Development of a high-throughput diagnostic screening tool to monitor the status of amitraz resistance and genotype in the cattle tick, Rhipicephalus microplus

Abstract

Acaricide resistance against all major classes of acaricides has been reported in tick populations around the world. In South Africa, amitraz is one of the most commonly used acaricides, with resistance being reported in both of the Rhipicephalus cattle tick species, namely Rhipicephalus microplus and R. decoloratus. Conventional methods to detect acaricide resistance, such as larval packet tests or adult immersion tests, are time-consuming assays that do not provide any information on tick genotype. This is essential for the identification of areas where resistance is emerging to allow for early intervention before homozygous resistant tick populations are established. Due to these limitations, new rapid detection approaches are needed to streamline resistance screening protocols to ensure knowledge-based intervention.

In this study, the current amitraz resistance status of R. microplus ticks in the Mnisi communal area was investigated, where it was found that 87% of the population was heterozygous. Results indicated that the heterozygous genotype is being maintained in the population despite the application of a continued selection pressure over time, which suggests that there could be a fitness cost associated with amitraz resistance. In addition, TaqMan SNP genotyping assays were designed and tested as a high-throughput diagnostic screening tool to improve the turnover time of genetic testing, where hundreds of samples can be analysed for an array of resistance associated genetic markers within hours. Two TaqMan SNP genotyping assays were designed to genotype the two SNPs in the octopamine/tyramine receptor gene that have been linked to amitraz resistance. The Oct 2 assay shows potential for the diagnostic screening of R. microplus field populations, although further testing is required. An ITS2 TaqMan SNP genotyping assay was also designed with the aim of rapidly differentiating between R. microplus and R. decoloratus ticks. This assay did not prove to be successful in initial testing but the concept may still hold potential for future studies.

This project will allow us to expand our database on the current status of amitraz resistance in the Mnisi communal area to put forward a strategy for improved acaricide use in collaboration with government and our industry partners. This will benefit the agricultural industry as well as the South African economy by reducing capital losses to farmers as they will be able to purchase the correct acaricides and use them in a more effective way. This will also impact veterinary health disciplines in South Africa by decreasing the tick burden on cattle, which will hinder the spread of resistant ticks and potentially lessen the transmission of tick-borne disease.