Comprehensive molecular profiling of the African swine fever virus in Korean wild boars between 2019 and 2024.

African swine fever (ASF), caused by the African swine fever virus (ASFV), is a lethal transboundary disease. Since its first detection in South Korea in 2019, ASFV has continuously infected wild boars, imposing major economic and ecological burdens. We investigated ASFV genetic diversity and transmission dynamics by analyzing 4,209 ASFV-positive wild boar samples collected between 2019 and 2024 using a multi-marker approach, including B646L (p72), the intergenic region (IGR) between I73R/I329L, MGF 360-1La, and MGF 505-9R/10R. All isolates were classified as Genotype II by p72 analysis. IGR II was predominant, while one case of IGR I and three of IGR III were detected in 2019-2020. IGR III reappeared in 2023 and increased in 2024. The Korean-specific MGF 360-1La mutation (L106P) emerged in 2020, peaked in 2021, and persisted thereafter at a low frequency. A shift in the MGF 505-9R/10R profile was observed: MGF-1 dominated until 2020, whereas MGF-5 emerged in 2021 and became predominant in 2024. Accordingly, ASFV strains were classified into six clusters with distinct spatial distributions and transmission trajectories. Cluster 1 represented the initial incursion and early spread, and two minor subclusters (1.1 and 1.2) were detected early on; Cluster 2 remained localized, Cluster 3 expanded southeastward and is emerging as dominant, and Cluster 4 was confined to eastern Gyeongsangbuk-do. These findings suggest that although ASFV in Korea originated from a single Genotype II introduction, the detection of multiple lineages reflects viral diversification or additional incursions. Continuous molecular surveillance using genetic markers, complemented by whole-genome analyses, is essential for detecting early variants and developing effective ASF control strategies.