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The tomato leafminer, Tuta absoluta, is an invasive crop pest that has evolved resistance to many of the insecticides used for its control. To facilitate the investigation of the underpinning mechanisms of resistance in this species we generated a contiguous genome assembly using long-read sequencing data. We leveraged this genomic resource to investigate the genetic basis of resistance to the diamide insecticide chlorantraniliprole in Spanish strains of T. absoluta that exhibit high levels of resistance to this insecticide. Transcriptomic analyses revealed that, in these strains, resistance is not associated with previously reported target-site mutations in the diamide target-site, the ryanodine receptor, but rather is associated with the marked overexpression (20-to >100-fold) of a gene encoding a UDP-glycosyltransferase (UGT). Functional expression of this UGT, UGT34A23, via ectopic expression in Drosophila melanogaster demonstrated that it confers strong and significant resistance in vivo. The genomic resources generated in this study provide a powerful resource for further research on T. absoluta. Our findings on the mechanisms underpinning resistance to chlorantraniliprole will inform the development of sustainable management strategies for this important pest.

The aphid Myzus persicae is a destructive agricultural pest that displays an exceptional ability to develop resistance to both natural and synthetic insecticides. To investigate the evolution of resistance in this species we generated a chromosome-scale genome assembly and living panel of >110 fully sequenced globally sampled clonal lines. Our analyses reveal a remarkable diversity of resistance mutations segregating in global populations of M. persicae. We show that the emergence and spread of these mechanisms is influenced by host–plant associations, uncovering the widespread co‐option of a host-plant adaptation that also offers resistance against synthetic insecticides. We identify both the repeated evolution of independent resistance mutations at the same locus, and multiple instances of the evolution of novel resistance mechanisms against key insecticides. Our findings provide fundamental insights into the genomic responses of global insect populations to strong selective forces, and hold practical relevance for the control of pests and parasites

Host shifts can lead to ecological speciation and the emergence of new pests and pathogens. However, the mutational events that facilitate the exploitation of novel hosts are poorly understood. Here, we characterize an adaptive walk underpinning the host shift of the aphid Myzus persicae to tobacco, including evolution of mechanisms that overcame tobacco chemical defenses. A series of mutational events added as many as 1.5 million nucleotides to the genome of the tobacco-adapted subspecies, M. p. nicotianae, and yielded profound increases in expression of an enzyme that efficiently detoxifies nicotine, both in aphid gut tissue and in the bacteriocytes housing the obligate aphid symbiont Buchnera aphidicola. This dual evolutionary solution overcame the challenge of preserving fitness of a mutualistic symbiosis during adaptation to a toxic novel host. Our results reveal the intricate processes by which genetic novelty can arise and drive the evolution of key innovations required for ecological adaptation.

BACKGROUND The tomato leafminer, Tuta absoluta, is an economically important pest of tomatoes in Europe, Africa, Asia and South America. In the UK this species is controlled using an integrated pest management (IPM) programme which incorporates the insecticides spinosad and chlorantraniliprole. In response to UK grower concerns of loss of efficacy of these compounds at certain sites, insecticide bioassays were performed on five populations collected from four commercial glasshouses and potential mechanisms of resistance investigated. RESULTS We observed high levels of resistance to spinosad in four of the strains, and in two of these tolerance to chlorantraniliprole. Selection of one of these strains with chlorantraniliprole rapidly resulted in a line exhibiting potent resistance to this compound. Sequencing of messenger RNA encoding the nicotinic acetylcholine receptor (nAChR) alpha 6 subunit, target of spinosad, revealed Ta alpha 6 transcripts in the spinosad-resistant strains that lack exon 4 and encode a highly truncated protein, or contain a triplet deletion in the predicted first transmembrane domain resulting in the loss of a highly conserved amino acid. Sequencing of the ryanodine receptor gene, encoding the target of diamide insecticides, of the chlorantraniliprole-selected line revealed an amino acid substitution (G4903V) that has been previously linked to diamide resistance in populations of T. absoluta in the Mediterranean and South America. CONCLUSION Taken together our results reveal emerging resistance in UK populations of T. absoluta to two of the most important insecticides used as part of IPM, with significant implications for the control of this species in the UK. (c) 2019 Society of Chemical Industry

BACKGROUND: Piperonyl butoxide (PBO) is a well-known insecticide synergist capable of interacting with phase 1 metabolic enzymes, specifically esterases and cytochrome P450s. In this study, structure activity relationship (SAR) analyses were used to characterise the interaction of around 30 analogues of PBO with the esterase FE4 and the P450 CYP6CY3 from insecticide resistant Myzus persicae (Sulzer), in order to predict the synthesis of more potent inhibitors. RESULTS: Enzyme inhibition studies were performed against esterase and oxidase activities and together with in silico modelling, key activity determinants of the analogues were identified and optimised. Novel analogues were then designed and synthesised, some of which showed greater inhibition against both enzymatic systems: specifically, dihydrobenzofuran moieties containing an alkynyl side chain and a butyl side chain against FE4, and benzodioxole derivatives with a propyl/butyl side chain and an alkynyl ether moiety for CYP6CY3. CONCLUSIONS: In vitro assays identified potential candidate synergists with high inhibitory potency. The in vivo confirmation of such results will allow consideration for a possible use in agriculture.

Many genes increase coding capacity by alternate exon usage. The gene encoding the insect nicotinic acetylcholine receptor (nAChR) a6 subunit, target of the bio-insecticide spinosad, is one example of this and expands protein diversity via alternative splicing of mutually exclusive exons. Here, we show that spinosad resistance in the tomato leaf miner, Tuta absoluta is associated with aberrant regulation of splicing of Taa6 resulting in a novel form of insecticide resistance mediated by exon skipping. Sequencing of the a6 subunit cDNA from spinosad selected and unselected strains of T. absoluta revealed all Taa6 transcripts of the selected strain were devoid of exon 3, with comparison of genomic DNA and mRNA revealing this is a result of exon skipping. Exon skipping cosegregated with spinosad resistance in survival bioassays, and functional characterization of this alteration using modified human nAChR a7, a model of insect a6, demonstrated that exon 3 is essential for receptor function and hence spinosad sensitivity. DNA and RNA sequencing analyses suggested that exon skipping did not result from genetic alterations in intronic or exonic cis-regulatory elements, but rather was associated with a single epigenetic modification downstream of exon 3a, and quantitative changes in the expression of trans-acting proteins that have known roles in the regulation of alternative splicing. Our results demonstrate that the intrinsic capacity of the a6 gene to generate transcript diversity via alternative splicing can be readily exploited during the evolution of resistance and identifies exon skipping as a molecular alteration conferring insecticide resistance.