Tokat ili Bağlarda Grapevine leafroll-associated virus -1,2,3,4'ün yaygınlığı ve Genetik Değişkenliği

Öz

Asma yaprak kıvırcıklığı hastalığı (GLRD), dünya çapında asmanın en önemli viral hastalıklarından biridir. Bu çalışma, Grapevine leafroll associated virus -1 (GLRaV-1), Grapevine leafroll associated virus -2 (GLRaV-2), Grapevine leafroll associated virus -3 (GLRaV-3), Grapevine leafroll-associated virus 4 (GLRaV-4), ve Grapevine leafroll-associated virus-9 (GLRaV-9) etmenlerinin yaygınlığının araştırılması amacıyla yapılmıştır. Bunun için Tokat ili Merkez, Erbaa, Niksar ve Pazar ilçelerindeki farklı bağ alanlarından virüs benzeri belirti gösteren asmalardan yaprak örnekleri ve bir yıllık bitki sürgünleri toplanmıştır. Surveyde elde edilen toplam 197 örnek GLRaV-1, GLRaV-2, GLRaV-3, GLRaV-4) ve GLRaV-9 etmenlerine özgü primerler ile iki aşamalı polimeraz zincir reaksiyonu (PCR) kullanılarak moleküler olarak test edilmiştir. 197 örnekten 106'sı (%53.80) bu virüslerle tekli veya çoklu enfeksiyon tespit edilmiştir. 103 (%52,28) örnek GLRaV-1 ile pozitif sonuçlanırken, enfeksiyon oranları 10 örnekle GLRaV-3 için %5,18, GLRaV-4 için 5 örnekle %2,53, GLRaV-2 ve GLRaV-9 için %1,03 olarak belirlenmiştir. GLRaV-1, GLRaV-2 ve GLRaV-3'ün pozitif PCR ürünleri çift yönlü sekans analizinden sonra MEGAX ile GenBank'taki referans diğer izolatlarla karşılaştırılması yapılmıştır. GLRaV-1 izolatlarının sekans analizine göre GenBank'taki referans izolatlarla %94-96, GLRaV-2 izolatları %96-99 ve GLRaV-3'ün UO3-3 izolatı %99-100 nükleotid benzerliği göstermiştir.

Anahtar Kelimeler

• Vitis vinifera
• virus
• RT-PCR
• Tokat
• Asma yaprak kıvırcıklığı hastalığı

Genetic variability of Grapevine Leafroll-Associated Virus-1,2,3,4 Infecting Vineyards in Tokat Province of Turkey

Abstract

Grapevine leafroll disease (GLRD) is one of the most important viral diseases of grapevine worldwide. The aim of this study was to investigate the prevelance of Grapevine leafroll associated virus-1 (GLRaV-1), Grapevine leafroll-associated virus-2 (GLRaV-2), Grapevine leafroll-associated virus-3 (GLRaV-3), Grapevine leafroll-associated virus 4 (GLRaV-4), and Grapevine leafroll-associated virus-9 (GLRaV-9) in Tokat province of Turkey. In 2018, leaf samples and one-year-old shoots of plants, showing virus-like symptoms, were collected from different vineyards in Central, Erbaa, Niksar, and Pazar districts of Tokat province. Totally 197 samples were obtained and tested molecularly using two-step polymerase chain reaction (PCR) by using virus-specific primers for GLRaV-1, GLRaV-2, GLRaV-3, GLRaV-4) and GLRaV-9. At least 106 (53.80%) samples were found to be infected showed single or multiple infections. The 103 (52.28%) samples resulted in positive with GLRaV-1 while the infection rates were %5.18 for GLRaV-3 with 10 samples, %2.53 for GLRaV-4 with 5 samples, %1.03 for GLRaV-2 and GLRaV-9 with 2 samples, respectively. The amplified PCR products of GLRaV-1, GLRaV-2 and GLRaV-3 were sequenced, aligned with MEGAX, and analyzed in BLAST for multiple comparisons of other isolates in the GenBank. Sequence analysis of viruses showed 94-96 % , 96-99 % and, 99-100 % nucleotide identity with references isolates in the GenBank for GLRaV-1, GLRaV-2, and UO3-3 isolate of GLRaV-3 respectively.

Keywords

• Vitis vinifera
• virus
• RT-PCR
• Tokat
• Grapevine leafroll disease

References

1. Al Rwahnih, M., Dolja, V. V., Daubert, S., Koonin, E., & Rowhani, A. (2011). Genomic and biological analysis of Grapevine leafroll-associated virus 7 reveals a possible new genus within the family Closteroviridae. Virus Research, 163(1), 302–309.
2. Alabi, O., Rwahnih, J., Karthikeyan, M. A., Poojari, G., Fuchs, S., Rowhani, M., & Naidu, A. (2011). Grapevine leafroll associated virus 1 occurs as genetically diverse populations. Phytopathology, 101(12), 1446-1456.
3. Alknowni, R., Rowhani, A., Daubert, S., & Golino, D. (2004). Partial characterization of a new ampelovirus associated with grapevine leafroll disease. Journal Plant Pathology, 86(2), 123–133.
4. Bertazzon, N., Borgo, M., Vanin, S., & Angelini, E. (2010). Genetic variability and pathological properties of Grapevine leafroll-associated virus 2 isolates. European Journal of Plant Pathology, 127(2), 185-197.
5. Beuve, M., Sempé, L., & Lemaire, O. (2007). A sensitive one-step realtime RT-PCR method for detecting Grapevine leafroll-associated virus 2 variants in grapevine. Journal of Virological Methods, 141(2), 117-124.
6. Buzkan, N., Karadağ, S., Kaya, A., Baloğlu, S., Minafra, A., & Ben-Dov, Y., (2012). Investigating the presence of mealybug species as vectors for viruses in grape growing areas in Turkey. Canadian Journal of Plant Pathology-Revue Canadienne de Phytopathologie, 34(2), 298-305.
7. Çelik, H. (2013). Türkiye bağciliğinda üretim hedefleri. Vizyon 2023 Bağcılık Çalıştayı, Tekirdağ Bağcılık Araştırma İstasyonu, Tekirdağ.
8. Çelik, H., Ağaoğlu, Y. S., Fidan, Y., Marasalı, B. & Söylemezoğlu, G. (1998). Genel Bağcılık. Sunfidan A.Ş. Mesleki Kitaplar Serisi, Ankara.

9. Elçi, E. (2019). Genomic Variability and Recombination Analysis of Grapevine leafroll-associated virus-1 Isolates from Turkey. Journal of Agricultural Sciences, 25(3), 319-327
10. Erkan, S., Gümüş, M., Paylan, I. C. & Sipahioğlu, H. M. (2011). Bitki Virüslerinin Tanılanmasında Kullanılan Serolojik Yöntemler. Elektronik Mikrobiyoloji Dergisi, 9(2), 35-49.
11. FAO. (2016). FAO-OIV FOCUS 2016. http://www.fao.org/3/a-i7042e.pdf. Access date: 18 Mayıs 2019.
12. FAO. (2017). Crops datas. https://www.fao.org/faostat/en/#data/QCL. Access date: 18 Mayıs 2019.
13. Ince, E., & Yavuz, S. (2015). Current status of Grapevine leafroll associated viruses in East Mediterranean Region of Turkey. Proceedings of the 18th Congress of the International Council for the Study of Virus and Virus-like Diseases of the Grapevine, Ankara, Turkey.
14. Karadeniz, A., Yagcı, A., Topkaya, S., & Yanar, Y. (2018). Serological Diagnosis of Several Grapevine Virus Agents in Tokat Province and District. Plant Protection Bulletin, 58(2), 103-110.
15. Kaya, A., & Erilmez, S. (2014). Detection of Viruses in Aegean Region Grapevines. The Journal of Turkish Phytopathology, 43(1-3), 45-57.
16. Kumar, S., Stecher, G., Li, M., Knyaz, C., & Tamura K. (2018). MEGA X: Molecular Evolutionary Genetics Analysis across computing platforms. Molecular Biology and Evolution, 35, 1547-1549.
17. Li, R., Mock, R., Huang, Q., Abad, J., Hartung, J., & Kinard, G. (2008). A reliable and inexpensive method of nucleic acid extraction for the PCR-based detection of diverse plant pathogens. Journal of Virological Methods, 154(1-2), 48–55.
18. Ling, K. S., Zhu, H. Y. & Gonsalves, D. (2004). Complete nucleotide sequence and genome organization of Grapevine leafroll-associated virus 3, type member of the genus Ampelovirus. Journal of General Virology, 85(7), 2099-2102.
19. Maliogka, V. I., Dovas, C. I. & Katis, N. I., (2008). Generic and Species-Specific Detection of Viruses Belonging to an Evolutionary Distinct Lineage within the Ampelovirus Genus. Journal of Virological Methods, 154(1-2), 41-47.
20. Maree, H. J., Pirie, M. D., Bester, R., Oosthuizen, K., & Burger, J. T. (2015). Phylogenomic analysis reveals deep divergence and recombination in an economically important grapevine virus. Public Library of Science 10(5), e0126819.
21. Martelli, G. P. (2014). Directory of Virus and Virus-Like Diseases of the Grapevine and Their Agents. Journal of Plant Pathology, 96(1), 1-136.
22. Martelli, G. P., Abou Ghanem-Sabanadzovic, N., Agranovsky, A. A., Al Rwahnih, M., Dolja, V. V., Dovas, C. I., Fuchs, M., Gugerli, P., Hu, J. S., Jelkmann, W., Katis, N. I., Maliogka, V. I., Melzer, M. J., Menzel, W., Minafra, A., Rott, M. E., Rowhani, A., Sabanadzovic, S., & Saldarelli, P. (2012). Taxonomic revision of the family Closteroviridae with special reference to the Grapevine Leafroll Associated members of the genus Ampelovirus and the putative species unassigned to the family. Journal of Plant Pathology, 94, 7-19.
23. Martelli, G. P. (2017). An overview on grapevine viruses, viroids, and the diseases they cause. In B. Meng, G. P. Martelli, D. A. Golino, & M. Fuchs (Eds.), Grapevine Viruses: Molecular Biology, Diagnostics and Management, Cham: Springer International Publishing, 31–46.
24. Martin, R. R., Eastwell, K. C., Wagner, A., Lamprecht, S., & Tzanetakis, I. E. (2005). Survey for Viruses of Grapevine in Oregon and Washington. Plant Disease, 89(7), 763-766.
25. Moura, C. J. M., Fajardo T. V. M., Eiras, M., Silva, F. N., & Nickel, O. (2018). Molecular characterization of GSyV-1 and GLRaV-3 and prevalence of grapevine viruses in a grape-growing area. Scientia Agricola, 75(1), 43-51.
26. Omeroglu, E. (2016). Determination of grapevine leafroll virus (GLRAVS) in some grape varieties in grapeyards in Tekirdağ province. MSc Thesis, Namık Kemal University Graduate School of Natural and Applied Sciences. Department of Plant Protection, Tekirdağ.
27. Onder, S. (2016). Detection and characterization of grapevine leafroll-associated viruses (GLRaVs) in Aegean Vineyards. PhD Thesis, Ege University. Graduate School of Natural and Applied Science, İzmir.
28. Onder, S., Paylan, I. C., & Gümüş, M. (2016). Detection of Several Important Viral Agents by Real-time PCR in Aegean Vineyards. The Journal of Turkish Phytopathology, 45, 21-29.
29. Türkmen, Y. (2014). The detection and prevalence of leafroll viruses in Turkish vineyard. MSc Thesis, Ankara University, Graduate School of Natural and Applied Science, Ankara.
30. Yakar, M. (2017). Molecular diversity of grapevine leafroll-associated virus-2 (glrav-2) ın autocthonous Grapevine varities in eastern mediterranean and southeast anatolia regions. MSc Thesis, Kahramanmaraş Sütçü İmam Universty Graduate School of Natural and Applied Sciences, Kahramanmaraş.