1a. Objectives (from AD-416):
ARS is interested in the role of pre-harvest sources of contaminations to fresh produce. The Cooperator is interested in improving the microbial safety of fresh produce.
1b. Approach (from AD-416):
Soil (soil amendments, compost) and irrigation water are major sources of pre-harvest contamination of fresh produce. We will utilize plant natural defense mechanism (glucosinolate-derived compounds) as a green manure to control attenuated strains of enteric pathogens in soil. Brassica plants will be grown in hoop houses. After crop harvest, remnant plant tissues will be tilled over as a green manure. Soil inoculated with attenuated strains will be sampled periodically for their persistence in presence of green manure.
3. Progress Report:
Antibacterial activity of GDC, mainly, isothiocyanates, indole methyl oxazolidinone, and methyl propyl pyrazole carboxylic acid were evaluated against E. coli O157:H7 and its mutants deficient in surface appendages, nonpathogenic E. coli, and Salmonella. Sterile filter disc (6 mm) impregnated with 10-20 µL of GDC (10 mg/ml) were placed on Tryptic soy agar (TSA) plates seeded with respective bacteria. After 48 h incubation at 37°C, the diameter (mm) of the inhibitory zone around the disc was recorded. Antibacterial effect of GDC was dose-dependent, increasing with the dose applied. Salmonella were more sensitive to these compounds than E. coli O157:H7 or non-pathogenic E. coli. Benzyl isothiocyanate exhibited significantly higher zone of inhibition than other compounds or gentamicin (positive control) against Salmonella strains. The antibacterial effects of benzyl- and phenethyl isothiocyanate against E. coli O157:H7 were comparable to that of gentamicin. The antibacterial effect of 3-Methyl-1-propyl-1H-pyrazole-4-carboxylic acid and 1H-Indol-3-ylmethyl-2-oxazolidinone was not evident on test pathogens. A nonpathogenic E. coli O157:H12 strain enriched in bovine fecal slurry (7 log CFU/ml) was sprayed on five plots (5’ x 4’ plot, 1 L per plot). Plots were marked as (1) inoculated control (2) acetic acid (3) cinnamaldehyde (4) Sporan® and (5) BIT, and treated with respective antimicrobials. Soil samples were analyzed periodically for E. coli O157:H12 populations by spiral plating on MacConkey agar containing 50 mg/L nalidixic acid or MPN procedure. Populations of E. coli O157:H12 reduced in soil with time irrespective of antimicrobial treatment. BIT was superior to acetic acid, cinnamaldehyde, and Sporan® in reducing E. coli O157:H12 in soil. None of these compounds had any effect on native microorganisms of soil.