Acibenzolar-S-methyl inhibits accumulation of replication-associated DNA of tomato yellow leaf curl virus

Acibenzolar-S-methyl (ASM), a functional analogue of salicylic acid, induces systemic acquired resistance and restricts infection by several RNA viruses, yet its mechanisms against DNA viruses remain unknown. By examining the accumulation of tomato yellow leaf curl virus (TYLCV) DNA in inoculated leaves of Nicotiana benthamiana, we investigated whether ASM suppresses begomovirus infection. TYLCV was inoculated by agroinfiltration, and virion-sense (VS) and complementary-sense (CS) strands of TYLCV DNA were quantified separately using two-step strand-specific qPCR. ASM pretreatment significantly reduced the total TYLCV DNA accumulation at 1, 2, and 3 days post-inoculation. To determine whether this reduction reflected a direct effect on TYLCV rather than impaired Agrobacterium-mediated T-DNA delivery, we used a replication-defective mutant (TYLCVΔC1) as a delivery control and confirmed that ASM did not affect T-DNA delivery. Both VS and CS strands of wild-type TYLCV were substantially reduced by ASM treatment, providing the first direct evidence that ASM suppresses begomovirus infection by targeting viral DNA accumulation at the replication stage.