A Multidrug-resistant Monophasic Salmonella Typhimurium Co-harboring mcr-1, fosA3, bla CTX-M-14 in a Transferable IncHI2 Plasmid from a Healthy Catering Worker in China.
ABSTRACT: Background:Polymyxins are currently regarded as a possible last-resort therapy to eradicate multidrug-resistant (MDR) gram-negative bacteria. Meanwhile, the old antimicrobial agent fosfomycin has recently been reintroduced into clinical use for the treatment of extended-spectrum ?-lactamase (ESBL)-producing and carbapenem-resistant Enterobacteriaceae. This study investigated a multidrug-resistant Salmonella 4,,12:i:- strain from a food catering handler, which had the potential to act as a vehicle for transmitting MDR foodborne pathogens. Methods:A Salmonella 4,,12:i:- YZU1189 strain was isolated from the fecal sample of a food catering worker according to the standard protocol of the Salmonella detection method from World Health Organization in 2003. Serotyping of YZU1189 was performed according to the Kauffmann-White scheme. The antimicrobial resistance phenotype of the strain was determined by the agar dilution method according to the instruction from Clinical and Laboratory Standards Institute (CLSI). Plasmid conjugation was performed between the donor strain Salmonella 4,,12:i:- YZU1189 and the recipient strain Escherichia coli C600. The genetic locations of mcr-1, bla CTX-M-14 and fosA3 genes were determined by the whole genome sequence analysis. Results:Salmonella 4,,12:i:- YZU1189 was an ESBL-producing stain isolated from a healthy catering worker. The strain displayed resistance to aminoglycosides, beta-lactams, polymyxins, fosfomycins, phenicols, trimethoprims, sulfonamides, tetracyclines and fluoroquinolones. Whole genome sequence analysis and plasmid conjugation revealed that the strain had a transferable IncHI2 plasmid carrying the mcr-1, bla CTX-M-14 and fosA3 genes. Sequence homology analysis showed that this plasmid possessed high sequence similarity to previously reported mcr-1, bla CTX-M-14 and fosA3 positive plasmids in China. Conclusion:This study reported a the multidrug-resistant Salmonella 4,,12:i:- isolate harboring mcr-1, bla CTX-M-14 and fosA3 from human for the first time in China. The occurrence of mcr-1 and fosA3 genes in the transferable IncHI2 plasmid pYZU1189 from the ESBL-producing Salmonella 4,,12:i:- isolate showed a potential threat to public health. Great concern should be taken for the spread of multidrug-resistant ESBL-producing Salmonella isolates from food catering workers to consumers.
Project description:The aim of this study was to investigate the characteristics of a fosA3 carrying IncC-IncN plasmid from a multidrug-resistant Salmonella isolate HNK130. HNK130 was isolated from a chicken and identified as ST17 Salmonella enterica serovar Indiana and exhibited resistance to 13 antibiotics including the cephalosporins and fosfomycin. S1 nuclease pulsed-field gel electrophoresis and Southern blot assays revealed that HNK130 harbored only one ?180-kb plasmid carrying fosA3 and bla CTX-M-14, which was not transferable via conjugation. We further examined 107 Escherichia coli electro-transformants and identified 3 different plasmid variants, pT-HNK130-1 (69), pT-HNK130-2 (15), and pT-HNK130-3 (23), in which pT-HNK130-1 seemed to be the same as the plasmid harbored in HNK130. We completely sequenced an example of each of these variants, and all three variants were IncC-IncN multi-incompatible plasmid and showed a mosaic structure. The fosA3 gene was present in all three and bounded by IS26 elements in the same orientation (IS26-322bp-fosA3-1758bp-IS26) that could form a minicircle containing fosA3. The bla CTX-M-14 gene was located within an IS15DI-?IS15DI-iroN-IS903B-bla CTX-M-14 -?ISEcp1-IS26 structure separated from the fosA3 gene in pT-HNK130-1, but was adjacent to fosA3 in pT-HNK130-3 in an inverted orientation. Linear comparison of the three variants showed that pT-HNK130-2 and pT-HNK130-3 resulted from the sequence deletion and inversion of pT-HNK130-1. Stability tests demonstrated that pT-HNK130-1 and pT-HNK130-3 could be stably maintained in the transformants without antibiotic selection but pT-HNK130-2 was unstable. This is the first description of an IncC-IncN hybrid plasmid from an ST17 S. Indiana strain and indicates that this plasmid may further facilitate dissemination of fosfomycin and cephalosporin resistance in Salmonella.
Project description:Extended-spectrum-?-lactamase (ESBL)-producing Enterobacteriaceae are critical-priority pathogens that cause substantial fatalities. With the emergence of mobile mcr genes mediating resistance to colistin in Enterobacteriaceae, clinicians are now left with few therapeutic options. Eleven clinical Enterobacteriaceae strains with resistance to cephems and/or colistin were genomically analyzed to determine their resistomes, mobilomes, and evolutionary relationships to global strains. The global phylogenomics of mcr genes and mcr-9.1-bearing genomes were further analyzed. Ten isolates were ESBL positive. The isolates were multidrug resistant and phylogenetically related to global clones but distant from local strains. Multiple resistance genes, including bla CTX-M-15 bla TEM-1, and mcr-9.1, were found in single isolates; ISEc9, IS19, and Tn3 transposons bracketed bla CTX-M-15 and bla TEM-1 Common plasmid types included IncF, IncH, and ColRNAI. mcr-9 was of close sequence identity to mcr-3, mcr-5, mcr-7, mcr-8, and mcr-10. Genomes bearing mcr-9.1 clustered into six main phyletic groups (A to F), with those of this study belonging to clade B. Enterobacter species and Salmonella species are the main hosts of mcr-9.1 globally, although diverse promiscuous plasmids disseminate mcr-9.1 across different bacterial species. Emergence of mcr-9.1 in ESBL-producing Enterobacteriaceae in South Africa is worrying, due to the restricted therapeutic options. Intensive One Health molecular surveillance might discover other mcr alleles and inform infection management and antibiotic choices.IMPORTANCE Colistin is currently the last-resort antibiotic for difficult-to-treat bacterial infections. However, colistin resistance genes that can move from bacteria to bacteria have emerged, threatening the safe treatment of many bacterial infections. One of these genes, mcr-9.1, has emerged in South Africa in bacteria that are multidrug resistant, further limiting treatment options for clinicians. In this work, we show that this new gene is disseminating worldwide through Enterobacter and Salmonella species through multiple plasmids. This worrying observation requires urgent action to prevent further escalation of this gene in South Africa and Africa.
Project description:A 139,622-bp IncI1 ST71 conjugative plasmid pEC012 from an avian Escherichia coli D-ST117 strain was sequenced, which carried five IS26-bracketed resistance modules: IS26-fosA3-orf1-orf2-?orf3-IS26, IS26-fip-?ISEcp1-bla CTX-M-65-IS903D-iroN-IS26, IS26-?tnpR-bla TEM-1-rmtB-IS26, IS26-oqxAB-IS26, and IS26-floR-aac(3)-IV-IS26. The backbone of pEC012 was similar to that of several other IncI1 ST71 plasmids: pV408, pM105, and pC271, but these plasmids had different arrangements of multidrug resistance region. In addition, the novel ISEc57 element was identified, which is in the IS21 family. The stepwise emergence of multi-resistance regions demonstrated the accumulation of different resistance determinants through homologous recombination. To the best of our knowledge, this is the first study to identify a multidrug-resistant IncI1 ST71 plasmid carrying bla CTX-M-65, rmtB, fosA3, floR, and oqxAB in an avian E. coli ST117 strain.
Project description:Background:An ESBL, carbapenemase- and MCR-1-producing Escherichia coli ST648 strain was isolated from the urine sample of a patient in a Chinese tertiary hospital in 2016. Methods:The strain was fully sequenced by GridION X5 platform of Oxford Nanopore Technology. Results:The sequence analysis showed that the extended-spectrum ?-lactamases CTX-M-65 and OXA-1, the carbapenemase NDM-5, the MCR-1 were encoded, respectively, by three different resistance plasmids. The pE648CTX-M-65-carrying bla CTX-M-65 was a novel conjugative plasmid belonging to IncHI2 type; except for the bla CTX-M-65, it also carried resistance genes ble, floR, sul1, aph(4)-Ia, aac(3)-VI, aac(6')-II, bla OXA-1, catB, arr3 and tetA. Besides, an IncX4 plasmid pE648MCR-1-carrying mcr-1 and an IncX3 plasmid pE648NDM-5-carrying bla NDM-5 were also identified. Conclusion:The three transferable resistance plasmids coexisting in the E. coli ST648 isolate indicated the high risk to disseminate the extensively-drug-resistance among Enterobacteriaceae.
Project description:The aim of this study was to investigate the occurrence and genomic characteristics of extended-spectrum-?-lactamase-producing Escherichia coli (ESBL-EC) in fennec fox imported from Sudan to China. We screened 88 fecal samples from fennec fox for ESBL-EC, using cefotaxime- and meropenem-supplemented selective medium. Antimicrobial susceptibility testing was performed by the agar dilution method except for colistin and tigecycline; for colistin and tigecycline, testing was conducted by the broth microdilution method. ESBL-EC bacteria were sequenced, and their genomes were characterized. Plasmid conjugation, S1 nuclease pulsed-field gel electrophoresis (PFGE), and Southern blotting were performed for a MCR-1-producing isolate. The genetic environment of mcr-1 and ESBL genes was also investigated. A total of 29 ESBL-EC bacteria were isolated from 88 fennec fox (32.9%), while no carbapenemase producers were found. The most prevalent genotypes were the bla CTX-M-55 and bla CTX-M-14 genes, followed by bla CTX-M-15 and bla CTX-M-64 We detected nine sequence types among 29 ESBL-EC. Furthermore, the mcr-1 gene was detected in isolate EcFF273. Conjugation analysis confirmed that the mcr-1 gene was transferable. S1 PFGE, Southern blotting, and whole-genome sequencing revealed that mcr-1 and bla CTX-M-64 were both located on a 65-kb IncI2 plasmid. This study reports for the first time the occurrence of ESBL-EC in fennec fox. The high prevalence of ESBL producers and the occurrence of MCR-1 producer in fennec fox imported into China from Sudan are unexpected. In addition, it clearly demonstrated that commensal E. coli strains can be reservoirs of bla CTX-M and mcr-1, potentially contributing to the dissemination and transfer of such genes to pathogenic bacteria among fennec fox. Our results support the implication of fennec fox as a biological vector for ESBL-producing members of the Enterobacteriaceae family.IMPORTANCE The extended-spectrum-?-lactamase (ESBL)-producing members of the Enterobacteriaceae family are a global concern for both animal and human health. There is some information indicating a high prevalence of ESBL producers in food animals. Moreover, there have been an increasing number of reports on ESBL-producing strains resistant to the last-resort antibiotic colistin with the global dissemination of the plasmid-mediated mcr-1 gene, which is believed to have originated in animal breeding. However, little is known regarding the burden of ESBL-producing Enterobacteriaceae on wild animals. No data were available on the prevalence of antimicrobial resistance (AMR) among wild animals imported into China. This is the first study to investigate the microbiological and genomics surveillance investigation of ESBL colonization among fennec fox (Vulpes zerda) imported from Sudan to China, and we uncovered a high prevalence of ESBL-EC. Furthermore, the underlying mechanism of colistin resistance in an isolate that harbored mcr-1 was also investigated. Results of characterization and analysis of 29 ESBL-producing E. coli may have important implications on our understanding of the transmission dynamics of these bacteria. We emphasize the importance of improved multisectoral surveillance for colistin-resistant E. coli in this region.
Project description:Salmonella enterica serovars have been isolated from Colombian broilers and broiler meat. The aim of this study was to investigate the diversity of ESBL/pAmpC genes in extended-spectrum cephalosporin resistant Salmonella enterica and the phylogeny of ESBL/pAmpC-carrying Salmonella using Whole Genome Sequencing (WGS). A total of 260 cefotaxime resistant Salmonella isolates, obtained between 2008 and 2013 from broiler farms, slaughterhouses and retail, were included. Isolates were screened by PCR for ESBL/pAmpC genes. Gene and plasmid subtyping and strain Multi Locus Sequence Typing was performed in silico for a selection of fully sequenced isolates. Core-genome-based analyses were performed per ST encountered. bla CMY-2-like was carried in 168 isolates, 52 carried bla CTX-M-2 group, 7 bla SHV, 5 a combination of bla CMY-2-like-bla SHV and 3 a combination of bla CMY-2-like-bla CTX-M-2 group. In 25 isolates no ESBL/pAmpC genes that were screened for were found. WGS characterization of 36 selected strains showed plasmid-encoded bla CMY-2 in 21, bla CTX-M-165 in 11 and bla SHV-12 in 7 strains. These genes were mostly carried on IncI1/ST12, IncQ1, and IncI1/ST231 plasmids, respectively. Finally, 17 strains belonged to S. Heidelberg ST15, 16 to S. Paratyphi B variant Java ST28, 1 to S. Enteritidis ST11, 1 to S. Kentucky ST152 and 1 to S. Albany ST292. Phylogenetic comparisons with publicly available genomes showed separate clustering of Colombian S. Heidelberg and S. Paratyphi B var. Java. In conclusion, resistance to extended-spectrum cephalosporins in Salmonella from Colombian poultry is mainly encoded by bla CMY-2 and bla CTX-M-165 genes. These genes are mostly associated with IncI1/ST12 and IncQ1 plasmids, respectively. Evolutionary divergence is observed between Colombian S. Heidelberg and S. Paratyphi B var. Java and those from other countries.
Project description:Present study characterized five Escherichia coli co-expressing ESBL and MCR-1 recovered from food, food-producing animals, and companion animals in China. Antimicrobial susceptibility tests, conjugation experiments, and plasmid typing were performed. Whole genome sequencing (WGS) was undertaken for all five isolates using either PacBio RS II or Illumina HiSeq 2500 platforms. The cefotaxime and colistin resistance encoded by bla CTX-M and mcr-1 genes, respectively, was transferable by conjugation either together or separately for all five strains. Interestingly, the ESBL and mcr-1 genes could be co-selected by cefotaxime, while the colistin only selected the mcr-1-carrying plasmids during the conjugation experiments. Five E. coli sequence types (ST88, ST93, ST602, ST162, and ST457) were detected. Although diverse plasmid profiles were identified, IncI2, IncFIB, and IncFII plasmid types were predominant. These five clonally unrelated isolates harbored the mcr-1 gene located on similar plasmid backbones, which showed high nucleotide similarity to plasmid pHNSHP45. The mcr-1 gene can be co-transmitted with bla CTX-M genes through IncI2 plasmids with or without ISApl1 in our study. Characterization of these co-existence ESBL and mcr-1 isolates extends our understanding on the dissemination of these resistance markers among bacteria of diverse origins.
Project description:Dissemination of extended-spectrum-cephalosporin (ESC)-resistant Salmonella, especially extended-spectrum-?-lactamase (ESBL)-producing Salmonella, is a concern worldwide. Here, we assessed Salmonella carriage by food workers in Japan to clarify the prevalence of ESC-resistant Salmonella harboring bla CTX-M We then characterized the genetic features, such as transposable elements, of bla CTX-M-harboring plasmids using whole-genome sequencing. A total of 145,220 stool samples were collected from food workers, including cooks and servers from several restaurants, as well as food factory workers, from January to October 2017. Isolated salmonellae were subjected to antimicrobial susceptibility testing (disk diffusion method), and whole-genome sequencing was performed for Salmonella strains harboring bla CTX-M Overall, 164 Salmonella isolates (0.113%) were recovered from 164 samples, from which we estimated that at least 0.113% (95% confidence interval [CI]: 0.096 to 0.132%) of food workers may carry Salmonella Based on this estimation, 3,473 (95% CI = 2,962 to 4,047) individuals among the 3,075,330 Japanese food workers are likely to carry Salmonella Of the 158 culturable isolates, seven showed resistance to ESCs: three isolates harbored bla CMY-2 and produced AmpC ?-lactamase, while four ESBL-producing isolates harbored bla CTX-M-14 (n?=?1, Salmonella enterica serovar Senftenberg) or bla CTX-M-15 (n?=?3, S. enterica serovar Haardt). bla CTX-M-15 was chromosomally located in the S Haardt isolates, which also contained ISEcp1, while the S Senftenberg isolate contained an IncFIA(HI1)/IncHI1A/IncHI1B(R27) hybrid plasmid carrying bla CTX-M-14 along with ISEcp1 This study indicates that food workers may be a reservoir of ESBL-producing Salmonella and associated genes. Thus, these workers may contribute to the spread of bla CTX-M via plasmids or mobile genetic elements such as ISEcp1 IMPORTANCE Antimicrobial-resistant Salmonella bacteria arise in farm environments through imprudent use of antimicrobials. Subsequently, these antimicrobial-resistant strains, such as extended-spectrum-?-lactamase (ESBL)-producing Salmonella, may be transmitted to humans via food animal-derived products. Here, we examined Salmonella carriage among food handlers in Japan. Overall, 164 of 145,220 fecal samples (0.113%) were positive for Salmonella Among the 158 tested isolates, four were identified as ESBL-producing isolates carrying ESBL determinants bla CTX-M-15 or bla CTX-M-14 In all cases, the genes coexisted with ISEcp1, regardless of whether they were located on the chromosome or on a plasmid. Our findings suggest that food workers may be a reservoir of ESBL-producing strains and could contribute to the spread of resistance genes from farm-derived Salmonella to other bacterial species present in the human gut.
Project description:Extended-spectrum ?-lactamase (ESBL)-producing Salmonella are one of the most important public health problems in developed countries. ESBL-producing Salmonella strains have been isolated from humans in Asian countries neighboring Japan, along with strains harboring the plasmid-mediated extended-spectrum cephalosporin (ESC)-resistance gene, ampC (pAmpC). However, only a few studies have investigated the prevalence of ESC-resistant Salmonella in chicken products in Japan, which are the main vehicle of Salmonella transmission. The aim of this study was to investigate the prevalence of ESBL-producing, pAmpC-harboring, or carbapenem-resistant Salmonella in chicken products in Japan. In total, 355 out of 779 (45.6%) chicken product samples collected from 1996-2010 contained Salmonella, resulting in 378 distinct isolates. Of these isolates, 373 were tested for resistance to ESCs, cephamycins, or carbapenems. Isolates that showed resistance to one or more of these antimicrobials were then examined by PCR and DNA sequence analysis for the presence of the bla(CMY), bla(CTX-M), bla(TEM), and bla(SHV) resistance genes. Thirty-five resistant isolates were detected, including 26 isolates that contained pAmpC (bla(CMY-2)), and nine ESBL-producing isolates harboring bla(CTX-M) (n = 4, consisting of two bla(CTX-M-2) and two bla(CTX-M-15 genes)), bla(TEM) (n = 4, consisting of one bla(TEM-20) and three bla(TEM-52) genes), and bla(SHV) (n = 1, bla(SHV-12)). All pAmpC-harboring and ESBL-producing Salmonella isolates were obtained from samples collected after 2005, and the percentage of resistant isolates increased significantly from 0% in 2004 to 27.9% in 2010 (P for trend = 0.006). This increase was caused in part by an increase in the number of Salmonella enterica subsp. enterica serovar Infantis strains harboring an approximately 280-kb plasmid containing bla(CMY-2) in proximity to ISEcp1. The dissemination of ESC-resistant Salmonella containing plasmid-mediated bla(CMY-2) in chicken products indicates the need for the development of continuous monitoring strategies in the interests of public health.
Project description:We report here the whole-genome sequence of the first extended-spectrum ?-lactamase (ESBL)-producing strain of Salmonella enterica subsp. enterica serovar Napoli, LC0541/17, isolated from the stools of an ambulatory pediatric patient in northern Italy. The strain was of sequence type 474 (ST474) and possessed a 90-kb IncI1 ST49 plasmid carrying the bla CTX-M-1 ESBL gene.