The In vitro Cultivation of Oesophagostomum radiatum, the Nodular Worm of Cattle. I. Development in Vitamin-Supplemented and Nonsupplemented Media

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Abstract
Using procedures described by Diamond and Douvres (1962. J. Parasit. in press), and certain modifications thereof, development of Oesophagostomum radiatum from the artificially exsheathed infective stage to the 4th molt was obtained free of bacteria and fungi. In addition to trials with a nonsupplemented basic medium, tests were made with the latter supplemented with 1 or more of the following: (1) a vitamin mixture, (2) a phosphate buffer mixture, and (3) buffer extracts of intestinal tissues from a 4-day-old parasite-free calf. There were no apparent differences in the morphology of larvae at comparable stages of development in 5 different media, although development to each stage (3d, 3d molt, 4th, 4th molt) required fewer days in supplemented media than in the basic medium. Maximal yield of larvae in late 4th-stage and 4th molt was obtained in the media containing the buffer extracts of intestinal tissues, but there were no differences in these yields in the other media. Comparisons with in vivo development showed that the larvae were stunted in the 3rd molt, 4th stage, and 4th molt; differentiation of the sexes was delayed; development of the genital primordium lagged behind somatic differentiation; and, in general, more time was required to reach the various stages. Development in different preparations of the same medium showed no differences ascribable to (1) the use of serum from helminth-free or infected calves or yearlings; (2) the age or number of exsheathed infective larvae in the inoculum; (3) liver extract from pigs of different ages; or (4) the frequency of larval transfers to fresh media. Larvae cultured in the vitamin-supplemented medium at 38.5[degree]C advanced to the 4th-stage and 4th molt much more rapidly than at 35 or 37[degree]C.