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In this study, we completed the whole genome sequence of a new tobamovirus isolated from tomato plants grown in greenhouses in Jordan during the spring of 2015. The 6393-nt single-stranded RNA (ssRNA) genome encodes four proteins, as do other tobamoviruses: two replication-related proteins of 126 kDa and 183 kDa, a 30-kDa movement protein (MP) and a 17.5-kDa coat protein (CP). Phylogenetic analysis showed that this virus does not group with either the tomato mosaic virus (ToMV) or the tobacco mosaic virus (TMV) clades. Instead, it stems from a branch leading to the TMV clade. Analysis of possible recombination events between this virus and representative isolates of closely related tomato-infecting tobamoviruses showed that at least one region originated by recombination. We provide evidence that we have identified a new tobamovirus, for which we propose the name “tomato brown rugose fruit virus”.

This study was conducted to evaluate the applicability of crAssphage, pepper mild mottle virus (PMMoV), and tobacco mosaic virus (TMV) as indicators of the reduction of human enteric viruses during wastewater treatment. Thirty-nine samples were collected from three steps at a wastewater treatment plant (raw sewage, secondary-treated sewage, and final effluent) monthly for a 13-month period. In addition to the three indicator viruses, eight human enteric viruses [human adenoviruses, JC and BK polyomaviruses, Aichi virus 1 (AiV-1), enteroviruses, and noroviruses of genogroups I, II, and IV] were tested by quantitative PCR. Indicator viruses were consistently detected in the tested samples, except for a few final effluents for crAssphage and TMV. The mean concentrations of crAssphage were significantly higher than those of most tested viruses. The concentrations of crAssphage in raw sewage were positively correlated with the concentrations of all tested human enteric viruses ( p <0.05), suggesting the applicability of crAssphage as a suitable indicator to estimate the concentrations of human enteric viruses in raw sewage. The reduction ratios of AiV-1 (1.8 ± 0.7 log 10 ) were the lowest among the tested viruses, followed by TMV (2.0 ± 0.3 log 10 ) and PMMoV (2.0 ± 0.4 log 10 ). Our findings suggested that the use of not only AiV-1 and PMMoV but also TMV as indicators of reductions in viral levels can be applicable during wastewater treatment.

The size of viral genomes is limited, thus the majority of encoded proteins possess multiple functions. The main function of tobamoviral movement protein (MP) is to perform plasmodesmata gating and mediate intercellular transport of the viral RNA. MP is a remarkable example of a protein that, in addition to the initially discovered and most obvious function, carries out numerous activities that are important both for the manifestation of its key function and for successful and productive infection in general. Briefly, MP binds the viral genome, delivers it to the plasmodesmata (PD) and mediates its intercellular transfer. To implement the transport function, MP interacts with diverse cellular factors. Each of these cellular proteins has its own function, which could be different under normal conditions and upon viral infection. Here, we summarize the data available at present on the plethora of cellular factors that were identified as tobamoviral MP partners and analyze the role of these interactions in infection development.

The bumblebee Bombus terrestris is a beneficial pollinator extensively used in tomato production. Our hypothesis was that bumblebee hives collected from a Tomato brown rugose fruit virus (ToBRFV) infected tomato greenhouse, preserve an infectious primary inoculum. Placing a bumblebee hive collected from a ToBRFV contaminated greenhouse, in a glass-/net-house containing only uninfected healthy tomato plants, spread ToBRFV disease. Control uninfected tomato plants grown in a glass-/net-house devoid of any beehive remained uninfected. ToBRFV-contaminated hives carried infectious viral particles as demonstrated in a biological assay on laboratory test plants of virus extracted from hive components. Viral particles isolated from a contaminated hive had a typical tobamovirus morphology observed in transmission electron microscopy. Assembly of ToBRFV genome was achieved by next generation sequencing analysis of RNA adhering to the bumblebee body. Bumblebee dissection showed that ToBRFV was mostly present in the abdomen suggesting viral disease spread via buzz pollination. These results demonstrate that bumblebee hives collected from ToBRFV-contaminated greenhouses carry a primary inoculum that reflects the status of viruses in the growing area. This new mode of ToBRFV spread by pollinators opens an avenue for detection of viruses in a growing area through analysis of the pollinators, as well as emphasizes the need to reevaluate the appropriate disease management protocols.

Monitoring of environmental water is crucial to protecting humans and animals from possible health risks. Although numerous human-specific viral markers have been designed to track the presence of human fecal contamination in water, they lack adequate sensitivity and specificity in different geographical regions. We evaluated the performances of six human-specific viral markers [Aichi virus 1 (AiV-1), human adenoviruses (HAdVs), BK and JC polyomaviruses (BKPyVs and JCPyVs), pepper mild mottle virus (PMMoV), and crAssphage] using 122 fecal-source samples collected from humans and five animal hosts in the Kathmandu Valley, Nepal. PMMoV and crAssphage showed high sensitivity (90–100%) with concentrations of 4.5–9.1 and 6.2–7.0 log10 copies/g wet feces (n = 10), respectively, whereas BKPyVs, JCPyVs, HAdVs, and AiV-1 showed poor performances with sensitivities of 30–40%. PMMoV and crAssphage were detected in 40–100% and 8–90%, respectively, of all types of animal fecal sources and showed no significantly different concentrations among most of the fecal sources (Kruskal–Wallis test, P > 0.05), suggesting their applicability as general fecal pollution markers. Furthermore, a total of 115 environmental water samples were tested for PMMoV and crAssphage to identify fecal pollution. PMMoV and crAssphage were successfully detected in 62% (71/115) and 73% (84/115) of water samples, respectively. The greater abundance and higher mean concentration of crAssphage (4.1 ± 0.9 log10 copies/L) compared with PMMoV (3.3 ± 1.4 log10 copies/L) indicated greater chance of detection of crAssphage in water samples, suggesting that crAssphage could be preferred to PMMoV as a marker of fecal pollution.

The genus Tobamovirus belongs to the family Virgaviridae, and the genome consists of monopartite, positive, single-strand RNA. Most species contain four open reading frames encoding four essential proteins. Transmission occurs primarily through mechanical contact between plants, and in some cases, via seed dispersal. Tobamovirus fructirugosum (tomato brown rugose fruit virus, ToBRFV), the most recently described species in the genus, was first reported in 2015. It overcame genetic resistance that had been effective in tomato for sixty years, causing devastating losses in tomato production worldwide, and highlights the importance of understanding Tobamovirus genomic variation and evolution. In this study, we measured and characterized nucleotide variation for the entire genome and for all species in the genus Tobamovirus. Additionally, we measured the selection pressure acting on each open reading frame. Results showed that low nucleotide diversity and negative selection pressure are general features of tobamoviruses, with values that are approximately the same across open reading frames and without hypervariable areas. A comparison of nucleotide diversity between T. fructirugosum and its close relatives, T. tomatotessellati (tomato mosaic virus, ToMV) and T. tabaci (tobacco mosaic virus, TMV), showed low nucleotide diversity in the movement protein region harboring the resistance-breaking mutation. Furthermore, phylogenetic and diversity analyses showed that T. fructirugosum continues to evolve, and geographical distribution and host influence genomic diversity.

Tomato brown rugose fruit virus (ToBRFV) is an emerging and rapidly spreading RNA virus that infects tomato and pepper, with tomato as the primary host. The virus causes severe crop losses and threatens tomato production worldwide. ToBRFV was discovered in greenhouse tomato plants grown in Jordan in spring 2015 and its first outbreak was traced back to 2014 in Israel. To date, the virus has been reported in at least 35 countries across four continents in the world. ToBRFV is transmitted mainly via contaminated seeds and mechanical contact (such as through standard horticultural practices). Given the global nature of the seed production and distribution chain, and ToBRFV's seed transmissibility, the extent of its spread is probably more severe than has been disclosed. ToBRFV can break down genetic resistance to tobamoviruses conferred by R genes Tm‐1, Tm‐2, and Tm‐22 in tomato and L1 and L2 alleles in pepper. Currently, no commercial ToBRFV‐resistant tomato cultivars are available. Integrated pest management‐based measures such as rotation, eradication of infected plants, disinfection of seeds, and chemical treatment of contaminated greenhouses have achieved very limited success. The generation and application of attenuated variants may be a fast and effective approach to protect greenhouse tomato against ToBRFV. Long‐term sustainable control will rely on the development of novel genetic resistance and resistant cultivars, which represents the most effective and environment‐friendly strategy for pathogen control. Taxonomy Tomato brown rugose fruit virus belongs to the genus Tobamovirus, in the family Virgaviridae. The genus also includes several economically important viruses such as Tobacco mosaic virus and Tomato mosaic virus. Genome and virion The ToBRFV genome is a single‐stranded, positive‐sense RNA of approximately 6.4 kb, encoding four open reading frames. The viral genomic RNA is encapsidated into virions that are rod‐shaped and about 300 nm long and 18 nm in diameter. Tobamovirus virions are considered extremely stable and can survive in plant debris or on seed surfaces for long periods of time. Disease symptoms Leaves, particularly young leaves, of tomato plants infected by ToBRFV exhibit mild to severe mosaic symptoms with dark green bulges, narrowness, and deformation. The peduncles and calyces often become necrotic and fail to produce fruit. Yellow blotches, brown or black spots, and rugose wrinkles appear on tomato fruits. In pepper plants, ToBRFV infection results in puckering and yellow mottling on leaves with stunted growth of young seedlings and small yellow to brown rugose dots and necrotic blotches on fruits. This pathogen profile summarizes current knowledge about ToBRFV, highlights recent research progress, discusses future research directions, and proposes short‐run and long‐term control strategies.

Tomato mosaic virus (ToMV) is one of the economically damageable Tobamovirus infecting the tomato in Egypt that has caused significant losses. It is therefore of great interest to trigger systemic resistance to ToMV. In this endeavor, we aimed to explore the capacity of ZnO-NPs (zinc oxide nanoparticles) to trigger tomato plant resistance against ToMV. Effects of ZnO-NPs on tomato (Solanum lycopersicum L.) growth indices and antioxidant defense system activity under ToMV stress were investigated. Noticeably that treatment with ZnO-NPs showed remarkably increased growth indices, photosynthetic attributes, and enzymatic and non-enzymatic antioxidants compared to the challenge control. Interestingly, oxidative damage caused by ToMV was reduced by reducing malondialdehyde, H2O2, and O2 levels. Overall, ZnO-NPs offer a safe and economic antiviral agent against ToMV.

Plant viruses can evolve towards new pathogenic entities that may eventually cause outbreaks and become epidemics or even pandemics. Seven years ago, tomato brown rugose fruit virus (ToBRFV) emerged, overcoming the genetic resistance that had been employed for more than sixty years against tobamoviruses in tomato. Since then, ToBRFV has spread worldwide, producing significant losses in tomato crops. While new resistances are deployed, the only means of control is the implementation of effective prevention and eradication strategies. For this purpose, in this work, we have designed, assessed, and compared an array of tests for the specific and sensitive detection of the ToBRFV in leaf samples. First, two monoclonal antibodies were generated against a singular peptide of the ToBRFV coat protein; antibodies were utilized to devise a double-antibody-sandwich enzyme-linked immunosorbent assay (DAS-ELISA) test that sensitively detects this virus and has no cross-reactivity with other related tobamoviruses. Second, a real-time quantitative PCR (RT-qPCR) test targeting the RNA-dependent replicase open reading frame (ORF) was designed, and its performance and specificity validated in comparison with the CaTa28 and CSP1325 tests recommended by plant protection authorities in Europe. Third, in line with the tendency to use field-deployable diagnostic techniques, we developed and tested two sets of loop-mediated isothermal amplification (LAMP) primers to double-check the detection of the movement protein ORF of ToBRFV, and one set that works as an internal control. Finally, we compared all of these methods by employing a collection of samples with different ToBRFV loads to evaluate the overall performance of each test.

Plant viruses drive widespread crop epidemics, yet the host plant responses across different cell types, particularly how these responses are influenced by cultivars with varying genetic backgrounds, including the presence of resistance (R) genes, remain poorly understood. Using tomato brown rugose fruit virus (ToBRFV) and two tomato cultivars, ‘Jinpeng No. 1’ (JP) and ‘Rutgers’ (RG), with different genetic backgrounds, this study used single‐cell RNA sequencing to explore infection dynamics and responses at the cellular level. Results showed that ToBRFV accumulated to different levels in the two cultivars, likely due to differences in their genetic backgrounds, particularly the distinct genotypes of the Tm‐22 and tm‐2 alleles. Following infection, the composition of cell types in tomato leaves also varied between the two cultivars. While the entry or movement of ToBRFV in the JP cultivar was not fully prevented early on, the viral accumulation in certain cell types of this cultivar was restricted. ToBRFV alters signalling pathways based on cell type and cultivars. Pseudotime analysis revealed that, in JP plants, ToBRFV reverses expression of brassinosteroid (BR) positive regulators during mesophyll cell development. Silencing positive BR regulators increased infection in JP plants, while suppressing it in RG plants, linking BR signalling to JP‐dependent resistance. Exogenous BR suppressed ToBRFV in JP but enhanced it in RG plants. This study reveals the differential involvement of BR signalling during viral infection in the two cultivars, offering a framework for future studies of plant‐virus interactions.