Identification of novel and known tobamoviruses in tomato and other solanaceous crops using a new pair of generic primers and development of a specific RT-qPCR for ToBRFV

Numerous tobamoviruses infect tomatoes and other cultivated solanaceous crops. Due to their transmission in contaminated seed and during mechanical cultivation activities, they occur frequently, often associated with considerable yield losses. While specific virus tests by ELISA or RT-PCR are available, generic methods covering a broad spectrum of virus species are preferred for screening. Here we present the development of a generic primer pair for the detection of a broad range of Tobamovirus species. The primers are degenerate at one base position and amplify a genome segment of ca. 560 bp. This allows an unambiguous assignment to a respective species upon sequencing. Isolate sequences of 14 different Tobamovirus species from the DSMZ plant virus collection were obtained to verify the spectrum of amplification. Routine virus indexing of tomato samples from Germany in 2018 revealed an amplicon for tobamoviruses that was then identified as Tomato brown rugose fruit virus (ToBRFV), previously known only from Jordan and Israel, and now detected for the first time infecting tomato in Europe. From the sequence information obtained, a specific RT-qPCR test for ToBRFV was developed. The generic RT-PCR for tobamoviruses is competent to detect known and novel tobamoviruses and is a convenient option for virus screening.