Source: PURDUE UNIVERSITY submitted to
PREVALENCE, PERSISTENCE, AND STRESS SURVIVAL OF FOODBORNE PATHOGENS IN FOOD SYSTEMS
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
TERMINATED
Funding Source
HATCH
Reporting Frequency
Annual
Accession No.
0226337
Grant No.
(N/A)
Project No.
IND011661
Proposal No.
(N/A)
Multistate No.
(N/A)
Program Code
(N/A)
Project Start Date
Apr 15, 2011
Project End Date
Sep 30, 2015
Grant Year
(N/A)
Project Director
Oliver, HA.
Recipient Organization
PURDUE UNIVERSITY
(N/A)
WEST LAFAYETTE,IN 47907
Performing Department
Food Science
Non Technical Summary
Characterizing foodborne pathogens and their ability to survive in food and food production/handling systems using integrated applied and basic research approaches is important to improving public health. This project will engage stakeholder advisory groups to consult studies aimed to investigate the human health risk attributable to foodborne pathogens in food and food production and handling systems, determine foodborne pathogen prevalence and contamination levels in food and food production and handling environments, and characterize the stress response mechanisms, molecular subtypes and pathogenic potential of foodborne pathogens isolated from food and food production and handling systems. Results will be communicated to stakeholders to encourage development of intervention strategies to prevent disease.
Animal Health Component
(N/A)
Research Effort Categories
Basic
(N/A)
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
7124010110090%
7124010302010%
Knowledge Area
712 - Protect Food from Contamination by Pathogenic Microorganisms, Parasites, and Naturally Occurring Toxins;

Subject Of Investigation
4010 - Bacteria;

Field Of Science
3020 - Education; 1100 - Bacteriology;
Goals / Objectives
The purpose of this project is to investigate foodborne pathogens and their ability to survive in food and food production/handling systems using integrated applied and basic research approaches. The objectives and approaches described hereafter have been developed to investigate human foodborne pathogens (e.g. Salmonella, E. coli, Listeria monocytogenes) in an effort to improve food safety and enhance public health. The specific objectives of this project include: (i) Objective I. Determine foodborne pathogen prevalence and contamination levels in food and food production and handling environments and(ii) Objective II. Characterize the stress response mechanisms, molecular subtypes and pathogenic potential of foodborne pathogens isolated from food and food production and handling systems. Outcomes: Objective I. Upon completion of this objective, we will have determined the prevalence and levels of foodborne pathogens (e.g., L. monocytogenes, E. coli, Salmonella) in food and food production and handling environments. These data will be used to update U.S. FDA/CFSAN, USDA/FSIS, and CDC risk assessments. By engaging an advisory board, this study will better meet the needs of the major stakeholder groups, notably industry, and will integrate our research and extension-based approach. Producer input will provide invaluable information on production practices and insight into feasibility of potential control strategies. As we will be collecting food and environmental samples from food production and handling systems, the food industry has a vested interest in understanding foodborne pathogen prevalence in products and their establishments. Engaging experts in policy and risk assessment will facilitate discussions between industry and regulators to ensure that data are interpreted accurately. Outreach/extension specialists will help us achieve our long-term goal of communicating the results to industry. Objective II. Upon completion of this objective, we will have identified environmental niches in food processing and handling environments that are persistently contaminated by foodborne pathogens and identified potential routes of transmission that contribute to cross-contamination of foods.
Project Methods
Objective I. Stakeholder advisory groups would be composed of representatives from food production, food processing, retail (grocery), government/policy, and academia/outreach sectors. The goals of advisory group meetings will be to discuss previous and ongoing research in foodborne pathogen prevalence and persistence along the food chain, discuss industry needs for pathogen control, and to identify additional long-term goals including intervention and control strategies Objective I. Detection and isolation of human foodborne pathogens. The FDA Bacterial Analytical Manual methods will be used to detect and isolate foodborne pathogens in food and environmental sponge samples (www.cfsan.fda.gov/~ebam/bam-10.html Objective III. RNA-Seq. RNA-Seq technology will be used to identify differences in genes expression among strains exposed to stress compared to a control. Specifically, RNA will be extracted and processed for sequencing as previously described by Oliver et al. 2009. Sequencing will be carried out on the Illumina Genome Analyzer or equivalent sequencer. To identify genes that showed significantly different transcript levels in the control strain and the test strain, statistical analyses will be performed using the normalized RNA-Seq coverage of each coding gene. Normalized RNA-Seq coverage (i.e. the number of reads that match an annotated CDS after normalization across runs) will be used for statistical analyses. Pulsed field gel electrophoresis (PFGE). PFGE typing will be performed using the standardized CDC PulseNet protocol (http://www.cdc.gov/pulsenet/protocols/pulsenet_listeria_protocol%20. pdf). DNA from isolates will be digested separately with the restriction enzymes which act as "molecular scissors" to yield DNA fingerprint patterns. PFGE patterns will be analyzed and compared using Bionumerics software (Applied Maths, Austin, TX). Single nucleotide polymorphism (SNP) genotyping. A multiplex SNP genotyping assay (SNaPshot Multiplex SNP Genotyping Kit; Applied Biosystems, Foster City, CA) will be used to determine the proportion of L. monocytogenes isolated from produce and other plant-based products and the retail environment that pose reduced human health risk. Briefly, the SNaPshot assay identifies changes (mutations) in DNA sequence that lead to nonfunctional proteins (typically virulence factors). Objective VI. Presentations of outcomes at professional and industry meetings. We will present our findings from Objectives I and II at professional meetings (e.g. International Association for Food Protection) and at relevant industry meetings (e.g. Food Marketing Institute) to reach a wider range of constituents. Webinars. We will conduct a series of webinars to disseminate knowledge on foodborne pathogen prevalence persistence, and virulence in foods and food processing and handling environments to producers and retailers. Delivery of webinars will allow us to reach a broad target audience across the U.S. (and beyond), including small and very small producers that would not typically attend symposia or workshops.

Progress 04/15/11 to 09/30/15

Outputs
Target Audience:Food safety experts, manufacturing and retail food industries, state and federal regulators Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?We worked directly with over 10 large retailers to investigate foodborne pathogens in retail food systems. Individual training programs were developed and delivered to corporate sanitarians and food safety managers. How have the results been disseminated to communities of interest?Our results have been and will continue to be published in peer reviewed journals (e.g., Journal of Food Protection, Foodborne Pathogens and Disease). We have presented this work the International Associate for Food Protection Annual meeting from 2010-2015, the Conference for Food Protection, Association for Food and Drug Officials, and federal public meetings in Washington D.C. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Objective I. Upon completion of this objective, we determined the prevalence and levels L. monocytogenes,in retail food production and handling environments. These data have beeb used to update U.S. FDA/CFSAN, USDA/FSIS, and CDC risk assessments. By engaging an advisory board, this study helped meet the needs of the major stakeholder groups, notably the retailindustry, to integreate our research findings. We engagedexperts in policy and risk assessment to facilitate discussions between industry and regulators to ensure that data are interpreted accurately. We developed extension-based trainings toachieve our long-term goal of communicating the results to industry. Objective II. Weidentified environmental niches in retail food processing and handling environments that are persistently contaminated by foodborne pathogens and identified potential routes of transmission that contribute to cross-contamination of foods. We also develop intervention strategies to reduce niches in retail.

Publications


    Progress 10/01/13 to 09/30/14

    Outputs
    Target Audience: The target audience for the work performed in 2013-2014 reporting period is retail food chains, government regulators, food safety experts and academics interested in food safety. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? 1) Retail food safety experts were training in food safety strateges in retail food systems 2) Four graduate students have continued their training in food safety research and outreach How have the results been disseminated to communities of interest? Six peer-reviewed publications and 1 book chapter have been submitted, accepted or published to commuincate our research resutls. What do you plan to do during the next reporting period to accomplish the goals? We are focusing our research efforts on transciptional profiling of foodborne pathogens from food systems to better understand genetic mechanisms that may attribute to persistent phenotypes.

    Impacts
    What was accomplished under these goals? Our group continues to improve understanding of foodborne pathogen prevalence, persistence, and control strategies. Objective I. Determine foodborne pathogen prevalence and contamination levels in food and food production and handling environments: In 2014 we finalized two studies that investigated prevalence and persistence of L. monocytogenesin retail food systems Objective II. Characterize the stress response mechanisms, molecular subtypes and pathogenic potential of foodborne pathogens isolated from food and food production and handling systems. In 2014 we finished a study investigating the biofilm formation capacity and sanitizer tolerance ofL. monocytogenes from retail food systems.

    Publications

    • Type: Book Chapters Status: Awaiting Publication Year Published: 2014 Citation: S. Hammons and H. F. Oliver*. 2014 Listeria monocytogenes at Retail in Retail Food Safety. Editors: Farber, Crichton, and Snyder. Springer Publishing, New York, NY
    • Type: Journal Articles Status: Published Year Published: 2014 Citation: 1. C. Simmons, M. J. Stasiewicz, E. Wright, S. Warchocki, S. Roof, J. R. Kause, N. Bauer, S. Ibrahim, M. Wiedmann, and H. F. Oliver*. 2014 Listeria monocytogenes and Listeria spp. Contamination patterns in retail delicatessen establishments in three US states. J Food Prot doi:10.4315/0362-028X.JFP-14-183
    • Type: Journal Articles Status: Published Year Published: 2014 Citation: I-H. Cho, A. D. Radadia, K. Farrokhzad, E. Ximenes, E. Bae, A. K. Singh, H. F. Oliver, M. Ladisch, A. Bhunia, B. Applegate, L. Mauer, R. Bashir, and J. Irudayaraj. 2014. Nano/Micro and spectroscopic approaches to food pathogen detection. Annu Rev Anal Chem. 7:65-88. Dr. Oliver contributed the introduction.
    • Type: Journal Articles Status: Published Year Published: 2014 Citation: A. M. Pleitner, V. Trinetta, M. T. Morgan, R. L. Linton, H. F. Oliver*. 2014. Transcriptional and phenotypic response of Listeria monocytogenes to chlorine dioxide. Appl Environ Microbiol 80(9).
    • Type: Journal Articles Status: Submitted Year Published: 2015 Citation: 1. S. R. Hammons, M. J. Stasiewicz, S. Roof, and H. F. Oliver*. Submitted. Environmental adenosine triphosphate levels correlate with probability of Listeria monocytogenes detection in retail delis. J Food Prot.
    • Type: Journal Articles Status: Accepted Year Published: 2014 Citation: J. Wang, A. Ray, S. Hammons, and H. F. Oliver*. Accepted. Biofilm formation, sanitizer tolerance, and virulence potential of persistent and transient Listeria monocytogenes isolate from retail delis. Foodborne Path Dis.
    • Type: Journal Articles Status: Accepted Year Published: 2015 Citation: S. McCoy, J. W. Chang, K. McNamara, H. F. Oliver+, and A. Deering. Accepted. Quality and safety attributes of Afghan raisins before and after processing in Afghanistan. J Food Process Preserve.


    Progress 10/01/12 to 09/30/13

    Outputs
    Target Audience: The target audience for the work performed in 2013 reporting period is retail food chains, government regulators, and academics interstined in food safety. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? The laboratory currently supports the work of 6 graduate students focused on the state objectives How have the results been disseminated to communities of interest? The results have been disseminated through peer reviewed publication, invited talks, and invited symposiums. What do you plan to do during the next reporting period to accomplish the goals? I will work to continue to meet the stated objects through conitiued research and publication. These include up to 4 publications on the prevalence and persistence ofL. monocytogenesin retail food systems.

    Impacts
    What was accomplished under these goals? (i) Objective I. Determine foodborne pathogen prevalence and contamination levels in food and food production and handling environments We have investigated the prevlaence and persistence ofL. monocytogenein retail >90 retail delis across the United States. We are currently analyzing the data for publication. (ii) Objective II. Characterize the stress response mechanisms, molecular subtypes and pathogenic potential of foodborne pathogens isolated from food and food production and handling systems. We have investigated the transcriptional affects of chlorine dioxide onL. monocytogene. This work is about to be submitted for publication. Further, a publication on a select strain ofL. monocytogeneswas published as reported previously

    Publications

    • Type: Journal Articles Status: Published Year Published: 2013 Citation: Oliver HF, Orsi RH, Wiedmann M, Boor KJ., SigmaB Plays a Limited Role in the Ability of Listeria monocytogenes Strain F2365 To Survive Oxidative and Acid Stress and in Its Virulence Characteristics. J Food Prot. 2013 Dec;76(12):2079-86. doi: 10.4315/0362-028X.JFP-12-542.
    • Type: Book Chapters Status: Submitted Year Published: 2013 Citation: Hammons S, Oliver HF. Submitted. Listeria monocytogenes: What the Retail Deli and Food Safety Manager Need to Know. In Farber, Crichton, and Synder (ed). Retail food Safety. Springer, New York.


    Progress 10/01/11 to 09/30/12

    Outputs
    OUTPUTS: The goal of this project was to develop and implement new control strategies for L. monocytogenes in commercial retail delis and to test the ability of these strategies to reduce and control L. monocytogenes contamination. Our specific objectives were to:Objective 1. Develop practical and feasible control strategies to (i) reduce L. monocytogenes in retail deli environments and (ii) reduce L. monocytogenes cross-contamination of RTE deli meats handled at retail. Objective 2. Conduct a survey to identify additional potential risk-factors (e.g. deli design, sanitation practices, equipment manufacturer) which may contribute to L. monocytogenes persistence in retail deli environments. Objective 3. Test the efficacy of control strategies developed in Obj.1 & 2 through implementation and follow-up testing in actual retail delis. The results of this study will be published in peer reviewed journals. The data and conclusions have been shared with stakeholders in the study and with retail food safety experts at national food safety meetings. PARTICIPANTS: Nothing significant to report during this reporting period. TARGET AUDIENCES: Food Industry safety and health specialists. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

    Impacts
    We found L. monocytogenes to be prevalent in retail delis. Non-food contact surfaces had higher L. monocytogenes prevalence than food contact surfaces. Further, we found that L. monocytogenes can persist over time but its continued persistence after implementation of enhanced control strategies may be due to failed execution of SSOPs. Interventions strategies employed in this study significantly decreased L. monocytogenes prevalence in stores with moderate prevalence (5-10% samples positive). Control strategies in the form of enhanced SSOPs were not sufficient to eliminate L. monocytogenes from highly prevalent and persistently contaminated stores indicating that more aggressive strategies (e.g., deep cleans, capital investment in redesign) may be necessary to fully mitigate persistent contamination. Efficient, effective SSOP verifications strategies are greatly needed in the retail deli food system. ATP testing may be a valuable tool to help ensure execution of SSOPs as well as useful as a training tool to demonstrate effective cleaning in retail delis. A positive impact of this study is that targeted and enhanced SSOPs can reduce L. monocytogenes in retail delis with moderate prevalence. Challenges remain around (i) efforts to reduce L. monocytogenes in delis with high prevalence and (ii) verification strategies to ensure execution of SSOPs in retail delis.

    Publications

    • K. Hoelzer, B. D. Sauders, M. D. Sanchez, P. T. Olsen, M. M Pickett, K. J. Mangione, D. H. Rice, J. Corby, S. Stich, E. D. Fortes, S. E. Roof, Y. T. Grohn, M. Wiedmann, and H. F. Oliver. 2011. Prevalence, distribution, and diversity of Listeria monocytogenes in retail environments, focusing on small establishments and establishments with a history of failed inspections. J Food Prot. 74:1083-1095.
    • K. Hoelzer, H. F. Oliver, L. Kohl, J. Hollingsworth, M. T. Well, and M. Wiedmann. 2011. Structured expert elicitation about Listeria monocytogenes cross-contamination in the environment of retail deli operations in the United States. Risk Anal. Epub doi: 10.1111/j.1539-6924.2011.01729.x


    Progress 10/01/10 to 09/30/11

    Outputs
    OUTPUTS: Dr. Oliver incorporated teaching modules on molecular methods in food microbiology in courses she teaches Purdue Univesrity. Dr. Oliver delivers lectures on molecular detection and subtyping of foodborne pathogens for the following courses: Food Microbiology, Food Microbiology Lab, and Graduate Food Microbiology. Furthermore, she taught a "back to class" microbiology seminar to Purdue Alumnae on Foodborne pathogens and food safety. Dr. Oliver taught at the third annual workshop on "Molecular Methods in Food Microbiology" at Colorado State University, which focused on molecular detection of foodborne pathogens and spoilage organisms, was conducted in Fort Collins, CO in June-July, 2010. This workshop is conducted in conjunction with Colorado State University (Dr. Kendra Nightingale), Cornell University (Dr. Martin Wiedmann), and Silliker, Inc. (Dr. Sarita Raengpradub-Wheeler). The workshop was attended by approximately 40 symposium participants and 20 people participated in the laboratory session. Participants indicated that they valued the different teaching styles of the instructors and the integration of complementary expertise. Dr. Oliver presented her research on L. monocytogenes in retail delis at the following conferences: Reciprocal Meat Conference, Manhattan, KS; American Meat Institute Foundation Listeria control workshop, Indianapolis, IN; Canadian Meat Council, Toronto, CA; EcoLab Annual Retail Symposium, Springfield, MA; International Association for Food Protection, Milwaukee, WI. PARTICIPANTS: Cornell University: Martin Wiedmann, Sherry Roof, Steven Worchocki, Courtenay Simmons, Emily Wright North Carolina A & T: Salam Ibrahim Colorado State University: Kendra Nightingale TARGET AUDIENCES: Target audiences include other academic researchers as well as food industry and government agencies that use molecular methods for detection and further characterization of foodborne pathogens. PROJECT MODIFICATIONS: Not relevant to this project.

    Impacts
    We will have tested approx 4,000 retail deli environmental samples to evaluate L. monocytogenes and Listeria. spp. prevalence and persistence post-intervention prevalence. We will compare L. monocytogenes and Listeria spp. prevalence in the current study to prevalence determined in the USDA-funded longitudinal study (i.e., baseline date collected before control strategies were implemented). Specifically, we evaluated differences in prevalence among each food, non-food, and transfer point contact surfaces sites as well as prevalence and persistence trends for each store pre- and post-intervention. We compared ATP bioluminescence values (collected daily in each establishment) with corresponding L. monocytogenes prevalence data for each store. Finally, molecular subtyping (PFGE) is being performed using the standardized CDC PulseNet protocol. Molecular subtyping of L. monocytogenes isolates from retail environments provide insight into remaining or additional harborage sites and routes of transmission associated with cross-contamination of deli meats at retail after our initial efforts to implement new control strategies. These data will be used to recommend best practices to the retail food industry and will be used to inform future iterations of the L. monocytogenes risk assessments and cross-contamination model

    Publications

    • K. Hoelzer, H.F. Oliver, L.R. Kohl, J. Hollingsworth, M. T. Wells, and M. Wiedmann. 2011. Structured Expert Elicitation About Listeria monocytogenes Cross-Contamination in the Environment of Retail Deli Operations in the United States. Risk Anal.
    • K. Hoelzer, B. D. Sauders, M. D. Sanchez, P. T. Olsen, M. M Pickett, K. J. Mangione, D. H. Rice, J Corby, S. Stich, E. D. Fortes, S. E. Roof, Y. T. Grohn, M. Wiedmann, and H. F. Oliver. 2011. Prevalence, distribution, and diversity of Listeria monocytogenes in potentially high risk retail environments. J Food Prot. 74: 1083-1095.
    • Y. Jones, H. F. Oliver, J. R. Deed, and H. F. Yancy. 2010. Real time assay for the detection of pufferfish products. J. Food Prot. 73(9):1698-70