An amino acid substitution in Fasciola hepatica P-glycoprotein associated with triclabendazole-resistant populations

Control of infections by the Common Liver Fluke (Fasciola hepatica) is threatened by the development of resistance to the anthelmintic of choice, triclabendazole (TCBZ). Enhanced drug efflux by ABC transporters such as P-glycoprotein (P-gp) has been implicated in the development of TCBZ- resistance. A putative full length cDNA coding for a P-gp expressed in adult Fasciola hepatica has been constructed and used to design a PCR primer set for amplifying a region encoding part of the second nucleotide binding domain of P-gp. Application of this primer set to genomic DNA derived from liver flukes from TCBZ-resistant and -susceptible field populations shows a significant difference in the alleles present in the two populations, supporting the hypothesis that drug efflux pumps play a role in resistance to TCBZ in the liver fluke. Analysis of an allele present at a three-fold higher frequency in the “resistant” population revealed that it is characterised by a single nucleotide polymorphism (SNP) producing a serine to arginine substitution at residue 1144. Homology modelling studies have been used to locate this site in the P-gp structure and hence assess its potential to modify functional activity. This SNP has the potential to form the basis of a molecular test for TCBZ-resistance based on genomic DNA extracted from fluke eggs.