The intake of probiotic lactic acid bacteria not only promotes digestion through the microbiome regulated host intestinal metabolism but also improves diseases such as irritable bowel syndrome and inflammatory bowel disease, and suppresses pathogenic harmful bacteria. This investigation aimed to evaluate the immunomodulatory effects in intestinal epithelial cells and to study the clinical efficacy of the selected the Bifidobacterium breve and Bifidobacterium longum groups. The physiological and biochemical properties were characterized, and immunomodulatory activity was measured against pathogenic bacteria. In order to find out the mechanism of inflammatory action of the eight viable and sonicated Bifidobacterium spp., we tried to confirm the changes in the pro-inflammatory cytokines (TNF-α, interleukin (IL)-6, IL-12) and anti-inflammatory cytokine (IL-10), and chemokines, (monocyte chemoattractant protein-1, IL-8) and inflammatory enzymatic mediator (nitric oxide) against Enterococcus faecalis ATCC 29212 infection in Caco-2 cells and RAW 264.7 cells. The clinical efficacy of the selected B. breve and B. longum group was studied as a probiotic adjuvant for acute diarrhea in children by oral administration. The results showed significant immunomodulatory effects on the expression levels of TNF-α, IL-6, IL-12, MCP-1, IL-8 and NO, in sonicated Bifidobacterium extracts and viable bifidobacteria. Moreover, each of the Bifidobacterium strains was found to react more specifically to different cytokines. However, treatment with sonicated Bifidobacterium extracts showed a more significant effect compared to treatment with the viable bacteria. We suggest that probiotics functions should be subdivided according to individual characteristics, and that personalized probiotics should be designed to address individual applications.