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Lactobacilli as Probiotics and Biopreservatives


<OL> <LI> To determine the influence of pH and concentration of Tween 80 for growing cultures of Lactobacillus acidophilus on subsequent stability of the culture in food/feed to ensure their survival on the food/feed until ingested. <LI> To compare eight strains of L. acidophilus for their effect on the immune response in young pigs. <LI> Evaluate the influence of encapsulation of cells of probiotic lactobacilli on their survival on feed/food. <LI> Determine if lactobacilli are internalized in the leaves of green vegetables such as spinach and, if so, can they exert control of food borne pathogens such as Escherichia coli O157:H7.

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NON-TECHNICAL SUMMARY: Today there is much interest in the role of probiotic microorganisms in health and/or nutrition. Among the potential benefits provided by lactobacilli used as probiotics are control of intestinal infections, improved carbohydrate utilization and control of serum cholesterol levels. The interest in the use of probiotics is high throughout the world both for humans and livestock. In addition to benefits for humans, probiotics are considered to control intestinal pathogens and as a potential alternative for use of subtherapeutic levels of antibiotics in livestock feed. Some of the lactobacilli which produce peroxide may be beneficial as food adjuncts in the control of undesirable microorganisms on foods at refrigeration temperatures. A number of species of the lactobacilli can produce sufficient hydrogen peroxide at refrigeration temperatures to exert this control. Lactobacillus delbrueckki ssp lactis is the species most often associated with high levels of peroxide production. These lactobacilli do not grow at refrigeration temperatures however they produce sufficient amounts of peroxide to be antagonistic toward other types of microorganisms. Studies in the past have shown that sufficient hydrogen peroxide is generated by some of these bacteria to exert antagonistic action toward undesirable microorganisms including psychrotrophic spoilage bacteria and some food borne pathogens. Thus these organisms may provide a very useful biopreservative for controlling not only food borne pathogens but also food spoilage organisms on refrigerated products such as meats. <P>

APPROACH: Objective One: Fatty acid profiles of each six strains of L. acidophilus grown in MRS broth will be determined by gas liquid chromatography. The cultures also will be grown in MRS broth, the cells harvested by centrifugation, resuspended in sterile milk and freeze dried and stored at -20C. The percentage survival based on the plate counts after two weeks storage will be compared to the percentages of lactobacillic and oleic acids in the cellular lipids to determine if there is a significant relationship The same cultures will be grown in MRS broth containing 0, 0.1, 0.2, 0.3, 0.4, and 0.5 percent Tween 80. Similar comparisons will be made with respect to survival during storage of the dried cultures and the fatty acid concentrations. To compare the effect of pH during growth of the same six cultures they will be grown at pH 5.0, 5.5, 6.0, and 6.5. Regression analyses will be used to determine if there are significant relationships between survival and fatty acids. Similar analyses will be done to determine if relationships exist between pH of growth and survival. <P>
Objective Two: Weaning age pigs will be used to compare the influence of feeding eight different strains of L. acidophilus on the immune response. On day 21 after samples have been taken each pig will be challenged by injection with salmonella lipopolysaccaride. Following injection, the body temperature and blood samples will be taken at 0, 1.5, and 3hr. The blood serum samples will be assayed for IgA, IgG, and IgM. <P>
Objective Three: Cultures of L. acidophilus (six different strains) will be grown to stationary phase, harvested by centrifugation and resuspended individually in sterile non-fat milk. The suspensions will be divided and 1% beta cyclodextrin added to one half. Portions of the control and beta cyclodextrin suspensions will be freeze dried and stored at room temperature. Survivors will be determined by plate counts. <P>
Objective Four: Bags of fresh spinach and/or lettuce obtained from local supermarkets and the leaves will be surface sterilized. The leaves will be macerated with a sterile mortar and pestal, the ground sample will be diluted and plated on MRS and LBS agars. An enrichment culture also will be made in MRS broth then plated onto MRS agar. Isolated colonies will be identified using methods used previously in our laboratory. They will be compared for inhibitory action toward E. coli 0157:H7 and Salmonella species following methods used previously in out laboratory.

McGlynn, William; Gilliland, Stanley
Oklahoma State University
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