Tomato spotted wilt virus impairs primary metabolism in tomato in a virus accumulation-dependent manner

Tomato spotted wilt virus (TSWV; Orthotospovirus tomatomaculae) is one of the major agricultural threats due to its worldwide distribution and broad host range. In the last decades, several tomato varieties carrying the Sw-5b gene for resistance to TSWV have been released. However, resistance conferred by Sw-5b can be overcome by the Sw5-Resistance-Breaking (SRB) strains. We investigated the interaction between Sw5b-carrying tomatoes and SRB TSWV to elucidate the molecular mechanisms underlying the resistance breakage. Eighteen samples from field-grown tomatoes naturally infected by TSWV in Italy were analyzed by transcriptome sequencing (RNA-Seq). Samples were categorized into four groups based on symptoms and virus titer: ASY (asymptomatic leaves with an average of 0.7×102 TSWV reads per million, RPM) and three symptomatic samples, i.e., SYM1 (3.3×104 RPM), SYM2 (7×104 RPM), and SYM3 (1.7×105 RPM). Remarkably, the higher the virus titer, the greater number of differentially expressed genes (DEGs) were found, which ranged from 33 to 44% of the whole transcriptome in SYM1 or SYM3 compared to ASY, respectively. Photosynthesis, protein biosynthesis and translocation were the main down-regulated biological processes, while enzyme such as oxidoreductases and transferases, genes related to the response to biotic stimuli, solute transport, and vesicle trafficking were overall up-regulated. Moreover, the expression of ca. 48% of genes of the whole transcriptome was significantly (P<0.05) correlated (positively or negatively) to the virus titer, and in 6% of cases (over 2000 genes) the correlation was very high (i.e., absolute value of R2>0.85). This finding was verified by a RT-qPCR assay in a greenhouse experiment with a Sw5b-tomato variety artificially inoculated with an SRB TSWV strain. The analysis of 15 genes in 12 samples with a TSWV titer ranging from 0 to 3.2×105-fold change confirmed the expected trend (positive or negative) of the gene expression × virus titer correlation. In conclusion, we elucidated the transcriptional changes induced by SRB TSWV on Sw5b-carrying tomatoes and showed that plant response to virus infection is strongly correlated to its accumulation level. Our finding highlights the importance of routinely determining the virus titer in plant-virus interaction studies to better interpret the results.