Bacteriophages of the Human Gut: The "Known Unknown" of the Microbiome.
Shkoporov AN, Hill C
Cell Host Microbe. Feb 2019
COMMENT: In this review, the authors summarize the taxonomic composition, dynamics, and spatial structure of the human gut phageome, they also review the available methods and challenges associated with metagenomic analysis pipelines and future research on the human phageome.
The review focus on some of the most significant biases and problems associated with metagenomic analysis of viral populations in the human gut, such as, the total viral counts in the gut and the DNA amplification bias due to the complex mixture of species with different genome sizes and different types of nucleic acids. Other limitations in viral metagenomics are the insufficiency of viral sequence databases and the bacterial DNA contamination
The first metagenomic studies of fecal viromes revealed that most bacterial viruses in the gut (81%–93%) are novel and can be neither assigned a taxonomic position nor linked to a bacterial host.
Viral metagenomics of the human gut yields between 75% and 99% of reads that do not produce significant alignments to any known viral genome. Given that the majority of reads cannot be aligned to a closed-reference database, the alternative is an open-reference approach where viral reads are assembled into contigs, which are then classified and annotated through reference-based and de novo annotation steps.
De novo identification of novel viral genomes in the metagenomic datasets against a background of bacterial and eukaryotic DNA contamination presents an extremely challenging task. Viral lineages are polyphyletic by origin and fast evolution of many viruses leads to further sequence diversification, sometimes to an extreme degree where no DNA sequence similarity can be captured even between members of the same viral family
A number software tools, databases, and websites have been specifically designed for processing high-throughput virome sequencing data. A selection of software tools, along with some general-purpose tools useful for the steps from read filtration, trimming, and assembly to gene prediction, host prediction, taxonomic classification, and multivariate analysis of community composition, are listed in the review.
Currently, high-throughput sequencing using short-read-based technologies remains the primary approach to characterizing unculturable viral communities in the gut. However, assembly, mapping, and classification of short reads arising from mostly novel and unknown viral genomes (viral dark matter) represent a bioinformatic challenge. Long-read sequencing technologies are becoming an interesting option, as they can be used to assist in scaffolding of large novel viral genomes.
Deep sequencing of bacteriophages in the gastrointestinal tract has uncovered previously overlooked viral populations of high complexity with potential roles in regulation of overall microbiome composition and in the onset, progression, and treatment of gut and systemic disorders.
A major obstacle toward application of advanced phage-based diagnostics and therapeutics is our incomplete understanding of the structure, dynamics, and function of the normal gut phageome.
In the light of recent discovery of bacteriophage transfer and even phage-mediated microbiota correction in fecal microbial transplantation, it seems important to study the ability of bacteriophages to spread horizontally or vertically (from mother to infant) in human populations, as well as consequences of such spread for microbiota composition.
While cross-sectional virome studies are important to understand population-level variance, longitudinal studies will be needed to identify persistent and accessory virome fractions per individual and relationships between temporal shifts in microbiome and virome in different life stages.
Isolation of previously uncultured but highly abundant and important phages, such as crAss-like bacteriophages and members of family Microviridae will help us understand their biological properties and roles in the maintenance of a dynamic equilibrium in the gut.
Novel sequencing technologies and improved bioinformatic pipelines in combination with phage culture techniques and phage proteomics methods will help to shed light on the viral dark matter, link previously uncultured bacteriophages to their hosts, and help us to explore the functional potential of phage genomes.