Consequences of squash (Cucurbita argyrosperma) domestication for plant defence and herbivore interactions.

MAIN CONCLUSION: Cucurbita argyrosperma domestication affected plant defence by downregulating the cucurbitacin synthesis-associated genes. However, tissue-specific suppression of defences made the cultivars less attractive to co-evolved herbivores Diabrotica balteata and Acalymma spp. Plant domestication reduces the levels of defensive compounds, increasing susceptibility to insects. In squash, the reduction of cucurbitacins has independently occurred several times during domestication. The mechanisms underlying these changes and their consequences for insect herbivores remain unknown. We investigated how Cucurbita argyrosperma domestication has affected plant chemical defence and the interactions with two herbivores, the generalist Diabrotica balteata and the specialist Acalymma spp. Cucurbitacin levels and associated genes in roots and cotyledons in three wild and four domesticated varieties were analysed. Domesticated varieties contained virtually no cucurbitacins in roots and very low amounts in cotyledons. Contrastingly, cucurbitacin synthesis-associated genes were highly expressed in the roots of wild populations. Larvae of both insects strongly preferred to feed on the roots of wild squash, negatively affecting the generalist's performance but not that of the specialist. Our findings illustrate that domestication results in tissue-specific suppression of chemical defence, making cultivars less attractive to co-evolved herbivores. In the case of squash, this may be driven by the unique role of cucurbitacins in stimulating feeding in chrysomelid beetles.

A beta-glucosidase of an insect herbivore determines both toxicity and deterrence of a dandelion defense metabolite.

Gut enzymes can metabolize plant defense compounds and thereby affect the growth and fitness of insect herbivores. Whether these enzymes also influence feeding preference is largely unknown. We studied the metabolization of taraxinic acid ß-D-glucopyranosyl ester (TA-G), a sesquiterpene lactone of the common dandelion (Taraxacum officinale) that deters its major root herbivore, the common cockchafer larva (Melolontha melolontha). We have demonstrated that TA-G is rapidly deglucosylated and conjugated to glutathione in the insect gut. A broad-spectrum M. melolontha ß-glucosidase, Mm_bGlc17, is sufficient and necessary for TA-G deglucosylation. Using cross-species RNA interference, we have shown that Mm_bGlc17 reduces TA-G toxicity. Furthermore, Mm_bGlc17 is required for the preference of M. melolontha larvae for TA-G-deficient plants. Thus, herbivore metabolism modulates both the toxicity and deterrence of a plant defense compound. Our work illustrates the multifaceted roles of insect digestive enzymes as mediators of plant-herbivore interactions.

Plants produce certain substances to fend off attackers like plant-feeding insects. To stop these compounds from damaging their own cells, plants often attach sugar molecules to them. When an insect tries to eat the plant, the plant removes the stabilizing sugar, 'activating' the compounds and making them toxic or foul-tasting. Curiously, some insects remove the sugar themselves, but it is unclear what consequences this has, especially for insect behavior. Dandelions, Taraxacum officinale, make high concentrations of a sugar-containing defense compound in their roots called taraxinic acid ß-D-glucopyranosyl ester, or TA-G for short. TA-G deters the larvae of the Maybug ­ a pest also known as the common cockchafer or the doodlebug ­ from eating dandelion roots. When Maybug larvae do eat TA-G, it is found in their systems without its sugar. However, it is unclear whether it is the plant or the larva that removes the sugar. A second open question is how the sugar removal process affects the behavior of the Maybug larvae. Using chemical analysis and genetic manipulation, Huber et al. investigated what happens when Maybug larvae eat TA-G. This revealed that the acidity levels in the larvae's digestive system deactivate the proteins from the dandelion that would normally remove the sugar from TA-G. However, rather than leaving the compound intact, larvae remove the sugar from TA-G themselves. They do this using a digestive enzyme, known as a beta-glucosidase, that cuts through sugar. Removing the sugar from TA-G made the compound less toxic, allowing the larvae to grow bigger, but it also increased TA-G's deterrent effects, making the larvae less likely to eat the roots. Any organism that eats plants, including humans, must deal with chemicals like TA-G in their food. Once inside the body, enzymes can change these chemicals, altering their effects. This happens with many medicines, too. In the future, it might be possible to design compounds that activate only in certain species, or under certain conditions. Further studies in different systems may aid the development of new methods of pest control, or new drug treatments.

When resistance is futile, tolerate instead: silicon promotes plant compensatory growth when attacked by above- and belowground herbivores.

Plants have evolved numerous herbivore defences that are resistance- or tolerance-based. Resistance involves physical and chemical traits that deter and/or harm herbivores whereas tolerance minimizes fitness costs of herbivory, often via compensatory growth. The Poaceae frequently accumulate large amounts of silicon (Si), which can be used for herbivore resistance, including biomechanical and (indirectly) biochemical defences. To date, it is unclear whether Si improves tolerance of herbivory. Here we report how Si enabled a cereal (Triticum aestivum) to tolerate damage inflicted by above- and belowground herbivores. Leaf herbivory increased Si concentrations in the leaves by greater than 50% relative to herbivore-free plants, indicating it was an inducible defensive response. In plants without Si supplementation, leaf herbivory reduced shoot biomass by 52% and root herbivory reduced root biomass by 68%. Si supplementation, however, facilitated compensatory growth such that shoot losses were more than compensated for (+14% greater than herbivore-free plants) and root losses were minimized to -16%. Si supplementation did not improve plant resistance since Si did not enhance biomechanical resistance (i.e. force of fracture) or reduce leaf consumption and herbivore relative growth rates. We propose that Si-based defence operates in wheat via tolerance either in addition or as an alternative to resistance-based defence.

Environmental gradients and the evolution of tri-trophic interactions.

Long-standing theory predicts herbivores and predators should drive selection for increased plant defences, such as the specific production of volatile organic compounds for attracting predators near the site of damage. Along elevation gradients, a general pattern is that herbivores and predators are abundant at low elevation and progressively diminish at higher elevations. To determine whether plant adaptation along such a gradient influences top-down control of herbivores, we manipulated soil predatory nematodes, root herbivore pressure and plant ecotypes in a reciprocal transplant experiment. Plant survival was significantly higher for low-elevation plants, but only when in the presence of predatory nematodes. Using olfactometer bioassays, we showed correlated differential nematode attraction and plant ecotype-specific variation in volatile production. This study not only provides an assessment of how elevation gradients modulate the strength of trophic cascades, but also demonstrates how habitat specialisation drives variation in the expression of indirect plant defences.

Terpene Synthase Genes in Quercus robur - Gene Characterization, Expression and Resulting Terpenes Due to Cockchafer Feeding.

Root herbivory caused by larvae of the forest cockchafer (Melolontha hippocastani) enhances the impact of drought on trees, particularly in oak forest rejuvenations. In Germany, geographically distant oak stands show differences in infestation strength by the forest cockchafer. While in Southwestern Germany this insect causes severe damage, oak forests in northern Germany are rarely infested. It is known that root-released volatile organic compounds (VOCs) are perceived by soil herbivores, thus guiding the larvae toward the host roots. In this work, we exposed seedlings of two distant oak provenances to forest cockchafer larvae and studied their population genetic properties, their root-based VOC chemotypes, their attraction for larvae and terpene synthase gene expression. Based on nuclear and chloroplast marker analysis, we found both oak populations to be genetically highly variable while showing typical patterns of migration from different refugial regions. However, no clear association between genetic constitution of the different provenances and the abundance of cockchafer populations on site was observed. In contrast to observations in the field, bioassays revealed a preference of the larvae for the northeastern oak provenance. The behavior of larvae was most likely related to root-released volatile terpenes and benzenoids since their composition and quantity differed between oak populations. We assume repellent effects of these compounds because the populations attractive to insects showed low abundance of these compounds. Five different oak terpene synthase (TPS) genes were identified at the genomic level which can be responsible for biosynthesis of the released terpenes. TPS gene expression patterns in response to larval feeding revealed geographic variation rather than genotypic variation. Our results support the assumption that root-released VOC are influencing the perception of roots by herbivores.

A Herbivore Tag-and-Trace System Reveals Contact- and Density-Dependent Repellence of a Root Toxin.

Foraging behavior of root feeding organisms strongly affects plant-environment-interactions and ecosystem processes. However, the impact of plant chemistry on root herbivore movement in the soil is poorly understood. Here, we apply a simple technique to trace the movement of soil-dwelling insects in their habitats without disturbing or restricting their interactions with host plants. We tagged the root feeding larvae of Melolontha melolontha with a copper ring and repeatedly located their position in relation to their preferred host plant, Taraxacum officinale, using a commercial metal detector. This method was validated and used to study the influence of the sesquiterpene lactone taraxinic acid ß-D-glucopyranosyl ester (TA-G) on the foraging of M. melolontha. TA-G is stored in the latex of T. officinale and protects the roots from herbivory. Using behavioral arenas with TA-G deficient and control plants, we tested the impact of physical root access and plant distance on the effect of TA-G on M. melolontha. The larvae preferred TA-G deficient plants to control plants, but only when physical root contact was possible and the plants were separated by 5 cm. Melolontha melolontha showed no preference for TA-G deficient plants when the plants were grown 15 cm apart, which may indicate a trade-off between the cost of movement and the benefit of consuming less toxic food. We demonstrate that M. melolontha integrates host plant quality and distance into its foraging patterns and suggest that plant chemistry affects root herbivore behavior in a plant-density dependent manner.

Volatiles released by Chinese liquorice roots mediate host location behaviour by neonate Porphyrophora sophorae (Hemiptera: Margarodidae).

BACKGROUND: The cochineal scale, Porphyrophora sophorae (Hemiptera: Coccoidea, Margarodidae), is one of the most serious arthropod pests of Chinese liquorice, Glycyrrhiza uralensis (Fabaceae), an important medicinal herb. The adult females tend to deposit the ovisacs in soil relatively far away from liquorice plants. After hatching, neonates move out of the soil and may use chemical cues to search for new hosts. RESULTS: We collected and analysed the volatiles from soils with and without liquorice roots, and chromatographic profiles revealed hexanal, ß-pinene and hexanol as potential host-finding cues for P. sphorae. The attractiveness of these compounds to neonates was studied in the laboratory using four-arm olfactometer bioassays. The larvae showed a clear preference for ß-pinene over hexanal and hexanol, as well as all possible combinations of the three compounds. In addition, a field experiment confirmed that ß-pinene was significantly more attractive than hexanal and hexanol. CONCLUSION: Newly eclosed larvae of P. sphorae exploit root volatiles as chemical cues to locate their host plant. ß-Pinene proved to be the major chemical cue used by P. sphorae neonates searching for roots of their host plant. © 2016 Society of Chemical Industry.

Amino acid-mediated impacts of elevated carbon dioxide and simulated root herbivory on aphids are neutralized by increased air temperatures.

Changes in host plant quality, including foliar amino acid concentrations, resulting from global climate change and attack from multiple herbivores, have the potential to modify the pest status of insect herbivores. This study investigated how mechanically simulated root herbivory of lucerne (Medicago sativa) before and after aphid infestation affected the pea aphid (Acyrthosiphon pisum) under elevated temperature (eT) and carbon dioxide concentrations (eCO2). eT increased plant height and biomass, and eCO2 decreased root C:N. Foliar amino acid concentrations and aphid numbers increased in response to eCO2, but only at ambient temperatures, demonstrating the ability of eT to negate the effects of eCO2. Root damage reduced aboveground biomass, height, and root %N, and increased root %C and C:N, most probably via decreased biological nitrogen fixation. Total foliar amino acid concentrations and aphid colonization success were higher in plants with roots cut early (before aphid arrival) than those with roots cut late (after aphid arrival); however, this effect was counteracted by eT. These results demonstrate the importance of amino acid concentrations for aphids and identify individual amino acids as being potential factors underpinning aphid responses to eT, eCO2, and root damage in lucerne. Incorporating trophic complexity and multiple climatic factors into plant-herbivore studies enables greater insight into how plants and insects will interact in the future, with implications for sustainable pest control and future crop security.

The role of carbon dioxide as an orientation cue for western corn rootworm larvae within the maize root system: implications for an attract-and-kill approach.

BACKGROUND: Western corn rootworm larvae use CO2 to locate maize roots. However, the importance of CO2 as a specific orientation cue close to maize roots has not been investigated unequivocally. This study aimed at elucidating the effect of CO2 -emitting capsules in combination with a soil insecticide (Tefluthrin = attract and kill) within the root system. We hypothesized that the capsules would result in aggregation of the larvae at the soil insecticide, thus increasing its efficacy. A nondestructive observation device was used to study larval distribution and behaviour. RESULTS: Spatial analysis of distance indices (SADIE) revealed aggregation of the larvae around the capsules in an attract-and-kill treatment after 4 h, which was not found with the conventional treatment without the capsules. However, larval mortality did not differ between treatments. CONCLUSION: CO2 is a weak attractant for western corn rootworm larvae within the root system. Consequently, an attract-and-kill strategy based on a CO2 product will not contribute to better control compared with conventional Tefluthrin applications. Host-specific compounds, combined with a CO2 source, should be used to target more larvae, making attract and kill a feasible management option against this pest.

Downstairs drivers--root herbivores shape communities of above-ground herbivores and natural enemies via changes in plant nutrients.

1. Terrestrial food webs are woven from complex interactions, often underpinned by plant-mediated interactions between herbivores and higher trophic groups. Below- and above-ground herbivores can influence one another via induced changes to a shared host plant, potentially shaping the wider community. However, empirical evidence linking laboratory observations to natural field populations has so far been elusive. 2. This study investigated how root-feeding weevils (Otiorhynchus sulcatus) influence different feeding guilds of herbivore (phloem-feeding aphids, Cryptomyzus galeopsidis, and leaf-chewing sawflies, Nematus olfaciens) in both controlled and field conditions. 3. We hypothesized that root herbivore-induced changes in plant nutrients (C, N, P and amino acids) and defensive compounds (phenolics) would underpin the interactions between root and foliar herbivores, and ultimately populations of natural enemies of the foliar herbivores in the field. 4. Weevils increased field populations of aphids by ca. 700%, which was followed by an increase in the abundance of aphid natural enemies. Weevils increased the proportion of foliar essential amino acids, and this change was positively correlated with aphid abundance, which increased by 90% on plants with weevils in controlled experiments. 5. In contrast, sawfly populations were 77% smaller during mid-June and adult emergence delayed by >14 days on plants with weevils. In controlled experiments, weevils impaired sawfly growth by 18%, which correlated with 35% reductions in leaf phosphorus caused by root herbivory, a previously unreported mechanism for above-ground-below-ground herbivore interactions. 6. This represents a clear demonstration of root herbivores affecting foliar herbivore community composition and natural enemy abundance in the field via two distinct plant-mediated nutritional mechanisms. Aphid populations, in particular, were initially driven by bottom-up effects (i.e. plant-mediated effects of root herbivory), but consequent increases in natural enemies triggered top-down regulation.