Background: A total of 94 lactic acid bacteria (LAB) were isolated from Tunisian artisanal (Ricotta cheese’s whey) and industrial (bactofugate) milk waste, identified and then screened for their antimicrobial activity against some bacteria implicated on nosocomial infections. </p><p> Objective: Bacterial genera and species identification was performed using molecular tools. The antimicrobial activity was tested against 7 strains of Gram-negative bacteria and 4 strains of Gram-positive bacteria as well as 6 yeasts. </p><p> Method: The Crude extract was found to have a narrow antimicrobial spectrum on Gram-positive bacteria mainly Listeria monocytogenes. Among the strains which showed antibacterial activity, four were determined to be bacteriocins-producers. They were identified as Lactococcus lactis. </p><p> Results: Brain Heart Infusion (BHI) Agar was found more adapted than Man, Rogosa and Sharpe (MRS) to investigate the antimicrobial activity of L. actococcus lactis against L. isteria monocytogenes. The genetic determinants encoding the antimicrobial peptides were targeted by specific PCR. </p><p> Conclusion: All L. lactis bacteriocin producing strains possessed the Nisine Z gene (nisZ) except for one, which contained both Nisine A and Nisine Z genes (nisA and nisZ). They have been identified as antilisterial agentS. </p><p>
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