Comparative chloroplast genomics reveals codon usage bias, simple sequence repeat dynamics, and phylogenomic...

Urticaceae is an ecologically diverse family, yet comparative studies on plastome structure and codon usage remain limited. To investigate the structural evolution and codon usage patterns in Urticaceae, we sequenced the complete chloroplast genome of Urtica cannabina and performed comparative analyses of plastome structure, codon usage bias, simple sequence repeats (SSRs), selective pressure, and phylogenetic relationships using plastome sequences from 48 Urticaceae species. The Urtica cannabina plastome (147,958 bp) displays a canonical quadripartite architecture, encoding a total of 128 genes (83 protein-coding, 37 tRNAs, and 8 rRNAs). Comparative analysis revealed a pronounced A/T bias (GC content: 36.56%), with a distinct decreasing GC gradient across codon positions (GC₁ > GC₂ > GC₃). Relative synonymous codon usage (RSCU) analysis indicated a preference for A/U-ending codons, and the average effective number of codons (ENC) values ranged from 45.59 to 47.22, indicating generally weak codon bias. Parity rule 2 (PR2) analysis indicated a consistent preference for G and T at the third codon position in 48 Urticaceae chloroplast genomes, with highly conserved base usage across all species. Furthermore, a total of 3,552 SSRs were identified family-wide, with mononucleotide repeats (71.85%) predominating in intergenic regions. Ka/Ks analysis indicated that 97.1% of protein-coding genes were under purifying selection (Ka/Ks < 0.5). Phylogenetic analysis showed that Hesperocnide tenella is closely related to the genus Urtica. This study characterizes the structural features and codon usage patterns of chloroplast genomes in Urticaceae, providing new insights into their evolutionary dynamics and establishing a basis for investigating the interplay among genomic traits, codon bias, and adaptive evolution.