Genetic Diversity, Evolutionary Origins, and Transmission Patterns of the ECSA Chikungunya Virus strain in Cambodia.

Abstract

Background: Chikungunya virus (CHIKV), a re-emerging arbovirus of global concern, caused amajor outbreak in Cambodia from 2020–2022. However, a critical gap remains in our understanding of its genomic diversity, evolutionary origins, and transmission patterns within the region, as comprehensive full-genome phylogenetic and phylodynamic analyses are limited. The phylogenetic investigation of this outbreak might provide critical lessons to inform public health interventions for managing future outbreaks in LMIC settings. Methods: This involved the bioinformatic analysis of 68 viral genomes collected from patients at four healthcare facilities in Cambodia. Following quality control (fastqc) and the exclusion criteria, 48 samples were selected for downstream analysis. Variant calling (using bcftools) and functional annotation (using snpEff) were conducted to characterize the mutational landscape of the outbreak CHIKV virus. Phylogenetic analysis (using Nextstrain) was performed to genotype the virus and determine their evolutionary relationship to global lineages. Phylodynamic and phylogeographic models were applied to estimate the time of the most recent common ancestor (TMRCA) and reconstruct transmission patterns in Nextstrain. Results: Of 48 patients, all had high fever, 32% had musculoskeletal symptoms, and 30% had a rash. All viral samples were of the ECSA-IOL genotype. These genomes lacked the Aedes albopictus-adapting E1:A226V mutation but featured other mutations (E1:K211E, E2:V264A, E1:D284E, E2:I211T) linked to increased replication in Ae. aegypti and mammalian adaptation. Phylogenetic analysis showed a close relationship to strains from Thailand. The estimated time of origin was June 2017, suggesting six years of cryptic circulation before the outbreak. Phylogeographic reconstruction identified Thailand as the likely source and Phnom Penh as a major national transmission hub, indicating a rapid superspreading event. Conclusion: The 2020-2022 Cambodian chikungunya outbreak was initiated by the ECSA-IOL lineage virus from Thailand circa 2017, which underwent a prolonged period of cryptic circulation. Our genomic findings confirm a transmission cycle primarily mediated by Aedes aegypti mosquitoes, characterized by adaptive mutations enhancing viral fitness. These results highlight the critical need for strengthened genomic surveillance systems in Cambodia and other LMICs to promptly detect and mitigate future outbreaks.