DataSheet_1_Cry1Ba1-mediated toxicity of transgenic Bergera koenigii and Citrus sinensis to the Asian citrus psyllid Diaphorina citri.pdf

The Asian citrus psyllid, Diaphorina citri, vectors the bacterial causative agent of citrus greening disease, which has severely impacted citrus production on a global scale. As the current repeated application of chemical insecticides is unsustainable for management of this insect and subsequent protection of groves, we investigated the potential use of the bacteria-derived pesticidal protein, Cry1Ba1, when delivered via transgenic citrus plants. Having demonstrated transformation of the Indian curry leaf tree, Bergera koenigii, for Cry1Ba1 expression for use as a trap plant, we produced transgenic plants of Duncan grapefruit, Citrus paridisi, Valencia sweet orange, Citrus sinensis, and Carrizo citrange, C. sinensis x Poncirus trifoliata, for expression of Cry1Ba1. The presence of the cry1ba1 gene, and cry1ba1 transcription were confirmed. Western blot detection of Cry1Ba1 was confirmed in most cases. When compared to those from wild-type plants, leaf discs from transgenic Duncan and Valencia expressing Cry1Ba1 exhibited a “delayed senescence” phenotype, similar to observations made for transgenic B. koenigii. In bioassays, significant reductions in the survival of adult psyllids were noted on transgenic B. koenigii and Valencia sweet orange plants expressing Cry1Ba1, but not on transgenic Duncan grapefruit or Carrizo citrange. In contrast to psyllids fed on wild type plants, the gut epithelium of psyllids fed on transgenic plants was damaged, consistent with the mode of action of Cry1Ba1. These results indicate that the transgenic expression of a bacterial pesticidal protein in B. koenigii and Valencia sweet orange offers a viable option for management of D. citri, that may contribute to solutions that counter citrus greening disease.

亚洲柑橘木虱（Diaphorina citri）是柑橘黄龙病致病菌的媒介昆虫，该病害已对全球柑橘产业造成严重冲击。当前通过重复施用化学杀虫剂防控该昆虫并保护果园的策略已不可持续，因此我们探究了通过转基因柑橘植株递送细菌源杀虫蛋白（Cry1Ba1）的应用潜力。我们已成功实现印度咖喱叶树（Bergera koenigii）的遗传转化，使其表达Cry1Ba1以作为诱集植物；随后培育了表达Cry1Ba1的邓肯葡萄柚（Citrus paridisi）、瓦伦西亚甜橙（Citrus sinensis）以及卡里佐枳橙（C. sinensis x Poncirus trifoliata）转基因植株。我们证实了cry1ba1基因的存在及其转录情况，且多数样本中可通过蛋白质印迹（Western blot）检测到Cry1Ba1蛋白。与野生型植株相比，表达Cry1Ba1的转基因邓肯葡萄柚和瓦伦西亚甜橙的叶盘呈现出“延迟衰老”表型，这与转基因印度咖喱叶树的观测结果一致。生物测定结果显示，取食表达Cry1Ba1的转基因印度咖喱叶树和瓦伦西亚甜橙植株的木虱成虫存活率显著降低，但取食转基因邓肯葡萄柚或卡里佐枳橙的木虱成虫存活率无显著变化。与取食野生型植株的木虱相比，取食转基因植株的木虱肠道上皮出现损伤，这与Cry1Ba1的作用模式相符。上述结果表明，在印度咖喱叶树和瓦伦西亚甜橙中转基因表达细菌源杀虫蛋白Cry1Ba1，可为亚洲柑橘木虱的防控提供可行方案，或可为柑橘黄龙病的防治提供助力。