Direct PCR amplification of hilA gene as specific, sensitive and rapid procedure for detecting Salmonella in minced beef.

摘要

PCR is a useful and precisely working tool and therefore recommended for rapid detection of pathogens in food; however, it faces by the presence of some PCR inhibitors in foods. To overcome this problem, an additional non-selective enrichment step (post-enrichment) in Luria-Bertani (LB) medium for 4, 6, 8, 10 or 12 h was suggested in the present work after the selective enrichment step as described in ISO-procedure for detection of Salmonella in food stuff. Minced beef samples were artificially contaminated either with Salmonella sp. alone or with a population of different organisms including Salmonella. The supernatants of lysed minced beef samples (95 degrees C for 20 min) were used directly in the PCR as DNA source, without extraction and purification of bacterial chromosomal DNA. Two specific primers were applied in PCR for detecting hilA gene of Salmonella. The obtained results demonstrated that 8 h post enrichment is sufficient to detect Salmonella at contamination level of 10 cfu per 25 g minced beef. Sensitivity and specificity of the described procedure were also evaluated. The present study provides useful information for directly applying the rapid PCR procedures for detecting pathogens in foods without prior DNA extraction and purification.