State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Hubei, Wuhan 430070, PR China.
Vet Parasitol. 2010 Jun 24;170(3-4):323-6. doi: 10.1016/j.vetpar.2010.02.009. Epub 2010 Feb 18.
In this study we investigated the prevalence and enzootic potential of Theileria spp. in water buffalo in the Hubei province in China. An indirect ELISA based on a recombinant major piroplasma surface protein was developed. The complete ORF of the 33-kDa major piroplasma surface protein (p33) was obtained from Theileria sergenti genomic DNA by PCR, cloned into the pET-28(a) vector and expressed in E. coli as a His-fusion protein. Then the recombinant p33 (rp33) was purified and used as the antigen to develop an iELISA. Specificity test showed that there was no cross-reaction with Babesia orientalis, Schistosoma japonicum, Anaplasma marginale and Toxoplasma gondii. 178 water buffaloes raised in different locations in Hubei province in China were detected by this iELISA, all samples were also examined by PCR and microscopy at the same time. The iELISA result showed a higher positive rate (27.5%) than PCR (22.5%) and microscopy (12.9%). This result indicated that the iELISA is a suitable method for the diagnosis of T. sergenti infection and could be used in serological surveys to map out the prevalence of the disease.
在本研究中，我们调查了中国湖北省水牛中泰勒虫属的流行情况和地方性流行潜力。建立了一种基于重组主要边缘体表面蛋白的间接 ELISA。通过 PCR 从瑟氏泰勒虫基因组 DNA 中获得了 33-kDa 主要边缘体表面蛋白 (p33) 的完整 ORF，将其克隆到 pET-28(a) 载体中，并在大肠杆菌中表达为 His 融合蛋白。然后纯化重组 p33(rp33) 并用作抗原开发 iELISA。特异性试验表明，与东方巴贝斯虫、日本血吸虫、边缘无浆体和刚地弓形虫无交叉反应。通过该 iELISA 检测了来自中国湖北省不同地点的 178 头水牛，同时所有样本也通过 PCR 和显微镜检查。iELISA 结果显示阳性率（27.5%）高于 PCR（22.5%）和显微镜检查（12.9%）。该结果表明，iELISA 是一种用于诊断瑟氏泰勒虫感染的合适方法，可用于血清学调查以绘制疾病流行情况图。
