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.

Ophiostomatalean fungi associated with wood boring beetles in South Africa including two new species.

Ambrosia beetles are small wood inhabiting members of the Curculionidae that have evolved obligate symbioses with fungi. The fungal symbionts concentrate nutrients from within infested trees into a usable form for their beetle partners, which then utilize the fungi as their primary source of nutrition. Ambrosia beetle species associate with one or more primary symbiotic fungal species, but they also vector auxiliary symbionts, which may provide the beetle with developmental or ecological advantages. In this study we isolated and identified ophiostomatalean fungi associated with ambrosia beetles occurring in a native forest area in South Africa. Using a modified Bambara beetle trap, living ambrosia beetle specimens were collected and their fungal symbionts isolated. Four beetle species, three Scolytinae and one Bostrichidae, were collected. Five species of ophiostomatalean fungi were isolated from the beetles and were identified using both morphological characters and DNA sequence data. One of these species, Raffaelea sulphurea, was recorded from South Africa for the first time and two novel species were described as Ceratocystiopsis lunata sp. nov. and Raffaelea promiscua sp. nov.

A selective fungal transport organ (mycangium) maintains coarse phylogenetic congruence between fungus-farming ambrosia beetles and their symbionts.

Thousands of species of ambrosia beetles excavate tunnels in wood to farm fungi. They maintain associations with particular lineages of fungi, but the phylogenetic extent and mechanisms of fidelity are unknown. We test the hypothesis that selectivity of their mycangium enforces fidelity at coarse phylogenetic scales, while permitting promiscuity among closely related fungal mutualists. We confirm a single evolutionary origin of the Xylosandrus complex-a group of several xyleborine genera that farm fungi in the genus Ambrosiella. Multi-level co-phylogenetic analysis revealed frequent symbiont switching within major Ambrosiella clades, but not between clades. The loss of the mycangium in Diuncus, a genus of evolutionary cheaters, was commensurate with the loss of fidelity to fungal clades, supporting the hypothesis that the mycangium reinforces fidelity. Finally, in vivo experiments tracked symbiotic compatibility throughout the symbiotic life cycle of Xylosandrus compactus and demonstrated that closely related Ambrosiella symbionts are interchangeable, but the probability of fungal uptake in the mycangium was significantly lower in more phylogenetically distant species of symbionts. Symbiont loads in experimental subjects were similar to wild-caught beetles. We conclude that partner choice in ambrosia beetles is achieved in the mycangium, and co-phylogenetic inferences can be used to predict the likelihood of specific symbiont switches.

Xyleborus volvulus (Coleoptera: Curculionidae): Biology and Fungal Associates.

The ambrosia beetle Xyleborus volvulus Fabricius has been reported as a potential vector of the plant pathogen Raffaelea lauricola T.C. Harr., Fraedrich & Aghayeva that is affecting avocado orchards in South Florida. In this study, we examined its life cycle, process of gallery formation, gallery structure, and fungal associates by rearing one generation on avocado sawdust medium under control conditions. The adult foundress excavated a vertical tunnel that constituted the main gallery with a length of 2.5 cm, followed by the construction of up to six secondary galleries with a total length of 4.4 cm. The time period for one generation (egg to adult) was 28 days. Teneral males emerged 3 days after the emergence of the first females. The F1 generation did not significantly contribute to gallery expansion. Four species of Raffaelea and nine yeast species were recovered from galleries and beetles. Raffaelea arxii and Candida berthetii were the most frequent symbionts recovered from new adults and galleries. Candida berthetii dominated during the early stages of the gallery development, whereas R. arxii was most frequent in later stages. Other Raffaelea species were inconsistently isolated from galleries, which suggests a strong association between Xyleborus volvulus and both R. arxii and C. berthetii These results suggest that R. arxii is the primary nutritional symbiont of X. volvulus and that yeast species may be pioneer colonizers that assist with the growth of fungal symbionts.IMPORTANCE Ambrosia beetles cultivate fungi in tunnels bored into weakened host trees. This obligate interaction is required for their survival as beetles feed on these symbiotic fungi, and the fungi benefit from transportation by the beetles. Xyleborus volvulus carries many nonpathogenic symbionts; however, recently the acquisition of Raffaelea lauricola (the causal agent of a lethal vascular disease of lauraceous trees) by this beetle has altered its status from wood degrader to potential pest in avocado. We conducted a study to understand the relationship of this beetle and its fungal associates. Our results show that X. volvulus has a multipartite flexible association with different Raffaelea species. The lack of fidelity in the mutualistic association may explain the acquisition of R. lauricola Knowing the beetle biology and its mutualistic interactions furthers an understanding of the beetle's role as a potential vector and in disease transmission.

Variation in Effects of Conophthorin on Catches of Ambrosia Beetles (Coleoptera: Curculionidae: Scolytinae) in Ethanol-Baited Traps in the United States.

In 2013, we examined the effects of conophthorin on flight responses of ambrosia beetles (Coleoptera: Curculionidae: Scolytinae) to multiple-funnel traps baited with ethanol in Georgia, Michigan, New Hampshire, and Oregon. Adventive species (=exotic, nonnative, immigrant, introduced) accounted for 91.4% of total catches of ambrosia beetles. Conophthorin increased catches of Xyleborinus saxesenii (Ratzeburg) in Georgia, New Hampshire, and Oregon. Catches of Cyclorhipidion pelliculosum (Eichhoff) were increased by conophthorin in New Hampshire but not in Michigan. In Oregon, conophthorin decreased catches of Xylosandrus germanus (Blandford) to ethanol-baited traps but not in Michigan and New Hampshire. In Georgia, conophthorin increased catches of Gnathotrichus materiarius (Fitch), Xyleborus spp., and Xylosandrus crassiusculus (Motschulsky) but decreased catches of Cnestus mutilatus (Blandford), Dryoxylon onoharaensum (Murayama), and Cyclorhipidion bodoanum (Reitter). Conophthorin had no effect on catches of Ambrosiophilus atratus (Eichhoff), Anisandrus dispar (F.), Anisandrus sayi (Hopkins), Gnathotrichus sulcatus (Leconte), Monarthrum fasciatum (Say), Monarthrum mali (Fitch), and Xylosandrus compactus (Eichhoff). Attraction of the bark beetle, Hypothenemus rotundicollis (Eichhoff), was interrupted by conophthorin in Georgia. Our results suggest that adding conophthorin lures to traps baited with ethanol may have utility in detection programs in North America and overseas. However, traps baited with ethanol alone should also be used due to interruption in attraction for some species of ambrosia beetles.

Ambrosiella roeperi sp. nov. is the mycangial symbiont of the granulate ambrosia beetle, Xylosandrus crassiusculus.

Isolations from the granulate ambrosia beetle, Xylosandrus crassiusculus (Coleoptera: Curculionidae: Scolytinae: Xyleborini), collected in Georgia, South Carolina, Missouri and Ohio, yielded an undescribed species of Ambrosiella in thousands of colony-forming units (CFU) per individual female. Partial sequences of ITS and 28S rDNA regions distinguished this species from other Ambrosiella spp., which are asexual symbionts of ambrosia beetles and closely related to Ceratocystis spp. Ambrosiella roeperi sp. nov. produces sporodochia of branching conidiophores with disarticulating swollen cells, and the branches are terminated by thick-walled aleurioconidia, similar to the conidiophores and aleurioconidia of A. xylebori, which is the mycangial symbiont of a related ambrosia beetle, X. compactus. Microscopic examinations found homogeneous masses of arthrospore-like cells growing in the mycangium of X. crassiusculus, without evidence of other microbial growth. Using fungal-specific primers, only the ITS rDNA region of A. roeperi was amplified and sequenced from DNA extractions of mycangial contents, suggesting that it is the primary or only mycangial symbiont of this beetle in USA.

Influence of Ethanol Grade on Captures of Ambrosia Beetles in Tree Fruit Orchards, Ornamental Nurseries, and Lumber Yards.

Ambrosia beetles, particularly invasive species within the tribe Xyleborini, such as Xylosandrus germanus (Blandford, 1894), pose significant threats to various ecosystems and managed habitats worldwide. Monitoring these invaders is vital for effective pest management, typically accomplished through ethanol-baited traps. We compared trap efficacy using denatured ethanol versus absolute ethanol in orchards, tree nurseries, and lumber yards in northeastern Ohio, USA, finding that absolute ethanol traps captured significantly more X. germanus. Analysis revealed acetone, ethanol, and methyl isobutyl ketone in the denatured ethanol, likely impacting trap efficacy. Our study underscores the importance of using pure denatured ethanol without acetone for effective monitoring, especially for X. germanus. Exotic xyleborines dominated trap captures across various habitats, emphasizing the need for tailored pest management strategies. Further research is warranted to explore the chemical ecology of ambrosia beetles and the influence of ethanol impurities on trap effectiveness.

 Tricosauniseriata, a new species of xyleborine ambrosia beetle from Thailand (Coleoptera, Curculionidae, Scolytinae, Xyleborini).

A new species, Tricosauniseriatasp. nov., is described here. A list of Tricosa species found in Thailand with distributions and an updated key to Tricosa are also provided.

Lethal and sublethal effects of synthetic and bioinsecticides toward the invasive ambrosia beetle Xylosandrus compactus.

BACKGROUND: Exotic ambrosia beetles are emerging widespread pests of several wild and managed trees and shrubs. Xylosandrus compactus (Eichhoff) is one of the most invasive species causing damage to a broad range of host plants. Little information is available on its control, including the impact of insecticides. Bioassays were conducted to evaluate the potential of four bioinsecticides and seven synthetic insecticides in controlling X. compactus. Beetle mortality and sublethal effects on tunneling, cultivation of the mutualist fungus and reproduction were assessed. RESULTS: Concentration-mortality curves were determined for all tested insecticides. Lambda-cyhalothrin was the most toxic insecticide, showing the lowest estimated 90% and 50% lethal concentrations (LC90 and LC50 ), followed by deltamethrin and thiamethoxam. Acetamiprid caused the highest levels of mortality and brood size reduction under extended laboratory conditions. Moreover, acetamiprid, thiamethoxam and lambda-cyhalothrin caused the greatest mortality and, together with deltamethrin, strongly affected progeny occurrence inside infested galleries and beetle brood size. Among the bioinsecticides, pyrethrins significantly affected beetle survival under laboratory conditions, but not brood size in extended laboratory bioassays. Some of the tested insecticides had significant lethal and sublethal effects only when beetles were exposed to fresher residues, highlighting differences in toxicity persistence. CONCLUSION: This study provides first baseline toxicity data for synthetic insecticides and bioinsecticides with different modes of action and origin toward X. compactus, and the first evidence that several insecticides can cause multiple sublethal effects on this pest. These findings can help in building suitable integrated pest management packages against this pest. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

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.

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.

Ecuadorian Coptoborus beetles harbor Fusarium and Graphium fungi previously associated with Euwallacea ambrosia beetles.

Ambrosia beetles from the scolytine tribe Xyleborini (Curculionidae) are important to the decomposition of woody plant material on every continent except Antarctica. These insects farm fungi on the walls of tunnels they build inside recently dead trees and rely on the fungi for nutrition during all stages of their lives. Such ambrosia fungi rely on the beetles to provide appropriate substrates and environmental conditions for growth. A small minority of xyleborine ambrosia beetle-fungal partnerships cause significant damage to healthy trees. The xyleborine beetle Coptoborus ochromactonus vectors a Fusarium (Hypocreales) fungus that is lethal to balsa (Ochroma pyramidale (Malvaceae)) trees in Ecuador. Although this pathogenic fungus and its associated beetle are not known to be established in the United States, several other non-native ambrosia beetle species are vectors of destructive plant diseases in this country. This fact and the acceleration of trade between South America and the United States demonstrate the importance of understanding fungal plant pathogens before they escape their native ranges. Here we identify the fungi accompanying Coptoborus ambrosia beetles collected in Ecuador. Classification based ribosomal internal transcribed spacer 1 (ITS) sequences revealed the most prevalent fungi associated with Coptoborus are Fusarium sp. and Graphium sp. (Microascales: Microascaceae), which have been confirmed as ambrosia fungi for xyleborine ambrosia beetles, and Clonostsachys sp. (Hypocreales), which is a diverse genus found abundantly in soils and associated with plants. Phylogenetic analyses of the Fusarium strains based on ITS, translation elongation factor (EF1-α), and two subunits of the DNA-directed RNA polymerase II (RPB1 and RPB2) identified them as Fusarium sp. AF-9 in the Ambrosia Fusarium Clade (AFC). This Fusarium species was previously associated with a few xyleborine ambrosia beetles, most notably the species complex Euwallacea fornicatus (Eichhoff 1868) (Curculionidae: Scolytinae: Xyleborini). Examination of ITS and EF1-α sequences showed a close affinity between the Graphium isolated from Coptoborus spp. and other xyleborine-associated Graphium as well as the soil fungus Graphium basitruncatum. This characterization of ambrosia fungi through DNA sequencing confirms the identity of a putative plant pathogen spread by Coptoborus beetles and expands the documented range of Fusarium and Graphium ambrosia fungi.

Growth temperature effect on mandibles' ontogeny and sexual dimorphism in the ambrosia beetle Xyleborus affinis (Curculionidae: Scolytinae).

Ambrosia beetles from the genus Xyleborus are important vectors of fungal pathogens in forest and agricultural systems, yet the influence of temperature on their morphological development has been poorly studied. Because host colonization and ambrosial fungi cultivation is mostly restricted to females, it is possible to speculate on strong sexual dimorphism expression in secondary sexual characters and ecological segregation between sexes. Here, we determined the effect of different growing temperatures (17, 23, 26 and 29 °C) on mandible ontogeny of larvae and adult individuals of X. affinis, and sexual dimorphism in adults, in shape and size variation using geometric morphometrics. Mandible shape change showed significant differences in magnitude and direction through larval ontogeny among temperature treatments. Sexual shape and size dimorphism were found in adult mandibles, and the degree of sexual dimorphism was dependent on growth temperature, with a significant effect of the interaction between temperature and sex on mandible shape and size variation. Higher morphological differences were observed at the base of mandibles among temperature treatments in adults and a gradual narrowing trend with temperature increments. These findings could have consequences on feeding performance and fungus cultivation inside colonies, potentially influencing their ability to establish populations in new geographical areas.

The granulate ambrosia beetle, Xylosandrus crassiusculus (Coleoptera: Curculionidae, Scolytinae), and its fungal symbiont found in South Africa.

Xylosandrus crassiusculus (Motchulsky) is a native Asian ambrosia beetle that has been accidentally introduced to many countries of the world, presumably through the international movement of nursery, timber, and wood products. The species is known in various tropical African countries but only as far south as Tanzania on the African continent. In this study, we report X. crassiusculus and its fungal symbiont for the first time from South Africa. The species was identified using both morphological characters and COI sequence data. Xylosandrus crassiusculus were obtained from three different provinces of South Africa and represent two distinct haplotypes. The fungal symbiont, Ambrosiella roeperi, was isolated and identified using DNA sequencing and morphological characterization.

First record of the black twig borer, Xylosandrus compactus (Coleoptera: Curculionidae, Scolytinae) in Spain.

We present the first record for Spain of the black twig borer, Xylosandrus compactus, an ambrosia beetle of Asian origin, collected from an infested carob tree located in Calvià (Majorca, Balearic Islands, Spain). X. compactus is included in the EPPO Alert List, and it has been recently reported causing damages in a Mediterranean maquis ecosystem in Italy and Southern France. Here, we discuss about the first steps of management of this Invasive Alien species (IAS), the eradication plan and the hypothesis of the path of introduction in this Western Mediterranean island.

Life Cycle and Development of Coptoborus ochromactonus (Coleoptera: Curculionidae: Scolytinae), a pest of balsa.

Coptoborus ochromactonus Smith and Cognato is a recently described xyleborine ambrosia beetle pest associated with balsa, Ochroma pyramidale (Cav. Ex Lam.) Urb., in Ecuador. This pest has caused significant loss of cultivated balsa in Ecuador, but little is known of its biology and ecology. Based on examination of multiple gallery systems, this study describes the gallery pattern and life cycle of C. ochromactonus and confirms the generic identity of the symbiotic fungus. Females initiated attack, excavating a gallery perpendicular to the bole. The primary tunnel branched into a secondary tunnel at a mean 3.13 mm. This first secondary tunnel was excavated in a horizontal plane between the phloem and xylem for a mean 18.0 mm. Along its length, two tertiary tunnels were constructed on each side, measuring a mean 26.0 and 20.0 mm, respectively. Conidiophores and conidia of an unidentified Fusarium sp. grew on the tunnel walls and were fed upon by the beetle and her progeny. Coptoborus ochromactonus exhibits sexual dimorphism. The female is 2.6 mm long and possesses a round pronotum, whereas the male is 2.2 mm long and has a quadrate pronotum. The developmental stages last a mean 6, 14, 6, 4 and 30 d for egg, larva, pupa, teneral adult, and adult, respectively, when reared at 25°C. Three larval instars were present, with mean head capsule widths of 0.23, 0.31, and 0.42 mm, respectively. In general, the life cycle of C. ochromactonus is similar to those described for other xyleborine ambrosia beetle species.

Xyleborus bispinatus Reared on Artificial Media in the Presence or Absence of the Laurel Wilt Pathogen (Raffaelea lauricola).

Like other members of the tribe Xyleborini, Xyleborus bispinatus Eichhoff can cause economic damage in the Neotropics. X. bispinatus has been found to acquire the laurel wilt pathogen Raffaelea lauricola (T. C. Harr., Fraedrich & Aghayeva) when breeding in a host affected by the pathogen. Its role as a potential vector of R. lauricola is under investigation. The main objective of this study was to evaluate three artificial media, containing sawdust of avocado (Persea americana Mill.) and silkbay (Persea humilis Nash.), for rearing X. bispinatus under laboratory conditions. In addition, the media were inoculated with R. lauricola to evaluate its effect on the biology of X. bispinatus. There was a significant interaction between sawdust species and R. lauricola for all media. Two of the media supported the prolific reproduction of X. bispinatus, but the avocado-based medium was generally more effective than the silkbay-based medium, regardless whether or not it was inoculated with R. lauricola. R. lauricola had a neutral or positive effect on beetle reproduction. The pathogen was frequently recovered from beetle galleries, but only from a few individuals which were reared on inoculated media, and showed limited colonization of the beetle's mycangia. Two media with lower water content were most effective for rearing X. bispinatus.