Walnut Witches'-Broom Disease Threatens Butternut Restoration Efforts in Indiana.

Due to the devastating effects of butternut canker disease, efforts to protect the endangered butternut () tree through resistance breeding have been a primary focus of forest restoration efforts. Walnut witches'-broom (WWB) disease poses a serious threat to these restoration efforts. This study sought to confirm the presence of the WWB disease phytoplasma, ' Phytoplasma pruni', in butternut research plantings in Indiana using molecular methods and Sanger sequencing and to identify butternut families affected by the disease. We also sought to better understand the incidence of the WWB phytoplasma in asymptomatic trees and asymptomatic branches of symptomatic trees to better direct management decisions. Sanger sequencing confirmed the presence of the WWB phytoplasma in the butternut restoration plantings, the first confirmation in Indiana based on sequencing to our knowledge, in both symptomatic and some asymptomatic trees. In addition, the WWB phytoplasma was detected in asymptomatic branches of symptomatic trees, indicating that phytoplasma infection is not necessarily localized to symptomatic tissues in a tree. Trees with positive molecular confirmation of the WWB phytoplasma consisted of 23 different butternut families and one family of Japanese walnut (), which is considered to be one of the most susceptible species to WWB disease. Based on these findings, future studies should prioritize identifying the hybridity and pedigrees of families and their susceptibility to WWB disease to aid in butternut restoration efforts.