Berbati, M.; Bousali, M.; Kaldis, A.; Moravec, T.; Karamitros, T.; Voloudakis, A. E.

Small RNA sequencing of healthy and ZYMV-infected watermelon (21 dpi) was performed and bioinformatics analysis identified 353 miRNAs from which 22 known and 331 new miRNAs. Important information about their precursors, their length, the loci of which they originated on watermelon genome are provided. The ZYMV genome could be a target for mir396a-3p, miR8706a, and miR1886i-5p from the miRbase, but none of them was identified in the watermelon miRNAome. Furthermore, watermelon miRNAome does not contain a miRNA targeting ZYMV genome with an expectation score [≤] 3.5. There were 34 resistance genes (CC-NBS-LRR, TIR-NBS-LRR, TIR-NBS) predicted as targets of 32 miRNAs in healthy watermelon. For nine differentially expressed miRNAs the respective target genes (10 in total) were bioinformatically predicted. For cla-new_miR307 (upregulated upon ZYMV infection) and cla-miR166h-3p (downregulated upon ZYMV infection) the targets were predicted to be ClaATRIP and ClaRBOHB, respectively, with ClaATRIP downregulated and ClaRBOHB upregulated upon ZYMV infection. ALSV-mediated VIGS of ClaATRIP rendered watermelon plants more resistant to ZYMV, whereas VIGS of ClaRBOHB resulted in higher levels of ZYMV titer in watermelon. These data suggest that ClaATRIP and ClaRBOHB are a susceptibility and resistant gene, respectively. Our results provide new insights in watermelon miRNAome and could propose new strategies for generating resistant watermelon to ZYMV.