Development and Verification of a Poultry Management Tool to Quantify Salmonella from Live to Final Product Utilizing RT-PCR.

The United States Department of Agriculture Food Safety and Inspection Service (USDA FSIS) does not maintain a zero-tolerance policy for Salmonella in poultry and poultry products, despite being a known food safety hazard throughout the poultry industry. In 2016, USDA FSIS established performance standards for a 52-week moving window with the maximum acceptable percent positive for comminuted turkey (325 g sample) at 13.5% (7 of 52 samples). Based upon FSIS verification sampling results from one 52-week moving window, the Salmonella prevalence for each poultry establishment in category 1 (below limit), 2 (meeting limit), or 3 (exceeding limit) are published for public viewing. Moreover, many poultry producers continue to have post-intervention samples test positive. Therefore, the use of quantification would be more valuable to determine the efficacy of process control interventions, corrective actions, and final product Log CFU/g of Salmonella to make rapid, within shift, food safety decisions. Therefore, the objectives of these studies are to develop, verify, and validate a rapid and reliable quantification method utilizing RT-PCR to enumerate Salmonella in the poultry industry from flock to final product and to utilize the method in an application study. BAX® System SalQuant® is an application of the BAX® System Real-Time PCR Assay for Salmonella to enumerate low levels of Salmonella with shortened enrichment times. Curve development encompassed inoculating poultry matrix samples at four levels with an ATCC strain of Salmonella, with three biological replicates per inoculation level, and five technical replicates being run on the BAX® System for various timepoints, gathering the data, and creating a linear-fit equation. A linear-fit equation was provided for each timepoint. The ideal timepoint, based on the statistical parameters surrounding the equation (R2 > 0.80, Log RMSE < 0.60, and enumerable range 0.00 to 4.00 Log CFU/mL (g)) that most accurately estimate Salmonella compared to most probable number (MPN), was chosen to be utilized for further studies.

Application of a Commercial Salmonella Real-Time PCR Assay for the Detection and Quantitation of Salmonella enterica in Poultry Ceca.

ABSTRACT: Foodborne salmonellosis is commonly associated with poultry and poultry products, necessitating continued development of pre- and postharvest food safety interventions and risk management strategies. Evaluation of technologies and strategies is limited by availability of cost-effective, rapid laboratory methods. The objective of this study was to evaluate a commercial qualitative PCR assay and its novel quantitative application to detect and enumerate Salmonella in poultry ceca as an analytical matrix. Ceca were collected at harvest, the contents were homogenized, and paired samples were evaluated with buffered peptone water (BPW) and BAX MP + Supplement (MPS) preenrichment broths followed by PCR screening with a BAX System Q7 PCR and by culture isolation. Additional ceca were inoculated with Salmonella to develop a standard curve for the BAX System SalQuant quantitative PCR application (QA), and estimates were obtained by the QA and most-probable-number (MPN) methods. For preenrichment media, PCR outcomes were equivalent to those of culture isolation for detecting Salmonella in ceca with 95.65 and 87.88% sensitivity and 82.00 and 100.00% specificity (P = 0.074) for BPW and MPS, respectively. However, at the sample level, BPW performed significantly worse (47.92%) than did MPS (68.75%) for overall isolation of Salmonella (P < 0.0001). After standard curve development, the mean QA estimates obtained for the inoculated samples were 1.14 (95% confidence interval [CI]: 0.62 to 1.66), 1.79 (1.50 to 2.08), 2.91 (2.65 to 3.17), and 3.76 (3.26 to 4.25) log CFU/mL for each targeted inoculation of 1.0, 2.0, 3.0, and 4.0 log CFU/mL, respectively, and were within or comparable to the 95% CI values of paired MPN estimates. These data support the use of MPS for the detection and isolation of Salmonella enterica from poultry ceca when screening with PCR and indicate that QA may be useful as an alternative tool to estimate Salmonella loads in poultry ceca, which may support preharvest food safety interventions.

Effect of deboning time on the growth of Salmonella, E. coli, aerobic, and lactic acid bacteria during beef sausage processing and storage.

The objective of the current study was to determine the effects of deboning time, three steps of sausage processing (grinding, salting, and batter formulation), and storage time (of raw materials and cooked sausage) on the growth (log CFU/g) of aerobic bacteria, lactic acid bacteria, and inoculated Salmonella and E. coli. Beef deboning time did not influence bacterial counts (P≥0.138). However, salting of raw ground beef resulted in a 0.4-log reduction in both aerobic plate count (APC) and Salmonella (P≤0.001). Lactic acid bacteria were increased from non-detectable concentration (0.54 log) on d 0 to 3.8 log on d 120 of vacuum storage (P≤0.019). Salmonella counts were increased (P<0.001) over storage time (3.2 to 3.3 log CFU/g from d 0 to 10). Results indicated that salting and batter formulation had a greater impact on bacterial counts than rigor state of raw beef.