Hélène Jean-Pierre

CTX-M β-Lactamase-Producing Escherichia coli in French Hospitals: Prevalence, Molecular Epidemiology, and Risk Factors

ABSTRACT In 2004, 65 CTX-M-producing Escherichia coli isolates were collected from infected patients in four French hospitals. The blaCTX-M-15 genes were predominant. Pulsed-field gel electrophoresis highlighted a clonal propagation of CTX-M-15-producing strains belonging to phylogenetic group B2, notably in the community. The main risk factors for acquiring these isolates were urinary tract infections or the presence of a urinary catheter in diabetic or renal failure patients.

qnrA in CTX-M-Producing Escherichia coli Isolates from France

ABSTRACT By PCR, we screened for qnr genes 112 clinical isolates of extended-spectrum β-lactamase-producing Escherichia coli collected from hospitals in France during 2004. For the first time, 7.7% of CTX-M-producing E. coli isolates presented a plasmid-mediated resistance to quinolones. All strains harbored a qnrA gene located on a sul1-type class 1 integron with similar structure to the In36 integron.

Anaeroglobus geminatus gen. nov., sp. nov., a novel member of the family Veillonellaceae.

A hitherto unknown anaerobic coccus isolated from a post-operative fluid collection was characterized by phenotypic and phylogenetic methods. 16S rDNA sequence analysis revealed an affiliation of this isolate to the family Veillonellaceae. Also, a high level of sequence similarity was observed to some oral clone sequences of Megasphaera spp. contained in the GenBank database under designations BB166, CS025 and BS073. These clones and the unknown bacterium form a well-separated phylogenetic branch that may represent a novel lineage within the family Veillonellaceae. Based on phenotypic and phylogenetic evidence, a new genus, Anaeroglobus gen. nov., is proposed for the unknown bacterium, with one species, Anaeroglobus geminatus gen. nov., sp. nov. The type strain of Anaeroglobus geminatus is strain AIP 313.00T (= CIP 106856T = CCUG 44773T). It is also suggested that the oral clones BB166, CS025 and BS073 belong to the genus Anaeroglobus.

Prosthetic joint infection due to Veillonella dispar

Abstract Described here is the first report of a monomicrobial joint infection caused by Veillonella dispar that resulted in loosening of a prosthesis. This clinical case shows that Veillonella spp. should not be disregarded as contaminant organisms, particularly when they are isolated in pure culture from clinical specimens involving cases of septic arthritis.

Extended-Spectrum β-Lactamase TEM-24 in an Aeromonas Clinical Strain: Acquisition from the Prevalent Enterobacter aerogenes Clone in France

The TEM-24 extended-spectrum β-lactamase (ESBL), initially characterized in Klebsiella pneumoniae (3), is currently the predominant ESBL in France (4). This could be related to the spread of a TEM-24-producing Enterobacter aerogenes clone present in most French hospitals (1) but also found in Belgium (5) and Spain (2). TEM-24 enzyme has been recovered from an increasing number of species of the family Enterobacteriaceae (6, 8) and in Pseudomonas aeruginosa (7). In contrast, resistance to expanded-spectrum cephalosporins mediated by ESBLs has never been described in the genus Aeromonas, for which β-lactam resistance involves three chromosomally mediated enzymes: a cephalosporinase, a penicillinase, and a carbapenemase (9). We report the first characterization of a TEM-24-producing clinical Aeromonas strain. 16S ribosomal DNA (rDNA) and gyrB sequencing (10) showed that the strain was most closely related to Aeromonas caviae. This strain was recovered together with an ESBL-producing E. aerogenes isolate from the diarrheal feces of a 76-year-old man admitted to Montpellier Hospital for intestinal ischemia. Based on the resistance phenotype observed after the disk diffusion assay, both E. aerogenes and Aeromonas sp. isolates were suspected to produce ceftazidimase-type ESBL and AAC(6′)-I enzyme. Synergy was clearly observed between expanded-spectrum cephalosporins and clavulanate. The resistance phenotype was transferred in Escherichia coli C600 by mating experiments using Mueller-Hinton agar (Sanofi Diagnostics Pasteur, Marnes-la-Coquette, France) containing ceftazidime (4 μg/ml). The MICs of β-lactams showed high levels of resistance with the E. aerogenes strain and the two transconjugants for penicillins (64 to >2,048 μg/ml), ceftazidime (512 to 1,048 μg/ml), and aztreonam (64 to 1,048 μg/ml). A low level of resistance or decrease in susceptibility was observed for cefotaxime (4 to 8 μg/ml). The Aeromonas sp. strain showed lower resistance levels (penicillins, 8 to 256 μg/ml; ceftazidime, 64 μg/ml; and aztreonam, 8 μg/ml). Clavulanate and tazobactam partially or completely restored the susceptibility to penicillins. ESBL characterization was performed for the two isolates by isoelectric focusing, ESBL-encoding gene amplification and sequencing, and plasmid content analysis (6, 7). The ESBLs were focused at a pI of 6.5. The sequence of ESBL-encoding genes shared 100% identity with the blaTEM-24 gene. Plasmids of approximately 180 kb, which displayed similar restriction patterns, were isolated from the two strains and their transconjugants. These results suggested an in vivo transfer of TEM-24-encoding plasmid from E. aerogenes to Aeromonas sp. in the intestinal tract. We have previously reported the transfer of 180-kb TEM-24-encoding plasmid from E. aerogenes to Providencia rettgeri CIP 107053 (6) and P. aeruginosa CIP 107051 (7). A comparative study of the restriction profiles obtained for the plasmids recovered from these organisms and from the strains analyzed here revealed similar restriction patterns (Fig. ​(Fig.1).1). Pulsed-field gel electrophoresis (PFGE) analysis of the E. aerogenes strains isolated in this study and previously (6, 7) revealed that they belong to the TEM-24-producing E. aerogenes clone prevalent in France. FIG. 1. EcoRI-SalI restriction patterns of 180-kb TEM-24-encoding plasmids obtained from transconjugants corresponding to the following clinical isolates: lane 2, P. rettgeri CIP 107053 (6); lane 3, P. aeruginosa CIP 107051 (7); lane 4, Aeromonas sp. (this study); ... This clone had disseminated the same TEM-24-encoding plasmid among various bacterial species for several years. The isolation of a TEM-24-producing Aeromonas sp. strain extends the list of TEM-24-harboring bacteria, and this wide host range is one of the factors responsible for the persistence and spread of the TEM-24-encoding plasmid.

Phylogeny, diversity and host specialization in the phylum Synergistetes with emphasis on strains and clones of human origin

Members of the phylum Synergistetes have been demonstrated in several environmental ecosystems and mammalian microflorae by culture-independent methods. In the past few years, the clinical relevance of some uncultivated phylotypes has been demonstrated in endodontic infections, and uncultured Synergistetes have been demonstrated in human mouth, gut and skin microbiota. However, Synergistetes are rarely cultured from human samples, and only 17 isolates are currently reported. Twelve members of Synergistetes isolated in the course of various infectious processes, including 3 Jonquetella anthropi, 2 Cloacibacillus evryensis, 2 Pyramidobacter piscolens and 5 unidentified strains, as well as 56 clones obtained by specific PCR from the normal vaginal microflora, were studied. 16S rRNA gene-based phylogeny showed that the clones were grouped into 3 clusters, corresponding to the genus Jonquetella, P. piscolens and one novel Synergistetes taxon. The presence and diversity of Synergistetes were reported for the first time in the vaginal microflora. Synergistetes were found in healthy patients, suggesting that they could play a functional role in human microflorae, but may also act as opportunistic pathogens. Studying the phylogenetic relationships between environmental and mammalian strains and clones revealed clearly delineated independent lineages according to the origin of the sequences.

Optimized PCR-Temporal Temperature Gel Electrophoresis compared to cultivation to assess diversity of gut microbiota in neonates.

Temporal Temperature Gel Electrophoresis of amplified 16S rRNA gene sequences (16S rDNA PCR-TTGE) constitutes a culture-independent molecular method used to study bacterial communities. All the technical steps are crucial for quality and exhaustiveness of the results obtained by such approach. Careful optimization of the protocols used is ideally needed for each ecosystem studied. We present here the strategy used to construct an optimized protocol for a 16S rDNA PCR-TTGE-based analysis of gut microflora in neonates. Improvement of the different steps, i.e. total DNA extraction, amplification in terms of efficiency and reduction of heteroduplex formation, TTGE migration conditions and bacterial identification from TTGE patterns, was performed. The optimized protocol was used for the subsequent analysis of 14 stool samples comparatively to a culture-based method. We showed that a specifically designed ladder representative of the diversity of the studied microflora is a useful tool for the identification of bacterial taxa despite biases inherent to 16S rRNA genes, including intra-genomic heterogeneity. Cultivation and PCR-TTGE gave congruent results but cultivation was more efficient for the detection of minor populations whereas PCR-TTGE gave a more complete description of the major populations. Finally, we demonstrated the reliability, the detection sensitivity and the convenience of the optimized 16S rDNA PCR-TTGE method compared with cultural approaches for studying the premature neonate gut microbiota.

Prevalence of mupirocin resistance among invasive coagulase-negative staphylococci and methicillin-resistant Staphylococcus aureus (MRSA) in France: emergence of a mupirocin-resistant MRSA clone harbouring mupA

OBJECTIVES Mupirocin is the cornerstone of decolonization regimens, a successful strategy to prevent healthcare-associated staphylococcal infections. Several recent studies have reported alarming results: (i) an extending reservoir of mupA, the ancestral mobile resistance gene, among coagulase-negative staphylococci (CoNS); (ii) the emergence of a new resistance gene (mupB); and (iii) a growing number of mupirocin-resistant methicillin-resistant Staphylococcus aureus (MRSA), including highly pathogenic clones. We performed a nationwide prospective study in France to detect such trends among invasive staphylococci. METHODS Between October 2011 and February 2012, 367 MRSA and 708 CoNS invasive isolates were collected from 37 hospitals and analysed centrally. Mupirocin MICs were determined using the broth microdilution method. mupA/B PCR was performed for resistant isolates (MIC >1 mg/L). Genetic relatedness between mupirocin-resistant MRSA isolates was determined by PFGE analysis and related isolates were tested by microarray. RESULTS Among MRSA isolates 2.2% (n = 8) were classified as mupirocin resistant; 1.4% (n = 5) showing low-level resistance (MIC ≤256 mg/L) and 0.8% (n = 3) high-level resistance (MIC >256 mg/L). Only the latter isolates carried mupA. A clonal relationship was identified between two mupA-negative MRSA from the same hospital and three mupA-positive MRSA from three distant towns; these three isolates belonged to the Lyon clone. Mupirocin resistance was identified in 10.3% of CoNS, mainly highly resistant mupA-positive isolates (5.6%). The mupB gene was not detected in mupirocin-resistant MRSA or CoNS. CONCLUSIONS This first large national study indicates the need for thorough epidemiological monitoring and a stewardship programme to prevent and detect mupirocin resistance in staphylococci.

Antimicrobial resistance in wildlife

1. The spread of antimicrobial resistance is of major concern for human health and leads to growing economic costs. While it is increasingly hypothesized that wildlife could play an important role in antimicrobial-resistant bacteria dynamics, empirical data remain scarce. 2. The present work builds on a systematic review of the available data in order to highlight the main information we have and to suggest research pathways that should be followed if we aim to fill the gaps in our current knowledge. 3. To achieve this goal, we address four questions: (i) Which resistant bacteria are the most frequently observed in wildlife? (ii) How are resistant bacteria exchanged between wildlife and the other hosts involved? (iii) In which habitats are those resistant bacteria found? (iv) Are resistances associated with certain ecological traits of the host? 4. Synthesis and applications. We highlight the strong link existing between the impact of human activities on natural habitats and the carriage of antimicrobial-resistant bacteria by wildlife. Furthermore, we underline that omnivorous, anthropophilic and carnivorous species are at high risk of being carriers and potentially spreaders of antimicrobial-resistant bacteria. Identifying among those groups key sentinel species may be of particular interest to implement ecosystem contamination surveillance. Finally, we discuss possible exchange routes for antimicrobial-resistant bacteria between humans and wildlife. Considering that water is of major importance in those exchanges, a critical way to control antimicrobial resistance spread may be to limit aquatic environment contamination by antimicrobial-resistant bacteria and antibiotics.

Staphylococcus aureus infective endocarditis versus bacteremia strains: Subtle genetic differences at stake.

Infective endocarditis (IE)((1)) is a severe condition complicating 10-25% of Staphylococcus aureus bacteremia. Although host-related IE risk factors have been identified, the involvement of bacterial features in IE complication is still unclear. We characterized strictly defined IE and bacteremia isolates and searched for discriminant features. S. aureus isolates causing community-acquired, definite native-valve IE (n=72) and bacteremia (n=54) were collected prospectively as part of a French multicenter cohort. Phenotypic traits previously reported or hypothesized to be involved in staphylococcal IE pathogenesis were tested. In parallel, the genotypic profiles of all isolates, obtained by microarray, were analyzed by discriminant analysis of principal components (DAPC)((2)). No significant difference was observed between IE and bacteremia strains, regarding either phenotypic or genotypic univariate analyses. However, the multivariate statistical tool DAPC, applied on microarray data, segregated IE and bacteremia isolates: IE isolates were correctly reassigned as such in 80.6% of the cases (C-statistic 0.83, P<0.001). The performance of this model was confirmed with an independent French collection IE and bacteremia isolates (78.8% reassignment, C-statistic 0.65, P<0.01). Finally, a simple linear discriminant function based on a subset of 8 genetic markers retained valuable performance both in study collection (86.1%, P<0.001) and in the independent validation collection (81.8%, P<0.01). We here show that community-acquired IE and bacteremia S. aureus isolates are genetically distinct based on subtle combinations of genetic markers. This finding provides the proof of concept that bacterial characteristics may contribute to the occurrence of IE in patients with S. aureus bacteremia.