Differences in Effects of Pyrrolizidine Alkaloids on Five Generalist Insect Herbivore Species

The evolution of the diversity in plant secondary compounds is often thought to be driven by insect herbivores, although there is little empirical evidence for this assumption. To investigate whether generalist insect herbivores could play a role in the evolution of the diversity of related compounds, we examined if (1) related compounds differ in their effects on generalists, (2) there is a synergistic effect among compounds, and (3) effects of related compounds differed among insect species. The effects of pyrrolizidine alkaloids (PAs) were tested on five generalist insect herbivore species of several genera using artificial diets or neutral substrates to which PAs were added. We found evidence that structurally related PAs differed in their effects to the thrips Frankliniella occidentalis, the aphid Myzus persicae, and the locust Locusta migratoria. The individual PAs had no effect on Spodoptera exigua and Mamestra brassicae caterpillars. For S. exigua, we found indications for synergistic deterrent effects of PAs in PA mixtures. The relative effects of PAs differed between insect species. The PA senkirkine had the strongest effect on the thrips, but had no effect at all on the aphids. Our results show that generalist herbivores could potentially play a role in the evolution and maintenance of the diversity of PAs.     

Development of an Insect Herbivore and its Pupal Parasitoid Reflect Differences in Direct Plant Defense

In nature, plants defend themselves by production of allelochemicals that are toxic to herbivores. There may be considerable genetic variation in the expression of chemical defenses because of various selection pressures. In this study, we examined the development of the small cabbage butterfly, Pieris rapae, and its gregarious pupal ectoparasitoid, Pteromalus puparum, when reared on three wild populations (Kimmeridge, Old Harry, Winspit) of cabbage, Brassica oleracea, and a Brussels sprout cultivar. Wild plant populations were obtained from seeds of plants that grow naturally along the south coast of Dorset, England. Significant differences in concentrations of allelochemicals (glucosinolates) were found in leaves of plants damaged by P. rapae. Total glucosinolate concentrations in Winspit plants, the population with the highest total glucosinolate concentration, were approximately four times higher than in the cultivar, the strain with the lowest total glucosinolate concentration. Pupal mass of P. rapae and adult body mass of Pt. puparum were highest when reared on the cultivar and lowest when developing on Kimmeridge plants, the wild strain with the lowest total glucosinolate concentration. Development of male parasitoids was also more negatively affected than female parasitoids. Our results reveal that plant quality, at least for the development of ‘adapted’ oligophagous herbivores, such as P. rapae, is not based on total glucosinolate content. The only glucosinolate compound that corresponded with the performance of P. rapae was the indole glucosinolate, neoglucobrassicin. Our results show that performance of ectoparasitoids may closely reflect constraints on the development of the host.     

Response of a Generalist Herbivore <Emphasis Type="Italic">Trichoplusia ni</Emphasis> to Jasmonate-Mediated Induced Defense in Tomato

The up-regulation of plant defense-related toxins or metabolic enzyme binding proteins often leads to a negative effect on herbivorous insects. These negative effects can manifest themselves at three points: changes in food ingestion, post-ingestive-changes, and post-digestive changes. Many studies have related the decrease in herbivore growth and/or survival with expression of chemicals that interfere with post-digestive effects such as the anti-nutritive effects of protease inhibitors. Nevertheless, it is unclear whether such compounds impact herbivores via earlier ingestive processes. We addressed this question by using a jasmonate-deficient mutant (Def-1), a jasmonate-overexpressor mutant (Prosystemin or Prosys), and wild-type tomato in three trials with 5th instar Trichoplusia ni. Decreases in relative growth rate (RGR) confirmed that T. ni fed on the Prosys plants developed poorly compared to those feeding on Def-1 plants (larvae on wild-types were intermediate). Preingestive and postingestive processes contributed to this effect. Total food ingested and the consumptive index were 25% lower on Prosys plants compared to Def-1 plants. Post-ingestive processes, measured by approximate digestibility, were 62% greater on Prosy plants. Post-digestive efficiency measured by conversion of ingested and digested food (ECI and ECD) decreased on Prosys plants two-fold compared to Def-1 plants. This post-digestive interference correlated well with the 2-fold decrease in activity of digestive enzymes, serine proteases, in Prosys-fed T. ni compared to those on Def-1 plants. No difference in detoxifying enzyme activity was detected.     

Effects of Arbuscular Mycorrhiza on Plant Chemistry and the Development and Behavior of a Generalist Herbivore

Arbuscular mycorrhiza (AM) formed between plants and AM fungi (AMF) can alter host plant quality and thus influence plant-herbivore interactions. While AM is known to affect the development of generalist chewing-biting herbivores, AM-mediated impacts on insect behavior have been neglected until now. In this study, the effects of Rhizophagus irregularis, a generalist AMF, on phenotypic and leaf metabolic traits of Plantago major plants were investigated. Further, the influence of AM-mediated host plant modifications on the development and on seven behavioral traits of larvae of the generalist Mamestra brassicae were recorded. Tests were carried out in the third (L3) and fourth (L4) larval instar, respectively. While shoot water content, specific leaf area, and foliar concentrations of the secondary metabolite aucubin were higher in AM-treated compared to non-mycorrhized (NM) plants, lower concentrations of the primary metabolites citric acid and isocitric acid were found in leaves of AM plants. Larvae reared on AM plants gained a higher body mass and tended to develop faster than individuals reared on NM plants. However, plant treatment had no significant effect on any of the behavioral traits. Instead, differences between larvae of different ages were detected in several behavioral features, with L4 being less active and less bold than L3 larvae. The results demonstrate that AM-induced modifications of host plant quality influence larval development, whereas the behavioral phenotype seems to be more fixed at least under the tested conditions.     

Density-Dependent Reduction and Induction of Milkweed Cardenolides by a Sucking Insect Herbivore

The effect of aphid population size on host-plant chemical defense expression and the effect of plant defense on aphid population dynamics were investigated in a milkweed-specialist herbivore system. Density effects of the aposematic oleander aphid, Aphis nerii, on cardenolide expression were measured in two milkweed species, Asclepias curassavica and A. incarnata. These plants vary in constitutive chemical investment with high mean cardenolide concentration in A. curassavica and low to zero in A. incarnata. The second objective was to determine whether cardenolide expression in these two host plants impacts mean A. nerii colony biomass (mg) and density. Cardenolide concentration (microgram/g) of A. curassavica in both aphid-treated leaves and opposite, herbivore-free leaves decreased initially in comparison with aphid-free controls, and then increased significantly with A. nerii density. Thus, A. curassavica responds to aphid herbivory initially with density-dependent phytochemical reduction, followed by induction of cardenolides to concentrations above aphid-free controls. In addition, mean cardenolide concentration of aphid-treated leaves was significantly higher than that of opposite, herbivore-free leaves. Therefore, A. curassavica induction is strongest in herbivore-damage tissue. Conversely, A. incarnata exhibited no such chemical response to aphid herbivory. Furthermore, neither host plant responded chemically to herbivore feeding duration time (days) or to the interaction between herbivore initial density and feeding duration time. There were also no significant differences in mean colony biomass or population density of A. nerii reared on high cardenolide (A. curassavica) and low cardenolide (A. incarnata) hosts.     

Induced Immunity Against Belowground Insect Herbivores- Activation of Defenses in the Absence of a Jasmonate Burst

Roots respond dynamically to belowground herbivore attack. Yet, little is known about the mechanisms and ecological consequences of these responses. Do roots behave the same way as leaves, or do the paradigms derived from aboveground research need to be rewritten? This is the central question that we tackle in this article. To this end, we review the current literature on induced root defenses and present a number of experiments on the interaction between the root herbivore Diabrotica virgifera and its natural host, maize. Currently, the literature provides no clear evidence that plants can recognize root herbivores specifically. In maize, mild mechanical damage is sufficient to trigger a root volatile response comparable to D. virgifera induction. Interestingly, the jasmonate (JA) burst, a highly conserved signaling event following leaf attack, is consistently attenuated in the roots across plant species, from wild tobacco to Arabidopsis. In accordance, we found only a weak JA response in D. virgifera attacked maize roots. Despite this reduction in JA-signaling, roots of many plants start producing a distinct suite of secondary metabolites upon attack and reconfigure their primary metabolism. We, therefore, postulate the existence of additional, unknown signals that govern induced root responses in the absence of a jasmonate burst. Surprisingly, despite the high phenotypic plasticity of plant roots, evidence for herbivore-induced resistance below ground is virtually absent from the literature. We propose that other defensive mechanisms, including resource reallocation and compensatory growth, may be more important to improve plant immunity below ground.     

Polymorphism in Plant Defense Against Herbivory: Constitutive and Induced Resistance in Cucumis sativus

Theory predicts that plant resistance to herbivores is determined by both genetic and environmentally induced components. In this study, we demonstrate that the phenotypic expression of plant resistance to spider mite herbivory in Cucumis sativus is determined by genetic and environmental factors and that there is an interaction between these factors. Previous feeding by spider mites induced systemic resistance to subsequent attack over several spatial scales within plants, reducing the population growth of mites compared to that on control plants. Effects of induction were effective locally over the short term, but resulted in local increased susceptibility to spider mite attack after several days. However, this local induced susceptibility on the damaged leaf was associated with induced resistance on newer leaves. Induced resistance was correlated with increases in cucurbitacin content of leaves, but was not associated with changes in the density of leaf trichomes. Induced resistance to herbivory was not detected in plants of a genotype lacking constitutive expression of cucurbitacins, which were in general highly susceptibile to mite attack. Allocation trade-offs between growth and defense are often invoked to explain the maintenance of variation in the levels of plant resistance. Contrary to current thinking, neither constitutive nor herbivore-induced plant resistance were associated with reductions in plant allocation to root and shoot growth. However, plants that had high levels of induced resistance to spider mites were the most susceptible to attack by a specialist beetle. Such ecological trade-offs between resistance to generalist herbivores and susceptibility to specialist herbivores may be important in the maintenance of variation of plant resistance traits. In summary, C. sativus exhibits strong genetic variation for constitutive and induced resistance to spider mites, and this variation in resistance is associated with ecological trade-offs.     

No Major Role for Binding by Salivary Proteins as a Defense Against Dietary Tannins in Mediterranean Goats

We investigated whether Mediterranean goats use salivary tannin-binding proteins to cope with tannin-rich forages by determining the affinity of salivary or parotid gland proteins for tannic acid or quebracho tannin. Mixed saliva, sampled from the oral cavity, or parotid gland contents were compared to the intermediate affinity protein bovine serum albumin with a competitive binding assay. Goats that consume tannin-rich browse (Damascus) and goats that tend to avoid tannins (Mamber) were sequentially fed high (Pistacia lentiscus L.), low (vetch hay), or zero (wheat hay) tannin forages. Affinity of salivary proteins for tannins did not differ between goat breeds and did not respond to presence or absence of tannins in the diet. Proteins in mixed saliva had slightly higher affinity for tannins than those in parotid saliva, but neither source contained proteins with higher affinity for tannins than bovine serum albumin. Similarly, 3 months of browsing in a tannin-rich environment had little effect on the affinity of salivary proteins for tannin in adult goats of either breed. We sampled mixed saliva from young kids before they consumed forage and after 3 months of foraging in a tannin-rich environment. Before foraging, the saliva of Mamber kids had higher affinity for tannic acid (but not quebracho tannin) than the saliva of Damascus kids, but there was no difference after 3 months of exposure to tannin-rich browse, and the affinity of the proteins was always similar to the affinity of bovine serum albumin. Our results suggest there is not a major role for salivary tannin-binding proteins in goats. Different tendencies of goat breeds to consume tannin-rich browse does not appear be related to differences in salivary tannin-binding proteins.     

Generalization of a Habituated Feeding Deterrent Response to Unrelated Antifeedants Following Prolonged Exposure in a Generalist Herbivore, Trichoplusia ni

The possibility of generalization of habituated response to unrelated feeding deterrents following prolonged exposure was examined in third instar Trichoplusia ni (Lepidoptera: Noctuidae) larvae by rearing them on antifeedants and then testing with other unrelated antifeedants. We introduced neonate larvae (<24-hr old) onto cabbage leaves treated with crude seed extracts of Melia volkensii (Meliaceae) or oil of Origanum vulgare (“oregano”) (Lamiaceae) and allowed them to feed until early in the third instar. Naïve larvae were reared on cabbage leaves treated with carrier solvent alone. Both experienced and naïve larvae were tested for feeding deterrent response with the same and the different extracts in a leaf disc choice bioassay. Habituation was generalized to both M. volkensii and oregano following prolonged exposure to either plant extract and also to a pure allelochemical, thymol, following prolonged exposure to either digitoxin or xanthotoxin. However, there was no generalization of the habituated response to oregano following prolonged exposure to digitoxin or thymol, or to thymol or xanthotoxin following prolonged exposure to oregano or M. volkensii. Our results demonstrate that habituated response to feeding deterrents in a polyphagous insect herbivore can be generalized among and between plant extracts and pure allelochemicals, but not in all situations. The implications of such behavioral plasticity in herbivorous insects for the use of antifeedants as crop protectants or for host plant shifts is discussed.     

Specific Polyphenols and Tannins are Associated with Defense Against Insect Herbivores in the Tropical Oak Quercus oleoides

The role of plant polyphenols as defenses against insect herbivores is controversial. We combined correlative field studies across three geographic regions (Northern Mexico, Southern Mexico, and Costa Rica) with induction experiments under controlled conditions to search for candidate compounds that might play a defensive role in the foliage of the tropical oak, Quercus oleoides. We quantified leaf damage caused by four herbivore guilds (chewers, skeletonizers, leaf miners, and gall forming insects) and analyzed the content of 18 polyphenols (including hydrolyzable tannins, flavan-3-ols, and flavonol glycosides) in the same set of leaves using high performance liquid chromatography and mass spectrometry. Foliar damage ranged from two to eight percent per region, and nearly 90% of all the damage was caused by chewing herbivores. Damage due to chewing herbivores was positively correlated with acutissimin B, catechin, and catechin dimer, and damage by mining herbivores was positively correlated with mongolinin A. By contrast, gall presence was negatively correlated with vescalagin and acutissimin B. By using redundancy analysis, we searched for the combinations of polyphenols that were associated to natural herbivory: the combination of mongolinin A and acutissimin B had the highest association to herbivory. In a common garden experiment with oak saplings, artificial damage increased the content of acutissimin B, mongolinin A, and vescalagin, whereas the content of catechin decreased. Specific polyphenols, either individually or in combination, rather than total polyphenols, were associated with standing leaf damage in this tropical oak. Future studies aimed at understanding the ecological role of polyphenols can use similar correlative studies to identify candidate compounds that could be used individually and in biologically meaningful combinations in tests with herbivores and pathogens.     

Influences of Plant Toxins and Their Spatial Distribution on Foraging by the Common Brushtail Possum, a Generalist Mammalian Herbivore

Generalist herbivores forage on a variety of plant species, allowing them to gain nutrients while limiting ingestion of harmful toxins. As the capacity to mix diets appears important for maximizing intake, the spatial scale in heterogeneity of food resources should influence the foraging behavior of herbivores. Our aim was to identify how the feeding strategy of a generalist mammalian herbivore, the common brushtail possum, responds to foods within a spatially defined environment. We evaluated foraging responses against increasing spatial separation between pairs of artificial diets that differed in flavor and toxin profile, to determine how distance and diet affect intake and behavior. Foraging responses were influenced by the type of diet or the degree of spatial separation between foods but not by their interaction. Diet influenced intake, time spent feeding, and feeding rate, but had no effect on nightly foraging interval, number of feeding bouts, or bout length. The number of switches between paired food resources and foraging efficiency (intake per unit distance, which accounts for the energetic costs of travelling), were influenced only by distance. Titrating foraging against a range of distances demonstrated how quickly foraging efficiency can decline in response to the spatial separation of food resources, highlighting the importance of spatial heterogeneity of plants within the home range of an herbivore.     

Trade-Offs in Plant Defense Against Pathogens and Herbivores: A Field Demonstration of Chemical Elicitors of Induced Resistance

Two signaling pathways, one involving salicylic acid and another involving jasmonic acid, participate in the expression of plant resistance to pathogens and insect herbivores. In this study, we report that stimulation of systemic acquired resistance in field-grown tomato plants with the salicylate mimic, benzothiadiazole: (1) attenuates the jasmonate-induced expression of the antiherbivore defense-related enzyme polyphenol oxidase, and (2) compromises host-plant resistance to larvae of the beet armyworm, Spodoptera exigua. Conversely, treatment of plants with jasmonic acid at concentrations that induce resistance to insects reduces pathogenesis-related protein gene expression induced by benzothiadiazole, and partially reverses the protective effect of benzothiadiazole against bacterial speck disease caused by Pseudomonas syringae pv. tomato. We conclude that effective utilization of induced plant resistance to the multiple pests typically encountered in agriculture will require understanding potential signaling conflicts in plant defense responses.     

The Significance of Ectomycorrhizas in Chemical Quality of Silver Birch Foliage and Above-Ground Insect Herbivore Performance

We tested whether the ectomycorrhizal (ECM) infection level of roots of silver birch (Betula pendula) affects performance of above-ground insect herbivores by increasing available plant biomass, by enhancing availability of nutrients, or by modifying concentration of defense compounds, i.e., phenolics, in birch foliage. Insect performance was determined for a phloem-feeding generalist (Lygus rugulipennis, the European tarnished plant bug), a phloem-feeding specialist (Calaphis flava, the birch aphid), and a chewing generalist (Epirrita autumnata, the autumnal moth larva). Silver birch plantlets had either natural ECM infection level (on average 24% of short roots with ECM), reduced ECM infection level with fungicide (F−, 9% ECM), or enhanced ECM infection level after inoculation with the fungus Paxillus involutus (PI+, 45% ECM) or Leccinum versipelle (LV+, 42% ECM). In general, the most pronounced effect of ECM was observed on growth of plantlets, i.e., stem growth decreased. In PI+-treated plants, leaf biomass also decreased. The effect of mycorrhizal colonization on the host plant’s nitrogen (N) and phosphorous (P) concentration was dependent on the mycorrhizal species and experiment. Fungicide treatment did not cause a consistent decrease in nutrients. Finally, defense of birch against herbivory, expressed as foliar phenolic concentration in plantlets, was not modified by ECM. However, E. autumnata had a significantly higher relative growth rate on PI+ plantlets with high leaf N concentration than on LV+ plantlets with low leaf N concentration. The birch aphid C. flava produced significantly less nymphs on birches with enhanced ECM infection levels (PI+ and LV+ plantlets) than on controls. In summary, our data show that the ECM infection level mainly affects the growth parameters of plantlets, whereas effects on leaf chemical quality are minor. Our data show that effects of ECM infection of birch roots on aboveground herbivores are multifaceted and depend on the fungal species forming ectomycorrhiza and also on the degree of specialization and feeding guild of insects.     

Intraspecific Variation in Plant Defense Alters Effects of Root Herbivores on Leaf Chemistry and Aboveground Herbivore Damage

Root herbivores can indirectly affect aboveground herbivores by altering the food quality of the plant. However, it is largely unknown whether plant genotypes differ in their response to root herbivores, leading to variable defensive phenotypes. In this study, we investigated whether root-feeding insect larvae (Agriotes sp. larvae, wireworms) induce different responses in Plantago lanceolata plants from lines selected for low and high levels of iridoid glycosides (IG). In the absence of wireworms, plants of the “high-IG line” contained approximately twofold higher levels of total IG and threefold higher levels of catalpol (one of the IG) in leaves than plants from the “low-IG line,” whereas both lines had similar levels of IG in roots. In response to wireworms, roots of plants from both lines showed increased concentrations of catalpol. Leaves of “low-IG line” plants increased catalpol concentrations in response to wireworms, whereas catalpol concentrations of leaves of “high-IG line” plants decreased. In contrast, glucose concentrations in roots of “low-IG” plants decreased, while they increased in “high-IG” plants after feeding by wireworms. The leaf volatile profile differed between the lines, but was not affected by root herbivores. In the field, leaf damage by herbivores was higher in wireworm-induced compared to noninduced “low-IG” plants and lower in wireworm-induced compared to noninduced “high-IG” plants, despite induction of catalpol in leaves of the “low-IG” plants and reduction in “high-IG” plants. This pattern might arise if damage is caused mainly by specialist herbivores for which catalpol may act as feeding stimulant rather than as deterrent. The present study documents for the first time that intraspecific variation in plant defense affects the outcome of plant-mediated interactions between root and shoot herbivores.     

Effects of Quantitative Variation in Allelochemicals in <Emphasis Type="Italic">Plantago lanceolata</Emphasis> on Development of a Generalist and a Specialist Herbivore and their Endoparasitoids

Studies in crop species show that the effect of plant allelochemicals is not necessarily restricted to herbivores, but can extend to (positive as well as negative) effects on performance at higher trophic levels, including the predators and parasitoids of herbivores. We examined how quantitative variation in allelochemicals (iridoid glycosides) in ribwort plantain, Plantago lanceolata, affects the development of a specialist and a generalist herbivore and their respective specialist and generalist endoparasitoids. Plants were grown from two selection lines that differed ca. 5-fold in the concentration of leaf iridoid glycosides. Development time of the specialist herbivore, Melitaea cinxia, and its solitary endoparasitoid, Hyposoter horticola, proceeded most rapidly when reared on the high iridoid line, whereas pupal mass in M. cinxia and adult mass in H. horticola were unaffected by plant line. Cotesia melitaearum, a gregarious endoparasitoid of M. cinxia, performed equally well on hosts feeding on the two lines of P. lanceolata. In contrast, the pupal mass of the generalist herbivore, Spodoptera exigua, and the emerging adult mass of its solitary endoparasitoid, C. marginiventris, were significantly lower when reared on the high line, whereas development time was unaffected. The results are discussed with regards to (1) differences between specialist and generalist herbivores and their natural enemies to quantitative variation in plant secondary chemistry, and (2) potentially differing selection pressures on plant defense.     

Developmental and Geographical Variation in the Chemical Defense of the Walkingstick Insect Anisomorpha buprestoides

Anisomorpha buprestoides, a walkingstick common in the southeastern United States, sprays chemicals that irritate and repel threatening insects, birds, or mammals. The active chemical in this substance was initially identified as a monoterpene dialdehyde. This compound can be present in several stereoisomeric forms, and subsequent studies have revealed that A. buprestoides produces at least three diastereomers: anisomorphal, dolichodial, and peruphasmal. However, no inquiry has been made to date into the geographical or developmental dependence of this variation. We report here that different populations of adult A. buprestoides spray either anisomorphal, or peruphasmal, or a mixture of the two stereoisomers. Additionally, offspring of a peruphasmal-producing population produced a variable mixture of anisomorphal and dolichodial but switched to peruphasmal upon reaching sexual maturity. This appears to be the first report of a developmentally regulated change in walkingstick insect chemical defense. Our results suggest a more complex role of these substances in the overall chemical ecology of walkingstick insects. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Formation of Simple Nitriles upon Glucosinolate Hydrolysis Affects Direct and Indirect Defense Against the Specialist Herbivore, <Emphasis Type="Italic">Pieris rapae</Emphasis>

The glucosinolate-myrosinase system, found in plants of the order Brassicales, has long been considered an effective defense system against herbivores. The defensive potential of glucosinolates is mainly due to the products formed after myrosinase-catalyzed hydrolysis upon tissue damage. The most prominent hydrolysis products, the isothiocyanates, are toxic to a wide range of organisms, including herbivorous lepidopterans. In contrast, little is known about the biological activities of alternative hydrolysis products such as simple nitriles and epithionitriles that are formed at the expense of isothiocyanates in the presence of epithiospecifier proteins (ESPs). Here, we used transgenic Arabidopsis thaliana (Brassicaceae) plants overexpressing ESP (35S:ESP plants) to investigate the effects of simple nitriles on direct and indirect defense against the specialist cabbage white butterfly Pieris rapae L. (Lepidoptera, Pieridae). In the 35S:ESP plants, glucosinolates are hydrolyzed mainly to simple nitriles upon tissue disruption, while isothiocyanates are the predominant hydrolysis products in Columbia-0 (Col-0) wild-type plants. The parasitoid Cotesia rubecula (Hymenoptera, Braconidae), a specialist on P. rapae larvae, was significantly more attracted to P. rapae-infested 35S:ESP plants than to P. rapae-infested Col-0 wild-type plants in a wind tunnel setup. Furthermore, female P. rapae butterflies laid more eggs on Col-0 wild-type plants than on 35S:ESP plants when the plants had been damaged previously. However, when given a choice to feed on 35S:ESP or Col-0 plants, caterpillars did not discriminate between the two genotypes. Growth rate and developmental time were not significantly different between caterpillars that were reared on 35S:ESP or Col-0 plants. Thus, the production of simple nitriles instead of isothiocyanates, as catalyzed by ESP, can promote both direct and indirect defense against the specialist herbivore P. rapae.     

Characterization of Mutations Causing CYP21A2 Deficiency in Brazilian and Portuguese Populations

Deficiency of 21-hydroxylase enzyme (CYP21A2) represents 90% of cases in congenital adrenal hyperplasia (CAH), an autosomal recessive disease caused by defects in cortisol biosynthesis. Computational prediction and functional studies are often the only way to classify variants to understand the links to disease-causing effects. Here we investigated the pathogenicity of uncharacterized variants in the CYP21A2 gene reported in Brazilian and Portuguese populations. Physicochemical alterations, residue conservation, and effect on protein structure were accessed by computational analysis. The enzymatic performance was obtained by functional assay with the wild-type and mutant CYP21A2 proteins expressed in HEK293 cells. Computational analysis showed that p.W202R, p.E352V, and p.R484L have severely impaired the protein structure, while p.P35L, p.L199P, and p.P433L have moderate effects. The p.W202R, p.E352V, p.P433L, and p.R484L variants showed residual 21OH activity consistent with the simple virilizing phenotype. The p.P35L and p.L199P variants showed partial 21OH efficiency associated with the non-classical phenotype. Additionally, p.W202R, p.E352V, and p.R484L also modified the protein expression level. We have determined how the selected CYP21A2 gene mutations affect the 21OH activity through structural and activity alteration contributing to the future diagnosis and management of CYP21A2 deficiency.     

Differential Response of a Local Population of Entomopathogenic Nematodes to Non-Native Herbivore Induced Plant Volatiles (HIPV) in the Laboratory and Field

Recent work has shown the potential for enhanced efficacy of entomopathogenic nematodes (EPN) through their attraction to herbivore induced plant volatiles. However, there has been little investigation into the utilization of these attractants in systems other than in those in which they were identified. We compared (E)-β-caryophyllene and pregeijerene in the highbush blueberry (Vaccinium corymbosum) agroecosystem in their ability to enhance the attraction of EPN to and efficacy against the system’s herbivore, oriental beetle (Anomala orientalis). The relative attractiveness of (E)-β-caryophyllene and pregeijerene to a local isolate of the EPN species Steinernema glaseri was tested in a six-arm olfactometer in the laboratory to gather baseline values of attraction to the chemicals alone in sand substrate before field tests. A similar arrangement was used in a V. corymbosum field by placing six cages with assigned treatments and insect larvae with and without compound into the soil around the base of 10 plants. The cages were removed after 72 h, and insect baits were retrieved and assessed for EPN infection. The lab results indicate that in sand alone (E)-β-caryophyllene is significantly more attractive than pregeijerene to the local S. glaseri isolate Conversely, there was no difference in attractiveness in the field study, but rather, native S. glaseri were more attracted to cages with G. mellonella larvae, no larvae, and cages with the blank control and G. mellonella larvae.