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Presence and Control of Food Borne Pathogens in Ready-to-Eat Foods


The long-term goal of this project is to contribute to the understanding and control of Salmonella, Escherichia coli O157:H7 and Listeria monocytogenes in specific ready-to-eat food products. <P>
We plan to conduct a series of experiments with pathogenic strains of each of these organisms and to test their survival against different physical, chemical and biological treatments. The specific goals of this proposal are to: <OL> <LI> Determine the effectiveness of bacteriophage treatment to reduce the viability of Escherichia coli O157 and Salmonella populations on the surface of lettuce, spinach and tomatoes. <LI> Characterize the thermal resistance of and identify treatments that would kill. Salmonella in low water activity foods. <LI> Identify combinations of antimicrobial GRAS ingredients that will inhibit and reduce the viable count of Listeria monocytogenes, E. coli O157 and Salmonella in a fresh Hispanic cheese (queso fresco). </ol> We expect that the outputs resulting from this project will include: laboratory research activities, mentoring of Food Science graduate and undergraduate students, scientific evidence that will support the idea of using bacteriophages, a time/temperature database for inactivation of Salmonella in low water activity foods, and GRAS addition combinations for treating Hispanic cheeses. <P>
This proposal will be at the core of the research program of the Principal Investigator, focused on understanding and seeking strategies to control food-borne pathogens. The PI has worked extensively in different projects related to these microorganisms, investigating pre-harvest control and prevalence of EHEC related to animal production, incidence of Salmonella and E. coli contamination in fresh vegetables, modeling the growth of L. monocytogenes in deli meats, among other projects.

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NON-TECHNICAL SUMMARY: The presence of potentially pathogenic bacteria in foods is a major public health concern and is a serious threat for the delivery of safe foods by the food industry. In recent years, however, an increasing number of food-borne disease outbreaks have seriously questioned the safety of our food supply systems. Many of those outbreaks have been caused by bacteria that contaminated ready-to-eat foods. Despite the fact that the food industry and the government have implemented a series of actions to prevent contamination, new outbreaks are still frequently reported. Salmonella outbreaks have been linked to a wide variety of food products, but in recent years the occurrence of several multi-state cases related to fresh produce and dry foods have heightened the concern for transmission to these types of products and have exposed weaknesses in our food supply. Some of the implicated foods have included chocolate, dried milk, almonds, peanut butter, peanut products, toasted oats cereal and dried spices. These cases demonstrate that these pathogenic bacteria have a very unique ability to remain viable at conditions of very low moisture for long periods of time. An increased number of outbreaks caused by EHEC O157 have also been linked to fresh produce in particular lettuce and spinach. Since some of the outbreaks with fresh produce were linked to pre-washed product, it suggests that serotype O157 may have unique mechanisms of survival associated with the plant tissue. L. monocytogenes has been a concern in ready-to-eat deli meats, but recent outbreaks linked to the consumption of fresh Hispanic style cheeses have revealed the relatively high risk of transmission via these products. These emerging foodborne challenges require the re-evaluation of current practices and the development of novel strategies to have an impact on preventing more human cases. This project presents a comprehensive and broad approach to address some of these food-pathogen risks. The proposed work may lead to a better understanding of the mechanisms of pathogen survival to identify intervention methods for control. The findings generated by this project will likely contribute to the advancement of a better understanding of the survival and control of pathogenic bacteria in ready-to-eat foods. The increase in knowledge on the effectiveness of the proposed control approaches will likely contribute to finding feasible intervention methods.


APPROACH: This project will be conducted by performing a series of microbiological, biochemical and molecular biology experiments using pure cultures of Salmonella, enterohemorrhagic Escherichia coli and Listeria monocytogenes. The experimental approach is described for each of the objectives below. Objective 1: Bacteriophages specific against E. coli O157 and Salmonella have been identified and partially characterized by our research group. Phages capable of inhibiting E. coli O157 will be used to treat lettuce and spinach leaves. Phages specific for infecting Salmonella Typhimurium will be used to treat tomato plants and fruits. The impact of a variety of factors that may influence the effectiveness of phage treatments will be studied. These factors would include: stage of plant growth, bacterial concentration, bacterial pre-inoculation conditions, phage concentration, temperature, pre-inoculation treatment, lettuce variety, relative humidity, strain specificity, presence of epiphytic microorganisms, treatment of internalized bacteria, and combination of phage/chemical antimicrobial treatments. Objective 2: We have developed an inoculation system to study the viability of bacteria at water activities of toasted oats cereal and we have adapted the capillary tube technique used for thermal inactivation in liquids, for powder dry matrices. We plan to perform a series of laboratory experiments using Salmonella serovars that have been associated with outbreaks of low water activity foods. We will be using different types of low water activity model food matrices: starch, casein, commercial cereal mixtures, spices, wheat flour, corn grits and vegetable oil. These materials will be inoculated with individual Salmonella strains, ground and re-dried where necessary and subjected to thermal treatments. Surviving bacterial counts will be determined by standard microbiological techniques. The range of temperatures that we plan to investigate is from 70 to 150 degree C. The data generated will be treated as a first-degree reaction order and the kinetic parameters for death rate, D value and Z value will be calculated. Objective 3: A fresh Hispanic cheese model ("queso fresco") was developed previously to study the growth of Listeria monocytogenes and recently we have identified that combinations of nisin and caprylic acid can be very inhibitory. For this objective we propose to use a similar model to optimize the treatment to enhance its antimicrobial effects on L. monocytogenes as well as E. coli O157 and Salmonella, and to minimize its impact on sensorial characteristics. Batches of queso fresco will be inoculated with mixtures of individual bacterial species. Cheese curds will be mixed with different ingredients. Queso fresco portions will be stored at refrigeration temperatures and the count of bacterial pathogens will be monitored periodically for 3 weeks. Samples will be subjected to microbiological tests for quantification of cell count. The GRAS ingredient treatments will include nisin, caprylic acid, potassium sorbate, sodium propionate, cinnamic acid, cinnamaldehyde, carvacrol, eugenol, monolaurin, monocaprylin, and other natural extracts.

Diez-Gonzalez, Francisco
University of Minnesota
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