Quantifying the fitness effects of resistance alleles with and without anthelmintic selection pressure usingCaenorhabditis elegans

Abstract

Albendazole and ivermectin are the two most commonly co-administered anthelmintic drugs in mass-drug administration programs worldwide. Despite emerging resistance, we do not fully understand the mechanisms of resistance to these drugs nor the consequences of delivering them in combination. Albendazole resistance has primarily been attributed to variation in the drug target, a beta-tubulin gene. Ivermectin targets glutamate-gated chloride channel (GluCl) genes, but it is unknown whether these genes are involved in ivermectin resistance in nature. Using Caenorhabditis elegans, we defined the fitness costs associated with loss of the drug target genes singly or in combinations of the genes that encode GluCl subunits. We quantified the loss-of-function effects on three traits: (i) multi-generational competitive fitness, (ii) fecundity, and (iii) development. In competitive fitness and development assays, we found that a deletion of the beta-tubulin gene ben-1 conferred albendazole resistance, but ivermectin resistance required loss of two GluCl genes (avr-14 and avr-15) or loss of three GluCl genes (avr-14, avr-15, and glc-1). The fecundity assays revealed that loss of ben-1 did not provide any fitness benefit in albendazole and that no GluCl deletion mutants were resistant to ivermectin. Next, we searched for evidence of multi-drug resistance across the three traits. Loss of ben-1 did not confer resistance to ivermectin, nor did loss of any single GluCl subunit or combination confer resistance to albendazole. Finally, we assessed the development of 124 C. elegans wild strains across six benzimidazoles and seven macrocyclic lactones to identify evidence of multi-drug resistance between the two drug classes and found a strong phenotypic correlation within a drug class but not across drug classes. Because each gene affects various aspects of nematode physiology, these results suggest that it is necessary to assess multiple fitness traits to evaluate how each gene contributes to anthelmintic resistance.

Quantitative tests of albendazole resistance in beta-tubulin mutants

“…Another caveat is that only a single trait, development, was measured. ABZ affects multiple traits, including fecundity and competitive fitness over multiple generations ( Shaver et al, 2024 ). Future studies should investigate multiple traits at different ABZ concentrations to fully understand the role of all beta-tubulin genes in the ABZ response.…”

“…High-throughput assay images were processed using CellProfiler ( https://github.com/AndersenLab/CellProfiler ). Processed image data were cleaned and processed using the easyXpress ( Nyaanga et al, 2021 ) R package as previously described ( Shaver et al, 2024 ). The two assays were cleaned and processed independently.…”