Unbiased tagging, isolation, and genomic characterization of bacteriophage transduction events in wastewater ecosystems

Investigators: Steven Ripp, Alice Layton, Shawn Hawkins, Courtney Johnson

Over the past few years there has been a revolutionary change in our understanding of the peculiar kinetics of the bacteriophage. Although we have known that phage parasitize bacterial hosts and in so doing mediate a horizontal gene transfer mechanism referred to as transduction, the magnitude of this gene transfer event is only now being realized. Much of the impact of gene transfer derives from the shear numbers of phage being found in the environment, now estimated at around 1031 – a 10-fold greater population than that of bacteria. A mere 500 or so of these phage have been sequenced, yet the information gained seems to suggest that phage are driving much of bacterial diversity and evolution. Phage genetic elements can be found in virtually all bacterial whole genome sequences, and with transduction occurring at rates approaching 2 × 1016 times per second, the magnitude of DNA turnover and exchange mediated by phage on a daily basis must be overwhelming. Additionally, some of this genetic exchange includes antibiotic resistance genes, exotoxins, mobile DNA elements, and even whole plasmids, making gene transfer risks and hazards especially prevalent. Adding to these risks is the poorly quantified number of bacterial hosts that a phage may potentially infect. Phage have always been considered to have very narrow host ranges, typically infecting only a few specific bacterial strains. Much of the evidence for this concept was obtained through highly biased laboratory plaque assays that enumerated only limited numbers of cultivable bacterial hosts. It is now being discovered that phage have much broader host ranges, often spanning across species. Thus, the interactions a phage may have with its global bacterial population may be extensive, as would the inter- and intra-species transfer of DNA within this population. To better elucidate this potential and to form a valid basis for gene transfer risk assessment, we have devised a method for tracking and isolating phage after infection of their host population. By introducing phage into a natural wastewater ecosystem, we believe the bacterial contacts and infectious cycles that it participates in will not be inclusive of only a few bacterial strains. Isolating the tagged phage at specific intervals and analyzing its accumulative genomic content will provide time series snapshots of where the phage has been and with whom it has been in contact with. In terms of risk assessment, these experiments will provide the first quantification of phage-host gene transfer dynamics in a natural, unadulterated ecosystem.

For more information, contact Steven Ripp.