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Evaluation of alternative detection methods for foodborne pathogens typically involves comparisons against a “gold standard” culture method, which may produce false-negative (FN) results, particularly under worst-case scenarios such as low contamination levels, difficult-to-detect strains, and challenging food matrices (e.g., matrices with a water activity of <0.6). We used extended enrichment times (up to 72 h for both primary and secondary enrichments) to evaluate a gold standard method for Salmonelladetection (the U.S. Food and Drug Administration Bacteriological Analytical Manual[BAM] method) in two low-water-activity foods (dry pet food and chocolate) inoculated at low contamination levels (most probable number ca. 1/25 g) with five Salmonellastrains. Strains were selected to include those with a poor ability to grow in enrichment media. Among the 100 pet food and 100 chocolate samples tested, 53 and 50, respectively, were positive with the standard BAM method, and 57 and 59, respectively, were positive with the extended BAM method. Thus, the FN probabilities for the standard BAM method were 7% for pet food and 15% for chocolate. An alternative enzyme immunoassay method for detection of Salmonellain chocolate produced FN probabilities of 6 and 20% when compared against the standard and extended BAM methods, respectively. Detection of SalmonellaMississippi was significantly reduced with the alternative method (P= 0.023) compared with the extended BAM method. We calculated a composite reference standard to further define FN probabilities based on variable results from multiple assays (the standard BAM, extended BAM, and alternative methods). Based on this standard, the enzyme immunoassay for Salmonelladetection in chocolate had a 28% FN probability and the standard and extended BAM methods had 23 and 9% FN probabilities, respectively. These results provide a framework for how inclusion of extended enrichment times can facilitate evaluation of alternative detection methods.

Management of Shiga toxin–producing Escherichia coli(STEC), including E. coliO157:H7, in food products is a major challenge for the food industry. Several interventions, such as irradiation, chemical disinfection, and pasteurization, have had variable success controlling STEC contamination. However, these interventions also indiscriminately kill beneficial bacteria in foods, may impact organoleptic properties of foods, and are not always environmentally friendly. Biocontrol using bacteriophage-based products to reduce or eliminate specific foodborne pathogens in food products has been gaining attention due to the specificity, safety, and environmentally friendly properties of lytic bacteriophages. We developed EcoShield PX, a cocktail of lytic bacteriophages, that specifically targets STEC. This study was conducted to examine the efficacy of this bacteriophage cocktail for reducing the levels of E. coliO157:H7 in eight food products: beef chuck roast, ground beef, chicken breast, cooked chicken, salmon, cheese, cantaloupe, and romaine lettuce. The food products were challenged with E. coliO157:H7 at ca. 3.0 log CFU/g and treated with the bacteriophage preparation at ca. 1 × 106, 5 × 106, or 1 × 107PFU/g. Application of 5 × 106and 1 × 107PFU/g resulted in significant reductions (P< 0.05) in E. coliO157:H7 levels of up to 97% in all foods. When bacteriophages (ca. 1 × 106PFU/g) were used to treat lower levels of E. coliO157:H7 (ca. 1 to 10 CFU/10 g) on beef chuck roast samples, mimicking the levels of STEC found under real-life conditions in food processing plants, the prevalence of STEC in the samples was significantly reduced (P< 0.05) by ≥80%. Our results suggest that this STEC-targeting bacteriophage preparation can result in significant reduction of both the levels and prevalence of STEC in various foods and, therefore, may help improve the safety and reduce the risk of recalls of foods at high risk for STEC contamination.

Although wildlife intrusion and untreated manure have been associated with microbial contamination of produce, relatively few studies have examined the survival of Escherichia colion produce under field conditions following contamination (e.g., via splash from wildlife feces). This experimental study was performed to estimate the die-off rate of E. colion preharvest lettuce following contamination with a fecal slurry. During August 2015, field-grown lettuce was inoculated via pipette with a fecal slurry that was spiked with a three-strain cocktail of rifampin-resistant nonpathogenic E. coli. Ten lettuce heads were harvested at each of 13 time points following inoculation (0, 2.5, 5, and 24 h after inoculation and every 24 h thereafter until day 10). The most probable number (MPN) of E. colion each lettuce head was determined, and die-off rates were estimated. The relationship between sample time and the log MPN of E. coliper head was modeled using a segmented linear model. This model had a breakpoint at 106 h (95% confidence interval = 69, 142 h) after inoculation, with a daily decrease of 0.70 and 0.19 log MPN for 0 to 106 h and 106 to 240 h following inoculation, respectively. These findings are consistent with die-off rates obtained in similar studies that assessed E. colisurvival on produce following irrigation. Overall, these findings provide die-off rates for E. colion lettuce that can be used in future quantitative risk assessments.