新发现的水椰八角铁甲天敌啮小蜂对寄主免疫抑制与调控机理

In the system of invasive pest Octodonta nipae Maulik and its endoparasitic wasps Tetrastichus brontispae Ferrière, the types and functions of pupa hemocyte in O. nipae will be studied with the technology of physiological ecology and molecular biology, to reveal invasive characteristics of invasive pest from the angle of celluar immunity. Kinds of parasitic factors and their timeliness will be analyzed to determine kinds of protein existed in venom and their regulations on host immune system. Changes of the number of total hemocyte cells (THC), and their extension, cytophagy and encapsulation in O. nipae pupa parasitized by T. brontispae will be studied, in order to clarify responses of the key types of hemocyte and their physiological mechanism of the hemocyte immune response. The correlations among antioxidase, phenoloxidases (PO) and ROS after the parasitism by T. brontispae will be investigated to expound the response of O. nipae parasitism by T. brontispae in the immune reactivity and its effect to the function such as humoral melanization. Also, this research will uncover the physiological and biochemistry process in the host which caused by the parasitic wasp for further explaining the mechanism that T. brontispae damages the host humoral immunity. This research will not only attribute to further analysis about the immune interaction between T. brontispae and host to enhance the nursing of parasitic wasp and improve the natural control effect in the field, but also provide a new idea and approach for the development of new-type pest inhibitor which takes the insect immune system as a target.

本项目以入侵害虫水椰八角铁甲-椰扁甲啮小蜂系统为对象，采用生理生态学和分子生物学技术，分析水椰八角铁甲蛹血细胞类型及其功能，从细胞免疫角度揭示入侵害虫的入侵特性；分析椰扁甲啮小蜂寄生因子的种类及其时效性，明确毒液中存在的蛋白种类及其对寄主免疫系统的调控；通过对啮小蜂寄生引起水椰八角铁甲蛹血细胞数量、延展、吞噬和包囊作用等的变化研究，明确关键血细胞类型的响应及对铁甲蛹血细胞免疫影响的生理机制；探明啮小蜂寄生后，铁甲蛹血淋巴中抗氧化酶、酚氧化酶PO和活性氧自由基ROS间相互关系，阐明害虫的免疫应激反应及其对黑化反应等功能的影响，揭示寄生蜂毒液对寄主生理和生物化学调控的过程，进一步解释啮小蜂破坏寄主的体液免疫反应的机理。该研究不仅有助于深入剖析椰扁甲啮小蜂与寄主的免疫互作效应，加强寄生蜂保育和提高田间自然控害效果，而且对于开发以昆虫免疫系统为靶标的新型害虫抑制剂等提供新的思路和途径。

本项目以入侵害虫水椰八角铁甲—椰扁甲啮小蜂系统为对象，采用生理生态学和分子生物学技术，分析了具有免疫活性的水椰八角铁甲蛹的浆血细胞、粒血细胞和类绛色细胞等的PO活性、延展、吞噬和包囊等特性，从细胞免疫角度揭示其入侵特性；明确了椰扁甲啮小蜂蜂卵表面不含PDVs和VLFs等因子，其主要的寄生因子是毒液、卵巢蛋白和VLPs；椰扁甲啮小蜂寄生后，其毒液关键蛋白可显著抑制寄主蛹血细胞的数量、延展、吞噬和包囊作用等，且寄主的免疫活性在寄生前期主要是局部活性调节，在寄生后期主要是系统免疫调节；进一步揭示了毒液蛋白抑制细胞免疫的分子机理，主要是啮小蜂寄生后，其毒液蛋白基因TbCRT、TbRhoGAP均显著抑制寄主蛹的钙网蛋白OnCRT、整合蛋白OnINT、OnRac1基因的表达，抑制寄主血细胞的肌动蛋白F-actin基因的表达，进而抑制寄主关键血细胞的粘附、延展、肌动蛋白聚合能力和包囊反应等，从而达到抑制寄主的细胞免疫功能的目的；阐明了椰扁甲啮小蜂寄生后，寄主水椰八角铁甲蛹血淋巴中抗氧化酶、酚氧化酶PO和活性氧自由基ROS间的相互关系，揭示了寄主的免疫应激反应及其对黑化反应等功能响应等，明确了啮小蜂毒液对寄主生理和生物化学调控的过程，啮小蜂寄生后抑制了寄主的酚氧化酶PO活性和血细胞的黑化效能，激活了寄主谷胱甘肽氧化酶（GSH-Px）、谷胱甘肽硫转移酶（GST）、谷胱甘肽还原酶（GR）等，但不影响血淋巴内的蛋白浓度；啮小蜂毒液活性蛋白基因serpin竞争寄主serpin与寄主serine（OnPPAE）的作用位点，从而抑制了寄主PO的激活和级联反应，阻止了寄主黑化反应等，进一步明确了毒液蛋白调控寄主体液免疫的反应途径及分子机理。该研究不仅有助于深入剖析椰扁甲啮小蜂与寄主的免疫互作效应，加强寄生蜂保育和提高田间自然控害效果，而且对于开发以昆虫免疫系统为靶标的新型害虫抑制剂等提供新的思路和途径。