Approaches to vaccines against bovine Theileriosis

牛泰勒虫病疫苗的研究方法

• 批准号: 61560357
• 负责人: TAKAHASHI Kiyoshi
• 金额: $ 1.09万
• 依托单位: Rakuno Gakuen University, Facuity of Dairy Science
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1986
• 资助国家: 日本
• 起止时间: 1986 至 1987
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-61560357/
• 关键词: Bovine; Theileriosis; Vaccine; モノクローナル抗体; ウシ; Theileria sergenti; シゾント; スポロゾイト; メロゾイト; モノクロナール抗体

Morphological changes of sporozoites, schizonts and merozoites in the life cycle of T.sergenti were observed to use as the vaccine sources, we attempted to produce monoclonal antibodies against each stage of T.sergent.1. Merozoites. Thirteen monoclonal antibodies against merozoites of T.sergenti were prepared for analysis of their structural proteins. All monoclonal antibodies stained the piroplasm, but not aninfected erythrocytes by indirect fluorescent antibody test. Western blot analysis showed that 6 hybridomas tested recongnized palypeptides with molecular weights of 32,000 or 23,000. These antibodies reacted with cell membrane of merozoites by immuno electron microscopical observations, but did not react with veil and bar-structure of infected erythrocytes. The onset of intraerythrocytic multiplication was marked by the appearance of rhopties, microneme and inner membrane. It was suggested that four dividing forms in intraerythrocytic merozoites had common anatigens and infective ability, because this merozoite had the same instructural features as the sporozoites.2. Sporozoites. In the lumen of the gut of vector ticks, gametocytes appeared with in 48 hours after repletion. The sporozoites in the salivary glands of infected adult and nymph ticks maintained at 25゜C were induced by feeding ticks on rabbit after 30 days repletion. Infection level of sporozoites was high in nymph than adult ticks. A kinete penetrated to acini in the salivary glands formed syncytium and it produced over ten thousand sporozoites by budding. We tried to prepared monolonal antibody against the sporozoites.3. Schizonts, 15 cattle were inoculated sporozoites to detect schizonts in lymphocytes, schizont like protozoa were observed only in 5 cases. Althouht attenuated schizont vaccine for T.annulata were reported, shizonts of T.sergenti were not harvested sufficient volume to use for a immunoligical studies.

本研究通过观察瑟氏锥虫生活史中的子孢子、裂殖子和孢子的形态变化，以其作为疫苗来源，尝试制备针对瑟氏锥虫各阶段的单克隆抗体.裂殖体制备了13株抗瑟氏锥虫裂殖子的单克隆抗体，并对其结构蛋白进行了分析。间接荧光抗体试验表明，所有单克隆抗体均能染色梨浆，而不染色红细胞。Westernblot分析表明，6株杂交瘤均能识别分子量为32，000或23，000的多肽。免疫电镜观察表明，这些抗体与裂殖子细胞膜发生反应，而与感染红细胞的透明膜和杆状结构不发生反应。红细胞内增殖的开始以出现棒状体、微线体和内膜为标志。红细胞内裂殖子的四种分裂形式具有共同的抗原和感染能力，因为这种裂殖子具有与子孢子相同的形态学特征.子孢子在媒介蜱肠道中，配子体在饱食后48小时出现。在25 ℃条件下，将感染蜱的成蜱和若蜱饲喂家兔30天后，其唾液腺中均能诱导出子孢子。蜱若虫的子孢子感染率高于成蜱。一个动合子侵入唾液腺的腺泡内形成合胞体，并通过出芽产生上万个子孢子。并尝试制备抗子孢子单克隆抗体. 15头牛接种子孢子后，淋巴细胞中检出寄生虫，仅5头牛检出寄生虫样原虫。虽然报道了环形毛癣菌的减毒活疫苗，但收获的瑟氏毛癣菌不足以用于免疫学研究。

项目成果

高橋清志: 日本獣医学雑誌. (1987)

高桥清：日本兽医杂志（1987）。

小林延竹: 日本獣医学雑誌. (1987)

小林信武：日本兽医杂志（1987）。

Kawamoto S.;Takahashi K.;Kawai S.Kurosawa T.;and Sonoda,M.: Parasitol.Res.

Kawamoto S.；Takahashi K.；Kawai S.Kurosawa T.；和 Sonoda,M.：Parasitol.Res。

Takahashi K.,Kawai S., and Nagahara A.: "Bar-structure in bovine erythrocytes infected with Theileria sergenti" Parasitol. Res.

Takahashi K.、Kawai S. 和 Nagahara A.：“感染塞氏泰勒虫的牛红细胞中的条形结构”Parasitol。

高橋清志: 臨床獣医. 5. 69-75 (1987)

高桥清：临床兽医。5. 69-75 (1987)