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item Matos, Anabelle

Submitted to: Journal of Food Protection
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/19/2004
Publication Date: 1/9/2005
Citation: Matos, A., Garland, J.L. 2005. Effects of community versus single strain inoculants on the biocontrol of salmonella and microbial community dynamics in alfalfa sprouts. Journal of Food Protection. 68(1):40-48.

Interpretive Summary: At least fifteen food poisoning outbreaks due to the consumption of sprouts contaminated with Salmonella have occurred in the past seven years in the United States. Alfalfa sprouts have been implicated in most of these sprout-related outbreaks. Biological control interventions may be an alternative or complementary approach to chemical and physical means for disinfecting seeds used for sprouting. We introduced a well-characterized single biocontrol bacterium, as well as mixtures of bacteria derived from fully grown alfalfa sprouts unto alfalfa seeds that were contaminated in the lab with the foodborne pathogen Salmonella. We then examined the feasibility of the use of these inoculants as biological control treatments to inhibit the growth of the foodborne pathogen Salmonella and examined the effect that these treatments may have on the microbial communities on the surfaces of sprouts. We found that although different modes of biological control may be involved, the use of single and mixtures of bacteria may inhibit the growth of Salmonella on alfalfa sprouts. This study provides critical information for the evaluation of effective biological control treatments on alfalfa sprouts. The results from this research will form the basis for regulatory approval of biological control agents and incorporation of novel intervention strategies that sprout growers can use to help ensure the microbiological safety of alfalfa sprouts, and in turn encourage consumers to include alfalfa sprouts as an option to meet their daily vegetable servings.

Technical Abstract: Potential biological control inoculants, Pseudomonas fluorescens 2-79, microbial communities derived from market sprouts and laboratory-grown alfalfa sprouts, were introduced into alfalfa seeds with and without a Salmonella inoculum to examine their ability to inhibit the growth of this food borne pathogen and assess the relative effects of the inoculants on the alfalfa microbial community structure and function. Alfalfa seeds contaminated with a Salmonella cocktail were soaked for 2 hours in bacterial suspensions from each inoculant tested. Inoculated alfalfa seeds were grown for 7 days and sampled during days 1, 3 and 7. At each sampling, alfalfa sprouts were sonicated for 7 min for 2 min to recover microflora from the surface, and the resulting suspensions were diluted and plated on selective and non-selective media. Total bacterial counts were obtained using acridine orange staining and the percent culturability was calculated. Phenotypic potential of sprout-associated microbial communities inoculated with biocontrol treatments was assessed using community-level physiological profiles (CLPP) based on patterns of utilization of 95 separate carbon sources in Biolog plates. CLPP were also determined using oxygen-sensitive fluorophore in BD microtiter plates to examine functional patterns in these communities. No significant differences in total and aerobic microbial cell density or microbial richness were observed due to the introduction of inoculants on alfalfa seeds with and without Salmonella. We found that P. fluorescens 2-79 significantly reduced the growth of Salmonella early during alfalfa growth (4.22 log), while the market sprout inoculum had the reverse effect, resulting in a 5.48 log reduction of Salmonella on day 7. CLPP analyses revealed that market sprout communities peaked higher and faster compared to the other inoculants tested. These results suggest that different modes of actions of single versus microbial consortia biocontrol treatments may be involved.