Rhodtestolin is a cardio-inhibitor that was first discovered in testes extracts of the blood-feeding insect, Rhodnius prolixus. Its role in reproduction remains unconfirmed, but if delivered to the female during spermatophore formation, it may serve to calm the female and/or relax the vaginal muscles to facilitate delivery and storage of the spermatophore. We describe here the anatomy of reproductive organs in R. prolixus and show that rhodtestolin is present in a low-molecular weight fraction of testes extracts separated by gel filtration, as well as in spermatophores delivered to the female during spermatophore formation. We also report that a rhodtestolin-like factor is present in the testes of R. brethesi, Triatoma dimidiata, T. klugi and Nesotriatoma bruneri, other Reduviidae, which are vectors of Chagas disease. Male secretions in insects are known to modify female behavior after copulation, and the presence of rhodtestolin in several genera of Reduviidae suggests that it plays an important role in reproductive success. Determining this role could lead to developing additional population control strategies for these bugs.