The vector-borne disease Huanglongbing (HLB) causes severe economic losses to citrus production worldwide with no available cure. Herein, we applied virus-induced gene silencing technology to engineer citrus that preferentially attracted and specifically killed Diaphorina citri, the vector associated with HLB. We engineered the infectious citrus tristeza virus (CTV-T36) clone to carry three truncated genes. The triple construct (CTV-tAwd-tWnt-tPDS) produces small interfering RNAs (siRNAs) against phytoene desaturase, PDS, to yield a phenotype with visual, olfactory, and gustatory cues that preferentially attracted D. citri. In addition, siRNAs targeted two genes related to flight in D. citri, abnormal wing disc (DcAwd) and wingless (DcWnt), that caused wing malformations and decreased survival in psyllids that fed on plants inoculated with the engineered virus. During two successive generations, D. citri reared on CTV-tAwd-tWnt-tPDS-inoculated plants exhibited higher mortality across life stages as well as reduced fecundity and fertility as compared with those reared on non-infected plants or CTV-wt-inoculated plants. Furthermore, CTV-tAwd-tWnt-tPDS-inoculated plants shortened the lifespan of D. citri by more than 20 days. Morphological abnormalities were noted in those adults that did successfully emerge on plants inoculated with CTV-tAwd-tWnt-tPDS, including cocked wings with a bowl-shaped depression and/or a convex shape. Phloem sap from CTV-tAwd-tWnt-tPDS-inoculated plants decreased the survival of D. citri adults, confirming that siRNAs were present in the sap of these plants. Collectively, we provide proof of concept for a novel variant of the attract-and-kill method where the cultivated crop is potentially transformed into a hyper-attractive population and transmission sink for a phytopathogen vector.

中文翻译：

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病毒诱导的基因沉默同时利用植物和昆虫的“吸引和杀死”特性来控制黄龙病


病媒传播疾病黄龙病 （HLB） 对全球柑橘生产造成了严重的经济损失，而且没有可用的治疗方法。在此，我们应用病毒诱导的基因沉默技术来设计柑橘，优先吸引并特异性杀死与 HLB 相关的载体 Diaphorina citri。我们设计了传染性柑橘 tristeza 病毒 （CTV-T36） 克隆，使其携带三个截短基因。三重构建体 （CTV-tAwd-tWnt-tPDS） 产生针对植物烯去饱和酶 （PDS） 的小干扰 RNA （siRNA），以产生具有视觉、嗅觉和味觉线索的表型，优先吸引柠檬果红。此外，siRNA 靶向柑橘木虱中与飞行相关的两个基因，异常翼盘 （DcAwd） 和无翼 （DcWnt），这会导致以接种工程病毒的植物为食的木虱出现翅膀畸形和存活率降低。在连续两代中，与在未感染植物或 CTV-wt 接种的植物上饲养的柑橘石楠相比，在 CTV-tAwd-tWnt-tPDS 接种的植物上饲养的柑橘石栉在生命阶段表现出更高的死亡率，以及繁殖力和生育能力降低。此外，接种 CTV-tAwd-tWnt-tPDS 的植物使柑橘 D. citri 的寿命缩短了 20 天以上。在接种 CTV-tAwd-tWnt-tPDS 的植物上成功出现的成虫中观察到形态学异常，包括具有碗状凹陷和/或凸形的翘起翅膀。来自 CTV-tAwd-tWnt-tPDS 接种植物的韧皮部汁液降低了柑橘石桐成虫的存活率，证实了这些植物的汁液中存在 siRNA。 总的来说，我们为吸引和杀死方法的新变体提供了概念验证，其中栽培作物有可能转化为极具吸引力的种群和植物病原体载体的传播汇。