Purpose: A new and rapid real-time PCR method was developed and evaluated against the FDA BAM culture method for detection of Salmonella Enteritidis in whole shell eggs.
Methods: Approximately 250 eggs were combined into a bulk sample for the uninoculated control, and a bulk lot consisting of approximately 900 eggs were inoculated with SE (ATCC 13706) at a concentration of 0.2-2 CFU per 1,000 g. Egg pools consisting of 20 eggs (1,000 g) were prepared. Twenty inoculated and 5 uninoculated egg pools were enriched according to FDA BAM. A second set of 20 inoculated and 5 uninoculated egg pools were supplemented with 100 ml of 10X TSB per pool, and then incubated at 35°C for 24 hours. Real-time PCR was performed on the 7500 Fast using standard conditions (95°C for 10 min; 40 cycles at 95°C for 15 seconds and 60°C for 60 seconds).
Results: Methods comparison showed that the real-time PCR method was equivalent to the FDA BAM method. For two independent studies, chi-square was 0 and 0.41. In addition, the two methods were compared on samples enriched according to the FDA BAM method on day 5 following overnight enrichment in TSB plus ferrous sulfate. Chi-square analysis indicated no difference between the two methods (χ2= 0 on two independent studies). No false-positive or false-negative results were observed using either enrichment method. Sample preparation was designed to be completely automated to give results in approximately 3 hours post enrichment.
Significance: The assay evaluated here was shown to be equivalent in performance to the FDA BAM culture procedure for the detection of Salmonella in eggs. Real-time PCR detection of SE in whole egg pools is simple, rapid, and more cost effective than the standard FDA BAM culture method.