Extensive, unprecedented wilt and mortality of the highly invasive tree-of-heaven (Ailanthus altissima) occurred recently within mixed-hardwood forests in south-central Pennsylvania. Until this study, the cause of the epidemic was unknown. Verticillium albo-atrum was consistently isolated from symptomatic Ailanthus seedlings and trees in areas having high levels of mortality, whereas V. dahliae was isolated from small scattered patches of diseased Ailanthus. Inoculations of potted Ailanthus seedlings in the greenhouse, as well as canopy trees in the field, revealed that both V. albo-atrum and V. dahliae were capable of infecting Ailanthus, but V. albo-atrum was more pathogenic. All Ailanthus seedlings and canopy trees inoculated with V. albo-atrum died within 3 months. In contrast, Ailanthus seedlings and canopy trees inoculated with V. dahliae became symptomatic, but 84% were still living 1 year following inoculation. We conclude that the major cause of Ailanthus wilt within forests of south-central Pennsylvania is V. albo-atrum. Pending further studies, including risk analysis, V. albo-atrum may be considered as a possible biocontrol agent for invasive Ailanthus.