Native and exotic Juglans nigra seedlings recruit distinct mycobiomes that differentiate with emergent disease variation

Abstract

Under global change, the transformation of landscapes and anthropogenic movement of organisms lead to novel biotic interactions that affect ecosystem health. A prior field-based metabarcoding survey revealed that Juglans nigra harbors a novel, low-diversity mycobiome outside of its native range that may contribute to the severity of thousand cankers disease (TCD), a destructive insect–disease complex affecting walnut in the western United States and Europe. The previous metabarcoding study also identified differences in mycobiome composition among plant genotypes and between naturally infected TCD-positive trees and healthy trees. In this study, seeds were germinated in the field in two locations: Martell Forest, Indiana (TCD negative) and Walla Walla, Washington (TCD positive). After one season, seedlings were transplanted to a greenhouse and inoculated with Geosmithia morbida, the fungal pathogen component of TCD. Fungi were isolated from roots and cankers and identified morphologically and with the internal transcribed spacer (ITS) and large subunit (LSU) regions of rDNA to test the extent to which trees in the two locations recruited different mycobiomes from their environment. Stem necrosis around inoculation points was measured to assess the extent to which necrosis was influenced by distinct mycobiomes and to examine covariation between necrosis and the mycobiome. Additionally, seedlings from Walla Walla were control inoculated to evaluate the effect of inoculation on the mycobiome. Root–shoot ratio was also measured to investigate the relationship between necrotic area and belowground versus aboveground investment. Canker area was marginally higher in Walla Walla, and covariation with the root–shoot ratio was generally negative but positive in one plot in Walla Walla. Roots from inoculated seedlings from Martell Forest had distinct and marginally more diverse mycobiomes than those from Walla Walla. Root and canker mycobiomes from inoculated seedlings differed between locations and with necrotic area. Mycobiome differences between seedlings with large and small cankers were higher in Martell Forest plots with greater variability in necrotic area, whereas in Walla Walla, plots with more variable necrotic area showed smaller mycobiome differences between seedlings with large and small cankers. These findings suggest that plants in novel habitats recruit low-diversity mycobiomes that influence disease etiology.