DETECTION OF SALMONELLA SPP., ESCHERICHIA COLI O157, LISTERIA MONOCYTOGENES AND CAMPYLOBACTER SPP. IN CHICKEN SAMPLES BY MULTIPLEX POLYMERASE CHAIN REACTION AND HYBRIDIZATION USING THE GENEGEN MAJOR FOOD PATHOGENS DETECTION KIT

Abstract

ABSTRACT We have applied an innovative commercial test system (the GeneGen Major Food Pathogens [MFP] Detection Kit) to detect four of the more important pathogenic microorganisms in food, Salmonella spp., Escherichia coli O157, Listeria monocytogenes and Campylobacter spp. by using multiplex polymerase chain reaction (PCR) combined with a shortened, simplified hybridization protocol on test strips. Testing 129 strains representing 23 different genera yielded no false‐positive or false‐negative results, and the limit of detection was 100 genome copies of each organism per multiplex PCR using purified genomic DNA. The kit was used for both analyses of chicken samples artificially contaminated with different numbers of the pathogens and of naturally contaminated samples. The results were compared with those obtained with standard microbiological techniques. Both methods yielded the same results in spiking experiments, detecting 1–10 cfu/25 g of the respective pathogen added. It was possible to verify the presence of all four pathogens simultaneously in one assay. The GeneGen MFP Detection Kit saves at least 2–4 days compared with standard methods. The protocol is easy to perform, and there is no need for any additional costly and sophisticated equipment, except for a thermocycler, because hybridization is carried out at ambient temperature and the results can be evaluated with the unaided eye. This enables the assay to be performed in relatively modestly equipped laboratories for both DNA‐based screening of food samples and/or confirmation of suspect colonies.