Epidemiological and Evolutionary Dynamics of Dengue Virus in Saudi Arabia: Insights from Three Decades of Molecular and Serological Surveillance.

Dengue fever represents a significant public health challenge in Saudi Arabia, yet comprehensive molecular characterization of circulating serotypes remains limited. This study combines epidemiological and phylogenetic analyses to understand dengue virus (DENV) dynamics in the Kingdom. A systematic review and meta-analysis of dengue epidemiological data from Saudi Arabia (1992-2026) was the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. All available DENV envelope (E) gene sequences from Saudi human cases (1992-2023) were retrieved from GenBank and Global Initiative on Sharing All Influenza Data (GISAID). Phylogenetic trees were constructed using maximum likelihood with 1000 bootstrap replicates and best-fit models. Selection pressure was analyzed using SLAC, FEL, FUBAR, and MEME methods, while glycosylation sites were predicted with NetNGlyc and NetOGlyc. The pooled seroprevalence from 25 studies (n = 32,393) was 40.71% (95% CI: 26.96-56.10%). DENV-2 predominated (80.25%), followed by DENV-1 and DENV-3, with DENV-4 remaining rare (0.42%). Males (67-78%) and adults aged 25-44 years were most affected. Phylogenetic analysis of 50 Saudi isolates showed DENV-1 strains clustered within American-African (1994) and Asian (2004-2011) genotypes, all DENV-2 within the Cosmopolitan genotype, and all DENV-3 within Genotype III (bootstrap support 99-100%). Selection pressure analysis indicated pervasive positive selection in DENV-2, episodic selection across serotypes, and strong purifying selection in the E gene. Several amino acid substitutions with potential functional importance were identified. No DENV-4 E gene sequences from Saudi Arabia are publicly available. Dengue in western Saudi Arabia is characterized by DENV-2 predominance, co-circulation of three serotypes, and multiple introductions. The absence of DENV-4 sequences highlights critical surveillance gaps. Sustained molecular surveillance, expanded genomic sequencing, and data sharing are essential for effective prevention and vaccine preparedness.