Isolation of spontaneous mutants of tomato brown rugose fruit virus that efficiently infect Tm-1 homozygote tomato plants

The tobamovirus, tomato brown rugose fruit virus (ToBRFV), is a significant concern in global tomato production due to the ineffectiveness of the widely used Tm-2² resistance gene. Our previous study showed that the tomato variety GCR237, a Tm-1 homozygote, resisted an Israeli isolate of ToBRFV (DSMZ PV-1241) for up to 35 days post inoculation (dpi), suggesting Tm-1-homozygous cultivars could control ToBRFV. In the present study, we inoculated GCR237 plants with ToBRFV and cultivated them for a longer period of time. The plants resisted systemic infection up to 50 dpi, but mosaic symptoms appeared on the upper leaves by 100 dpi. We retrieved the virus from symptomatic leaves and established four single local lesion isolates. These isolates had several amino acid (AA) substitutions in the helicase domain of 126-kDa/183-kDa replication proteins, where the Tm-1 protein likely binds to inhibit viral RNA replication. Back-inoculating these isolates onto GCR237 plants confirmed they had acquired the ability to overcome GCR237’s resistance and induced mosaic symptoms as early as 14 dpi. About 90% of 229 ToBRFV isolates in the NCBI database had identical AA sequences in the corresponding region to DSMZ PV-1241, while ~ 10% inherently had AA substitutions that would confer complete breaking ability to the Tm-1 resistance. These results suggest that while Tm-1 can inhibit ToBRFV RNA replication, ToBRFV can easily overcome Tm-1 homozygotes.