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Gerard Demangeat - One of the best experts on this subject based on the ideXlab platform.

  • From a Movement-Deficient Grapevine Fanleaf Virus to the Identification of a New Viral Determinant of Nematode Transmission
    Viruses, 2019
    Co-Authors: Lorène Belval, Veronique Komar, Olivier Lemaire, Emmanuelle Vigne, Aurelie Marmonier, Sophie Gersch, Corinne Schmitt-keichinger, Peggy Andret-link, Gerard Demangeat
    Abstract:

    Grapevine fanleaf Virus (GFLV) and Arabis Mosaic Virus (ArMV) are nepoViruses responsible for grapevine degeneration. They are specifically transmitted from grapevine to grapevine by two distinct ectoparasitic dagger nematodes of the genus Xiphinema. GFLV and ArMV move from cell to cell as virions through tubules formed into plasmodesmata by the self-assembly of the viral movement protein. Five surface-exposed regions in the coat protein called R1 to R5, which differ between the two Viruses, were previously defined and exchanged to test their involvement in Virus transmission, leading to the identification of region R2 as a transmission determinant. Region R4 (amino acids 258 to 264) could not be tested in transmission due to its requirement for plant systemic infection. Here, we present a fine-tuning mutagenesis of the GFLV coat protein in and around region R4 that restored the Virus movement and allowed its evaluation in transmission. We show that residues T258, M260, D261, and R301 play a crucial role in Virus transmission, thus representing a new viral determinant of nematode transmission.

  • The backbone model of the Arabis Mosaic Virus reveals new insights into functional domains of NepoVirus capsid
    Journal of Structural Biology, 2013
    Co-Authors: J. Lai-kee-him, Pascale Schellenberger, Gerard Demangeat, C. Dumas, E. Richard, S. Trapani, V. Komar, C. Ritzenthaler, P. Bron
    Abstract:

    Arabis Mosaic Virus (ArMV) and Grapevine fanleaf Virus (GFLV) are two picorna-like Viruses from the genus NepoVirus, consisting in a bipartite RNA genome encapsidated into a 30 nm icosahedral viral particle formed by 60 copies of a single capsid protein (CP). They are responsible for a severe degeneration of grapevines that occurs in most vineyards worldwide. Although sharing a high level of sequence identity between their CP, ArMV is transmitted exclusively by the ectoparasitic nematode Xiphinema diversicaudatum whereas GFLV is specifically transmitted by the nematode X. index. The structural determinants involved in the transmission specificity of both Viruses map solely to their respective CP. Recently, reverse genetic and crystallographic studies on GFLV revealed that a positively charged pocket in the CP B domain located at the Virus surface may be responsible for vector specificity. To go further into delineating the coat protein determinants involved in transmission specificity, we determined the 6.5 Å resolution cryo-electron microscopy structure of ArMV and used homology modeling and flexible fitting approaches to build its pseudo-atomic structure. This study allowed us to resolve ArMV CP architecture and delineate connections between ArMV capsid shell and its RNA. Comparison of ArMV and GFLV CPs reveals structural differences in the B domain pocket, thus strengthening the hypothesis of a key role of this region in the viral transmission specificity and identifies new potential functional domains of NepoVirus capsid.

  • First detection of Tomato black ring Virus (TBRV) in a French vineyard
    Journal International des Sciences de la Vigne et du Vin, 2013
    Co-Authors: Coralie Laveau, Daniel Esmenjaud, Maarten Van Helden, Guillaume Darrieutort, Gerard Demangeat
    Abstract:

    Aim : Grapevine plants from the Bordeaux wine region (France) showing symptoms of fanleaf degeneration, but negative for the two main fanleaf Viruses were screened by ELISA for other nepoViruses that could explain the symptoms. Methods and results : ELISA tests were performed over a 3-year period (2009-2011) on leaves and woody canes. A total of 665 grapevine plants grafted with Merlot, Cabernet franc and Cabernet-Sauvignon, were found free from Grapevine fanleaf Virus (GFLV) and Arabis Mosaic Virus (ArMV) but infected with Tomato black ring Virus (TBRV). The Longidorid nematode species Longidorus attenuatus, known as the TBRV vector in grapevine, was detected from soil samples collected in the infected area. Conclusion : Both the Virus and its vector might have originated from a vegetable garden established prior to vine planting, considering that the TBRV-infected area with the most fanleaf degeneration symptoms co-localizes with this previous garden. Significance and impact of the study: This is the first record of TBRV infection in a grapevine plot in France.

  • the coat protein determines the specificity of Virus transmission by xiphinema diversicaudatum
    Journal of Plant Pathology, 2010
    Co-Authors: Aurelie Marmonier, Marc Fuchs, Pascale Schellenberger, Olivier Lemaire, Daniel Esmenjaud, P Andretlink, Corinne Schmittkeichinger, C. Ritzenthaler, Gerard Demangeat
    Abstract:

    Arabis Mosaic Virus (ArMV) and Grapevine fanleaf Virus (GFLV), two closely related members of the genus NepoVirus, family Secoviridae, are responsible for fanleaf degeneration disease of grapevines. ArMV and GFLV are specifically transmitted by the ectoparasitic soilborne nematodes Xiphinema diversicaudatum and X. index, respectively. Previous work has shown that the coat protein determines the transmission specificity of GFLV. No information is available on the determinants of the specific transmission of ArMV by X. diversicaudatum. We addressed this issue by testing the transmissibility of three chimeric Viruses comprising GFLV RNA-1 and GFLV/ArMV chimeric RNA-2 molecules coding for a coat protein from ArMV. The three recombinant Viruses and wild type ArMV, unlike GFLV, were transmitted by X. diversicaudatum but not by X. index. In addition, RT-PCR assays indicated that ArMV and the three chimeric Viruses seemed to be retained by X. diversicaudatum but not by X. index following nematode feeding on infected plants. Conversely, GFLV was retained by X. index but not by X. diversicaudatum. These results are consistent with the notion that the coat protein carries the determinants for the specific transmission of Ar- MV by X. diversicaudatum and that transmission specificity of both ArMV and GFLV is associated with specific Virus retention by nematode vectors.

  • Cross-protection as control strategy against grapevine fanleaf Virus in naturally infected vineyards
    Plant Disease, 2008
    Co-Authors: Veronique Komar, Gerard Demangeat, Emmanuelle Vigne, Olivier Lemaire
    Abstract:

    The efficacy of cross-protection at mitigating the impact of Grapevine fanleaf Virus (GFLV) on grapevines (Vitis vinifera) was assessed in two naturally infected vineyard sites. Test vines consisted of scions grafted onto rootstocks that were healthy or infected by mild protective strains GFLV-GHu or Arabis Mosaic Virus (ArMV)-Ta. Challenge GFLV infection via the nematode Xiphinema index was monitored over nine consecutive years in control and ArMV-Ta crossprotected vines by double-antibody sandwich–enzyme-linked immunosorbent assay using GFLV-specific antibodies, and in GFLV-GHu cross-protected vines by characterizing the coat protein gene of superinfecting isolates by immunocapture–reverse transcription–polymerase chain reaction–restriction fragment length polymorphism. Results were consistent with a significantly reduced challenge infection rate in cross-protected vines compared with control vines, more so in those protected with GFLV-GHu (19 versus 90%) than with ArMV-Ta (40 versus 65% in field A and 63 versus 90% in field B). However, the two mild strains significantly reduced fruit yield by 9% (ArMV-Ta) and 17% (GFLV-GHu) over 8 years and had a limited effect on fruit quality. Therefore, in spite of a great potential at reducing the incidence of challenge field isolates, cross-protection with natural mild protective strains GFLV-GHu and ArMV-Ta is not attractive to control GFLV because the negative impact on yield is a limiting factor for its deployment.

L Pinck - One of the best experts on this subject based on the ideXlab platform.

  • Replication of grapevine fanleaf Virus satellite RNA transcripts in Chenopodium quinoa protoplasts
    2013
    Co-Authors: F. Hans, L Pinck
    Abstract:

    A set of full-length cDNA clones of the satellite RNA of grapevine fanleaf nepoVirus isolate Fl3 (GFLV-F13) was constructed with a variable number of additional, non-viral nucleotides at the 5 ' and 3 ' ends. The biological activity of the RNAs transcribed from these constructs was tested in Chenopodium quinoa protoplasts using a helper Virus. When inoculated with Arabis Mosaic Virus S (ArMV-S) RNA as helper, transcripts with 33 non-viral nucleotides at the 5 ' end (tr45p4) did not replicate, whereas transcripts with only one non-viral nucleotide at the 5 ' end (tr3S and tr3M) did replicate. Capping of the transcripts enhanced their replication. On the other hand, the presence of extra nucleotides at the 3 ' end had little influence on the biological activity of the in vitro transcripts. In contrast with ArMV-S, GFLV isolate 24 was not a helper for tr3M transcripts, indicating a specific interaction between the helper strain and the satellite RNA

  • involvement of rna2 encoded proteins in the specific transmission of grapevine fanleaf Virus by its nematode vector xiphinema index
    Virology, 2001
    Co-Authors: Christophe Belin, Gerard Demangeat, Veronique Komar, Corinne Schmitt, L Pinck
    Abstract:

    Abstract The nepoVirus Grapevine fanleaf Virus (GFLV) is specifically transmitted by the nematode Xiphinema index. To identify the RNA2-encoded proteins involved in X. index -mediated spread of GFLV, chimeric RNA2 constructs were engineered by replacing the 2A, 2B MP , and/or 2C CP sequences of GFLV with their counterparts in Arabis Mosaic Virus (ArMV), a closely related nepoVirus which is transmitted by Xiphinema diversicaudatum but not by X. index. Among the recombinant Viruses obtained from transcripts of GFLV RNA1 and chimeric RNA2, only those which contained the 2C CP gene (504 aa) and 2B MP contiguous 9 C-terminal residues of GFLV were transmitted by X. index as efficiently as natural and synthetic wild-type GFLV, regardless of the origin of the 2A and 2B MP genes. As expected, ArMV was not transmitted probably because it is not retained by X. index. These results indicate that the determinants responsible for the specific spread of GFLV by X. index are located within the 513 C-terminal residues of the polyprotein encoded by RNA2.

  • the nine c terminal residues of the grapevine fanleaf nepoVirus movement protein are critical for systemic Virus spread
    Journal of General Virology, 1999
    Co-Authors: Christophe Belin, Gerard Demangeat, Corinne Schmitt, B Walter, F Gaire, L Pinck
    Abstract:

    The grapevine fanleaf Virus (GFLV) RNA2-encoded polyprotein P2 is proteolytically cleaved by the RNA1-encoded proteinase to yield protein 2A, 2B(MP) movement protein and 2C(CP) coat protein. To further investigate the role of the 2B(MP) and 2C(CP) proteins in Virus movement, RNA2 was engineered by alternatively replacing the GFLV 2B(MP) and 2C(CP) genes with their counterparts from the closely related Arabis Mosaic Virus (ArMV). Transcripts of all chimeric RNA2s were able to replicate in Chenopodium quinoa protoplasts and form tubules in tobacco BY-2 protoplasts in the presence of the infectious transcript of GFLV RNA1. Virus particles were produced when the GFLV 2C(CP) gene was replaced with its ArMV counterpart, but systemic Virus spread did not occur in C. quinoa plants. In addition, chimeric RNA2 containing the complete ArMV 2B(MP) gene was neither encapsidated nor infectious on plants, probably because polyprotein P2 was incompletely processed. However, chimeric RNA2 encoding ArMV 2B(MP), in which the nine C-terminal residues were those of GFLV 2B(MP), formed Virus particles and were infectious in the presence of GFLV but not ArMV 2C(CP). These results suggest that the nine C-terminal residues of 2B(MP) must be of the same Virus origin as the proteinase for efficient proteolytic processing of polyprotein P2 and from the same Virus origin as the 2C(CP) for systemic Virus spread.

  • the 119 kda and 124 kda polyproteins of Arabis Mosaic nepoVirus isolate s are encoded by two distinct rna2 species
    Journal of General Virology, 1995
    Co-Authors: A M Loudes, C. Ritzenthaler, M. A. Serghini, Monique Pinck, L Pinck
    Abstract:

    Arabis Mosaic Virus (ArMV) is a nepoVirus that is serologically distantly related to grapevine fanleaf Virus (GFLV). Both ArMV and GFLV induce grapevine degeneration disease. Several ArMV isolates, unlike isolates of GFLV, produce upon in vitro translation of RNA2 a polyprotein (P2) that forms a double band in polyacrylamide-SDS gels. Cloning of full-length copies of RNA2 of an ArMV isolate from grapevine (ArMV-S) revealed that this isolate contained two RNA2s of different length, called RNA2-U and RNA2-L. The two species were not readily separated by electrophoresis of the virion RNA under denaturing gel electrophoresis conditions but could be distinguished by analysis of primer extension and in vitro translation products. The size difference of the two RNA2s is due mostly if not exclusively to differences in their coding regions. The 124 kDa RNA2-U-encoded polyprotein P2′ and the 119 kDa RNA2-L-encoded polyprotein P2″, which comigrate, respectively, with the upper and lower polyprotein bands produced by RNA2 of ArMV-S, were more than 95% identical except in their N-terminal domains. In vitro maturation experiments and sequence comparisons indicate that the N-terminal products of P2′ and P2″ have a molecular mass of 31 kDa and 26 kDa. The genomic organization proposed is similar to that of GFLV RNA2.

  • Protection against Virus infection in tobacco plants expressing the coat protein of grapevine fanleaf nepoVirus.
    Plant cell reports, 1994
    Co-Authors: N. Bardonnet, F. Hans, M. A. Serghini, L Pinck
    Abstract:

    Grapevine fanleaf nepoVirus (GFLV) is responsible for the economically significant “court-noue” disease in vineyards. Its genome is made up of two single-stranded RNA molecules (RNA1 and RNA2) which direct the synthesis of polyproteins P1 and P2 respectively. A chimeric coat protein gene derived from the C-terminal part of P2 was constructed and subsequently introduced into a binary transformation vector. Transgenic Nicotiana benthamiana plants expressing the coat protein under the control of the CaMV 35S promoter were engineered by Agrobacterium tumefaciens-mediated transformation. Protection against infection with virions or viral RNA was tested in coat protein-expressing plants. A significant delay of systemic invasion was observed in transgenic plants inoculated with Virus compared to control plants. This effect was also observed when plants were inoculated with viral RNA. No coat protein-mediated cross-protection was observed when transgenic plants were infected with Arabis Mosaic Virus (ArMV), a closely related nepoVirus also responsible for a “court-noue” disease.

Emmanuelle Vigne - One of the best experts on this subject based on the ideXlab platform.

  • From a Movement-Deficient Grapevine Fanleaf Virus to the Identification of a New Viral Determinant of Nematode Transmission
    Viruses, 2019
    Co-Authors: Lorène Belval, Veronique Komar, Olivier Lemaire, Emmanuelle Vigne, Aurelie Marmonier, Sophie Gersch, Corinne Schmitt-keichinger, Peggy Andret-link, Gerard Demangeat
    Abstract:

    Grapevine fanleaf Virus (GFLV) and Arabis Mosaic Virus (ArMV) are nepoViruses responsible for grapevine degeneration. They are specifically transmitted from grapevine to grapevine by two distinct ectoparasitic dagger nematodes of the genus Xiphinema. GFLV and ArMV move from cell to cell as virions through tubules formed into plasmodesmata by the self-assembly of the viral movement protein. Five surface-exposed regions in the coat protein called R1 to R5, which differ between the two Viruses, were previously defined and exchanged to test their involvement in Virus transmission, leading to the identification of region R2 as a transmission determinant. Region R4 (amino acids 258 to 264) could not be tested in transmission due to its requirement for plant systemic infection. Here, we present a fine-tuning mutagenesis of the GFLV coat protein in and around region R4 that restored the Virus movement and allowed its evaluation in transmission. We show that residues T258, M260, D261, and R301 play a crucial role in Virus transmission, thus representing a new viral determinant of nematode transmission.

  • Nanobody-mediated resistance to Grapevine fanleaf Virus in plants
    Plant Biotechnology Journal, 2018
    Co-Authors: Caroline Hemmer, Veronique Komar, Shahinez Garcia, Emmanuelle Vigne, Aurelie Marmonier, Lea Ackerer, Kamal Hleibieh, Samia Djennane, Sophie Gersch, Mireille Perrin
    Abstract:

    Summary Since their discovery, single-domain antigen-binding fragments of camelid-derived heavy-chain-only antibodies, also known as nanobodies (Nbs), have proven to be of outstanding interest as therapeutics against human diseases and pathogens including Viruses, but their use against phytopathogens remains limited. Many plant Viruses including Grapevine fanleaf Virus (GFLV), a nematode-transmitted icosahedral Virus and causal agent of fanleaf degenerative disease, have worldwide distribution and huge burden on crop yields representing billions of US dollars of losses annually, yet solutions to combat these Viruses are often limited or inefficient. Here, we identified a Nb specific to GFLV that confers strong resistance to GFLV upon stable expression in the model plant Nicotiana benthamiana and also in grapevine rootstock, the natural host of the Virus. We showed that resistance was effective against a broad range of GFLV isolates independently of the inoculation method including upon nematode transmission but not against its close relative, Arabis Mosaic Virus. We also demonstrated that Virus neutralization occurs at an early step of the Virus life cycle, prior to cell-to-cell movement. Our findings will not only be instrumental to confer resistance to GFLV in grapevine, but more generally they pave the way for the generation of novel antiviral strategies in plants based on Nbs.

  • Genetic structure and molecular variability of Grapevine fanleaf Virus populations
    Virus Research, 2010
    Co-Authors: J. E. Olivier, Emmanuelle Vigne, Marc Fuchs
    Abstract:

    To gain insights into the evolutionary mechanisms of Grapevine fanleaf Virus (GFLV) from the genus NepoVirus, family Secoviridae, the sequences of the complete coding region of RNA2, including genes 2AHP, 2BMP and 2CCP, and partial sequence from the RNA1-encoded gene 1EPol of 14 GFLV isolates from three naturally infected California vineyards were characterized. Phylogenetic analyses suggested two to three evolutionarily divergent lineages that did not reflect the vineyard origin of the isolates or an association with rootstock genotype or scion cultivar. Examination of the genetic variability of the California isolates alongside isolates worldwide, for which three RNA1 and 44 RNA2 coding sequences are available, revealed similar patterns of molecular evolution for the different regions within the GFLV genome but distinct selection constraints with the strongest pressure exerted on genes 2CCP and 2BMP, an intermediate level of pressure exerted on gene 1EPol, and the weakest pressure exerted on gene 2AHP. Some of the California isolates resulted from interspecies recombination events between GFLV and Arabis Mosaic Virus with crossover sites suspected in gene 1EPol and identified in genes 2AHP and 2BMP; and intraspecies recombination events inferred in the four target genes but most frequently observed within gene 2CCP. This study suggested that purifying selection and recombination are important evolutionary mechanisms in the genetic diversification of GFLV.

  • Genetic structure and variability of Virus populations in cross-protected grapevines superinfected by Grapevine fanleaf Virus
    Virus Research, 2009
    Co-Authors: Emmanuelle Vigne, Veronique Komar, Aurelie Marmonier, Olivier Lemaire
    Abstract:

    Recombination was assessed in a vineyard site in which grapevines cross-protected with mild strains GHu of Grapevine fanleaf Virus (GFLV) or Ta of Arabis Mosaic Virus (ArMV) were superinfected with GFLV field isolates following transmission by the nematode vector Xiphinema index. The genetic structure and variability within RNA2 of isolates from grapevines co-infected with GFLV field isolates and either GFLVGHu or ArMV-Ta were characterized to identify intra- and interspecies recombinants. Sequence analysis and phylogenetic relationships inferred intraspecies recombination among GFLV field isolates but not between field isolates and GFLV-GHu. SISCAN analysis confirmed a Mosaic structure for two GFLV field isolates for which recombination siteswere located in the movement protein and coat protein genes. One of the recombinantswas found in eight grapevines thatwere in close spatial proximity within the vineyard site, suggesting its transmission by X. index. No interspecies recombination was detected between GFLV field isolates and ArMV-Ta. Altogether, our findings suggest that mild protective strains GFLV-GHu and ArMV-Ta did not assist the emergence of viable recombinants to detectable level during a 12-year crossprotection trial. To our knowledge, this is the first extensive characterization of the genetic structure and variability of Virus isolates in cross-protected plants.

  • multiple interspecies recombination events within rna2 of grapevine fanleaf Virus and Arabis Mosaic Virus
    Archives of Virology, 2008
    Co-Authors: Emmanuelle Vigne, Aurelie Marmonier
    Abstract:

    Sequence alignments and SISCAN analyses inferred multiple interspecies recombination events within RNA2 of strains GHu of Grapevine fanleaf Virus (GFLV) and Ta of Arabis Mosaic Virus (ArMV), two closely related subgroup A nepoViruses in the family Comoviridae. Interspecies recombination events were identified in the 5′ untranslated region, the putative homing protein and movement protein genes but not in the coat protein gene and 3′ untranslated region. These findings suggest a dynamic relationship between GFLV and ArMV, and a differential selection pressure on RNA2-encoded proteins with constraints in terms of function and co-adaptation that limit interspecies recombination to certain gene segments.

Marc Fuchs - One of the best experts on this subject based on the ideXlab platform.

  • genetic variability evolution and biological effects of grapevine fanleaf Virus satellite rnas
    Phytopathology, 2013
    Co-Authors: J Gottula, D Lapato, Keiran K Cantilina, S Saito, B Bartlett, Marc Fuchs
    Abstract:

    Large satellite RNAs (type B satRNAs) of Grapevine fanleaf Virus (GFLV) from the genus NepoVirus, family Secoviridae were identified in a naturally infected vineyard and a grapevine germplasm collection. These GFLV satRNA variants had a higher nucleotide sequence identity with satRNAs of Arabis Mosaic Virus (ArMV) strains NW and J86 (93.8 to 94.6%) than with the satRNA of GFLV strain F13 and those of other ArMV strains (68.3 to 75.0%). Phylogenetic analyses showed no distinction of GFLV and ArMV satRNAs with respect to the identity of the helper Virus. Seven stretches of 8 to 15 conserved nucleotides (I-VII) were identified in the 5' region of subgroup A nepoVirus genomic RNAs GFLV, ArMV, and Grapevine deformation Virus) and nepoVirus type B satRNAs, including previously reported motif I, suggesting that large satRNAs might have originated from recombination between an ancestral subgroup A nepoVirus RNA and an unknown RNA sequence with the 5' region acting as a putative cis-replication element. A comparative analysis of two GFLV strains carrying or absent of satRNAs showed no discernable effect on Virus accumulation and symptom expression in Chenopodium quinoa, a systemic herbaceous host. This work sheds light on the origin and biological effects of large satRNAs associated with subgroup A nepoViruses.

  • Arabis Mosaic Virus in grapevines in new york state
    Plant Disease, 2013
    Co-Authors: F Celebitoprak, Keith L. Perry, Jeremy R. Thompson, Marc Fuchs
    Abstract:

    In a limited survey of commercial vineyards and a germplasm repository in Ontario County, NY, 20 vines of Vitis sp. were tested in fall and spring 2010 to 2012 for Viruses using a double-antibody sandwich (DAS)-ELISA and macroarray with oligonucleotide probes for grapevine Viruses ((3) and unpublished). The plants selected for analysis included those showing atypical growth including leaf deformation, yellowing, cupping or spotting, vein clearing, shortening of internodes, and reduced vigor. Arabis Mosaic Virus (ArMV; genus NepoVirus, family Secoviridae) was detected in leaf tissue and wood scrapings in two vines using the DAS-ELISA with antibodies from Bioreba (Reinach, Switzerland). The ArMV positive vines were from Vitis hybrid cultivars Noah and Geisenheim 26. ArMV was also detected in these two vines using the macroarray, with hybridization observed to 24 of 32 oligonucleotide probes specific to this Virus. To confirm the identification of the Virus, total RNAs were extracted from leaf tissues, hybri...

  • the coat protein determines the specificity of Virus transmission by xiphinema diversicaudatum
    Journal of Plant Pathology, 2010
    Co-Authors: Aurelie Marmonier, Marc Fuchs, Pascale Schellenberger, Olivier Lemaire, Daniel Esmenjaud, P Andretlink, Corinne Schmittkeichinger, C. Ritzenthaler, Gerard Demangeat
    Abstract:

    Arabis Mosaic Virus (ArMV) and Grapevine fanleaf Virus (GFLV), two closely related members of the genus NepoVirus, family Secoviridae, are responsible for fanleaf degeneration disease of grapevines. ArMV and GFLV are specifically transmitted by the ectoparasitic soilborne nematodes Xiphinema diversicaudatum and X. index, respectively. Previous work has shown that the coat protein determines the transmission specificity of GFLV. No information is available on the determinants of the specific transmission of ArMV by X. diversicaudatum. We addressed this issue by testing the transmissibility of three chimeric Viruses comprising GFLV RNA-1 and GFLV/ArMV chimeric RNA-2 molecules coding for a coat protein from ArMV. The three recombinant Viruses and wild type ArMV, unlike GFLV, were transmitted by X. diversicaudatum but not by X. index. In addition, RT-PCR assays indicated that ArMV and the three chimeric Viruses seemed to be retained by X. diversicaudatum but not by X. index following nematode feeding on infected plants. Conversely, GFLV was retained by X. index but not by X. diversicaudatum. These results are consistent with the notion that the coat protein carries the determinants for the specific transmission of Ar- MV by X. diversicaudatum and that transmission specificity of both ArMV and GFLV is associated with specific Virus retention by nematode vectors.

  • Genetic structure and molecular variability of Grapevine fanleaf Virus populations
    Virus Research, 2010
    Co-Authors: J. E. Olivier, Emmanuelle Vigne, Marc Fuchs
    Abstract:

    To gain insights into the evolutionary mechanisms of Grapevine fanleaf Virus (GFLV) from the genus NepoVirus, family Secoviridae, the sequences of the complete coding region of RNA2, including genes 2AHP, 2BMP and 2CCP, and partial sequence from the RNA1-encoded gene 1EPol of 14 GFLV isolates from three naturally infected California vineyards were characterized. Phylogenetic analyses suggested two to three evolutionarily divergent lineages that did not reflect the vineyard origin of the isolates or an association with rootstock genotype or scion cultivar. Examination of the genetic variability of the California isolates alongside isolates worldwide, for which three RNA1 and 44 RNA2 coding sequences are available, revealed similar patterns of molecular evolution for the different regions within the GFLV genome but distinct selection constraints with the strongest pressure exerted on genes 2CCP and 2BMP, an intermediate level of pressure exerted on gene 1EPol, and the weakest pressure exerted on gene 2AHP. Some of the California isolates resulted from interspecies recombination events between GFLV and Arabis Mosaic Virus with crossover sites suspected in gene 1EPol and identified in genes 2AHP and 2BMP; and intraspecies recombination events inferred in the four target genes but most frequently observed within gene 2CCP. This study suggested that purifying selection and recombination are important evolutionary mechanisms in the genetic diversification of GFLV.

  • Transgenic grapevine rootstocck clones expressing the coat protein or movement protein genes of Grapevine fanleaf Virus: Characterization and reaction to Virus infection upon protoplast electroporation
    Plant Science, 2006
    Co-Authors: Laure Valat, Marc Fuchs, Monique Burrus
    Abstract:

    The reaction to Grapevine fanleaf Virus (GFLV) infection in 42 independent transgenic grapevine rootstock 41B clones expressing the coat protein (CP) or movement protein (MP) gene of GFLV was assayed by protoplast electroporation. Two of the 26 transgenic clones expressing the CP gene did not support the accumulation of GFLV MP to detectable levels, 12 accumulated substantially lower levels of MP, and 12 accumulated equivalent levels of MP relative to protoplasts of nontransformed controls at 72 h post-electroporation, as shown by Western blots with anti-MP γ-globulins. Interestingly, inhibition of MP accumulation was achieved against virions but not viral RNAs, and was dependent on the inoculum dose. No interference was observed with the multiplication of Arabis Mosaic Virus, which is closely related to GFLV, likely due to low nucleotide identity between the CP genes. Also, one of the 16 transgenic clones expressing the MP gene significantly reduced the accumulation level of GFLV CP at 72 h post-electroporation, as shown by DAS-ELISA with anti-GFLV γ-globulins. The potential of protoplast electroporation as rapid identification of GFLV-resistant grapevine clones at the cell level will be discussed relative to field screening for resistance at the plant level by nematode-mediated GFLV transmission.

Veronique Komar - One of the best experts on this subject based on the ideXlab platform.

  • From a Movement-Deficient Grapevine Fanleaf Virus to the Identification of a New Viral Determinant of Nematode Transmission
    Viruses, 2019
    Co-Authors: Lorène Belval, Veronique Komar, Olivier Lemaire, Emmanuelle Vigne, Aurelie Marmonier, Sophie Gersch, Corinne Schmitt-keichinger, Peggy Andret-link, Gerard Demangeat
    Abstract:

    Grapevine fanleaf Virus (GFLV) and Arabis Mosaic Virus (ArMV) are nepoViruses responsible for grapevine degeneration. They are specifically transmitted from grapevine to grapevine by two distinct ectoparasitic dagger nematodes of the genus Xiphinema. GFLV and ArMV move from cell to cell as virions through tubules formed into plasmodesmata by the self-assembly of the viral movement protein. Five surface-exposed regions in the coat protein called R1 to R5, which differ between the two Viruses, were previously defined and exchanged to test their involvement in Virus transmission, leading to the identification of region R2 as a transmission determinant. Region R4 (amino acids 258 to 264) could not be tested in transmission due to its requirement for plant systemic infection. Here, we present a fine-tuning mutagenesis of the GFLV coat protein in and around region R4 that restored the Virus movement and allowed its evaluation in transmission. We show that residues T258, M260, D261, and R301 play a crucial role in Virus transmission, thus representing a new viral determinant of nematode transmission.

  • Nanobody-mediated resistance to Grapevine fanleaf Virus in plants
    Plant Biotechnology Journal, 2018
    Co-Authors: Caroline Hemmer, Veronique Komar, Shahinez Garcia, Emmanuelle Vigne, Aurelie Marmonier, Lea Ackerer, Kamal Hleibieh, Samia Djennane, Sophie Gersch, Mireille Perrin
    Abstract:

    Summary Since their discovery, single-domain antigen-binding fragments of camelid-derived heavy-chain-only antibodies, also known as nanobodies (Nbs), have proven to be of outstanding interest as therapeutics against human diseases and pathogens including Viruses, but their use against phytopathogens remains limited. Many plant Viruses including Grapevine fanleaf Virus (GFLV), a nematode-transmitted icosahedral Virus and causal agent of fanleaf degenerative disease, have worldwide distribution and huge burden on crop yields representing billions of US dollars of losses annually, yet solutions to combat these Viruses are often limited or inefficient. Here, we identified a Nb specific to GFLV that confers strong resistance to GFLV upon stable expression in the model plant Nicotiana benthamiana and also in grapevine rootstock, the natural host of the Virus. We showed that resistance was effective against a broad range of GFLV isolates independently of the inoculation method including upon nematode transmission but not against its close relative, Arabis Mosaic Virus. We also demonstrated that Virus neutralization occurs at an early step of the Virus life cycle, prior to cell-to-cell movement. Our findings will not only be instrumental to confer resistance to GFLV in grapevine, but more generally they pave the way for the generation of novel antiviral strategies in plants based on Nbs.

  • Genetic structure and variability of Virus populations in cross-protected grapevines superinfected by Grapevine fanleaf Virus
    Virus Research, 2009
    Co-Authors: Emmanuelle Vigne, Veronique Komar, Aurelie Marmonier, Olivier Lemaire
    Abstract:

    Recombination was assessed in a vineyard site in which grapevines cross-protected with mild strains GHu of Grapevine fanleaf Virus (GFLV) or Ta of Arabis Mosaic Virus (ArMV) were superinfected with GFLV field isolates following transmission by the nematode vector Xiphinema index. The genetic structure and variability within RNA2 of isolates from grapevines co-infected with GFLV field isolates and either GFLVGHu or ArMV-Ta were characterized to identify intra- and interspecies recombinants. Sequence analysis and phylogenetic relationships inferred intraspecies recombination among GFLV field isolates but not between field isolates and GFLV-GHu. SISCAN analysis confirmed a Mosaic structure for two GFLV field isolates for which recombination siteswere located in the movement protein and coat protein genes. One of the recombinantswas found in eight grapevines thatwere in close spatial proximity within the vineyard site, suggesting its transmission by X. index. No interspecies recombination was detected between GFLV field isolates and ArMV-Ta. Altogether, our findings suggest that mild protective strains GFLV-GHu and ArMV-Ta did not assist the emergence of viable recombinants to detectable level during a 12-year crossprotection trial. To our knowledge, this is the first extensive characterization of the genetic structure and variability of Virus isolates in cross-protected plants.

  • Cross-protection as control strategy against grapevine fanleaf Virus in naturally infected vineyards
    Plant Disease, 2008
    Co-Authors: Veronique Komar, Gerard Demangeat, Emmanuelle Vigne, Olivier Lemaire
    Abstract:

    The efficacy of cross-protection at mitigating the impact of Grapevine fanleaf Virus (GFLV) on grapevines (Vitis vinifera) was assessed in two naturally infected vineyard sites. Test vines consisted of scions grafted onto rootstocks that were healthy or infected by mild protective strains GFLV-GHu or Arabis Mosaic Virus (ArMV)-Ta. Challenge GFLV infection via the nematode Xiphinema index was monitored over nine consecutive years in control and ArMV-Ta crossprotected vines by double-antibody sandwich–enzyme-linked immunosorbent assay using GFLV-specific antibodies, and in GFLV-GHu cross-protected vines by characterizing the coat protein gene of superinfecting isolates by immunocapture–reverse transcription–polymerase chain reaction–restriction fragment length polymorphism. Results were consistent with a significantly reduced challenge infection rate in cross-protected vines compared with control vines, more so in those protected with GFLV-GHu (19 versus 90%) than with ArMV-Ta (40 versus 65% in field A and 63 versus 90% in field B). However, the two mild strains significantly reduced fruit yield by 9% (ArMV-Ta) and 17% (GFLV-GHu) over 8 years and had a limited effect on fruit quality. Therefore, in spite of a great potential at reducing the incidence of challenge field isolates, cross-protection with natural mild protective strains GFLV-GHu and ArMV-Ta is not attractive to control GFLV because the negative impact on yield is a limiting factor for its deployment.

  • the specific transmission of grapevine fanleaf Virus by its nematode vector xiphinema index is solely determined by the viral coat protein
    Virology, 2004
    Co-Authors: P Andretlink, Gerard Demangeat, Corinne Schmittkeichinger, Veronique Komar
    Abstract:

    Abstract The viral determinants involved in the specific transmission of Grapevine fanleaf Virus (GFLV) by its nematode vector Xiphinema index are located within the 513 C-terminal residues of the RNA2-encoded polyprotein, that is, the 9 C-terminal amino acids of the movement protein (2BMP) and contiguous 504 amino acids of the coat protein (2CCP) [Virology 291 (2001) 161]. To further delineate the viral determinants responsible for the specific spread, the four amino acids that are different within the 9 C-terminal 2BMP residues between GFLV and Arabis Mosaic Virus (ArMV), another nepoVirus which is transmitted by Xiphinema diversicaudatum but not by X. index, were subjected to mutational analysis. Of the recombinant Viruses derived from transcripts of GFLV RNA1 and RNA2 mutants that systemically infected herbaceous host plants, all with the 2CCP of GFLV were transmitted by X. index unlike none with the 2CCP of ArMV, regardless of the mutations within the 2BMP C-terminus. These results demonstrate that the coat protein is the sole viral determinant for the specific spread of GFLV by X. index.