title: Analysis of putative inhibitors of anthelmintic resistance mechanisms in cattle gastrointestinal nematodes.
abstract: Effects of the cytochrome P450 inhibitor piperonyl butoxide and the P-glycoprotein inhibitor verapamil on the efficacy of ivermectin and thiabendazole were studied in vitro in susceptible and resistant isolates of the cattle parasitic nematodes Cooperia oncophora and Ostertagia ostertagi. The effects of combined use of drug and piperonyl butoxide/verapamil, respectively, were investigated in the Egg Hatch Assay, the Larval Development Assay and the Larval Migration Inhibition Assay. The effects of piperonyl butoxide and verapamil as inhibitors of thiabendazole and ivermectin responses were particularly marked for larval development, where both inhibitors were able to completely eliminate all differences between susceptible and resistant isolates. Even the lowest concentrations of anthelmintics used in combination with inhibitors caused complete inhibition of development. Differences and/or similarities among responses in different isolates were only obtained in the two other assays: in the Egg Hatch Assay piperonyl butoxide caused a shift in concentration-response curves obtained with thiabendazole to the left for all isolates tested, changing relative differences between isolates. In contrast, an effect of verapamil in the Egg Hatch Assay was only apparent for benzimidazole-resistant isolates. In the Larval Migration Inhibition Assay only ivermectin was tested and piperonyl butoxide shifted the concentration-response curves for all isolates to the left, again eliminating differences in EC50 values between susceptible and resistant isolates. This was not the case using verapamil as an inhibitor, where curves for both susceptible and benzimidazole-resistant isolates shifted to the left in Ostertagia isolates. In Cooperia the picture was more complex with ivermectin-resistant isolates showing a larger shift than the susceptible isolate. Single nucleotide polymorphisms in the β-tubulin isotype 1 gene were investigated. Significantly increased frequencies of resistance-associated alleles were observed for the codons 167 and 200 in one benzimidazole-resistant isolate but not in an isolate selected for benzimidazole resistance at an early stage of selection.
authors.affiliation: Institute for Parasitology and Tropical Veterinary Medicine, Freie Universität Berlin, Robert-von-Ostertag-Str. 7-13, 14136 Berlin, Germany.
authors.forename: Salha
authors.initials: S
authors.lastname: AlGusbi
authors.affiliation: Institute for Parasitology and Tropical Veterinary Medicine, Freie Universität Berlin, Robert-von-Ostertag-Str. 7-13, 14136 Berlin, Germany.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Krücken
authors.affiliation: Institute for Parasitology and Tropical Veterinary Medicine, Freie Universität Berlin, Robert-von-Ostertag-Str. 7-13, 14136 Berlin, Germany.
authors.forename: Sabrina
authors.initials: S
authors.lastname: Ramünke
authors.affiliation: Institute for Parasitology and Tropical Veterinary Medicine, Freie Universität Berlin, Robert-von-Ostertag-Str. 7-13, 14136 Berlin, Germany.
authors.forename: Georg
authors.initials: G
authors.lastname: von Samson-Himmelstjerna
authors.affiliation: Institute for Parasitology and Tropical Veterinary Medicine, Freie Universität Berlin, Robert-von-Ostertag-Str. 7-13, 14136 Berlin, Germany. Electronic address: J.demeler@fu-berlin.de.
authors.forename: Janina
authors.initials: J
authors.lastname: Demeler
mesh_terms: D015766:Albendazole / Q000008:administration & dosage / Q000494:pharmacology; D000818:Animals; D000871:Anthelmintics / Q000494:pharmacology*; D001681:Biological Assay; D002121:Calcium Channel Blockers / Q000008:administration & dosage / Q000494:pharmacology; D002417:Cattle; D004351:Drug Resistance / Q000235:genetics; D004359:Drug Therapy, Combination; D005767:Gastrointestinal Diseases / Q000469:parasitology / Q000662:veterinary*; D007559:Ivermectin / Q000494:pharmacology; D007814:Larva / Q000187:drug effects; D009348:Nematoda / Q000187:drug effects* / Q000235:genetics; D009349:Nematode Infections / Q000469:parasitology / Q000662:veterinary*; D010063:Ovum / Q000187:drug effects; D010574:Pesticide Synergists / Q000008:administration & dosage / Q000494:pharmacology; D010882:Piperonyl Butoxide / Q000008:administration & dosage / Q000494:pharmacology; D020641:Polymorphism, Single Nucleotide; D013827:Thiabendazole / Q000008:administration & dosage / Q000494:pharmacology; D014700:Verapamil / Q000008:administration & dosage / Q000494:pharmacology
chemical_list: D000871:Anthelmintics; D002121:Calcium Channel Blockers; D010574:Pesticide Synergists; D007559:Ivermectin; D014700:Verapamil; D015766:Albendazole; D010882:Piperonyl Butoxide; D013827:Thiabendazole
keywords: Anthelmintic resistance; Cattle nematodes; Cytochrome P450; In vitro assay; P-glycoprotein; Pyrosequencing; β-Tubulin