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。Babesiamicroti在小鼠中产生自限感染，并且恢复的小鼠对重新感染具有抗性。在本研究中，使用巨噬细胞清道夫受体（SR），可诱导的一氧化氮合酶（INOS）和伽马干扰素（IFN-γ）敲除小鼠检查了针对芽孢杆菌的保护机制。在SR - / - 小鼠中，堆积的细胞体积的减小和每个+小鼠的体重的增加和增加的小鼠。 iNOS缺陷小鼠的寄生虫血症比在感染的早期阶段比对照小鼠的寄生虫增加了，尽管它们从感染中恢复了。 IFN-γ敲除小鼠无法控制原发性感染，也不能防止挑战感染2。在大肠杆菌和感染了重组杆状病毒的昆虫细胞中表达了rodhaini抗原p26。 BALB/C小鼠用重组抗原和Freund的调节剂免疫，显示了40-100％的生存率，以抗挑战感染。 MAB 1-5H识别出58 kDa蛋白的b.microti蛋白，并被发现与Rodhaini的60 kDa蛋白交叉反应。编码p58蛋白的完整核苷酸序列包含1,629碱基对（BP）核苷酸的开放式阅读框，该核苷酸由542个氨基酸保留，该基因被称为p58基因。蛋白质同源性搜索显示链蛋白的η亚基的明显氨基酸身份。3。 MAB BEG3识别B.Equi的19kDa抗原，并抑制了B.Equi在体外的繁殖。 mAb bclld识别出b.caballi merozoite的48 kDa蛋白Rhoptry蛋白。克隆了编码50kDa蛋白的cDNA，并确定完整的核苷酸序列。发现B.Equi（MEA-1），B.Caballi（BC48）和B.Gibsoni（p50）的重组蛋白很有用，可作为抗原用于血清诊断方法的抗原，例如ELISA或乳胶凝集测试。

项目成果

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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)

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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)

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