Sybille B. Unsicker

Protective perfumes: the role of vegetative volatiles in plant defense against herbivores

Herbivore damage to leaves and other vegetative tissues often stimulates the emission of volatile compounds, suggesting that these substances have a role in plant defense. In fact, ample evidence has accumulated in the last few years indicating that volatiles from vegetative plant parts can directly repel herbivores, such as ovipositing butterflies and host-seeking aphids. Volatiles have also been demonstrated to protect plants by attracting herbivore enemies, such as parasitic wasps, predatory arthropods and possibly even insectivorous birds. Even below ground herbivory results in the release of volatiles that attract herbivore enemies. However, plant volatiles are also known to attract enemies of plants. Hence, to determine the true value of these substances in defense, more research is needed especially in natural communities with non-agricultural species.

The specificity of herbivore-induced plant volatiles in attracting herbivore enemies

Plants respond to herbivore attack by emitting complex mixtures of volatile compounds that attract herbivore enemies, both predators and parasitoids. Here, we explore whether these mixtures provide significant value as information cues in herbivore enemy attraction. Our survey indicates that blends of volatiles released from damaged plants are frequently specific depending on the type of herbivore and its age, abundance and feeding guild. The sensory perception of plant volatiles by herbivore enemies is also specific, according to the latest evidence from studies of insect olfaction. Thus, enemies do exploit the detailed information provided by plant volatile mixtures in searching for their prey or hosts, but this varies with the diet breadth of the enemy.

Complementarity effects through dietary mixing enhance the performance of a generalist insect herbivore

The ontogenetic niche concept predicts that resource use depends on an organism’s developmental stage. This concept has been investigated primarily in animals that show differing resource use strategies as juveniles and as adults, such as amphibians. We studied resource use and performance in the grasshopper Chorthippus parallelus (Orthoptera, Acrididae) provided with food plant mixtures of either one, three or eight plant species throughout their development. C. parallelus survival and fecundity was highest in the food plant mixture with eight plant species and lowest in the treatments where only one single plant species was offered as food. C. parallelus’ consumption throughout its ontogeny depended on sex, and feeding on different plant species was dependent on a grasshopper’s developmental stage. To depict grasshopper foraging in food plant mixtures compared to foraging on single plant species, we introduce the term “relative forage total” (RFT) based on an approach used in biodiversity research by Loreau and Hector (Nature 413:548–274, 2001). RFT of grasshoppers in food plant mixtures was always higher than what would have been expected from foraging in monocultures. The increase in food consumption was due to an overall increase in feeding on plant species in mixtures compared to consumption of the same species offered as a single diet. Thus we argue that grasshopper foraging exhibits complementarity effects. Our results reinforce the necessity to consider development-related changes in insect herbivore feeding. Thorough information on the feeding ontogeny of insect herbivores could not only elucidate their nutritional ecology but also help to shed light on their functional role in plant communities.

The Effects of Arbuscular Mycorrhizal Fungi on Direct and Indirect Defense Metabolites of Plantago lanceolata L.

Arbuscular mycorrhizal fungi can strongly influence the metabolism of their host plant, but their effect on plant defense mechanisms has not yet been thoroughly investigated. We studied how the principal direct defenses (iridoid glycosides) and indirect defenses (volatile organic compounds) of Plantago lanceolata L. are affected by insect herbivory and mechanical wounding. Volatile compounds were collected and quantified from mycorrhizal and non-mycorrhizal P. lanceolata plants that underwent three different treatments: 1) insect herbivory, 2) mechanical wounding, or 3) no damage. The iridoids aucubin and catalpol were extracted and quantified from the same plants. Emission of terpenoid volatiles was significantly higher after insect herbivory than after the other treatments. However, herbivore-damaged mycorrhizal plants emitted lower amounts of sesquiterpenes, but not monoterpenes, than herbivore-damaged non-mycorrhizal plants. In contrast, mycorrhizal infection increased the emission of the green leaf volatile (Z)-3-hexenyl acetate in untreated control plants, making it comparable to emission from mechanically wounded or herbivore-damaged plants whether or not they had mycorrhizal associates. Neither mycorrhization nor treatment had any influence on the levels of iridoid glycosides. Thus, mycorrhizal infection did not have any effect on the levels of direct defense compounds measured in P. lanceolata. However, the large decline in herbivore-induced sesquiterpene emission may have important implications for the indirect defense potential of this species.

Invertebrate herbivory along a gradient of plant species diversity in extensively managed grasslands

Increasing plant diversity has long been hypothesized to negatively affect levels of invertebrate herbivory due to a lower number of specialist insect herbivores in more diverse sites, but studies of natural systems have been rare. We used a planned comparison to study herbivory in a set of 19 semi-natural montane grasslands managed as hay meadows. Herbivory was measured in transects through the plant communities, and in individuals of Plantago lanceolata and Trifolium pratense that were transplanted into each meadow. In addition, plant community biomass and arthropod abundances were determined in the grasslands. Before the first mowing in June, mean herbivory levels correlated negatively with plant species richness, as predicted by theory, but they were also significantly affected by plant community biomass and plant community composition. After mowing, herbivory levels were only significantly related to plant community composition. Damage levels in the transplants were lower than herbivory levels in the established plant communities. Most insect herbivores were generalists and not specialists. The number of insect herbivores and spiders were positively correlated and tended to increase with increasing plant species richness. Herbivory levels were correlated negatively with spider abundances. We conclude that while the predicted negative relationship between plant species richness and insect herbivory can be found in grasslands, the underlying mechanism involves generalist rather than specialist herbivores. Our data also suggest a role of natural enemies in generalist herbivore activities.

Specific bottom–up effects of arbuscular mycorrhizal fungi across a plant–herbivore–parasitoid system

The majority of plants are involved in symbioses with arbuscular mycorrhizal fungi (AMF), and these associations are known to have a strong influence on the performance of both plants and insect herbivores. Little is known about the impact of AMF on complex trophic chains, although such effects are conceivable. In a greenhouse study we examined the effects of two AMF species, Glomus intraradices and G. mosseae on trophic interactions between the grass Phleum pratense, the aphid Rhopalosiphum padi, and the parasitic wasp Aphidius rhopalosiphi. Inoculation with AMF in our study system generally enhanced plant biomass (+5.2%) and decreased aphid population growth (−47%), but there were no fungal species-specific effects. When plants were infested with G. intraradices, the rate of parasitism in aphids increased by 140% relative to the G. mosseae and control treatment. When plants were associated with AMF, the developmental time of the parasitoids decreased by 4.3% and weight at eclosion increased by 23.8%. There were no clear effects of AMF on the concentration of nitrogen and phosphorus in plant foliage. Our study demonstrates that the effects of AMF go beyond a simple amelioration of the plants’ nutritional status and involve rather more complex species-specific cascading effects of AMF in the food chain that have a strong impact not only on the performance of plants but also on higher trophic levels, such as herbivores and parasitoids.

Emission of volatile organic compounds after herbivory from Trifolium pratense (L.) under laboratory and field conditions

Plants emit a wide range of volatile organic compounds in response to damage by herbivores, and many of the compounds have been shown to attract the natural enemies of insect herbivores or serve for inter- and intra-plant communication. Most studies have focused on volatile emission in the laboratory while little is known about emission patterns in the field. We studied the emission of volatiles by Trifolium pratense (red clover) under both laboratory and field conditions. The emission of 24 compounds was quantified in the laboratory, of which eight showed increased emission rates after herbivory by Spodoptera littoralis caterpillars, including (E)-β-ocimene, the most abundant compound, (Z)-β-ocimene, linalool, (E)-β-caryophyllene, (E,E)-α-farnesene, 4,8-dimethyl-1,3,7-nonatriene (DMNT), 1-octen-3-ol, and methyl salicylate (MeSA). While most of these compounds have been reported as herbivore-induced volatiles from a wide range of plant taxa, 1-octen-3-ol seems to be a characteristic volatile of legumes. In the field, T. pratense plants with varying herbivore damage growing in established grassland communities emitted only 13 detectable compounds, and the correlation between herbivore damage and volatile release was more variable than in the laboratory. For example, the emission of (E)-β-ocimene, (Z)-β-ocimene, and DMNT actually declined with damage, while decanal exhibited increased emission with increasing herbivory. Elevated light and temperature increased the emission of many compounds, but the differences in light and temperature conditions between the laboratory and the field could not account for the differences in emission profiles. Our results indicate that the release of volatiles from T. pratense plants in the field is likely to be influenced by additional biotic and abiotic factors not measured in this study. The elucidation of these factors may be important in understanding the physiological and ecological functions of volatiles in plants.

Impact of invertebrate herbivory in grasslands depends on plant species diversity

Invertebrate herbivores are ubiquitous in most terrestrial ecosystems, and theory predicts that their impact on plant community biomass should depend on diversity and productivity of the associated plant communities. To elucidate general patterns in the relationship between invertebrate herbivory, plant diversity, and productivity, we carried out a long-term herbivore exclusion experiment at multiple grassland sites in a mountainous landscape of central Germany. Over a period of five years, we used above- and belowground insecticides as well as a molluscicide to manipulate invertebrate herbivory at 14 grassland sites, covering a wide range of plant species diversity (13-38 species/m2) and aboveground plant productivity (272-1125 g x m(-2) x yr(-1)), where plant species richness and productivity of the sites were not significantly correlated. Herbivore exclusion had significant effects on the plant communities: it decreased plant species richness and evenness, and it altered plant community composition. In particular, exclusion of belowground herbivores promoted grasses at the expense of herbs. In contrast to our expectation, herbivore effects on plant community biomass were not influenced by productivity. However, effect size of invertebrate herbivores was negatively correlated with plant diversity of the grasslands: the effect of herbivory on biomass tended to be negative at sites of high diversity and positive at sites of low diversity. In general, the effects of aboveground herbivores were relatively small as compared to belowground herbivores, which were important drivers of plant community composition. Our study is the first to show that variation in the effects of invertebrate herbivory on plant communities across a landscape is significantly influenced by plant species richness.

Four terpene synthases produce major compounds of the gypsy moth feeding-induced volatile blend of Populus trichocarpa

After herbivore damage, many plants increase their emission of volatile compounds, with terpenes usually comprising the major group of induced volatiles. Populus trichocarpa is the first woody species with a fully sequenced genome, enabling rapid molecular approaches towards characterization of volatile terpene biosynthesis in this and other poplar species. We identified and characterized four terpene synthases (PtTPS1-4) from P. trichocarpa which form major terpene compounds of the volatile blend induced by gypsy moth (Lymantria dispar) feeding. The enzymes were heterologously expressed and assayed with potential prenyl diphosphate substrates. PtTPS1 and PtTPS2 accepted only farnesyl diphosphate and produced (-)-germacrene D and (E,E)-α-farnesene as their major products, respectively. In contrast, PtTPS3 and PtTPS4 showed both mono- and sesquiterpene synthase activity. They produce the acyclic terpene alcohols linalool and nerolidol but exhibited opposite stereospecificity. qRT-PCR analysis revealed that the expression of the respective terpene synthase genes was induced after feeding of gypsy moth caterpillars. The TPS enzyme products may play important roles in indirect defense of poplar to herbivores and in mediating intra- and inter-plant signaling.

Herbivore-induced volatile emission in black poplar: regulation and role in attracting herbivore enemies

After herbivory, plants release volatile organic compounds from damaged foliage as well as from nearby undamaged leaves that attract herbivore enemies. Little is known about what controls the volatile emission differences between damaged and undamaged tissues and how these affect the orientation of herbivore enemies. We investigated volatile emission from damaged and adjacent undamaged foliage of black poplar (Populus nigra) after herbivory by gypsy moth (Lymantria dispar) caterpillars and determined the compounds mediating the attraction of the gypsy moth parasitoid Glyptapanteles liparidis (Braconidae). Female parasitoids were more attracted to gypsy moth-damaged leaves than to adjacent non-damaged leaves. The most characteristic volatiles of damaged versus neighbouring undamaged leaves included terpenes, green leaf volatiles and nitrogen-containing compounds, such as aldoximes and nitriles. Electrophysiological recordings and olfactometer bioassays demonstrated the importance of nitrogenous volatiles. Under field conditions, parasitic Hymenoptera were more attracted to traps baited with these substances than most other compounds. The differences in volatile emission profiles between damaged and undamaged foliage appear to be regulated by jasmonate signalling and the local activation of volatile biosynthesis. We conclude that characteristic volatiles from damaged black poplar foliage are essential cues enabling parasitoids to find their hosts.