Cyanophage thioredoxin and virulence enhancer genes differentially contribute to phage fitness

Abstract Viruses carry many homologs of cellular genes as well as small genes of unknown function, often called viral dark matter. One cellular-like gene present in viruses is the redox protein thioredoxin, found in all domains of cellular life. However, the functional relevance and evolutionary benefit of viral-encoded thioredoxins, and small genes of unknown function, remain unclear. Both types of genes are present in marine cyanophages, viruses that infect the globally important marine cyanobacteria. In the T7-like cyanophage, Syn5, the phage thioredoxin overlaps with a small gene of unknown function that we named cve for cyanophage virulence enhancer. While thioredoxin genes are common across a wide variety of viruses infecting diverse host types, we found that cve is restricted to cyanophages. Genetic inactivation of thioredoxin and cve in the Syn5 cyanophage revealed that thioredoxin enhanced phage fitness, increasing phage genome replication and progeny production. Furthermore, redox active Syn5 thioredoxin negatively impacted host growth, indicating that its activity is detrimental to host metabolism. The cve gene increased phage virulence yet reduced phage burst size, and is thus a double-edged sword. Despite this trade-off, cve significantly enhanced overall phage fitness emphasizing the importance of virulence for viral fitness. Our findings indicate that viral-encoded thioredoxin and cve genes differentially alter infection properties to provide an evolutionary advantage to the Syn5 phage. They further reveal the function of a small, cyanophage-specific gene of unknown function, unveiling the first known host-type-specific virulence enhancer in viruses.