First report of <i>Tomato brown rugose fruit virus</i> in tomato in France

Abstract

In February, 2020 a sample of tomato (Solanum lycopersicon cv. Torino) was submitted to Fera Science Ltd. from a grower in Finistère, France. The symptoms seen were mosaic of the leaves and curling of the plant head. The sample was tested by ELISA for Tomato brown rugose fruit virus (ToBRFV), Tomato mosaic virus (DSMZ, Germany), Alfalfa mosaic virus, Pepino mosaic virus, Potato virus Y, (Bioreba, Switzerland), Cucumber mosaic virus (Agdia, USA) and Potato virus X (Fera, UK). The sample was positive for ToBRFV but negative for the other viruses. To confirm the diagnosis, the sample was tested for ToBRFV by real-time RT-PCR, using the CaTa28 assay (International Seed Federation, 2020) and by conventional RT-PCR with tobamovirus primers F-5476/R-6287 (Levitsky et al., 2019). The sample tested positive by real time RT-PCR for ToBRFV. A product of the expected size (c. 800 bp) was obtained in the RT-PCR and confirmed as ToBRFV by nucleotide sequence comparison (GenBank Accession No. MW284988). Genome sequences for ToBRFV were also obtained by high throughput sequencing as described for historic isolates by Hammond et al. (2020). The genome of ToBRFV (MW284987) had between 99.94-99.95% nucleotide identity to genomes of ToBRFV isolates from Turkey (MT107885) and Israel (KX619418), respectively. Following notification to the NPPO in France, the French national reference laboratory (Plant Health Laboratory, ANSES) received ten symptomatic tomato samples, from cultivars Admiro (1), Goutine (3), Plaisance (2), Sweetelle (1), Torino (2) and Uriburi (1), collected in one greenhouse located in Finistère, Britanny. Symptoms consisted of leaf mosaic, narrowing (needle-like) and embossing of leaves. All samples were tested using the CaTa28 and CSP1325 real-time PCR primer sets (International Seed Federation, 2020) and were positive for ToBRFV for both targets. Two of the ten samples having a Ct below 10 were also amplified by conventional RT-PCR using generic tobamovirus primers TobamodF/TobamodR (Li et al., 2011). Products of the expected size (c. 880 bp) were obtained for the RT-PCR and were confirmed as ToBRFV by nucleotide sequence comparison. The sequence was deposited in GenBank (MZ542763) and had a nucleotide identity >99% with isolates from the UK (MN182533) and Jordan (MK648157). Brittany being a key region for tomato production, phytosanitary measures were implemented, and all plants and growing media located in the greenhouse (2.4 ha) were destroyed and the facility disinfected. The French NPPO set up an extensive survey in all parts of the country and more than 1600 samples of tomato and pepper (Capsicum spp) were collected and tested. None of the samples tested positive supporting the implemented phytosanitary measures. The tomato plants from the outbreak site were propagated in the UK from seed of Dutch origin. Trace-back investigations found no evidence of ToBRFV at the UK propagation site. This work was partially funded under the Defra-Fera Long Term Service Agreement