Herbivorous insects are responsible for large losses to agriculture through feeding damage itself but also by vectoring pathogens such as viruses. Plants have evolved various mechanisms to defend themselves against insects, including the production of specialised metabolites that act as natural insecticides. Throughout history, plants and plant extracts harbouring specialised metabolites have been exploited by man for their antiinsect properties. Also, commercial insecticides are often based on plant-specialised metabolites and adapted further e.g. to give them the physicochemical properties that render them systemic. The development and application of these synthetic pesticides has made it possible to scale up the production and to increase the yield of food crops worldwide. However, their adversary effects on the environment and on non-target organisms, first discussed in Rachel Carson's Silent Spring, continue to cause debate and recently resulted in a ban on the use of certain pesticides in the European Union. The trait of insect defence has been largely overlooked during crop domestication and resulted in basically unarmed crop plants. Endogenous production of specialised metabolites could be reintroduced in modern crop varieties to retrieve self-defences. However, despite the fact that a plethora of specialised metabolites from plants has been described, studies that validate their deterrent or insecticidal effect in planta (crop), let alone in field situations, are sparse. To answer the question whether it is feasible to reintroduce endogenous insecticides to bring about insect resistance, the defence metabolites of plants, their synthesis and regulatory mechanisms underlying their production are examined.

Secretions from glandular trichomes potentially protect plants against a variety of aggressors. In the tomato clade of the Solanum genus, glandular trichomes of wild species produce a rich source of chemical diversity at the leaf surface. Previously, 7-epi-zingiberene produced in several accessions of Solanum habrochaites was found to confer resistance to whiteflies (Bemisia tabaci) and other insect pests. Here, we report the identification and characterisation of 9-hydroxy-zingiberene (9HZ) and 9-hydroxy-10,11-epoxyzingiberene (9H10epoZ), two derivatives of 7-epi-zingiberene produced in glandular trichomes of S. habrochaites LA2167. Using a combination of transcriptomics and genetics, we identified a gene coding for a cytochrome P450 oxygenase, ShCYP71D184, that is highly expressed in trichomes and co-segregates with the presence of the zingiberene derivatives. Transient expression assays in Nicotiana benthamiana showed that ShCYP71D184 carries out two successive oxidations to generate 9HZ and 9H10epoZ. Bioactivity assays showed that 9-hydroxy-10,11-epoxyzingiberene in particular exhibits substantial toxicity against B. tabaci and various microorganisms including Phytophthora infestans and Botrytis cinerea. Our work shows that trichome secretions from wild tomato species can provide protection against a wide variety of organisms. In addition, the availability of the genes encoding the enzymes for the pathway of 7-epi-zingiberene derivatives makes it possible to introduce this trait in cultivated tomato by precision breeding.

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Background Plant-produced specialised metabolites are a powerful part of a plant’s first line of defence against herbivorous insects, bacteria and fungi. Wild ancestors of present-day cultivated tomato produce a plethora of acylsugars in their type-I/IV trichomes and volatiles in their type-VI trichomes that have a potential role in plant resistance against insects. However, metabolic profiles are often complex mixtures making identification of the functionally interesting metabolites challenging. Here, we aimed to identify specialised metabolites from a wide range of wild tomato genotypes that could explain resistance to vector insects whitefly (Bemisia tabaci) and Western flower thrips (Frankliniella occidentalis). We evaluated plant resistance, determined trichome density and obtained metabolite profiles of the glandular trichomes by LC-MS (acylsugars) and GC-MS (volatiles). Using a customised Random Forest learning algorithm, we determined the contribution of specific specialised metabolites to the resistance phenotypes observed. Results The selected wild tomato accessions showed different levels of resistance to both whiteflies and thrips. Accessions resistant to one insect can be susceptible to another. Glandular trichome density is not necessarily a good predictor for plant resistance although the density of type-I/IV trichomes, related to the production of acylsugars, appears to correlate with whitefly resistance. For type VI-trichomes, however, it seems resistance is determined by the specific content of the glands. There is a strong qualitative and quantitative variation in the metabolite profiles between different accessions, even when they are from the same species. Out of 76 acylsugars found, the random forest algorithm linked two acylsugars (S3:15 and S3:21) to whitefly resistance, but none to thrips resistance. Out of 86 volatiles detected, the sesquiterpene α-humulene was linked to whitefly susceptible accessions instead. The algorithm did not link any specific metabolite to resistance against thrips, but monoterpenes α-phellandrene, α-terpinene and β-phellandrene/D-limonene were significantly associated with susceptible tomato accessions. Conclusions Whiteflies and thrips are distinctly targeted by certain specialised metabolites found in wild tomatoes. The machine learning approach presented helped to identify features with efficacy toward the insect species studied. These acylsugar metabolites can be targets for breeding efforts towards the selection of insect-resistant cultivars.

Main conclusion Cultivated tomatoes harboring the plastid-derived sesquiterpenes from S. habrochaites have altered type-VI trichome morphology and unveil additional genetic components necessary for piercing-sucking pest resistance. Abstract Arthropod resistance in the tomato wild relative Solanum habrochaites LA1777 is linked to specific sesquiterpene biosynthesis. The Sesquiterpene synthase 2 (SsT2) gene cluster on LA1777 chromosome 8 controls plastid-derived sesquiterpene synthesis. The main genes at SsT2 are Z-prenyltransferase (zFPS) and Santalene and Bergamotene Synthase (SBS), which produce α-santalene, β-bergamotene, and α-bergamotene in LA1777 round-shaped type-VI glandular trichomes. Cultivated tomatoes have mushroom-shaped type-VI trichomes with much smaller glands that contain low levels of monoterpenes and cytosolic-derived sesquiterpenes, not presenting the same pest resistance as in LA1777. We successfully transferred zFPS and SBS from LA1777 to cultivated tomato (cv. Micro-Tom, MT) by a backcrossing approach. The trichomes of the MT-Sst2 introgressed line produced high levels of the plastid-derived sesquiterpenes. The type-VI trichome internal storage-cavity size increased in MT-Sst2, probably as an effect of the increased amount of sesquiterpenes, although it was not enough to mimic the round-shaped LA1777 trichomes. The presence of high amounts of plastid-derived sesquiterpenes was also not sufficient to confer resistance to various tomato piercing-sucking pests, indicating that the effect of the sesquiterpenes found in the wild S. habrochaites can be insect specific. Our results provide for a better understanding of the morphology of S. habrochaites type-VI trichomes and paves the way to obtain insect-resistant tomatoes.