Papillomaviruses are a heterogeneous group of DNA viruses with closed circular double-stranded DNA genomes of about 8 kb in size that contain three general regions. An upstream regulatory region (URR) contains sequences that control transcription and replication, an early region contains genes (e.g., E6, E7, E1, E2, E4, and E5) involved primarily in enzymatic activities, and a structural region produces the L1 capsid protein and L2, which facilitates packaging of the viral DNA. Human papillomaviruses (HPVs) are classified by the sequence similarity of their genomes. A cloned HPV genome whose L1 open reading frame (ORF) displays less than 90% similarity to previously designated types is defined as a novel type. To date, more than 90 different genotypes of the HPV have been fully characterized (21). Intratypic variants and subtypes are defined as HPVs that vary by less than 10% in their L1 DNA sequences (5, 21).HPVs are causally involved in the etiology of cervical cancer and its precursor lesions (16,17,43,49). Of the high-risk HPV types associated with cervical cancer, HPV16 is the most prevalent and is found in approximately half of all cancers (7, 49). Numerous variants of HPV16 have been identified in different geographic locations and ethnic groups (35,42,53,68). Although all HPV16 isolates are closely related, previous studies inferred five distinct phylogenetic branches among HPV16 variants: European (E), Asian (As), Asian-American (As-Am), African-1 (Af-1), and African-2 (Af-2), corresponding to the geographic locations from which the samples were obtained (12,34). Subsequent studies by sequence analyses of the HPV16 variants in other genomic regions (e.g., E6, L2, and L1) expanded and complemented this phylogenetic hypothesis (69).Although HPV16 variants are an important focus of phylogenetic studies and the molecular variants of E2, L2, L1, the URR, and especially the E6 region have been described in detail previously (28), covariation among different ORFs belonging to the same lineage or isolate have not been studied in great detail. HPV16 variants have demonstrable differences in biological properties in vitro which may be responsible, in part, for differences in pathogenicity, carcinogenic risk, and perhaps immunogenicity (28). Furthermore, HPV16 variants are associated with different cervical cancer risks (6,32,65,67). Although the diversifying selection in the HPV16 E6 and E7 oncogenes has been described recently (20), the evolutionary basis of the entire genome, coevolutionary mechanisms among different HPV16 genes, and their underlying biological significance remain unknown. Obtaining whole-genome sequences representative of the major HPV16 variants allowed us to determine with certainty nucleotide and amino acid sequence changes that are of potential evolutionary importance.Comparison of synonymous (silent; d S ) and nonsynonymous