Phytochemical drivers of insect herbivory: a functional toolbox to support agroecological diversification.

Plant metabolism is a key feature of biodiversity that remains underexploited in functional frameworks used in agroecology. Here, we study how phytochemical diversity considered at three organizational levels can promote pest control. In a factorial field experiment, we manipulated plant diversity in three monocultures and three mixed crops of oilseed rape to explore how intra- and interspecific phytochemical diversity affects pest infestation. We combined recent progress in metabolomics with classic metrics used in ecology to test a box of hypotheses grounded in plant defence theory. According to the hypothesis of 'phytochemically mediated coevolution', our study stresses the relationships between herbivore infestation and particular classes of specialized metabolites like glucosinolates. Among 178 significant relationships between metabolites and herbivory rates, only 20% were negative. At the plant level, phytochemical abundance and richness had poor predictive power on pest regulation. This challenges the hypothesis of 'synergistic effects'. At the crop cover level, in line with the hypothesis of 'associational resistance', the phytochemical dissimilarity between neighbouring plants limited pest infestation. We discuss the intricate links between associational resistance and bottom-up pest control. Bridging different levels of organization in agroecosystems helps to dissect the multi-scale relationships between phytochemistry and insect herbivory.

Bumble bees damage plant leaves and accelerate flower production when pollen is scarce.

Maintaining phenological synchrony with flowers is a key ecological challenge for pollinators that may be exacerbated by ongoing environmental change. Here, we show that bumble bee workers facing pollen scarcity damage leaves of flowerless plants and thereby accelerate flower production. Laboratory studies revealed that leaf-damaging behavior is strongly influenced by pollen availability and that bee-damaged plants flower significantly earlier than undamaged or mechanically damaged controls. Subsequent outdoor experiments showed that the intensity of damage inflicted varies with local flower availability; furthermore, workers from wild colonies of two additional bumble bee species were also observed to damage plant leaves. These findings elucidate a feature of bumble bee worker behavior that can influence the local availability of floral resources.

Plant volatiles induced by herbivore eggs prime defences and mediate shifts in the reproductive strategy of receiving plants.

Plants can detect cues associated with the risk of future herbivory and modify defence phenotypes accordingly; however, our current understanding is limited both with respect to the range of early warning cues to which plants respond and the nature of the responses. Here we report that exposure to volatile emissions from plant tissues infested with herbivore eggs promotes stronger defence responses to subsequent herbivory in two Brassica species. Furthermore, exposure to these volatile cues elicited an apparent shift from growth to reproduction in Brassica nigra, with exposed plants exhibiting increased flower and seed production, but reduced leaf production, relative to unexposed controls. Our results thus document plant defence priming in response to a novel environmental cue, oviposition-induced plant volatiles, while also showing that plant responses to early warning cues can include changes in both defence and life-history traits.

Plant responses to butterfly oviposition partly explain preference-performance relationships on different brassicaceous species.

The preference-performance hypothesis (PPH) states that herbivorous female insects prefer to oviposit on those host plants that are best for their offspring. Yet, past attempts to show the adaptiveness of host selection decisions by herbivores often failed. Here, we tested the PPH by including often neglected oviposition-induced plant responses, and how they may affect both egg survival and larval weight. We used seven Brassicaceae species of which most are common hosts of two cabbage white butterfly species, the solitary Pieris rapae and gregarious P. brassicae. Brassicaceous species can respond to Pieris eggs with leaf necrosis, which can lower egg survival. Moreover, plant-mediated responses to eggs can affect larval performance. We show a positive correlation between P. brassicae preference and performance only when including the egg phase: 7-day-old caterpillars gained higher weight on those plant species which had received most eggs. Pieris eggs frequently induced necrosis in the tested plant species. Survival of clustered P. brassicae eggs was unaffected by the necrosis in most tested species and no relationship between P. brassicae egg survival and oviposition preference was found. Pieris rapae preferred to oviposit on plant species most frequently expressing necrosis although egg survival was lower on those plants. In contrast to the lower egg survival on plants expressing necrosis, larval biomass on these plants was higher than on plants without a necrosis. We conclude that egg survival is not a crucial factor for oviposition choices but rather egg-mediated responses affecting larval performance explained the preference-performance relationship of the two butterfly species.

Divergence in Glucosinolate Profiles between High- and Low-Elevation Populations of Arabidopsis halleri Correspond to Variation in Field Herbivory and Herbivore Behavioral Preferences.

Variation in local herbivore pressure along elevation gradients is predicted to drive variation in plant defense traits. Yet, the extent of intraspecific variation in defense investment along elevation gradients, and its effects on both herbivore preference and performance, remain relatively unexplored. Using populations of Arabidopsis halleri (Brassicaceae) occurring at different elevations in the Alps, we tested for associations between elevation, herbivore damage in the field, and constitutive chemical defense traits (glucosinolates) assayed under common-garden conditions. Additionally, we examined the feeding preferences and performance of a specialist herbivore, the butterfly Pieris brassicae, on plants from different elevations in the Alps. Although we found no effect of elevation on the overall levels of constitutive glucosinolates in leaves, relative amounts of indole glucosinolates increased significantly with elevation and were negatively correlated with herbivore damage in the field. In oviposition preference assays, P. brassicae females laid fewer eggs on plants from high-elevation populations, although larval performance was similar on populations from different elevations. Taken together, these results support the prediction that species distributed along elevation gradients exhibit genetic variation in chemical defenses, which can have consequences for interactions with herbivores in the field.

Early herbivore alert matters: plant-mediated effects of egg deposition on higher trophic levels benefit plant fitness.

Induction of plant defences, specifically in response to herbivore attack, can save costs that would otherwise be needed to maintain defences even in the absence of herbivores. However, plants may suffer considerable damage during the time required to mount these defences against an attacker. This could be resolved if plants could respond to early cues, such as egg deposition, that reliably indicate future herbivory. We tested this hypothesis in a field experiment and found that egg deposition by the butterfly Pieris brassicae on black mustard (Brassica nigra) induced a plant response that negatively affected feeding caterpillars. The effect cascaded up to the third and fourth trophic levels (larval parasitoids and hyperparasitoids) by affecting the parasitisation rate and parasitoid performance. Overall, the defences induced by egg deposition had a positive effect on plant seed production and may therefore play an important role in the evolution of plant resistance to herbivores.

To be in time: egg deposition enhances plant-mediated detection of young caterpillars by parasitoids.

Animals use information from their environment while foraging for food or prey. When parasitic wasps forage for hosts, they use plant volatiles induced by herbivore activities such as feeding and oviposition. Little information is available on how wasps exploit specific plant volatiles over time, and which compounds indicate changes in host quality. In experiments investigating the role of herbivore-induced plant volatiles in wasp foraging, induction of plant response is usually achieved by placing larvae on clean plants instead of allowing the natural sequence of events: to let eggs deposited by the herbivore develop into larvae. We compared the attraction of the parasitoid Cotesia glomerata to volatiles emitted by black mustard (Brassica nigra) plants induced by eggs and successive larval stages of the Large Cabbage White butterfly (Pieris brassicae) to the attraction of this parasitoid to black mustard plant volatiles induced only by larval feeding in a wind tunnel setup. We show that wasps are attracted to plants infested with eggs just before and shortly after larval hatching. However, wasp preference changed at later time points towards plants induced only by larval feeding. These temporal changes in parasitoid attraction matched with changes in the chemical compositions of the blends of plant volatiles. Previous studies have shown that host quality/suitability decreases with caterpillar age and that P. brassicae oviposition induces plant defences that negatively affect subsequently feeding caterpillars. We investigated parasitoid performance in hosts of different ages. Wasp performance was positively correlated with preference. Moreover, parasitism success decreased with time and host stage. In conclusion, the behaviour of Cotesia glomerata is fine-tuned to exploit volatiles induced by eggs and early host stages that benefit parasitoid fitness.

Phenotypic plasticity of plant response to herbivore eggs: effects on resistance to caterpillars and plant development.

Herbivory induces direct resistance responses in plants that negatively affect subsequently colonizing herbivores. Moreover, eggs of herbivorous insects can also activate plant resistance, which in some cases prevents hatching larvae from feeding. Until now, plant-mediated effects of eggs on subsequent herbivory, and the specificity of such responses, have remained poorly understood. We studied the specificity and effects of plant resistance induced by herbivore egg deposition against lepidopteran larvae of species with different dietary breadths, feeding on a wild annual plant, the crucifer Brassica nigra. We examined whether this plant-mediated response affects the growth of caterpillars of a specialist (Pieris brassicae) that feeds on B. nigra leaves and flowers, and a generalist (Mamestra brassicae) that rarely attacks this wild crucifer. We measured growth rates of neonate larvae to the end of their second instar after the larvae had hatched on plants exposed to eggs vs. plants without eggs, under laboratory and semi-field conditions. Moreover, we studied the effects of egg deposition by the two herbivore species on plant height and flowering rate before and after larval hatching. Larvae of both herbivore species that developed on plants previously infested with eggs of the specialist butterfly P. brassicae gained less mass compared with larvae that developed on egg-free plants. Plants exposed to butterfly eggs showed accelerated plant growth and flowering compared to egg-free plants. Egg deposition by the generalist moth M. brassicae, in contrast, had no effect on subsequent performance by either herbivore species, or on plant development. Our results demonstrate that B. nigra plants respond differently to eggs of two herbivore species in terms of plant development and induced resistance to caterpillar attack. For this annual crucifer, the retardation of caterpillar growth in response to deposition of eggs by P. brassicae in combination with enhanced growth and flowering likely result in reproductive assurance, after being exposed to eggs from an herbivore whose larvae rapidly reduce the plant's reproductive potential through florivory.

Reward value determines memory consolidation in parasitic wasps.

Animals can store learned information in their brains through a series of distinct memory forms. Short-lasting memory forms can be followed by longer-lasting, consolidated memory forms. However, the factors determining variation in memory consolidation encountered in nature have thus far not been fully elucidated. Here, we show that two parasitic wasp species belonging to different families, Cotesia glomerata (Hymenoptera: Braconidae) and Trichogramma evanescens (Hymenoptera; Trichogrammatidae), similarly adjust the memory form they consolidate to a fitness-determining reward: egg-laying into a host-insect that serves as food for their offspring. Protein synthesis-dependent long-term memory (LTM) was consolidated after single-trial conditioning with a high-value host. However, single-trial conditioning with a low-value host induced consolidation of a shorter-lasting memory form. For Cotesia glomerata, we subsequently identified this shorter-lasting memory form as anesthesia-resistant memory (ARM) because it was not sensitive to protein synthesis inhibitors or anesthesia. Associative conditioning using a single reward of different value thus induced a physiologically different mechanism of memory formation in this species. We conclude that the memory form that is consolidated does not only change in response to relatively large differences in conditioning, such as the number and type of conditioning trials, but is also sensitive to more subtle differences, such as reward value. Reward-dependent consolidation of exclusive ARM or LTM provides excellent opportunities for within-species comparison of mechanisms underlying memory consolidation.

Plant volatiles induced by herbivore egg deposition affect insects of different trophic levels.

Plants release volatiles induced by herbivore feeding that may affect the diversity and composition of plant-associated arthropod communities. However, the specificity and role of plant volatiles induced during the early phase of attack, i.e. egg deposition by herbivorous insects, and their consequences on insects of different trophic levels remain poorly explored. In olfactometer and wind tunnel set-ups, we investigated behavioural responses of a specialist cabbage butterfly (Pieris brassicae) and two of its parasitic wasps (Trichogramma brassicae and Cotesia glomerata) to volatiles of a wild crucifer (Brassica nigra) induced by oviposition of the specialist butterfly and an additional generalist moth (Mamestra brassicae). Gravid butterflies were repelled by volatiles from plants induced by cabbage white butterfly eggs, probably as a means of avoiding competition, whereas both parasitic wasp species were attracted. In contrast, volatiles from plants induced by eggs of the generalist moth did neither repel nor attract any of the tested community members. Analysis of the plant's volatile metabolomic profile by gas chromatography-mass spectrometry and the structure of the plant-egg interface by scanning electron microscopy confirmed that the plant responds differently to egg deposition by the two lepidopteran species. Our findings imply that prior to actual feeding damage, egg deposition can induce specific plant responses that significantly influence various members of higher trophic levels.

Anti-aphrodisiac compounds of male butterflies increase the risk of egg parasitoid attack by inducing plant synomone production.

During mating in many butterfly species, males transfer spermatophores that contain anti-aphrodisiacs to females that repel conspecific males. For example, males of the large cabbage white, Pieris brassicae (Lepidoptera: Pieridae), transfer the anti-aphrodisiac, benzyl cyanide (BC) to females. Accessory reproductive gland (ARG) secretion of a mated female P. brassicae that is deposited with an egg clutch contains traces of BC, inducing Brussels sprouts plants (Brassica oleracea var. gemmifera) to arrest certain Trichogramma egg parasitoids. Here, we assessed whether deposition of one egg at a time by the closely related small cabbage white, Pieris rapae, induced B. oleracea var. gemmifera to arrest Trichogramma wasps, and whether this plant synomone is triggered by substances originating from male P. rapae seminal fluid. We showed that plants induced by singly laid eggs of P. rapae arrest T. brassicae wasps three days after butterfly egg deposition. Elicitor activity was present in ARG secretion of mated female butterflies, whereas the secretion of virgin females was inactive. Pieris rapae used a mixture of methyl salicylate (MeSA) and indole as an anti-aphrodisiac. We detected traces of both anti-aphrodisiacal compounds in the ARG secretion of mated female P. rapae, whereas indole was lacking in the secretion of virgin female P. rapae. When applied onto the leaf, indole induced changes in the foliar chemistry that arrested T. brassicae wasps. This study shows that compounds of male seminal fluid incur possible fitness costs for Pieris butterflies by indirectly promoting egg parasitoid attack.

Hitch-hiking parasitic wasp learns to exploit butterfly antiaphrodisiac.

Many insects possess a sexual communication system that is vulnerable to chemical espionage by parasitic wasps. We recently discovered that a hitch-hiking (H) egg parasitoid exploits the antiaphrodisiac pheromone benzyl cyanide (BC) of the Large Cabbage White butterfly Pieris brassicae. This pheromone is passed from male butterflies to females during mating to render them less attractive to conspecific males. When the tiny parasitic wasp Trichogramma brassicae detects the antiaphrodisiac, it rides on a mated female butterfly to a host plant and then parasitizes her freshly laid eggs. The present study demonstrates that a closely related generalist wasp, Trichogramma evanescens, exploits BC in a similar way, but only after learning. Interestingly, the wasp learns to associate an H response to the odors of a mated female P. brassicae butterfly with reinforcement by parasitizing freshly laid butterfly eggs. Behavioral assays, before which we specifically inhibited long-term memory (LTM) formation with a translation inhibitor, reveal that the wasp has formed protein synthesis-dependent LTM at 24 h after learning. To our knowledge, the combination of associatively learning to exploit the sexual communication system of a host and the formation of protein synthesis-dependent LTM after a single learning event has not been documented before. We expect it to be widespread in nature, because it is highly adaptive in many species of egg parasitoids. Our finding of the exploitation of an antiaphrodisiac by multiple species of parasitic wasps suggests its use by Pieris butterflies to be under strong selective pressure.

Influence of grain type on the insecticidal efficacy of two diatomaceous earth formulations against Rhyzopertha dominica (F) (Coleoptera: Bostrychidae).

Laboratory bioassays were carried out in order to evaluate the effectiveness of two diatomaceous earth (DE) formulations, Insecto and SilicoSec, against adults of Rhyzopertha dominica (F) in eight different grain commodities. The adherence of the two DEs to each grain was also measured. The eight grains tested were wheat, whole barley, peeled barley, oats, rye, triticale, rice and maize. These commodities were treated with the DEs at three rates, 0.75, 1.0 and 1.5 g DE kg(-1) grain. The mortality of R dominica adults was measured after 24 h, 48 h, 7 days and 14 days of exposure in the treated grains at 26 degrees C and 55% RH. After the 14-day mortality count, all adults were removed and the treated grains retained under the same conditions for a further 60 days. The treated grains were subsequently examined for F1 progeny. Significant differences were recorded among the eight grain types as well as between the DE formulations tested. After 14 days of exposure, even at the lowest DE rate for both formulations, adult mortality was high (>90%) in wheat and triticale. In contrast, adult mortality was significantly lower in peeled barley. Increasing the rate improved the efficacy of the DEs significantly in only some of the grains tested. Reproductive capacity in all the treated grains was significantly suppressed when compared with untreated grains. Generally, more F1 individuals of R dominica were noted in the treated peeled barley than in the other commodities. Significant differences in the percentage of DE retention were noted among the eight grains. The highest retention level was noted in rice (>87%) and the lowest in maize (<6%). However, the degree of DE adherence to a given grain was not always indicative of the effectiveness of DE against R dominica.