Ghugare Gaurav S, Nimkande Vijay D, Khairnar Krishna
Environmental Virology Cell, Council of Scientific and Industrial Research-National Environmental Engineering Research Institute (CSIR NEERI), Nehru Marg, Nagpur, Maharashtra, India.
Curr Protoc Cell Biol. 2018 Jun;79(1):e41. doi: 10.1002/cpcb.41. Epub 2018 May 18.
The method described here enables rapid bacteriophage isolation and enrichment of host-specific bacteriophages from an environmental sample. This is achieved by using a simple 0.45-µm Millipore membrane where a specific host is immobilized on the membrane and a sample suspected of containing bacteriophages is exposed to the immobilized cells with the help of a membrane filtration unit. This filtration step facilitates host-specific interaction of bacteriophages with the host and maximization of this interaction using a classic membrane filtration method. Under the effect of vacuum from a vacuum pump, a filter assembly provides a chance for every bacteriophage in the sample to interact with the specific host on the membrane filter. Our technique allows retaining specific bacteriophages on the membrane along with its host cells via adsorption; these adsorbed bacteriophages (along with their hosts) on a filter disc are then enriched in regular nutritive broth, tryptone soya broth (TSB), by incubation. With help of a plaque assay method, host-specific phages of various bacterial species can be isolated, segregated, and enriched. © 2018 by John Wiley & Sons, Inc.
本文所述方法能够从环境样本中快速分离噬菌体并富集宿主特异性噬菌体。这是通过使用一个简单的0.45微米密理博膜来实现的,在该膜上固定有特定宿主,借助膜过滤装置将疑似含有噬菌体的样本暴露于固定化细胞。此过滤步骤有助于噬菌体与宿主进行宿主特异性相互作用,并使用经典膜过滤方法使这种相互作用最大化。在真空泵产生的真空作用下,过滤组件为样本中的每个噬菌体提供了与膜过滤器上的特定宿主相互作用的机会。我们的技术允许通过吸附将特定噬菌体与其宿主细胞一起保留在膜上;然后将这些吸附在滤盘上的噬菌体(及其宿主)在常规营养肉汤——胰蛋白胨大豆肉汤(TSB)中培养进行富集。借助噬菌斑测定法,可以分离、区分并富集各种细菌物种的宿主特异性噬菌体。© 2018约翰威立父子出版公司