Chronic arsenic-fluoride co-exposure impairs spermatogenesis with gut microbiota-associated bile acid and lipid metabolic remodeling.

Arsenic and fluoride frequently co-occur in groundwater, but the gut-associated metabolic changes accompanying male reproductive toxicity from their co-exposure remain unclear. We established a chronic developmental drinking water co-exposure model in Sprague-Dawley rats using 100 mg/L NaF and 75 mg/L NaAsO2. Reproductive outcomes were evaluated at postnatal day (PND) 60 and PND120, and reciprocal fecal transfer was performed between control and arsenic-fluoride (AsF) backgrounds. Developmental AsF exposure impaired sperm production and quality, disrupted reproductive hormone homeostasis, altered spermatogenic-stage markers, and induced testicular and epididymal injury. Repeated administration of AsF-donor fecal material to control recipients was accompanied by selected adverse reproductive changes, whereas control-donor material was accompanied by partial changes in the opposite direction in AsF-exposed recipients. Endpoint 16S rRNA sequencing demonstrated exposure-associated microbial community differences, and donor-material administration was accompanied by changes in selected host phenotypes. Untargeted serum metabolomics revealed prominent remodeling of bile acid- and lipid-related signals, including reciprocal Fecal microbiota transplantation-associated patterns and exploratory relationships between bile-acid-related features and reproductive hormones. These metabolic findings were accompanied by differences in colonic junction- and inflammation-related markers, intestinal-hepatic FXR-FGF15-related proteins, hepatic bile acid and lipid endpoints, and testicular autophagy-related markers. Collectively, the findings characterize reproductive hazards at the tested concentrations and support a nonexclusive gut-liver-testis framework in which donor-material-associated microbial and metabolic changes accompany partial modification of recipient phenotypes.