It is proposed to continue studies on the developmental biology of helminths using in vitro culture techniques as a tool. Nutritional requirements will focus on sterols, particularly for the free-living stages of microbial- feeding species. Studies will be made of the morphologic changes exhibited by sterol-deficient nematodes using light and electron microscopy. The growth and differentiation of coelomocytes during all stages of the life cycle of Nipprostrongylus brasiliensis and other nematodes will be described by light, transmission and scanning electron microscopy as a baseline study to obtain information on the function(s) of this cell. The uptake and release of vitamin B12 by coelomocytes will be investigated using H3 - and Co57-B12 in autoradiographic studies, and quantitatively by use of a gamma spectrometer, and by assay with Ochromonas and the radiosorbent procedure. The effect of larval and adult Spirometra mansonoides on the vitamin B12 content of the host wil be determined, and the role of vitamin B12 in cestode metabolism will be studied, particularly with regard to propionate metabolism and energy producing reactions. Vitamin B12-cestode relations will be explored in greater depth to determine if a true dichotomy exists in uptake and concentration between different orders. An analysis will be made to relate the data obtained to cestode evolutionary patterns. An important aim of the study is to improve techniques for the culture of parasitic nematodes, so that the entire life cycle of selected species will develop in vitro and in sequential passage.