Acibenzolar-S-methyl induces resistance against ambrosia beetle attacks in dogwoods exposed to simulated flood stress.

Xylosandrus spp. ambrosia beetles (Coleoptera: Curculionidae: Scolytinae) are important wood-boring pests of nursery trees weakened by abiotic and biotic stressors. Acibenzolar-S-methyl (ASM), a plant defense elicitor, was tested for inhibiting Xylosandrus spp. tunneling (i.e., attacks) into flood-stressed flowering dogwoods (Cornus florida L. (Cornales: Cornaceae)). Container-grown dogwoods were treated with ASM substrate drench + flooding, ASM foliar spray + flooding, ASM drench + no flooding, ASM foliar + no flooding, no ASM + flooding, or no ASM + no flooding at 3 days before flood stress in a completely randomized design under field conditions. Trees were flooded for 14 days and then drained and watered as needed. Attacks were counted every 2 days for 28 days. Plant tissue samples were collected at 7 and 14 days after flooding to determine ethanol content using solid-phase microextraction-gas chromatography-mass spectrometry. Trees were dissected to determine gallery formation and depth, fungal colonization, and the presence of eggs, larvae, and adults. The highest number of Xylosandrus beetle species attacks were recorded from plants exposed to no ASM + flooding, but attacks were reduced in ASM treated trees (drench or foliar) + flooding. Trees treated with drenches had fewer attacks than foliar sprays. Plants assigned to no flood had the fewest beetle attacks. Moreover, ASM reduced Xylosandrus spp. gallery formation and depth, fungal colonization, and presence of eggs, larvae, and adults. All flooded trees produced ethanol. In conclusion, ASM induced a plant defense response to Xylosandrus spp. tunneling in dogwoods under flood stress conditions.

Volatiles of fungal cultivars act as cues for host-selection in the fungus-farming ambrosia beetle Xylosandrus germanus.

Many wood-boring insects use aggregation pheromones during mass colonization of host trees. Bark beetles (Curculionidae: Scolytinae) are a model system, but much less is known about the role of semiochemicals during host selection by ambrosia beetles. As an ecological clade within the bark beetles, ambrosia beetles are obligately dependent on fungal mutualists for their sole source of nutrition. Mass colonization of trees growing in horticultural settings by exotic ambrosia beetles can occur, but aggregation cues have remained enigmatic. To elucidate this mechanism, we first characterized the fungal associates of the exotic, mass-aggregating ambrosia beetle Xylosandrus germanus in Southern Germany. Still-air olfactometer bioassays documented the attraction of X. germanus to its primary nutritional mutualist Ambrosiella grosmanniae and to a lesser extent another common fungal isolate (Acremonium sp.). During two-choice bioassays, X. germanus was preferentially attracted to branch sections (i.e., bolts) that were either pre-colonized by conspecifics or pre-inoculated with A. grosmanniae. Subsequent analyses identified microbial volatile organic compounds (MVOCs) that could potentially function as aggregation pheromones for X. germanus. To our knowledge, this is the first evidence for fungal volatiles as attractive cues during host selection by X. germanus. Adaptive benefits of responding to fungal cues associated with an infestation of conspecifics could be a function of locating a suitable substrate for cultivating fungal symbionts and/or increasing the likelihood of mating opportunities with the flightless males. However, this requires solutions for evolutionary conflict arising due to potential mixing of vertically transmitted and horizontally acquired symbiont strains, which are discussed.

Attraction of Myzus persicae (Hemiptera: Aphididae) to Volatiles Emitted From the Entomopathogenic Fungus Beauveria bassiana.

Beauveria bassiana (Balsamo) Vuillemin infects a wide variety of insects, including the green peach aphid, Myzus persicae (Sulzer). Volatiles emitted from B. bassiana can act as semiochemical attractants or repellents, with most responses reported to date resulting in insects avoiding B. bassiana. Since insects can detect 'enemy-specific volatile compounds', we hypothesized the preference behavior of M. persicae would be influenced by volatile emissions from B. bassiana. We conducted Petri dish and Y-tube olfactometer bioassays to characterize the preference of M. persicae to B. bassiana strain GHA. During Petri dish bioassays, more apterous and alate M. persicae were recorded in the vicinity of agar colonized by B. bassiana compared to agar, or Fusarium proliferatum (Matsushima) Nirenberg and Ambrosiella grosmanniae Mayers, McNew, & Harrington as representatives of nonentomopathogenic fungi. Petri dish bioassays also determined that apterous and alate M. persicae preferred filter paper saturated with 1 × 107, 1 × 106, and 1 × 105B. bassiana conidia/ml compared to Tween 80. Y-tube bioassays documented that more apterous and alate M. persicae oriented upwind to volatiles from B. bassiana mycelia compared to agar. Apterous and alate Myzus persicae were also preferentially attracted to 1 × 107 and 1 × 106B. bassiana conidia/ml compared to Tween-80 during Y-tube bioassays. These results complement a previous finding that the mosquito Anopheles stephensi (Diptera: Culicidae) Liston is attracted to volatiles from B. bassiana. Future studies aimed at characterizing the olfactory mechanism leading to the attraction of M. persicae to B. bassiana could aid in optimizing lure-and-kill strategies.

Type and duration of water stress influence host selection and colonization by exotic ambrosia beetles (Coleoptera: Curculionidae).

Fungus-farming ambrosia beetles in the tribe Xyleborini tunnel into plants and trees to establish chambers for cultivating their nutritional fungal mutualists and rearing offspring. Some xyleborine ambrosia beetles preferentially infest and perform better in living but weakened trees. Flood stress predisposes horticultural tree crops to infestation, but the impact of drought stress has not been well studied. Our objectives were to compare the effects of flood stress vs. drought stress on host selection and colonization by xyleborine ambrosia beetles and to assess the duration of flooding. Container-grown Cornus florida L. trees were flood stressed using a pot-in-pot system to submerge the roots in water while drought-stressed conditions were imposed by withholding irrigation and precipitation. When experimental trees were held under field conditions for 14 days, 7.5 × more ambrosia beetles landed on stems of the flood-stressed than on the drought-stressed trees. During two additional experiments over 14 and 22 days, ambrosia beetles tunneled into the flood-stressed trees but not the drought-stressed or standard irrigation trees. By simultaneously deploying trees that were flood stressed for varying lengths of time, it was found that more tunnel entrances, and xyleborine adults and offspring were recovered from trees that were flooded for 1-16 days and 7-22 days than from trees that were flooded for 14-29 days and 28-43 days. These results indicate that acute and severe drought stress does not predispose C. florida to infestation, but flood stress and the duration of flooding influence ambrosia beetle host selection and colonization. Understanding the role of host quality on ambrosia beetle preference behavior will assist with predicting the risk of infestation of these opportunistic insects in horticultural tree crops.

Stability of Nuclear and Mitochondrial Reference Genes in Selected Tissues of the Ambrosia Beetle Xylosandrus germanus.

The fungus-farming ambrosia beetle Xylosandrus germanus (Blandford) uses a pouch-like structure (i.e., mycangium) to transport spores of its nutritional fungal mutualist. Our current study sought to identify reference genes necessary for future transcriptome analyses aimed at characterizing gene expression within the mycangium. Complementary DNA was synthesized using selected tissue types from laboratory-reared and field-collected X. germanus consisting of the whole body, head + thorax, deflated or inflated mycangium + scutellum, inflated mycangium, and thorax + abdomen. Quantitative reverse-transcription PCR reactions were performed using primers for 28S ribosomal RNA (28S rRNA), arginine kinase (AK), carbamoyl-phosphate synthetase 2-aspartate transcarbamylase-dihydroorotase (CAD), mitochondrial cytochrome oxidase 1 (CO1), and elongation factor-1α (EF1α). Reference gene stability was analyzed using GeNorm, NormFinder, BestKeeper, ΔCt, and a comprehensive final ranking by RefFinder. The gene CO1 was identified as the primary reference gene since it was generally ranked in first or second position among the tissue types containing the mycangium. Reference gene AK was identified as a secondary reference gene. In contrast, EF1α was generally ranked in the last or penultimate place. Identification of two stable reference genes will aid in normalizing the expression of target genes for subsequent gene expression studies of X. germanus' mycangium.

Microbial Control Agents for Fungus Gnats (Diptera: Sciaridae: Lycoriella) Affecting the Production of Oyster Mushrooms, Pleurotus spp.

Infestations of fungus gnats (Diptera: Sciaridae) can reduce the production of oyster mushrooms (Pleurotus spp.) grown as food crops within controlled environments. The objectives of this study were to assess the efficacy of Bacillus thuringiensis var. israelensis (Bti) and Steinernema feltiae against fungus gnat larvae. A bioassay was developed, whereby pasteurized straw was inoculated with Pleurotus columbinus and treated with Bti (Gnatrol®), S. feltiae (Nemashield®), or water. Fungus gnats (Lycoriella sp.) were released into each bioassay container for ovipositing onto the straw, thereby exposing the F1 larvae to treated or untreated substrate. Sticky cards within the containers entrapped fungus gnats emerging from the substrate as an indicator of larval survivorship. Following three bioassays, fewer fungus gnats emerged from straw treated with Bti compared to S. feltiae and the water control. Three additional bioassays using Pleurotus ostreatus also demonstrated that fewer fungus gnats emerged from straw treated with Bti compared to S. feltiae and the untreated control. Steinernema feltiae was generally ineffective. Monitoring substrate weight in the bioassay containers over time indicated that Bti and S. feltiae did not impede colonization by P. ostreatus. Incorporating Bti into straw substrate is a promising approach for managing fungus gnats infesting Pleurotus spp.

Electrophysiological and Behavioral Responses of an Ambrosia Beetle to Volatiles of its Nutritional Fungal Symbiont.

Ambrosia beetles (Coleoptera: Scolytinae) cultivate their fungal symbiont within host substrates as the sole source of nutrition on which the larvae and adults must feed. To investigate a possible role for semiochemicals in this interaction, we characterized electrophysiological and behavioral responses of Xylosandrus germanus to volatiles associated with its fungal symbiont Ambrosiella grosmanniae. During still-air walking bioassays, X. germanus exhibited an arrestment response to volatiles of A. grosmanniae, but not antagonistic fungi Beauveria bassiana, Metarhizium brunneum, Trichoderma harzianum, the plant pathogen Fusarium proliferatum, or malt extract agar. Solid phase microextraction-gas chromatography-mass spectrometry identified 2-ethyl-1-hexanol, 2-phenylethanol, methyl benzoate and 3-methyl-1-butanol in emissions from A. grosmanniae; the latter two compounds were also detected in emissions from B. bassiana. Concentration-responses using electroantennography documented weak depolarizations to A. grosmanniae fungal volatiles, unlike the comparatively strong response to ethanol. When tested singly in walking bioassays, volatiles identified from A. grosmanniae elicited relatively weak arrestment responses, unlike the responses to ethanol. Xylosandrus germanus also exhibited weak or no long-range attraction to the fungal volatiles when tested singly during field trials in 2016-2018. None of the fungal volatiles enhanced attraction of X. germanus to ethanol when tested singly; in contrast, 2-phenylethanol and 3-methyl-1-butanol consistently reduced attraction to ethanol. Volatiles emitted by A. grosmanniae may represent short-range olfactory cues that could aid in distinguishing their nutritional fungal symbiont from other fungi, but these compounds are not likely to be useful as long-range attractants for improving detection or mass trapping tactics.

Colonization of Trees by Ambrosia Beetles (Coleoptera: Curculionidae: Scolytinae) Is Influenced by Duration of Flood Stress.

The ambrosia beetles Xylosandrus germanus (Blandford) and Xylosandrus crassiusculus (Motschulsky) bore into flood-stressed trees to establish colonies, but the influence of flooding duration on colonization is unknown. This relationship was examined by flooding trees for various time periods and evaluating colonization. In one experiment, X. germanus bored into 20 dogwood (Cornus florida L.) trees during a 3-d flood treatment. Ten trees dissected that season had no offspring present in tunnels; the remaining trees appeared healthy and bloomed the following spring. In another experiment, dogwood trees were flooded for 3 or 7 d and then dissected to assess colonization. The incidence of superficial (short unbranched) and healed (callus tissue in entrance) tunnels was greater in the 3-d trees, while the incidence of tunnels with X. germanus or offspring was greater in the 7-d trees. Four experiments (three in Ohio and one in Virginia) had flood treatments of 0 (nonflooded), 3, 5, 7, and 10 d. Numbers of tunnel entrances, tunnels with X. germanus, and incidence of tunnels with offspring or live foundresses tended to increase as flood duration increased on apple (Malus × domestica Borkh.), dogwood, and redbud (Cercis canadensis L.) in Ohio and redbud in Virginia. Nonflooded trees in Ohio had no boring activity, but ambrosia beetles bored into three nonflooded trees in Virginia. Indicators of unsuccessful colonization, such as superficial tunnels and healing, decreased as flood duration increased. These results suggest tree crops may recover from boring by ambrosia beetles following short-duration flood events, and not necessarily require culling.

Effects of chlorantraniliprole residual on Helicoverpa zea in Bt and non-Bt cotton.

BACKGROUND: Helicoverpa zea is managed with foliar applications of chlorantraniliprole in cotton varieties that do not express the Vip3Aa19 toxin in the US Cotton Belt. Foliar insecticides and Bt could interact to influence larval susceptibility. Therefore, it has been suggested that chlorantraniliprole can be used as a tool for Bt resistance management. We designed field and laboratory studies to test the hypothesis that the interaction of Bt toxin and chlorantraniliprole application would result in lower H. zea larval survival when compared to the individual effect of Bt or chlorantraniliprole alone. We also tested for these interactions over time, since chlorantraniliprole residual has not been studied in cotton. RESULTS: Results from two field experiments and two laboratory experiments were similar. We found no interactions with Bt and chlorantraniliprole using data not corrected for natural mortality in untreated plots, indicating that these factors did not interact to influence survival. Moreover, we found that Bt and chlorantraniliprole did not interact to influence larval weight and instar. Chlorantraniliprole had lethal and sublethal effects on H. zea larval growth parameters feeding on cotton leaves up to 22 days after application, the final time period that we tested. Finally, concentration of chlorantraniliprole in the leaf was associated with larval survival for the duration of this study, but not larval growth or instar. CONCLUSION: Our findings complement the recommendation to use chlorantraniliprole for managing H. zea in cotton, given its long-residual effects. However, the utility of chlorantraniliprole as a Bt-resistance management tool for H. zea remains unclear. © 2021 Society of Chemical Industry.

Thiamethoxam Differentially Impacts the Survival of the Generalist Predators, Orius insidiosus (Hemiptera: Anthocoridae) and Hippodamia convergens (Coleoptera: Coccinellidae), When Exposed via the Food Chain.

Insect predators are seldom considered during toxicological trophic assessments for insecticide product development. As a result, the ecological impact of novel insecticides on predators is not well understood, especially via the food chain, i.e., when their prey is exposed to insecticides. Neonicotinoids are systemic insecticides widely used in agriculture to control herbivorous insects, but their effects on predatory insects via the food chain have not been well characterized. In this study, we documented the time-course effects of the neonicotinoid thiamethoxam on the survival of two predators, the insidiosus flower bug Orius insidiosus (Say) and the convergent lady beetle Hippodamia convergens Guérin-Méneville, when preying upon the aphids Aphis glycines Matsumura (Hemiptera: Aphididae), Aphis gossypii Glover (Hemiptera: Aphididae), and Myzus persicae (Sulzer) (Hemiptera: Aphididae). Aphids were exposed to thiamethoxam-treated or untreated plants every week over the course of 5 wk. After transferring aphids to Petri dishes, predators were allowed to feed on aphids. We found that the survival of the insidiosus flower bug, but not the convergent lady beetle, was reduced after consuming aphids reared on thiamethoxam-treated plants compared to untreated plants. Survival reduction of the insidiosus flower bug was observed only during the first weeks after thiamethoxam application; no reduction occurred 28 d after treatment or beyond. These results demonstrate that a systemic application of thiamethoxam could be compatible with convergent lady beetles and insidiosus flower bugs, if the time of predator release does not coincide with thiamethoxam activity. These findings are critical for the development of future pest control programs that integrate biological and chemical control.

Attraction of Invasive Ambrosia Beetles (Coleoptera: Curculionidae: Scolytinae) to Ethanol-Treated Tree Bolts.

Ethanol-treated bolts (tree stem sections) have potential as monitoring and pesticide screening tools for ambrosia beetles (Coleoptera: Curculionidae: Scolytinae). Bolts were infused with ethanol by immersing them for at least 24 h. Attacks on ethanol-treated bolts by Xylosandrus species were compared with captures in ethanol-baited traps. Bolts infused in ethanol were usually as attractive or more attractive to Xylosandrus germanus (Blandford) than ethanol-baited bottle traps. Xylosandrus crassiusculus (Motschulsky) were more attracted to bolts than trap in some experiments, but numbers were low and differences were usually not significant. Two techniques for treating bolts with ethanol were compared. Attraction of ambrosia beetles to ethanol-infused bolts were compared with bolts with a drilled cavity filled with ethanol. Drilled bolts filled with ethanol were attractive to X. germanus and were reliably attacked, but numbers of beetles were often lower than in traps and infused bolts. Aged and fresh ethanol-infused bolts were compared with evaluate residual attractiveness. Bolts aged 7 d usually had fewer X. germanus than fresh bolts and traps, and bolts aged 14 d had no beetles. Ethanol-infused bolts from different species of trees were compared. Xylosandrus germanus attacked all species tested with more attacks usually in red maple (Acer rubrum L.). Anisandrus maiche Stark was attracted to ethanol-infused bolts indicating it may attack trees emitting ethanol. Bolts attracted fewer nontarget species than traps, but residual attraction was much less. The selectivity of ethanol-treated bolts for Xylosandrus species should make them useful for monitoring and screening pesticides against those species.

Long-Lasting Insecticide Netting for Protecting Tree Stems from Attack by Ambrosia Beetles (Coleoptera: Curculionidae: Scolytinae).

Ambrosia beetles (Coleoptera: Curculionidae: Scolytinae) are destructive wood-boring insects of horticultural trees. We evaluated long-lasting insecticide netting for protecting stems against ambrosia beetles. Container-grown eastern redbud, Cercis canadensis, trees were flood-stressed to induce ambrosia beetle attacks, and deltamethrin-treated netting was wrapped from the base of the stem vertically to the branch junction. Trees were deployed under field conditions in Ohio, Virginia, Tennessee, and Mississippi with the following treatments: (1) flooded tree; (2) flooded tree with untreated netting; (3) flooded tree with treated 'standard mesh' netting of 24 holes/cm2; (4) flooded tree with treated 'fine mesh' netting of 28 holes/cm2; and/or (5) non-flooded tree. Treated netting reduced attacks compared to untreated netting and/or unprotected trees in Mississippi in 2017, Ohio and Tennessee in 2018, and Virginia in 2017-2018. Inconsistent effects occurred in Mississippi in 2018. Fewer Anisandrus maiche, Xylosandrus germanus, and Xyleborinus saxesenii were dissected from trees deployed in Ohio protected with treated netting compared to untreated netting; trees deployed in other locations were not dissected. These results indicate long-lasting insecticide netting can provide some protection of trees from ambrosia beetle attacks.

Weekly Survivorship Curves of Soybean Aphid Biotypes 1 and 4 on Insecticidal Seed-Treated Soybean.

Thiamethoxam, an insecticide used in soybean seed treatments, effectively suppresses soybean aphids (Aphis glycines) Matsumura (Hemiptera: Aphididae) for a short time after planting. However, exactly when and how quickly soybean aphid populations could increase is unknown. Likewise, we lack data on virulent soybean aphid biotypes (that can overcome soybean resistance) when fed on seed-treated soybean. Determining the survival of soybean aphids over time on insecticidal seed-treated soybean is critical for improving soybean aphid management and may provide insights to manage aphid virulence to aphid resistant-soybean. In greenhouse and field experiments, aphid-susceptible soybean plants (with and without an insecticidal seed treatment) were infested at 7, 14, 21, 28, 35, and 42 days after planting (DAP). We compared aphid survival among biotypes 1 (avirulent) and 4 (virulent) and insecticide treatment 72 h after infestation. We also measured thiamethoxam concentrations in plant tissue using liquid chromatography-tandem mass spectrometry. As expected, soybean aphid survival was significantly lower on seed-treated soybean up to 35 DAP for both biotypes, which correlates with the decrease of thiamethoxam in the plant over time. Moreover, we found no significant difference between avirulent and virulent biotype survivorship on insecticidal seed-treated soybean plants, although we did find significantly greater survival for the virulent biotype compared with the avirulent biotype on untreated soybean in the field. In conclusion, our study further characterized the relative short duration of seed treatment effectiveness on soybean aphid and showed that survivorship of virulent aphids on seed-treated soybean is similar to avirulent aphids.

Trap Tree and Interception Trap Techniques for Management of Ambrosia Beetles (Coleoptera: Curculionidae: Scolytinae) in Nursery Production.

The majority of wood-boring ambrosia beetles are strongly attracted to ethanol, a behavior which could be exploited for management within ornamental nurseries. A series of experiments was conducted to determine if ethanol-based interception techniques could reduce ambrosia beetle pest pressure. In two experiments, trap trees injected with a high dose of ethanol were positioned either adjacent or 10-15 m from trees injected with a low dose of ethanol (simulating a mildly stressed tree) to determine if the high-dose trap trees could draw beetle attacks away from immediately adjacent stressed nursery trees. The high-ethanol-dose trees sustained considerably higher attacks than the low-dose trees; however, distance between the low- and high-dose trees did not significantly alter attack rates on the low-dose trees. In a third experiment, 60-m length trap lines with varying densities of ethanol-baited traps were deployed along a forest edge to determine if immigrating beetles could be intercepted before reaching sentinel traps or artificially stressed sentinel trees located 10 m further in-field. Intercept trap densities of 2 or 4 traps per trap line were associated with fewer attacks on sentinel trees compared to no traps, but 7 or 13 traps had no impact. None of the tested intercept trap densities resulted in significantly fewer beetles reaching the sentinel traps. The evaluated ethanol-based interception techniques showed limited promise for reducing ambrosia beetle pressure on nursery trees. An interception effect might be enhanced by applying a repellent compound to nursery trees in a push-pull strategy.

Symbiont selection via alcohol benefits fungus farming by ambrosia beetles.

Animal-microbe mutualisms are typically maintained by vertical symbiont transmission or partner choice. A third mechanism, screening of high-quality symbionts, has been predicted in theory, but empirical examples are rare. Here we demonstrate that ambrosia beetles rely on ethanol within host trees for promoting gardens of their fungal symbiont and producing offspring. Ethanol has long been known as the main attractant for many of these fungus-farming beetles as they select host trees in which they excavate tunnels and cultivate fungal gardens. More than 300 attacks by Xylosandrus germanus and other species were triggered by baiting trees with ethanol lures, but none of the foundresses established fungal gardens or produced broods unless tree tissues contained in vivo ethanol resulting from irrigation with ethanol solutions. More X. germanus brood were also produced in a rearing substrate containing ethanol. These benefits are a result of increased food supply via the positive effects of ethanol on food-fungus biomass. Selected Ambrosiella and Raffaelea fungal isolates from ethanol-responsive ambrosia beetles profited directly and indirectly by (i) a higher biomass on medium containing ethanol, (ii) strong alcohol dehydrogenase enzymatic activity, and (iii) a competitive advantage over weedy fungal garden competitors (Aspergillus, Penicillium) that are inhibited by ethanol. As ambrosia fungi both detoxify and produce ethanol, they may maintain the selectivity of their alcohol-rich habitat for their own purpose and that of other ethanol-resistant/producing microbes. This resembles biological screening of beneficial symbionts and a potentially widespread, unstudied benefit of alcohol-producing symbionts (e.g., yeasts) in other microbial symbioses.

Residue Age and Attack Pressure Influence Efficacy of Insecticide Treatments Against Ambrosia Beetles (Coleoptera: Curculionidae).

Management of ambrosia beetles in ornamental nurseries relies, in part, on insecticide treatments to prevent beetles from boring into trees. However, data on residual efficacy of commonly used pyrethroid insecticides is needed to gauge the duration that trees are protected during spring when peak beetle pressure occurs. Residual efficacy of bifenthrin and permethrin trunk sprays was examined in field trials which used trees injected with 10% ethanol to ensure host attack pressure. Permethrin consistently reduced attacks by Xylosandrus germanus (Blandford; Coleoptera: Curculionidae) and other ambrosia beetles for at least 4 wk, while efficacy of bifenthrin was inconsistent and lasted only about 10 d. Since previous studies demonstrated attacks are positively correlated with host ethanol emissions, we injected trees with 2.5, 5, and 10% ethanol to determine if residual efficacy was affected by attack pressure. Preventive treatments with bifenthrin reduced ambrosia beetle attacks at all concentrations of injected ethanol compared to non-sprayed controls. There was no interaction between attack pressure and insecticide treatment with respect to total attacks or attacks by X. germanus. However, increasing attack pressure did increase the probability of attacks on insecticide treated trees by X. germanus and other Scolytinae. Results from our current study will improve the ability of growers to make decisions on frequency of protective sprays, but residual efficacy of insecticide treatments may decline as attack pressure increases. Cultural practices should therefore maximize host vigor and minimize attack pressure associated with stress-induced ethanol emissions.

Interaction of Insecticide and Media Moisture on Ambrosia Beetle (Coleoptera: Curculionidae) Attacks on Selected Ornamental Trees.

Exotic ambrosia beetles, particularly Xylosandrus crassiusculus (Motschulsky) (Coleoptera: Curculionidae: Scolytinae) and Xylosandrus germanus (Blandford) (Coleoptera: Curculionidae: Scolytinae), are among the most damaging pests of ornamental trees in nurseries. Growers have had few tactics besides insecticide applications to reduce ambrosia beetle attacks but recent research has shown that attacks may be reduced by maintaining media moisture below a 50% threshold thereby reducing flood stress. We compared the efficacy of managing media moisture and insecticide applications for reducing ambrosia beetle attacks on three ornamental tree species in North Carolina. During trials in spring 2013 and 2015, flooded Cornus florida and Cornus kousa were heavily attacked despite sprays with permethrin, but nonflooded C. kousa or C. florida were not attacked. In spring 2015 trials, both nonflooded and flooded Styrax japonicus were heavily attacked regardless of permethrin applications. Although ethanol emissions were not measured, the apparently healthy nonflooded S. japonicus trees may have been exposed to an unknown physiological stress, such as low temperature injury, the previous winter, which predisposed them to beetle attack. However, ethanol levels within host tissues were not measured as part of the current study. X. crassiusculus (75%), Xyloborinus saxesenii Ratzburg (13%), and X. germanus (9%) were the most abundant species collected in ethanol baited traps deployed in 2015, while X. crassiusculus (63%) and X. germanus (36%) were the predominant species reared from attacked trees. Results indicate that managing media moisture levels at or below 50%, and maximizing tree health overall, may provide significant protection against Xylosandrus spp. attacks in flood intolerant tree species.

Trap Style, Bait, and Height Deployments in Black Walnut Tree Canopies Help Inform Monitoring Strategies for Bark and Ambrosia Beetles (Coleoptera: Curculionidae: Scolytinae).

Knowledge about which bark and ambrosia beetle species are active and at what heights in black walnut canopies is not well understood. Neither is the role of these beetles in spreading Thousand Cankers Disease. To assist with future planned research, which will assess the extent to which these beetle species are associated with Geosmithia morbida Kolarík, Freeland, Utley, and Tisserat (Ascomycota: Hypocreales: Bionectriaceae), experiments were undertaken to monitor bark and ambrosia beetles in urban landscapes and parks in Tennessee between 2011 and 2013. Within mature walnut tree canopies, sticky panel, modified soda bottle, and Lindgren traps were deployed at different heights, with and without ethanol as an attractant and with and without walnut stem sections, or in situ limbs that had been girdled or injection with ethanol to simulate stressed tree tissues. Bark and ambrosia beetle species (Coleoptera: Curculionidae: Scolytinae) collected in greatest abundance included Ambrosiodmus rubricollis (Eichhoff), Ambrosiophilus atratus (Eichhoff), Cnestus mutilatus (Blandford), Dryoxylon onoharaense (Murayama), Euwallacea validus (Eichhoff), Monarthrum fasciatum (Say), Monarthrum mali (Fitch), Xyleborinus saxesenii (Ratzeburg), Xyleborus affinis Eichhoff, Xyleborus ferrugineus (Fabricius), Xylosandrus crassiusculus (Motschulsky), and Xylosandrus germanus (Blandford). C. mutilatus, X. saxesenii, and X. crassiusculus were more active higher in trees than most other species and were strongly attracted to ethanol via all means of lure deployment. C. mutilatus, which were captured from April through October and increased in abundance across the 3-yr study, were most abundant in late May with a second activity period in late August.

Flood Stress as a Technique to Assess Preventive Insecticide and Fungicide Treatments for Protecting Trees against Ambrosia Beetles.

Ambrosia beetles tunnel into the heartwood of trees where they cultivate and feed upon a symbiotic fungus. We assessed the effectiveness of flood stress for making Cercis canadensis L. and Cornus florida L. trees attractive to attack as part of insecticide and fungicide efficacy trials conducted in Ohio and Virginia. Since female ambrosia beetles will not begin ovipositing until their symbiotic fungus is established within the host, we also assessed pre-treatment of trees with permethrin, azoxystrobin, and potassium phosphite on fungal establishment and beetle colonization success. Permethrin reduced attacks on flooded trees, yet no attacks occurred on any of the non-flooded trees. Fewer galleries created within flooded trees pre-treated with permethrin, azoxystrobin, and potassium phosphite contained the purported symbiotic fungus; foundress' eggs were only detected in flooded but untreated trees. While pre-treatment with permethrin, azoxystrobin, and potassium phosphite can disrupt colonization success, maintaining tree health continues to be the most effective and sustainable management strategy.

Developing a Media Moisture Threshold for Nurseries to Reduce Tree Stress and Ambrosia Beetle Attacks.

Exotic ambrosia beetles are among the most damaging pests of trees grown in nurseries. The primary pests Xylosandrus crassiusculus Motschulsky and Xylosandrus germanus Blandford use ethanol to locate vulnerable trees. Research, primarily with X. germanus, has shown that flood-stressed trees emit ethanol and are preferentially attacked by ambrosia beetles. Our goal was to develop a media (also called potting soil) moisture threshold as an integrated pest management (IPM) tactic and assess grower practices that lead to ambrosia beetle attacks. Flooded Cornus florida L., Cornus kousa Burg., and Magnolia grandiflora L. trees incurred more attacks than unflooded trees that were not attacked. To determine optimal media moisture levels, we grew flood-tolerant Acer rubrum L. and flood-intolerant C. florida in containers with 10, 30, 50, 70, or 90% media moisture. No flooded or unflooded A. rubrum were attacked. However, C. florida grown in 70 or 90% moisture were attacked and died, whereas trees at 30 and 50% moisture were not attacked. Thus, we suggest an upper moisture threshold of 50% when growing C. florida and other flood-intolerant trees. However, during peak ambrosia beetle flight activity in spring 2013 and 2014, we found that media moisture levels in commercial nurseries were often between 50 and 90%. Implementing a media moisture threshold, as a new IPM tool, could reduce ambrosia beetle attacks and the need for insecticide applications, which is currently the only available management tactic. Future research should focus on how changes in substrates, irrigation, and other practices could help growers meet this threshold.