Genotipicação de Clostridium perfringens isolados de frangos de corte através da PCR múltipla

Abstract

Clostridium perfringens (Cp) is an anaerobic gram-positive bacterium which causes gaseous gangrene and enterotoxaemias in humans and domestic animals, besides being the primary cause of necrotic enteritis in poultry. Cp isolates were preliminary identified according to the lecithinase test on agar TSC-egg yolk, colony with double haemolysis in desfibrinated horse blood agar, Gram staining and biochemical tests. Cp isolates (171) were obtained from the intestinal content of broiler chickens sampled in a slaughterhouse in Pará de Minas city, MG, Brazil. Cp was isolated in 62/125 (49.6%) strains from jejunum content and in 109/125 (87.2%) of ileum. Cp strains were classified into five toxigenic types (A-E), using multiplex PCR assay for genotyping of the principal and lethal toxins in the detection of genes coding for toxins alfa, beta, epsilon e iota, nomely genes cpa, cpb, etx e iA genes, beta2 toxin (cpb2) and enterotoxin (cpe). From a total of 62 Cp jejunum isolates obtained 42/62 (67.7%) were type A, 1/62 (1.6%) type A with the amplification of products for beta2 toxin gene (A/B2), 0/62 (0%) type B, 17/62 (27.4%) type C and 1/62 (1.6%) type D. A total of 109 ileum Cp isolates were obtained being 62/109 (56.9%) type A, 3/109 (2.7%) type A/B2 toxin gene, 1/62 (0.9%) type B, 38/109 (34.9%) type C, 1/109 (0.9%) type D. Cp A (60.8%) and Cp C (32.2%) toxigenic types were the most prevalent types in the analyzed intestinal contents of broiler chickens Cp A 104/171 (60.8%) and 55/171 (32.2%) toxigenic types which were the most prevalent types analyzed into two partes of the intestinal content of broiler chickens. Five (2.9%) out of 171 Cp isolates were not typified. The iota toxin (iA) and enterotoxin gene (cpe) codifying genes were not identified.