ivermectin

Development of the larval migration inhibition test for comparative analysis of ivermectin sensitivity in cyathostomin populations

Citation

Claire McArthur, Ian G Handel, Ailie Robinson, Jane Hodgkinson, Barend MdeC Bronsvoort, Faith A. Burden, Ray Kaplan, Jacqui. B. Matthews. June 2015. Development of the larval migration inhibition test for comparative analysis of ivermectin sensitivity in cyathostomin populations. Veterinary Parasitology.

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Publication date: 
20 June 2015
DOI number: 
10.1016/j.vetpar.2015.06.019
Abstract

Cyathostomins are the most prevalent parasitic pathogens of equids worldwide. These nematodes have been controlled using broad-spectrum anthelmintics; however, cyathostomin resistance to each anthelmintic class has been reported and populations insensitive to more than one class are relatively commonplace. The faecal egg count reduction test (FECRT) is considered the most suitable method for screening anthelmintic sensitivity in horses, but is subject to variation and is relatively time-consuming to perform. Here, we describe a larval migration inhibition test (LMIT) to assess ivermectin (IVM) sensitivity in cyathostomin populations. This test measures the paralysing effect of IVM on the ability of third stage larvae (L3) to migrate through a pore mesh. When L3 from a single faecal sample were examined on multiple occasions, variation in migration was observed: this was associated with the length of time that the L3 had been stored before testing but the association was not significant. Half maximal effective concentration (EC50) values were then obtained for cyathostomin L3 from six populations of horses or donkeys that showed varying sensitivity to IVM in previous FECRTs. Larvae from populations indicated as IVM resistant by FECRT displayed significantly higher EC50 values in the LMIT than L3 from populations classified as IVM sensitive or L3 from populations that had not been previously exposed to IVM or had limited prior exposure. The analysis also showed that EC50 values obtained using L3 from animals in which IVM faecal egg count reduction (FECR) levels had been recorded as <95% were significantly higher than EC50 values obtained using L3 from animals for which FECR was measured as >95%. For one of the populations, time that had elapsed since IVM administration had an effect on the EC50 value obtained, with a longer time since treatment associated with lower EC50 values. These results indicate that the LMIT has value in discriminating IVM sensitivity amongst cyathostomin populations, but several factors were identified that need to be taken into account when executing the test and interpreting the derived data.

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The P-glycoprotein inhibitor ketoconazole causes a reversion to ivermectin sensitivity in cyathostomins in vitro

Citation

Laura Peachey, Jacqui. B. Matthews, Gina L. Pinchbeck, Faith A. Burden, Nikki Stradling, Jane Hodgkinson. The P-glycoprotein inhibitor ketoconazole causes a reversion to ivermectin sensitivity in cyathostomins in vitro. Presented at British Society for Parasitology Spring Meeting.

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Date presented: 
Monday 7 April 2014
Abstract

Anthelmintic resistance is a major veterinary and public health issue globally, of most concern is the level of resistance to the macrocyclic lactones. Recent studies have identified a role in resistance for the ATP binding cassette (ABC) drug transporters, P-glycoproteins (P-gps). This study demonstrates the effect of the P-gp inhibitor ketoconazole on the efficacy of ivermectin (IVM) against equid cyathostomin larvae using the larval migration inhibition test (LMIT). Third stage cyathostomin larvae (L3) were cultured from two populations; 1) with recent history of IVM resistance in vivo and 2) naive to anthelmintic exposure. The sensitivity to IVM in each group (n=8) was characterised using the LMIT. The IVM LMIT was repeated for each sample with and without the addition of 10µM ketoconazole. Probit analysis was performed on grouped data from each population to give LC-50 values. The LC-50 value for IVM in Populations 1 and 2 was 4.9 and 2.4µg/ml respectively indicating that Population 1 has a resistant phenotype in comparison to Population 2. Addition of 10µM ketoconazole to IVM in Population 1 caused a drop in LC-50 value from 5.8 to 1.6µg/ml. In Population 2 the effect of the addition of ketoconazole was negligible (1.1 to 0.9µg/ml). This study demonstrates that the P-gp inhibitor ketoconazole causes reversion to a sensitive phenotype in IVM-resistant cyathostomins, inferring that P-gps play a role in their resistance to IVM. This work will be corroborated by investigation into P-gp genes and their expression in cyathostomins.

Online references

The P-glycoprotein inhibitor ketoconazole causes a reversion to sensitivity in ivermectin resistant cyathostomins in vitro

Citation

Laura Peachey, Jacqui. B. Matthews, Gina L. Pinchbeck, Faith A. Burden, Jane Hodgkinson. The P-glycoprotein inhibitor ketoconazole causes a reversion to sensitivity in ivermectin resistant cyathostomins in vitro. Presented at Anthelmintics: From Discovery to Resistance. (5 February - 7 February 2014). San Francisco, USA.

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Presentation details
Date presented: 
Thursday 6 February 2014
Abstract

Anthelmintic resistance is a growing problem in both the developed and developing world; of most concern is the level of resistance detected against the potent macrocylic lactone (ML) anthelmintics. To identify and target a common mechanism of resistance to anthelmintics would allow potential modification of existing drugs, and may even enable the prediction and prevention of the development of resistance to novel drugs. There is a growing body of evidence that P-glycoproteins (P-gps) are involved in resistance to the MLs in many parasitic nematodes of humans and veterinary species (Ardelli et al, 2011). P-gps belong to class two of the ATP binding cassette (ABC) transporter protein superfamily; they are responsible for the active removal of xenobiotic compounds from cells. The cyathostomins are gastrointestinal nematodes of equids that cause significant pathology. Recently resistance to MLs has been described in cyathostomins (Molento et al, 2008), its mechanisms have not yet been elucidated.

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