Dataset Information

Absolute Quantification of Grapevine Red Blotch Virus in Grapevine Leaf and Petiole Tissues by Proteomics

ABSTRACT: Grapevine red blotch is a recently identified viral disease that was first recognized in the Napa Valley of California. Infected plants showed foliar symptoms similar to leafroll, another grapevine viral disease, on vines testing negative for known grapevine leafroll-associated virus. Later, the Grapevine red blotch virus (GRBV) was independently discovered in the US states of California and New York and was demonstrated to be the causal agent of red blotch disease. Due to its wide occurrence in the US, vector transmission and impacts on grape industry, this virus has the potential to cause serious economic losses. Despite numerous attempts, it was not possible to isolate or visualize viral particles from GRBV infected plants. Consequently, this has hampered the development of a serological assay that would facilitate GRBV detection in grapevine. We therefore decided to explore mass spectrometry approaches in order to quantify GRBV in infected plants and to identify potential biomarkers for viral infection. We present for the first time the physical detection on the protein level of the two GRBV genes V1 (coat protein) and V2 in grapevine tissue lysates. The GRBV coat protein load in leaf petioles was determined to be in the range of 100 to 900 million copies per milligram wet weight by using three heavy isotope labeled reference peptides as internal standards. The V1 copy number per unit wet tissue weight in leaves appeared to be about six times lower, and about 200-times lower in terms of protein concentration in the extractable protein mass than in petioles. We found a consistent upregulation of several enzymes involved in flavonoid biosynthesis in leaf and petiole extracts of GRBV-infected plants by label-free shotgun proteomics, indicating the activation of a defense mechanism against GRBV, a plant response already described for grapevine leafroll associated virus infection on the transcriptome level. Last but not least, we identified some other microorganisms belonging to the grapevine leaf microbiota, two bacterial species (Novosphingobium sp. Rr 2-17 and Methylobacterium) and one virus, Grapevine rupestris stem pitting associated virus.

ORGANISM(S): Grapevine Leafroll-associated Virus Vitis Vinifera

SUBMITTER: Sophie Lagache

PROVIDER: PXD011138 | panorama | Sat Jan 23 00:00:00 GMT 2021

REPOSITORIES: PanoramaPublic

ACCESS DATA

Publications

Absolute Quantification of <i>Grapevine Red Blotch Virus</i> in Grapevine Leaf and Petiole Tissues by Proteomics.

Buchs Natasha N Braga-Lagache Sophie S Uldry Anne-Christine AC Brodard Justine J Debonneville Christophe C Reynard Jean-Sébastien JS Heller Manfred M

Frontiers in plant science 20181129

Grapevine red blotch is a recently identified viral disease that was first recognized in the Napa Valley of California. Infected plants showed foliar symptoms similar to leafroll, another grapevine viral disease, on vines testing negative for known grapevine leafroll-associated virus. Later, the <i>Grapevine red blotch virus</i> (GRBV) was independently discovered in the US states of California and New York and was demonstrated to be the causal agent of red blotch disease. Due to its wide occurr ...[more]

PMID: 30555495

Similar Datasets

Project description:Grapevine red blotch is a recently identified viral disease that was first recognized in the Napa Valley of California. Infected plants showed foliar symptoms similar to leafroll, another grapevine viral disease, on vines testing negative for known grapevine leafroll-associated virus. Later, the Grapevine red blotch virus (GRBV) was independently discovered in the US states of California and New York and was demonstrated to be the causal agent of red blotch disease. Due to its wide occurrence in the US, vector transmission and impacts on grape industry, this virus has the potential to cause serious economic losses. Despite numerous attempts, it was not possible to isolate or visualize viral particles from GRBV infected plants. Consequently, this has hampered the development of a serological assay that would facilitate GRBV detection in grapevine. We therefore decided to explore mass spectrometry approaches in order to quantify GRBV in infected plants and to identify potential biomarkers for viral infection. We present for the first time the physical detection on the protein level of the two GRBV genes V1 (coat protein) and V2 in grapevine tissue lysates. The GRBV coat protein load in leaf petioles was determined to be in the range of 100 to 900 million copies per milligram wet weight by using three heavy isotope labeled reference peptides as internal standards. The V1 copy number per unit wet tissue weight in leaves appeared to be about six times lower, and about 200-times lower in terms of protein concentration in the extractable protein mass than in petioles. We found a consistent upregulation of several enzymes involved in flavonoid biosynthesis in leaf and petiole extracts of GRBV-infected plants by label-free shotgun proteomics, indicating the activation of a defense mechanism against GRBV, a plant response already described for grapevine leafroll associated virus infection on the transcriptome level. Last but not least, we identified some other microorganisms belonging to the grapevine leaf microbiota, two bacterial species (Novosphingobium sp. Rr 2-17 and Methylobacterium) and one virus, Grapevine rupestris stem pitting associated virus.

Project description:The overall goal of the study was to use in vivo data combined with functional genomics to define gene expression signatures representative of a spectrum of HSV CNS infections. Innate immune deficiencies result in a spectrum of severe clinical outcomes following infection. In particular, there is a strong association between loss of the signal transducer and activator of transcription (Stat) pathway, breach of the blood-brain barrier (BBB), and virus-induced neuropathology. The gene signatures that characterize resistance, disease, and mortality in the virus-infected nervous system have not been defined. Herpes simplex virus type 1 (HSV-1) is commonly associated with encephalitis in humans, and humans and mice lacking Stat1 display increased susceptibility to HSV central nervous system (CNS) infections. In this study, two HSV-1 strains were used, KOS (wild type [WT]), and Δvhs, an avirulent recombinant lacking the virion host shutoff (vhs) function. In addition, two mouse strains were used: strain 129 (control) and a Stat1-deficient (Stat1(-/-)) strain. Using combinations of these virus and mouse strains, we established a model of infection resulting in three different outcomes: viral clearance without neurological disease (Δvhs infection of control mice), neurological disease followed by viral clearance (Δvhs infection of Stat1(-/-) mice and WT infection of control mice), or neurological disease followed by death (WT infection of Stat1(-/-) mice). Through the use of functional genomics on the infected brain stem and liver, we determined gene signatures that were representative of the three infection outcomes. Gender matched, 6- to 8- week old immunocompetent, control 129S6 and 129S6 Stat1 knockout mice were infected corneally with 2x10^6 PFU of either wild type HSV-1, a vhs-null HSV virus, or mock-infected. Brain stems and liver of individual mice were isolated at days 1, 3, 5 and 7 post-inoculation for microarray analysis. For microarray analysis, samples were collected from n=2 animals (1 male, 1 female) per mouse strain and virus strain for each time point. Equal masses of tissue were pooled from two mock-infected mice per time point and run on microarray.

Project description:Agaricus bisporus is a soil-inhabiting fungus which is cultivated for production of white button mushrooms. A disease of A. bisporus has been previously described with a range of disease symptoms (yield loss, pinning delay, cap distortions and cap browning) which has been given collective name of “Mushroom Virus X” (MVX). The causes of this disease are not clear however prior to this research an association was found between the disease and double-stranded RNA molecules in the mushroom fruitbodies. The experiment was designed to examine causes and host responses of the disease causing the Brown Cap symptom in the cultivated mushroom A. bisporus. This microarray experiment was performed before the Agaricus bisporus genome was sequenced. The gene sequences used to design probes were from known and novel A. bisporus sequences and sequences of transcript fragments identified by Suppression Subtractive Hybridization of non-symptomatic and virus-diseased A. bisporus mushroom fruitbodies. The A. bisporus mushroom fruitbodies were grown on composted wheat straw using commercial cultivation procedures. The gene expression comparison was made of RNA isolated from 32 mushroom fruitbodies (Agaricus bisporus) samples: 20 samples from 5 separate virus-infected commercial mushroom farms with crops displaying the brown symptom (4 replicate samples per farm) and 12 samples from a non-infected crop grown at the University of Warwick. The precise composition of the viral load was the subject of this and future research/papers. Abstract of Manuscript submitted to Applied and Environmental Microbiology: Characterizing the viral agents causing brown cap mushroom disease of Agaricus bisporus by Daniel Eastwood, Julian Green, Helen Grogan, and Kerry Burton (Paper #AEM01093-15). The symptoms of viral infections of fungi range from cryptic to severe but there is little knowledge of the factors involved in this transition of fungal/viral interactions. Brown Cap Mushroom Disease of the cultivated Agaricus bisporus is economically important and represents a model system to describe this transition. Differentially expressed transcript fragments between mushrooms showing the symptoms of Brown Cap Mushroom Disease and control white non-infected mushrooms have been identified and sequenced. Ten of these RNA fragments have been found to be up-regulated over a thousand-fold between diseased and non-diseased tissue but are absent from the Agaricus bisporus genome sequence and hybridise to double-stranded RNA’s extracted from diseased tissue. We hypothesize these transcript fragments are viral and represent components of the disease-causing agent, a bipartite virus with similarities to the family Partitiviridae. The virus fragments were found at two distinct levels within infected mushrooms, at raised levels in infected, non-symptomatic, white coloured mushrooms and much greater levels (3,500-87,000 times greater) in infected mushrooms exhibiting brown colouration. In addition, differential screening revealed 9 up-regulated and 32 down-regulated host Agaricus bisporus transcripts. Chromametric analysis was able to distinguish colour differences between non-infected white mushrooms and white infected mushrooms at an early stage of mushroom growth. This method may be the basis for an ‘on-farm’ disease detection assay.

Dataset Information

Absolute Quantification of Grapevine Red Blotch Virus in Grapevine Leaf and Petiole Tissues by Proteomics

Publications

Absolute Quantification of <i>Grapevine Red Blotch Virus</i> in Grapevine Leaf and Petiole Tissues by Proteomics.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets