Isolation, screening, and molecular identification of new lactic acid bacteria as potential probiotics

Abstract

Background and objective Lactic acid bacteria (LAB) are the most useful microorganisms to society, LAB are used in the production of many fermented food products that stimulate digestion and immunity. Therefore, the objective study aimed to isolate, characterize, and identify new LAB members and determining their potential probiotic properties. Materials and methods This study was performed through isolation, selection, and identification of LAB strains morphologically, biochemically, and genetically. Then, to determine their probiotics traits, which include acid resistance, bile-salt tolerance, β-galactosidase activity, heat sensitivity, salinity, cholesterol reduction, and antioxidant activity. Results and discussion Eight LAB isolates were isolated and purified from milk and cheeses, out of eight isolates, K2 showed the highest resistance to acidic pH 2.0, where Log CFU/ml reduced from the initial count 8.79±0.17 at pH 7.0 to 4.57±0.10 at pH 2.0 with 51.99% survival rate. Also, the six bile-tolerant isolates (K1, K2, K3, K4, A11, and A5) were found tolerant to 1% bile-salt concentrations for 3 h with 92.50, 71.10, 64.35, 60.72, 71.11, and 72.21% rate of survival, respectively. Also, antibiotic susceptibility of A11 isolate was the most sensitive, but isolate A2 was the most resistant against antibiotics. Besides, heat sensitivity of selected LAB isolates (K1, K2, A11, and B1) were able to survive at temperatures 15, 37, and 45°C, but isolates (K3, K4, A5, and A2) did not grow at 45°C. Additionally, NaCl tolerance clearly revealed that both isolates (K2 and A11) were able to tolerate a wide range of NaCl (1–8% w/v), then the growth declined with the increase of salt (10%) concentration. The results showed that the cholesterol assimilation was reduced by 32.66 and 30.20% for A11 and K1 isolates, respectively. Also, K2 isolate showed the highest antioxidant activity (86.93%), and the best β-galactosidase producer (24.65 U/ml). Therefore, the 16S-rRNA sequencing was carried out to identify four promising LAB isolates that could be employed as a probiotic strain. Phylogenetic analysis showed that isolates belonged to three genera, including Lactobacillus, Lactococcus, and Enterococcus. Strains were Lactobacillus casei (A5) KU510332, Enterococcus faecium (A11) KU510333, Lactococcus lactis (K1) KU510334, and E. faecium (K2) KU510335. Conclusion Our results presenting four new isolates as promising potential probiotics for further application in processing probiotic products.