Resurrection of a Global, Metagenomically Defined Microvirus

The Gokushovirinae (family Microviridae) are a group of single-stranded, circular DNA bacteriophages that have been detected in metagenomic datasets from every ecosystem on the planet. Despite their abundance, little is known about their biology or their bacterial hosts: isolates are exceedingly rare, known only from a very small number of obligate intracellular bacteria. By synthesizing circularized phage genomes from prophages embedded in diverse enteric bacteria, we produced viable gokushovirus phage particles that could reliably infect E. coli, thereby allowing experimental analysis of its life cycle and growth characteristics. Revived phages integrate into host genomes by hijacking a phylogenetically conserved chromosome-dimer resolution system, in a manner reminiscent of cholera phage CTX. Sequence motifs required for lysogeny are detectable in other metagenomically defined gokushoviruses, but we show that even partial motifs enable phages to persist in a state of pseudolysogeny by continuously producing viral progeny inside hosts without leading to collapse of their host culture. This ability to employ multiple, disparate survival strategies is likely key to the long-term persistence and global distribution of Gokushovirinae. The capacity to harness gokushoviruses as an experimentally tractable model system thus substantially changes our knowledge of the nature and biology of these ubiquitous phages.