Development of a novel in vitro equine anthelmintic assay

An in vitro assay involving the use of a horse strongyle (Strongylus edentatus) and the micromotility meter has been developed to test for equine anthelmintic activity. Three commercially available equine anthelmintics (dichlorvos, ivermectin, and pyrantel pamoate) and an investigational drug (p-toluoyl chloride phenylhydrazone) were evaluated in this assay at four concentrations. After a 24-h incubation, greater than or equal to 10 micrograms/ml of all four drug treatments significantly (P less than or equal to 0.05) reduced the motility of ensheathed L-3 S. edentatus larvae, thereby indicating anthelmintic activity. Pyrantel pamoate also reduced motility at 1 microgram/ml, while the hydrazone significantly increased movement at this level. At 0.1 microgram/ml, none of the treatments significantly reduced motility; one treatment (dichlorvos) significantly increased larval motility. Incubation for 48 h resulted in significant activity (reduction in motility) at greater than or equal to 1 microgram/ml with two drugs (ivermectin, pyrantel pamoate); dichlorvos and the hydrazone reduced motility at greater than or equal to 10 micrograms/ml. None of the treatments significantly reduced motility at the lowest concentration (0.1 microgram/ml); however, at 48 h, two treatments (dichlorvos, hydrazone) significantly increased motility at the lowest concentration (0.1 microgram/ml). The in vitro S. edentatus motility assay proved to be sensitive, accurate and rapid. This assay system should be a valuable addition to tests used to identify potential equine anthelmintics, monitor helminth resistance to drugs, and perhaps define the kinetics and mode of action for drugs.