Macrocyclic lactones differ in interaction with recombinant P-glycoprotein 9 of the parasitic nematode Cylicocylus elongatus and ketoconazole in a yeast growth assay.

Macrocyclic lactones (MLs) are widely used parasiticides against nematodes and arthropods, but resistance is frequently observed in parasitic nematodes of horses and livestock. Reports claiming resistance or decreased susceptibility in human nematodes are increasing. Since no target site directed ML resistance mechanisms have been identified, non-specific mechanisms were frequently implicated in ML resistance, including P-glycoproteins (Pgps, designated ABCB1 in vertebrates). Nematode genomes encode many different Pgps (e.g. 10 in the sheep parasite Haemonchus contortus). ML transport was shown for mammalian Pgps, Pgps on nematode egg shells, and very recently for Pgp-2 of H. contortus. Here, Pgp-9 from the equine parasite Cylicocyclus elongatus (Cyathostominae) was expressed in a Saccharomyces cerevisiae strain lacking seven endogenous efflux transporters. Pgp was detected on these yeasts by flow cytometry and chemiluminescence using the monoclonal antibody UIC2, which is specific for the active Pgp conformation. In a growth assay, Pgp-9 increased resistance to the fungicides ketoconazole, actinomycin D, valinomycin and daunorubicin, but not to the anthelmintic fungicide thiabendazole. Since no fungicidal activity has been described for MLs, their interaction with Pgp-9 was investigated in an assay involving two drugs: Yeasts were incubated with the highest ketoconazole concentration not affecting growth plus increasing concentrations of MLs to determine competition between or modulation of transport of both drugs. Already equimolar concentrations of ivermectin and eprinomectin inhibited growth, and at fourfold higher ML concentrations growth was virtually abolished. Selamectin and doramectin did not increase susceptibility to ketoconazole at all, although doramectin has been shown previously to strongly interact with human and canine Pgp. An intermediate interaction was observed for moxidectin. This was substantiated by increased binding of UIC2 antibodies in the presence of ivermectin, moxidectin, daunorubicin and ketoconazole but not selamectin. These results demonstrate direct effects of MLs on a recombinant nematode Pgp in an ML-specific manner.

Decreased strongyle egg re-appearance period after treatment with ivermectin and moxidectin in horses in Belgium, Italy and The Netherlands.

The objective of the present study was to evaluate the efficacy of an oral treatment with ivermectin (IVM) or moxidectin (MOX) against gastro-intestinal strongyles in naturally infected horses by performing a faecal egg count reduction test (FECRT) and by monitoring the egg reappearance period (ERP) after treatment. Therefore, a field efficacy study with a randomised complete block design for each study site was conducted, with the individual animal as the experimental unit. At least 10 study sites in Italy, Belgium and The Netherlands were selected and animals were allocated to one of the two treatment groups based on the pre-treatment faecal egg counts (FEC). Animals were treated on Day 0 with an oral paste containing either IVM (at 0.2mg/kg bodyweight) or MOX (at 0.4 mg/kg bodyweight). After treatment, faecal samples were collected at least every fortnight during 56 days after treatment with IVM and during 84 days after MOX treatment. In total, 320 horses on 32 farms were examined. The FECRT on Day 14 indicated a 100% efficacy in 59 of the 64 treatment groups and >92% efficacy in the remaining 5 groups. The ERP was decreased for at least one of the anthelmintics on 17 out of 32 study sites (15 sites or 47% for MOX and 17 sites or 53% for IVM) and on 9 sites (28%) the ERP was decreased for both anthelmintics. On some of these study sites the efficacy declined at the end of the expected ERP, often with good efficacy 2 weeks earlier. Nevertheless, on 1, 3 and 5 study sites in Italy, Belgium and The Netherlands respectively, an efficacy below 90% for IVM and MOX was identified as soon as Day 42 or Day 56. In The Netherlands, the efficacy of IVM was below 90% from Day 28 or Day 35 after treatment on 1 site each. The present study reports a high efficacy of MOX and IVM in a FECRT 14 days after treatment, yet does indicate a shortened ERP for these treatments in more than half of the selected study sites.

Efficacy of major anthelmintics against horse cyathostomins in France.

This paper reports a survey conducted in France during 2011 to evaluate the efficacy of commonly used anthelmintics against horse cyathostomins. A total of 40 farms and 1089 horses were screened for the presence of cyathostomins. All farms but one were positive, with an overall animal infection rate of 53.7%, ranging from 9% to 83% on individual farms. On 445 horses from 30 of these farms, a faecal egg count reduction test (FECRT) was performed to evaluate the efficacy of oral formulations of fenbendazole (FBZ), pyrantel embonate (PYR), ivermectin (IVM) and moxidectin (MOX). Calculation of the mean FECR and 95% confidence intervals (95% CI) around the mean was performed using bootstrap analysis. Resistance to FBZ was found on 17 of 18 farms investigated, with a mean reduction of 57% (95% CI: 38.5-71.2%). Suspected resistance for PYR was found on 6 of 30 farms, and confirmed on another 3 of 30 farms, with a mean reduction for PYR of 94.7% (95% CI: 88.9-98.5%). Reduced efficacy simultaneously of FBZ and PYR was found in 7 farms. Reduced efficacy of IVM was found in one animal on one farm and of MOX in one animal on another farm, and was combined with resistance against FBZ and/or PYR. These results indicate that single and multiple drug resistance and reduced efficacy in equine cyathostomins is present in France. Macrocylic lactones proved to be highly effective compounds against cyathostomins, with reduced efficacy for IVM and MOX in two farms only. These results extend present knowledge on the occurrence of drug resistant cyathostomins in Europe, and illustrate the necessity to use anthelmintics in appropriate worm control programmes.