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149400.9862Characterization 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
200210.9840IncHI1 plasmids mediated the tet(X4) gene spread in Enterobacteriaceae in porcine. The tigecycline resistance gene tet(X4) was widespread in various bacteria. However, limited information about the plasmid harboring the tet(X4) gene spread among the different species is available. Here, we investigated the transmission mechanisms of the tet(X4) gene spread among bacteria in a pig farm. The tet(X4) positive Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae and Enterobacter hormaeche were identified in the same farm. The whole genome sequencing (WGS) analysis showed that the K. pneumoniae belonged to ST727 (n = 11) and ST3830 (n = 1), E. cloacae and E. hormaeche belonged to ST524 (n = 1) and ST1862 (n = 1). All tet(X4) genes were located on the IncHI1 plasmids that could be conjugatively transferred into the recipient E. coli C600 at 30°C. Moreover, a fusion plasmid was identified that the IncHI1 plasmid recombined with the IncN plasmid mediated by ISCR2 during the conjugation from strains B12L to C600 (pB12L-EC-1). The fusion plasmid also has been discovered in a K. pneumoniae (K1L) that could provide more opportunities to spread antimicrobial resistance genes. The tet(X4) plasmids in these bacteria are derived from the same plasmid with a similar structure. Moreover, all the IncHI1 plasmids harboring the tet(X4) gene in GenBank belonged to the pST17, the newly defined pMLST. The antimicrobial susceptibility testing was performed by broth microdilution method showing the transconjugants acquired the most antimicrobial resistance from the donor strains. Taken together, this report provides evidence that IncHI1/pST17 is an important carrier for the tet(X4) spread in Enterobacteriaceae species, and these transmission mechanisms may perform in the environment.202337065147
520920.9838Complete 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
302930.9837Antibiotic multiresistance plasmid pRSB101 isolated from a wastewater treatment plant is related to plasmids residing in phytopathogenic bacteria and carries eight different resistance determinants including a multidrug transport system. Ten different antibiotic resistance plasmids conferring high-level erythromycin resistance were isolated from an activated sludge bacterial community of a wastewater treatment plant by applying a transformation-based approach. One of these plasmids, designated pRSB101, mediates resistance to tetracycline, erythromycin, roxythromycin, sulfonamides, cephalosporins, spectinomycin, streptomycin, trimethoprim, nalidixic acid and low concentrations of norfloxacin. Plasmid pRSB101 was completely sequenced and annotated. Its size is 47 829 bp. Conserved synteny exists between the pRSB101 replication/partition (rep/par) module and the pXAC33-replicon from the phytopathogen Xanthomonas axonopodis pv. citri. The second pRSB101 backbone module encodes a three-Mob-protein type mobilization (mob) system with homology to that of IncQ-like plasmids. Plasmid pRSB101 is mobilizable with the help of the IncP-1alpha plasmid RP4 providing transfer functions in trans. A 20 kb resistance region on pRSB101 is located within an integron-containing Tn402-like transposon. The variable region of the class 1 integron carries the genes dhfr1 for a dihydrofolate reductase, aadA2 for a spectinomycin/streptomycin adenylyltransferase and bla(TLA-2) for a so far unknown Ambler class A extended spectrum beta-lactamase. The integron-specific 3'-segment (qacEDelta1-sul1-orf5Delta) is connected to a macrolide resistance operon consisting of the genes mph(A) (macrolide 2'-phosphotransferase I), mrx (hydrophobic protein of unknown function) and mphR(A) (regulatory protein). Finally, a putative mobile element with the tetracycline resistance genes tetA (tetracycline efflux pump) and tetR was identified upstream of the Tn402-specific transposase gene tniA. The second 'genetic load' region on pRSB101 harbours four distinct mobile genetic elements, another integron belonging to a new class and footprints of two more transposable elements. A tripartite multidrug (MDR) transporter consisting of an ATP-binding-cassette (ABC)-type ATPase and permease, and an efflux membrane fusion protein (MFP) of the RND-family is encoded between the replication/partition and the mobilization module. Homologues of the macrolide resistance genes mph(A), mrx and mphR(A) were detected on eight other erythromycin resistance-plasmids isolated from activated sludge bacteria. Plasmid pRSB101-like repA amplicons were also obtained from plasmid-DNA preparations of the final effluents of the wastewater treatment plant indicating that pRSB101-like plasmids are released with the final effluents into the environment.200415528650
149240.9837Characterization 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
200550.9837Chromosomal 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
304760.9834Formaldehyde-resistance in Enterobacteriaceae and Pseudomonas aeruginosa: identification of resistance genes by DNA-hybridization. A 4.1. Kb large DNA fragment of a E. coli plasmid pVU 3695, on which the genes for formaldehyde-resistance are located, was used as a DNA probe to identify bacteria that carry this segment among formaldehyde-resistant bacteria. It was shown by Southern Blot-, Dot Blot-, and Colony Blot- Hybridization studies that the DNA of all formaldehyde-resistant E. coli, Serratia marcescens, Enterobacter cloacae, Citrobacter freundii and Klebsiella pneumoniae strains tested hybridize with the DNA probe from E. coli. In contrast the E. coli DNA probe does not hybridize with the DNA from formaldehyde-resistant Pseudomonas aeruginosa strains.19911909132
149870.9832Resistance of Klebsiella pneumoniae Strains Carrying bla (NDM-1) Gene and the Genetic Environment of bla (NDM-1). OBJECTIVE: Regional dissemination is the major cause of the widespread prevalence of a plasmid-encoding NDM-1 enzyme. We investigated the drug resistance, joint efficiency, and gene environment of a Klebsiella pneumoniae strain carrying bla (NDM-1) gene. MATERIALS AND METHODS: Carbapenem-non-susceptible strains were analyzed using the VITEK 2 Compact. Strains carrying bla (NDM-1) were identified using polymerase chain reaction and sequencing. Antimicrobial susceptibility testing and plasmid conjugation experiments were then conducted. Strains carrying bla (NDM-1) were subjected to Southern blot analysis. After the gene mapping of bla (NDM-1), library construction, and sequencing, plasmids were subsequently spliced and genotyped using the software Glimmer 3.0, and then analyzed using Mauve software. RESULTS: Among 1735 carbapenem-non-susceptible strains, 54 strains of bla (NDM-1)-positive bacteria were identified, which consisted of 44 strains of K. pneumoniae, 8 strains of Acinetobacter baumannii and 2 strains of Escherichia coli. Strains carrying bla (NDM-1) had a resistance rate of more than 50% in most antibiotics. Plasmid conjugation between strains carrying bla (NDM-1) and E. coli strain J53 had a success rate of 50%. Southern blot analysis indicated that each strain had multiple plasmids containing bla (NDM-1). Among the five plasmids containing bla (NDM-1) in K. pneumoniae for sequencing, two plasmids with complete sequences were obtained. The findings were as follows: (i) The p11106 and p12 plasmids were highly similar to pNDM-BTR; (ii) the p11106 and p12 plasmids showed differences in the 20-30 kb region (orf00032-orf00043) from the other six plasmids; and (iii) bla (NDM-1) was located at orf00037, while ble was found at orf00038. Two tnpA genes were located in the upstream region, and orf00052 (tnpA) in the 36 kb region was in the downstream sequence. CONCLUSION: bla (NDM-1)-containing bacteria exhibit multidrug resistance, which rapidly spreads and is transferred through efficient plasmid conjugation; the multidrug resistance of these bacteria may be determined by analyzing their drug-resistant plasmids. The presence of ble and tnpA genes suggests a possible hypothesis that bla (NDM-1) originates from A. baumannii, which is retained in K. pneumoniae over a long period by transposition of mobile elements.202032425903
152980.9831Emergence and Characterization of a Novel IncP-6 Plasmid Harboring bla (KPC-2) and qnrS2 Genes in Aeromonas taiwanensis Isolates. The dissemination of Klebsiella pneumoniae carbapenemases (KPCs) among Gram-negative bacteria is an important threat to global health. However, KPC-producing bacteria from environmental samples are rarely reported. This study aimed to elucidate the underlying resistance mechanisms of three carbapenem-resistant Aeromonas taiwanensis isolates recovered from river sediment samples. Pulsed-field gel electrophoresis (PFGE) and whole genome sequencing (WGS) analysis indicated a close evolutionary relationship among A. taiwanensis isolates. S1-PFGE, Southern blot and conjugation assays confirmed the presence of bla (KPC-) (2) and qnrS2 genes on a non-conjugative plasmid in these isolates. Plasmid analysis further showed that pKPC-1713 is an IncP-6 plasmid with a length of 53,205 bp, which can be transformed into DH5α strain and mediated carbapenems and quinolones resistance. The plasmid backbone of p1713-KPC demonstrated 99% sequence identity to that of IncP-6-type plasmid pKPC-cd17 from Aeromonas spp. and IncP-6-type plasmid: 1 from Citrobacter freundii at 74% coverage. A 14,808 bp insertion sequence was observed between merT gene and hypothetical protein in p1713-KPC, which include the quinolone resistance qnrS2 gene. Emergence of plasmid-borned bla (KPC) and qnrS2 genes from A. taiwanensis isolates highlights their possible dissemination into the environment. Therefore, potential detection of such plasmids from clinical isolates should be closely monitored.201931572337
150590.9831New 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
1174100.9831Identification 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
3036110.9831Complete 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
2007120.9830Novel 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
2448130.9829Emerging coexistence of three PMQR genes on a multiple resistance plasmid with a new surrounding genetic structure of qnrS2 in E. coli in China. BACKGROUND: Quinolones are commonly used for treatment of infections by bacteria of the Enterobacteriaceae family. However, the rising resistance to quinolones worldwide poses a major clinical and public health risk. This study aimed to characterise a novel multiple resistance plasmid carrying three plasmid-mediated quinolone resistance genes in Escherichia coli clinical stain RJ749. METHODS: MICs of ceftriaxone, cefepime, ceftazidime, ciprofloxacin, and levofloxacin for RJ749 and transconjugant c749 were determined by the Etest method. Conjugation was performed using sodium azide-resistant E. coli J53 strain as a recipient. The quinolone resistance-determining regions of gyrA, gyrB, parC, and parE were PCR-amplified. RESULTS: RJ749 was highly resistant to quinolones, while c749 showed low-level resistance. S1-nuclease pulsed-field gel electrophoresis revealed that RJ749 and c749 both harboured a plasmid. PCR presented chromosomal mutation sites of the quinolone resistance-determining region, which mediated quinolone resistance. The c749 genome comprised a single plasmid, pRJ749, with a multiple resistance region, including three plasmid-mediated quinolone resistance (PMQR) genes (aac (6')-Ib-cr, qnrS2, and oqxAB) and ten acquired resistance genes. One of the genes, qnrS2, was shown for the first time to be flanked by two IS26s. Three IS26-mediated circular molecules carrying the PMQR genes were detected. CONCLUSIONS: We revealed the coexistence of three PMQR genes on a multiple resistance plasmid and a new surrounding genetic structure of qnrS2 flanked by IS26 elements. IS26 plays an important role in horizontal spread of quinolone resistance.202032293532
1247140.9829Antibiotic resistance determinants of multidrug-resistant Acinetobacter baumannii clinical isolates in Algeria. Antibiotic susceptibility testing was performed on 71 Acinetobacter baumannii clinical isolates, and presence of antibiotic resistance genes was screened for by PCR amplification and sequencing. Resistance rates were very high for aminoglycosides (22-80%), fluoroquinolones (>90%), and cephalosporins (>90%) but remained low for rifampin (2.8%) or null for colistin. Antibiotic resistance encoding genes detected were as follows: blaTEM-128 gene (74.6%), aph(3')-VI (50.7 %), aadA (63.4%), ant(2″)-I (14.1%), aac(3)-Ia (91.1%), aac(6')-Ib (4.2%), mutation Ser83Leu in gyrA (94.4%), double mutations Ser83Leu and Ser80Leu (or Ser84Leu) in gyrA and parC (69.0%), and mutation I581N in RRDR of the rpoB gene.201323688522
2451150.9828Acquisition and transfer of antibiotic resistance genes in association with conjugative plasmid or class 1 integrons of Acinetobacter baumannii. Conjugation is a type of horizontal gene transfer (HGT) that serves as the primary mechanism responsible for accelerating the spread of antibiotic resistance genes in Gram-negative bacteria. The present study aimed to elucidate the mechanisms underlying the conjugation-mediated gene transfer from the extensively drug-resistant Acinetobacter baumannii (XDR-AB) and New Delhi Metallo-beta-lactamase-1-producing Acinetobacter baumannii (NDM-AB) to environmental isolates of Acinetobacter spp. Conjugation experiments demonstrated that resistance to ticarcillin and kanamycin could be transferred from four donors to two sodium azide-resistant A. baumannii strains, namely, NU013R and NU015R. No transconjugants were detected on Mueller-Hinton Agar (MHA) plates containing tetracycline. Plasmids obtained from donors as well as successful transconjugants were characterized by PCR-based replicon typing and S1-nuclease pulsed-field gel electrophoresis (S1-PFGE). Detection of antibiotic resistance genes and integrase genes (int) was performed using PCR. Results revealed that the donor AB364 strain can transfer the blaOXA-23 and blaPER-1 genes to both recipients in association with int1. A 240-kb plasmid was successfully transferred from the donor AB364 to recipients. In addition, the aphA6 and blaPER-1 genes were co-transferred with the int1 gene from the donor strains AB352 and AB405. The transfer of a 220-kb plasmid from the donors to recipient was detected. The GR6 plasmid containing the kanamycin resistance gene (aphA6) was successfully transferred from the donor strain AB140 to both recipient strains. However, the blaNDM-1 and tet(B) genes were not detected in all transconjugants. Our study is the first to demonstrate successful in vitro conjugation, which indicated that XDR-AB contained combination mechanisms of the co-transfer of antimicrobial resistance elements with integron cassettes or with the plasmid group GR6. Thus, conjugation could be responsible for the emergence of new types of antibiotic-resistant strains.201830521623
3007160.9828Analysis 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
3040170.9827Similarity in the Structure of tetD-Carrying Mobile Genetic Elements in Bacterial Strains of Different Genera Isolated from Cultured Yellowtail. Structure analysis was performed on the antibiotic-resistance-gene region of conjugative plasmids of four fish farm bacteria.The kanamycin resistance gene, IS26, and tetracycline resistance gene (tetA(D)) were flanked by two IS26s in opposite orientation in Citrobacter sp. TA3 and TA6, and Alteromonas sp. TA55 from fish farm A. IS26-Inner was disrupted with ISRSB101. The chloramphenicol resistance gene, IS26 and tetA (D) were flanked by two IS26s in direct orientation in Salmonella sp. TC67 from farm C. Structures of tetA (D) and IS26 were identical among the four bacteria, but there was no insertion within the IS26-Inner of Salmonella sp. TC67. Horizontal gene transfer between the strains of two different genera in fish farm A was suggested by the structure homologies of mobile genetic elements and antibiotic resistance genes.201627667524
1528180.9827First Report of Coexistence of bla (SFO-1) and bla (NDM-1) β-Lactamase Genes as Well as Colistin Resistance Gene mcr-9 in a Transferrable Plasmid of a Clinical Isolate of Enterobacter hormaechei. Many antimicrobial resistance genes usually located on transferable plasmids are responsible for multiple antimicrobial resistance among multidrug-resistant (MDR) Gram-negative bacteria. The aim of this study is to characterize a carbapenemase-producing Enterobacter hormaechei 1575 isolate from the blood sample in a tertiary hospital in Wuhan, Hubei Province, China. Antimicrobial susceptibility test showed that 1575 was an MDR isolate. The whole genome sequencing (WGS) and comparative genomics were used to deeply analyze the molecular information of the 1575 and to explore the location and structure of antibiotic resistance genes. The three key resistance genes (bla (SFO-1), bla (NDM-1), and mcr-9) were verified by PCR, and the amplicons were subsequently sequenced. Moreover, the conjugation assay was also performed to determine the transferability of those resistance genes. Plasmid files were determined by the S1 nuclease pulsed-field gel electrophoresis (S1-PFGE). WGS revealed that p1575-1 plasmid was a conjugative plasmid that possessed the rare coexistence of bla (SFO-1), bla (NDM-1), and mcr-9 genes and complete conjugative systems. And p1575-1 belonged to the plasmid incompatibility group IncHI2 and multilocus sequence typing ST102. Meanwhile, the pMLST type of p1575-1 was IncHI2-ST1. Conjugation assay proved that the MDR p1575-1 plasmid could be transferred to other recipients. S1-PFGE confirmed the location of plasmid with molecular weight of 342,447 bp. All these three resistant genes were flanked by various mobile elements, indicating that the bla (SFO-1), bla (NDM-1), and mcr-9 could be transferred not only by the p1575-1 plasmid but also by these mobile elements. Taken together, we report for the first time the coexistence of bla (SFO-1), bla (NDM-1), and mcr-9 on a transferable plasmid in a MDR clinical isolate E. hormaechei, which indicates the possibility of horizontal transfer of antibiotic resistance genes.202134220761
2009190.9827Aminoglycoside resistance genes aph(2")-Ib and aac(6')-Im detected together in strains of both Escherichia coli and Enterococcus faecium. Escherichia coli SCH92111602 expresses an aminoglycoside resistance profile similar to that conferred by the aac(6')-Ie-aph(2")-Ia gene found in gram-positive cocci and was found to contain the aminoglycoside resistance genes aph(2")-Ib and aac(6')-Im (only 44 nucleotides apart). aph(2")-Ib had been reported previously in Enterococcus faecium SF11770. aac(6')-Im had not been detected previously in enterococci and was found to be present also 44 nucleotides downstream from aph(2")-Ib in E. faecium SF11770. aph(2")-Ib and aac(6')-Im are separate open reading frames, each with its own putative ribosome binding site, whereas aac(6')-Ie-aph(2")-Ia appears to be a fusion of two genes with just one start and one stop codon. The deduced AAC(6')-Im protein exhibits 56% identity and 80% similarity to the AAC(6')-Ie domain of the bifunctional enzyme AAC(6')-APH(2"). Our results document the existence of a member of the aph(2") family of genes in gram-negative bacteria and provide evidence suggesting the horizontal transfer of aph(2")-Ib and aac(6')-Im as a unit between gram-positive and gram-negative bacteria.200111557456