ABSTRACT
INTRODUCTION: Staphylococcus spp. is an important healthcare-associated pathogen and the identification of methicillin-resistant strains in samples of colonization may provide data to assist in the antimicrobial therapy success. OBJECTIVES: To determine the occurrence of colonization by methicillin-resistant Staphylococcus spp. (MRS), through the detection of the mecA gene and to evaluate different phenotypic methods for the presumptive detection of methicillin resistance in samples of the anterior nasal cavity and hands of the health care personnel of a university hospital in the state of Pernambuco, Brazil. METHODS: We selected the 28 isolates of Staphylococcus spp., which showed an intermediate or resistant phenotypic profile for oxacillin, detected by the Kirby Bauer technique. The methods used were disk-diffusion tests for cefoxitin, minimal inhibitory concentration by E-test for oxacillin, screening for oxacillin resistance and mecA gene detection by polymerase chain reaction (PCR). RESULTS: About the phenotypic methods utilized, only the E-test of oxacillin did not show a statistically significant difference in relation to PCR for the mecA gene detection, considered the gold standard. CONCLUSION: The E-test of oxacillin was the best of the phenotypic methods utilized. It is necessary to correctly detect MRS in healthy individuals, because they can act as carriers and can therefore be a potential source of microorganisms involved in hospital infections.
INTRODUÇÃO: Staphylococcus spp. é um importante patógeno associado aos cuidados em saúde, e a identificação de isolados resistentes à meticilina em amostras de colonização pode fornecer dados para auxiliar no sucesso da terapia antimicrobiana. OBJETIVOS: Determinar a ocorrência de colonização por Staphylococcus spp. resistentes à meticilina (MRS) por meio da detecção do gene mecA e avaliar diferentes métodos fenotípicos para a detecção presuntiva da resistência à meticilina em amostras da cavidade nasal anterior e das mãos de profissionais de saúde de um hospital universitário no Estado de Pernambuco, Brasil. MÉTODOS: Foram selecionados 28 isolados de Staphylococcus spp. que mostraram perfil intermediário ou resistente à oxacilina, detectado pela técnica de Kirby Bauer. Os métodos utilizados foram o teste de disco difusão de cefoxitina, concentração inibitória mínima pelo E-test de oxacilina, screening para avaliação da resistência à oxacilina e reação em cadeia da polimerase (PCR) para detecção do gene mecA. RESULTADOS: Dos métodos fenotípicos utilizados, apenas o E-test de oxacilina não mostrou diferença estatística significante em relação à PCR para a detecção do gene mecA, considerado o método padrão-ouro. CONCLUSÃO: O E-test de oxacilina foi o melhor método fenotípico utilizado. É necessário detectar corretamente o MRS em indivíduos saudáveis, pois eles podem atuar como portadores, sendo uma fonte potencial de microrganismos envolvidos em infecções hospitalares.
Subject(s)
Humans , Health Personnel , Methicillin Resistance , Polymerase Chain Reaction , Staphylococcus epidermidis , Methicillin-Resistant Staphylococcus aureus/isolation & purificationABSTRACT
Twenty-eight Klebsiella pneumoniae clinical isolates that exhibited an extended-spectrum cephalosporin-resistance profile from a city in the Northeast of Brazil were analysed by PCR and DNA sequencing in order to determine the occurrence of blaCTX-M genes and class 1 integrons. We determined the occurrence of the blaCTX-M-2 gene in six K. pneumoniae isolates and describe the first detection of the blaCTX-M-28 gene in South America. Seven isolates carried class 1 integrons. Partial sequencing analysis of the 5'-3'CS variable region in the class 1 integrons of three isolates revealed the presence of aadA1, blaOXA-2 and dfr22 gene cassettes.
Subject(s)
Humans , DNA, Bacterial/genetics , Integrons/genetics , Klebsiella pneumoniae/genetics , beta-Lactamases/genetics , Anti-Bacterial Agents/pharmacology , Base Sequence , Brazil , Drug Resistance, Bacterial/genetics , Klebsiella pneumoniae/drug effects , Klebsiella pneumoniae/enzymology , Molecular Sequence Data , Polymerase Chain Reaction , Sequence Analysis, DNAABSTRACT
PCR analysis of 16S-23S internal transcribed spacer (PCR ribotyping) and tRNA intergenic spacer (tDNA-PCR) were evaluated for their effectiveness in identification of clinical strains of Klebsiella pneumoniae and differentiation with related species. For this purpose both methods were applied to forty-three clinical isolates biochemically identified as K. pneumoniae subsp. pneumoniae isolated from patients clinical specimens attended at five hospitals in three Brazilian cities. References strains of K. pneumoniae subsp. pneumoniae, K. pneumoniae subsp. ozaenae, K. oxytoca, K. planticola and Enterobacter aerogenes were also analyzed. Both PCR methods showed specific patterns for each species. A conserved PCR ribotype pattern was observed for all clinical K. pneumoniae isolates, while differing from other related analyzed species. tDNA-PCR revealed five distinct patterns among the K. pneumoniae clinical isolates studied, demonstrating a predominant group with 90,6 percent of isolates presenting the same pattern of K. pneumoniae type strain. Both PCR-based methods were not able to differentiate K. pneumoniae subspecies. On the basis of the results obtained, both methods were efficient to differentiate the Klebsiella species analyzed, as well as E. aerogenes. Meanwhile tDNA-PCR revealed different tRNA arrangements in K. pneumoniae, suggesting intra-species heterogeneity of their genome organization, the polymorphism of the intergenic spacers between 16S and 23S rRNA genes appears to be highly conserved whithin K. pneumoniae clinical isolates, showing that PCR ribotyping can be an useful tool for identification of K. pneumoniae isolates.