Molecular diagnosis of Globodera rostochiensis Wollenweber (Tylenchida: Heteroderidae) in the potato growing areas of Aegean Region, Turkey

Abstract

DNA isolation was done by the 8 different cyst populations collected from the potato growing areas of Aegean Region (Turkey) between 2006-2008. As a result of Multiplex PCR conducted with species-specific primer, composing of band is observed in 435 bp as specific to Globodera rostochiensis Wollenweber (Tylenchida: Heteroderidae). By doing PCR with rDNA2 and rDNA1.58S primers of the same sample, as a consequence of cropping with Hinf l enzyme, in 233bp and 522bp band were confirmed as specific to G. rostochiensis. In this study, other potato cyst nematodes, Globodera pallida Stone, 1973 was not determined. Globodera rostochiensis was determined previously by using morphometric methods in Turkey. The identification of G. rostochiensis was confirmed by molecular methods with this study. 

Keywords

• Potato cyst nematode
• Globodera rostochiensis
• potato
• molecular diagnosis

Ege Bölgesi patates alanlarında Globodera rostochiensis Wollenweber, (Tylenchida: Heteroderidae)’in moleküler yöntemlerle saptanması

Öz

Ege Bölgesi’nde patates alanlarından 2006-2008 yıllarında 8 adet kist nematodu populasyonu toplanarak, elde edilen kistlerden DNA izolasyonu yapılmıştır. Türe özgü spesifik primerler ile gerçekleştirilen Multipleks PCR sonucunda Globodera rostochiensis Wollenweber, (Tylenchida: Heteroderidae)’e özgü 435 bp de bant oluşumu gözlenmiştir. Aynı örneklerin rDNA2 ve rDNA1.58S primerleri ile PCR yapılarak Hinf l enzimi kesimi sonucunda G. rostochiensis’e özgü 233bp ve 522bp.de bant verdiği tespit edilmiştir. Bu çalışmada patates kist nematodlarından Globodera pallida Stone, 1973 tespit edilmemiştir. Türkiye’de daha önce morfometrik yöntemlerle saptanan, G. rostochiensis’ in varlığı moleküler yöntemlerle de ilk defa belirlenmiştir

Anahtar Kelimeler

• Patates kist nematodu
• Globodera rostochiensis
• patates
• moleküler teşhis

Kaynakça

1. Anonymous, 2009. Türkiye İstatistik Kurumu, Ankara (web sayfası: http://tuikrapor.tuik.gov.tr), (erişim Tarihi: Mart 2011).
2. Bates, J. A., E. J. Taylor, P. T. Gans, & J. E. Thomas, 2002. Determination of relative proportions of Globodera species in mixed populations of potato cyst nematodes using PCR product melting peak analysis. Molecular Plant Pathology, 3: 153–161.
3. Brown, E. B., 1969. Assessment of the damage caused to potatoes by potato cyst eelworm, Heterodera rostochiensis Woll. Annals of Applied Biology, 63 (3): 493-502.
4. Bulman, S. R., & J. W. Marshall, 1997. Differentiation of Australian potato cyst nematode (PCN) populations using the polymerase chain reaction (PCR). New Zealand Journal of Crop and Horticultural Science, 25:123–129.
5. Enneli, S. & G. Öztürk, 1996. “Orta Anadolu Bölgesinde patateslerde zarar yapan, önemli bitki paraziti nematodlar, 396-403”. Türkiye 3. Entomoloji Kongresi (24-28 Eylül, Ankara) Bildirileri, Ankara Üniversitesi Basımevi, 716s.
6. EPPO, 2004. Diagnostic protocols for regulated pests, Globodera rostochiensis and Globodera pallida. EPPO Bulletin 34:309–314.
7. EPPO, 2011. Data Sheets on Quarantine Pests Globodera rostochiensis and Globodera pallida. (web sayfası: http://www.eppo.org/QUARANTINE/nematodes/Globodera_pallida/ HETDSP_ds.pdf), (erişim tarihi: Mart, 2011).
8. Evans, D. & A. R. Stone, 1977. A review of the distribution and biology of the potato cyst nematodes Globodera rostochiensis and G. pallida. PANS 23(2), 178-189.

9. Evans, K. & J. A. Rowe, 1998. “Distribution and Economic Importance, 1–30”. In: The Cyst Nematodes (Ed: S.B. Sharma), Kluwer Academic Editors, Dordrecht, The Netherlands.
10. Fenwick, D. W., 1940. Methods for the recovery and counting of cysts of Heterodera schachtii from soil. Journal of Helminthology, 18: 155-172.
11. Fleming, C. C., S. J. Turner, T. O. Powers & A. L. Szalansky, 1998. Diagnostics of cyst nematodes: Use of the polymerase chain reaction to determine species and estimate population levels. Aspects of Applied Biology, 52: 375.382.
12. Franco, J., R. Oros, G. Main & N. Ortuno, 1998. “Potato Cyst Nematodes (Globodera species) in South America, 239-269”. In: Potato Cyst Nematodes: Biology, Distribution and Control (Eds: R.J. Marks & B.B. Brodie), CAB International, Wallingford, UK.
13. Franklin, M. T., 1951. The Cyst Forming Species of Heterodera. Commonwealth Agricultural Bureaux, Farnham Royal, 147 pp.
14. Fullaondo, A., E. Barrena, M. Viribay, I. Barrena, A. Salazar & E. Ritter, 1999. Identification of potato cyst nematode species Globodera rostochiensis and G. pallida by PCR using specific primer combinations. Nematology 1 (2): 157-163.
15. Jones, F. G. W., 1970. The control of the potato cyst nematode. Journal of the Royal Society of Arts, 118: 179-199.
16. Phillis, I., 1992. Assessment of potato yield loss caused by the potato cyst nematode, Globodera rostochiensis, (web page: http://www.cababstractsplus.org/ abstracts/ Abstract. aspx?AcNo=19922319811), (Erişim tarihi: Mart 2010).
17. Phillips, M. S., C. A. Hackett & D. L. Trudgill, 1991. The relationship between the initial and final population densities of the potato cyst nematode Globodera pallida for partially resistant potatoes. Journal of Applied Ecology, 28: 109- 119.
18. Powers, T. 2004. Nematode molecular diagnostics: from bands to barcodes. Annual Review of Phytopathology, 42:367-383.
19. Pylypenko, L. A. 1998. Distribution of Globodera rostochiensis Wollenweber, 1923 (Tylenchida, Heteroderidae) in Ukraine. Vesnik Zoologii, 32: 139-142.
20. Pylypenko, L. A. 1999. The Interrelations in A System Parasite - Host Plant at the Potato Globoderosis. Ph.D. Thesis, National Agrarian University, Kyiv, Ukraine, 136 pp.
21. Pylypenko, L. A., T. Uehara, M. S. Phillips, D. D. Sigareva & V. C. Blok, 2005. Identification of Globodera rostochiensis and G. pallida in the Ukraine by PCR. European Journal of Plant Pathology, 111: 39–46.
22. Skantar, A. M., Z. A. Handoo, L. K. Carta & D. J. Chitwood, 2007. Morphological and molecular identification of Globodera pallida associated with potato in Idaho. Journal of Nematology, 39 (2): 133-144.
23. Syracuse, A. J., C. S. Johnson, J. D. Eisenback, C. L. Nessler & E. P. Smith, 2004. Intraspecific variability within Globodera tabacum solanacearum using random amplified polymorphic DNA. Journal of Nematology, 36: 433- 439.
24. Szalanskj, A. L., D. D. Sui, T. S. Harris & T. O. Powers, 1997. Identification of cyst nematodes of agronomic and regulatory concern with PCR-RFLP of ITS I. Journal of Nematology, 29:255-267.
25. Ulutaş, E, 2010. Ege Bölgesi Patates Üretim Alanlarında Bulunan Önemli Bitki Paraziti Nematodların Belirlenmesi ve Bitki Gelişimine Etkileri. Ege Üniversitesi Fen Bilimleri Enstitüsü, (Basılmamış) Doktora Tezi, Bornova, İzmir, 92+XVIII s.
26. Vejl, P., S. Skupinová, P. Sedlak & J. Domkarova, 2002. Identification of PCN species (Globodera rostochiensis, G. pallida) by using of ITS-1 region´s polymorphism. Rostlinna Vyroba, 48: 486-489.
27. Vrain, T. C., D. A. Wakarchuk, A. C. Levesque & R. I. Hamilton, 1992. Intraspecific rDNA restriction fragment length polymorphisms in the Xiphinema americanum group. Fundamental and Applied Nematology, 15: 563–573.
28. Winslow, R. D & R. J. Willis, 1972. "Nematode Diseases of Potatoes. II. Potato Cyst Nematode, Heterodera rostochiensis, 18-34”. In: Economic Nematology (Ed: J. Webster), Academic Press, New York., 563pp.
29. White, T. J., T. Bruns, S. Lee & J. Taylor, 1990. “Amplification and Direct Sequencing of Fungal Ribosomal RNA Genes for Phylogenetics, 315–322”. In:. PCR Protocols. A Guide to Methods and Applications (Eds: M. A. Innes, D. H. Gelfard, J. J. Sninsky & T. J. White). Academic Press, San Diego.