An attempt was made, to characterize natural antibiotics or lantibiotics from unconventional sources and its antibacterial spectrum against food borne pathogens and drug resistant bacteria. Six different traditional fermented foods i.e., fermented fish, fermented soybeans, Soibum (fermented bamboo shoots), milk, idly and dosa batter were used for the isolation of bacteriocin producing Lactic acid bacteria (LAB). Among all bacterial cultures isolated from the various sources, 129 cultures have found to produce antimicrobial compounds. Nisin specific reporter bacteria was utilized as biosensor to identify the Nisin like bacteriocin, where 10 cultures found to be positive Nisin producer. Identified Nisin like bacteriocin was partially concentrated by using ammonium sulphate followed by butanol extraction. Minimum inhibitory concentration (MIC) was analyzed against food borne pathogen and drug resistant bacteria. MIC of partially purified Nisin (pp-Nisin) of all the LAB isolates against food-borne pathogens are ranged between 0.5 and 92µg/ml respected to various Gram-positive bacteria. Similarly, the drug resistant bacteria were also inhibited by pp-Nisin (MIC ranged between 15 and 175µg/ml). All samples of ppnisin exhibited auto induction ability. Taxonomic identification of the nisin producers was done by whole genome sequencing which reveals that cultures belongs to Lactococcus lactis ssp. lactis. Also it was found that Lactococcus lactis ssp. lactis C2d and Lactococcus lactis ssp. lactis SP2C4 harbor nisA gene and Lactococcus lactis ssp. lactis FS2 (L. lactis FS2) harbor nisQ gene. The finding of this study highlights the first case of L. lactis FS2 isolated from fermented fish harbor nisQ gene. Antibacterial activity of pp-Nisin against drug resistant LAB is also reported.