A team of researchers from Rice University has discovered previously unknown relationships between bacteriophages—viruses that infect bacteria—and their bacterial hosts. This discovery could become a powerful tool for developing next-generation microbiome engineering technologies.

A study published in the journal Nature Communications uses an RNA barcoding system developed at the university, which allows scientists to identify which bacteria receive genetic material from bacteriophages in complex microbial communities. Using this approach, the researchers identified a previously undescribed group of bacterial hosts for the well-studied bacteriophage P1 and also investigated how minor changes in the virus's structure influence the microorganisms it can infect.

"Phages are ubiquitous, and they play a vital role in shaping microbial communities and transferring genes between bacteria," noted study author Lauren Stadler. "However, identifying which phages interact with which bacteria in real microbial ecosystems has long been challenging. Our work provides a scalable way to directly observe these interactions."

WHY IS TRACKING PHAGES SO DIFFICULT?

Bacteriophages are the most numerous biological entities on Earth, outnumbering all other life forms. They influence microbial ecosystems by killing bacteria, altering their metabolism, and transferring genes between organisms. Scientists are increasingly interested in using phages as an alternative to antibiotics and as tools for microbiome management. However, traditional methods for determining which bacteria a particular phage can infect often require culturing the bacteria in the laboratory, are labor-intensive, or fail to distinguish between simple viral attachment to a cell and successful DNA transfer.

To overcome these limitations, a team from Rice University, also including James Chappell and Jonathan Silberg, adapted a synthetic biology platform called RNA-addressable modification. It was originally developed to track gene transfer between bacteria through conjugation.

The system uses a specially engineered ribozyme—an RNA molecule capable of catalyzing specific biochemical reactions. After a bacterium receives DNA from a phage, the ribozyme inserts a unique "barcode" into its 16S ribosomal RNA. Scientists can then identify the recipient bacterium using targeted RNA sequencing.

"Rather than isolating each interaction individually, we allow the phage to leave a molecular signature on the cells it reaches," Stadler explained. "This gives us a sensitive and high-throughput way to map the host spectrum directly within microbial communities."

CHECKING THE SYSTEM IN WASTEWATER

The researchers embedded a barcoding system into bacteriophage P1, a virus known to transfer DNA between enterobacteria and likely facilitate the spread of antibiotic resistance genes. They then tested this approach in both laboratory-grown microbial communities and wastewater samples collected from a wastewater treatment plant in the Houston area.

Experiments with wastewater yielded an even more unexpected result. Among the organisms receiving genetic material from P1 were members of the Aeromonadales order, including the bacterium Aeromonas hydrophila—a common wastewater inhabitant that had never previously been considered a host for the P1 phage.

"Discovering an entirely new host group in a complex natural environment demonstrates the power of this approach," Stadler noted. "It's likely that there are many important interactions between phages and bacteria that remain hidden simply because we haven't had the tools to uncover them yet."

HOW TAIL FIBERS CHANGE THE HOST SPECTRUM

The team also used the new technology to investigate how different viral tail fibers—the protein structures that phages use to recognize and attach to bacteria—affect host range.

By engineering phage particles with alternative tail fiber variants and using an RNA barcoding system, the researchers showed that each tail fiber type targets different groups of microorganisms in wastewater communities.

"These experiments allowed us to see how relatively small genetic changes in a phage can dramatically alter the range of bacteria it interacts with," Stadler said. "This information is extremely valuable for designing phages with targeted properties—whether delivering beneficial genes or selectively killing harmful bacteria."

In the future, this method could accelerate the development of genetically modified bacteriophages for medicine, environmental cleanup, and industrial biotechnology. Because the approach relies on common molecular biology techniques such as amplicon sequencing rather than labor-intensive bacterial cultivation, it could also pave the way for large-scale studies of viral ecology in a wide variety of microbiomes.