Investigation of symptom types and viral proteins produced by Beet necrotic yellow vein virus isolates on the model plant Nicotiana benthamiana

Öz

Beet necrotic yellow vein virus (BNYVV) causes a disease ‘rhizomania’ in sugar beet areas in the world. In this study, 15 BNYVV-infested soil samples were selected according to geographic locations. Firstly, BNYVV was propagated as bait plant technique. Then, systemic host Nicotiana benthamiana was mechanically inoculated by 15 BNYVV isolates. Some BNYVV isolates induced very strong symptom (dwarfing and mosaic) the others induced mild symptom (mosaic) on N. benthamiana. After RNA isolation, RT-PCR studies were conducted by using gene specific primers. This study showed that the P31 gene could be detected in N. benthamiana plants infected with 15 BNYVV isolates, the CP gene with only 10 isolates, the P25 gene with one isolate and the P26 gene with four isolates. This result supported that the P31 gene is involved symptom expression on N. benthamiana of BNYVV, the P25 and P26 genes were not possible to be detect with some BNYVV isolates during virus replication.

Anahtar Kelimeler

• CP
• ELISA
• P25
• P26
• P31
• Rhizomania
• RT-PCR
• Sequencing

Model bitki Nicotiana benthamiana’da Beet necrotic yellow vein virus izolatlarının simptom tipleri ve bazı viral proteinlerin araştırılması

Öz

Beet necrotic yellow vein virus (BNYVV), dünyada şeker pancarı alanlarında ‘rhizomania’ olarak adlandırılan hastalığa neden olmaktadır. Bu çalışmada, Türkiye’nin farklı coğrafik bölgelerinde şeker pancarı üretim alanlarından elde edilmiş BNYVV ile bulaşık olduğu bilinen 15 toprak örneği kullanılmıştır. Öncelikle, tuzak bitki testi yöntemine göre BNYVV izolatlarının yeniden çoğaltımı sağlanmıştır. Daha sonra, 15 BNYVV izolatı virüsün sistemik konukçusu Nicotiana benthamiana’ya mekanik olarak inokule edilmiştir. Şiddetli (cücelik ve mozayik) ya da zayıf (mozayik) simptom oluşumu gösteren bitkilerin sistemik yapraklarından RNA izolasyonunu takiben, gen spesifik primerler kullanılarak RT-PCR çalışmaları gerçekleştirilmiştir. Bu çalışmalar sonucunda; 15 BNYVV izolatı ile enfekteli N. benthamiana’da P31 ile ilgili gen bölgesi saptanmasına ragmen, kılıf protein bölgesi 10 izolatta, P25 bölgesi bir izolatta, P26 bölgesi ise dört izolatta belirlenmiştir. Bu sonuç, N. benthamina’da BNYVV’nin simptom oluşumunda P31 geninin rol oynadığını desteklemiş, P25 ve P26 gen bölgelerinin ise bazı izolatlarda virüsün replikasyonu esnasında tespit edilemediğini göstermiştir.

Anahtar Kelimeler

• CP
• ELISA
• Rhizomania
• P25
• P26
• P31
• RT-PCR
• sekans analizi

Kaynakça

1. Acosta-Leal R, Fawley MW, Rush CM, 2008. Changes in the intra isolate genetic structure of Beet necrotic yellow vein virus populations associated with plant resistance breakdown. Virology 376: 60-68.
2. Adams MJ, Antoniv JF, Mullins JGL, 2001. Plant virus transmission by plasmodiophorid fungi is associated with distinctive transmembrane regions of virus-encoded proteins. Arch. Virol. 146: 1139-1153. Andika IB, Kondo H, Tamada T, 2005. Evidence that RNA-silencing-mediated resistance to Beet necrotic yellow vein virus is less effective in roots than in leaves. Mol. Plant-Microbe Interact. 18: 194-204.
3. Bally J, Jung H, Mortimer C, Naim F, Philips JG, Hellens R, Bombarely A, Goodin MM, Waterhouse PM, 2018. The rise and rise of Nicotiana benthamiana: A plant for all reasons. Annu. Rev. Phytopathol. 56: 405-426.
4. Bouzoubaa S, Quillet L, Guilley H, Jonard GT, Richards K, 1987. Nucleotide sequence of beet necrotic yellow vein virus RNA-1. J. Gen. Virol. 68: 615-626.
5. Chiba S, Kondo H, Miyanishi M, Andika IB, Han C, Tamada T, 2011. The evolutionary history of Beet necrotic yellow vein virus deduced from genetic variation, geographic origin and spread, and the breaking of host resistance. Mol. Plant-Microbe Interact. 24 (2): 207-218.
6. Chiba S, Hleibieh K, Delbianco A, Klein E, Ratti C, Ziegler-Graff V, Bouzoubaa S, Gilmer D, 2013. The benyvirus RNA silencing suppressor is essential for long-distance movement, requires both zinc-finger and NoLS basic residues but not a nucleolar localization for its silencing-suppression activity. Mol. Plant-Microbe Interact. 26: 168-181.
7. Erkan S, Gümüş M, Paylan İC, Sipahioğlu HM, 2011. Bitki virüslerinin tanılanmasında kullanılan serolojik yöntemler. Elektronik Mikrobiyoloji Dergisi, 9 (2): 35-49.
8. Gilmer D, Bouzoubaa S, Hehn A, Guilley H, Richards K, Jonard G, 1992. Efficent cell-to-cell movement of Beet necrotic yellow vein virus requires 3' proximal genes located on RNA 2. Virology, 189: 40-47.

9. Gilmer D, 2016. Molecular biology and replication of Beet necrotic yellow vein virus. In: Rhizomania. (eds. Biancardi E, Tamada T). Springer International Publishing Switzerland, pp. 85-107.
10. Gilmer D, Ratti C, ICTV Report Consortium, 2017. ICTV Virus Taxonomy Profile: Benyviridae. J. Gen. Virol. 98 (7): 1571-1572.
11. Goodin MM, Chakrabarty R, Banerjee R, Yelton S, De Bolt S, 2007. New Gateways to discovery. Plant Physiol. 145 (4): 1100-1109.
12. Goodin MM, Zaitlin D, Naidu R A, Lommel SA, 2008. Nicotiana benthamiana: its history and future as a model for plant-pathogen interactions. Mol. Plant-Microbe Interact. 21: 1015-1026.
13. Güngör M, Erkan E, Kutluk Yılmaz ND, 2022. Türkiye’de şeker pancarı üretim alanlarında enfeksiyon oluşturan Beet necrotic yellow vein virus izolatlarının p31 geninin moleküler karakterizasyonu. Ondokuz Mayıs Üniversitesi, Ziraat Fakültesi, Anadolu Tarım Bilimleri Dergisi, 37 (3): 505-524.
14. Haeberle AM, Sutissi-Garaud C, Schmitt C, Garaud JC, Richards KE, Guilley H, Jonard G, 1994. Detection by immunogold labelling of P75 readthrough protein near an extremity of beet necrotic yellow vein virus particles. Arch. Virol. 134: 195-203.
15. Horváth J, 1994. Beet necrotic yellow vein furovirus 1. New hosts. Acta Phytopathologica Et Entomologica Hungarica, 29:109-118.
16. Hugo SA, Henry CM, Harju V, 1996. The role of alternative hosts of Polymyxa betae in transmission of Beet necrotic yellow vein virus (BNYVV) in England. Plant Pathol. 45: 662-666.
17. Kaya R, 2009. Distribution of rhizomania disease in sugar beet growing areas of Turkey. Tarım Bilimleri Dergisi, 15 (4): 332-340.
18. Keskin B, 1964. Polymyxa betae n.sp. ein parasit in den wurzein von Beta vulgaris Tournefort, besonders wahrend den jugendent wicklung den zuckerrübe. Archives of Microbiology, 49: 348-374.
19. Koch F, 1987. Bericht über eine in verschiedene zuckerrübenanbaugebiete der Turkseker in Anatolien und Thrazien zum stadium von wurzelerkrankungen. KWS Kleinwanzlebener Saatzucht, AG, Einbeck, Germany. Koenig R, Haeberle AM, Commandeur U, 1997. Detection and characterization of a distinct type of Beet necrotic yellow vein virus RNA-5 in sugar beet growing area in Europe. Arch. Virol. 142:1499-1504.
20. Kruse M, Koenig R, Hoffman A, Kaufmann A, Commandeur U, Solevyev AG, Savenkov I, Burgermeister W, 1994. Restriction fragment length polymorphism analysis of reverse transcription-PCR products reveals the existance of two major strain groups of beet necrotic yellow vein virus. J. Gen. Virol. 75: 1835-1842.
21. Kuszala M, Putz C, 1977. Rhizomania of sugar beet in Alsace. Host range and biological properties of Beet necrotic yellow vein virus. Annales de Phytopathologie, 9 (4): 435-446.
22. Kutluk Yılmaz ND, Meunier A, Schmit JF, Stas A, Bragard C, 2007. Partial nucleotide sequence analysis of Turkish isolates of Beet necrotic yellow vein virus (BNYVV) RNA-3. Plant Pathol. 56: 311-316.
23. Kutluk Yılmaz ND, Arlı Sokmen M, 2010. Occurrence of soilborne sugar beet viruses transmitted by Polymyxa betae northern and central parts of Turkey. Journal of Plant Pathology, 92 (2): 497-500.
24. Kutluk Yılmaz ND, 2016a. New natural weed host Raphanus raphanistrum L. (Brassicaceae) for Beet necrotic yellow vein virus and it’s vector Polymyxa betae Keskin. Turk J Agric For. 40: 120-126.
25. Kutluk Yilmaz ND, Arli-Sokmen M, Kaya R, Sevik MA, Tunali B, Demirtas S, 2016b. The widespread occurrences of Beet soil borne virus and RNA-5 containing Beet necrotic yellow vein virus isolates in sugar beet production areas in Turkey. Eur. J. Plant Pathol. 144 (2): 443-455.
26. Kumar S, Stecher G, Tamura K, 2016. MEGA7: Molecular Evolutionary Genetics Analysis Version 7.0 for bigger datasets. Mol. Biol. Evol. 33: 1870-1874.
27. Lauber E, Bleykasten-Grosshans C, Erhardt M, Bouzoubaa S, Jonard G, Richards KE, Guilley H, 1998a. Cell-to-cell movement of Beet necrotic yellow vein virus: I. Heterologous complementation experiments provide evidence for specific interactions among the triple gene block proteins. Mol. Plant-Microbe Interact. 11: 618-625.
28. Lauber E, Guilley H, Tamada T, Richards KE, Jonard G, 1998b. Vascular movement of Beet necrotic yellow vein virus in Beta macrocarpa is probably dependent on an RNA-3 sequence domain rather that a gene product. J. Gen. Virol. 79: 385-393.
29. Lennefors BL, Savenkoc EI, Mukasa SB, Valkonen JPT, 2005. Sequence diversity of four soilborne sugarbeet-infecting viruses. Virus Genes, 31: 57-64.
30. Link D, Schmidlin L, Schirmer A, Klein E, Erhardt M, Geldreich A, Lemaire O, Gilmer D, 2005. Functional characterization of the Beet necrotic yellow vein virus RNA-5 encoded p26 protein: evidence for structural pathogenicity determinants. J. Gen. Virol. 86: 2115-2125.
31. Liu T, Han C, Li D, Cai Z, Yu J, Liu Y, 2003. Effect of the RNA5 component on pathogenicity of beet necrotic yellow vein virus. Chinese Science Bulletin, 48: 796-799.
32. Liu HY, Sears JL, Lewellen RT, 2005. Occurrence of resistance-breaking beet necrotic yellow vein virus of sugar beet. Plant Dis. 89: 464-468.
33. Liebe S, Varrelmann M, 2022. Ongoing evolution of Beet necrotic yellow vein virus towards Rz1-resistance breaking in Europe. Plant Pathol. 71:1647–1659.
34. Link D, Schmidlin L, Schirmer A, Klein E, Erhardt M, Geldreich A, Lemaire O, Gilmer D, 2005. Functional characterization of the Beet necrotic yellow vein virus RNA-5 encoded p26 protein: evidencefor structural pathogenicity determinants. J. Gen. Virol. 86: 2115-2125.
35. Meunier A, Schmit JF, Stas A, Kutluk N, Bragard C, 2003. Multiplex reverse transcription for simultaneous detection of beet necrotic yellow vein virus, beet soilborne virus, and beet virus Q and their vector Polymyxa betae KESKIN on sugar beet. Appl. Environ. Microbiol. 2356-2360.
36. Miyanishi M, Kusume T, Saito M, Tamada T, 1999. Evidence for three groups of sequence variants of beet necrotic yellow vein virus RNA 5. Arch. Virol. 144: 879-892.
37. Özer G, Ertunç F, 2005. Detection of rhizomania disease in sugar beet plantations of Amasya sugar refinery. Journal of Agricultural Sciences, 11 (3): 339-343.
38. Özmen CY, Khabbazi SD, Khabbazi AD, Gürel S, Kaya R, Oğuz MÇ, Turan F, Rezaei F, Kibar U, Ergül A, 2020. Genome composition analysis of multipartite BNYVV reveals the occurrence of genetic re-assortment in the isolates of Asia Minor and Thrace. Nature Scientific Reports, 10, 4129.
39. Rahim MD, Andika IB, Han C, Kondo H, Tamada T, 2007. RNA4-encoded P31 of Beet necrotic yellow vein virus is involved in efficient vector transmission, symptom severity and silencing suppression in roots. J. Gen. Virol. 88: 1611-1619.
40. Satoh K, Kondoh H, Sasaya T, Shimizu T, Choi IR, 2010. Selective modification of rice (Oryza sativa) gene expression by Rice stripe virus infection. J. Gen. Virol. 91: 294-305.
41. Schirmer A, Link D, Cognat V, Moury B, Beuve M, Meunier A, Bragard C, Gilmer D, Lemaire O, 2005. Phylogenetic analysis of isolates of Beet necrotic yellow vein virus collected wordwide. J. Gen. Virol. 86: 2897-2911.
42. Scholten OE, Lange W, 2000. Breeding for resistance to rhizomania in sugar beet: a review. Euphytica, 112: 219-231.
43. Tamada T, Baba T, 1973. Beet necrotic yellow vein virus from rhizomania affected sugar beet in Japan. Ann. Phytopathol. Soc. Jpn., 39: 325-332.
44. Tamada T, 1975. Beet necrotic yellow vein virus. CMI/ABB. Description of Plant Viruses, Association of Applied Biologists, Wellesbourne, UK, 144: 4.
45. Tamada T, Abe H, 1989. Evidence that beet necrotic yellow vein virus RNA-4 is essential for efficient transmission by the fungus Polymyxa betae. J. Gen. Virol. 70: 3391-3398.
46. Tamada T, Abe H, Saito M, Kiguchi T, Harada T, 1989. Production and pathogenicity of isolates of Beet necrotic yellow vein virus with different numbers of RNA components. J. Gen. Virol. 70: 3399-3409.
47. Tamada T, Saito M, Kiguchi T, Kusume T, 1990. Effect of isolates of Beet necrotic yellow vein virus with different of RNA components on the development of rhizomania symtoms. Proc. Ist Symp. Int. Work. Group. Plant Viruses Fungal Vectors, 41-44, 21-24 Ağustos, Braunscweig, Germany.
48. Tamada T, Kusume T, Uchino H, Kiguchi T, Saito M, 1996. Evidence that Beet necrotic yellow vein virus RNA 5 is involved in symtom development of sugar beet roots. Proc. 3rd Symp. Int. Work. Group Plant Viruses Fungal Vectors, 49-52, 6-7 Ağustos, Dundee, Scotland.
49. Tamada T, 2002. Beet necrotic yellow vein virus. CMA/ABB descriptions of plant viruses. Association of Applied Biologists. Wellesbourne UK.
50. Tamada T, 2007. Suscepbility and resistance of Beta vulgaris subsp. maritima to foliar rub-inoculation with beet necrotic yellow vein virus. J. Gen. Plant Pathol. 73 (1): 76-80.
51. Tamada T, Kondo H, 2013. Biological and genetic diversity of plasmodiophorid-transmitted viruses and their vectors. J. Gen. Plant Pathol. 79: 307-320.
52. Tamada T, 2016. General features of Beet necrotic yellow vein virus. In: Rhizomania. (eds. Biancardi E, Tamada T), Springer International Publishing, Switzerland, pp. 55-83.
53. Tamada T, Uchino H, Kusume T, Iketani-Saito M, Chiba S, Andika IB, Kondo H, 2020. Pathogenic roles of beet necrotic yellow vein virus RNA5 in the exacerbation of symptoms and yield reduction, development of scab-like symptoms, and Rz1-resistance breaking in sugar beet. Plant Pathol. 70: 219-232.
54. Thompson JD, Higgins DG, Gibson TJ, 1994. Clustal W: Improving the sensitivity of proggosive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Research, 22: 4673-4680.
55. Yardımcı N, Çulal Kılıç H, 2011. Identification of Beet necrotic yellow vein virus in lakes district: A major beet growing area in Turkey. Indian Journal of Virology, 22 (2): 127-130.
56. Wang Y, Fan H, Wang X-B, Li M, Li D, Yu J, 2011. Detection and characterization of spontaneous internal deletion mutants of Beet necrotic yellow vein virus RNA3 from systemic host Nicotiana benthamiana. Virology Journal, 8: 335-344.