Staphylococcal Panton-Valentine Leucocidin as a Major Virulence Factor Associated to Furuncles

Panton-Valentine Leucocidin (PVL), one of the β-barrel pore-forming staphylococcal leucotoxins, is known to be associated to furuncles and some severe community pneumonia. However, it is still uncertain how many other virulence factors are also associated to furuncles and what the risk factors of furuncles are in immuno-compromised status of patients, especially the HIV (+) patients. In this paper, we use antigen immunoprecipitation and multiplex PCR approach to determine the presence of 19 toxins, 8 adhesion factors and the PFGE profiles associated to furuncles in three independent patient study groups of S. aureus (SA) isolates collected from the Cayenne General Hospital (French Guiana). The patient groups were made of: 16 isolates from HIV (−) patients, 9 from HIV (+) patients suffering from furuncles, and 30 control isolates from patients with diverse secondary infected dermatitis. Our data reveals that the majority (96%) of SA strains isolated from HIV patient-derived furuncles significantly produced PVL (p<10−7), whereas only 10% of SA strains produced this toxin in secondary infected dermatosis. A high prevalence of LukE-LukD-producing isolates (56 to 78%) was recorded in patient groups. Genes encoding clumping factor B, collagen- and laminin-binding proteins (clfB, cna, lbp, respectively) were markedly frequent (30 to 55%), without being associated to a specific group. Pulse field gel electrophoresis evidenced 24 overall pulsotypes, whereas the 25 PVL-producing isolates were distributed into 15 non clonal fingerprints. These pulsotypes were not specific PVL-producing isolates. PVL appears to be the major virulence factor associated to furuncles in Europe and in South America regardless of the immune status of the HIV patients

Évaluation de la contamination bactériologique des eaux usées des stations d épuration du Grand Agadir (Isolement, caractérisation moléculaire et antibiorésistance des espèces du genre Vibrio)

Au Maroc et principalement dans la Wilaya d Agadir trois stations d épuration des eaux usées fonctionnant selon la technique d infiltration-percolation sur sable ont été mises en place. L objectif de cette étude est de déterminer les caractéristiques physico-chimiques, bactériologiques, ainsi que l étude des souches de Vibrio isolées de 3 stations d épuration de la ville. Les résultats d abattement des bactéries indicatrices de contamination fécale entre l entrée et la sortie des 3 stations d épuration dépassent les 99%. La qualité des eaux épurées issues des 3 STEPs échantillonnées Ben Sergao, Drarga et L Mzar, est de type A selon les recommandations de l OMS (1989), mais il faut que le nombre des oeufs d helminthes soit moins de 1/L pour une réutilisation en irrigation de façon non restrictive. Au cours de cette étude et après identification par le système Vitek 2 et/ou par PCR, sérotypage, 58 souches de Vibrio ont été isolées et analysées. Cette population est représentée par quatre espèces : 31 souches de V. cholerae non-O1, 17 souches de V. alginolyticus, 9 souches de V. fluvialis et une souche de V. metschnikovii. 53,44% des souches de Vibrio isolées ont un phénotype sauvage, 46,56% des souches présentent le phénotype pénicillinase à bas niveau ou phénptype céphalosporinase à bas niveau. L étude des profils de migration électrophorètique de l ADN total après macro restriction enzymatique par NotI, a révélé un degré important d hétérogénéité des souches de Vibrio. La spectrométrie de masse MALDI TOF a été utilisée pour identifier et classer des souches de Vibrio. La qualité d identification par cette technique est fonction de la richesse de la banque de spectres enregistrés.In countries such as Morocco, characterized by scarce rainfall, lack of freshwater resources and high groundwater salinity where agriculture reuse of municipal wastewater is becoming a compulsory choice for water resources management. This study evaluated the efficiency of the three wastewater treatment plants (WTPs) for the removal of physico-chemical and microbiological contaminants. Resistance to antibiotics, pulsed field gel electrophoresis and MALDI-TOF mass spectrometry were used to characterize Vibrio isolates. Mean values were used to determine treatment performances of the 3 WTPs, electric conductivity didn t showed any difference between raw and treated wastewater and the mean values varying from 2900 S/cm to 3300, the pH varying from 6,66 to 8,6, and the temperature varying between 16C and 26.4C. Removal efficiencies of fecal coliforms, enterococci and spores of sulphite reducing anaerobic bacteria were betwenn 3 and 4 log unit for the 3WTPs. 4 species of Vibrio were identified, among the 58 Vibrio sp. isolated, 53,44% were identified as V. cholerae, 29,31% as V. alginolyticus, 17,78% as V. fluvialis and 1,74% as V. metschnikovii. Of the total 58 Vibrio isolates, 53,44% were susceptible to all antibiotics. Of the resistant (46,56%) Vibrio strains, 39,83% were resistant against one to three antibiotics. PFGE with NotI digestion produced patterns with higher level of heterogeneity, and about 31% of Vibrio isolates were untypeable. MALDI-TOF-MS-based fingerprinting of Vibrio isolates has potential as a rapid for identification and finest differences between strains can readily be evaluated by the dendrogram based on percentage identity of MALDI-TOF mass spectra of Vibrio isolates, but requires further development for database of BioTyper.STRASBOURG-Sc. et Techniques (674822102) / SudocSudocFranceF

High-resolution melting analysis for rapid characterization of qnr alleles in clinical isolates and detection of two novel alleles, qnrB25 and qnrB42.

International audienceOBJECTIVES: qnr genes are plasmid-mediated quinolone resistance genes. Five qnr families have been described with several alleles (7 alleles of qnrA, 53 alleles of qnrB, 1 allele of qnrC, 1 allele of qnrD and 6 alleles of qnrS). Their detection requires a PCR specific for each qnr family and further sequencing for allele characterization. METHODS: High-resolution melt curve analysis (HRMA) was coupled to multiplex and simplex real-time PCR assays for detection and characterization of qnrA, qnrB and qnrS alleles. The protocol was set using 27 reference strains harbouring the most frequent alleles and was applied to 55 clinical isolates unknown for qnr positivity. RESULTS: Out of the 27 reference strains tested, 21 alleles showed distinct profiles using HRMA: 6 qnrA, 12 qnrB and 3 qnrS. For the qnrB alleles showing similar profiles, we gathered them into four groups that were easily distinguished. For the alleles that we could not test, in silico analysis showed that they would be identified using the HRMA protocol set. Among the clinical isolates, 28 qnr-positive isolates were detected and the qnr allele was characterized as 8 qnrA1, 4 qnrB1, 5 qnrB2, 3 qnrB4, 1 qnrB8, 1 qnrB5, 3 qnrS1 and 1 qnrS2, with concordant results with PCR sequencing. Two new qnrB alleles were detected and distinguished using HMRA. They were further designated as qnrB25 and qnrB42. CONCLUSIONS: We developed an HRMA assay for characterizing the qnr alleles in clinical isolates. This high-throughput method can be used to screen a large number of isolates. This method allowed the detection of new qnrB alleles

Pulsed field gel electrophoresis (PFGE) dependent dendogram of isolated <i>Staphylococcus aureus</i>.

<p>Pulsed field gel electrophoresis (PFGE) proves no specific clonal relationship between PVL-producing isolates issued from furuncles or secondary dermatosis. The similarity of the different pulsotypes was established by using Molecular Analyst™ software. Twenty four pulsotypes corresponded to the 55 isolates and their distribution is given according to the groups of isolates issued from secondary infected dermatosis (1), furuncles from HIV (−)(2) or HIV (+)(3) patients.</p

Production of toxins and identification of genes encoding toxins in <i>Staphylococcus aureus</i>.

<p>The majority of <i>SA</i> strains isolated from HIV patient-derived furuncles significantly produced PVL (<i>p</i><0.05), whereas only 10% of <i>SA</i> strains produced this toxin in secondary infected dermatosis. A high prevalence of LukE-LukD-producing isolates (56 to 78%) was recorded in patient groups.</p

Primers used for PCR detection of genes encoding virulence factors.

<p>Other virulence factors have been detected by multiplex PCR onto purified total DNA (Qiagen). Presence of genes encoding enterotoxins E, G, H, K, L, T, epidermolysin D (Etd), genes encoding Edin A and EDIN B and the genes encoding seven adhsesion factors (Cna, FnbA, FnbB Bbp, Clfb, Fib, Ebp and Lbp) was checked in 8 set in function of base size.</p