AimsThis study evaluated the antibiotic-induced changes in microbial ecology, intestinal dysbiosis and low-grade inflammation; and the combined effect of four different Lactobacillus species on recovery of microbiota composition and improvement of gut barrier function in mice. Methods and ResultsAdministration of the antibiotic ampicillin for 2weeks decreased microbial community diversity, induced caecum tumefaction and increased gut permeability in mice. Application of a probiotic cocktail of four Lactobacillus species (JUP-Y4) modulated the microbiota community structure and promoted the abundance of potentially beneficial bacteria such as Akkermansia. Ampicillin administration led to a decline in Bacteroidetes from 466391% to 026400362%; the addition of JUP-Y4 restored this to 414 +/- 287%. This probiotic supplementation was more effective than natural restoration, where the levels of Bacteroidetes were only restored to 293 +/- 207%. Interestingly, JUP-Y4 treatment was more effective in the restoration of microbiota in faecal samples than in caecal samples. JUP-Y4 also significantly reduced the levels of d-lactate and endotoxin (lipopolysaccharide, LPS) in the serum of mice, and increased the expression of tight-junction proteins while reducing the production of inflammatory cytokines (TNF-, IL-6, MCP-1, IFN- and IL-1) in the ileum and the colon of antibiotic-treated mice. ConclusionsJUP-Y4 not only promoted recovery from antibiotic-induced gut dysbiosis, but also enhanced the function of the gut barrier, reduced inflammation and lowered levels of circulating endotoxin in mice. Significance and Impact of the StudyConsumption of a mixture of Lactobacillus species may encourage faster recovery from antibiotic-induced gut dysbiosis and gut microbiota-related immune disturbance.