Synergistic effects of acetic acid and ethanol on offspring production and gallery expansion by fungus-farming ambrosia beetles

Authors: Ranger, Christopher; BANISZEWSKI, JULIE - Former ARS Employee; PATWA, NISHA - Louisiana State University; WEI, HSIN-HO - Bowling Green State University; PACCHAIN, SUDHAN - Bowling Green State University; RAUBENOLT, JESSI - The Ohio State University; ALTLAND, MADELINE - The Ohio State University; CAMBRONERO-HEINRICHS, JUAN - Universita Di Padova; PHUNTUMART, VIPAPORN - Bowling Green State University; Reding, Michael; RASSATI, DAVIDE - Universita Di Padova; BIEDERMANN, PETER - University Of Freiburg

Submitted to: Journal of Chemical Ecology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/23/2026
Publication Date: 5/30/2026
Citation: Ranger, C.M., Baniszewski, J.A., Patwa, N., Wei, H., Pacchain, S., Raubenolt, J.A., Altland, M., Cambronero-Heinrichs, J.C., Phuntumart, V., Reding, M.E., Rassati, D., Biedermann, P.H. 2026. Synergistic effects of acetic acid and ethanol on offspring production and gallery expansion by fungus-farming ambrosia beetles. Journal of Chemical Ecology. 52. Article 50. https://doi.org/10.1007/s10886-026-01725-3.
DOI: https://doi.org/10.1007/s10886-026-01725-3

Interpretive Summary: Wood-boring ambrosia beetles are destructive insect pests of trees growing in U.S. ornamental nurseries and tree fruit orchards. Some species of invasive ambrosia beetles specifically infest trees weakened by physiological stressors. The offspring of ambrosia beetles depend on mutualistic fungi that they introduce and cultivate within stressed trees. These trees produce ethanol and acetic acid as by-products under stress, which generally suppresses fungal growth. Ethanol within stressed trees facilitates the growth of ambrosia beetles’ fungal mutualists and thus aids in host colonization, while acetic acid’s effects are unknown. Here, we evaluated the effects of acetic acid on tunneling and offspring by two invasive species, Xylosandrus germanus and Anisandrus maiche. Cut stems (i.e., bolts) of dogwood and sugar maple were infused with water, 5% ethanol, or a mixture of 5% ethanol plus acetic acid at various dilutions. Infestations by these two species of ambrosia beetles showed that more frass, larvae, pupae, and adults were found in bolts infused with 5% ethanol and 5% acetic acid treatment than in those treated with 5% ethanol and water. X-ray analysis revealed that bolts infused with a 5% ethanol-acetic acid mixture had larger gallery volumes than those infused with 5% ethanol, which were larger than the water control. A positive relationship was found between frass ejected by these ambrosia beetles and gallery volume for bolts infused with water, 5% ethanol, and a mixture of 5% ethanol and 5% acetic acid. Our study finds that acetic acid and ethanol in host tree tissues enhance the colonization by ambrosia beetles, contrary to the expectation that they would inhibit their fungal mutualists. By understanding the critical roles of acetic acid and ethanol in the colonization of trees by exotic ambrosia beetles and their associated fungi, targeted treatments for U.S. farmers of horticultural trees can be developed to disrupt this process.

Technical Abstract: Wood-boring ambrosia beetles and their offspring exhibit facultatively social behaviors while cultivating their nutritional fungal mutualists within host trees. While these beetles utilize ethanol-rich, stressed trees for farming, the specific chemical cues that influence cooperative behavior and colony fitness remain uncharacterized. Here, we demonstrate that stress-induced metabolites of anaerobic respiration—acetic acid and ethanol—act as critical signals that elicit larval cooperation and gallery expansion in Xylosandrus germanus and Anisandrus maiche. We infused wood bolts with water, 5% ethanol, or a synergistic mixture of 5% ethanol plus acetic acid at various dilutions. Infestations in the combined 5% ethanol-acetic acid treatment resulted in significantly greater gallery excavation, evidenced by increased ejected frass, as well as higher egg-laying and brood counts, compared to ethanol-only or water controls. Furthermore, because larvae obtain nutrients from both fungal gardens and fungus-infected wood, the synergistic effect of ethanol and acetic acid likely functions as a feeding stimulant that promotes excavation. X-ray micro-computed tomography revealed that galleries in the ethanol-acetic acid treatment had significantly larger volumes than those in other treatments. A positive relationship between ejected frass and gallery volume confirmed that the increased space was due to active excavation. Crucially, because these volumes were measured while only a single foundress and her immature offspring were present, the expansion highlights the specific contribution of the larvae. Our findings reveal that host-plant chemistry is a fundamental driver of multigenerational cooperation, demonstrating that larvae are active engineers in the gallery’s physical environment in response to resource quality.