Search

Herbivorous thrips are considered as one of the most economically important pests worldwide. They cause direct damage by feeding on plants of at least 60 families including many vegetable and ornamental crops, and indirect damage by transferring devastating plant viruses. Their high and parthenogenetic reproductive capacity and hidden lifestyle (pupation in the soil, thigmotactic behavior) make thrips hard to control. Current pest management control relies mainly on the use of chemical pesticides, to which thrips can develop resistance. Exploiting the natural defense mechanisms of plants against thrips can provide the opportunity towards more sustainable resistance breeding. The major aim of this PhD research was to explore the molecular mechanisms underlying natural defenses of the model plant Arabidopsis thaliana (Arabidopsis) in response to the Western flower thrips (Frankliniella occidentalis). To achieve this, several non-destructive bioassays were designed to assess thrips performance on single Arabidopsis leaves or whole plants. Using RNA-sequencing in a high-density time series, combined with bioinformatic analyses, we were able to capture the transcriptional dynamics and chronology of genes within the gene regulatory network that were induced in single Arabidopsis leaves upon thrips infestation. We confirmed that jasmonic acid (JA) is the predominant phytohormone modulating the induced defense response against thrips as (1) most up-regulated genes showed an overrepresentation of bHLH TF binding motifs which correlated with the activation of JA-associated processes, (2) JA-insensitive mutant coi1-34 and the MYC triple mutant myc2,3,4 showed significantly more feeding damage and a higher number of oviposited eggs compared to the Col-0 wild-type and (3) the novel regulators discovered in the thrips-induced gene regulatory network (GRN), although having distinct roles in the thrips-GRN, are closely connected to the JA pathway. However, depending on the thrips developmental stage, the predominant JA-induced response is modulated differently. For example, L1 larvae induced a relatively higher expression level of the ethylene (ET)-coregulated JA-inducible ERF-branch marker gene PDF1.2, which correlated with defense repression, while the older developmental larval stages activated higher levels of the abscisic acid (ABA)-coregulated JA-inducible MYC-branch marker gene VSP2, which was associated with enhanced plant resistance. Interestingly, although herbivory by L2 stage larvae of thrips mainly activated the expression of the MYC-branch marker gene VSP2 in the locally infested leaves, the ERF-branch marker gene PDF1.2 was mainly activated in the systemic, undamaged leaves of Arabidopsis, which correlated with enhanced preference of subsequently attacking thrips for the youngest systemic leaf tissue. In contrast, the tissue-chewing caterpillar Mamestra brassicae induced relatively higher levels of VSP2 in the local as well as the systemic, undamaged leaves of Arabidopsis, which was associated with enhanced deterrence of subsequently attacking thrips. Collectively, the work described in this thesis provides a better understanding of thrips-induced plant defense responses, which can facilitate the development of more sustainable thrips-resistance breeding.

Thrips are tiny, cell-content–feeding insects that are a major pest on crops and ornamentals. Besides causing direct feeding damage, thrips may also cause indirect damage by vectoring tospoviruses. Novel resistance mechanisms to thrips need to be discovered and validated. Induction of jasmonic acid–dependent defenses has been demonstrated to be essential for resistance to thrips, but underlying mechanisms still need to be discovered. For this, it is vital to use robust plant-thrips assays to analyze plant defense responses and thrips performance. In recently developed high-throughput phenotyping platforms, the feeding damage that is visible as silver spots, and the preference of thrips in a two-choice setup is assessed, using leaf discs. Here, we describe whole-plant thrips assays that are essential for (1) validation of findings obtained by the leaf disc assays, (2) assessment of longer-term effects on thrips feeding success and fecundity, (3) determination of spatial-temporal effects induced by primary thrips infestation on a secondary attack by thrips or other insects or pathogens, and (4) assessment of gene expression and metabolite changes. We present detailed methods and tips and tricks for (a) rearing and selection of thrips at different developmental stages, (b) treatment of the whole plant or an individual leaf with thrips, and (c) determination of feeding damage and visualization of thrips oviposition success in leaves.

Diphosphorylated inositol polyphosphates, also referred to as inositol pyrophosphates, are important signaling molecules that regulate critical cellular activities in many eukaryotic organisms, such as membrane trafficking, telomere maintenance, ribosome biogenesis, and apoptosis. In mammals and fungi, two distinct classes of inositol phosphate kinases mediate biosynthesis of inositol pyrophosphates: Kcs1/IP6K- and Vip1/PPIP5K-like proteins. Here, we report that PPIP5K homologs are widely distributed in plants and that Arabidopsis thaliana VIH1 and VIH2 are functional PPIP5K enzymes. We show a specific induction of inositol pyrophosphate InsP8 by jasmonate and demonstrate that steady state and jasmonate-induced pools of InsP8 in Arabidopsis seedlings depend on VIH2. We identify a role of VIH2 in regulating jasmonate perception and plant defenses against herbivorous insects and necrotrophic fungi. In silico docking experiments and radioligand binding-based reconstitution assays show high-affinity binding of inositol pyrophosphates to the F-box protein COI1-JAZ jasmonate coreceptor complex and suggest that coincidence detection of jasmonate and InsP8 by COI1-JAZ is a critical component in jasmonate-regulated defenses.