Bacteriocin-producing lactic acid bacteria isolated from mangrove forests in southern Thailand as potential bio-control agents in food: Isolation, screening and optimization
Noraphat Hwanhlem, Jean-Marc Chobert, Aran H-Kittikun
A total of 386 isolates of lactic acid bacteria isolated from mangrove forests (soil, water, leaf, twig and fruit) in southern Thailand were screened for bacteriocin production. Only 4 strains that produced bacteriocin-like inhibitory substance (BLIS) in MRS broth, named KT2W2G, KT2W2L, TS9S17 and TS9S19 showed an inhibition zone against Lactobacillus sakei subsp. sakei JCM 1157, Listeria monocytogenes DMST 17303 and Brochothrix thermosphacta DSM 20171 as indicators by using agar well diffusion assay. None of the inhibitory activities were related to the production of either organic acids or hydrogen peroxide. The BLISs produced by these strains were not affected by heating but were sensitive to proteolytic enzymes. The isolate KT2W2L was identified as Lactococcus lactis subsp. lactis while the isolates KT2W2G, TS9S17 and TS9S19 were identified as Enterococcus faecalis. These BLISs showed a wide range of antibacterial activity against similar bacterial strains, food-spoilage and food-borne pathogens, but were inactive against the Gram-negative bacteria tested. Statistical experimental designs, based on the Plackett–Burman protocol, were applied to optimize the bacteriocin production by Ent faecalis KT2W2G in flask cultures. By using Plackett–Burman protocol, lactose and temperature were found to be the most important factors for bacteriocin production. The effects of the two main factors on bacteriocin activity were further investigated using a central composite design (CCD) and the optimum composition was found to be lactose 14.85 g/l and temperature 25.59°C. Optimum conditions were validated by experiment in which bacteriocin activity (Arbitrary Unit/ml) was increased 8-fold (640 AU/ml) in 18 h fermentation.