Visualization of Viral Infection Dynamics in a Unicellular Eukaryote and Quantification of Viral Production Using Virus Fluorescence in situ Hybridization

11 pages, 7 figures, supplementary material https://www.frontiersin.org/articles/10.3389/fmicb.2020.01559/full#supplementary-material.-- All datasets generated for this study are included in the article/Supplementary Material ; One of the major challenges in viral ecology is to assess the impact of viruses in controlling the abundance of specific hosts in the environment. To this end, techniques that enable the detection and quantification of virus-host interactions at the single-cell level are essential. With this goal in mind, we implemented virus fluorescence in situ hybridization (VirusFISH) using as a model the marine picoeukaryote Ostreococcus tauri and its virus Ostreococcus tauri virus 5 (OtV5). VirusFISH allowed the visualization and quantification of the proportion of infected cells during an infection cycle in experimental conditions. We were also able to quantify the abundance of free viruses released during cell lysis, discriminating OtV5 from other mid-level fluorescence phages in our non-axenic infected culture that were not easily distinguishable with flow cytometry. Our results showed that although the major lysis of the culture occurred between 24 and 48 h after OtV5 inoculation, some new viruses were already produced between 8 and 24 h. With this work, we demonstrate that VirusFISH is a promising technique to study specific virus-host interactions in non-axenic cultures and establish a framework for its application in complex natural communities ; YC was supported by Ministerio de Economía, Industria y Competitividad, Gobierno de España (MINECO), grant number: BES-2014-067849 during her PhD. MS was supported by Agencia Canaria de Investigación, Innovación y Sociedad de la Información (ACIISI), grant: Viera y Clavijo 2016. Ministerio de Economía, Industria y Competitividad, Gobierno de España (MINECO), grant numbers: CTM2013‐43767‐P and CTM2016‐75083‐R supported the project and paid for the investigation (reagents, material, etc.). NG was supported by Agence Nationale de la Recherche (ANR), ...