An Inexpensive System for Rapid and Accurate On-site Detection of Garlic-Infected Viruses by Agarose Gel Electrophoresis Followed by Array Assay.

Garlic can be infected by a variety of viruses, but mixed infections with leek yellow stripe virus, onion yellow dwarf virus, and allexiviruses are the most damaging, so an easy, inexpensive on-site method to simultaneously detect at least these three viruses with a certain degree of accuracy is needed to produce virus-free plants. The most common laboratory method for diagnosis is multiplex reverse transcription polymerase chain reaction (RT-PCR). However, allexiviruses are highly diverse even within the same species, making it difficult to design universal PCR primers for all garlic-growing regions in the world.

Generation and Retention of Defective RNA3 from Cucumber Mosaic Virus and Relevance of the 2b Protein to Generation of the Subviral RNA.

A defective RNA3 (D3Yα) of strain Y of cucumber mosaic virus (CMV-Y) was examined on host-specific maintenance, experimental conditions, and a viral factor required for its generation in plants. D3Yα was stably maintained in cucumber but not in tomato plants for 28 days post inoculation (dpi). D3Yα was generated in Nicotiana tabacum or N.

Metagenomic Analyses of Viruses in the Orchid Mycorrhizal Interaction Using Improved Assemble Tools.

In nature, mycorrhizal association with soil-borne fungi is indispensable for orchid species. Compatible mycorrhizal fungi form endo-mycorrhizal structures in orchid cells, and the fungal structures are digested in orchid cells to be supplied to orchids as nutrition. Because orchid seeds lack the reserves for germination, they keep receiving nutrition through mycorrhizal formation from seed germination until nonphotosynthetic young seedlings develop leaves and become photoautotrophic.

Host-specific adaptation drove the coevolution of leek yellow stripe virus and plants.

Host adaptation plays a crucial role in virus evolution and is a consequence of long-term interactions between virus and host in a complex arms race between host RNA silencing and viral RNA silencing suppressor (RSS) as counterdefense. Leek yellow stripe virus (LYSV), a potyvirus causing yield loss of garlic, infects several species of plants. The unexpected discovery of an interspecific hybrid of garlic, leek, and great-headed (GH) garlic motivated us to explore the host-adaptive evolution of LYSV.

VIGS as a strategy to reverse aphid wing induction by Y-satellite RNA of cucumber mosaic virus.

Y-satellite RNA (Y-sat) of cucumber mosaic virus upregulates the expression of the aphid ABCG4 gene, which promotes aphid wing formation. We used ABCG4 virus-induced gene silencing (VIGS) to prevent the wing-induction mechanism of Y-sat and thus inhibited aphid wing formation. Of the aphids on plants with VIGS of ABCG4, only about 30% had wings, and 60-70% of the winged aphids were small and likely impaired in flying ability.

Two mutually exclusive evolutionary scenarios for allexiviruses that overcome host RNA silencing and autophagy by regulating viral CRP expression.

The genus Allexivirus currently includes eight virus species that infect allium plants. Previously, we showed that there are two distinct groups of allexiviruses (deletion [D]-type and insertion [I]-type) based on the presence or absence of a 10- to 20-base insert (IS) between the coat protein (CP) and cysteine rich protein (CRP) genes. In the present study of CRPs to analyze their functions, we postulated that evolution of allexiviruses may have been largely directed by CRPs and thus proposed two evolutionary scenarios for allexiviruses based mainly on the presence or absence of IS and determined by how the allexiviruses challenge host resistance mechanisms (RNA silencing and autophagy).

Battle for control of anthocyanin biosynthesis in two Brassicaceae species infected with turnip mosaic virus.

It has previously been found that turnip mosaic virus (TuMV) greatly suppresses anthocyanin accumulation (AA) in Brassica rapa leaves, and that such leaves become infected whilst anthocyanin-enriched leaves on the same plants are rarely infected. To clarify whether AA is a defense against TuMV, in this study we examined tissue-level patterns of spontaneous AA in relation to the cellular localization of a TuMV strain that expresses a yellow fluorescent protein. We found that TuMV infection was significantly blocked by AA, suggesting that it functions as a chemical barrier against TuMV.

Diversity of viral RNA silencing suppressors and their involvement in virus-specific symptoms.

RNA silencing is an evolutionarily conserved and homology-dependent gene inactivation system that regulates most biological processes at either the transcriptional or post-transcriptional level. In plants, insects and certain mammalian systems, RNA silencing constitutes the basis of the antiviral defense mechanism. To counteract RNA silencing-based antiviral responses viruses adopt strategies of replication and host invasion that include mechanisms of RNA silencing suppression.

Leek Yellow Stripe Virus Can Adjust for Host Adaptation by Trimming the N-Terminal Domain to Allow the P1 Protein to Function as an RNA Silencing Suppressor.

In Japan, the P1 protein (S-type) encoded by leek yellow stripe virus (LYSV) isolates detected in Honshu and southward is shorter than the P1 (N-type) of LYSV isolates from garlic grown in Hokkaido due to a large deletion in the N-terminal half. In garlic fields in Hokkaido, two types of LYSV isolate with N- and S-type P1s are sometimes found in mixed infections. In this study, we confirmed that N- and S-type P1 sequences were present in the same plant and that they belong to different evolutionary phylogenetic groups.

Coat protein of partitiviruses isolated from mycorrhizal fungi functions as an RNA silencing suppressor in plants and fungi.

Orchid seeds depend on colonization by orchid mycorrhizal (OM) fungi for their germination; therefore, the orchids and OM fungi have long maintained a close relationship (e.g., formation of the hyphal mass structure, peloton) during their evolution.

Importin/exportin-mediated nucleocytoplasmic shuttling of cucumber mosaic virus 2b protein is required for 2b's efficient suppression of RNA silencing.

The 2b protein (2b) of cucumber mosaic virus (CMV), an RNA-silencing suppressor (RSS), is a major pathogenicity determinant of CMV. 2b is localized in the nucleus and cytoplasm, and its nuclear import is determined by two nuclear localization signals (NLSs); a carrier protein (importin [IMPα]) is predicted to be involved in 2b's nuclear transport. Cytoplasmic 2bs play a role in suppression of RNA silencing by binding to small RNAs and AGO proteins.

Progressive DNA demethylation in epigenetic hybrids between parental plants with and without methylation of the transgene promoter.

Crosses of parents that differ in their DNA methylation states leads to progressive demethylation in the F hybrids. In plant breeding research, hybrid vigor in F hybrids is known to be a very important phenomenon. Hybrid vigor, or heterosis, refers to the fact that F hybrids from crosses with a certain combination of parents have traits that are superior to those of the parents.

A plant virus satellite RNA directly accelerates wing formation in its insect vector for spread.

Cucumber mosaic virus (CMV) often accompanies a short RNA molecule called a satellite RNA (satRNA). When infected with CMV in the presence of Y-satellite RNA (Y-sat), tobacco leaves develop a green mosaic, then turn yellow. Y-sat has been identified in the fields in Japan.

Resistance induction based on the understanding of molecular interactions between plant viruses and host plants.

Background: Viral diseases cause significant damage to crop yield and quality. While fungi- and bacteria-induced diseases can be controlled by pesticides, no effective approaches are available to control viruses with chemicals as they use the cellular functions of their host for their infection cycle. The conventional method of viral disease control is to use the inherent resistance of plants through breeding.

Artificially Edited Alleles of the Eukaryotic Translation Initiation Factor 4E1 Gene Differentially Reduce Susceptibility to Cucumber Mosaic Virus and Potato Virus Y in Tomato.

Eukaryotic translation initiation factors, including eIF4E, are susceptibility factors for viral infection in host plants. Mutation and double-stranded RNA-mediated silencing of tomato genes can confer resistance to viruses, particularly members of the genus. Here, we artificially mutated the gene on chromosome 3 of a commercial cultivar of tomato ( L.

RNA silencing-related genes contribute to tolerance of infection with potato virus X and Y in a susceptible tomato plant.

Background: In plants, the RNA silencing system functions as an antiviral defense mechanism following its induction with virus-derived double-stranded RNAs. This occurs through the action of RNA silencing components, including Dicer-like (DCL) nucleases, Argonaute (AGO) proteins, and RNA-dependent RNA polymerases (RDR). Plants encode multiple AGOs, DCLs, and RDRs.

Apple latent spherical virus (ALSV)-induced gene silencing in a medicinal plant, Lithospermum erythrorhizon.

Lithospermum erythrorhizon is a medicinal plant that produces shikonin, a red lipophilic naphthoquinone derivative that accumulates exclusively in roots. The biosynthetic steps required to complete the naphthalene ring of shikonin and its mechanism of secretion remain unclear. Multiple omics studies identified several candidate genes involved in shikonin production.

Virus-Induced Gene Silencing in Lilies Using Cucumber Mosaic Virus Vectors.

Virus-induced gene silencing (VIGS) systems are effective for rapid analysis of gene functions in plants that require a long period of growth such as Lilium. We successfully developed a VIGS system using the cucumber mosaic virus (HL strain, CMV-HL) vector to induce RNA silencing of the L. leichtlinii phytoene desaturase gene (LlPDS), where at 30 days postinoculation (dpi), photo-bleaching was observed in the upper leaves of L.

Application of Virus-Induced Gene Silencing to Arbuscular Mycorrhizal Fungi.

Downregulation of AM fungal genes using a plant viral vector is feasible. A partial sequence of a target fungal gene is cloned into the multicloning site of CMV2-A1 vector developed from RNA2 of Cucumber mosaic virus Y strain, and the RNA2, together with RNA1 and RNA3 of the virus, are in vitro-transcribed. Inoculation of Nicotiana benthamiana with these viral RNAs results in reconstitution of the virus in the plant, which triggers silencing of the fungal gene.

Application of cucumber mosaic virus to efficient induction and long-term maintenance of virus-induced gene silencing in spinach.

Virus-induced gene silencing (VIGS) is a useful tool for functional genomics in plants. In this study, we tried to apply cucumber mosaic virus (CMV) to efficient induction of VIGS in spinach. Although VIGS for spinach had been previously developed based on two viruses (beet curly top virus and tobacco rattle virus), they still have some problems with systemic movement and long-term maintenance of VIGS in spinach.

Role of salicylic acid glucosyltransferase in balancing growth and defence for optimum plant fitness.

Salicylic acid (SA), an essential secondary messenger for plant defence responses, plays a role in maintaining a balance (trade-off) between plant growth and resistance induction, but the detailed mechanism has not been explored. Because the SA mimic benzothiadiazole (BTH) is a more stable inducer of plant defence than SA after exogenous application, we analysed expression profiles of defence genes after BTH treatment to better understand SA-mediated immune induction. Transcript levels of the salicylic acid glucosyltransferase (SAGT) gene were significantly lower in BTH-treated Nicotiana tabacum (Nt) plants than in SA-treated Nt control plants, suggesting that SAGT may play an important role in SA-related host defence responses.

Transcriptional silencing of 35S driven-transgene is differentially determined depending on promoter methylation heterogeneity at specific cytosines in both plus- and minus-sense strands.

Background: De novo DNA methylation triggered by short interfering RNAs is called RNA-directed DNA methylation (RdDM). Transcriptional gene silencing (TGS) through RdDM can be induced using a viral vector. We have previously induced RdDM on the 35S promoter in the green fluorescent protein (GFP)-expressing Nicotiana benthamiana line 16c using the cucumber mosaic virus vector.

Rapid transient protein production by the coat protein-deficient cucumber mosaic virus vector: non-packaged CMV system, NoPaCS.

We developed a non-packaged CMV system (NoPaCS) for CMV-agroinfection with a virus-inescapable transgenic plant platform, enabling rapid, high production of a large-sequence target protein. For rapidly producing high levels of a desirable protein, many plant virus vectors have been developed. However, there is always a concern that such recombinant viruses may escape into the environment.

Metagenomic Analyses of the Viruses Detected in Mycorrhizal Fungi and Their Host Orchid.

In nature, mycorrhizal association with soilborne fungi is indispensable for orchid families. Fungal structures from compatible endo-mycorrhizal fungi in orchid cells are digested in cells to be supplied to orchids as nutrition. Because orchid seeds lack the reserves for germination, they keep receiving nutrition through mycorrhizal formation from seed germination until shoots develop (leaves) and become photoautotrophic.