应用表达微小隐孢子虫P23的重组干酪乳杆菌建立微小隐孢子虫IFAT诊断方法

畜牧兽医学报 ›› 2010, Vol. 41 ›› Issue (10): 1296-1300.doi:

应用表达微小隐孢子虫P23的重组干酪乳杆菌建立微小隐孢子虫IFAT诊断方法

格日勒图^1，2，石泉¹，王艳霞¹，兰丽³，满达¹，张和平²*

1. 内蒙古农业大学兽医学院，呼和浩特 010018；2. 内蒙古农业大学乳品生物技术与工程教育部重点实验室，呼和浩特 010018；3. 内蒙古农业大学生命科学学院，呼和浩特 010018

收稿日期:1900-01-01 修回日期:1900-01-01 出版日期:2010-10-25 发布日期:2010-10-25
通讯作者: 张和平

Establishment of the Diagnostic Method of IFAT for Cryptosporidium parvum Using the Recombinant Lactobacillus casei Expressing C. parvum P23

Geriletu^1，2， SHI Quan¹, WANG Yan-xia¹, LAN Li³, MAN Da¹, ZHANG He-ping²*

1. College of Veterinary Medicine, Inner Mongolia Agricultural University， Hohhot 010018, China;2. Key Laboratory of Dairy Biotechnology and Engineering of Ministry of Education, Inner Mongolia Agricultural University， Hohhot 010018, China；3. College of Life Sciences, Inner Mongolia Agricultural University, Hohhot 010018，China

Received:1900-01-01 Revised:1900-01-01 Online:2010-10-25 Published:2010-10-25

摘要/Abstract

摘要： 本试验旨在建立IFAT和间接ELISA方法作为微小隐孢子虫病诊断方法。应用DNA重组技术，构建原核表达载体pGEX-P23，经IPTG诱导表达出GST-P23融合蛋白，并进行纯化，将该重组融合蛋白作为包被抗原，建立ELISA诊断方法；同样构建重组载体pMG-P23，用电转化方法将其转入干酪乳杆菌中，称为重组活干酪乳杆菌（含pMG-P23），以此为已知抗原，建立IFAT诊断方法。应用ELISA和IFAT方法检测来自内蒙古地区的508份牛血清、187份绵羊血清和197份山羊血清：其中ELISA方法在各种动物血清中阳性率依次分别为0.59%、5.88%和4.57%，而IFAT方法则为0.19%、5.88%和4.57%。2种检测方法阳性符合率在牛血清样品中为33.33%，在绵羊和山羊中则均为100%。试验结果表明，重组抗原P23具有良好的反应原性，2种诊断方法的结果基本吻合，显示出良好的一致性和敏感性，可在微小隐孢子虫病的诊断实践中推广应用。

Abstract: The present experiment was performed with the objective of establishing the diagnostic methods of IFAT and ELISA for detection of the Cryptosporidium parvum. The prokaryotic expression vector pGEX-P23 was constructed by DNA recombinant technology and expression of the GST-P23 fusion protein by IPTG induction. After purification, the fusion protein was used as coating antigen in the ELISA. Using the same method, the recombinant vector pMGP23 has been constructed and electrotransformed into the Lactobacillus casei, named as recombinant Lactobacillus casei, which was used as a known antigen in IFAT. 508 samples of bovine serum, 187 samples of sheep serum and 197 samples of goat serum collected from the animals in Inner Mongolia were detected by ELISA and IFAT. The positive rates were 0.59%(bovine), 5.88%(sheep) and 4.57%(goat) by ELISA and 0.19%, 5.88% and 4.57% by IFAT. Accordance rate of these two diagnostic methods in bovine serum samples was 33.33% and the rate in sheep and goat was 100%. The results showed that the recombinant P23 has a good reactionogenicity. The positive results of two diagnostic methods were basically identical, which showed good identity and sensitivity. The methods could be widely applied in the diagnosis of Ｃryptosporidiosis.

格日勒图;石泉;王艳霞;兰丽;满达;张和平. 应用表达微小隐孢子虫P23的重组干酪乳杆菌建立微小隐孢子虫IFAT诊断方法[J]. 畜牧兽医学报, 2010, 41(10): 1296-1300.

Geriletu;SHI Quan;WANG Yan-xia;LAN Li;MAN Da;ZHANG He-ping. Establishment of the Diagnostic Method of IFAT for Cryptosporidium parvum Using the Recombinant Lactobacillus casei Expressing C. parvum P23[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2010, 41(10): 1296-1300.

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