Dataset Information

Dissemination of NDM-1-Producing Enterobacteriaceae Mediated by the IncX3-Type Plasmid.

ABSTRACT: The emergence and spread of NDM-1-producing Enterobacteriaceae have resulted in a worldwide public health risk that has affected some provinces of China. China is an exceptionally large country, and there is a crucial need to investigate the epidemic of blaNDM-1-positive Enterobacteriaceae in our province. A total of 186 carbapenem-resistant Enterobacteriaceae isolates (CRE) were collected in a grade-3 hospital in Zhejiang province. Carbapenem-resistant genes, including blaKPC, blaIMP, blaVIM, blaOXA-48 and blaNDM-1 were screened and sequenced. Ninety isolates were identified as harboring the blaKPC-2 genes, and five blaNDM-1-positive isolates were uncovered. XbaI-PFGE revealed that three blaNDM-1-positive K. pneumoniae isolates belonged to two different clones. S1-PFGE and southern blot suggested that the blaNDM-1 genes were located on IncX3-type plasmids with two different sizes ranging from 33.3 to 54.7 kb (n=4) and 104.5 to 138.9 kb (n=1), respectively, all of which could easily transfer to Escherichia coli by conjugation and electrotransformation. The high-throughput sequencing of two plasmids was performed leading to the identification of a smaller 54-kb plasmid, which had high sequence similarity with a previously reported pCFNDM-CN, and a larger plasmid in which only a 7.8-kb sequence of a common gene environment around blaNDM-1 (blaNDM-1-trpF- dsbC-cutA1-groEL-?InsE,) was detected. PCR mapping and sequencing demonstrated that four smaller blaNDM-1 plasmids contained a common gene environment around blaNDM-1 (IS5-blaNDM-1-trpF- dsbC-cutA1-groEL). We monitored the CRE epidemic in our hospital and determined that KPC-2 carbapenemase was a major risk to patient health and the IncX3-type plasmid played a vital role in the spread of the blaNDM-1 gene among the CRE.

SUBMITTER: Yang Q

PROVIDER: S-EPMC4457825 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Dissemination of NDM-1-Producing Enterobacteriaceae Mediated by the IncX3-Type Plasmid.

Yang Qing Q Fang Lanfang L Fu Ying Y Du Xiaoxing X Shen Yuqin Y Yu Yunsong Y

PloS one 20150605 6

The emergence and spread of NDM-1-producing Enterobacteriaceae have resulted in a worldwide public health risk that has affected some provinces of China. China is an exceptionally large country, and there is a crucial need to investigate the epidemic of blaNDM-1-positive Enterobacteriaceae in our province. A total of 186 carbapenem-resistant Enterobacteriaceae isolates (CRE) were collected in a grade-3 hospital in Zhejiang province. Carbapenem-resistant genes, including blaKPC, blaIMP, blaVIM, b ...[more]

PMID: 26047502

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Project description:The continuous emergence of novel New Delhi metallo-β-lactamase-5 (NDM-5)-producing Enterobacteriaceae isolates is receiving more and more public attention. Twenty-two NDM-5-producing strains were identified from 146 carbapenemase-producing Enterobacteriaceae (CRE) strains isolated from pediatric patients between January and March 2017, indicating that the blaNDM-5 gene has spread to children. All 22 isolates, including 16 Klebsiella pneumoniae strains, four Klebsiella aerogenes strains, and two Escherichia coli strains, showed significantly high resistance to β-lactam antibiotics (except aztreonam) but remained susceptible to tigecycline and colistin. K. pneumoniae and K. aerogenes strains were respectively defined as homologous clonal isolates by pulsed-field gel electrophoresis (PFGE). Multilocus sequence typing (MLST) results confirmed the genetic relatedness with all K. pneumoniae strains belonging to sequence type (ST) 48. Two E. coli isolates (ST617 and ST1236) were considered genetically unrelated. Twenty-two blaNDM-5 plasmids were positive for the IncX3 amplicon and showed almost identical profiles after digestion with HindIII and EcoRI. Four representative strains (K. pneumoniae K725, K. aerogenes CR33, E. coli Z214, and E. coli Z244) were selected for further study. Plasmids harboring blaNDM-5 showed strong stability in both clinical isolates and transconjugants, without apparent plasmid loss after 100 serial generations. S1-PFGE followed by Southern blot analysis demonstrated that the blaNDM-5 gene was located on an ∼46-kb plasmid. Plasmid sequences of pNDM-K725, pNDM-CR33, and pNDM-Z214 were almost identical but were slightly different from that of pNDM-Z244. Compared with pNDM-Z244, ΔISAba125 and partial copies of IS3000 were missing. The genetic backgrounds of the blaNDM-5 gene in four strains were slightly different from that of the typical pNDM_MGR194. This study comprehensively characterized the horizontal gene transfer of the blaNDM-5 gene among different Enterobacteriaceae isolates in pediatric patients, and the IncX3-type plasmid was responsible for the spread.IMPORTANCE The emergence of CRE strains resistant to multiple antibiotics is considered a substantial threat to human health. Therefore, all the efforts to provide a detailed molecular transmission mechanism of specific drug resistance can contribute positively to prevent the further spread of multidrug-resistant bacteria. Although the new superbug harboring blaNDM-5 has been reported in many countries, it was mostly identified among E. coli strains, and the gene transfer mechanism has not been fully recognized and studied. In this work, we identified 22 blaNDM-5-positive strains in different species of Enterobacteriaceae, including 16 Klebsiella pneumoniae strains, four Klebsiella aerogenes strains, and two Escherichia coli strains, which indicated the horizontal gene transfer of blaNDM-5 among Enterobacteriaceae strains in pediatric patients. Moreover, blaNDM-5 was located on a 46-kb IncX3 plasmid, which is possibly responsible for this widespread horizontal gene transfer. The different genetic contexts of the blaNDM-5 gene indicated some minor evolutions of the plasmid, based on the complete sequences of the blaNDM-5 plasmids. These findings are of great significance to understand the transmission mechanism of drug resistance genes, develop anti-infection treatment, and take effective infection control measures.

Project description:NDM-5 carbapenemase was mainly identified in Escherichia coli, while the rapid transmission of bla NDM-5 among Enterobacteriaceae has raised serious public attention. This study identified 14 NDM-5-producing Klebsiella pneumoniae isolates from 107 carbapenem-resistant K. pneumoniae isolates, recovered from blood, urine, and normally sterile body fluids of pediatric patients from January 2016 to December 2018. All NDM-5-producing isolates were highly resistant to ?-lactams, while tigecycline and polymyxin B exhibited excellent antimicrobial activity. These 14 strains belonged to 9 different sequence types (STs) and displayed various pulsed-field gel electrophoresis (PFGE) patterns, suggesting that they were not clonally related. S1-PFGE followed by Southern blotting showed that the bla NDM-5 gene was located on an ?46-kb IncX3 plasmid in all strains. All bla NDM-5-carrying plasmids were successfully transferred into recipient E. coli J53. PCR-based sequencing demonstrated that all of the bla NDM-5-carrying plasmids shared highly similar backbones, with nucleotide sequence identity of >99%. Moreover, this plasmid displayed high sequence similarity to the previously reported epidemic IncX3 bla NDM-5-carrying plasmids, with dynamic changes observed only in bla NDM-5-surrounding elements. Interestingly, the IncX3 bla NDM-5-carrying plasmids showed strong stability in clinical isolates when cultured in antibiotic-free medium. However, after the conjugation inhibitor linoleic acid was added, a gradual increase in the level of IncX3 plasmid loss could be observed. Clinical isolates displayed 10% to 15% bla NDM-5-carrying plasmid loss after coculture with linoleic acid for 5?days. These results showed that the IncX3 plasmid facilitated the dissemination of bla NDM-5 among multiclonal K. pneumoniae strains in children and that conjugal transfer contributed significantly to IncX3 plasmid stability within K. pneumoniae IMPORTANCE The emergence and spread of New Delhi metallo-?-lactamase (NDM)-producing Enterobacteriaceae have been a serious challenge to public health, and NDM-5 shows increased resistance to carbapenems compared with other variants. NDM-5 has been identified mostly in E. coli but has rarely been described in K. pneumoniae and other Enterobacteriaceae isolates. Here, we present the dissemination of highly similar 46-kb IncX3 bla NDM-5-carrying plasmids among multiclonal K. pneumoniae strains in children, highlighting the horizontal gene transfer of bla NDM-5 among K. pneumoniae strains via the IncX3 plasmid. Moreover, the IncX3 bla NDM-5-carrying plasmids displayed strong stability in clinical strains when cultured in antibiotic-free medium, and the plasmid maintenance was attributed partly to conjugal transfer. Plasmid conjugation is mediated by the type IV secretion system (T4SS), and T4SS is conserved among all epidemic IncX3 bla NDM-5-carrying plasmids. Therefore, combining conjugation inhibition and promotion of plasmid loss would be an effective strategy to limit the conjugation-assisted persistence of IncX3 bla NDM-5-carrying plasmids.

Project description:The emergence of New Delhi metallo-?-lactamase (NDM) in common enterobacterial species is a major concern for healthcare. Early reports have revealed that the spread of NDM involved diverse and heterogeneous plasmids. Recently, the involvement of a rare, IncX3 subtype plasmid has been increasingly recognized. Here, we studied the prevalence of IncX plasmid subtypes in 198 carbapenem-resistant Enterobacteriaceae, originating from a territory-wide active surveillance in Hong Kong in 2016. The complete sequences and biological features of the bla NDM-carrying plasmids were investigated. A total of 62 NDM-type, 21 OXA-48 type, 14 IMP-type, 8 KPC-type, 4 IMI-type producers, and 89 non-carbapenemase-producers were tested for presence of IncX subtypes. IncX3 (n = 60) was the most common subtype, followed by IncX4 (n = 6) and IncX1 (n = 2). The prevalence of IncX3 subtype in isolates producing NDM, other carbapenemase types and non-carbapenemase producers were 75.8, 21.3, and 3.4%, respectively (P < 0.001). An IncX3 plasmid (size ?50 kb) was confirmed to carry bla NDM in 47 isolates of different enterobacterial species. Thirteen IncX3 plasmids originating from six healthcare regions in Hong Kong were completely sequenced. The results showed that the IncX3 plasmids carrying bla NDM share a high degree of sequence identity with a previously reported plasmid, pNDM-HN380 (GenBank accession JX104760), over the backbone and genetic load regions. A blast search further revealed the occurrence of identical or nearly identical IncX3 plasmids carrying bla NDM in other part of China, Korea, Myanmar, India, Oman, Kuwait, Italy, and Canada. Two IncX3 carrying bla NDM were investigated further. Conjugation experiments demonstrated that the IncX3 plasmids could be efficiently transferred to multiple enterobacterial species at frequencies that are comparable or higher than the epidemic IncFII plasmid carrying bla CTX-M (pHK01). In addition, efficient transfer of the NDM plasmids occurred over a range of temperatures. In conclusion, this study demonstrated the important role played by IncX3 in the dissemination of NDM and the occurrence of pNDM-HN380-like plasmids in geographically widespread areas. The high mobility of IncX3 plasmid across different enterobacterial species highlights the ability of this plasmid replicon to be an important vehicle in worldwide dissemination of NDM.

Project description:BackgroundThe emergence and spread of New Delhi metallo-β-lactamase-producing Enterobacteriaceae has been a serious challenge to manage in the clinic due to its rapid dissemination of multi-drug resistance worldwide. As one main type of carbapenemases, New Delhi metallo-β-lactamase (NDM)is able to confer resistance to almost all β-lactams, including carbapenems, in Enterobacteriaceae. Recently, New Delhi metallo-β-lactamase-5 attracted extensive attention because of increased resistance to carbapenems and widespread dissemination. However, the dissemination mechanism of blaNDM-5 gene remains unclear.MethodsA total of 224 carbapenem-resistant Enterobacteriaceae isolates (CRE) were collected from different hospitals in Zhejiang province. NDM-5-positive isolates were identified and subjected to genotyping, susceptibility testing, and clinical data analysis. We established the genetic location of blaNDM-5 with southern blot hybridisation, and analysed plasmids containing blaNDM-5 with filter mating and DNA sequencing.ResultsEleven New Delhi metallo-β-lactamase-5 (NDM-5)-producing strains were identified, including 9 Escherichia coli strains, 1 Klebsiella pneumoniae strain, and 1 Citrobacter freundii strain. No epidemiological links for E. coli isolates were identified by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). S1-PFGE and southern blot suggested that the blaNDM-5 gene was located on a 46-kb IncX3-type plasmid in all isolates. Nine of the 11 isolates (81.8%) tested could successfully transfer their carbapenem-resistant phenotype to E. coli strain C600. Moreover, sequence analysis further showed that this plasmid possessed high sequence similarity to most of previously reported blaNDM-5-habouring plasmids in China.ConclusionThe present data in this study showed the IncX3 type plasmid played an important role in the dissemination of blaNDM-5 in Enterobacteriaceae. In addition, to the best of our knowledge, this report is the first to isolate both E. coli and C. freundii strains carrying blaNDM-5 from one single patient, which further indicated the possibility of blaNDM-5 transmission among diverse species. Close surveillance is urgently needed to monitor the further dissemination of NDM-5-producing isolates.

Dataset Information

Dissemination of NDM-1-Producing Enterobacteriaceae Mediated by the IncX3-Type Plasmid.

Publications

Dissemination of NDM-1-Producing Enterobacteriaceae Mediated by the IncX3-Type Plasmid.

Similar Datasets

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