preventing autonomous replication of this region. The hybrid mob gene was functional in conjugative mobilization of plasmids between strains of Bacillus subtilis.19911946749
149360.8531Coexistence of blaKPC-2 and blaNDM-1 in one IncHI5 plasmid confers transferable carbapenem resistance from a clinical isolate of Klebsiella michiganensis in China. OBJECTIVES: This study firstly identified an IncHI5 plasmid pK254-KPC_NDM co-carrying two different class carbapenemase genes blaKPC-2 and blaNDM-1 in Klebsiella michiganensis K254. METHODS: The strain K254 was sequenced by high-throughput genome sequencing. A detailed genomic and phenotypic characterization of pK254-KPC_NDM was performed. RESULTS: pK254-KPC_NDM displayed the conserve IncHI5 backbone and carried a resistant accessory region: Tn1696-related transposon Tn7414 containing blaKPC-2 and blaNDM-1. A sequence comparison was applied to a collection of four Tn1696-related transposons (Tn7414-Tn7417) harbouring carbapenemase genes. For all these four transposons, the blaNDM-1 was carried by Tn125 derivatives within three different mobile genetic elements. Tn7414 further acquired another carbapenemase gene, blaKPC-2, because of the integration of the local blaKPC-2 genetic environment from Tn6296, resulting in the high-level carbapenem resistance of K. michiganensis K254. The conjugal transfer and plasmid stability experiments confirmed that pK254-KPC_NDM could be transferred intercellularly and keep the stable vertical inheritance in different bacteria, which would contribute to the further dissemination of multiple carbapenemase genes and enhance the adaption and survival of K. michiganensis under complex and diverse antimicrobial selection pressures. CONCLUSION: This study was the first to report the K. michiganensis isolate coharbouring blaKPC-2 and blaNDM-1 in the Tn1696-related transposon in IncHI5 plasmid. The emergence of novel transposons simultaneously carrying multiple carbapenemase genes might contribute to the further dissemination of high-level carbapenem resistance in the isolates of the hospital settings and pose new challenges for the treatment of nosocomial infection.202337714378
53570.8527Improved broad-host-range plasmids for DNA cloning in gram-negative bacteria. Improved broad-host-range plasmid vectors were constructed based on existing plasmids RSF1010 and RK404. The new plasmids pDSK509, pDSK519, and pRK415, have several additional cloning sites and improved antibiotic-resistance genes which facilitate subcloning and mobilization into various Gram-negative bacteria. Several new polylinker sites were added to the Escherichia coli plasmids pUC118 and pUC119, resulting in the new plasmids, pUC128 and pUC129. These plasmids facilitate the transfer of cloned DNA fragments to the broad-host-range vectors. Finally, the broad-host-range cosmid cloning vector pLAFR3 was improved by the addition of a double cos casette to generate the new plasmid, pLAFR5. This latter cosmid simplifies vector preparation and has permitted the rapid cloning of genomic DNA fragments generated with Sau3A. The resulting clones may be introduced into other Gram-negative bacteria by conjugation.19882853689
599980.8526Food isolate Listeria monocytogenes harboring tetM gene plasmid-mediated exchangeable to Enterococcus faecalis on the surface of processed cheese. The genetic basis of tetracycline resistance in a food isolate Listeria monocytogenes (Lm16) was evaluated. Resistance to tetracycline was associated with the presence of the tetM gene in plasmid DNA. The sequence of tetM showed 100% of similarity with the Enterococcus faecalis sequences found in the EMBL database, suggesting that Lm16 received this gene from E. faecalis. Various size bands were detected in the DNA plasmid analysis, the largest being approximately 54.38 kb. Transferability of the tetM gene was achieved in vitro by agar matings between Lm16 and E. faecalis JH2-2, proving the potential for the spread of tetM by horizontal gene transfer. Furthermore, the conjugation experiments were performed on the surface of processed cheese, confirming the transferability in a food matrix. PCR assays were used to confirm the identity of E. faecalis and to detect the tetM gene in transconjugant bacteria. Additionally, the minimal inhibitory concentration for tetracycline and rifampicin and plasmid profiling were performed. This is the first report of a food isolate L. monocytogenes carrying the tetM gene in plasmid DNA, and it highlights the potential risk of spreading antimicrobial resistance genes between different bacteria.201829580513
299890.8523Membrane vesicles derived from Enterococcus faecalis promote the co-transfer of important antibiotic resistance genes located on both plasmids and chromosomes. BACKGROUND: Bacterial membrane vesicles (BMVs) are novel vehicles of antibiotic resistance gene (ARG) transfer in Gram-negative bacteria, but their role in the spread of ARGs in Gram-positive bacteria has not been defined. The purpose of this study was to evaluate the role of MVs in the transmission of antimicrobial resistance in Gram-positive bacteria. METHODS: A linezolid-resistant Enterococcus faecalis CQ20 of swine origin was selected as the donor strain. Linezolid-susceptible E. faecalis SC032 of human origin, Enterococcus faecium BM4105 and Escherichia coli were selected as recipient strains. The presence of plasmids (pCQ20-1 and pCQ20-2) and an optrA-carrying transposon Tn6674 in CQ20, MVs and vesiculants was verified by WGS or PCR. MVs were isolated with density gradient centrifugation, and MV-mediated transformation was performed to assess the horizontal transferability of MVs. The MICs for CQ20 and its vesiculants were determined by the broth microdilution method. RESULTS: CQ20-derived MVs (CQ20-MV) were isolated, and PCR identified the presence of two plasmids and the optrA gene in the CQ20-MVs. MV-mediated transformation to E. faecalis SC032 and E. faecium BM4105 was successfully performed, and the WGS data also showed that both plasmids pCQ20-1 and pCQ20-2 and optrA-carrying transposon Tn6674 were transferred to E. faecalis SC032 and E. faecium BM4105, but failed for E. coli. Additionally, vesiculants that had acquired ARGs still had the ability to spread these genes via MVs. CONCLUSIONS: To our knowledge, this is the first report of MV-mediated co-transfer of ARG-carrying plasmids and transposons in the Gram-positive bacterium E. faecium.202438109479
5211100.8522Pediococcus pentosaceus IMI 507025 genome sequencing data. The genome sequence data for the pickled cucumbers isolate, Pediococcus pentosaceus IMI 507025, is reported. The raw reads and analysed genome reads were deposited at NCBI under Bioproject with the accession number PRJNA814992. The number of contigs before and after trimming were 17 and 12 contigs, respectively. The total size of the genome was 1,795,439 bp containing 1,811 total genes, of which 1,751 were coding sequences. IMI 507025 identity was determined via average nucleotide identity (ANI), obtaining an identity value of 99.5994% between IMI 507025 and the type strain P. pentosaceus ATCC 33316, identifying the strain as P. pentosaceus. Screening for the antimicrobial resistance (AMR) and virulence genes in the genome of IMI 507025 showed no hits, confirming the safety of the tested strain. Presence of plasmids was not found.202235864877
5868110.8521Evaluation of plasmid content and tetracycline resistance conjugative transfer in Enterococcus italicus strains of dairy origin. Five Enterococcus italicus strains harbouring tet genes responsible for the tetracycline resistance were subjected to plasmid profile determination studies. For four strains tested the profiles showed between three and six plasmid bands, the size of which ranged between 1.6 and 18.5 kb. Southern hybridization experiments associated tetS and tetK genes with chromosomal DNA in all strains and tetM gene with plasmids of around the same size (18.5 kb) in two of the tested strains. The ability of the new species to transfer tetM gene was studied by transfer experiments with the tetracycline-susceptible recipient strains E. faecalis JH2-2 and OG1RF; mobilization experiments were performed with E. faecalis JH 2-2 harbouring the conjugative plasmid pIP501as helper plasmid. The results obtained show that the new enterococcal species was able to acquire antibiotic resistance by conjugation, but not to transfer its plasmids to other bacteria. Further PCR and hybridization experiments carried out to assess the presence of mobilization sequences also suggest that the tetM plasmid from E. italicus is a non-mobilizable plasmid.200919484299
5873120.8519pDB2011, a 7.6 kb multidrug resistance plasmid from Listeria innocua replicating in Gram-positive and Gram-negative hosts. pDB2011, a multidrug resistance plasmid isolated from the foodborne Listeria innocua strain TTS-2011 was sequenced and characterized. Sequence analysis revealed that pDB2011 had a length of 7641 bp and contained seven coding DNA sequences of which two were annotated as replication proteins, one as a recombination/mobilization protein and one as a transposase. Furthermore, pDB2011 harbored the trimethoprim, spectinomycin and macrolide-lincosamide-streptogramin B resistance genes dfrD, spc and erm(A), respectively. However, pDB2011 was only associated with trimethoprim and spectinomycin resistance phenotypes and not with phenotypic resistance to erythromycin. A region of the plasmid encoding the resistance genes spc and erm(A) plus the transposase was highly similar to Staphylococcus aureus transposon Tn554. The dfrD gene was 100% identical to dfrD found in a number of Listeria monocytogenes isolates. Additionally, assessment of the potential host range of pDB2011 revealed that the plasmid was able to replicate in Lactococcus lactis subsp. cremoris MG1363 as well as in Escherichia coli MC1061 and DH5α. This study reports the first multidrug resistance plasmid in L. innocua. A large potential for dissemination of pDB2011 is indicated by its host range of both Gram-positive and Gram-negative bacteria.201323774482
825130.8516Attaching effacement of the rabbit enterocyte brush border is encoded on a single 96.5-kilobase-pair plasmid in an enteropathogenic Escherichia coli O111 strain. An enteropathogenic Escherichia coli (EPE) O111 serotype a,b,H- strain carried the following four plasmids: pLV501 (96.5 kilobase pairs [kbp]) specifying resistance to chloramphenicol, tetracycline, and kanamycin; pLV502 (8 kbp) specifying ampicillin resistance; pLV503 (1.9 kbp) specifying streptomycin resistance; and pLV504 (80 kbp) with no resistance markers. This EPEC attached to HEp-2 cells to produce localized clumps of bacteria (localized adhesion) and attached intimately to the enterocyte surface, leading to loss of the brush border (attaching effacement). Plasmid pLV501 was also found to specify the ability to produce localized adhesion on HEp-2 cells and attaching effacement in a rabbit ileal explant model system. Restriction maps showed considerable dissimilarities between pLV501 and pMAR-2, an EPEC plasmid carrying the EPEC adherence factor (EAF) genes. Furthermore, pLV501 did not hybridize with the EAF probe, whereas pLV504 did. There was sequence homology between pLV501 and large plasmids in all seven other well-characterized EPEC, only five of which hybridized with the EAF probe. These findings indicate that pLV501 carries at least one of the genes responsible for production of the brush border damage characteristic of EPEC.19902182541
5209140.8514Complete Nucleotide Sequence of pGA45, a 140,698-bp IncFIIY Plasmid Encoding bla IMI-3-Mediated Carbapenem Resistance, from River Sediment. Plasmid pGA45 was isolated from the sediments of Haihe River using Escherichia coli CV601 (gfp-tagged) as recipients and indigenous bacteria from sediment as donors. This plasmid confers reduced susceptibility to imipenem which belongs to carbapenem group. Plasmid pGA45 was fully sequenced on an Illumina HiSeq 2000 sequencing system. The complete sequence of plasmid pGA45 was 140,698 bp in length with an average G + C content of 52.03%. Sequence analysis shows that pGA45 belongs to IncFIIY group and harbors a backbone region which shares high homology and gene synteny to several other IncF plasmids including pNDM1_EC14653, pYDC644, pNDM-Ec1GN574, pRJF866, pKOX_NDM1, and pP10164-NDM. In addition to the backbone region, plasmid pGA45 harbors two notable features including one bla IMI-3-containing region and one type VI secretion system region. The bla IMI-3-containing region is responsible for bacteria carbapenem resistance and the type VI secretion system region is probably involved in bacteria virulence, respectively. Plasmid pGA45 represents the first complete nucleotide sequence of the bla IMI-harboring plasmid from environment sample and the sequencing of this plasmid provided insight into the architecture used for the dissemination of bla IMI carbapenemase genes.201626941718
5867150.8513Molecular analysis of florfenicol-resistant Pasteurella multocida isolates in Germany. OBJECTIVES: Three florfenicol-resistant Pasteurella multocida isolates from Germany, two from swine and one from a calf, were investigated for the genetics and transferability of florfenicol resistance. METHODS: The isolates were investigated for susceptibility to antimicrobial agents and plasmid content. Florfenicol resistance plasmids carrying the gene floR were identified by transformation and PCR. Plasmids were mapped, and a novel plasmid type was sequenced completely. PFGE served to determine the clonality of the isolates. RESULTS: In one porcine and the bovine P. multocida isolate, florfenicol resistance was associated with the plasmid pCCK381 previously described in a bovine P. multocida isolate from the UK. The remaining porcine isolate harboured a new type of floR-carrying plasmid, the 10 226 bp plasmid pCCK1900. Complete sequence analysis identified an RSF1010-like plasmid backbone with the mobilization genes mobA, mobB and mobC, the plasmid replication genes repA, repB and repC, the sulphonamide resistance gene sul2 and the streptomycin resistance genes strA and strB. The floR gene area was integrated into a region downstream of strB, which exhibited homology to the floR flanking regions found in various bacteria. PFGE revealed that the floR-carrying P. multocida strains from Germany were unrelated and also different from the UK strain. CONCLUSIONS: After the UK and France, floR-mediated florfenicol resistance has now also been identified in target bacteria from Germany. PFGE data and the analysis of plasmids strongly suggested that the spread of florfenicol resistance is due to the horizontal transfer of plasmids rather than the clonal dissemination of a resistant P. multocida isolate.200818786941
5407160.8511Resistance mechanisms and tedizolid susceptibility in clinical isolates of linezolid-resistant bacteria in Japan. OBJECTIVES: Studies combining linezolid resistance mechanisms and tedizolid susceptibility in linezolid-resistant clinical isolates are scarce. This study investigated the linezolid resistance mechanisms and tedizolid susceptibility of linezolid-resistant strains isolated clinically in Japan. METHODS: We analysed 25 linezolid-resistant strains of Enterococcus faecium and Enterococcus faecalis isolated from Japanese hospitals between 2015 and 2021. MICs of linezolid and tedizolid were determined using the agar plate dilution method. Each 23S rRNA copy was amplified by PCR, sequenced and analysed for mutations. The linezolid resistance genes cfr, poxtA, optrA, fexA and fexB were also detected by PCR. RESULTS: Drug susceptibility tests revealed that five linezolid-resistant E. faecium isolates had low (≤1 mg/L) tedizolid MICs. Resistance mechanisms included the G2576T mutation in 23S rRNA, the T2504A mutation and the resistance genes optrA, fexA and fexB. The T2504A mutation was identified in one E. faecium isolate, which exhibited linezolid and tedizolid MICs of 64 and 32 mg/L, respectively. CONCLUSIONS: Some linezolid-resistant isolates demonstrated low (≤1 mg/L) tedizolid MICs. To determine whether tedizolid susceptibility testing should be performed on linezolid-resistant isolates, more linezolid-resistant isolates should be collected and tested for tedizolid MICs. Tedizolid MICs were 2-3 doubling dilutions lower than linezolid MICs. The results of this study suggest that future research should investigate whether the T2504A mutation contributes to tedizolid resistance. To our knowledge, this is the first study to report tedizolid susceptibility in E. faecium with the T2504A mutation and in isolate harbouring this mutation.202540463587
9868170.8511The mosaic architecture of Aeromonas salmonicida subsp. salmonicida pAsa4 plasmid and its consequences on antibiotic resistance. Aeromonas salmonicida subsp. salmonicida, the causative agent of furunculosis in salmonids, is an issue especially because many isolates of this bacterium display antibiotic resistances, which limit treatments against the disease. Recent results suggested the possible existence of alternative forms of pAsa4, a large plasmid found in A. salmonicida subsp. salmonicida and bearing multiple antibiotic resistance genes. The present study reveals the existence of two newly detected pAsa4 variants, pAsa4b and pAsa4c. We present the extensive characterization of the genomic architecture, the mobile genetic elements and the antimicrobial resistance genes of these plasmids in addition to the reference pAsa4 from the strain A449. The analysis showed differences between the three architectures with consequences on the content of resistance genes. The genomic plasticity of the three pAsa4 variants could be partially explained by the action of mobile genetic elements like insertion sequences. Eight additional isolates from Canada and Europe that bore similar antibiotic resistance patterns as pAsa4-bearing strains were genotyped and specific pAsa4 variants could be attributed to phenotypic profiles. pAsa4 and pAsa4c were found in Europe, while pAsa4b was found in Canada. In accordance with their content in conjugative transfer genes, only pAsa4b and pAsa4c can be transferred by conjugation in Escherichia coli. The plasticity of pAsa4 variants related to the acquisition of antibiotic resistance indicates that these plasmids may pose a threat in terms of the dissemination of antimicrobial-resistant A. salmonicida subsp. salmonicida bacteria.201627812409
3036180.8510Complete nucleotide sequences of 84.5- and 3.2-kb plasmids in the multi-antibiotic resistant Salmonella enterica serovar Typhimurium U302 strain G8430. The multi-antibiotic resistant (MR) Salmonella enterica serovar Typhimurium phage type U302 strain G8430 exhibits the penta-resistant ACSSuT-phenotype (ampicillin, chloramphenicol, streptomycin, sulfonamides and tetracycline), and is also resistant to carbenicillin, erythromycin, kanamycin, and gentamicin. Two plasmids, 3.2- and 84.5-kb in size, carrying antibiotic resistance genes were isolated from this strain, and the nucleotide sequences were determined and analyzed. The 3.2-kb plasmid, pU302S, belongs to the ColE1 family and carries the aph(3')-I gene (Kan(R)). The 84.5-kb plasmid, pU302L, is an F-like plasmid and contains 14 complete IS elements and multiple resistance genes including aac3, aph(3')-I, sulII, tetA/R, strA/B, bla(TEM-1), mph, and the mer operon. Sequence analyses of pU302L revealed extensive homology to various plasmids or transposons, including F, R100, pHCM1, pO157, and pCTX-M3 plasmids and TnSF1 transposon, in regions involved in plasmid replication/maintenance functions and/or in antibiotic resistance gene clusters. Though similar to the conjugative plasmids F and R100 in the plasmid replication regions, pU302L does not contain oriT and the tra genes necessary for conjugal transfer. This mosaic pattern of sequence similarities suggests that pU302L acquired the resistance genes from a variety of enteric bacteria and underscores the importance of a further understanding of horizontal gene transfer among the enteric bacteria.200716828159
820190.8510Nucleotide sequence analysis of a transposon (Tn5393) carrying streptomycin resistance genes in Erwinia amylovora and other gram-negative bacteria. A class II Tn3-type transposable element, designated Tn5393 and located on plasmid pEa34 from streptomycin-resistant strain CA11 of Erwinia amylovora, was identified by its ability to move from pEa34 to different sites in plasmids pGEM3Zf(+) and pUCD800. Nucleotide sequence analysis reveals that Tn5393 consists of 6,705 bp with 81-bp terminal inverted repeats and generates 5-bp duplications of the target DNA following insertion. Tn5393 contains open reading frames that encode a putative transposase (tnpA) and resolvase (tnpR) of 961 and 181 amino acids, respectively. The two open reading frames are separated by a putative recombination site (res) consisting of 194 bp. Two streptomycin resistance genes, strA and strB, were identified on the basis of their DNA sequence homology to streptomycin resistance genes in plasmid RSF1010. StrA is separated from tnpR by a 1.2-kb insertion element designated IS1133. The tnpA-res-tnpR region of Tn5393 was detected in Pseudomonas syringae pv. papulans Psp36 and in many other gram-negative bacteria harboring strA and strB. Except for some strains of Erwinia herbicola, these other gram-negative bacteria lacked insertion sequence IS1133. The prevalence of strA and strB could be accounted for by transposition of Tn5393 to conjugative plasmids that are then disseminated widely among gram-negative bacteria.19938380801