Molecular cloning and characterization of glutamate-gated chloride channels and implications of ivermectin resistance mechanisms in the cattle tick, Rhipicephalus microplus

Rhipicephalus microplus is a primary vector for Babesia bigemina and Babesia bovis, the agents of bovine babesiosis, as well as Anaplasma marginale, the agent of bovine anaplasmosis. These tick-borne diseases cause substantial economic losses in the livestock industry, especially in tropical and subtropical regions. Effective tick control is challenged by the emergence of acaricide resistance, particularly to ivermectin (IVM), which targets the glutamate-gated chloride (GluCl) channels in ticks. In this study, we conducted molecular cloning and characterization of the GluCl channel in R. microplus, and first reported the existence of alternative splicing at the 5’ end of the GluCl channel. We identified two new exon regions and six different alternative splicing isoforms. We then examined the distribution of these isoforms in an IVM-resistant El Zamora strain, a Mexico-imported Merida strain and a susceptible laboratory-maintained Deutch strain. Homological modeling and docking analysis indicated differential binding affinities of these isoforms to IVM and L-glutamate. We also identified several SNPs of the GluCl channel only found in IVM-resistant strain. Our findings suggest that specific alternative splicing patterns and several SNP candidates of the GluCl channel might contribute to IVM resistance in R. microplus. Understanding these mechanisms is crucial for developing new strategies to manage acaricide resistance and improve tick control measures.