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83400.9868Molecular diversity of class 2 integrons in antibiotic-resistant gram-negative bacteria found in wastewater environments in China. The molecular architecture of class 2 integrons among gram-negative bacteria from wastewater environments was investigated in Jinan, China. Out of the 391 antibiotic-resistant bacteria found, 38 isolates harboring class 2 integrons encoding potentially transferrable genes that could confer antibiotic resistance were found. These isolates were classified into 19 REP-PCR types. These strains were identified using 16S rRNA gene sequencing and found to be as follows: Proteus mirabilis (16), Escherichia coli (7), Providencia spp. (7), Proteus spp. (2), P. vulgaris (3), Shigella sp. (1), Citrobacter freundii (1), and Acinetobacter sp. (1). Their class 2 integron cassette arrays were amplified and then either analyzed using PCR-RFLP or sequenced. The typical array dfrA1-sat2-aadA1 was detected in 27 isolates. Six atypical arrays were observed, including three kinds of novel arrangements (linF2(∆attC1)-dfrA1(∆attC2)-aadA1-orf441 or linF2(∆attC1)-dfrA1(∆attC2)-aadA1, dfrA1-catB2-sat2-aadA1, and estX(Vr)-sat2-aadA1) and a hybrid with the 3'CS of class 1 integrons (dfrA1-sat2-aadA1-qacH), and dfrA1-sat1. Twenty-four isolates were also found to carry class 1 integrons with 10 types of gene cassette arrays. Several non-integron-associated antibiotic resistance genes were found, and their transferability was investigated. Results showed that water sources in the Jinan region harbored a diverse community of both typical and atypical class 2 integrons, raising concerns about the overuse of antibiotics and their careless disposal into the environment.201323264021
149410.9864Characterization 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
200720.9863Novel ISCR1-linked resistance genes found in multidrug-resistant Gram-negative bacteria in southern China. Non-duplicate multidrug-resistant (MDR) Gram-negative bacteria (n=1329) isolated from southern China between January 2008 and December 2009 were investigated for the presence of ISCR1 as well as characterisation of ISCR1-linked resistance genes. Of 433 ISCR1-positive strains, 151 appeared to carry ISCR1-linked resistance genes. Seven different ISCR1-linked resistance gene arrays were identified by restriction fragment length polymorphism (RFLP) and DNA sequencing analysis. Many of these arrays are reported in some species for the first time. A total of 12 genes, including a novel ABC transporter (GenBank accession no. GU944725), qnrA1, qnrB2, qnrB6, bla(DHA-1), ampR, bla(CTX-M-9), bla(PER-1), insB, sapA-like peptide transport periplasmic protein, putative glutathione S-transferase and short-chain dehydrogenase/reductase, were detected. This study was the first to employ PCR-RFLP using HinfI and RsaI to analyse ISCR1-linked genes. ISCR1 was widely disseminated among MDR Gram-negative bacteria and was in close association with quinolone resistance and β-lactamase genes (class A and class C) in southern China.201222890194
123530.9862Characterization of integrons and antimicrobial resistance genes in clinical isolates of Gram-negative bacteria from Palestinian hospitals. Sixty Gram-negative bacterial isolates were collected from Palestinian hospitals in 2006. Thirty-two (53.3%) isolates showed multidrug resistance phenotypes. PCR and DNA sequencing were used to characterize integrons and antimicrobial resistance genes. PCR screening showed that 19 (31.7%) and five (8.3%) isolates were positive for class 1 and class 2 integrons, respectively. DNA-sequencing results for the captured antimicrobial resistance gene cassettes within class 1 integrons identified the following genes: dihydrofolate reductases, dfrA1, dfrA5, dfrA7, dfrA12, dfrA17 and dfrA25; aminoglycoside adenyltransferases, aadA1, aadA2, aadA5, aadA12 and aadB; aminoglycoside acetyltransferase, aac(6')-Ib; and chloramphenicol resistance gene, cmlA1. ESBL were identified in 25 (41.7%) isolates. The identified ESBL were bla(CTX-M-15), bla(CTX-M-56), bla(OXA-1), bla(SHV-1), bla(SHV-12), bla(SHV-32) and bla(TEM-1) genes. Moreover, we characterized the plasmid-mediated quinolone resistance genes, aac(6')-Ib-cr and qnrB2, which were detected in seven (11.7%) and two (3.3%) isolates, respectively. In this study various types of antibiotic resistance genes have been identified in Gram-negative bacteria from Palestinian hospitals, many of which are reported in the Middle East area for the first time.200919903259
117440.9861Identification of plasmid-mediated quinolone resistance qnr genes in multidrug-resistant Gram-negative bacteria from hospital wastewaters and receiving waters in the Jinan area, China. We investigated the prevalence of plasmid-mediated quinolone resistance (PMQR) qnr genes by the polymerase chain reaction (PCR) in antibiotic-resistant bacteria isolates collected from aquatic environments in Jinan during 2 years (2008.3-2009.11). Genes were identified to variant level by PCR restriction fragment length polymorphism analysis or sequencing. qnrA1, qnrB2, qnrB4, qnrB6, qnrB9, qnrS1, and the new qnrB variant qnrB26 were detected in 31 strains from six genera (Klebsiella spp., Escherichia coli, Enterobacter spp., Proteus spp., Shigella spp., and Citrobacter spp.), four of which contained double qnr genes. Other PMQR genes, aac(6')-Ib-cr and qepA, were found in 12 (38.7%) and 5 (16.1%) of 31 isolates, respectively; while qepA was found in Shigella spp. for the first time. Eight types of β-lactamase genes and eight other types of resistance genes were also present in the 31 qnr-positive isolates. The detection rate for five β-lactamase genes (blaTEM, blaCTX, ampR, blaDHA, and blaSHV) was >45%. Class 1 integrons and complex class 1 integrons were prevalent in these strains, which contained 15 different gene cassette arrays and 5 different insertion sequence common region 1 (ISCR1)-mediated downstream structures. qnrA1, qnrB2, and qnrB6 were present in three ISCR1-mediated downstream structures: qnrA1-ampR, sapA-like-qnrB2, and sdr-qnrB6. We also analyzed the horizontal transferability of PMQR genes and other resistance determinants. The qnr genes and some integrons and resistance genes from 18 (58.1%) of the 31 qnr-positive strains could be transferred to E. coli J53 Azi(R) or E. coli DH5α recipient strains using conjugation or transformation methods. The results showed that a high number of qnr genes were associated with other resistance genes in aquatic environments in Jinan. This suggests that we should avoid over-using antibiotics and monitor aquatic environments to control the spread of antibiotic resistance genes.201323844849
150550.9860New insights on mcr-1-harboring plasmids from human clinical Escherichia coli isolates. Mobile colistin resistance (mcr) genes were described recently in Gram-negative bacteria including carbapenem-resistant Enterobacterales. There are ten mcr genes described in different Gram-negative bacteria, however, Escherichia coli harboring mcr-1 gene is by far the most frequent combination. In Argentina, mcr-1 gene was characterized only on plasmids belonging to IncI2 group. The aim of this work was to get new insights of mcr-1-harboring plasmids from E. coli. Eight E. coli isolates from a larger collection of 192 clinical E. coli isolates carrying the mcr-1 gene were sequenced using next generation technologies. Three isolates belonged to ST131 high-risk clone, and five to single ST, ST38, ST46, ST226, ST224, and ST405. Eight diverse mcr-1-harboring plasmids were analyzed: IncI2 (1), IncX4 (3), IncHI2/2A (3) and a hybrid IncFIA/HI1A/HI1B (1) plasmid. Plasmids belonging to the IncI2 (n = 1) and IncX4 (n = 3) groups showed high similarity with previously described plasmids. Two IncHI2/HI2A plasmids, showed high identity between them, while the third, showed several differences including additional resistance genes like tet(A) and floR. One IncFIA/H1A/H1B hybrid plasmid was characterized, highly similar to pSRC27-H, a prototype plasmid lacking mcr genes. mcr-1.5 variant was found in four plasmids with three different Inc groups: IncI2, IncHI2/HI2A and the hybrid FIA/HI1A/HI1B plasmid. mcr-1.5 variant is almost exclusively described in our country and with a high frequency. In addition, six E. coli isolates carried three allelic variants codifying for CTX-M-type extended-spectrum-β-lactamases: blaCTX-M-2 (3), blaCTX-M-65 (2), and blaCTX-M-14 (1). It is the first description of mcr-1 harboring plasmids different to IncI2 group in our country. These results represents new insights about mcr-1 harboring plasmids recovered from E. coli human samples from Argentina, showing different plasmid backbones and resistance gene combinations.202438408071
138860.9860Snapshot Study of Whole Genome Sequences of Escherichia coli from Healthy Companion Animals, Livestock, Wildlife, Humans and Food in Italy. Animals, humans and food are all interconnected sources of antimicrobial resistance (AMR), allowing extensive and rapid exchange of AMR bacteria and genes. Whole genome sequencing (WGS) was used to characterize 279 Escherichia coli isolates obtained from animals (livestock, companion animals, wildlife), food and humans in Italy. E. coli predominantly belonged to commensal phylogroups B1 (46.6%) and A (29%) using the original Clermont criteria. One hundred and thirty-six sequence types (STs) were observed, including different pandemic (ST69, ST95, ST131) and emerging (ST10, ST23, ST58, ST117, ST405, ST648) extraintestinal pathogenic Escherichia coli (ExPEC) lineages. Eight antimicrobial resistance genes (ARGs) and five chromosomal mutations conferring resistance to highest priority critically important antimicrobials (HP-CIAs) were identified (qnrS1, qnrB19, mcr-1, bla(CTX-M1,15,55), bla(CMY-2), gyrA/parC/parE, ampC and pmrB). Twenty-two class 1 integron arrangements in 34 strains were characterized and 11 ARGs were designated as intI1 related gene cassettes (aadA1, aadA2, aadA5, aad23, ant2_Ia, dfrA1, dfrA7, dfrA14, dfrA12, dfrA17, cmlA1). Notably, most intI1 positive strains belonged to rabbit (38%) and poultry (24%) sources. Three rabbit samples carried the mcr-1 colistin resistance gene in association with IS6 family insertion elements. Poultry meat harbored some of the most prominent ExPEC STs, including ST131, ST69, ST10, ST23, and ST117. Wildlife showed a high average number of virulence-associated genes (VAGs) (mean = 10), mostly associated with an ExPEC pathotype and some predominant ExPEC lineages (ST23, ST117, ST648) were identified.202033172096
137570.9860Characterization of integrons and their cassettes in Escherichia coli and Salmonella isolates from poultry in Korea. Ninety-nine Escherichia coli and 33 Salmonella isolates were assessed for antimicrobial susceptibility (disc diffusion test). Sulfonamide and tetracycline resistance genes were identified through PCR, and class 1 and class 2 integrons with resistance gene cassettes were identified with PCR followed by sequencing. Salmonella (63.6%) and E. coli (85.8%) isolates were multidrug resistant (resistance to 3 or more antimicrobials), and the highest incidences of resistance were observed for tetracycline, nalidixic acid, and sulfamethoxazole. The sul1, sul2, tetA, and tetB resistance determinant genes were predominant in E. coli, whereas only sul2 and tetA were identified in Salmonella isolates. In the E. coli isolates, 54 (54.5%) class 1 integrons, 6 (6.1%) class 2 integrons, and 5 (5.1%) class 1 and class 2 integrons together were detected, whereas only 3 (9.1%) integrons were found in the Salmonella serovars. Around 87% of the integrons in E. coli harbored resistance gene cassettes conferring resistance to streptomycin/spectinomycin (aadA, aminoglycoside resistance gene), trimethoprim (dfrA, dihydrofolate reductase gene), streptothricin [sat1 and sat2 (streptothricin acetyltransferase), and estX (putative esterases)]. The most common gene cassettes were aadA1+dfrA1 and dfrA1+sat2+aadA1 in class 1 and class 2 integrons, respectively. Other cassettes including aadA5+dfrA7, dfrA12+aadA2, aadA2+aadA1+dfrA12, and aadA5+aadA2/dfrA7 were also identified. Among the Salmonella serovars, Salmonella Malmoe harbored aadA1+dfrA1 and dfrA12+sat2+aadA1 genes. The aadA1, aadA2, sat2, and dfrA1 had wide variation in similarity among themselves and from previously reported genes worldwide. The diverse gene cassettes could be responsible for the prominent resistance profiles observed and a potential source for dissemination of antimicrobial resistance determinants to other bacteria.201324135609
149280.9859Characterization of the tet(M)-bearing transposon Tn7125 of Escherichia coli strain A13 isolated from an intensive pig farm located in Henan province, China. BACKGROUND: Transposons carrying tet(M) in Gram-positive bacteria have been reported extensively, while there is a paucity of data on the transmission characteristics of tet(M) in Gram-negative bacteria. Therefore, the aim of this study was to investigate the genetic characteristics of the tet(M)-bearing transposon Tn7125, and to clarify the transmission mechanism of the plasmids pTA13-1 and pTA13-3 in Escherichia coli strain A13. METHODS: Plasmids from strain A13 and a corresponding transconjugant were determined by whole genome sequencing and analyzed using bioinformatics tools. The plasmids pTA13-1 and pTA13-3 of the transconjugant TA13 were characterized by S1-pulse-field gel electrophoresis, Southern hybridization, stability experiments, and direct competition assays. RESULTS: The conjugated IncF2:A6:B20 plasmid pTA13-1 co-transferred with the 41-kb plasmid pTA13-3, which carried no resistance genes; plasmid pTA13-2, which harbored the replication initiator PO111; and the IncX4 plasmid pTA13-4, which harbored the antibiotic resistance gene mcr-1. The novel IS26-bracked composite transposon Tn7125 was located on plasmid pTA13-1, which mainly consists of three resistance modules: IS26-ctp-lp-tet(M)-hp-IS406tnp, qac-aadA1-cmlA1-aadA2-DUF1010-dfrA12, and ∆ISVSa3-VirD-floR-LysR-ISVSa3. The plasmid pTA13-1 was highly stable in E. coli strain J53 with no fitness cost to the host or disadvantage in growth competition. CONCLUSION: Evolution of co-integrated transposons, such as Tn7125, may convey antibiotic resistance to a wide spectrum of hosts via the plasmids pTA13-1 and pTA13-3, which acts as an adaptable and mobile multidrug resistance reservoir to accelerate dissemination of other genes by co-selection, thereby posing a potentially serious barrier to clinical treatment regimens.202540639501
263690.9859Characterisation of plasmids harbouring extended-spectrum cephalosporin resistance genes in Escherichia coli from French rivers. Antimicrobial resistance is a "One Health" issue that requires improved knowledge of the presence and abundance of resistant bacteria in the environment. Extended-spectrum cephalosporins (ESCs) are critically important antibiotics (CIAs), and resistance to these CIAs is often encoded by beta-lactamase genes borne on conjugative plasmids. We thus decided to characterise 21 plasmids of ESC-resistant Escherichia coli randomly selected from isolates previously obtained from river water collected in a rural area in western France. The plasmids encoding ESC resistance were sequenced to investigate the diversity of the genes encoding ESC resistance and their genetic context. Sequences revealed that eleven IncI1 pMLST3 plasmids carried the bla(CTX-M-1) and sul2 genes, and some of them also had the tet(A), aadA5 or dfrA17 genes. The bla(CTX-M-1) gene was also detected on an IncN plasmid. Five plasmids obtained from four rivers contained bla(CTX-M-14), either on IncI1 or on IncFII plasmids. Two strains from two rivers contained bla(CTX-M-15) on IncN pMLST7 plasmids, with qnrS1 and dfrA14 genes. One plasmid contained the bla(CTX-M-55), a bla(TEM-1B)-like, and fosA genes. One plasmid contained the bla(CMY-2) gene. The diversity of the genes and plasmids of the resistant bacteria isolated from French rivers is probably related to the various animal and human origins of the isolated bacteria.202032273005
3008100.9859Sequence of conjugative plasmid pIP1206 mediating resistance to aminoglycosides by 16S rRNA methylation and to hydrophilic fluoroquinolones by efflux. Self-transferable IncFI plasmid pIP1206, isolated from an Escherichia coli clinical isolate, carries two new resistance determinants: qepA, which confers resistance to hydrophylic fluoroquinolones by efflux, and rmtB, which specifies a 16S rRNA methylase conferring high-level aminoglycoside resistance. Analysis of the 168,113-bp sequence (51% G+C) revealed that pIP1206 was composed of several subregions separated by copies of insertion sequences. Of 151 open reading frames, 56 (37%) were also present in pRSB107, isolated from a bacterium in a sewage treatment plant. pIP1206 contained four replication regions (RepFIA, RepFIB, and two partial RepFII regions) and a transfer region 91% identical with that of pAPEC-O1-ColBM, a plasmid isolated from an avian pathogenic E. coli. A putative oriT region was found upstream from the transfer region. The antibiotic resistance genes tet(A), catA1, bla(TEM-1), rmtB, and qepA were clustered in a 33.5-kb fragment delineated by two IS26 elements that also carried a class 1 integron, including the sulI, qacEDelta1, aad4, and dfrA17 genes and Tn10, Tn21, and Tn3-like transposons. The plasmid also possessed a raffinose operon, an arginine deiminase pathway, a putative iron acquisition gene cluster, an S-methylmethionine metabolism operon, two virulence-associated genes, and a type I DNA restriction-modification (R-M) system. Three toxin/antitoxin systems and the R-M system ensured stabilization of the plasmid in the host bacteria. These data suggest that the mosaic structure of pIP1206 could have resulted from recombination between pRSB107 and a pAPEC-O1-ColBM-like plasmid, combined with structural rearrangements associated with acquisition of additional DNA by recombination and of mobile genetic elements by transposition.200818458128
2005110.9856Chromosomal 16S Ribosomal RNA Methyltransferase RmtE1 in Escherichia coli Sequence Type 448. We identified rmtE1, an uncommon 16S ribosomal methyltransferase gene, in an aminoglycoside- and cephalosporin-resistant Escherichia coli sequence type 448 clinical strain co-harboring bla(CMY-2). Long-read sequencing revealed insertion of a 101,257-bp fragment carrying both resistance genes to the chromosome. Our findings underscore E. coli sequence type 448 as a potential high-risk multidrug-resistant clone.201728418308
5872120.9856Characterization 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
1535130.9856Complete Genome Sequencing of Acinetobacter baumannii AC1633 and Acinetobacter nosocomialis AC1530 Unveils a Large Multidrug-Resistant Plasmid Encoding the NDM-1 and OXA-58 Carbapenemases. Carbapenem-resistant Acinetobacter spp. are considered priority drug-resistant human-pathogenic bacteria. The genomes of two carbapenem-resistant Acinetobacter spp. clinical isolates obtained from the same tertiary hospital in Terengganu, Malaysia, namely, A. baumannii AC1633 and A. nosocomialis AC1530, were sequenced. Both isolates were found to harbor the carbapenemase genes bla(NDM-1) and bla(OXA-58) in a large (ca. 170 kb) plasmid designated pAC1633-1 and pAC1530, respectively, that also encodes genes that confer resistance to aminoglycosides, sulfonamides, and macrolides. The two plasmids were almost identical except for the insertion of ISAba11 and an IS4 family element in pAC1633-1, and ISAba11 along with relBE toxin-antitoxin genes flanked by inversely orientated pdif (XerC/XerD) recombination sites in pAC1530. The bla(NDM-1) gene was encoded in a Tn125 composite transposon structure flanked by ISAba125, whereas bla(OXA-58) was flanked by ISAba11 and ISAba3 downstream and a partial ISAba3 element upstream within a pdif module. The presence of conjugative genes in plasmids pAC1633-1/pAC1530 and their discovery in two distinct species of Acinetobacter from the same hospital are suggestive of conjugative transfer, but mating experiments failed to demonstrate transmissibility under standard laboratory conditions. Comparative sequence analysis strongly inferred that pAC1633-1/pAC1530 was derived from two separate plasmids in an IS1006-mediated recombination or transposition event. A. baumannii AC1633 also harbored three other plasmids designated pAC1633-2, pAC1633-3, and pAC1633-4. Both pAC1633-3 and pAC1633-4 are cryptic plasmids, whereas pAC1633-2 is a 12,651-bp plasmid of the GR8/GR23 Rep3-superfamily group that encodes the tetA(39) tetracycline resistance determinant in a pdif module.IMPORTANCE Bacteria of the genus Acinetobacter are important hospital-acquired pathogens, with carbapenem-resistant A. baumannii listed by the World Health Organization as the one of the top priority pathogens. Whole-genome sequencing of carbapenem-resistant A. baumannii AC1633 and A. nosocomialis AC1530, which were isolated from the main tertiary hospital in Terengganu, Malaysia, led to the discovery of a large, ca. 170-kb plasmid that harbored genes encoding the New Delhi metallo-β-lactamase-1 (NDM-1) and OXA-58 carbapenemases alongside genes that conferred resistance to aminoglycosides, macrolides, and sulfonamides. The plasmid was a patchwork of multiple mobile genetic elements and comparative sequence analysis indicated that it may have been derived from two separate plasmids through an IS1006-mediated recombination or transposition event. The presence of such a potentially transmissible plasmid encoding resistance to multiple antimicrobials warrants vigilance, as its spread to susceptible strains would lead to increasing incidences of antimicrobial resistance.202133504662
3009140.9856Identification of a novel conjugative plasmid carrying the multiresistance gene cfr in Proteus vulgaris isolated from swine origin in China. The multiresistance gene cfr has a broad host range encompassing both Gram-positive and Gram-negative bacteria, and can be located on the chromosomes or on plasmids. In this study, a novel conjugative plasmid carrying cfr, designated as pPvSC3, was characterized in a Proteus vulgaris strain isolated from swine in China. Plasmid pPvSC3 is 284,528 bp in size and harbors 10 other antimicrobial resistance genes, making it a novel plasmid that differs from all known plasmids due to its unique backbone and repA gene. BLAST analysis of the plasmid sequence shows no significant homology to any known plasmid backbone, but shows high level homology to Providencia rettgeri strain CCBH11880 Contig_9, a strain isolated from surgical wound in Brazil, 2014. There are two resistance-determining regions in pPvSC3, a cfr-containing region and a multidrug-resistant (MDR) region. The cfr-containing region is flanked by IS26, which could be looped out via IS26-mediated recombination. The MDR region harbors 10 antimicrobial resistance genes carried by various DNA segments that originated from various sources. Plasmid pPvSC3 could be successfully transferred to Escherichia coli by conjugation. In summary, we have characterized a novel conjugative plasmid pPvSC3 carrying the multiresistance gene cfr and 10 other antimicrobial resistance genes, and consider that this novel type of plasmid deserves attention.201931499097
2004150.9855Deciphering the Structural Diversity and Classification of the Mobile Tigecycline Resistance Gene tet(X)-Bearing Plasmidome among Bacteria. The emergence of novel plasmid-mediated resistance genes constitutes a great public concern. Recently, mobile tet(X) variants were reported in diverse pathogens from different sources. However, the diversity of tet(X)-bearing plasmids remains largely unknown. In this study, the phenotypes and genotypes of all the tet(X)-positive tigecycline-resistant strains isolated from a slaughterhouse in China were characterized by antimicrobial susceptibility testing, conjugation, pulsed-field gel electrophoresis with S1 nuclease (S1-PFGE), and PCR. The diversity and polymorphism of tet(X)-harboring strains and plasmidomes were investigated by whole-genome sequencing (WGS) and single-plasmid-molecule analysis. Seventy-four tet(X4)-harboring Escherichia coli strains and one tet(X6)-bearing Providencia rettgeri strain were identified. The tet(X4)-bearing elements in 27 strains could be transferred to the recipient strain via plasmids. All tet(X4)-bearing plasmids isolated in this study and 15 tet(X4)-bearing plasmids reported online were analyzed. tet(X4)-bearing plasmids ranged from 9 to 294 kb and were categorized as ColE2-like, IncQ, IncX1, IncA/C2, IncFII, IncFIB, and hybrid plasmids with different replicons. The core tet(X4)-bearing genetic contexts were divided into four major groups: ISCR2-tet(X4)-abh, △ISCR2-abh-tet(X4)-ISCR2, ISCR2-abh-tet(X4)-ISCR2-virD2-floR, and abh-tet(X4)-ISCR2-yheS-cat-zitR-ISCR2-virD2-floR Tandem repeats of tet(X4) were universally mediated by ISCR2 Different tet(X)-bearing strains existed in the same microbiota. Reorganization of tet(X4)-bearing multidrug resistance plasmids was found to be mediated by IS26 and other homologous regions. Finally, single-plasmid-molecule analysis captured the heterogenous state of tet(X4)-bearing plasmids. These findings significantly expand our knowledge of the tet(X)-bearing plasmidome among microbiotas, which establishes a baseline for investigating the structure and diversity of human, animal, and environmental tigecycline resistomes. Characterization of tet(X) genes among different microbiotas should be performed systematically to understand the evolution and ecology.IMPORTANCE Tigecycline is an expanded-spectrum tetracycline used as a last-resort antimicrobial for treating infections caused by superbugs such as carbapenemase-producing or colistin-resistant pathogens. Emergence of the plasmid-mediated mobile tigecycline resistance gene tet(X4) created a great public health concern. However, the diversity of tet(X4)-bearing plasmids and bacteria remains largely uninvestigated. To cover this knowledge gap, we comprehensively identified and characterized the tet(X)-bearing plasmidome in different sources using advanced sequencing technologies for the first time. The huge diversity of tet(X4)-bearing mobile elements demonstrates the high level of transmissibility of the tet(X4) gene among bacteria. It is crucial to enhance stringent surveillance of tet(X) genes in animal and human pathogens globally.202032345737
1189160.9855Detection of the carbapenemase gene bla(VIM-5) in members of the Pseudomonas putida group isolated from polluted Nigerian wetlands. There are increasing concerns about possible dissemination of clinically relevant antibiotic resistance genes, including genes encoding for carbapenemases in the environment. However, little is known about environmental distribution of antibiotic resistance in Africa. In this study, four polluted urban wetlands in Nigeria were investigated as potential reservoirs of carbapenem-resistant bacteria (CRB). CRB were isolated from the wetlands, characterized by Blue-Carba test, MIC determinations and whole genome sequencing (WGS). Nine of 65 bacterial isolates identified as members of the Pseudomonas putida group (P. plecoglossicida and P. guariconensis, respectively) harboured the metallo-beta-lactamase gene bla(VIM-5). WGS revealed the bla(VIM-5) in three novel Tn402-like class 1 integron structures containing the cassette arrays aadB|bla(VIM-5)|bla(PSE-1), aadB|bla(VIM-5)|aadB|bla(PSE-1), and bla(VIM-5)|aadB|tnpA|bla(PSE-1)|smr2|tnpA, respectively. Strains carrying the aadB|bla(VIM-5)|bla(PSE-1) cassette also carried an identical integron without bla(VIM-5). In addition(,) the strains harboured another Tn402-like class 1 integron carrying bcr2, several multidrug resistance efflux pumps, and at least one of ampC, aph(3")-lb, aph(6)-ld, tetB, tetC, tetG, floR, and macAB. This is the first report of a carbapenemase gene in bacteria from environmental sources in Nigeria and the first report of bla(VIM-5) in environmental bacteria isolates. This result underscores the role of the Nigerian environment as reservoir of bacteria carrying clinically relevant antibiotic resistance genes.201830310126
1509170.9855Characterization of plasmids harbouring qnrS1, qnrB2 and qnrB19 genes in Salmonella. OBJECTIVES: The aim of this study was to identify and characterize plasmids carrying qnrS1, qnrB2 and qnrB19 genes identified in Salmonella strains from The Netherlands. The identification of plasmids may help to follow the dissemination of these resistance genes in different countries and environments. METHODS: Plasmids from 33 qnr-positive Salmonella strains were transferred to Escherichia coli and analysed by restriction, Southern blot hybridization, PCR and sequencing of resistance determinants. They were also assigned to incompatibility groups by PCR-based replicon typing, including three additional PCR assays for the IncU, IncR and ColE groups. The collection included isolates from humans and one from chicken meat. RESULTS: Five IncN plasmids carrying qnrS1, qnrB2 and qnrB19 genes were identified in Salmonella enterica Bredeney, Typhimurium PT507, Kentucky and Saintpaul. qnrS1 genes were also located on three further plasmid types, belonging to the ColE (in Salmonella Corvallis and Anatum), IncR (in Salmonella Montevideo) and IncHI2 (in Salmonella Stanley) groups. CONCLUSIONS: Multiple events of mobilization, transposition and replicon fusion generate the complexity observed in qnr-positive isolates that are emerging worldwide. Despite the fact that the occurrence of qnr genes in bacteria from animals is scarcely reported, these genes are associated with genetic elements and located on plasmids that are recurrent in animal isolates.200919001452
3060180.9855Integron mobilization unit as a source of mobility of antibiotic resistance genes. Antibiotic resistance genes are spread mostly through plasmids, integrons (as a form of gene cassettes), and transposons in gram-negative bacteria. We describe here a novel genetic structure, named the integron mobilization unit (IMU), that has characteristics similar to those of miniature inverted transposable elements (MITEs). Two IMUs (288 bp each) were identified from a carbapenem-resistant Enterobacter cloacae isolate that formed a composite structure encompassing a defective class 1 integron containing the carbapenem resistance gene bla(GES-5). This beta-lactamase gene was located on a 7-kb IncQ-type plasmid named pCHE-A, which was sequenced completely. The plasmid pCHE-A was not self conjugative but was mobilizable, and it was successfully transferred from E. cloacae to Pseudomonas aeruginosa. The in silico analysis of the extremities of the IMU elements identified similarities with those of insertion sequence ISSod9 from Shewanella oneidensis MR-1. The mobilization of the IMU composite structure was accomplished by using the transposase activity of ISSod9 that was provided in trans. This is the first identification of MITE-type structures as a source of gene mobilization, implicating here a clinically relevant antibiotic resistance gene.200919332679
1087190.9854Characterization and Comparative Genomics Analysis of lncFII Multi-Resistance Plasmids Carrying bla (CTX) (-) (M) and Type1 Integrons From Escherichia coli. This research aimed to investigate the presence and transferability of the extended-spectrum β-lactamase resistance genes to identify the genetic context of multi-drug resistant (MDR) loci in two Escherichia coli plasmids from livestock and poultry breeding environment. MICs were determined by broth microdilution. A total of 137 E. coli resistant to extended-spectrum β-lactam antibiotics were screened for the presence of the ESBL genes by PCR. Only two E. coli out of 206 strains produced carbapenemases, including strain 11011 that produced enzyme A, and strain 417957 that produced enzyme B. The genes were bla (KPC) and bla (NDM) , respectively. The plasmids containing bla (CTX) (-) (M) were conjugatable, and the plasmids containing carbapenem resistance gene were not conjugatable. Six extended-spectrum β-lactamase resistance genes were detected in this research, including bla (TEM), bla (CTX) (-) (M), bla (SHV), bla (OAX) (-) (1), bla (KPC), and bla (NDM) , and the detection rates were 94.89% (130/137), 92.7% (127/137), 24.81% (34/137), 20.43% (28/137), 0.72% (1/137), and 0.72% (1/137), respectively. Two conjugative lncFII multi-resistance plasmids carrying bla (CTX) (-) (M), p11011-fosA and p417957-CTXM, were sequenced and analyzed. Both conjugative plasmids were larger than 100 kb and contained three accessory modules, including MDR region. The MDR region of the two plasmids contained many antibiotic resistance genes, including bla (CTX) (-) (M), mph (A), dfrA17, aadA5, sul1, etc. After transfer, both the transconjugants displayed elevated MICs of the respective antimicrobial agents. A large number of resistance genes clusters in specific regions may contribute to the MDR profile of the strains. The presence of mobile genetic elements at the boundaries can possibly facilitate transfer among Enterobacteriaceae through inter-replicon gene transfer. Our study provides beta-lactam resistance profile of bacteria, reveals the prevalence of β-lactamase resistance genes in livestock and poultry breeding environment in Zhejiang Province, and enriches the research on IncFII plasmids containing bla (CTX) (-) (M).202134867876