Date of Award
Doctor of Philosophy
Jeffrey K. Beetham
Treatment of parasitic nematodes infections is generally limited to one of three major drug classes; resistance to these is an increasing problem. Because development of new drugs and drug classes is expensive and slow it is important to understand how resistance to current drugs occurs. Nicotinic acetylcholine receptors provide drug targets for both the nicotinic agonist and amino-acetonitrile derivative anthelminthic classes. Much of the research on resistance to nicotinic agonists has been performed in the free-living nematode Caenorhabditis elegans. This research has by necessity looked at limited gene sets which present an incomplete picture of what is believed to be a polygenic trait. Attempts to reproduce this research in parasitic species have shown that research in C. elegans does not always translate to parasites.
We used second generation sequencing to obtain a broader view of resistance in a parasitic nematode, Oesophagostomum dentatum, than is easily accomplished with traditional molecular methods. Because O. dentatum lacks a sequenced genome it was first necessary to identify mRNA sequences for genes shown in other genera/species to associate with resistance. We developed a method of assembly that produces longer sequences than traditional assembly methods and used that to identify mRNAs for 34 genes associated with resistance to levamisole and other major anthelminthics.
With this sequence information we assessed the expression levels and sequence changes in the levamisole resistance associated genes between levamisole-sensitive and -resistant nematode isolates. We identified 9 mRNAs exhibiting at least a 2-fold decrease in expression between the two isolates and 72 non-synonymous SNPs. We have used this information to propose that levamisole resistance in this parasitic model associates with decreased abundance of functional receptors containing UNC-38 and/or UNC-63, as well as decreased signal transduction moderated by LEV-10, NRA-1, RIC-3, LEV-11, UNC-22, and UNC-68ry.
Romine, Nathan, "Transcriptomic identification and characterization of levamisole resistance associated genes in the swine nodular worm Oesophagostomum dentatum" (2012). Graduate Theses and Dissertations. 12836.