Much attention and interest have recently been directed toward application of natural antimicrobial compounds in foods and food packaging. Lysozyme is one of the most often used naturally occuring food preservatives. Lysozyme catalyzes the breakage of of β-1,4-linkages of peptidoglycan in bacterial cell walls, and hence kills bacteria. Although lysozymes are generally produced from most organisms, the source has been commercially limited to egg whites. In this study, we screened 26 bacterial strains for the production of lysozyme, and found 7 strains that possessed extracellular lysozyme activity. The lysozyme activities were 342 U from Thermomonas koreensis, 290 U from Lactobacillus rhamnosus, 178 U from Lb. paracasei, 118 U from Lb. reuteri, 90 U from Bacillus subtilis and only 2.26 U from T. hydrothermalis. The cell-free supernatants from lysozyme producing T. koreensis (CFS-Tko), Lb. paracasei (CFS-Lpa), Lb. reuteri (CFS-Lre) and Lb. rhamnosus (CFS-Lrh) showed antibacterial activity against Micrococcus luteus, B. cereus, Staphylococcus aureus, Streptococcus mutans, Listeria monocytogenes, Escherichia coli and Vibrio parahaemolyticus. The inhibitory effect of CFS-Tko was stronger than that of egg white lysozyme (HEWL) against B. cereus. Moreover, the CFS of lysozyme producing lactobacilli, cultured for 48 h, completely inhibited the growth of the test strains. Our results suggest that newly found CFS can be used to extend shelf-life of food and to enhance food safety in food packaging.