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The intricate relationship between herbivorous insects and plants has evolved over millions of years, central to this dynamic interaction are salivary proteins (SPs), which mediate key processes ranging from nutrient acquisition to plant defense manipulation. SPs, sourced from salivary glands, intestinal regurgitation or acquired through horizontal gene transfer, exhibit remarkable functional versatility, influencing insect development, behavior, and adhesion mechanisms. Moreover, SPs play pivotal roles in modulating plant defenses, to induce or inhibit plant defenses as elicitors or effectors. In this review, we delve into the multifaceted roles of SPs in herbivorous insects, highlighting their diverse impacts on insect physiology and plant responses. Through a comprehensive exploration of SP functions, this review aims to deepen our understanding of plant-insect interactions and foster advancements in both fundamental research and practical applications in plant-insect interactions.

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Herbivorous insects can identify their host plants by sensing plant secondary metabolites as chemical cues. We previously reported the two-factor host acceptance system of the silkworm Bombyx mori larvae. The chemosensory neurons in the maxillary palp (MP) of the larvae detect mulberry secondary metabolites, chlorogenic acid (CGA), and isoquercetin (ISQ), with ultrahigh sensitivity, for host plant recognition and feeding initiation. Nevertheless, the molecular basis for the ultrasensitive sensing of these compounds remains unknown. In this study, we demonstrated that two gustatory receptors (Grs), BmGr6 and BmGr9, are responsible for sensing the mulberry compounds with attomolar sensitivity for host plant recognition by silkworm larvae. Calcium imaging assay using cultured cells expressing the silkworm putative sugar receptors (BmGr4-10) revealed that BmGr6 and BmGr9 serve as receptors for CGA and ISQ with attomolar sensitivity in human embryonic kidney 293T cells. CRISPR/Cas9-mediated knockout (KO) of BmGr6 and BmGr9 resulted in a low probability of making a test bite of the mulberry leaves, suggesting that they lost the ability to recognize host leaves. Electrophysiological recordings showed that the loss of host recognition ability in the Gr-KO strains was due to a drastic decrease in MP sensitivity toward ISQ in BmGr6-KO larvae and toward CGA and ISQ in BmGr9-KO larvae. Our findings have revealed that the two Grs, previously considered to be sugar receptors, are molecules responsible for detecting plant phenolics in host plant recognition.

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Recent studies on insect interactions on plants have revealed that herbivorous insects indirectly interact with each other through changes in plant traits following herbivory. However, less attention has been given to plant biomass relative to plant quality in relation to indirect interactions among herbivores. We explored the extent to which the larval food demand of two specialist butterflies (Sericinus montela and Atrophaneura alcinous) explains their interaction on a host plant, Aristolochia debilis. A laboratory experiment showed that plant mass consumption by A. alcinous larvae was 2.6 times greater than that by S. montela. We predicted that A. alcinous, which requires more food, is more vulnerable to food shortages than S. montela. In a cage experiment, an asymmetric interspecific interaction was detected between the two specialist butterflies; S. montela larval density significantly decreased the survival and prolonged the development time of A. alcinous, but A. alcinous density affected neither the survival nor the development time of S. montela. The prediction based on the food requirement was partly supported by the fact that increasing A. alcinous density likely caused a food shortage, which more negatively affected A. alcinous survival than S. montela survival. Conversely, increasing the density of S. montela did not reduce the remaining food quantity, suggesting that the negative effect of S. montela density on A. alcinous was unlikely to be due to food shortage. Although aristolochic acid I, a defensive chemical specific to Aristolochia plants, did not influence the food consumption or growth of either butterfly larva, unmeasured attributes of plant quality may have mediated an indirect interaction between the two butterflies. Consequently, our study suggests that not only the quality but also the quantity of plants should be considered to fully understand the characteristics, such as symmetry, of interspecific interactions among herbivorous insects on the same host plant.

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In the field, plants usually have to face the combined effects of abiotic and biotic stresses. In our study, two spring wheat cultivars-Septima and Quintus-were subjected to three water regimes [70%, 50%, and 40% soil water capacity (SWC)], aphid (Metopolophium dirhodum) infestation, or the combination of both stresses, i.e., water deficit (50%, 40% SWC) and aphids. The study has a 2 × 3 × 2 factorial design with three biological replicates. In the present study, the results of proteomic analysis using 2D-DIGE followed by MALDI-TOF/TOF protein identification are presented. Water deficit but also aphid infestation led to alterations in 113 protein spots including proteins assigned to a variety of biological processes ranging from signaling via energy metabolism, redox regulation, and stress and defense responses to secondary metabolism indicating a long-term adaptation to adverse conditions. The absence of specific proteins involved in plant response to herbivorous insects indicates a loss of resistance to aphids in modern wheat cultivars during the breeding process and is in accordance with the "plant vigor hypothesis." Septima revealed enhanced tolerance with respect to Quintus as indicated by higher values of morphophysiological characteristics (fresh aboveground biomass, leaf length, osmotic potential per full water saturation) and relative abundance of proteins involved in mitochondrial respiration and ATP biosynthesis.

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Horizontal gene transfer (HGT) provides an evolutionary shortcut for recipient organisms to gain novel functions. Although reports of HGT in higher eukaryotes are rapidly accumulating, in most cases the evolutionary trajectory, metabolic integration, and ecological relevance of acquired genes remain unclear. Plant cell wall degradation by HGT-derived enzymes is widespread in herbivorous insect lineages. Pectin is an abundant polysaccharide in the walls of growing parts of plants. We investigated the significance of horizontally acquired pectin-digesting polygalacturonases (PGs) of the leaf beetle Phaedon cochleariae. Using a CRISPR/Cas9-guided gene knockout approach, we generated a triple knockout and a quadruple PG-null mutant in order to investigate the enzymatic, biological, and ecological effects. We found that pectin-digestion 1) is exclusively linked to the horizontally acquired PGs from fungi, 2) became fixed in the host genome by gene duplication leading to functional redundancy, 3) compensates for nutrient-poor diet by making the nutritious cell contents more accessible, and 4) facilitates the beetles development and survival. Our analysis highlights the selective advantage PGs provide to herbivorous insects and demonstrate the impact of HGT on the evolutionary success of leaf-feeding beetles, major contributors to species diversity.

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The immunotoxicity induced by heavy metals on herbivorous insects reflect the alterations of the susceptibility to entomopathogenic agents in herbivorous insects exposed to heavy metal. In the present study, the susceptibility of gypsy moth larvae to Bacillus thuringiensis under Cd treatment at low and high dosages was investigated, and the gut microbiome-hemolymph metabolome responses that affected larval disease susceptibility caused by Cd exposure were examined. Our results showed that mortality of gypsy moth larvae caused by B. thuringiensis was significantly higher in larvae pre-exposed to Cd stress, and there was a synergistic effect between Cd pre-exposure and bacterial infection. Exposure to Cd significantly decreased the abundance of several probiotics (e.g., Serratia for the low Cd dosage and Weissella, Aeroonas, and Serratia for the high Cd dosage) and increased the abundances of several pathogenic bacteria (e.g., Stenotrophomonas, Gardnerella, and Cutibacterium for the low Cd dosage and Pluralibacter and Tsukamurella for the high Cd dosage) compared to the controls. Moreover, metabolomics analysis indicated that amino acid biosynthesis and metabolism were significantly perturbed in larval hemolymph under Cd exposure at both the low and high dosages. Correlation analysis demonstrated that several altered metabolites in larval hemolymph were significantly correlated with changes in the gut microbial community. The results demonstrate that prior exposure to Cd increases the susceptibility of gypsy moth larvae to B. thuringiensis in a synergistic fashion due to gut microbiota dysbiosis and hemolymph metabolic disorder, and thus microbial-based biological control may be the best pest control strategy in heavy metal-polluted areas.

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Plant sphingolipids are important membrane components and bioactive molecules in development and defense responses. However, the function of sphingolipids in plant defense, especially against herbivores, is not fully understood. Here, we report that Spodoptera exigua feeding affects sphingolipid metabolism in Arabidopsis, resulting in increased levels of sphingoid long-chain bases, ceramides, and hydroxyceramides. Insect-induced ceramide and hydroxyceramide accumulation is dependent on the jasmonate signaling pathway. Loss of the Arabidopsis alkaline ceramidase ACER increases ceramides and decreases long-chain base levels in plants; in this work, we found that loss of ACER enhances plant resistance to S. exigua and improves response to mechanical wounding. Moreover, acer-1 mutants exhibited more severe root-growth inhibition and higher anthocyanin accumulation than wild-type plants in response to methyl jasmonate treatment, indicating that loss of ACER increases sensitivity to jasmonate and that ACER functions in jasmonate-mediated root growth and secondary metabolism. Transcript levels of ACER were also negatively regulated by jasmonates, and this process involves the transcription factor MYC2. Thus, our findings reveal that ACER is involved in mediating jasmonate-related plant growth and defense and that jasmonates function in regulating the expression of ACER.

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This study investigated whether lead (Pb), at concentrations allowed for soil, affects the community of insects that live in the aerial part of plants. We evaluated the effect of Pb concentrations on accumulated species richness, composition, and abundance of different functional groups of insects. Kale plants were grown in soil experimentally contaminated with four concentrations of lead nitrate: 0 (control), 144 (T1), 360 (T2), and 600 (T3) mg/kg of soil. The experiment was conducted in an open greenhouse for the natural colonization of insects. Insects were collected twice using trap bags attached to the plant leaf and by direct removal. The concentration of Pb in the stem and leaf samples increased with the increased soil contamination, even showing values above the limit allowed by the legislation for this plant species. Control plants showed a higher richness of accumulated insect species. In addition, the treatments had an effect on the community composition, in which Diaeretiella rapae (primary parasitoid) was found as an indicator of the control + T1 treatments and the top species Pachyneuron sp. (parasitoid of predators) was associated with the control. The abundance of chewing and sucking herbivores, their respective parasitoids, predators, and parasitoids of predators were negatively affected. Hyperparasitoid abundance was not affected, but their accumulated species richness was. This study was innovative in demonstrating that soil contamination by different concentrations of a heavy metal (Pb) can negatively affect the community of plant-associated insects, even at concentrations allowed for soil, reflecting possible damage to the ecosystem.

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The study of herbivorous insects underpins much of the theory that concerns the evolution of species interactions. In particular, Pieridae butterflies and their host plants have served as a model system for studying evolutionary arms races. To learn more about the coevolution of these two clades, we reconstructed ancestral ecological networks using stochastic mappings that were generated by a phylogenetic model of host-repertoire evolution. We then measured if, when, and how two ecologically important structural features of the ancestral networks (modularity and nestedness) evolved over time. Our study shows that as pierids gained new hosts and formed new modules, a subset of them retained or recolonised the ancestral host(s), preserving connectivity to the original modules. Together, host-range expansions and recolonisations promoted a phase transition in network structure. Our results demonstrate the power of combining network analysis with Bayesian inference of host-repertoire evolution to understand changes in complex species interactions over time.

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Herbivore-induced plant volatiles play important ecological roles in defense against stresses. However, if and which volatile(s) are involved in the plant-plant communication in response to herbivorous insects in tea plants remains unknown. Here, plant-plant communication experiments confirm that volatiles emitted from insects-attacked tea plants can trigger plant resistance and reduce the risk of herbivore damage by inducing jasmonic acid (JA) accumulation in neighboring plants. The emission of six compounds was significantly induced by geometrid Ectropis obliqua, one of the most common pests of the tea plant in China. Among them, (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT) could induce the accumulation of JA and thus promotes the resistance of neighboring intact plants to herbivorous insects. CsCYP82D47 was identified for the first time as a P450 enzyme, which catalyzes the final step in the biosynthesis of DMNT from (E)-nerolidol. Down-regulation of CsCYP82D47 in tea plants resulted in a reduced accumulation of DMNT and significantly reduced the release of DMNT in response to the feeding of herbivorous insects. The first evidence for plant-plant communication in response to herbivores in tea plants will help to understand how plants respond to volatile cues in response to herbivores and provide new insight into the role(s) of DMNT in tea plants.

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Cadmium as a common environmental stressor may exert highly toxic effects on herbivorous insects. The question was whether possible elevation of an oxidative stress and imbalance of energetic reserves in insects may depend on developmental stage, sex and insect population's multigenerational history of exposure to cadmium. So, the aim of this study was to compare of the development traits, total antioxidant capacity, lipid peroxidation, RSSR to RSH ratio and the concentration of carbohydrates, glycogen, lipids and proteins in whole individuals (larvae or pupae) of Spodoptera exigua originating from two strains: control and selected over 120 generations with sublethal metal concentration (44 Cd mg per dry weight of diet). Generally, the increase of the protein, carbohydrates, glycogen concentration and lipid peroxidation decrease with age of the larvae were found. Revealed cases of a higher mobilisation of carbohydrates and proteins, and changes in total antioxidant capacity or lipid peroxidation, in individuals being under metal exposure, occurred in strain-depended mode. Short-term Cd exposure effect was connected with possible higher engagement of proteins and glycogen in detoxification processes, but also higher concentration of lipid peroxidation. In turn, for long-term Cd exposure effect lower lipids concentration and higher thiols usage seemed to be more specific.

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Heavy metal pollution is one of the thorniest issues in the world, which is a serious threat to ecosystems and food security. As an important link of food chain and food web in the ecosystem, herbivorous insects play important role in the transfer and accumulation of heavy metals. Consequently, more and more attentions have been paid on the potential effects of heavy metal pollution on herbivorous insects. Here, we reviewed the effects of heavy metal pollution on herbivorous insects with literature publshied during 2007 to 2018. Herbivorous insects were exposed to heavy metals pollution via four ways, including insect diets or food added with inorganic heavy metals, field exposure with heavy metals, "soil/substrate-plants-herbivorous insect" food chain and in vitro injection. Excessive accumulation of heavy metals in insect body results in decreased survival rate, reproductive capacity and population growth, the retarded growth and development. Physiological and biochemical toxicity of heavy metal pollution to herbivorous insects included cell ultra-structure destruction and DNA damage, decreased amount of energy materials, and changes in enzymes acti-vity and gene expression levels. However, herbivorous insects can resist heavy metal stress by producing metallothionein and enhancing activity of detoxification enzymes etc., which probably results in their adaptive evolution to heavy metal stress at low intensity or long-term and even improve their tolerance to other stresses (e.g., pesticides).

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Compared with the highly diverse microbiota of leaves, herbivorous insects exhibit impoverished gut microbial communities. Research to date has focused on the bacterial component of these gut microbiomes, neglecting the fungal component. As caterpillar gut bacterial microbiomes are derived strongly from their diet, we hypothesized that their mycobiomes would reflect the host leaf mycobiomes. Using the ITS2 rDNA and V5-V6 16S rRNA gene regions for DNA metabarcoding of caterpillar gut and host leaf sample pairs we compared their mycobiome genus diversity and compositions and identified genera associated with caterpillar guts. Leaves and caterpillar guts harbored different mycobiomes with quite low qualitative similarity (Jaccard index = 38.03%). The fungal genera most significantly associated with the caterpillar gut included Penicillium, Mucor and unidentified Saccharomycetales, whereas leaf-associated genera included Holtermanniella, Gibberella (teleomorph of Fusarium) and Seimatosporium. Although caterpillar gut and leaf mycobiomes had similar genus richness overall, this indicator was not correlated for individual duplets. Moreover, as more samples entered the analysis, mycobiome richness increased more rapidly in caterpillar guts than in leaves. The results suggest that the mycobiota of the caterpillar gut differs from that of their feeding substrate; further, the mycobiomes appear to be richer than the well-studied bacterial microbiotas.

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Seminatural grasslands are important biodiversity hotspots, but they are increasingly degraded by intensive agriculture. Grassland restoration is considered to be promising in halting the ongoing loss of biodiversity, but this evaluation is mostly based on plant communities. Insect herbivores contribute substantially to grassland biodiversity and to the provisioning of a variety of ecosystem functions. However, it is unclear how they respond to different measures that are commonly used to restore seminatural grasslands from intensively used agricultural land. We studied the long-term success of different restoration techniques, which were originally targeted at reestablishing seminatural grassland plant communities, for herbivorous insect communities on taxonomic as well as functional level. Therefore, we sampled insect communities 22 yr after the establishment of restoration measures. These measures ranged from harvest and removal of biomass to removal of the topsoil layer and subsequent seeding of plant propagules. We found that insect communities in restored grasslands had higher taxonomic and functional diversity compared to intensively managed agricultural grasslands and were more similar in composition to target grasslands. Restoration measures including topsoil removal proved to be more effective, in particular in restoring species characterized by functional traits susceptible to intensive agriculture (e.g., large-bodied species). Our study shows that long-term success in the restoration of herbivorous insect communities of seminatural grasslands can be achieved by different restoration measures and that more invasive approaches that involve the removal of the topsoil layer are more effective. We attribute these restoration successes to accompanying changes in the plant community, resulting in bottom-up control of the herbivore community. Our results are of critical importance for management decisions aiming to restore multi-trophic communities, their functional composition and consequently the proliferation of ecosystem functions.

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Increased frequency and severity of drought, as a result of climate change, is expected to drive critical changes in plant-insect interactions that may elevate rates of tree mortality. The mechanisms that link water stress in plants to insect performance are not well understood. Here, we build on previous reviews and develop a framework that incorporates the severity and longevity of drought and captures the plant physiological adjustments that follow moderate and severe drought. Using this framework, we investigate in greater depth how insect performance responds to increasing drought severity for: (i) different feeding guilds; (ii) flush feeders and senescence feeders; (iii) specialist and generalist insect herbivores; and (iv) temperate versus tropical forest communities. We outline how intermittent and moderate drought can result in increases of carbon-based and nitrogen-based chemical defences, whereas long and severe drought events can result in decreases in plant secondary defence compounds. We predict that different herbivore feeding guilds will show different but predictable responses to drought events, with most feeding guilds being negatively affected by water stress, with the exception of wood borers and bark beetles during severe drought and sap-sucking insects and leaf miners during moderate and intermittent drought. Time of feeding and host specificity are important considerations. Some insects, regardless of feeding guild, prefer to feed on younger tissues from leaf flush, whereas others are adapted to feed on senescing tissues of severely stressed trees. We argue that moderate water stress could benefit specialist insect herbivores, while generalists might prefer severe drought conditions. Current evidence suggests that insect outbreaks are shorter and more spatially restricted in tropical than in temperate forests. We suggest that future research on the impact of drought on insect communities should include (i) assessing how drought-induced changes in various plant traits, such as secondary compound concentrations and leaf water potential, affect herbivores; (ii) food web implications for other insects and those that feed on them; and (iii) interactions between the effects on insects of increasing drought and other forms of environmental change including rising temperatures and CO2 levels. There is a need for larger, temperate and tropical forest-scale drought experiments to look at herbivorous insect responses and their role in tree death.

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The enemy-release hypothesis is one of the most popular but also most discussed hypotheses to explain invasion success. However, there is a lack of explicit, experimental tests of predictions of the enemy-release hypothesis (ERH), particularly regarding the effects of above- and belowground herbivory. Long-term studies investigating the relative effect of herbivores on invasive vs. native plant species within a community are still lacking. Here, we report on a long-term field experiment in an old-field community, invaded by Solidago canadensis s. l., with exclusion of above- and belowground insect herbivores. We monitored population dynamics of the invader and changes in the diversity and functioning of the plant community across eight years. Above- and belowground insects favoured the establishment of the invasive plant species and thereby increased biomass and decreased diversity of the plant community. Effects of invertebrate herbivores on population dynamics of S. canadensis appeared after six years and increased over time, suggesting that long-term studies are needed to understand invasion dynamics and consequences for plant community structure. We suggest that the release from co-evolved trophic linkages is of importance not only for the effect of invasive species on ecosystems, but also for the functioning of novel species assemblages arising from climate change.

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Under natural conditions, aboveground herbivory and plant-soil feedbacks (PSFs) are omnipresent interactions strongly affecting individual plant performance. While recent research revealed that aboveground insect herbivory generally impacts the outcome of PSFs, no study tested to what extent the intensity of herbivory affects the outcome. This, however, is essential to estimate the contribution of PSFs to plant performance under natural conditions in the field. Here, we tested PSF effects both with and without exposure to aboveground herbivory for four common grass species in nine grasslands that formed a gradient of aboveground invertebrate herbivory. Without aboveground herbivores, PSFs for each of the four grass species were similar in each of the nine grasslands-both in direction and in magnitude. In the presence of herbivores, however, the PSFs differed from those measured under herbivory exclusion, and depended on the intensity of herbivory. At low levels of herbivory, PSFs were similar in the presence and absence of herbivores, but differed at high herbivory levels. While PSFs without herbivores remained similar along the gradient of herbivory intensity, increasing herbivory intensity mostly resulted in neutral PSFs in the presence of herbivores. This suggests that the relative importance of PSFs for plant-species performance in grassland communities decreases with increasing intensity of herbivory. Hence, PSFs might be more important for plant performance in ecosystems with low herbivore pressure than in ecosystems with large impacts of insect herbivores.

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BACKGROUND: Much evolutionary theory predicts that diversity arises via both adaptive radiation (diversification driven by selection against niche-overlap within communities) and divergence of geographically isolated populations. We focus on tropical fruit flies (Blepharoneura, Tephritidae) that reveal unexpected patterns of niche-overlap within local communities. Throughout the Neotropics, multiple sympatric non-interbreeding populations often share the same highly specialized patterns of host use (e.g., flies are specialists on flowers of a single gender of a single species of host plants). Lineage through time (LTT) plots can help distinguish patterns of diversification consistent with ecologically limited adaptive radiation from those predicted by ecologically neutral theories. Here, we use a time-calibrated phylogeny of Blepharoneura to test the hypothesis that patterns of Blepharoneura diversification are consistent with an "ecologically neutral" model of diversification that predicts that diversification is primarily a function of time and space. RESULTS: The Blepharoneura phylogeny showed more cladogenic divergence associated with geography than with shifts in host-use. Shifts in host-use were associated with ~ 20% of recent splits (< 3 Ma), but > 60% of older splits (> 3 Ma). In the overall tree, gamma statistic and maximum likelihood model fitting showed no evidence of diversification rate changes though there was a weak signature of slowing diversification rate in one of the component clades. CONCLUSIONS: Overall patterns of Blepharoneura diversity are inconsistent with a traditional explanation of adaptive radiation involving decreases in diversification rates associated with niche-overlap. Sister lineages usually use the same host-species and host-parts, and multiple non-interbreeding sympatric populations regularly co-occur on the same hosts. We suggest that most lineage origins (phylogenetic splits) occur in allopatry, usually without shifts in host-use, and that subsequent dispersal results in assembly of communities composed of multiple sympatric non-interbreeding populations of flies that share the same hosts.

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Primary consumers are under strong selection from resource ('bottom-up') and consumer ('top-down') controls, but the relative importance of these selective forces is unknown. We performed a meta-analysis to compare the strength of top-down and bottom-up forces on consumer fitness, considering multiple predictors that can modulate these effects: diet breadth, feeding guild, habitat/environment, type of bottom-up effects, type of top-down effects and how consumer fitness effects are measured. We focused our analyses on the most diverse group of primary consumers, herbivorous insects, and found that in general top-down forces were stronger than bottom-up forces. Notably, chewing, sucking and gall-making herbivores were more affected by top-down than bottom-up forces, top-down forces were stronger than bottom-up in both natural and controlled (cultivated) environments, and parasitoids and predators had equally strong top-down effects on insect herbivores. Future studies should broaden the scope of focal consumers, particularly in understudied terrestrial systems, guilds, taxonomic groups and top-down controls (e.g. pathogens), and test for more complex indirect community interactions. Our results demonstrate the surprising strength of forces exerted by natural enemies on herbivorous insects, and thus the necessity of using a tri-trophic approach when studying insect-plant interactions.

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Woody encroachment in grasslands has become increasingly problematic globally. Grazing by domestic animals can facilitate woody encroachment by reducing competition from herbaceous plants and fire frequency. Herbivorous insects and parasitic plants can each exert forces that result in the natural biological control of encroaching woody plants through reducing seeding of their host woody plants. However, the interplay of grazing and dynamics of herbivorous insects or parasitic plants, and its effects on the potential biological control of woody encroachment in grasslands remains unclear. We investigated the flower and pod damage by herbivorous insects, and the infection rates of a parasitic plant on the shrub Caragana microphylla, which is currently encroaching in Inner Mongolia Steppe, under different grazing management treatments (33-year non-grazed, 7-year non-grazed, currently grazed). Our results showed that Caragana biomass was highest at the currently grazed site, and lowest at the 33-year non-grazed site. Herbaceous plant biomass followed the opposite pattern, suggesting that grazing is indeed facilitating the encroachment of Caragana plants in Inner Mongolia Steppe. Grazing also reduced the abundance of herbivorous insects per Caragana flower, numbers of flowers and pods damaged by insect herbivores, and the infection rates of the parasitic plant on Caragana plants. Our results suggest that grazing may facilitate woody encroachment in grasslands not only through canonical mechanisms (e.g. competitive release via feeding on grasses, reductions in fires, etc.), but also by limiting natural biological controls of woody plants (herbivorous insects and parasitic plants). Thus, management efforts must focus on preventing overgrazing to better protect grassland ecosystems from woody encroachment.