Real-Time PCR Method for Detection of Salmonella spp. in Environmental Samples.

The methods currently used for detecting in environmental samples require 2 days to produce results and have limited sensitivity. Here, we describe the development and validation of a real-time PCR screening method that produces results in 18 to 24 h. Primers and probes specific to the gene , group D, and serovar Enteritidis organisms were designed and evaluated for inclusivity and exclusivity using a panel of 329 isolates representing 126 serovars and 22 non- organisms. The - and group D-specific sets identified all the isolates accurately. The PCR method had 100% inclusivity and detected 1 to 2 copies of DNA per reaction. Primers specific for -differentiating fragment 1 (Sdf-1) in conjunction with the group D set had 100% inclusivity for 32 Enteritidis isolates and 100% exclusivity for the 297 non-Enteritidis isolates. Single-laboratory validation performed on 1,741 environmental samples demonstrated that the PCR method detected 55% more positives than the itek mmunoiagnostic ssay ystem (VIDAS) method. The PCR results correlated well with the culture results, and the method did not report any false-negative results. The receiver operating characteristic (ROC) analysis documented excellent agreement between the results from the culture and PCR methods (area under the curve, 0.90; 95% confidence interval of 0.76 to 1.0) confirming the validity of the PCR method. This validated PCR method detects 55% more positives for in half the time required for the reference method, VIDAS. The validated PCR method will help to strengthen public health efforts through rapid screening of spp. in environmental samples.