The worldwide invasion history of a pest ambrosia beetle inferred using population genomics.

Xylosandrus crassiusculus, a fungus-farming wood borer native to Southeastern Asia, is the most rapidly spreading invasive ambrosia species worldwide. Previous studies focusing on its genetic structure suggested the existence of cryptic genetic variation in this species. Yet, these studies used different genetic markers, focused on different geographical areas and did not include Europe. Our first goal was to determine the worldwide genetic structure of this species based on both mitochondrial and genomic markers. Our second goal was to study X. crassiusculus' invasion history on a global level and identify the origins of the invasion in Europe. We used a COI and RAD sequencing design to characterize 188 and 206 specimens worldwide, building the most comprehensive genetic data set for any ambrosia beetle to date. The results were largely consistent between markers. Two differentiated genetic clusters were invasive, albeit in different regions of the world. The markers were inconsistent only for a few specimens found exclusively in Japan. Mainland USA could have acted as a source for further expansion to Canada and Argentina through stepping stone expansion and bridgehead events. We showed that Europe was only colonized by Cluster II through a complex invasion history including several arrivals from multiple origins in the native area, and possibly including bridgehead from the United States. Our results also suggested that Spain was colonized directly from Italy through intracontinental dispersion. It is unclear whether the mutually exclusive allopatric distribution of the two clusters is due to neutral effects or due to different ecological requirements.

Effects of permethrin on ambrosia beetles (Coleoptera: Curculionidae: Scolytinae) in ornamental nurseries.

Exotic ambrosia beetles (Coleoptera: Curculionidae: Scolytinae), such as Xylosandrus crassiusculus (Motschulsky), Xylosandrus germanus (Blandford), and Xylosandrus compactus (Eichoff) are serious pests in southeastern ornamental nurseries. Preventative pyrethroid trunk sprays effectively reduce boring damage. However, it is unclear how pyrethroids such as permethrin prevent attack. Thus, the objective was to determine how permethrin-treated bolts interact with invading ambrosia beetles. In 2022, a study with 2 independent trials was conducted in a nursery on red maple (Acer rubrum L.), bolts during March and April, respectively. The treatments were (i) nonbaited, nontreated bolt, (ii) ethanol baited bolt, (iii) nonbaited bolt + glue [painted on bolt], (iv) ethanol baited bolt + glue, (v) ethanol baited bolt + glue + permethrin, (vi) ethanol baited bolt + glue + permethrin + verbenone, and (vii) ethanol baited bolt + glue + verbenone. Ambrosia beetles trapped on glue, beetles which fell into the pail with soap solution under the bolts, and entry holes on bolts were quantified. Permethrin prevented beetle attacks but did not reduce the number of ambrosia beetles landing on the treated bolts. Verbenone reduced ambrosia beetles from landing on the bolts but did not prevent boring into bolts. The numbers of ambrosia beetles in soapy water were not significantly different among treatments. Ambrosia beetles are landing on permethrin-treated bolts but not boring into the bolts, implying that fresh permethrin residues may not be necessary for ambrosia beetle management.

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.

Erwiniaceae bacteria play defensive and nutritional roles in two widespread ambrosia beetles.

Ambrosia beetles are fungal-growing insects excavating galleries deep inside the wood. Their success as invaders increased scientific interest towards them. However, most studies on their microbiota targeted their fungal associates whereas the role of bacterial associates is understudied. To explore the role of abundant microbial associates, we isolated bacteria from active galleries of two widespread ambrosia beetles, Xylosandrus crassiusculus and X. germanus. These isolates were classified within the Erwiniaceae family and through a phylogenetic analysis including isolates from other insects we showed that they clustered with isolates obtained from ambrosia and bark beetles, including Erwinia typographi. The whole genome analysis of the isolate from active galleries of X. crassiusculus suggested that this bacterium plays both a nutritional role, by providing essential amino acids and enzymes for the hydrolysis of plant biomass, and a defensive role, by producing antibiotics. This defensive role was also tested in vitro against fungi, including mutualists, common associates, and parasites. The bacteria inhibited the growth of some of the common associates and parasites but did not affect mutualists. Our study supported the hypothesis of a mutualist role of Erwiniaceae bacteria in ambrosia beetles and highlighed the importance of bacteria in maintaining the symbiosis of their host with nutritional fungi.

Cerambycid Pheromones 3,2-Hydroxyketones Affect Catches of Some Bark and Ambrosia Beetles (Coleoptera: Curculionidae) in Ethanol-Baited Multiple-Funnel Traps in Southeastern United States.

In 2012-2013, we assessed the interactive effects of the cerambycid pheromones syn-2,3-hexanediol, 3-hydroxyhexan-2-one, and 3-hydroxyoctan-2-one on catches of bark and ambrosia beetles (Coleoptera: Curculionidae) in ethanol-baited multiple-funnel traps in north Georgia and South Carolina. We found that catches for nine of eleven species of ambrosia beetles in ethanol-baited traps were either unaffected or enhanced by the addition of 3,2-hydroxyketones. Similarly catches of five species of bark beetles were either unaffected or enhanced by the addition of 3,2-hydroxyketones. In particular, catches of Xylosandrus crassiusculus (Motschulsky), Cnestus mutilatus (Blandford), and Monarthrum fasciatum (Say) in ethanol-baited traps increased with the addition of 3-hydroxyhexan-2-one and/or 3-hydroxyoctan-2-one. Catches of the bark beetles Hylocurus rudis (LeConte) and Hypothenemus rotundicollis (Eichhoff) were enhanced by the addition of 3-hydroxyhexan-2-one and 3-hydroxyoctan-2-one, respectively. syn-2,3-Hexanediol had no effect on catches of bark and ambrosia beetles in ethanol-baited traps. Our data provide support for the use of ethanol + cerambycid pheromones for targeting non-native species of bark and ambrosia beetles as well as cerambycids in detection programs.

Effects of Trap, and Ethanol Lure Type and Age on Attraction of Ambrosia Beetles (Coleoptera: Curculionidae).

Ambrosia beetles (Xylosandrus spp.) are pests of woody ornamental nurseries and tree nut orchards. Growers use ethanol-infused bolts and ethanol-mediated bottle traps for monitoring ambrosia beetles in the spring. To refine these monitoring tools, we investigated the 1) sensitivity of the commercially available ethanol pouches placed on bolts compared with that of ethanol-infused bolts for ambrosia beetle attacks; and 2) the effect of aging of ethanol pouches on ambrosia beetle captures in bottle traps. Experiments were conducted in ornamental nurseries and pecan orchards in 2019 and 2020. For objective 1, the treatments included a bolt infused with locally purchased ethanol, an ethanol pouch attached to a bolt, and a control (bolt without ethanol). For objective 2, ethanol pouches were initially field-aged for 0, 1, 2, 3, and 4 wk and ambrosia beetle captures were documented weekly for 4 wk. Ethanol pouches older than 5 wk were replaced with fresh pouches at the pecan sites while ethanol pouches at nursery sites were aged up to 8 wk. The ethanol-pouch comparison study showed that the number of ambrosia beetle attacks was significantly greater on ethanol-infused bolt traps than with the ethanol pouches on bolts at the nursery and pecan sites in both years. The age of the ethanol pouches did not affect captures of Xylosandrus crassiusculus (Motschulsky) and X. germanus (Blandford) in bottle traps within 8 wk of deployment. Results provide practical implications into the sensitivity of the different ethanol lure types in association with bolt traps and the longevity of the ethanol pouches for grower use.

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.

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.