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1.
Arch Virol ; 168(12): 296, 2023 Nov 20.
Article En | MEDLINE | ID: mdl-37985520

Neckar River virus (NRV), first isolated from a water sample of the Neckar River (Germany) in the 1980s, was serologically characterized as a novel tombusvirus. In this study, the complete genome sequence was determined, and an infectious full-length cDNA clone was constructed. The genome organization of NRV (DSMZ PV-0270) resembles that of tombusviruses. The genome consists of 4739 nucleotides and contains five open reading frames (ORFs) and one additional putative ORF (pX) in the 3'-terminal region. Phylogenetic analysis and sequence comparisons confirmed NRV to be a member of the species Tombusvirus neckarfluminis in the genus Tombusvirus. The infectious full-length cDNA clone was constructed using Gibson assembly and subsequent infection of Nicotiana benthamiana plants by Rhizobium radiobacter inoculation. The virus derived from the full-length cDNA clone caused symptoms resembling those caused by the wild-type virus, but slightly milder.


Tombusviridae , Tombusvirus , Tombusvirus/genetics , Tombusviridae/genetics , DNA, Complementary , Phylogeny , Genome, Viral , Open Reading Frames , RNA, Viral/genetics
2.
Front Plant Sci ; 14: 1187563, 2023.
Article En | MEDLINE | ID: mdl-37600206

Asparagus samples were examined from growing areas of Germany and selected European as well as North, Central and South American countries. Overall, 474 samples were analyzed for Asparagus virus 1 (AV1) using DAS-ELISA. In our survey, 19 AV1 isolates were further characterized. Experimental transmission to 11 species belonging to Aizoaceae, Amarantaceae, Asparagaceae, and Solanaceae succeeded. The ultrastructure of AV1 infection in asparagus has been revealed and has been compared with the one in indicator plants. The cylindrical inclusion (CI) protein, a core factor in viral replication, localized within the cytoplasm and in systemic infections adjacent to the plasmodesmata. The majority of isolates referred to pathotype I (PI). These triggered a hypersensitive resistance in inoculated leaves of Chenopodium spp. and were incapable of infecting Nicotiana spp. Only pathotype II (PII) and pathotype III (PIII) infected Nicotiana benthamiana systemically but differed in their virulence when transmitted to Chenopodium spp. The newly identified PIII generated amorphous inclusion bodies and degraded chloroplasts during systemic infection but not in local lesions of infected Chenopodium spp. PIII probably evolved via recombination in asparagus carrying a mixed infection by PI and PII. Phylogeny of the coat protein region recognized two clusters, which did not overlap with the CI-associated grouping of pathotypes. These results provide evidence for ongoing modular evolution of AV1.

3.
Mol Plant Pathol ; 24(10): 1319-1329, 2023 10.
Article En | MEDLINE | ID: mdl-37410356

In the field of plant virology, the usage of reverse genetic systems has been reported for multiple purposes. One is understanding virus-host interaction by labelling viral cDNA clones with fluorescent protein genes to allow visual virus tracking throughout a plant, albeit this visualization depends on technical devices. Here we report the first construction of an infectious cDNA full-length clone of beet mosaic virus (BtMV) that can be efficiently used for Agrobacterium-mediated leaf inoculation with high infection rate in Beta vulgaris, being indistinguishable from the natural virus isolate regarding symptom development and vector transmission. Furthermore, the BtMV clone was tagged with the genes for the monomeric red fluorescent protein or the Beta vulgaris BvMYB1 transcription factor, which activates the betalain biosynthesis pathway. The heterologous expression of BvMYB1 results in activation of betalain biosynthesis genes in planta, allowing visualization of the systemic BtMV spread with the naked eye as red pigmentation emerging throughout beet leaves. In the case of BtMV, the BvMYB1 marker system is stable over multiple mechanical host passages, allows qualitative as well as quantitative virus detection and offers an excellent opportunity to label viruses in plants of the order Caryophyllales, allowing an in-depth investigation of virus-host interactions on the whole plant level.


Beta vulgaris , Potyvirus , Transcription Factors/genetics , Transcription Factors/metabolism , Betalains , Beta vulgaris/metabolism , DNA, Complementary/genetics , Potyvirus/genetics , Plant Diseases
4.
Front Plant Sci ; 14: 1098786, 2023.
Article En | MEDLINE | ID: mdl-37063189

Beet necrotic yellow vein virus (BNYVV) causes rhizomania disease in sugar beet (Beta vulgaris), which is controlled since more than two decades by cultivars harboring the Rz1 resistance gene. The development of resistance-breaking strains has been favored by a high selection pressure on the soil-borne virus population. Resistance-breaking is associated with mutations at amino acid positions 67-70 (tetrad) in the RNA3 encoded pathogenicity factor P25 and the presence of an additional RNA component (RNA5). However, natural BNYVV populations are highly diverse making investigations on the resistance-breaking mechanism rather difficult. Therefore, we applied a reverse genetic system for BNYVV (A type) to study Rz1 resistance-breaking by direct agroinoculation of sugar beet seedlings. The bioassay allowed a clear discrimination between susceptible and Rz1 resistant plants already four weeks after infection, and resistance-breaking was independent of the sugar beet Rz1 genotype. A comprehensive screen of natural tetrads for resistance-breaking revealed several new mutations allowing BNYVV to overcome Rz1. The supplementation of an additional RNA5 encoding the pathogenicity factor P26 allowed virus accumulation in the Rz1 genotype independent of the P25 tetrad. This suggests the presence of two distinct resistance-breaking mechanisms allowing BNYVV to overcome Rz1. Finally, we showed that the resistance-breaking effect of the tetrad and the RNA5 is specific to Rz1 and has no effect on the stability of the second resistance gene Rz2. Consequently, double resistant cultivars (Rz1+Rz2) should provide effective control of Rz1 resistance-breaking strains. Our study highlights the flexibility of the viral genome allowing BNYVV to overcome host resistance, which underlines the need for a continuous search for alternative resistance genes.

5.
Arch Microbiol ; 205(4): 129, 2023 Mar 22.
Article En | MEDLINE | ID: mdl-36947239

Since the first report in 2009, at least ten additional viruses have been identified and assigned to the proposed virus family Alternaviridae. Here we report two new mycoviruses tentatively assigned to this family, both identified as members of the fungal family Nectriaceae, which were isolated from surface-disinfected apple roots (Malus x domestica, Borkh.) affected by apple replant disease (ARD). ARD is a highly complex, worldwide-occurring disease resulting from plant reactions to a disturbed (micro)-biome and leads to high economic losses every year. The first alternavirus characterized in this study was identified in a Dactylonectria torresensis isolate. The virus was tentatively named dactylonectria torresensis alternavirus 1 (DtAV1) as the first member of the proposed new species Alternavirus dactylonectriae. The second virus was identified in an isolate of Ilyonectria robusta and was tentatively named ilyonectria robusta alternavirus 1 (IrAV1) as the first member of the proposed new species Alternavirus ilyonectriae. Full genomic sequences of the viruses were determined and are presented. Further, we found hints for putative components of a methyl transferase machinery using in silico approaches. This putative protein domain is encoded by segment 2. However, this result only establishes the basis for subsequent studies in which the function must be confirmed experimentally in vitro. Thus, this is the first study where a function is predicted to all three genomic segments within the group of the alternaviruses. These findings provide further insights into the virome of ARD-associated fungi and are therefore another brick in the wall of understanding the complexity of the disease.


Hypocreales , Hypocreales/genetics , Genomics , Plants , Phylogeny
6.
Arch Virol ; 167(11): 2411-2415, 2022 Nov.
Article En | MEDLINE | ID: mdl-35963918

The fungus Ilyonectria pseudodestructans belongs to the family Nectriaceae and was found to be part of the endophytic microbiome of apple trees (Malus x domestica, Borkh.) with apple replant disease (ARD). After dsRNA extraction, a mycoviral infection became evident. Here, we report the identification of a new virus, tentatively named "Ilyonectria pseudodestructans chrysovirus 1" (IpCV1), as the first member of the proposed new species "Alphachrysovirus ilyonectriae" within the genus Alphachrysovirus. This is the first report of a chrysovirus infecting a member of the fungal genus Ilyonectria. IpCV1 has a tripartite dsRNA genome with a total length of 8944 bp. The segments are 3439 bp, 2850 bp, and 2655 bp in length, and each dsRNA carries a single ORF. The encoded viral proteins are a 125.92-kDa RNA-dependent RNA polymerase, a 100.75-kDa coat protein, and one protein of unknown function with a predicted molecular mass of 93.04 kDa. The 5´ and 3´ UTRs are comparatively short and are 79 to 91 bp and 62 to 148 bp in length, respectively. This study provides the basis for further investigations of the impact of IpCV1 on its host and the etiology of ARD.


Fungal Viruses , RNA Viruses , 3' Untranslated Regions , Fungal Viruses/genetics , Genome, Viral , Hypocreales , Open Reading Frames , Phylogeny , RNA Viruses/genetics , RNA, Double-Stranded/genetics , RNA, Viral/genetics , RNA-Dependent RNA Polymerase , Viral Proteins/genetics
7.
J Gen Virol ; 103(8)2022 08.
Article En | MEDLINE | ID: mdl-35947097

The A-type of beet necrotic yellow vein virus (BNYVV) is widely distributed in Europe and is one of the major virus types causing rhizomania disease in sugar beet. The closely related P-type is mainly limited to a small region in France (Pithiviers). Both virus types possess four RNAs (RNA1-4), but the P-type harbours an additional fifth RNA species (RNA5). The P-type is associated with stronger disease symptoms and resistance-breaking of Rz1, one of the two resistance genes which are used to control BNYVV infection. These characteristics are presumably due to the presence of RNA5, but experimental evidence for this is lacking. We generated the first infectious cDNA clone of BNYVV P-type to study its pathogenicity in sugar beet in comparison to a previously developed A-type clone. Using this tool, we confirmed the pathogenicity of the P-type clone in the experimental host Nicotiana benthamiana and two Beta species, B. macrocarpa and B. vulgaris. Independent of RNA5, both the A- and the P-type accumulated in lateral roots and reduced the taproot weight of a susceptible sugar beet genotype to a similar extent. In contrast, only the P-type clone was able to accumulate a virus titre in an Rz1-resistant variety whereas the A-type clone failed to infect this variety. The efficiency of the P-type to overcome Rz1 resistance was strongly associated with the presence of RNA5. Only a double resistant variety, harbouring Rz1 and Rz2, prevented an infection with the P-type. Reassortment experiments between the P- and A-type clones demonstrated that both virus types can exchange whole RNA components without losing the ability to replicate and to move systemically in sugar beet. Although our study highlights the close evolutionary relationship between the two virus types, we were able to demonstrate distinct pathogenicity properties that are attributed to the presence of RNA5 in the P-type.


Beta vulgaris , Plant Viruses , Clone Cells , DNA, Complementary/genetics , Plant Diseases , Plant Viruses/genetics , RNA , Sugars , Virulence/genetics
8.
Virus Genes ; 58(5): 423-435, 2022 Oct.
Article En | MEDLINE | ID: mdl-35841525

In this study, three new mycoviruses were identified co-infecting the apple replant disease (ARD)-associated root endophyte Rugonectria rugulosa. After dsRNA extraction, six viral fragments were visualized. Four fragments belong to a quadrivirus, which has a genome size of 17,166 bp. Each of the fragments of this quadrivirus has a single ORF encoding a protein. Two of these proteins are coat protein subunits, one ORF encodes the RdRp, and one protein has an unknown function. This virus was tentatively named rugonectria rugulosa quadrivirus 1 (RrQV1) as a member of the proposed new species Quadrivirus rugonectria. Another fragment represents the dsRNA intermediate form of a + ssRNA mitovirus with a genome size of 2410 nt. This virus encodes an RdRp and is tentatively called rugonectria rugulosa mitovirus 1 (RrMV1). RrMV1 is suggested as a member of a new species with the proposed name Mitovirus rugonectria. The sixth fragment belongs to the genome of an unclassified dsRNA virus tentatively called rugonectria rugulosa dsRNA virus 1 (RrV1). The monopartite dsRNA genome of RrV1 has a length of 8964 bp and contains two ORFs encoding a structure/gag protein and an RdRp. Full genomic sequences were determined and the genome structure as well as molecular properties are presented. After phylogenetic studies and sequence identity analyses, all three isolates are proposed as new mycoviruses. The results help to improve the understanding of the complexity of the factors involved in ARD and support the interest in mycoviral research. Subsequent analyses need to focus on the impact of mycoviruses on the biology and pathogenicity of ARD-associated fungi. The results of such studies could contribute to the development of mitigation strategies against the disease.


Fungal Viruses , Hypocreales , Malus , RNA Viruses , Gene Products, gag/genetics , Genome, Viral/genetics , Hypocreales/genetics , Malus/genetics , Open Reading Frames/genetics , Phylogeny , Protein Subunits/genetics , RNA, Double-Stranded/genetics , RNA, Viral/genetics , RNA-Dependent RNA Polymerase
9.
Arch Virol ; 165(10): 2385-2388, 2020 Oct.
Article En | MEDLINE | ID: mdl-32647930

Spartina mottle virus (SpMV), an unassigned member of the family Potyviridae, has been known since 1980, when it was first described in England and Wales in symptomatic plants of the genus Spartina. In infected cells, flexuous particles and pinwheel inclusion bodies were found that resemble those of potyvirids. To date, the NCBI database contains only two partial sequences of a German (Nessmersiel) and an Italian (Assisi) isolate, suggesting that SpMV could be the first member of a new genus, called "Sparmovirus", in the family Potyviridae. In this study, the first complete genome sequence of the German SpMV isolate (SpMV Ger) was determined. The genome of SpMV is a single-stranded, monopartite, polyadenylated RNA consisting of 9376 nucleotides. Sequence analysis revealed a genome organization similar to that of classical potyviruses, including many conserved features. In phylogenetic analysis, SpMV could not be assigned to any of the known genera, but it showed the closest relationship to rymoviruses and common reed chlorotic stripe virus (CRCSV, unassigned). Sequence comparisons confirmed that a new genus should be established containing SpMV, CRCSV, and three Bermuda grass mosaic virus isolates, which are considered divergent strains of SpMV.


Genome, Viral , Poaceae/virology , Polyproteins/genetics , Potyviridae/genetics , RNA, Viral/genetics , Germany , Open Reading Frames , Phylogeny , Plant Diseases/virology , Potyviridae/classification , Potyviridae/isolation & purification , Whole Genome Sequencing
10.
Sci Rep ; 10(1): 683, 2020 01 20.
Article En | MEDLINE | ID: mdl-31959799

Rice bacterial blight (BB) caused by Xanthomonas oryzae pv. oryzae (Xoo) implies substantial yield loss to rice. In times of climate change, increasing temperatures are observed and further acceleration is expected worldwide. Increasing temperature often turns into inhibition of host plant defense to pathogens. Recently, a reduced resistance in rice IRBB4 carrying Xa4, but an increase in resistance in IRBB7 carrying Xa7 resistance by increasing temperature has been reported. Influence of high temperature on both R genes (Xa4+Xa7) combined in IRBB67 was analyzed under growth chamber conditions and transcriptomic analysis performed. The pyramided line IRBB67 showed no differences in lesion length between both temperature regimes, demonstrating that non-effectiveness of Xa4 at high temperature did not affect IRBB67 resistance. Moreover, Xa4 complements Xa7 resistance with no Xoo spread in planta beyond the symptomatic area under both temperature regimes in IRBB67. Time course transcriptomic analysis revealed that temperature enhanced IRBB67 resistance to combined heat and Xoo. Our findings highlight altered cellular compartments and point at a role of the cell wall involved in Xoo resistance and heat stress tolerance in both susceptible (IR24) and the resistant (IRBB67) NILs. Interestingly, up-regulation of trehalose-6-phosphatase gene and low affinity cation transporter in IRBB67 suggest that IRBB67 maintained a certain homeostasis under high temperature which may have enhanced its resistance. The interplay of both heat stress and Xoo responses as determined by up-regulated and down-regulated genes demonstrates how resistant plants cope with combined biotic and abiotic stresses.


Disease Resistance , Oryza/growth & development , Plant Proteins/genetics , Cell Wall/genetics , Climate Change , Gene Expression Profiling , Gene Expression Regulation, Plant , Homeostasis , Hot Temperature , Oryza/genetics , Oryza/microbiology , Xanthomonas/pathogenicity
11.
Arch Virol ; 164(7): 1883-1887, 2019 Jul.
Article En | MEDLINE | ID: mdl-31079213

Using next-generation sequencing to characterize agents associated with a severe stunting disease of parsley from Germany, we identified a hitherto undescribed virus. We sequenced total RNA and rolling-circle-amplified DNA from diseased plants. The genome sequence of the virus shows that it is a member of the genus Nanovirus, but it lacks DNA-U4. In addition to the seven genomic DNAs of the virus, we identified a second DNA-R and seven distinct alphasatellites associated with the disease. We propose the name "parsley severe stunt associated virus" (PSSaV) for this novel nanovirus.


DNA, Viral/genetics , Nanovirus/genetics , Nanovirus/isolation & purification , Petroselinum/virology , Plant Diseases/virology , Base Sequence , DNA, Circular/genetics , DNA, Satellite/genetics , DNA, Single-Stranded/genetics , Genome, Viral/genetics , Germany , High-Throughput Nucleotide Sequencing , Nanovirus/classification
12.
J Gen Virol ; 100(2): 308-320, 2019 02.
Article En | MEDLINE | ID: mdl-30667354

Celery latent virus (CeLV) is an incompletely described plant virus known to be sap and seed transmissible and to possess flexuous filamentous particles measuring about 900 nm in length, suggesting it as a possible member of the family Potyviridae. Here, an Italian isolate of CeLV was transmitted by sap to a number of host plants and shown to have a single-stranded and monopartite RNA genome being 11 519 nucleotides (nts) in size and possessing some unusual features. The RNA contains a large open reading frame (ORF) that is flanked by a short 5' untranslated region (UTR) of 13 nt and a 3' UTR consisting of 586 nt that is not polyadenylated. CeLV RNA shares nt sequence identity of only about 40 % with other members of the Potyviridae (potyvirids). The CeLV polyprotein is notable in that it starts with a signal peptide, has a putative P3N-PIPO ORF and shares low aa sequence identity (about 18 %) with other potyvirids. Although potential cleavage sites were not identified for the N-terminal two-thirds of the polyprotein, the latter possesses a number of sequence motifs, the identity and position of which are characteristic of other potyvirids. Attempts at constructing an infectious full-length cDNA clone of CeLV were successful following Rhizobium radiobacter infiltration of Nicotiana benthamiana and Apium graveolens. CeLV appears to have the largest genome of all known potyvirids and some unique genome features that may warrant the creation of a new genus, for which we propose the name 'celavirus'.


Apium/virology , DNA, Complementary , Potyviridae/growth & development , Potyviridae/genetics , 3' Untranslated Regions , 5' Untranslated Regions , Agrobacterium tumefaciens/genetics , Genetic Vectors , Italy , Open Reading Frames , Plant Diseases/virology , Polyproteins/genetics , Potyviridae/isolation & purification , RNA, Viral/genetics , Sequence Homology, Amino Acid , Sequence Homology, Nucleic Acid , Nicotiana , Viral Proteins/genetics
13.
Front Plant Sci ; 10: 1703, 2019.
Article En | MEDLINE | ID: mdl-32010172

Beet necrotic yellow vein virus (BNYVV) is causal agent of rhizomania disease, which is the most devastating viral disease in sugar beet production leading to a dramatic reduction in beet yield and sugar content. The virus is transmitted by the ubiquitous distributed soil-borne plasmodiophoromycete Polymyxa betae that infects the root tissue of young sugar beet plants. Rz1 is the major resistance gene widely used in most sugar beet varieties to control BNYVV. The strong selection pressure on the virus population promoted the development of strains that can overcome Rz1 resistance. Resistance-breaking has been associated with mutations in the RNA3-encoded pathogenicity factor P25 at amino acid positions 67-70 (tetrad) as well as with the presence of an additional RNA component (RNA5). However, respective studies investigating the resistance-breaking mechanism by a reverse genetic system are rather scarce. Therefore, we studied Rz1 resistance-breaking in sugar beet using a recently developed infectious clone of BNYVV A-type. A vector free infection system for the inoculation of young sugar beet seedlings was established. This assay allowed a clear separation between a susceptible and a Rz1 resistant genotype by measuring the virus content in lateral roots at 52 dpi. However, mechanical inoculation of sugar beet leaves led to the occurrence of genotype independent local lesions, suggesting that Rz1 mediates a root specific resistance toward BNYVV that is not active in leaves. Mutation analysis demonstrated that different motifs within the P25 tetrad enable increased virus replication in roots of the resistant genotype. The resistance-breaking ability was further confirmed by the visualization of BNYVV in lateral roots and leaves using a fluorescent-labeled complementary DNA clone of RNA2. Apart from that, reassortment experiments evidenced that RNA5 enables Rz1 resistance-breaking independent of the P25 tetrad motif. Finally, we could identify a new resistance-breaking mutation, which was selected by high-throughput sequencing of a clonal virus population after one host passage in a resistant genotype. Our results demonstrate the feasibility of the reverse genetic system for resistance-breaking analysis and illustrates the genome plasticity of BNYVV allowing the virus to adapt rapidly to sugar beet resistance traits.

14.
Mol Plant Pathol ; 19(10): 2333-2348, 2018 10.
Article En | MEDLINE | ID: mdl-30011123

Rhizomania of sugar beet, caused by Beet necrotic yellow vein virus (BNYVV), is characterized by excessive lateral root (LR) formation leading to dramatic reduction of taproot weight and massive yield losses. LR formation represents a developmental process tightly controlled by auxin signaling through AUX/IAA-ARF responsive module and LATERAL ORGAN BOUNDARIES DOMAIN (LBD) transcriptional network. Several LBD transcription factors play central roles in auxin-regulated LR development and act upstream of EXPANSINS (EXPs), cell wall (CW)-loosening proteins involved in plant development via disruption of the extracellular matrix for CW relaxation and expansion. Here, we present evidence that BNYVV hijacks these auxin-regulated pathways resulting in formation LR and root hairs (RH). We identified an AUX/IAA protein (BvAUX28) as interacting with P25, a viral virulence factor. Mutational analysis indicated that P25 interacts with domains I and II of BvAUX28. Subcellular localization of co-expressed P25 and BvAUX28 showed that P25 inhibits BvAUX28 nuclear localization. Moreover, root-specific LBDs and EXPs were greatly upregulated during rhizomania development. Based on these data, we present a model in which BNYVV P25 protein mimics action of auxin by removing BvAUX28 transcriptional repressor, leading to activation of LBDs and EXPs. Thus, the evidence highlights two pathways operating in parallel and leading to uncontrolled formation of LRs and RHs, the main manifestation of the rhizomania syndrome.


Beta vulgaris/metabolism , Beta vulgaris/virology , Plant Viruses/pathogenicity , Transcription Factors/metabolism , Plant Diseases/genetics , Plant Diseases/virology , Transcription Factors/genetics , Viral Proteins/genetics , Viral Proteins/metabolism , Virulence Factors/genetics , Virulence Factors/metabolism
15.
J Gen Virol ; 2018 Jul 30.
Article En | MEDLINE | ID: mdl-30058995

Infectious full-length clones of Beet necrotic yellow vein virus (BNYVV) and Beet soil-borne mosaic virus (BSBMV), both genus Benyvirus, were used for fluorescent labelling with the objective to study their interaction in coinfection and superinfection experiments. Fluorescent labelling was achieved by replacing a part of the RNA2 encoded coat protein read-through domain with either GFP or mRFP fluorescent marker proteins. This resulted in a translational fusion comprising the coat and the fluorescent protein. The labelled viruses were infectious and moved systemically in Nicotiana benthamiana, producing wild-type-like symptoms. Virus particles could be observed by electron microscopy, demonstrating that the viral read-through domain is dispensable for particle formation. Coinfection experiments revealed a spatial separation of differentially labelled populations of both identical and different Benyvirus species after N. benthamiana agro-inoculation. Identical observations were obtained when Tobacco rattle virus (TRV) was differentially labelled and used for coinfection. In contrast, coinfections of BSBMV with Potato virus X (PVX) or TRV resulted in many co-infected cells lacking spatial separation. Micro-projectile co-bombardment of N. benthamiana leaves revealed that two differently labelled populations of the same virus co-infected only a few cells before starting to separate. In superinfection experiments with N. benthamiana, BSBMV and BNYVV were unable to establish a secondary infection in plants that were previously infected with BNYVV or BSBMV. Taken together, this is the first work to describe the interaction between two economically important Benyviruses using fluorescence-labelled full-length clones.

16.
Virology ; 518: 25-33, 2018 05.
Article En | MEDLINE | ID: mdl-29453056

Two members of the Benyviridae family and genus Benyvirus, Beet soil-borne mosaic virus (BSBMV) and Beet necrotic yellow vein virus (BNYVV), possess identical genome organization, host range and high sequence similarity; they infect Beta vulgaris with variable symptom expression. In the US, mixed infections are described with limited information about viral interactions. Vectors suitable for agroinoculation of all genome components of both viruses were constructed by isothermal in vitro recombination. All 35S promoter-driven cDNA clones allowed production of recombinant viruses competent for Nicotiana benthamiana and Beta macrocarpa systemic infection and Polymyxa betae transmission and were compared to available BNYVV B-type clone. BNYVV and BSBMV RNA1 + 2 reassortants were viable and spread long-distance in N. benthamiana with symptoms dependent on the BNYVV type. Small genomic RNAs were exchangeable and systemically infected B. macrocarpa. These infectious clones represent a powerful tool for the identification of specific molecular host-pathogen determinants.


Beta vulgaris/virology , DNA, Complementary/genetics , Mosaic Viruses/genetics , Plant Diseases/virology , Plant Viruses/genetics , Reassortant Viruses/genetics , Cloning, Molecular , Gene Expression Regulation, Viral , Plant Leaves/virology , RNA Viruses/genetics , RNA, Viral/genetics , RNA, Viral/metabolism
17.
Arch Virol ; 163(4): 1107-1111, 2018 Apr.
Article En | MEDLINE | ID: mdl-29327236

The complete genome sequence of a German isolate of celery mosaic virus (CeMV, a potyvirus) from Quedlinburg (DSMZ PV-1003) was determined (MF962880). This represents the second fully sequenced genome of this virus, along with a Californian isolate (HQ676607.1). The positive-sense single-stranded RNA is 10,000 nucleotides in length and shows the typical organization of potyviruses but has a shorter PIPO than CeMV California. In comparison to CeMV isolates from different origins, CeMV-Quedlinburg and isolates from the Netherlands (AF203531.1) and Aschersleben, Germany (AJ271087.1) show a NAG instead of DAG in the region of the coat protein responsible for aphid transmission. In this study the first infectious full-length clone of celery mosaic virus was obtained and the infectivity confirmed by Rhizobium radiobacter infiltration of Apium species.


Apium/virology , Capsid Proteins/genetics , Genome, Viral , Phylogeny , Potyvirus/genetics , RNA, Viral/genetics , Agrobacterium tumefaciens/genetics , Amino Acid Sequence , Gene Expression , Germany , Open Reading Frames , Plant Diseases/virology , Potyvirus/classification , Potyvirus/isolation & purification , Sequence Alignment , Sequence Analysis, DNA , Sequence Homology, Amino Acid , Whole Genome Sequencing
18.
J Gen Virol ; 99(1): 17-18, 2018 Jan.
Article En | MEDLINE | ID: mdl-29214972

The Partitiviridae is a family of small, isometric, non-enveloped viruses with bisegmented double-stranded (ds) RNA genomes of 3-4.8 kbp. The two genome segments are individually encapsidated. The family has five genera, with characteristic hosts for members of each genus: either plants or fungi for genera Alphapartitivirus and Betapartitivirus, fungi for genus Gammapartitivirus, plants for genus Deltapartitivirus and protozoa for genus Cryspovirus. Partitiviruses are transmitted intracellularly via seeds (plants), oocysts (protozoa) or hyphal anastomosis, cell division and sporogenesis (fungi); there are no known natural vectors. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the taxonomy of the Partitiviridae, which is available at www.ictv.global/report/partitiviridae.


Genome, Viral , Phylogeny , RNA Viruses/genetics , RNA, Viral/genetics , Virion/genetics , Alveolata/virology , Fungi/virology , Plants/virology , RNA Viruses/classification , RNA Viruses/ultrastructure , Terminology as Topic , Virion/ultrastructure , Virus Replication
19.
Arch Virol ; 162(6): 1519-1528, 2017 Jun.
Article En | MEDLINE | ID: mdl-28190200

A Luminex xTAG-based assay for plant-infecting tospoviruses was developed. The test enables the detection of tospoviruses in general and the differentiation of the four important member species of this genus: Tomato spotted wilt virus, Impatiens necrotic spot virus, the proposed 'Capsicum chlorosis virus' and Watermelon silver mottle virus. The generic tospovirus primers used in this method are also applicable for detection of tospoviruses by basic RT-PCR. We also describe an economic alternative method for the distinction of the four tospoviruses mentioned and of additional member viruses, based on a restriction fragment length polymorphism (RFLP). The sophisticated Luminex xTAG technology allows the simultaneous detection of various targets. This study is part of a project that aims to develop a method for the simultaneous detection of various plant pathogens (viral, bacterial and fungal) in plant material.


Immunoassay/methods , Multiplex Polymerase Chain Reaction/methods , Tospovirus/genetics , Tospovirus/isolation & purification , DNA Primers , Plant Diseases/virology , Plants/virology , Polymorphism, Restriction Fragment Length , RNA, Viral/analysis , Restriction Mapping/methods , Tospovirus/classification
20.
Plant Dis ; 100(2): 380-387, 2016 Feb.
Article En | MEDLINE | ID: mdl-30694136

Rice bacterial blight (BB) is caused by Xanthomonas oryzae pv. oryzae and is responsible for substantial yield loss worldwide. Host resistance remains the most feasible control measure. However, pathogen variability leads to the failure of certain resistance genes to control the disease, and climate change with high amplitudes of heat predisposes the host plant to pathogen invasion. Due to pressure in natural selection, landrace species often carry a wide range of unique traits conferring tolerance of stress. Therefore, exploring their genetic background for host resistance could enable the identification of broad-spectrum resistance to combined abiotic and biotic stresses. Nineteen Oryza glaberrima accessions and O. sativa rice variety SUPA were evaluated for BB resistance under high temperature (35 and 31°C day and night, respectively) using 14 X. oryzae pv. oryzae strains originated from the Philippines. Under normal temperature, most of the accessions showed resistance to 9 strains (64.3%) and accession TOG6007 showed broad-spectrum resistance to 12 strains (85.7%). Under high temperature, most accessions showed a reduction in BB disease, whereas, accession TOG5620 showed disease reduction from all the X. oryzae pv. oryzae strains under high temperature. Molecular characterization using gene-based and linked markers for BB resistance genes Xa4, xa5, Xa7, xa13, and Xa21 revealed the susceptible alleles of Xa4, xa5, xa13, and Xa21 in O. glaberrima. However, no allele of Xa7 was detected among O. glaberrima accessions. Our results suggest that O. glaberrima accessions contain a BB resistance different from the Xa gene type. Genome-wide association mapping could be used to identify quantitative trait loci that are associated with BB resistance or combined BB resistance and high-temperature tolerance.

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