Genome-resolved viral and cellular metagenomes revealed potential key virus-host interactions in a deep freshwater lake

Abstract

The use of metagenomics has dramatically expanded the known virosphere, but freshwater viral diversity and the ecological relationships between viruses and hosts remain poorly understood. In this study, we conducted a large-scale metagenomic exploration of planktonic dsDNA prokaryotic viruses by sequencing both virion (<0.22 μm) and cellular (0.22-5.0 μm) fractions collected spatiotemporally from a deep freshwater lake (Lake Biwa, Japan). This simultaneously reconstructed 183 complete (i.e., circular) viral genomes and 57 bacterioplankton metagenome-assembled genomes. Spatiotemporal, intra-and extra-cellular dynamics of individual viruses assessed through metagenomic read coverage revealed a hypolimnion-specific viral community analogous to the vertically-stratified bacterioplankton community. The hypolimnetic community was generally stable during stratification, but occasionally shifted abruptly due to lysogenic induction, presumably reflecting low bacterial productivity in the hypolimnion. Genes involved in assimilatory sulfate reduction were encoded in 20 (10.9%) viral genomes, including those of dominant viruses, and may aid viral propagation in sulfurlimited freshwater systems. Hosts were predicted for 40 (21.9%) viral genomes, encompassing 10 phyla (or classes of Proteobacteria) including ubiquitous freshwater bacterioplankton lineages (e.g., LD12 and Ca. Nitrosoarchaeum). Comparison with published metagenomes revealed phylogeographic connectivity of viral communities in geographically isolated habitats, probably following the phylogeographic connectivity of their hosts. Notably, analogous to their hosts, actinobacterial viruses were among the most diverse, ubiquitous, and abundant viral groups in freshwater systems, with high lytic replication rates in surface waters. This result suggested that freshwater Actinobacteria are under high viral lytic pressure, which likely facilitated their genomic micro-diversification to elude viral recognition. Significance StatementMetagenomics allows for the reconstruction of bacterial and viral genomes without cultivation, contributing substantially to elucidation of bacterial and viral diversity and their key ecological roles.In this study, we report the largest-scale metagenomic exploration of freshwater prokaryotic viruses to date. We investigated the hypolimnion of a deep freshwater lake and tracked viruses throughout their life cycles, including both intra-and extra-cellular phases. This method allowed us to reconstruct both viral and host genomes and characterize numerous novel virus-host interactions, including those involving ubiquitous freshwater bacterial lineages. When the data were compared with published metagenomic datasets, we uncovered genome-resolved phylogeographic connectivity among viruses from geographically isolated habitats and identified ecologically important viral groups that are ubiquitous in aquatic habitats.In total, 12 water samples were collected at a pelagic site on Lake Biwa, the largest freshwater lake in Japan. The epilimnion (5 m) was ...

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

“…Rank abundance in each sample (Fig. S4) seems to follow the power distribution as reported in ocean and lake virome studies (32, 33). For top 10% abundant viral sequences in each sample (“top 10% among category 1, 2, 4, 5” in Table S1), we conducted a recently developed gene calling specifically designed for phage genomes (34) that are very compact and often have overlapping adjacent genes (10.5-39.4% in the top 10% abundant viral sequences).…”

Microdiversity and phylogeographic diversification of bacterioplankton in pelagic freshwater systems revealed through long-read amplicon sequencing

“…To evaluate the performance of our single nucleotide-resolved analysis, we tested whether the single nucleotide polymorphisms (SNPs) predicted from ASVs can be reproduced in shotgun metagenomic reads. We focused on sequences of the CL500-11 lineage in Lake Biwa, and used the assembly and read mapping results generated from a sample collected in the hypolimnion of the lake in September 2016 [60]. Reads mapped to the contig containing the rRNA gene operon of CL500-11 were analyzed using IGV v. 2.4.10 software [61] to determine the base frequency for each nucleotide position.…”

“…To test the accuracy and sensitivity of our analysis, we focused on the CL500-11 lineage in the hypolimnion of Lake Biwa-the OTU and sample with the greatest number of reads (2606 reads) in the present study-and investigated whether the SNPs expected from ASVs could be reproduced through metagenomic read mapping. Although the metagenomic sample was collected a year after the main sampling period for the present study [60], metagenomic reads (i.e., randomly fragmented total DNA) reflect real sequence variance in the environment and thus are a good reference for assessing the performance of our approach. We identified 30 base positions with SNPs by aligning all 24 ASVs of CL500-11 detected in the sample (Fig.…”

“…6C). Notably, the original metagenomic study [60] assembled only one (ASV_2) of the ASVs present in the lake (Fig. 4).…”

Viral metagenomes of Lake Soyang, the largest freshwater lake in South Korea

“…Advances in viral metagenome (virome) studies have facilitated access to a vast amount of phage genomes without the need for phage cultivation. Although most of the virome sequences remain uninterpretable in terms of viral taxonomy and their host information due to a dearth of reference phage genomes across public databases, viral community structures within diverse environments, including ocean 3 , freshwater 4 , soil 5 , and gut 6 , have been accessed and predicted. The virome sequences are valuable for quantifying the abundance of specific phage genomes within the environment in silico 3 .…”