Xi Huang

Prevalence, risk factors, outcomes, and molecular epidemiology of mcr-1-positive Enterobacteriaceae in patients and healthy adults from China: an epidemiological and clinical study

BACKGROUND The mcr-1 gene confers transferable colistin resistance. mcr-1-positive Enterobacteriaceae (MCRPE) have attracted substantial medical, media, and political attention; however, so far studies have not addressed their clinical impact. Herein, we report the prevalence of MCRPE in human infections and carriage, clinical associations of mcr-1-positive Escherichia coli (MCRPEC) infection, and risk factors for MCRPEC carriage. METHODS We undertook this study at two hospitals in Zhejiang and Guangdong, China. We did a retrospective cross-sectional assessment of prevalence of MCRPE infection from isolates of Gram-negative bacteria collected at the hospitals from 2007 to 2015 (prevalence study). We did a retrospective case-control study of risk factors for infection and mortality after infection, using all MCRPEC from infection isolates and a random sample of mcr-1-negative E coli infections from the retrospective collection between 2012 and 2015 (infection study). We also did a prospective case-control study to assess risk factors for carriage of MCRPEC in rectal swabs from inpatients with MCRPEC and mcr-1 negative at the hospitals and collected between May and December, 2015, compared with mcr-1-negative isolates from rectal swabs of inpatients (colonisation study). Strains were analysed for antibiotic resistance, plasmid typing, and transfer analysis, and strain relatedness. FINDINGS We identified 21 621 non-duplicate isolates of Enterobacteriaceae, Acinetobacter spp, and Pseudomonas aeruginosa from 18 698 inpatients and 2923 healthy volunteers. Of 17 498 isolates associated with infection, mcr-1 was detected in 76 (1%) of 5332 E coli isolates, 13 (<1%) of 348 Klebsiella pneumoniae, one (<1%) of 890 Enterobacter cloacae, and one (1%) of 162 Enterobacter aerogenes. For the infection study, we included 76 mcr-1-positive clinical E coli isolates and 508 mcr-1-negative isolates. Overall, MCRPEC infection was associated with male sex (209 [41%] vs 47 [63%], adjusted p=0·011), immunosuppression (30 [6%] vs 11 [15%], adjusted p=0·011), and antibiotic use, particularly carbapenems (45 [9%] vs 18 [24%], adjusted p=0·002) and fluoroquinolones (95 [19%] vs 23 [30%], adjusted p=0·017), before hospital admission. For the colonisation study, we screened 2923 rectal swabs from healthy volunteers, of which 19 were MCRPEC, and 1200 rectal swabs from patients, of which 35 were MCRPEC. Antibiotic use before hospital admission (p<0·0001) was associated with MCRPEC carriage in 35 patients compared with 378 patients with mcr-1-negative E coli colonisation, whereas living next to a farm was associated with mcr-1-negative E coli colonisation (p=0·03, univariate test). mcr-1 could be transferred between bacteria at high frequencies (10-1 to 10-3), and plasmid types and MCRPEC multi-locus sequence types (MLSTs) were more variable in Guangdong than in Zhejiang and included the human pathogen ST131. MCRPEC also included 17 unreported ST clades. INTERPRETATION In 2017, colistin will be formally banned from animal feeds in China and switched to human therapy. Infection with MRCPEC is associated with sex, immunosuppression, and previous antibiotic exposure, while colonisation is also associated with antibiotic exposure. MLST and plasmid analysis shows that MCRPEC are diversely spread throughout China and pervasive in Chinese communities. FUNDING National Key Basic Research Program of China, National Natural Science Foundation of China/Zhejiang, National Key Research and Development Program, and MRC, UK.

Emergence of the Plasmid-Mediated mcr-1 Gene in Colistin-Resistant Enterobacter aerogenes and Enterobacter cloacae

The gene mcr-1 was reported as the first plasmid-mediated colistin resistance gene in Escherichia coli isolates from food animals, food, and patients in China ([1][1]). Since then, detection of mcr-1 -positive strains has been reported in Enterobacteriaceae worldwide ([2][2][–][3][4][4]). The

Possible Transmission of mcr-1–Harboring Escherichia coli between Companion Animals and Human

To the Editor: Plasmid-mediated, colistin-resistance mechanism gene mcr-1 was first identified in Escherichia coli isolates from food, food animals, and human patients in November 2015 (1). Reports on detection of mcr-1 in Enterobacteriaceae from humans and food animals soon followed from ≈12 countries (2–5). Here we report detection of mcr-1 in colistin-resistant E. coli isolated from companion animals and the possible transmission of mcr-1–harboring E. coli between companion animals and a person.

CTX-M-137, a hybrid of CTX-M-14-like and CTX-M-15-like β-lactamases identified in an Escherichia coli clinical isolate

OBJECTIVES To characterize a novel CTX-M chimera, CTX-M-137, from Escherichia coli clinical isolates in China. METHODS Isolates were collected from five hospitals between 22 February 2009 and 20 December 2011. Resistance genes were investigated by PCR. blaCTX-M-137 was cloned and purified for kinetic measurements. Conjugation experiments, S1-PFGE and Southern blotting were performed to study the plasmid harbouring blaCTX-M-137. The genetic environment of blaCTX-M-137 was determined by genomic cloning and sequencing. RESULTS A total of 247 cephalosporin-resistant E. coli were identified. blaCTX-M group genes were the most prevalent extended-spectrum β-lactamase (ESBL) genes, with 71 isolates harbouring blaCTX-M-1 group genes and 137 isolates harbouring blaCTX-M-9 group genes. A novel chimera of CTX-M-14-like and CTX-M-15-like ESBLs, designated CTX-M-137, was identified from a 60-year-old man with a urinary tract infection. The N-terminus of CTX-M-137 matched CTX-M-14 and the C-terminus matched CTX-M-15. CTX-M-137 conferred resistance to ceftazidime, cefotaxime and aztreonam. Purified CTX-M-137 showed good hydrolytic activity against ceftazidime and cefotaxime, and was inhibited by clavulanic acid. The blaCTX-M-137 was carried on an ∼83 kb IncI1 plasmid. blaCTX-M-137 was carried on a complete transposition unit ISEcp1-blaCTX-M-137-Δorf477 inserted into yagA, which is part of the IncI1 plasmid backbone. CONCLUSIONS We identified a novel CTX-M chimera, CTX-M-137, with a CTX-M-14-like N-terminus and a CTX-M-15-like C-terminus. Our findings suggest an ongoing diversification of CTX-M-type ESBLs through recombination events.

OprD mutations and inactivation in imipenem-resistant Pseudomonas aeruginosa isolates from China

To investigate the mechanisms involved in imipenem resistance of Pseudomonas aeruginosa in southern China, 61 imipenem-resistant P. aeruginosa clinical isolates were collected from 4 hospitals between October 2011 and June 2012. All isolates were resistant to imipenem, whereas 21.3% were susceptible or intermediate to meropenem. Variable degrees of resistance to other β-lactam and non-β-lactam antimicrobials were observed. PFGE revealed high-level of clonal diversity. Among the 61 isolates, 50 isolates had OprD loss by disrupted oprD mutations, including 43 with frameshift mutations of oprD and 7 with a premature stop codon by single point mutation. Six isolates were oprD-negative by PCR, suggestive of a major disruption of oprD genes. Five isolates had intact oprD but had reduced expression of oprD genes. In addition, only one isolate with disrupted oprD mutation by a premature stop codon was confirmed to be a metallo-β-lactamase producer (IMP-9). Our results show that the loss of OprD, as well as reduced expression of oprD and MBL production, were the predominant mechanisms of imipenem resistance in P. aeruginosa in southern China.

MCR-1-producing Klebsiella pneumoniae outbreak in China

In January, 2017, The Lancet Infectious Diseases published our finding that Enterobacteriaceae carrying mcr-1, a plasmid-mediated colistin resistance gene, are highly heterogenous in sequence type (ST) grouping and plasmid types indicating the diversity of mcr-1-carrying bacteria in China, and mcr-1-positive Enterobacteriaceae infections were associated with male sex, immunosuppression and antibiotics use before hospitalisation. Here we report a hospital outbreak of the MCR-1-producing Klebsiella pneumoniae. Six clinical isolates including one Escherichia coli and five K pneumoniae were identified from six patients with pneumonia admitted to a paediatric leukaemia ward at a hospital in Guangzhou, China, during 2 months between December, 2015, and January, 2016. These patients were treated with different combinations of vancomycin, imipenem, lincomycin, cefotaxime, and caspofungin acetate. Two of the six patients died from their infections. All six isolates were resistant to colistin, polymyxin B, cefotaxime, and gentamicin; and the five K pneumoniae isolates were additionally resistant to ceftazidime, cefepime, amikacin, fosfomycin, and ciprofloxacin. Various resistance genes were identified by PCR and sequencing in these isolates, including mcr-1, blaTEM-1, blaCTX-M-3, aac(3)-IIa, aph(3’)-Ia, fosA3 in E coli and mcr-1, blaTEM-1, blaSHV-11, blaCTX-M-3, aac(3)-IVa, aphA6, armA, fosA3, qnrB4, and qnrS1 in K pneumoniae (appendix). All six isolates were subjected to multilocus sequence typing (MLST) and pulsed field gel electrophoresis (PFGE). As a result, five mcr-1-positive K pneumonia isolates were identified as ST11, and the mcr-1-positive E coli from patient 1 belonged to ST156. Results of PFGE were consistent with MLST and indicated that the five K pneumonia were clonally related. Colistin resistance was successfully transferred to E coli C6 00 with high conjugation frequencies (101 to 102) in all isolates, suggesting that mcr-1 was located on transferable plasmids. The results of S1-PFGE and plasmid replicon typing for mcr-1-carrying plasmid in the transconjugants showed that mcr-1 was located at IncX4 plasmids of approximately 50 kb in size in all six isolates, which harboured blaTEM-1 and blaCTX-M-3 (appendix). In conclusion, we identified a hospital outbreak of an MCR-1-producing K pneumoniae ST11 strain among children with acute leukaemia. Our finding suggests that mcr-1 can spread in the hospital environment in the absence of colistin use.

Coproduction of MCR-1 and NDM-1 by Colistin-Resistant Escherichia coli Isolated from a Healthy Individual

The first transferable plasmid-mediated colistin resistance gene, mcr-1 was reported in Escherichia coli isolates from food animals, food, and patients in China and now has been reported worldwide ([1][1]). Furthermore, cocarriage of mcr-1 and bla NDM has been reported in E. coli and other members

NDM-1-Producing Citrobacter freundii, Escherichia coli, and Acinetobacter baumannii Identified from a Single Patient in China

ABSTRACT We identified New Delhi metallo-β-lactamase (NDM-1)-producing Citrobacter freundii GB032, Escherichia coli GB102, and Acinetobacter baumannii GB661 in urine and stool samples from a single patient in China. Plasmid profiling and Southern blotting indicated that blaNDM-1 from GB032 and that from GB102 were likely located on the same plasmid, while blaNDM-1 from GB661 was located on a very large (>400-kb) plasmid. This case underscores the broad host range of blaNDM-1 and its potential to spread between members of the family Enterobacteriaceae and A. baumannii.

Outbreak of PER-1 and diversity of β-lactamases among ceftazidime-resistant Pseudomonas aeruginosa clinical isolates.

A growing number of β-lactamases have been reported in Pseudomonas aeruginosa clinical isolates. The aim of this study was to investigate the diversity of β-lactamases in the collection of 51 ceftazidime-resistant P. aeruginosa clinical isolates in four hospitals of southern China. Among these isolates, variable degrees of resistance to other β-lactam and non-β-lactam agents were observed. Pulsed-field gel electrophoresis (PFGE) revealed a high degree of clonality with five main genotypes. Of the 51 isolates tested, 35 (68.6%) were identified as extended-spectrum β-lactamase (ESBL) producers, with 35 producing PER-1, 1 CTX-M-3, 7 CTX-M-15 and 1 CTX-M-14. Most (82.9%, 29/35) PER-1-producing isolates were collected from two hospitals between January and April in 2008 and belonged to the same PFGE pattern (pattern B) with similar antibiogram and β-lactamase profiles, which suggested an outbreak of this clone at the time. The prevalence of CTX-M-type ESBL (17.6%, 9/51) was unexpectedly high. One isolate was identified as producing VIM-2. Furthermore, we also reported an occurrence of a novel OXA-10 variant, OXA-246, in 14 P. aeruginosa isolates. In addition, AmpC overproduction was found to be the β-lactamase-mediated mechanism responsible for ceftazidime resistance in 6 isolates (11.8%). Our results revealed an overall diversity of β-lactamases and outbreak of a PER-1-producing clone among ceftazidime-resistant P. aeruginosa in southern China.

High Rates of Human Fecal Carriage of mcr-1–Positive Multidrug-Resistant Enterobacteriaceae Emerge in China in Association With Successful Plasmid Families

Background mcr-1-mediated colistin resistance in Enterobacteriaceae is concerning, as colistin is used in treating multidrug-resistant Enterobacteriaceae infections. We identified trends in human fecal mcr-1-positivity rates and colonization with mcr-1-positive, third-generation cephalosporin-resistant (3GC-R) Enterobacteriaceae in Guangzhou, China, and investigated the genetic contexts of mcr-1 in mcr-1-positive 3GC-R strains. Methods Fecal samples were collected from in-/out-patients submitting specimens to 3 hospitals (2011-2016). mcr-1 carriage trends were assessed using iterative sequential regression. A subset of mcr-1-positive isolates was sequenced (whole-genome sequencing [WGS], Illumina), and genetic contexts (flanking regions, plasmids) of mcr-1 were characterized. Results Of 8022 fecal samples collected, 497 (6.2%) were mcr-1 positive, and 182 (2.3%) harbored mcr-1-positive 3GC-R Enterobacteriaceae. We observed marked increases in mcr-1 (0% [April 2011] to 31% [March 2016]) and more recent (since January 2014; 0% [April 2011] to 15% [March 2016]) increases in human colonization with mcr-1-positive 3GC-R Enterobacteriaceae (P < .001). mcr-1-positive 3GC-R isolates were commonly multidrug resistant. WGS of mcr-1-positive 3GC-R isolates (70 Escherichia coli, 3 Klebsiella pneumoniae) demonstrated bacterial strain diversity; mcr-1 in association with common plasmid backbones (IncI, IncHI2/HI2A, IncX4) and sometimes in multiple plasmids; frequent mcr-1 chromosomal integration; and high mobility of the mcr-1-associated insertion sequence ISApl1. Sequence data were consistent with plasmid spread among animal/human reservoirs. Conclusions The high prevalence of mcr-1 in multidrug-resistant E. coli colonizing humans is a clinical threat; diverse genetic mechanisms (strains/plasmids/insertion sequences) have contributed to the dissemination of mcr-1, and will facilitate its persistence.