Perez J M, Martinez D, Debus A, Sheikboudou C, Bensaid A
Département Elevage et Médecine Vétérinaire, Centre International en Recherche Agronomique pour le Développement, Pointe à Pitre, Guadeloupe, French West Indies.
Clin Diagn Lab Immunol. 1997 Sep;4(5):620-3. doi: 10.1128/cdli.4.5.620-623.1997.
Cowdria ruminantium is a tick-borne rickettsia which causes severe disease in ruminants. All studies with C. ruminantium reported so far were carried out with stocks consisting of infective blood collected from reacting animals or from the same stocks propagated in vitro. Cloned isolates are needed to conduct studies on immune response of the host, on genetic diversity of the parasite, and on mechanisms of attenuation and the development of vaccines. A method of cloning based on the particular chlamydia life cycle of Cowdria was developed. Instead of cloning extracellular elementary bodies, it appeared more convenient to clone endothelial cells infected by one morula resulting from the infection of the cell by one elementary body of Cowdria. Two hundred and sixteen clones were obtained by limiting dilution of infected cells. The method was experimentally validated by comparing randomly amplified polymorphic DNA fingerprints from individual clones obtained from endothelial cell cultures coinfected with two different stocks of C. ruminantium.
反刍动物考德里氏体是一种由蜱传播的立克次氏体，可在反刍动物中引发严重疾病。迄今为止，所有关于反刍动物考德里氏体的研究都是使用从有反应的动物身上采集的感染性血液或在体外繁殖的同一批次样本进行的。需要克隆分离株来开展宿主免疫反应、寄生虫遗传多样性、减毒机制及疫苗研发等方面的研究。基于考德里氏体独特的衣原体生命周期，开发了一种克隆方法。不是克隆细胞外的原体，而是克隆被考德里氏体一个原体感染细胞后形成的一个桑葚体感染的内皮细胞，这样似乎更方便。通过对感染细胞进行有限稀释获得了216个克隆。通过比较从共同感染两种不同反刍动物考德里氏体批次的内皮细胞培养物中获得的单个克隆的随机扩增多态性DNA指纹图谱，对该方法进行了实验验证。
