[Show abstract][Hide abstract] ABSTRACT: Multidrug-resistant and highly virulent Klebsiella pneumoniae isolates are emerging, but the clonal groups (CGs) corresponding to these high-risk strains have remained imprecisely defined. We aimed to identify K. pneumoniae CGs on the basis of genome-wide sequence variation and to provide a simple bioinformatics tool to extract virulence and resistance gene data from genomic data. We sequenced 48 K. pneumoniae isolates, mostly of serotypes K1 and K2, and compared the genomes with 119 publicly available genomes. A total of 694 highly conserved genes were included in a core-genome multilocus sequence typing scheme, and cluster analysis of the data enabled precise definition of globally distributed hypervirulent and multidrug-resistant CGs. In addition, we created a freely accessible database, BIGSdb-Kp, to enable rapid extraction of medically and epidemiologically relevant information from genomic sequences of K. pneumoniae. Although drug-resistant and virulent K. pneumoniae populations were largely nonoverlapping, isolates with combined virulence and resistance features were detected.
[Show abstract][Hide abstract] ABSTRACT: Hypervirulent Klebsiella pneumoniae isolates of capsular serotype K2 (hvKP-K2) that cause community-acquired invasive infections represent several unrelated clones, which all belong to phylogenetic group KpI. These clones were recognized using multilocus sequence typing (MLST) and genomic analyses, but no rapid method currently exists to differentiate them. In this work, a multiplex PCR assay was developed for identification of three hvKP-K2 groups: (i) sequence type (ST) 86; (ii) ST380 and ST679 (i.e. clonal group 380); and (iii) ST65 and ST375. A specific genetic marker, Kp50233, allowing to distinguish K. pneumoniae sensu stricto (corresponding to phylogroup KpI) from closely related species, was included in the assay. This PCR will be useful to better define the epidemiology and clinical features of emerging virulent K. pneumoniae clones.
[Show abstract][Hide abstract] ABSTRACT: In Klebsiella pneumoniae, overexpression of the AcrAB efflux pump and the more recently described OqxAB efflux pump has been linked to an antibiotic cross-resistance phenotype, but the mechanisms of regulation are largely unknown. Moreover, while AcrAB has been shown to participate in K. pneumoniae virulence, the contribution of OqxAB has not yet been assessed.
Journal of Antimicrobial Chemotherapy 09/2014; · 5.34 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The prevalence of Escherichia coli sequence type 131 (ST131) and its subclone H30 was assessed among a collection of 490 E. coli isolated in 2013 in a French university hospital. The prevalence of ST131 was 4% among bloodstream isolates (regardless of antimicrobial resistance) and 17.2% among extended-spectrum β-lactamase (ESBL)-producing isolates. Although a much lower prevalence of ST131 was found among bloodstream E. coli isolates compared with other countries, a large predominance of H30 subclone within ST131 was confirmed. It was also confirmed that, among ESBL-producing E. coli, ST131 isolates were more frequently resistant to amoxicillin/clavulanic acid, ceftazidime, fluoroquinolones and aminoglycosides than non-ST131 isolates.
International Journal of Antimicrobial Agents 09/2014; · 4.42 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In 2008, a previously unknown Escherichia coli clonal group, sequence type 131 (ST131), was identified on three continents. Today, ST131 is the predominant E. coli lineage among extraintestinal pathogenic E. coli (ExPEC) isolates worldwide. Retrospective studies have suggested that it may originally have risen to prominence as early as 2003. Unlike other classical group B2 ExPEC isolates, ST131 isolates are commonly reported to produce extended-spectrum β-lactamases, such as CTX-M-15, and almost all are resistant to fluoroquinolones. Moreover, ST131 E. coli isolates are considered to be truly pathogenic, due to the spectrum of infections they cause in both community and hospital settings and the large number of virulence-associated genes they contain. ST131 isolates therefore seem to contradict the widely held view that high levels of antimicrobial resistance are necessarily associated with a fitness cost leading to a decrease in pathogenesis. Six years after the first description of E. coli ST131, this review outlines the principal traits of ST131 clonal group isolates, based on the growing body of published data, and highlights what is currently known and what we need to find out to provide public health authorities with better information to help combat ST131.
[Show abstract][Hide abstract] ABSTRACT: We characterized 53 OXA-48-producing Klebsiella pneumoniae (OXA-48-Kp) isolated between 2011 and 2013 in 21 French hospitals. All the isolates were genotyped using MLST and PFGE and the population structure of the species was determined by a nucleotide-based analysis of the entire K. pneumoniae MLST database. Most of the OXA-48-Kp isolates also produced CTX-M-15 and remained susceptible to imipenem and meropenem. The isolates were distributed into 20 STs, of which five were dominant (ST15, ST101, ST147, ST395, and ST405). All the OXA-48-Kp clustered in the major clade of K. pneumoniae KpI. This article is protected by copyright. All rights reserved.
Clinical Microbiology and Infection 06/2014; · 4.58 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Determining the prevalence of children in day-care centres (DCCs) carrying faecal extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae and molecularly characterizing those belonging to the Escherichia coli species.
Stools were collected from children's diapers (January-April 2012) in randomly chosen DCCs and plated onto ChromID(™) ESBL. Colonies growing on this medium were identified by the Vitek 2(®) system and tested for antibiotic susceptibility and for ESBL production by the double-disc synergy test. ESBL genotypes were determined as well as phylogenetic groups, ERIC-2 (enterobacterial repetitive intergenic consensus) PCR profiles and sequence types (STs) for the E. coli isolates. Serotypes, virotypes, fimH alleles, ESBL-carrying plasmids and PFGE patterns were determined for the ST131 E. coli isolates.
Among 419 children from 25 participating DCCs, 1 was colonized by CTX-M-15-producing Klebsiella pneumoniae and 27 (6.4%) by E. coli, which all produced CTX-M enzymes [CTX-M-15 (37%), CTX-M-1 (26%), CTX-M-14 (22%), CTX-M-27 (11%) and CTX-M-22 (4%)]. The 27 E. coli isolates, 55.5% belonging to group B2, displayed 20 ERIC-2 PCR profiles and 16 STs. The ST131 E. coli isolates were dominant (44%), displayed serotypes O25b:H4 and O16:H5, fimH alleles 30 and 41 and virotypes A and C. According to the PFGE patterns, one strain of E. coli ST131 producing a CTX-M-15 enzyme carried by an IncF F2:A1:B- plasmid had spread within one DCC.
This study shows a notable prevalence (6.4%) of DCC children with faecal CTX-M-producing E. coli isolates comprising a high proportion of E. coli ST131 isolates, suggesting that these children might be a reservoir of this clone.
Journal of Antimicrobial Chemotherapy 01/2014; · 5.34 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Escherichia coli classification into phylogroups reflects the diversity of their pathogenicity and their ecological niche, B2 isolates being the most virulent among extra-intestinal strains. MALDI-TOF MS allows a quick, automated, simple and inexpensive bacterial identification. We evaluated the MALDI-TOF MS as a tool for E. coli phylogroup differentiation.
We used 656 E. coli isolates, previously assigned to phylogroup A, B1, B2, and D by multiplex PCR, to constitute independent training and validation sets. We then defined two phylogrouping strategies, both validated on spectra obtained by the ‘direct transfer method’.
The first strategy used the MALDI Biotyper software (Bruker Daltonik) that identified a single peak shift between isolates of phylogroup B2 and those of groups A, B1 and D. It accurately classified 89% of the isolates. The second strategy used the ClinProTools software (Bruker Daltonik) and was based on three successive models. The model 1 adequately differentiated 92% of phylogroup B2-isolates from those belonging to phylogroups A, B1, D. The model 2 adequately discriminated 87% of phylogroup D-isolates from those of phylogroups A and B1. The model 3 correctly sorted 69% of A and B1-isolates.
We concluded that clinical laboratories could routinely and very quickly assign E. coli isolates to phylogroups with MALDI-TOF MS. These methods could (i) expedite the detection of the most virulent strains belonging to phylogroup B2 and (ii) be a first-line tool to monitor the epidemiology of extra-intestinal pathogenic E. coli.
International Journal of Medical Microbiology. 01/2014;
[Show abstract][Hide abstract] ABSTRACT: A total of 1,021 extended-spectrum-β-lactamase-producing Escherichia coli (ESBLEC) isolates obtained in 2006 during a Spanish national survey conducted in 44 hospitals were analyzed for the presence of the O25b:H4-B2-ST131 (sequence type 131) clonal group. Overall, 195 (19%) O25b-ST131 isolates were detected, with prevalence rates ranging from 0% to 52% per hospital. Molecular characterization of 130 representative O25b-ST131 isolates showed that 96 (74%) were positive for CTX-M-15, 15 (12%) for CTX-M-14, 9 (7%) for SHV-12, 6 (5%) for CTX-M-9, 5 (4%) for CTX-M-32, and 1 (0.7%) each for CTX-M-3 and the new ESBL enzyme CTX-M-103. The 130 O25b-ST131 isolates exhibited relatively high virulence scores (mean, 14.4 virulence genes). Although the virulence profiles of the O25b-ST131 isolates were fairly homogeneous, they could be classified into four main virotypes based on the presence or absence of four distinctive virulence genes: virotypes A (22%) (afa FM955459 positive, iroN negative, ibeA negative, sat positive or negative), B (31%) (afa FM955459 negative, iroN positive, ibeA negative, sat positive or negative), C (32%) (afa FM955459 negative, iroN negative, ibeA negative, sat positive), and D (13%) (afa FM955459 negative, iroN positive or negative, ibeA positive, sat positive or negative). The four virotypes were also identified in other countries, with virotype C being overrepresented internationally. Correspondingly, an analysis of XbaI macrorestriction profiles revealed four major clusters, which were largely virotype specific. Certain epidemiological and clinical features corresponded with the virotype. Statistically significant virotype-specific associations included, for virotype B, older age and a lower frequency of infection (versus colonization), for virotype C, a higher frequency of infection, and for virotype D, younger age and community-acquired infections. In isolates of the O25b:H4-B2-ST131 clonal group, these findings uniquely define four main virotypes, which are internationally distributed, correspond with pulsed-field gel electrophoresis (PFGE) profiles, and exhibit distinctive clinical-epidemiological associations.
Journal of Clinical Microbiology. 10/2013; 51(10).
[Show abstract][Hide abstract] ABSTRACT: To determine factors associated with CTX-M-producing ST131 Escherichia coli which is the worldwide predominant lineage among CTX-M-producing E. coli isolates.
Consecutive inpatients with a clinical sample positive for CTX-M-producing E. coli and considered as cases in a previous 8-month (2008-2009) case-control study performed in ten university hospitals in the Paris area were included in the present sub-population study. Patients with a CTX-M-producing ST131 E. coli clinical isolate were compared with those with a CTX-M-producing non-ST131 E. coli clinical isolate with regard to 66 variables. Variables were first compared using univariate logistic regression, then a multivariate analysis using a backward selection with variables with p-value <0.1 in univariate analysis was carried out.
Fifty-five patients with a CTX-M-producing ST131 E. coli clinical isolate were compared to 97 patients with a CTX-producing non-ST131 E. coli clinical isolate. Multivariate analysis showed that only previous residence in long term care facilities (OR = 4.4; 95% CI = 1.3-14.7) was positively associated with a CTX-M-producing ST131 E. coli isolate. However, it also showed that regular consumption of poultry products (OR = 0.2; 95% CI = 0.1-0.6), having had at least one device in the preceding 6 months (OR = 0.3; 95% CI = 0.1-0.7) and stay in ICU (OR = 0.2; 95% CI = 0.05-0.8) were negatively associated with isolation of CTX-M-producing ST131 E. coli from clinical samples.
This study provides more insight into the epidemiological features of ST131 and non-ST131 E. coli producing CTX-M enzymes. It shows, for the first time, that isolation of CTX-M-producing ST131 E. coli from clinical samples is not linked to consumption of various foods and confirms that residence in long term care facilities is a predictor of these isolates.
PLoS ONE 09/2013; 8(9):72191-. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Background. Fluoroquinolone-resistant Escherichia coli are increasingly prevalent. Their clonal origins-potentially critical for control efforts-remain undefined.Methods. Antimicrobial resistance profiles and fine clonal structure were determined for 236 diverse-source historical (1967-2009) E. coli isolates representing sequence type ST131 and 853 recent (2010-2011) consecutive E. coli isolates from five clinical laboratories in Seattle, WA, and Minneapolis, MN. Clonal structure was resolved based on fimH sequence (fimbrial adhesin gene: H sub-clone assignments), multi-locus sequence typing, gyrA and parC sequence (fluoroquinolone resistance-determining loci), and pulsed-field gel electrophoresis.Results. Of the recent fluoroquinolone-resistant clinical isolates, 52% represented a single ST131 sub-clonal lineage, H30, which expanded abruptly after 2000. This sub-clone had a unique and conserved gyrA/parC allele combination, supporting its tight clonality. Unlike other ST131 sub-clones, H30 was significantly associated with fluoroquinolone resistance and was the most prevalent sub-clone among current E. coli clinical isolates, overall (10.4%) and within every resistance category (11-52%).Conclusions. Most current fluoroquinolone-resistant E. coli clinical isolates, and the largest share of multidrug-resistant isolates, represent a highly clonal sub-group that likely originated from a single rapidly expanded and disseminated ST131 strain. Focused attention to this strain will be required to control the fluoroquinolone and multidrug-resistant E. coli epidemic.
The Journal of Infectious Diseases 01/2013; · 5.85 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Imipenem (IPM) is a carbapenem antibiotic frequently used in severe hospital infections. Several reports have mentioned the emergence of resistant isolates exhibiting membrane modifications. A study was conducted between September 2005 and August 2007 to survey infections due to Enterobacter aerogenes in patients hospitalised in a French university hospital. Resistant E. aerogenes clinical isolates obtained from patients treated with IPM and collected during the 3 months following initiation of treatment were phenotypically and molecularly characterised for β-lactamases, efflux pumps activity and outer membrane proteins. Among the 339 patients infected with E. aerogenes during the study period, 41 isolates (12.1%) were resistant to extended-spectrum cephalosporins and 17 patients (5.0%) were treated with IPM. The isolates from these 17 patients presented TEM-24 and basal efflux expression. Following IPM treatment, an IPM-intermediate-susceptible (IPM-I) isolate emerged in 11 patients and an IPM-resistant (IPM-R) isolate in 6 patients. A change in the porin balance (Omp35/Omp36) was observed in IPM-I isolates exhibiting ertapenem resistance. Finally, a porin deficiency (Omp35 and Omp36 absence) was detected in IPM-R isolates associated with efflux pump expression. This study indicates that the alteration in porin expression, including the shift of porin expression and lack of porins, contribute to the E. aerogenes adaptive response to IPM treatment.
International journal of antimicrobial agents 12/2012; · 3.03 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: OBJECTIVES: In 2006, 0.6% of healthy subjects living in the Paris area had extended-spectrum β-lactamase (ESBL)-producing Escherichia coli in their gut. To assess the evolution of this rate, a study identical to that of 2006 was conducted in 2011. PARTICIPANTS AND METHODS: Healthy adults who visited the IPC check-up centre in February-March 2011 and agreed to participate, provided stools and answered a questionnaire on the visit day. Stools were analysed to detect ESBL producers and to isolate the dominant E. coli population. ESBLs were molecularly characterized. For the subjects harbouring ESBL-producing E. coli, the phylogenetic group and sequence type (ST) were determined for both ESBL-producing and dominant E. coli isolates. PFGE profiles were also determined when two types of isolates had the same ST. RESULTS: Among the 345 subjects included, 21 (6%) had ESBL-producing E. coli faecal carriage. None of the previously published risk factors was identified. CTX-M accounted for 86% and SHV-12 for 14%. Dominant and ESBL-producing E. coli were similarly distributed into phylogenetic groups (A, 52%-48%; B1, 5%; B2, 24%-14%; and D, 19%-33%). Dominant and ESBL-producing E. coli displayed a polyclonal structure (18 STs each). However, ST10 and ST131 were identified in dominant and ESBL-producing E. coli isolates from different subjects. Most (20/21) ESBL producers were subdominant and belonged (16/21) to STs different from that of the corresponding dominant E. coli. CONCLUSIONS: The 10-fold increase in the rate of healthy subjects with ESBL-producing E. coli faecal carriage over a 5 year period suggests wide dissemination of these isolates in the Parisian community.
Journal of Antimicrobial Chemotherapy 11/2012; · 5.34 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In Klebsiella pneumoniae, overexpression of non-specific efflux pumps causes low-level cross-resistance to β-lactams (mainly cefoxitin) and to antibiotics belonging to other families, such as quinolones, chloramphenicol and tetracycline (11, 12).…
Antimicrobial Agents and Chemotherapy 10/2012; · 4.57 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Escherichia coli is the species most frequently associated with clinical infections by extended-spectrum-β-lactamase (ESBL)-producing isolates, with the CTX-M ESBL enzymes being predominant and found in genetically diverse E. coli isolates. The main objective of this study was to compare, on the basis of a case-control design, the phylogenetic diversity of 152 CTX-M-producing and 152 non-ESBL-producing clinical E. coli isolates. Multilocus sequence typing revealed that even though CTX-M enzymes were largely disseminated across the diversity of E. coli isolates, phylogenetic group B2 showed a particularly heterogeneous situation. First, clone ST131 of group B2 was strongly associated with CTX-M production (55 [79%] of 70 isolates), with CTX-M-15 being predominant. Second, the remaining members of group B2 were significantly less frequently associated with CTX-M production (9 [12%] of 75) than E. coli phylogenetic groups A, B1, and D (88 [55%] of 159). CTX-M-producing ST131 E. coli isolates were significantly more frequent in patients hospitalized in geriatric wards or long-term care facilities. Besides, the non-ESBL ST131 isolates significantly more frequently showed resistance to penicillins than the non-ESBL, non-ST131 isolates did. In conclusion, the present study emphasizes the particular antimicrobial resistance and epidemiologic characteristics of clone ST131 within group B2, which could result from the higher antibiotic exposure of this clone, as it is the predominant clone of group B2 carried in the human gut.
Journal of clinical microbiology 07/2012; 50(9):2974-81. · 4.16 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Extended-spectrum β-lactamase-producing Enterobacteriaceae isolates (ESBLE) are emerging pathogens that confer resistance to antimicrobial drugs. We conducted a 10-year study in France (January 2001-April 2010) to investigate the incidence of and risk factors for ESBLE infections after liver transplant. Of 710 transplant patients screened preoperatively for ESBLE fecal carriage, 5.5% had ESBLE infection develop within 4 months after surgery; patients with pretransplant ESBLE fecal carriage were more likely to have infection develop than were noncarriers. Typing showed extensive genetic diversity, with a large predominance of CTX-M enzymes. Independent predictors of ESBLE infection were pretransplant fecal carriage, Model for End Stage Liver Disease score >25, and return to surgery. Our results indicate that the influx of preoperatively acquired ESBLE isolates into the hospital outweighs cross-transmission in the epidemiology of ESBLE infections after liver transplant. Transplant candidates should be systematically screened for carriage, and posttransplant infection in carriers should be treated with carbapenems.
[Show abstract][Hide abstract] ABSTRACT: Escherichia coli sequence type 131 (ST131), an emerging disseminated public health threat, causes multidrug-resistant extraintestinal infections. Among 579 diverse E. coli ST131 isolates from 1967-2009, we compared pulsotypes (>94% similar XbaI pulsed-field gel electrophoresis profiles) by collection year, geographic origin, source, and antimicrobial drug-resistance traits. Of 170 pulsotypes, 65 had >2 isolates and accounted for 85% of isolates. Although extensively dispersed geographically, pulsotypes were significantly source specific (e.g., had little commonality between humans vs. foods and food animals). The most prevalent pulsotypes were associated with recent isolation, humans, and antimicrobial drug resistance. Predominant pulsotype 968 was associated specifically with fluoroquinolone resistance but not with extended-spectrum β-lactamase production or bla(CTX-M-15). Thus, several highly successful antimicrobial drug-resistant lineages within E. coli ST131 have recently emerged and diffused extensively among locales while maintaining a comparatively restricted host/source range. Identification of factors contributing to this behavior of ST131 could help protect public health.
[Show abstract][Hide abstract] ABSTRACT: Recently, the worldwide propagation of clonal CTX-M-15-producing Escherichia coli isolates, namely ST131 and O25b:H4, has been reported. Like the majority of extra-intestinal pathogenic E. coli isolates, the pandemic clone ST131 belongs to phylogenetic group B2, and has recently been shown to be highly virulent in a mouse model, even though it lacks several genes encoding key virulence factors (Pap, Cnf1 and HlyA). Using two animal models, Caenorhabditis elegans and zebrafish embryos, we assessed the virulence of three E. coli ST131 strains (2 CTX-M-15- producing urine and 1 non-ESBL-producing faecal isolate), comparing them with five non-ST131 B2 and a group A uropathogenic E. coli (UPEC). In C. elegans, the three ST131 strains showed intermediate virulence between the non virulent group A isolate and the virulent non-ST131 B2 strains. In zebrafish, the CTX-M-15-producing ST131 UPEC isolates were also less virulent than the non-ST131 B2 strains, suggesting that the production of CTX-M-15 is not correlated with enhanced virulence. Amongst the non-ST131 B2 group isolates, variation in pathogenic potential in zebrafish embryos was observed ranging from intermediate to highly virulent. Interestingly, the ST131 strains were equally persistent in surviving embryos as the non-ST131-group B2 strains, suggesting similar mechanisms may account for development of persistent infection. Optical maps of the genome of the ST131 strains were compared with those of 24 reference E. coli strains. Although small differences were seen within the ST131 strains, the tree built on the optical maps showed that these strains belonged to a specific cluster (86% similarity) with only 45% similarity with the other group B2 strains and 25% with strains of group A and D. Thus, the ST131 clone has a genetic composition that differs from other group B2 strains, and appears to be less virulent than previously suspected.
PLoS ONE 01/2012; 7(3):e34294. · 3.53 Impact Factor