Optimized protocol for viral metagenomics studies
Next Generation Sequencing (NGS) has revolutionized the study of the human gut microbiome – the community of microorganisms (including bacteria, fungi, and viruses) present within the human gut. While in recent years, our understanding of the human microbiome has greatly improved, most studies have focused on bacteria, ignoring viral genomes. For this reason, little is known regarding the human virome –the viral component of the human microbiome. One of the main challenges in studying the role of the human gut virome in health is the absence of validated methods for high throughput and reproducible virome analysis.
In this study, the quantitative effects of different steps of sample preparation for virome analysis were evaluated with qPCR and next-generation sequencing (NGS). Several procedures were tested for the homogenization, centrifugation, filtration, and random amplification steps, using a mock-virome (including 9 highly diverse viruses among which coronaviruses) and a bacterial mock-community. As a result, an optimized protocol for fecal sample preparation was created, the NetoVIR (Novel enrichment technique of VIRomes). The NetoVIR protocol makes use of the Minilys (Bertin Technologies, France) to efficiently and uniformly homogenize samples. Thanks to Minilys-powered homogenization and other optimized steps.