High-Yield Purification of Giardia intestinalis Cysts from Fecal Samples

Current Protocols in Microbiology Pub Date : 2020-10-09 DOI:10.1002/cpmc.117

Paul Ogbuigwe, Anthony B. Pita, Matthew A. Knox, Niluka Velathanthiri, David T. S. Hayman

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Abstract

Giardia is an enteric protozoan parasite that causes gastroenteritis in all classes of vertebrates. It is ranked among the leading causes of death in children under 5 years of age. Giardiasis affects approximately 280 million people worldwide annually, a situation exacerbated by the low availability of effective treatments and the lack of a vaccine. In addition, the parasite is difficult to manipulate in in vitro environments, which hampers the development of effective disease management strategies. This article highlights the development of a method for the purification of viable Giardia cysts from fecal samples, verified by a trypan blue dye exclusion test. This protocol produces a 10-fold increase in yield over current methods. By combining sucrose flotation with gated filtration, the protocol significantly reduces the amount of debris in the purified cysts suspension. Cyst viability is verified by a trypan blue dye exclusion test. The ability to purify large quantities of Giardia from fecal samples could advance the development of effective treatments to target this worldwide prevalent parasite. © 2020 Wiley Periodicals LLC.

Basic Protocol: Purification of Giardia cysts from fecal samples

Support Protocol: Cyst viability test

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粪样中肠贾第虫囊的高效纯化

贾第鞭毛虫是一种肠道原生动物寄生虫，可引起所有脊椎动物的肠胃炎。它是造成5岁以下儿童死亡的主要原因之一。贾第虫病每年影响全世界约2.8亿人，有效治疗方法缺乏和缺乏疫苗加剧了这一情况。此外，寄生虫难以在体外环境中操作，这阻碍了有效疾病管理策略的发展。本文重点介绍了一种从粪便样品中纯化活贾第鞭毛虫囊的方法，并通过台盼蓝染料排除试验进行了验证。该方案比目前的方法产量增加了10倍。通过将蔗糖浮选与门控过滤相结合，该方案显著减少了纯化囊悬浮液中的碎屑量。通过台盼蓝染料排除试验证实囊肿的生存能力。从粪便样本中纯化大量贾第鞭毛虫的能力可以促进针对这种全球流行寄生虫的有效治疗方法的发展。©2020 Wiley期刊有限公司基本方案:从粪便样本中纯化贾第鞭毛虫囊肿支持方案:囊肿活力测试

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来源期刊

Current Protocols in Microbiology Immunology and Microbiology-Parasitology

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期刊介绍： Current Protocols in Microbiology provides detailed, step-by-step instructions for analyzing bacteria, animal and plant viruses, fungi, protozoans and other microbes. It offers updated coverage of emerging technologies and concepts, such as biofilms, quorum sensing and quantitative PCR, as well as proteomic and genomic methods. It is the first comprehensive source of high-quality microbiology protocols that reflects and incorporates the new mandates and capabilities of this robust and rapidly evolving discipline.