Integrated Phenotypic-Genotypic Analysis of Candidate Probiotic Weissella Cibaria Strains Isolated from Dairy Cows in Kuwait

Abstract

Probiotics represent a possible strategy for controlling intestinal infections in livestock. Members of the Weissella genus are increasingly being studied for health-related applications in animals and humans. Here we investigated the functional properties of two Weissella cibaria strains isolated from cows reared in Kuwait breeding facilities by combining phenotypic with genomic analyses. W. cibaria SP7 and SP19 exhibited good growth in vitro under acidic conditions and in the presence of bile salts compared to the reference probiotic Lacticaseibacillus (formerly Lactobacillus) rhamnosus GG. Both strains were able to adhere to Caco-2 and HT-29 cell lines, as well as to mucin. The cell-free supernatants of the two isolates exhibited inhibitory activity towards Escherichia coli ATCC 25,922 and Salmonella enterica UC3605, which was ultimately due to the low pH of supernatants. W. cibaria SP19 showed a co-aggregation ability similar to that of L. rhamnosus GG when incubated with S. enterica. Whole genome sequencing and analysis revealed that both strains harbored several genes involved in carbohydrate metabolism and general stress responses, indicating bacterial adaptation to the gastrointestinal environment. We also detected genes involved in the adhesion to host epithelial cells or extracellular matrix. No evidence of acquired antibiotic resistance or hemolytic activity was found in either strain. These findings shed light on the potential of W. cibaria for probiotic use in livestock and on the mechanisms underlying host-microbe interaction in the gut. W. cibaria` strain SP19 exhibited the best combination of in vitro probiotic properties and genetic markers, and is a promising candidate for further investigation.

Keywords: Antagonistic activity; Cattle; Genomics; Intestinal disease; Probiotics; Weissella.

Fig. 1

Survival of W. cibaria SP7 and SP19 at various pH as determined by viable bacteria counts. The probiotic L. rhamnosus GG was included as a control strain. Results are presented as mean±SD of three experiments; *P<0.05, ***P<0.001

Fig. 2

Survival of W. cibaria SP7 and SP19 at different bile salt concentrations as determined by viable bacteria counts. The probiotic L. rhamnosus GG was included as a control strain. Results are presented as mean ± SD of three experiments; **P < 0.01

Fig. 3

Aggregation and co-aggregation of W. cibaria SP7 and SP19 with S. enterica UC3605. The probiotic L. rhamnosus GG was included as a control strain. Results are presented as mean percentage ± SD of three experiments; *P < 0.05, ***P < 0.001

Fig. 4

Adhesion of W. cibaria SP7 and SP19 to Caco-2, HT-29, and mucin. The probiotic L. rhamnosus GG was included as a control strain. Results are presented as mean percentage ± SD of two experiments; **P < 0.01, ***P < 0.001