Genetic Diversity of Beet Necrotic Yellow Vein Virus in Serbia: First Detection of J-Type RNA5 and P25 Tetrad Associated with Resistance-Breaking Isolates

Beet necrotic yellow vein virus (BNYVV), the causal agent of rhizomania, is a major threat to sugar beet production worldwide. In Serbia, during a three-year monitoring period (2023–2025) conducted across 31 localities (72 fields) representing the main sugar beet production areas, BNYVV was detected in 26.5% of analyzed samples. Of the 88 ELISA-positive samples, 60 originated from cultivars carrying the Rz1, whereas 28 were obtained from cultivars carrying the combined Rz1+Rz2 resistance genes. This study presents the first analysis of the genetic diversity and population structure of BNYVV in Serbia, based on P25 tetrad variability and RNA5 sequence analysis. Sequence analysis revealed five distinct P25 tetrad variants, with the ALHG tetrad being the most prevalent. Moreover, in one locality in the Bačka region, the VHHG variant, previously associated with Rz1-resistance-breaking isolates, was also detected. Phylogenetic analysis based on p25 sequences grouped all Serbian isolates within the P25-I cluster, corresponding to the A-type virus population, and showed close relationship with European isolates, particularly those from Italy. Importantly, RNA5 was detected in all selected isolates, representing the first report of RNA5 in Serbia. The results of the p26sequence analysis revealed high conservation among Serbian isolates, which clustered within the J-type in phylogenetic analysis. Overall, the results demonstrate that Serbian BNYVV populations are genetically diverse and include variants associated with Rz1-resistance breaking. These findings highlight the need for continuous molecular surveillance to monitor the emergence and spread of virulent strains and to support effective disease management strategies in sugar beet cultivation.