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Characterization of Antibiotic Resistant Foodborne Pathogens in Fresh Produce


The potential development of environmental reservoirs of antibiotic resistance in farmland has recently been a concern. The origin of antibiotic resistance and the implications of the findings are unclear, suggesting a dire need to identify potential major sources of antibiotic resistant pathogens. Therefore, the overall goal of this project is to ascertain the occurrence of antibiotic resistant pathogenic microorganisms in farm environment, fresh produce, and deliver educational programs on safe handling of fresh produce.

The overall plan of operation will involve: <OL> <LI> collecting, characterizing and analyzing microbiological samples from organic and non-organic farms, and fresh produce <LI> developing educational programs on safe handling of fresh produce to minimize the potential for foodborne illnesses;<LI>collaborating with United States Department of Agriculture (USDA) and in food safety research to address foodborne pathogens in fresh produce<LI>training students in food safety research.

The specific objectives are to: <OL> <LI>characterize microorganisms isolated from organic and conventional farms, pre-harvest and post-harvest fresh produce<LI>assess the occurrence, profiles, patterns, and persistence of antibiotic resistant pathogenic microorganisms in soils, irrigation water, raw manure, and produce samples<LI> educate farmers and consumers on hygienic ways of growing, handling, and storing fresh produce<LI> increase student participation in food safety research and outreach.

</ol> The anticipated outcome and impacts of this proposed project include:<OL> <LI> provision of scientific data to USDA Food Safety and Inspection Service (FSIS), Food and Drug Administration (FDA), and Centers for Disease Control (CDC). This will provide insight on profiles of antibiotic resistant pathogens in fresh produce, which is essential in implementing prevention and control measures<LI>improved hygienic fresh produce handling practices by farmers and consumers<LI>a proficient food safety workforce for the 21st century<LI>a stronger partnership between Tennessee State University (TSU), USDA and FDA agencies in food safety research. </ol>

Research findings on occurrence, profiles, patterns, and persistence of antibiotic resistant microorganisms in fresh produce chain; and educational programs can be used in subsequent risk assessment exercises to elucidate the role of raw produce in the dissemination of antibiotic resistance to human populations. The proposed project complements continuing efforts at TSU to strengthen food safety research and outreach.

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NON-TECHNICAL SUMMARY: Recently, there has been alarming increase of bacteria resistant to antimicrobial drugs world-wide. Annual cost of treating infections caused by antibiotic-resistant bacteria is estimated to be 4-5 billion dollars. World-wide increase of foodborne infections with antibiotic resistant pathogens is of growing concern in human medicine. Since antimicrobials are used in livestock and crop production to control pathogens, there is concern about antibiotic resistance development in these pathogens and subsequent transfer to humans through contaminated food. Antibiotic resistance among bacteria associated with food animals has been well documented; research regarding resistance profiles of bacteria isolated from raw produce is lacking. Many outbreaks have been traced to produce and this will continue to occur until fresh produce growers, retail stores, and consumers increase their knowledge and awareness of the risks and consequences of foodborne pathogens. There is data gap between research and the level of fresh produce safety awareness in growers, retail stores, and consumers. Food safety doesn't begin at the grocery store or in the kitchen. It begins on the farm. Research data is needed to determine all major sources of antibiotic foodborne pathogens in fresh produce. Educational programs on safe ways of handling and storing fresh produce are essential to improve fresh produce safety. Data on the prevalence and types of antibiotic resistance in microorganisms isolated from fresh produce may help explicate the role of foods in the transmission of antibiotic-resistant strains to human populations. The aim of the proposed work is to obtain patterns and profiles of antibiotic resistant microorganisms isolated from farms and fresh produce. This data will be integrated with previous microbiological data (in our laboratory) from foods of animal origin; chicken, beef, pork, turkey, lamb, guinea fowls, and goat. The integrated data will represent database for foodborne pathogens and their resistance to antibiotics. This will provide insight on profiles of pathogenic microorganisms in the environment which is essential in implementing prevention and control measures. Educational programs will lead to improved hygienic fresh produce handling practices by farmers and consumers. Therefore, research findings and educational programs will provide useful model information which in turn will guide mitigation efforts and will also be an asset to outbreak investigations.

<P>APPROACH: Environmental sampling sites in farms will include soil, irrigation water, fresh produce, and composted or raw manure. Samples will be labeled with the date, ID numbers, and immediately placed on ice and transported to Tennessee State University for analyses. Questionnaires will be administered to collect data on management practice of farms, retail stores, and consumer handling of fresh produce. Collected samples (25 grams) will be blended in 225 ml of lactose broth and enriched at 35 degrees Celsius for 24 hours. After incubation, the samples will be pummeled in a Stomacher at normal speed setting for 2 min. After homogenization, each sample will be streaked in MacConkey agar plates and incubated at 37 degrees Celsius for 24 hours. Single colonies on each plate will be subjected to antibiotic susceptibility testing. API 20E test kit (bioMerieux, Mo.) will be used to identify the bacteria isolates. DNA will be isolated using the Aquapure genomic DNA isolation kit (Bio-Rad Laboratories). DNA concentration and relative purity will be determined using a fluorometer. The PCR assays will be performed to determine the presence of foodborne isolates from the samples. The educational program will be designed to educate the farmers on safe agricultural practices such as, avoiding planting crops on land prone to flooding, to avoid animal feeding units coming down stream for irrigating farms, testing of irrigation water before application process, and to avoid application of animal manure on the fields within 120 days before harvesting, and use of stainless steel cutter for harvesting which prevents any handling by staff. The education program will highlight the importance of keeping fresh produce under controlled temperatures from farms to processing plants and retail stores. Information learned from the survey and the laboratory research will be used to try and improve the practices found to be deficient in farms, retail stores, and consumers. Therefore we will develop and test educational materials targeted to the at-risk fresh produce safety practices identified in farms, retail stores and in the results of the microbial studies. In keeping with the rapidly expanding usage of the Internet, a comprehensive web site on prevalence of foodborne pathogens in farm environment, fresh produce, retail stores; and educational materials on fresh produce safety practices will be developed. Care will be given to optimize ease of use through highly visible navigation aids, easily-read text, use of appropriate colors, and graphic elements. Those who do not have computers or who choose to receive information via other techniques can read the same basic information in a prepared brochure and advertisements. In addition to being distributed to the study participants, these will be made available through local agencies and offices such as extension, health clinics, area agency on aging, displayed at point of sale such as supermarkets, retail stores and schools. Pre- and post-surveys will be conducted to evaluate the effectiveness of the educational materials and delivery methods at improving participants knowledge and practices.

Kilonzo-Nthenge, Agnes
Tennessee State University
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