Gilleard, John StuartRezansoff, Andrew Mischa2018-05-082018-05-082018-04-30Rezansoff, A. M. (2018). Investigating the genetic basis of ivermectin resistance in Haemonchus contortus (Doctoral thesis, University of Calgary, Calgary, Canada). Retrieved from https://prism.ucalgary.ca. doi:10.11575/PRISM/31897http://hdl.handle.net/1880/106615Parasitic nematodes have a major impact on human and animal health and their control is threatened by the emergence of resistance to the anthelmintic drugs on which control depends. Although resistance to ivermectin is widespread in many livestock parasites, the genetic mechanisms remain elusive. In this thesis, a number of approaches are undertaken to investigate the genetics of ivermectin resistance in Haemonchus contortus, an important parasitic nematode research model. A transcriptomic comparison of the susceptible MHco3(ISE) and two ivermectin resistant strains, MHco4(WRS) and MHco10(CAVR), was undertaken to investigate if differences in gene expression revealed insights into ivermectin resistance mechanisms. Although this approach did not reveal clear ivermectin resistance gene candidates, the results have some important implications for RNAseq analysis in genetically diverse organisms. First, extremely high levels of genetic diversity in H. contortus had a major impact on RNAseq analysis and needed to be accounted for reliable identification of differentially expressed genes. Second, there were a remarkably large number of genes differentially expressed between the three H. contortus strains. In a novel genetic crossing approach, two independent serially backcrossed H. contortus populations were utilized in which ivermectin resistance loci had been introgressed into the MHco3(ISE) genome. Markers for six leading candidate ivermectin resistance genes from the literature were examined and none were found to show evidence of introgression in the backcross populations. A seventh marker, Hcms8a20 showed clear evidence of introgression and was later confirmed by collaborators using a whole genome sequencing approach to show its location in a 11.2 Mb introgressed region. A set of 25 in vivo or in vitro phenotyped ivermectin resistant H. contortus field populations were then used to investigate evidence of selection on the Hcms8a20 marker and four other loci distributed across the 11.2 Mb introgressed region. A deep amplicon sequencing approach revealed that, of the loci tested, only Hcms8a20 showed consistent evidence of purifying selection in the ivermectin resistant field populations. Overall, the results provide strong evidence that a major ivermectin resistance locus is located within a few megabases of the Hcms8a20 locus in many different ivermectin resistant H. contortus field populations.engUniversity of Calgary graduate students retain copyright ownership and moral rights for their thesis. You may use this material in any way that is permitted by the Copyright Act or through licensing that has been assigned to the document. For uses that are not allowable under copyright legislation or licensing, you are required to seek permission.Haemonchus contortusH. contortusDrug ResistanceIvermectinAnthelminticParasiteNematodeLivestockSheepRNAseqDeep amplicon sequencingPopulation geneticsBioinformaticsGeneticsBiology--MolecularParasitologyVeterinary Sciencedoctoral thesis10.11575/PRISM/31897