fbl-Typing of Staphylococcus lugdunensis: A Frontline Tool for Epidemiological Studies, but Not Predictive of Fibrinogen Binding Ability

Staphylococcus lugdunensis is increasingly recognized as a potent pathogen, responsible for severe infections with an outcome resembling that of Staphylococcus aureus. Here, we developed and evaluated a tool for S. lugdunensis typing, using DNA sequence analysis of the repeat-encoding region (R-domain) in the gene encoding the fibrinogen (Fg)-binding protein Fbl (fbl-typing). We typed 240 S. lugdunensis isolates from various clinical and geographical origins. The length of the R-domain ranged from 9 to 52 repeats. fbl-typing identified 54 unique 18-bp repeat sequences and 92 distinct fbl-types. The discriminatory power of fbl-typing was higher than that of multilocus sequence typing (MLST) and equivalent to that of tandem repeat sequence typing. fbl-types could assign isolates to MLST clonal complexes with excellent predictive power. The ability to promote adherence to immobilized human Fg was evaluated for 55 isolates chosen to reflect the genetic diversity of the fbl gene. We observed no direct correlation between Fg binding ability and fbl-types. However, the lowest percentage of Fg binding was observed for isolates carrying a 5′-end frameshift mutation of the fbl gene and for those harboring fewer than 43 repeats in the R-domain. qRT-PCR assays for some isolates revealed no correlation between fbl gene expression and Fg binding capacity. In conclusion, this study shows that fbl-typing is a useful tool in S. lugdunensis epidemiology, especially because it is an easy, cost-effective, rapid and portable method (http://fbl-typing.univ-rouen.fr/). The impact of fbl polymorphism on the structure of the protein, its expression on the cell surface and in virulence remains to be determined

In vitro activity of daptomycin against Enterococcus faecalis under various conditions of growth-phases, inoculum and pH

Enterococcus faecalis (E. faecalis) has become a major leading cause of nosocomial endocarditis. Treatment of such infections remains problematic and new therapeutic options are needed. Nine E. faecalis strains were tested: six obtained from patients presenting endocarditis, one with isolated bacteremia, and two reference strains. Antibiotics included daptomycin, alone or in combination, linezolid, tigecycline, rifampicin, gentamicin, teicoplanin, ceftriaxone and amoxicillin. Time-kill studies included colony counts at 1, 4 and 24 h of incubation. Significant bactericidal activity was defined as a decrease of ≥3log10CFU/ml after 24 h of incubation. Antibiotics were tested at a low (10(6) CFU/ml) and high (10(9) CFU/ml) inoculum, against exponential- and stationary-phase bacteria. We also performed time kill studies of chemically growth arrested E. faecalis. Various pH conditions were used during the tests. In exponential growth phase and with a low inoculum, daptomycin alone at 60 µg/ml and the combination amoxicillin-gentamicin both achieved a 4-log10 reduction in one hour on all strains. In exponential growth phase with a high inoculum, daptomycin alone was bactericidal at a concentration of 120 µg/ml. All the combinations tested with this drug were indifferent. In stationary phase with a high inoculum daptomycin remained bactericidal but exhibited a pH dependent activity and slower kill rates. All combinations that did not include daptomycin were not bactericidal in conditions of high inoculum, whatever the growth phase. The results indicate that daptomycin is the only antibiotic that may be able of overcoming the effects of growth phase and high inoculum

Etude de la virulence de Staphylococcus lugdunensis

Staphylococcus lugdunensis is a coagulase negative staphylococcus species that may causes various infections of unusual severity. We conducted a translational study with a prospective clinical trial that aimed to describe S. lugdunensis infections and its real pathogenicity, associated with a systematic research of in vitro putative virulence factors. The final objective was to determine the statistical relationship between those two and find a putative real virulence factor. This trial was conducted between 2013 and 2016. It showed that S. lugdunensis displayed a high level of pathogenicity as 37.2% of all strains isolated came from infected patients and most of those infections were osteoarticular infections. We discovered a new protease that we named lugdulysin and that was strongly associated with osteoarticular infections. This secreted protein of 37 kDa was purified and sequenced, we characterized its chemical properties and the nucleotide environment of the coding sequence. We also achieved de novo sequencing of 7 strains of S. lugdunensis and found that several mobile genetic elements belonged to the sequences as plasmids and prophages.Staphylococcus lugdunensis est un staphylocoque à coagulase négative qui présente de nombreuses particularités sur le plan clinique et microbiologique. Cette bactérie commensale de la peau est impliquée dans des infections humaines d’une particulière gravité. Ce travail de thèse nous a permis de déterminer que S. lugdunensis présente bien une pathogénicité tout à fait inhabituelle pour un SCN car 37.2% de toutes les souches recueillies entre 2013 et 2016 étaient impliquées dans un processus infectieux. Nous avons aussi observé que les infections ostéo-articulaires en étaient la première manifestation. Nous avons découvert une nouvelle protéase excrétée par 61.7% des souches qui présente une association statistique forte avec les infections ostéo-articulaires. Il s’agit d’une métalloprotéase de 37 kDa que nous avons pu purifier puis séquencer afin de caractériser ses propriétés biochimiques et son environnement génique. Enfin, nous avons aussi réalisé un séquençage de novo de 7 souches de S. lugdunensis et démontrer l’existence de multiples éléments génétiques mobiles

Comparative Genomics and Identification of an Enterotoxin-Bearing Pathogenicity Island, SEPI-1/SECI-1, in Staphylococcus epidermidis Pathogenic Strains

Staphylococcus epidermidis is a leading cause of nosocomial infections, majorly resistant to beta-lactam antibiotics, and may transfer several mobile genetic elements among the members of its own species, as well as to Staphylococcus aureus; however, a genetic exchange from S. aureus to S. epidermidis remains controversial. We recently identified two pathogenic clinical strains of S. epidermidis that produce a staphylococcal enterotoxin C3-like (SEC) similar to that by S. aureus pathogenicity islands. This study aimed to determine the genetic environment of the SEC-coding sequence and to identify the mobile genetic elements. Whole-genome sequencing and annotation of the S. epidermidis strains were performed using Illumina technology and a bioinformatics pipeline for assembly, which provided evidence that the SEC-coding sequences were located in a composite pathogenicity island that was previously described in the S. epidermidis strain FRI909, called SePI-1/SeCI-1, with 83.8–89.7% nucleotide similarity. Various other plasmids were identified, particularly p_3_95 and p_4_95, which carry antibiotic resistance genes (hsrA and dfrG, respectively), and share homologies with SAP085A and pUSA04-2-SUR11, two plasmids described in S. aureus. Eventually, one complete prophage was identified, ΦSE90, sharing 30 out of 52 coding sequences with the Acinetobacter phage vB_AbaM_IME200. Thus, the SePI-1/SeCI-1 pathogenicity island was identified in two pathogenic strains of S. epidermidis that produced a SEC enterotoxin causing septic shock. These findings suggest the existence of in vivo genetic exchange from S. aureus to S. epidermidis

Virulence study of Staphylococcus lugdunensis

Staphylococcus lugdunensis est un staphylocoque à coagulase négative qui présente de nombreuses particularités sur le plan clinique et microbiologique. Cette bactérie commensale de la peau est impliquée dans des infections humaines d’une particulière gravité. Ce travail de thèse nous a permis de déterminer que S. lugdunensis présente bien une pathogénicité tout à fait inhabituelle pour un SCN car 37.2% de toutes les souches recueillies entre 2013 et 2016 étaient impliquées dans un processus infectieux. Nous avons aussi observé que les infections ostéo-articulaires en étaient la première manifestation. Nous avons découvert une nouvelle protéase excrétée par 61.7% des souches qui présente une association statistique forte avec les infections ostéo-articulaires. Il s’agit d’une métalloprotéase de 37 kDa que nous avons pu purifier puis séquencer afin de caractériser ses propriétés biochimiques et son environnement génique. Enfin, nous avons aussi réalisé un séquençage de novo de 7 souches de S. lugdunensis et démontrer l’existence de multiples éléments génétiques mobiles.Staphylococcus lugdunensis is a coagulase negative staphylococcus species that may causes various infections of unusual severity. We conducted a translational study with a prospective clinical trial that aimed to describe S. lugdunensis infections and its real pathogenicity, associated with a systematic research of in vitro putative virulence factors. The final objective was to determine the statistical relationship between those two and find a putative real virulence factor. This trial was conducted between 2013 and 2016. It showed that S. lugdunensis displayed a high level of pathogenicity as 37.2% of all strains isolated came from infected patients and most of those infections were osteoarticular infections. We discovered a new protease that we named lugdulysin and that was strongly associated with osteoarticular infections. This secreted protein of 37 kDa was purified and sequenced, we characterized its chemical properties and the nucleotide environment of the coding sequence. We also achieved de novo sequencing of 7 strains of S. lugdunensis and found that several mobile genetic elements belonged to the sequences as plasmids and prophages

Virulence study of Staphylococcus lugdunensis

Staphylococcus lugdunensis est un staphylocoque à coagulase négative qui présente de nombreuses particularités sur le plan clinique et microbiologique. Cette bactérie commensale de la peau est impliquée dans des infections humaines d’une particulière gravité. Ce travail de thèse nous a permis de déterminer que S. lugdunensis présente bien une pathogénicité tout à fait inhabituelle pour un SCN car 37.2% de toutes les souches recueillies entre 2013 et 2016 étaient impliquées dans un processus infectieux. Nous avons aussi observé que les infections ostéo-articulaires en étaient la première manifestation. Nous avons découvert une nouvelle protéase excrétée par 61.7% des souches qui présente une association statistique forte avec les infections ostéo-articulaires. Il s’agit d’une métalloprotéase de 37 kDa que nous avons pu purifier puis séquencer afin de caractériser ses propriétés biochimiques et son environnement génique. Enfin, nous avons aussi réalisé un séquençage de novo de 7 souches de S. lugdunensis et démontrer l’existence de multiples éléments génétiques mobiles.Staphylococcus lugdunensis is a coagulase negative staphylococcus species that may causes various infections of unusual severity. We conducted a translational study with a prospective clinical trial that aimed to describe S. lugdunensis infections and its real pathogenicity, associated with a systematic research of in vitro putative virulence factors. The final objective was to determine the statistical relationship between those two and find a putative real virulence factor. This trial was conducted between 2013 and 2016. It showed that S. lugdunensis displayed a high level of pathogenicity as 37.2% of all strains isolated came from infected patients and most of those infections were osteoarticular infections. We discovered a new protease that we named lugdulysin and that was strongly associated with osteoarticular infections. This secreted protein of 37 kDa was purified and sequenced, we characterized its chemical properties and the nucleotide environment of the coding sequence. We also achieved de novo sequencing of 7 strains of S. lugdunensis and found that several mobile genetic elements belonged to the sequences as plasmids and prophages

Cinétique de bactéricidie de diverses associations d'antibiotiques sur des souches d'Enterococcus faecalis évaluée in vitro dans diverses conditions de croissance bactérienne, d'inoculum et de pH

STRASBOURG-Medecine (674822101) / SudocSudocFranceF

Coagulase-Negative Staphylococci Pathogenomics

Coagulase-negative Staphylococci (CoNS) are skin commensal bacteria. Besides their role in maintaining homeostasis, CoNS have emerged as major pathogens in nosocomial settings. Several studies have investigated the molecular basis for this emergence and identified multiple putative virulence factors with regards to Staphylococcus aureus pathogenicity. In the last decade, numerous CoNS whole-genome sequences have been released, leading to the identification of numerous putative virulence factors. Koch’s postulates and the molecular rendition of these postulates, established by Stanley Falkow in 1988, do not explain the microbial pathogenicity of CoNS. However, whole-genome sequence data has shed new light on CoNS pathogenicity. In this review, we analyzed the contribution of genomics in defining CoNS virulence, focusing on the most frequent and pathogenic CoNS species: S. epidermidis, S. haemolyticus, S. saprophyticus, S. capitis, and S. lugdunensis