In Vitro and in Vivo Selection of Potentially Probiotic Lactobacilli From Nocellara del Belice Table Olives

Abstract

Table olives are increasingly recognized as a vehicle as well as a source of probiotic bacteria, especially those fermented with traditional procedures based on the activity of indigenous microbial consortia, originating from local environments. In the present study, we report characterization at the species level of 49 Lactic Acid Bacteria (LAB) strains deriving from Nocellara del Belice table olives fermented with the Spanish or Castelvetrano methods, recently isolated in our previous work. Ribosomal 16S DNA analysis allowed identification of 4 <i>Enterococcus gallinarum</i>, 3 <i>E. casseliflavus</i>, 14 <i>Leuconostoc mesenteroides</i>, 19 <i>Lactobacillus pentosus</i>, 7 <i>L. coryniformis</i>, and 2 <i>L. oligofermentans</i>. The <i>L. pentosus</i> and <i>L. coryniformis</i> strains were subjected to further screening to evaluate their probiotic potential, using a combination of <i>in vitro</i> and <i>in vivo</i> approaches. The majority of them showed high survival rates under <i>in vitro</i> simulated gastro-intestinal conditions, and positive antimicrobial activity against <i>Salmonella enterica</i> serovar Typhimurium, <i>Listeria monocytogenes</i> and enterotoxigenic <i>Escherichia coli</i> (ETEC) pathogens. Evaluation of antibiotic resistance to ampicillin, tetracycline, chloramphenicol, or erythromycin was also performed for all selected strains. Three <i>L. coryniformis</i> strains were selected as very good performers in the initial <i>in vitro</i> testing screens, they were antibiotic susceptible, as well as capable of inhibiting pathogen growth <i>in vitro</i>. Parallel screening employing the simplified model organism <i>Caenorhabditis elegans</i>, fed the <i>Lactobacillus</i> strains as a food source, revealed that one <i>L. pentosus</i> and one <i>L. coryniformis</i> strains significantly induced prolongevity effects and protection from pathogen-mediated infection. Moreover, both strains displayed adhesion to human intestinal epithelial Caco-2 cells and were able to outcompete foodborne pathogens for cell adhesion. Overall, these results are suggestive of beneficial features for novel LAB strains, which renders them promising candidates as starters for the manufacturing of fermented table olives with probiotic added value.