This research was conducted to evaluate the safety and effects of two probiotic candidate strains, Enterococcus faecium (E. faecium) and B. fragilis (BF), which were isolated from healthy sheep, on various factors such as intestinal structure, growth performance, organ indices, and gut microbiota in female C57BL/6J mice. A total of fifty mice were categorized into five groups: BF108, BF109, EF108, EF109, and a control group. The results indicated that mice in the EF108 and BF109 groups experienced a significant increase in body weight by day 28 compared to the control group (p < 0.05). Additionally, the liver index in the BF108 and EF108 groups was significantly lower (p < 0.05) than that of the control group. The length of the colon was also significantly greater (p < 0.05) in the EF109 and BF109 groups relative to the control group. According to the linear discriminant analysis effect size (LEfSe), the presence of probiotics led to an increased relative abundance of Pediococcus acidilactici in the EF109 and BF109 groups, while Lactiplantibacillus plantarum was the most prevalent strain in the EF108 and BF108 groups. Finally, Spearman correlation analysis demonstrated a positive association between Bacillota and Bacteroidota as well as Verrucomicrobiota. Additionally, Bacteroidota displayed positive relationships with Verrucomicrobiota and Planctomycetota, while showing a negative correlation with Actinomycetota and Gemmatimonadota, supporting its potential use as next-generation probiotics. In summary, our results indicate probiotics may have a role in promoting intestinal health by enhancing intestinal shape and histological characteristics.