ABSTRACT: Biofilms from floor drains in food processing facilities with a recent history of no detectable Listeria monocytogenes were cultured for microorganisms producing antilisterial metabolites. A total of 413 microbial isolates obtained from 12 drain biofilm samples were assayed at 15°C and 37°C for antagonistic activity to L. monocytogenes by two agar plate detection methods. Twenty-one of 257 bacterial isolates and 3 of 156 yeast isolates had anti- listerial activity. All 24 microbial isolates, which produced metabolites inhibitory to L. monocytogenes, were assayed for anti- listerial activity in coinoculated broth cultures containing tryptic soy broth with 0.6% yeast extract (TSB-YE).
A 5-strain mixture of 103 L. monocytogenes/ml and 105 cfu of the candidate microorganism/ml were combined in TSB-YE and incubated at 37°C for 24 h, 15°C for 14 d, 8°C for 21 d and 4°C for 28 d. Substantial inhibition of L. monocytogenes growth (4 to 5 log cfu/ml) was observed for 9 isolates at 37°C, 2 at 15°C and 8°C, and 3 at 4°C. None of the 3 yeast isolates was even moderately inhibitory to L. monocytogenes (£ 1 log10 cfu/ml growth inhibition) at 37°C. The nine inhibitory bacterial isolates were identified as Enterococcus durans (6 isolates), Lactococcus lactis subsp. lactis (2 isolates), and Lactobacillus plantarum (1 isolate). The anti-L. monocytogenes properties of these isolates were evaluated in biofilms of L. monocytogenes on stainless steel coupons at 37°, 15°, 8° and 4°C. Results revealed that all of nine isolates were highly inhibitory (>5 log10 L. monocytogenes cfu/cm2 growth inhibition to a undetectable level as determined by a direct plating method) at 37°C for 24 h when compared with a control biofilm of L. monocytogenes only. At 4° and 8ºC, three isolates, E. durans 141-1 and 152 and Lc. lactis subsp. lactis C-1-92, were highly inhibitory to L. monocytogenes through 35 days (>4 log10 L. monocytogenes/cm2 growth inhibition). These three bacterial isolates appear to be especially promising competitive inhibition candidates to control L. monocytogenes in biofilms at environmental temperatures of 4°, 8º and 37°C.
A 5-strain mixture of 103 L. monocytogenes/ml and 105 cfu of the candidate microorganism/ml were combined in TSB-YE and incubated at 37°C for 24 h, 15°C for 14 d, 8°C for 21 d and 4°C for 28 d. Substantial inhibition of L. monocytogenes growth (4 to 5 log cfu/ml) was observed for 9 isolates at 37°C, 2 at 15°C and 8°C, and 3 at 4°C. None of the 3 yeast isolates was even moderately inhibitory to L. monocytogenes (£ 1 log10 cfu/ml growth inhibition) at 37°C. The nine inhibitory bacterial isolates were identified as Enterococcus durans (6 isolates), Lactococcus lactis subsp. lactis (2 isolates), and Lactobacillus plantarum (1 isolate). The anti-L. monocytogenes properties of these isolates were evaluated in biofilms of L. monocytogenes on stainless steel coupons at 37°, 15°, 8° and 4°C. Results revealed that all of nine isolates were highly inhibitory (>5 log10 L. monocytogenes cfu/cm2 growth inhibition to a undetectable level as determined by a direct plating method) at 37°C for 24 h when compared with a control biofilm of L. monocytogenes only. At 4° and 8ºC, three isolates, E. durans 141-1 and 152 and Lc. lactis subsp. lactis C-1-92, were highly inhibitory to L. monocytogenes through 35 days (>4 log10 L. monocytogenes/cm2 growth inhibition). These three bacterial isolates appear to be especially promising competitive inhibition candidates to control L. monocytogenes in biofilms at environmental temperatures of 4°, 8º and 37°C.
Keywords: Listeria monocytogenes, biofilm, competitive inhibition, lactic acid bacteria
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