Samples of fermented millet (koozh) prepared traditionally, were collected from different places in and around Thiruvanamalai and Vellore district (South India) including the tribal inhabited areas of Jawadhu hills. About 75 different organisms were isolated of which seven were found to be lactic acid bacteria (LAB).Out of these seven strains, a single strain that produce bacteriocin was identified, characterized and confirmed by 16s rDNA sequencing as Enterococcus faecium. The bacteriocin inhibited the growth of Lactobacillus sp and other food borne pathogens like Leuconostoc mesenteroides and Listeria monocytogenes and found to exhibit bactericidal activity of 52.66 IU/mL (nisin equivalents). Bacteriocin was treated with proteinase K, pronase, papain, chymotrypsin, trypsin and pepsin, but the action of protease alone was observed, indicates that the bacteriocin was Proteinaceous in nature. This bacteriocin was found to be stable at pH range of 2-12 and tolerant to temperature for 2 hrs upto 110°C at acidic pH. Such molecules of high stability and broad spectral antibacterial property has potential application in food industry as probiotic supplements. © Global Science Publications.