Despite achieving endoscopic remission, a substantial number of patients with inflammatory bowel disease (IBD), particularly females, continue to experience chronic abdominal pain. The mecha- nisms underlying these sex differences in pain perception remain unclear, but the gut microbiota and sex hormones are potential factors influencing pain sensitivity. The study utilizes a post- inflammatory DSS mouse model of IBD to examine sex-specific differences in the gut microbiota and visceral pain. We hypothesized that the gut microbiota plays a crucial role in modulating chronic visceral pain in females, leading to increased pain compared to ovariectomized females and male mice. Male and female mice were given DSS to induce colitis and thereafter allowed to recover to achieve a post-inflammatory state. Visceral pain was assessed using the visceral motor reflex (VMR) to colorectal distention (CRD). The results demonstrate that cycling female mice exhibit increased visceral pain compared to males and ovariectomized females, despite dis- playing decreased colitis severity compared to males. DSS-induced colitis altered the gut mi- crobial community in both male and cycling female mice, with sex-specific differences observed in metabolic profiles. Short-chain fatty acid (SCFA) levels were increased in post-inflammatory male mice but decreased in females. Furthermore, specific metabolite levels, such as adenine and d-tryptophan, exhibit sex-specific differences in post-inflammatory mice. Using fecal micro- biota transplant (FMT) to colonize germ-free mice with sex-hormone-appropriate and opposite sex-hormone microbiota, showed that the female post-inflammatory microbiota increased visceral hypersensitivity in both female and male recipient mice. These findings highlight the importance of sex-specific differences in the post-inflammatory gut microbiota and their influence on chronic pain. Understanding role of these differences in post-inflammatory visceral pain in IBD is crucial and may lead to the development of novel therapeutic approaches for managing chronic pain.