Identification and characterization of Pepino mosaic potexvirus in tomato

At the beginning of 1999, a new virus disease occurred in protected tomato crops in The Netherlands. Initial diagnostic tests revealed the presence of a potexvirus but serological tests ruled out the presence of Potato X potexvirus (PVX). Tests for other potexviruses reported from solanaceous crops provisionally identified the virus as Pepino mosaic potexvirus (PepMV). The virus was purified, and an antiserum was produced, which showed strong reactions with both the type isolate of PepMV from pepino and two other isolates from tomato. Host range and symptomatology of the pepino and tomato isolates of PepMV revealed clear differences from PVX. However, differences were also observed between the pepino and tomato isolates of PepMV. Sequence alignment of DNA fragments of 584 bp derived from the RNA polymerase cistron showed almost 95% identity with the pepino isolate, whereas the identity with PVX appeared to be < 60%. Together, these results identified PepMV as the causal agent of the new virus disease in tomato. Based on the differences from the type isolate from pepino ( Solanum muricatum ), the isolates from tomato should be considered as a distinct strain of PepMV for which the name tomato strain is proposed.     

The use of attenuated isolates of <Emphasis Type="Italic">Pepino mosaic virus</Emphasis> for cross-protection

Pepino mosaic virus (PepMV) has recently emerged as a highly infectious viral pathogen in tomato crops. Greenhouse trials were conducted under conditions similar to commercial tomato production. These trials examined whether tomato plants can be protected against PepMV by a preceding infection with an attenuated isolate of this virus. Two potential attenuated isolates that displayed mild leaf symptoms were selected from field isolates. Two PepMV isolates that displayed severe leaf symptoms were also selected from field isolates to challenge the attenuated isolates. The isolates with aggressive symptoms were found to reduce bulk yields by 8 and 24% in single infections, respectively. Yield losses were reduced to a 0–3% loss in plants that were treated with either one of the attenuated isolates, while no effects were observed on the quality of the fruits. After the challenge infection, virus accumulation levels and symptom severity of the isolates with aggressive symptoms were also reduced by cross-protection. Infection with the attenuated isolates alone did neither affect bulk yield, nor quality of the harvested tomato fruits.     

Genetic variation and evolutionary analysis of Pepino mosaic virus in Sicily: insights into the dispersion and epidemiology

Pepino mosaic virus (PepMV) is a highly infectious potexvirus that causes a severe disease in tomato (Solanum lycopersicum) crops worldwide. In Sicily, the first outbreak was detected in a single greenhouse in 2005 and it was promptly eradicated. However, in 2008, a large number of greenhouses were simultaneously affected, and it was impossible to eradicate or control the virus. This study addressed the dispersion and the genetic diversity of PepMV isolates obtained from the outbreak in Sicily, in comparison with worldwide PepMV isolates, to gain insight into the factors determining the evolution and epidemiology of the virus. A total of 1800 samples from plants with and without symptoms were collected in the Sicilian provinces of Agrigento, Caltanissetta, Palermo, Ragusa, Siracusa and Trapani. Three isolates collected at different times were biologically characterized. The incidence of the virus increased rapidly from 13% in 2011 to 63% in 2013, and phylogenetic analysis showed that all Sicilian isolates of PepMV belonged to the CH2 strain, one of the six strains previously described. Nucleotide diversity of the Sicilian isolates was low, thus suggesting rapid spread and genetic stability.     

Susceptibility of different plant species and tomato cultivars to two isolates of <Emphasis Type="Italic">Pepino mosaic virus</Emphasis>

As Pepino mosaic virus has become a pathogen of major importance in worldwide tomato production, information is needed on possible differences between the sensitivity of cultivars towards infection. Furthermore, it is important what hosts other than Solanaceae may be virus reservoirs and are, therefore, threats for tomato cultivation. Two PepMV isolates (PepMV-Sav, E397, a European tomato isolate and PV-0554, a Peruvian pepino isolate) differing in their origin and virulence were used for several experiments to investigate these issues. The response to mechanical inoculation with PepMV was studied using 25 tomato cultivars, seven indicator plant species, and nine other possible horticultural host plants. Symptom development after infection with PepMV was monitored and the virus was detected by DAS-ELISA and IC-RT-PCR. Garlic and broad bean were shown to be additional hosts of PepMV depending on the virus isolate. Nicotiana benthamiana seems to be the most sensitive indicator among all tested indicator plants developing symptoms. Both PepMV isolates infected all tested tomato cultivars. Development of disease symptoms depended on the cultivar and the virus isolate but symptoms were not visible in all cases. None of the cultivars showed tolerance against the two isolates but two responded with a lower susceptibility at an absorbance level of 0.2 (healthy control 0.09). It was observed that some cultivars grown hydroponically showed also lower losses in biomass and yield. Data indicated a correlation between absorbance level in DAS-ELISA and reduction in total tomato growth.     

Ultrastructural aspects of tomato leaves infected by Tomato torrado virus (ToTV) and co‐infected by other viruses

Optical and electron microscopy studies were carried out to investigate the cytopathology induced in tomato leaves infected by Tomato torrado virus (ToTV), a new picorna‐like virus associated with the ‘Torrado’ disease. Infected leaves, showing typical Torrado disease symptoms were surveyed in commercial greenhouses in the main tomato production areas of Spain. The effect of the co‐infection of ToTV with other viruses which commonly infect tomato crops was also studied. Ultra‐thin sections of ToTV‐infected tomato leaves did not show a strong cellular alteration. However, crystalline arrays of isometric virus‐like particles (VLPs) of 20–30 nm in the inclusion bodies were observed in phloem parenchyma cells of the infected tissues. Tissues co‐infected by ToTV and either Tomato chlorosis virus (ToCV) or Pepino mosaic virus (PepMV) presented more severe cellular alterations. The most deleterious consequences for tomato cells were found in triple infections of ToTV, PepMV and Tomato spotted wilt virus (TSWV), where characteristic cell wall overgrowth was distinguishable, together with a large amount of necrotic cells.     

Genetic characterization of <Emphasis Type="Italic">Pepino mosaic virus</Emphasis> isolates from Belgian greenhouse tomatoes reveals genetic recombination

Over a period of a few years, Pepino mosaic virus (PepMV) has become one of the most important viral diseases in tomato production worldwide. Infection by PepMV can cause a broad range of symptoms on tomato plants, often leading to significant financial losses. At present, five PepMV genotypes (EU, LP, CH2, US1 and US2) have been described, three of which (EU, LP and US2) have been reported in Europe. Thus far, no correlation has been found between different PepMV genotypes and the symptoms expressed in infected plants. In this paper, the genetic diversity of the PepMV population in Belgian greenhouses is studied and related to symptom development in tomato crops. A novel assay based on restriction fragment length polymorphism (RFLP) was developed to discriminate between the different PepMV genotypes. Both RFLP and sequence analysis revealed the occurrence of two genotypes, the EU genotype and the CH2 genotype, within tomato production in Belgium. Whereas no differences were observed in symptom expression between plants infected by one of the two genotypes, co-infection with both genotypes resulted in more severe PepMV symptoms. Furthermore, our study revealed that PepMV recombinants frequently occur in mixed infections under natural conditions. This may possibly result in the generation of viral variants with increased aggressiveness.     

Characterization of weeds and rotational crops as alternative hosts of Spongospora subterranea,the causal agent of powdery scab in Israel

Alternative hosts of Spongospora subterranea may allow multiplication and survival of the pathogen over time; thus, host range is important from an epidemiological aspect. Weeds and rotational crops, such as wheat and barley, were sampled from potato fields with a history of powdery scab (PS) and examined for the presence of S. subterranea by root staining followed by microscopic observations and by qPCR analysis after DNA extraction. The pathogen was detected in plants of 16 weed species from eight families and in volunteer plants of potato and wheat. The ability of the pathogen to infect weeds and rotational crops was further examined by artificial inoculations with sporosori in pot experiments. Successful inoculations occurred with 13 weed species from eight families and with 12 rotational crops from five families. The findings of this study indicate a wide host range in Israel; the families Malvaceae and Zygophyllaceae and the following species are reported for the first time as S. subterranea hosts: Solanum elaeagnifolium, Triticum aestivum, Cynodon dactylon, Phalaris paradoxa, Phalaris minor, Setaria verticillata, Rostaria cristata, Sinapis nigra, Arachis hypogaea, Medicago sativa, Astragalus hauraensis, Amaranthus albus, Chenopodium murale, Chenopodium opulifolium, Salsola soda, Malva nicaeensis, Chrysanthemum segetum, Verbesina encelioides, Ammi majus and Tribulus terrestris. Controlling weeds and avoiding the relevant rotational crops observed to be S. subterranea-positive and thus potential hosts, should be taken into consideration in the management of PS, to reduce pathogen inoculum build-up.     

Effects of alternative winter cover cropping systems on weed suppression in organically grown tomato (<Emphasis Type="Italic">Solanum lycopersicum</Emphasis>)

Weed control is a major concern for organic farmers around the world and non-chemical weed control methods are now the subject of many investigations. Field studies were conducted in tomato (Solanum lycopersicum L.) from 2004 to 2006 at the Black Sea Agricultural Research Institute experiment field to determine the weed suppressive effects of winter cover crops. Treatments consisted of ryegrass (Lolium multiflorum L.), oat (Avena sativa L.), rye (Secale cereale L.), wheat (Triticum aestivum L.), gelemen clover (Trifolium meneghinianum Clem.), Egyptian clover (Trifolium alexandrinum L.), common vetch (Vicia sativa L.), hairy vetch (Vicia villosa Roth.) and a control with no cover crop. Treatments were arranged in a randomized complete block design with four replications. To determine the weed suppressive effects of the cover crops, weed density and total weed dry biomass were assessed at 14, 28, and 56 days after termination (DAT) of the cover crops from all plots using a 50 × 50 cm quadrat placed randomly in each plot. After cover crop kill and incorporation into soil, tomato seedlings variety ‘H2274’ were transplanted. Broadleaved weed species were the most prominent species in both years. Total weed biomass measured just prior to cover crop incorporation into the soil was significantly lower in S. cereale plots than in the others. The number of weed species was lowest at 14 DAT and later increased at 28 and 56 DAT, and subsequently remained constant during harvest. This research indicates that cover crops such as L. multiflorum, S. cereale, V. sativa and V. villosa could be used in integrated weed management programs to manage some weeds in the early growth stages of organic tomato.     

Phylodynamics of Pepino mosaic virus in Spain

Pepino mosaic virus (PepMV) is an emerging pathogen that causes severe economic losses in tomato crops in the Northern hemisphere. After its first identification, the new viral strain PepMV-CH2 has been isolated in several countries worldwide. In order to further understand the evolutionary dynamics of PepMV before and after PepMV-CH2 emergence, we analyzed a collection of PepMV isolates from southeastern Spain, estimating the rate of PepMV molecular evolution and the coalescence process for the effective number of PepMV infections using a Bayesian phylogenetic approach. Our results show that the rate of PepMV molecular evolution was 5.570?×?10^?3 substitutions/site/year, a value which is approximately an order of magnitude higher than the rates recently reported for other plant RNA viruses. Moreover, PepMV-CH2 was estimated to have originated in 2000, coincident with the onset of PepMV-CH2 infections in southeastern Spain, its population following now an expansion process. This further illustrates that genetic and ecological interactions among different viral strains can modulate the evolutionary dynamics of PepMV and determine its epidemiological profile.     

Weed size hierarchies in Denmark

In 962 field trials in different crops, the size of a number of weed species was investigated. The weeds were ranked according to their size on a single occasion. The ranks of the species were examined relative to two weed species, Viola arvensis and Matricaria inodora. The ranking was, however, independent of which of the two species were used as standard. The correlation between the rankings with Viola arvensis as standard, and the rankings with Matricaria inodora as standard were r= 0-93 for spring sown crops, and r= 0.84 for autumn sown crops. Sinapis arvensis was the largest weed species in spring sown crops, with a weight of 14.5 times that of Viola arvensis, followed by Brassica napus, Galeopsis spp., Thlaspi arvense and Amsinckia spp. Viola arvensis. Veronica spp. and Lamium spp. were the smallest weed species in spring sown crops. Galium aparine was the largest species in autumn sown crops, whereas spring germinating species such as Polygonum spp. were among the smallest. The ranks of Viola arvensis, Myosotis arvensis, Lapsana communis, Matricaria inodora and Lamium spp. were almost identical in spring sown crops and in autumn sown crops.     

Weed hosts of <Emphasis Type="Italic">Verticillium dahliae</Emphasis> in cotton fields in Turkey and characterization of <Emphasis Type="Italic">V. dahliae</Emphasis> isolates from weeds

A weed survey conducted in 2004 and 2005 in Aydin province of Turkey showed that Solanum nigrum, Xanthium strumarium, Amaranthus retroflexus, Portulaca oleracea, Sonchus oleraceus and Datura stramonium were the most prevalent weeds in the cotton fields exhibiting Verticillium wilt. Verticillium dahliae Kleb. was recovered from A. retroflexus and X. strumarium in those cotton fields. This is the first report of V. dahliae occurring naturally in A. retroflexus in Turkey. Pathogenicity tests on cotton and weeds showed that the virulence of V. dahliae isolates from weeds was higher on cotton plants than on weeds, with the disease severity ranging from 31.7% to 98.0%. Disease severity of V. dahliae isolates was 54.7–93.9% on eggplant, 23.7–51.6% on cucumber and 11.0–16.4% on tomato, whereas it did not cause any disease symptoms, or only low levels, on pepper and bell pepper. Two vegetative compatibility groups (VCGs) were identified among seven tested weed isolates: VCG2A (two isolates) and VCG2B (three isolates) using international reference strains.     

Competition between Ridolfia segetum and sunflower

Studies on competition between Ridolfia segetum Maris, and sunflower (Helianthemum annuus L.) were conducted at eight locations in southern Spain in 1990 and 1991. in order to define competition models and to estimate from these economic thresholds as affected by crop inputs and potential yields. Competition losses in sunflower crops ranged from 19% to 56% of weed–free yields. There were slightly better correlations between percentage sunflower reduction and weed density than with weed dry weight, (?0.66 and ?0.59, respectively). The weed competitive index, or sunflower crop dry weight reduction per unit dry weight of R. segetum, was 1.09. The percentage yield losses due to weed density (NPR_t) were fitted to multiple linear, quadratic, exponential and hyperbolic models. The hyperbolic equation, %RSY=100 (1+1/b*NPR_t)^?1, where b=0.14 and is the R. segetum competitive ability index, had the lowest error sum of squares (SSE), and gave the best biological explanation for the competition response. Early emergence (before mid–March) made weeds about 1.5 times more competitive than late emergence. The economic threshold to offset the cost of a shallow post–emergence tillage, assuming 70% control efficiency, ranged from about 2.5 plants m ^?2 for low–yielding crops(1200kgha^?1) to less than one plant m^?2 for higher–yielding crops (2800 kg ha^?1).     

Mapping Ridolfia segetum patches in sunflower crop using remote sensing

Ridolfia segetum is a frequent umbelliferous weed in sunflower crops in the Mediterranean basin. Field and remote sensing research was conducted in 2003 and 2004 over two naturally infested fields to determine the potential of multispectral imagery for discrimination and mapping of R. segetum patches in sunflower crops. The efficiency of the four wavebands blue (B), green (G), red (R) and near‐infrared (NIR), selected vegetation indices and the spectral angle mapper (SAM) classification method were studied using aerial photographs taken in the late vegetative (mid‐May), flowering (mid‐June) and senescence (mid‐July) crop growth stages. Discrimination efficiency of R. segetum patches in sunflower crops is consistently affected by their phenological stages, in this order: flowering > senescence > vegetative. In both fields, R. segetum patches were efficiently discriminated in mid‐June, corresponding to the flowering phase, by using the waveband G, the ratio R/B or SAM with overall accuracies ranging from 85% to 98%. The application of the median‐filtering algorithm to any of the classified images improved the accuracy. Our results suggest that mapping R. segetum weed patches in sunflower to implement site‐specific weed management techniques is feasible with aerial photography when images are taken from 8 to 10 weeks before harvesting.     

Genetic Structure of the Population of Pepino mosaic virus Infecting Tomato Crops in Spain

ABSTRACT The population structure of Pepino mosaic virus (PepMV), which has caused severe epidemics in tomato in Spain since 2000, was analyzed. Isolates were characterized by the nucleotide sequence of the triple gene block and coat protein gene and, for a subset of isolates, a part of the RNA-dependent RNA polymerase gene. The full-length sequence of the genomic RNA of a Solanum muricatum isolate from Peru also was determined. In spite of high symptom diversity, the Spanish population of PepMV mostly comprised highly similar isolates belonging to the strain reported in Europe (European tomato strain), which has been the most prevalent genotype in Spain. The Spanish PepMV population was not structured spatially or temporally. Also, isolates highly similar to those from nontomato hosts from Peru (Peruvian strain) or to isolate US2 from the United States (US2 strain) were detected at lower frequency relative to the European strain. These two strains were detected in peninsular Spain only in 2004, but the Peruvian strain has been detected in the Canary Islands since 2000. These results suggest that PepMV was introduced into Spain more than once. Isolates from the Peruvian and US2 strains always were found in mixed infections with the European tomato strain, and interstrain recombinants were detected. The presence of different strains of the virus, and of recombinant isolates, should be considered for the development of control strategies based on genetic resistance.     

Simultaneous detection and identification of <Emphasis Type="Italic">Pepino mosaic virus</Emphasis> (PepMV) isolates by multiplex one-step RT-PCR

A RT-PCR was developed for the simultaneous detection and identification of three groups of Pepino mosaic virus (PepMV): European/Peruvian, Chilean 1/US1 and Chilean 2/US2 groups, followed by a restriction analysis that allowed the separation of the European, Peruvian, Chilean 2 and US2 isolates (patent pending). The multiplex RT-PCR reaction was performed by a mix of six primers that amplified a part of the RNA-dependent RNA polymerase gene of PepMV plus an internal control. Amplifications resulted in a 980 bp, 703 bp or 549 bp PCR product for European/Peruvian, Chilean 1/US1 or Chilean 2/US2 groups, respectively. For the identification of the isolates present within the European/Peruvian and Chilean 2/US2 groups, the amplified PCR fragments were directly digested with SacI enzyme. The multiplex RT-PCR method presented higher sensitivity to detect CH1/US1 isolates in field samples than the RFLP-PCR method described by Hanssen et al. (European Journal of Plant Pathology 121:131–146, 2008). The detection limit observed with the multiplex RT-PCR was equal to or 3,125 times higher when compared to single RT-PCR or ELISA-DAS and molecular hybridisation methods, respectively. The use of the multiplex RT-PCR method in routine analysis of field tomato samples allowed the detection of 36.2 and 33.4% more positives when compared to the serological and molecular hybridisation methods, respectively, and the identification of plants infected with one, two or three isolates of PepMV.     

New sources of resistance to PepMV in tomato

Journal of Plant Diseases and Protection - Pepino mosaic virus (PepMV) disease causes important losses in tomato (Solanum lycopersicum) yield around the world. In order to find new sources of...     

Spread and interaction of Pepino mosaic virus (PepMV) and Pythium aphanidermatum in a closed nutrient solution recirculation system: effects on tomato growth and yield

Pepino mosaic virus (PepMV) was shown to be efficiently transmitted between tomato plants grown in a closed recirculating hydroponic system. PepMV was detected in all plant parts after transmission via contaminated nutrient solution using ELISA, immunocapture RT‐PCR, RT‐PCR, electron microscopy, and by inoculation to indicator plants. Detection of PepMV in nutrient solution was only possible after concentration by ultracentrifugation followed by RT‐PCR. Roots tested positive for PepMV 1–3 weeks after inoculation, and subsequently a rapid spread from the roots into the young leaves and developing fruits was found within 1 week. PepMV was only occasionally detected in the older leaves. None of the infected plants showed any symptoms on fruits, leaves or other organs. Pre‐infection of roots of tomato cv. Hildares with Pythium aphanidermatum significantly delayed PepMV root infections. When mechanically inoculated with PepMV at the 2–4 leaf stage, yield loss was observed in all plants. However, only plants of cv. Castle Rock recorded significant yield losses when infected via contaminated nutrient solution. Yield losses induced by infection with PepMV and/or P. aphanidermatum ranged from 0·4 up to 40% depending on experimental conditions.     

Molecular characterisation of <Emphasis Type="Italic">Turnip mosaic virus</Emphasis> isolates from Brassicaceae weeds

Eight provinces of Iran were surveyed during 2003–2008 to find Brassicaceae reservoir weed hosts of Turnip mosaic virus (TuMV). A total of 532 weed samples were collected from plants with virus-like symptoms. The samples were tested for the presence of TuMV by enzyme-linked immunosorbent assay using specific antibodies. Among those tested, 340 samples (64%) were found to be infected with TuMV. Rapistrum rugosum, Sisymberium loeselii, S. irio and Hirschfeldia incana were identified as the Brassicaceae weed hosts of TuMV, and the former two plant species were found to be the most important weed hosts for the virus in Iran. The full-length sequences of the genomic RNAs of IRN TRa6 and IRN SS5 isolates from R. rugosum and S. loeselii were determined. No evidence of recombination was found in both isolates using different recombination-detecting programmes. Phylogenetic analyses of the weed isolates with representative isolates from the world showed that the IRN TRa6 and IRN SS5 isolates fell into an ancestral basal-Brassica group. This study shows for the first time the wide distribution and phylogenetic relationships of TuMV from weeds in the mid-Eurasia of Iran.