Laboratory assays reveal diverse phenotypes among microfilariae ofDirofilaria immitisisolates with known macrocyclic lactone susceptibility status

Prevention of canine heartworm disease caused byDirofilaria immitisrelies on chemoprophylaxis with macrocyclic lactone anthelmintics. Alarmingly, there are increased reports ofD.immitisisolates with resistance to macrocyclic lactones and the ability to break through prophylaxis. Yet, there is not a well-established laboratory assay that can utilize biochemical phenotypes of microfilariae to predict drug resistance status. In this study we evaluated laboratory assays measuring cell permeability, metabolism, and P-glycoprotein-mediated efflux. Our assays revealed that trypan blue, propidium iodide staining, and resazurin metabolism could detect differences amongD.immitisisolates but none of these approaches could accurately predict drug susceptibility status for all resistant isolates tested. P-glycoprotein assays suggested that the repertoire of P-gp expression is likely to vary among isolates, and investigation of pharmacological differences among different P-gp genes is warranted. Further research is needed to investigate and optimize laboratory assays forD.immitismicrofilariae, and caution should be applied when adapting cell death assays to drug screening studies for nematode parasites.