Single worm genotyping demonstrates that Onchocerca ochengi females simultaneously produce progeny sired by different males

Onchocerca ochengi is a filarial nematode parasite of African cattle and most closely related to Onchocerca volvulus, the causing agent of river blindness. O. ochengi females induce the formation of a nodule in the dermis of the host, in which they remain sedentary in very close association with the host's tissue. Males, which do not adhere to the host's tissue, are also found within the nodules at an average number of about one male per nodule. Young O. ochengi females tend to avoid the immediate proximity of existing nodules. Therefore, O. ochengi nodules are dispersed in the ventral inguinal skin at considerable distances from each other. It has been speculated that males avoid the risk of leaving a female once they have found one and remain in the nodule as territorial males rendering the reproductive strategy of O. ochengi essentially monogamous. We developed a protocol that allows reliable PCR amplification of single copy loci from different developmental stages of O. ochengi including embryos and microfilariae. From 32 O. ochengi nodules, we genotyped the female worms and the 67 adult male worms, found in these nodules, together with a fraction of the progeny from within the uteri of females. In 18 of 32 gravid females progeny derived from multiple males were found. In five nodules, the males isolated from the same nodule as the female were not sufficient to explain the genotypes of the entire progeny. We conclude that frequently O. ochengi females simultaneously produce progeny sired by different males and that most but not all males are still present in the nodule when their offspring is ready to hatch.