In vitro drug susceptibility of Theileria equi and the role of abc transporters in resistance to imidocarb dipropionate

The primary work presented involves evaluation of the apicomplexan hemoparasite Theileria equi for imidocarb dipropionate resistance, assessment of possible alternative treatment options, and investigation of the potential role for ATP-binding cassette transporters in drug resistance. An in vitro drug susceptibility assay was developed and adapted to multiple different drugs. The susceptibility of two T. equi isolates and a lab generated variant to both imidocarb dipropionate and the experimental bumped kinase inhibitor 1294 was assessed. Two distinct isolates of T. equi had an almost four-fold difference in imidocarb dipropionate susceptibility, and a variant generated through continuous in vitro imidocarb dipropionate exposure developed a further 15-fold decrease in susceptibility. In contrast, there was no difference in susceptibility to bumped kinase inhibitor 1294, suggesting that these drugs may represent a possible alternative for resistant T. equi parasites. Also presented is the failure of four T. equi infected ponies to clear infection with the USDA Florida strain despite two rounds of imidocarb dipropionate treatment, a demonstration of drug resistance in vivo.;This collective data raises concern for the existence and emergence of drug resistance at the parasite level. An investigation was undertaken of the 45 putative T. equi ATP-binding cassette transporters as potential mediators of imidocarb dipropionate efflux leading to drug resistance. Selection of the most likely candidate transporters based on differential transcription was accomplished using RNA sequencing. No experiments performed in this study supported imidocarb dipropionate as a substrate for efflux by the most differentially expressed ABC transporter BEWA_032300, however there are other transporters of interest and other mechanisms by which ABC transporters can contribute to drug resistance for further investigation.;The described drug assay was utilized in a collaborative project to examine the effect of environmental exposures on the stability and efficacy of buparvaquone and parvaquone. T. equi was used as a model organism for Theileria parva, for which these drugs are vital in prophylaxis and treatment. It was found that air exposure causes significant drug degradation and subsequent loss of efficacy, a finding that has implications for successful treatment as well as the prevention of drug resistance development.