Detection of lumpy skin disease virus belonging to cluster 2.5 in flies collected during the outbreak in the Republic of Korea

Lumpy skin disease (LSD) is a vector-borne viral disease of cattle caused by lumpy skin disease virus (LSDV), which has recently expanded into Asia. In 2023, the Republic of Korea (ROK) experienced its first outbreak of LSD; however, information on the involvement of arthropods and the genetic lineage of circulating LSDV during the initial outbreak remained limited. This study aimed to investigate the presence of LSDV in flies collected from outbreak farms and to determine the genetic lineage of the detected virus. 672 arthropods, including biting and non-biting flies, and mosquitoes were collected from 94 cattle farms affected during the first LSD outbreak in the ROK between October and November 2023. Sample pools were initially screened for LSDV using a commercial DIVA (differentiating infected from vaccinated animals) qPCR assay. A total of four pools (2.67%) out of 150 pools tested positive for LSDV. qPCR-positive pools were confirmed by conventional PCR targeting the LW126 gene. Genetic sequences were successfully obtained from the four confirmed LSDV-positive pools. Phylogenetic analysis of the confirmed LSDV-positive pools was performed using sequencing of the G-protein-coupled chemokine receptor (GPCR) and RNA polymerase 30 kDa subunit (RPO30) genes. LSDV DNA was detected and confirmed in both biting and non-biting fly pools collected from outbreak farms. Phylogenetic analyses based on GPCR and RPO30 gene sequences revealed that the detected virus clustered with Asian recombinant LSDV strains belonging to cluster 2.5, which are genetically distinct from classical African field strains and Neethling-based vaccine strains. This study provides the first molecular detection of LSDV belonging to the Asian recombinant lineage detected in field-collected flies during the first LSD outbreak in the ROK. Although virus infectivity and transmission were not assessed, the detection of LSDV genetic material in flies associated with outbreak farms supports the inclusion of arthropod surveillance as part of LSD outbreak investigations in newly affected regions.