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.
Performances of two commercial test systems for the detection of Salmonella spp. and genus Listeria/Listeria monocytogenes (GeneGen Salmonella Detection Kit and GeneGen Listeria Detection Kit, respectively) were tested. The technology combines multiplex polymerase chain reaction with a shortened, simplified hybridization protocol using test strips. In specificity tests using a collection of over 80 bacterial strains representing 22 genera, no false positive or false negative results were observed. The limit of detection was 100 and 10 genome copies for the Salmonella and Listeria assays, respectively. The kits were used for both analyses of spiked and naturally contaminated chicken samples, and their performances were compared with standard methods. An amount of 1–10 cfu/25 g of the respective pathogen added could be detected. Results achieved with the GeneGen Salmonella Detection Kit were in 100% concordance with the microbiological protocol. The GeneGen Listeria Detection Kit proved to be more sensitive than the standard method in the spiking experiments, as well as for chicken samples naturally contaminated with both Listeria innocua and L. monocytogenes. The kits provided a time saving of several days compared to conventional methods (30‐h detection time for Salmonella, 30–54 h for Listeria). The protocol is simple and does not require any high‐tech equipment for evaluation or documentation after DNA amplification. Hybridization can be carried out at ambient temperature, and results are evaluated with the unaided eye. Thus, these kits could be a valuable tool for the screening of food samples and confirmation of suspect colonies, especially in modestly equipped laboratories.
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