Cross-order detection of bacteriophage transduction in microbial communities using RNA barcoding

Bacteriophages (phages) facilitate gene transfer and microbial evolution in all ecosystems and have applications as tools for engineering microbiomes and as antimicrobials. Historic efforts to map phage hosts, such as plaque assays, are limited to cultured bacteria, are low throughput, and are hard to apply in microbial communities and environmentally-relevant contexts. To overcome these limitations, we integrate a synthetic ribozyme that stores information about participation in horizontal gene transfer in 16S ribosomal RNA (rRNA) into the phage-plasmid P1, and perform targeted 16S rRNA sequencing following transduction to identify phage-host interactions. Experiments in synthetic and wastewater communities reveal Aeromonadales as a previously unreported P1 host order and show P1 transduction into pathogens. In wastewater, host range varies across phagemids having different origins of replication and phage-derived particles having different tail fibers. This work shows how autonomous barcoding can be used in phages to identify the molecular controls on their host range in microbial communities. Bacteriophages are the most abundant life form on earth and can be applied to eliminate or engineer bacteria. Here, authors demonstrate RNA barcoding as a high throughput tool to measure bacteriophage host range in natural microbial communities and inform bacteriophage ecology and applications.