Maintenance of infectivity of Trypanosoma congolense in vitro with explants of infected skin at 37 degrees C

Glossina morsitans infected with two stocks of Trypanosoma congolense were fed on rabbits and calves to produce local skin reactions containing trypanosomes. Areas of infected skin were removed from the animals and used to prepare dermal explant cultures in Eagle's MEM and RPMI 1640 culture medium, supplemented with foetal bovine serum and containing penicillin and streptomycin. Cultures were incubated at 37 °C and media were changed at 24 to 48 hour intervals to maintain pH 7·0 to 7·2. There was evidence of trypanosome multiplication in explant cultures set up in both media; one trypanosome stock was maintained equally well in both Eagle's MEM and RPMI 1640, but the other stock survived better in Eagle's MEM. Explant cultures prepared from calf tissues generally yielded more trypanosomes at 24 hours than those prepared from rabbit tissues. The numbers of parasites present near the explants at 24 hours were maintained for up to 14 to 15 days before a decline in parasite concentration occurred. The organisms retained typical blood stream trypomastigote morphology and were infective for mice for periods up to 21 days. The trypanosomes growing in primary explant cultures could not be subpassaged in culture media alone or on to monolayers of fibroblast-like cells of bovine, murine or buffalo origin. Attempts to establish primary cultures by placing infected skin explants directly on to similar monolayers were also unsuccessful.