Ralstonia solanacearum species complex strains are the causative agents for wilting diseases of many plants, including the economically important brown rot of potato. We developed a high-throughput virulence screen that is implemented in 96-well microtiter plates using seedlings grown in soft water agar to save space, effort, and resources. Nicotiana glutinosa was determined to be the most effective host for this assay, and we confirmed bacterial growth and systemic spread in inoculated seedlings. In our assay, N. glutinosa seeds were sown quickly and easily on top of individual water agar wells of a 96-well plate by pipetting out desired number of seeds in an aqueous suspension. They were inoculated on the same day by first touching a bacterial colony with an autoclaved toothpick and then stabbing the toothpick into the center of the water agar well. Such inoculation method resulted in inocula above a threshold of 2 × 10⁴ CFU per well achieving consistent virulence results and enabling reduction of inoculum preparation efforts to facilitate high-throughput screening. Our assay is suitable for forward genetic screening of a large number of strains, isolates or mutants for disease symptoms under both cool (20 °C) and warm (28 °C) temperature conditions before detailed studies can be narrowed down to a manageable number of desired candidates. Our virulence screen method provides a valuable tool for future work in understanding genetics of virulence of Rssc, especially cool virulence of the highly regulated race 3 biovar 2 group of R. solanacearum, leading toward development of effective control strategies.

Ralstonia solanacearum种复杂菌株是许多植物枯萎病害的病原体，包括具有重要经济意义的马铃薯褐腐病。我们开发了一种高通量毒力筛选，该筛选使用软水琼脂中生长的幼苗在 96 孔微量滴定板中实施，以节省空间、精力和资源。Nicotiana glutinosa被确定为最有效的宿主，我们证实了接种幼苗中的细菌生长和系统传播。在我们的检测中， N. glutinosa通过移出水悬浮液中所需数量的种子，将种子快速轻松地播种到 96 孔板的单个水琼脂孔顶部。他们在同一天接种，首先用高压灭菌的牙签接触细菌菌落，然后将牙签刺入水琼脂井的中心。这种接种方法导致接种超过 2 × 10 ⁴  CFU的阈值每孔实现一致的毒力结果，并能够减少接种准备工作以促进高通量筛选。我们的检测适用于在凉爽 (20 °C) 和温暖 (28 °C) 温度条件下对大量菌株、分离株或突变体的疾病症状进行前向遗传筛选，然后才能将详细研究范围缩小到可管理的数量期望的候选人。我们毒性筛选方法提供了在毒力的理解遗传学今后工作的宝贵工具RSSC ，特别是冷却高度管制的比赛中3生化型2组的致病力青枯雷尔氏菌，对有效控制策略发展的主导。