SCREENING OF ANTI-CLOSTRIDIAL LACTIC ACID BACTERIA STRAINS ISOLATED FROM URUGUAYAN DAIRY FARMS

A total of 130 strains of lactic acid bacteria (starter lactic acid bacteria -SLAB- and non-starter lactic acid bacteria -NSLAB-) were isolated from milk, whey and cheese. Anti-clostridial activity was detected in 56 isolates against Clostridium tyrobutyricum ATCC 25755. Seven strains with the highest inhibitory activity were selected to determine their inhibitory mechanisms. Anti-clostridial activity of strains Streptococcus macedonicus 23, S. macedonicus 24, Lactobacillus casei/paracasei 29 and Lactobacillus rhamnosus 104 was due to organic acids production. Inhibitory mechanisms of strains L. casei 26 and 95 were acid production, bacteriocin and hydrogen peroxide. While anti-clostridial activity of Lactobacillus delbrueckii subsp. bulgaricus 76 was due to acid and hydrogen peroxide production. Cell-free supernatants obtained from strains 26, 95 and 76 conserved their anti-clostridial activity after thermal treatments at 70 and 100 °C. Supernatants obtained from bacteriogenic strains 26 and 95 were analyzed by RP-HPLC. RP-HPLC fractions with anti-clostridial activity were analyzed by mass spectrometry (MALDI-TOF). A peptide with molecular weight of 1162.54 Da was detected in HPLC fraction of L. casei 95. No results were obtained for MALDI-TOF analysis of another HPLC fractions. Based on the thermostability and low molecular weight, L. casei 95 anti-clostridial peptide would belong to a class I or II bacteriocin according to Gram-positive bacteria antimicrobial peptides classification.