PROJECT SUMMMARY: Antibiotics are essential medicines to treat bacterial infections. Many broad-spectrum antibiotics, such as oral ciprofloxacin, are commonly prescribed to treat many gastrointestinal infections. Antibiotics also affect the microbiota by altering its diversity and function and sometimes promoting dysbiosis. Although previous studies implied that some beneficial microbes, such as Lactobacilli, have preexisting resistance to various antibiotics, it is still unclear how commonly prescribed oral antibiotics, like ciprofloxacin, selects for Lactobacilli species. This proposal addresses this knowledge gap by first examining the effect of ciprofloxacin exposure in Lactobacilli species. In Aim 1, we will examine the effects of oral ciprofloxacin on the microbiota by identifying changes in the bacterial taxa from ileum and fecal contents through 16SrRNA sequencing and assess the Lactobacilli abundance by qPCR. Then, we will examine the resistance phenotype to ciprofloxacin of different Lactobacilli species using colony forming units and population analysis profile assays. Our preliminary data suggests that Lactobacillus relative abundance increases during ciprofloxacin treatment. Additionally, preliminary in vitro data implies that Lactobacillus tends to have preexisting resistance to ciprofloxacin. Although studies have demonstrated that antibiotics affect the microbiota and host immune responses, it is still unclear how oral ciprofloxacin alters Lactobacillus species which affect the regulatory immune response. Foxp3+ Regulatory T cells (Tregs) and intraepithelial lymphocytes (IELs) are known for their immunoregulatory functions. These cells mediate tolerance against various stimuli, such as nutrients and microbiota, via anti-inflammatory cytokine expression, like IL-10. Many beneficial microbes induce Tregs and other IL-10-producing cells. Lactobacillus has been shown to influence the development of IELs. Additionally, previous studies using antibiotic cocktails have shown that IL-10 production from colonic regulatory T cells decrease during treatment. However, it is still unknown if ciprofloxacin-selected Lactobacilli induces IL-10 production in IELs, especially in the small intestine. In Aim 2, we will examine the changes in IL-10 production of intestinal T cells during ciprofloxacin treatment by treating IL-10 reporter mice with ciprofloxacin. Our preliminary flow cytometry data suggests that ciprofloxacin treatment increases IL-10 production in IELs. Furthermore, preliminary in vitro data suggested that Lactobacillus induces IL-10 production from macrophages. Therefore, Lactobacillus could also promote IL-10 production in IELs during ciprofloxacin treatment. Overall, this project will assess how ciprofloxacin selects for Lactobacillus species that promote IL-10 production in the small intestine. In addition, this work will provide insight on how commonly prescribed antibiotics affect the microbiota and intestinal regulatory immune ...