Skip to main content
ARS Home » Northeast Area » Wyndmoor, Pennsylvania » Eastern Regional Research Center » Food Safety and Intervention Technologies Research » Research » Research Project #429829

Research Project: Bacterial Pathogens in Regulated Foods and Processing Technologies for Their Elimination

Location: Food Safety and Intervention Technologies Research

2019 Annual Report

1: Determine the prevalence, levels, types, and locations of pathogens at various points from production through to consumption of raw, further processed, and/or RTE foods. 1.1. Determine the prevalence and levels of L. monoctyogenes, STEC, and Salmonella spp. in RTE foods at retail, as well as at abattoirs/processing plants. 1.2. Determine the relatedness of L. monoctyogenes, STEC, and Salmonella spp. recovered from foods using molecular typing methods such as PFGE and MLGT. 1.3. Assess perceptions, food safety attitudes, and self-reported behaviors related to observed food safety hazards by consumers who shop at grocery stores. 2: Develop, optimize, and validate processing technologies for eliminating pathogens. 2.1 - Determine the transfer and survival of STEC and Salmonella spp. in ground and tenderized (i.e., non-intact) red meat, pork, pet, and poultry products. 2.2 - Determine cook dwell times for ground poultry products using common consumer preparation methods such as cooking on gas or electric grills at internal instantaneous temperatures ranging from 100° to 160°F for lethality towards Salmonella and STEC and for consumer acceptability. 2.3 - Determine the effectiveness of food grade antimicrobials applied via electrostatic spray and Sprayed Lethality in Container (SLIC®) methods on pork offal and on chicken necks and frames for control of Salmonella and STEC. 2.4 - Validate fermentation and cooking of dry-fermented sausages for control of STEC, Salmonella, and other pathogens. 3: Develop and/or validate strategies to deliver antimicrobials to raw and packaged foods from production through to consumption to control L. monocytogenes, STEC, Salmonella spp., and other pathogens.

We will exploit the tools of microbiology, molecular biology, and food science to recover, characterize, and control food borne pathogens from production through to consumption for a variety of foods, with emphasis on specialty/ethnic and higher volume, higher risk foods. We will identify where pathogens enter the food supply, determine how they persist, and investigate biological, chemical, and physical interventions to eliminate or better manage them to improve public health. The target pathogens of greatest concern for this project are Listeria monocytogenes, Salmonella spp., Shiga toxin-producing Escherichia coli, Trichinella spiralis, and Toxoplasma gondii. Targeted foods would include, but not be limited to, raw and ready-to-eat (RTE) meat, poultry, pet, and dairy foods, as well as raw and further processed non-intact meats. One focus of the proposed research is to identify sources and niches of the above mentioned pathogens in foods and food processing environments, as well as at retail and food service establishments, to gain insight on factors contributing to their survival and persistence. Multiple isolates recovered from each sample testing positive from such surveys will be retained for further characterization by phenotypic and genotypic (e.g., pulsed-field gel electrophoresis, PCR-based methods, and/or whole genome sequencing) methods to establish relatedness of isolates and their source and succession. As another focus of our research, efforts will be made to validate processes and interventions such as fermentation, high pressure processing, food grade chemicals, and heat, alone or in combination, to inhibit/remove undesirable bacteria from the food supply and to better manage their presence, populations, and/or survival during manufacture and storage of target foods/feed. The proposed research to find, characterize, and kill pathogens along the food chain continuum will expand our knowledge of the most prevalence/potent food borne pathogens and help us to elaborate better methods for controlling them in foods prior to human contact or consumption, thereby enhancing the safety of our global food supply.

Progress Report
We continue to make measurable progress on our stated milestones via productive collaborations with CRADA partners and food safety professionals from academia, government, industry, and consumer groups. Programmatically, we quantified the prevalence, levels, and types of target pathogens along the food chain continuum from farm to flush and developed and validated biological, chemical, and physical interventions to control Listeria monocytogenes (Lm), Shiga toxin-producing Escherichia coli (STEC), and Salmonella spp. (Sal) in a variety of foods. Regarding the former, we established the recovery rate of STEC in raw, non-intact veal and beef purchased at food retailers in the Mid-Atlantic states in the U.S. In related experiments, the inability to recover viable cells of STEC displaying serogroup-specific surface antigens for at least one of the seven regulated serotypes of STEC in combination with the stx and eae virulence genes suggested that STEC are not common in raw marinades (fresh and spent) from specialty grocers or in raw, non-intact pork obtained from grocery stores in NC, PA, DE, or NJ. We also quantified the prevalence of Sal in raw chicken livers from food retailers, research farms, and abattoirs. Whereas the pathogen was recovered quite often from raw chicken livers purchased at food retailers (ca. 60%; 6.4 MPN to 2.4 log CFU/g), Sal was recovered far less frequently from livers harvested from birds on a research farm (ca. 5.8%; 0.4 to 2.2 MPN/g) or from livers obtained at a poultry slaughter facility (6.7%). Studies are ongoing to subtype the isolates retained from the abovementioned surveys. Collectively, these data provide insight on the true prevalence of pathogens in higher volume and/or higher risk foods, as well as their levels and types, and lead to better management of pathogens and lower public health risks. Regarding pathogen control and interventions, we monitored the viability or improved the safety and/or extended the shelf life of raw, further processed, and/or fermented foods by applying high pressure or heat or by treating such products with food grade chemicals as surface agents or ingredients. Examples include the use of buffered vinegar, a “clean label” food grade chemical, to control Lm in rotisserie chicken salad, monitoring the fate of STEC, Lm, and/or Sal on slices of a dry-cured meat to determine if such products provide a favorable environment for persistence or outgrowth of these pathogens during the extended (up to 12 months) shelf life, and assessing viability of STEC in “Soupie”, a home-made soppressata, to validate safe processes for this specialty/ethnic product popular in the coal regions of PA. These experiments analyzed the effect of experimental parameters such as cooking appliance, volume and type of cooking oil, product formulation, levels and types of antimicrobials, and cooking/fermentation times and temperatures. As expected, the higher the temperature and the longer the time for application of heat or fermentation/drying, the greater the reduction in pathogen levels. These data have real world application for maintaining the safety of our Nation’s food supply via our frequent input from regulators and the food industry and our use of pathogenic strains and pilot-scale processing equipment.

1. Prevalence of Salmonella spp. in/on raw chicken livers. There was demonstrable increase in chicken liver-associated outbreaks over the last 10 years due to Campylobacter and Salmonella. To assess the potential for further outbreaks from liver, ARS researchers in Wyndmoor, Pennsylvania, surveyed 249 raw chicken livers from food retailers for the presence and levels of Salmonella. The pathogen was recovered from 148 of 249 (59.4%) chicken livers purchased at retail stores in DE, NJ, and PA over an about 9-month period; pathogen levels ranged from 6.4 MPN/g to 2.4 log CFU/g. Of note, researchers were more likely (P = 0.019) to recover Salmonella from livers that were packaged by retailers (81 of 121 livers; 66.9%) compared to those packaged directly by processors (67 of 128 livers; 52.3%). Although proper cooking would eliminate low levels of Salmonella in/on raw chicken livers, further interventions for processors are needed to lower the prevalence and levels of this pathogen on poultry liver. Thus, these data will assist the industry in producing safer chicken products, therby protecting the health of the American consumer.

2. Screening raw, non-intact pork for Shiga toxin-producing Escherichia coli (STEC). Although pork is not a common vehicle for food borne illness due to STEC, over the last 25 years there have been a handful of STEC outbreaks caused by pork products. Thus, ARS researchers in Wyndmoor, Pennsylvania, screened 514 raw pork samples (395 ground/non-intact and 119 intact samples) purchased at grocery stores in New Jersey, Delaware, and Pennsylvania, between July and December of 2017 to assess the recovery rate of cells of the seven regulated serogroups of STEC (STEC-7; O26, O45, O103, O111, O121, O145, or O157:H7). Using a PCR-based method for screening purposes, 5.3% (21 of 395 samples) of the non-intact and 3.4% (3 of 119 samples) of the intact pork samples tested positive for the requisite virulence genes (i.e., stx and eae) and for the somatic O antigens for at least one of the targeted STEC serogroups: zero of these 24 presumptive positive pork samples subsequently yielded a viable isolate of STEC displaying a STEC-7 serogroup-specific surface antigen along with the stx and eae genes. These data suggest that STEC-7 are not common in retail raw pork samples in the Mid-Atlantic region of the U.S. Thus, these results will be useful for assisting the industry to develop strategies to lower the risks of STEC attributed to pork products, and in turn, enhancing the safety of our Nation’s food supply.

3. Analysis of marinades from food retailers for Shiga toxin-producing Escherichia coli (STEC). STEC have caused human illness from ingestion of poorly stored, handled, processed, or prepared raw beef products, including both non-intact and marinated products. To better quantify the recovery rate and attendant risk of STEC from such products, ARS researchers at Wyndmoor, Pennsylvania, in collaboration with collaborators at North Carolina State University (NCSU), screened marinades from food retailers for the presence of cells of the seven regulated serotypes of STEC. Of 115 marinade samples (58 fresh marinades and 57 spent/used marinades) collected over 52 weeks from four food retailers within 60 miles of NCSU, zero samples tested positive for STEC using a PCR-based method. Thus, although our findings would suggest that marinades do not present an appreciable risk from STEC, retailers must continue to prepare and maintain marinade solutions and meat at 4 deg C or less, as well as frequently and properly clean and disinfect the equipment and environment in both the processing area and deli case to ensure the safety and quality of red meat and poultry products subjected to marinization for the purposed of enhancing tenderness or flavor. These results will assist the industry and food retailers to mitigate critical risk factors associated with the marination process and marinade formulation, so that contaminated meats are not available for purchase by consumers.

4. Inactivation of Shiga toxin-producing Escherichia coli (STEC) in meat bars. Consumer preferences for foods that are more healthy, lower in calories, and higher in protein have fueled the development of meat-based snack products such as meat bars, a nutrient-dense, protein-rich, hand held portable snack food. Although very popular among consumers, relatively little data has been published to establish if meat bars can support the growth or survival of microbial pathogens, if present, during preparation, handling, or extended storage. Thus, ARS researchers at Wyndmoor, Pennsylvania, mentored and collaborated with student interns to validate commercially-relevant formulations and time/temperature dehydration conditions for lethality of STEC in meat bars. Meat bar batter (i.e., ground beef, pecans, flaxseed flour, cranberries, sunflower seeds, sea salt, black pepper, and celery powder) prepared with or without encapsulated citric acid (ECA) was inoculated with an eight-strain cocktail of cells of STEC to a target level of 3.0 million cells per gram of meat bar. Meat bars (40 g each) were separately cooked/dried, without the addition of humidity, in a commercial, stainless-steel dehydrator set at either 62.8 deg C for 6 h, 71.1 deg C for 4 h, or 62.8 deg C for 2 h and then at 71.1deg C for 2 h. Regardless of formulation, the cooking/drying time/temperature regimens tested herein resulted in reductions of ca. greater than 160,000 cells of STEC per gram. This study provides valuable information for producers and policy makers to enhance the safety of this expanding line of high-protein, meat-based, snack products.

5. Behavior of food pathogens in a specialty, dry-cured beef product. Dry-cured, ready-to-eat (RTE) meats are an increasingly popular food choice, but there is a scarcity of information about the microbiological safety of such products. Thus, ARS researchers at Wyndmoor, Pennsylvania, monitored the behavior of multi-strain cocktails of cells of Listeria monocytogenes (Lm) or Shiga toxin-producing Escherichia coli (STEC) on bresaola, an Italian dry-cured beef product, during extended storage at refrigeration and abusive temperatures. In brief, two slices (ca. 8 g each) of commercially-sliced bresaola were layered horizontally into a nylon-polyethylene bag and then inoculated with a multi-strain cocktail of STEC or Lm to a target level of 1,000 cells per slice. Bags were vacuum-sealed and stored at 4 or 10 deg C. Bresaola did not support growth of Lm or STEC during extended storage: pathogen numbers decreased by ca. 15 to 65 cells per package after 150 or 90 days when bresaola was stored at 4 or 10 deg C, respectively. Thus, bresaola does not provide a favorable environment for outgrowth of Lm or STEC if present on the surface from inadequate processing and/or post-process contamination. These data will assist the food industry and regulatory agencies for both enhancing and assuring microbial food safety of the American food supply.

Review Publications
Jung, Y.N., Porto Fett, A.C., Shoyer, B.A., Henry, E.D., Shane, L.E., Osoria, M., Luchansky, J.B. 2019. Prevalence, levels and viability of Salmonella in/on raw chicken livers. Journal of Food Protection. 82(5):834-843.
Shane, L.E., Porto Fett, A.C., Shoyer, B.A., Phebus, R.K., Thippareddi, H., Hallowell, A.M., Miller, K., Foster-Bey, L., Campano, S.G., Taorimina, P., Glowski, D., Tompkin, R.B., Luchansky, J.B. 2018. Effect of fermentation and post-fermentation heating times and temperatures for controlling Shiga toxin-producing Escherichia coli in a dry-fermented-type sausage. Italian Journal of Food Safety.
Hill, D.E., Luchansky, J.B., Porto Fett, A.C., Gamble, H., Urban Jr, J.F., Fournet, V.M., Hawkins Cooper, D.S., Gajadhar, A., Holley, R., Juneja, V.K., Dubey, J.P. 2018. Rapid inactivation of Toxoplasma gondii bradyzoites in dry cured sausage. Food and Waterborne Parasitology.
Porto Fett, A.C., Shoyer, B.A., Shane, L.E., Osoria, M., Henry, E.D., Jung, Y.N., Luchansky, J.B. 2019. Thermal inactivation of Salmonella spp. in pate made from chicken livers. Journal of Food Protection. 82(6):980-987.
Luchansky, J.B., Mayhew, M., Jung, Y.N., Klinedinst, A., Harkins, L., Shane, L.E., Osoria, M., Mcgeary, L., Traugher, Z., Shoyer, B.A., Chapman, B., Cope, S.J., Campano, S.G., Porto Fett, A.C. 2019. Fate of Shiga toxin-producing Escherichia coli in meat bars during processing and storage: a consumers' perspective. Journal of Food Protection. 82(7):1249-1264.
Fredericks, J.N., Hawkins Cooper, D.S., Hill, D.E., Luchansky, J.B., Porto Fett, A.C., Gamble, H.R., Fournet, V.M., Urban Jr, J.F., Gajadhar, A.A., Holley, R., Dubey, J.P. 2019. Low salt exposure results in inactivation of Toxoplasma gondii bradyzoites during formulation of dry cured ready-to-eat pork sausage. Food and Waterborne Parasitology. 15:e00047.