Mangle Chen Donghai Wang Jiali Yang Yuhao Cao Song Xuemei Yu Wen Lu Hongying Zheng Jiejun Peng Guanwei Wu Jian Wu Jun-Min Li Jianping Chen Fei Yan ^* Shaofei Rao ^*

• Institute of Plant Virology, Ningbo University, Ningbo, China

Tomato brown rugose fruit virus (ToBRFV) is an emerging, rapidly spreading virus belonging to the genus Tobamovirus which seriously decreases tomato yields.RNA silencing is an evolutionarily conserved antiviral mechanism. In virus-infected plants, virus-derived small interfering RNAs (vsiRNAs) are one of the key components involved in the RNA silencing-based antiviral activity in plants. The main function of vsiRNAs is to target and degrade viral RNA. Studies have found that some vsiRNAs can also target host transcripts, further regulating host responses and symptoms, promoting viral survival and spread. In this study, the vsiRNA profiles of ToBRFVinfected tomato plants were obtained by deep sequencing. VsiRNAs were mainly 21 and 22 nucleotides in length and had a U-bias at the 5' end. The single-nucleotide resolution profile shows that vsiRNAs exhibit a heterogeneous continuous distribution in the ToBRFV genomic RNA, with hotspot regions on the antisense strand located at the 5' end of the RdRP and in the coding regions of MP and CP. The presence of vsiRNAs was confirmed in tomato plants infected with ToBRFV through RT-PCR, and GO and KEGG enrichment analyses were performed on the predicted vsiRNA target genes (with an expectation value less than or equal to 2.5). Seven potential target genes were selected for qRT-PCR analysis, confirming that their transcript accumulation significantly decreased in the leaves of tomato plants infected with ToBRFV. These genes may play an important role in the process of viral infection in tomato. Our results suggest a role for vsiRNAs in the ToBRFV-tomato interaction.