Gene expression changes in a P-glycoprotein ( Tci-pgp-9) putatively associated with ivermectin resistance in Teladorsagia circumcincta

Abstract

Anthelmintic resistance in parasitic nematodes of small ruminants is widespread and, in some parts of the world, threatens the sustainability of sheep production. The genetic changes underlying resistance to anthelmintics, particularly ivermectin (IVM), remain to be determined. The majority of studies to date have investigated target site mutations; relatively little attention has been paid to the role of changes in gene expression. In this study, we investigated the expression of putative drug transporter molecules, P-glycoproteins (Pgps), in Teladorsagia circumcincta, the predominant parasitic nematode species of sheep in the UK and the major anthelmintic resistant species. Utilising a degenerate PCR approach, 11 partial Pgp sequences were identified. Constitutive differences in gene expression between an IVM-susceptible (MTci2) and a multidrug-resistant (MTci5) isolate were determined for 10 of the Pgps using the ΔΔCt TaqMan® real-time PCR method. Gene expression differences were particularly marked in one of these genes, namely Tci-pgp-9. In the MTci5 isolate, statistically significant increases in Tci-pgp-9 expression, at the mRNA level, were observed across all life-cycle stages and most notably in eggs (55-fold increase). Comparison of the partial Tci-pgp-9 nucleotide sequences from MTci2 and MTci5 also identified high levels of polymorphism. This work has shown that constitutively increased expression in Tci-pgp-9, coupled with increased sequence polymorphism, could play a role in allowing multidrug-resistant T. circumcincta to survive IVM exposure. The genetic changes underpinning these gene expression changes remain to be elucidated and need to be investigated in other isolates. These changes could form the basis of an IVM resistance marker to monitor the spread of resistance and to evaluate management practices aimed at delaying its spread.