RESUMO
Background: Molecular characterization of staphylococcal cassette chromosome mec (SCCmec) types of methicillin-resistant Staphylococcus aureus (MRSA) is very essential for studying the epidemiology of MRSA. Objectives: This study reports two multiplex PCR for molecular typing of MRSA collected from Jeddah, Kingdom of Saudi Arabia. Materials and Methods: A total of 101 clinical isolates of strains were collected from major hospital laboratories and public health centres, Jeddah, Kingdom of Saudi Arabia during the period from August 2009 to May 2011. All the strains were tested phenotypically by conventional methods and genotypically by a novel multiplex PCR targeting at the same time S. aureus 16S rRNA, Panton - valentine leucocidin (PVL) and mecA resistance genes. All the strains were tested also by multiplex PCR for typing of SCC mec types. Results: All the 101 strains previously identified phenotypically as S. aureus with bacteriological examination were positive for amplification of 756 base pair fragments specific for 16S rRNA of S. aureus. Moreover, all the strains were positive for amplification of 1339 base pair fragments specific for mecA gene, while only 38 strains (37.6%) showed positive amplification of 433 base pair fragments specific for PVL gene. The most predominant SCC mec type among the examined isolates is type V 43 (42.5) followed by SCCmec type III 39 (38.6%). Conclusion: The newly modified multiplex PCR is rapid and sensitive method for detection of MRSA. Moreover, the most predominant SCC mec type among the examined isolates from Jeddah, King Saudi Arabia is type V (42.5%), followed by Type III (38.6%).
Assuntos
Proteínas de Bactérias/genética , Toxinas Bacterianas/genética , Exotoxinas/genética , Genótipo , Humanos , Leucocidinas/genética , Staphylococcus aureus Resistente à Meticilina/classificação , Staphylococcus aureus Resistente à Meticilina/genética , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Testes de Sensibilidade Microbiana , Tipagem Molecular/métodos , Reação em Cadeia da Polimerase Multiplex/métodos , Fenótipo , Arábia Saudita , Sensibilidade e Especificidade , Infecções Estafilocócicas/microbiologia , Fatores de TempoRESUMO
Fourteen isolates of Corynebacteruim pseudotuberculosis of them 7 were isolated from sheep with Caseous Lymphadenitis "biotype 1" and 7 isolated from buffaloes with Oedematous Skin Disease "biotype 2". All isolates were identified by standard microbiological techniques and by polymerase chain reaction targeting, 16S rRNA and phospholipase D genes. Synergistic haemolytic titers of all isolates were assayed by plate technique. The presences of phospholipase D gene in supernatants of all isolates were performed by sodium dodecyl sulfate polyacrylamide gel electrophoresis immunoblot technique by using hyperimmune serum raised in rabbit immunized with recombinant phospholipase D gene antigen. The concentration of phospholipase D gene was assayed by scanning the bound phospholipase D gene with specific antibodies that appeared at 31.5 kDa. Results presented that there is no correlation between titer of Synergistic haemolytic activity and the actual phospholipase D genes concentration in culture supernatants. Also results presented that Synergistic haemolytic activity and phospholipase D genes produced by biotype 2 (buffalo isolates) was generally higher than those by biotype 1(sheep isolates).
Assuntos
Animais , Bovinos , Coelhos , Infecções por Corynebacterium , Corynebacterium pseudotuberculosis/enzimologia , Corynebacterium pseudotuberculosis/isolamento & purificação , Fosfolipase D/genética , Fosfolipase D/isolamento & purificação , Regulação da Expressão Gênica , Técnicas In Vitro , Linfadenite , RNA , Búfalos , Eletroforese , Ativação Enzimática , Métodos , Coelhos , OvinosRESUMO
Purpose: Vancomycin-resistant enterococci (VRE) pose an emerging problem in hospitals worldwide. The present study was undertaken to determine the occurrence, species prevalence, antibacterial resistance, and phenotypic and genetic characteristics of VRE isolated in Riyadh hospitals, KSA. Materials and Methods: Two hundred and six isolates of enterococcal species were obtained from clinical samples. The antibiotic susceptibility of isolates and minimum inhibitory concentration (MIC) tests for vancomycin and teicoplanin were determined. Molecular typing of VRE isolates was carried out by using pulsed field gel electrophoresis (PFGE) and the resistance genotype was determined by polymerase chain reaction (PCR). Results: VRE accounted for 3.9% of the isolates and were detected mostly in urine, wound and blood specimens isolated from ICU, internal medicine and surgical wards. All strains were identified to species level and were found to consist of E. faecalis (69.2%), E. faecium (11.3%), E. avium (2.1%), E. hirae (0.8%), E. casseliflavus (1.3%) and E. gallinarum (1.3%) species. According to the susceptibility data obtained, 8 (3.9%) out of 206 isolates were found to be VRE (MICs > 32 μg/ml). The vanA, vanB and vanC gene fragments of E. faecalis, E. faecium and E. gallinarum were amplified from isolates and were detected. PFGE patterns of the VRE isolates revealed homogenous patterns with dominant clone suggesting that the strains intrinsic resistance is independent. Conclusions: This study shows an emergence of VRE along with increased rate of multidrug-resistant enterococci in the area of the study. Regular surveillance of antimicrobial susceptibilities should be done regularly and the risk factors should be determined.