PCR multiplexes discriminate Fusarium symbionts of invasive Euwallacea ambrosia beetles that inflict damage on numerous tree species throughout the United States

Authors: SHORT, DYLAN P - West Virginia University; O Donnell, Kerry; STAJICH, JASON - University Of California; HULCR, JIRI - University Of Florida; KIJIMOTO, TEIYA - West Virginia University; BERGER, MATTHEW - West Virginia University; MACIAS, ANGIE - West Virginia University; SPAHR, ELLIE - West Virginia University; BATEMAN, CRAIG - University Of Florida; ESKALEN, AKIF - University Of California

Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/12/2016
Publication Date: 1/9/2017
Citation: Short, D.P.G., O'Donnell, K., Stajich, J.E., Hulcr, J., Kijimoto, T., Berger, M.C., Macias, A.M., Spahr, E.J., Bateman, C.C., Eskalen, A., et al. 2017. PCR multiplexes discriminate Fusarium symbionts of invasive Euwallacea ambrosia beetles that inflict damage on numerous tree species throughout the United States. Plant Disease. 101(1):233-240. https://doi.org/10.1094/PDIS-07-16-1046-RE.
DOI: https://doi.org/10.1094/PDIS-07-16-1046-RE

Interpretive Summary: Six different species of the exotic Asian ambrosia beetle genus Euwallacea have invaded the U.S. within the past 15 years. These beetles farm closely related phytopathogenic fungi within the Fusarium solani species complex as a source of food. These invasive mutualists pose a serious threat to urban landscapes, forests, and avocado production worldwide. To date, these pests have caused canker and dieback disease on over one hundred native and non-native tree species within the U.S., including avocado in Florida and California. Current methods for identifying the plant pathogenic Fusarium species these beetles carry around and cultivate in their galleries are time consuming and require costly DNA sequence data. To address this problem, we mined the whole genome sequence of the six invasive Fusarium species within the U.S. and used it to design species specific molecular diagnostics. The assays were shown to rapidly and accurately identify the six foreign Fusarium species. The molecular diagnostic assays will be of interest to plant pathologists, entomologists, and plant quarantine officials who are interested in tracking the geographic distribution and host range of the phytopathogenic Fusarium species that are currently within the U.S. as well as state and federal efforts to prevent further introductions.

Technical Abstract: Asian Euwallacea ambrosia beetles vector Fusarium mutualists. The ambrosial fusaria are all members of the Ambrosia Fusarium Clade (AFC) within the Fusarium solani species complex (FSSC). Several Euwallacea-Fusarium mutualists have been introduced into non-native regions and have caused varying degrees of damage to orchard, landscape, and forest trees. Knowledge of symbiont fidelity is limited by current identification methods, which typically requires analysis of DNA sequence data from beetles and the symbionts cultured from their oral mycangia. Here, polymerase chain reaction (PCR)-based diagnostic tools were developed to identify the six Fusarium symbionts of exotic Euwallacea currently known within the U.S. Whole genome sequences were generated for representatives of six AFC species plus F. ambrosium and aligned to the annotated genome of F. euwallaceae. Taxon-specific binding sites were identified that rapidly distinguish the AFC species currently within the U.S. PCR specificity, reliability, and sensitivity were validated using a panel of 72 Fusarium isolates including 47 reference cultures. Culture-independent multiplex assays accurately identified the six AFC fusaria using DNA isolated from beetle heads. The PCR assays were used to show that E. validus is exclusively associated with AF-4 throughout its sampled range within eastern North America. The rapid assay will support federal and state agency efforts to monitor spread of these invasive pests and mitigate further introductions.