Analysis on protective antigen of Babesia parasites by developmental engineering

巴贝虫寄生虫保护性抗原的发育工程分析

• 批准号: 09460143
• 负责人: IGARASHI Ikuo
• 金额: $ 8.83万
• 依托单位: Obihiro University Agriculture and Veterinary Medicine
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B).
• 财政年份: 1997
• 资助国家: 日本
• 起止时间: 1997 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-09460143/
• 关键词: Babesia parasites; protective immunity; cytokine knock-out mice; recombinant antigen; gene engineering; diagnostic antigens; Monoclonal antibody; developmental engineering; マウスバベシア; ウマバベシア; イヌバベシア; ELISA; ノックアウトマウス; サイトカイン; 防御抗原

1. Babesia microti produces a self-limiting infection in mice, and recovered mice are resistant to reinfection. In the present study, the protective mechanisms against B.microti were examined using macrophage scavenger receptor (SR), inducible nitric oxide synthase (iNOS), and gamma interferon (IFN-γ) knock-out mice. Decrease of packed cell volume and increase of spleen weight were less obvious in SR-/- mice than in SR+/+ mice. Parasitemia in iNOS-deficient mice increased more rapidly than those of control mice during the early stage of infection, although they recovered from the infection. IFN-γ knock-out mice could not control primary infection, nor protect against challenge infection.2. B.rodhaini antigen p26 was expressed in Escherichia coli and in insect cells infected with a recombinant baculovirus. BALB/c mice immunized with both recombinant antigens and Freund's adjuvants showed 40- 100% survival rate against challenge infection. MAb 1-5H recognized a 58-kDa protein of B.microti and was found to cross-react with a 60-kDa protein of B.rodhaini. The complete nucleotide sequence encoding p58 protein contained an open reading frame of 1,629-base pair (bp) nucleotides that consists of 542 amino acid residues and the gene was named as p58 gene. A protein homology search showed significant amino acid identities to the η subunit of the chaperonin.3. MAb BEG3 recognized 19kDa antigen of B.equi and inhibited the multiplication of B.equi in vitro. MAb BCllD recognized a 48-kDa protein rhoptry protein of B.caballi merozoite. A cDNA encoding a 50kDa protein of B.gibsoni was cloned and the complete nucleotide sequence was determined. Recombinant proteins of B.equi (MEA-1), B.caballi (BC48) and B.gibsoni(p50) were found to be useful as antigens for serodiagnostic methods such as ELISA or latex agglutination test.

1、田鼠巴贝虫对小鼠产生自限性感染，康复后的小鼠对再次感染有抵抗力。在本研究中，使用巨噬细胞清道夫受体（SR）、诱导型一氧化氮合酶（iNOS）和γ干扰素（IFN-γ）敲除小鼠检查了针对田鼠小芽孢杆菌的保护机制。 SR-/-小鼠的细胞堆积体积的减少和脾脏重量的增加不如SR+/+小鼠明显。在感染早期，iNOS 缺陷小鼠的寄生虫血症比对照小鼠增加得更快，尽管它们从感染中恢复了。 IFN-γ敲除小鼠不能控制原发感染，也不能抵抗攻击性感染。2． B.rodhaini 抗原 p26 在大肠杆菌和感染重组杆状病毒的昆虫细胞中表达。用重组抗原和弗氏佐剂免疫的BALB/c小鼠显示出针对攻击感染的40-100%的存活率。 MAb 1-5H 可识别田鼠芽孢杆菌 (B.microti) 的 58 kDa 蛋白，并被发现与罗德海尼芽孢杆菌 (B.rodhaini) 的 60 kDa 蛋白发生交叉反应。编码p58蛋白的完整核苷酸序列含有1,629个碱基对(bp)核苷酸的开放阅读框，由542个氨基酸残基组成，该基因被命名为p58基因。蛋白质同源性搜索显示与伴侣蛋白的 η 亚基具有显着的氨基酸同一性。3。 MAb BEG3 识别马 B.equi 的 19kDa 抗原并抑制 B.equi 体外增殖。 MAb BCllD 识别 B.caballi 裂殖子的 48 kDa 蛋白棒状体蛋白。克隆了编码 B.gibsoni 50kDa 蛋白的 cDNA，并确定了完整的核苷酸序列。研究发现，B.equi (MEA-1)、B.caballi (BC48) 和 B.gibsoni(p50) 的重组蛋白可用作血清诊断方法（例如 ELISA 或乳胶凝集试验）的抗原。

项目成果

Kamada, T., Igarashi, I., Inoue, N., Nagasawa, H., et al.: "The role of scavenger receptor type A(SR-A) during infection with Babesia microti."J.Protozool.Res.. 7. 81-89 (1997)

Kamada, T.、Igarashi, I.、Inoue, N.、Nagasawa, H. 等人：“A 型清道夫受体 (SR-A) 在田鼠巴贝虫感染过程中的作用。”J.Protozool.Res。

Igarashi, I., Suzuki, R., Waki, S., Tagawa. Y., Seng. S., et al.: "Roles of CD4+ T cells and gamma interferon for protective immunity against Babesia microti infection in mice."Infect.Immun.. 67. 4143-4148 (1999)

五十岚，I.，铃木，R.，胁，S.，田川。

Igarashi,I et al.: "Immunization with recombinant surface antigens p26 with Freund's adjuvants against Babesia rodhaini infection."J.Vet.Med.Sci.. 62. 717-723 (2000)

Igarashi,I 等人：“用重组表面抗原 p26 与弗氏佐剂对抗罗德海尼巴贝虫感染进行免疫。”J.Vet.Med.Sci.. 62. 717-723 (2000)

Avarzed,A.,Igarashi,I.et al.: "Monoclonal antibody against Babesia equi:its characterization and poteitial use of antigen recognized by monoclonal antibodiy for diagnosis." J.Clini.Micorbiology. 36. 1835-1839 (1998)

Avarzed,A.,Igarashi,I.等人：“针对马巴贝虫的单克隆抗体：其特征以及单克隆抗体识别的抗原在诊断中的潜在用途。”

Igarashi,I.et al.: "Babesia bigemina :In vitro and in vivo effects of curdlan sulfate on growth of parasites." Experimental Parasitology. 90. 290-293 (1998)

Igarashi,I.et al.：“二联巴贝斯虫：硫酸凝胶多糖对寄生虫生长的体外和体内影响。”