Research Brief

Trypanosoma congolense: In Vitro Susceptibility of Bloodstream Forms to Diminazene and Isometamidium

Trypanosoma congolense bloodstream forms were propagated in vitro axenically in a simplified cultivation medium at 34°C. Viability of a drug-sensitive and a drug-resistant clone were examined for 10 days following exposure to 0.1, 1.0 and 10.0 μg ml⁻¹ of diminazene aceturate and 0.1, 1.0 and 10.0 ng ml⁻¹ of isometamidium chloride for various time intervals. Drug-sensitive T. congolense were irreversibly damaged after incubation with 10 μg ml⁻¹ or 1 μg ml⁻¹ diminazene aceturate for 30 min or 2 h, respectively, while drug-resistant trypanosomes were not affected. Exposure to 10 ng ml⁻¹ isometamidium chloride eliminated drug-sensitive trypanosomes after 24 h and drug-resistant trypanosomes after 96 h. The data obtained on in vitro time-dose responses of T. congolense were related to pharmacokinetic data of diminazene and isometamidium in cattle plasma.

In vitro test methods have been developed both for determining the sensitivity of isolates of pathogenic African trypanosomes and for evaluating new compounds for antitrypanosomal activity. The principles of the assays and their main advantages or drawbacks are presented and discussed. In vitro assays which do not require trypanosomes pre-adapted to culture conditions are the Drug Incubation Infectivity Test (DIIT) and the [³H]Hypoxanthine Incorporation Assay. Chemosensitivity tests which do require continuous growth of trypanosomes in vitro include photometric, fluorescence, growth inhibition, long-term viability and metacyclic incubation assays. Evidence is presented to use metacyclic or bloodstream forms in such assays but to avoid procyclic trypanosomes. The drug sensitivity of a homogeneous trypanosome population can be quantified by using photometric, fluorescence or growth inhibition assays lasting 1–3 days. Small numbers of resistant organisms hidden in a sensitive population can be detected employing long-term viability assays (7–10 days). Final selection of the assay to be employed will depend on the parameter to be investigated, and the equipment available.