Clades of huge phages from across Earth’s ecosystems

Abstract

Bacteriophages typically have small genomes 1 and depend on their bacterial hosts for replication 2 . Here we sequenced DNA from diverse ecosystems and found hundreds of phage genomes with lengths of more than 200 kilobases (kb), including a genome of 735 kb, which is-to our knowledge-the largest phage genome to be described to date. Thirty-five genomes were manually curated to completion (circular and no gaps). Expanded genetic repertoires include diverse and previously undescribed CRISPR-Cas systems, transfer RNAs (tRNAs), tRNA synthetases, tRNA-modification enzymes, translation-initiation and elongation factors, and ribosomal proteins. The CRISPR-Cas systems of phages have the capacity to silence host transcription factors and translational genes, potentially as part of a larger interaction network that intercepts translation to redirect biosynthesis to phage-encoded functions. In addition, some phages may repurpose bacterial CRISPR-Cas systems to eliminate competing phages. We phylogenetically define the major clades of huge phages from human and other animal microbiomes, as well as from oceans, lakes, sediments, soils and the built environment. We conclude that the large gene inventories of huge phages reflect a conserved biological strategy, and that the phages are distributed across a broad bacterial host range and across Earth's ecosystems.Phages-viruses that infect bacteria-are considered distinct from cellular life owing to their inability to carry out most biological processes required for reproduction. They are agents of ecosystem change because they prey on specific bacterial populations, mediate lateral gene transfer, alter host metabolism and redistribute bacterially derived compounds through cell lysis 2-4 . They spread antibiotic resistance 5 and disperse pathogenicity factors that cause disease in humans and animals 6,7 . Most knowledge about phages is based on laboratorystudied examples, the vast majority of which have genomes that are a few tens of kb in length. Widely used isolation-based methods select against large phage particles, and they can be excluded from phage concentrates obtained by passage through 100-nm or 200-nm filters 1 . In 2017, only 93 isolated phages with genomes that were more than 200 kb in length were published 1 . Sequencing of whole-community DNA can uncover phage-derived fragments; however, large genomes can still escape detection owing to fragmentation 8 . A new clade of human-and animal-associated megaphages was recently described on the basis of genomes that were manually curated to completion from metagenomic datasets 9 . This finding prompted us to carry out a more-comprehensive analysis of microbial communities to evaluate the prevalence, diversity and ecosystem distribution of phages with large genomes. Previously, phages with genomes of more than 200 kb have been referred to as 'jumbophages' 1 or, in the case of phages with genomes of more than 500 kb, as megaphages 9 . As the set reconstructed here span both size ranges we refer to them simply as 'huge phage...

Accurate and complete genomes from metagenomes

“…Another seemingly rare error involves the artificial concatenation of an identical sequence, sometimes of hundreds of base pairs in length, repeated up to (or more than) three times. This has been a problem with some sequences of seemingly large phage deposited in public databases, as discussed by Devoto et al (2019) and Al-Shayeb et al (2020). This phenomenon is easily identified by running a repeat finder, a step that should also be included in the curation to completion pipeline (see below).…”

Long-read shotgun metagenome sequencing using PromethION uncovers novel bacteriophages, their abundance, and interaction with host bacterial immunity in the oral microbiota

“…For the first time in oral microbiota publications, we discovered six phages with genomes larger than 200 kb (Fig. S6, Table 1), that were classified as jumbo, and had been rarely found but recently began to be identified across Earth’s ecosystems (37, 38). We also discovered five jumbo (>200 kb) prophages.…”

“…3), and identified as much as ten jumbo oral phages/prophages (including the plasmid-like element) in an oral environment, significantly increasing our knowledge about “who is there” in the human oral cavity. Such jumbo phages were previously found among approximately 20 host bacterial genus (38, 46, 47) and recently began to be identified across Earth’s ecosystems (37). However, so far, there is no publication of those in the human oral cavity.…”

Making sense of virus size and the tradeoffs shaping viral fitness

“…By nature, viruses are smaller than the cells they infect, but the range of virus sizes is nonetheless substantial, with lengths of viral particles (virions) varying from 17 nm to ~1.5 µm, and genome size varying from ~1 kb to 2.5 Mb (Campillo-Balderas et al 2015). The largest 'giant' viruses have primarily been isolated from unicellular protists (Campillo-Balderas et al 2015, Wilhelm et al 2017, although there is metagenomic evidence for 'megaphages' of prokaryotes with genomes up to 716 kb (Devoto et al 2019, Al-Shayeb et al 2020, and a chaetognath appears to be infected by viruses 1.25 µm in length (Shinn and Bullard 2018). In contrast, the known viruses of plants and fungi have genomes < 30 kb (Campillo-Balderas et al 2015).…”