Abstract
BACKGROUND: We observed a discrepancy between dengue NS1 antigen test and molecular diagnostics, with the emergence of dengue virus (DENV) serotype 3 in Sri Lanka, and sought to understand the cause for the rise in cases and high failure rates of molecular diagnostics. METHODS: Whole-genome sequencing was carried out in 22 DENV-3 samples. Phylogenetic and molecular clock analyses were done for genotype assignment and to understand the rate of evolution. Mutation analysis was done to understand the reasons for polymerase chain reaction (PCR) nondetection. RESULTS: We identified 2 DENV-3 genotypes (I and III) cocirculating. DENV-3 genotype III strains shared a common ancestor with a sequence from India collected in 2022, while DENV-3 genotype I, was found to share a common ancestor with DENV-3 sequences from China. DENV-3 genotype III was detected by the modified Centers for Disease Control and Prevention DENV-3 primers, whereas genotype I evaded detection due to key mutations at forward and reverse primer binding sites. We identified point mutations C744T and A756G in the forward primer binding sites and G795A in the reverse primer binding sites, which were not identified in DENV-3 genotype III. Furthermore, our Sri Lankan DENV-3 strains demonstrated a high root to tip ratio compared to the previous DENV-3 sequences, indicating a high mutation rate during the time of sampling (2017 to 2023). CONCLUSIONS: The cocirculation of multiple genotypes associated with an increase in cases highlights the importance of continuous surveillance of DENVs to identify mutations resulting in nondetection by diagnostics and differences in virulence.