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Problem statement: This study reports the isolation of lactic acid bacteria from 13 honey samples produced in Malaysia, Libya and Saudi Arabia and their antibacterial activity against three Gram negative pathogenic bacteria. Approach: A modified MRS agar with 0.8% CaCO3 and MRS with 1% glucose was found to facilitate isolation of LAB compared to MRS, tomato juice agar and modified tomato juice agar. 32 isolates were confirmed LAB by catalase test and Gram staining. Six isolates were screened for antibacterial activity and identified as strains of Lactobacillus acidophilus 1 by API CH50. Results: All the isolates showed very good inhibitory activity against target Gram negative bacteria as indicated by the diameter of inhibition zone: Salmonella Typhimurium (23-30 mm), Escherichia coli (7-18 mm) and Enterobacter aerogenes (10-18 mm) after 24 h incubation at 30°C. Supernatants of L. acidophilus 1 strains showed good antibacterial activity against all target bacteria. Heating the supernatants at 90 and 121°C for 1 h enhanced the antibacterial activity against all target bacteria except supernatants H006-A and H010-G against S. Typhimurium. Antibacterial activity of supernatants were maintained after pH adjustment to 3, but at pH5 supernatants H006-A, H008-D and H010-G lost the activity against S. Typhimurium and E. coli within 48 h of incubation while at pH 6 all supernatants lost activity except against E. aerogenes. Enzymes treatments of supernatants with RNase II and Proteinase K for 1 h inhibited all target bacteria except supernatants H008-D, H008-E and H006-A which were relatively sensitive to both enzymes against S. Typhimurium and E. coli. Conclusion/Recommendations: In conclusion, honey from different sources contains strains of L. acidophilus 1 that produced compounds with good antibacterial activity which may be responsible for the antibacterial properties of honey

Evaluation on Antibacterial Activity of <i>Lactobacillus acidophilus</i> Strains Isolated from Honey