Assessment of pleiotropic transcriptome perturbations in Arabidopsis engineered for indirect insect defence.

BACKGROUND: Molecular characterization is an essential step of risk/safety assessment of genetically modified (GM) crops. Holistic approaches for molecular characterization using omics platforms can be used to confirm the intended impact of the genetic engineering, but can also reveal the unintended changes at the omics level as a first assessment of potential risks. The potential of omics platforms for risk assessment of GM crops has rarely been used for this purpose because of the lack of a consensus reference and statistical methods to judge the significance or importance of the pleiotropic changes in GM plants. Here we propose a meta data analysis approach to the analysis of GM plants, by measuring the transcriptome distance to untransformed wild-types. RESULTS: In the statistical analysis of the transcriptome distance between GM and wild-type plants, values are compared with naturally occurring transcriptome distances in non-GM counterparts obtained from a database. Using this approach we show that the pleiotropic effect of genes involved in indirect insect defence traits is substantially equivalent to the variation in gene expression occurring naturally in Arabidopsis. CONCLUSION: Transcriptome distance is a useful screening method to obtain insight in the pleiotropic effects of genetic modification.

Genetic engineering of plant volatile terpenoids: effects on a herbivore, a predator and a parasitoid.

BACKGROUND: Most insect-resistant transgenic crops employ toxins to control pests. A novel approach is to enhance the effectiveness of natural enemies by genetic engineering of the biosynthesis of volatile organic compounds (VOCs). Before the commercialisation of such transgenic plants can be pursued, detailed fundamental studies of their effects on herbivores and their natural enemies are necessary. The linalool/nerolidol synthase gene FaNES1 was constitutively expressed from strawberry in three Arabidopsis thaliana accessions, and the behaviour of the aphid Brevicoryne brassicae L., the parasitoid Diaeretiella rapae McIntosh and the predator Episyrphus balteatus de Geer was studied. RESULTS: Transgenic FaNES1-expressing plants emitted (E)-nerolidol and larger amounts of (E)-DMNT and linalool. Brevicoryne brassicae was repelled by the transgenic lines of two of the accessions, whereas its performance was not affected. Diaeretiella rapae preferred aphid-infested transgenic plants over aphid-infested wild-type plants for two of the accessions. In contrast, female E. balteatus predators did not differentiate between aphid-infested transgenic or wild-type plants. CONCLUSION: The results indicate that the genetic engineering of plants to modify their emission of VOCs holds considerable promise for facilitating biological control of herbivores. Validation for crop plants is a necessary next step to assess the usefulness of modified volatile emission in integrated pest management.

Three-step pathway engineering results in more incidence rate and higher emission of nerolidol and improved attraction of Diadegma semiclausum.

The concentration and ratio of terpenoids in the headspace volatile blend of plants have a fundamental role in the communication of plants and insects. The sesquiterpene (E)-nerolidol is one of the important volatiles with effect on beneficial carnivores for biologic pest management in the field. To optimize de novo biosynthesis and reliable and uniform emission of (E)-nerolidol, we engineered different steps of the (E)-nerolidol biosynthesis pathway in Arabidopsis thaliana. Introduction of a mitochondrial nerolidol synthase gene mediates de novo emission of (E)-nerolidol and linalool. Co-expression of the mitochondrial FPS1 and cytosolic HMGR1 increased the number of emitting transgenic plants (incidence rate) and the emission rate of both volatiles. No association between the emission rate of transgenic volatiles and their growth inhibitory effect could be established. (E)-Nerolidol was to a large extent metabolized to non-volatile conjugates.

Characterization of the natural variation in Arabidopsis thaliana metabolome by the analysis of metabolic distance.

Metabolite fingerprinting is widely used to unravel the chemical characteristics of biological samples. Multivariate data analysis and other statistical tools are subsequently used to analyze and visualize the plasticity of the metabolome and/or the relationship between those samples. However, there are limitations to these approaches for example because of the multi-dimensionality of the data that makes interpretation of the data obtained from untargeted analysis almost impossible for an average human being. These limitations make the biological information that is of prime importance in untargeted studies be partially exploited. Even in the case of full exploitation, current methods for relationship elucidation focus mainly on between groups variation and differences. Therefore, a measure that is capable of exploiting both between- and within-group biological variation would be of great value. Here, we examined the natural variation in the metabolome of nine Arabidopsis thaliana accessions grown under various environmental conditions and established a measure for the metabolic distance between accessions and across environments. This data analysis approach shows that there is just a minor correlation between genetic and metabolic diversity of the nine accessions. On the other hand, it delivers so far in Arabidopsis unexplored chemical information and is shown to be biologically relevant for resistance studies. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11306-011-0375-3) contains supplementary material, which is available to authorized users.

Effects of glucosinolates on a generalist and specialist leaf-chewing herbivore and an associated parasitoid.

Glucosinolates (GLS) are secondary plant metabolites that as a result of tissue damage, for example due to herbivory, are hydrolysed into toxic compounds that negatively affect generalist herbivores. Specialist herbivores have evolved specific adaptations to detoxify GLS or inhibit the formation of toxic hydrolytic products. Although rarely studied, GLS and their breakdown products may also affect parasitoids. The objectives were to test the effects of GLS in a multitrophic system consisting of the generalist herbivore Spodoptera exigua, the specialist herbivore Pieris rapae, and the endoparasitoid Hyposoter ebeninus. Three ecotypes of Arabidopsis thaliana that differ in their GLS composition and concentrations and one transformed line that constitutively produces higher concentrations of aliphatic GLS were used, the latter allowing a direct assessment of the effects of aliphatic GLS on insect performance. Feeding by the generalist S. exigua and the specialist P. rapae induced both higher aliphatic and indole GLS concentrations in the A. thaliana ecotypes, although induction was stronger for indole than aliphatic GLS. For both herbivores a negative correlation between performance and aliphatic GLS concentrations was observed. This suggests that the specialist, despite containing a nitrile-specifier protein (NSP) that diverts GLS degradation from toxic isothiocyanates to less toxic nitriles, cannot completely inhibit the formation of toxic GLS hydrolytic products, or that the costs of this mechanism are higher at higher GLS concentrations. Surprisingly, performance of the parasitoid was positively correlated with higher concentrations of aliphatic GLS in the plant, possibly caused by negative effects on host immune responses. Our study indicates that GLS can not only confer resistance against herbivores directly, but also indirectly by increasing the performance of the parasitoids of these herbivores.

Herbivore-mediated effects of glucosinolates on different natural enemies of a specialist aphid.

The cabbage aphid Brevicoryne brassicae is a specialist herbivore that sequesters glucosinolates from its host plant as a defense against its predators. It is unknown to what extent parasitoids are affected by this sequestration. We investigated herbivore-mediated effects of glucosinolates on the parasitoid wasp Diaeretiella rapae and the predator Episyrphus balteatus. We reared B. brassicae on three ecotypes of Arabidopsis thaliana that differ in glucosinolate content and on one genetically transformed line with modified concentrations of aliphatic glucosinolates. We tested aphid performance and the performance and behavior of both natural enemies. We correlated this with phloem and aphid glucosinolate concentrations and emission of volatiles. Brevicoryne brassicae performance correlated positively with concentrations of both aliphatic and indole glucosinolates in the phloem. Aphids selectively sequestered glucosinolates. Glucosinolate concentration in B. brassicae correlated negatively with performance of the predator, but positively with performance of the parasitoid, possibly because the aphids with the highest glucosinolate concentrations had a higher body weight. Both natural enemies showed a positive performance-preference correlation. The predator preferred the ecotype with the lowest emission of volatile glucosinolate breakdown products in each test combination, whereas the parasitoid wasp preferred the A. thaliana ecotype with the highest emission of these volatiles. The study shows that there are differential herbivore-mediated effects of glucosinolates on a predator and a parasitoid of a specialist aphid that selectively sequesters glucosinolates from its host plant.

Genetic variation in strigolactone production and tillering in rice and its effect on Striga hermonthica infection.

Tillering in cereals is a complex process in the regulation of which also signals from the roots in the form of strigolactones play an important role. The strigolactones are signalling molecules that are secreted into the rhizosphere where they act as germination stimulants for root parasitic plants and hyphal branching factors for arbuscular mycorrhizal fungi. On the other hand, they are also transported from the roots to the shoot where they inhibit tillering or branching. In the present study, the genetic variation in strigolactone production and tillering phenotype was studied in twenty rice varieties collected from all over the world and correlated with S. hermonthica infection. Rice cultivars like IAC 165, IAC 1246, Gangweondo and Kinko produced high amounts of the strigolactones orobanchol, 2'-epi-5-deoxystrigol and three methoxy-5-deoxystrigol isomers and displayed low amounts of tillers. These varieties induced high S. hermonthica germination, attachment, emergence as well as dry biomass. In contrast, rice cultivars such as Super Basmati, TN 1, Anakila and Agee displayed high tillering in combination with low production of the aforementioned strigolactones. These varieties induced only low S. hermonthica germination, attachment, emergence and dry biomass. Statistical analysis across all the varieties confirmed a positive correlation between strigolactone production and S. hermonthica infection and a negative relationship with tillering. These results show that genetic variation in tillering capacity is the result of genetic variation in strigolactone production and hence could be a helpful tool in selecting rice cultivars that are less susceptible to S. hermonthica infection.