Etude de la première et de la dernière étape du processus catalytique des b-lactamases à spectre étendu de type CTX-M

L'utilisation intensive des céphalosporines de 3ème génération en thérapeutique humaine a conduit à l'émergence de b-lactamases à spectre étendu (BLSE) capables d'hydrolyser ces molécules. Parmi elles, les enzymes de type CTX-M constituent actuellement un véritable problème de santé publique de part leur dissémination mondiale et leur isolement préférentiel chez des bactéries virulentes retrouvées aussi bien à l'hôpital qu'en communautaire. Une meilleure connaissance des interactions moléculaires entre les b-lactamines et ces b-lactamases pourrait contribuer à lutter efficacement contre ces enzymes. L'objectif de ce travail était donc d'analyser le 1er et le dernier stade du processus catalytique des CTX-M. La structure de l'enzyme mutée inactive CTX-M-9 S70G a été obtenue à haute résolution en complexe avec différentes b-lactamines permettant d'étudier d'une part la phase de reconnaissance du substrat (pénicilline G, pipéracilline, céfotaxime et céfoxitine) et d'autre part la phase précédant la libération du produit de l'hydrolyse (pénicilline G et pipéracilline hydrolysées). Il émerge de ces travaux : 1) la présence d'interactions conservées entre les enzymes de type CTX-M et les b-lactamines au cours du processus catalytique, 2) une densité importante de liaisons hydrogène et un élargissement de la poche catalytique permettant une reconnaissance et une accomodation optimale du céfotaxime, 3) un mécanisme d'entrée des b-lactamines dans le site actif probablement assisté par le résidu arginine 276, et 4) un réarrangement moléculaire consécutif à l'hydrolyse du cycle b-lactame favorisant un phénomène d'auto-expulsion de la b-lactamine hydrolysée. Ces travaux permettent donc de mieux comprendre les aspects moléculaires du mécanisme d'hydrolyse des b-lactamines par les BLSE de type CTX-M et pourraient aider, à terme, à la conception de nouvelles molécules inhibitrices de ces enzymes ou de nouvelles b-lactamines plus résistantes à leur action hydrolytique.CLERMONT FD-BCIU-Santé (631132104) / SudocLYON1-BU Santé (693882101) / SudocSudocFranceF

Identification of a Variety of Staphylococcus Species by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry ▿

Whole-cell fingerprinting by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) in combination with a dedicated bioinformatic software tool (MALDI Biotyper 2.0) was used to identify 152 staphylococcal strains corresponding to 22 staphylococcal species. Spectra of the 152 isolates, previously identified at the species level using a sodA gene-based oligonucleotide array, were analyzed against the main spectra of 3,030 microorganisms. A total of 151 strains out of 152 (99.3%) were correctly identified at the species level; only one strain was identified at the genus level. The MALDI-TOF MS method revealed different clonal lineages of Staphylococcus epidermidis that were of either human or environmental origin, which suggests that the MALDI-TOF MS method could be useful in the profiling of staphylococcal strains. The topology of the dendrogram generated by the MALDI Biotyper 2.0 software from the spectra of 120 Staphylococcus reference strains (representing 36 species) was in general agreement with that inferred from the 16S rRNA gene-based analysis. Our findings indicate that the MALDI-TOF MS technology, associated with a broad-spectrum reference database, is an effective tool for the swift and reliable identification of Staphylococci

Multirecombinant Enterovirus A71 Subgenogroup C1 Isolates Associated with Neurologic Disease, France, 2016–2017

International audienceIn 2016, an upsurge of neurologic disease associated with infection with multirecombinant enterovirus A71 subgenogroup C1 lineage viruses was reported in France. These viruses emerged in the 2000s; 1 recombinant is widespread. This virus lineage has the potential to be associated with a long-term risk for severe disease among children

Inventory of extended-spectrum-β-lactamase-producing Enterobacteriaceae in France as assessed by a multicenter study

The objective of this study was to perform an inventory of the ESBL-producing Enterobacteriaceae isolates responsible for infections in French hospitals, and to assess the mechanisms associated with ESBL diffusion.200 non-redundant ESBL-producing Enterobacteriaceae strains isolated from clinical samples were collected during a multi-centric study performed in 18 representative French hospitals. Antibiotic resistance genes were identified by PCR and sequencing experiments. The clonal relatedness between isolates was investigated by the Diversilab system. ESBL-encoding plasmids were compared by PCR-based-replicon-typing and plasmid-multi-locus-sequence-typing.CTX-M-15, CTX-M-1, CTX-M-14 and SHV-12 were the most prevalent ESBLs (8 to 46.5%). The three CTX-M-type EBSLs were significantly observed in Escherichia coli (37.1%, 24.2% and 21.8% respectively), and CTX-M-15 was the predominant ESBL in Klebsiella pneumoniae (81.1%). SHV-12 was associated with ESBL-encoding Enterobacter cloacae strains (37.9%). qnrB, aac(6')-Ib-cr and aac (3)-II genes were the main plasmid-mediated resistance genes, with prevalence varying between 19.5 and 45% according to the ESBLs. Molecular typing did not identify wide clonal diffusion. Plasmid analysis suggested the diffusion of few numbers of ESBL-encoding plasmids, especially in K. pneumoniae and E. cloacae However, the ESBL-encoding genes were observed in different plasmid replicons according to the bacterial species.The prevalence of ESBL subtypes is different according to the Enterobacteriaceae species. Plasmid spread is a key determinant of this epidemiology and the link observed between the ESBL-encoding plasmids and the bacterial host explain the differences observed in the Enterobacteriaceae species

Staphylococcus capitis isolated from bloodstream infections: a nationwide 3-month survey in 38 neonatal intensive care units

International audienceTo increase the knowledge about S. capitis in the neonatal setting, we conducted a nationwide 3-month survey in 38 neonatal intensive care units (NICUs) covering 56.6% of French NICU beds. We demonstrated 14.2% of S. capitis BSI (S.capBSI) among nosocomial BSIs. S.capBSI incidence rate was 0.59 per 1000 patient-days. A total of 55.0% of the S.capBSIs were late onset catheter-related BSIs. The S. capitis strains infected preterm babies (median gestational age 26 weeks, median birth weight 855 g). They were resistant to methicillin and aminoglycosides and belonged to the NRCS-A clone. Evolution was favorable in all but one case, following vancomycin treatment