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:70mer probes were designed to detect plant viruses infection in genus level. This microarray platform is able to detect 169 plant virus species of 13 virus genera.
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:70mer probes were designed to detect plant viruses infection in genus level. This microarray platform is able to detect 169 plant virus species of 13 virus genera. Virus sampels were extracted from infected plant hosts. Genomic RNA was extracted and hybridized to the microarray.
Project description:Phloem localization of plant viruses is advantageous for acquisition by sap-sucking vectors but hampers host-virus protein interaction studies. In this study, Potato leafroll virus (PLRV)-host protein complexes were isolated from systemically infected potato, a natural host of the virus. Comparing two different co-immunoprecipitation support matrices coupled to mass spectrometry, we identified 44 potato proteins and one viral protein (P1) specifically associated with virus isolated from infected phloem. An additional 142 proteins interact in complex with virus at varying degrees of confidence. Greater than 80% of these proteins were previously found to form high confidence interactions with PLRV isolated from the model host Nicotiana benthamiana. Bioinformatics revealed that these proteins are enriched for functions related to plasmodesmata, organelle membrane transport, translation and mRNA processing. Our results show that model system proteomics experiments are extremely valuable for understanding protein interactions regulating infection in recalcitrant pathogens such as phloem-limited viruses.
Project description:Potato leafroll virus (PLRV) is an aphid-borne, RNA virus in the Luteoviridae that causes significant loss to potato production worldwide. Precision methods to block virus infection in plants or the transmission of a virus by the insect represent new approaches to virus disease management and require an in depth knowledge of the protein interactions used by viruses in their plant hosts and aphid vectors. In this study, we evaluated the feasibility of using an inexpensive, bead-free assay to identify plant proteins that interact with wild-type (WT) and PLRV mutant form. Comparison of host and viral proteins identified as enriched ³2-fold in both WT and mutant experiments using the bead-free method revealed a ~65% overlap with proteins identified as forming high-confident interactions with PLRV using antibody coated-magnetic beads with 19 of these proteins also detected as significant interactions in the bead-free assay. An additional 18 virus-host interactions were only identified in the bead-free assay. Two prey proteins, a 14-3-3 signal transduction protein and malate dehydrogenase 2, were identified as having a weakened or lost association with the mutant form of the virus in both immunoprecipitation assays, showing that the method is sensitive enough to detect quantitative differences between different, yet closely related viral bait proteins. Collectively, our analysis shows that the bead-free platform is a low-cost alternative that can be used by core facilities and other investigators to reproducibly identify plant and viral proteins interacting with virions.
Project description:Parasitoid wasps are one of the most species-rich groups of animals on Earth, due to their ability to successfully develop as parasites of nearly all types of insects. Unlike most known parasitoid wasps that specialize within one or a few host species, Diachasmimorpha longicaudata is a generalist that can survive within multiple genera of tephritid fruit fly hosts, including many globally important pest species. D. longicaudata has therefore been widely released to suppress pest populations as part of biological control efforts in tropical and subtropical agricultural systems. In this study, we investigated the role of a mutualistic poxvirus in shaping the host range of D. longicaudata across three genera of agricultural pest species: two of which are permissive hosts for D. longicaudata parasitism and one that is a nonpermissive host. We found that permissive hosts Ceratitis capitata and Bactrocera dorsalis were highly susceptible to virus infection, displaying rapid virus replication and abundant fly mortality. However, the nonpermissive host Zeugodacus cucurbitae largely overcame virus infection, exhibiting substantially lower mortality and no virus replication. Further investigation of transcriptional dynamics during virus infection demonstrated hindered viral gene expression and limited changes in fly gene expression within the nonpermissive host compared to the permissive species, indicating that the host range of the viral symbiont may dictate the host range of D. longicaudata wasps. These findings also reveal that viral symbiont activity may be a major contributor to the success of D. longicaudata as a generalist parasitoid species and a globally successful biological control agent.
Project description:To understand the molecular basis of viral diseases, transcriptome profiling has been widely used to correlate host gene expression change patterns with disease symptoms during viral infection in many plant hosts. We used infection of apple by Apple stem grooving virus (ASGV), which produces no disease symptoms, to assess the significance of host gene expression changes in disease development. We specifically asked the question whether such asymptomatic infection is attributed to limited changes in host gene expression. Using RNA-seq, we identified a total of 184 up-regulated and 136 down-regulated genes in apple shoot cultures permanently infected by ASGV in comparison with virus-free shoots cultures. As in most plant hosts showing disease symptoms during viral infection, these differentially expressed genes encode known or putative proteins involved in cell cycle, cell wall biogenesis, response to biotic and abiotic stress, development and fruit ripening, phytohormone function, metabolism, signal transduction, transcription regulation, translation, transport, and photosynthesis. Our data suggest that current approaches to correlate host gene expression changes under viral infection conditions to specific infection processes or disease symptom development, based on the interpretation of individual gene functions, have severe limitations. Integrative approaches that can take into account plant development stages, gene threshold levels as well as compensatory, synergistic and antagonistic effects may be necessary to develop a sound systems understanding of the biological significance of host gene expression changes during infection. Compare the transcript profiling of ASGV-infected asymptomatic apple planlets (AP-Vinfect) and virus-free apple plantlets (AP-Vfree) by deep sequencing using Illumina RNA-Seq to check whether lots of genes were modulated by ASGV infection.
Project description:Grapevine rupestris stem pitting-associated virus (GRSPaV) is a widespread virus affecting Vitis spp. Although it has established a compatible viral interaction in Vitis vinifera L. without the development of phenotypic alterations, it can occur as distinct variants that show different symptoms in diverse Vitis species. We investigated the changes induced by GRSPaV in V. vinifera cv Bosco, an Italian white grape variety, by combining agronomic, physiological, and molecular approaches, in order to provide comprehensive information about the global effects of GRSPaV. In two consecutive years, this virus caused a moderate decrease in physiological efficiency, yield performance, and sugar content in berries associated with several transcriptomic alterations. Transcript profiles were analysed by microarray techniques in petiole, leaf, and berry samples collected at véraison and by quantitative real-time RT-PCR (qRT-PCR) in a time course carried out at five relevant grapevine developmental stages. Global gene expression analyses showed that transcriptomic changes were highly variable among the different organs and the different phenological phases. GRSPaV triggers some unique responses in the grapevine at véraison, never reported before for other plant-virus interactions, such as an increase in transcripts involved in photosynthesis and CO2 fixation, associated with a moderate reduction in the photosynthesis rate and some defence mechanisms, and to an overlap with responses to water and salinity stresses. We hypothesise that the long co-existence between grapevine and GRSPaV has resulted in the evolution of a form of mutual adaptation between the virus and its host. This study contributes to elucidating alternative mechanisms used by infected plants to contend with viruses.
2012-10-05 | GSE36632 | GEO
Project description:Genetic diversity of Grapevine leafroll-associated virus 3 in Greek vineyards