Human Papillomavirus (HPV) is a major contributor to various human cancers, particularly cervical cancer. Despite its significant impact, the codon usage bias in high-risk HPV types has not been extensively studied. Understanding this bias, however, could provide valuable insights into the virus itself and inform the optimization of vaccine design. This study explores codon usage bias within the genomes of 17 high-risk HPV types (HPV-16, 18, 26, 31, 33, 35, 39, 45, 51, 52, 53, 56, 58, 59, 66, 68, and 82) through comparative analysis. While overall codon usage preference across these genotypes is not highly significant, a notable trend emerges in the preference for codons ending in A or U, with 24 out of 26 favored codons (Relative Synonymous Codon Usage >
1) ending in A or U. Moreover, no common optimal codons are shared among the 17 genomes. The study also identifies the underrepresentation of CpG and ApA dinucleotides, alongside the overrepresentation of CpA and UpG, which likely contribute to codon usage preferences that may influence viral replication and immune evasion strategies. Integrated analysis further suggests that natural selection is the primary force driving codon usage bias in these high-risk HPV genomes. Additionally, these HPVs exhibit a limited set of favored codons shared with humans, potentially minimizing competition for translation resources. This study offers new insights into codon usage bias in high-risk HPVs and underscores the importance of this understanding for optimizing vaccine design.