我国羊泰勒虫的生物学特性研究
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摘要
羊泰勒虫病(ovine and caprine theileriosis)是由媒介蜱传播的由泰勒科泰勒属的原虫寄生于绵羊和山羊巨噬细胞、淋巴细胞和红细胞内所引起的疾病的总称。泰勒虫与巴贝斯虫、无浆体、可厥体可分别引起家畜的泰勒虫病、巴贝斯虫病、无浆体病及心水病，将这些疾病统称为蜱传病(Tick-borne diseases)，在全世界每年造成约70亿美元的经济损失。羊泰勒虫病危害严重，可引起羔羊和外地引进羊大量死亡，使慢性发病的山羊和绵羊发育变缓，产肉量和产毛量显著下降，从而给养羊业造成重大经济损失。在我国，羊泰勒虫病是一种重要而常见的蜱传性原虫病，主要分布于我国西部地区，包括甘肃、宁夏、内蒙古、青海、四川和陕西等省。然而，目前国内对羊泰勒虫病的研究尚不全面。本项研究以我国分布的两种羊泰勒虫为主要研究对象，对其生物学特性进行进一步的研究，主要研究结果如下：
1．利用传播试验，确定青海血蜱和长角血蜱可以作为我国新定名的吕氏泰勒虫的传播媒介。从感染吕氏泰勒虫的羊体饱血的青海血蜱幼蜱和若蜱发育而来的若蜱和成蜱，能分别传播这种泰勒虫给除脾绵羊；同时，从感染吕氏泰勒虫的羊体饱血的长角血蜱幼蜱和若蜱发育而来的长角血蜱若蜱和成蜱，也能分别传播这种泰勒虫给除脾绵羊。实验结果表明，吕氏泰勒虫至少能被2种硬蜱—青海血蜱和长角血蜱传播，传播方式为阶段性传播。
2．利用传播试验，确定青海血蜱和长角血蜱可以作为我国新定名的尤氏泰勒虫的传播媒介。从感染尤氏泰勒虫的羊体饱血的青海血蜱幼蜱和若蜱发育而来的若蜱和成蜱，能分别传播这种泰勒虫给除脾和未除脾绵羊；同时，从感染尤氏泰勒虫的羊体饱血的长角血蜱若蜱发育而来的成蜱，能将这种泰勒虫传播给除脾和未除脾绵羊；但是，从感染尤氏泰勒虫的羊体饱血的长角血蜱幼蜱发育而来的若蜱，对尤氏泰勒虫的传播没有成功。实验表明，尤氏泰勒虫至少能被2种硬蜱—青海血蜱和长角血蜱传播，传播方式为阶段性传播。
3．用液氮保存的吕氏泰勒虫和尤氏泰勒虫的虫血分别感染绵羊，待虫体出现后，每日做血液涂片、空气干燥、甲醇固定和姬姆萨染色，进行两种虫体的形态学观察。观察内容包括虫体形态种类、各类虫体出现的先后次序、最高染虫率和染虫率高峰时各类虫体所占比例。结果表明，这两种泰勒虫的虫体胞浆呈浅蓝色，细胞核成紫色，形态类型主要包括圆形、卵圆形、针形、杆状、三叶草形、十字架形、逗点形和不规则等多种形态。在整个病程中，以梨籽形、圆形(卵圆形)、针形3类虫体占优势；而且这2个虫种在形态学上几乎没有明显著异。
4．对尤氏泰勒虫人工感染的试验绵羊进行了病理学观察。结果显示：体表少毛部位、眼结膜、皮下脂肪和大网膜轻度黄染；淋巴结和肺脏肿胀，切面湿润多汁；心、肝、肺、肠道利瘤胃表面可见针尖至粟粒大小出血点或出血斑。主要的组织学病变为淋巴结充血、出血及淋巴窦网状内皮细胞增生；肺脏间质、网状细胞增生，肺泡隔增厚；肾小球内皮细胞、血管内皮细胞增生，淋巴网状细胞灶状积累，甚至形成结节；心肌细胞变性坏死，染色不均；肠道肌层、粘膜层和固有膜明显充血、出血；肝细胞颗粒变性和有散在细胞坏死，汇管区和小叶间有淋巴网状细胞增生；瘤胃黏膜下层有散在性出血，肌内层见灶状出血。
5．建立了一种从感染有羊泰勒虫的羊血液细胞分离羊泰勒虫裂殖子的快速方法。用嗜水气单孢菌产生的细胞毒素—溶血素Ah-1，裂解寄生有羊泰勒虫的红细胞，继而用Percoll密度梯度离心分离和纯化羊泰勒虫裂殖子。光学显微镜检验表明，从感染羊泰勒虫的红细胞分离的裂殖子仍然保持正常的虫体形态，并从宿主细胞成分中游离出来。
6．构建了吕氏泰勒虫cDNA表达文库，文库的重组率为100％，初级文库库容量约为1.0×10~6PFU，扩增文库滴度约为8.2×10~8 PFU／mL。本研究用RNAlater RNA Stabilization Reagent保存的吕氏泰勒虫虫体直接提取并纯化mRNA，采用oligo(dT)引物合成双链cDNA，并在其两端加EcoRⅠ／HindⅢ定向接头。将所产生的cDNA分子定向克隆到具有EcoRⅠ／HindⅢ粘性末端的λSCREEN载体的两臂之间。用PhageMaker extract对以上连接产物进行体外包装以形成完整的噬菌体，并用之转染大肠杆菌ER1647，从而构建成吕氏泰勒虫的cDNA表达文库。
7．筛选出9个具有免疫反应性的基因。将吕氏泰勒虫cDNA表达文库与宿主菌BL21(DE3)pLysE按一定比例混合孵育并铺板，当噬斑出现后将NC膜覆盖于顶层胶表面并继续培养3.5 h。随后，将此NC膜依次与吕氏泰勒虫阳性血清和碱性磷酸酶标记的兔抗绵羊IgG反应，最后加底物显色，初步筛选得到蓝紫色的阳性克隆，并复筛直至整个膜上均为阳性信号为止。用所得阳性噬菌体转染宿主菌BM25.8使之自动亚克隆为重组质粒，用此亚克隆质粒转化宿主菌JM109并从中提取重组质粒进行PCR、测序和蛋白表达分析，共获得24个新基因，10个基因能成功地诱导表达，9个具有免疫反应性。
8．建立了吕氏泰勒虫的PCR检测方法。利用从吕氏泰勒虫筛选的T1401基因，根据其序列设计了其特异性引物，用PCR方法检测吕氏泰勒虫感染血样中虫体基因组DNA。从自然感染吕氏泰勒虫的基因组中，用该引物扩增出368bp大小的DNA片段；而用该引物不能从尤氏泰勒虫，羊巴贝斯虫未定种、绵羊无浆体和健康羊基因组DNA中扩增出368bp大小的DNA片段。PCR检测方法能从1％吕氏泰勒虫虫血稀释到10~(-5)倍，仍能检测出虫体DNA。该法比光学显微镜检200个视野敏感。为我国吕氏泰勒虫的流行病学调查提供了一种可靠的检测和诊断方法。
Ovine and caprine theileriosis is caused by genus Theileria, which is transmitted by ticks and parasitized in macrophage, lymphblast and eryhrocytes of a variety of sheep and goats. Theileria, Babesia, Anaplasma and Cowdria were able to respectively cause serious theileriosis, babesiosis, anaplasmosis and heartwater, and all of them were known as tick-borne diseases. Tick-borne diseases caused a loss of 7 billion $ per year. The disease seriously inflences the sheep industry and results in many exotic sheep (or goats) and local lambs to death each year, and caused a huge economic loss of sheep industry. Theileriosis caused by Theileria luwenshuni and/or Theileria uilenbergi is an important tick-borne protozoan disease, and very common in sheep and goats in China. Investigations carried out over many years indicated that it is mainly distributed in Western China, including Gansu, Ningxia, Inner Mongolia, Qiahai, Sicuan and Shaanxi. However, the disease and causative pathogen have not been systematically studied in China. In this study, Theileria spp. infective to sheep and goats is characterized biologically. The major results were presented below:
1. Experimental transmission of a recently identified new Theileria luwenshuni pathogenic for sheep in northern China is described. Haemaphysalis qinghaiensis nymphs and adults developed from larvae and nymph engorged on sheep infected with T. luwenshuni were able to respectively transmit the T. luwenshuni to splenectomized sheep. Meanwhile, H. longicornis nymphs and adults developed from larvae and nymphs engorged on sheep infected with T. luwenshuni were also able to respectively transmit this new T. luwenshuni to splenectomized sheep. This experiment suggested that the T. luwenshuni could be transmitted by at least two species of Haemaphysalis sp. ticks, H. longicornis and H. qinghaiensis, and the mode of transmission is stage to stage.
2. Experimental transmission of a recently identified new Theileria uilenbergi pathogenic for sheep in northern China is described. H. qinghaiensis nymphs and adults developed from larvae and nymph engorged on sheep infected with T. uilenbergi were able to respectively transmit the T. uilenbergi to splenectomized and intact sheep. Meanwhile, H. longicornis adults developed from nymphs engorged on sheep infected with T. uilenbergi were also able to respectively transmit this new T. uilenbergi to splenectomized sheep, but H. longicornis nymphs developed from larvae engorged on sheep infected with T. uilenbergi failed to respectively transmit this Theileria sp. to sheep. This experiment suggested that the T. uilenbergi could be transmitted by at least two species of Haemaphysalis sp. ticks, H. longicornis and H. qinghaiensis, and the mode of transmission is stage to stage.
3. Description of morphology of T. luwenshuni and T. uilenbergi. The splenectomized sheep were respectively injected jugularly with the cryopreserved blood containing T. luwenshuni and T. uilenbergi. When piroplasms appeared in the blood smear from the ears of two sheep, the morphology and dimensions of parasites was examined from thin blood smears, air-dried, fixed with methanol, and stained with 10% Giemsa solution. The dimensions of the intra-erythrocytic stages of T. luwenshuni and T. uilenbergi were determined from a splenectomized sheep 1246 and sheep 2239 with aid of an ocular micrometer thin blood smears. The parasites selected for measurements were of various morphologies, and 200 parasites were measured. The resulted showed that the protoplasma was slightly blue, and the nucleus was purple by Giemsa staining. The morphology of the two designated strain of Theileria spp. appears mostly as single pyriform, needle, rod, round, oval, cross, amoeboid, etc. forms. In the infected sheep, the pyriform and needle forms were found first, then the rod, round, and oval forms, in the end, three-leaf and cross shaped. The pyriform, the round and the needle almost exist in every stage of the disease and seem to have a dominant shape. Meanwhile, there are no differences in morphology and sizes between T. luwenshui and T. uilenbergi.
4. Pathological observation of experimental theileriosis caused by Theileria uilenbergi. Sheep were infected experimentally with T. uilenbergi by subcutaneous injection. The results showed that some tissues, including the less wool part, eye conjunctiva, subcutaneous fat and greater omentum of sheep were slightly yellowish. The heart, liver and lung were generally enlarged, and the cut surfaces of lymph nodes and lung were generally moist and juiceness. Meanwhile, various sizes of haemorrhage were seen in the heart, liver, lung, kidney, especially worst in intestine etc.. The principal microscopic founding was hyperaemia, hemorrhage and proliferation of reticuloendothetial cell in lymph nodes, proliferation of interstitium and reticulocytes in lung, and alveolar septum become wider, proliferation of renal glomerulus intima cells and blood vessel intima cells in ren. Lymph reticulocytes were accumulated into a tubercle in ren. Cardiac muscle cells showed apomorphosis and uneven dyeing. The tunica propria was obvious hyperaemia and hemorrhage in the intestinal tract. Additionally, the hepatocytes showed apomorphosis and necrosis. various sizes of haemorrhage were seen in submucosa and muscle layer of rumen.
5. A simple and rapid method for purifying merozoites of Theileria spp. from infected erythrocytes in sheep or goats. Infected erythrocytes were effectively lysed using hemolysin Ah-1 produced from Aeromonas hydrophila, and the lysate was subjected to centrifugation in a Percoll~(TM) discontinuous density gradient at common speed. Light microscopical study showed that the merozoites purified retained their intact basic structure and were free of erythrocyte membrance and other host-cell components.
6. Establishment of cDNA expression library of T. luwenshuni. Purified mRNA was isolated from the T. luwenshuni parasites, which were cryopreserved in RNAlater RNA Stabilization Reagent at-70℃. A library of oligo(dT)-primed cDNA with added directional EcoRⅠ/HindⅢlinkers was constructed and ligated to the EcoRⅠ/HindⅢarms of the screen vector. The recombinant phage DNA was packaged by using PhageMaker packaging extracts, resulting in a primary cDNA library with a size of 1.0×10 PFU and amplified cDNA library with a phage titer of 8.2×10~8PFU/mL.
7. Immunoscreening of nine new genes. Recombinant phages were plated on a host BL21(DE3)pLysE, and when plaques begin to form, the plates were overlaid with nitrocellulose filters and incubated for an additional 3.5 hours. The filters were processed by successive incubations with anti-Theileria luwenshuni serum, rabbit anti- sheep IgG alkaline phosphatase conjugate, and color development substrates and thus positive plaques with intense purple color were found. Pure phage stocks were obtained by plaque purification and converted to plasmid subclones by plating phage on host strain BM25.8. Recombinant plasmids that were subcloned in BM25.8 were isolated and then transformed into host strain JM109. Twenty-four new genes were obtained after analysis of the plasmids exstracted from JM109 by PCR, sequencing and expression, ten genes of them can be expressed successfully, and nine genes were positive against Theileria luwenshuni sera with Western Blotting.
8. Development of PCR for detection of T. luwenshuni. A PCR method was developed with a pair of specific primers, which were designed according to the partial nucleotide sequence of T1401 gene immunoscreened from cDNA expression library of Theileria luwenshuni. A 368-bp DNA fragment was specifically amplified from genomic DNA purified from blood samples from sheep and goats, naturally infected with T. luwenshuni. No PCR products resulted from T. uilenbergi, unidentified Babesia sp., Anaplasma ovis, or normal sheep leucocytes DNA using these specific primers. The PCR was sensitive enough to detect parasite DNA from the dilution of 10~(-5) with 0.00001% parasitemia. This is more sensitive than examining 200 fields under light microscopy. These results indicate that the PCR provides a useful diagnostic tool for the detection of T. luwenshuni infection and epidemiological survey in small ruminants in China.
1．利用传播试验，确定青海血蜱和长角血蜱可以作为我国新定名的吕氏泰勒虫的传播媒介。从感染吕氏泰勒虫的羊体饱血的青海血蜱幼蜱和若蜱发育而来的若蜱和成蜱，能分别传播这种泰勒虫给除脾绵羊；同时，从感染吕氏泰勒虫的羊体饱血的长角血蜱幼蜱和若蜱发育而来的长角血蜱若蜱和成蜱，也能分别传播这种泰勒虫给除脾绵羊。实验结果表明，吕氏泰勒虫至少能被2种硬蜱—青海血蜱和长角血蜱传播，传播方式为阶段性传播。
2．利用传播试验，确定青海血蜱和长角血蜱可以作为我国新定名的尤氏泰勒虫的传播媒介。从感染尤氏泰勒虫的羊体饱血的青海血蜱幼蜱和若蜱发育而来的若蜱和成蜱，能分别传播这种泰勒虫给除脾和未除脾绵羊；同时，从感染尤氏泰勒虫的羊体饱血的长角血蜱若蜱发育而来的成蜱，能将这种泰勒虫传播给除脾和未除脾绵羊；但是，从感染尤氏泰勒虫的羊体饱血的长角血蜱幼蜱发育而来的若蜱，对尤氏泰勒虫的传播没有成功。实验表明，尤氏泰勒虫至少能被2种硬蜱—青海血蜱和长角血蜱传播，传播方式为阶段性传播。
3．用液氮保存的吕氏泰勒虫和尤氏泰勒虫的虫血分别感染绵羊，待虫体出现后，每日做血液涂片、空气干燥、甲醇固定和姬姆萨染色，进行两种虫体的形态学观察。观察内容包括虫体形态种类、各类虫体出现的先后次序、最高染虫率和染虫率高峰时各类虫体所占比例。结果表明，这两种泰勒虫的虫体胞浆呈浅蓝色，细胞核成紫色，形态类型主要包括圆形、卵圆形、针形、杆状、三叶草形、十字架形、逗点形和不规则等多种形态。在整个病程中，以梨籽形、圆形(卵圆形)、针形3类虫体占优势；而且这2个虫种在形态学上几乎没有明显著异。
4．对尤氏泰勒虫人工感染的试验绵羊进行了病理学观察。结果显示：体表少毛部位、眼结膜、皮下脂肪和大网膜轻度黄染；淋巴结和肺脏肿胀，切面湿润多汁；心、肝、肺、肠道利瘤胃表面可见针尖至粟粒大小出血点或出血斑。主要的组织学病变为淋巴结充血、出血及淋巴窦网状内皮细胞增生；肺脏间质、网状细胞增生，肺泡隔增厚；肾小球内皮细胞、血管内皮细胞增生，淋巴网状细胞灶状积累，甚至形成结节；心肌细胞变性坏死，染色不均；肠道肌层、粘膜层和固有膜明显充血、出血；肝细胞颗粒变性和有散在细胞坏死，汇管区和小叶间有淋巴网状细胞增生；瘤胃黏膜下层有散在性出血，肌内层见灶状出血。
5．建立了一种从感染有羊泰勒虫的羊血液细胞分离羊泰勒虫裂殖子的快速方法。用嗜水气单孢菌产生的细胞毒素—溶血素Ah-1，裂解寄生有羊泰勒虫的红细胞，继而用Percoll密度梯度离心分离和纯化羊泰勒虫裂殖子。光学显微镜检验表明，从感染羊泰勒虫的红细胞分离的裂殖子仍然保持正常的虫体形态，并从宿主细胞成分中游离出来。
6．构建了吕氏泰勒虫cDNA表达文库，文库的重组率为100％，初级文库库容量约为1.0×10~6PFU，扩增文库滴度约为8.2×10~8 PFU／mL。本研究用RNAlater RNA Stabilization Reagent保存的吕氏泰勒虫虫体直接提取并纯化mRNA，采用oligo(dT)引物合成双链cDNA，并在其两端加EcoRⅠ／HindⅢ定向接头。将所产生的cDNA分子定向克隆到具有EcoRⅠ／HindⅢ粘性末端的λSCREEN载体的两臂之间。用PhageMaker extract对以上连接产物进行体外包装以形成完整的噬菌体，并用之转染大肠杆菌ER1647，从而构建成吕氏泰勒虫的cDNA表达文库。
7．筛选出9个具有免疫反应性的基因。将吕氏泰勒虫cDNA表达文库与宿主菌BL21(DE3)pLysE按一定比例混合孵育并铺板，当噬斑出现后将NC膜覆盖于顶层胶表面并继续培养3.5 h。随后，将此NC膜依次与吕氏泰勒虫阳性血清和碱性磷酸酶标记的兔抗绵羊IgG反应，最后加底物显色，初步筛选得到蓝紫色的阳性克隆，并复筛直至整个膜上均为阳性信号为止。用所得阳性噬菌体转染宿主菌BM25.8使之自动亚克隆为重组质粒，用此亚克隆质粒转化宿主菌JM109并从中提取重组质粒进行PCR、测序和蛋白表达分析，共获得24个新基因，10个基因能成功地诱导表达，9个具有免疫反应性。
8．建立了吕氏泰勒虫的PCR检测方法。利用从吕氏泰勒虫筛选的T1401基因，根据其序列设计了其特异性引物，用PCR方法检测吕氏泰勒虫感染血样中虫体基因组DNA。从自然感染吕氏泰勒虫的基因组中，用该引物扩增出368bp大小的DNA片段；而用该引物不能从尤氏泰勒虫，羊巴贝斯虫未定种、绵羊无浆体和健康羊基因组DNA中扩增出368bp大小的DNA片段。PCR检测方法能从1％吕氏泰勒虫虫血稀释到10~(-5)倍，仍能检测出虫体DNA。该法比光学显微镜检200个视野敏感。为我国吕氏泰勒虫的流行病学调查提供了一种可靠的检测和诊断方法。
Ovine and caprine theileriosis is caused by genus Theileria, which is transmitted by ticks and parasitized in macrophage, lymphblast and eryhrocytes of a variety of sheep and goats. Theileria, Babesia, Anaplasma and Cowdria were able to respectively cause serious theileriosis, babesiosis, anaplasmosis and heartwater, and all of them were known as tick-borne diseases. Tick-borne diseases caused a loss of 7 billion $ per year. The disease seriously inflences the sheep industry and results in many exotic sheep (or goats) and local lambs to death each year, and caused a huge economic loss of sheep industry. Theileriosis caused by Theileria luwenshuni and/or Theileria uilenbergi is an important tick-borne protozoan disease, and very common in sheep and goats in China. Investigations carried out over many years indicated that it is mainly distributed in Western China, including Gansu, Ningxia, Inner Mongolia, Qiahai, Sicuan and Shaanxi. However, the disease and causative pathogen have not been systematically studied in China. In this study, Theileria spp. infective to sheep and goats is characterized biologically. The major results were presented below:
1. Experimental transmission of a recently identified new Theileria luwenshuni pathogenic for sheep in northern China is described. Haemaphysalis qinghaiensis nymphs and adults developed from larvae and nymph engorged on sheep infected with T. luwenshuni were able to respectively transmit the T. luwenshuni to splenectomized sheep. Meanwhile, H. longicornis nymphs and adults developed from larvae and nymphs engorged on sheep infected with T. luwenshuni were also able to respectively transmit this new T. luwenshuni to splenectomized sheep. This experiment suggested that the T. luwenshuni could be transmitted by at least two species of Haemaphysalis sp. ticks, H. longicornis and H. qinghaiensis, and the mode of transmission is stage to stage.
2. Experimental transmission of a recently identified new Theileria uilenbergi pathogenic for sheep in northern China is described. H. qinghaiensis nymphs and adults developed from larvae and nymph engorged on sheep infected with T. uilenbergi were able to respectively transmit the T. uilenbergi to splenectomized and intact sheep. Meanwhile, H. longicornis adults developed from nymphs engorged on sheep infected with T. uilenbergi were also able to respectively transmit this new T. uilenbergi to splenectomized sheep, but H. longicornis nymphs developed from larvae engorged on sheep infected with T. uilenbergi failed to respectively transmit this Theileria sp. to sheep. This experiment suggested that the T. uilenbergi could be transmitted by at least two species of Haemaphysalis sp. ticks, H. longicornis and H. qinghaiensis, and the mode of transmission is stage to stage.
3. Description of morphology of T. luwenshuni and T. uilenbergi. The splenectomized sheep were respectively injected jugularly with the cryopreserved blood containing T. luwenshuni and T. uilenbergi. When piroplasms appeared in the blood smear from the ears of two sheep, the morphology and dimensions of parasites was examined from thin blood smears, air-dried, fixed with methanol, and stained with 10% Giemsa solution. The dimensions of the intra-erythrocytic stages of T. luwenshuni and T. uilenbergi were determined from a splenectomized sheep 1246 and sheep 2239 with aid of an ocular micrometer thin blood smears. The parasites selected for measurements were of various morphologies, and 200 parasites were measured. The resulted showed that the protoplasma was slightly blue, and the nucleus was purple by Giemsa staining. The morphology of the two designated strain of Theileria spp. appears mostly as single pyriform, needle, rod, round, oval, cross, amoeboid, etc. forms. In the infected sheep, the pyriform and needle forms were found first, then the rod, round, and oval forms, in the end, three-leaf and cross shaped. The pyriform, the round and the needle almost exist in every stage of the disease and seem to have a dominant shape. Meanwhile, there are no differences in morphology and sizes between T. luwenshui and T. uilenbergi.
4. Pathological observation of experimental theileriosis caused by Theileria uilenbergi. Sheep were infected experimentally with T. uilenbergi by subcutaneous injection. The results showed that some tissues, including the less wool part, eye conjunctiva, subcutaneous fat and greater omentum of sheep were slightly yellowish. The heart, liver and lung were generally enlarged, and the cut surfaces of lymph nodes and lung were generally moist and juiceness. Meanwhile, various sizes of haemorrhage were seen in the heart, liver, lung, kidney, especially worst in intestine etc.. The principal microscopic founding was hyperaemia, hemorrhage and proliferation of reticuloendothetial cell in lymph nodes, proliferation of interstitium and reticulocytes in lung, and alveolar septum become wider, proliferation of renal glomerulus intima cells and blood vessel intima cells in ren. Lymph reticulocytes were accumulated into a tubercle in ren. Cardiac muscle cells showed apomorphosis and uneven dyeing. The tunica propria was obvious hyperaemia and hemorrhage in the intestinal tract. Additionally, the hepatocytes showed apomorphosis and necrosis. various sizes of haemorrhage were seen in submucosa and muscle layer of rumen.
5. A simple and rapid method for purifying merozoites of Theileria spp. from infected erythrocytes in sheep or goats. Infected erythrocytes were effectively lysed using hemolysin Ah-1 produced from Aeromonas hydrophila, and the lysate was subjected to centrifugation in a Percoll~(TM) discontinuous density gradient at common speed. Light microscopical study showed that the merozoites purified retained their intact basic structure and were free of erythrocyte membrance and other host-cell components.
6. Establishment of cDNA expression library of T. luwenshuni. Purified mRNA was isolated from the T. luwenshuni parasites, which were cryopreserved in RNAlater RNA Stabilization Reagent at-70℃. A library of oligo(dT)-primed cDNA with added directional EcoRⅠ/HindⅢlinkers was constructed and ligated to the EcoRⅠ/HindⅢarms of the screen vector. The recombinant phage DNA was packaged by using PhageMaker packaging extracts, resulting in a primary cDNA library with a size of 1.0×10 PFU and amplified cDNA library with a phage titer of 8.2×10~8PFU/mL.
7. Immunoscreening of nine new genes. Recombinant phages were plated on a host BL21(DE3)pLysE, and when plaques begin to form, the plates were overlaid with nitrocellulose filters and incubated for an additional 3.5 hours. The filters were processed by successive incubations with anti-Theileria luwenshuni serum, rabbit anti- sheep IgG alkaline phosphatase conjugate, and color development substrates and thus positive plaques with intense purple color were found. Pure phage stocks were obtained by plaque purification and converted to plasmid subclones by plating phage on host strain BM25.8. Recombinant plasmids that were subcloned in BM25.8 were isolated and then transformed into host strain JM109. Twenty-four new genes were obtained after analysis of the plasmids exstracted from JM109 by PCR, sequencing and expression, ten genes of them can be expressed successfully, and nine genes were positive against Theileria luwenshuni sera with Western Blotting.
8. Development of PCR for detection of T. luwenshuni. A PCR method was developed with a pair of specific primers, which were designed according to the partial nucleotide sequence of T1401 gene immunoscreened from cDNA expression library of Theileria luwenshuni. A 368-bp DNA fragment was specifically amplified from genomic DNA purified from blood samples from sheep and goats, naturally infected with T. luwenshuni. No PCR products resulted from T. uilenbergi, unidentified Babesia sp., Anaplasma ovis, or normal sheep leucocytes DNA using these specific primers. The PCR was sensitive enough to detect parasite DNA from the dilution of 10~(-5) with 0.00001% parasitemia. This is more sensitive than examining 200 fields under light microscopy. These results indicate that the PCR provides a useful diagnostic tool for the detection of T. luwenshuni infection and epidemiological survey in small ruminants in China.
引文
邓国藩，姜在阶。中国经济昆虫志，第三十九册，蜱螨亚纲，硬蜱科．北京：科学出版社，1991，pp43-34。
郭固，徐显增．新疆硬蜱调查．全国寄生虫科研工作第一次会议论文汇编。成都，1964，23-52。
郭淑珍，马登录，牟永娟，杨树猛，王文飙，格桂花，尕旦吉．方百乾，赵春生，甘南地区羊泰勒虫病的预防与治疗试验。中国兽医寄生虫病，2006，14(3)：48-51。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。真核细胞mRNA的提取、纯化和分析。分子克隆试验指南。科学出版社，2002a，516-588。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。cDNA文库制备及基因经鉴定。分子克隆试验指南。科学出版社，2002b，857-944。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。从血清中去除交叉反应抗体。分子克隆试验指南。科学出版社，2002c，1190-1194。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。用IPTG诱导启动子在大肠杆菌种表达克隆化基因。分子克隆试验指南。科学出版社，2002d，1128-1132。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。中性聚丙烯酰胺凝胶电泳。分子克隆试验指南。科学出版社，2002e，418-424。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。cDNA文库制备及基因鉴定(反转录酶)。分子克隆试验指南。科学出版社，2002f, 866-867。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。分子克隆试验指南。科学出版社，2002f, 855。
刘光远，陈怀涛，白启，李有全，才学鹏，试验性羊泰勒虫病的病理学观察。中国兽医科技，2004，34(6)：59-61。
吕文顺，魏宝瑛，黄守云，白启。宁夏牛环形泰勒焦虫的传播试验及形态学观察。见：吕文顺等编：牛泰勒焦虫病的研究。银川：宁夏人民出版社，1980，pp26-33。
吕文顺。泰勒虫科。见：赵辉元主编．家畜寄生虫病与防治学。长春：吉林科学技术出版社，1996，pp 117-125。
李作民，郝有奎，陈廷和，余喜桥，席怀琏，左永让，陈永庆，刘静。隆德县奠安公社羊的泰勒焦虫病报告。甘肃畜牧兽医，1984，(1)：13～14。
李智民，牛玉荣。小尾寒羊泰勒虫病的诊断和治疗。中国兽医科技，1995，25(10)：32～33。
罗建勋，殷宏，张其才，等。羊泰勒虫不同虫株的分离及致病性观察。中国兽医科技，1998，28(12)：34—35。
青海畜牧兽医科学研究所。山羊泰勒焦虫病的诊断。青海畜牧兽医科学研究所资料汇编，1968，5：273-276。
王心娥。甘肃蜱类。兰州大学学报，1980，3：87-92。
王奉先，李欧，李伟，王文祥，华旦，王洛藏，郭仁民，史卫祖，李忠民。湟源、门源、尖扎三县羊泰勒虫病调查。畜牧兽医科技通讯，1980，3：29～32。
王长吉。牛梨形虫病。见：陈耀春．《中国动物疫病志》。北京：科学出版社，1993，381～390。
温进坤，韩梅。SDS-聚丙烯酰胺凝胶电泳。医学分子生物学理论与研究技术。科学出版社，2002a，220-228。
温进坤，韩梅。Western印记。医学分子生物学理论与研究技术。科学出版社，2002b，228-238。
沈倍奋。cDNA基因文库技术。分子文库。科学出版社，2001，1-23。
孙彩琴。青海血蜱体内牛羊梨形虫PCR检测技术的建立和应用[硕士学位论文]。2005，38-41。
殷宏。大巴贝斯虫的蜱传播特性及蛋白分析[硕士论文]。北京：中国农业科学院研究生院，1996，35-40。
殷宏。羊的泰勒虫生物学及分子生物学特性研究[博士论文]。2002d，pp 6-7。
殷宏。羊的泰勒虫生物学及分子生物学特性研究[博士论文]。2002e，112-124。
殷宏。羊的泰勒虫生物学及分子生物学特性研究[博士学位论文]。2002f,93-124。
殷宏。大巴贝斯虫的蜱传播特性及蛋白分析[D]。北京：中国农业科学院研究生院，1996，35～40。
杨辅国，冯泽光，余广海，刘家雄，卫召勤，何锡池．甘孜藏族自治州乾宁农牧站绵羊泰勒焦虫病报告。中国兽医杂志，1958，(2)：33～37。
Ahmed JS, Luo J, Schnittger L, Seitzer U, Jongejan F, Yin H. Phylogenetic position of small-ruminant infecting piroplasms. Ann N Y Acad Sci. 2006 Oct, 1081: 498-504.
Alani A J, Konrad J, Herbert Ⅳ. Serodiagnosis of Babesia rnotasi (Wales), Theileria recondita (Wales) and Cytoecetes phagocytophila infection in sheep. Research in Veterinary Science 1987, 43(1): 104-8.
Alani AJ, Herbert Ⅳ. Morphology and transmission of Theileria recondita (Theileriidae: Sporozoa) isolated from Haemaphysalis punctata from north Wales.Vet Parasitol. 1988a Jun; 28(4):283-91.
Alani AJ, Herbert Ⅳ. Pathogenesis of infection with Theileria recondita (Wales) isolated from Haemaphysalis punctata from north Wales.Vet Parasitol. 1988b Jun; 28(4): 293-301.
Alani AJ. and Herbert Ⅳ. Investigation on the transmission of Babesia motasi (Babesiidae: Sporozoa) by Haemaphysalis punctata (Canestrini and Fanzago 1877). Vet. Parasitol, 1988c. in press.
Altay K, Dumanli N, Holman PJ, Aktas M. Detection of Theileria ovis in naturally infected sheep by nested PCR.Vet Parasitol. 2005 Jan 20; 127(2): 99-104.
Aktas M, Dumanli N, Attay K. Survey of Theileria ovis in sheep and goats in the Elazig region using the polymerase chain reaction. Turkiye Parazitol Derg. 2005; 29(1): 17-21. Turkish.
Aktas M, Altay K, Dumanl N. Development of a polymerase chain reaction method for diagnosis of Babesia ovis infection in sheep and goats. Veterinary Parasitology, 2005, 133 : 277-281.
Aktas M, Dumanli N, Cetinkaya B, Cakmak A. Field evaluation of PCR in detecting Theileria annulata infection in cattle in the east of Turkey. Vet. Rec. 2002, 150, 548-549.
Alhassan A, Pumidonming W, Okamura M, Hirita H, Battsetseg B, Fujisaki F, Yokoyama N, Igarashi I. Development of a single-round and multiplex PCR method for the simultaneous detection of Babesia caballi and Babesia equi in horse blood. Vet. Parasitol. 2005,129, 43-49.
ANDERS RF. MULTIPLE CROSS-REACTIVITIES AMONGST ANTIGENS OF PLASMODIUM- FALCIPARUM IMPAIR THE DEVELOPMENT OF PROTECTIVE IMMUNITY AGAINST MALARIA PARASITE. IMMUNOLOGY. 1986, 8 (6): 529-539.
Bishop R,Gobright E,Nene V,Morzaria S,Musoke A,Sohanpal B.Polymorphic open reading frames encoding secretory proteins are located less than 3 kilobases from Theileria parva telomeres.Mol Biochem Parasitol.2000,110(2) :359-71
Broemer M,Krappmann D,Scheidereit C.Requirement of Hsp90 activity for IkappaB kinase(IKK)biosynthesis and for constitutive and inducible IKK and NF-kappaB activation.Oncogene.2004 Jul 8;23(31) :5378-86．
Barnett SF.Theileriosis.In:Weinmann D,Ristic M.Infectious Blood Diseases of Man and Animals,Vol.2． New York:Academic Press,1968,pp269-328．
Barnett SF.Theileria.In:Kreier J P.Parasitic Protozoa,Vol 4． New York:Academic Press,1977,pp77-113．
Baumann R.Die kleinasiatische schaftheileriose.Berlin und Munchener Tierarzliche Wochenscrift,1939,30:469-474．
Brown CGD.Dynamics and impact of tick borne diseases of cattle.Tropical animal Health and Production,1997,29:1s-3s.
Brown CGD,Ilhan T,Kirvar E,Thomas M,Wilkie G,Leemans I,Hooshmand-Rad P.Theileria lestoquardi and T.annulata in cattle,sheep,and goats-in vitro and in vivo studies.Ann N Y Acad Sci,1998,849:44-51
Bishop R,Sohanpal B,Kariuki DP,Young AS,Nene V,Baylis H,Allsopp BA,Spooner PR,Dolan TT,Morzaria SP.Detection of a carrier state in Theileria parva-infected cattle by the polymerase chain reaction.Parasitology,1992,104:215-232．
Chen HR,Jan SP,Tsao T Y,Sheu YJ,Banroques J,and Cheng SC.Snt309p,a component of the Prp19p-associated complex that interacts with Prp19p and associates with the spliceosome simultaneously with or immediately after dissociation of U4 in the same manner as Prp19p.Mol.Cell.Biol.1998,18:2196-2204．
Chen G,Cao P,Goeddel DV.TNF-induced recruitment and activation of the IKK complex require Cdc37 and Hsp90． Mol Cell.2002 Feb;9(2) :401-10．
Dschunkowsky E.Urodschevich V.Theileriasis in goats,sheep and cattle with a description of Theileria hirci n.sp.from Serbia.Parasitology,1924,16:107-110．
Daniel Nagore,Josune Garci'a-Sanmarti'n,Ana L.Garci'a-Pe'rez,Ramo'n AJ,Ana Hurtado.Identification,genetic diversity and prevalence of Theileria and Babesia species in a sheep population from Northern Spain.International Journal for Parasitology.2004,34,1059-1067．
d'Oliveira C,Van der Weide M,Habela MA,Jacquiet P,Jongejan F.Detection of Theileria annulata in blood samples of carrier cattle by PCR.J.Clin.Microbiol.1995,33,2665-2669．
Enigk K.Das Auftreten der Schafpiroplasmose in Deutschland.Z.Tropenmed.Parasitol,1953,4:175-183．
Enigk,K.Die Schafpiroplasmose in Deutschland.Dtsch.Tierarztl.Wochenschr,1956,63:161-162．
Enigk K,Friedhoff K.and Wirahadiredja S.Zur empfanglichkeit yon haus-und wildwieder kauern flit Theileria ovis(Piroplasmidea).Z.Parasitenk,1964,24:276-287．
Eustace BK,Sakurai T,Stewart JK,YimLamai D,Unger C,Zehetmeier C,Lain B,Torella C,Henning SW,Beste G,Scroggins BT,Neckers L,Ilag LL,Jay DG.Functional proteomic screens reveal an essential extracellular role for hsp90 alpha in cancer cell invasiveness.Nat Cell Biol.2004 Jun,6(6) :507-14．
Escalante AA and Ayala FJ.Evolutionary Origin of Plasmodium and Other Apicomplexa Based on rRNA Genes.Proc.Natl.Acad.Sci.U.S.A.1995,92,5793-5797．
Friedhoff KT.Tick-borne diseases of sheep and goats caused by Babesia,Theileria or Anaplasma spp.Parassitologia,1997,39:99-109．
Frevert U,Reinwald E,Dietrich C,et al.Isolation and immunoelectromicroscopical characterization of Theileria annulata macroschizonts[J].Z Parasitenkd.1986;72(5) :617-30．
Fukumoto S,Xuenan Xuan,Shigeno S,Kimbita E,Igarashi I,Nagasawa H,Fujisaki K,Mikami T.Development of a polymerase chain reaction method for diagnosis Babesia gibsoni infection in dog.J.Vet.Med.Sci.2001,63,977-981．
Guo,SZ,Yuan ZP,Wu GX,et al.Epidemiology of ovine theileriosis in Gannan region,Gansu Province,China [J].Parasitol.Res.2002,88,S36-S37．
Gentra.Genomic DNA Purification Kit,2005,16-17．
Gerard GF,D'alessio JM,and Kotewicz ML.cDNA synthesis by cloned Moloney murine leukemia virus reverse transcriptase lacking ribonuclease H activity.Focus(Life Technology).1989,11:66-69．
Gerard GF,Schmidt BJ,Kotewicz ML,and Campbell JH.cDNA synthesis by cloned Moloney murine leukemia virus reverse transcriptase processing full DNA polymerase activity.Focus(Life Technology),1992,14:91-93．
Gerard GF,D'alessio JM.Reverse transcriptase.(EC2． 7． 7． 49) :The use of Moloney murine leukemia virus reverse transcriptase to synthesize DNA from RNA.Methods Mol.Biol.1993,16:73-94．
Gubbels JM,de Vos AP,van der Weide M,Viseras J,Schouls LM,de Vries E,Jongejan F.Simultaneous detection of bovine Theileria and Babesia species by reverse line blot hybridization.Journal of Clinical Microbiology,1999,37(6) :1782-1789．
Hermann P,Dobbelaere DA.Theileria-induced constitutive IKK activation is independent of functional Hsp90． FEBS Lett.2006 Sep 18;580(21) :5023-8． Epub 2006 Aug 22．
Hooshmand-Rad P,Hawa NJ.Malignant theileriosis of sheep and goats.Tropical Animal.Health Production,1973a,5:97-102．
Hooshmand-Rad P,Hawa NJ.Transmission of Theileria hirci in sheep by Hyalomma anatolicum anatolicum.Tropical Animal Health Production,1973b,5:103-109．
Irvin AD.Characterization of species and strain of Theileria.Advance in Parasitology,1987,26:145-197．
Jansen BC and Neitz WO.Onderstepoort Journal of Veterinary Research.1956,27,3．
Jochen Deckert,Klaus Hartmuth,Daniel Boehringer,Nastaran Behzadnia,Cindy L.Will,Berthold Kastner,Holger Stark,Henning Urlaub,and Reinhard Lu(?)hrmann.Protein Composition and Electron Microscopy Structure of Affinity-Purified Human Spliceosomal B Complexes Isolated under Physiological Conditions.MOLECULAR AND CELLULAR BIOLOGY,July 2006,p.5528-5543 Vol.26,No.14．
Josune García-Sanmartín,Daniel Nagore a,Ana L.García-Pérez,Ramón A.Juste and Ana Hurtado。Molecular diagnosis of Theileria and Babesia species infecting cattle in Northern Spain using reverse line blot macroarrays。BMC Veterinary Research。BMC Veterinary Research 2006,2:16．
Jochen Deckert,Klaus Hartmuth,Daniel Boehringer,Nastaran Behzadnia,Cindy L.Will,Berthold Kastner,Holger Stark,Henning Urlaub,and Reinhard Lu(?)hrmann.Protein Composition and Electron Microscopy Structure of Affinity-Purified Human Spliceosomal B Complexes Isolated under Physiological Conditions.MOLECULAR AND CELLULAR BIOLOGY,July 2006,p.5528-5543 Vol.26,No.14．
Kjemtrup AM,Robinson T,Conrad PA.Description and epidemiology of Theileria youngi n.sp.from a northern Californian dusky-footed wood rat(Neotoma Fuscipes)population.Journal of Parasitology,2001,87(2) :373-378．
Khayyat SM,Gilder AA.Ovine piroplasmosis in Iraq.Transaction of the Royal Society of Tropical Medicine and Hygiene,1947,41:119-126．
Kirvar E,Wilkie G,Katzer F,Brown CG.Theileria lestoquardi--maturation and quantification in Hyalomma anatolicum anatolicum ticks.Parasitology.1998 Sep;117(Pt 3) :255-63．
Kirvar E,Ilhan T,Katzer F,Wilkie G,Hooshmand-Rad,Brown CGD.Detection of Theileria lestoquardi(hirci)in ticks,sheep,goats using polymerase chain reaction.Ann.NY Acad.Sci.1998,49,52-62．
Kirvar E,Ilhan T,Katzer F,Wilkie G,Hooshmand-Rad,P Zweygarth E,estenberg C,Phipps P,Brown CGD.Detection of Theileria annulata in cattle and vector ticks by PCR using the Tams1 gene sequences.Parasitology,2000,120,245-254．
Kotewicz Ml,Sampson CM,D'alessio,and Gerard GF.Isolation of cloned Moloney murine leukemia virus reverse transscriptase lacking ribonuclease H activity.Nucletic Acids Res.1988,16:265-277．
Lestoquard F.Les piroplasmosis du mouton en Algerie.Bulletin de la societe de Pathologie Exotique,1926,4:222-315．
Littlewood W.Annual report for.Ministry of Agriculture,Egypt Veterinary Service.1914．
Lewis D,Purnell RE and Beavan WJ.The piroplasm Theileria ovis detected in sheep in South Wales.Vet.Rec,1981,108:56-57．
Levine ND.Toxonomy of the piroplasmas.Transaction of the American Microscopical Scociety,1971,90:2-33．
Levine ND.Protozoan parasites of domestic animals and of man.Minneapolis,Minnesota:Burgess Publishing Company,1973,pp 342-343
Levine ND,Corliss JO,Cox F EG,Grain J,Honigberg BM,Leedale GF,Loeblich AR,Lorn J,Lynn D,Merinfeld EG, Page FC,Poljansky G,Sprague V,Vavra J,and Wallace FG,A newly revised classification of the Protozoa,Journal of Protozoology 1980,27:37-58．
Levine ND.The protozoan phylum apicomplexa,Vol Ⅱ,Beca Raton:CRC Press,1988,pp35-45．
Luo JX,Yin H.Theileriosis of sheep and goats in China.Tropical Animal Health and Production,1997,29:8s-10s.
Leemans I,Brown D,Hooshmand-Rad P,Kirvar E,Uggla A.Infectivity and cross-immunity studies of Theileria lestoquardi and Theileria annulata in sheep and cattle.I.In vivo responses.Vet Parasitol.1999a,82:179-192．
Leemans I,Brown D,Fossum C,Hooshmand-Rad P,Kirvar E,Wilkie G,Uggla A.Infectivity and cross-immunity studies of Theileria lestoquardi and Theileria annulata in sheep and cattle.Ⅱ.In vitro studies.Vet Parasitol,1999b,82:193-204．
Leemans I.Coparative aspect of Theileria lestoquardi and Theileria annulata infections of sheep and cattle.Ph D thesis,Swedish University of Agricultural Sciences,Uppsala,2001,pp 13-23．
Li Y,Luo J,Liu Z,Guan G,Gao J,Ma M,Dang Z,Liu A,Ren Q,Lu B,Liu J,Zhao H,Li J,Liu G,Bai Q,Yin H..Experimental transmission of Theileria sp.(China 1) infective for small ruminants by Haemaphysalis longicornis and Haemaphysalis qinghaiensis.Parsitol Res.2007 Mar 17;[Epub ahead of print].
Melanie D.Ohi,Craig W.Vander Kooi,Joshua A.Rosenberg,Liping Ren,Justin P.Hirsch,Walter J.Chazin,Thomas Walz.Structural and Functional Analysis of Essential pre-mRNA Splicing Factor Prp19p.MOLECULAR AND CELLULAR BIOLOGY,Jan.2005,p.451-460．
Malcolm J.Gardner,et al.Pathogen That Transforms Lymphocytes Genome Sequence of Theileria parva,a Bovine.Science,2005,309,134-137．
Mazlum Z.Vector of Theileria hirci in south and southern Iran.Annale de Parasitologie Humaine et Comparee,1970,45:523-525．
Morel PC,Uilenberg GThe nomenclature of some Theileria species (Sporozoa,Babesioidea) of domestic ruminants.Revue d'Elevage et de Medecine Veterinaire des Pays Tropicaux,1981,34( 2) :139-143．
Mehlhorn H,Schein E,Ahmed JS.Theileria.In:Kreier JP.Parasitic Protozoa,Vol 7． London:Academic Press,1994,pp217-304．
Mehlhorn H,Schein E.Redescription of Babesia equi Laveran,1901 as Theileria equi Mehlhorn,Schein 1998． Parasitology Research,1998,84(6) :467-75．
Minami T,Kawazu S,Shimizu K,Fujisaki K,Kamio T,Ito S.Detection of the schizont stage in experimentally infected cattle with Japanese Theileria sergenti.Jpn J Vet Sci,1994,52(3) :601-604．
Aktas M,Altay K and Dumahli N.PCR-based detection of Theileria ovis in Rhipicephalus bursa adult ticks.Veterinary Parasitology.Volume 140,Issues 3-4,10 September 2006,Pages 259-263．
Neitz WO.Theileriosis,gonderiosis and cytauzoonosis:a review.Onderstepoort Journal of Veterinary Research,1957,27:257-430．
Neitz WO.The experimental transmission of Theileria ovis by Rhipicephalus evertsi mimeticus and R.bursa.Onderstepoort J Vet Res.1972 Jun;39(2) :83-86．
Nyormoi O,Bwayo JJ,Hirumi H.Theileria parva:isolation of macroschizonts from in vitro propagated lymphoblastoid cells of cattle[J].Exp Parasitol.1981,Dec;52(3) :303-311．
Novagen.OrientExpress cDNA Library Construction Systems.1999． 1-13．
Novagen.Ascreen~(TM) Vector Manual.1999． 1-19．
Ohi MD,andGould KL.Characterization of interactions among the Ceflp-Prp19p-associated splicing complex.RNA,2002． 8:798-815．
Osman AM.Major tick-borne diseases of sheep and goats in the Sudan.Parassitologia,1997,39:143-144．
Oura CAL,Bishop RP,Wampande EM,Lubega GW,Tait A.Application of a reverse line blot assay to the study of haemoparasites in cattle in Uganda.Int.J.Parasitol.2004,34,603-613．
Oliveira-Sequira TCG,Oliveira MCS,Araujo JP,Amarante AFT.PCR-based detection of Babesia bovis and Babesia bigemina in their natural host Boophilus microplus and cattle.Int.J.Parasitol.2005,35,105-111．
Passos LMF,Bell-Sakyi L,Brown CGD.Immunochemical charecterization of in vitro culture-derived antigens of Babesia bovis and Babesia bigemina.Vet.Parasitol.1998,76,239-249．
Pratt WB,Toft DO.Regulation of signaling protein function and trafficking by the hsp90/hsp70-based chaperone machinery.Exp Biol Med (Maywood).2003,228(2) :111-133．
Pain A,Renauld H,Berriman M.Genome of the Host-Cell Transforming Parasite Theileria annulata Compared with T.parva.Science.2005,309,131-133．
QIAGEN.Oligotex Handbook.2002a,6-16,34-45,68-78．
QIAGEN.RNAlater~(TM)Handbook.2002b,4-12．
QIAGEN.Storage,quantification,and Dermination of quality of polyA~(+) mRNA.Oligotex Handbook,2002c,78．
Robinson PM.Theileriosis annulata and its transmission-a review.Tropical Animal Health and Production,1982,14:3-12．
Raestegaiff AF,Timofeeff PS and Matschulsy SN.Zur Frage der endoglobularen parasiten der Schafe und Ziegen und ihre Uebertrager Vorlaufige.Mitt.Berliner Tierarzt.Wochenschr.1936,52:584-585．
Schnitter L,Yin H,Jianxun L,Ludwig W,Shayan P,Rahbari S,Voss-Holtmann A,Ahmed JS.Ribosomal small-subunit RNA gene-sequence analysis of Theileria lestoquardi and a Theileria species highly pathogenic for Theileria buffeli (syn T.orientalis) in cattle.Res Vet Sci,2000,86:352-358．
Schnittger L,Yin H,Qi B,Gubbels MJ,Beyer D,Niemann S,Jongejan F,Ahmed JS.Simultaneous detection and differentiation of Theileria and Babesia parasites infecting small ruminants by reverse line blotting.Parasitol Res.2004 Feb;92(3) :189-96．
Sasmal NK,Biswas SS,Bhattacharyya B,Banerjee CD.Maitra DN.Theileria hirci-studies on transmission from Sahabadi sheep to black Bengal goat.Indian Veterinary Journal,1983,60:599-602．
Sparagano OAE Allsopp MTEP,Mank RA,Rijpkema SG.,Figueroa JV,Jongejan F.Molecular detection of pathogen DNA in ticks (Acari:Ixodidae):a review.Exp.Appl.Acarol.1999,23,929-960．
Schnittger L,Yin H,Qi B,Gubbels MJ,Beyer D,Niemann S,Jongejan F,Ahmed JS.Simultaneous detection and differentiation of Theileria and Babesia parasites infecting small ruminants by reverse line blotting.Parasitol.Res.2004,92,189-196．
Spitalska E,Torina A,Cannella V,Caracappa S,Sparagano OA.Discrimination between Theileria lestoquardi and Theileria annulata in their vectors and hosts by RFLP based on the 18S rRNA gene.Parasitol Res.2004,94(4) :318-20． Epub 2004 Sep15．
Shayan P,Rahbari S.Simultaneous differentiation between Theileria spp.and Babesia spp.on stained blood smear using PCR.Parasitol Res.2005 Oct;97(4) :281-6． Epub 2005 Jul 9．
Shao-yao Ying.Complementary DNA Library.Generation of cDNA Library.2003． Humana Press,Totowa,New Jersey,1-12．
Sugimoto C,Conrad PA,Ito S,et al.Isolation of Theileria parva schizonts from infected lymphoblastoid cells.Acta Trop.1988 Sep;45(3) :203-216．
Sugimoto C,Sato M,Kawazu S,et al.Purification of merozoites of Theileria sergenti from infected bovine erythrocytes[J].Parasitol Res.1991;77(2) :129-231．
Sohanpal B,Wasawo D,Bishop R.Cloning of telomere-associated DNA using single-specific-primer polymerase chain reaction provides evidence for a conserved sequence directly adjacent to Theileria parva telomeric repeats.Gene.2000,19;255(2) :401-9．
Thomson JG.and Hall GN.The occurrence of Babesia motasi (Wenyon,1926) ,in sheep in Northern Nigeria,with a discussion on the classification of the piroplasms.J.Comp.Pathol.Ther.1933,46:218-231．
Tarn WY,Lee KR,and Cheng SC.Yeast precursor mRNA processing protein PRP19 associates with the spliceosome concomitant with or just after dissociation of U4 small nuclear RNA.Proc.Natl.Acad.Sci.USA,1993． 90:10821-10825．
Tarn WY,Hsu CH,Huang KT,Chen H R,Kao HY,Lee KR,andCheng SC.Functional association of essential splicing factor(s) with PRP19 in a protein complex.EMBO J.1994,13:2421-2431．
TaKaRa.MiniBEST Plasmid Purification Kit manual.2006,1-3．
Uilenberg G,Andreasen MP.Haematoxenus separatus sp.n.(sporozoa,Theileriidae) of sheep in Tazania.Revue d'Elevage et de Medecine Veterinaire des Pays Tropicaux.1974,27:459-465．
Uilenberg G,Schreuder BEC.Further studies on Haematoxenus separatus (Sporozoa,Theileriidae) of sheep in Tanzania.Revue d'Elevage et de Medecine Veterinaire des Pays Tropicaux,1976,29:119-126．
Uilenberg G.Theilerial species of domestic livestock.In:A.D.Irvin,M.P.Cunningham and A.S.Young (Editors),Advances in the Control of Theileriosis.Current Topics in Veterinary Medicine and Animal Science,1980,Volume 14． Martinus Nijhoff,the Hague,pp.4-37．
Uilenberg G.Theileria species of domestic livestock.In:Irvin AD,Cunningham MP,Young AS.Advances in the Control of Theileriosis.The Hague:Martinus Nijhoff Publishers,1981,pp 4-37．
Uilenberg G,Perie NM,Sparjer AAM,Franssen FFJ.Theileria orientalis,a cosmopolitan blood parasite of cattle:demonstration of the schizont stage.Research Veterinary Science,1985,38:352-357．
Uilenburg G.International collaborative research:significance of tick-borne hemoparasitic diseases to world animal health.Veterinary Parasitology,1995,57:19-41．
Workman P.Altered states:selectively drugging the Hsp90 cancer chaperone.Trends Mol Med.2004 Feb;10(2) :47-51．
Will CL,and Lu"hrmann R.Spliceosome structure and function.In R.F.Gesteland,T.R.Cech,and A.J.F.Atkins (ed.),The RNA world,3rd ed.2006． Cold Spring Harbor Laboratory Press,Cold Spring Harbor,N.Y..p.369-400．
Watt D,Sparagano O,Brown CGD,Walke AR.Use of the polymerase chain reaction for identification and quantification of Theileria parva protozoa in Rhipicephalus appendiculatus ticks.Parasitol.Res.1997,83,359-363．
Yakimoff Wl.Endoglobulare parasiten der schafe im Nordkaukasus.Achive fur Protisenkunde,1929,65:151-164．
Young AS,Mchinja SJ.Observations on Haematoxenus separatus Uilenberg & Andreasen 1974 in the erythrocytes of Kenyan sheep.Research in Veterinary Science,1977,23:387-388．
Yin H,Jianxun L,Zhang QC,Guiquan G,Schnittger L,Ahmed J,Wensun L.Preliminary biological and molecular characterization of a putative new species of Theileria infective to small ruminants.Proceedings of the Fifth Biennial Conference of the Society for Tropical Veterinary Medicine,programs and abstract,Key West,Florida,12-16 June,Paul Gibbs,Gainesville,Florida,1999,pp77．
Yin H,Guan G,Ma M,Luo J,Lu B,Yuan G,Bai Q,Lu C,Yuan Z,Preston P.Haemaphysalis qinghaiensis ticks transmit at least two different Theileria species:one is infective to yaks,one is infective to sheep.Vet Parasitol,2002a,107:29-35．
Yin H,Luo J,Guan G,Lu B,Ma M,Zhang Q,Lu W,Lu C,Ahmed J.Experiments on transmission of an unidentified Theileria sp.to small ruminants with Haemaphysalis qinghaiensis and Hyalomma anatolicum anatolicum.Vet Parasitol,2002b,108:21-30．
Yin H,Luo JX,Guan GQ,Gao JL,Lu BY,Zhang QC,Ma ML,Lu WS,Lu WX,Yuan ZP,Guo SZ,Wang B,Du HD,Schnittger L,Ahmed J,Jongejian F.Transmission of an unidentified Theileria species to small ruminants with Haemaphysalis qinghaiensis ticks collected from field.Parasitol Res,2002c,88:S25-S27．
Yin H,Luo JX,Schnittger L,Lu BY,Beyer D,Ma ML,Guan GQ,Bai Qi,Lu CP,Ahmed J.Phylogenetic analysis of Theileria species transmitted by Haemaphysalis qinghaiensis.Parasitol Res,2004,92(1) :36-42．
郭固，徐显增．新疆硬蜱调查．全国寄生虫科研工作第一次会议论文汇编。成都，1964，23-52。
郭淑珍，马登录，牟永娟，杨树猛，王文飙，格桂花，尕旦吉．方百乾，赵春生，甘南地区羊泰勒虫病的预防与治疗试验。中国兽医寄生虫病，2006，14(3)：48-51。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。真核细胞mRNA的提取、纯化和分析。分子克隆试验指南。科学出版社，2002a，516-588。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。cDNA文库制备及基因经鉴定。分子克隆试验指南。科学出版社，2002b，857-944。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。从血清中去除交叉反应抗体。分子克隆试验指南。科学出版社，2002c，1190-1194。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。用IPTG诱导启动子在大肠杆菌种表达克隆化基因。分子克隆试验指南。科学出版社，2002d，1128-1132。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。中性聚丙烯酰胺凝胶电泳。分子克隆试验指南。科学出版社，2002e，418-424。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。cDNA文库制备及基因鉴定(反转录酶)。分子克隆试验指南。科学出版社，2002f, 866-867。
J．萨姆布鲁克，D．W．拉塞尔 著(黄培堂 等译)。分子克隆试验指南。科学出版社，2002f, 855。
刘光远，陈怀涛，白启，李有全，才学鹏，试验性羊泰勒虫病的病理学观察。中国兽医科技，2004，34(6)：59-61。
吕文顺，魏宝瑛，黄守云，白启。宁夏牛环形泰勒焦虫的传播试验及形态学观察。见：吕文顺等编：牛泰勒焦虫病的研究。银川：宁夏人民出版社，1980，pp26-33。
吕文顺。泰勒虫科。见：赵辉元主编．家畜寄生虫病与防治学。长春：吉林科学技术出版社，1996，pp 117-125。
李作民，郝有奎，陈廷和，余喜桥，席怀琏，左永让，陈永庆，刘静。隆德县奠安公社羊的泰勒焦虫病报告。甘肃畜牧兽医，1984，(1)：13～14。
李智民，牛玉荣。小尾寒羊泰勒虫病的诊断和治疗。中国兽医科技，1995，25(10)：32～33。
罗建勋，殷宏，张其才，等。羊泰勒虫不同虫株的分离及致病性观察。中国兽医科技，1998，28(12)：34—35。
青海畜牧兽医科学研究所。山羊泰勒焦虫病的诊断。青海畜牧兽医科学研究所资料汇编，1968，5：273-276。
王心娥。甘肃蜱类。兰州大学学报，1980，3：87-92。
王奉先，李欧，李伟，王文祥，华旦，王洛藏，郭仁民，史卫祖，李忠民。湟源、门源、尖扎三县羊泰勒虫病调查。畜牧兽医科技通讯，1980，3：29～32。
王长吉。牛梨形虫病。见：陈耀春．《中国动物疫病志》。北京：科学出版社，1993，381～390。
温进坤，韩梅。SDS-聚丙烯酰胺凝胶电泳。医学分子生物学理论与研究技术。科学出版社，2002a，220-228。
温进坤，韩梅。Western印记。医学分子生物学理论与研究技术。科学出版社，2002b，228-238。
沈倍奋。cDNA基因文库技术。分子文库。科学出版社，2001，1-23。
孙彩琴。青海血蜱体内牛羊梨形虫PCR检测技术的建立和应用[硕士学位论文]。2005，38-41。
殷宏。大巴贝斯虫的蜱传播特性及蛋白分析[硕士论文]。北京：中国农业科学院研究生院，1996，35-40。
殷宏。羊的泰勒虫生物学及分子生物学特性研究[博士论文]。2002d，pp 6-7。
殷宏。羊的泰勒虫生物学及分子生物学特性研究[博士论文]。2002e，112-124。
殷宏。羊的泰勒虫生物学及分子生物学特性研究[博士学位论文]。2002f,93-124。
殷宏。大巴贝斯虫的蜱传播特性及蛋白分析[D]。北京：中国农业科学院研究生院，1996，35～40。
杨辅国，冯泽光，余广海，刘家雄，卫召勤，何锡池．甘孜藏族自治州乾宁农牧站绵羊泰勒焦虫病报告。中国兽医杂志，1958，(2)：33～37。
Ahmed JS, Luo J, Schnittger L, Seitzer U, Jongejan F, Yin H. Phylogenetic position of small-ruminant infecting piroplasms. Ann N Y Acad Sci. 2006 Oct, 1081: 498-504.
Alani A J, Konrad J, Herbert Ⅳ. Serodiagnosis of Babesia rnotasi (Wales), Theileria recondita (Wales) and Cytoecetes phagocytophila infection in sheep. Research in Veterinary Science 1987, 43(1): 104-8.
Alani AJ, Herbert Ⅳ. Morphology and transmission of Theileria recondita (Theileriidae: Sporozoa) isolated from Haemaphysalis punctata from north Wales.Vet Parasitol. 1988a Jun; 28(4):283-91.
Alani AJ, Herbert Ⅳ. Pathogenesis of infection with Theileria recondita (Wales) isolated from Haemaphysalis punctata from north Wales.Vet Parasitol. 1988b Jun; 28(4): 293-301.
Alani AJ. and Herbert Ⅳ. Investigation on the transmission of Babesia motasi (Babesiidae: Sporozoa) by Haemaphysalis punctata (Canestrini and Fanzago 1877). Vet. Parasitol, 1988c. in press.
Altay K, Dumanli N, Holman PJ, Aktas M. Detection of Theileria ovis in naturally infected sheep by nested PCR.Vet Parasitol. 2005 Jan 20; 127(2): 99-104.
Aktas M, Dumanli N, Attay K. Survey of Theileria ovis in sheep and goats in the Elazig region using the polymerase chain reaction. Turkiye Parazitol Derg. 2005; 29(1): 17-21. Turkish.
Aktas M, Altay K, Dumanl N. Development of a polymerase chain reaction method for diagnosis of Babesia ovis infection in sheep and goats. Veterinary Parasitology, 2005, 133 : 277-281.
Aktas M, Dumanli N, Cetinkaya B, Cakmak A. Field evaluation of PCR in detecting Theileria annulata infection in cattle in the east of Turkey. Vet. Rec. 2002, 150, 548-549.
Alhassan A, Pumidonming W, Okamura M, Hirita H, Battsetseg B, Fujisaki F, Yokoyama N, Igarashi I. Development of a single-round and multiplex PCR method for the simultaneous detection of Babesia caballi and Babesia equi in horse blood. Vet. Parasitol. 2005,129, 43-49.
ANDERS RF. MULTIPLE CROSS-REACTIVITIES AMONGST ANTIGENS OF PLASMODIUM- FALCIPARUM IMPAIR THE DEVELOPMENT OF PROTECTIVE IMMUNITY AGAINST MALARIA PARASITE. IMMUNOLOGY. 1986, 8 (6): 529-539.
Bishop R,Gobright E,Nene V,Morzaria S,Musoke A,Sohanpal B.Polymorphic open reading frames encoding secretory proteins are located less than 3 kilobases from Theileria parva telomeres.Mol Biochem Parasitol.2000,110(2) :359-71
Broemer M,Krappmann D,Scheidereit C.Requirement of Hsp90 activity for IkappaB kinase(IKK)biosynthesis and for constitutive and inducible IKK and NF-kappaB activation.Oncogene.2004 Jul 8;23(31) :5378-86．
Barnett SF.Theileriosis.In:Weinmann D,Ristic M.Infectious Blood Diseases of Man and Animals,Vol.2． New York:Academic Press,1968,pp269-328．
Barnett SF.Theileria.In:Kreier J P.Parasitic Protozoa,Vol 4． New York:Academic Press,1977,pp77-113．
Baumann R.Die kleinasiatische schaftheileriose.Berlin und Munchener Tierarzliche Wochenscrift,1939,30:469-474．
Brown CGD.Dynamics and impact of tick borne diseases of cattle.Tropical animal Health and Production,1997,29:1s-3s.
Brown CGD,Ilhan T,Kirvar E,Thomas M,Wilkie G,Leemans I,Hooshmand-Rad P.Theileria lestoquardi and T.annulata in cattle,sheep,and goats-in vitro and in vivo studies.Ann N Y Acad Sci,1998,849:44-51
Bishop R,Sohanpal B,Kariuki DP,Young AS,Nene V,Baylis H,Allsopp BA,Spooner PR,Dolan TT,Morzaria SP.Detection of a carrier state in Theileria parva-infected cattle by the polymerase chain reaction.Parasitology,1992,104:215-232．
Chen HR,Jan SP,Tsao T Y,Sheu YJ,Banroques J,and Cheng SC.Snt309p,a component of the Prp19p-associated complex that interacts with Prp19p and associates with the spliceosome simultaneously with or immediately after dissociation of U4 in the same manner as Prp19p.Mol.Cell.Biol.1998,18:2196-2204．
Chen G,Cao P,Goeddel DV.TNF-induced recruitment and activation of the IKK complex require Cdc37 and Hsp90． Mol Cell.2002 Feb;9(2) :401-10．
Dschunkowsky E.Urodschevich V.Theileriasis in goats,sheep and cattle with a description of Theileria hirci n.sp.from Serbia.Parasitology,1924,16:107-110．
Daniel Nagore,Josune Garci'a-Sanmarti'n,Ana L.Garci'a-Pe'rez,Ramo'n AJ,Ana Hurtado.Identification,genetic diversity and prevalence of Theileria and Babesia species in a sheep population from Northern Spain.International Journal for Parasitology.2004,34,1059-1067．
d'Oliveira C,Van der Weide M,Habela MA,Jacquiet P,Jongejan F.Detection of Theileria annulata in blood samples of carrier cattle by PCR.J.Clin.Microbiol.1995,33,2665-2669．
Enigk K.Das Auftreten der Schafpiroplasmose in Deutschland.Z.Tropenmed.Parasitol,1953,4:175-183．
Enigk,K.Die Schafpiroplasmose in Deutschland.Dtsch.Tierarztl.Wochenschr,1956,63:161-162．
Enigk K,Friedhoff K.and Wirahadiredja S.Zur empfanglichkeit yon haus-und wildwieder kauern flit Theileria ovis(Piroplasmidea).Z.Parasitenk,1964,24:276-287．
Eustace BK,Sakurai T,Stewart JK,YimLamai D,Unger C,Zehetmeier C,Lain B,Torella C,Henning SW,Beste G,Scroggins BT,Neckers L,Ilag LL,Jay DG.Functional proteomic screens reveal an essential extracellular role for hsp90 alpha in cancer cell invasiveness.Nat Cell Biol.2004 Jun,6(6) :507-14．
Escalante AA and Ayala FJ.Evolutionary Origin of Plasmodium and Other Apicomplexa Based on rRNA Genes.Proc.Natl.Acad.Sci.U.S.A.1995,92,5793-5797．
Friedhoff KT.Tick-borne diseases of sheep and goats caused by Babesia,Theileria or Anaplasma spp.Parassitologia,1997,39:99-109．
Frevert U,Reinwald E,Dietrich C,et al.Isolation and immunoelectromicroscopical characterization of Theileria annulata macroschizonts[J].Z Parasitenkd.1986;72(5) :617-30．
Fukumoto S,Xuenan Xuan,Shigeno S,Kimbita E,Igarashi I,Nagasawa H,Fujisaki K,Mikami T.Development of a polymerase chain reaction method for diagnosis Babesia gibsoni infection in dog.J.Vet.Med.Sci.2001,63,977-981．
Guo,SZ,Yuan ZP,Wu GX,et al.Epidemiology of ovine theileriosis in Gannan region,Gansu Province,China [J].Parasitol.Res.2002,88,S36-S37．
Gentra.Genomic DNA Purification Kit,2005,16-17．
Gerard GF,D'alessio JM,and Kotewicz ML.cDNA synthesis by cloned Moloney murine leukemia virus reverse transcriptase lacking ribonuclease H activity.Focus(Life Technology).1989,11:66-69．
Gerard GF,Schmidt BJ,Kotewicz ML,and Campbell JH.cDNA synthesis by cloned Moloney murine leukemia virus reverse transcriptase processing full DNA polymerase activity.Focus(Life Technology),1992,14:91-93．
Gerard GF,D'alessio JM.Reverse transcriptase.(EC2． 7． 7． 49) :The use of Moloney murine leukemia virus reverse transcriptase to synthesize DNA from RNA.Methods Mol.Biol.1993,16:73-94．
Gubbels JM,de Vos AP,van der Weide M,Viseras J,Schouls LM,de Vries E,Jongejan F.Simultaneous detection of bovine Theileria and Babesia species by reverse line blot hybridization.Journal of Clinical Microbiology,1999,37(6) :1782-1789．
Hermann P,Dobbelaere DA.Theileria-induced constitutive IKK activation is independent of functional Hsp90． FEBS Lett.2006 Sep 18;580(21) :5023-8． Epub 2006 Aug 22．
Hooshmand-Rad P,Hawa NJ.Malignant theileriosis of sheep and goats.Tropical Animal.Health Production,1973a,5:97-102．
Hooshmand-Rad P,Hawa NJ.Transmission of Theileria hirci in sheep by Hyalomma anatolicum anatolicum.Tropical Animal Health Production,1973b,5:103-109．
Irvin AD.Characterization of species and strain of Theileria.Advance in Parasitology,1987,26:145-197．
Jansen BC and Neitz WO.Onderstepoort Journal of Veterinary Research.1956,27,3．
Jochen Deckert,Klaus Hartmuth,Daniel Boehringer,Nastaran Behzadnia,Cindy L.Will,Berthold Kastner,Holger Stark,Henning Urlaub,and Reinhard Lu(?)hrmann.Protein Composition and Electron Microscopy Structure of Affinity-Purified Human Spliceosomal B Complexes Isolated under Physiological Conditions.MOLECULAR AND CELLULAR BIOLOGY,July 2006,p.5528-5543 Vol.26,No.14．
Josune García-Sanmartín,Daniel Nagore a,Ana L.García-Pérez,Ramón A.Juste and Ana Hurtado。Molecular diagnosis of Theileria and Babesia species infecting cattle in Northern Spain using reverse line blot macroarrays。BMC Veterinary Research。BMC Veterinary Research 2006,2:16．
Jochen Deckert,Klaus Hartmuth,Daniel Boehringer,Nastaran Behzadnia,Cindy L.Will,Berthold Kastner,Holger Stark,Henning Urlaub,and Reinhard Lu(?)hrmann.Protein Composition and Electron Microscopy Structure of Affinity-Purified Human Spliceosomal B Complexes Isolated under Physiological Conditions.MOLECULAR AND CELLULAR BIOLOGY,July 2006,p.5528-5543 Vol.26,No.14．
Kjemtrup AM,Robinson T,Conrad PA.Description and epidemiology of Theileria youngi n.sp.from a northern Californian dusky-footed wood rat(Neotoma Fuscipes)population.Journal of Parasitology,2001,87(2) :373-378．
Khayyat SM,Gilder AA.Ovine piroplasmosis in Iraq.Transaction of the Royal Society of Tropical Medicine and Hygiene,1947,41:119-126．
Kirvar E,Wilkie G,Katzer F,Brown CG.Theileria lestoquardi--maturation and quantification in Hyalomma anatolicum anatolicum ticks.Parasitology.1998 Sep;117(Pt 3) :255-63．
Kirvar E,Ilhan T,Katzer F,Wilkie G,Hooshmand-Rad,Brown CGD.Detection of Theileria lestoquardi(hirci)in ticks,sheep,goats using polymerase chain reaction.Ann.NY Acad.Sci.1998,49,52-62．
Kirvar E,Ilhan T,Katzer F,Wilkie G,Hooshmand-Rad,P Zweygarth E,estenberg C,Phipps P,Brown CGD.Detection of Theileria annulata in cattle and vector ticks by PCR using the Tams1 gene sequences.Parasitology,2000,120,245-254．
Kotewicz Ml,Sampson CM,D'alessio,and Gerard GF.Isolation of cloned Moloney murine leukemia virus reverse transscriptase lacking ribonuclease H activity.Nucletic Acids Res.1988,16:265-277．
Lestoquard F.Les piroplasmosis du mouton en Algerie.Bulletin de la societe de Pathologie Exotique,1926,4:222-315．
Littlewood W.Annual report for.Ministry of Agriculture,Egypt Veterinary Service.1914．
Lewis D,Purnell RE and Beavan WJ.The piroplasm Theileria ovis detected in sheep in South Wales.Vet.Rec,1981,108:56-57．
Levine ND.Toxonomy of the piroplasmas.Transaction of the American Microscopical Scociety,1971,90:2-33．
Levine ND.Protozoan parasites of domestic animals and of man.Minneapolis,Minnesota:Burgess Publishing Company,1973,pp 342-343
Levine ND,Corliss JO,Cox F EG,Grain J,Honigberg BM,Leedale GF,Loeblich AR,Lorn J,Lynn D,Merinfeld EG, Page FC,Poljansky G,Sprague V,Vavra J,and Wallace FG,A newly revised classification of the Protozoa,Journal of Protozoology 1980,27:37-58．
Levine ND.The protozoan phylum apicomplexa,Vol Ⅱ,Beca Raton:CRC Press,1988,pp35-45．
Luo JX,Yin H.Theileriosis of sheep and goats in China.Tropical Animal Health and Production,1997,29:8s-10s.
Leemans I,Brown D,Hooshmand-Rad P,Kirvar E,Uggla A.Infectivity and cross-immunity studies of Theileria lestoquardi and Theileria annulata in sheep and cattle.I.In vivo responses.Vet Parasitol.1999a,82:179-192．
Leemans I,Brown D,Fossum C,Hooshmand-Rad P,Kirvar E,Wilkie G,Uggla A.Infectivity and cross-immunity studies of Theileria lestoquardi and Theileria annulata in sheep and cattle.Ⅱ.In vitro studies.Vet Parasitol,1999b,82:193-204．
Leemans I.Coparative aspect of Theileria lestoquardi and Theileria annulata infections of sheep and cattle.Ph D thesis,Swedish University of Agricultural Sciences,Uppsala,2001,pp 13-23．
Li Y,Luo J,Liu Z,Guan G,Gao J,Ma M,Dang Z,Liu A,Ren Q,Lu B,Liu J,Zhao H,Li J,Liu G,Bai Q,Yin H..Experimental transmission of Theileria sp.(China 1) infective for small ruminants by Haemaphysalis longicornis and Haemaphysalis qinghaiensis.Parsitol Res.2007 Mar 17;[Epub ahead of print].
Melanie D.Ohi,Craig W.Vander Kooi,Joshua A.Rosenberg,Liping Ren,Justin P.Hirsch,Walter J.Chazin,Thomas Walz.Structural and Functional Analysis of Essential pre-mRNA Splicing Factor Prp19p.MOLECULAR AND CELLULAR BIOLOGY,Jan.2005,p.451-460．
Malcolm J.Gardner,et al.Pathogen That Transforms Lymphocytes Genome Sequence of Theileria parva,a Bovine.Science,2005,309,134-137．
Mazlum Z.Vector of Theileria hirci in south and southern Iran.Annale de Parasitologie Humaine et Comparee,1970,45:523-525．
Morel PC,Uilenberg GThe nomenclature of some Theileria species (Sporozoa,Babesioidea) of domestic ruminants.Revue d'Elevage et de Medecine Veterinaire des Pays Tropicaux,1981,34( 2) :139-143．
Mehlhorn H,Schein E,Ahmed JS.Theileria.In:Kreier JP.Parasitic Protozoa,Vol 7． London:Academic Press,1994,pp217-304．
Mehlhorn H,Schein E.Redescription of Babesia equi Laveran,1901 as Theileria equi Mehlhorn,Schein 1998． Parasitology Research,1998,84(6) :467-75．
Minami T,Kawazu S,Shimizu K,Fujisaki K,Kamio T,Ito S.Detection of the schizont stage in experimentally infected cattle with Japanese Theileria sergenti.Jpn J Vet Sci,1994,52(3) :601-604．
Aktas M,Altay K and Dumahli N.PCR-based detection of Theileria ovis in Rhipicephalus bursa adult ticks.Veterinary Parasitology.Volume 140,Issues 3-4,10 September 2006,Pages 259-263．
Neitz WO.Theileriosis,gonderiosis and cytauzoonosis:a review.Onderstepoort Journal of Veterinary Research,1957,27:257-430．
Neitz WO.The experimental transmission of Theileria ovis by Rhipicephalus evertsi mimeticus and R.bursa.Onderstepoort J Vet Res.1972 Jun;39(2) :83-86．
Nyormoi O,Bwayo JJ,Hirumi H.Theileria parva:isolation of macroschizonts from in vitro propagated lymphoblastoid cells of cattle[J].Exp Parasitol.1981,Dec;52(3) :303-311．
Novagen.OrientExpress cDNA Library Construction Systems.1999． 1-13．
Novagen.Ascreen~(TM) Vector Manual.1999． 1-19．
Ohi MD,andGould KL.Characterization of interactions among the Ceflp-Prp19p-associated splicing complex.RNA,2002． 8:798-815．
Osman AM.Major tick-borne diseases of sheep and goats in the Sudan.Parassitologia,1997,39:143-144．
Oura CAL,Bishop RP,Wampande EM,Lubega GW,Tait A.Application of a reverse line blot assay to the study of haemoparasites in cattle in Uganda.Int.J.Parasitol.2004,34,603-613．
Oliveira-Sequira TCG,Oliveira MCS,Araujo JP,Amarante AFT.PCR-based detection of Babesia bovis and Babesia bigemina in their natural host Boophilus microplus and cattle.Int.J.Parasitol.2005,35,105-111．
Passos LMF,Bell-Sakyi L,Brown CGD.Immunochemical charecterization of in vitro culture-derived antigens of Babesia bovis and Babesia bigemina.Vet.Parasitol.1998,76,239-249．
Pratt WB,Toft DO.Regulation of signaling protein function and trafficking by the hsp90/hsp70-based chaperone machinery.Exp Biol Med (Maywood).2003,228(2) :111-133．
Pain A,Renauld H,Berriman M.Genome of the Host-Cell Transforming Parasite Theileria annulata Compared with T.parva.Science.2005,309,131-133．
QIAGEN.Oligotex Handbook.2002a,6-16,34-45,68-78．
QIAGEN.RNAlater~(TM)Handbook.2002b,4-12．
QIAGEN.Storage,quantification,and Dermination of quality of polyA~(+) mRNA.Oligotex Handbook,2002c,78．
Robinson PM.Theileriosis annulata and its transmission-a review.Tropical Animal Health and Production,1982,14:3-12．
Raestegaiff AF,Timofeeff PS and Matschulsy SN.Zur Frage der endoglobularen parasiten der Schafe und Ziegen und ihre Uebertrager Vorlaufige.Mitt.Berliner Tierarzt.Wochenschr.1936,52:584-585．
Schnitter L,Yin H,Jianxun L,Ludwig W,Shayan P,Rahbari S,Voss-Holtmann A,Ahmed JS.Ribosomal small-subunit RNA gene-sequence analysis of Theileria lestoquardi and a Theileria species highly pathogenic for Theileria buffeli (syn T.orientalis) in cattle.Res Vet Sci,2000,86:352-358．
Schnittger L,Yin H,Qi B,Gubbels MJ,Beyer D,Niemann S,Jongejan F,Ahmed JS.Simultaneous detection and differentiation of Theileria and Babesia parasites infecting small ruminants by reverse line blotting.Parasitol Res.2004 Feb;92(3) :189-96．
Sasmal NK,Biswas SS,Bhattacharyya B,Banerjee CD.Maitra DN.Theileria hirci-studies on transmission from Sahabadi sheep to black Bengal goat.Indian Veterinary Journal,1983,60:599-602．
Sparagano OAE Allsopp MTEP,Mank RA,Rijpkema SG.,Figueroa JV,Jongejan F.Molecular detection of pathogen DNA in ticks (Acari:Ixodidae):a review.Exp.Appl.Acarol.1999,23,929-960．
Schnittger L,Yin H,Qi B,Gubbels MJ,Beyer D,Niemann S,Jongejan F,Ahmed JS.Simultaneous detection and differentiation of Theileria and Babesia parasites infecting small ruminants by reverse line blotting.Parasitol.Res.2004,92,189-196．
Spitalska E,Torina A,Cannella V,Caracappa S,Sparagano OA.Discrimination between Theileria lestoquardi and Theileria annulata in their vectors and hosts by RFLP based on the 18S rRNA gene.Parasitol Res.2004,94(4) :318-20． Epub 2004 Sep15．
Shayan P,Rahbari S.Simultaneous differentiation between Theileria spp.and Babesia spp.on stained blood smear using PCR.Parasitol Res.2005 Oct;97(4) :281-6． Epub 2005 Jul 9．
Shao-yao Ying.Complementary DNA Library.Generation of cDNA Library.2003． Humana Press,Totowa,New Jersey,1-12．
Sugimoto C,Conrad PA,Ito S,et al.Isolation of Theileria parva schizonts from infected lymphoblastoid cells.Acta Trop.1988 Sep;45(3) :203-216．
Sugimoto C,Sato M,Kawazu S,et al.Purification of merozoites of Theileria sergenti from infected bovine erythrocytes[J].Parasitol Res.1991;77(2) :129-231．
Sohanpal B,Wasawo D,Bishop R.Cloning of telomere-associated DNA using single-specific-primer polymerase chain reaction provides evidence for a conserved sequence directly adjacent to Theileria parva telomeric repeats.Gene.2000,19;255(2) :401-9．
Thomson JG.and Hall GN.The occurrence of Babesia motasi (Wenyon,1926) ,in sheep in Northern Nigeria,with a discussion on the classification of the piroplasms.J.Comp.Pathol.Ther.1933,46:218-231．
Tarn WY,Lee KR,and Cheng SC.Yeast precursor mRNA processing protein PRP19 associates with the spliceosome concomitant with or just after dissociation of U4 small nuclear RNA.Proc.Natl.Acad.Sci.USA,1993． 90:10821-10825．
Tarn WY,Hsu CH,Huang KT,Chen H R,Kao HY,Lee KR,andCheng SC.Functional association of essential splicing factor(s) with PRP19 in a protein complex.EMBO J.1994,13:2421-2431．
TaKaRa.MiniBEST Plasmid Purification Kit manual.2006,1-3．
Uilenberg G,Andreasen MP.Haematoxenus separatus sp.n.(sporozoa,Theileriidae) of sheep in Tazania.Revue d'Elevage et de Medecine Veterinaire des Pays Tropicaux.1974,27:459-465．
Uilenberg G,Schreuder BEC.Further studies on Haematoxenus separatus (Sporozoa,Theileriidae) of sheep in Tanzania.Revue d'Elevage et de Medecine Veterinaire des Pays Tropicaux,1976,29:119-126．
Uilenberg G.Theilerial species of domestic livestock.In:A.D.Irvin,M.P.Cunningham and A.S.Young (Editors),Advances in the Control of Theileriosis.Current Topics in Veterinary Medicine and Animal Science,1980,Volume 14． Martinus Nijhoff,the Hague,pp.4-37．
Uilenberg G.Theileria species of domestic livestock.In:Irvin AD,Cunningham MP,Young AS.Advances in the Control of Theileriosis.The Hague:Martinus Nijhoff Publishers,1981,pp 4-37．
Uilenberg G,Perie NM,Sparjer AAM,Franssen FFJ.Theileria orientalis,a cosmopolitan blood parasite of cattle:demonstration of the schizont stage.Research Veterinary Science,1985,38:352-357．
Uilenburg G.International collaborative research:significance of tick-borne hemoparasitic diseases to world animal health.Veterinary Parasitology,1995,57:19-41．
Workman P.Altered states:selectively drugging the Hsp90 cancer chaperone.Trends Mol Med.2004 Feb;10(2) :47-51．
Will CL,and Lu"hrmann R.Spliceosome structure and function.In R.F.Gesteland,T.R.Cech,and A.J.F.Atkins (ed.),The RNA world,3rd ed.2006． Cold Spring Harbor Laboratory Press,Cold Spring Harbor,N.Y..p.369-400．
Watt D,Sparagano O,Brown CGD,Walke AR.Use of the polymerase chain reaction for identification and quantification of Theileria parva protozoa in Rhipicephalus appendiculatus ticks.Parasitol.Res.1997,83,359-363．
Yakimoff Wl.Endoglobulare parasiten der schafe im Nordkaukasus.Achive fur Protisenkunde,1929,65:151-164．
Young AS,Mchinja SJ.Observations on Haematoxenus separatus Uilenberg & Andreasen 1974 in the erythrocytes of Kenyan sheep.Research in Veterinary Science,1977,23:387-388．
Yin H,Jianxun L,Zhang QC,Guiquan G,Schnittger L,Ahmed J,Wensun L.Preliminary biological and molecular characterization of a putative new species of Theileria infective to small ruminants.Proceedings of the Fifth Biennial Conference of the Society for Tropical Veterinary Medicine,programs and abstract,Key West,Florida,12-16 June,Paul Gibbs,Gainesville,Florida,1999,pp77．
Yin H,Guan G,Ma M,Luo J,Lu B,Yuan G,Bai Q,Lu C,Yuan Z,Preston P.Haemaphysalis qinghaiensis ticks transmit at least two different Theileria species:one is infective to yaks,one is infective to sheep.Vet Parasitol,2002a,107:29-35．
Yin H,Luo J,Guan G,Lu B,Ma M,Zhang Q,Lu W,Lu C,Ahmed J.Experiments on transmission of an unidentified Theileria sp.to small ruminants with Haemaphysalis qinghaiensis and Hyalomma anatolicum anatolicum.Vet Parasitol,2002b,108:21-30．
Yin H,Luo JX,Guan GQ,Gao JL,Lu BY,Zhang QC,Ma ML,Lu WS,Lu WX,Yuan ZP,Guo SZ,Wang B,Du HD,Schnittger L,Ahmed J,Jongejian F.Transmission of an unidentified Theileria species to small ruminants with Haemaphysalis qinghaiensis ticks collected from field.Parasitol Res,2002c,88:S25-S27．
Yin H,Luo JX,Schnittger L,Lu BY,Beyer D,Ma ML,Guan GQ,Bai Qi,Lu CP,Ahmed J.Phylogenetic analysis of Theileria species transmitted by Haemaphysalis qinghaiensis.Parasitol Res,2004,92(1) :36-42．