Physical fractionation of aquatic viral assemblages
Physical fractionation of aquatic viral assemblages
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DOI：
10.4319/lom.2011.9.150
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发表时间：
2011-04-01
影响因子：
2.7
通讯作者：
Steward, Grieg F.
中科院分区：
文献类型：
--
作者：
Brum, Jennifer R.;Steward, Grieg F.
水生病毒非常多样，对浮游生物生态和进化有重大影响，但这些病毒中的大多数仍未确定其特征。大多数病毒尚未培养，探索病毒多样性的独立于培养的手段只提供了对病毒基因组信息的零散看法。在本研究中，根据水生病毒不同的物理性质，对超速离心法和层析方法分离水生病毒的能力进行了评估，目的是为进一步研究水生病毒的亚群提供物理上的丰富。根据水生病毒的浮力密度、表面电荷和大小不同，连续氯化铯梯度离心法、强阴离子交换层析和弱阴离子交换层析以及Sephacryl S-1000尺寸排除层析分别可分离水生病毒。蔗糖密度梯度、阳离子交换层析和使用可控孔玻璃的尺寸排除层析被发现不足以有效地分离病毒组合。使用上面列出的成功方法，我们能够将复杂的病毒组合分离成通过脉冲场凝胶电泳法确定的具有原始群落中存在的病毒的可区分子集的组分。强阴离子交换层析在分离样品中的病毒方面特别有效，并在大多数组分中产生一至六条主要的病毒基因组条带。我们的结果表明，单独的病毒种群可能在没有培养的情况下从复杂的组合中获得物理上的丰富，并可能被分离出来。这些分离方法有望提高我们鉴定水环境中大多数未培养病毒的基因类型和表型的能力。
Aquatic viruses are extraordinarily diverse and have major influences on plankton ecology and evolution, but the majority of these viruses remain uncharacterized. Most viruses have not been cultivated and cultivation-independent means to explore viral diversity have provided only a fragmented view of viral genomic information. In this study, ultracentrifugation and chromatographic methods were evaluated for their ability to fractionate aquatic viruses based on their differing physical properties with the aim of physically enriching subsets of aquatic viruses for further study. Centrifugation in continuous cesium chloride gradients, strong and weak anion-exchange chromatography, and size-exclusion chromatography with Sephacryl S-1000 were found to separate aquatic viruses based on their differing buoyant densities, surface charges, and sizes, respectively. Sucrose density gradients, cation-exchange chromatography, and size-exclusion chromatography using controlled-pore glass were found to have insufficient resolution to effectively separate viral assemblages. Using the successful methods listed above, we were able to separate complex assemblages of viruses into fractions that had distinguishable subsets of the viruses present in the original community as determined by pulsed-field gel electrophoresis. Strong anion-exchange chromatography was particularly effective at separating viruses in the samples and resulted in one to six dominant viral genome bands in most fractions. Our results suggest that individual populations of viruses may be physically enriched, and possibly isolated, from complex assemblages without cultivation. These fractionation methods are expected to improve our ability to characterize the genotypes and phenotypes of the uncultivated majority of viruses in aquatic environments.