Multiplex PCR for detection of virulence genes in Yersinia enterocolitica from food and faecal samples in San Luis city

FAVIER GABRIELA ISABEL; ESCUDERO MARÍA ESTHER; LUCERO ESTRADA CECILIA STELLA MARYS; STEFANINI DE GUZMÁN ANA MARÍA

San Luis

Jornada; XXIV Reunión Científica Anual de la Sociedad de Biología de Cuyo. IV Reunión Científica de la Sociedad Argentina de Microscopía (SAMIC).; 2006

Sociedad de Biología de Cuyo

Of the six biotypes (B) of Y. enterocolitica, five (1B, 2, 3, 4 and 5) are considered pathogenic in humans and contain markers associated with virulence. These are located on the chromosome (ail, inv, htrA) and on the pYV virulence plasmid (yadA, virF). Multiplex PCR assays for the detection of plasmid- and chromosome-borne virulence genes of Y. enterocolitica isolated from a variety of foods (skin chicken, bovine and pork meat) and human patients(faecal samples) in San Luis city were developed. Besides, the reference Y. enterocolitica strain W 1024 B2 0:9 was assayed. Specific PCR primers were obtained from sequences available in the GeneBank database (GeneDoc and Primer 3 software). Each multiplex PCR assay was performed in 50 ul of reaction mixture containing 3 ul of DNA, 1mM of dNTP mix, 1.5 mM of CI2Mg, 0.5 X reaction buffer, 10 pmol of each forward and reverse primer and 0.25 U of Taq DNA polymerase. The samples were amplified for 35 cycles, and each cycle consisted of 1.5 min at 94°C, 1.15 min at 58°C and 1.5 min at 72°C. Y. enterocolitica B2 strains were positive for virF and invA genes, while Y. enterocolitica B 1A strains were positive for htrA and yst genes. Multiplex PCR assays showed simultaneous amplification of virulence genes from different biotypes of Y. enterocolitica isolated from a variety of foods and human patients in San Luis city.