Purpose: The research objectives were to isolate and identify native bacteria on leafy greens exhibiting antagonistic activity against Salmonella enterica Typhimurium LT2 and Escherichia coli O157:H7 ATCC 700728.
Methods: Field samples of spinach and Endive lettuce (n = 30) were collected from two south Texas farms in June 2012. Of each sample, 25 g were taken and homogenized with 225 ml of 0.1% peptone water for 1 min. Aerobic mesophiles, lactic acid bacteria (LAB), fungi, enterococci, and coliforms were enumerated using appropriate media. From each sample, 4-12 isolated colonies from each medium were subjected to biochemical identification by Vitek 2 (bioMérieux N.A.). Pathogen-antagonizing isolates were identified using the Agar Spot method using de Man, Rogosa and Sharpe agar for LAB, or tryptic soy agar (TSA) for all other bacteria. Surface-spotted plates were incubated at 35°C for 24h; then overlaid with molten TSA containing 6.0 log CFU/ml S. Typhimurium or E. coli O157:H7. Plates were incubated at 35°C for 24h; inhibition halo diameters produced by isolates were measured by caliper, baseline corrected and averaged. Isolates producing a mean inhibition halo >1.0 mm were designated antagonistic.
Results: Populations of aerobic mesophiles, fungi, enterococci, LAB and coliforms were 6.7 ± 0.7, 5.3 ± 0.6, 3.4 ± 1.5, 4.7 ± 1.3 and 5.5 ± 0.9 log CFU/cm2, respectively, for spinach; 6.1 ± 0.4, 5.0 ± 0.6, 2.7 ± 1.2, 4.6 ± 0.9 and 5.0 ± 0.5 log CFU/cm2, respectively, for lettuce. Spinach-recovered isolates of Staphylococcus intermedius exhibited maximum antagonism against pathogens with inhibition halos ranging from 2.2 to 13.9 mm. Lettuce-recovered isolates of Lactococcus garvieae exhibited maximum antagonism with inhibition halos ranging from 1.1 to 17.8 mm.
Significance: These data suggest that native microorganisms present on the surfaces of leafy greens can inhibit pathogen growth and may be useful in ensuring produce safety.