Host-plant resistance to western flower thrips in Arabidopsis
Thoen, Manus P.M. - \ 2016
Wageningen University. Promotor(en): Marcel Dicke; Harro Bouwmeester, co-promotor(en): Maarten Jongsma. - Wageningen : Wageningen University - ISBN 9789462578807 - 191
arabidopsis thaliana - host plants - insect pests - frankliniella occidentalis - defence mechanisms - pest resistance - genomics - genome analysis - host-seeking behaviour - optical tracking - data analysis - insect plant relations - arabidopsis thaliana - waardplanten - insectenplagen - frankliniella occidentalis - verdedigingsmechanismen - plaagresistentie - genomica - genoomanalyse - gedrag bij zoeken van een gastheer - optisch sporen - gegevensanalyse - insect-plant relaties
Western flower thrips is a pest on a large variety of vegetable, fruit and ornamental crops. The damage these minute slender insects cause in agriculture through feeding and the transmission of tospoviruses requires a sustainable solution. Host-plant resistance is a cornerstone of Integrated Pest Management (IPM). Plants have many natural defense compounds and morphological features that aid in the protection against herbivorous insects. However, the molecular and physiological aspects that control host-plant resistance to thrips are largely unknown.
A novel and powerful tool to study host-plant resistance to insects in natural populations is genome-wide association (GWA) mapping. GWA mapping provides a comprehensive untargeted approach to explore the whole array of plant defense mechanisms. The development of high-throughput phenotyping (HTP) systems is a necessity when large plant panels need to be screened for host-plant resistance to insects. An automated video-tracking platform that could screen large plant panels for host-plant resistance to thrips, and dissect host-plant resistance to thrips in component traits related to thrips behavior, was developed. This phenotyping platform allows the screening for host-plant resistance against thrips in a parallel two-choice setup using EthoVision tracking software. The platform was used to establish host-plant preference of thrips with a large plant population of 345 wild Arabidopsis accessions (the Arabidopsis HapMap population) and the method was optimized with two extreme accessions from this population that differed in resistance to thrips. This method can be a reliable and effective high throughput phenotyping tool to assess host-plant resistance to thrips in large plant populations. EthoAnalysis, a novel software package was developed to improve the analyses of insect behavior. There were several benefits from using EthoAnalysis to analyze EthoVision data. The detailed event statistics that could be extracted from EthoAnalysis allows researchers to distinguish detailed differences in moving and feeding behavior of thrips. The potential of this additional information is discussed in the light of quantitative genetic studies.
Stress resistance was studied in the HapMap population on a total of 15 different biotic and abiotic stresses ranging from biotic stresses like insects and nematodes, to abiotic stresses like drought and salt. A multi-trait GWA study to unravel the genetic architecture underlying plant responses to the different stresses was performed. A genetic network in this study revealed little correlation between the plant responses to the different insect herbivores studied (aphids, whiteflies, thrips and caterpillars). For thrips resistance a weak positive correlation with resistance to drought stress and Botrytis, and a negative correlation with resistance to parasitic plants were observed. One of the surprising outcomes of this study was the absence of shared major QTLs for host-plant resistance and abiotic stress tolerance mechanisms. RESISTANCE METHYLATED GENE 1 (RMG1) was one of the candidate genes in this multi-trait GWA study that could be controlling shared resistance mechanisms against many different stresses in Arabidopsis. RMG1 is a nucleotide-binding site Leucine-rich repeat (NB-LRR) disease resistance protein and its potential relation to several resistance/tolerance traits was successfully demonstrated with T-DNA insertion lines.
The 15 stresses were used in a comparison with a metabolomics dataset on this Arabidopsis HapMap population. It was discovered that levels of certain aliphatic glucosinolates correlated positively with the levels of resistance to thrips. This correlation was further investigated with the screening of a RIL (Recombinant Inbred Line) population for resistance to thrips, several knockout mutants and the analysis of co-localization of GWA mapping results between glucosinolates genes and thrips resistance. In a GWA analysis, the C4 alkenyl glucosinolates that correlated the strongest with thrips resistance mapped to the genomic regions containing genes known to regulate the biosynthesis of these compounds. However, thrips resistance did not co-localize with any of the GSL genes, unless a correction for population stratification was omitted. Additional screening of a Cvi x Ler RIL population showed a QTL for thrips resistance on chromosome 2, but no co-localization with any known glucosinolate genes, nor with thrips resistance loci identified by GWA mapping. Knock-out mutants and overexpressors of glucosinolate synthesis genes could also not confirm a causal link between glucosinolates and resistance to thrips. It is possible that the crucial factors that control resistance to thrips may not have been present in sufficient quantities or in the right combinations in the mutants, RILs and NIL screened in this study. Alternatively, the correlation between thrips feeding damage and glucosinolate profiles could be based on independent geographical clines. More research should be conducted to assess which of these explanations is correct.
In the general discussion, the results from this thesis are discussed in a broader perspective. Some prototypes of new phenotyping platforms that could further aid screening for resistance to thrips in the future are presented. Natural variation in host-plant resistance to thrips is compared to the variation in host-plant resistance to aphids and caterpillars. The geographic distribution of host-plant resistance to thrips is not evident in the other insects, in line with the distribution of glucosinolate profiles and other climate factors. The chapter concludes with some suggestions for future research in the field of host-plant resistance to thrips.
Plant responses to multiple herbivory : phenotypic changes and their ecological consequences
Li, Yehua - \ 2016
Wageningen University. Promotor(en): Marcel Dicke, co-promotor(en): Rieta Gols. - Wageningen : Wageningen University - ISBN 9789462578043 - 165
brassica oleracea - brevicoryne brassicae - aphidoidea - caterpillars - insect pests - pest resistance - defence mechanisms - phenotypes - insect plant relations - parasitoids - natural enemies - herbivore induced plant volatiles - plant-herbivore interactions - genetic variation - brassica oleracea - brevicoryne brassicae - aphidoidea - rupsen - insectenplagen - plaagresistentie - verdedigingsmechanismen - fenotypen - insect-plant relaties - parasitoïden - natuurlijke vijanden - herbivoor-geinduceerde plantengeuren - plant-herbivoor relaties - genetische variatie
This thesis explores whether aphid-infestation interferes with the plant response to chewing herbivores and whether this impacts performance and behaviour of individual chewing insect herbivores and their natural enemies, as well as the entire insect community. I investigated this using three wild cabbage populations (Brassica oleracea) that are known to differ in inducible secondary chemistry, to reveal whether patterns were consistent.
A literature review on recent developments in the field of plant interactions with multiple herbivores (Chapter 2) addressed how plant traits mediate interactions with various species of the associated insect community and their dynamics. In addition, the mechanisms underlying phenotypic changes in response to different herbivores were discussed from the expression of defence-related genes, phytohormones and secondary metabolites in plants to their effects on the performance and behaviour of individual insects as well as the entire insect community. In Chapter 3, I investigated the effects of early-season infestation by the aphid Brevicoryne brassicae on the composition and dynamics of the entire insect community throughout the season in a garden experiment replicated in two consecutive years. Aphid infestation in the early season only affected a subset of the community, i.e. the natural enemies of aphids, but not the chewing herbivores and their natural enemies. Moreover, the effects were only significant in the first half (June & July), but waned in the second half of the season (August & September). The effect of aphid infestation on the community of natural enemies also varied among the cabbage populations. Chapter 4 investigated the effects of aphid infestation on plant direct defences against chewing herbivores in laboratory experiments by comparing the performance of chewing herbivores and their parasitoids on aphid-infested and aphid-free plants. The performance of the specialist herbivore Plutella xylostella and its parasitoid Diadegma semiclausum was better on plants infested with aphids than on aphid-free plants, whereas the performance of the generalist herbivore Mamestra brassicae and its parasitoid Microplitis mediator was not affected by aphid infestation. These results suggest that aphid induced changes in plant traits may differentially affect the performance of leaf-chewing herbivore species attacking the same host plant, and also varied among the cabbage populations. Chapter 5 examined the effects of B. brassicae aphid infestation on plant indirect defences against chewing herbivores. In a two-choice olfactometer bioassay, preference behaviour for volatiles emitted by plants infested with hosts alone and those emitted by plants infested with aphids and hosts was compared for D. semiclausum and M mediator, larval endoparasitoids of caterpillars of P. xylostella and M. brassicae, respectively. In addition, the headspace volatiles emitted by host-infested and dually-infested plants were collected and analyzed. Co-infestation with aphids differentially affected volatile-mediated foraging behaviour of the two parasitoid species in an infestation period-dependent manner. Diadegma semiclausum preferred dually infested plants over host-infested plants when aphids infested the plants for a short time period, i.e. 7 days, but the volatile preference of D. semiclausum was reversed when aphid infestation was extended to 14 days. In contrast, M. mediator consistently preferred volatiles emitted by the dually-infested plants over those emitted by host-infested plants. The patterns of preference behaviour of the two wasp species were consistent across the three cabbage populations. Interestingly, the emission rate of most volatile compounds was reduced in plants dually-infested with caterpillars and aphids compared to singly-infested with caterpillars. This study showed that aphid infestation increased plant indirect defences against caterpillars, but depended on the aphid infestation period and specific caterpillar-parasitoid association. We hypothesized a negative interference of aphid infestation on plant defences against chewing herbivores based on previously reported SA-JA antagonism. In Chapter 6, we assessed the activation of SA and JA signaling pathways in plants infested by both aphids (B. brassicae) and various caterpillar species (P. xylostella, M. brassicae and Pieris brassicae) in different time sequences by quantifying transcription levels of the SA- and JA-responsive marker genes, PR-1 and LOX respectively. The results did not provide support for SA-JA antagonism. Compared to single infestation with each of the herbivore species, dual infestation with aphid and caterpillars had no interactive effects on the transcription levels of the SA- and JA-responsive maker genes, regardless of the temporal sequence of aphid and caterpillar attack, or the identity of the attacking caterpillar species.
The findings of this thesis contribute to our understanding of plant responses to herbivory by insect species belonging to different feeding guilds and their ecological effects on other associated community members. Aphid infestation may interfere with plant direct and indirect defences against leaf-chewing herbivores at the individual species level, but the effects are species-specific and also depend on the infestation period of aphids. Early-season aphid infestation may further affect the composition of the insect community, but the effect is smaller influencing only a subset of the community compared to early infestation by chewing herbivores. The molecular mechanism underlying plant responses to both phloem-feeding and leaf-chewing herbivores are complex and require the investigation of a range of genes involved in JA- and SA-mediated defence signal transduction. Plant interact with multiple herbivores at different levels of biological organization ranging from the subcellular level to the individual and the community level, and an integrated multidisciplinary approach is required to investigate plant-insect interactions.
Unraveling molecular mechanisms underlying plant defense in response to dual insect attack : studying density-dependent effects
Kroes, A. - \ 2016
Wageningen University. Promotor(en): Marcel Dicke; Joop van Loon. - Wageningen : Wageningen University - ISBN 9789462577756 - 265
016-3953 - arabidopsis thaliana - insect pests - herbivory - pest resistance - defence mechanisms - insect plant relations - molecular plant pathology - density - arabidopsis thaliana - insectenplagen - herbivorie - plaagresistentie - verdedigingsmechanismen - insect-plant relaties - moleculaire plantenziektekunde - dichtheid
In the field, plants suffer from attack by herbivorous insects. Plants have numerous adaptations to defend against herbivory. Not only do these defense responses reduce performance of the feeding herbivore, they also result in the attraction of natural enemies of herbivores.
The majority of studies investigating plant-insect interactions addressed mainly the effects of attack by a single herbivore species on induced plant defenses. However, because plants are members of complex communities, plants are exposed to different insect attackers at the same time. Moreover, attacks by different herbivores interact at different levels of biological organization, ranging from the level of gene expression, phytohormone production and biochemical changes up to the individual level. Effects of plant responses to feeding by two or more herbivore species simultaneously might cascade through the community and thereby affect insect community composition.
The induction of plant defense responses is regulated by a network of signaling pathways that mainly involve the phytohormones jasmonic acid (JA), salicylic acid (SA) and ethylene (ET). The signaling pathways of the two phytohormones SA and JA interact antagonistically, whereas JA and ET signaling pathways can interact both synergistically and antagonistically in regulating plant defense responses. In general, JA-mediated signaling underlies defense responses against leaf-chewing herbivores, such as caterpillars, whereas phloem-feeding insects, such as aphids, mainly induce SA-regulated defenses.
When caterpillars and aphids simultaneously feed on the same host plant, crosstalk between phytohormonal signaling pathways may affect the regulation of plant defenses. Consequently, multiple insect herbivores feeding on plants interact indirectly through plant-mediated effects. Studies investigating molecular mechanisms underlying interference by multiple attacking insects with induced plant defenses will benefit studies on the ecological consequences of induced plant responses.
The aim of this thesis was to elucidate molecular mechanisms that underlie plant-mediated interactions between attacking herbivores from different feeding guilds, namely Brevicoryne brassicae aphids and Plutella xylostella caterpillars.
Because herbivore density affects the regulation of plant defense responses, it may also influence the outcome of multiple insect-plant interactions. To study if modulation of induced plant defenses in response to dual insect attack depends on insect density, plants were infested with two densities of aphids.
Responses of Arabidopsis thaliana plants to simultaneous feeding by aphids and caterpillars were investigated by combining analyses of phytohormone levels, defense gene expression, volatile emission, insect performance and behavioral responses of parasitoids. To better predict consequences of interactions between plants and multiple insect attackers for herbivore communities, the regulation of defense responses against aphids and caterpillars was also studied in the ecological model plant wild Brassica oleracea.
Transcriptomic changes of plants during multiple insect attack and their consequences for the plant’s interactions with members of the associated insect community take place at different time scales. Direct correlation of transcriptomic responses with community development is, therefore, challenging. However, detailed knowledge of subcellular mechanisms can provide tools to address this challenge.
One of the objectives of this thesis, therefore, was to investigate the involvement of phytohormonal signaling pathways and their interactions during defense responses against caterpillars or aphids at different densities, when feeding alone or simultaneously on the model plant A. thaliana. The studies show that aphids at different densities interfere in contrasting ways with caterpillar-induced defenses, which required both SA- and JA-signal-transduction pathways. Transcriptional analysis revealed that expression of the SA transcription factor gene WRKY70 was differentially affected upon infestation by aphids at low or high densities. Interestingly, the expression data indicated that a lower expression level of WRKY70 led to significantly higher MYC2 expression through SA-JA crosstalk. Based on these findings, it is proposed that by down-regulating WRKY70 expression, the plant activates JA-dependent defenses which could lead to a higher resistance against aphids and caterpillars.
Plutella xylostella caterpillars also influenced plant defense responses when feeding simultaneously with aphids. Caterpillar feeding affected aphid-induced defenses which had negative consequences for aphid performance. Induction of both ET- and JA-mediated defense responses is required for this interference. Moreover, aphid density also played an important role in the modulation by P. xylostella of aphid-induced defenses: P. xylostella caterpillars induced changes in levels of JA and its biologically active from, JA-Ile, only when feeding simultaneously with aphids at a high density.
To study the overall effect of dual herbivory on induced plant defenses, not only interference with induced direct defense, but also with induced indirect defenses was addressed in A. thaliana. We found a significant preference of the aphid parasitoid Diaeretiella rapae for volatiles from aphid-infested A. thaliana wild-type plants and ein2-1 (ET-insensitive) mutants. Interestingly, simultaneous feeding by P. xylostella caterpillars on wild-type plants increased D. rapae’s preference for odors from aphid-infested plants. However, upon disruption of the ET-signaling pathway, D. rapae did not distinguish between ein2-1 mutants infested by aphids or by both aphids and caterpillars. This showed that intact ET signaling is needed for caterpillar modulation of the attraction of D. rapae parasitoids.
On the other hand, attraction of the caterpillar parasitoid Diadegma semiclausum to volatiles emitted by A. thaliana plants simultaneously infested by caterpillars and aphids was influenced by the density of the feeding aphids. Biosynthesis and emission of the terpene (E,E)-α-farnesene could be linked to the observed preference of D. semiclausum parasitoids for the HIPV blend emitted by plants dually infested by caterpillars and aphids at a high density, compared to dually infested plants with a low aphid density.
Transcriptomic changes in the response of A. thaliana wild-type plants to simultaneous feeding by P. xylostella caterpillars and B. brassicae aphids compared to plants infested by P. xylostella caterpillars alone were assessed using a microarray analysis. I particularly addressed the question whether the transcriptomic response to simultaneously attacking aphids and caterpillars was dependent on aphid density and time since initiation of herbivory. The data show that in response to simultaneous feeding by P. xylostella caterpillars and B. brassicae aphids the number of differentially expressed genes was higher compared to plants on which caterpillars had been feeding alone. Additionally, specific genes were differentially expressed in response to aphids feeding at low or high density. Cluster analysis showed that the pattern of gene expression over the different time points in response to dual infestation was also affected by the density of the attacking aphids. These results suggest that insects attacking at a high density cause an acceleration in plant responses compared to insects attacking at low density.
As a next step in the study of multiple interacting herbivores, I studied whether plant responses to dual herbivory have consequences for the performance of a subsequently arriving herbivore, Mamestra brassicae caterpillars. The ecological consequences of plant responses to dual herbivory cascading into a chain of interactions affecting other community members have remained unstudied so far. We used wild B. oleracea plants to evaluate dual herbivore-induced plant adaptations for subsequent herbivory. We found that simultaneous feeding by P. xylostella and B. brassicae resulted in different plant defense-related gene expression and differences in plant hormone levels compared to single herbivory, and this had a negative effect on subsequently arriving M. brassicae caterpillars. Differential induction of JA-regulated transcriptional responses to dual insect attack was observed which could have mediated a decrease in M. brassicae performance. The induction of plant defense signaling also affected both P. xylostella and B. brassicae performance. This study further helps to understand herbivore community build-up in the context of plant-mediated species interactions.
Altogether, findings from this thesis reveal a molecular basis underlying plant responses against multiple herbivory and provide insight in plant-mediated interactions between aphids and caterpillars feeding on plants growing in the field or used in agriculture.
Mapping moves on Arabidopsis : from natural variation to single genes affecting aphid behaviour
Kloth, K.J. - \ 2016
Wageningen University. Promotor(en): Marcel Dicke; Harro Bouwmeester, co-promotor(en): Maarten Jongsma. - Wageningen : Wageningen University - ISBN 9789462576483 - 269
016-3933 - arabidopsis thaliana - insect pests - aphidoidea - pest resistance - genetic mapping - gene expression - quantitative traits - functional genomics - feeding behaviour - insect plant relations - arabidopsis thaliana - insectenplagen - aphidoidea - plaagresistentie - genetische kartering - genexpressie - kwantitatieve kenmerken - functionele genomica - voedingsgedrag - insect-plant relaties
Adaptation of the brown planthopper, Nilaparvata lugens (Sta°l), to resistant rice varieties
Ferrater, J.B. - \ 2015
Wageningen University. Promotor(en): Marcel Dicke, co-promotor(en): F.G. Horgan; Peter de Jong. - Wageningen : Wageningen University - ISBN 9789462575592 - 200
insectenplagen - nilaparvata lugens - adaptatie - oryza sativa - rijst - cultivars - plaagresistentie - symbionten - gisten - endosymbionten - insect pests - nilaparvata lugens - adaptation - oryza sativa - rice - cultivars - pest resistance - symbionts - yeasts - endosymbionts
This thesis examines the three-way interaction between yeast-like symbionts, an insect herbivore [Nilaparvata lugens (Stål)] and its rice (Oryza sativa L.) host, during adaptation of the herbivore to resistant rice varieties. A long-term selection study (20 generations of continuous rearing, ca. 24 months) was conducted with N. lugens populations on four rice varieties (IR22, a susceptible variety and IR65482, IR62, and PTB33, three resistant varieties). Planthopper performance and the abundance of yeast-like symbionts (YLS) were monitored throughout the selection process. N. lugens populations adapted to the resistant varieties as noted by increasing body size and increased egglaying. Xylem feeding was observed as a possible behavioural adaptation of N. lugens: planthoppers on resistant plants had relatively high levels of xylem feeding compared with planthoppers on susceptible plants. Planthoppers selected on resistant varieties, had clear differences in YLS densities that were not related to fitness on the varieties and, therefore, did not support a YLS density-mediated adaptation hypothesis.
Furthermore, this study examined whether YLS density affected the capacity of planthoppers to switch between hosts on which they have been selected for several generations (natal plant) to new varieties (exposed plants) under normal YLS densities (symbiotic) and after reduction of YLS densities by heat treatment (aposymbiotic). The results suggested that YLS do not mediate host plant switching in planthoppers as removal of symbionts influenced body weight but not the relative capacity of nymphs to feed on different plants. This study also tested if virulence is acquired by shared feeding sites with virulent and avirulent planthoppers. In the study, planthoppers with varying levels of virulence affected the host plants differently: The most virulent hoppers appeared to suppress rice defences to a greater extent than non-virulent planthoppers. Planthoppers attained highest weights on those plants on which virulent planthoppers had previously fed which suggests that feeding by the virulent planthoppers facilitated subsequent planthopper feeding on the same plant. Our preliminary results indicate that feeding by mixed virulent-avirulent populations could potentially accelerate adaptation by N. lugens to resistant rice varieties.
The capacity of virulent and avirulent planthoppers to feed on a range of 24 resistant rice varieties was examined using a series of bioassays. Planthoppers were observed to feed and lay eggs on all the varieties tested, many of which have never been widely deployed in the field. Furthermore, planthoppers selected on resistant varieties often had increased fitness on other resistant varieties, even when these possess different resistance genes. However, there was no strong evidence that once planthoppers have adapted to a resistant variety, they will exhibit fitness costs on other varieties with dissimilar genes. The mechanisms underlying insect virulence are complex and further research on planthopper adaptation is necessary to help conserve genetic resources and prolong the durability of available resistant varieties.
Resistente biologische aardappelrassen in de etalage
Sikkema, A. ; Lammerts Van Bueren, E. - \ 2015
Resource: nieuwssite voor studenten en medewerkers van Wageningen UR 9 (2015)14. - ISSN 1389-7756 - p. 10 - 10.
aardappelen - rassen (planten) - biologische landbouw - akkerbouw - phytophthora - resistentieveredeling - resistentie van variëteiten - plaagresistentie - biologische plantenveredeling - gewasbescherming - potatoes - varieties - organic farming - arable farming - phytophthora - resistance breeding - varietal resistance - pest resistance - organic plant breeding - plant protection
Er zijn zes nieuwe biologische aardappelrassen gepresenteerd die resistent zijn tegen phytophthora.
Bodembacterie helpt plant tegen rupsenvraat
Sikkema, A. ; Pangesti, N.P.D. - \ 2015
Wageningen : St. voor Duurzame Ontwikkeling
arabidopsis - bodembacteriën - rizosfeerbacteriën - pseudomonas - gewasbescherming - rupsen - plaagresistentie - biologische bestrijding - landbouwkundig onderzoek - arabidopsis - soil bacteria - rhizosphere bacteria - pseudomonas - plant protection - caterpillars - pest resistance - biological control - agricultural research
Bodembacteriën die in het wortelmilieu van planten leven, verminderen de vatbaarheid van planten voor rupsenvraat. Dat blijkt uit onderzoek van Wageningse entomologen. In de modelplant Arabidopsis konden ze aantonen dat rhizobacteriën de plant in verhoogde staat van paraatheid brengen.
Resistentie toets van aardappel tegen Meloidogyne Chitwoodi 2010-2013 : overzicht van het onderzoek medegefinancierd door Productschap Akkerbouw
Molendijk, L.P.G. ; Been, T.H. - \ 2014
Lelystad : Praktijkonderzoek Plant & Omgeving B.V. - 18
solanum tuberosum - aardappelen - plaagresistentie - resistentie van variëteiten - meloidogyne chitwoodi - plantenparasitaire nematoden - detectie - nederland - akkerbouw - solanum tuberosum - potatoes - pest resistance - varietal resistance - meloidogyne chitwoodi - plant parasitic nematodes - detection - netherlands - arable farming
Induction of indirect plant defense in the context of multiple herbivory : gene transcription, volatile emission, and predator behavior
Menzel, T.R. - \ 2014
Wageningen University. Promotor(en): Marcel Dicke; Joop van Loon. - Wageningen : Wageningen University - ISBN 9789462571297 - 146
planten - plaagresistentie - geïnduceerde resistentie - verdedigingsmechanismen - multitrofe interacties - phaseolus lunatus - mijten - tetranychus urticae - roofmijten - phytoseiulus persimilis - voedingsgedrag - genen - transcriptie - genexpressie - herbivoor-geinduceerde plantengeuren - plants - pest resistance - induced resistance - defence mechanisms - multitrophic interactions - phaseolus lunatus - mites - tetranychus urticae - predatory mites - phytoseiulus persimilis - feeding behaviour - genes - transcription - gene expression - herbivore induced plant volatiles
Plants live in complex environments and are under constant threat of being attacked by herbivorous arthropods. Consequently plants possess an arsenal of sophisticated mechanisms in order to defend themselves against their ubiquitous attackers. Induced indirect defenses involve the attraction of natural enemies of herbivores, such as predators and parasitoids. Predators and parasitoids use odors emitted by damaged plants that serve as a “cry for help” to find their respective prey or host herbivore. The aim of this thesis was to use a multidisciplinary approach, with focus on molecular and chemical methods, combined with behavioral investigations, to elucidate the mechanisms of plant responses to multiple herbivory that affect a tritrophic system consisting of a plant, an herbivore and a natural enemy.
Induced plant defenses are regulated by a network of defense signaling pathways in which phytohormones act as signaling molecules. Accordingly, simulation of herbivory by exogenous application of phytohormones and actual herbivory by the two-spotted spider mite Tetranychus urticae affected transcript levels of a defense gene involved in indirect defense in Lima bean. However, two other genes involved in defense were not affected at the time point investigated. Moreover, application of a low dose of JA followed by minor herbivory by T. urticae spider mites affected gene transcript levels and emissions of plant volatiles commonly associated with herbivory. Only endogenous phytohormone levels of jasmonic acid (JA), but not salicylic acid (SA), were affected by treatments. Nevertheless, the low-dose JA application resulted in a synergistic effect on gene transcription and an increased emission of a volatile compound involved in indirect defense after herbivore infestation.
Caterpillar feeding as well as application of caterpillar oral secretion on mechanically inflicted wounds are frequently used to induce plant defense against biting-chewing insects, which is JA-related. Feeding damage by two caterpillar species caused mostly identical induction of gene transcription, but combination of mechanical damage and oral secretions of caterpillars caused differential induction of the transcription of defense genes. Nevertheless, gene transcript levels for plants that subsequently experienced an infestation by T. urticae were only different for a gene potentially involved in direct defense of plants that experienced a single event of herbivory by T. urticae. Indirect defense was not affected. Also sequential induction of plant defense by caterpillar oral secretion and an infestation by T. urticae spider mites did not interfere with attraction of the specialist predatory mite P. persimilis in olfactometer assays. The predator did distinguish between plants induced by spider mites and plants induced by the combination of mechanical damage and caterpillar oral secretion but not between plants with single spider mite infestation and plants induced by caterpillar oral secretion prior to spider mite infestation. The composition of the volatile blends emitted by plants induced by spider mites only or by the sequential induction treatment of caterpillar oral secretion followed by spider mite infestation were similar. Consequently, the induction of plant indirect defense as applied in these experiments was not affected by previous treatment with oral secretion of caterpillars. Moreover, herbivory by conspecific T. urticae mites did not affect gene transcript levels or emission of volatiles of plants that experienced two bouts of herbivore attack by conspecific spider mites compared to plants that experienced only one bout of spider mite attack. This suggests that Lima bean plants do no increase defense in response to sequential herbivory by T. urticae.
In conclusion, using a multidisciplinary approach new insights were obtained in the mechanisms of induction of indirect plant defense and tritrophic interactions in a multiple herbivore context, providing helpful leads for future research on plant responses to multiple stresses.
Whitefly resistance in tomato: from accessions to mechanisms
Lucatti, A.F. - \ 2014
Wageningen University. Promotor(en): Richard Visser, co-promotor(en): Ben Vosman; Sjaak van Heusden. - Wageningen : Wageningen University - ISBN 9789462570153 - 143
solanum lycopersicum - tomaten - wilde verwanten - insectenplagen - bemisia tabaci - plaagresistentie - verdedigingsmechanismen - plantenveredeling - insect-plant relaties - solanum lycopersicum - tomatoes - wild relatives - insect pests - bemisia tabaci - pest resistance - defence mechanisms - plant breeding - insect plant relations
Tomato (Solanum lycopersicum) is affected by a wide range of biotic stresses, of which Bemisia tabaci is one of the most important.Bemisia tabaci affects tomato directly through phloem sap feeding, and indirectly through its ability to be the vector of a large number of viruses. Different methods are available for whitefly control, and although several biological control agents are used against whiteflies in greenhouse cultivation, chemical control still is an essential component in open field tomato production. Breeding for host plant resistance is considered as one of the most promising methods in insect pest control in crop plants, and especially it is a promising alternative in whitefly control. Resistance to whiteflies was found in several wild relatives of tomato like Solanum peruvianum, S. pennellii, S. habrochaites, S. lycopersicum var. cerasiforme, S. pimpinellifolium andS. galapagense. In spite of previous breeding efforts, whiteflies are still a problem in tomato cultivation. The aim of my research was to identify and understand resistance mechanisms targeting specific stages of the whitefly life cycle in order to provide breeders with tools for developing whitefly resistant varieties.
I assessed the natural variation and whitefly resistance in Solanum galapagense and S. cheesmaniae, two wild tomato species endemic to the Galapagos Islands. Previously, Solanum galapagense and S. cheesmaniae were classified as two species based on a morphological species concept, but with molecular markers no clear separation could be made. So far, only a limited number of accessions/populations of S. galapagense and S. cheesmaniae have been evaluated for insect resistance and therefore it was unknown if the insect resistance coincides with the morphological species boundaries. Neither was there any knowledge about the relation between geographical and climatic conditions today on the Galapagos and the occurrence of the two species. We characterized twelve accessions of S. galapagense, 22 of S. cheesmaniae, and as reference one of S. lycopersicum for whitefly resistance using no-choice experiments. Whitefly resistance was found in S. galapagense only and was associated with the presence of relatively high levels of acyl sugars and the presence of glandular trichomes of type I and IV.It is likely that a minimum level of acyl sugars and the presence of glandular trichomes type IV are needed to achieve an effective level of resistance. Genetic fingerprinting using 3316 polymorphic SNP markers did not show a clear differentiation between the two species endemic to the Galapagos. Acyl sugar accumulation as well as the climatic and geographical conditions at the collection sites of the accessions did not follow the morphological species boundaries. Altogether, our results suggest that S. galapagense and S. cheesmaniae might be considered as morphotypes rather than two species and that their co-existence is likely the result of selective pressure.
Plants possess several resistance mechanisms acting at different time points during the interaction with herbivorous insect. Before any contact with the insects, plants emit an array of volatile organic compounds that can act as attractant or repellent of insects.Bemisia tabaci use a set of plant-derived cues in the process of host plant selection. It recognizes mainly monoterpenes (p-cymene, γ-terpinene and β-myrcene, α-phellandrene) and sesquiterpenes (7-epizingiberene and R-curcumene). Previously the line FCN93-6-2, which was derived from a cross between a susceptible tomato cultivar (Uco Plata INTA) and S. habrochaites (FCN3-5) was proved to be non-preferred by the greenhouse whitefly Trialeurodes vaporariorum. We identified chemical cues produced by FCN93-6-2 and S. habrochaites that can affect the preference of the whitefly B. tabaci as well as the potential chromosomal region(s) of S. habrochaites harbouring the genes involved in the preference. Two S. habrochaites accessions (CGN1.1561 and in FCN3-5) and the line FCN93-6-2 were non-preferred by B. tabaci when the whiteflies could get in direct contact with the plant and also when the whiteflies were offered olfactory cues only. The non-preference was independent of trichome type IV and of the presence of methyl-ketones but associated to the presence of monoterpenes in lower concentrations. Functional validation of the candidate metabolites and of the different introgressions is still needed.
Once the insect has landed on a plant, another set of resistance mechanisms enter into action. We have described a 3.06 Mbp introgression on top of Chromosome 5 (OR-5) from the wild tomato species S. habrochaites (CGN1.1561). For the identification of OR-5, we went from the selection of specific F2 plants to the development of F2BC4S1 and F2BC4S2 families. This introgression was sufficient to reduce whitefly fecundity without an evident effect on whitefly survival. The identification of mechanisms exclusively affecting whitefly fecundity and independent of trichomes type IV opens new doors for resistance breeding to whiteflies that may be especially interesting in greenhouse cultivation combination with natural enemies of the whitefly.
As an additional layer of defences, plants can perceive stress signals and respond to them in a specific way through induction of their immune system. This induction can also be triggered by exposing the plants to priming agents like hormones, some xenobiotic chemicals, like benzothiadiazole (BTH), β-aminobutyric acid (BABA), and sugars. Although the effect of priming agents was shown in laboratory and field studies, little is known about the effect of the genetic background of tomato on the extent of the priming, e.g. do genotypes varying in their level of resistance to insects and pathogens respond in the same way to a priming agent. We assessed the effect of selected priming agents on the effectiveness of natural defence in tomato. A set of no-choice and choice bioassays was conducted using tomato genotypes varying in their level of basal resistance to Bemisia tabaci and pathogens. We observed that whitefly survival and oviposition were not affected by the priming treatment in no-choice assays. Overall, in choice assays, fructose treated plants were more preferred by whiteflies than control plants. A genotype specific effect of priming was seen for the line FCN93-6-2. On this tomato line, JA and BABA applications decreased the number of whiteflies, e.g. making them less preferred.
In this thesis, I have gone from the screening of wild relatives of tomatoes to in depth characterization of resistance mechanisms. I have identified resistance mechanisms targeting specific stages of the whitefly life cycle, thus providing new tools for breeding durable whitefly resistance in tomato.
Genetics of insect resistance to plant defence
Vermeer, K.M.C.A. - \ 2014
Wageningen University. Promotor(en): Marcel Dicke, co-promotor(en): Peter de Jong. - Wageningen : Wageningen University - ISBN 9789461738363 - 199
insect-plant relaties - co-evolutie - planten - plaagresistentie - verdedigingsmechanismen - insectenplagen - secundaire metabolieten - phyllotreta nemorum - barbarea vulgaris - genetisch bepaalde resistentie - genetische analyse - insect plant relations - coevolution - plants - pest resistance - defence mechanisms - insect pests - secondary metabolites - phyllotreta nemorum - barbarea vulgaris - genetic resistance - genetic analysis
Plants are chemically defended against insect herbivory in various ways. They produce a broad range of secondary metabolites that may be toxic or deterrent to insects. Specialist insects, however, are often capable of overcoming these defences. The yellow striped flea beetle (Phyllotreta nemorumL.) is a specialist that feeds on crucifers (Brassicaceae) such as Sinapis arvensisand Barbarea vulgaris. In Denmark, two types of Barbarea vulgarisvar. arcuataare distinguished: one with pubescent leaves (P-type) and one with glabrous leaves (G-type). All individuals of P. nemorumcan feed on B. vulgarisP-type. Barbarea vulgarisG-type, on the other hand, is chemically defended against most P. nemorumindividuals during the flea beetle reproductive season. The defence compounds are hypothesized to be saponins, a class of compounds with various biological effects and insecticidal properties. Despite high levels of these saponins during summer, some flea beetles can and do feed on B. vulgaris G-type. The ability of P. nemorumto feed on B. vulgarisG-type is heritable; resistance against the defence of B. vulgarisG-type is controlled by dominant major resistance genes (R-genes). One dominant R-allele of an R-gene is enough to convert a susceptible beetle into a resistant one. Despite knowledge of the inheritance patterns of resistance in the flea beetles, which have been demonstrated to be variable, the underlying mechanism of flea beetle resistance has, so far, remained unclear. This prompted me to investigate, as an initial part of my thesis, the genetic basis of the flea beetle adaptation to the defence of B. vulgarisG-type.
The interaction between B. vulgarisand the flea beetle is a unique natural model system to study chemical defences in plants and counter-adaptations in insects. Plant and insect are both polymorphic with respect to the trait involved in resistance and hereby provide an excellent opportunity to study the geographic aspects of the evolution of the resistance trait in both interacting species. In this thesis, I focus on the resistance of the flea beetle, and take the presence of different genotypes of the plant as a given. Phyllotreta nemorumis a major pest, for example in oil seed rape. Understanding how resistance evolves in P. nemorumwill not only benefit flea beetle control, but also control of other pest insects. Understanding insect resistance includes knowledge of seasonal, geographic and genetic variation in both plant defense and herbivore adaptation.The R-gene has a remarkable distribution. Flea beetle populations living on B. vulgarisG-type consist solely of resistant individuals, but on host plant patches nearby B. vulgarisG-type lower frequencies of resistant beetles are found than one would expect with the amount of gene flow found at the neutral level between these subpopulations.
The aim of this thesis was to find the gene that is held responsible for the resistance of P. nemorumto the defences of B. vulgaris, investigate the distribution of this resistance trait and explain the distribution of this trait in natural populations. The following questions were addressed: (1) what is the genetic basis of the adaptation under study? (2) how is the resistance distributed across flea beetle populations in Denmark? and (3) which factors underlie this distribution?
In order to answer these questions, I used an integrated approach. I have combined a candidate gene approach (CHAPTER3) with an empirical approach via the study of variation in resistance in flea beetle populations (CHAPTER4), and a population genomics approach by using molecular markers to gain insight in the genomic make-up of the population and its connection with the resistance trait (CHAPTERS5 and 6). The population genomics approach is a recent advance in methods to detect the involvement of selection in the distribution of alleles at presumably adaptive loci. Using this approach one can distinguish locus-specific effects, like directional selection, from genome-wide effects, on the distribution of alleles at loci of interest.
The population genomics approach is introduced in CHAPTER2 together with the Geographic Mosaic Theory of Coevolution. I illustrate how processes underlying this theory of coevolution can be investigated with the population genomics approach. According to the geographic mosaic theory of coevolution, reciprocal selection between interacting species only happens in so-called hot-spots. Hot spots can be identified using population genomics and genetic variation found at specific loci can be attributed to locus-specific processes such as directional selection. For the B. vulgaris- flea beetle system this means that with a population genomics approach we can examine whether the distribution of resistant flea beetles on alternative host plants is only influenced by migration, or also by selection (CHAPTER5). Another valuable utility of the population genomics approach is to investigate whether a candidate gene for the R-gene is under selection, by looking whether a candidate gene is experiencing locus-specific effects beside genome-wide effects when comparing flea beetle populations living on B. vulgarisG-type with populations living on alternative host plants (CHAPTER6).
However, before using a population genomics approach to compare the resistance trait or a candidate gene with parts of the genome that only experience genome-wide effects, I have tried to identify the genetic basis of the flea beetle adaptation to the defence ofB. vulgarisG-type. In CHAPTER3, I have addressed this question by using a candidate gene approach to examine the involvement of a possible detoxifying enzyme in P. nemorum. Genes coding for β-glucosidase were a candidate for genes underlying the difference between resistant and susceptible beetles, because β-glucosidase is used as detoxifying enzyme by other organisms resistant to saponin defence. Three different β-glucosidase cDNA sequences were cloned from Danish flea beetle lines. We named them β-glucosidase A, B and C. β-glucosidase C was only found in resistant lines and not in the susceptible line. We then tested if recombinant β-glucosidase C breaks down the most abundant and most effective defence compound in B. vulgarisG-type, hederagenin cellobioside. β-glucosidase C was able to deglycosylate one glucose unit of hederagenin cellobioside, when expressed in an insect cell line. This suggests that expressed β-glucosidase C can deglycosylate antifeedant saponins and may play a role in the resistant flea beetle’s ability to overcome the defence of B. vulgaris. Next, a segregating family was created in which offspring differed in resistance genotype. Again β-glucosidase cDNA sequences were cloned to find a difference in the presence of these β-glucosidases between resistant and susceptible individuals. This time cDNA sequences of β-glucosidases A, B and C were present in both resistant and susceptible individuals although significantly fewer β-glucosidase C cDNA sequence variants were found in susceptible individuals than in resistant individuals. Thus, the genetic basis of flea beetle resistance remains unclear. Further investigation is needed to explore if the β-glucosidase C protein is also capable of inactivating hederagenin cellobioside by hydrolysizing the second glucose unit from the saponin and if there is a difference in enzyme activity of β-glucosidase C between resistant and susceptible beetles.
Subsequently, in CHAPTER4 I have investigated whether the frequency of resistant beetles decreased in populations living on other host plant patches than B. vulgarisG-type and whether the change in frequency was significant within the flea beetle season. I found that the frequency of resistant beetles varied significantly among years, but there was no evidence for a decrease in the frequency of resistant beetles, the latter being expected if selection acts against the resistance on other host plants than B. vulgarisG-type. Furthermore, I found that the frequency of resistant beetles varied significantly within a flea beetle season. This study demonstrates that relative frequencies of different resistance phenotypes of P. nemorumon other host plants than B. vulgarisG-type are highly dynamic, both within and across years. It is, therefore, important to sample season-wide when one wants to monitor the changes in frequencies of insect resistance in natural systems.
In CHAPTERS5 and 6 I took a population genomics approach to investigate if the observed geographicaldistribution of resistance of P. nemorumto chemically defendedB. vulgarisin flea beetle populations could be explained by factors that are solely associated with genome-wide effects, such as migration, or also by locus-specific factors like selection at the resistance locus. First, neutral microsatellites were used to reveal the genetic differentiation at parts of the genome that are only influenced by genome-wide processes. Next, the level of neutral genetic differentiation was compared with the genetic differentiation found for the resistance trait. The resistance trait was an outlier in pairwise comparisons between flea beetle populations on B. vulgarisand S. arvensis, meaning that the level of genetic differentiation was significantly higher than expected if the resistance trait experiences only genome-wide effects. The resistance trait was also an outlier in the pairwise comparison between populations on S. arvensis, which suggests that the resistance trait is also under directional selection on other host plants than B. vulgarisG-type.
Additionally, I examined in CHAPTER6 if the homologous β-glucosidases B and C sequences found in CHAPTER3 correspond to two alleles of the major resistance gene, because of their similarity and their presence in flea beetle lines. The sequence of β-glucosidases C had so far only been found in resistant individuals, so we hypothesized it to be the dominant resistance allele and the sequence of β-glucosidases B would then correspond to the susceptible allele. In order to find out if this hypothesized PneR-gene (Phyllotreta nemorum R-gene) is the resistance gene, we first directly compared resistance phenotypes of beetles collected from populations on B. vulgarisG-type and S. arvensiswith genotypes derived with primers developed for β-glucosidase B and C. The phenotype of the flea beetles did not match the genotype derived with the β-glucosidase primers. Additionally, the frequency of heterozygotes and homozygotes of the PneR-gene genotype was not significantly deviating from Hardy-Weinberg Equilibrium which implies that there are no locus-specific effects involved when both sequences are seen as one gene with two alleles. A population approach was taken like in CHAPTER5, this time including the genetic differentiation estimated for the candidate gene as well. The candidate gene behaved similar to the neutral loci while the resistance trait was an outlier in most pairwise comparisons between flea beetle populations. If both sequences are alleles of the same gene, then the candidate gene is not directly responsible for the flea beetle resistance to B. vulgarisG-type defence.
The results presented in this thesis show the complexity of genetic processes (either genome-wide or locus specific) affecting local adaptation and the distribution of a resistance trait in insects in natural populations. Furthermore, the present study shows that when studying coevolution between insect and host plant by means of adaptive traits, also geographical and seasonal variation in allele frequencies should be considered. A multidisciplinary approach to study adaptation in plant-insect interactions such as used in this thesis, will benefit research on plant-insect interactions, including applied research such as studying the potential of host plants as dead-end traps for pest insects and preventing/diminishing the development of resistance by pest insects to crop defences.
Bruine rat resistent tegen rattengif : alternatieven zijn dringend gewenst
Schoelitsz, B. ; Meerburg, B.G. - \ 2013
Zoogdier 24 (2013)3. - ISSN 0925-1006 - p. 6 - 8.
fauna - ratten - plaagresistentie - pesticidenwerking - fauna - rats - pest resistance - pesticidal action
Overlast door ratten wordt sinds jaar en dag tegengegaan met rodenticiden (beter bekend als rattengif). Ze zijn toegelaten voor de bestrijding van deze knaagdieren. Maar wat als bruine ratten niet meer doodgaan van deze middelen en hiervoor dus resistent zijn? Wat weten we allemaal over resistentie bij deze soort? En wat zijn de consequenties hiervan voor de bestrijding van de bruine rat?
Identification of Arabidopsis thaliana genes that can increase resistance towards phloem feeding insects
Chen, X. - \ 2013
Wageningen University. Promotor(en): Richard Visser, co-promotor(en): Ben Vosman. - [S.l.] : S.n. - ISBN 9789461737649 - 96
arabidopsis thaliana - insectenplagen - myzus persicae - plaagresistentie - genkartering - genexpressie - mutanten - plantenveredeling - turnip yellows virus - vectoren, ziekten - arabidopsis thaliana - insect pests - myzus persicae - pest resistance - gene mapping - gene expression - mutants - plant breeding - turnip yellows virus - disease vectors
Phloem feeding insects are among the most devastating pests worldwide. They not only cause damage by feeding from the phloem, but also by vectoring plant viruses. During their evolution plants have developed a variety of defense traits to combat insects. These plant resistance traits can be antixenotic and/or antibiotic. Antixenosis is the first line of defense that prevents insects from landing and settling, while antibiosis reduces the population development of the colonizing insects.In this project we aimed at identifying genes that can increase resistance towards phloem feeding insects and also prevent, as far as possible, transmission of viruses. Acknowledging that changing the expression level or expression localization of genes might increase resistance, we screened an Arabidopsis thaliana activation tag gain-of-function mutant collection for increased resistance towards the green peach aphid (Myzus persicae). In these mutants, tagged genes are overexpressed by the strong 35S enhancer adjacent to the natural promoter that results in a dominant gain-of-function phenotype. The overexpression of a particular gene in such mutants may result in enhanced resistance to aphids and other phloem feeding insects.
To identify mutants with increased insect resistance efficient and reproducible screening methods needed to be developed first. Based on the hypothesis that there is a trade-off between plant fitness and plant resistance, we first screened a subset of 170 mutants that were previously selected based on their reduced growth to increase the chance of identifying mutants with increasedresistance. In this screening we usedchoice assays and selected one mutant that displays enhanced antixenosis based resistance towards aphids. Further characterization of this mutant revealed that that the antixenosis is phloem based and requires intact plants.
To evaluate aphid resistance of a larger number (>5000) of activation tag mutants, we established a high throughput screening system in which plant resistance against aphids is inferred from a reduced transmission of the circulative Turnip yellows virus(TuYV). This virus can only be transmitted into a plant after virus-infected aphids feed for a prolonged (> 10min) time from the phloem sap. In the initial screening 13 virus-free mutant lines were identified. The putative candidate mutant lines were re-evaluated and characterized, resulting in nine mutants on which aphids showed a reduced population development.
Molecular analysis of two of these mutants revealed that the genes underlying the resistance were IRM1(Increased ResistancetoMyzus persicae1,At5g65040)and SKS13 (SKU5Similar13, At3g13400). In wild type plants,IRM1is strongly expressed in xylem and extremely low expressed in other plant tissue whereas SKS13 is exclusively expressed in pollen. We show that constitutive overexpression of these genes in all plant tissues confers enhanced resistance towards aphids. Analysis of aphid feeding behavior showed that the resistance conferred by IRM1and SKS13affect the aphids differently. On the IRM1 overexpressing mutant aphids encounter difficulties in reaching the phloem, indicating that resistance factors are located between the cell surface and the phloem. On the SKS13overexpressingmutant the phloem feeding of aphids is severely affected, indicating that resistance factors are phloem based. Further analysis strongly suggests the involvement of Reactive Oxygen Species (ROS) in the reduced aphid performance on the SKS13overexpressingmutant. We also show that the resistances are not aphid specific, as the performance of the cabbage aphid (Brevicoryne brassicae)is also affectedon both overexpressing mutants.
The results obtained in this thesis show that plant resistance to insects can be increased by expressing genes that are assigned for other biological functions. Characterization of the identified mutants revealed twogenes conferring enhanced aphid resistance via different mechanisms. These findings lead to a better understanding of plant-aphid interactions on the molecular level. Furthermore, such knowledge obtained from the model plant A.thalianashould be applied in crop plants, which can be achieved by transgenic and genetic studies in combination with newly developed techniques, such as RNAi and TILLING.
Resistance to thrips in pepper
Maharijaya, A. - \ 2013
Wageningen University. Promotor(en): Richard Visser, co-promotor(en): Roeland Voorrips; Ben Vosman. - Wageningen : Wageningen UR - ISBN 9789461737397 - 110
capsicum - capsicum annuum - capsicum chinense - paprika's - spaanse pepers - insectenplagen - vectoren, ziekten - frankliniella occidentalis - thrips - plaagresistentie - loci voor kwantitatief kenmerk - soortkruising - plantenveredeling - capsicum - capsicum annuum - capsicum chinense - sweet peppers - chillies - insect pests - disease vectors - frankliniella occidentalis - thrips - pest resistance - quantitative trait loci - interspecific hybridization - plant breeding
Pepper (Capsicum) production is constrained by heavy infestations of thrips, causing direct and indirect (by transmitting viruses) damage. Thrips control using chemical insecticides, biological agents, culture practices and integrated pest management has limited success. The availability of thrips-resistant varieties would increase the effectiveness of thrips control and may also delay and reduce the transmission of viruses. This thesis is aimed at obtaining more knowledge regarding thrips resistance in pepper, including the identification of new sources of resistance, the elucidation of resistance mechanisms, identification of factors contributing to resistance and a QTL analysis.
We developed several test methods to evaluate plant resistance to thrips and showed that in vitro tests correlate well with greenhouse tests. We used these methods to test a collection of Capsicum accessions of widely different origin and crop types. This resulted in the identification of a few accessions (mostly C. annuum) with high levels of resistance to two thrips species: Frankliniella occidentalis and Thrips parvispinus. Since C. annuum is the most widely cultivated species, the finding of resistance in C. annuum is means that the resistance can be easily introgressed through conventional crossing and selection.
The effect of resistance in pepper on thrips reproduction and development was studied using three highly resistant, three medium resistant and three susceptible accessions selected based on damage ratings. Adult and pre-adult survival, developmental time and reproduction rate were assessed in a detached leaf system. Resistance factors in leaves of resistant pepper accessions were shown to have significant effects on oviposition rate, larval mortality and life-cycle period, indicating that this resistance is based on antibiosis.
In order to map QTL for resistance we developed an F2 population from the cross between a susceptible C. chinense accession and the resistant C. annuum AC 1979. A genetic linkage map for this population was based on AFLP and SSR markers, where the SSR markers served to assign and orient most linkage groups to pepper chromosomes. As larval stages were highly affected by resistance in pepper leaves, damage caused by larvae and larval survival were used as parameters to detect QTLs conferring resistance to thrips. Interval mapping detected one QTL for each of these parameters, all co-localizing near the same marker on chromosome 6. This QTL explained about 50% of the genetic variation, and the resistance allele of this QTL was inherited from the resistant parent. No other resistance QTLs were detected in this population.
Since resistance to thrips was clearly expressed in pepper leaves we proceeded to study leaf traits that may contribute to resistance. Morphological leaf characters and metabolites have frequently been linked with resistance to thrips in other plant species. However, we found no convincing evidence that any of these traits played a role in thrips resistance in pepper. In the F2 mapping population we found no correlation and no QTL co-localization of resistance with leaf morphological characters previously linked to resistance in pepper against insect pest and in other plant species against thrips e.g. color, toughness, trichome density, and cuticula thickness. GC-MS (Gass Chromatography – Mass Spectrometry) analysis of the three resistant, three intermediate and three susceptible accessions mentioned above showed that seven metabolites were correlated with resistance to thrips and six compounds with susceptibility. However, when we applied GC-MS and LC-MS (Liquid Chromatography – Mass Spectrometry) to leaves of the F2 mapping population, we found no strong correlation between resistance and any detected metabolites. Two metabolite QTLs co-localized with the resistance QTL. However, these QTLs explained only a small proportion of the variance and the co-localization was not supported by strong correlations of the metabolites with resistance. This suggests that the major resistance factor(s) in pepper against thrips may not or only partially be determined by the presence or absence of specific metabolites.
This thesis provides a strong basis for the development of thrips resistant pepper varieties through introgression of the resistance QTL region on chromosome 6 originating from resistant C. annuum accessions. However, the effect of resistance QTL on chromosome 6 should be confirmed in another population such as a population of F3 lines. In vitro leaf assay can be used as evaluation methods in pepper breeding program. This has the advantages of minimizing the risk of contamination and of controlled environmental conditions. Elucidation of factors contributing to resistance should be continued by giving attention to other possibilities such as proteins, specifically proteinase inhibitors, or other leaf anatomical and morphological traits. Also other extraction and detection methods may be used to discover other metabolites that might be related to resistance. Finally, for practical applications it is necessary study how to use the antibiosis based mechanism against thrips found in this thesis in thrips control and/or management practices.
Resistance mechanisms against Bemisia tabaci in wild relatives of tomato
Elsen, F.H.W. van den - \ 2013
Wageningen University. Promotor(en): Marcel Dicke, co-promotor(en): Ben Vosman; Sjaak van Heusden. - S.l. : s.n. - ISBN 9789461737298 - 179
solanum lycopersicum - wilde verwanten - insectenplagen - bemisia tabaci - plaagresistentie - verdedigingsmechanismen - plantenveredeling - solanum lycopersicum - wild relatives - insect pests - bemisia tabaci - pest resistance - defence mechanisms - plant breeding
The silverleaf whitefly (Bemisia tabaciGenn.) poses a serious threat to tomato cultivation. A large part of the damage is done directly through heavy host plant colonization. Colonization has a negative impact on the plant, as the whitefly takes up nutrients from the phloem and induces phytotoxic responses, which result in irregular ripening of the fruits. However, most damage is done indirectly as the silverleaf whitefly vectors a broad range of plant pathogenic viruses.
The silverleaf whitefly can successfully be controlled biologically in greenhouse cultivations, but control of the whitefly in the field is mainly based on the application of pesticides. The use of pesticides can have a negative effect on non-harmful or beneficial organisms in the field. Moreover, the effectiveness of pesticides can decline or even completely disappear through adaptation of the whitefly. An effective alternative for the use of pesticides could be the deployment of resistant cultivars. Nowadays, genetic factors responsible for whitefly resistance can be transferred faster and more efficiently into tomato cultivars through marker-assisted backcross breeding programs. Complete resistance against the whitefly is present in some crossable wild relatives of the cultivated tomato and the literature reports extensively about accessions with a high level of resistance against the whitefly.
In this work, I have studied different populations that were developed by interspecific crosses between cultivated tomato and the tomato wild relativesS. habrochaitesLYC4 and S. pennelliiLA3791. By integrating datasets from different research disciplines, I have studied the background of whitefly resistance in these populations. Furthermore, these data were used to identify the chromosomal loci in the wild tomato relatives that harbor genes responsible for the resistance and that can be bred into cultivated tomato.
The mechanisms underlying the resistance in S. pennelliiLA3791 were studied through phenotypic resistance assays that demonstrated that survival and oviposition of the whitefly were not possible on this wild relative. Through removal of glandular cells, present on the leaf trichomes, the resistance was almost completely lost and only adult survival was still significantly different from the wild type. This result led to the hypothesis that glandular trichomes play an important role in the resistance. This was confirmed in a segregating population based on a cross between S. pennelliiLA3791 and a susceptible cultivated tomato. Plants that lacked glandular trichomes type I and IV, had the same resistance level as the susceptible parent. Further analyses of the segregating population showed that the presence of glandular trichomes was not the only factor determining resistance, but that the composition and quantity of the metabolites in the glandular trichomes also played an important role. To gain more knowledge on the role of individual metabolites on whitefly resistance and susceptibility, we analyzed the total metabolite content of extreme phenotypes of the F2 population. Gas Chromatography-Mass Spectrometry (GC-MS) and Liquid Chromatography Time-Of-Flight Mass Spectrometry (LC-TOF-MS) were employed for the analyses of the total metabolite content. Analyses revealed that on basis of the total metabolite profiles the extreme phenotypes (susceptible versus resistant for the silverleaf whitefly) could be discriminated into two groups that were correlated with resistance or susceptibility. A number of these metabolites could be annotated, but for the majority of the components this was not possible on the basis of available literature and databases. Subsequently, I have studied the genetic basis of the phenotypic resistance parameters as well as the genetic basis of the metabolites from the GC-MS and LC-TOF-MS analyses. A genetic linkage map of the F2 mapping population was developed using DNA markers (Amplification Fragment Length Polymorphisms,AFLPs and Single Nucleotide Polymorphisms, SNPs). QTLs (Quantitative Trait Loci) were identified between the majority of the metabolites and the genetic markers (>90%) and also we found genetic linkages between whitefly resistance parameters and markers. The QTLs for metabolites and phenotypic parameters partly co-localized at the same positions on the genetic map. Several metabolite QTLs (mQTLs) co-localized with each other in so-called ‘hotspots’. Remarkably, the results of the individual phenotypic QTLs (phQTLs) for adult survival and oviposition as well as the mQTLs for the individual components did not give high explained variances (<20%), which was supported by an analysis of individual metabolite profiles, that showed a high variation in composition between F2genotypes with an identical resistance level.
On the basis of these results I hypothesized that resistance could not be explained by a specific composition of metabolites, but that multiple metabolic profiles can result in the same level of resistance in a plant. To support this hypothesis, a backcross population was developed, an F2BC1,by backcrossing a completely resistant F2plant with the recurrent parent. The complete F2BC1population was analyzed by LC-TOF-MS to characterize the metabolite content of the progeny lines alongside resistance assays for adult survival and oviposition on these plants. Again, in this population we identified genotypes that possessed a level of resistance equal to the S. pennelliiLA3791 donor parent. From the analyses it became clear that the complexity of the chemical profiles was reduced and that only a few components were correlated with whitefly resistance or susceptibility. A genetic linkage map with a large number of SNP markers enabled the identification of new QTLs alongside the QTLs from the previous F2mapping that were confirmed in the F2BC1 populations. The reduction in complexity of the chemical profile was accompanied by an increase in explained variances of both the phenotypic as well as the metabolite QTLs. The results indicate that performing phenotyping assays by scoring resistance parameters in a population along with analyzing the chemical profiles is required to identify resistance loci, which can subsequently be used in marker-assisted breeding programs.
Finally I have studied an Introgression Line (IL) population, consisting of 30 lines, which each contained a different introgression of S. habrochaitesLYC4, a whitefly-resistant wild relative of cultivated tomato. Survival and oviposition assays of the whole population revealed that there were a few lines that showed a slightly reduced susceptibility for the silverleaf whitefly. Completely resistant lines were not identified, which indicates that the resistance in this wild relative is complex and governed by the interaction of several genes at different locations on the tomato genome. Such genetic interactions, also referred to as epistatic interactions, complicate the identification of genes involved in resistance and the underlying resistance mechanisms. Therefore, I concluded that IL populations are not suitable for the elucidation of a complex trait as whitely resistance in tomato.
In conclusion, this thesis demonstrates the most important aspects of susceptibility and resistance against the silverleaf whitefly in a S. pennellii accession and provides strong evidence for the underlying resistance mechanisms. Furthermore, we were capable of reducing the complex phenotypic and genotypic variation, which was present in the F2 population, via a backcross with the recurrent parent. This made it possible to identify three genetic loci in S. pennellii that play a role in whitefly resistance. A logical next step of this research would be the fine mapping of these three loci in order to enable the transfer of these loci/genes into cultivated tomato lines. By doing so, an important step towards sustainable control of the silverleaf whitefly in tomato cultivation could be made.
Unravelling the resistance mechanism of lettuce against Nasonovia ribisnigri
Broeke, C.J.M. ten - \ 2013
Wageningen University. Promotor(en): Joop van Loon; Marcel Dicke. - S.l. : s.n. - ISBN 9789461735782 - 228
lactuca virosa - lactuca sativa - slasoorten - insectenplagen - nasonovia ribisnigri - plaagresistentie - verdedigingsmechanismen - gedrag bij zoeken van een gastheer - insect-plant relaties - lactuca virosa - lactuca sativa - lettuces - insect pests - nasonovia ribisnigri - pest resistance - defence mechanisms - host-seeking behaviour - insect plant relations
Aphids are serious pests of crop plant species, and host plant resistance is often the most effective and environmentally friendly control strategy to control these pests. One of these aphid pests is the black currant - lettuce aphid, Nasonovia ribisnigri (Mosely), an economically important pest of cultivated lettuce, Lactuca sativa L. Host plant resistance has been used since 1982 to control this aphid species and is mediated by the Nr-gene, originating from wild lettuce Lactuca virosa L. However, this resistance is not effective anymore, since N. ribisnigri aphids virulent to the Nr-resistance have been reported since 2007. The aim of this thesis was to unravel the mechanism of resistance mediated by the Nr-gene against N. ribisnigri, by behavioural studies on the aphids on both resistant and susceptible lettuce, to allow lettuce breeders to accelerate their resistance breeding programmes. Although the exact mechanism of Nr-mediated resistance remains unknown, the data in this thesis provide insight into this mechanism. The active site of the Nr-mediated resistance is mainly located in the phloem and some resistance might be encountered by the aphids along the pathway to the phloem. The inability of the avirulent aphids to feed from the resistant plant could be caused by the failure of aphids to suppress the wound response of the sieve element. The resistance factor(s) are only produced in the shoot, because grafts with resistant shoots and susceptible roots were resistant, whereas grafts with susceptible shoots and resistant roots remained susceptible. An intact vascular system is needed for full resistance, because both detached leaves and leaf disks of resistant lettuce plants were less resistant.
Variation in virulence was observed among populations of different geographical origin. Aphids from a highly virulent population performed equally well on both resistant and susceptible lettuce plants, whereas semi-virulent aphids performed better on susceptible lettuce plants. Both short-term and long-term virulence loss were observed for virulent aphid populations differing in virulence level, which indicates this virulence is associated with fitness costs. A possible mechanism underlying virulence in N. ribisnigri to the Nr-resistance is the presence of an effector protein in the salivary secretion of the aphids suppressing resistance. Virulent aphids seemed to actively suppress the resistance in lettuce against the avirulent aphids.
The original donor or the Nr-resistance, L. virosa accession IVT 280, was tested as possible source of new resistance against the virulent biotypes of N. ribisnigri and was foundfully resistant against virulent aphids, and can be exploited as a source of resistance in breeding for new resistance in cultivated lettuce.
Groene Veredeling ; Veredelingsonderzoek tripsresistentie in prei
Scholten, O.E. ; Henken, B. ; Burger, K. ; Vosman, B. - \ 2013
resistentieveredeling - thrips - plaagresistentie - preien - vollegrondsgroenten - plantenveredeling - biologische plantenveredeling - genetisch bepaalde resistentie - resistance breeding - thrips - pest resistance - leeks - field vegetables - plant breeding - organic plant breeding - genetic resistance
Brochure met onderzoeksinformatie. Het Groene Veredelings-onderzoek naar tripsresistentie in prei levert een bijdrage aan de ontwikkeling van prei met een hoger niveau van tripsresistentie; voor zowel de biologische als de gangbare sector.
Biosynthesis of monoterpene alcohols, derivatives and conjugates in plants : roles in resistance to western flower thrips
Yang, T. - \ 2013
Wageningen University. Promotor(en): Marcel Dicke; Harro Bouwmeester, co-promotor(en): Maarten Jongsma. - S.l. : s.n. - ISBN 9789461735317 - 116
planten - plaagresistentie - verdedigingsmechanismen - insectenplagen - frankliniella occidentalis - monoterpenen - vluchtige verbindingen - biosynthese - pyrethrinen - plants - pest resistance - defence mechanisms - insect pests - frankliniella occidentalis - monoterpenes - volatile compounds - biosynthesis - pyrethrins
Western flower thrips (WFT), Frankliniella occidentalis, is one of the most serious pests in several vegetable and flower crops worldwide. It is a highly polyphagous insect and a vector of several plant viruses of which the Tomato Spotted Wilt Virus and the Impatiens Necrotic Spot Virus are the most important. Feeding by WFT causes light coloured patches on leaves, petals and fruits, stunted plant growth, and flower and fruit deformation. Synthetic pesticides has been widely used to control WFT. However, the frequent use of these pesticides leads to rapid resistance in WFT, and they are a threat to the environment. Therefore, it is desirable to identify natural sources of resistance effective against WFT to allow breeders to improve resistance in crop species.
Monoterpenes, as constituents of floral scents and plant resins, play an important role in pollinator attraction and in direct and indirect plant defence against pest insects and pathogens. For example, linalool is a common floral scent constituent and found to be emitted from the leaves by many plant species after herbivore attack. In earlier work, linalool-overexpressing Arabidopsis has been tested for resistance to the pest aphid, Myzus persicae, in dual-choice assays, and transgenic plants significantly repelled or deterred the aphids. A linalool synthase (LIS) was overexpressed in chrysanthemum plants and studied the effect of transgenic plants on WFT (Chapter 2). The volatiles from leaves of transgenic plants were significantly attractive to WFT, however, WFT were significantly deterred by the content of leaf discs from transgenic plants. The headspace analysis showed that the volatiles of LIS chrysanthemum leaves were strongly dominated by linalool,but, they also emitted small amounts of the C11-homoterpene, (3E)-4,8-dimethyl-1,3,7-nonatriene, a derivative of nerolidol. In addition, LC-MS analysis showed that several non-volatile linalool glycosides were significantly increased in the leaves of LIS chrysanthemum compared with leaves of wild-type plants. A geraniol synthase (GES) was overexpressed in maize to see whether WFT could be affected by geraniol or its derivatives (Chapter 3). However, geraniol produced in transgenic maize was all efficiently converted to non-volatile glycoside, geranoyl-6-O-malonyl-β-D-glucopyranoside, and GES maize had no effect on WFT behaviour. These studies demonstrate complex effects of terpene engineering on the metabolic changes in transgenic plants. These results suggest that the release/glycosylation of terpenes should be controlled to improve plant resistance against WFT upon metabolic engineering with terpene synthases.
The research subsequently focused on a well-known natural pesticide—pyrethrins. Pyrethrins comprise a group of six closely related esters, derived from the monoterpene alcohol chrysanthemol. Pyrethrins are the economically most important natural insecticide with broad uses in homes, agriculture and stored products for more than 150 years. The effect of pyrethrins against WFT was evaluated on its survival, feeding behaviour, and reproduction both in vitro and in planta (infiltrated chrysanthemum leaves) (Chapter 4). Pyrethrins at 0.1% (w/v) and 1% (w/v) exhibited a significantly negative effect on feeding, and the effects of natural concentrations of pyrethrins in pyrethrum leaves can explain the observed high mortality of WFT feeding on pyrethrum leaves. After the finding of this strong effect of pyrethrins on WFT, the study on the biosynthetic pathway of pyrethrins was continued in order to introduce pyrethrin biosynthesis in transgenic plants. A second function of the published enzyme, chrysanthemyl diphosphate synthase (CDS) was identified (Chapter 5). CDS has been reported to catalyse the formation of chrysanthemyl diphosphate (CPP). However, CDS was demonstrated to also catalyse the next step of CPP into chrysanthemol both in vitro and in vivo. CDS was proposed to be renamed as a chrysanthemol synthase (CHS) using DMAPP as substrate. The gene involved in the next step converting chrysanthemol to chrysanthemic acid has also been characterized (Ramirez, 2013). A chrysanthemic acid:CoA
ligase, which is involved in the final stage of pyrethrin biosynthesis was also studied (Chapter 6). The function of this enzyme was confirmed in vitro and the encoding gene showed a similar expression pattern as CHS in several different tissues and flower developmental stages. The gene responsible for making the final esters is a GDSL-lipase-like acyltransferase (Kikuta et al., 2012). We assume still three to four enzymes are required for the biosynthesis of the basic one of the six pyrethrin esters, jasmolin I, from the precursors DMAPP and jasmonic acid which are universal in plants. And four to five extra genes are required for the complete biosynthesis of all six pyrethrin esters.
In this study, new insights were gained for the biosynthesis of monoterpenes and their derivatives and conjugates, as well as for plant resistance to WFT mediated by these compounds. The characterization of genes involved in pyrethrin biosynthesis paves the way for metabolic engineering of this natural pesticide in other crops.
Groene veredeling: Veredelingsonderzoek tripsresistentie prei
Scholten, O.E. ; Burger, K. ; Henken, G. ; Vosman, B. - \ 2012
resistentieveredeling - preien - plantenveredeling - thrips - plaagresistentie - vollegrondsgroenten - resistance breeding - leeks - plant breeding - thrips - pest resistance - field vegetables
Poster met onderzoeksinformatie.
Abscisinezuur speelt rol bij resistentie tegen stress-omstandigheden : je kunt de plant ook te veel 'pamperen'
Heuvelink, E. ; Kierkels, T. - \ 2012
Onder Glas 9 (2012)11. - p. 26 - 27.
glastuinbouw - plantenziekten - abscisinezuur - hormonen - resistentiemechanismen - cultuurmethoden - plaagresistentie - weerstand - greenhouse horticulture - plant diseases - abscisic acid - hormones - resistance mechanisms - cultural methods - pest resistance - resistance
Abscisinezuur zorgt ervoor dat planten ongunstige omstandigheden kunnen overleven. In de kas proberen we zulke situaties juist te vermijden. Maar je kunt de plant ook te veel ‘pamperen’. Dat kan leiden tot een ongevoeligheid voor dit ‘stress-hormoon’ die zich later wreekt.