Characterization of Italian resistance-breaking isolates of tomato spotted wilt virus reveals double resistance-breaking strains in tomato and pepper and new insights into the occurrence of the recently reported D122G substitution in the NSm protein

Tomato spotted wilt virus (TSWV) is a generalist plant virus that infects over 1000 plant species, causing significant damage to cultivated crops, including tomato and pepper, two of the most widely grown crops globally. Strategies for TSWV control in these hosts rely on different approaches, including the availability of resistance genes (Sw-5 in tomato, Tsw in pepper), encoding intracellular nucleotide-binding leucine-rich repeat (NLR) receptors that interact with TSWV NSm (in tomato) or NSs (in pepper) protein. However, the emergence of natural resistance-breaking (RB) isolates of TSWV has been reported worldwide for both crops and is currently of serious concern in the global scenario. The determinants of RB phenotypes have been well-documented, particularly for tomato, where single amino acid substitutions in the N-terminal region of the NSm protein are sufficient to induce RB. Through biological characterization and high-throughput sequencing of various Italian TSWV isolates collected in field conditions, we identified and characterized, for the first time, TSWV isolates capable of overcoming resistance in both tomato and pepper (referred to as double resistance-breaking, D-RB). Additionally, our investigation evidenced the occurrence in the field of: (i) a tomato RB isolate coding for RB-inducing NSm variants with distinct substitutions and (ii) tomato RB isolates coding for a NSm protein carrying a substitution (D > G) so far not revealed in Italian agricultural system.