Location: Food Safety and Intervention Technologies Research
2020 Annual Report
Objectives
The overall goal of this project is to determine the growth and inactivation kinetics of foodborne pathogens suspended in foods treated using thermal and nonthermal process interventions, with a strong emphasis on ExPEC.
1. Develop and validate models to simulate pathogen behavior under both growth and inactivation conditions.
2. Developing and validating non-thermal and thermal intervention technologies to inactivate pathogens and spoilage microorganisms in raw and ready-to-eat foods and food contact surfaces.
3. Examine any relationship between genotype (virulence factors) and pathogen resistance to interventions.
The results of this research will be transferred to regulatory agencies (USDA Food Safety Inspection Service (FSIS), US Food and Drug Administration (FDA)) to develop genomic-based risk assessments. In addition, results will be transferred to women’s health groups, commercial entities, and the meat and poultry industry. This approach may be ultimately expanded to include other thermal and nonthermal intervention technologies and extraintestinal foodborne pathogens.
Approach
Extraintestinal Escherichia coli (ExPEC) are common contaminants in food which includes fresh produce, fish, meat and poultry. Illness occurs after contaminated food is consumed, the ExPEC colonize the gastrointestinal tract, and are then accidentally transferred to the urethra. They then cause urinary tract infections (UTI), sepsis, and meningitis. Approximately 6-8 million cases of UTI and 23,000 deaths annually are attributed to ExPEC. ExPEC and other extraintestinal foodborne pathogens which are found in meat and poultry have been directly traced to illness in humans. In addition, these emerging foodborne pathogens are resistant to multiple antibiotics and are considered a national research priority as noted in the President’s Council of Advisors on Science and Technology (PCAST, 2014). As specifically noted by regulatory agencies this project addresses “an area of growing concern to FSIS and the public health community” which will help: 1) improve the ability to develop safe processing procedures and to evaluate the impact of processing deviations on pathogen growth in the impacted products; 2) provide insights into mechanisms that contribute to the survival of pathogens to commonly used microbial intervention and mechanisms that affect the severity of illness in humans, and antibiotic resistance in outbreak strains; and 3) provide a scientific foundation for the development of new Agency food safety policies. The Centers for Disease Control and Prevention (CDC) recommends the use of foods treated with appropriate intervention technologies to lessen the risk of foodborne illness for “at risk” individuals. There is little if any information about growth or inactivation kinetics of the ExPEC in food using both thermal and nonthermal food safety intervention technologies or how pathogen genotype affects their resistance to intervention technologies, hence we will generate such data to fill the informational void regarding these emerging pathogens as part of this unique food safety project.
Progress Report
This project focuses on assessing, characterizing, and killing the emerging pathogen extraintestinal pathogenic E. coli (ExPEC) in meat and poultry, with a genomics component to investigate the role that virulence factors and antibiotic resistance play in pathogen resistance to intervention technologies to aid in metagenomic risk assessments conducted by USDA Food Safety Inspection Service (FSIS). ExPEC are a diverse set of emerging pathogens that are present in poultry and red meat, in addition to produce. The association of ExPEC with disease in humans can be traced directly from food animals, produce, and food to humans through modern genetic analysis techniques. They are associated with illnesses such as sepsis (the 6th leading cause of death), ulcerative colitis and Crohn’s Disease (ca. 1 million cases), urinary tract infections (11 million case annually, and 23,000 deaths), and meningitis (ca. 500 deaths) annually. The ExPEC that cause illness in humans are categorized as Uropathogenic E. coli (UPEC), Sepsis- Associated Pathogenic E. coli (SEPEC) and Neonatal Meningococcal E. coli (NMEC). There is now very strong evidence these disease conditions are a form of foodborne illness. Many ExPEC are also resistant to multiple antibiotics, including the antibiotics of last resort. The total cost of ExPEC in foods may be as high as $25 billion, annually. In contrast, there were only six confirmed cases of deaths associated with Shiga toxin-producing E. coli such as O157:H7 in 2016.
In continuing research on the presence of ExPEC in retail chicken meat we isolated ExPEC Sequence Type (ST) 131 and ST 117 isolates which are involved in urinary tract infections. Both isolates are resistant to multiple antibiotics. We also isolated Klebsiella pneumoniae isolates which contained virulence factors necessary for infection in humans which are resistant to multiple antibiotics as well as sanitizers used by both the hospital and food industries. Microbial Resource Announcements were published on these isolates and inactivation kinetic data presented at multiple scientific meetings and distributed to our stakeholders in FSIS as well as women’s health groups. We conducted a survey of E. coli types isolated from fresh herbs (e.g. mint, cilantro, basil, parsley) and found the E. coli ST were those associated with disease in humans, carried multiple ExPEC virulence factors, were resistant to multiple antibiotics, and industrial sanitizers. It appears E. coli on produce are being transferred there through animal waste as they are typically associated with both food and wild animals. The incidence of E. coli on the retail herbs ranged from zero to >10,000 per gram. The presence of E. coli with these virulence factors and antibiotics are concerning as each of the fore-mentioned fresh herbs are typically eaten raw.
In cooperation with our collaborators at National Taiwan University (Taipei, Taiwan) and Tunghai University (Taichung, Taiwan), we have achieved the model development of HPP in combination with several food grade antimicrobials, e.g. acetic acid, allyl isothiocyanate (AITC) and trans-cinnamaldehyde (tCinn), to assess/predict shiga toxin-producing Escherichia coli O157:H7 (STEC) and UPEC inactivation. We found that at high-pressure treatment (250-350 MPa; 10-20 min), AITC (0.03-0.09%, w/w) and tCinn (0.10-0.20%), a 5-log (i.e. 99.999%) reduction may be achieved. Therefore, a modest pressure level (i.e. 300-400 MPa vs. 500-600 MPa currently in use) hurdled with 0.05-0.15% (w/w) of several antimicrobials can deliver the 5-log reduction of ExPEC, UPEC and STEC in chicken meat. The 5-log reduction is required by FDA to validate the non-thermal intervention technology. The effective high-pressure is much lower than that typically required at 500-600 MPa level which can further reduce the food texture damages and HPP operation cost. Regression models to predict inactivation also have been developed and validated to include STEC, UPEC, Salmonella and Listeria monocytogenes. All models have R^2 > 0.9, which indicated good predictions vs. experimental data. We also found that the foodborne pathogen resistance to HPP stress is (in order from high to low) STEC > UPEC > Salmonella > L. monocytogenes. Those models may assist the risk assessment task to control and enhance food safety.
The HPP operation temperature impact on pathogen lethality was investigated and evaluated from -10 to +20 degrees Centigrade. We found the best temperature range to deliver inactivation is around 0- 6 degrees Centigrade for raw ground meats. This finding is in line with the meat process for safety concern in terms of microbial growth potential. To reduce the meat quality damaged by high pressure (> 450 MPa), enzyme applications to improve meat texture has been studied and results demonstrated much improved meat color (appearance) and texture can be achieved [via. Scanning Electron Microscopy (SEM) images observation/evaluation]. Food industry may use those texture results and developed models to optimize the product, processing development and cost reduction, in the meanwhile the microbial risk (foodborne pathogens) can be properly assessed to meet the food safety concerns.
Those results have been or prepared to be published in peer-review journals (e.g. Food Control with Impact Factor ca. 4) in 2020/2021.
Accomplishments
Review Publications
Chuang, S., Sheen, S., Sommers, C.H., Zhou, S., Sheen, L. 2020. Survival evaluation for salmonella spp. and listeria monocytogenes in ground chicken meat subject to high hydrostatic pressure and carvacrol using selective and nonselective media. Journal of Food Protection. 83(1):37-44. https://doi.org/10.4315/0362-028X.JFP-19-075.
Hussain, S.A., Xu, A., Sommers, C.H., Sarker, M.I. 2020. Draft genome sequence of red heat-causing halomonas eurihalina MS1, a moderately halophilic bacterium isolated from saline soil in Alicante, Spain. Microbiology Resource Announcements. https://doi.org/10.1128/MRA.01426-19.
Zhou, S., Sheen, S., Zhao, G., Chuang, S., Han, L., Liu, L.S. 2020. Inactivation of Salmonella in tomato by combination treatment of high pressure and trans-cinnamaldeyde. Food Control. https://doi.org/10.1016/j.foodcont.2020.107441.
