Drug-resistant Drosophila indicate glutamate-gated chloride channels are targets for the antiparasitics nodulisporic acid and ivermectin

The fruit fly Drosophila melanogaster was used to examine the mode of actio n of the novel insecticide and acaricide nodulisporic acid. Flies resistant to nodulisporic acid were selected by stepwise increasing the dose of drug in the culture media. The resistant strain, glc(1), is at least 20-fold re sistant to nodulisporic acid and 3-fold cross-resistant to the parasiticide ivermectin, and exhibited decreased brood size, decreased locomotion, and bang sensitivity. Binding assays using glc(1) head membranes showed a marke d decrease in the affinity for nodulisporic acid and ivermectin. A combinat ion of genetics and sequencing identified a proline to serine mutation (P29 9S) in the gene coding for the glutamate-gated chloride channel subunit DmG luCl alpha. To examine the effect of this mutation on the biophysical prope rties of DmGluCl alpha channels, it was introduced into a recombinant DmGlu Cl alpha, and RNA encoding wild-type and mutant subunits was injected into Xenopus oocytes. Nodulisporic: acid directly activated wild-type and mutant DmGluCl alpha channels. However. mutant channels were approximate to 10-fo ld less sensitive to activation by nodulisporic acid, as well as ivermectin and the endogenous ligand glutamate, providing direct evidence that noduli sporic acid and ivermectin act on DmGluCl alpha channels.