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Previous issue date: 2010-03-31 / Objetivo geral: Caracterizacao fenotipica e genotipica de S. aureus resistentes a oxacilina (MRSA), coletados de hemoculturas provenientes de diversos centros participantes do projeto SCOPE - Brasil, no periodo de Junho de 2007 a Julho de 2009. Objetivos especificos: (i) avaliar a distribuicao dos tipos de SCCmec atraves das tecnicas de PCR convencional e PCR multiplex das diferentes instituicoes participantes; (ii) detectar a possivel presenca do gene lukF codificador da toxina PVL (Panton Valentine Leukocidin); (iii) avaliar a similaridade genetica das amostras utilizando as tecnicas de PFGE e MLST e (iv) determinar as concentracoes inibitorias minimas a oxacilina e vancomicina. Material e Metodos: Foram avaliados 62 isolados de MRSA provenientes do primeiro episodio de infeccao de corrente sanguinea (ICS) de pacientes hospitalizados em 11 centros medicos de diferentes regioes do pais. Os isolados resistentes a oxacilina foram submetidos a PCR convencional para a deteccao do gene nuc e lukF e PCR multiplex para identificar os tipos de SCCmec. Foi feito diluicao em agar e EtestR para vancomicina, PFGE e MLST. Resultados: Do total de 62 amostras, 29 amplificaram para o SCCmec tipo III, 16 amplificaram para o SCCmec tipo II, 9 amostras amplificaram para o SCCmec tipo IV, 6 amplificaram para SCCmec tipo I e 2 amostras nao foram identificadas pelas metodologias de PCR multuplex utilizadas. A amostra com SCCmec subtipo IVc apresentou o gene que codifica PVL. As amostras com SCCmec tipo I, II e III apresentaram CIM >256 ƒÊg/ml para oxacilina por EtestR com excecao de uma delas com SCCmec tipo II com CIM igual a 128 ƒÊg/ml. Seis amostras do total de nove com SCCmec tipo IV apresentaram CIM.48 ƒÊg/ml. Observamos CIMs de 1,0, 1,5 e 2,0 ƒÊg/m para vancomicina por EtestR. Quando feito a diluicao em agar, duas amostras tiveram CIM igual a 2,0 ƒÊg/ml, todas as outras apresentaram valores entre 0,5 e 1,0 ƒÊg/ml. Dentre as 8 amostras de MRSA que foram submetidas a tecnica de MLST, verificou-se ST105 em amostras com SCCmec tipo I; ST5 e ST105 para amostras com SCCmec tipo II; ST239 para amostras carreando SCCmec tipo III e ST5 e ST1176 relacionados a amostras apresentando SCCmec tipo IV. Conclusoes: Houve predominio de amostras de MRSA carreadoras de SCCmec tipo III e relacionadas geneticamente ao clone CEB. Foram detectadas amostras portando SCCmec tipo II e IV relacionados aos clones Nova Iorque/Japao e Pediatrico em diferentes hospitais e regioes do pais e ausencia do clone Chile/Cordoba. Apenas uma amostra com SCCmec tipo IV (subtipo IVc) nao relacionada a nenhum clone foi produtora de PVL. Apenas dois ancestrais geneticos comuns (CC5 e CC8) nas amostras estudadas foram observados pela tecnica de MLST, sendo caracterizado um novo ST1176. Nao foram detectadas amostras resistentes a vancomicina. / General objective: Phenotypic and genotypic characterization of methicillin-resistant S. aureus (MRSA) collected from bloodcultures from several centers participants of Brazilian SCOPE Project, from June 2007 to July 2009. Specific Objectives: (i) to assess the different types of SCCmec distribution through conventional and multiplex PCR for the different participant institutions; (ii) to detect the possible presence of lukF gene, which codes for PVL toxin (Panton Valentine Leukocidin); (iii) to assess the genetic similarity of these samples through PFGE and MLST and (iv) to determine the oxacillin and vancomycin minimal inhibitory concentration (MIC). Material and Methods: Sixty two MRSA isolated from the first episode of bloodstream infection (BSI) were evaluated. These samples came from patients hospitalized at the 11 medical centers from different regions of the country. The oxacillin-resistant isolated were submitted to conventional PCR to detect nuc and lukF genes and to multiplex PCR to identify SCCmec types. Agar dilution and E-test for vancomycin were performed. The strains were molecular typed by PFGE and MLST. Results: From the total of 62 samples, 29 amplified SCCmec type III, 16 SCCmec type II, 9 SCCmec type IV, 6 SCCmec type I and 2 could not be identified. A sample with SCCmec subtype IVc carried the gene which codifies for PVL. Samples with SCCmec types I, II and III showed MIC > 256 ƒÊg/ml for oxalicin by EtestR. Just one of them, with SCCmec type II, presented MIC = 128 ƒÊg/ml. From the nine samples with SCCmec type IV, six presented MIC . 48 ƒÊg/ml. MICs of 1.0, 1.5 e 2.0 ƒÊg/ml were also observed for vancomycin by EtestR. Regarding the agar dilution, two samples presented MICs of 2.0 ƒÊg/ml and all the others showed values between 0.5 e 1.0 ƒÊg/ml. From the 8 MRSA samples typed by MLST, it was observed ST105 in a sample with SCCmec type I; ST5 and ST105 in samples with SCCmec type II, ST239 with SCCmec type III and ST5 and ST1176 with SCCmec type IV. Conclusion: The majority of samples were MRSA carrying SCCmec type III and genetically related to CEB clone. Also were detected samples SCCmec type II and IV, related to New York/Japan and Pediatric clones in different hospitals at different regions of the country. Only one sample SCCmec type IV (subtype IVc) not related to any clone was positive for PVL. Only two common genetic ancestral (CC5 and CC8) were observed through MLST and a new ST1176 was characterized. No vancomycin-resistant isolate was detected. / TEDE / BV UNIFESP: Teses e dissertações
| Identifer | oai:union.ndltd.org:IBICT/oai:repositorio.unifesp.br:11600/9825 |
| Date | 31 March 2010 |
| Creators | Paschoal, Loren [UNIFESP] |
| Contributors | Universidade Federal de São Paulo (UNIFESP), Pignatari, Antonio Carlos Campos [UNIFESP] |
| Publisher | Universidade Federal de São Paulo (UNIFESP) |
| Source Sets | IBICT Brazilian ETDs |
| Language | Portuguese |
| Detected Language | English |
| Type | info:eu-repo/semantics/publishedVersion, info:eu-repo/semantics/masterThesis |
| Format | 101 p. |
| Source | reponame:Repositório Institucional da UNIFESP, instname:Universidade Federal de São Paulo, instacron:UNIFESP |
| Rights | info:eu-repo/semantics/openAccess |
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