Metagenomics, or sequencing of the genetic material from a complete microbial community, is a promising tool to discover novel microbes and viruses. Viral metagenomes typically contain many unknown sequences. Here we describe the discovery of a previously unidentified bacteriophage present in the majority of published human fecal metagenomes, which we refer to as crAssphage. Its ~97 kbp genome is six times more abundant in publicly available metagenomes than all other known phages together; comprises up to 90% and 22% of all reads in virus-like particle (VLP)-derived metagenomes and total community metagenomes, respectively; and totals 1.68% of all human fecal metagenomic sequencing reads in the public databases. The majority of crAssphage-encoded proteins match no known sequences in the database, which is why it was not detected before. Using a new co-occurrence profiling approach, we predict a Bacteroides host for this phage, consistent with Bacteroides-related protein homologs and a unique carbohydrate-binding domain encoded in the phage genome,.

Metagenomics has changed the face of virus discovery by enabling the accurate identification of viral genome sequences without requiring isolation of the viruses. As a result, metagenomic virus discovery leaves the first and most fundamental question about any novel virus unanswered: What host does the virus infect? The diversity of the global virosphere and the volumes of data obtained in metagenomic sequencing projects demand computational tools for virus–host prediction. We focus on bacteriophages (phages, viruses that infect bacteria), the most abundant and diverse group of viruses found in environmental metagenomes. By analyzing 820 phages with annotated hosts, we review and assess the predictive power of in silico phage–host signals. Sequence homology approaches are the most effective at identifying known phage–host pairs. Compositional and abundance-based methods contain significant signal for phage–host classification, providing opportunities for analyzing the unknowns in viral metagenomes. Together, these computational approaches further our knowledge of the interactions between phages and their hosts. Importantly, we find that all reviewed signals significantly link phages to their hosts, illustrating how current knowledge and insights about the interaction mechanisms and ecology of coevolving phages and bacteria can be exploited to predict phage–host relationships, with potential relevance for medical and industrial applications.

Microbial viruses can control host abundances via density-dependent lytic predator-prey dynamics. Less clear is how temperate viruses, which coexist and replicate with their host, influence microbial communities. Here we show that virus-like particles are relatively less abundant at high host densities. This suggests suppressed lysis where established models predict lytic dynamics are favoured. Meta-analysis of published viral and microbial densities showed that this trend was widespread in diverse ecosystems ranging from soil to freshwater to human lungs. Experimental manipulations showed viral densities more consistent with temperate than lytic life cycles at increasing microbial abundance. An analysis of 24 coral reef viromes showed a relative increase in the abundance of hallmark genes encoded by temperate viruses with increased microbial abundance. Based on these four lines of evidence, we propose the Piggyback-the-Winner model wherein temperate dynamics become increasingly important in ecosystems with high microbial densities; thus 'more microbes, fewer viruses'.

Motivation: Bacteriophages have two distinct lifestyles: virulent and temperate. The virulent lifestyle has many implications for phage therapy, genomics and microbiology. Determining which lifestyle a newly sequenced phage falls into is currently determined using standard culturing techniques. Such laboratory work is not only costly and time consuming, but also cannot be used on phage genomes constructed from environmental sequencing. Therefore, a computational method that utilizes the sequence data of phage genomes is needed.Results: Phage Classification Tool Set (PHACTS) utilizes a novel similarity algorithm and a supervised Random Forest classifier to make a prediction whether the lifestyle of a phage, described by its proteome, is virulent or temperate. The similarity algorithm creates a training set from phages with known lifestyles and along with the lifestyle annotation, trains a Random Forest to classify the lifestyle of a phage. PHACTS predictions are shown to have a 99% precision rate.Availability and implementation: PHACTS was implemented in the PERL programming language and utilizes the FASTA program (Pearson and Lipman, 1988) and the R programming language library ‘Random Forest’ (Liaw and Weiner, 2010). The PHACTS software is open source and is available as downloadable stand-alone version or can be accessed online as a user-friendly web interface. The source code, help files and online version are available at http://www.phantome.org/PHACTS/.Contact: katelyn@rohan.sdsu.edu; redwards@sciences.sdsu.eduSupplementary information: Supplementary data are available at Bioinformatics online.

Microbiomes are vast communities of microbes and viruses that populate all natural ecosystems. Viruses have been considered the most variable component of microbiomes, as supported by virome surveys and examples of high genomic mosaicism. However, recent evidence suggests that the human gut virome is remarkably stable compared to other environments. Here we investigate the origin, evolution, and epidemiology of crAssphage, a widespread human gut virus. Through a global collaboratory, we obtained DNA sequences of crAssphage from over one-third of the world's countries, and showed that its phylogeography is locally clustered within countries, cities, and individuals. We also found colinear crAssphage-like genomes in both Old-World and New-World primates, challenging genomic mosaicism and suggesting that the association of crAssphage with primates may be millions of years old. We conclude that crAssphage is a benign globetrotter virus that may have co-evolved with the human lineage and an integral part of the normal human gut virome.

MotivationCurrently there are no tools specifically designed for annotating genes in phages. Several tools are available that have been adapted to run on phage genomes, but due to their underlying design, they are unable to capture the full complexity of phage genomes. Phages have adapted their genomes to be extremely compact, having adjacent genes that overlap and genes completely inside of other longer genes. This non-delineated genome structure makes it difficult for gene prediction using the currently available gene annotators. Here we present PHANOTATE, a novel method for gene calling specifically designed for phage genomes. Although the compact nature of genes in phages is a problem for current gene annotators, we exploit this property by treating a phage genome as a network of paths: where open reading frames are favorable, and overlaps and gaps are less favorable, but still possible. We represent this network of connections as a weighted graph, and use dynamic programing to find the optimal path.ResultsWe compare PHANOTATE to other gene callers by annotating a set of 2133 complete phage genomes from GenBank, using PHANOTATE and the three most popular gene callers. We found that the four programs agree on 82% of the total predicted genes, with PHANOTATE predicting more genes than the other three. We searched for these extra genes in both GenBank’s non-redundant protein database and all of the metagenomes in the sequence read archive, and found that they are present at levels that suggest that these are functional protein-coding genes.Availability and implementation https://github.com/deprekate/PHANOTATE Supplementary information Supplementary data are available at Bioinformatics online.

The approximately 10 11 viruses and microbial cells per gram of fecal matter (dry weight) in the large intestine are important to human health. The responses of three common gut bacteria species, and one opportunistic pathogen, to 117 commonly consumed foods, chemical additives, and plant extracts were tested. Many compounds, including Stevia rebaudiana and bee propolis extracts, exhibited species-specific growth inhibition by prophage induction. Overall, these results show that various foods may change the abundances of gut bacteria by modulating temperate phage and suggests a novel path for landscaping the human gut microbiome.

BackgroundDiversity-generating retroelements (DGRs) are genetic cassettes that selectively mutate target genes to produce hypervariable proteins. First characterized in Bordetella bacteriophage BPP-1, the DGR creates a hypervariable phage tail fiber that enables host tropism switching. Subsequent surveys for DGRs conclude that the majority identified to date are bacterial or archaeal in origin. This work examines bacteriophage and bacterial genomes for novel phage-encoded DGRs.ResultsThis survey discovered 92 DGRs that were only found in phages exhibiting a temperate lifestyle. The majority of phage-encoded DGRs were identified as prophages in bacterial hosts from the phyla Bacteroidetes, Proteobacteria, and Firmicutes. Sequence reads from these previously unidentified prophages were present in viral metagenomes (viromes), indicating these prophages can produce functional viruses. Five phages possessed hypervariable proteins with structural similarity to the tail fiber of BPP-1, whereas the functions of the remaining DGR target proteins were unknown. A novel temperate phage that harbors a DGR cassette targeting a protein of unknown function was induced from Bacteroides dorei. This phage, here named Bacteroides dorei Hankyphage, lysogenizes 13 different Bacteroides species and was present in 34% and 21% of whole-community metagenomes and human-associated viromes, respectively.ConclusionsHere, the number of known DGR-containing phages is increased from four to 92. All of these phages exhibit a temperate lifestyle, including a cosmopolitan human-associated phage. Targeted hypervariation by temperate phages may be a ubiquitous mechanism underlying phage-bacteria interaction in the human microbiome.Electronic supplementary materialThe online version of this article (10.1186/s40168-018-0573-6) contains supplementary material, which is available to authorized users.