1a. Objectives (from AD-416):
ARS EMFSL will focus on the objective to determine the factors influencing bacterial attachment to leafy greens surfaces and internalization of E. coli 0157:H7.
1b. Approach (from AD-416):
Foodborne outbreaks due to microbial contamination of fresh produce have been increasing in recent years. This proposal addresses methods to prevent, detect, monitor, control, and respond to potential food safety hazards in the production and processing of specialty crops, including fresh produce. The goals target identification of routes of introduction; factors that influence persistence and survival of E. coli and S. enterica in leafy greens and melons from production to finished product preparation; and dissemination of information across the nation. The results obtained during this study will help in developing alternative technologies to enhance the safety of produce and to educate/train different sectors of the produce industry and different groups of consumers on food safety practices to avoid consumption of contaminated produce. Ultimately, it will help specialty crop industries recover and retain consumer confidence and reduce food safety risks. For the proposed project, ARS, EMFSL, will focus on part of objective (1) to determine the factors influencing bacterial attachment to leafy greens surfaces and internalization of E. coli O157:H7 while the cooperator will conduct research to address part of objectives 1, 2 and 3-6.
3. Progress Report:
We conducted a “proof of concept study” to determine if Shiga toxigenic Escherichia coli can be internalized into roots of spinach grown on a hydroponic system, under strictly controlled conditions that will prevent the contact of the upper parts of the plant (leaves and stems) with inocula. In addition, the effect of the spinach cultivar was also evaluated. The inoculums level strongly affected the incidence of internalization on both liquid and semisolid system. At the lower inoculums (5 log CFU/ml) the internalization was rare for all strains tested during the entire experiment. In contrast, at high inoculums level (7 log CFU/ml), the incidence of internalization ranged between 19 and 29%. Although all internalization observed was through the root system, the highest incidence of contamination (40% from all positives) was detected in stem portion of the plant. A possible explanation for a lower incidence of bacteria detection in roots could be conditioned by the morphophysiological difference of the root from other plant parts. After root are detached from the spinach plant they continue to actively uptake water, thus probably during the 10 min disinfection treatment in solution containing 0.1 % mercury chloride, root tissue is subjected to a greater degree of disinfection as compared to leaves and shoots. In addition, roots are lacking the hydrophobic wax layer as compared to leaves, which also contributes to a greater degree of mercury chloride penetration into the vegetal tissue, and as a result lower numbers of bacteria recovered from roots. Further, the mechanical wounding of the spinach roots strongly increased the probability of internalization of bacteria into hydroponically grown spinach plants on media containing 5 log CFU/ml. On intact plants grown on the same inoculated media the internalization was rare during the entire experiment (up to 35 days). E. coli O157:H7 populations recovered from leaves and shoots of root-wounded plants were similar or marginally higher than in hydroponically grown spinach plants on media containing 7 log CFU/ml. Both spinach cultivars, Space and Waitiki, were contaminated with the similar probability suggesting that spinach cultivar does not influence bacteria internalization into root system of the plant. Curli expression ability of STEC strain had no effect its internalization into spinach at the root level.