Interaction dynamics and virus–host range for estuarine actinophages captured by epicPCR

Viruses impact microbial diversity, gene flow and function through virus–host interactions. Although metagenomics surveys are rapidly cataloguing viral diversity, methods are needed to capture specific virus–host interactions in situ. Here, we leveraged metagenomics and repurposed emulsion paired isolation-concatenation PCR (epicPCR) to investigate viral diversity and virus–host interactions in situ over time in an estuarine environment. The method fuses a phage marker, the ribonucleotide reductase gene, with the host 16S rRNA gene of infected bacterial cells within emulsion droplets providing single-cell resolution for dozens of samples. EpicPCR captured in situ virus–host interactions for viral clades with no closely related database representatives. Abundant freshwater Actinobacteria lineages, in particular Rhodoluna sp., were the most common hosts for these poorly characterized viruses, with interactions correlated with environmental factors. Multiple methods used to identify virus–host interactions, including epicPCR, identified different and largely non-overlapping interactions within the vast virus–host interaction space. Tracking virus–host interaction dynamics also revealed that multi-host viruses had significantly longer periods with observed virus–host interactions, whereas single-host viruses were observed interacting with hosts at lower minimum abundances, suggesting more efficient interactions. Capturing in situ interactions with epicPCR revealed environmental and ecological factors shaping virus–host interactions, highlighting epicPCR as a valuable technique in viral ecology. Using epicPCR, a method that fuses a phage marker gene with the 16S rRNA gene of infected bacterial cells at single-cell resolution, the authors identify host–virus interactions and dynamics over time in an estuarine environment.