Development of international standard diagnostic method for equine babesiosis

马巴贝斯虫病国际标准诊断方法的制定

• 批准号: 13356007
• 负责人: IGARASHI Ikuo
• 金额: $ 28.29万
• 依托单位: Obihiro University of Agriculture and Veterinary Medicine
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (A)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13356007/
• 关键词: Equine babesiosis; Babesia equi; Babesia caballi; Serological diagnosis; Recombinant antigen; ELISA; Immunochromatographic test; International standard method; 血清診断; 金コロイド粒子; ディップスティック法; 国際標準診断法; ウマバベシア; ラテックス凝集反応; 国際標準

Equine babesiosis, caused by Babesia equi and Babesia caballi, is characterized by fever, anemia, icterus, hepatosplenomegaly, lethargy, and, in some cases, leading to great economic losses in the horse industry. The infected horses often remain carriers of the parasites for a long period and are known to act as sources for subsequent infections to other horses via tick vectors. Therefore, the development of a high-quality system has been required for the serological diagnosis of babesial infection. In the present study, development of international standard diagnostic methods with high specificity and sensitivity for equine babesiosis was examined by gene cloning of Babesia DNA with immune sera, production of recombinant antigens, evaluation of ELISA and imunochromatographic test using those antigens. Novel genes, Be82, Be158 and Bc134, were cloned from B.equi and B.caballi, respectively. ELISA using these recombinant antigens showed the high specificity and sensitivity for the detect … More ion of antibodies against both babesial infections. Productions of recombinant antigens in Escherichia coli, EMA-2 and BC48, were also improved by the construction of truncated genes or modification of culture or induction factors. ELISA using these truncated EMA-2 and BC48 showed high specificity and sensitivity for the detection of antibodies. An immunochromotographic test for simultaneous detection of both B.caballi and B.equi specific antibodies was developed. Evaluation in the detection of sera from uninfected horses and experimentally infected horses showed high sensitivity and specificity as shown in ELISA. A differential single-round and multiplex polymerase chain reaction (PCR) method was also developed for the simultaneous detection of B.caballi and B.equi, by targeting 18S ribosomal RNA genes. In conclusion, EMA-2 for B.equi and BC48 for B.caballi were most excellent as antigens for the diagnosis. ELISA and immunochromatographic test using both antigens have shown to have high specificity and sensitivity for the detection of antibodies. The worldwide experimental comparison and evaluation of these serological tests and genetic analysis will would be feasible tests for internationally recognized diagnosis for equine babesiosis in the world. Less

由Babesia Equi和Babesia caballi引起的马巴巴病的特征是发烧，贫血，红冰，肝肾上腺肿，嗜血杆菌，嗜睡，在某些情况下，导致马行业的经济损失巨大。被感染的马很长一段时间都是寄生虫的携带者，被称为随后通过tick矢量向其他马的感染来源。因此，需要开发高质量的系统的血清学诊断Babesial感染。在本研究中，通过使用免疫剂的贝贝西亚DNA的基因克隆，重组抗原的产生，ELISA的评估和不抗色谱术测试来检查具有高特异性和对马的敏感性的国际标准诊断方法的开发。新型基因BE82，BE158和BC134分别从B.Equi和b.cavalli克隆。使用这些重组抗原的ELISA显示出对检测的高特异性和敏感性……更多的抗体与两种贝氏感染的离子。通过截短基因的构造或培养因子的修饰或诱导因子的修饰，也改善了大肠杆菌，EMA-2和BC48中重组抗原的生产。使用这些截短的EMA-2和BC48的ELISA对检测抗体的检测表现出很高的特异性和敏感性。开发了用于简单检测b.cavalli和B. equi特异性抗体的免疫色谱测试。如ELISA所示，在未感染马和实验感染马的血清检测中的评估表现出高灵敏度和特异性。还通过靶向18S核糖体RNA基因，开发了一种差异的单一单轮和多聚合酶链反应（PCR）方法，以简单地检测b.cavalli和b.equi。总之，B.Equi和BC48的EMA-2对于诊断剂的抗原是最出色的。使用两种抗原的ELISA和免疫色谱测试都证明对检测抗体具有很高的特异性和敏感性。这些血清学测试和遗传分析的全球实验比较和评估将是对世界上马巴比斯病的国际公认诊断的可行测试。较少的

项目成果

ウマバベシア感染用診断キット

乌马巴贝虫感染诊断试剂盒

• 发表时间: 2005

Bork S, Yokoyama N, Ikehara Y, Kumar S, Sugimoto C. Igarashi I.: "Growth-inhibitory effect of heparin on Babesia parasites"Antimicrob Agents Chemother.. 48(1). 236-241 (2004)

Bork S、Yokoyama N、Ikehara Y、Kumar S、Sugimoto C. Igarashi I.：“肝素对巴贝虫寄生虫的生长抑制作用”Antimicrob Agents Chemother.. 48(1)。

Serodiagnosis of Babesia gibsoni infection in dogs by an improved enzyme-linked immunosorbent assay with recombinant truncated P50.

通过使用重组截短 P50 的改进酶联免疫吸附测定法对犬吉布氏巴贝虫感染进行血清诊断。

• 发表时间: 2004
• 期刊: J.Vet.Med.Sci. 66
• 作者: Verdida;R.A.;Hara;O.A.;Xuan;X.;Fukumoto;S.;Igarashi;I.;Zhang;S.;Dong;J.;Inokuma;H.;Kabeya;H.;Sato;Y.;Moritomom T.;Maruyama;S.;Claveria F;Nagasawa;H.
• 通讯作者: H.

Cloning of a truncated Babesia equi gene encoding an 82-kilodalton protein and its potential use in an enzyme-linked immunosorbent assay.

编码 82 千道尔顿蛋白质的截短马巴贝虫基因的克隆及其在酶联免疫吸附测定中的潜在用途。

• 发表时间: 2002
• 期刊: J Clin Microbiol. 40
• 作者: Hirata H;Ikadai H;Yokoyama N;Xuan X;Fujisaki K;Suzuki N;Mikami T;Igarashi I.
• 通讯作者: Igarashi I.

Xuan, X., Larsen, A., Ikadai, H., Tanaka, T., Igarashi, I. Et al.: "Expression of Babesia equi merozoite antigen-1 in insect cells by recombinant baculovirus and evaluation of its diagnostic potential in an enzyme-linked immunosorbent assay"J. Clin. Micor

Xu, X., Larsen, A., Ikadai, H., Tanaka, T., Igarashi, I. 等人：“通过重组杆状病毒在昆虫细胞中表达马贝斯虫裂殖子抗原 1 并评估其诊断潜力