During 2019, tomato fruits showing viral-like symptoms of marbled yellow spots were abundant in Israel. The new symptoms were distinctive from those typical of tomato brown rugose fruit virus (ToBRFV) infection but resembled symptoms of pepino mosaic virus (PepMV) infection. RT-PCR analysis and the serological tests (enzyme linked immunosorbent assay, western blot and in situ immunofluorescence) revealed and confirmed the presence of both the tobamovirus ToBRFV and the potexvirus PepMV in the symptomatic fruits. A mixture of rod-like and filamentous particles, characteristic of viruses belonging to tobamovirus and potexvirus genera, was visualized by transmission electron microscopy of the tomato fruit viral extract. Sanger sequencing of amplified PepMV-coat protein gene segments showed ~98% sequence identity to the Chilean (CH2)-strain. In a biological assay testing the contribution of traded infected tomatoes to the establishment of tomato plant disease, we applied direct and indirect inoculation modes using Tm-2²-resistant tomato plants. The results, assessed by disease symptom development along with serological and molecular analyses, showed that the ToBRFV and PepMV co-infected fruits were an effective inoculum source for disease spread only when fruits were damaged. Importantly, intact fruits did not spread the viral disease. These results added a new factor to disease epidemiology of these viruses.

The tobamovirus tomato brown rugose fruit virus (ToBRFV), a major threat to tomato production worldwide, has recently been documented in mixed infections with the potexvirus pepino mosaic virus (PepMV) CH2 strain in traded tomatoes in Israel. A study of greenhouse tomato plants in Israel revealed severe new viral disease symptoms including open unripe fruits and yellow patched leaves. PepMV was only detected in mixed infections with ToBRFV in all 104 tested sites, using serological and molecular analyses. Six PepMV isolates were identified, all had predicted amino acids characteristic of CH2 mild strains excluding an isoleucine at amino acid position 995 of the replicase. High-throughput sequencing of viral RNA extracted from four selected symptomatic plants showed solely the ToBRFV and PepMV, with total aligned read ratios of 40.61% and 11.73%, respectively, indicating prevalence of the viruses. Analyses of interactions between the co-infecting viruses by sequential and mixed viral inoculations of tomato plants, at various temperatures, showed a prominent increase in PepMV titers in ToBRFV pre-inoculated plants and in mixed-infected plants at 18–25 °C, compared to PepMV-single inoculations, as analyzed by Western blot and quantitative RT-PCR tests. These results suggest that Israeli mild PepMV isolate infections, preceded by ToBRFV, could induce symptoms characteristic of PepMV aggressive strains.

The tobamovirus tomato brown rugose fruit virus (ToBRFV) infects tomato plants harboring the Tm-2² resistance allele, which corresponds with tobamoviruses’ avirulence (Avr) gene encoding the movement protein to activate a resistance-associated hypersensitive response (HR). ToBRFV has caused severe damage to tomato crops worldwide. Unlike tomato plants, pepper plants harboring the L resistance alleles, which correspond with the tobamovirus Avr gene encoding the coat protein, have shown HR manifestations upon ToBRFV infection. We have found that ToBRFV inoculation of a wide range of undefined pepper plant varieties could cause a “hypersensitive-like cell death” response, which was associated with ToBRFV transient systemic infection dissociated from disease symptom manifestations on fruits. Susceptibility of pepper plants harboring L¹, L³, or L⁴ resistance alleles to ToBRFV infection following HRs was similarly transient and dissociated from disease symptom manifestations on fruits. Interestingly, ToBRFV stable infection of a pepper cultivar not harboring the L gene was also not associated with disease symptoms on fruits, although ToBRFV was localized in the seed epidermis, parenchyma, and endothelium, which borders the endosperm, indicating that a stable infection of maternal origin of these tissues occurred. Pepper plants with systemic ToBRFV infection could constitute an inoculum source for adjacently grown tomato plants.

Tomato brown rugose fruit virus (ToBRFV) is a soil-borne virus showing a low percentage of ca. 3% soil-mediated infection when the soil contains root debris from a previous 30–50 day growth cycle of ToBRFV-infected tomato plants. We designed stringent conditions of soil-mediated ToBRFV infection by increasing the length of the pre-growth cycle to 90–120 days, adding a ToBRFV inoculum as well as truncating seedling roots, which increased seedling susceptibility to ToBRFV infection. These rigorous conditions were employed to challenge the efficiency of four innovative root-coating technologies in mitigating soil-mediated ToBRFV infection while avoiding any phytotoxic effect. We tested four different formulations, which were prepared with or without the addition of various virus disinfectants. We found that under conditions of 100% soil-mediated ToBRFV infection of uncoated positive control plants, root-coating with formulations based on methylcellulose (MC), polyvinyl alcohol (PVA), silica Pickering emulsion and super-absorbent polymer (SAP) that were prepared with the disinfectant chlorinated-trisodium phosphate (Cl-TSP) showed low percentages of soil-mediated ToBRFV infection of 0%, 4.3%, 5.5% and 0%, respectively. These formulations had no adverse effect on plant growth parameters when compared to negative control plants grown under non ToBRFV inoculation conditions.

Invasive weeds cause significant crop yield and economic losses in agriculture. The highest in-direct impact may be attributed to the role of invasive weeds as virus reservoirs especially within the commercial growing areas. The new tobamovirus tomato brown rugose fruit virus (ToBRFV), recently identified in the Middle-East, overcomes the Tm-22 resistance allele in the cultivated tomato varieties grown in greenhouses. In this study, we determined the role of invasive weed species as potential hosts for ToBRFV and pepino mosaic virus (PepMV). Out of all tested weed species, the invasive species So-lanum elaeagnifolium and S. rostratum, mechanically inoculated with ToBRFV, were positive for To-BRFV in both enzyme linked immunosorbent assay (ELISA) and RT-PCR tests. S. rostratum was also positive for PepMV. No conspicuous phenotype was observed on ToBRFV infected S. elaeagnifolium plants suggesting a hostplant associated defense response. S. rostratum plants inoculated with either ToBRFV alone or a mixture of ToBRFV and PepMV-IL, contained high ToBRFV levels. In addition, when inoculated with ToBRFV or PepMV-IL disease symptom manifestations were observed in S. rostratum plants and the symptoms were exacerbated upon mixed infections with both viruses. The distribution and abundance of both Solanaceae species increase the risks of virus transmission between species.