In vitro probiotic–associated, safety, and anti–listerial properties of indigenous dairy–derived lactobacilli: functional screening and virulence–related gene modulation

Abstract

The search for autochthonous food–derived lactobacilli with complementary probiotic–associated and bioprotective characteristics is of considerable interest for functional food development and microbial control. In the present study, lactic acid bacteria (LAB) isolated from traditional Iranian dairy products were screened for in vitro gastrointestinal tolerance, adhesion–related properties, cholesterol removal capacity, antioxidant potential, safety–related phenotypes, and antimicrobial activity. Among 62 initial isolates, ten rod–shaped candidates were selected and taxonomically identified by 16 S rRNA gene sequencing as Levilactobacillus brevis , Limosilactobacillus fermentum , Lactiplantibacillus plantarum , or Lactiplantibacillus pentosus . The selected strains exhibited strain–dependent tolerance to simulated gastrointestinal conditions and bile exposure, accompanied by varying degrees of cell–surface hydrophobicity (CSH), auto–aggregation, and adhesion to Caco–2 cells. Culture supernatants (CSs) derived from these strains effectively inhibited the growth of Listeria monocytogenes and its biofilm–related phenotypes under in vitro conditions; notably, SW16 maintained complete growth inhibition at the highest two–fold dilution evaluated (1:16 of the original CS). Furthermore, exposure to SW16 CS resulted in reduced relative expression of several virulence– and stress–associated genes in L. monocytogenes , including hly , plcB , and sigB . None of the selected candidates displayed detectable hemolytic, DNase, or gelatinase activity. Antibiotic susceptibility profiles obtained by disc diffusion were interpreted solely as a preliminary phenotypic safety screen, given the absence of validated species–specific breakpoints for several of the tested strain–antibiotic combinations; acquired resistance was not assessed genetically. Collectively, the findings demonstrated that these dairy–derived lactobacilli exhibit diverse in vitro probiotic–associated and anti–listerial properties. Nevertheless, because the antimicrobial assays were conducted using laboratory–generated CSs under static experimental conditions; therefore, chemical characterization of the active components, validation in relevant food–matrix models, comprehensive genomic assessment of safety determinants, and in vivo studies are warranted before probiotic functionality or food–level biocontrol efficacy can be established.