Design and preliminary analytical evaluation of an in-house primer panel for dengue virus whole-genome sequencing: Evidence from Chhattisgarh, India.

Next-generation sequencing offers higher resolution than targeted gene sequencing for genomic surveillance and phylogenetic analysis of dengue virus (DENV). However, limited availability of commercial whole genome sequencing assays restrict comprehensive DENV genomic analysis. This study thus aimed to design, develop and evaluate an in-house DENV NGS primer panel and workflow, compatible with all four DENV serotypes, to investigate the circulating serotypes and genotypes in Chhattisgarh, India. The in-house primers panels were used to generate library and subsequent sequencing in Illumina MiniSeq using paired-end sequencing. A total of 30 archived DENV positive serum samples confirmed by NS1/Dengue IgM/IgG assay were included to check efficacy of in-house primer panel. Complete genome consensus sequences were successfully obtained for all 30 samples using the in-house DENV NGS primer panel and workflow processed in repetition. Sequencing-based serotyping demonstrated 100% concordance with RT-PCR results, without observing any discrepancies. Phylogenetic analysis revealed the circulation of DENV-1 (n = 1, 3.3%), DENV-2 (n = 23, 76.6%), and DENV-3 (n = 6, 20.0%) in Chhattisgarh. Notably, DENV-2 strains exhibited 20 distinct amino acid substitutions in the E protein, indicating potential genetic diversity within circulating strains. The docking analysis suggests that mutations within the DENV-2 E protein may induce structural changes capable of modulating interaction with DC-SIGN. The developed in-house DENV WGS primer panel provides a cost-effective, high-resolution approach enabling comprehensive genomic surveillance, mutation analysis, and epidemiological insights in resource-limited settings with reliable sequencing depth and coverage. Mutation-induced structural changes in the E protein alter receptor-binding interfaces and conformation, potentially affecting viral attachment and host-cell entry.