Abstract
The microbiota of the human gut is a complex and rich community where bacteria and their viruses, the bacteriophages, are dominant. There are few studies on the phage community and no clear standard for isolating them, sequencing and analysing their genomes. Since this makes comparisons between studies difficult, we aimed at defining an easy, low-cost, and reproducible methodology. We analysed five different techniques to isolate phages from human adult faeces and developed an approach to analyse their genomes in order to quantify contamination and classify phage contigs in terms of taxonomy and lifestyle. We chose the polyethylene glycol concentration method to isolate phages because of its simplicity, low cost, reproducibility, and of the high number and diversity of phage sequences that we obtained. We also tested the reproducibility of this method with multiple displacement amplification (MDA) and showed that MDA severely decreases the phage genetic diversity of the samples and the reproducibility of the method. Lastly, we studied the influence of sequencing depth on the analysis of phage diversity and observed the beginning of a plateau for phage contigs at 20,000,000 reads. This work contributes to the development of methods for the isolation of phages in faeces and for their comparative analysis.
Highlights
The microbiota of the human gut is a complex and rich community where bacteria and their viruses, the bacteriophages, are dominant
Our understanding of human gut microbiota has drastically increased in the past decade, but the majority of gut metagenomic studies have focused on the bacterial component of the microbiota in healthy subjects and in patients suffering from various pathological conditions
In the context of a project where we study the bacterial microbiota of the faeces of several healthy individuals during several months[13], we looked for a simple, efficient, reproducible and inexpensive methodology for (i) phage isolation from faeces, (ii) DNA extraction and sequencing and (iii) computational analyses of the sequencing data
Summary
The microbiota of the human gut is a complex and rich community where bacteria and their viruses, the bacteriophages, are dominant. There are few studies on the phage community and no clear standard for isolating them, sequencing and analysing their genomes. There is a high inter-individual variation between the phageome of healthy individuals[9], but a recent study showed that a small set of phages are found in the majority of healthy people[10]. One of these is the crAssphage, a 97 kbp Podoviridae phage that is highly abundant and ubiquitous in the human gut metagenome[11]. Our limited knowledge about gut phages is reflected in the limited host taxonomic www.nature.com/scientificreports/
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
