Abstract Onchocerciasis, a debilitating eye and skin disease, is caused by an obligatory human parasitic filarial worm Onchocerca volvulus. It is the world's second leading infectious cause of blindness in humans, primarily in Sub-Saharan Africa, that is being targeted for elimination. The strategy for elimination of O. volvulus focuses currently on controlling transmission through ivermectin-based mass drug administration programs. However, this treatment does not kill the adult parasites, which can live and reproduce for more than 10 years within their infected host. Recent reports suggest that onchocerciasis cannot be eliminated through mass drug administration with ivermectin solely. In addition, ivermectin is contraindicated in areas of marked co- endemicity with Loa loa, where the risk of severe adverse events is associated with high levels of circulating L. loa microfilariae. Furthermore, the potential for ivermectin resistance, the lack of its macrofilaricidal activity, and the long time line (>20 years) for transmission interruption has prompted a call for research into the development of new tools (macrofilaricidal drugs that kill adult worms, diagnostics and vaccines), the basis of which relies on a comprehensive understanding of the parasite biology and parasite-human host interaction. Humans are the only definitive host for O. volvulus, where the adult worms reside within highly vascularized and encapsulated nodules. Because there are no existing small animal models for propagating the life cycle of O. volvulus, the adult parasites must be obtained surgically from subcutaneous nodules. Moreover, no one has yet been able to culture O. volvulus in vitro from the vector-derived infective stage larvae (L3) to the adult stages. Using optimized two-dimensional (2-D) culturing systems, we were able to demonstrate for the first time that O. volvulus larvae can molt twice and develop to the early pre-adult male and female L5 stages. The 2-D cell culture, although promising, does not adequately reproduce the proper microenvironment for the parasite's development. We therefore propose to develop 3-D in vitro culturing systems that would better mimic the in vivo mammalian subcutaneous tissues where this obligatory human parasite resides. Thus, the major goal of this exploratory, high-risk proposal is to integrate our knowledge of the biology of O. volvulus with the existing expertise in tissue engineering, and adapt various in vitro skin tissue models for optimum worm development and long term survival. Our proposed studies will lead to novel in vitro culturing 3-D systems for an important human parasite. It will also provide a new screening platform to ascertain the activity of lead macrofilaricidal candidates. Moreover, it will provide the filarial research community with an access to well- developed O. volvulus worms at different developmental stages, and thus provide valuable source of material to address and accelerate innovative unsolved biomedical inquiries in parasite's biology that could not have been tackled before.