記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Viruses  

Partitiviruses are one of the most prevalent double-stranded RNA viruses that have been identified mostly in filamentous fungi and plants. Partitiviruses generally infect host fungi asymptomatically but infrequently exert significant effect(s) on morphology and virulence, thus being considered a potential source of biological control agents against pathogenic fungi. In this study, we performed a screening for mycoviruses of a collection of Thai isolates of rice fungal pathogen Rhizoctonia oryzae-sativae, a causal agent of rice aggregated sheath spot disease. As a result, 36% of tested isolates carried potentially viral double-stranded RNAs with sizes ranging from 2 to 3 kbp. By conventional cDNA library construction and RNA-seq, we determined six new alphapartitiviruses that infected three isolates: tentatively named Rhizoctonia oryzae-sativae partitivirus 1 to 6 (RosPV1-6). Furthermore, RT-PCR detection of each virus revealed their omnipresent nature in different R. oryzaesativae isolates. Although virus-curing of basidiomycetous fungi is generally difficult, our repeated attempts successfully obtained virus-free (for RosPV1, RosPV2, and uncharacterized partitiviruses), isogenic strain of R. oryzae-sativae TSS190442. The virus-cured strain showed slightly faster colony growth on the synthetic media and severe symptom development on the rice sheath compared to its virus-infected counterpart. Overall, this study shed light on the distribution of partitiviruses in R. oryzae-sativae in a paddy environment and exemplified a virus-curing protocol that may be applicable for other basidiomycetous fungi.

DOI： 10.3390/v13112269

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Frontiers in Microbiology  

An increasing number of viruses are continuously being found in a wide range of organisms, including fungi. Recent studies have revealed a wide viral diversity in microbes and a potential importance of these viruses in the natural environment. Although virus exploration has been accelerated by short-read, high-throughput sequencing (HTS), and viral de novo sequencing is still challenging because of several biological/molecular features such as micro-diversity and secondary structure of RNA genomes. This study conducted de novo sequencing of multiple double-stranded (ds) RNA (dsRNA) elements that were obtained from fungal viruses infecting two Fusarium sambucinum strains, FA1837 and FA2242, using conventional HTS and long-read direct RNA sequencing (DRS). De novo assembly of the read data from both technologies generated near-entire genomic sequence of the viruses, and the sequence homology search and phylogenetic analysis suggested that these represented novel species of the Hypoviridae, Totiviridae, and Mitoviridae families. However, the DRS-based consensus sequences contained numerous indel errors that differed from the HTS consensus sequences, and these errors hampered accurate open reading frame (ORF) prediction. Although with its present performance, the use of DRS is premature to determine viral genome sequences, the DRS-mediated sequencing shows great potential as a user-friendly platform for a one-shot, whole-genome sequencing of RNA viruses due to its long-reading ability and relative structure-tolerant nature.

DOI： 10.3389/fmicb.2021.641484

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Virology  

Cryphonectria nitschkei chrysovirus 1 (CnCV1), was described earlier from an ascomycetous fungus, Cryphonectria nitschkei strain OB5/11, collected in Japan; its partial sequence was reported a decade ago. Complete sequencing of the four genomic dsRNA segments revealed molecular features similar to but distinct from previously reported members of the family Chrysoviridae. Unique features include the presence of a mini-cistron preceding the major large open reading frame in each genomic segment. Common features include the presence of CAA repeats in the 5′-untranslated regions and conserved terminal sequences. CnCV1-OB5/11 could be laterally transferred to C. nitschkei and its relatives C. radicalis and C. naterciae via coculturing, virion transfection and protoplast fusion, but not to fungal species other than the three species mentioned above, even within the genus Cryphonectria, suggesting a very narrow host range. Phenotypic comparison of a few sets of CnCV1-infected and -free isogenic strains showed symptomless infection in new hosts.

DOI： 10.1016/j.virol.2020.11.011

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Pathogens  

Rice orange leaf phytoplasma (ROLP) causes clear orange to yellowish leaf discoloration and severe stunting in rice seedlings. The ecological and biological characteristics of ROLP are largely unknown because the disease has not widely caused serious problems in rice cultivated areas, thereby leading to the low accumulation of research data. However, in the past decade, the disease became a threat to rice production, particularly in South China and India; it has also been recognised in other Asian countries, such as Vietnam, Thailand and the Philippines. Here, we observed the occurrence of ROLP in paddies of the Southeast Asian counties (Cambodia, Vietnam and the Philippines) and found that the isolates in the Philippines and Vietnam were monophyletic, while those in India, Thailand and Cambodia were more diverse, suggesting their potential origins. In Cambodia, it was revealed that following polymerase chain reaction (PCR) detection, the known ROLP-insect vectors, N. virescens Distant and Recilia dorsalis Motchulsky, were ROLP-positive, indicating their roles in pathogen dispersal. Moreover, fluorescent and scanning electron microscopy revealed the intensive accumulation of the phytoplasma in phloem tissues and massive accumulation of storage starch in vascular bundle sheath and parenchyma. Altogether, this study illustrated the genetic variability of global ROLP isolates and the pathogen’s biological impact on rice tissue.

DOI： 10.3390/pathogens10020169

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記述言語：日本語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Plant Pathology  

Beet necrotic yellow vein virus (BNYVV) generally has a four-segmented positive-sense RNA genome (RNAs 1–4), but some European and most Asian strains have an additional segment, RNA5. This study examined the effect of RNA5 and RNA3 on different sugar beet cultivars using a Polymyxa-mediated inoculation system under field and laboratory conditions. In field tests, the degree of sugar yield served as an index for assessing the virulence of BNYVV strains. Japanese A-II type isolates without RNA5 caused mostly 15%–90% sugar yield reductions, depending on the susceptibility of sugar beet cultivars, whereas the isolates with RNA5 induced more than 90% yield losses in the seven susceptible cultivars, but small yield losses in one Rz1-resistant and Rizor cultivars. However, a laboratory-produced isolate containing RNA5 but lacking RNA3 caused higher yield losses in Rizor than in susceptible plants, and induced scab-like symptoms on the root surface of both susceptible and resistant plants. In laboratory tests, A-II type isolates without RNA5 had low viral RNA accumulation levels in roots of Rizor and Rz1-resistant plants at early stages of infection, but in the presence of RNA5, viral RNA3 accumulation levels increased remarkably. This increased RNA3 accumulation was not observed in roots of the WB42 accession with the Rz2 gene. In contrast, the presence of RNA3 did not affect RNA5 accumulation levels. Collectively, this study demonstrated that RNA5 is involved in the development of scab-like symptoms and the enhancement of RNA3 accumulation, and suggests these characteristics of RNA5 are associated with Rz1-resistance breaking.

DOI： 10.1111/ppa.13266

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記述言語：日本語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Pathogens  

Rice orange leaf phytoplasma (ROLP) belongs to the “Candidatus Phytoplasma asteris” 16SrI-B subgroup, which is solely transmitted by the zigzag-striped leafhopper (Recilia dorsalis Motchulsky) and the green leafhopper (Nephotettix cincticeps Uhler) (Hemiptera: Cicadellidae). Recently, rice plants showing orange leaf discoloration have become ubiquitous in several paddies of two provinces in the Philippines. In total of 98 symptomatic rice plants, 82% (Laguna) and 95% (Mindanao) were ROLP-positive by nested PCR detection. These plants showed more varying symptoms than previously reported. The vector insect R. dorsalis was scarcely present but green paddy leafhopper, N. virescens Distant (Hemiptera: Cicadellidae), was commonly observed in the paddies, thus the ability of N. virescens to transmit ROLP was thoroughly investigated. Newly emerged adult N. virescens, which fed on ROLD-source rice plants, were used to inoculate a susceptible rice seedling and was serially transferred into a new healthy seedling. Resultant positive transmission rates varied from 5.1% to 17.8%. The transmission ability of the insects was generally decreased over time. These findings suggest that N. virescens is an alternative vector of ROLP in the Philippines. Altogether, this study highlighted the increasing importance of ROLD-reemergence in Southeast and East Asia and proved the need for careful management of this alternative vector insect.

DOI： 10.3390/pathogens9120990

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記述言語：日本語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Journal of General Plant Pathology  

In Solanaceae plants, the major phytoalexins produced during the induction of plant defense are sesquiterpenoids, such as capsidiol for Nicotiana species and rishitin for Solanum species, which are produced via the mevalonate (MVA) pathway. Eight enzymes are involved in the production of farnesyl pyrophosphate (FPP), the common precursor of phytosterols for maintaining membrane integrity and sesquiterpenoid phytoalexins for plant defense. In this study, expression profiles of N. benthamiana genes for the production of capsidiol during the induction of disease resistance were investigated. In the genome of N. benthamiana, multiple copies of genes for each enzyme in the MVA pathway were identified, and the expression of some, but not all MVA genes, was significantly upregulated after inoculation with Phytophthora infestans, or treatment with the INF1 elicitor, a secretory protein of P. infestans. For genes encoding enzymes involved in capsidiol production, 10 copies of 5-epi-aristolochene synthase (NbEAS) and six copies of 5-epi-aristolochene dihydroxylase (NbEAH) were identified, and all copies were significantly upregulated during the induction of disease resistance. Gene silencing of MAP kinase genes NbWIPK, NbSIPK, and NbNTF4 compromised INF1-induced production of phytoalexins. Expression analysis of control and NbWIPK/SIPK/NTF4-silenced plants indicated that most of the MVA genes are not under the control of these MAP kinases. In contrast, the expression pattern of NbWIPK/SIPK/NTF4 and all copies of NbEAH genes showed significant correlation, suggesting that MAP kinases are critical regulators of transcriptional upregulation of specific genes for capsidiol production.

DOI： 10.1007/s10327-020-00931-5

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記述言語：日本語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Crop Protection  

Rice blast (Pyricularia oryzae Cavara) is one of the most devastating diseases affecting the rice crop in Cambodia and other countries in the world. The fungus Trichoderma spp. is known as one of biological control agents applied as a soil treatment, seed treatment and foliar application, that is used for suppression of various diseases caused by fungal pathogens. Trichoderma harzianum strain BTB 022 is one of the commercial biological control products available in Cambodian markets. The combined use of T. harzianum and a resistant variety, to manage blast disease, are considered as sustainable approaches to reduce yield losses and to cope with recent restrictions on fungicide use. A series of consecutive experiments was conducted to examine the effectiveness of T. harzianum on suppression of rice blast incidence in Koktrap and Polors agricultural research stations during wet and dry seasons in 2016 and 2017. In both years, the treatments consisted of the use of Trichoderma on susceptible and resistant rice varieties. In 2017 the two treatments were combined with conventional practice treatments representing the average farmers' practice. The experiments were arranged in randomized complete block design with three replications in 2016 and four replications in 2017. Leaf blast incidence was assessed at five and four growth stages in 2016 and 2017, respectively, and the area under the leaf blast progress curve (AULBPC) was computed for each year and location. Neck blast (NB) incidence was assessed at dough stage and grain yield (GY) was measured at ripening stage. T. harzianum reduced the incidence of leaf blast and neck blast on IR504 (susceptible strain), but its efficacy was not consistent. The magnitude of disease suppression by T. harzianum was higher for neck blast than for leaf blast. GY variation was correlated with AULBPC and NB incidence, which suggests that disease reduction corresponded to an increase in yield (AULBPC: r = −0.877, P < 0.001; NB incidence: r = −0.567, P < 0.001). T. harzianum effectively reduced neck blast at high disease pressure. Growing a resistant variety, e.g. CAR14, effectively reduced AULBPC and NB incidence compared to T. harzianum and farmers’ practice of fungicidal use but the association of T. harzianum and resistant variety did not increase the effect in the control of disease.

DOI： 10.1016/j.cropro.2019.104864

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Toxins  

Fusarium head blight (FHB) of cereals is a severe disease caused by the Fusarium graminearum species complex. It leads to the accumulation of the mycotoxin deoxynivalenol (DON) in grains and other plant tissues and causes substantial economic losses throughout the world. DON is one of the most troublesome mycotoxins because it is a virulence factor to host plants, including wheat, and exhibits toxicity to plants and animals. To control both FHB and DON accumulation, a biological control approach using DON-degrading bacteria (DDBs) is promising. Here, we performed a disease control assay using an in vitro petri dish test composed of germinated wheat seeds inoculated with F. graminearum (Fg) and DDBs. Determination of both grown leaf lengths and hyphal lesion lengths as a measure of disease severity showed that the inoculation of seeds with the DDBs Devosia sp. strain NKJ1 and Nocardioides spp. strains SS3 or SS4 were protective against the leaf growth inhibition caused by Fg. Furthermore, it was as effective against DON accumulation. The inoculation with strains SS3 or SS4 also reduced the inhibitory effect on leaves treated with 10 μg mL-1 DON solution (without Fg). These results indicate that the DDBs partially suppress the disease by degrading DON.

DOI： 10.3390/toxins12060399

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： doi.org/10.1093/pcp/pcae011

記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： doi.org/10.1111/ppl.14052

記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： doi: 10.3389/fpls.2023.1177060

記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： doi.org/10.1128/mra.00259-23

担当区分：筆頭著者   記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： doi.org/10.1099/jgv.0.001848

記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： doi.org/10.1007/s10327-023-01116-6

記述言語：英語   掲載種別：研究論文（研究会，シンポジウム資料等）  

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担当区分：責任著者   記述言語：英語   掲載種別：研究論文（国際会議プロシーディングス）  

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1007/s10327-019-00855-9

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.virol.2019.05.008

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.3389/fpls.2019.00222

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.virusres.2017.11.020

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

記述言語：英語   掲載種別：研究論文（学術雑誌）  

記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1099/jgv.0.001152

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.3390/v10110584

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1007/s10327-018-0801-8

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1099/jgv.0.000985

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1128/mBio.02350-17

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1080/14735903.2016.1174811

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1080/15592324.2017.1300733

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1094/PBIOMES-01-17-0002-R

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ELSEVIER SCIENCE BV  

The family Partitiviridae comprises of five genera with bi-segmented dsRNA genomes that accommodate members infecting plants, fungi or protists. All partitiviruses with only a few exceptions cause asymptomatic infections. We report the characterization of a novel betapartitivirus termed Rosellinia necatrix partitivirus 6 (RnPV6) from a field isolate of a plant pathogenic fungus, white root rot fungus. RnPV6 has typical partitivirus features: dsRNA1 and dsRNA2 are 2462 and 2499 bps in length encoding RNA dependent RNA polymerase and capsid protein. Purified particles are spherical with a diameter of 30 nm. Taking advantage of infectivity as virions, RnPV6 was introduced into a model filamentous fungal host, chestnut blight fungus to investigate virus/host interactions. Unlike other partitiviruses tested previously, RnPV6 induced profound phenotypic alterations with symptoms characterized by a reduced growth rate and enhanced pigmentation and was tolerant to host RNA silencing. In addition, a variety of defective RNAs derived from dsRNA1 appear after virion transfection. These sub-viral RNAs were shown to interfere with RnPV6 replication, at least for that of cognate segment dsRNA1. Presence of these sub-viral elements resulted in reduced symptom expression by RnPV6, suggesting their nature as defective-interfering RNAs. The features of RnPV6 are similar to but distinct from those of a previously reported alphapartitivirus, Rosellinia necatrix partitivirus 2 that is susceptible to RNA silencing. (C) 2015 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.virusres.2015.10.017

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.virusres.2016.05.011

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ELSEVIER SCIENCE BV  

The identification of mycoviruses contributes greatly to understanding of the diversity and evolutionary aspects of viruses. Powdery mildew fungi are important and widely studied obligate phytopathogenic agents, but there has been no report on mycoviruses infecting these fungi. In this study, we used a deep sequencing approach to analyze the double-stranded RNA (dsRNA) segments isolated from field collected samples of powdery mildew fungus-infected red clover plants in Japan. Database searches identified the presence of at least ten totivirus (genus Totivirus)-like sequences, termed red clover powdery mildew-associated totiviruses (RPaTVs). The majority of these sequences shared moderate amino acid sequence identity with each other (&lt;44%) and with other known totiviruses (&lt;59%). Nine of these identified sequences (RPaTV1a, 1b and 2-8) resembled the genome of the prototype totivirus, Saccharomyces cerevisiae virus-L-A (ScV-L-A) in that they contained two overlapping open reading frames (ORFs) encoding a putative coat protein (CP) and an RNA dependent RNA polymerase (RdRp), while one sequence (RPaTV9) showed similarity to another totivirus, Ustilago maydis virus H1 (UmV-H1) that encodes a single polyprotein (CP-RdRp fusion). Similar to yeast totiviruses, each ScV-L-A-like RPaTV contains a-1 ribosomal frameshift site downstream of a predicted pseudoknot structure in the overlapping region of these ORFs, suggesting that the RdRp is translated as a CP-RdRp fusion. Moreover, several ScV-L-A-like sequences were also found by searches of the transcriptome shotgun assembly (TSA) libraries from rust fungi, plants and insects. Phylogenetic analyses show that nine ScV-L-A-like RPaTVs along with ScV-L-A-like sequences derived from TSA libraries are clustered with most established members of the genus Totivirus, while one RPaTV forms a new distinct Glade with UmV-H1, possibly establishing an additional genus in the family. Taken together, our results indicate the presence of diverse, novel totiviruses in the powdery mildew fungus populations infecting red clover plants in the field. (C) 2015 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.virusres.2015.11.015

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記述言語：英語   掲載種別：論文集(書籍)内論文   出版者・発行元：Springer International Publishing  

Beet necrotic yellow vein virus (BNYVV) is a member of the genus Benyvirus in the family Benyviridae with multipartite positive-sense single-stranded RNA genomes encapsidated in rigid rod-shaped particles. The members of Benyvirus including four species are similar to those of the family Virgaviridae, in respect of viral particle assembly, movement, and plasmodiophorid transmissibility. Recent studies revealed that ancestors and/or relatives of benyviruses may have infected a wide range of hosts such as plants, insects, algae, and fungi. For phylogenetic analyses of BNYVV genes, worldwide BNYVV isolates form four clades, A-I, A-II, A-III, and B, from which at least ten subgroup isolates (strains) have derived. These original BNYVV types and their progeny strains might have existed in East Asia, and each source had introduced infection to cultivated sugar beet plants and might have spread worldwide only in the last half century. Along with the growth of resistant varieties in rhizomania-infested areas since the 1980s, strong selection pressure has been imposed on the RNA3-encoded p25 gene, and, consequently, resistance-breaking variants that have single amino acid changes in the p25 protein have been generated. RNA5-encoded p26 gene is also associated with resistance breaking as well as symptom severity in sugar beet roots.

DOI： 10.1007/978-3-319-30678-0_5

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：NATL ACAD SCIENCES  

Viruses often coinfect single host organisms in nature. Depending on the combination of viruses in such coinfections, the interplay between them may be synergistic, apparently neutral with no effect on each other, or antagonistic. RNA silencing is responsible for many cases of interference or cross-protection between viruses, but such antagonistic interactions are usually restricted to closely related strains of the same viral species. In this study, we present an unprecedented example of RNA silencing-mediated one-way interference between unrelated viruses in a filamentous model fungus, Cryphonectria parasitica. The replication of Rosellinia necatrix victorivirus 1 (RnVV1; Totiviridae) was strongly impaired by coinfection with the prototypic member of the genus Mycoreovirus (MyRV1) or a mutant of the prototype hypovirus (Cryphonectria hypovirus 1, CHV1) lacking the RNA silencing suppressor (CHV1-Delta p69). This interference was associated with marked transcriptional induction of key genes in antiviral RNA silencing, dicer-like 2 (dcl2) and argonaute-like 2 (agl2), following MyRV1 or CHV1-Delta p69 infection. Interestingly, the inhibition of RnVV1 replication was reproduced when the levels of dcl2 and agl2 transcripts were elevated by transgenic expression of a hairpin construct of an endogenous C. parasitica gene. Disruption of dcl2 completely abolished the interference, whereas that of agl2 did not always lead to its abolishment, suggesting more crucial roles of dcl2 in antiviral defense. Taken altogether, these results demonstrated the susceptible nature of RnVV1 to the antiviral silencing in C. parasitica activated by distinct viruses or transgene-derived double-stranded RNAs and provide insight into the potential for broad-spectrum virus control mediated by RNA silencing.

DOI： 10.1073/pnas.1509151112

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：SPRINGER WIEN  

Cymbidium chlorotic mosaic virus (CyCMV), isolated from a spring orchid (Cymbidium goeringii), was characterized molecularly. CyCMV isometric virions comprise a single, positive-strand RNA genome of 4,083 nucleotides and 30-kDa coat protein. The virus genome contains five overlapping open reading frames with a genomic organization similar to that of sobemoviruses. BLAST searches and phylogenetic analysis revealed that CyCMV is most closely related to papaya lethal yellowing virus, a proposed dicot-infecting sobemovirus (58.8 % nucleotide sequence identity), but has a relatively distant relationship to monocot-infecting sobemoviruses, with only modest sequence identities. This suggests that CyCMV is a new monocot-infecting member of the floating genus Sobemovirus.

DOI： 10.1007/s00705-015-2460-9

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE  

The chestnut blight fungus, Cryphonectria parasitica, is an important plant pathogenic ascomycete. The fungus hosts a wide range of viruses and now has been established as a model filamentous fungus for studying virus/host and virus/virus interactions. This is based on the development of methods for artificial virus introduction and elimination, host genome manipulability, available host genome sequence with annotations, host mutant strains, and molecular tools. Molecular tools include subcellular distribution markers, gene expression reporters, and vectors with regulatable promoters that have been long available for unicellular organisms, cultured cells, individuals of animals and plants, and certain filamentous fungi. A comparison with other filamentous fungi such as Neurospora crassa has been made to establish clear advantages and disadvantages of C parasitica as a virus host. In addition, a few recent studies on RNA silencing vs. viruses in this fungus are introduced. (C) 2014 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.virol.2014.09.024

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

Endogenous non-retroviral RNA like sequences (NRVSs) have been discovered in the genome of a wide range of eukaryotes. These are considered as fossil RNA viral elements integrated into host genomes by as-yet-known mechanisms, and in many cases, those fossils are estimated to be millions-of-years-old. It is likely that the number of NRVS records will increase rapidly due to the growing availability of whole-genome sequences for many kinds of eukaryotes. Discovery of the novel NRVSs and understanding of their phylogenetic relationship with modern viral relatives provide important information on deep evolutionary history of RNA virus-host interactions. In this chapter, therefore, the common strategies for the identification and characterization of endogenous NRVSs from plants, insects, and fungi are described.

DOI： 10.1007/978-1-4939-1743-3_7

PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：FRONTIERS RESEARCH FOUNDATION  

Here we report a biological and molecular characterization of a novel positive-sense RNA virus isolated from a field isolate (NW10) of a filamentous phytopathogenic fungus, the white root rot fungus that is designated as Rosellinia necatrix fusarivirus 1 (RnFV1). A recently developed technology using zinc ions allowed us to transfer RnFV1 to two mycelially incompatible Rosellinia necatrix strains. A biological comparison of the virus-free and -recipient isogenic fungal strains suggested that RnFV1 infects latently and thus has no potential as a virocontrol agent. The virus has an undivided positive-sense RNA genome of 6286 nucleotides excluding a poly (A) tail. The genome possesses two non-overlapping open reading frames (ORFs): a large ORF1 that encodes polypeptides with RNA replication functions and a smaller ORF2 that encodes polypeptides of unknown function. A lack of coat protein genes was suggested by the failure of virus particles from infected mycelia. No evidence was obtained by Northern analysis or classical 5'-RACE for the presence of subgenomic RNA for the downstream ORE Sequence similarities were found in amino-acid sequence between RnFV1 putative proteins and counterparts of a previously reported mycovirus, Fusarium graminearum virus 1 (FgV1). Interestingly, several related sequences were detected by BLAST searches of independent transcriptome assembly databases one of which probably represents an entire virus genome. Phylogenetic analysis based on the conserved RNA-dependent RNA polymerase showed that RnFV1, FgV1, and these similar sequences are grouped in a cluster distinct from distantly related hypoviruses. It is proposed that a new taxonomic family termed Fusariviridae be created to include RnFV1 and FgV1.

DOI： 10.3389/fmicb.2014.00360

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PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：SOC GENERAL MICROBIOLOGY  

Rosellinia necatrix megabirnavirus 1 (RnMBV1) with a bipartite dsRNA genome (dsRNA1 and dsRNA2) confers hypovirulence to its natural host, the white root rot fungus, and is thus regarded as a potential virocontrol (biocontrol) agent. Each segment has two large ORFs: ORF1 and partially overlapping ORF2 on dsRNA1 encode the major capsid protein (CP) and RNA-dependent RNA polymerase (RdRp), whilst ORF3 and ORF4 on dsRNA2 encode polypeptides with unknown functions. Here, we report the biological and molecular characterization of this virus in the chestnut blight fungus, Cryphonectria parasitica, a filamentous fungus that has been used as a model for mycovirus research. Transfection with purified RnMBV1 particles into an RNA-silencing-defective strain (Delta dcl-2) of C. parasitica and subsequent anastomosis with the WT strain (EP155) resulted in stable persistent infection in both host strains. However, accumulation levels in the two strains were different, being similar to 20-fold higher in Delta dcl-2 than in EP155. Intriguingly, whilst RnMBV1 reduced both virulence and growth rate in Delta dcl-2, it attenuated virulence without affecting significantly other traits in EP155. Western blot analysis using antiserum against recombinant proteins encoded by either ORF1 or ORF2 demonstrated the presence of a 250 kDa protein in purified virion preparations, suggesting that RdRp is expressed as a CP fusion product via a - 1 frameshift. Antiserum against the ORF3-encoded protein allowed the detection of 150, 30 and 23 kDa polypeptides specifically in RnMBV1-infected mycelia. Some properties of an RnMBV1 mutant with genome rearrangements, which occurred after transfection of Delta dcl-2 and EP155, were also presented. This study provides an additional example of C. parasitica serving as a versatile, heterologous fungus for exploring virus host interactions and virus gene expression strategies.

DOI： 10.1099/vir.0.058164-0

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PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE  

The transcriptional strategy of orchid fleck virus (OFV), which has a two-segmented negative-strand RNA genome and resembles plant nucleorhabdoviruses, remains unexplored. In this study, the transcripts of six genes encoded by OFV RNA1 and RNA2 in the poly(A)-enriched RNA fraction from infected plants were molecularly characterized. All of the OFV mRNAs were initiated at a start sequence 3'-UU-5' with one to three non-viral adenine nucleotides which were added at the 5' end of each mRNA, whereas their 3' termini ended with a 5'-AUUUAAA(U/G)AAAA(A)n-3' sequence. We also identified the presence of polyadenylated short transcripts derived from the 3'-terminal leader regions of both genomic and antigenomic strands, providing the first example of plus- and minus-strand leader RNAs in a segmented minus-strand RNA virus. The similarity in the transcriptional strategy between this bipartite OFV and monopartite rhabdoviruses, especially nucleorhabdoviruses (family Rhabdoviridae) is additional support for their close relationship. (C) 2014 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.virol.2014.01.007

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PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：SPRINGER WIEN  

The complete genomic sequence of Habenaria mosaic virus (HaMV), which infects terrestrial orchids (Habenaria radiata), has been determined. The genome is composed of 9,499 nucleotides excluding the 3'-terminal poly(A) tail, encoding a large polyprotein of 3,054 amino acids with the genomic features typical of a potyvirus. Putative proteolytic cleavage sites were identified by sequence comparison to those of known potyviruses. The HaMV polyprotein showed 58 % amino acid sequence identity to that encoded by the most closely related potyvirus, tobacco vein banding mosaic virus. Phylogenetic analysis of the polyprotein amino acid sequence and its coding sequences confirmed that HaMV formed a cluster with the chilli veinal mottle virus group, most of which infect solanaceous plants. These results suggest that HaMV is a distinct member of the genus Potyvirus.

DOI： 10.1007/s00705-013-1784-6

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PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.2222/jsv.64.225

PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）  

The complete nucleotide sequence of the burdock mottle virus (BdMoV) isolated from an edible burdock plant ( Arctium lappa) in Japan has been determined. BdMoV has a bipartite genome, whose organization is similar to RNA1 and RNA2 of benyviruses, beet necrotic yellow vein virus (BNYVV), beet soil-borne mosaic virus (BSBMV), and rice stripe necrosis virus (RSNV). BdMoV RNA1 (7038 nt) contains a single open reading frame (ORF) encoding a 249-kDa polypeptide that consists of methyl-transferase, helicase, papain-like protease, AlkB-like, and RNA-dependent RNA polymerase domains. The AlkB-like domain sequence is not present in the proteins encoded by other known benyviruses, but is found in replication-associated proteins of viruses mainly belonging to the families Alfaflexiviridae and Betaflexiviridae. BdMoV RNA2 (4315 nt) contains six ORFs that are similar to those of benyviruses: these are coat protein (CP), CP readthrough, triple gene block movement and cysteine-rich proteins. Phylogenetic analyses showed that BdMoV is more closely related to BNYVV and BSBMV than to RSNV. Database searches showed that benyvirus replicase-related sequences are present in the chromosomes of a chickpea plant ( Cicer arietinum) and a blood-sucking insect ( Rhodnius prolixus). Some other benyvirus-related sequences are found in the transcriptome shotgun libraries of a few species of plants and a bark beetle. Our results show that BdMoV is a distinct species of the genus Benyvirus and that ancestral and extant benyviruses may have infected or currently infect a wide range of hosts, including plants and insects. © 2013 Elsevier B.V.

DOI： 10.1016/j.virusres.2013.07.015

Scopus

PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）  

Orchid fleck virus (OFV) has a unique two-segmented negative-sense RNA genome that resembles that of plant nucleorhabdoviruses. In infected plant cells, OFV and nucleorhabdoviruses induce an intranuclear electron-lucent viroplasm that is believed to be the site for virus replication. In this study, we investigated the molecular mechanism by which OFV viroplasms are produced in vivo. Among OFV-encoded proteins, the nucleocapsid protein (N) and the putative phosphoprotein (P) were present in nuclear fractions of OFV-infected Nicotiana benthamiana plants. Transient coexpression of N and P, in the absence of virus infection, was shown to be sufficient for formation of an intranuclear viroplasm-like structure in plant cells. When expressed independently as a fluorescent protein fusion product in uninfected plant cells, N protein accumulated throughout the cell, while P protein accumulated in the nucleus. However, the N protein, when coexpressed with P, was recruited to a subnuclear region to induce a large viroplasm-like focus. Deletion and substitution mutagenesis demonstrated that the P protein contains a nuclear localization signal (NLS). Artificial nuclear targeting of the N-protein mutant was insufficient for formation of viroplasm-like structures in the absence of P. A bimolecular fluorescence complementation assay confirmed interactions between the N and P proteins within subnuclear viroplasm-like foci and interactions of two of the N. benthamiana importin-α homologues with the P protein but not with the N protein. Taken together, our results suggest that viroplasm formation by OFV requires nuclear accumulation of both the N and P proteins, which is mediated by P-NLS, unlike nucleorhabdovirus viroplasm utilizing the NLS on protein N. © 2013, American Society for Microbiology.

DOI： 10.1128/JVI.00270-13

Scopus

PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER SOC MICROBIOLOGY  

A novel victorivirus, termed Rosellinia necatrix victorivirus 1 (RnVV1), was isolated from a plant pathogenic ascomycete, white root rot fungus Rosellinia necatrix, coinfected with a partitivirus. The virus was molecularly and biologically characterized using the natural and experimental hosts (chestnut blight fungus, Cryphonectria parasitica). RnVV1 was shown to have typical molecular victorivirus attributes, including a monopartite double-stranded RNA genome with two open reading frames (ORFs) encoding capsid protein (CP) and RNA-dependent RNA polymerase (RdRp), a UAAUG pentamer presumed to facilitate the coupled termination/reinitiation for translation of the two ORFs, a spherical particle structure similar to 40 nm in diameter, and moderate levels of CP and RdRp sequence identity (34 to 58%) to those of members of the genus Victorivirus within the family Totiviridae. A reproducible transfection system with purified RnVV1 virions was developed for the two distinct fungal hosts. Transfection assay with purified RnVV1 virions combined with virus elimination by hyphal tipping showed that the effects of RnVV1 on the phenotype of the natural host were negligible. Interestingly, comparison of the RNA silencing-competent (standard strain EP155) and -defective (Delta dcl-2) strains of C. parasitica infected with RnVV1 showed that RNA silencing acted against the virus to repress its replication, which was restored by coinfection with hypovirus or transgenic expression of an RNA silencing suppressor, hypovirus p29. Phenotypic changes were observed in the Delta dcl-2 strain but not in EP155. This is the first reported study on the host range expansion of a Totiviridae member that is targeted by RNA silencing.

DOI： 10.1128/JVI.00557-13

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PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER SOC MICROBIOLOGY  

A novel mycovirus termed Rosellinia necatrix partitivirus 2 (RnPV2), isolated from a phytopathogenic fungus, Rosellinina necatrix strain W57, was molecularly and biologically characterized in both natural and experimental host fungi. Three double-stranded RNA (dsRNA) segments, dsRNA1, dsRNA2, and defective interfering dsRNA1 (DI-dsRNA1), whose sizes were approximately 2.0, 1.8, and 1.7 kbp, respectively, were detected in W57. While the dsRNA2 sequence, encoding the coat protein, was reported previously, dsRNA1 and DI-dsRNA1 were shown to encode competent and defective (truncated) RNA-dependent RNA polymerase, respectively. Artificial introduction of RnPV2 into an RNA silencing-defective, Dicer-like 2 knockout mutant (Delta dcl-2) of a nonnatural host, Cryphonectria parasitica (chestnut blight fungus), resulted in successful infection by the DI-dsRNA1-carrying and -free RnPV2. The DI-dsRNA1-free RnPV2 strain was characterized by a higher ratio of accumulation of the intact dsRNA1 to dsRNA2, enhanced replication and severer symptom expression, compared with the DI-carrying strain. These findings confirmed the nature of DI-dsRNA1 as a DI-RNA. Both viral strains replicated to higher levels in a Delta dcl-2 mutant than in a wild-type C. parasitica fungal strain (EP155) and induced severe symptoms in the Delta dcl-2 mutant but subtle symptoms in EP155, indicating that the host RNA silencing targets the partitivirus. No obvious phenotypic effects of infection by either virus strain were detected in the natural host fungus. These combined results represent the first example of a partitivirus with DI-RNA that alters viral symptom induction in a host-dependent manner.

DOI： 10.1128/JVI.02835-12

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PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER PHYTOPATHOLOGICAL SOC  

The RNA silencing-suppression properties of Beet necrotic yellow vein virus (BNYVV) and Beet soil-borne mosaic virus (BSBMV) cysteine-rich p14 proteins have been investigated. Suppression of RNA silencing activities were made evident using viral infection of silenced Nicotiana benthamiana 16C, N. benthamiana agroinfiltrated with green fluorescent protein (GFP), and GF-FG hairpin triggers supplemented with viral suppressor of RNA silencing (VSR) constructs or using complementation of a silencing-suppressor-defective BNYVV virus in Chenopodium quinoa. Northern blot analyses of small-interfering RNAs (siRNAs) in agroinfiltration tests revealed reduced amounts of siRNA, especially secondary siRNA, suggesting that benyvirus VSR act downstream of the siRNA production. Using confocal laser-scanning microscopy imaging of infected protoplasts expressing functional p14 protein fused to an enhanced GFP reporter, we showed that benyvirus p14 accumulated in the nucleolus and the cytoplasm independently of other viral factors. Site-directed mutagenesis showed the importance of the nucleolar localization signal embedded in a C4 zinc-finger domain in the VSR function and intrinsic stability of the p14 protein. Conversely, RNA silencing suppression appeared independent of the nucleolar localization of the protein, and a correlation between BNYVV VSR expression and long-distance movement was established.

DOI： 10.1094/MPMI-06-12-0142-R

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PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）  

Fungal viruses comprise two groups: a major group of five families with double-stranded RNA genomes and a minor group with positive-sense single-stranded (ss)RNA genomes. Although many fungal viruses have been identified, no negative-stranded (-)ssRNA mycoviruses have been reported. Here we present two lines of evidence suggesting the presence of (-)ssRNA viruses in filamentous fungi based on an exhaustive search using extant (-)ssRNA viruses as queries. This revealed (-)ssRNA virus L protein-like sequences in the genome of a phytopathogenic obligate ascomycete, Erysiphe pisi. A similar search for (-)ssRNA viruses in fungal transcriptome shotgun assembly libraries demonstrated that two independent libraries from Sclerotinia homoeocarpa, another phytopathogenic ascomycete, contained several sequences considered to correspond to the entire mononegavirus L gene and likely originating from an infecting (-)ssRNA virus. These results provide strong evidence for both ancient and extant (-)ssRNA virus infections in fungi. © 2012 Elsevier Inc.

DOI： 10.1016/j.virol.2012.10.002

Scopus

PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE  

Here we report the biological and molecular attributes of a novel dsRNA virus isolated from Rosellinia necatrix, a filamentous phytopathogenic fungus. The virus, termed Rosellinia necatrix quadrivirus 1 (RnQV1), forms rigid spherical particles approximately 45 nm in diameter in infected mycelia. The particles contain 4 dsRNA segments, dsRNA1 to dsRNA4, with a size range of 4.9 to 3.7 kbp, each possessing a single large ORF. A comparison of the virus-infected and -cured isogenic fungal strains suggested that RnQV1 infection has no appreciable phenotypic effects. Phylogenetic analysis using the dsRNA3-encoded RdRp sequence revealed that RnQV1 is more distantly related to quadripartite chrysoviruses than to monopartite totiviruses, and is placed in a distinct group from other mycoviruses. No significant sequence similarities were evident between known proteins and RnQV1 structural proteins shown to be encoded by dsRNA2 or dsRNA4. These suggest that RnQV1 is a novel latent virus, belonging to a new family. (C) 2012 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.virol.2012.01.013

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PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：PUBLIC LIBRARY SCIENCE  

Non-retroviral RNA virus sequences (NRVSs) have been found in the chromosomes of vertebrates and fungi, but not plants. Here we report similarly endogenized NRVSs derived from plus-, negative-, and double-stranded RNA viruses in plant chromosomes. These sequences were found by searching public genomic sequence databases, and, importantly, most NRVSs were subsequently detected by direct molecular analyses of plant DNAs. The most widespread NRVSs were related to the coat protein (CP) genes of the family Partitiviridae which have bisegmented dsRNA genomes, and included plant-and fungus-infecting members. The CP of a novel fungal virus (Rosellinia necatrix partitivirus 2, RnPV2) had the greatest sequence similarity to Arabidopsis thaliana ILR2, which is thought to regulate the activities of the phytohormone auxin, indole-3-acetic acid (IAA). Furthermore, partitivirus CP-like sequences much more closely related to plant partitiviruses than to RnPV2 were identified in a wide range of plant species. In addition, the nucleocapsid protein genes of cytorhabdoviruses and varicosaviruses were found in species of over 9 plant families, including Brassicaceae and Solanaceae. A replicase-like sequence of a betaflexivirus was identified in the cucumber genome. The pattern of occurrence of NRVSs and the phylogenetic analyses of NRVSs and related viruses indicate that multiple independent integrations into many plant lineages may have occurred. For example, one of the NRVSs was retained in Ar. thaliana but not in Ar. lyrata or other related Camelina species, whereas another NRVS displayed the reverse pattern. Our study has shown that single-and double-stranded RNA viral sequences are widespread in plant genomes, and shows the potential of genome integrated NRVSs to contribute to resolve unclear phylogenetic relationships of plant species.

DOI： 10.1371/journal.ppat.1002146

Web of Science

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER PHYTOPATHOLOGICAL SOC  

Beet necrotic yellow vein virus (BNYVV) is an economically important pathogen of sugar beet and has been found worldwide, probably as the result of recent worldwide spread. The BNYVV genome consists of four or five RNA components. Here, we report analysis of sequence variation in the RNA3-p25, RNA4-p31, RNA2-CP, and RNA5-p26 genes of 73 worldwide isolates. The RNA3-p25 gene encodes virulence and avirulence factors. These four sets of gene sequences each fell into two to four groups, of which the three groups of p25 formed eight subgroups with different geographical distributions. Each of these subgroup isolates (strains) could have arisen from four original BNYVV population and their mixed infections. The genetic diversity for BNYVV was relatively small. Selection pressure varied greatly depending on the BNYVV gene and geographical location. Isolates of the Italy strain, in which p25 was subject to the strongest positive selection, were able to overcome the Rz1-host resistance gene to differing degrees, whereas other geographically limited strains could not. Resistance-breaking variants were generated by p25 amino acid changes at positions 67 and 68. Our studies suggest that BNYVV originally evolved in East Asia and has recently become a pathogen of cultivated sugar beet followed by the emergence of new resistance-breaking variants.

DOI： 10.1094/MPMI-10-10-0241

Web of Science

記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.2222/jsv.60.163

PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER SOC MICROBIOLOGY  

White root rot, caused by the ascomycete Rosellinia necatrix, is a devastating disease worldwide, particularly in fruit trees in Japan. Here we report on the biological and molecular properties of a novel bipartite double-stranded RNA (dsRNA) virus encompassing dsRNA-1 (8,931 bp) and dsRNA-2 (7,180 bp), which was isolated from a field strain of R. necatrix, W779. Besides the strictly conserved 5&apos; (24 nt) and 3&apos; (8 nt) terminal sequences, both segments show high levels of sequence similarity in the long 5&apos; untranslated region of approximately 1.6 kbp. dsRNA-1 and -2 each possess two open reading frames (ORFs) named ORF1 to -4. Although the protein encoded by 3&apos;-proximal ORF2 on dsRNA-1 shows sequence identities of 22 to 32% with RNA-dependent RNA polymerases from members of the families Totiviridae and Chrysoviridae, the remaining three virus-encoded proteins lack sequence similarities with any reported mycovirus proteins. Phylogenetic analysis showed that the W779 virus belongs to a separate clade distinct from those of other known mycoviruses. Purified virions similar to 50 nm in diameter consisted of dsRNA-1 and -2 and a single major capsid protein of 135 kDa, which was shown by peptide mass fingerprinting to be encoded by dsRNA-1 ORF1. We developed a transfection protocol using purified virions to show that the virus was responsible for reduction of virulence and mycelial growth in several host strains. These combined results indicate that the W779 virus is a novel bipartite dsRNA virus with potential for biological control (virocontrol), named Rosellinia necatrix megabirna-virus 1 (RnMBV1), that possibly belongs to a new virus family.

DOI： 10.1128/JVI.01830-09

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PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：OXFORD UNIV PRESS  

Cryphonectria hypovirus 1 (CHV1), associated with the picorna-like superfamily, infects the chestnut blight fungus and attenuates the virulence of the host fungus. The genomic RNA of the virus has two continuous open reading frames, A and B, separated by the pentanucleotide UAAUG. We present here evidence suggesting that ORF B is translated from genome-sized virus mRNA by a coupled termination/reinitiation mechanism mediated by the pentamer. In the coupled translation, the overlapping UAA and AUG triplets serve as the stop codon of ORF A and the initiator of ORF B, respectively. This was established by the use of a luciferase assay with a basic construct containing the ORF A sequence and the firefly luciferase gene while retaining the pentamer between the two coding sequences. The proportion of ribosomes reinitiating translation after terminating was determined to be 2.54.4 by three independent assay systems in fungal and insect cells. Use of a series of mutant constructs identified two sequence elements, the pentamer and the p40 sequence, that affect the efficiency of coupled translation and virus replication. Together, these results provide the first example of coupled translation facilitated by the pentanucleotide UAAUG in the kingdom Fungi. The mechanism by which the preceding p40-coding sequence promotes reinitiation is discussed.

DOI： 10.1093/nar/gkp224

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PubMed

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：MICROBIOLOGY SOC  

The RNA3-encoded p25 protein of beet necrotic yellow vein virus (BNYVV) is responsible for the production of rhizomania symptoms of sugar beet roots (Beta vulgaris subsp. vulgaris). Here, it was found that the presence of the p25 protein is also associated with the resistance response in rub-inoculated leaves of sugar beet and wild beet (Beta vulgaris subsp. maritima) plants. The resistance phenotype displayed a range of symptoms from no visible lesions to necrotic or greyish lesions at the inoculation site, and only very low levels of virus and viral RNA accumulated. The susceptible phenotype showed large, bright yellow lesions and developed high levels of virus accumulation. In roots after Polymyxa betae vector inoculation, however, no drastic differences in virus and viral RNA accumulation levels were found between plants with susceptible and resistant phenotypes, except at an early stage of infection. There was a genotype-specific interaction between BNYVV strains and two selected wild beet lines (MR1 and MR2) and sugar beet cultivars. Sequence analysis of natural BNYVV isolates and site-directed mutagenesis of the p25 protein revealed that 3 aa residues at positions 68, 70 and 179 are important in determining the resistance phenotype, and that host-genotype specificity is controlled by single amino acid changes at position 68. The mechanism of the occurrence of resistance-breaking BNYVV strains is discussed.

DOI： 10.1099/vir.0.83624-0

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