Determination of some probiotic properties of lactic acid bacteria isolated from different sources

Lactic acid bacteria (LAB) are a group of commercially important organisms that play an important role in the fermentation process of hexose sugars into lactic acid and they are widely used in industrial applications and as probiotics since they were designated as "generally recognized as safe" organisms. In order to isolate lactic acid bacteria (LAB), different food, donkey milk, and breast milk samples were collected and 45 bacterial strains were purified from those samples. Gram staining, catalase test, methylene blue staining were used for morphologic identification, RAPD-PCR, and 16S rRNA gene sequencing methods were used for molecular identification. According to morphological characteristics 21 isolates were classified as gram (+), catalase (-) cocci, 7 isolates were classified as gram (+), catalase (-) lactobacilli, and grown well on MRS agar plates. A total of 17 isolates were not grown on MRS agar and excluded from the rest of the experimental process. For molecular identification of isolates, OPA-7 primer was used with RAPD-PCR. According to the RAPD-PCR bant profiles, 15 isolates which have the same bant profiles were eliminated and 13 isolates were used for 16s rRNA gene sequence analysis. Lactobacillus sakei MH1, Lactococcus garvieae MH3, Lactococcus lactis subs. cremoris MH4, Enterococcus faeceium MH5, 3 isolates Lactococcus lactis MH3, MH8, MH9, 2 isolates Weissella confusa MH6, MH7 4 isolates Pediococcus acidilactici MH10, MH11, MH12, MH13 were designated according to 16s rRNA gene sequence analysis results. The 13 isolates were characterized for antibiotic susceptibility, survival under pepsin, pancreatin, bile salts, low pH, bacteriocin production, and hemolytic activity. Disc diffusion test results show that all of the isolates were resistant to kanamycin, and most of resistant to gentamicin. MIC test results show that all of the isolates were resistant to streptomycin. Lactobacillus sakei MH1, Lactococcus garvieae MH3, Lactococcus lactis subsp. cremoris MH3, Weissella confusa MH7, Lactococcus lactis MH8, Lactococcus lactis MH9 and Pediococcus acidilactici MH10 survived in pH:2 at 1 hour, and in pH: 3 all of the isolates except Lactococcus lactis subsp. cremoris were survived three hours. All strains did not show antimicrobial effect against indicator microorganisms according to the bacteriocin production test. All strains were able to survive in pancreatin and different bile salt (%0.3, %0.5, %1) concentration, but a low percentage survived in the presence of pepsin (pH:3) Enterococcus faeceium MH5; Weissella confusa MH6, MH7; Lactococcus lactis MH8 and Lactococcus lactis MH9. The tested strains showed γ-hemolytic activity. According to these results, our isolates is probiotic potential candidates for application in the food and pharmaceutical industries, however, isolates should screen for the other probiotic selection criteria.