Evaluation of culture enrichment procedures for use with Salmonella detection immunoassay

Abstract

To design efficient culture strategies for use with immunoassays to detect Salmonella in food, the growth of these organisms was investigated according to the Bacteriological Analytical Manual (BAM) and enrichment-immunoassay (EI) culture procedures. The cultures were further evaluated using a commercial enzyme-linked immunosorbent assay (ELISA) kit . The BAM procedure includes pre-enrichment in nutrient broth (NB) for 16 h followed by selective enrichment in either Rappaport–Vassiliadis (RV) or tetrathionate brilliant green (TBG) broth for 16 h. The EI procedure includes pre-enrichment in NB for 4 h, selective enrichment in RV for 16 h and post-enrichment in NB for 4 h. The effects of different incubation times for pre- and post-enrichment, and different culture media for selective enrichment (TBG and RV) and post-enrichment in NB and Brain Heart Infusion broth (BHI) on the growth of the bacteria and ELISA titers in the EI procedure were also investigated. Salmonella enteritidis and S. typhimurium inoculated at different initial concentrations between 0.1 and 35 CFU/ml grew to similar concentrations of 10 7 to 10 8 colony forming unit (CFU)/ml in pure culture and generally 2 to 4 fold lower concentrations ( P<0.05) in mixed culture using spiked chicken rinse. In the BAM procedure, the concentration of Salmonella cultured in RV was higher ( P<0.01) than that in TBG. The cultures in TBG showed positive results for ELISA, but those in RV were generally negative. In the EI procedure, the ELISA titers from cultures post-enriched in NB or BHI were higher ( P<0.01) when TBG, as compared to RV, was used for selective enrichment. Post-enrichment in BHI yielded higher numbers of Salmonella and higher ELISA titers than those in NB ( P<0.05) for post-enrichment. This study demonstrated that in both culture procedures small numbers of Salmonella could be increased to at least 10 7 CFU/ml which is detectable by most ELISAs, and that the type of the culture media used may have a significant impact on ELISA results.