Genetic characterization of Pepino mosaic virus 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 aggressivenes

Higher plasma sclerostin and lower Wnt signaling gene expression in white adipose tissue of prediabetic South Asian men compared with white Caucasian men

Background: South Asians generally have an unfavourable metabolic phenotype compared with white Caucasians, including central obesity and insulin resistance. The Wnt protein family interacts with insulin signaling, and impaired Wnt signaling is associated with adiposity and type 2 diabetes mellitus. We aimed to investigate Wnt signaling in relation to insulin signaling in South Asians compared with white Caucasians.Methods: Ten Dutch South Asian men with prediabetes and overweight or obesity and 10 matched Dutch white Caucasians were included. Blood samples were assayed for the Wnt inhibitor sclerostin. Subcutaneous white adipose tissue (WAT) and skeletal muscle biopsies were assayed for Wnt and insulin signaling gene expression with quantitative reverse transcription polymerase chain reaction (Clinicaltrials.gov NCT02291458).Results: Plasma sclerostin was markedly higher in South Asians compared with white Caucasians (+65%, P < 0.01). Additionally, expression of multiple Wnt signaling genes and key insulin signaling genes were lower in WAT in South Asians compared with white Caucasians. Moreover, in WAT in both ethnicities, Wnt signaling gene expression strongly positively correlated with insulin signaling gene expression. In skeletal muscle, WNT10B expression in South Asians was lower, but expression of other Wnt signaling and insulin signaling genes was comparable between ethnicities. Wnt and insulin signaling gene expression also positively correlated in skeletal muscle, albeit less pronounced.Conclusion: South Asian men with overweight or obesity and prediabetes have higher plasma sclerostin and lower Wnt signaling gene expression in WAT compared with white Caucasians. We interpret that reduced Wnt signaling could contribute to impaired insulin signaling in South Asians.Diabetes mellitus: pathophysiological changes and therap

Violence and publicity: constructions of political responsibility after 9/11

Focussing upon recent political philosophical reflections on the War on Terror, this paper asks whether violence can be understood without undermining the empirical and normative potential of public action to curtail it. Explanations of political violence are often either dismissed as mere exoneration, or end up reducing all forms of power to violence. Negotiating a path between these poles requires developing a double sense of responsibility that affirms that while violence can be comprehended as a rational phenomenon this does not in principle undermine the potential of public action to democratize violence. Theoretical approaches that ontologize violence as an ineradicable feature of 'the political' are criticized for leaving little space for thinking about the possibility of legitimate public action in curtailing political violence. The public dimensions of the relationships between justification, legitimacy and political violence are considered by relating contemporary left-liberal arguments about terrorism and legitimate violence to recent feminist conceptualizations of the practice of taking responsibility

Pepino mosaic virus: an endemic pathogen of tomato crops

Owing to their large population size and short generation time, viruses generally have a huge potential to evolve and adapt under natural selection pressure. Despite tremendous efforts in human, animal and plant health management, viral diseases remain difficult to control and eradicate. Moreover, existing control strategies are compromised by the continuous emergence of new viruses. In Chapter 1 emerging viruses of tomato crops are reviewed. This includes Pepino mosaic virus (PepMV), a Potexvirus with a single stranded RNA genome, a rapidly emerging virus which has become one of the most important viral diseases in tomato production worldwide over the recent years. Infection by PepMV can cause a broad range of symptoms on tomato plants, of which especially the typical fruit marbling can lead to significant economical losses. Presently, five PepMV genotypes (EU, LP, CH2, US1 and US2) have been described worldwide, three of which (EU, LP and US2) have previously been reported to occur in Europe. As nature and severity of PepMV symptoms are highly variable, economical damage caused by PepMV is difficult to assess and the identification of factors contributing to symptom severity is warranted. In Chapter 2 the genetic diversity of the PepMV population in Belgian greenhouses is studied and related to the symptom development in tomato crops. Previously, no correlation has been found between different PepMV genotypes and the symptomatology of infected plants. A novel assay based on restriction fragment length polymorphism (RFLP) was developed to discriminate the different PepMV genotypes. Both RFLP and sequence analysis revealed the occurrence of two genotypes, the EU genotype as well as the CH2 genotype, within the tomato production in Belgium. Surprisingly, a clear dominance of the CH2 genotype in the Belgian PepMV population was found, although this genotype has previously not been found in commercial tomato production. 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 such mixed infections. So far, it remained unclear whether different PepMV isolates can cause differential symptom severity. Therefore, PepMV symptomatology of different isolates was studied in Chapter 3. Based on the survey described in Chapter 2, four isolates that differed in symptom expression in the crop of origin were selected for greenhouse trials. The selected isolates were inoculated onto tomato plants grown in separate plastic tunnels. PepMV symptom development was assessed regularly and extensive sampling followed by ELISA analyses, genotyping and nucleotide sequencing was performed to study viral presence and variation in PepMV sequences throughout the trial period. Two isolates (EU mild and CH2 mild) that were selected based on mild symptom expression in the crop of origin caused only mild symptoms in the trial, while two other isolates (CH2 aggressive and EU+CH2) that were selected for severe symptom display, caused considerably more severe symptoms. Sequence identity between the mild and the aggressive CH2 isolates was as high as 99.4%. Results of this study show that differential symptom expression can, at least partially, be attributed to the PepMV isolate, which may be related to minor differences at the nucleotide level between isolates. In Chapter 4, seed transmission of PepMV in tomato is demonstrated. Fruit was harvested from the greenhouse trials described in Chapter 3 and more than 100,000 seeds were extracted and cleaned using an enzymatic treatment without disinfection. Infection assays using indicator plants confirmed the presence of infectious virus particles on the seeds. In the framework of a European project, seeds were distributed to 10 different laboratories in three separate batches, and germinated for seedling analyses by ELISA. In total over 87,000 plants were tested, and 23 PepMV-infected plants were detected, indicating an overall transmission rate of 0.026 %. Our results clearly show that PepMV can be transmitted from contaminated seeds to seedlings, highlighting the risk of using seeds from PepMV-infected plants, and revealed the potential for seed transmission to contribute to spread of PepMV. In Chapter 5, the potential of three mild PepMV isolates, belonging to the CH2, EU and LP genotypes, to protect a tomato crop against the aggressive CH2 isolate (Chapter 3) as the challenge isolate, was assessed in greenhouse trials. After challenge infection, enhanced symptom display was recorded in plants that were pre-inoculated with a protector isolate that belonged to a different genotype (EU, LP) than the challenge isolate. A quantitative genotype-specific TaqMan assay revealed that in these plants, the accumulation of the challenge isolate only temporarily slowed down. By contrast, efficient cross-protection was obtained using the mild isolate of the CH2 genotype, and in this case the challenge isolate was barely detectable in the pre-inoculated plants. These results suggest that the interaction between PepMV isolates largely depends on RNA sequence homology and that post-transcriptional gene silencing plays an important role in cross-protection. As plant viruses are obligate intracellular parasites that hijack host cellular functions and resources for their replication and movement, they generally induce a wide variety of alterations in host gene expression and cell physiology. In Chapter 6, we used a custom-designed Affymetrix tomato GeneChip array that contains probe sets to interrogate over 22,000 tomato transcripts to study transcriptional changes in response to inoculation with the highly similar (99.4% nucleotide sequence identity) mild and aggressive CH2 isolates that are characterized in Chapter 3. Interestingly, our results show that both isolates induce differential transcriptomic responses in the tomato host despite accumulation to similar viral titers. PepMV inoculation resulted in an extensive transient repression of host genes which clearly affected primary metabolism. Especially the defense response intensity was higher upon inoculation with the aggressive isolate, and defense was mediated by salicylic acid signaling rather than by jasmonate signaling. Our results furthermore show that PepMV differentially regulates the RNA silencing pathway, suggesting a role for PepMV-encoded silencing suppressors, and the ubiquitination pathway. In addition, perturbation of pigment biosynthesis, as monitored by differential regulation of the flavonoid/anthocyanin and lycopene biosynthesis pathways, was monitored, which can be associated with the typical PepMV-induced marbling of tomato fruit. Finally, Chapter 7, the general discussion, is a PepMV pathogen profile in which the results obtained in this work are discussed and integrated into a review on the current knowledge on this highly successful pathogen of tomato crops. <br/

Pepino mosaic virus: a successful pathogen that rapidly evolved from emerging to endemic in tomato crops

Taxonomy: Pepino mosaic virus (PepMV) belongs to the Potexvirus genus of the Flexiviridae family. Physical properties: PepMV virions are nonenveloped flexuous rods that contain a monopartite, positive-sense, single-stranded RNA genome of 6.4 kb with a 3' poly-A tail. The genome contains five major open reading frames (ORFs) encoding a 164-kDa RNA-dependent RNA polymerase (RdRp), three triple gene block proteins of 26, 14 and 9 kDa, and a 25-kDa coat protein. Genome diversity: Four PepMV genotypes, with an intergenotype RNA sequence identity ranging from 78% to 95%, can be distinguished: the original Peruvian genotype (LP); the European (tomato) genotype (EU); the American genotype US1; and the Chilean genotype CH2. Transmission: PepMV is very efficiently transmitted mechanically, and a low seed transmission rate has been demonstrated. In addition, bumblebees have been associated with viral transmission. Host range: Similar to other Potexviruses, PepMV has a rather narrow host range that is thought to be largely restricted to species of the Solanaceae family. After originally being isolated from pepino (Solanum muricatum), PepMV has been identified in natural infections of the wild tomato species S. chilense, S. chmielewskii, S. parviflorum and S. peruvianum. PepMV is causing significant problems in the cultivation of the glasshouse tomato Solanum lycopersicum, and has been identified in weeds belonging to various plant families in the vicinity of tomato glasshouses. Symptomatology: PepMV symptoms can be very diverse. Fruit marbling is the most typical and economically devastating symptom. In addition, fruit discoloration, open fruit, nettle-heads, leaf blistering or bubbling, leaf chlorosis and yellow angular leaf spots, leaf mosaic and leaf or stem necrosis have been associated with PepMV. The severity of PepMV symptoms is thought to be dependent on environmental conditions, as well as on the properties of the viral isolate. Minor nucleotide sequence differences between isolates from the same genotype have been shown to lead to enhanced aggressiveness and symptomatology. Control: Prevention of infection through strict hygiene measures is currently the major strategy for the control of PepMV in tomato production. Cross-protection can be effective, but only under well-defined and well-controlled conditions, and the effectiveness depends strongly on the PepMV genotype

Emerging Viral Diseases of Tomato Crops

Viral diseases are an important limiting factor in many crop production systems. Because antiviral products are not available, control strategies rely on genetic resistance or hygienic measures to prevent viral diseases, or on eradication of diseased crops to control such diseases. Increasing international travel and trade of plant materials enhances the risk of introducing new viruses and their vectors into production systems. In addition, changing climate conditions can contribute to a successful spread of newly introduced viruses or their vectors and establishment of these organisms in areas that were previously unfavorable. Tomato is economically the most important vegetable crop worldwide and many viruses infecting tomato have been described, while new viral diseases keep emerging. Pepino mosaic virus is a rapidly emerging virus which has established itself as one of the most important viral diseases in tomato production worldwide over recent years. Begomovirus species and other whitefly-transmitted viruses are invading into new areas, and several recently described new viruses such as Tomato torrado virus and new Tospovirus species are rapidly spreading over large geographic areas. In this article, emerging viruses of tomato crops are discussed

Rapid evolutionary dynamics of the Pepino mosaic virus - status and future perspectives

Pepino mosaic virus (PepMV) has emerged as an important pathogen of greenhouse tomato crops and is currently distributed worldwide. Population genetic studies have revealed a shift in the dominant PepMV genotype from European (EU) to Chilean 2 (CH2) in North America and several European countries. New genetic variants are constantly being created by mutation and recombination events. Single nucleotide substitutions in different parts of the genome were found to affect on development of symptoms resulting in new pathotypes and accumulation of viral RNA. The variability of the PepMV population has a great impact on designing specific diagnostic tools and developing efficient and durable strategies of disease control. In this paper we review the current knowledge about the PepMV population, the evolutionary dynamics of this highly infective virus, methods for its detection and plant protection strategies

Pepino mosaic virus isolates and differential symptomatology in tomato

Based on a survey conducted in commercial tomato production in Belgium in 2006, four Pepino mosaic virus (PepMV) isolates that differed in symptom expression in the crop of origin were selected for greenhouse trials. The selected isolates were inoculated onto tomato plants grown in four separate plastic tunnels. PepMV symptom development was assessed regularly and extensive sampling followed by ELISA analyses, genotyping and sequencing was performed to study viral presence and variation in PepMV sequences throughout the trial period. Two isolates (EU-mild and CH2-mild) that were selected based on mild symptom expression in the crop of origin caused only mild symptoms in the trial, while two other isolates (CH2-aggressive and EU + CH2) that were selected for severe symptom display, caused considerably more severe symptoms. Sequence homology between CH2-mild and CH2-aggressive was as high as 99·4%. Results of this study show that differential symptom expression can, at least partially, be attributed to the PepMV isolate, which may be related to minor differences at the nucleotide level between isolates