Limnology-Aquatic Ecology and Evolution, Limnological Institute, University of Konstanz, 78464 Konstanz, Germany.
Department of Biomolecular Mechanisms, Max Planck Institute for Medical Research, 69120 Heidelberg, Germany.
Viruses. 2022 May 16;14(5):1056. doi: 10.3390/v14051056.
Viruses are an abundant component of aquatic systems, but their detection and quantification remain a challenge. Virophages co-replicate with giant viruses in the shared host cell, and can inhibit the production of new giant virus particles, thereby increasing the survival of the infected host population. Here, we present a protocol for Droplet Digital PCR (ddPCR) to quantify simultaneously giant virus and virophage in a mixed sample, enabling the rapid, culture-free and high throughput detection of virus and virophage. As virophage can be present as free virus particles or integrated into the virus host's genome as well as associated with organic particles, we developed a simple method that enables discrimination between free and particle-associated virophages. The latter include aggregated virophage particles as well as virophage integrated into the host genome. We used, for our experiments, a host-virus-virophage system consisting of , CroV and mavirus. Our results show that ddPCR can be an efficient method to quantify virus and virophage, and we discuss potential applications of the method for studying ecological and evolutionary processes of virus and virophages.
病毒是水生系统中丰富的组成部分，但它们的检测和定量仍然是一个挑战。噬病毒体与巨型病毒在宿主细胞中共复制，并能抑制新的巨型病毒颗粒的产生，从而增加受感染宿主群体的存活率。在这里，我们提出了一种使用液滴数字 PCR（ddPCR）同时定量混合样本中巨型病毒和噬病毒体的方案，能够快速、无培养和高通量地检测病毒和噬病毒体。由于噬病毒体可以以游离病毒颗粒的形式存在，也可以整合到病毒宿主的基因组中，或者与有机颗粒相关联，我们开发了一种简单的方法，能够区分游离的和颗粒相关的噬病毒体。后者包括聚集的噬病毒体颗粒以及整合到宿主基因组中的噬病毒体。我们在实验中使用了一个由 CroV 和 mavirus 组成的宿主-病毒-噬病毒体系统。我们的结果表明，ddPCR 可以是一种有效的定量病毒和噬病毒体的方法，我们讨论了该方法在研究病毒和噬病毒体的生态和进化过程中的潜在应用。
