Biocontrol of <i>Ailanthus altissima</i>: inoculation protocol and risk assessment for <i>Verticillium nonalfalfae</i> (Plectosphaerellaceae: Phyllachorales)

<i>Verticillium nonalfalfae</i> has been proposed as a biocontrol for invasive <i>Ailanthus altissima</i> (tree-of-heaven) in Ohio, Pennsylvania and Virginia. However, previous studies evaluating this potential biocontrol utilised a conidial suspension with a short shelf life as inoculum. Anticipating future expanded use of <i>V. nonalfalfae</i>, we evaluated other inoculum formulations, inoculation protocols and sensitivity of non-target (non-<i>Ailanthus</i>) plant species within Pennsylvania. The most effective inoculum formulation, with an extended shelf life, was prepared by mixing water with stored, refrigerated soil containing <i>V. nonalfalfae</i>. Less successful, but positive infections were obtained by simply using infected <i>Ailanthus</i> wood and leaves as inoculum. Monthly inoculation of <i>Ailanthus</i> trees demonstrated that the optimal time for successful inoculations was April to May, but limited infections were achieved during all months, including the winter. The health of <i>Ailanthus</i> and non-target species was evaluated within a decade-old natural Verticillium wilt epicentre, where all mature <i>Ailanthus</i> trees had been killed by <i>V. nonalfalfae</i>. Verticillium wilt was observed on a few small <i>Ailanthus</i> trees, likely newly established seedlings, whereas non-target species were asymptomatic. Our findings reveal that soil formulated and natural inocula are effective biocontrols against <i>Ailanthus</i>, and <i>V. nonalfalfae</i> appears to pose little threat to non-target plants.

非苜蓿黄萎病菌（Verticillium nonalfalfae）已被提议作为美国俄亥俄州、宾夕法尼亚州及弗吉尼亚州入侵物种臭椿（Ailanthus altissima，又名天堂树）的生物防治制剂。然而，此前评估该潜在生防制剂的研究均采用保存期较短的分生孢子悬浮液作为接种体。鉴于未来有望扩大非苜蓿黄萎病菌的应用范围，我们在宾夕法尼亚州针对其他接种体制剂、接种规程以及非靶标（非臭椿）植物的敏感性开展了相关评估。本研究筛选出的高效接种体制剂可延长保存期，其制备方式为将清水与冷藏保存的、携带有非苜蓿黄萎病菌的土壤混合。仅使用受侵染的臭椿木材与叶片作为接种体也可成功实现侵染，但防治效果稍逊。对臭椿植株的月度接种试验表明，成功接种的最佳时段为4月至5月，但全年（包括冬季）均可实现有限侵染。我们在一处已有10年历史的天然黄萎病（Verticillium wilt）发病中心内评估了臭椿与非靶标植物的健康状况，该疫源地内所有成熟臭椿植株均已被非苜蓿黄萎病菌致死，研究仅在少量小型臭椿植株（大概率为新近定植的实生苗）上观察到黄萎病症状，而非靶标植物均未表现出染病迹象。本研究结果显示，土壤制剂与天然接种体均可作为防治臭椿的有效生物防治手段，且非苜蓿黄萎病菌对非靶标植物几乎无威胁。