Lifestyle-associated divergence in genomic features, codon usage, and predicted translational adaptation across Yersinia enterocolitica phages

Virulent and temperate bacteriophages (phages), infecting Yersinia enterocolitica , experience distinct evolutionary pressures because of their contrasting lifestyles. However, how lifestyle-associated differences shape genome composition, codon usage, translational adaptation to the host, and the energetic costs of protein biosynthesis in these phage groups remains poorly understood. We compared experimentally validated Y. enterocolitica virulent and temperate phages across these genomic and predicted translational features. Virulent phages showed greater nucleotide heterogeneity, with 91.4% of within-group genome pairs lacking megablast-detectable similarity versus 76.2% for temperate phages. Their genomes also spanned a broader size range (39,261–352,596 bp versus 29,546–47,467 bp), although strongly influenced by three jumbo phages; the rank-based difference remained significant after their exclusion (Mann–Whitney U = 136.0, p = 0.0248; Cliff’s δ = 0.511). CDS length and predicted molecular weight did not differ significantly between lifestyles. Temperate-phage CDSs had higher median overall GC content (48.0% versus 37.8%), GC3 content (48.0% versus 31.1%), and observed and expected effective numbers of codons, indicating broader synonymous codon usage. Virulent-phage CDSs showed higher predicted codon adaptation (median CAI2, 0.731 versus 0.703) and translation-elongation efficiency (ITE, 0.590 versus 0.567). Iterative balanced downsampling yielded a median PERMANOVA R² of 0.189 (empirical 95% range, 0.178–0.200), with p = 0.001 in all 1,000 iterations; PERMDISP was consistently significant, indicating differences in multivariate location and dispersion. Primary phage-level permutation analysis supported lifestyle effects for nine of 11 variables and lifestyle × functional-group interactions for seven, although no interaction remained significant after Holm correction in the CR2 sensitivity analysis. Thirty-two codons remained significant after false-discovery-rate correction, with greater codon-usage dispersion among virulent phages. Y. enterocolitica virulent and temperate phages exhibited distinct genomic-compositional and predicted translational signatures. Virulent phages showed greater genomic and codon-usage heterogeneity, lower GC content, more restricted codon usage, and higher predicted translational-adaptation indices, whereas temperate phages showed broader codon usage and GC content closer to that of the bacterial host. These findings support an association between phage lifestyle, genome composition, codon usage, and predicted translational adaptation, while recognizing contributions from phage lineage, genome size, gene function, mutational bias, and host-associated selective pressures.