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179400.8789Insights into the evolution of gene organization and multidrug resistance from Klebsiella pneumoniae plasmid pKF3-140. Plasmid-mediated transfer of drug-resistance genes among various bacterial species is considered one of the most important mechanisms for the spread of multidrug resistance. To gain insights into the evolution of gene organization and antimicrobial resistance in clinical bacterial samples, a complete plasmid genome of Klebsiella pneumoniae pKF3-140 is determined, which has a circular chromosome of 147,416bp in length. Among the 203 predicted genes, 142 have function assignment and about 50 appear to be involved in plasmid replication, maintenance, conjugative transfer, iron acquisition and transport, and drug resistance. Extensive comparative genomic analyses revealed that pKF3-140 exhibits a rather low sequence similarity and structural conservation with other reported K. pneumoniae plasmids. In contrast, the overall organization of pKF3-140 is highly similar to Escherichia coli plasmids p1ESCUM and pUTI89, which indicates the possibility that K. pneumoniae pKF3-140 may have a potential origin in E. coli. Meanwhile, interestingly, several drug resistant genes show high similarity to the plasmid pU302L in Salmonella enterica serovar Typhimurium U302 strain G8430 and the plasmid pK245 in K. pneumoniae. This mosaic pattern of sequence similarities suggests that pKF3-140 might have arisen from E. coli and acquired the resistance genes from a variety of enteric bacteria and underscores the importance of a further understanding of horizontal gene transfer among enteric bacteria.201323402892
306110.8787Tetracycline-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
303620.8774Complete 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
82630.8715Sequence identity with type VIII and association with IS176 of type IIIc dihydrofolate reductase from Shigella sonnei. An uncommon dihydrofolate reductase (DHFR), type IIIc, was coded for by Shigella sonnei that harbors plasmid pBH700 and that was isolated in North Carolina. The trimethoprim resistance gene carried on pBH700 was subcloned and sequenced. The nucleotide sequence of the gene encoding type IIIc DHFR was identical to the gene encoding type VIII DHFR. The type IIIc amino acid sequence was approximately 50% similar to those of DHFRs commonly found in enteric bacteria. Furthermore, this gene was flanked by IS176 (IS26), an insertion sequence usually associated with those of aminoglycoside resistance genes. The gene for type IIIc DHFR was located by hybridization within a 1,993-bp PstI fragment in each of eight conjugative plasmids from geographically diverse strains of S. sonnei. Each plasmid also conferred resistance to ampicillin, streptomycin, and sulfamethoxazole and belonged to incompatibility group M. Plasmids carrying this new trimethoprim resistance gene, which is uniquely associated with IS176, have disseminated throughout the United States.19957695291
587340.8714pDB2011, 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
81250.8711Characterization of plQ5 plasmid originating fromKlebsiella pneumoniae. plQ5 plasmid consists of a group of genes specifying resistance to ampicillin, chloramphenicol, carbencillin, kanamycin and trimethoprim-sulphamethoxazole. It is isolated inKlebslella pneumoniae ZD532, is about 26.8 Kb and is freely transmissible to various bacterial species of Gram-negative bacteria. Physical characterization revealed that plQ5 plasmid has a single site forHindill,BamHI,EcoRI and two sites forBglII restriction enzyme.199024429982
521460.8710Comparative genomic analysis of a new tellurite-resistant Psychrobacter strain isolated from the Antarctic Peninsula. The Psychrobacter genus is a cosmopolitan and diverse group of aerobic, cold-adapted, Gram-negative bacteria exhibiting biotechnological potential for low-temperature applications including bioremediation. Here, we present the draft genome sequence of a bacterium from the Psychrobacter genus isolated from a sediment sample from King George Island, Antarctica (3,490,622 bp; 18 scaffolds; G + C = 42.76%). Using phylogenetic analysis, biochemical properties and scanning electron microscopy the bacterium was identified as Psychrobacter glacincola BNF20, making it the first genome sequence reported for this species. P. glacincola BNF20 showed high tellurite (MIC 2.3 mM) and chromate (MIC 6.0 mM) resistance, respectively. Genome-wide nucleotide identity comparisons revealed that P. glacincola BNF20 is highly similar (>90%) to other uncharacterized Psychrobacter spp. such as JCM18903, JCM18902, and P11F6. Bayesian multi-locus phylogenetic analysis showed that P. glacincola BNF20 belongs to a polyphyletic clade with other bacteria isolated from polar regions. A high number of genes related to metal(loid) resistance were found, including tellurite resistance genetic determinants located in two contigs: Contig LIQB01000002.1 exhibited five ter genes, each showing putative promoter sequences (terACDEZ), whereas contig LIQB1000003.2 showed a variant of the terZ gene. Finally, investigating the presence and taxonomic distribution of ter genes in the NCBI's RefSeq bacterial database (5,398 genomes, as January 2017), revealed that 2,623 (48.59%) genomes showed at least one ter gene. At the family level, most (68.7%) genomes harbored one ter gene and 15.6% exhibited five (including P. glacincola BNF20). Overall, our results highlight the diverse nature (genetic and geographic diversity) of the Psychrobacter genus, provide insights into potential mechanisms of metal resistance, and exemplify the benefits of sampling remote locations for prospecting new molecular determinants.201829479501
81770.8710Mercury resistance transposons in Bacilli strains from different geographical regions. A total of 65 spore-forming mercury-resistant bacteria were isolated from natural environments worldwide in order to understand the acquisition of additional genes by and dissemination of mercury resistance transposons across related Bacilli genera by horizontal gene movement. PCR amplification using a single primer complementary to the inverted repeat sequence of TnMERI1-like transposons showed that 12 of 65 isolates had a transposon-like structure. There were four types of amplified fragments: Tn5084, Tn5085, Tn(d)MER3 (a newly identified deleted transposon-like fragment) and Tn6294 (a newly identified transposon). Tn(d)MER3 is a 3.5-kb sequence that carries a merRETPA operon with no merB or transposase genes. It is related to the mer operon of Bacillus licheniformis strain FA6-12 from Russia. DNA homology analysis shows that Tn6294 is an 8.5-kb sequence that is possibly derived from Tn(d)MER3 by integration of a TnMERI1-type transposase and resolvase genes and in addition the merR2 and merB1 genes. Bacteria harboring Tn6294 exhibited broad-spectrum mercury resistance to organomercurial compounds, although Tn6294 had only merB1 and did not have the merB2 and merB3 sequences for organomercurial lyases found in Tn5084 of B. cereus strain RC607. Strains with Tn6294 encode mercuric reductase (MerA) of less than 600 amino acids in length with a single N-terminal mercury-binding domain, whereas MerA encoded by strains MB1 and RC607 has two tandem domains. Thus, Tn(d)MER3 and Tn6294 are shorter prototypes for TnMERI1-like transposons. Identification of Tn6294 in Bacillus sp. from Taiwan and in Paenibacillus sp. from Antarctica indicates the wide horizontal dissemination of TnMERI1-like transposons across bacterial species and geographical barriers.201626802071
520980.8708Complete 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
303590.8706Molecular characterization of plasmids with antimicrobial resistant genes in avian isolates of Pasteurella multocida. The complete nucleotide sequences of two plasmids from avian isolates of Pasteurella multocida that caused outbreaks of fowl cholera in Taiwan were determined. The entire sequences of the two plasmids, designated as pJR1 and pJR2, were 6792 bp and 5252 bp. Sequence analysis showed that the plasmid pJR1 contained six major genes: the first gene (sulII) encoded a type II sulfonamide resistant dihydropteroate synthase, the second gene (tetG) encoded a tetracycline resistance protein, the third gene (catB2) encoded a chloramphenicol acetyltransferase, the fourth gene (rep) encoded a replication protein, and the fifth and sixth genes (mbeCy and deltambeAy) encoded proteins involved in the mobilization of plasmid. The plasmid pJR2 contained five major genes: the first gene (deltaintI1) encoded a truncated form of a type I integrase, the second gene (aadA1) encoded an aminoglycoside adenylyltransferase that confers resistance to streptomycin and spectinomycin, the third gene (blaP1) encoded a beta-lactamase that confers resistance to ampicillin and carbenicillin, and the fourth and fifth genes might encode proteins involved in the plasmid replication or segregation. Sequence comparisons showed that the antibiotic resistance genes found in pJR1 and pJR2 exhibited a high degree of sequence homology to the corresponding genes found in a great variety of gram-negative bacteria, including Escherichia coli, Salmonella enterica Typhimurium DT104, Psedomonas spp., P. multocida, Mannheimia spp., and Actinobacills pleuropneumoniae, which suggests that these resistance genes were disseminated in these bacteria. Although sulII and tetG genes were found previously in P. multocida or Mannheimia spp., this is the first report on the presence of catB2, aadA1, and blaP1 genes in bacteria of the family Pasturellaceae. Moreover, the aadA1 and blaP1 genes found in pJR2 were organized into an integron structure, which is a site-specific recombination system capable of capturing and mobilizing antibiotic resistance genes. This is also the first report on the presence of an integron in bacteria of the family Pasteurellaceae. The presence of a P. multocida integron might facilitate the spreading of antibiotic resistance genes between P. multocida and other gram-negative bacteria.200314708986
2997100.8706Genomic Characterization of Multidrug-Resistant Escherichia coli BH100 Sub-strains. The rapid emergence of multidrug-resistant (MDR) bacteria is a global health problem. Mobile genetic elements like conjugative plasmids, transposons, and integrons are the major players in spreading resistance genes in uropathogenic Escherichia coli (UPEC) pathotype. The E. coli BH100 strain was isolated from the urinary tract of a Brazilian woman in 1974. This strain presents two plasmids carrying MDR cassettes, pBH100, and pAp, with conjugative and mobilization properties, respectively. However, its transposable elements have not been characterized. In this study, we attempted to unravel the factors involved in the mobilization of virulence and drug-resistance genes by assessing genomic rearrangements in four BH100 sub-strains (BH100 MG2014, BH100 MG2017, BH100L MG2017, and BH100N MG2017). Therefore, the complete genomes of the BH100 sub-strains were achieved through Next Generation Sequencing and submitted to comparative genomic analyses. Our data shows recombination events between the two plasmids in the sub-strain BH100 MG2017 and between pBH100 and the chromosome in BH100L MG2017. In both cases, IS3 and IS21 elements were detected upstream of Tn21 family transposons associated with MDR genes at the recombined region. These results integrated with Genomic island analysis suggest pBH100 might be involved in the spreading of drug resistance through the formation of resistance islands. Regarding pathogenicity, our results reveal that BH100 strain is closely related to UPEC strains and contains many IS3 and IS21-transposase-enriched genomic islands associated with virulence. This study concludes that those IS elements are vital for the evolution and adaptation of BH100 strain.202033584554
3018110.8704The 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
3007120.8702Analysis 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
491130.8700Class II broad-spectrum mercury resistance transposons in Gram-positive bacteria from natural environments. We have studied the mechanisms of the horizontal dissemination of a broad-spectrum mercury resistance determinant among Bacillus and related species. This mer determinant was first described in Bacillus cereus RC607 from Boston Harbor, USA, and was then found in various Bacillus and related species in Japan, Russia and England. We have shown that the mer determinant can either be located at the chromosome, or on a plasmid in the Bacillus species, and is carried by class II mercury resistance transposons: Tn5084 from B. cereus RC607 and B. cereus VKM684 (ATCC10702) and Tn5085 from Exiguobacterium sp. TC38-2b. Tn5085 is identical in nucleotide sequence to TnMERI1, the only other known mer transposon from Bacillus species, but it does not contain an intron like TnMERI1. Tn5085 is functionally active in Escherichia coli. Tn5083, which we have isolated from B. megaterium MK64-1, contains an RC607-like mer determinant, that has lost some mercury resistance genes and possesses a merA gene which is a novel sequence variant that has not been previously described. Tn5083 and Tn5084 are recombinants, and are comprised of fragments from several transposons including Tn5085, and a relative of a putative transposon from B. firmus (which contains similar genes to the cadmium resistance operon of Staphylococcus aureus), as well as others. The sequence data showed evidence for recombination both between transposition genes and between mer determinants.200111446519
1494140.8698Characterization of a Novel Chromosomal Class C β-Lactamase, YOC-1, and Comparative Genomics Analysis of a Multidrug Resistance Plasmid in Yokenella regensburgei W13. Yokenella regensburgei, a member of the family Enterobacteriaceae, is usually isolated from environmental samples and generally resistant to early generations of cephalosporins. To characterize the resistance mechanism of Y. regensburgei strain W13 isolated from the sewage of an animal farm, whole genome sequencing, comparative genomics analysis and molecular cloning were performed. The results showed that a novel chromosomally encoded class C β-lactamase gene with the ability to confer resistance to β-lactam antibiotics, designated bla (YOC) (-) (1), was identified in the genome of Y. regensburgei W13. Kinetic analysis revealed that the β-lactamase YOC-1 has a broad spectrum of substrates, including penicillins, cefazolin, cefoxitin and cefotaxime. The two functionally characterized β-lactamases with the highest amino acid identities to YOC-1 were CDA-1 (71.69%) and CMY-2 (70.65%). The genetic context of the bla (YOC) (-) (1) -ampR-encoding region was unique compared with the sequences in the NCBI nucleotide database. The plasmid pRYW13-125 of Y. regensburgei W13 harbored 11 resistance genes (bla (OXA) (-) (10), bla (LAP) (-) (2), dfrA14, tetA, tetR, cmlA5, floR, sul2, ant(3″)-IIa, arr-2 and qnrS1) within an ∼34 kb multidrug resistance region; these genes were all related to mobile genetic elements. The multidrug resistance region of pYRW13-125 shared the highest identities with those of two plasmids from clinical Klebsiella pneumoniae isolates, indicating the possibility of horizontal transfer of these resistance genes between bacteria of various origins.202032973731
3021150.8698Sequencing and comparative analysis of IncP-1α antibiotic resistance plasmids reveal a highly conserved backbone and differences within accessory regions. Although IncP-1 plasmids are important for horizontal gene transfer among bacteria, in particular antibiotic resistance spread, so far only three plasmids from the subgroup IncP-1α have been completely sequenced. In this study we doubled this number. The three IncP-1α plasmids pB5, pB11 and pSP21 were isolated from bacteria of two different sewage treatment plants and sequenced by a combination of next-generation and capillary sequencing technologies. A comparative analysis including the previously analysed IncP-1α plasmids RK2, pTB11 and pBS228 revealed a highly conserved plasmid backbone (at least 99.9% DNA sequence identity) comprising 54 core genes. The accessory elements of the plasmid pB5 constitute a class 1 integron interrupting the parC gene and an IS6100 copy inserted into the integron. In addition, the tetracycline resistance genes tetAR and the ISTB11-like element are located between the klc operon and the trfA-ssb operon. Plasmid pB11 is loaded with a Tn5053-like mercury resistance transposon between the parCBA and parDE operons and contains tetAR that are identical to those identified in plasmid pB5 and the insertion sequence ISSP21. Plasmid pSP21 harbours an ISPa7 element in a Tn402 transposon including a class 1 integron between the partitioning genes parCBA and parDE. The IS-element ISSP21 (99.89% DNA sequence identity to ISSP21 from pB11), inserted downstream of the tetR gene and a copy of ISTB11 (identical to ISTB11 on pTB11) inserted between the genes pncA and pinR. On all three plasmids the accessory genes are almost always located between the backbone modules confirming the importance of the backbone functions for plasmid maintenance. The striking backbone conservation among the six completely sequenced IncP-1α plasmids is in contrast to the much higher diversity within the IncP-1β subgroup.201121115076
5871160.8698Plasmid-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
3017170.8695The 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
3015180.8695Genetic structure and biological properties of the first ancient multiresistance plasmid pKLH80 isolated from a permafrost bacterium. A novel multidrug-resistance plasmid, pKLH80, previously isolated from Psychrobacter maritimus MR29-12 found in ancient permafrost, was completely sequenced and analysed. In our previous studies, we focused on the pKLH80 plasmid region containing streptomycin and tetracycline resistance genes, and their mobilization with an upstream-located ISPpy1 insertion sequence (IS) element. Here, we present the complete sequence of pKLH80 and analysis of its backbone genetic structure, including previously unknown features of the plasmid's accessory region, notably a novel variant of the β-lactamase gene blaRTG-6. Plasmid pKLH80 was found to be a circular 14 835 bp molecule that has an overall G+C content of 40.3 mol% and encodes 20 putative ORFs. There are two distinctive functional modules within the plasmid backbone sequence: (i) the replication module consisting of repB and the oriV region; and (ii) the mobilization module consisting of mobA, mobC and oriT. All of the aforementioned genes share sequence identities with corresponding genes of different species of Psychrobacter. The plasmid accessory region contains antibiotic resistance genes and IS elements (ISPsma1 of the IS982 family, and ISPpy1 and ISAba14 of the IS3 family) found in environmental and clinical bacterial strains of different taxa. We revealed that the sequences flanking blaRTG-6 and closely related genes from clinical bacteria are nearly identical. This fact suggests that blaRTG-6 from the environmental strain of Psychrobacter is a progenitor of blaRTG genes of clinical bacteria. We also showed that pKLH80 can replicate in different strains of Acinetobacter and Psychrobacter genera. The roles of IS elements in the horizontal transfer of antibiotic resistance genes are examined and discussed.201425063046
5872190.8694Characterization 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