Species coexistence is shaped by a range of biotic and abiotic factors. Beyond predation, parasitism, and competition, one species may interfere with another’s reproduction to induce sexual exclusion from a habitat. Here, we test for reproductive interference from inter-species mating between sympatric nematodes Caenorhabditis macrosperma and C. nouraguensis. Higher intrinsic population growth of C. nouraguensis arises from greater fecundity of both sexes, predicting it to be superior in resource competition. Mate discrimination between species is incomplete, however, with inter-species mating errors reducing lifespan and reproductive fitness of female C. nouraguensis only. These asymmetric costs arise within hours, due to ectopic migration of C. macrosperma’s giant sperm cells. We modelled the population dynamic impacts of reproductive interference, then confirmed rapid sexual exclusion in mixed-species communities with multi-generation experiments. These findings demonstrate the profound ecological implications of reproductive interference for demographic parameters and species coexistence through a cell-mediated mechanism of inter-species harm.