Growth Inhibitory Effects of Lactobacillus acidophilus, Lactobacillus reuteri, and Bifidobacterium bifidum Supernatant on Blastocystis Subtypes 1 and 3 Via the TLR4 Axis

Background: Blastocystis is a common protozoan parasite found in the human intestinal tract and is associated with several gastrointestinal symptoms as well as inflammatory conditions in the bowel. Probiotics are groups of beneficial microorganisms with a substantial impact on overall human health. Objectives: This study aimed to investigate the growth inhibitory effects of Lactobacillus acidophilus, Lactobacillus reuteri, and Bifidobacterium bifidum on Blastocystis subtypes (ST) 1 and 3 through the evaluation of expression changes in toll-like receptor 4 (TLR4) in Caco2 cell culture. Materials and Methods: The parasite and Caco2 cells were cultured and maintained, and the supernatant of bacteria was prepared. The viability of parasites and cell cultures exposed to supernatants was measured separately by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, in comparison with metronidazole. In addition, TLR4 expression changes in cell cultures and co-cultures exposed to the supernatants were measured using real-time polymerase chain reaction (PCR). Results: The mean viability of Caco2 cells exposed to the highest and lowest (100 vs. 10 μg/mL) concentrations of supernatants was in a similar range. The survival of Blastocystis ST1 and ST3 exposed to the 15, 25, and 35 μg/mL of probiotics supernatants significantly decreased during 24, 48, and 72 hours. Although not statistically significant, the findings indicated a decrease in TLR4 expression in Caco2 cells and an increase in gene expression in co-cultures exposed to the probiotic supernatants (15, 25, and 35 μg/mL). Conclusion: This novel therapeutic field of study using probiotics compounds deserves further exploration to find unprecedented therapies against Blastocystis infection.