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The pepino (Solanum muricatum) is an Andean vegetable crop closely related to tomato. In the last decades, it has been introduced in the Mediterranean region and other parts of the world as a potential new crop. However, several tomato major pathogens may threaten the expansion of pepino cultivation. We identified Fusarium oxysporum f. sp. lycopersici (FOL), Verticillium dahliae (VE), pepino mosaic virus (PepMV) and tomato mosaic virus (ToMV) as four of the most likely pathogens to cause damage to pepino crops in Mediterranean climates. In order to evaluate the response of the pepino genepool against these pathogens, as well as to identify sources of tolerance, we inoculated six accessions of cultivated pepino, nine accessions of seven pepino wild relatives, and one interspecific hybrid with FOL, VE, PepMV and ToMV and followed its symptomatology for 30 days (FOL and VE) or 60 days (PepMV and ToMV). ELISA tests were also performed for PepMV and ToMV. Susceptible tomato materials were used as controls. The pepino genepool displayed fewer symptoms than susceptible tomato controls after inoculation with FOL, with most accessions being tolerant or resistant. Regarding VE, a wide variation of values for the symptoms index (SI) was observed, with three cultivated pepino accessions displaying tolerance. For PepMV a wide variation for SI was also observed, with one accession of S. caripense being resistant, and several accessions of pepino and other wild relatives displaying different degrees of tolerance. PepMV absorbance values obtained by ELISA tests followed a pattern similar to that of SI. For ToMV no resistances were found, although two wild accessions and the interspecific hybrid displayed low values for the SI and were considered as moderately tolerant. ELISA tests against ToMV revealed that the virus replicated well in all materials. None of the accessions evaluated displayed resistance or high levels of tolerance to the four pathogens, but some of them were complementary for resistance or high levels of tolerance. Although the interspecific hybrid tested was not resistant to any of the pathogens, it was tolerant to FOL and PepMV and moderately tolerant to VE and ToMV. A multivariate hierarchical clustering revealed similar patterns among accessions in the response to the two fungal diseases (FOL and VE) on one side and to the two viral ones (PepMV and ToMV) on the other. The information generated in this study has allowed identifying materials within the pepino genepool for the development of multi‐resistant pepino cultivars to major diseases threatening its expansion in the Mediterranean region. [ABSTRACT FROM AUTHOR]

Keywords: TVBMV; ChiVMV; Solanum muricatum; RT-PCR; China EN TVBMV ChiVMV Solanum muricatum RT-PCR China 1147 1148 2 08/23/22 20220801 NES 220801 Supplementary information The online version contains supplementary material available at https://doi.org/10.1007/s42161-022-01120-3. Leaf samples from 15 symptomatic pepino fruit plants were analyzed for the presence of cucumber mosaic virus (CMV), potato virus Y (PVY), tobacco vein banding mosaic virus (TVBMV), chilli veinal mottle virus (ChiVMV), tobacco mosaic virus (TMV) and tomato mosaic virus (ToMV) by ELISA using commercial kits (Nano diagnostics, USA). [Extracted from the article]

Waterborne and seedborne Pepino mosaic virus (Pep MV) and Potato spindle tuber viroid ( PSTVd) pose serious threats to tomato production due to seed transmission and mechanical transmission, coupled with their long-term stability outside the host plant. Therefore, rapid and sensitive diagnostic procedures are needed to prevent the spread of these quarantine pathogens. In particular, water and seed contamination are difficult to detect and confirm without efficient concentration methods. This study presents procedures that improve detection of PSTVd from tomato seeds and leaf tissue, and Pep MV from water and tomato leaf tissue. For efficient concentration of Pep MV from water samples, a procedure was optimized using convective interaction media monolithic chromatography columns, which provides concentration by three orders of magnitude. For concentration of PSTVd from seed extracts, an easy-to-use and efficient method was developed based on RNA binding to positively charged anion-exchange resin beads that provides up to 100-fold more sensitive detection in comparison with procedures without a concentration step. This thus allows confirmation of RT- qPCR results with sequencing of RT- PCR products in samples with low viroid levels. In addition, reverse-transcription loop-mediated isothermal amplification assays for detection of PSTVd and Pep MV were optimized and adapted to both laboratory and on-site testing requirements. This allows rapid detection of these pathogens in crude leaf homogenates, in under 30 min. These procedures of concentration and detection are shown to be efficient and to fill the gaps in diagnostics of Pep MV and PSTVd, especially when these pathogens are present at low levels in difficult matrices such as water and seeds. [ABSTRACT FROM AUTHOR]

The viral pathogen Pepino mosaic virus (PepMV) can cause serious problems in the crop tomato ( Solanum lycopsericum ). One strategy to control infections of this virus is the application of hygienic measures. Alternatively, tomato plants can be cross-protected by attenuated isolates of a PepMV genotype. After two weeks, these plants are cross-protected against virulent isolates of the same genotype. In this study, the effectivity of a multi-genotype cross-protection was assessed. In greenhouse experiments young tomato plants were inoculated with a mixture of attenuated isolates of the Peruvian (LP) and Chili-2 (CH2) genotype, named VX1 and VC1, respectively. Inoculated plants were challenged with virulent isolates of the EU and CH2 genotype. The mixture of attenuated isolates, named V10, prevented almost completely the appearance of viral symptoms like leaf and stem necrosis, stunting, loss of fruit quality and loss of yield, caused by the virulent isolates. In contrast, when only a single genotype (CH2) was used to cross-protect the plants, symptoms were not prevented after the challenge with the mixture of virulent isolates. This study offers perspectives for a large-scale application of a multi-genotype cross-protection against PepMV in tomato. [ABSTRACT FROM AUTHOR]

The potato Rx gene provides resistance against Pepino mosaic virus (PepMV) in tomato; however, recent work has suggested that the resistance conferred may not be durable. Resistance breaking can probably be attributed to multiple mutations observed to accumulate in the capsid protein (CP) region of resistancebreaking isolates, but this has not been confirmed through directed manipulation of an infectious PepMV clone. The present work describes the introduction of two specific mutations, A-T78 and A-T114, into the coat protein minimal elicitor region of an Rx-controlled PepMV isolate of the EU genotype. Enzyme-linked immunosorbent assay (ELISA) and phenotypic evaluation were conducted in three Rx-expressing and wild-type solanaceous hosts: Nicotiana benthamiana, Nicotiana tabacum and Solanum lycopersicum. Mutation A-T78 alone was sufficient to confer Rx-breaking activity in N. benthamiana and S. lycopersicum, whereas mutation A-T114 was found to be associated, in most cases, with a secondary A-D100 mutation to break Rx-mediated resistance in S. lycopersicum. These results suggest that the need for a second, fitness-restoring mutation may be dependent on the PepMV mutant under consideration. Both mutations conferred Rx breaking in S. lycopersicum, whereas neither conferred Rx breaking in N. tabacum and only A-T78 allowed Rx breaking in N. benthamiana, suggesting that Rx may function in a different manner depending on the genetic background in which it is present. [ABSTRACT FROM AUTHOR]

This paper presents studies on an ultrastructural analysis of plant tissue infected with different pathotypes of Pepino mosaic virus (PepMV) and the immunolocalization of viral coat proteins. Because the PepMV virus replicates with a high mutation rate and exhibits significant genetic diversity, therefore, isolates of PepMV display a wide range of symptoms on infected plants. In this work, tomato plants of the Beta Lux cultivar were inoculated mechanically with three pathotypes representing the Chilean 2 (CH2) genotype: mild (PepMV-P22), necrotic (PepMV-P19) and yellowing (PepMV-P5-IY). The presence of viral particles in all infected plants in the different compartments of various cell types (i.e. spongy and palisade mesophyll, sieve elements and xylem vessels) was revealed via ultrastructural analyses. For the first time, it was possible to demonstrate the presence of crystalline inclusions, composed of virus-like particles. In the later stage of PepMV infection (14 dpi) various pathotype-dependent changes in the structure of the individual organelles (i.e. mitochondria, chloroplasts) were found. The strongest immunogold labeling of the viral coat proteins was also observed in plants infected by necrotic isolates. A large number of viral coat proteins were marked in the plant conductive elements, both xylem and phloem. [ABSTRACT FROM AUTHOR]

Mild variants of many viruses are able to protect infected plants from subsequent invasion by more severe variants of the same viruses through a process known as cross-protection. In the past, the cross-protective viral variants were commonly derived from mild field isolates that were sometimes genetically heterogeneous, providing variable levels of cross-protection. Here, we report a novel approach to rapidly generate cross-protective variants of the tomato-infecting Pepino mosaic virus (PepMV) independently of the availability of mild field isolates. Our approach sought to attenuate PepMV by mutating less conserved amino acid residues of the abundantly produced capsid protein (CP). These less-conserved amino acid residues were identified through multiple alignments of CPs of six potexviruses including PepMV, and were altered systematically to yield six PepMV mutants. These mutants were subsequently inoculated onto the model plant Nicotiana benthamiana, as well as tomato, to evaluate their accumulation levels, symptom severities, and cross-protection potentials. The mutant KD, in which the threonine (T) and alanine (A) residues at CP positions 66 and 67 were replaced with lysine (K) and aspartic acid (D), respectively, were found to accumulate to low levels in infected plants, cause very mild symptoms, and effectively protect both N. benthamiana and tomato against secondary infections by wild-type PepMV. These data suggest that our approach represents a simple, fast, and reliable way of generating attenuated viral variants capable of cross-protection. [ABSTRACT FROM AUTHOR]

The conclusions of the European Food Safety Authority (EFSA) following the peer review of the initial risk assessments carried out by the competent authority of the rapporteur Member State Belgium for the pesticide active substance Pepino mosaic virus strain CH2 isolate 1906, and the assessment of the application to include Pepino mosaic virus strain CH2 isolate 1906 in Annex IV of Regulation (EC) No 396/2005, are reported. The context of the peer review was that required by Regulation (EC) No 1107/2009 of the European Parliament and of the Council. The conclusions were reached on the basis of the evaluation of the representative use of Pepino mosaic virus strain CH2 isolate 1906 as a virus inoculation for cross protection of tomatoes under glasshouse application. The reliable endpoints concluded as being appropriate for use in regulatory risk assessment, derived from the available studies and literature in the dossier peer reviewed, are presented. Missing information identified as being required by the regulatory framework is listed. [ABSTRACT FROM AUTHOR]

In just a few short years, pepino mosaic disease has quickly become endemic in greenhouse tomatoes around the world. Although three genotypes of Pepino mosaic virus (PepMV) were identified in the United States, genetic composition of PepMV in greenhouse tomato crops in North America has not been determined. In this study, genetic variability and population structure of PepMV were evaluated through nucleotide sequence comparison and phylogenetic analysis of two genomic regions (helicase domain and TGB2-3) derived from 91 cDNA clones that were derived from 31 field-collected samples. These samples were collected from several major greenhouse tomato facilities in five states in the United States and two provinces in Canada. All four major genotypes of PepMV (EU, US1, US2, and CH2) were found in North America. Three distinct genotypes (EU, US1, and US2) were found in mixed infection in samples collected from Arizona and Colorado, two genotypes (EU and CH2) in Texas, and a single genotype (EU) in Alabama and California and the provinces of British Columbia and Ontario in Canada. The complexity of population genetics of PepMV in the United States poses an additional challenge to the greenhouse tomato industry because a tomato cultivar with durable resistance to multiple genotypes of PepMV may be harder to develop. [ABSTRACT FROM AUTHOR]