Arvind Varsani (@viralnomad.bsky.social)

A functional cyanophage thioredoxin increases competitive phage fitness Thioredoxins are ubiquitous redox proteins that are found in all domains of life. These conserved proteins are also found in many phages, including marine cyanophages that infect the ecologically important marine cyanobacteria. However, their role in phage infection is not known. Cyanophages also carry many small genes lacking homology to known functional domains. Whether these have a functional role or not remains unknown. Here, we explore the distribution and role of a cyanophage thioredoxin (trxA), and that of a small gene directly downstream of it (g26), in phage infection. For this we used the T7-like cyanophage, Syn5, which infects an open-ocean marine Synechococcus strain, WH8109. We found that thioredoxin genes are common in phage genomes, including in cyanophages. The g26 gene, however, is restricted in it distribution to the cyanophages. The cyanophage thioredoxin is catalytically active and it increases phage DNA replication, progeny production and competitive fitness. It also negatively impacts host growth. The g26 gene product is translationally coupled to, and thus dependent on, translation of the thioredoxin gene. This gene itself significantly increases phage virulence and fitness, yet reduces burst size. Our findings demonstrate that cyanophage thioredoxins impact phage fitness and infection physiology and that small viral genes with no homology to known genes can play an important role in the infection process. These findings provide insights into the importance of unusual genes in phage genomes and show that they are likely to play an important role in the interactions between abundant cyanobacteria and cyanophages in ocean ecosystems. ### Competing Interest Statement The authors have declared no competing interest. https://www.biorxiv.org/content/10.1101/2025.10.05.680603v1