COMPARATIVE WHOLE-GENOME ANALYSIS OF DENV-1 DENV-3 REVEALS SEROTYPE-SPECIFIC GENETIC DIVERSITY AND PROTEIN-LEVEL EVOLUTIONARY PATTERNS

Abstract

Background: Dengue virus (DENV) is a major mosquito-borne pathogen characterized by extensive genetic diversity and continuous viral evolution. Understanding the distribution of genomic variation among circulating serotypes is important for molecular surveillance and for identifying conserved and variable regions of the viral proteome. Methods: Whole-genome sequencing datasets comprising 166 DENV genomes, including DENV-1 (n=14), DENV-2 (n=105), and DENV-3 (n=47), were analyzed using a comparative genomic framework. High-quality sequence reads were subjected to quality control, reference-guided alignment, variant detection and functional annotation. Missense mutations were characterized at the protein and conserved-domain levels. For extended comparative analysis, mutation burden was normalized according to genome sampling and amino-acid sequence length, while sequence variability was assessed using Shannon entropy. Phylogenetic and genotype analyses were used to investigate evolutionary clustering, and codon-based selection analyses were considered for characterization of evolutionary constraints. Results: A total of 721 missense mutations were identified across the analyzed DENV proteomes, comprising 180 mutations in DENV-1, 377 in DENV-2, and 164 in DENV-3. DENV-2 therefore exhibited the highest absolute mutation count, although interpretation of serotype-specific mutation burden requires normalization because of unequal sampling. Protein-level analysis revealed heterogeneous mutation distribution, with NS5 showing pronounced variability, whereas the M protein was comparatively conserved. Domain-level analysis further demonstrated non-uniform distribution of sequence variation across functional regions. The extended analytical framework incorporating normalized mutation burden, entropy, phylogenetic relationships and selection pressure provides complementary measures for evaluating DENV genomic diversity and evolutionary constraint. Conclusions: This study establishes a comparative genome- and proteome-level framework for characterizing genetic diversity among DENV-1, DENV-2 and DENV-3. The identification of variable and conserved viral proteins and domains provides a foundation for genomic surveillance and for prioritizing regions for subsequent structural, functional, antiviral and vaccine-related investigations. Further analysis using the underlying genome sequences will be necessary to statistically validate normalized diversity, genotype assignments and site-specific selection signatures.