Characterization of Lactic Acid Bacteria in Raw Buffalo Milk: a Screening for Novel Probiotic Candidates and Their Transcriptional Response to Acid Stress.

Lactic acid bacteria (LAB) are important microorganisms for the food industry due to their functional activity, as starters and potential probiotic strains. With that in mind, we explored the LAB diversity in raw buffalo milk, screening for novel potential probiotic strains. A total of 11 strains were identified by combination of MALDI-TOF and partial 16S rDNA sequencing and selected as potential probiotic candidates. Bacteria innocuity assessment was performed by determining antimicrobial susceptibility and the presence of virulence factors. Antagonism activity against Escherichia coli, Pseudomonas aeruginosa, Listeria monocytogenes and Staphylococcus aureus was assessed, as well as milk proteolytic activity and exopolysaccharides production. Seven strains were identified as innocuous and two of them, Lactobacillus rhamnosus LB1.5 and Lactobacillus paracasei LB6.4 were selected for further probiotic potential analyses. Both strains demonstrated adhesion ability to Caco-2 cells, coaggregated with S. aureus and E. coli and maintained cell viability after gastrointestinal simulation in vitro, suggesting their probiotic potential. Furthermore, the transcriptional response of Lact. rhamnosus LB1.5 and Lact. paracasei LB6.4 to in vitro acid stress was assessed by RT-qPCR targeting seven genes related to adhesion, aggregation, stress tolerance, DNA repair and central metabolism. The association between the transcriptional responses and the maintenance of cell viability after gastrointestinal simulation highlights the genetic ability as probiotic of the two selected strains. Finally, we have concluded that Lact. rhamnosus LB1.5 and Lact. paracasei LB6.4 are important probiotic candidates to further in vivo studies.