Pesticide resistance arises rapidly in arthropod herbivores, as can host plant adaptation, and both are significant problems in agriculture. These traits have been challenging to study as both are often polygenic and many arthropods are genetically intractable. Here, we examined the genetic architecture of pesticide resistance and host plant adaptation in the two-spotted spider mite, Tetranychus urticae, a global agricultural pest. We show that the short generation time and high fecundity of T. urticae can be readily exploited in experimental evolution designs for high-resolution mapping of quantitative traits. As revealed by selection with spirodiclofen, an acetyl-CoA decarboxylase inhibitor, in populations from a cross between a spirodiclofen resistant and a susceptible strain, and which also differed in performance on tomato, we found that a limited number of loci could explain quantitative resistance to this compound. These were resolved to narrow genomic intervals, suggesting specific candidate genes, including acetyl-CoA decarboxylase itself, clustered and copy variable cytochrome P450 genes, and NADPH cytochrome P450 reductase, which encodes a redox partner for cytochrome P450s. For performance on tomato, candidate genomic regions for response to selection were distinct from those responding to the synthetic compound and were consistent with a more polygenic architecture. In accomplishing this work, we exploited the continuous nature of allele frequency changes across experimental populations to resolve the existing fragmented T. urticae draft genome to pseudochromosomes. This improved assembly was indispensable for our analyses, as it will be for future research with this model herbivore that is exceptionally amenable to genetic studies. .

Arthropod herbivores cause dramatic crop losses, and frequent pesticide use has led to widespread resistance in numerous species. One such species, the two-spotted spider mite, Tetranychus urticae, is an extreme generalist herbivore and a major worldwide crop pest with a history of rapidly developing resistance to acaricides. Mitochondrial Electron Transport Inhibitors of complex I (METI-Is) have been used extensively in the last 25 years to control T. urticae around the globe, and widespread resistance to each has been documented. METI-I resistance mechanisms in T. urticae are likely complex, as increased metabolism by cytochrome P450 monooxygenases as well as a target-site mutation have been linked with resistance. To identify loci underlying resistance to the METI-I acaricides fenpyroximate, pyridaben and tebufenpyrad without prior hypotheses, we crossed a highly METI-I-resistant strain of T. urticae to a susceptible one, propagated many replicated populations over multiple generations with and without selection by each compound, and performed bulked segregant analysis genetic mapping. Our results showed that while the known H92R target-site mutation was associated with resistance to each compound, a genomic region that included cytochrome P450-reductase (CPR) was associated with resistance to pyridaben and tebufenpyrad. Within CPR, a single nonsynonymous variant distinguished the resistant strain from the sensitive one. Furthermore, a genomic region linked with tebufenpyrad resistance harbored a noncanonical member of the nuclear hormone receptor 96 (NHR96) gene family. This NHR96 gene does not encode a DNA-binding domain (DBD), an uncommon feature in arthropods, and belongs to an expanded family of 47 NHR96 proteins lacking DBDs in T. urticae. Our findings suggest that although cross-resistance to METI-Is involves known detoxification pathways, structural differences in METI-I acaricides have also resulted in resistance mechanisms that are compound-specific.

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