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Plasma and serum from Cape buffalo (Syncerus caffer) kill bloodstream stages of all species of African trypanosomes in vitro. The trypanocidal serum component was isolated by sequential chromatography on hydroxylapatite, protein A-G, Mono Q, and Superose 12. The purified trypanocidal protein had a molecular mass of 150 kDa, and activity correlated with the presence of a 146-kDa polypeptide detected upon reducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Amino acid sequences of three peptide fragments of the 146-kDa reduced polypeptide, ligand affinity and immunoaffinity chromatography of the native protein, and sensitivity to pharmacological inhibitors, identified the trypanocidal material as xanthine oxidase (EC 1.1.3.22). Trypanocidal activity resulted in the inhibition of trypanosome glycolysis and was due to H2O2 produced during catabolism of extracellular xanthine and hypoxanthine by the purine catabolic enzyme.

The high ability of African buffalo, as compared to domestic cattle, to control infections with Trypanosoma brucei brucei ILTat 1.4 organisms did not correlate with the timing or magnitude of parasite surface coat-specific antibody responses and may have resulted from the constitutive presence in buffalo blood of a novel trypanocidal factor. Buffalo plasma and serum contained material that killed bloodstream stage T. b. brucei, T. b. rhodesiense, T. b. gambiense, T. evansi, T. congolense, and T. vivax organisms during four h of incubation at 37 degrees C in vitro. Serum from eland was also trypanocidal whereas serum from oryx, waterbuck, yellow-back duiker, cattle, horse, sheep, goat, mouse, rat, and rabbit was not trypanocidal. The buffalo serum trypanocidal material was not lipoprotein, or IgG, and had the following properties: 1) a density of > 1.24 g/ml determined by flotation ultracentrifugation; 2) insolubility in 50% saturated ammonium sulphate; 3) non-reactivity with anti-bovine IgM, and anti-bovine IgG; 4) non-reactivity with protein G, and protein A; 5) a relative molecular mass of 152 kDa determined by chromatography on Sephacryl S 300, and of 133 kDa determined by chromatography of the 50% SAS cut of IgG-depleted buffalo serum on Superose 12; 6) no associated cholesterol; and 7) inactivation by digestion with proteinase K that was immobilized on agarose.