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Many herbivorous Lepidoptera accumulate plant toxins within their own tissues as a defensive strategy. Pioneering research in this area was conducted by Miriam Rothschild and Deane Bowers, who showed that the cycad-feeding butterfly Eumaeus atala sequester the toxic plant compound cycasin and thereby deter vertebrate and invertebrate predators. The current study focuses on another cycad compound, beta-methylamino-L-alanine (BMAA), that is known to accumulate in the tissues of insects and other herbivores, and which has been shown to have neurotoxic effects in humans. Chemical analyses revealed that BMAA accumulates in both immature and adult tissues of E. atala, as well as adult tissues of another cycad-feeding lepidopteran, Seirarctia echo. However, the distribution of BMAA across life stages and tissues did not conform to patterns predicted for defensive sequestration, and subsequent behavioral experiments with ants showed that these invertebrate predators were not deterred by BMAA. Our results suggest that high levels of BMAA in the tissues of cycad-feeding insects likely reflect passive bioaccumulation rather than defensive sequestration. Combined with the previous work by Rothschild and Bowers, these results provide an example in which two different plant toxins accumulate within the tissues of a single herbivore species via different mechanisms and with different implications for ecology and evolution. They thereby lay the groundwork for further investigation into the processes underlying active sequestration and non-adaptive bioaccumulation., Frontiers in Ecology and Evolution, 11, ISSN:2296-701X

Our understanding of how novel warning color traits evolve in natural populations is largely based on studies of reproductive stages and organisms with endogenously produced pigmentation. In these systems, genetic drift is often required for novel alleles to overcome strong purifying selection stemming from frequency-dependent predation and positive assortative mating. Here, we integrate data from field surveys, predation experiments, population genomics, and phenotypic correlations to explain the origin and maintenance of geographic variation in a diet-based larval pigmentation trait in the redheaded pine sawfly (Neodiprion lecontei), a pine-feeding hymenopteran. Although our experiments confirm that N. lecontei larvae are indeed aposematic-and therefore likely to experience frequency-dependent predation-our genomic data do not support a historical demographic scenario that would have facilitated the spread of an initially deleterious allele via drift. Additionally, significantly elevated differentiation at a known color locus suggests that geographic variation in larval color is currently maintained by selection. Together, these data suggest that the novel white morph likely spread via selection. However, white body color does not enhance aposematic displays, nor is it correlated with enhanced chemical defense or immune function. Instead, the derived white-bodied morph is disproportionately abundant on a pine species with a reduced carotenoid content relative to other pine hosts, suggesting that bottom-up selection via host plants may have driven divergence among populations. Overall, our results suggest that life stage and pigment source can have a substantial impact on the evolution of novel warning signals, highlighting the need to investigate diverse aposematic taxa to develop a comprehensive understanding of color variation in nature.

The impact of different diets on chemical defense has been extensively studied in animals that sequester defensive chemicals from food. However, there are fewer studies of diet-mediated variation in autogenously produced defenses. Ladybird beetles, which use autogenously synthesized defensive alkaloids, are used as models in a wide diversity of studies of chemical defense, specifically in studies of intraspecific variation in color pattern and chemical defense. Many aphidophagous ladybirds consume a wide diversity of aphid prey, which vary in quality and thus could affect the synthesis of chemical defense. We measured alkaloid recovery rate after reflex bleeding by the ladybird Adalia bipunctata on two different aphid diets, the high quality Acyrthosiphon pisum and the lower quality Aphis fabae. Alkaloids reaccumulated in ladybirds more slowly when they were fed A. fabae than when they were fed A. pisum and females generally had more alkaloid than males, but reaccumulated alkaloid more slowly. Recovery times were more than 12 days. There appeared to be a weak positive relationship between alkaloid level and time since reflex bleeding for eggs of A. pisum- but not A. fabae-fed females. Our findings on diet and alkaloid synthesis in ladybirds suggest that chemical defense levels are very dynamic, indicating that studies conducted at a single point in time, such as those focused on ladybird color pattern, fail to consider a wide diversity of temporal variation that occurs in the field. This is likely true for many autogenously produced chemical defense systems in a diversity of other organisms.

Marine invasive species and their bioactive metabolites have become critical ecological issues in the Mediterranean Sea. In particular, the highly invasive green algae Caulerpa taxifolia and Caulerpa cylindracea are known to contain the bioactive sesquiterpene caulerpenyne (1) and the bisindolic alkaloid caulerpin (2), potentially acting as chemical stressors for native species. The recent spread of a variety of C. taxifolia, Caulerpa taxifolia var. distichophylla, also raises urgent questions about its chemical composition. Indeed, the only chemical data available for this alga are limited to the seasonal variations of caulerpenyne (1) in samples collected in the Eastern Mediterranean. In this study, we confirmed the presence of 1 also in C. taxifolia var. distichophylla collected along the Sicilian coast, while 2 was not detected in the alga. However, caulerpin(2) was found both in a Mediterranean specimen of C. taxifolia, and at a much higher level in the congeneric C. cylindracea. This suggests that C. taxifolia var. distichophylla differs from C. taxifolia in its secondary metabolism, potentially exerting dissimilar chemically mediated impacts on native biota. Further chemical investigations on the terpenoidic content of C. taxifolia var. distichophylla led to isolate and identify squalene 2,3 oxide (3), phytol (4), and plastoquinone (5), along with the two unreported sesquiterpene lactones 6 and 7. Finally, chemoecological assays clarified that caulerpenyne (1) at its natural concentration in C. taxifolia var. distichophylla elicits avoidance responses in native shrimp, although sensitization was a prerequisite to significantly induce food rejection.

Cyanobacteria are photosynthetic microorganisms that compose phytoplankton and therefore have a trophic relationship with zooplankton, which represent an important link for energy flux in aquatic food webs. Several species can form blooms and produce bioactive metabolites known as cyanotoxins. However, the ecological and adaptative role of these toxins are still under debate. Many studies have addressed the cyanotoxins’ function in defense against herbivory when grazing pressure by zooplankton plays a role in phytoplankton top-down control. Thus, the present study evaluated the ecophysiological responses of the cyanobacterial strain Microcystis aeruginosa NPLJ-4 underlying the chemical induced defense against the cladoceran Daphnia gessneri. Exposure to predator infochemicals consisted of cultures established in ASM-1 medium prepared in a filtrate from a culture of adults of D. gessneri at an environmentally relevant density. Daphnia infochemicals promoted a significant increase in toxin production by M. aeruginosa. However, no differences in growth were observed, despite a significant increase in both maximum photosynthetic efficiency and electron transport rate in response to zooplankton. Additionally, there was no significant variation in the production of exopolysaccharides. Overall, although a grazer-induced defense response was demonstrated, there were no effects on M. aeruginosa fitness, which maintained its growth in the presence of Daphnia alarm cues.

Antipredation strategies contribute to the lifetime reproductive success of organisms, particularly in more vulnerable life stages that look to survive until reproduction. In insects, eggs and larval stages are often immobile or unable to rapidly flee and hide from predators. Understanding what alternative antipredation strategies they use, but also how these change over development time, is required to fully appreciate how species adapt to biotic threats. Murgantia histrionica is a stink bug, conspicuously colored from egg to adult, known to sequester defensive glucosinolates from its cruciferous hosts as adults. We sought to assess whether this chemical defense is also present in its eggs and early nymphal instars and quantified how it fluctuates among life stages. In parallel, we looked at an alternative antipredation strategy, described for the first time in this species: tonic immobility (i.e., death feigning). We also qualitatively investigated ultraviolet reflectance in eggs and adults as a proxy of conspicuousness against UV-absorbing leaves. Our results show that the eggs are significantly more chemically defended than the first two but not third mobile life stages, yet compound concentrations do not statistically differ across nymphal instars. Tonic immobility is favored by hatchlings, but less so by subsequent instars. Eggs also had obvious ultraviolet reflectance, suggesting that they would contrast against a leaf substrate and, considering their chemical load, that they may be aposematic. We argue that there are two possible interpretations of our results. One is that, throughout ontogeny, tonic immobility is a useful defensive strategy until adequate chemical protection is achieved over an extended feeding period. The other is that both aposematism and tonic immobility are used by this species, but variation in strategy use throughout ontogeny is decoupled.

Aphids are abundant in natural and managed vegetation, supporting a diverse community of organisms and causing damage to agricultural crops. Due to a changing climate, periods of drought are anticipated to increase, and the potential consequences of this for aphid–plant interactions are unclear.Using a meta‐analysis and synthesis approach, we aimed to advance understanding of how increased drought incidence will affect this ecologically and economically important insect group and to characterize any potential underlying mechanisms. We used qualitative and quantitative synthesis techniques to determine whether drought stress has a negative, positive, or null effect on aphid fitness and examined these effects in relation to (a) aphid biology, (b) geographical region, and (c) host plant biology.Across all studies, aphid fitness is typically reduced under drought. Subgroup analysis detected no difference in relation to aphid biology, geographical region, or the aphid–plant combination, indicating the negative effect of drought on aphids is potentially universal. Furthermore, drought stress had a negative impact on plant vigor and increased plant concentrations of defensive chemicals, suggesting the observed response of aphids is associated with reduced plant vigor and increased chemical defense in drought‐stressed plants.We propose a conceptual model to predict drought effects on aphid fitness in relation to plant vigor and defense to stimulate further research., Using meta‐analysis, we provide the first comprehensive analysis of drought stress on aphid fitness, showing negative effects across different host plant families, aphid tribes, and geographic regions. These negative effects were observed for different aphid fitness parameters and host specialisms and likely result from drought‐induced reduction in plant vigor and increased tissue concentrations of plant‐defensive compounds.

In natural and agricultural ecosystems, plants are often simultaneously or sequentially exposed to combinations of stressors. Here we tested whether limited water availability (LWA) affects plant response to insect herbivory using two populations of Eruca sativa from desert and Mediterranean habitats that differ in their induced defenses. Considering that such differences evolved as responses to biotic and possibly abiotic stress factors, the two populations offered an opportunity to study ecological aspects in plant response to combined stresses. Analysis of chemical defense mechanisms showed that LWA significantly induced total glucosinolate concentrations in the Mediterranean plants, but their concentrations were reduced in the desert plants. However, LWA, with and without subsequent jasmonate elicitation, significantly induced the expression of proteinase inhibitor in the desert plants. Results of a no-choice feeding experiment showed that LWA significantly increased desert plant resistance to Spodoptera littoralis larvae, whereas it did not affect the relatively strong basal resistance of the Mediterranean plants. LWA and subsequent jasmonate elicitation increased resistance against the generalist insect in Mediterranean plants, possibly due to both increased proteinase inhibitor expression and glucosinolate accumulation. The effect of LWA on the expression of genes involved in phytohormone signaling, abscisic acid (ABA-1) and jasmonic acid (AOC1), and the jasmonate responsive PDF1.2, suggested the involvement of abscisic acid in the regulation of defense mechanisms in the two populations. Our results indicate that specific genotypic responses should be considered when estimating general patterns in plant response to herbivory under water deficiency conditions.

Plants constantly cope with insect herbivory, which is thought to be the evolutionary driver for the immense diversity of plant chemical defenses. Herbivorous insects are in turn restricted in host choice by the presence of plant chemical defense barriers. In this study, we analyzed whether butterfly host–plant patterns are determined by the presence of shared plant chemical defenses rather than by shared plant evolutionary history. Using correlation and phylogenetic statistics, we assessed the impact of host–plant chemical defense traits on shaping northwestern European butterfly assemblages at a macroevolutionary scale. Shared chemical defenses between plant families showed stronger correlation with overlap in butterfly assemblages than phylogenetic relatedness, providing evidence that chemical defenses may determine the assemblage of butterflies per plant family rather than shared evolutionary history. Although global congruence between butterflies and host–plant families was detected across the studied herbivory interactions, cophylogenetic statistics showed varying levels of congruence between butterflies and host chemical defense traits. We attribute this to the existence of multiple antiherbivore traits across plant families and the diversity of insect herbivory associations per plant family. Our results highlight the importance of plant chemical defenses in community ecology through their influence on insect assemblages., Using correlation and phylogenetic statistics, we assessed the impact of host‐plant chemical defense traits on shaping Northwestern European butterfly assemblages at a macroevolutionary scale. Shared chemical defenses between plant families showed stronger correlation with overlap in butterfly assemblages than phylogenetic relatedness, providing evidence that chemical defenses may determine the assemblage of butterflies per plant family rather than shared evolutionary history. ​

Communication behaviors play a critical role in both an individual’s fitness as well as the viability of populations. Solitary animals use chemical communication (i.e., scent marking) to locate mates and defend their territory to increase their own fitness. Previous research has suggested that striped skunks (Mephitis mephitis) do not perform scent-marking behaviors, despite being best known for using odor as chemical defense. We used video camera traps to document behaviors exhibited by striped skunks at a remote site in coastal California between January 2012 and April 2015. Our camera traps captured a total of 71 visits by striped skunks, the majority of which (73%) included a striped skunk exhibiting scent-marking behaviors. Overall, we documented 8 different scent-marking behaviors. The most frequent behaviors we documented were cheek rubbing (45.1%), investigating (40.8%), and claw marking (35.2%). The behaviors exhibited for the longest durations on average were grooming ( $$ \overline{x} $$ = 34. 4 s) and investigating ( $$ \overline{x} $$ = 21.2 s). Although previous research suggested that striped skunks do not scent mark, we documented that at least some populations do and our findings suggest that certain sites are used for communication via scent marking. Our study further highlights how camera traps allow researchers to discover previously undocumented animal behaviors.

Based on the hypothesis that the variation of the metabolomes of latex is a response to selective pressure and should thus be affected differently from other organs, their variation could provide an insight into the defensive chemical selection of plants. Metabolic profiling was used to compare tissues of three Euphorbia species collected in diverse regions. The metabolic variation of latexes was much more limited than that of other organs. In all the species, the levels of polyisoprenes and terpenes were found to be much higher in latexes than in leaves and roots of the corresponding plants. Polyisoprenes were observed to physically delay the contact of pathogens with plant tissues and their growth. A secondary barrier composed of terpenes in latex and in particular, 24-methylenecycloartanol, exhibited antifungal activity. These results added to the well-known role of enzymes also present in latexes, show that these are part of a cooperative defense system comprising biochemical and physical elements. Supplementary Information The online version contains supplementary material available at 10.1007/s10886-021-01274-x.

Of the approximately one million described insect species, ground beetles (Coleoptera: Carabidae) have long captivated the attention of evolutionary biologists due to the diversity of defensive compounds they synthesize. Produced using defensive glands in the abdomen, ground beetle chemicals represent over 250 compounds including predator-deterring formic acid, which has evolved as a defensive strategy at least three times across Insecta. Despite being a widespread method of defense, formic acid biosynthesis is poorly understood in insects. Previous studies have suggested that the folate cycle of one-carbon (C1) metabolism, a pathway involved in nucleotide biosynthesis, may play a key role in defensive-grade formic acid production in ants. Here, we report on the defensive gland transcriptome of the formic acid-producing ground beetle Harpalus pensylvanicus. The full suite of genes involved in the folate cycle of C1 metabolism are significantly differentially expressed in the defensive glands of H. pensylvanicus when compared to gene expression profiles in the rest of the body. We also find support for two additional pathways potentially involved in the biosynthesis of defensive-grade formic acid, the kynurenine pathway and the methionine salvage cycle. Additionally, we have found an array of differentially expressed genes in the secretory lobes involved in the biosynthesis and transport of cofactors necessary for formic acid biosynthesis, as well as genes presumably involved in the detoxification of secondary metabolites including formic acid. We also provide insight into the evolution of the predominant gene family involved in the folate cycle (MTHFD) and suggest that high expression of folate cycle genes rather than gene duplication and/or neofunctionalization may be more important for defensive-grade formic acid biosynthesis in H. pensylvanicus. This provides the first evidence in Coleoptera and one of a few examples in Insecta of a primary metabolic process being co-opted for defensive chemical biosynthesis. Our results shed light on potential mechanisms of formic acid biosynthesis in the defensive glands of a ground beetle and provide a foundation for further studies into the evolution of formic acid-based chemical defense strategies in insects.

Plant defense theory explores how plants invest in defenses against natural enemies but has focused primarily on the traits expressed by juvenile and mature plants. Here we describe the diverse ways in which seeds are chemically and physically defended. We suggest that through associations with other traits, seeds are likely to exhibit defense syndromes that reflect constraints or trade-offs imposed by selection to attract dispersers, enable effective dispersal, ensure appropriate timing of seed germination, and enhance seedling performance. We draw attention to seed and reproductive traits that are analogous to defense traits in mature plants and describe how the effectiveness of defenses is likely to differ at pre- and postdispersal stages. We also highlight recent insights into the mutualistic and antagonistic interactions between seeds and microbial communities, including fungi and endohyphal bacteria, that can influence seed survival in the soil and subsequent seedling vigor.

Since most species are collections of genetically variable populations distributed to habitats differing in their abiotic/biotic environmental factors and community composition, the pattern and strength of natural selection imposed by species on each others' traits are also expected to be highly spatially variable. Here, we used genomic and quantitative genetic approaches to understand how spatially variable selection operates on the genetic basis of plant defenses to herbivores. To this end, an F2 progeny was generated by crossing Datura stramonium (Solanaceae) parents from two populations differing in their level of chemical defense. This F2 progeny was reciprocally transplanted into the parental plants’ habitats and by measuring the Identity by Descent (IBD) relationship of each F2 plant to each parent, we were able to elucidate how spatially variable selection imposed by herbivores operated on the genetic background (IBD) of resistance to herbivory, promoting local adaptation. The results highlight that plants possessing the highest total alkaloid concentrations (sum of all alkaloid classes) were not the most well‐defended or fit. Instead, specific alkaloids and their linked loci/alleles were favored by selection imposed by different herbivores. This has led to population differentiation in plant defenses and thus, to local adaptation driven by plant‐herbivore interactions. This article is protected by copyright. All rights reserved

Toads of the genus Bufo synthesize and accumulate bufadienolides (BDs) in their parotoid glands. BDs are cardiotonic steroids that play an important role in defense against the toads' predators. Three bufonid taxa occur in mainland Japan, Bufo japonicus formosus, B. j. japonicus, and B. torrenticola. The chemical structures of BDs isolated from B. j. formosus were studied several decades ago, but there is no further information on the toxic components of Japanese toads and their metabolism. In this study, we analyzed BDs of toads from throughout Japan and compared the BD profiles by liquid chromatography/mass spectrometry (LC/MS) and hierarchical cluster analysis (HCA). We observed BDs in three taxa of Japanese toads, and identified five of the most common BDs by nuclear magnetic resonance (NMR) analyses. Of the five BDs, only bufalin was detected in all individuals. HCA of individual BD profiles divided the three taxa into five primary clusters and several subclusters. This result indicates that BD profiles differ both among and within the taxa. The clustering pattern of BDs is generally concordant with a phylogenetic tree reconstructed from the mitochondrial cytochrome b gene of Japanese toads. Our results suggest that the BDs of Japanese toads have diversified not in response to specific selective pressures, but simply due to population structuring over evolutionary time.

Temperature influences all biochemical and biophysiological processes within an organism. By extension, it also affects those ecological interactions that are mediated by gland-produced chemical compounds, such as reservoir-based chemical defense. Herein, we investigate how environmental temperature affects the regeneration of defensive secretions and influences the efficacy of chemical defense in a model predator-prey system: the oribatid mite Archegozetes longisetosus and the predaceous rove beetle Stenus juno. Through a combination of chemical analyses, non-linear regression modeling and theoretical simulations we show that the amount of defensive secretion responded to temperature in a unimodal optimum curve: the regeneration rate followed a positive, linear relationship up to 35 °C, but rapidly broke down beyond this temperature (“tipping point” effect). Using functional response simulations, there is an initially positive dampening effect on the predation rate when regeneration is optimal, but at higher temperatures chemical defense does not counteract the previously described effects of elevated predatory pressure. In a larger context, our results demonstrate the need to integrate relevant environmental factors in predator-prey modeling approaches.

Insects evolved adaptive plasticity to minimize the effects of the chemical defenses of their host plants. Nevertheless, the expressional response and adaptation of phytophagous specialists for long‐term adaption and short‐term response to host phytochemicals remains largely unexplored. The mulberry (Morus alba)–silkworm (Bombyx mori) interaction is an old and well‐known model of plant–insect interaction. In this study, we examined the long‐term adaption and short‐term response of the mulberry‐specialist silkworm to two sugar‐mimic alkaloids in mulberry: the commonly encountered 1‐deoxynojirimycin (1‐DNJ) and occasionally encountered 1,4‐dideoxy‐1,4‐imino‐D‐arabinitol (D‐AB1), respectively. Global transcriptional patterns revealed that the physiological responses induced by the selective expression of genes involved in manifold cellular processes, including detoxification networks, canonical digestion processes, target enzymes, and other fundamental physiological processes, were crucial for regulating metabolic homeostasis. Comparative network analysis of the effects of exposure to D‐AB1 and 1‐DNJ supported the contention that B. mori produced similar and specific trajectories of changed gene expression in response to different sugar‐mimic alkaloids. D‐AB1 elicited a substantial proportion of downregulated genes relating to carbohydrate metabolism, catabolic process, lipid metabolism, and glycan biosynthesis and metabolism. This study dramatically expands our knowledge of the physiological adaptations to dietary sugar‐mimic alkaloid intake and uncovered both metabolic evolutionarily responses and unique adaptive mechanisms previously unknown in insects., Insects evolved adaptive plasticity to minimize the effects of the chemical defenses of their host plants. In this study, we found that the transcriptional plasticity in genes related to metabolism, detoxification, digestion, and general cellular processes played a key role in both long‐term adaptation and short‐term responses of the mulberry‐specialist insect to changing sugar‐mimic alkaloids.

Birds are important predators of insects and insects often incorporate chemical defenses that may make themselves distasteful or toxic to the predators. Predators can respond to chemically defended prey in multiple ways, the predator psychology approach to predation often treats predation as a general process despite the possibility for multiple responses among species. The effectiveness of a prey’s chemical defense at reducing predation might also vary depending on what predator is attacking the prey. Here, we compared the attack strategies of three different species of avian predators (Japanese bush warblers [Horornis diphone], narcissus flycatchers [Ficedula narcissina], and Japanese tits [Parus minor]) which are found in the temperate forests of Japan. We found overall, that undefended prey was preferred over the defended prey, but the different predator species had different preferences and handled prey differently from one another. This suggests that different predator species might exert different selection pressures on chemically defended prey and this adds to our growing appreciation that predator behavior can vary among predator species. Moreover, our findings emphasize the importance of understanding differences in behavior among free-living predator species in studies of aposematism and mimicry.

Evolutionary convergence of color pattern in mimetic species is tightly linked with the evolution of chemical defenses. Yet, the evolutionary forces involved in natural variations of chemical defenses in aposematic species are still understudied. Herein, we focus on the evolution of chemical defenses in the butterfly tribe Heliconiini. These neotropical butterflies contain large concentrations of cyanogenic glucosides, cyanide‐releasing compounds acting as predator deterrent. These compounds are either de novo synthesized or sequestered from their Passiflora host plant, so that their concentrations may depend on host plant specialization and host plant availability. We sampled 375 wild Heliconiini butterflies across Central and South America, covering 43% species of this clade, and quantify individual variations in the different CGs using liquid chromatography coupled with tandem mass spectrometry. We detected new compounds and important variations in chemical defenses both within and among species. Based on the most recent and well‐studied phylogeny of Heliconiini, we show that ecological factors such as mimetic interactions and host plant specialization have a significant association with chemical profiles, but these effects are largely explained by phylogenetic relationships. Our results therefore suggest that shared ancestries largely contribute to chemical defense variation, pointing out at the interaction between historical and ecological factors in the evolution of Müllerian mimicry., Within Heliconiini butterflies, we show that ecological factors such as mimetic interactions and host plant specialization have a significant effect on chemical profiles, but these effects are largely explained by phylogenetic relationships. Our results suggest that shared ancestries largely contribute to chemical defence variation, pointing out at the interaction between historical and ecological factors in the evolution of Müllerian mimicry.

Many specialist herbivores have evolved strategies to cope with plant defences, with gut microbiota potentially participating to such adaptations. In this study, we assessed whether the history of plant use (population origin) and microbiota may interact with plant defence adaptation. We tested whether microbiota enhance the performance of Melitaea cinxia larvae on their host plant, Plantago lanceolata and increase their ability to cope the defensive compounds, iridoid glycosides (IGs). The gut microbiota were significantly affected by both larval population origin and host plant IG level. Contrary to our prediction, impoverishing the microbiota with antibiotic treatment did not reduce larval performance. As expected for this specialized insect herbivore, sequestration of one of IGs was higher in larvae fed with plants producing higher concentration of IGs. These larvae also showed metabolic signature of intoxication (i.e. decrease in Lysine levels). However, intoxication on highly defended plants was only observed when larvae with a history of poorly defended plants were simultaneously treated with antibiotics. Our results suggest that both adaptation and microbiota contribute to the metabolic response of herbivores to plant defence though complex interactions. Read the free Plain Language Summary for this article on the Journal blog.

Fungi are widely distributed in the terrestrial environment, freshwater, and marine habitat. Only approximately 100,000 of these have been classified although there are about 5.1 million characteristic fungi all over the world. These eukaryotic microbes produce specialized metabolites and participate in a variety of ecological functions, such as quorum detection, chemical defense, allelopathy, and maintenance of symbiosis. Fungi therefore remain an important resource for the screening and discovery of biologically active natural products. Sesquiterpenoids are arguably the richest natural products from plants and micro-organisms. The rearrangement of the 15 high-ductility carbons gave rise to a large number of different skeletons. At the same time, abundant structural variations lead to a diversification of biological activity. This review examines the isolation, structural determination, bioactivities, and synthesis of sesquiterpenoids that were specially produced by fungi over the past five years (2015–2020).

Aposematic organisms warn predators of their unprofitability using a combination of defenses, including visual warning signals, startling sounds, noxious odors, or aversive tastes. Using multiple lines of defense can help prey avoid predators by stimulating multiple senses and/or by acting at different stages of predation. We tested the efficacy of three lines of defense (color, smell, taste) during the predation sequence of aposematic wood tiger moths (Arctia plantaginis) using blue tit (Cyanistes caeruleus) predators. Moths with two hindwing phenotypes (genotypes: WW/Wy = white, yy = yellow) were manipulated to have defense fluid with aversive smell (methoxypyrazines), body tissues with aversive taste (pyrrolizidine alkaloids) or both. In early predation stages, moth color and smell had additive effects on bird approach latency and dropping the prey, with the strongest effect for moths of the white morph with defense fluids. Pyrrolizidine alkaloid sequestration was detrimental in early attack stages, suggesting a trade-off between pyrrolizidine alkaloid sequestration and investment in other defenses. In addition, pyrrolizidine alkaloid taste alone did not deter bird predators. Birds could only effectively discriminate toxic moths from non-toxic moths when neck fluids containing methoxypyrazines were present, at which point they abandoned attack at the consumption stage. As a result, moths of the white morph with an aversive methoxypyrazine smell and moths in the treatment with both chemical defenses had the greatest chance of survival. We suggest that methoxypyrazines act as context setting signals for warning colors and as attention alerting or “go-slow” signals for distasteful toxins, thereby mediating the relationship between warning signal and toxicity. Furthermore, we found that moths that were heterozygous for hindwing coloration had more effective defense fluids compared to other genotypes in terms of delaying approach and reducing the latency to drop the moth, suggesting a genetic link between coloration and defense that could help to explain the color polymorphism. Conclusively, these results indicate that color, smell, and taste constitute a multimodal warning signal that impedes predator attack and improves prey survival. This work highlights the importance of understanding the separate roles of color, smell and taste through the predation sequence and also within-species variation in chemical defenses.

In nature some successful invasive plants grow faster and are better chemical defenders than native plants, which appears to contradict the conventional theory that plant growth and chemical defense are trade-offs in terms of the allocation of limited resources. Using greenhouse experiments we compared the growth, chemical defensive matters (condensed tannins and total phenolics) and energy-use strategies (construction cost and energy-use efficiency) of four highly noxious invasive plants to their congeneric or co-occurring native species in subtropical China. We hypothesized that the trade-offs between growth and chemical defense in invasive plants are alleviated, and this alleviation benefits from their high photosynthetic capacities and energy-use strategies. Our results showed that all the four invasive plants grew faster than their native counterparts, and three of them had higher condensed tannins and the remaining one had higher total phenolics, indicating the invasive plants achieved both faster growth and stronger chemical defense. Moreover, the growth rate positively related to condensed tannins but negatively to total phenolics in both invasive and native plants, indicating the chemical-specific pattern of the trade-offs. Our results demonstrate that the higher photosynthetic capacity and energy-use efficiency together with lower biomass construction cost facilitated the faster growth and stronger chemical defense in invasive plants, providing an insight for their chemical-specific trade-offs between growth and chemical defense.

Plants are chemically-complex organisms; each individual contains diverse tissue-types, has the ability to differentially allocate secondary metabolites to these tissues and can change this allocation through time. The interaction of variation in chemical defense of different tissue types and variation in chemical defense through time, however, is rarely examined and has not been studied for iridoid glycoside-producing woody plants. In this study, we quantified allocation of iridoid glycosides (IGs) to the leaves, flowers, fruits, and seeds of 25 individuals of a long-lived shrub (Lonicera x bella Zabel, Caprifoliaceae), at five important phenological timepoints (leaf-out, flowering, fruit appearance, fruit ripening, and fruit dispersal) throughout a growing season. We found that leaves had 2x higher IG concentrations during flowering and fruiting than earlier in the season (after leaf-out), and later in the season (after fruit dispersal). The individual IG driving this increase in leaves during reproduction, secologanin, was also the most abundant IG in semiripe fruits. Flowers and seeds were composed of different proportions of individual IGs than fruits or leaves, but did not change across time and had overall low concentrations of IGs. In L. x bella, phenological events such as flowering and fruiting lead to an increase in leaf chemical defense that is likely to influence interactions with leaf-feeders. Our results stress the importance of considering phenology when sampling plants for the quantification of chemical defenses.

Much of the world’s biodiversity is held within tropical rainforests, which are increasingly fragmented by agricultural practices. In these threatened landscapes, there are many organisms that acquire chemical defenses from their diet and are therefore intimately connected with their local food webs. Poison frogs (Family Dendrobatidae) are one such example, as they acquire alkaloid-based chemical defenses from their diet of leaf litter ants and mites. It is currently unknown how habitat fragmentation impacts chemical defense across trophic levels, from arthropods to frogs. We examined the chemical defenses and diets of the Diablito poison frog (Oophaga sylvatica), and the diversity of their leaf litter ant communities in secondary forest and reclaimed cattle pasture. Notably, this research was performed in collaboration with two high school science classrooms. We found that the leaf litter of forest and pasture frog habitats differed significantly in ant community structure. We also found that forest and pasture frogs differed significantly in diet and alkaloid profiles, where forest frogs contained more of specific alkaloids and ate more ants in both number and volume. Finally, ant species composition of frog diets resembled the surrounding leaf litter, but diets were less variable. This suggests that frogs tend to consume particular ant species within each habitat. To better understand how ants contribute to the alkaloid chemical profiles of frogs, we chemically profiled several ant species and found some alkaloids to be common across many ant species while others are restricted to a few species. At least one alkaloid (223H) found in ants from disturbed sites was also found in skins from pasture. Our experiments are the first to link anthropogenic land use changes to dendrobatid poison frog chemical defenses through variation in leaf litter communities, which has implications for conservation management of these threatened amphibians.

We review a recently discovered white spruce (Picea glauca) chemical defense against spruce budworm (Choristoneura fumiferana)involving hydroxyacetophenones. These defense metabolites detected in the foliage accumulate variably as the aglycons, piceol and pungenol, or the corresponding glucosides, picein and pungenin. We summarize current knowledge of the genetic, genomic, molecular, and biochemical underpinnings of this defense and its effects onC.fumiferana. We present an update with new results on the ontogenic variation and the phenological window of this defense, including analysis of transcript responses inP.glaucatoC.fumiferanaherbivory. We also discuss this chemical defense from an evolutionary and a breeding context.

Frogs in the genus Phyllobates are known for the presence of batrachotoxin, a highly toxic alkaloid, in their skin. Nevertheless, Phyllobates frogs from Costa Rica and Panama (P. lugubris and P. vittatus) are considered non-toxic, as they have been reported to harbor low concentrations of this alkaloid. However, the potential toxicity of Central American Phyllobates has not been assessed experimentally. Our goal was to determine the toxicity of the whole skin of P. vittatus, an endemic species from the Southeastern Pacific region of Costa Rica. We performed median lethal dose (LD50) tests in mice to determine general toxicity, and an irritant assay based on the behavioral responses of mice to subcutaneous injection, to determine differences in irritability, as a measure of toxicity, among three study localities. Using UPLC-ESI-QTOF, we obtained chemical profiles of the methanolic extract of frog skins. Due to the absence of mortality at the studied doses, we were unable to estimate LD50. However, we recorded a list of toxicity symptoms in mice that are consistent with cardiotoxic effects, and found that mice presented more symptoms at higher concentrations of skin extracts during the first hour of the LD50 assays, recovering completely at all doses by the end of the assay. On the other hand, we did not detect differences in irritability among studied localities. Additionally, we putatively identified three toxic alkaloids (Batrachotoxinin A, DHQ 251A and Lehm 275A). This study provides the first experimental data on the toxicity and associated symptoms in mice, as well as the chemical profile of the skin of P. vittatus. We suggest that the skin alkaloids of P. vitattus may confer a chemical defense towards predators.

Most transplant experiments across species geographic range boundaries indicate that adaptation to stressful environments outside the range is often constrained. However, the mechanisms of these constraints remain poorly understood. We used extended generation crosses from diverged high and low elevation populations. In experiments across low elevation range boundaries, there was selection on the parental lines for abiotic stress‐tolerance and resistance to herbivores. However, in support of a defense‐tolerance trade‐off, extended generation crosses showed nonindependent segregation of these traits in the laboratory across a drought‐stress gradient and in the field across the low elevation range boundary. Genotypic variation in a marker from a region of the genome containing a candidate gene (MYC2) was associated with change in the genetic trade‐off. Thus, using crosses and forward genetics, we found experimental genetic and molecular evidence for a pleiotropic trade‐off that could constrain the evolution of range expansion., Evolutionary constraints are important to understand range limits, yet the mechanisms are poorly understood. Here we used an integrative approach to show that a trade‐off between abiotic stress‐tolerance and chemical defense against herbivores may inhibit the evolution of low elevation range expansion in the ecological model Boechera stricta.

Previous studies have shown a causal link between mammalian herbivory, tolerance, and chemical defense in Arabidopsis thaliana, driven by the process of endoreduplication (replication of the genome without mitosis). Removal of the apical meristem by mammalian herbivores lowers auxin, which triggers entry into the endocycle. Increasing chromosome number through endoreduplication, and therefore gene copy number, provides a means of increasing gene expression promoting rapid regrowth rates, higher defensive chemistry and enhanced fitness. Here, we assess whether insect leaf-feeding elicits the same compensatory response as the removal of apical dominance. Insect feeding has been shown to downregulate auxin production, which should trigger endoreduplication. Results here support this contention; insect leaf-feeding by Trichoplusia ni elicited a compensatory response similar to that of mammalian herbivores-an ecotype-specific response consistent with the level of endoreduplication. The interactive effects of mammalian and insect herbivory were also assessed to determine whether interactions were additive (pairwise) or non-additive (diffuse) on tolerance (fitness). Specifically, results indicate that herbivory is either diffuse (a significant clipping × T. ni interaction) or pairwise (no significant interaction between clipping and T. ni herbivory), dependent upon plant genotype and compensatory ability. In general, herbivore-induced changes in plant quality appear to be responsible for the observed differences in herbivory and fitness compensation. We discuss the importance of evaluating endoreduplication among plants within a population to avoid masking the association between tolerance and resistance and the fitness consequences of multi-herbivore interactions.

Salicylates and jasmonates may act synergically or antagonically depending on the plant growth conditions, biotic and abiotic stress factors to which plants are exposed to, and the tissue and plant species. Although possible tradeoffs between salicylate and jasmonate accumulation have been documented in model plants, information is still scarce in Mediterranean plant species, particulary in studies considering plants growing in their natural habitat. We evaluated to what extent inter-individual variability, sun orientation and diurnal variations in environmental conditions modulate not only the accumulation of salicylic acid and jasmonates (both free and conjugated forms), but also the contents of abscisic acid, hydrolyzable tannins and free phenolics, in white-leaved rockrose (Cistus albidus L.), a semi-deciduous shrub typically found in the Mediterranean basin. We found that jasmonates, most particularly the biologically active jasmonoyl-isoleucine and the jasmonic acid precursor, 12-oxo-phytodienoic acid showed a much more marked inter-individual variability than salicylic acid, abscicic acid, hydrolyzable tannins and free phenolics. East orientation reduced both abscisic acid and salicylic acid contents, while increased the accumulation of both jasmonates and hydrolyzable tannins, without affecting total free phenolic contents. Furthermore, salicylic acid and jasmonates contents, in particular those of 12-oxo-phytodienoic acid, changed throughout the day, while those of abscisic acid, hydrolyzable tannins and free phenolics remained constant. We conclude that sun orientation and inter-individual variability strongly influence stress-related phytohormones and chemical defense accumulation, with East-oriented individuals presenting higher amounts of jasmonates and hydrolyzable tannins, but less abscisic acid and salicylic acid than West-oriented ones. Results suggest a tradeoff between salicylic acid and jasmonates accumulation under stress conditions in C. albidus growing in their natural habitat.

Many organisms use inducible defenses as protection against predators. In animals, inducible defenses may manifest as changes in behavior, morphology, physiology, or life history, and prey species can adjust their defensive responses based on the dangerousness of predators. Analogously, prey may also change the composition and quantity of defensive chemicals when they coexist with different predators, but such predator‐induced plasticity in chemical defenses remains elusive in vertebrates. In this study, we investigated whether tadpoles of the common toad (Bufo bufo) adjust their chemical defenses to predation risk in general and specifically to the presence of different predator species; furthermore, we assessed the adaptive value of the induced defense. We reared tadpoles in the presence or absence of one of four caged predator species in a mesocosm experiment, analyzed the composition and quantity of their bufadienolide toxins, and exposed them to free‐ranging predators. We found that toad tadpoles did not respond to predation risk by upregulating their bufadienolide synthesis. Fishes and newts consumed only a small percentage of toad tadpoles, suggesting that bufadienolides provided protection against vertebrate predators, irrespective of the rearing environment. Backswimmers consumed toad tadpoles regardless of treatment. Dragonfly larvae were the most voracious predators and consumed more predator‐naïve toad tadpoles than tadpoles raised in the presence of dragonfly cues. These results suggest that tadpoles in our experiment had high enough toxin levels for an effective defense against vertebrate predators even in the absence of predator cues. The lack of predator‐induced phenotypic plasticity in bufadienolide synthesis may be due to local adaptation for constantly high chemical defense against fishes in the study population and/or due to the high density of conspecifics.

In the 1930s, outbreaks of the wasting disease pathogen Labyrinthula zosterae caused a severe reduction of the eelgrass Zostera marina meadows in the Atlantic Ocean. Many surviving populations were found in low-salinity environments, and low-salinity environments have therefore been hypothesized to act as a refuge for eelgrass against L. zosterae infection. Here, we investigated L. zosterae pathogen load and wasting disease symptoms in eelgrass over a similar to 970 km salinity gradient (6-25 PSU) along the Swedish coast. Furthermore, laboratory infection experiments and studies of inhibitory compounds were carried out to investigated whether resistance against the pathogen is correlated to differences in natural pathogen pressure among eelgrass populations. The degree of L. zosterae infection was positively correlated to salinity and the pathogen was absent in several of the eelgrass meadows in lower salinity (7-8 PSU). However, a low L. zosterae pathogen load was also found in some eelgrass populations in the lowest salinity (6 PSU). No correlation between resistance and pathogen pressure in situ was detected, and all eelgrass shoots produced chemical compounds that inhibited L. zosterae growth. These results imply that positive correlations between L. zosterae and salinity are not due to eelgrass resistance, but rather to the poor ability of L. zosterae to cope with low salinity. However, our results also indicate that some strains of L. zosterae may adapt to low salinity, and therefore there may also be a risk of wasting disease outbreaks in low-salinity eelgrass meadows, in contrast to what so far has been the general hypothesis.

PREMISE The optimal defense theory (ODT) predicts that the allocation of chemical defenses in plants will be concentrated in parts or tissues that are of higher fitness value for the individuals that produce them. Chemicals are known to be allocated to certain parts of aquatic plants, and the morphological architecture of Nymphoides humboldtiana, a species that exposes its parts to different environmental factors and consumers, may be an excellent model to evaluate within-plant susceptibility to consumers according to the ODT. METHODS Under laboratory experimental conditions, we evaluated the defensive properties of extracts from vegetative (leaves, rhizomes, roots) and reproductive (long stem internodes, flowers, fruits) parts of N. humboldtiana against consumption by the generalist herbivorous gastropod Biomphalaria glabrata. Extracts were also subjected to chemical analysis by high-performance liquid chromatography, principal component analysis, and analysis of their relationships to defensive actions. RESULTS Extracts of all vegetative and internode (reproductive) parts of N. humboldtiana exhibited defensive properties against B. glabrata, but the long stem internodes exhibited the highest percentage of inhibition. Chemical profiles of these parts were qualitatively and quantitatively different, but a major unidentified compound is presumably responsible for the higher defensive property found in internodes. CONCLUSIONS Our results support the ODT, since chemical defense was more effective in long stem internodes, which have a high fitness value for N. humboldtiana to keep the flowers emerged on the water surface in response to the rapid and dynamic changes in water levels typical of freshwater environments.

Plants heavily rely on chemical defense systems against a variety of stressors. The glucosinolates in the Brassicaceae and some allies are the core molecules of one of the most researched such pathways. These natural products are enzymatically converted into isothiocyanates (ITCs) and occasionally other defensive volatile organic constituents (VOCs) upon fungal challenge or tissue disruption to protect the host against the stressor. The current review provides a comprehensive insight on the effects of the isothiocyanates on fungi, including, but not limited to mycorrhizal fungi and pathogens of Brassicaceae. In the review, our current knowledge on the following topics are summarized: direct antifungal activity and the proposed mechanisms of antifungal action, QSAR (quantitative structure-activity relationships), synergistic activity of ITCs with other agents, effects of ITCs on soil microbial composition and allelopathic activity. A detailed insight into the possible applications is also provided: the literature of biofumigation studies, inhibition of post-harvest pathogenesis and protection of various products including grains and fruits is also reviewed herein.

15 Pág. Instituto de Ciencias Forestales (ICIFOR), Intraspecific grafting with selected scions and rootstocks was used to study drought tolerance in the recalcitrant Mediterranean conifer P. pinaster. Conifers store different terpene types in needles, stems and roots, which can be modified by drought, but most studies have focused on a single organ and over selected terpenes which limits understanding of the overall plant response. In this work, terpene profile (160 components among mono-, sesqui-, di-terpenes and resin acids) as well as eco-physiological traits were assessed in needles, roots, and scion and rootstock stems to analyze the drought sensibility of scions and rootstocks on grafted P. pinaster trees. We used two scions from wet (Galicia) and dry (Oria) provenances grafted onto rootstocks showing differences in the efficiency of water use, grown under two different water regimes, well-watered and water stressed. Constitutively, organs showed clearly different qualitative and quantitative terpene profiles, such as a long-term optimized resources allocation, being neutral and acid diterpenes those more discriminant. Environmental factors may selectively affect the regulation of diterpene biosynthesis in needles, since scion provenances determined the variation of their profiles, mainly regarding again neutral and acid diterpenes. Different scion-conferred terpene phenotypes were observed in each organ, having a relevant role isoabienol content. Drought triggered needle/root resin acids redistribution, which may provide higher protection to roots, correlated to the effective water stress endured by plants. Drought sensitivity of scion-rootstock combinations determined the intensity of drought-triggered changes in terpene levels in all organs, suggesting that chemical defense and drought tolerance are somehow interconnected., This study was supported by the projects AGL2015-66048-C2-1-R (pROMPTeR) and RTI2018-098015-B-I00 (PErDURE) from Ministerio de Economía y Competitividad‐Fondos Europeos de Desarrollo Regional (Spain)

Made available in DSpace on 2021-06-25T10:56:57Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-06-01 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Several secondary metabolites are associated with plant resistance against herbivores. Cassava genotypes present a wide variety of metabolites with insecticidal potential, but little is known about the identity of these compounds. The present work was conducted for the identification of insecticidal molecules present in cassava genotypes. To this purpose, we firstly evaluated the development of the generalist herbivore Spodoptera frugiperda (J.E. Smith, 1797) (Lepidoptera: Noctuidae) fed with leaves from six cassava genotypes (IAC-14, IAC-90, IAC-12, IAC-Caapora, Baianinha and MEcu 72). Following bioassays, we measured the levels of two anti-herbivory related groups of compounds (phenolics and steroidal saponins) in all genotypes using colorimetric methods. The metabolic fingerprinting (GC–MS) of three contrasting cassava genotypes selected by their biological interferences over S. frugiperda was performed. The most unsuitable nutritional indices were observed for larvae fed with MEcu 72. Lower reproductive indices were observed for adults in which larval stages were fed with Baianinha or MEcu 72. Steroidal saponin content was similar in all genotypes, but phenolic content was 25% higher in MEcu 72. GC–MS metabolite fingerprinting of MEcu 72, Baianinha and IAC-Caapora resulted in the annotation of 53 metabolites in which 20 presented different relative abundance among the evaluated genotypes. Molecules such as myo-inositol-2-monophosphate, p-coumaric acid, chlorogenic acid, p-hydroxybenzoic acid, octadecadienoic acid and n-hexacosane were more abundant in MEcu 72 or Baianinha than in IAC-Caapora. The possible roles of these molecules in the development of S. frugiperda are discussed. School of Agricultural and Veterinarian Sciences São Paulo State University (UNESP) Institute of Biosciences São Paulo State University (UNESP) Institute of Chemistry São Paulo State University (UNESP) Departamento de Ciências da Produção Agrícola Setor de Fitossanidade Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista “Júlio de Mesquita Filho” (UNESP), Via de Acesso Prof. Paulo Donato Castellane s/n School of Agricultural and Veterinarian Sciences São Paulo State University (UNESP) Institute of Biosciences São Paulo State University (UNESP) Institute of Chemistry São Paulo State University (UNESP) Departamento de Ciências da Produção Agrícola Setor de Fitossanidade Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista “Júlio de Mesquita Filho” (UNESP), Via de Acesso Prof. Paulo Donato Castellane s/n CAPES: Finance Code 001

The capability to respond to wounding is a process shared by organisms of different kingdoms that can result in the regeneration of whole-body parts or lost structures or organs. Filamentous fungi constitute a rich food source that ensures survival and reproduction of their predators and are therefore continuously exposed to mechanical damage. Nevertheless, our understanding of how fungi respond to wounding and predators is scarce. Fungi like plants and animals respond to injury recognizing Damage- and Microbe-Associated Molecular Patterns (DAMPs/MAMPs) that activate Ca(2+) and Mitogen-Activated Protein Kinase dependent signaling for the activation of defense mechanisms. During herbivory, plants, in addition to activating pathways related to injury, activate specific responses to combat their predators. Using a transcriptional approach, we studied the capacity of the filamentous fungus Trichoderma atroviride to activate specific responses to injury and attack by different arthropods. Attack by Drosophila melanogaster inhibited the transcriptional activation of genes required for hyphal regeneration, and the fungal innate immune and chemical defense responses. We also provide mechanistic insight of this inhibition involving components of the D. melanogaster salivary glands that repress the expression of a set of genes and block hyphal regeneration.

Recent research shows that earthworms can alter defense traits of plants against herbivores and pathogens by affecting soil biochemistry. Yet, the effects of invasive earthworms on defense traits of native plants from previously earthworm‐free ecosystems as well as the consequences for multitrophic interactions are virtually unknown.Here we use a combination of an observational study and a complementary experimental study to investigate the effects of invasive earthworms on leaf defense traits, herbivore damage and pathogen infection in two poplar tree species (Populus balsamifera and Populus tremuloides) native to North American boreal forests.Our observational study showed that earthworm invasion was associated with enhanced leaf herbivory (by leaf‐chewing insects) in saplings of both tree species. However, we only detected significant shifts in the concentration of chemical defense compounds in response to earthworm invasion for P. balsamifera. Specifically, leaf phenolic concentrations, including salicinoids and catechin, were lower in P. balsamifera from earthworm‐invaded sites.Our experimental study confirmed an earthworm‐induced reduction in leaf defense levels in P. balsamifera for one of the defense compounds, tremulacin. The experimental study additionally showed that invasive earthworms reduced leaf dry matter content, potentially increasing leaf palatability, and enhanced susceptibility of trees to infection by a fungal pathogen, but not to aphid infestation, in the same tree species. Synthesis. Our results show that invasive earthworms can decrease the concentrations of some chemical defense compounds in P. balsamifera, which could make them susceptible to leaf‐chewing insects. Such potential impacts of invasive earthworms are likely to have implications for tree survival and competition, native tree biodiversity and ecosystem functioning., Invasive earthworms can dramatically alter native biodiversity and ecosystem functioning. Yet, we know little how invasive earthworms affect the chemical defense of native trees. Here using a combination of observational and experimental studies, we show that invasive earthworms from Europe can decrease the concentrations of some chemical defense compounds in a native North American deciduous tree, which could make them susceptible to leaf‐chewing insects.

Este artículo contiene 9 páginas, 6 figuras, 2 tablas., Defensive mechanisms in blister beetles (Coleoptera: Meloidae) include a wide variety of behavioral responses, chemical defense, and conspicuous external colorations. Although some of these mechanisms have been previously described, proctodeal extrusion, a defensive behavior involving the extrusion of inner abdominal membranes from the proctodeal region which appear intensely red or orange colored when the hemolymph is seen through them, has not been reported to date. Here, we tested the ability to display proctodeal extrusion in response to threat stimuli in wild populations of three blister beetle species inhabiting Central Spain: Berberomeloe majalis (Linnaeus, 1758), Berberomeloe comunero Sánchez-Vialas, García-París, Ruiz & Recuero, 2020, and Physomeloe corallifer (Germar, 1818). In addition, we observed and recorded various other defensive behaviors such as immobility, antennal threat display, autohemorrhage (reflex bleeding), defecation, and thanatosis (death feigning). The frequency at which proctodeal extrusion was observed differed among species, as did the stress intensity needed for extrusion and the probability of proctodeal extrusion in response to a particular threatening stimulus. Our findings indicate that, although proctodeal extrusion might be a widespread potential defensive mechanism in Meloidae, the ability to elicit it is not generalized across lineages. Physomeloe and Berberomeloe are endemic to the semiarid Mediterranean region, and species adapted to such a climate would have developed strategies that limit hydric stress such as proctodeal extrusion, which mirrors the effect of autohemorrhage but without the fluid loss., Field work and additional resources for this study were supported by the Spanish government (Ministerio de Ciencia, Innovación y Universidades) and the European Regional Development Fund (ERDF), under grant PID2019-110243 GB-I00/AEI/10.13039/501100011033 to MG-P

The composition of chemical weaponry of termite soldiers show interspecific and intraspecific variation. However, spatial effects on the qualitative and quantitative compositions of these substances in Neotropical termites are poorly known. Hexane extracts of heads and the defensive secretion of soldiers of Constrictotermes cyphergaster from four localities in Northeast Brazil were analyzed by gas chromatography and mass spectrometry. Chemical analysis allowed the detection of 54 compounds from the head extract and from the direct extraction of the defensive secretion of soldiers, and the percentage of common substances and the presence of exclusive substances varied depending on the spatial distance between colonies. The profile of the chemical armament of soldiers consists basically of terpenoids: monoterpenes (45.53%-71.97 - for head extract and 57.41% - 78.56 for secretion) and sesquiterpenes (19.69% - 35.78% for head extract and 18.41% - 33.31%for secretion). In general, the main component of the chemical arsenal, regardless of the methodology used for extraction, was α-pinene (27.98-50.44%). Two chemotypes were identified based on chemical differences between populations of ecoregions with distinct spatial-environmental and climate characteristics: (1) α-pinene33%; (2) α-pinene33% for both extracts (head and secretion). The results reveal a similar pattern of chemical differentiation for soldiers in both extracts (head and secretion), with increasing differences as a function of distance between the analyzed colonies.

Bombardier beetles (Coleoptera, Carabidae, Brachininae) possess a remarkable defense mechanism where a hot chemical spray is released from the tip of their abdomen, with an audible explosive sound. To date, the repellent properties of these chemicals have been tested against a limited number of taxa, such as amphibians and insects. To investigate the impact of bombardier beetle defenses on avian predators, feeding trials were conducted using the bombardier beetle (Pheropsophus jessoensis) and the Japanese quail (Coturnix japonica), a sympatric and generalist predator. All naive, hand-reared quail attacked live beetles, indicating the absence of an innate aversion to them. However, most of the quail rejected consuming the beetles whether or not the beetles sprayed them with chemicals. Naive quail also rejected dead P. jessoensis individuals. These results support the recent hypothesis that it is not essential for P. jessoensis to spray noxious chemicals to deter predators. We also found that some of the quail exposed to live P. jessoensis remembered to avoid them for up to 5 weeks. Our results provide the first evidence of the repelling effects of bombardier beetle defense mechanisms on avian predators.

The activation of plant defense systems in response to herbivory or experimentally applied methyl jasmonate (MeJA) involves the production of chemical defense substances functioning as warning signals to repel herbivores and protect against pathogens. They also serve as signals detectable by undamaged neighboring plants, a phenomenon called plant-plant communication. We studied how altitudinal variation in temperature and timing of snowmelt affected herbivore resistance, growth and reproduction of untreated bilberry (Vaccinium myrtillus L.) 20–500 cm from MeJA-treated ramets. Across two years, responses of MeJA-treated and untreated bilberry ramets were recorded twice per season along an elevational gradient in a boreal system. At low and medium altitudes, untreated bilberry showed increased herbivore resistance and reduced growth and reproduction up to 500 cm from MeJA-treated ramets. In the warmer sites at these altitudes, the effects persisted for two years for ramets up to 100 cm from the treated ramets. At high altitudes, however, only untreated ramets growing 20–100 cm from the treated ramets showed increased resistance to insect herbivores and reduced reproduction, but these effects did not persist into the second year. Altitudinal variation in climate affected trade-offs between plant defense, growth, and reproduction. Our findings indicate that plant-plant communication is also influenced by the combination of changes in climate and time after induction. Adaptations of plants growing under increasing temperature in high-latitude environments can profoundly impact ecosystem functioning, especially where bilberry, the key plant species in the boreal system, interacts with its herbivores.

The volatile alkylpyrazines methyl‐ and methoxypyrazines (MPs) present in the reflex bleeds of coccinellid beetles such as the harlequin ladybird beetle Harmonia axyridis are important semiochemicals that function in antipredatory defense behavior. Pyrazines have also been coadapted from a primarily defensive role into pheromones that function in intraspecific communication, attraction, and aggregation behavior. However, the biosynthesis of MPs in ladybird beetles is poorly understood. Here, we tested the hypothesis that MPs could be produced by microbial symbionts in H. axyridis, which generates four different MPs. The evaluation of tissue‐specific MP production showed that MP concentrations were highest in the gut tissue and hemolymph of the beetles rather than the fat body tissue as the presumed site of MP biosynthesis. Furthermore, manipulation of gut microbiota by antibiotic‐containing diets resulted in a lower MP content in adult beetles. The analysis of the bacterial community of the digestive tract revealed the presence of bacteria of the genera Serratia and Lactococcus which are reportedly able to produce MPs. In line with the known diet‐dependent production of MP in H. axyridis, we determined that the presence or relative abundance of some of the potential MP producers (Enterococcus and Staphylococcus) is also diet‐dependent. We hypothesize a potential role of the microbiota in MP production in H. axyridis as a possible example for outsourcing the synthesis of ecologically important semiochemicals to its gut bacteria.

Toxicity is widespread among living organisms, and evolves as a multimodal phenotype. Part of this phenotype is the ability to avoid self-intoxication (autoresistance). Evolving toxin resistance can involve fitness tradeoffs, so autoresistance is often expected to evolve gradually and in tandem with toxicity, resulting in a correlation between the degrees of toxicity and autoresistance among toxic populations. We investigate this correlation in Phyllobates poison frogs, notorious for secreting batrachotoxin (BTX), a potent neurotoxin that targets sodium channels, using ancestral sequence reconstructions of BTX-sensing areas of the muscular voltage-gated sodium channel. Reconstructions suggest that BTX resistance arose at the root of Phyllobates, coinciding with the evolution of BTX secretion. After this event, little or no further evolution of autoresistance seems to have occurred, despite large increases in toxicity throughout the history of these frogs. Our results, therefore, provide no evidence in favor of an evolutionary correlation between toxicity and autoresistance, which conflicts with previous work. Future research on the functional costs and benefits of mutations putatively involved in BTX resistance, as well as their prevalence in natural populations, should shed light on the evolutionary mechanisms driving the relationship between toxicity and autoresistance in Phyllobates frogs.

Insects manifest phenotypic plasticity in their development and behavior in response to plant defenses, via molecular mechanisms that produce tissue-specific changes. Phenotypic changes might vary between species that differ in their preferred hosts and these effects could extend beyond larval stages. To test this, we manipulated the diet of southern armyworm (SAW; Spodoptera eridania) and fall armyworm (FAW; Spodoptera frugiperda) using a tomato mutant for jasmonic acid plant defense pathway (def1), and wild type plants, and then quantified gene expression of Troponin t (Tnt) and flight muscle metabolism of the adult insects. Differences in Tnt spliceform ratios in insect flight muscles correlate with changes to flight muscle metabolism and flight muscle output. We found that SAW adults reared on induced def1 plants had higher relative abundances (RA) of the A isoform of Troponin t (Tnt A) in their flight muscles; in contrast, FAW adults reared on induced def1 plants had lower RAs of Tnt A in their flight muscles compared to adults reared on def1 and controls. Although mass-adjusted flight metabolic rates showed no independent host plant effects in either species, higher flight metabolic rates in SAW correlated with increased RAs of Tnt A. Flight muscle metabolism also showed an interaction of host plants with Tnt A in both species, suggesting that host plants might be influencing flight muscle metabolic output by altering Tnt. This study illustrates how insects respond to variation in host plant's chemical defense by phenotypic modifications to their flight muscle proteins, with possible implications for dispersal.

International audience; With the rise of large herbivore populations in most northern hemisphere forests, browsing is becoming an increasingly important driver of forest regeneration dynamics. Among other processes affecting the regeneration, the concept of plant-herbivore feedback loops holds that browsed saplings are more subject to subsequent herbivory. This phenomenon is interpreted as a consequence of compensatory growth following browsing since fast growth is generally associated with higher digestibility and lower defense against herbivores. However, studies linking browsing-induced trait variations to subsequent attractiveness to herbivores are still lacking, especially in the forest context. In this study, we experimentally examine the existence of a feedback loop between oak (Quercus robur L.) and roe deer (Capreolus capreolus) and investigate its underlying morphological and chemical traits. We simulated single and repeated roe deer browsing on nursery-grown oak saplings and measured the changes in sapling height growth, lateral branching, leaf traits and winter shoot traits over two years. We conducted winter feeding trials with tame roe deer one year after the first treatment to test the effect of simulated browsing on sapling attractiveness. Simulated browsing reduced sapling height growth but had no effect on branching. Simulated browsing had no effect on leaf traits after half a year, but decreased the phenolic content and increased the fiber content of winter shoots the following winter. Contrary to our predictions, roe deer preferentially browsed control saplings over saplings previously browsed. After two years, repeated browsing promoted fast carbon acquisition leaf traits (high chlorophyll, high specific leaf area and low fiber content), reduced leaf phenolic content and increased leaf digestibility. We showed that a reduction in 1-year-old oak sapling height growth following browsing, combined with increased structural defense at the expense of chemical defense in winter shoots the following winter, was correlated with reduced browsing pressure, thereby challenging the feedback loop hypothesis. However, we also demonstrated that repeated browsing promoted fast carbon acquisition leaf traits in 2.5-year-old saplings, which tend to support the existence of a feedback loop on older and more intensively browsed saplings. As such, our study provides empirical evidence that morphological and physiological trait responses to browsing influence oak sapling attractiveness, but that the direction and magnitude of this effect depend on the ontogenic stage of the sapling and on the number of browsing events.

Elevated CO2 alters C3 plant tolerance to insect herbivory, as well as the induction kinetics of defense hormones salicylic acid (SA) and jasmonic acid (JA), but the underlying physiological mechanism causing this response is not well understood. In principle, SA could be induced under elevated CO2 by reactive oxygen signals generated in photosynthesis, ultimately influencing chemical defense. To test whether the effects of elevated CO2 on C3 plant chemical defense against herbivorous insects are modulated by photosynthesis, Arabidopsis thaliana var. Col-0 plants were grown in two 2 × 2 × 2 nested factorial combinations of ambient (400 ppm) and elevated (800 ppm) CO2, and two dimensions of light regimes comprising intensity (‘mild’ 150 μmol E m−2 s−1 vs. ‘low’ light, 75 μmol E m−2 s−1) and periodicity (‘continuous’, 150 μmol E m−2 s−1 vs. ‘dynamic’, in which lights were turned off, then on, for 15 min every 2 h). Plants were challenged with herbivore damage from third instar Trichoplusia ni (cabbage looper). Consistent with experimental predictions, elevated CO2 interacted with light as well as herbivory to induce foliar concentration of SA, while JA was suppressed. Under dynamic light, foliar content of total glucosinolates was reduced. Under combination of elevated CO2 and dynamic light, T. ni removed significantly more leaf tissue relative to control plants. The observations that CO2 and light interactively modulate defense against T. ni in A. thaliana provide an empirical argument for a role of photosynthesis in C3 plant chemical defense.

Host plant chemical composition critically shapes the performance of insect herbivores feeding on them. Some insects have become specialized on plant secondary metabolites, and even use them to their own advantage such as defense against predators. However, infection by plant pathogens can seriously alter the interaction between herbivores and their host plants. We tested whether the effects of the plant secondary metabolites, iridoid glycosides (IGs), on the performance and immune response of an insect herbivore are modulated by a plant pathogen. We used the IG-specialized Glanville fritillary butterfly Melitaea cinxia, its host plant Plantago lanceolata, and the naturally occurring plant pathogen, powdery mildew Podosphaera plantaginis, as model system. Pre-diapause larvae were fed on P. lanceolata host plants selected to contain either high or low IGs, in the presence or absence of powdery mildew. Larval performance was measured by growth rate, survival until diapause, and by investment in immunity. We assessed immunity after a bacterial challenge in terms of phenoloxidase (PO) activity and the expression of seven pre-selected insect immune genes (qPCR). We found that the beneficial effects of constitutive leaf IGs, that improved larval growth, were significantly reduced by mildew infection. Moreover, mildew presence downregulated one component of larval immune response (PO activity), suggesting a physiological cost of investment in immunity under suboptimal conditions. Yet, feeding on mildew-infected leaves caused an upregulation of two immune genes, lysozyme and prophenoloxidase. Our findings indicate that a plant pathogen can significantly modulate the effects of secondary metabolites on the growth of an insect herbivore. Furthermore, we show that a plant pathogen can induce contrasting effects on insect immune function. We suspect that the activation of the immune system toward a plant pathogen infection may be maladaptive, but the actual infectivity on the larvae should be tested.