Ping Shen

ST11, the dominant clone of KPC-producing Klebsiella pneumoniae in China

OBJECTIVESnKlebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae has spread rapidly in China. In this study, we aimed to investigate the molecular epidemiology of KPC-producing K. pneumoniae isolates in China.nnnMETHODSnNinety-five carbapenem-resistant K. pneumoniae isolates from 13 hospitals in nine cities covering five provinces in China were analysed. Antibiotic susceptibility was determined by the Etest. Multilocus sequence typing (MLST) and PFGE were used for epidemiological analysis. The genetic structure around bla(KPC) and the encoding genes of extended-spectrum β-lactamases and plasmid-mediated AmpC enzymes were detected by PCR and sequencing. Plasmids were analysed by transformation, restriction and Southern blot.nnnRESULTSnAll isolates harboured the bla(KPC-2) gene. Seven sequence types (STs) were obtained. The dominant clone was ST11 (61/95), which was identified in isolates from Zhejiang province (four hospitals in Hangzhou and one hospital in Ningbo), Jiangsu province (one hospital in Nanjing) and Anhui province (one hospital in Hefei). Isolates with ST11 showed >80% similarity in PFGE patterns. Plasmids from 14 selected transformants, their original isolates representing different STs and/or regions, had a diversity of HindIII restriction maps.nnnCONCLUSIONSnThe dominant clone of KPC-producing K. pneumoniae in China is ST11, which is closely related to ST258, which has been reported worldwide.

Plasmid-mediated KPC-2 in a Klebsiella pneumoniae isolate from China.

ABSTRACT A carbapenem-resistant isolate of Klebsiella pneumoniae producing class A carbapenemase KPC-2 was identified in Zhejiang, China. The KPC-2 gene was located on an approximately 60-kb plasmid in a genetic environment partially different from that of blaKPC-2 in the isolates from the United States and Colombia.

Novel Genetic Environment of the Carbapenem-Hydrolyzing β-Lactamase KPC-2 among Enterobacteriaceae in China

ABSTRACT Thirty-nine blaKPC-producing isolates of the family Enterobacteriaceae with carbapenem resistance or reduced carbapenem susceptibility were obtained from inpatients from eight hospitals in six cities of three provinces in eastern China. The pulsed-field gel electrophoresis analysis of all 36 Klebsiella pneumoniae isolates revealed six major patterns. The resistant plasmids of most isolates were successfully transferred by conjugation and evaluated experimentally to be 40 to 180 kb in size. A 20.2-kb blaKPC-surrounding nucleotide sequence from plasmid pKP048 has been obtained and contains an integration structure of a Tn3-based transposon and partial Tn4401 segment, with the gene order Tn3-transposase, Tn3-resolvase, ISKpn8, the blaKPC-2 gene, and the ISKpn6-like element. The chimera of several transposon-associated elements indicated a novel genetic environment of the K. pneumoniae carbapenemase β-lactamase gene in isolates from China.

Epidemiology and characteristics of antimicrobial resistance in China

A comprehensive surveillance system for bacterial resistance in tertiary hospitals has been established in China that involves tertiary hospitals in distinct regions nationwide, enabling the collection of a large amount of antimicrobial surveillance data. Antimicrobial resistance in China has become a serious healthcare problem, with high resistance rates of most common bacteria to clinically important antimicrobial agents. Methicillin-resistant S. aureus, ESBL-producing Enterobacteriaceae and carbapenem-resistant Acinetobacter baumannii represent more than 50% of microbial isolates. Additionally, bacterial resistance to fluoroquinolones, macrolides and third-generation cephalosporins is of serious concern. The molecular epidemiology and resistance mechanisms of the antimicrobial strains in China exhibited regional specificity, as well as the influence of dissemination of international clonal complexes. The molecular characteristics of MRSA, ESBL- and carbapenemase-producing Enterobacteriaceae, and macrolide-resistant gram-positive Streptococci in China were significantly different from those in other countries and regions, while S. pneumoniae serotypes appear to have been affected by the global spread of prevalent clones in other parts of the world. Moreover, important antimicrobial resistant bacteria such as community-acquired-MRSA, multidrug-resistant P. aeruginosa and extensive-resistant A. baumannii, and the antimicrobial resistance in primary healthcare and outpatient setting should be intensely monitored and investigated in the future.

Complete nucleotide sequence of Klebsiella pneumoniae multidrug resistance plasmid pKP048, carrying blaKPC-2, blaDHA-1, qnrB4, and armA.

ABSTRACT The Klebsiella pneumoniae multidrug resistance plasmid pKP048 was completely sequenced. This plasmid carries several important resistance determinants, such as blaKPC-2, blaDHA-1, qnrB4, and armA, which confer resistance to carbapenems, cephalosporins, fluoroquinolones, and aminoglycosides, respectively. Analysis of the finished 151,188-bp sequence data revealed 163 putative genes, 108 of which were assigned functions such as replication, stable inheritance, antibiotic resistance, a mobile element, conjugal transfer, and a restriction-modification system, showing the strong phylogenetic mosaicism and plasticity of the plasmid.

Molecular epidemiology and mechanisms of carbapenem resistance in Pseudomonas aeruginosa isolates from Chinese hospitals

We investigated the molecular epidemiology and carbapenem resistance mechanisms of 258 non-duplicate carbapenem-resistant clinical isolates of Pseudomonas aeruginosa collected from 2006 to 2007 at 28 hospitals in China. Up to 88% of the carbapenem-resistant isolates were multidrug-resistant. Pulsed-field gel electrophoresis (PFGE) revealed that levels of intrahospital and interhospital dissemination of clones were low. To assess the mechanisms leading to resistance, all 258 carbapenem-resistant isolates were analysed for expression of the chromosomal beta-lactamase (AmpC), the porin important for entry of carbapenems (OprD) and an efflux system (MexAB-OprM) known to extrude some beta-lactams. Carbapenem resistance was driven mainly by mutational inactivation of OprD, accompanied or not by hyperexpression of AmpC or MexAB-OprM. Metallo-beta-lactamase genes were detected in 22 carbapenem-resistant isolates in China, belonging to eight pulsotypes. The bla(OXA-50) gene was detected among all of the carbapenem-resistant isolates, whereas the bla(GES-5) gene was detected in only one carbapenem-resistant isolate.

Complete nucleotide sequence of pKP96, a 67 850 bp multiresistance plasmid encoding qnrA1, aac(6′)-Ib-cr and blaCTX-M-24 from Klebsiella pneumoniae

OBJECTIVESnThe multiresistance plasmid pKP96 from Klebsiella pneumoniae was sequenced completely and analysed concerning its genetic environment and distributing of antimicrobial resistance genes.nnnMETHODSnThe complete sequence of the plasmid was determined using a whole-genome shotgun approach. MICs of 13 antimicrobial agents were determined using Etests. A conjugation experiment was performed in liquid medium.nnnRESULTSnpKP96 is a circularly closed 67 850 bp multiresistance plasmid with an IncN incompatibility group. Seventy putative genes were identified according to the annotation of the finished sequence. The backbone region of the plasmid, comprising the conjugal transfer and plasmid replication regions, showed 91% identity to the IncN plasmid R46. Several mobile elements were found to be inserted into pKP96 together with antimicrobial resistance genes, including qnrA1, aac(6)-Ib-cr and bla(CTX-M-24).nnnCONCLUSIONSnPlasmid pKP96 is a chimera that has acquired its multiple antimicrobial resistance determinants horizontally from different sources. It may have evolved from an ancestor plasmid similar to R46 through the stepwise events of integration or recombination.

Dissemination of a clone carrying a fosA3-harbouring plasmid mediates high fosfomycin resistance rate of KPC-producing Klebsiella pneumoniae in China

Fosfomycin has been proposed as an adjunct to other active agents for treating KPC-producing Klebsiella pneumoniae infections. This study aimed to investigate the prevalence of fosfomycin resistance and plasmid-mediated resistance determinants among KPC-producing K. pneumoniae isolates from clinical samples in China. In total, 278 KPC-producing and 80 extended-spectrum β-lactamase (ESBL)-producing (non-KPC-producing) clinical K. pneumoniae isolates were collected in 12 hospitals from 2010 to 2013. Fosfomycin susceptibility testing was carried out using the agar dilution method. Phylogenetic clonal patterns were revealed by pulsed-field gel electrophoresis (PFGE). Isolates were screened for plasmid-mediated fosfomycin resistance genes (fosA, fosA3 and fosC2) by PCR amplification. A plasmid was completely sequenced by next-generation sequencing. The fosfomycin resistance rate in KPC-producers (60.8%; 169/278) was significantly higher than in ESBL-producers (12.5%; 10/80). In addition, 94 KPC-producing isolates were positive for fosA3 and most of them were clonally related. A 23939-bp plasmid (pFOS18) co-harbouring fosA3 and bla(KPC-2) was completely sequenced, revealing that the fosA3 gene was flanked by two copies of IS26; however, bla(KPC-2) was located on a Tn3-Tn4401 integration structure. Although the fosA3 and blaKPC-2 genes are located on different transposon systems, they are able to spread together worldwide through plasmid transfer. Dissemination of the clone carrying the fosA3-harbouring plasmid mediates the high fosfomycin resistance rate of KPC-producing K. pneumoniae in China. Fosfomycin as an alternative option for treating infections caused by KPC-producing K. pneumoniae should not be recommended in hospitals in which fosfomycin-resistant clonal dissemination is emerging.

Molecular Epidemiology and Genetic Diversity of Fluoroquinolone-Resistant Escherichia coli Isolates from Patients with Community-Onset Infections in 30 Chinese County Hospitals

ABSTRACT The high frequency of fluoroquinolone resistance in Escherichia coli is a feature of clinical bacteriology in China, where the molecular epidemiology and genetic characteristics of this resistance in county hospitals remain unclear. A total of 590 nonduplicate E. coli isolates from 30 county hospitals located across seven Chinese regions were examined for plasmid-mediated quinolone resistance (PMQR) genes and mutations in quinolone resistance-determining regions (QRDRs). Multilocus sequence typing (MLST) and phylogenetic analysis of fluoroquinolone-resistant isolates were used to determine their genetic relatedness. The ciprofloxacin resistance rate of community-onset E. coli was 51.2%, and at least one PMQR gene was carried by 220 (37.3%) isolates. These included qnr (3.7%), aac(6′)-Ib-cr (19.7%), qepA (14.4%), and oqxAB (3.8%). Two novel oqxB mutants were identified and named oqxB20 and oqxB29. From 60 sequence types (STs) isolated, 5 novel STs (ST4499 to ST4503) were identified. ST1193 (7.9%) was the second most abundant ST among fluoroquinolone-resistant isolates (ST131 was the most common, with 14.6%), and this is the first report of it in China. This is also the first report of ST2115 and ST3014 isolates from human samples. Ciprofloxacin-resistant E. coli isolates fell mainly into phylogroups B2 and D. The rates of fluoroquinolone resistance and the prevalence of PMQR genes in community-onset E. coli isolates from Chinese county hospitals were high. The wide-ranging molecular epidemiology of E. coli isolates from scattered locations across China indicates that fluoroquinolone resistance evolved from different sources.

Identification of key taxa that favor intestinal colonization of Clostridium difficile in an adult Chinese population.

Fecal microbial transplantation provides a high curative rate for recurrent Clostridium difficile infection (CDI). However, limitations associated with FMT drive the need to identify key taxa for selective probiotic therapy for prevention, treatment and cure of human CDI. CDI-associated changes in gut microbiota were investigated in adult patients in the Western countries and among infant population in China. However, there has been no such study involving adult patients in China. Therefore, using high throughput sequencing of the 16S ribosomal RNA V3 region and real-time quantitative polymerase chain reaction, we identified CDI-associated key taxa by comparing the fecal microbiota composition of 15 adult patients with CDI with those of 18 individuals with C. difficile-negative nosocomial diarrhea (CDN) and 25 healthy control subjects. Reduced fecal bacterial diversity and dramatic shifts of intestinal microbial composition in CDI and CDN groups were observed compared with healthy controls. Putative butyrate-producing anaerobic bacteria were significantly depleted whereas endotoxin-producing opportunistic pathogens and lactate-producing phylotypes increased dramatically in patients with CDI compared with healthy controls. Further screening of specific microbes causing diarrheal diseases and resistance against CDI is necessary.