ردیابی و تعیین خصوصیات ملکولی ویروئید فرورفتگی میوه سیب(Apple Dimple Viroid) در منطقه مراغه

نوع مقاله : مقاله پژوهشی

نویسنده

استادیار گروه گیاهپزشکی، دانشکده کشاورزی، دانشگاه مراغه.

چکیده

چکیده
در تابستان سال­های 1395 و 1396 تعداد 73 نمونه با علائم مشابه با ویروئید فرورفتگی میوه سیب از ارقام گالا و رد دلیشیز منطقه­ی مراغه جمع­آوری شد. RNA کل از پوست ساقه و برگ نمونه­های جمع­آوری شده حاوی علایم لکه­های فرورفته و خال­های زرد با حاشیه نامنظم روی پوست میوه، استخراج گردید و با استفاده از آغازگرهای اختصاصی ژنوم کامل ویروئید به طول 307-306 جفت باز با آزمون RT-PCR تکثیر و ترادف 10 نمونه تعیین گردید. جستجوی ترادف ها با BLASTn و هم­ردیف­سازی آنها با ترادف­های مربوطه در بانک ژن نشان داد که جدایه­های مورد مطالعه دارای بیشترین و کمترین یکسانی ترادف به میزان 100 و 4/74 درصد بترتیب با رس شمارهای EF088662 (ایتالیا) و KF788291 (ایتالیا) می­باشند. بیشترین تفاوت نوکلئوتیدی در بین نواحی  CCRو TR قرار داشتند. آنالیز فیلوژنتیک با روش حداکثر احتمال نشان داد که جدایه های ایرانی به همراه جدایه­ها و واریانت­های گزارش شده از ایتالیا در یک گروه قرار می گیرند.
 
 

کلیدواژه‌ها


عنوان مقاله [English]

Detection and Molecular Characterization of Apple Dimple Fruit Viroid in Apple Trees in Maragheh Area

نویسنده [English]

  • Vahid Roumi
Assistant Professor, Department of Plant Protection, Faculty of Agriculture, University of Maragheh.
چکیده [English]

Abstract
During years 2016-2017 surveys in apple orchards of Maragheh region, 73 samples of Gala and Red delicious cultivars with symptoms resembling to those caused by Apple dimple fruit viroid (ADFVd) were collected and subjected to two-step RT-PCR. Total RNA from stem bark and leaves of symptomatic plants showing dimple fruit and irregular yellow spots on fruit was extracted and full-length 306-307 base pairs genome of the isolates were amplified by using specific primer pairs. Ten amplicons with expected size were directly sequenced on both strands. Blast analysis and multiple sequence alignment of the sequences with those full-length ADFVd sequences deposited in GenBank, showed identities ranging from 74.4-100%. Iranian ADFVd isolates had the highest and lowest identity with two Italian isolates, accession Nos. EF088662 and KF788291, respectively. The region between CCR and TR domains was the most divergent. In phylogenetic analysis by maximum likelihood method, all Iranian isolates were clustered together with the Italian isolates and variants.
 

کلیدواژه‌ها [English]

  • Keywords: Detection
  • Viroid
  • Dimple fruit
  • Apple
  • Maragheh
احمدی ک، قلیزاده ح، عبادزاده ح­ر، حاتمی ف، حسینپور ر، کاظمی فرد ر و عبدشاه ه، 1395. آمارنامه کشاورزی سال زراعی 94-93، جلد سوم :محصولات باغبانی. وزارت جهادکشاورزی، معاونت برنامه­ریزی و اقتصادی، مرکز فناوری اطلاعات و ارتباطات. 240 صفحه.
Barba M, Hosakawa M, Wang QC, Taglienti A and Zhang Z, 2017. Viroid Elimination by Thermotherapy, Cold Therapy, Tissue Culture, In Vitro Micrografting, or Cryotherapy. Chapter 40. In: Viroids and Satellites. Elsevier Science. Pages: 425-435.
Chiumenti M, Torchetti EM, Di Serio F and Minafra A, 2014. Identification and characterization of a viroid resembling Apple dimple fruit viroid in fig (Ficus carica L.) by next generation sequencing of small RNAs. Virus Research 188: 5-59.
Choueiri E, El Zammar S, Jreijiri F, Hobeika C, Myrta A and Di Serio F, 2007. First report of Apple dimple fruit viroid in Lebanon. Journal of Plant Pathology 89 (2): 304.
Dalakouras A, Dadami E, Wassenegger M, Krczal G and Wassenegger M. 2016. RNA-directed DNA methylation efficiency depends on trigger and target sequence identity. Plant Journal 87: 202-214.
Di Serio F, Aparicio F, Alioto D, Ragozzino A, and Flores R 1996. Identification and molecular properties of a 306 nucleotide viroid associated with apple dimple fruit disease. Journal of General Virology 77: 2833–2837.
Di Serio F, Flores R, Verhoeven J Th J, Li SF, Pallás V, Randles JW, Sano T, Vidalakis G and Owens RA, 2014. Current status of viroid taxonomy. Archives of Virology, 159 (12): 3467-3478.
Di Serio F, Malfitano M, Alioto D, Ragozzino A and Flores R, 2002. Apple dimple fruit viroid: sequence variability and its specific detection by multiplex fluorescent RT-PCR in the presence of Apple scar skin viroid. Journal of PlantPathology84(1): 27-34.
Diener TO 1971. Potato spindle tuber ‘‘virus’’ IV. A replicating low molecular weight RNA. Virology 45: 411–428.
Ding B, 2009. The biology of viroid-host interactions. Annual Review of Phytopathology 47: 105–131
EPPO Standards, 1999. Pathogen-tested material of Malus, Pyrus and Cydonia. Certification schemes PM, 4/27(1): 16.
Felsenstein J, 1985. Confidence limits on phylogenies: An approach using the bootstrap. Evolution 39: 783-791.
Flores R and Owens RA, 2008. Viroids. In: Mahy BWJ, Van Regenmortel MHV (eds) Encyclopedia of Virology. Elsevier, Oxford, Pages: 332–342.
Flores R, Hernandez C, Martınez de Alba AE, Daro`s JA and Di Serio F, 2005. Viroids and viroid-host interactions. Annual Review of Phytopathology 43: 117–139.
Flores R, Randles JW, Bar-Joseph M and Diener TO, 1998. A proposed scheme for viroid classification and nomenclature. Archives of Virology 143: 623–629.
Giguere T, Adkar-Purushothama CR and Perreault JP, 2014. Comprehensive Secondary Structure Elucidation of Four Genera of the Family Pospiviroidae. PLOS ONE, 9 (6): e98655.
Giguere T and Perreault JP, 2017. Classification of the Pospiviroidae based on their structural hallmarks. PLoS ONE, 12(8): e0182536.
Hadidi A, Flores R, Randles JW and Palukaitis P, 2017. Viroids and Satellites. Elsevier Science, 716 Pages.
Hall TA, 1999. BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symposium Series 41: 95-98.
Hao L, Xie J, Chen S, Wang S, Gong Z, Ling KS, Guoa L, Fana Z and Zhou. T, 2016. A multiple RT-PCR assay for simultaneous detection and differentiation of latent viruses and apscarviroids in apple trees. Journal of Virological Methods 234: 16–21.
He YH, Isono S, Kawaguchi-Ito Y, Taneda A, Kondo K, Iijima A, Tanaka K and Sano T, 2010. Characterization of a new Apple dimple fruit viroid variant that causes yellow dimple fruit formation in ‘Fuji’ apple trees. Journal of General Plant Pathology 76(5): 324-330.
Ito T, Ieki H and Ozaki K. 2000. A population of variants of a viroid closely related
to citrus viroid-I in citrus plants. Archives of Virology 145: 2105–2114.
Keese P and Symons RH, 1985. Domains in viroids: evidence of intermolecular RNA rearrangements and their contribution to viroid evolution. Proceedings of the National Academy of Sciences 82: 4582–4586.
Kimura M, 1980. A simple method for estimating evolutionary rate of base substitutions through comparative studies of nucleotide sequences. Journal of Molecular Evolution 16: 111-120.
Kumar S, Stecher G, and Tamura K, 2016. MEGA7: Molecular Evolutionary Genetics Analysis version 7.0 for bigger datasets. Molecular Biology and Evolution 33(7): 1870-4.
Ling KS, 2017. Decontamination Measures to Prevent Mechanical Transmission of Viroids. Chapter 41 in: Viroids and Satellites. Elsevier Science. Pages: 437-445.
Malfitano M, Alioto D, Ragozzino A, Di Serio F, and Flores R, 2004. Experimental evidence that Apple dimple fruit viroid does not spread naturally. Acta Horticulturae 657: 357-360.
Owens RA, Flores R, Di Serio F, Li S-F, Palla´s V, Randles JW, Sano T and Vidalakis G, 2012. Viroids. In King AMQ, Adams MJ, Carstens EB, Lefkowitz EJ (eds), Virus taxonomy: ninth report of the International Committee on Taxonomy of Viruses, Elsevier Academic Press, London UK, Pages: 1221–1234.
Roumi V, Gazel M, and Caglayan K, 2017.First report of Apple dimple fruit viroid in apple trees in Iran. New Disease Reports 35: 3.
Takeda R and Ding B. 2009. Viroid intercellular trafficking: RNA motifs, cellular factors and broad impacts. Viruses 1: 210-21.
Thompson JD, Higgins DG, and Gibson TJ, 1994. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Research 22(22): 4673–4680.
Vamenani R, Rahimian H, Alavi SM and Borhani B, 2015. A Viroid Resembling Hop stunt viroid in infected apple trees with apple scar skin disease in the Northeast of Iran. Iranian Journal of Virology 9(1): 32-35.
Ye T, Chen SY, Wang R, Hao L, Chen H, Wang N, Guo LY, Fan ZF, Li SF and Zhou T, 2013. Identification and molecular characterization of Apple dimple fruit viroid in China. Journal of Plant Pathology 95: 637–641.