Genome Wide Analysis Reveals Diverse and Novel Prophages in Tsukamurella

Prophages play key roles in bacterial evolution, yet their diversity in the genus Tsukamurella remains poorly characterized. In this study, we analyzed 55 publicly available Tsukamurella genomes and identified eight high-quality prophages, including seven high‑quality draft genomes and one complete circular genome, distributed among T. asaccharolytica (n = 1), T. paurometabola (n = 1), T. tyrosinosolvens (n = 2), T. soli (n = 1), and T. ocularis (n = 3). Comparative genomic analyses revealed three major clusters: the previously described Tsukamurella phages TIN2, TIN3, TIN4; the three T. ocularis prophages; and prophages from T. paurometabola and T. tyrosinosolvens. The remaining prophages showed limited similarities to known phages or prophages, highlighting their novelty. Despite substantial genomic divergence, key lysis-associated modules, particularly endolysin and holin families, were relatively conserved among phage and prophage genomes. These findings expand current understanding of Tsukamurella phage diversity and provide new insights into the evolutionary dynamics of actinobacteriophages.