Abstract
Background:
Through the production of microbial metabolites, particularly short-chain fatty acids (SCFAs) and tryptophan-derived compounds, the gut microbiome plays a central role in host metabolism and intestinal health. Probiotic bacteria such as Lactobacillus plantarum and Bifidobacterium breve are known to influence the composition of microbial communities and their metabolic activity in the gastrointestinal tract. However, the combined effects of these probiotics on gut microbial families, phylum-level structure, and metabolite production remain unclear.

Aim:
This study aimed to determine whether probiotic treatments using Lactobacillus plantarum, Bifidobacterium breve, and their combination could stimulate broader shifts in the gut microbial community by promoting the abundance of other beneficial probiotic-associated bacterial families and phyla, and to evaluate how these microbial changes influence key gut metabolites.

Methods:
Experimental mice (24 mice) were divided into four groups (6 mice each): Control, Lactobacillus plantarum, Bifidobacterium breve, and Combination treatment groups. Gut microbial composition was analyzed using next generation sequencing (NGS) of the 16S rRNA amplicons, and relative abundances of dominant bacterial families and phyla were calculated. Metabolomic analysis was performed to quantify major microbial metabolites including butyrate, acetate, propionate, indole-3-propionic acid (IPA), and trimethylamine-N-oxide (TMAO).

Results:
At the family level, Lactobacillaceae showed a significant increase in the Combination group compared with the other groups (P < 0.05), while other families including Oscillospiraceae, Lachnospiraceae, Muribaculaceae, Prevotellaceae, Bacteroidaceae, and Akkermansiaceae showed variations that were not statistically significant (P > 0.05). At the phylum level, Bacillota remained the dominant phylum across all groups without significant differences (P > 0.05). Metabolomic analysis demonstrated significant differences in key microbial metabolites. Butyrate, acetate, propionate, indole-3-propionic acid, and trimethylamine-N-oxide showed significant variation among treatments (P < 0.05), indicating that probiotic supplementation influenced microbial metabolic activity in the gut.

Conclusion:
This study showed that probiotic treatments using Lactobacillus plantarum, Bifidobacterium breve, and their combination stimulate broader shifts in the gut microbial community by promoting the abundance of other beneficial probiotic-associated bacterial families and phyla, and to evaluate how these microbial changes influence key gut metabolites.

Key words: Acetate; Gut microbiome; Metabolomics; Probiotics; Short-chain fatty acids.