Isolates of avian trypanosomes from nestling sparrowhawks (Accipiter nisus) and from Simulium latipes were compared by isoenzyme electrophoresis with the previously described avian trypanosomes Trypanosoma corvi and T. everetti. Simulium isolates developed into trypomastigotes in semi-defined medium at 37 degrees C confirming that they belong to the genus Trypanosoma. Cellulose acetate electrophoresis (CAE) and polyacrylamide gel electrophoresis (PAGE) methods were employed. A total of 11 enzymes was examined, of which 8 gave satisfactory results for all lysates (MDH, ICD, PGM, 6PGD, SOD, PEP I, PEP II and PEP D). The zymograms of 3 isolates from Simulium and 4 from A. nisus were similar indicating that they were the same organism and the isoenzyme patterns coincided with those of T. corvi. It is concluded that A. nisus and S. latipes are infected with T. corvi. Flagellate infections were found in the midgut, hindgut and rectum of Simulium where large numbers of epimastigotes were found in the lumen and attached to the cuticular intima of the rectal ampullae by hemidesmosomes at the distal end of an expanded flagellum. Out of 285 nestling sparrowhawks 13 (5%) were infected with trypanosomes. In view of their age and inability to preen themselves it is suggested they became infected via infected prey.