Although probiotics are increasingly used in irritable bowel syndrome (IBS), their mechanism of action has not been elucidated sufficiently. We aimed to evaluate the impact of a multispecies probiotic on enteric microbiota composition in women with diarrhea-predominant-IBS (IBS-D) and to determine whether these effects are associated with changes in IBS symptoms or inflammatory markers. In a double-blind, placebo-controlled study, Rome III IBS-D women completed a two-week run-in period and eligible women were assigned at random to a probiotic capsule (BIO-25) or an indistinguishable placebo, twice daily for 8weeks. IBS symptoms and stool consistency were rated daily by visual analogue scales and the Bristol stool scale. High sensitivity C-reactive protein, fecal calprotectin and microbial composition were tested at baseline and at 4 and 8weeks. Microbial sequencing of the 16S rRNA was performed and data were analyzed to compare patients who responded to treatment with those who did not. 172 IBS-D patients were recruited and 107 eligible patients were allocated to the intervention (n=54) or placebo (n=53) group. Compared to placebo, BIO-25 did not result in changes in microbial diversity or taxa proportions, except for higher relative proportions of Lactobacillus in the BIO-25 group (P=0.002). Symptomatic responders to BIO-25 showed a reduction in the proportion of Bilophila(P=0.003) posttreatment. Patients with beneficial inflammatory-marker changes had higher baseline proportions of Faecalibacterium(P=0.03), Leuconostoc (P=0.03), and Odoribacter (P=0.05) compared to corresponding non-responders. Identifying patients with a more amenable microbiome at treatment initiation may result in better treatment response.