Repurposing of currently marketed compounds with proven efficacy against apicomplexan parasites was used as an approach to define novel candidate
therapeutics for bovine
besnoitiosis. Besnoitia besnoiti tachyzoites grown in MARC-145 cells were exposed to different concentrations of
toltrazuril,
diclazuril,
imidocarb,
decoquinate,
sulfadiazine and
trimethoprim alone or in combination with
sulfadiazine. Drugs were added either just prior to
infection of MARC-145 cells (0 h post
infection, hpi) or at 6 hpi. A primary evaluation of
drug effects was done by direct immunofluorescence staining and counting. Potential effects on the host cells were assessed using a XTT kit for cell proliferation. Compounds displaying promising efficacy were selected for IC50 and IC99 determination by qPCR. In addition, the impact of drugs on the tachyzoite ultrastructure was assessed by TEM and long-term treatment assays were performed. Cytotoxicity assays confirmed that none of the compounds affected the host cells.
Decoquinate and
diclazuril displayed invasion inhibition rates of 90 and 83% at 0 h pi and 73 and 72% at 6 h pi, respectively. The remaining drugs showed lower efficacy and were not further studied.
Decoquinate and
diclazuril exhibited IC99 values of 100 nM and 29.9 μM, respectively. TEM showed that
decoquinate primarily affected the parasite mitochondrium, whilst
diclazuril interfered in cytokinesis of daughter zoites. The present study demonstrates the efficacy of
diclazuril and
decoquinate against B. besnoiti in vitro and further assessments of safety and efficacy of both drugs should be performed in the target species.