A pangenomic approach to understanding the evolution of insecticide resistance

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The evolution of insect resistance to insecticides represents a growing threat to the sustainable control of many important insect crop pests and disease vectors, threatening global food security and human health. To effectively combat resistance, it is critically important to understand its underlying genomic architecture, including the genetic variants underpinning resistance and their affect on phenotype. Advances in genome sequencing technology over the last two decades have greatly facilitated investigation of the genetic basis of insecticide resistance. However, key knowledge gaps remain on the type and number of genetic variants affecting resistance and their relative contribution to phenotype. Many of these deficits in understanding relate to the current paradigm of using a single reference genome as the starting point for most genomic analyses. This approach means that certain types of genetic variation not present in the reference typically remain undiscovered. This problem is particularly acute for structural variation (SV) encompassing presence-absence variation (PAV), copy number variation (CNV) and chromosomal rearrangements, which, as a consequence, have been referred to as the 'dark matter' of the genome. The failure to effectively characterise SV is important, as in humans SVs have been shown to affect more of the genome per nucleotide change than any other class of sequence variant. Furthermore, research by ourselves and others has provided clear evidence that SV can be a key source of genetic variation in the evolution of insecticide resistance.The overarching objective of this project is to develop a new paradigm for the genomic analysis of adaptive traits such as insecticide resistance in pest insects. We will leverage recent advances in sequencing technology, in combination with an exceptional biological resource comprising a living library of globally sampled clones of the damaging aphid crop pest Myzus persicae, to assemble the pangenome, the collection of all the DNA sequences that occur in a species, of this pest. This unprecedented resource will allow us to characterise the complete spectrum of genetic variation in a global crop pest species for the first time, and address multiple key knowledge gaps on the insect pangenome and the role of SV in adaptive evolution. These include understanding the percentage of the pangenome that is structurally variant in pest populations and how this differs across different SV types, how often and how many SVs lead to changes in gene function or expression, and crucially, their overall relevance and contribution to key phenotypic traits such as insecticide resistance. Our analyses will focus on three types of SV that have been frequently implicated in insecticide resistance, comprising CNV, PAV and chromosomal rearrangements. We will then test for association between the full spectrum of genetic variation identified in our aphid clone library and insecticide resistance. Finally, we will examine the impact of different types of SVs on gene expression and gene function, and validate the role of a selection of candidate SVs in resistance using functional approaches. Together, these analyses will allow us to fundamentally and systematically interrogate the genetic determinants of insecticide resistance in a way that has never been previously possible.The knowledge and tools generated in this project will provide both fundamental advances in our understanding of the genetic variation that provides the substrate for natural selection in insects, and powerful resources to develop strategies for the sustainable control of highly damaging, globally distributed crop pests.