Colette Broekgaarden

Early season herbivore differentially affects plant defence responses to subsequently colonizing herbivores and their abundance in the field

Induction of plant defences by early season herbivores can mediate interspecific herbivore competition. We have investigated plant‐mediated competition between three herbivorous insects through studies at different levels of biological integration. We have addressed (i) gene expression; (ii) insect behaviour and performance under laboratory conditions; and (iii) population dynamics under field conditions. We studied the expression of genes encoding a trypsin inhibitor and genes that are involved in glucosinolate biosynthesis in response to early season herbivory by Pieris rapae caterpillars in Brassica oleracea plants. Furthermore, we studied the interaction of these transcriptional responses with responses to secondary herbivory by the two specialist herbivores, P. rapae and Plutella xylostella, and the generalist Mamestra brassicae. P. rapae‐induced responses strongly interacted with plant responses to secondary herbivory. Sequential feeding by specialist herbivores resulted in enhanced or similar expression levels of defence‐related genes compared to primary herbivory by specialists. Secondary herbivory by the generalist M. brassicae resulted in lower gene expression levels than in response to primary herbivory by this generalist. Larval performance of both specialist and generalist herbivores was negatively affected by P. rapae‐induced plant responses. However, in the field the specialist P. xylostella was more abundant on P. rapae‐induced plants and preferred these plants over undamaged plants in oviposition experiments. In contrast, the generalist M. brassicae was more abundant on control plants and preferred undamaged plants for oviposition. P. rapae did not discriminate between plants damaged by conspecifics or undamaged plants. Our study shows that early season herbivory differentially affects transcriptional responses involved in plant defence to secondary herbivores and their population development dependent upon their degree of host plant specialization.

Jasmonate and ethylene signaling mediate whitefly‐induced interference with indirect plant defense in Arabidopsis thaliana

Upon herbivore attack, plants activate an indirect defense, that is, the release of a complex mixture of volatiles that attract natural enemies of the herbivore. When plants are simultaneously exposed to two herbivore species belonging to different feeding guilds, one herbivore may interfere with the indirect plant defense induced by the other herbivore. However, little is understood about the mechanisms underlying such interference. Here, we address the effect of herbivory by the phloem-feeding whitefly Bemisia tabaci on the induced indirect defense of Arabidopsis thaliana plants to Plutella xylostella caterpillars, that is, the attraction of the parasitoid wasp Diadegma semiclausum. Assays with various Arabidopsis mutants reveal that B. tabaci infestation interferes with indirect plant defense induced by P. xylostella, and that intact jasmonic acid and ethylene signaling are required for such interference caused by B. tabaci. Chemical analysis of plant volatiles showed that the composition of the blend emitted in response to the caterpillars was significantly altered by co-infestation with whiteflies. Moreover, whitefly infestation also had a considerable effect on the transcriptomic response of the plant to the caterpillars. Understanding the mechanisms underlying a plants responses to multiple attackers will be important for the development of crop protection strategies in a multi-attacker context.

Natural variation in herbivore-induced volatiles in Arabidopsis thaliana

To study whether natural variation in Arabidopsis thaliana could be used to dissect the genetic basis of responses to herbivory in terms of induced volatile emissions, nine accessions were characterized upon herbivory by biting-chewing Pieris rapae caterpillars or after treatment with the phytohormone jasmonic acid (JA). Analysis of 73 compounds in the headspace showed quantitative differences in the emission rates of several individual compounds among the accessions. Moreover, variation in the emission of volatile compounds after JA treatment was reflected in the behaviour of the parasitoid Diadegma semiclausum when they were offered the headspace volatiles of several combinations of accessions in two-choice experiments. Accessions also differ in transcript levels of genes that are associated with the emission of plant volatiles. The genes BSMT1 and Cyp72A13 could be connected to the emission of methyl salicylate and (E,E)-4,8,12-trimethyltrideca-1,3,7,11-tetraene (TMTT), respectively. Overall, Arabidopsis showed interesting phenotypic variations with respect to the volatile blend emitted in response to herbivory that can be exploited to identify genes and alleles that underlie this important plant trait.

Responses of Brassica oleracea cultivars to infestation by the aphid Brevicoryne brassicae: an ecological and molecular approach

Intraspecific variation in resistance or susceptibility to herbivorous insects has been widely studied through bioassays. However, few studies have combined this with a full transcriptomic analysis. Here, we take such an approach to study the interaction between the aphid Brevicoryne brassicae and four white cabbage (Brassica oleracea var. capitata) cultivars. Both under glasshouse and field conditions, two of the cultivars clearly supported a faster aphid population development than the other two, indicating that aphid population development was largely independent of the environmental conditions. Genome-wide transcriptomic analysis using 70-mer oligonucleotide microarrays based on the Arabidopsis thaliana genome showed that only a small number of genes were differentially regulated, and that this regulation was highly cultivar specific. The temporal pattern in the expression behaviour of two B. brassicae-responsive genes in all four cultivars together with targeted studies employing A. thaliana knockout mutants revealed a possible role for a trypsin-and-protease inhibitor in defence against B. brassicae. Conversely, a xyloglucan endotransglucosylase seemed to have no effect on aphid performance. Overall, this study shows clear intraspecific variation in B. brassicae susceptibility among B. oleracea cultivars under glasshouse and field conditions that can be partly explained by certain differences in induced transcriptional changes.

Genotypic variation in genome-wide transcription profiles induced by insect feeding: Brassica oleracea--Pieris rapae interactions.

BackgroundTranscriptional profiling after herbivore attack reveals, at the molecular level, how plants respond to this type of biotic stress. Comparing herbivore-induced transcriptional responses of plants with different phenotypes provides insight into plant defense mechanisms. Here, we compare the global gene expression patterns induced by Pieris rapae caterpillar attack in two white cabbage (Brassica oleracea var. capitata) cultivars. The two cultivars are shown to differ in their level of direct defense against caterpillar feeding. Because Brassica full genome microarrays are not yet available, 70-mer oligonucleotide microarrays based on the Arabidopsis thaliana genome were used for this non-model plant.ResultsThe transcriptional responses of the two cultivars differed in timing as characterized by changes in their expression pattern after 24, 48 and 72 hours of caterpillar feeding. In addition, they also differed qualitatively. Surprisingly, of all genes induced at any time point, only one third was induced in both cultivars. Analyses of transcriptional responses after jasmonate treatment revealed that the difference in timing did not hold for the response to this phytohormone. Additionally, comparisons between Pieris rapae- and jasmonate-induced transcriptional responses showed that Pieris rapae induced more jasmonate-independent than jasmonate-dependent genes.ConclusionThe present study clearly shows that global transcriptional responses in two cultivars of the same plant species in response to insect feeding can differ dramatically. Several of these differences involve genes that are known to have an impact on Pieris rapae performance and probably underlie different mechanisms of direct defense, present in the cultivars.

Exploiting natural variation to identify insect‐resistance genes

Herbivorous insects are widespread and often serious constraints to crop production. The use of insect-resistant crops is a very effective way to control insect pests in agriculture, and the development of such crops can be greatly enhanced by knowledge on plant resistance mechanisms and the genes involved. Plants have evolved diverse ways to cope with insect attack that has resulted in natural variation for resistance towards herbivorous insects. Studying the molecular genetics and transcriptional background of this variation has facilitated the identification of resistance genes and processes that lead to resistance against insects. With the development of new technologies, molecular studies are not restricted to model plants anymore. This review addresses the need to exploit natural variation in resistance towards insects to increase our knowledge on resistance mechanisms and the genes involved. We will discuss how this knowledge can be exploited in breeding programmes to provide sustainable crop protection against insect pests. Additionally, we discuss the current status of genetic research on insect-resistance genes. We conclude that insect-resistance mechanisms are still unclear at the molecular level and that exploiting natural variation with novel technologies will contribute greatly to the development of insect-resistant crop varieties.

Ethylene: Traffic Controller on Hormonal Crossroads to Defense.

The hormone ethylene plays an important modulating role in the immune signaling network that regulates defense against microbial pathogens and insect herbivores. Ethylene (ET) is an important hormone in plant responses to microbial pathogens and herbivorous insects, and in the interaction of plants with beneficial microbes and insects. Early ET signaling events during these biotic interactions involve activities of mitogen-activated protein kinases and ETHYLENE RESPONSE FACTOR transcription factors. Rather than being the principal regulator, ET often modulates defense signaling pathways, including those regulated by jasmonic acid and salicylic acid. Hormonal signal integrations with ET steer the defense signaling network to activate specific defenses that can have direct effects on attackers, or systemically prime distant plant parts for enhanced defense against future attack. ET also regulates volatile signals that attract carnivorous enemies of herbivores or warn neighboring plants. Conversely, ET signaling can also be exploited by attackers to hijack the defense signaling network to suppress effective defenses. In this review, we summarize recent findings on the significant role of ET in the plants’ battle against their enemies.

Transcriptional responses of Brassica nigra to feeding by specialist insects of different feeding guilds

Abstract  Plants show phenotypic changes when challenged with herbivorous insects. The mechanisms underlying these changes include the activation of transcriptional responses, which are dependent on the attacking insect. Most transcriptomic studies on crucifer–insect interactions have focused on the model plant Arabidopsis thaliana, a species that faces low herbivore pressure in nature. Here, we study the transcriptional responses of plants from a wild black mustard (Brassica nigra) population to herbivores of different feeding guilds using an A. thaliana‐based whole‐genome microarray that has previously been shown to be suitable for transcriptomic analyses in Brassica. Transcriptional responses of B. nigra after infestation with either Pieris rapae caterpillars or Brevicoryne brassicae aphids are analyzed and compared. Additionally, the insect‐induced expression changes of some individual genes are analyzed through quantitative real‐time polymerase chain reaction. The results show that feeding by both insect species results in the accumulation of transcripts encoding proteins involved in the detoxification of reactive oxygen species, defensive proteins and glucosinolates and this is correlated with experimental evidence in the literature on such biochemical effects. Although genes encoding proteins involved in similar processes are regulated by both insects, there was little overlap in the induction or repression of individual genes. Furthermore, P. rapae and B. brassicae seem to affect different phytohormone signaling pathways. In conclusion, our results indicate that B. nigra activates several defense‐related genes in response to P. rapae or B. brassicae feeding, but that the response is dependent on the attacking insect species.

Genetic architecture of plant stress resistance: multi‐trait genome‐wide association mapping

Summary Plants are exposed to combinations of various biotic and abiotic stresses, but stress responses are usually investigated for single stresses only. Here, we investigated the genetic architecture underlying plant responses to 11 single stresses and several of their combinations by phenotyping 350 Arabidopsis thaliana accessions. A set of 214 000 single nucleotide polymorphisms (SNPs) was screened for marker‐trait associations in genome‐wide association (GWA) analyses using tailored multi‐trait mixed models. Stress responses that share phytohormonal signaling pathways also share genetic architecture underlying these responses. After removing the effects of general robustness, for the 30 most significant SNPs, average quantitative trait locus (QTL) effect sizes were larger for dual stresses than for single stresses. Plants appear to deploy broad‐spectrum defensive mechanisms influencing multiple traits in response to combined stresses. Association analyses identified QTLs with contrasting and with similar responses to biotic vs abiotic stresses, and below‐ground vs above‐ground stresses. Our approach allowed for an unprecedented comprehensive genetic analysis of how plants deal with a wide spectrum of stress conditions.

High throughput phenotyping for aphid resistance in large plant collections

BackgroundPhloem-feeding insects are among the most devastating pests worldwide. They not only cause damage by feeding from the phloem, thereby depleting the plant from photo-assimilates, but also by vectoring viruses. Until now, the main way to prevent such problems is the frequent use of insecticides. Applying resistant varieties would be a more environmental friendly and sustainable solution. For this, resistant sources need to be identified first. Up to now there were no methods suitable for high throughput phenotyping of plant germplasm to identify sources of resistance towards phloem-feeding insects.ResultsIn this paper we present a high throughput screening system to identify plants with an increased resistance against aphids. Its versatility is demonstrated using an Arabidopsis thaliana activation tag mutant line collection. This system consists of the green peach aphid Myzus persicae (Sulzer) and the circulative virus Turnip yellows virus (TuYV). In an initial screening, with one plant representing one mutant line, 13 virus-free mutant lines were identified by ELISA. Using seeds produced from these lines, the putative candidates were re-evaluated and characterized, resulting in nine lines with increased resistance towards the aphid.ConclusionsThis M. persicae-TuYV screening system is an efficient, reliable and quick procedure to identify among thousands of mutated lines those resistant to aphids. In our study, nine mutant lines with increased resistance against the aphid were selected among 5160 mutant lines in just 5 months by one person. The system can be extended to other phloem-feeding insects and circulative viruses to identify insect resistant sources from several collections, including for example genebanks and artificially prepared mutant collections.