【作者】 张焘；

【导师】 叶恭银；

【作者基本信息】 浙江大学 ， 农业昆虫与害虫防治， 2003， 硕士

【摘要】 为了探明丽蝇蛹集金小蜂如何调控寄主免疫的机理，本论文就该蜂毒腺结构，毒液成份以及寄生对寄主血细胞数量和组成，血淋巴可溶性蛋白组份、含量，以及糖含量的变化作了初步的研究，结果如下： 未寄生的棕尾别麻蝇Boettcherisca peregrina蛹血淋巴中主要存在两种血细胞，即浆血细胞和颗粒细胞。棕尾别麻蝇蛹被丽蝇蛹集金小蜂Nasonia nitripennis寄生后1d浆血细胞的比例显著高于同期未寄生蛹，颗粒细胞比例也显著高于同期未寄生蛹。但寄生2～5d的蛹与同期未寄生蛹的颗粒细胞、浆血细胞占总细胞数的比例没有显著差异。数量方面，被寄生蛹的颗粒细胞和浆血细胞数量在寄生后1～5 d均显著高于同期未寄生蛹。寄生1 d后寄生蛹与未寄生蛹的血细胞总数（THC）间没有明显差异。寄生后2～5d，被寄生蛹的THC显著高于同期未寄生蛹，分别为同期为寄生蛹的3.2、1.8、2.1和2.3倍。 毒液器官的超微结构研究结果表明，毒囊包括一层外肌肉层，单层分泌细胞层和平行于内腔，贴近内膜的单层鳞状细胞层。内膜内陷到各个分泌细胞间，形成一个外面由管状细胞包裹的管腔。而毒腺则是由立方形分泌细胞形成的单细胞层，和平行于内腔，贴近内膜的单层鳞状细胞层共同构成。丽蝇蛹集金小蜂毒囊肌肉层是由纵向的肌肉组成的，相对较厚，毒囊没有分泌细胞，并受神经支配，寄生时毒液的注射是由肌肉层的收缩而造成的。丽蝇蛹金小蜂毒腺由单立方形分泌细胞层，和内部贴近内腔的鳞片状的细胞层共同构成的。分泌细胞核相对较大，并且有分散的染色质，常常可以看到分泌的毒液颗粒。围绕着分泌细胞的粗面内质网及高尔基体形成的小泡浓度充分表明了分泌器官具有活跃的分泌和运输功能，但在毒囊细胞中则很少可以看到，充分表明分泌功能主要在毒腺，而毒囊的主要功能则是贮藏，分泌功能则较弱。 利用PDQuest^TM二维电泳图像分析软件分析毒液的双向电泳图谱，可以识别到100多个毒液蛋白质点。从蛋白质的分子量分布范围来分析，分子量从2KDa到145KDa的区域均有蛋白质点，大于70KDa的蛋白质点很少，分子量在2KDa-40KDa蛋白质点较密。从蛋白质点的等电点分别范围来分析，偏于酸性端的蛋白质点明显多于碱性端的蛋白质点，大多数蛋白质点的等电点为中性或酸性。 丽蝇蛹集金小蜂羽化后0．5．6d毒液含量整体呈逐渐上升的趋势，其中第6d蛋白质浓度达到最高。而第7d蛋白质浓度则急剧下降，显著低于第6d的浓度。 寄生后 l－4d蛹血淋巴蛋白浓度相对稳定，在22．95－43．sling／ffil之间变化。未寄生血淋巴蛋白浓度均极显著高于同期寄生蛹，分别是寄生蛹血淋巴蛋白浓度的2．2、1．9、2．4和1．5倍。未寄生蛹血淋巴蛋白浓度1～4d逐渐下降；而寄生蛹则与此不同，寄生后l—3d逐渐下降，到了4d则稍稍上升，保持在一个稳定的水平。100 KDa的蛋白在棕尾别麻蝇蛹寄生初期含量较低，寄生后第 3 d寄生蛹中此蛋白的含量明显升高，而到同期未寄生蛹中第4d此蛋白的含量才有所上升，但远达不到被寄生蛹中的水平。贮藏蛋白在寄生蛹体内降解明显，同时，32 KDa和 28 KDa蛋白在寄生 2 d后含量下降。 寄生后 1叫 d蛹血淋巴中可溶性糖含量较为稳定，而未寄生蛹血淋巴中可溶性糖的含量在寄生后上升，寄生Zd后达到最高而后又下降。寄生蛹血淋巴中可溶性糖含量在寄生后1～4d显著低于同期未寄生蛹；未寄生蛹血淋巴可溶性糖含量在Zd后达到最高，而寄生蛹则在4d后达到最高。更多还原

【Abstract】 In order to investigate the mechanism how to regulate the host’s immunity by Nasonia nitripennis ,this article makes the preliminary study in the following topics: the structure of venom gland, change in constitution and count of hemocyte population, and the concentration of soluble proteins and soluble sugars after parasitization. The result is summarized as following:There are two kinds of hemocyte in hemolymph of Boettcherisca peregrina pupae, plasmatoctes(PLs) and granular cells(GRs). In the first day after parasitization, the percentage of PLs is significantly higher in parasitized pupae than unparasitized ones, so as the percentage of GRs. But from the second day to the fifth day after parasitized, the percentage of GRs and PLs aren’t significantly different between the parasitized pupaes and in unparasitized pupaes.From the first day to the fifth day after parasitized, the count of GRs and PLs is significantly higher in parasitized pupaes than in unparasitized pupaes. After the first day parasitzed, total hemocyte count (THC) isn’t significantly different between parasitized pupaes and unparasitized ones, and from the second day to the fifth day, it is significantly higher in parasitized pupaes than unparasitized ones, and THC of parasitized pupaes is 3.2, 1.8, 2.1 and 2.3 times than that of unparasitized pupaes.The venom reservoir is consisted of an outer musular sheath, a single layer of secretory cells and an inner single layer of squamous cells adjacents to the intima which lines the lumen. The intima invaginates into each secretory cell and forms a duct which is surrounded by duct cells. And the gland filaments consist of a single layer of cuboidal secretory cells and an inner layer of squamous cells adjacent to the intima which lines the lumen. The reservoir apparatus are bordered distally by longitudinal muscles which form a relatively thick muscular sheath, and are innervated, the rapid injection of the venom may be facilitated by muscular contraction of the reservoir. In the gland filaments, the secretory cells of venom apparatus are cuboidal and form a continuous layer which is limited basically by a granular lamella. Nuclei of secretory cells are relatively large and contain scatteredchromatin. Concentration of sER and Golgi bodies around the secretory appartus indicate the possibility of active transport through the secretory appartus. But there can’t be seen in the reservoir’s cells. From the fact above, we can make a conclusion that the secretory function mainly acted by the gland filament, and the main function of reservoir is storing the venom.From the utilization of the software PDQuest檛o exmaine the photo of venom’s electrophoresis, there are 100 more protein dots can be deteced. The protein’s weight are from 2 KDa to 145 KDa, the dots mainly scattered in the weight from 2 KDa to 40 KDa, only several dots above the 70 KDa. Using 2-DE we know that venom protein are mainly acid protein, and molecular weight is mainly between 2-40 KDa.From the zero point fifth day to the sixth day, the concentration of venom is ascending, and there comes the highest concentration. But on the seventh day, the concentration is sharply droping, and high lower than the concentration of sixth day.The concentration of protein is relatively stably in the hemolymph from the first day to the fourth day, changing between 22.95mg/ml and 43. 51mg/ml, it is is 3.2, 1.8, 2.1, 2.3 times higher in unparasitized pupaes than the in parasitized pupaes. The concentration of protein in the hemolymph from the unparasitized pupae is droping from the first day to the fourth day. The concentration of protein in the hemolymph from parasitized pupaes are droping from the first day to the third day, but on the fourth day , concentration are rising. The concentration of the protein of 100 KDa is lower in the hemolymph of former stage, the concentration is rising on the third day in the parasitized pupaes, but rising on the fourth day in the unparasitized pupaes, much lower than that in parasitized pupaes. The c更多还原