Malarial development in mosquito and vaccine development. Mechanism of malarial sporozoite accumulation in the salivary glands of Anopheline mosquitoes.

我们研究了伯氏疟原虫子孢子在斯氏按蚊唾液腺中聚集的机制。通过对感染伯氏疟原虫的中肠进行透射电子显微镜观察，结果表明：(1)摄血24小时后，Ookinate通过细胞连接到达中肠基膜；(2)感染后两天，卵母细胞停止运动，变圆，直径为6微米。(3)感染后第8天，卵囊内可见大量核发育成子孢子。(4)感染后14d，卵囊发育成熟卵囊，核心周围有大量稀释子存在。为了观察子孢子进入唾液腺的情况，我们用2.5%的臭氧固定活蚊子，并在2小时后解剖取出唾液腺。扫描电子显微镜和透射电子显微镜观察结果表明：(1)表面有子孢子存在，部分子孢子穿透唾液腺。(2)在唾液腺内，子孢子从细胞外进入细胞质，聚集在分泌腔内。我们从蚊子唾液腺中提取了mRNA，合成了cDNA，并将其连接到噬菌体中，构建了cDNA库。用唾液腺单抗进行筛选，筛选出4个阳性克隆。4个克隆的插入片段大小分别为1450bp、360bp、220bp和790bp。

We studied the mechanism of malarial sporozoite accumulation of Plasmodium berghei in the salivery gland of Anopheles stephensi.1. We got following results with the observation of midgut infected with Plasmodium berghei by transmission electron microscope (TEM).(1) Ookinate reached the basal lamina of midgut through the cell junction 24 hours after blood was ingested. (2) Two days after infection, ookinate whose diameter was 6 um has stopped moving and became round. (3) Eight days after infection, there were many nuclei which developed into sporozoites inside the oocyst. (4) Fourteen days after infection, oocyst developed matured oocyst, and many sparozoite exsisted around the core.2. To observe the penetration of the sporozoites into salivary glands, we fixed living mosquitoes by 2.5% osmium and dissected them to remove the salivary glands after 2 hours. We got following results by scanning electron microscope and TEM.(1) Sporozoites exsisted on the surface and some of them were penetrating in the salivary glands. (2) Inside of the salivary glands, sporozoites entered the cytoplasm of the cell from outside and gathered in the secretory cavity.3. We extracted mRNA from salivery gland of mosquito and synthesized cDNA.The cDNA was ligated lambdagtll phage and cDNA libraries were produced. These were screened by the monoclonal antibody of salivery gland and 4 possitive clones were selected. The size of insert of 4 clones were 1450bp, 360bp, 220bp and 790bp.