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1
Moskalev, A. V., V. B. Sboychakov, A. V. Apchel und N. V. Tsygan. „Вiological effects of viroids“. Bulletin of the Russian Military Medical Academy 20, Nr. 2 (15.12.2018): 209–14. http://dx.doi.org/10.17816/brmma12333.
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Possible potential microbial agents of a human which represent by ring highly complementary one-chained ribonucleic acid with an absent of protein coating, as for viruses containing a ribonucleic acid, are characterized. Pathogenic effects of viroids in comparison with viruses and defective viruses are considered. Intimate viroid’s mechanisms on a host cell are described. So viroids do not code any proteins, their action on a plant should be consequence of direct interaction of viroid’s ribonucleic acid and host cell’s contents. However the molecular mechanism by which viroids causes plants’ diseases still remains not completely determined. It is considered, that the first targets of viroid are the nucleic acids and proteins of the host cell. Genomes of some viroids contain areas complementary to some cellular RNA. In connection with it supposed, that disease begins because of inhibition of functions of these cellular ribonucleic acids or their cutting directed viroids by a ribonucleic acid. Pathogenicity of viroids can be also a result of mimicry at a molecular level. Because of features of structure or sequence nucleotides the viroid’s ribonucleic acid can replace by itself some cellular ribonucleic acids. In viroid’s pathogenesis also can be involved and an interference of ribonucleic acids. All this allows to assume, that viroids can become the reason of infectious diseases of the human.
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Hadidi, Ahmed, Liying Sun und John W. Randles. „Modes of Viroid Transmission“. Cells 11, Nr. 4 (18.02.2022): 719. http://dx.doi.org/10.3390/cells11040719.
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Studies on the ways in which viroids are transmitted are important for understanding their epidemiology and for developing effective control measures for viroid diseases. Viroids may be spread via vegetative propagules, mechanical damage, seed, pollen, or biological vectors. Vegetative propagation is the most prevalent mode of spread at the global, national and local level while further dissemination can readily occur by mechanical transmission through crop handling with viroid-contaminated hands or pruning and harvesting tools. The current knowledge of seed and pollen transmission of viroids in different crops is described. Biological vectors shown to transmit viroids include certain insects, parasitic plants, and goats. Under laboratory conditions, viroids were also shown to replicate in and be transmitted by phytopathogenic ascomycete fungi; therefore, fungi possibly serve as biological vectors of viroids in nature. The term “mycoviroids or fungal viroids” has been introduced in order to denote these viroids. Experimentally, known sequence variants of viroids can be transmitted as recombinant infectious cDNA clones or transcripts. In this review, we endeavor to provide a comprehensive overview of the modes of viroid transmission under both natural and experimental situations. A special focus is the key findings which can be applied to the control of viroid diseases.
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Ito, Takao, Hiroyuki Ieki, Katsumi Ozaki, Toru Iwanami, Kenji Nakahara, Tatsuji Hataya, Tsutae Ito, Masahiro Isaka und Takeshi Kano. „Multiple Citrus Viroids in Citrus from Japan and Their Ability to Produce Exocortis-Like Symptoms in Citron“. Phytopathology® 92, Nr. 5 (Mai 2002): 542–47. http://dx.doi.org/10.1094/phyto.2002.92.5.542.
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Sequential polyacrylamide gel electrophoresis analyses showed many viroid-like RNAs in samples collected from citrus trees in Japan. Reverse transcription polymerase chain reaction and sequencing analyses of the amplified fragments verified that they were derived from variants of six citrus viroids, Citrus exocortis viroid (CEVd), Citrus bent leaf viroid (CBLVd) including CVd-I-LSS (a distinct variant of CBLVd), Hop stunt viroid, Citrus viroid III, Citrus viroid IV, and Citrus viroid OS. The samples induced symptoms with variable severity in Arizona 861-S1 ‘Etrog’ citrons (Citrus medica L.) likely due to the varying accumulation patterns produced by the different viroids. Some of the symptoms caused by the samples harboring the citrus viroids other than CEVd were as severe as those caused by CEVd. Some source citrus trees showing the severe bark scaling characteristic of exocortis disease in trifoliate orange (Poncirus trifoliata (L.) Raf.) rootstocks contained only citrus viroids other than CEVd in complex. This indicates that certain exocortis-like diseases in Japan were caused by some combination of citrus viroids not including CEVd.
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Kunta, Madhurababu, J. V. da Graça und Mani Skaria. „Molecular Detection and Prevalence of Citrus Viroids in Texas“. HortScience 42, Nr. 3 (Juni 2007): 600–604. http://dx.doi.org/10.21273/hortsci.42.3.600.
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Viroids are graft- or mechanically transmissible agents, disseminated through budding. Biological indexing of commercially important citrus cultivars grown in the Lower Rio Grande Valley of Texas showed that many are infected with citrus viroids. Most of these trees carried more than one viroid. In most cases, the infected trees are asymptomatic carriers because sour orange, the predominant rootstock used in Texas, does not show symptoms of viroid infection. Detection of viroids through biological indexing on sensitive indicator plants followed by sequential polyacrylamide gel electrophoresis (sPAGE) is the gold standard but is time-consuming and requires plants to be kept at optimum conditions. A conditional use of reverse transcriptase–polymerase chain reaction (RT-PCR) provides an efficient and alternative detection of viroids for use in the Texas virus-free citrus budwood certification program. RT-PCR could be useful in Texas to help expedite the evaluation for the presence of viroids before conducting the final biologic indexing. Using RT-PCR, we could detect, clone, and sequence full-length viroids of Citrus exocortis viroid (CEVd), Hop stunt viroid (HSVd) (both cachexia and noncachexia variants), Citrus viroid-III (Citrus dwarfing viroid), and Citrus viroid-IV (Citrus bark cracking viroid) from a collection of viroid-inoculated grapefruit plants. The source plants were previously shown to be viroid-infected by biological indexing on Etrog citron plants. Based on our results, RT-PCR can be a conditional substitute for biological indexing of mother trees in foundation blocks and shoot tip-grafted trees in the virus-free budwood program. A positive RT-PCR result has a serendipitous value because those trees can be discarded from the pool before expensive biological indexing.
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Lee, Benjamin D., und Eugene V. Koonin. „Viroids and Viroid-like Circular RNAs: Do They Descend from Primordial Replicators?“ Life 12, Nr. 1 (12.01.2022): 103. http://dx.doi.org/10.3390/life12010103.
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Viroids are a unique class of plant pathogens that consist of small circular RNA molecules, between 220 and 450 nucleotides in size. Viroids encode no proteins and are the smallest known infectious agents. Viroids replicate via the rolling circle mechanism, producing multimeric intermediates which are cleaved to unit length either by ribozymes formed from both polarities of the viroid genomic RNA or by coopted host RNAses. Many viroid-like small circular RNAs are satellites of plant RNA viruses. Ribozyviruses, represented by human hepatitis delta virus, are larger viroid-like circular RNAs that additionally encode the viral nucleocapsid protein. It has been proposed that viroids are direct descendants of primordial RNA replicons that were present in the hypothetical RNA world. We argue, however, that much later origin of viroids, possibly, from recently discovered mobile genetic elements known as retrozymes, is a far more parsimonious evolutionary scenario. Nevertheless, viroids and viroid-like circular RNAs are minimal replicators that are likely to be close to the theoretical lower limit of replicator size and arguably comprise the paradigm for replicator emergence. Thus, although viroid-like replicators are unlikely to be direct descendants of primordial RNA replicators, the study of the diversity and evolution of these ultimate genetic parasites can yield insights into the earliest stages of the evolution of life.
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Barbosa, C. J., J. A. Pina, J. Pérez-Panadés, L. Bernad, P. Serra, L. Navarro und N. Duran-Vila. „Mechanical Transmission of Citrus Viroids“. Plant Disease 89, Nr. 7 (Juli 2005): 749–54. http://dx.doi.org/10.1094/pd-89-0749.
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Preliminary transmission assays conducted under greenhouse conditions demonstrated that Citrus exocortis viroid (CEVd), Citrus bent leaf viroid (CBLVd), Hop stunt viroid (HSVd), Citrus viroid III (CVd-III), and Citrus viroid IV (CVd-IV) can be mechanically transmitted from citron to citron (Citrus medica) by a single slash with a knife blade. The impact of mechanical transmission of viroids by pruning and harvesting operations was also demonstrated in experimental and commercial field plots. Transmission efficiency under field conditions ranged from 4% in ‘Nules’ clementine to 10% in ‘Navelina’ sweet orange and 21% in ‘Verna’ lemon. Transmission efficiency varied only slightly with viroid and donor hosts. The impact of viroid transmission on tree height, canopy volume, and crop harvest was minimal. When the donor host was coinfected with several viroids, the viroids were not necessarily cotransmitted. Considerations regarding viroid transmission in other climates are discussed. Measures to control viroid spread in nurseries should be mandatory in certification programs.
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Wei, Shuang, Ruiling Bian, Ida Bagus Andika, Erbo Niu, Qian Liu, Hideki Kondo, Liu Yang et al. „Symptomatic plant viroid infections in phytopathogenic fungi“. Proceedings of the National Academy of Sciences 116, Nr. 26 (10.06.2019): 13042–50. http://dx.doi.org/10.1073/pnas.1900762116.
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Viroids are pathogenic agents that have a small, circular noncoding RNA genome. They have been found only in plant species; therefore, their infectivity and pathogenicity in other organisms remain largely unexplored. In this study, we investigate whether plant viroids can replicate and induce symptoms in filamentous fungi. Seven plant viroids representing viroid groups that replicate in either the nucleus or chloroplast of plant cells were inoculated to three plant pathogenic fungi,Cryphonectria parasitica,Valsa mali, andFusarium graminearum. By transfection of fungal spheroplasts with viroid RNA transcripts, each of the three, hop stunt viroid (HSVd), iresine 1 viroid, and avocado sunblotch viroid, can stably replicate in at least one of those fungi. The viroids are horizontally transmitted through hyphal anastomosis and vertically through conidia. HSVd infection severely debilitates the growth ofV. malibut not that of the other two fungi, while inF. graminearumandC. parasitica, with deletion of dicer-like genes, the primary components of the RNA-silencing pathway, HSVd accumulation increases. We further demonstrate that HSVd can be bidirectionally transferred betweenF. graminearumand plants during infection. The viroids also efficiently infect fungi and induce disease symptoms when the viroid RNAs are exogenously applied to the fungal mycelia. These findings enhance our understanding of viroid replication, host range, and pathogenicity, and of their potential spread to other organisms in nature.
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Lee, Benjamin D., Uri Neri, Caleb J. Oh, Peter Simmonds und Eugene V. Koonin. „ViroidDB: a database of viroids and viroid-like circular RNAs“. Nucleic Acids Research 50, Nr. D1 (09.11.2021): D432—D438. http://dx.doi.org/10.1093/nar/gkab974.
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Abstract We introduce ViroidDB, a value-added database that attempts to collect all known viroid and viroid-like circular RNA sequences into a single resource. Spanning about 10 000 unique sequences, ViroidDB includes viroids, retroviroid-like elements, small circular satellite RNAs, ribozyviruses, and retrozymes. Each sequence's secondary structure, ribozyme content, and cluster membership are predicted via a custom pipeline optimized for handling circular RNAs. The data can be explored via a purpose-built user interface that features visualizations, multiple sequence alignments, and a portal for downloading bulk data. Users can browse the data by sequence type, taxon, or typo-tolerant search of metadata fields. The database is freely accessible at https://viroids.org.
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Venkataraman, Srividhya, Uzma Badar, Erum Shoeb, Ghyda Hashim, Mounir AbouHaidar und Kathleen Hefferon. „An Inside Look into Biological Miniatures: Molecular Mechanisms of Viroids“. International Journal of Molecular Sciences 22, Nr. 6 (10.03.2021): 2795. http://dx.doi.org/10.3390/ijms22062795.
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Viroids are tiny single-stranded circular RNA pathogens that infect plants. Viroids do not encode any proteins, yet cause an assortment of symptoms. The following review describes viroid classification, molecular biology and spread. The review also discusses viroid pathogenesis, host interactions and detection. The review concludes with a description of future prospects in viroid research.
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SEČNIK, Andrej, Sebastjan RADIŠEK, Nataša ŠTAJNER und Jernej JAKŠE. „Študij polarnosti verig različnih viroidov in njihovih kombinacij pri okuženih rastlinah hmelja“. Acta agriculturae Slovenica 115, Nr. 1 (05.03.2020): 193. http://dx.doi.org/10.14720/aas.2020.115.1.1319.
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<p>Hop plant (<em>Humulus lupulus </em>L.) is an important industrial crop, grown for harvesting hop cones however, it is a host to four different viroids as well. The nature of viroid infections is not entirely clarified. In our work, we focused on analyzing viroid accumulation and their strands polarity through RNA sequencing and reverse transcription polymerase chain reaction in real time. RNA-seq data indicate that viroids amplify until saturation further demonstrating plant's biological capacity. Negative fold changes in accumulation of individual viroids between hop samples with single and multiple infections are suggesting an antagonistic relationship amongst viroids, where citrus bark cracking viroid seems to be the least and hop stunt viroid the most sensitive to the other two. RNA-seq data also show that on average (−) viroid strand is dominating over (+), especially for the citrus bark cracking viroid. Reverse transcription polymerase chain reaction in real time results from strand polarity analysis seem to be less consistent between different combinations of infection but are showing level of conformity with RNA-seq in the case of citrus bark cracking viroid.</p>
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Seo, Hyesu, Ying Wang und Woong June Park. „Time-Resolved Observation of the Destination of Microinjected Potato Spindle Tuber Viroid (PSTVd) in the Abaxial Leaf Epidermal Cells of Nicotiana benthamiana“. Microorganisms 8, Nr. 12 (20.12.2020): 2044. http://dx.doi.org/10.3390/microorganisms8122044.
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Viroids are single-stranded noncoding RNA molecules of 250–400 nucleotides that cause plant diseases. One of the two families of viroids is Pospiviroidae, the members of which replicate in the nuclei of host cells. To replicate in plants, viroids of Pospiviroidae must enter the nucleus. However, the nuclear import of viroids remains understudied. In this work, we documented the time-dependent characteristics of the changes in microinjected fluorescently labeled potato spindle tuber viroid (PSTVd). The cytoplasmic fluorescence disappeared gradually, with only nuclear fluorescence remaining as the PSTVd injected in the cytoplasm was imported into the nucleus. Through this work, we determined that the time for half-maximal nuclear accumulation of the viroid was about 23 min. Interestingly, we found some cells where the nuclear import did not occur, despite the high level of cytosolic viroid injected. In some cells, the injected viroids disappeared within 10–20 min. The nuclear import of PSTVd is not a simple concentration-dependent process but was probably under the regulation of diverse factors that may be missing from some cells used for our observation.
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Fuji, Shin-ichi, Tomofumi Mochizuki, Mitsuru Okuda, Shinya Tsuda, Satoshi Kagiwada, Ken-Taro Sekine, Masashi Ugaki et al. „Plant viruses and viroids in Japan“. Journal of General Plant Pathology 88, Nr. 2 (17.01.2022): 105–27. http://dx.doi.org/10.1007/s10327-022-01051-y.
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AbstractAn increasing number of plant viruses and viroids have been reported from all over the world due largely to metavirogenomics approaches with technological innovation. Herein, the official changes of virus taxonomy, including the establishment of megataxonomy and amendments of the codes of virus classification and nomenclature, recently made by the International Committee on Taxonomy of Viruses were summarized. The continued efforts of the plant virology community of Japan to index all plant viruses and viroids occurring in Japan, which represent 407 viruses, including 303 virus species and 104 unclassified viruses, and 25 viroids, including 20 species and 5 unclassified viroids, as of October 2021, were also introduced. These viruses and viroids are collectively classified into 81 genera within 26 families of 3 kingdoms (Shotokuvirae, Orthornavirae, Pararnavirae) across 2 realms (Monodnaviria and Riboviria). This review also overviewed how Japan’s plant virus/viroid studies have contributed to advance virus/viroid taxonomy.
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Gillings, MR, P. Broadbent und BI Gollnow. „Viroids in Australian Citrus: Relationship to Exocortis, Cachexia and Citrus Dwarfing“. Functional Plant Biology 18, Nr. 5 (1991): 559. http://dx.doi.org/10.1071/pp9910559.
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Viroids are the smallest infectious agents known, being unencapsidated RNAs of 240-380 bases. Citrus exocortis viroid (CEV) causes poor tree performance, especially when infected scions are grafted to trifoliate orange or citrange rootstocks. To eliminate infected budwood trees, various methods were used to detect CEV, including field inspections, bud inoculation of Etrog citron indicators (in which CEV causes severe epinasty) and hybridisation with CEV cDNA. A large number of trees with exocortis- like symptoms such as dwarfing and/or bud union abnormalities produced only mild epinasty when grafted on Etrog citron and did not hybridise to the CEV probe. Upon purification and electrophoretic analysis, the presence of viroids other than CEV was demonstrated. Screening of over 1800 trees resulted in the detection of four groups of viroids as determined by RNA homology and length in nucleotides. CEV was the only member of the first group and was the largest viroid detected, with 371 bases. CEV almost invariably occurred with other viroids of lower molecular weight. The second group, CV I, contained two viroids, of 325 and 332 bases, which were not homologous to CEV or hop stunt viroid (HSVd). Group CV II contained three viroids, of 297, 299 and 302 bases, all of which were homologous to HSVd. The CV III group contained two viroids, of 290 and 295 bases, which were not homologous to CEV or HSVd. The CV II and CV III groups were strongly associated with various field symptoms, including the disease cachexia (CV IIb), and dwarfing of trees on trifoliate orange rootstock. To confirm any causal relationship between individual viroids and field symptoms, viroids representative of each group were purified and used to inoculate Etrog citron. Tissue from these citrons was used to inoculate young field trees with viroids, singly and in combination. Parson's Special mandarin inoculated with CV IIb showed symptoms of cachexia after 2 years, confirming CV IIb as a causal agent of this disease. It will probably be 5 years before the effect of other inoculations can be assessed.
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Diener, Theodor. „Of Viroids and Prions“. Viruses 10, Nr. 12 (23.11.2018): 663. http://dx.doi.org/10.3390/v10120663.
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In 2017, Hadidi et al. edited a voluminous monograph entitled “Viroids and Satellites”, in which each known viroid and viroid-like satellite species was described in detail from many perspectives by more than 100 experts from 24 countries. In its 700+ pages, the book is a much needed detailed and reliable compendium of a subject, which, undoubtedly, is still little known by many potential readers. Because most users of the book may be expected to be practical plant pathologists, it appears essential that the book contain, in addition to the detailed viroid and satellite descriptions, one chapter, in which the basic molecular biology of viroids and satellites is described.
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Matsushita, Yosuke, Hironobu Yanagisawa und Teruo Sano. „Vertical and Horizontal Transmission of Pospiviroids“. Viruses 10, Nr. 12 (12.12.2018): 706. http://dx.doi.org/10.3390/v10120706.
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Viroids are highly structured, single-stranded, non-protein-coding circular RNA pathogens. Some viroids are vertically transmitted through both viroid-infected ovule and pollen. For example, potato spindle tuber viroid, a species that belongs to Pospiviroidae family, is delivered to the embryo through the ovule or pollen during the development of reproductive tissues before embryogenesis. In addition, some of Pospiviroidae are also horizontally transmitted by pollen. Tomato planta macho viroid in pollen infects to the ovary from pollen tube during pollen tube elongation and eventually causes systemic infection, resulting in the establishment of horizontal transmission. Furthermore, fertilization is not required to accomplish the horizontal transmission. In this review, we will overview the recent research progress in vertical and horizontal transmission of viroids, mainly by focusing on histopathological studies, and also discuss the impact of seed transmission on viroid dissemination and seed health.
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Thomas, Tina P., Madhurababu Kunta, John V. da Graça, Mamadou Sétamou, Mani Skaria und Apurba Bhattacharya. „Suppression of Phytophthora Infection in Citrus Infected with Viroids“. HortScience 45, Nr. 7 (Juli 2010): 1069–72. http://dx.doi.org/10.21273/hortsci.45.7.1069.
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Citrus viroid-induced resistance to Phytophthora infection in citrus was measured by the number of Phytophthora sporangia in ‘Rio Red’ grapefruit (Citrus paradisi Macf.) bait tissue infected with citrus viroids compared with non-inoculated controls. Different viroid isolates containing mixtures of viroids [Citrus exocortis viroid (CEVd), Hop stunt viroid (HSVd), Citrus viroid III (CVd-III), Citrus viroid IV (CVd-IV)] were designated by plant numbers and sources. Source 13E was associated with the lowest number of sporangia in bark, leaves, and roots used as baits, whereas CEVd E9, a known severe CEVd isolate, significantly reduced the number of sporangia in leaves and bark. Sources 1A, 2E, 3E, 4D, and 6E showed a significantly reduced number of sporangia on bark, leaves, and roots compared with healthy plants and 44A; however, their effect was not as pronounced as that of E9 and 13E. Sources 12E and 44A did not suppress sporangia production. Previous reverse transcriptase–polymerase chain reaction analysis showed that all source plants had mixed infections with several viroids, whereas 12E and 44A contained no viroids. In addition to confirming the earlier reports on the suppression of Phytophthora infection in general, our study showed significantly reduced Phytophthora sporangia development resulting from a number of viroids in mixed infection, but there did not appear to be any effect related to viroid species. To determine if concentration affected resistance to Phytophthora, phenolic acids were extracted. Extraction of phenolic acids with 80% ethanol was more efficient compared with 100% methanol and an acetonitrile–water mixture. High-performance liquid chromatography revealed no notable detection of salicylic acid in healthy and viroid-infected plants, but there was a small peak corresponding to salicylic acid in Phytophthora-infected and both viroid and Phytophthora-infected plants. Flavone was detected in all the source plants with a slight increase in Phytophthora-infected and both viroid and Phytophthora-infected plants. A peak corresponding to quercetin dehydrate was detected in Phytophthora-infected plants. Efficient use of the right viroid isolate(s) can result in suppression of Phytophthora infection of citrus.
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Sänger, Heinz L. „VIROIDS AND VIROID DISEASES“. Acta Horticulturae, Nr. 234 (Dezember 1988): 79–88. http://dx.doi.org/10.17660/actahortic.1988.234.9.
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Vernière, C., X. Perrier, C. Dubois, A. Dubois, L. Botella, C. Chabrier, J. M. Bové und N. Duran Vila. „Interactions Between Citrus Viroids Affect Symptom Expression and Field Performance of Clementine Trees Grafted on Trifoliate Orange“. Phytopathology® 96, Nr. 4 (April 2006): 356–68. http://dx.doi.org/10.1094/phyto-96-0356.
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Citrus exocortis viroid (CEVd), Citrus bent leaf viroid (CBLVd), a noncachexia variant of Hop stunt viroid (HSVd), Citrus viroid III (CVd-III), and Citrus viroid IV (CVd-IV) were co-inoculated as two-, three-, four-, and five-viroid mixtures to Clementine trees grafted on trifoliate orange to evaluate their effect on symptom expression, tree growth, and fruit yield. Most trees infected with CEVd-containing viroid mixtures developed exocortis scaling symptoms, as did CEVd alone, whereas most trees infected with HSVd- or CVd-IV-containing mixtures developed bark-cracking symptoms. Trees infected with mixtures containing both CEVd and CVd-IV revealed the existence of antagonism between these two viroids in terms of the expected bark-scaling and cracking symptoms. Synergistic interactions also were identified in trees infected with certain viroid combinations that, in spite of lacking CEVd, expressed exocortis-like scaling symptoms. Viroid interactions also affected the expected response of trees in terms of vegetative growth and fruit yield. Trees infected with viroid combinations containing CEVd or CVd-III were smaller and produced less fruit than trees infected with mixtures not containing these viroids. Viroid interactions on scion circumference and cumulative fruit yield, in terms of additivity of their effects, were statistically confirmed using a factorial analysis of variance model with two mean estimation approaches. In single-viroid infections, CEVd, CVd-III, and, to a lesser extent, CBLVd consistently and significantly reduced tree size and fruit yield. Conversely, HSVd and CVd-IV slightly increased fruit yield and reduced scion circumference. Rare and not consistent significant interactions were detected with the five-, four-, and three-viroid combinations. Antagonistic interactions between CEVd and CVd-III or CBLVd and CVd-III were revealed over the years with consistent significance. The antagonistic interaction between CEVd and CVd-IV was highly significant over the years when additional viroids were present; however, this antagonism appeared much later in the case of an exclusive interaction. HSVd and CVd-IV showed a consistent and significant synergistic interaction on yield only when both viroids were exclusively present. These results demonstrate antagonistic or synergistic relationships between citrus viroids depending on the viroid mixtures present in the host.
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Hadidi, Ahmed. „Next-Generation Sequencing and CRISPR/Cas13 Editing in Viroid Research and Molecular Diagnostics“. Viruses 11, Nr. 2 (29.01.2019): 120. http://dx.doi.org/10.3390/v11020120.
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Viroid discovery as well as the economic significance of viroids and biological properties are presented. Next-generation sequencing (NGS) technologies combined with informatics have been applied to viroid research and diagnostics for almost a decade. NGS provides highly efficient, rapid, low-cost high-throughput sequencing of viroid genomes and of the 21–24 nt vd-sRNAs generated by the RNA silencing defense of the host. NGS has been utilized in various viroid studies which are presented. The discovery during the last few years that prokaryotes have heritable adaptive immunity mediated through clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated Cas proteins, have led to transformative advances in molecular biology, notably genome engineering and most recently molecular diagnostics. The potential application of the CRISPR-Cas13a system for engineering viroid interference in plants is suggested by targeting specific motifs of three economically important viroids. The CRISPR-Cas13 system has been utilized recently for the accurate detection of human RNA viruses by visual read out in 90 min or less and by paper-based assay. Multitarget RNA tests by this technology have a good potential for application as a rapid and accurate diagnostic assay for known viroids. The CRISPR/Cas system will work only for known viroids in contrast to NGS, but it should be much faster.
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Di Serio, Francesco, Enza Maria Torchetti, José-Antonio Daròs und Beatriz Navarro. „Reassessment of Viroid RNA Cytosine Methylation Status at the Single Nucleotide Level“. Viruses 11, Nr. 4 (18.04.2019): 357. http://dx.doi.org/10.3390/v11040357.
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Composed of a few hundreds of nucleotides, viroids are infectious, circular, non-protein coding RNAs able to usurp plant cellular enzymes and molecular machineries to replicate and move in their hosts. Several secondary and tertiary RNA structural motifs have been implicated in the viroid infectious cycle, but whether modified nucleotides, such as 5C-methylcytosine (m5C), also play a role has not been deeply investigated so far. Here, the possible existence of m5C in both RNA polarity strands of potato spindle tuber viroid and avocado sunblotch viroid -which are representative members of the nucleus- and chloroplast-replicating viroids, respectively- has been assessed at single nucleotide level. We show that a standard bisulfite protocol efficiently used for identifying m5C in cellular RNAs may generate false positive results in the case of the highly structured viroid RNAs. Applying a bisulfite conversion protocol specifically adapted to RNAs with high secondary structure, no m5C was identified in both polarity strands of both viroids, indicating that this specific nucleotide modification does not likely play a role in viroid biology.
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Adkar-Purushothama, Charith Raj, und Jean-Pierre Perreault. „Impact of Nucleic Acid Sequencing on Viroid Biology“. International Journal of Molecular Sciences 21, Nr. 15 (01.08.2020): 5532. http://dx.doi.org/10.3390/ijms21155532.
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The early 1970s marked two breakthroughs in the field of biology: (i) The development of nucleotide sequencing technology; and, (ii) the discovery of the viroids. The first DNA sequences were obtained by two-dimensional chromatography which was later replaced by sequencing using electrophoresis technique. The subsequent development of fluorescence-based sequencing method which made DNA sequencing not only easier, but many orders of magnitude faster. The knowledge of DNA sequences has become an indispensable tool for both basic and applied research. It has shed light biology of viroids, the highly structured, circular, single-stranded non-coding RNA molecules that infect numerous economically important plants. Our understanding of viroid molecular biology and biochemistry has been intimately associated with the evolution of nucleic acid sequencing technologies. With the development of the next-generation sequence method, viroid research exponentially progressed, notably in the areas of the molecular mechanisms of viroids and viroid diseases, viroid pathogenesis, viroid quasi-species, viroid adaptability, and viroid–host interactions, to name a few examples. In this review, the progress in the understanding of viroid biology in conjunction with the improvements in nucleotide sequencing technology is summarized. The future of viroid research with respect to the use of third-generation sequencing technology is also briefly envisaged.
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Góra-Sochacka, Anna. „Viroids: unusual small pathogenic RNAs.“ Acta Biochimica Polonica 51, Nr. 3 (30.09.2004): 587–607. http://dx.doi.org/10.18388/abp.2004_3546.
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Viroids are small (about 300 nucleotides), single-stranded, circular, non-encapsidated pathogenic RNA molecules. They do not code for proteins and thus depend on plant host enzymes for their replication and other functions. They induce plant diseases by direct interaction with host factors but the mechanism of pathogenicity is still unknown. They can alter the expression of selected plant genes important for growth and development. Viroids belong to two families, the Avsunviroidae and the Pospiviroidae. Viroids of the Avsunviroidae family adopt a branched or quasi rod-like secondary structure in their native state. Members of the Pospiviroidae family adopt a rod-like secondary structure. In such native structures five structural/functional domains have been identified: central (C), pathogenicity, variable and two terminal domains. The central conserved region (CCR) within the C domain characterizes viroids of the Pospiviroidae. Specific secondary structures of this region play an important role in viroid replication and processing. Viroids of the Avsunviroidae family lack a CCR but possess self-cleaving properties by forming hammerhead ribozyme structures; they accumulate and replicate in chloroplasts, whereas members of the Pospiviroidae family have a nuclear localization. Viroid replication occurs via a rolling circle mechanism using either a symmetric or asymmetric pathway in three steps, RNA transcription, processing and ligation.
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Joubert, Melissa, Noëlani van den Berg, Jacques Theron und Velushka Swart. „Transcriptomics Advancement in the Complex Response of Plants to Viroid Infection“. International Journal of Molecular Sciences 23, Nr. 14 (12.07.2022): 7677. http://dx.doi.org/10.3390/ijms23147677.
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Viroids are the smallest plant pathogens, consisting of a single-stranded circular RNA of less than 500 ribonucleotides in length. Despite their noncoding nature, viroids elicit disease symptoms in many economically important plant hosts, and are, thus, a class of pathogens of great interest. How these viroids establish disease within host plants, however, is not yet fully understood. Recent transcriptomic studies have revealed that viroid infection influences the expression of genes in several pathways and processes in plants, including defence responses, phytohormone signalling, cell wall modification, photosynthesis, secondary metabolism, transport, gene expression and protein modification. There is much debate about whether affected pathways signify a plant response to viroid infection, or are associated with the appearance of disease symptoms in these interactions. In this review, we consolidate the findings of viroid–host transcriptome studies to provide an overview of trends observed in the data. When considered together, changes in the gene expression of different hosts upon viroid infection reveal commonalities and differences in diverse interactions. Here, we discuss whether trends in host gene expression can be correlated to plant defence or disease development during viroid infection, and highlight avenues for future research in this field.
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Aviña-Padilla, Katia, Erika Janet Zamora-Macorra, Daniel Leobardo Ochoa-Martínez, Flor Citlally Alcántar-Aguirre, Maribel Hernández-Rosales, Loranda Calderón-Zamora und Rosemarie W. Hammond. „Mexico: A Landscape of Viroid Origin and Epidemiological Relevance of Endemic Species“. Cells 11, Nr. 21 (03.11.2022): 3487. http://dx.doi.org/10.3390/cells11213487.
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Viroids are single-stranded, circular RNA molecules (234-406 nt) that infect a wide range of crop species and cause economic losses in agriculture worldwide. They are characterized by the existence of a population of sequence variants, attributed to the low fidelity of RNA polymerases involved in their transcription, resulting in high mutation rates. Therefore, these biological entities exist as quasispecies. This feature allows them to replicate within a wide range of host plants, both monocots and dicots. Viroid hosts include economically important crops such as tomato, citrus, and fruit trees such as peach and avocado. Given the high risk of introducing viroids to viroid disease-free countries, these pathogens have been quarantined globally. As discussed herein, Mexico represents a geographical landscape of viroids linked to their origin and comprises considerable biodiversity. The biological features of viroid species endemic to Mexico are highlighted in this communication. In addition, we report the phylogenetic relationships among viroid and viroid strains, their economic impact, geographical distribution, and epidemiological features, including a broad host range and possible long-distance, seed, or insect-mediated transmission. In summary, this review could be helpful for a better understanding of the biology of viroid diseases and future programs on control of movement and spread to avoid economic losses in agricultural industries.
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Steinbachová, Lenka, Jaroslav Matoušek, Gerhard Steger, Helena Matoušková, Sebastjan Radišek und David Honys. „Transformation of Seed Non-Transmissible Hop Viroids in Nicotiana benthamiana Causes Distortions in Male Gametophyte Development“. Plants 10, Nr. 11 (06.11.2021): 2398. http://dx.doi.org/10.3390/plants10112398.
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Viroids are small, non-coding, parasitic RNAs that promote developmental distortions in sensitive plants. We analyzed pollen of Nicotiana benthamiana after infection and/or ectopic transformation with cDNAs of citrus bark cracking viroid (CBCVd), apple fruit crinkle viroid (AFCVd) and potato spindle tuber viroid (PSTVd) variant AS1. These viroids were seed non-transmissible in N. benthamiana. All viroids propagated to high levels in immature anthers similar to leaves, while their levels were drastically reduced by approximately 3.6 × 103, 800 and 59 times in mature pollen of CBCVd, AFCVd and PSTVd infected N. benthamiana, respectively, in comparison to leaves. These results suggest similar elimination processes during male gametophyte development as in the Nicotiana tabacum we presented in our previous study. Mature pollen of N. benthamiana showed no apparent defects in infected plants although all three viroids induced strong pathological symptoms on leaves. While Nicotiana species have naturally bicellular mature pollen, we noted a rare occurrence of mature pollen with three nuclei in CBCVd-infected N. benthamiana. Changes in the expression of ribosomal marker proteins in AFCVd-infected pollen were detected, suggesting some changes in pollen metabolism. N. benthamiana transformed with 35S-driven viroid cDNAs showed strong symptoms including defects in pollen development. A large number of aborted pollen (34% and 62%) and a slight increase of young pollen grains (8% and 15%) were found in mature pollen of AFCVd and CBCVd transformants, respectively, in comparison to control plants (3.9% aborted pollen and 0.3% young pollen). Moreover, pollen grains with malformed nuclei or trinuclear pollen were found in CBCVd-transformed plants. Our results suggest that “forcing” overexpression of seed non-transmissible viroid led to strong pollen pathogenesis. Viroid adaptation to pollen metabolism can be assumed as an important factor for viroid transmissibility through pollen and seeds.
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Flores, Ricardo, Carmen Hernández, A. Emilio Martínez de Alba, José-Antonio Daròs und Francesco Di Serio. „Viroids and Viroid-Host Interactions“. Annual Review of Phytopathology 43, Nr. 1 (September 2005): 117–39. http://dx.doi.org/10.1146/annurev.phyto.43.040204.140243.
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Hodgson, R. A. J., G. C. Wall und J. W. Randles. „Specific Identification of Coconut Tinangaja Viroid for Differential Field Diagnosis of Viroids in Coconut Palm“. Phytopathology® 88, Nr. 8 (August 1998): 774–81. http://dx.doi.org/10.1094/phyto.1998.88.8.774.
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Tinangaja is a widespread lethal disease of putative viroid etiology affecting coconut palm on the island of Guam. Determination of its distribution and mode of spread requires a simple and reliable diagnostic procedure that is specific for the associated coconut tinangaja viroid (CTiVd). A method of extracting tissue followed by analytical agarose gel electrophoresis for CTiVd detection has been developed and used to identify the viroid in leaf samples of suspect symptomatic palms growing in the field. Two-dimensional polyacrylamide gel electrophoresis showed that the viroid band contained circular molecules that are typical for viroids. Confirmation of the identity of CTiVd and detection of low levels of viroid below the threshold of detection by agarose gel electrophoresis was achieved either by diagnostic oligonucleotide-probe (DOP) hybridization assay or by reverse-transcription polymerase chain reaction (RT-PCR) with the oligonucleotide probe as one of the two PCR primers. RT-PCR was not substantially more sensitive than DOP-hybridization assay. This procedure also was applicable to coconut cadang-cadang viroid (CCCVd), and oligonucleotide probes designed to be specific for either CTiVd or CCCVd distinguished between these two viroids in coconut leaf extracts. This strategy provides a rapid and specific indexing procedure for the two characterized viroids of coconut palm and will be applicable to further studies on the viroid-like sequences previously reported in tropical monocotyledons.
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Bang, Yoon Hyun, Eun Gyeong Song, Younghye Lee und Ki Hyun Ryu. „Occurrence of Viruses and Viroids in Chrysanthemum Plants (Dendranthema morifolium) Cultivated in Yesan-gun, Chungcheongnam-do in Korea“. Research in Plant Disease 28, Nr. 4 (31.12.2022): 237–44. http://dx.doi.org/10.5423/rpd.2022.28.4.237.
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Chrysanthemum plants are one of the most economically important plants in South Korea. Both virus and viroid can cause diseases and economic damage to the plants. In this study, we investigated the detection of seven viruses and two viroids in 350 chrysanthemum plants cultivated in Yesan-gun, Chungcheongnam-do. Two viruses, chrysanthemum virus B (CVB) and tomato aspermy virus (TAV), and two viroids, chrysanthemum chlorotic mottle viroid (CChMVd) and chrysanthemum stunt viroid (CSVd), were detected in this study. The two viruses were detected in six samples and one sample, respectively. The two viroids were detected in 97 samples and 21 samples, respectively. The nucleotide sequences of the CVB-CN-Y, TAV-CN-Y, CChMVd-CN-Y, and CSVd-CN-Y obtained in this study showed 83.7–86.9%, 99.2–100.0%, 94.4–99.5%, and 95.7–99.7% identity, respectively, compared to their other strains/isolates. The CVB-CN-Y and TAV-CN-Y showed the greatest nucleotide sequence homology to CVB-GS1 and three TAV isolates (TAV-V, TAV-P, and TAV-ChJ), respectively. The CChMVd-CN-Y and CSVd-CN-Y showed the greatest nucleotide sequence homology to CChMVd-Horst and four CSVd isolates (Au1.1, K4pop, Sagae, and Tochigi), respectively. This study is the report on the infection rate of viruses and viroids in chrysanthemum plants cultivated in Yesan-gun in 2021.
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Cottilli, Patrick, Borja Belda-Palazón, Charith Raj Adkar-Purushothama, Jean-Pierre Perreault, Enrico Schleiff, Ismael Rodrigo, Alejandro Ferrando und Purificación Lisón. „Citrus exocortis viroid causes ribosomal stress in tomato plants“. Nucleic Acids Research 47, Nr. 16 (08.08.2019): 8649–61. http://dx.doi.org/10.1093/nar/gkz679.
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Abstract Viroids are naked RNAs that do not code for any known protein and yet are able to infect plants causing severe diseases. Because of their RNA nature, many studies have focused on the involvement of viroids in RNA-mediated gene silencing as being their pathogenesis mechanism. Here, the alterations caused by the Citrus exocortis viroid (CEVd) on the tomato translation machinery were studied as a new aspect of viroid pathogenesis. The presence of viroids in the ribosomal fractions of infected tomato plants was detected. More precisely, CEVd and its derived viroid small RNAs were found to co-sediment with tomato ribosomes in vivo, and to provoke changes in the global polysome profiles, particularly in the 40S ribosomal subunit accumulation. Additionally, the viroid caused alterations in ribosome biogenesis in the infected tomato plants, affecting the 18S rRNA maturation process. A higher expression level of the ribosomal stress mediator NAC082 was also detected in the CEVd-infected tomato leaves. Both the alterations in the rRNA processing and the induction of NAC082 correlate with the degree of viroid symptomatology. Taken together, these results suggest that CEVd is responsible for defective ribosome biogenesis in tomato, thereby interfering with the translation machinery and, therefore, causing ribosomal stress.
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Seo, Hyesu, Kyunghee Kim und Woong June Park. „Effect of VIRP1 Protein on Nuclear Import of Citrus Exocortis Viroid (CEVd)“. Biomolecules 11, Nr. 1 (13.01.2021): 95. http://dx.doi.org/10.3390/biom11010095.
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Before replicating, Pospiviroidae viroids must move into the plant nucleus. However, the mechanisms of viroid nuclear import are not entirely understood. To study the nuclear import of viroids, we established a nuclear import assay system using onion cell strips and observed the import of Alexa Fluor-594-labeled citrus exocortis viroid (CEVd). To identify the plant factors involved in the nuclear import of viroids, we cloned the Viroid RNA-binding Protein 1 (VIRP1) gene from a tomato cultivar, Seokwang, and heterologously expressed and purified the VIRP1 protein. The newly prepared VIRP1 protein had alterations of amino acid residues at two points (H52R, A277G) compared with a reference VIRP1 protein (AJ249595). VIRP1 specifically bound to CEVd and promoted its nuclear import. However, it is still uncertain whether VIRP1 is the only factor required for the nuclear import of CEVd because CEVd entered the plant nuclei without VIRP1 in our assay system. The cause of the observed nuclear accumulation of CEVd in the absence of VIRP1 needs to be further clarified.
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Rizza, S., A. Catara, X. F. Ma und Z. Deng. „Detection of Multiple Infections of Citrus exocortis viroid, Citrus viroid III, and Hop stunt viroid Variants in Hunan Province, China“. Plant Disease 91, Nr. 9 (September 2007): 1205. http://dx.doi.org/10.1094/pdis-91-9-1205a.
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Citrus cultivation in China has increased since the late 1970s, with China now having the largest area of citrus in culture in the world that is spread in 22 provinces and municipalities. Hunan Province has undergone a program to become one of the major citrus producers in China. Poncirus trifoliata is the main rootstock, so citrus viroids are a limiting factor for further citriculture development. In mainland China, only the presence of Citrus exocortis viroid (CEVd) has been reported from Etrog citron indexing, sPAGE (sequential polyacrylamide gel electrophoresis) analysis (2), and reverse transcription (RT)-PCR (3). Three viroid-like RNAs, a1, b1, and d, based on sPAGE patterns were detected years ago in our laboratory in budsticks received from Sichuan Province. To identify different viroids and determine their distribution, a survey has been undertaken. Field trees showing stunting, bark scaling and cracking of the rootstock, and poor yield were tested using biological indexing and PCR for the most frequent citrus viroids. Samples from six trees of a local sweet orange variety and three of a Clementine variety introduced from abroad, both grafted on P. trifoliata and showing a variable degree of bark scaling and cracking, were collected near Changsha and in the County of Xin Ning at the end of summer 2006. Small pieces of bark were inserted in stems of young E. citron budwood grafted on rough lemon and maintained in a warm greenhouse (24 to 32°C). Indexing on E. citron showed mild epinasty and leaf roll typical of citrus viroid infections. To identify specific viroids, bark was ground to a fine powder with liquid nitrogen and total RNA was extracted with TRIZOL Reagent (Invitrogen, San Diego, CA) and tested by RT-PCR to detect CEVd, Hop Stunt viroid (HSVd), and Citrus viroid III (CVd-III), as well as to identify the cachexia variants of HSVd. Four primer pairs were used to test the RNA extracts by RT-PCR (1). All samples were infected by HSVd, eight with CVd-III, and six with CEVd. The cachexia variants of HSVd were detected in four of nine samples. Mixed infections were as follows: one sample had CEVd and HSVd, eight had HSVd and CVd-III, and five were infected by the three viroids. A second sampling 3 months after inoculation gave the same amplification patterns. The results show that at least three viroids are present in citrus orchards in Hunan Province. To our knowledge, this is the first report of cachexia variants of HSVd and CVd-III in China. The common occurrence of these viroids supports the need for proper indexing of mother trees and a specific shoot tip grafting program to create healthy budwood sources to provide healthy plants. References: (1) L. Bernard and N. Duran-Vila. Mol. Cell. Probes, 20:105, 2006. (2) L. Han et al. Viroids. CSIRO Publishing, Melbourne, 283, 2003. (3). Q. Hu et al. Acta Bot. Sin. 39:613, 1997.
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Stuchi, Eduardo Sanches, Simone Rodrigues da Silva, Luiz Carlos Donadio, Otávio Ricardo Sempionato und Eduardo Toller Reiff. „Field performance of "marsh seedless" grapefruit on trifoliate orange inoculated with viroids in Brazil“. Scientia Agricola 64, Nr. 6 (Dezember 2007): 582–88. http://dx.doi.org/10.1590/s0103-90162007000600004.
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Some viroids reduce citrus tree growth and may be used for tree size control aiming the establishment of orchards with close tree spacing that may provide higher productivity than conventional ones. To study the effects of citrus viroids inoculation on vegetative growth, yield and fruit quality of 'Marsh Seedless' grapefruit (Citrus paradisi Macf.) grafted on trifoliate orange [Poncirus trifoliata (L.) Raf.], an experiment was set up in January 1991, in Bebedouro, São Paulo State, Brazil. The experimental design was randomized blocks with four treatments with two plants per plot: viroid isolates Citrus Exocortis Viroid (CEVd) + Hop stunt viroid (HSVd - CVd-II, a non cachexia variant) + Citrus III viroid (CVd-III) and Hop stunt viroid (HSVd - CVd-II, a non cachexia variant) + Citrus III viroid (CVd-III) and controls: two healthy buds (control), and no grafting (absolute control). Inoculation was done in the field, six months after planting by bud grafting. Both isolates reduced tree growth (trunk diameter, plant height, canopy diameter and volume). Trees not inoculated yielded better (average of eleven harvests) than inoculated ones but the productivity was the same after 150 months. Fruit quality was affected by viroids inoculation but not in a restrictive way. The use of such severe dwarfing isolates for high density plantings of grapefruit on trifoliate orange rootstock is not recommended.
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Najar, A., und N. Duran-Vila. „Viroid Prevalence in Tunisian Citrus“. Plant Disease 88, Nr. 11 (November 2004): 1286. http://dx.doi.org/10.1094/pdis.2004.88.11.1286b.
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The citrus industry in Tunisia is based mainly on the production of local cultivars of sweet orange (Citrus sinensis), common mandarin (C. reticulata), clementine (C. clementina), and lemon (C. limon). Sour orange (C. aurantium) is the only rootstock presently being used in the major growing area located at Cap Bon where 80% of citrus is being produced. The presence of tristeza disease in the Mediterranean basin is a threat to the Tunisian citrus industry, and new rootstocks giving tristeza tolerant rootstock/scion combinations are urgently needed as an alternative to sour orange. Since some promising rootstocks are known to be sensitive to viroids (1), a survey was conducted to determine if the cultivars grown presently in Tunisia were infected with viroids. Following a preliminary report (2), an extensive survey was conducted from 1995-2001 that included 174 symptomless sources being grown at Cap Bon: 26 Maltaise demi-sanguine, 9 Maltaise sanguine, 20 Maltaise blonde, 4 orange doublefine, 16 Washington navel, 12 Valencia late, 29 common mandarin, 42 Cassar clementine, 5 Lunari lemon, and 11 Eureka lemon. These sources were graft-inoculated into Etrog citron that subsequently developed symptoms characteristic of viroid infection. Sequential polyacrylamide gel electrophoresis analysis and molecular hybridization using viroid-specific probes (1) revealed that all sources were infected with at least two viroids. Citrus exocortis viroid (CEVd), Hop stunt viroid (HSVd), and Citrus viroid III (CVd-III) were widespread and accounted for 68.4, 67.8, and 81.0% of the sources tested, respectively. Citrus bent leaf viroid (CBLVd) and Citrus viroid IV (CVd-IV) were only found in 32.7 and 2.3% of the sources. The most frequent viroid combinations were CEVd+HSVd+CVd-III (17.8%) and CEVd+CVd-III (17,2%), whereas HSVd+CVd-IV and CEVd+CBLVd+CVd-III+CVd-IV were found in a single source (0.6%). References: (1) N. Duran-Vila and J. S. Semancik. Pages 178–194 in: Viroids. CSIRO Publishing, Australia, 2003. (2) A. Najar et al. Pages 398–400 in: Proc. 15th Conf. Int. Org. Citrus Virol, 2002.
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Singh, R. P., A. D. Dilworth, V. K. Baranwal und K. N. Gupta. „Detection of Citrus exocortis viroid, Iresine viroid, and Tomato chlorotic dwarf viroid in New Ornamental Host Plants in India“. Plant Disease 90, Nr. 11 (November 2006): 1457. http://dx.doi.org/10.1094/pd-90-1457a.
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Columnea latent viroid, originating from ornamental plants, is known to be harmful to crop plants (2). Despite the potential threat to crop plants, the importance of ornamental plants in viroid evolution is not fully appreciated. Availability of a Pospiviroid genus-specific primer pair (1) to detect the most prevalent viroids in ornamental plants and a simplified nucleic acid preparation protocol (3) for use in reverse-transcription polymerase chain reaction (RT-PCR) have facilitated surveys of ornamental plants for pospiviroids. Using the above protocol in India, leaf and shoot samples were collected randomly from roadside beds consisting of ground covers or creepers/trailing plants at the IARI campus, New Delhi. These were extracted in 50 mM NaOH + 2.5 mM EDTA solution, centrifuged to sediment the coarse debris from sap, and 10 μl of the supernatant was spotted on a nitrocellulose membrane. Individual spots were eluted with distilled sterile water (30 μl) and the eluates were used for RT-PCR detection of viroids (3). Amplified fragments or subsequently cloned plasmids were also purified using NaOH-EDTA membrane protocol. Cloning and sequencing of amplicons (195 to 224 bp) revealed a very high sequence identity with specific viroids from the viroid sequence database (NCBI). Among the 19 plant samples assayed, only three plant species were infected by viroids, although none of them exhibited any symptoms. The three plant species included: (i) moss verbena, Glandularia puchella (Verbenaceae, native to Argentina and Chile, now established in several regions of the world), infected with a viroid (Accession No. DQ846884) having 99% sequence identity to Citrus exocortis viroid (CEVd) (Accession No. S67446); (ii) trailing verbena, Verbena × hybrida (Verbenaceae, ornamental plant), doubly infected with a viroid (Accession No. DQ846885) having 95% sequence identity to CEVd (Accession No. DQ094297) and infected with another viroid (Accession No. DQ846883) having 98% sequence identity to Tomato chlorotic dwarf viroid (TCDVd) (Accession No. AF162131); and (iii) red joyweed, Alternanthera sessilis (Amaranthaceae, a perennial weed herb) infected with a viroid (Accession No. DQ846886) having 96% sequence identity to Iresine viroid (IrVd) (Accession No. DQ094293). CEVd and TCDVd were mechanically transferred to tomato seedlings causing reduced growth of plants, smaller leaves, and bunchy-top appearance of plants, symptoms similar to those typically observed with other isolates of these viroids. As expected from previous studies, IrVd was not transmitted to tomato plants. Natural infection of verbena with CEVd has been detected in North America (2) but this was a novel observation in India. Additional novel observations include: A. sessilis as a new host for IrVd; and TCDVd is the first crop viroid to be isolated from a naturally infected ornamental plant. The significance of these viroid findings in ground cover and widely grown ornamental plants may lie in their potential role in spreading the viroids to citrus plants in citrus-growing countries such as India. References: (1) H. Bostan et al. J. Virol. Methods 116:189, 2004, (2) R. P. Singh and J. A. Teixeira da Silva. Floriculture, Ornamental Plant Biotechnol. 3:531, 2006. (3) R. P. Singh et al. J. Virol. Methods 132:204, 2006.
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Tian, Mengyuan, Shuang Wei, Ruiling Bian, Jingxian Luo, Haris Ahmed Khan, Huanhuan Tai, Hideki Kondo, Ahmed Hadidi, Ida Bagus Andika und Liying Sun. „Natural Cross-Kingdom Spread of Apple Scar Skin Viroid from Apple Trees to Fungi“. Cells 11, Nr. 22 (20.11.2022): 3686. http://dx.doi.org/10.3390/cells11223686.
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Viroids are the smallest known infectious agents that are thought to only infect plants. Here, we reveal that several species of plant pathogenic fungi that were isolated from apple trees infected with apple scar skin viroid (ASSVd) carried ASSVd naturally. This finding indicates the spread of viroids to fungi under natural conditions and further suggests the possible existence of mycoviroids in nature. A total of 117 fungal isolates were isolated from ASSVd-infected apple trees, with the majority (85.5%) being an ascomycete Alternaria alternata and the remaining isolates being other plant-pathogenic or -endophytic fungi. Out of the examined samples, viroids were detected in 81 isolates (69.2%) including A. alternata as well as other fungal species. The phenotypic comparison of ASSVd-free specimens developed by single-spore isolation and ASSVd-infected fungal isogenic lines showed that ASSVd affected the growth and pathogenicity of certain fungal species. ASSVd confers hypovirulence on ascomycete Epicoccum nigrum. The mycobiome analysis of apple tree-associated fungi showed that ASSVd infection did not generally affect the diversity and structure of fungal communities but specifically increased the abundance of Alternaria species. Taken together, these data reveal the occurrence of the natural spread of viroids to plants; additionally, as an integral component of the ecosystem, viroids may affect the abundance of certain fungal species in plants. Moreover, this study provides further evidence that viroid infection could induce symptoms in certain filamentous fungi.
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Wang, X. F., C. Y. Zhou, K. Z. Tang und Z. A. Li. „Occurrence of Four Citrus Viroids in Chongqing, China“. Plant Disease 92, Nr. 6 (Juni 2008): 978. http://dx.doi.org/10.1094/pdis-92-6-0978b.
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Chongqing Municipality, located in the superior citrus belt of the upper-middle Yangtze River, is one of the most important citrus-producing areas in China. A survey was performed to evaluate the occurrence and distribution of citrus viroids in this area, where Poncirus trifoliata is the main rootstock. From 2002 to 2006, 72 samples of sweet oranges (Citrus sinensis), lemons (C. jambhiri), mandarins (C. reticulata), and mandarin hybrids, which showed stunting, bark scaling, and cracking symptoms on the rootstock, were collected and graft inoculated into Arizona 861-S1 Etrog citron (C. medica) on rough lemon rootstock and maintained in a greenhouse at 28 to 32°C. Fifty-one of the seventy-two samples were cultivars imported from abroad, and the remaining samples were all local cultivars. Sixty samples induced symptoms typical of citrus viroids on the Etrog indicator plants. To identify the causal agent(s), a one-step reverse transcription (RT)-PCR protocol, using five primer pairs (1) targeting the complete genome sequences, was used to detect Citrus exocortis viroid (CEVd), Citrus bent leaf viroid (CBLVd), Hop stunt viroid (HSVd), Citrus viroid III (CVd-III), and Citrus viroid IV (CVd-IV) (2). All 72 samples were infected by citrus viroids. Sixty-five and thirty-five of the seventy-two samples were positive for HSVd and CVd-III, respectively. CEVd and CBLVd were found, respectively, in 20 and 11 of 72 samples, whereas CVd-IV was not detected. Of 72 samples, 12 without typical symptoms on Etrog citrons were infected by HSVd and CBLVd. Nearly all (70 of 72) infected citrus plants harbored more than one viroid species, and two plants were both infected by CEVd, CBLVd, HSVd, and CVd-III. RT-PCR products were purified and ligated into pGEM T-Easy Vector (Promega, Madison, WI) and three clones for each of the four viroid species were sequenced and deposited in GenBank: CEVd (Accession Nos. EU382202, U382203, and EU382204); CBLVd (Accession Nos. EU382211, EU382212, and EU382213); HSVd (Accession Nos. EU382208, EU382209, and EU382210); and CVd-III (Accession Nos. EU382205, EU382206, and EU382207). BLAST analysis showed that these nucleotide sequences had greater than 94% nucleotide identity to the most similar genome sequences in GenBank. Sweet orange was more frequently infected by viroids than the other citrus cultivars. To our knowledge, this is the first report of CBLVd, HSVd, and CVd-III in Chongqing and the first report of CBLVd in China. The high incidence of citrus viroids in Chongqing necessitates rapid development of a system of propagation and testing to reduce the incidence of viroids and the associated loss of citrus production. References: (1) L. Bernard and N. Duran-Vila. Mol. Cell. Probes 20:105, 2006. (2) K. Z. Tang et al. Acta Hortic. Sin. 32:408, 2005.
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Eiras, Marcelo, Maria Luisa P. N. Targon, Thor V. M. Fajardo, Ricardo Flores und Elliot W. Kitajima. „Citrus exocortis viroid and Hop Stunt viroid Doubly infecting grapevines in Brazil“. Fitopatologia Brasileira 31, Nr. 5 (Oktober 2006): 440–46. http://dx.doi.org/10.1590/s0100-41582006000500002.
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Viroids, non-protein-coding small (246-401 nt) circular single-stranded RNAs with autonomous replication, are currently classified into two families. Within the family Pospiviroidae, Citrus exocortis viroid (CEVd) belongs to the genus Pospiviroid while Hop stunt viroid (HSVd) is the single member of the genus Hostuviroid. These pathogens are distributed worldwide and infect a large number of hosts. In Brazil, isolates of CEVd and HSVd have been detected in both citrus and grapevine. To characterize and study the genetic variability of these viroids, total RNA from leaves of grapevine Vitis vinifera 'Cabernet Sauvignon' and V. labrusca 'Niagara Rosada' from Bento Gonçalves, RS, was used as a template for RT-PCR amplification with specific primers for the five viroids described infecting grapevines [HSVd, CEVd, Grapevine yellow speckle viroid 1 (GYSVd-1), Grapevine yellow speckle viroid 2 (GYSVd-2) and Australian grapevine viroid (AGVd)]. Leaf samples of Citrus medica infected with CEVd from São Paulo were also analyzed. The resulting products were separated by agarose gel electrophoresis and DNA fragments of the expected size were eluted, cloned and sequenced. The grapevine samples analyzed were doubly infected by CEVd and HSVd. A phylogenetic analysis showed that the Brazilian grapevine HSVd variants clustered with other grapevine HSVd variants, forming a specific group separated from citrus variants, whereas the Brazilian CEVd variants clustered with other citrus and grapevine variants.
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Flores, Ricardo, Beatriz Navarro, Sonia Delgado, Pedro Serra und Francesco Di Serio. „Viroid pathogenesis: a critical appraisal of the role of RNA silencing in triggering the initial molecular lesion“. FEMS Microbiology Reviews 44, Nr. 3 (01.05.2020): 386–98. http://dx.doi.org/10.1093/femsre/fuaa011.
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ABSTRACT The initial molecular lesions through which viroids, satellite RNAs and viruses trigger signal cascades resulting in plant diseases are hotly debated. Since viroids are circular non-protein-coding RNAs of ∼250–430 nucleotides, they appear very convenient to address this issue. Viroids are targeted by their host RNA silencing defense, generating viroid-derived small RNAs (vd-sRNAs) that are presumed to direct Argonaute (AGO) proteins to inactivate messenger RNAs, thus initiating disease. Here, we review the existing evidence. Viroid-induced symptoms reveal a distinction. Those attributed to vd-sRNAs from potato spindle tuber viroid and members of the family Pospiviroidae (replicating in the nucleus) are late, non-specific and systemic. In contrast, those attributed to vd-sRNAs from peach latent mosaic viroid (PLMVd) and other members of the family Avsunviroidae (replicating in plastids) are early, specific and local. Remarkably, leaf sectors expressing different PLMVd-induced chloroses accumulate viroid variants with specific pathogenic determinants. Some vd-sRNAs containing such determinant guide AGO1-mediated cleavage of mRNAs that code for proteins regulating chloroplast biogenesis/development. Therefore, the initial lesions and the expected phenotypes are connected by short signal cascades, hence supporting a cause-effect relationship. Intriguingly, one virus satellite RNA initiates disease through a similar mechanism, whereas in the Pospiviroidae and in plant viruses the situation remains uncertain.
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Matoušek, J. „Viroids: sequence variability and evolution of pathogenic RNA“. Plant Protection Science 38, SI 1 - 6th Conf EFPP 2002 (01.01.2002): 173–76. http://dx.doi.org/10.17221/10348-pps.
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Viroids as the smallest pathogenic circular single-stranded pathogenic RNAs form populations of quasi-species, which<br />has been recently identified by thermodynamic methods like TGGE pre-selection and heteroduplex analysis. It was found<br />that replication under thermal stress led to enormously high level of viroid mutagenesis. Mostly multiple mutants having<br />non-random distribution of base changes were found. A specific “hot spots” were identified in the regions, where<br />a characteristic “pathogenicity domains” are localised in different viroids of the pospiviroidae family. Specific viroid<br />microevolution was observed upon artificial inoculation of non-host plant species. Our results suggest that viroid propagation<br />under physiological stress can be assumed as important factor, which is among others, responsible for an appearance<br />of viroid quasi-species in the nature. Evolution and new viroid patotypes could accumulate due to environmental stress<br />including various pollutants may be a potential danger for cultured plants.
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Di Serio, Francesco, Silvia Ambrós, Teruo Sano, Ricardo Flores und Beatriz Navarro. „Viroid Diseases in Pome and Stone Fruit Trees and Koch’s Postulates: A Critical Assessment“. Viruses 10, Nr. 11 (07.11.2018): 612. http://dx.doi.org/10.3390/v10110612.
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Composed of a naked circular non-protein-coding genomic RNA, counting only a few hundred nucleotides, viroids—the smallest infectious agents known so far—are able to replicate and move systemically in herbaceous and woody host plants, which concomitantly may develop specific diseases or remain symptomless. Several viroids have been reported to naturally infect pome and stone fruit trees, showing symptoms on leaves, fruits and/or bark. However, Koch’s postulates required for establishing on firm grounds the viroid etiology of these diseases, have not been met in all instances. Here, pome and stone fruit tree diseases, conclusively proven to be caused by viroids, are reviewed, and the need to pay closer attention to fulfilling Koch’s postulates is emphasized.
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Itaya, Asuka, Yoshie Matsuda, Robert A. Gonzales, Richard S. Nelson und Biao Ding. „Potato spindle tuber viroid Strains of Different Pathogenicity Induces and Suppresses Expression of Common and Unique Genes in Infected Tomato“. Molecular Plant-Microbe Interactions® 15, Nr. 10 (Oktober 2002): 990–99. http://dx.doi.org/10.1094/mpmi.2002.15.10.990.
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Viroids are the smallest plant pathogens. These RNAs do not encode proteins and are not encapsidated, and yet they can replicate autonomously, move systemically, and cause diseases in infected plants. Notably, strains of a viroid with subtle differences in nucleotide sequences can cause dramatically different symptoms in infected plants. These features make viroids unique probes to investigate the role of a pathogenic RNA genome in triggering host responses. We conducted a comprehensive analysis of the differential gene expression patterns of tomato plants at various stages of infection by a mild and severe strain of Potato spindle tuber viroid (PSTVd). We also compared tomato gene expression altered by the PSTVd strains with that altered by Tobacco mosaic virus (TMV). Our analyses revealed that the two PSTVd strains altered expression of both common and unique tomato genes. These genes encode products involved in defense/stress response, cell wall structure, chloroplast function, protein metabolism, and other diverse functions. Five genes have unknown functions. Four genes are novel. The expression of some but not all of these genes was also altered by TMV infection. Our results indicate that viroids, although structurally simple, can trigger complex host responses. Further characterization of viroid-altered gene expression in a host plant should help understand viroid pathogenicity and, potentially, the mechanisms of RNA-mediated regulation of plant gene expression.
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Bessolicyna, E., und A. Harina. „Development of a PCR-test system for detection of the Potato spindle tuber viroid“. Proceedings of the Komi Science Centre of the Ural Division of the Russian Academy of Sciences, Nr. 6 (28.12.2022): 84–88. http://dx.doi.org/10.19110/1994-5655-2022-6-84-88.
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The Potato spindle tuber viroid infects potato plants, causing a serious damage to agriculture by reducing yields. Therefore, there is a need to identify this pathogen. Viroids are RNA molecules and so cannot be detected with the immunological methods and the classical PCR. The aim of this work is to develop a method for detection of the Potato spindle tuber viroid. Primers have been selected for the reverse transcription reaction and the subsequent PCR, the annealing temperatures and the size of the amplified fragment have been calculated. Primers and reaction conditions have been tested on plant material. PCR products of the calculated size have been obtained. Determination of their nucleotide sequence confirmed the identification of the genetic material of the Potato spindle tuber viroid. Thus, this PCR test system can be used for detection of the Potato viroid and become the basis for detection of other
vegetable crops’ viroids.
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Matoušek, Jaroslav, und Gerhard Steger. „The Splicing Variant TFIIIA-7ZF of Viroid-Modulated Transcription Factor IIIA Causes Physiological Irregularities in Transgenic Tobacco and Transient Somatic Depression of “Degradome” Characteristic for Developing Pollen“. Cells 11, Nr. 5 (23.02.2022): 784. http://dx.doi.org/10.3390/cells11050784.
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Viroids are small, non-coding, pathogenic RNAs with a significant ability of adaptation to several basic cellular processes in plants. TFIIIA-7ZF, a splicing variant of transcription factor IIIA, is involved in replication of nuclear-replicating viroids by DNA-dependent polymerase II. We overexpressed NbTFIIIA-7ZF from Nicotiana benthamiana in tobacco (Nicotiana tabacum) where it caused morphological and physiological deviations like plant stunting, splitting of leaf petioles, pistils or apexes, irregular branching of shoots, formation of double-blade leaves, deformation of main stems, and modification of glandular trichomes. Plant aging and senescence was dramatically delayed in transgenic lines. Factors potentially involved in viroid degradation and elimination in pollen were transiently depressed in transgenic leaves. This depressed “degradome” in young plants involved NtTudor S-like nuclease, dicers, argonoute 5, and pollen extracellular nuclease I showing expression in tobacco anthers and leaves. Analysis of the “degradome” in tobacco leaves transformed with either of two hop viroids confirmed modifications of the “degradome” and TFIIIA expression. Thus, the regulatory network connected to TFIIIA-7ZF could be involved in plant pathogenesis as well as in viroid adaptation to avoid its degradation. These results support the hypothesis on a significant impact of limited TFIIIA-7ZF on viroid elimination in pollen.
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Patzak, Josef, Alena Henychová, Karel Krofta, Petr Svoboda und Ivana Malířová. „The Influence of Hop Latent Viroid (HLVd) Infection on Gene Expression and Secondary Metabolite Contents in Hop (Humulus lupulus L.) Glandular Trichomes“. Plants 10, Nr. 11 (26.10.2021): 2297. http://dx.doi.org/10.3390/plants10112297.
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Viroids are small infectious pathogens, composed of a short single-stranded circular RNA. Hop (Humulus lupulus L.) plants are hosts to four viroids from the family Pospiviroidae. Hop latent viroid (HLVd) is spread worldwide in all hop-growing regions without any visible symptoms on infected hop plants. In this study, we evaluated the influence of HLVd infection on the content and the composition of secondary metabolites in maturated hop cones, together with gene expression analyses of involved biosynthesis and regulation genes for Saaz, Sládek, Premiant and Agnus cultivars. We confirmed that the contents of alpha bitter acids were significantly reduced in the range from 8.8% to 34% by viroid infection. New, we found that viroid infection significantly reduced the contents of xanthohumol in the range from 3.9% to 23.5%. In essential oils of Saaz cultivar, the contents of monoterpenes, terpene epoxides and terpene alcohols were increased, but the contents of sesquiterpenes and terpene ketones were decreased. Secondary metabolites changes were supported by gene expression analyses, except essential oils. Last-step biosynthesis enzyme genes, namely humulone synthase 1 (HS1) and 2 (HS2) for alpha bitter acids and O-methytransferase 1 (OMT1) for xanthohumol, were down-regulated by viroid infection. We found that the expression of ribosomal protein L5 (RPL5) RPL5 and the splicing of transcription factor IIIA-7ZF were affected by viroid infection and a disbalance in proteosynthesis can influence transcriptions of biosynthesis and regulatory genes involved in of secondary metabolites biosynthesis. We suppose that RPL5/TFIIIA-7ZF regulatory cascade can be involved in HLVd replication as for other viroids of the family Pospiviroidae.
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Jiang, Dong-Mei, Meng Wang, Shi-Fang Li und Zhi-Xiang Zhang. „High-Throughput Sequencing Analysis of Small RNAs Derived from Coleus Blumei Viroids“. Viruses 11, Nr. 7 (05.07.2019): 619. http://dx.doi.org/10.3390/v11070619.
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Characterization of viroid-derived small RNAs (vd-sRNAs) is important to understand viroid–host interactions; however, vd-sRNAs belonging to the genus Coleviroid are yet to be identified and characterized. Herein, we used coleus plants singly infected with coleus blumei viroid (CbVd)-1, -5, or -6 and doubly infected with CbVd-1 and -5 to identify and analyze their vd-sRNAs. We found sense and antisense vd-sRNAs for CbVd-1, -5 and -6, and 22-nt vd-sRNAs were the most abundant; moreover, the 5′-terminal nucleotides (nts) of CbVd-1, -5, and -6 were biased toward U and C, and sRNAs derived from these three viroids were unevenly distributed along their genomes. We also noted that CbVd-5 and -6 share a fragment that forms the right half of the rod-like secondary structure of these viroids, which implied that they generated almost the same type of vd-sRNAs. This finding indicated that vd-sRNA biogenesis is mainly determined by the primary sequence of their substrates. More importantly, we found two complementary vd-sRNAs (22 nt) that were generated from the central conserved region (CCR) of these three viroids, suggesting an important role of CCR in vd-sRNA biogenesis. In conclusion, our results provide novel insight into the biogenesis of vd-sRNAs and the biological roles of CCR.
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Singh, Rudra P., Xianzhou Nie und Mathuresh Singh. „Tomato chlorotic dwarf viroid: an evolutionary link in the origin of pospiviroids“. Journal of General Virology 80, Nr. 11 (01.11.1999): 2823–28. http://dx.doi.org/10.1099/0022-1317-80-11-2823.
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Over 40 isolates of potato spindle tuber viroid (PSTVd) have been reported from potato, other Solanum species and greenhouse tomato. These isolates have sequence similarities in the range 95–99%. A viroid which caused chlorotic leaves and severe dwarfing of plants in greenhouse tomato crops was detected. The viroid was found to hybridize readily with PSTVd probes. It migrated faster than PSTVd in return-polyacrylamide gel electrophoresis and was not amplified in RT–PCR by a primer pair based on the lower strand of the central conserved region of PSTVd. Nucleotide sequencing of the viroid indicated that it is a circular RNA of 360 nt, with less than 90% sequence similarities with PSTVd isolates. The Variable domain (V) has less than 60% and the Terminal Right domain less than 90% sequence similarity, while the remainder of the molecule has greater than 97% similarity with PSTVd. Because of its less-than 90% sequence similarities, unique V domain, lack of seed-transmission and lack of cross-protection by PSTVd, the viroid from tomato is proposed to be a distinct viroid species (tomato chlorotic dwarf viroid; TCDVd) which also differs from two viroids infecting tomato in nature. TCDVd may be an evolutionary link in the development of crop viroids, with Mexican papita viroid as the ancestral viroid.
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Jevremović, Darko, und Bojana Vasilijević. „Apple viruses and viroids“. Biljni lekar 50, Nr. 6 (2022): 509–17. http://dx.doi.org/10.5937/biljlek2206509j.
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Apples are the third most produced fruit in the world, behind bananas and oranges, in terms of volume. Over the past ten years, Serbia's apple production has nearly doubled. Apples are less subjected to viral and viroid attacks than other fruit species. Most viruses cause no visible symptoms on the plant's organs, and the trees either seem to be in good condition or, in the case of sensitive cultivars, their yield is reduced. Apple stem pitting virus (ASPV), apple stem grooving virus (ASGV), apple chlorotic leaf spot virus (ACLSV), and apple mosaic virus (ApMV) are four apple economically important and widely distributed viruses. Among the viroids, the apple scar skin viroid (ASSVd) is particularly destructive for apple production.
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Broadbent, P., und CM Dephoff. „Virus indexing in the New South Wales citrus improvement scheme“. Australian Journal of Experimental Agriculture 32, Nr. 4 (1992): 493. http://dx.doi.org/10.1071/ea9920493.
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A virus indexing program was initiated in the 1950s to test local citrus clones with commercial potential, for the presence of citrus exocortis viroid and psorosis. As indexing techniques improved, clones have also been tested for the presence of citrus cachexia viroid and other viroids, citrus infectious variegation virus, citrus tatter leaf virus, and strain severity of citrus tristeza virus. Of 123 candidate clones tested, only 2 were infected with psorosis virus. Crinkly leaf (infectious variegation virus) was widespread in lemons but has been eliminated by nucellar embryony. The widespread incidence of citrus exocortis and other lower molecular weight viroids has been reduced by screening mother trees on Poncirus trifoliata stocks, biological indexing, and, more recently, by sequential polyacrylamide gel electrophoresis of RNA extracts. Cachexia is rarely found in Australian citrus clones.
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Ortolá, Beltrán, und José-Antonio Daròs. „Viroids: Non-Coding Circular RNAs Able to Autonomously Replicate and Infect Higher Plants“. Biology 12, Nr. 2 (21.01.2023): 172. http://dx.doi.org/10.3390/biology12020172.
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Viroids are a unique type of infectious agent, exclusively composed of a relatively small (246–430 nt), highly base-paired, circular, non-coding RNA. Despite the small size and non-coding nature, the more-than-thirty currently known viroid species infectious of higher plants are able to autonomously replicate and move systemically through the host, thereby inducing disease in some plants. After recalling viroid discovery back in the late 60s and early 70s of last century and discussing current hypotheses about their evolutionary origin, this article reviews our current knowledge about these peculiar infectious agents. We describe the highly base-paired viroid molecules that fold in rod-like or branched structures and viroid taxonomic classification in two families, Pospiviroidae and Avsunviroidae, likely gathering nuclear and chloroplastic viroids, respectively. We review current knowledge about viroid replication through RNA-to-RNA rolling-circle mechanisms in which host factors, notably RNA transporters, RNA polymerases, RNases, and RNA ligases, are involved. Systemic movement through the infected plant, plant-to-plant transmission and host range are also discussed. Finally, we focus on the mechanisms of viroid pathogenesis, in which RNA silencing has acquired remarkable importance, and also for the initiation of potential biotechnological applications of viroid molecules.
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Licciardello, Grazia, Rosario Ferraro, Giuseppe Scuderi, Marcella Russo und Antonino F. Catara. „A Simulation of the Use of High Throughput Sequencing as Pre-Screening Assay to Enhance the Surveillance of Citrus Viruses and Viroids in the EPPO Region“. Agriculture 11, Nr. 5 (27.04.2021): 400. http://dx.doi.org/10.3390/agriculture11050400.
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Citrus are affected by many viruses and viroids, some globally widespread and some restricted to particular countries or areas. In this study, we simulated the use of high throughput sequencing (HTS) and the bioinformatic analysis of small interfering RNAs (siRNA) as a pre-screening method to guide bioindexing and molecular detection to enhance the surveillance survey of some key or emerging citrus viruses, such as non-European citrus tristeza virus isolates (non-EU CTV), citrus tatter leaf virus, citrus leprosis virus, citrus yellow mosaic virus, and citrus bark cracking viroid, present in the EPPO lists, and the citrus yellow vein clearing virus. The HTS’s ability to detect other citrus viroids was also evaluated. The results demonstrate that HTS provides a comprehensive phytosanitary status of citrus samples either in single and multiple infections of viruses and viroids. It also provides effective information on citrus tristeza virus mixed infections despite not being able to identify the non-EU variants of the virus. Bioindexing checks each single virus infection but does not differentiate viroids on the Etrog citron indicator and is time-consuming. Molecular assays are valuable as confirmation tests of viruses and viroids but many pairs of primers are needed for a full screening and new or non-target pathogens remain undetected. In addition, the genomes of two isolates of the citrus yellow vein clearing virus and the citrus tatter leaf virus, detected in a sample from China, are described.