Pre-egg hatch efficacy of dormant applications against Lymantria dispar (Lepidoptera: Erebidae).

Lymantria dispar L. is an invasive, non-native defoliating Lepidopteran established in North America that feeds on forest and urban trees. While many products are available to manage L. dispar post-emergence, few exist to prevent egg hatch when applied to egg masses. Here, we present the results of 3 separate experiments aimed at determining the efficacy of pre-emergent insecticides against L. dispar egg hatch. We found that the labeled rate (1:1) of Golden Pest Spray Oil (GPSO; AI: 93% soybean oil) can prevent L. dispar larvae from emerging in both field and lab assays. In large public spaces, we found that this treatment was ineffective at preventing L. dispar emergence or defoliation. Acelepryn (AI: 18.4% chlorantraniliprole) resulted in some suppression of egg hatch at a very low rate (.06 ml/ 3.8 liter) in both lab and field settings and the efficacy of higher rates should be further investigated. We also tested GPSO against Lepitect (97.4% acephate) in a public area that also received a Foray 48B (12.65% Bacillus thuringiensis, subsp. kurstaki) aerial application. On large oak trees in public areas, GPSO and Lepitect were not effective at reducing defoliation. Dormant pesticide applications generally reduce the risk of affecting negatively predator and parasitoid communities and are therefore desirable. Lymantria dispar pre-egg hatch applications will not work in every situation but should be considered as part of an integrated pest management (IPM) strategy for individual homeowner trees where thorough coverage can be obtained.

Impact of chronic stylet-feeder infestation on folivore-induced signaling and defenses in a conifer.

Our understanding of how conifers respond biochemically to multiple simultaneous herbivore attacks is lacking. Eastern hemlock (Tsuga canadensis; 'hemlock') is fed on by hemlock woolly adelgid (Adelges tsugae; 'adelgid') and by later-instar gypsy moth (Lymantria dispar; 'gypsy moth') caterpillars. The adelgid is a stylet-feeding insect that causes a salicylic acid (SA)-linked response in hemlock, and gypsy moth larvae are folivores that presumably cause a jasmonic acid (JA)-linked response. This system presents an opportunity to study how invasive herbivore-herbivore interactions mediated through host biochemical responses. We used a factorial field experiment to challenge chronically adelgid-infested hemlocks with gypsy moth caterpillars. We quantified 17 phytohormones, 26 phenolic and terpene metabolites, and proanthocyanidin, cell wall-bound (CW-bound) phenolic, and lignin contents. Foliage infested with adelgid only accumulated gibberellins and SA; foliage challenged by gypsy moth only accumulated JA phytohormones. Gypsy moth folivory on adelgid-infested foliage reduced the accumulation of JA phytohormones and increased the SA levels. Both herbivores increased CW-bound phenolics and gypsy moth increased lignin content when feeding alone but not when feeding on adelgid-infested foliage. Our study illustrates the importance of understanding the biochemical mechanisms and signaling antagonism underlying tree responses to multiple stresses and of disentangling local and systemic stress signaling in trees.

Impact of Hemlock Woolly Adelgid (Hemiptera: Adelgidae) Infestation on the Jasmonic Acid-Elicited Defenses of Tsuga canadensis (Pinales: Pinaceae).

Hemlock woolly adelgid is an invasive piercing-sucking insect in eastern North America, which upon infestation of its main host, eastern hemlock ('hemlock'), improves attraction and performance of folivorous insects on hemlock. This increased performance may be mediated by hemlock woolly adelgid feeding causing antagonism between the the jasmonic acid and other hormone pathways. In a common garden experiments using hemlock woolly adelgid infestation and induction with methyl jasmonate (MeJA) and measures of secondary metabolite contents and defense-associated enzyme activities, we explored the impact of hemlock woolly adelgid feeding on the local and systemic induction of jasmonic acid (JA)-elicited defenses. We found that in local tissue hemlock woolly adelgid or MeJA exposure resulted in unique induced phenotypes, whereas the combined treatment resulted in an induced phenotype that was a mixture of the two individual treatments. We also found that if the plant was infested with hemlock woolly adelgid, the systemic response of the plant was dominated by hemlock woolly adelgid, regardless of whether MeJA was applied. Interestingly, in the absence of hemlock woolly adelgid, hemlock plants had a very weak systemic response to MeJA. We conclude that hemlock woolly adelgid infestation prevents systemic induction of JA-elicited defenses. Taken together, compromised local JA-elicited defenses combined with weak systemic induction could be major contributors to increased folivore performance on hemlock woolly adelgid-infested hemlock.

Light Limitation Impacts Growth but Not Constitutive or Jasmonate Induced Defenses Relevant to Emerald Ash Borer (Agrilus planipennis) in White Fringetree (Chionanthus virginicus) or Black Ash (Fraxinus nigra).

White fringetree is a host for the invasive emerald ash borer (EAB) but is of lower quality than the related and highly susceptible black ash. Field observations suggest that host trees grown in full sun are more resistant to EAB than those in shade, however the impact of light limitation on chemical defenses has not been assessed. We quantified constitutive and jasmonate-induced phloem defenses and growth patterns of white fringetree and black ash under differential light conditions and related them to EAB larval performance. White fringetree had significantly lower constitutive and induced activities of peroxidase, polyphenol oxidase, ß-glucosidase, chitinase and lignin content, but significantly higher gallic acid equivalent soluble phenolic, soluble sugar, and oleuropein concentrations compared to black ash. Multivariate analyses based on tissue chemical attributes displayed clear separation of species and induced defense responses. Further, EAB performed significantly worse on white fringetree than black ash, consistent with previous studies. Light limitation did not impact measured defenses or EAB larval performance, but it did decrease current year growth and increase photosynthetic efficiency. Overall our results suggest that phenolic profiles, metabolite abundance, and growth traits are important in mediating white fringetree resistance to EAB, and that short-term light limitation does not influence phloem chemistry or larval success.

Chronic impacts of invasive herbivores on a foundational forest species: a whole-tree perspective.

Forests make up a large portion of terrestrial plant biomass, and the long-lived woody plants that dominate them possess an array of traits that deter consumption by forest pests. Although often extremely effective against native consumers, invasive species that avoid or overcome these defenses can wreak havoc on trees and surrounding ecosystems. This is especially true when multiple invasive species co-occur, since interactions between invasive herbivores may yield non-additive effects on the host. While the threat posed by invasive forest pests is well known, long-term field experiments are necessary to explore these consumer-host interactions at appropriate spatial and temporal scales. Moreover, it is important to measure multiple variables to get a "whole-plant" picture of their combined impact. We report the results of a 4-yr field experiment addressing the individual and combined impacts of two invasive herbivores, the hemlock woolly adelgid (Adelges tsugae) and elongate hemlock scale (Fiorinia externa), on native eastern hemlock (Tsuga canadensis) in southern New England. In 2011, we planted 200 hemlock saplings into a temperate forest understory and experimentally manipulated the presence/absence of both herbivore species; in 2015, we harvested the 88 remaining saplings and assessed plant physiology, growth, and resource allocation. Adelgids strongly affected hemlock growth: infested saplings had lower above/belowground biomass ratios, more needle loss, and produced fewer new needles than control saplings. Hemlock scale did not alter plant biomass allocation or growth, and its co-occurrence did not alter the impact of adelgid. While both adelgid and scale impacted the concentrations of primary metabolites, adelgid effects were more pronounced. Adelgid feeding simultaneously increased free amino acids local to feeding sites and a ~30% reduction in starch. The cumulative impact of adelgid-induced needle loss, manipulation of nitrogen pools, and the loss of stored resources likely accelerates host decline through disruption of homeostatic source-sink dynamics occurring at the whole-plant level. Our research stresses the importance of considering long-term impacts to predict how plants will cope with contemporary pressures experienced in disturbed forests.

Variation in the Volatile Profiles of Black and Manchurian Ash in Relation to Emerald Ash Borer Oviposition Preferences.

Emerald ash borer (EAB; Agrilus planipennis) is a devastating pest of ash (Fraxinus spp.) in its invaded range in North America. Its coevolved Asian hosts are more resistant and less preferred for oviposition than susceptible North American species. We compared EAB oviposition preferences and bark and canopy volatile organic compound (VOC) emissions of resistant Manchurian ash and susceptible black ash, and examined relationships between VOC profiles and oviposition. In the field, black ash was highly preferred for oviposition while no eggs were laid on Manchurian ash, and we found clear differences in the VOC profiles of Manchurian and black ash. We detected 78 compounds emitted from these species, including 16 compounds that elicited EAB antennal activity in prior studies. Four compounds were unique to black and 11 to Manchurian ash. Emission rates of 14 canopy and 19 bark volatiles varied among the two species, including four previously reported as antennally active. Specifically, 7-epi-sesquithujene (bark) emissions were greater from black ash, while ß-caryophyllene (canopy), linalool (bark), and α-cubebene (bark) were emitted at higher rates by Manchurian ash. No relationships were found between the emission rate of any single compound or group of compounds (e.g. monoterpenes) suggesting that preference may be based on complex profile combinations. This is the first study to directly compare VOCs of black and Manchurian ash as well as the first to examine bark- and canopy-specific VOCs. The unique bark and canopy VOC profiles of these two species implicates potentially important variation in VOCs between a closely related resistant and susceptible species that provides a foundation for future studies of host preferences of EAB.

Feeding and Development of Emerald Ash Borer (Coleoptera: Buprestidae) on Cultivated Olive, Olea europaea.

We examined the suitability of cultivated olive, Olea europaea L., as a host for emerald ash borer, Agrilus planipennis Fairmaire. In a bioassay using cut stems from a field-grown olive tree (cv. 'Manzanilla') we found that 45% of larvae that had emerged from eggs used to inoculate stems, were recovered alive, many as larvae or prepupae, during periodic debarking of a subset of stems. Three intact stems that 19 larvae successfully entered were exposed to a simulated overwintering treatment. Four live adults emerged afterwards, and an additional pupa and several prepupae were discovered after debarking these stems. Cultivated olive joins white fringetree as one of the two species outside of the genus Fraxinus capable of supporting the development of emerald ash borer from neonate to adult.

Pretty Picky for a Generalist: Impacts of Toxicity and Nutritional Quality on Mantid Prey Processing.

Prey have evolved a number of defenses against predation, and predators have developed means of countering these protective measures. Although caterpillars of the monarch butterfly, Danaus plexippus L., are defended by cardenolides sequestered from their host plants, the Chinese mantid Tenodera sinensis Saussure guts the caterpillar before consuming the rest of the body. We hypothesized that this gutting behavior might be driven by the heterogeneous quality of prey tissue with respect to toxicity and/or nutrients. We conducted behavioral trials in which mantids were offered cardenolide-containing and cardenolide-free D. plexippus caterpillars and butterflies. In addition, we fed mantids starved and unstarved D. plexippus caterpillars from each cardenolide treatment and nontoxic Ostrinia nubilalis Hübner caterpillars. These trials were coupled with elemental analysis of the gut and body tissues of both D. plexippus caterpillars and corn borers. Cardenolides did not affect mantid behavior: mantids gutted both cardenolide-containing and cardenolide-free caterpillars. In contrast, mantids consumed both O. nubilalis and starved D. plexippus caterpillars entirely. Danaus plexippus body tissue has a lower C:N ratio than their gut contents, while O. nubilalis have similar ratios; gutting may reflect the mantid's ability to regulate nutrient uptake. Our results suggest that post-capture prey processing by mantids is likely driven by a sophisticated assessment of resource quality.

Impact of Consuming 'Toxic' Monarch Caterpillars on Adult Chinese Mantid Mass Gain and Fecundity.

Predators that feed on chemically-defended prey often experience non-lethal effects that result in learned avoidance of the prey species. Some predators are able to consume toxic prey without ill-effect. The Chinese mantid is able to consume cardenolide-containing monarch caterpillars without immediate adverse effects. Although they discard the caterpillars' gut contents, mantids consume sequestered cardenolides. Although consumption of these cardenolides does not elicit an acute response, there may be long-term costs associated with cardenolide consumption. We tested the hypothesis that consumption of monarch caterpillars will adversely affect adult mantid biomass gain and reproductive condition. We reared mantids from egg to adult and assigned them to one of four toxicity groups that differed in the number of monarch caterpillars offered over a 15-day period. Mantids consumed similar amounts of prey biomass during the experiment. Yet, mantids in the high-toxicity group had a higher conversion efficiency and gained more biomass than mantids in other groups. Mantids in all treatment groups produced similar numbers of eggs. However, mantids in the high-toxicity group produced heavier eggs and devoted a greater portion of their biomass toward egg production than those in the control group. This increase in reproductive condition is probably driven by variation in prey nutritional value and/or the nutritional advantages inherent in eating multiple food types. Our results demonstrate the mantid is able to incorporate 'toxic' monarch prey into its diet without acute or chronic ill-effects.

Higher Activities of Defense-Associated Enzymes may Contribute to Greater Resistance of Manchurian Ash to Emerald Ash Borer Than A closely Related and Susceptible Congener.

Emerald ash borer (EAB) is an invasive beetle native to Asia that infests and kills ash (Fraxinus spp.) in North America. Previous experiments indicated that larvae feeding on co-evolved, resistant Manchurian ash (F. mandshurica) have increased antioxidant and quinone-protective enzyme activities compared to larvae feeding on susceptible North American species. Here, we examined mechanisms of host-generated oxidative and quinone-based stress and other putative defenses in Manchurian ash and the closely related and chemically similar, but susceptible, black ash (F. nigra), with and without exogenous application of methyl jasmonate (MeJA) to induce resistance mechanisms. Peroxidase activities were 4.6-13.3 times higher in Manchurian than black ash, although both species appeared to express the same three peroxidase isozymes. Additionally, peroxidase-mediated protein cross-linking activity was stronger in Manchurian ash. Polyphenol oxidase, ß-glucosidase, chitinase, and lipoxygenase activities also were greater in Manchurian ash, but only lipoxygenase activity increased with MeJA application. Phloem H2O2 levels were similar and were increased by MeJA application in both species. Lastly, trypsin inhibitor activity was detected in methanol and water extracts that were not allowed to oxidize, indicating the presence of phenolic-based trypsin inhibitors. However, no proteinaceous trypsin inhibitor activity was detected in either species. In response to MeJA application, Manchurian ash had higher trypsin inhibitor activity than black ash using the unoxidized water extracts, but no treatment effects were detected using methanol extracts. Based on these results we hypothesize that peroxidases, lignin polymerization, and quinone generation contribute to the greater resistance to EAB displayed by Manchurian ash.

Incidence of Infestation and Larval Success of Emerald Ash Borer (Agrilus planipennis) on White Fringetree (Chionanthus virginicus), Chinese Fringetree (Chionanthus retusus), and Devilwood (Osmanthus americanus).

We compared the incidence of infestation by emerald ash borer (EAB) and lilac borer on white fringetree to that of its Asian congener, Chinese fringetree, Chionanthus retusus, and a North American relative, devilwood, Osmanthus americanus. We also conducted laboratory bioassays to determine the suitability of these hosts for EAB larvae. At Spring Grove Cemetery and Arboretum in Cincinnati, Ohio, 9 of 28 white fringetrees examined were infested by EAB. Most of the white fringetrees had lilac borer infestation, and most of the trees infested by EAB also had lilac borer infestation. None of the 11 Chinese fringetrees examined were infested by either EAB or lilac borer. Each of the five devilwood individuals examined was infested by lilac borer, but not EAB. At The Morton Arboretum in Lisle, Illinois, 7 of 16 white fringetrees examined were infested by EAB, while none of the seven Chinese fringetrees examined were infested by either insect. A 40-d bioassay confirmed that white fringetree was an acceptable host, producing fourth-instar larvae that were smaller than those produced on a highly susceptible cultivar of green ash, Fraxinus pennsylvanica. No larvae survived on Chinese fringetree, and neonates were largely incapable of feeding on it. Two larvae survived on devilwood, reaching the second instar and excavating extensive galleries. Future work should be aimed at biotic and abiotic factors influencing the susceptibility of white fringetree, as well as further examination of close relatives for their vulnerability to EAB.

Water conservation features of ova of Agrilus planipennis (Coleoptera: Buprestidae).

The emerald ash borer, Agrilus planipennis Fairmaire, has destroyed millions of ash trees (Fraxinus spp.) in North America since first identified in Detroit in 2002. With species of ash distributed throughout North America, it is easy to speculate the extinction of all susceptible species of ash on the continent given a lack of physical, environmental, or climactic barrier for dispersal of the insect. We investigated water balance characteristics of emerald ash borer ova by using gravimetric methods in an effort to measure their response to heat- and water-stress and explore possible influences this stress may have on the ecology and physiology of the ovum. We also explored the possible water balance benefit of a peculiar, "clustering," oviposition behavior, as well as the difference in responses to stress between ova from a laboratory colony and ova from two wild populations. We found no evidence of water vapor absorption as a water balance strategy; rather enhanced water retention, resistance to desiccation, and viability with low water content were important survival strategies for these ova. Surface lipids resist thermal breakdown as indicated by ova having no detectable critical transition temperature, maintaining their water-proofing function as temperature rises. The observed "clustering" behavior had no desiccation-avoidance benefit and ova from the wild populations behaved almost identically to the ova from the lab colony, although the lab ova were slightly larger and more sensitive to dehydration. Given this new information, there appears to be no heat- or water-stress barriers for the dispersal of this devastating pest at the ovum stage.

Microbes as targets and mediators of allelopathy in plants.

Studies of allelopathy in terrestrial systems have experienced tremendous growth as interest has risen in describing biochemical mechanisms responsible for structuring plant communities, determining agricultural and forest productivity, and explaining invasive behaviors in introduced organisms. While early criticisms of allelopathy involved issues with allelochemical production, stability, and degradation in soils, an understanding of the chemical ecology of soils and its microbial inhabitants has been increasingly incorporated in studies of allelopathy, and recognized as an essential predictor of the outcome of allelopathic interactions between plants. Microbes can mediate interactions in a number of ways with both positive and negative outcomes for surrounding plants and plant communities. In this review, we examine cases where soil microbes are the target of allelopathic plants leading to indirect effects on competing plants, provide examples where microbes play either a protective effect on plants against allelopathic competitors or enhance allelopathic effects, and we provide examples where soil microbial communities have changed through time in response to allelopathic plants with known or potential effects on plant communities. We focus primarily on interactions involving wild plants in natural systems, using case studies of some of the world's most notorious invasive plants, but we also provide selected examples from agriculturally managed systems. Allelopathic interactions between plants cannot be fully understood without considering microbial participants, and we conclude with suggestions for future research.