Viruses, including bacteriophage and archaeal viruses, are the most abundant form of life on earth (1031). They interact with all life and shape the global ecosystem through their impacts on community composition and horizontal gene transfer. However, phage-host relationships have proven challenging to identify without use of culture-based experiments to generate unambiguous evidence for a phage’s presence in a given host. These experiments inherently require that all hosts are culturable, typically restricting the scope and microbial diversity that can be surveyed and limiting our understanding of potentially valuable phage-host relationships.

Proximity ligation sequencing is a powerful genomic method for associating viruses with their hosts directly in native microbial communities. Proximity ligation captures, in vivo, physical interactions between the host microbial genome and the genetic material of both lytic and lysogenic phage. Similar to culturing experiments, these linkages offer direct evidence that phage sequences were present within an intact host cell, thereby establishing a phage-host pair. However, unlike culturing experiments, proximity-ligation methods do not require the propagation of living bacterial cells and unlike single cell sequencing experiments, only capture phage-host interactions inside cells. The combination of intra-phage and phage-host signal enables us to simultaneously deconvolve viral genome bins (vMAGs) directly from metagenomes and to assign microbial hosts to large numbers of vMAGs without culturing.

Our application of this technology to hundreds of complex microbiome samples has yielded thousands of novel phage and archaeal virus genomes with host assignments, as well as large numbers of new microbial genomes. Through broad-scale application of proximity ligation sequencing, we are creating a global-scale database of highly diverse phage-host interactions from samples from across the world. We will present published and unpublished work highlighting the power of this approach in the field of metagenomic discovery.