Isolation and phenotypic and molecular Identification of thermophilic species of Campylobacter from chilled chicken carcasses

Nas últimas quatro décadas espécies de Campylobacter spp. têm sido reconhecidas como patógenos emergentes e despontaram como importantes agentes de gastrenterites de origem alimentar em várias partes do mundo. No Brasil, cabe aos órgãos normatizadores e à Vigilância Sanitária assegurar o cumprimento das legislações a fim de garantir a segurança dos alimentos comercializados e preservar a saúde do consumidor. O objetivo deste estudo foi verificar a ocorrência das espécies termofílicas de Campylobacter em 30 amostras de carcaças resfriadas de frango adquiridas no comércio do Município do Rio de Janeiro. A preparação das amostras e o isolamento de colônias suspeitas de Campylobacter spp. foram realizados de acordo com os protocolos descritos no Compendium of Methods for the Microbiological Examination of Foods, APHA. A confirmação de Campylobacter spp. foi feita pelo teste de aglutinação em látex e a diferenciação das espécies foi realizada pela bioquímica convencional e pela abordagem molecular. Foi proposto, neste estudo, a utilização da Duplex PCR para amplificar os genes 16S rRNA e o gene da oxirredutase, para a detecção das espécies do gênero Campylobacter e da espécie C. jejuni respectivamente. O plaqueamento direto das amostras, utilizando meios de cultivo seletivos, foi mais eficiente no isolamento de Campylobacter spp. do que a etapa de enriquecimento seletivo.During the last four decades, different species of Campylobacter spp. have been recognized as emerging pathogens and important agents of gastroenteritis from food source worldwide. In Brazil, health authorities are responsible for the surveillance of foods in order to preserve the health of the population. The aim of this study was to determine the proportion of thermophilic Campylobacter species among 30 chilled chicken carcasses samples purchased in the Rio de Janeiro City. Sample preparation and isolation of suspected Campylobacter spp. colonies were performed according to the protocols described in the Compendium of Methods for the Microbiological Examination of Foods, APHA. Campylobacter spp. confirmation was performed with the latex agglutination test and species differentiation was carried out through biochemistry tests and molecular approach. A Duplex-PCR approach has been proposed through amplification of 16S rRNA and oxidoreductase genes, for the detection of the genera Campylobacter and C. jejuni respectively. Direct plating of samples using selective culture media, was more efficient for the isolation of Campylobacter spp. then the selective enrichment step. We detected the presence of Campylobacter spp. in 21 samples (70%), six (28,6%) from slaughterhouses, eight (38,1%) from supermarkets and seven (33,3%) from street markets. Two (9,6%) out of 21 isolates were identified as C. coli, 18 (85,7%) as C. jejuni and one isolate showed after 7 days an ambiguous result by biochemical tests. Duplex PCR confirmed the biochemical results for C. coli and detected 19 (90,48%) C. jejuni after 4 hours. Our results suggest that the implementation of rapid and reliable methods for the detection of this food pathogen in poultry may contribute for the improvement of food surveillance systems and public health surveillance