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1.
Nat Commun ; 12(1): 1007, 2021 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-33579946

RESUMO

Plant viruses cause massive crop yield loss worldwide. Most plant viruses are RNA viruses, many of which contain a functional tRNA-like structure. RNase P has the enzymatic activity to catalyze the 5' maturation of precursor tRNAs. It is also able to cleave tRNA-like structures. However, RNase P enzymes only accumulate in the nucleus, mitochondria, and chloroplasts rather than cytosol where virus replication takes place. Here, we report a biotechnology strategy based on the re-localization of plant protein-only RNase P to the cytosol (CytoRP) to target plant viruses tRNA-like structures and thus hamper virus replication. We demonstrate the cytosol localization of protein-only RNase P in Arabidopsis protoplasts. In addition, we provide in vitro evidences for CytoRP to cleave turnip yellow mosaic virus and oilseed rape mosaic virus. However, we observe varied in vivo results. The possible reasons have been discussed. Overall, the results provided here show the potential of using CytoRP for combating some plant viral diseases.


Assuntos
Resistência à Doença/fisiologia , Ribonuclease P/genética , Ribonuclease P/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Cloroplastos/metabolismo , Vírus do Mosaico/genética , Vírus do Mosaico/metabolismo , Vírus de Plantas/genética , Protoplastos/metabolismo , Precursores de RNA/metabolismo , RNA de Transferência/genética , Ribonuclease P/química
2.
Arch Virol ; 165(11): 2541-2548, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32851430

RESUMO

Morphological, biological, serological, and molecular tests underpin the description of costus stripe mosaic virus (CoSMV) as a new member of the genus Potyvirus, family Potyviridae. Found affecting the native ornamental Costus spiralis in Brazil, the pathogen showed a severely restricted natural and experimental host range. Excluding the poly(A) tail, the CoSMV genome contains a large open reading frame (ORF) of 9,446 nucleotides that encodes a polyprotein with 3,046 amino acids, which is potentially cleaved into ten products, and a small ORF (77 amino acids) knows as PIPO. Genome analysis demonstrated the highest CoSMV nucleotide sequence identity to onion yellow dwarf virus (51.79%). No evidence of recombination was detected in the CoSMV genome, and phylogenetic analysis revealed its basal position in a group formed by members of the genus Potyvirus, along with Cyrtanthus elatus virus A (Vallota speciosa virus) and canna yellow streak virus. CoSMV was not transmitted by aphids of the species Aphis solanella, Myzus persicae or Uroleucon sonchi, which could be due to mutations in the HC-Pro motifs required for aphid transmission. A divergence in the P1 protein cleavage site was found when compared to other members of the family Potyviridae. Based on its unique biological and molecular characteristics and the current species demarcation criteria, we propose CoSMV to be a new tentative member of the genus Potyvirus.


Assuntos
Costus/virologia , Genoma Viral , Vírus do Mosaico/classificação , Filogenia , Sequência de Aminoácidos , Sequência de Bases , Brasil , Cisteína Endopeptidases/genética , Fases de Leitura Aberta , RNA Viral/genética , Análise de Sequência de DNA , Proteínas Virais/genética
3.
Plant Dis ; 104(10): 2713-2719, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32716275

RESUMO

Apple mosaic disease is widespread in the major apple-producing areas in China and is frequently associated with the presence of the newly identified Apple necrotic mosaic virus (ApNMV), belonging to subgroup 3 of Ilarvirus genus in the family of Bromoviridae. Mosaic symptoms were also observed in a hawthorn tree. Deep sequencing revealed the hawthorn tree with mosaic symptom was infected by ApNMV, which was confirmed by RT-PCR. The complete nucleotide sequences of RNA1 (3,378 nt), RNA2 (2,778 nt), and RNA3 (1,917 nt) of ApNMV from the hawthorn were obtained, sharing 93.8 to 96.8%, 89.7 to 96.1%, and 89.8 to 94.6% nucleotide identities with those from apples and crabapples, respectively. Two hypervariable regions were found, which showed 59.2 to 85.7% and 64.0 to 89.3% sequence identities at position 142 to 198 aa and at position 780 to 864 aa in the POL protein, respectively, between the hawthorn isolate and other isolates (apple, crabapple). A grafting test demonstrated that ApNMV was easily transmissible from hawthorns to apple trees, with severe chlorosis, yellowing, mosaic, curling, and necrosis. In addition, a total of 11,685 hawthorn trees were surveyed for the incidence of mosaic disease from five provinces in China, and only six were found showing typical mosaic symptoms. A total of 145 individual trees (six symptomatic, 68 asymptomatic, and 71 other symptoms) were tested for the presence or absence of ApNMV by RT-PCR. Among them, six symptomatic, four asymptomatic, and 10 other symptomatic trees tested positive for ApNMV. Taken together, these results demonstrated that the hawthorn tree was identified as a new natural host for ApNMV with a relatively low frequency (13.8%, 20 out of 145) in the main producing areas, and it was likely to be the causal pathogen of hawthorn mosaic disease.


Assuntos
Crataegus , Vírus do Mosaico , China , Filogenia , Doenças das Plantas , RNA-Seq , Árvores
4.
PLoS One ; 15(4): e0229196, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32294099

RESUMO

Citrus mosaic virus (CiMV) is one of the causal viruses of citrus mosaic disease in satsuma mandarins (Citrus unshiu). Prompt detection of trees infected with citrus mosaic disease is important for preventing the spread of this disease. Although rabbit monoclonal antibodies (mAbs) exhibit high specificity and affinity, their applicability is limited by technical difficulties associated with the hybridoma-based technology used for raising these mAbs. Here, we demonstrate a feasible CiMV detection system using a specific rabbit mAb against CiMV coat protein. A conserved peptide fragment of the small subunit of CiMV coat protein was designed and used to immunize rabbits. Antigen-specific antibody-producing cells were identified by the immunospot array assay on a chip method. After cloning of variable regions in heavy or light chain by RT-PCR from these cells, a gene set of 33 mAbs was constructed and these mAbs were produced using Expi293F cells. Screening with the AlphaScreen system revealed eight mAbs exhibiting strong interaction with the antigen peptide. From subsequent sequence analysis, they were grouped into three mAbs denoted as No. 4, 9, and 20. Surface plasmon resonance analysis demonstrated that the affinity of these mAbs for the antigen peptide ranged from 8.7 × 10-10 to 5.5 × 10-11 M. In addition to CiMV, mAb No. 9 and 20 could detect CiMV-related viruses in leaf extracts by ELISA. Further, mAb No. 20 showed a high sensitivity to CiMV and CiMV-related viruses, simply by dot blot analysis. The anti-CiMV rabbit mAbs obtained in this study are envisioned to be extremely useful for practical applications of CiMV detection, such as in a virus detection kit.


Assuntos
Anticorpos Monoclonais/biossíntese , Citrus/virologia , Vírus do Mosaico/isolamento & purificação , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/química , Especificidade de Anticorpos/imunologia , Proteínas do Capsídeo/imunologia , Cinética , Folhas de Planta/virologia , Coelhos
5.
Int J Mol Sci ; 21(6)2020 Mar 13.
Artigo em Inglês | MEDLINE | ID: mdl-32183174

RESUMO

The plant nonexpressor of pathogenesis-related 1 (NPR1) and pathogenesis-associated 1 (PR1) genes play fundamental roles in plant immunity response, as well as abiotic-stress tolerance. Nevertheless, comprehensive identification and characterization of NPR1 and PR1 homologs has not been conducted to date in Cymbidium orchids, a valuable industrial crop cultivated as ornamental and medicinal plants worldwide. Herein, three NPR1-like (referred to as CsNPR1-1, CsNPR1-2, and CsNPR1-3) and two PR1-like (CsPR1-1 and CsPR1-2) genes were genome-widely identified from Cymbidium orchids. Sequence and phylogenetic analysis revealed that CsNPR1-1 and CsNPR1-2 were grouped closest to NPR1 homologs in Zea mays (sharing 81.98% identity) and Phalaenopsis (64.14%), while CsNPR1-3 was classified into a distinct group with Oryza sativa NPR 3 (57.72%). CsPR1-1 and CsPR1-2 were both grouped closest to Phalaenopsis PR1 and other monocot plants. Expression profiling showed that CsNPR1 and CsPR1 were highly expressed in stem/pseudobulb and/or flower. Salicylic acid (SA) and hydrogen peroxide (H2O2) significantly up-regulated expressions of CsNPR1-2, CsPR1-1 and CsPR1-2, while CsNPR1-3, CsPR1-1 and CsPR1-2 were significantly up-regulated by abscisic acid (ABA) or salinity (NaCl) stress. In vitro transcripts of entire Cymbidium mosaic virus (CymMV) genomic RNA were successfully transfected into Cymbidium protoplasts, and the CymMV infection up-regulated the expression of CsNPR1-2, CsPR1-1 and CsPR1-2. Additionally, these genes were transiently expressed in Cymbidium protoplasts for subcellular localization analysis, and the presence of SA led to the nuclear translocation of the CsNPR1-2 protein, and the transient expression of CsNPR1-2 greatly enhanced the expression of CsPR1-1 and CsPR1-2. Collectively, the CsNPR1-2-mediated signaling pathway is SA-dependent, and confers to the defense against CymMV infection in Cymbidium orchids.


Assuntos
Ácido Abscísico/farmacologia , Orchidaceae/genética , Reguladores de Crescimento de Planta/farmacologia , Proteínas de Plantas/genética , Estresse Salino , Regulação da Expressão Gênica de Plantas , Peróxido de Hidrogênio/farmacologia , Vírus do Mosaico/patogenicidade , Orchidaceae/efeitos dos fármacos , Orchidaceae/virologia , Proteínas de Plantas/metabolismo , Salicilatos/farmacologia , Homologia de Sequência , Transcriptoma
6.
Phytopathology ; 110(7): 1352-1361, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32202482

RESUMO

Shiso (Perilla frutescens var. crispa) is widely grown as an important vegetable or herb crop in Japan. Beginning around the year 2000, occurrences of severe mosaic symptoms on shiso were documented and gradually spread across Kochi Prefecture, one of four major shiso production areas in Japan. Next generation sequencing and cloning indicated the presence of a previously unknown virus related to the members of the genus Emaravirus, for which we proposed the name Perilla mosaic virus (PerMV). The genome of PerMV consists of 10 RNA segments, each encoding a single protein in the negative-sense orientation. Of these proteins, P1, P2, P3a, P3b, P4, and P5 show amino acid sequence similarities with those of known emaraviruses, whereas no similarities were found in P6a, P6b, P6c, and P7. Characteristics of the RNA segments as well as phylogenetic analysis of P1 to P4 indicate that PerMV is a distinct and highly divergent emaravirus. Electron microscopy observations and protein analyses corresponded to presence of an emaravirus. Transmission experiments demonstrated that an eriophyid mite, Shevtchenkella sp. (family Eriophyidae), transmits PerMV with a minimum 30-min acquisition access period. Only plants belonging to the genus Perilla tested positive for PerMV, and the plant-virus-vector interactions were evaluated. The nucleotide sequences reported here are available in the DDBJ/ENA/GenBank databases under accession numbers LC496090 to LC496099.


Assuntos
Vírus do Mosaico , Perilla , Animais , Japão , Filogenia , Doenças das Plantas
7.
Phytopathology ; 110(1): 68-79, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31631806

RESUMO

High-throughput sequencing technologies were used to identify plant viruses in cereal samples surveyed from 2012 to 2017. Fifteen genome sequences of a tenuivirus infecting wheat, oats, and spelt in Estonia, Norway, and Sweden were identified and characterized by their distances to other tenuivirus sequences. Like most tenuiviruses, the genome of this tenuivirus contains four genomic segments. The isolates found from different countries shared at least 92% nucleotide sequence identity at the genome level. The planthopper Javesella pellucida was identified as a vector of the virus. Laboratory transmission tests using this vector indicated that wheat, oats, barley, rye, and triticale, but none of the tested pasture grass species (Alopecurus pratensis, Dactylis glomerata, Festuca rubra, Lolium multiflorum, Phleum pratense, and Poa pratensis), are susceptible. Taking into account the vector and host range data, the tenuivirus we have found most probably represents European wheat striate mosaic virus first identified about 60 years ago. Interestingly, whereas we were not able to infect any of the tested cereal species mechanically, Nicotiana benthamiana was infected via mechanical inoculation in laboratory conditions, displaying symptoms of yellow spots and vein clearing evolving into necrosis, eventually leading to plant death. Surprisingly, one of the virus genome segments (RNA2) encoding both a putative host systemic movement enhancer protein and a putative vector transmission factor was not detected in N. benthamiana after several passages even though systemic infection was observed, raising fundamental questions about the role of this segment in the systemic spread in several hosts.


Assuntos
Genoma Viral , Vírus do Mosaico , Vírus de Plantas , Animais , Grão Comestível/virologia , Genoma Viral/genética , Hemípteros/virologia , Vírus do Mosaico/genética , Noruega , Doenças das Plantas/virologia , Vírus de Plantas/genética , Suécia
8.
Theor Appl Genet ; 133(1): 217-226, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31587088

RESUMO

KEY MESSAGE: A cytological map of Haynaldia villosa chromosome arm 4VS was constructed to facilitate the identification and utilization of beneficial genes on 4VS. Induction of wheat-alien chromosomal structure aberrations not only provides new germplasm for wheat improvement, but also allows assignment of favorable genes to define physical regions. Especially, the translocation or introgression lines carrying alien chromosomal fragments with different sizes are useful for breeding and alien gene mapping. Chromosome arm 4VS of Haynaldia villosa (L.) Schur (syn. Dasypyrum villosum (L.) P. Candargy) confers resistances to eyespot and wheat yellow mosaic virus (WYMV). In this research, we used both irradiation and the pairing homoeologous gene (Ph) mutant to induce chromosomal aberrations or translocations. By using the two approaches, a structural aberration library of chromosome arm 4VS was constructed. In this library, there are 57 homozygous structural aberrations, in which, 39 were induced by the Triticum aestivum cv. Chinese Spring (CS) ph1b mutant (CS ph1b) and 18 were induced by irradiation. The aberrations included four types, i.e., terminal translocation, interstitial translocation, deletion and complex structural aberration. The 4VS cytological map was constructed by amplification in the developed homozygous aberrations using 199 4VS-specific markers, which could be allocated into 39 bins on 4VS. These bins were further assigned to their corresponding physical regions of chromosome arm 4DS based on BLASTn search of the marker sequences against the reference sequence of Aegilops tauschii Cosson. The developed genetic stocks and cytological map provide genetic stocks for wheat breeding as well as alien gene tagging.


Assuntos
Mapeamento Cromossômico , Cromossomos de Plantas/genética , Biblioteca Gênica , Triticum/citologia , Triticum/genética , Análise Citogenética , Resistência à Doença/genética , Genes de Plantas , Loci Gênicos , Marcadores Genéticos , Íons , Vírus do Mosaico/fisiologia , Doenças das Plantas/genética , Doenças das Plantas/virologia , Recombinação Genética/genética , Homologia de Sequência do Ácido Nucleico , Triticum/virologia
9.
Plant Biotechnol J ; 18(6): 1384-1395, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-31769589

RESUMO

Isoflavonoids, which include a variety of secondary metabolites, are derived from the phenylpropanoid pathway and are distributed predominantly in leguminous plants. These compounds play a critical role in plant-environment interactions and are beneficial to human health. Isoflavone synthase (IFS) is a key enzyme in isoflavonoid synthesis and shares a common substrate with flavanone-3-hydroxylase (F3H) and flavone synthase II (FNS II). In this study, CRISPR/Cas9-mediated multiplex gene-editing technology was employed to simultaneously target GmF3H1, GmF3H2 and GmFNSII-1 in soya bean hairy roots and plants. Various mutation types and frequencies were observed in hairy roots. Higher mutation efficiencies were found in the T0 transgenic plants, with a triple gene mutation efficiency of 44.44%, and these results of targeted mutagenesis were stably inherited in the progeny. Metabolomic analysis of T0 triple-mutants leaves revealed significant improvement in isoflavone content. Compared with the wild type, the T3 generation homozygous triple mutants had approximately twice the leaf isoflavone content, and the soya bean mosaic virus (SMV) coat protein content was significantly reduced by one-third after infection with strain SC7, suggesting that increased isoflavone content enhanced the leaf resistance to SMV. The isoflavone content in the seeds of T2 triple mutants was also significantly increased. This study provides not only materials for the improvement of soya bean isoflavone content and resistance to SMV but also a simple system to generate multiplex mutations in soya bean, which may be beneficial for further breeding and metabolic engineering.


Assuntos
Isoflavonas , Vírus do Mosaico , Sistemas CRISPR-Cas/genética , Engenharia Metabólica , Plantas Geneticamente Modificadas/genética , Soja/genética
10.
Mol Plant Microbe Interact ; 33(3): 412-422, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31841359

RESUMO

Viral suppressors of RNA silencing (VSRs) are a cluster of viral proteins that have evolved to counteract eukaryotic antiviral RNA silencing pathways, thereby contributing to viral pathogenicity. In this study, we revealed that the matrix protein P4 encoded by rice stripe mosaic virus (RSMV), which is an emerging cytoplasmic rhabdovirus, is a weak RNA silencing suppressor. By conducting yeast two-hybrid, bimolecular fluorescence complementation, and subcellular colocalization assays, we proved that P4 interacts with the rice endogenous suppressor of gene silencing 3 (OsSGS3). We also determined that P4 overexpression has no effect on OsSGS3 transcription. However, P4 can promote the degradation of OsSGS3 via ubiquitination and autophagy. Additionally, a potato virus X-based expression system was used to confirm that P4 enhances the development of mosaic symptoms on Nicotiana benthamiana leaves by promoting hydrogen peroxide accumulation but not cell death. To verify whether P4 is a pathogenicity factor in host plants, we generated transgenic P4-overexpressing rice plants that exhibited disease-related developmental defects including decreased plant height and excessive tillering. Our data suggest that RSMV-encoded P4 serves as a weak VSR that inhibits antiviral RNA silencing by targeting OsSGS3.


Assuntos
Inativação Gênica , Vírus do Mosaico/patogenicidade , Doenças das Plantas/virologia , Interferência de RNA , Proteínas da Matriz Viral/genética , Autofagia , Oryza/genética , Oryza/virologia , Proteínas de Plantas , Plantas Geneticamente Modificadas , Potexvirus , Tabaco , Ubiquitinação
11.
Arch Virol ; 165(2): 491-494, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31784911

RESUMO

A new macluravirus infecting Pinellia ternata in China was identified by high-throughput sequencing (HTS) and tentatively named "crow-dipper mosaic-associated virus" (CrdMV). The complete genome sequence of CrdMV was determined by reverse transcription (RT) PCR and rapid amplification of cDNA ends (RACE) PCR. The genomic RNA of CrdMV consists of 8,454 nucleotides (nt), excluding the poly(A) tail at the 3' end. CrdMV has a genomic structure typical of macluraviruses, with large open reading frame encoding a polyprotein of 2,696 amino acids (aa). CrdMV shares 54.40%-59.37% nt sequence identity at the genome sequence level, 48.00%-58.58% aa sequence identity, at the polyprotein sequence level and 37.27%-49.22% aa sequence identity at the CP sequence level with other members of the genus Macluravirus. These values are well below the species demarcation threshold for the family Potyviridae. Phylogenetic analysis based on the amino acid sequences of polyproteins confirmed that CrdMV clusters closely with broad-leafed dock virus A (BDVA, GenBank accession no. KU053507). These results suggest that CrdMV should be considered a distinct member of the genus Macluravirus.


Assuntos
Genoma Viral/genética , Vírus do Mosaico/genética , Pinellia/virologia , Potyviridae/genética , Vírus Satélites/genética , Sequência de Aminoácidos , Animais , China , Corvos/virologia , Tamanho do Genoma/genética , Genômica/métodos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Filogenia , Doenças das Plantas/virologia , Poliproteínas/genética , RNA Viral/genética , Proteínas Virais/genética
12.
Cienc. tecnol. salud ; 6(2): 98-106, jul dic 2019. ^c27 cmilus
Artigo em Espanhol | LILACS | ID: biblio-1095831

RESUMO

El complejo de mosca blanca (Hemiptera: Aleyrodidae) incluye algunas de las principales plagas del ejote francés (Phaseolus vulgaris L.). Dentro de las cuales, Bemisia tabaci es vector del virus del mosaico dorado que afecta la calidad y rendimiento del cultivo, con pérdidas hasta del 100% y un control difícil debido a la resistencia adquirida por las plagas hacia algunos agroquímicos. El ejote francés ocupa el segundo lugar entre de los productos no tradicionales de exportación de Guatemala. Su manejo agronómico ha sido principalmente a través del control químico, el cual afecta insectos y otros organismos que no son el objetivo del control, tales como: polinizadores, insectos benéficos, humanos y fauna silvestre. Los objetivos del estudio fueron: determinar la presencia de enemigos naturales nativos de la mosca blanca e identificar las especies de mosca blanca presentes en el cultivo del ejote francés en Chimaltenango. Para el estudio se establecieron cuatro parcelas de 300 m², se realizaron muestreos semanales durante dos ciclos del cultivo. En cada parcela se muestrearon cinco sitios y en cada sitio cinco plantas. Las especies de parasitoides nativos encontrados fueron: Encarsia Formosa Gahan, Eretmocerus eremicus Rose y Zolnerowuch y Amitus fuscipennis MacGown y Nebeker, la especie más abundante fue A. fuscipennis. Los depredadores identificados fueron Chrysoperla carnea (Stephens) e Hippodamia convergens Guerin-Meneville. La especie más abundante fue H. convergens. Estas especies podrían ser herramientas valiosas para ser empleadas en programas de control biológico, producciones orgánicas o en programas de manejo integrado de plagas.


The whitefly complex (Hemiptera: Aleyrodidae) includes some of the main pests of the French green bean (Phaseolus vulgaris L.). Among which, Bemisia tabaci is a vector of the golden mosaic virus that affects the quality and yield of the crop, with losses up to 100% and difficult control due to the resistance acquired by pests towards some agrochemicals. The French green bean ranks second among the non-traditional export products of Guatemala. Its agronomic management has been mainly through chemical control, which affects insects and other organisms that are not the objective of the control, such as: pollinators, beneficial insects, humans and wildlife. The objectives of the study were: to determine the presence of natural enemies native to the whitefly and identify the species of whitefly present in the cultivation of the French bean in Chimaltenango. For the study, four 300 m² plots were established, weekly sampling was carried out during two crop cycles. Five sites were sampled on each plot and five plants on each site. The native parasitoid species found were: Encarsia Formosa Gahan, Eretmocerus eremicus Rose and Zolnerowuch and Amitus fuscipennis MacGown and Nebeker, the most abundant species was A. fuscipennis. The predators identified were Chrysoperla carnea (Stephens) and Hippodamia convergens Guerin-Meneville. The most abundant species was H. convergens. These species could be valuable tools to be used in biological control programs, organic productions or in integrated pest management programs.


Assuntos
Animais , Phaseolus/crescimento & desenvolvimento , Phaseolus/parasitologia , Hemípteros/crescimento & desenvolvimento , Hemípteros/parasitologia , Controle Biológico de Vetores/métodos , Vírus do Mosaico
13.
Virus Genes ; 55(6): 854-858, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31605266

RESUMO

A virus isolate from tabasco pepper (Capsicum frutescens) has been reported as a strain of the comovirus Andean potato mottle virus (APMoV). Using the replicative intermediate viral dsRNA, the pepper virus strain was sequenced by Illumina MiSeq. The viral genome was de novo assembled resulting in two RNAs with lengths of 6028 and 3646 nt. Nucleotide sequence analysis indicated that they corresponded to the RNA-1 and RNA-2 of a novel comovirus which we tentatively named pepper mild mosaic virus (PepMMV). Predictions of the open reading frame (ORF) of RNA-1 resulted in a single ORF of 5871 nt with five cistrons typical of comoviruses, cofactor proteinase, helicase, viral protein genome-linked, 3C-like proteinase (Pro), and RNA-dependent RNA polymerase (RdRP). Similarly, sequence analysis of RNA-2 resulted in a single ORF of 3009 nt with two cistrons typical of comoviruses: movement protein and coat protein (large coat protein and small coat proteins). In pairwise amino acid sequence alignments using the Pro-Pol protein, PepMMV shared the closest identities with broad bean true mosaic virus and cowpea mosaic virus, 56% and 53.9% respectively. In contrast, in alignments of the amino acid sequence of the coat protein (small and large coat proteins) PepMMV shared the closest identities to APMoV and red clover mottle virus, 54% and 40.9% respectively. A phylogenetic tree constructed using the conserved domains for the Pro-Pol from all members of the family Secoviridae confirmed the comovirus nature of the virus. Phylogenetic and sequence analyses supports proposing PepMMV as a new species of the genus Comovirus.


Assuntos
Comovirus/genética , Genoma Viral/genética , Sequenciamento Completo do Genoma , Sequência de Aminoácidos/genética , Capsicum/genética , Capsicum/virologia , Anotação de Sequência Molecular , Vírus do Mosaico/genética , Fases de Leitura Aberta/genética , Filogenia , RNA de Cadeia Dupla/genética , RNA Viral/genética , /genética
14.
Arch Virol ; 164(11): 2805-2810, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31451963

RESUMO

Physalis peruviana is a perennial solanaceous plant that has recently been established as a commercial crop in Brazil. This work reports the near-complete genome sequence, particle morphology, and plant host responses to a putative new sobemovirus, named "physalis rugose mosaic virus". The virus, characterized by isometric particles of ca. 30 nm in diameter, causes foliar symptoms of mosaic, malformation and blistering, accompanied by stunting. The near-complete genome sequence comprises 4175 nucleotides and contains five open reading frames that are similar to those of other sobemoviruses. In addition to P. peruviana, the new virus systemically infected Capsicum annuum, Nicotiana tabacum and Solanum lycopersicum by mechanical inoculation. Thus, this virus may cause disease in these crops in the field.


Assuntos
Genoma Viral/genética , Vírus do Mosaico/classificação , Vírus do Mosaico/crescimento & desenvolvimento , Physalis/virologia , Doenças das Plantas/virologia , Vírus de Plantas/classificação , Vírus de Plantas/genética , Brasil , Capsicum/virologia , Lycopersicon esculentum/virologia , Vírus do Mosaico/genética , Vírus de Plantas/crescimento & desenvolvimento , RNA Viral/genética , Tabaco/virologia
15.
Int J Mol Sci ; 20(13)2019 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-31252649

RESUMO

The synergistic infection of maize chlorotic mottle virus (MCMV) and sugarcane mosaic virus (SCMV) causes maize lethal necrosis, with considerable losses to global maize production. microRNAs (miRNAs) are conserved non-coding small RNAs that play essential regulatory roles in plant development and environmental stress responses, including virus infection. However, the characterization of maize miRNAs in response to synergistic infection of MCMV and SCMV remains largely unknown. In this study, the profiles of small RNAs from MCMV and SCMV single- and co-infected (S + M) maize plants were obtained by high-throughput sequencing. A total of 173 known miRNAs, belonging to 26 miRNA families, and 49 novel miRNAs were profiled. The expression patterns of most miRNAs in S + M-infected maize plants were similar to that in SCMV-infected maize plants, probably due to the existence of RNA silencing suppressor HC-Pro. Northern blotting and quantitative real-time PCR were performed to validate the accumulation of miRNAs and their targets in different experimental treatments, respectively. The down-regulation of miR159, miR393, and miR394 might be involved in antiviral defense to synergistic infection. These results provide novel insights into the regulatory networks of miRNAs in maize plants in response to the synergistic infection of MCMV and SCMV.


Assuntos
MicroRNAs/genética , Vírus do Mosaico/patogenicidade , Doenças das Plantas/genética , Tombusviridae/patogenicidade , Zea mays/genética , Regulação da Expressão Gênica de Plantas , Doenças das Plantas/virologia , Zea mays/virologia
16.
Virology ; 535: 20-31, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31254744

RESUMO

High Plains wheat mosaic virus (genus Emaravirus), an octapartite negative-sense RNA virus, encodes two RNA silencing suppressors, P7 and P8. In this study, we found that P7 and P8 efficiently delayed the onset of dsRNA-induced transitive pathway of RNA silencing. Electrophoretic mobility shift assays (EMSA) revealed that only P7 protected long dsRNAs from dicing in vitro and bound weakly to 21- and 24-nt PTGS-like ds-siRNAs. In contrast, P8 bound strongly and relatively weakly to 21- and 24-nt ds-siRNAs, respectively, suggesting size-specific binding. In EMSA, neither protein bound to 180-nt and 21-nt ssRNAs at detectable levels. Sequence analysis revealed that P7 contains a conserved GW motif. Mutational disruption of this motif resulted in loss of suppression of RNA silencing and pathogenicity enhancement, and failure to complement the silencing suppression-deficient wheat streak mosaic virus. Collectively, these data suggest that P7 and P8 proteins utilize distinct mechanisms to overcome host RNA silencing for successful establishment of systemic infection in planta.


Assuntos
Interações entre Hospedeiro e Microrganismos , Evasão da Resposta Imune , Vírus do Mosaico/imunologia , Vírus do Mosaico/patogenicidade , Interferência de RNA , Triticum/virologia , Proteínas Virais/metabolismo , Análise Mutacional de DNA , Ensaio de Desvio de Mobilidade Eletroforética , Proteínas Virais/genética
17.
Nat Nanotechnol ; 14(7): 712-718, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31110265

RESUMO

Large doses of chemical pesticides are required to achieve effective concentrations in the rhizosphere, which results in the accumulation of harmful residues. Precision farming is needed to improve the efficacy of pesticides, but also to avoid environmental pollution, and slow-release formulations based on nanoparticles offer one solution. Here, we tested the mobility of synthetic and virus-based model nanopesticides by combining soil column experiments with computational modelling. We found that the tobacco mild green mosaic virus and cowpea mosaic virus penetrate soil to a depth of at least 30 cm, and could therefore deliver nematicides to the rhizosphere, whereas the Physalis mosaic virus remains in the first 4 cm of soil and would be more useful for the delivery of herbicides. Our experiments confirm that plant viruses are superior to synthetic mesoporous silica nanoparticles and poly(lactic-co-glycolic acid) for the delivery and controlled release of pesticides, and could be developed as the next generation of pesticide delivery systems.


Assuntos
Agricultura/métodos , Preparações de Ação Retardada/metabolismo , Vírus do Mosaico/metabolismo , Praguicidas/metabolismo , Microbiologia do Solo , Comovirus/metabolismo , Nanopartículas/metabolismo , Solo/química , Vírus do Mosaico do Tabaco/metabolismo , Tymovirus/metabolismo
18.
Phytopathology ; 109(8): 1475-1480, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30951441

RESUMO

Wheat yellow mosaic virus (WYMV) belongs to the genus Bymovirus in the family Potyviridae and has a bipartite genome (RNA1 and RNA2). WYMV in Japan is classified into three pathotypes (I to III) based on its pathogenicity to wheat cultivars. Among these three, pathotypes I and II are discriminated by their pathogenicity to the wheat cultivar Fukuho; pathotype I infects Fukuho but pathotype II does not. In the present study, the genomic regions that are involved in such pathogenicity were examined using infectious viral cDNA clones of pathotypes I and II. Reassortant experiments between viral RNA1 and RNA2 revealed the presence of a viral factor related to pathogenicity in RNA1. A chimeric pathotype II virus harboring a cylindrical inclusion (CI) cistron from pathotype I facilitated systemic infection of Fukuho, indicating that CI protein is involved in pathogenicity. Furthermore, analysis of chimeric and site-directed mutants revealed that three amino acids at the N-terminal region of CI protein were involved in pathogenicity to Fukuho. On the other hand, at the single-cell level, pathotype II replicated in protoplasts of Fukuho similar to that of pathotype I virus. These data suggest that differential pathogenicity between pathotypes I and II was considered to depend on the ability of cell-to-cell or long-distance viral movement, in which CI protein is involved. To the best of our knowledge, this is the first report to show the involvement of the bymoviral CI protein in pathogenicity.


Assuntos
Vírus do Mosaico , Potyviridae , Triticum/virologia , Japão , Doenças das Plantas , RNA Viral
19.
Virol J ; 16(1): 23, 2019 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-30786887

RESUMO

BACKGROUND: Polyadenylation influences many aspects of mRNA as well as viral RNA. variable polyadenylation at the 3' end have been reported in RNA viruses. It is interesting to identify the characteristic and potential role of 3' polyadenylation of Wheat yellow mosaic virus (WYMV), which has been reported to contain two genomic RNAs with 3' poly(A) tails and caused severe disease on wheat in East Asia region. METHODS: 3' RACE was used to identify sequences of the 3' end in WYMV RNAs from naturally infected wheat by WYMV. In vitro translation assay was performed to analyze effect of UTRs of WYMV with or without 3'polyadenylation on translation. In vitro replication mediated by WYMV NIb protein were performed to evaluate effect of variable polyadenylation on replication. RESULTS: Variable polyadenylation in WYMV RNAs was identified via 3' RACE. WYMV RNAs in naturally infected wheat in China simultaneously present with regions of long, short, or no adenylation at the 3' ends. The effects of variable polyadenylation on translation and replication of WYMV RNAs were evaluated. 5'UTR and 3'UTR of WYMV RNA1 or RNA2 synergistically enhanced the translation of the firefly luciferase (Fluc) gene in in vitro WGE system, whereas additional adenylates had an oppositive effect on this enhancement on translation mediated by UTRs of WYMV. Additional adenylates remarkably inhibited the synthesis of complementary strand from viral genome RNA during the in vitro replication mediated by WYMV NIb protein. CONCLUSIONS: 3' end of WYMV RNAs present variable polyadenylation even no polyadenylation. 3' polyadenylation have opposite effect on translation mediated by UTRs of WYMV RNA1 or RNA2. 3' polyadenylation have negative effect on minus-strand synthesis of WYMV RNA in vitro. Variable polyadenylation of WYMV RNAs may provide sufficient selection on the template for translation and replication.


Assuntos
Vírus do Mosaico/genética , Poliadenilação , Triticum/virologia , Replicação Viral , China , Vírus do Mosaico/fisiologia , Doenças das Plantas/virologia , Sinais de Poliadenilação na Ponta 3' do RNA/genética , RNA Viral/genética
20.
Mol Biotechnol ; 61(2): 84-92, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30484145

RESUMO

The aim of this study is to assess the effect of methyl jasmonate (MeJA) and temperature on the valuable pharmaceuticals expression in a virus-mediated transient expression system, and so the Zuchini Yellow Mosaic Virus (ZYMV) based vector was used for transferring the GFP reporter gene and recombinant tissue plasminogen activator (rtPA) gene (K2S) to cucurbit (Cucurbita pepo L.). MeJA, temperature and time (days after inoculation), were evaluated as a factorial experiment in a completely randomized design (CRD). At first, the effect of all treatment combinations on GFP expression was assessed. At this step, the ELISA test was used to select the optimum treatment combination. ELISA method revealed the significant difference between applied treatments. The optimized treatment significantly increased the expression of rtPA compared to the control. The Real-Time PCR reaction for both GFP and rtPA genes showed no significant differences between optimum and control treatments, however, transcripts of the small subunit of RuBisCO were extremely down-regulated in optimum treatment condition. Reduction in RuBisCO expression at protein level was tangible under treatment condition based on the ELISA test. Therefore, it can be inferred that suppressing the expression of RuBisCO, probably resulted in higher access of expression system to free amino acids inside the cell. In this study, MeJA has been shown to be a positive factor, but the low temperature (17 °C), unlike previous studies, suppressed the expression of recombinant protein unexpectedly, probably due to the incompatibility of the viral construct with low temperature. In conclusion, the use of a suitable gene construct, which is not sensitive to temperature, is likely to result in a more favorable outcome.


Assuntos
Acetatos/farmacologia , Cucurbita/genética , Ciclopentanos/farmacologia , Expressão Gênica/efeitos dos fármacos , Oxilipinas/farmacologia , Reguladores de Crescimento de Planta/farmacologia , Ativador de Plasminogênio Tecidual/genética , Regulação da Expressão Gênica de Plantas , Genes Reporter/genética , Vetores Genéticos , Agricultura Molecular , Vírus do Mosaico/genética , Plantas Geneticamente Modificadas , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Ribulose-Bifosfato Carboxilase/genética , Ribulose-Bifosfato Carboxilase/metabolismo , Temperatura , Ativador de Plasminogênio Tecidual/metabolismo
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