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Unidentified soluble factors secreted by  E. coli , a frequently isolated microorganism in genitourinary infections, have been reported to inhibit mitochondrial membrane potential (ΔΨm), motility and vitality of human spermatozoa. Here we explore the mechanisms involved in the adverse impact of  E. coli  on sperm motility, focusing mainly on sperm mitochondrial function and possible membrane damage induced by mitochondrial-generated reactive oxygen species (ROS). Furthermore, as lactobacilli, which dominate the vaginal ecosystem of healthy women, have been shown to exert anti-oxidant protective effects on spermatozoa, we also evaluated whether soluble products from these microorganisms could protect spermatozoa against the effects of  E. coli . We assessed motility (by computer-aided semen analysis), ΔΨm (with JC-1 dye by flow cytometry), mitochondrial ROS generation (with MitoSOX red dye by flow cytometry) and membrane lipid-peroxidation (with the fluorophore BODIPY C 11  by flow cytometry) of sperm suspensions exposed to  E. coli  in the presence and in the absence of a combination of 3 selected strains of lactobacilli ( L. brevis ,  L. salivarius ,  L. plantarum ). A Transwell system was used to avoid direct contact between spermatozoa and microorganisms. Soluble products of  E. coli  induced ΔΨm loss, mitochondrial generation of ROS and membrane lipid-peroxidation, resulting in motility loss. Soluble factors of lactobacilli prevented membrane lipid-peroxidation of  E. coli -exposed spermatozoa, thus preserving their motility. In conclusion, sperm motility loss by soluble products of  E. coli  reflects a mitochondrial dysfunction-related membrane lipid-peroxidation. Lactobacilli could protect spermatozoa in the presence of vaginal disorders, by preventing ROS-induced membrane damage.

Soluble Products of Escherichia coli Induce Mitochondrial Dysfunction-Related Sperm Membrane Lipid Peroxidation Which Is Prevented by Lactobacilli