Jossé Lyne, Bones Alexander J, Purton Tracey, Michaelis Martin, Tsaousis Anastasios D
Laboratory of Molecular & Evolutionary Parasitology, RAPID group, School of Biosciences, University of Kent, Canterbury, United Kingdom.
Current address: Department of Plant Sciences, University of Oxford, Oxford, Oxfordshire, United Kingdom.
Curr Protoc Microbiol. 2019 Jun;53(1):e80. doi: 10.1002/cpmc.80. Epub 2019 Feb 8.
Cryptosporidium is a genus of ubiquitous unicellular parasites belonging to the phylum Apicomplexa. Cryptosporidium species are the second largest cause of childhood diarrhea and are associated with increased morbidity. Accompanying this is the low availability of treatment and lack of vaccines. The major barrier to developing effective treatment is the lack of reliable in vitro culture methods. Recently, our lab has successfully cultivated C. parvum in the esophageal cancer-derived cell line COLO-680N, and has been able to maintain infection for several weeks. The success of this cell line was assessed with a combination of various techniques including fluorescent microscopy and qPCR. In addition, to tackle the issue of long-term oocyst production in vitro, a simple, low-cost bioreactor system using the COLO-680N cell line was established, which produced infectious oocysts for 4 months. This chapter provides details on the methodologies used to culture, maintain, and assess Cryptosporidium infection and propagation in COLO-680N. © 2019 by John Wiley & Sons, Inc.
隐孢子虫是顶复门中一类广泛存在的单细胞寄生虫。隐孢子虫物种是儿童腹泻的第二大病因，并与发病率增加有关。与此同时，治疗方法有限且缺乏疫苗。开发有效治疗方法的主要障碍是缺乏可靠的体外培养方法。最近，我们实验室已成功在源自食管癌的细胞系COLO-680N中培养了微小隐孢子虫，并能够维持感染数周。通过包括荧光显微镜和定量聚合酶链反应在内的多种技术组合评估了该细胞系的成功情况。此外，为了解决体外长期产生卵囊的问题，建立了一种使用COLO-680N细胞系的简单、低成本生物反应器系统，该系统可产生传染性卵囊达4个月。本章详细介绍了用于在COLO-680N中培养、维持和评估隐孢子虫感染及繁殖的方法。© 2019约翰威立国际出版公司
