Rt-PCR Analysis and Evolutionary Relationship of Some Hungarian Grapevine leafroll associated virus 1 and 3 Isolates

Eszter Cseh, András Péter Takács, Richard Gáborjányi, László Palkovics, László Kocsis

Department of Horticulture, Georgikon Faculty, University of Pannonia, Keszthely, Hungary;.
Department of Plant Pathology, Faculty of Horticultural Science, Corvinus University of Budapest, Budapest, Hungary..
Plant Protection Institute, Georgikon Faculty, University of Pannonia, Keszthely, Hungary;.

DOI: 10.4236/ajps.2013.410250                

Abstract

Hungarian isolates of Grapevine leafroll associated virus 1 and 3 (GLRaV-1, GLRaV-3) were tested using serological (DAS-ELISA) and molecular (RT-PCR) methods. Five hundred bp long PCR products of the part of HSP70 gene of one serologically positive GLRaV-1 and four GLRaV-3 isolates were sequenced. These sequences were applied for phylogenetic analysis and compared to foreign virus isolates of NCBI GenBank. Phylogenetic analysis of GLRaV-1 HSP70 gene supported the earlier results that it could be divided into two clusters: E and A. The Hungarian isolate 6.4.1 belonged to the group E. This isolate showed the highest homology with the AY754914 isolate from the Czech Republic. GLRaV-3 sequence data could cluster five groups. Hungarian 2.2; 3.5 and 4.2 isolates were estimated belonging to the group II. The 1.4 isolate from the same vineyard as 2.2 varied in sequence data so it belonged to the other, IV. variant group with two South African, two Austrian and a Syrah isolate. According to the phylogenetic analysis, two variant groups occurred in Hungary. These isolates related with each other, but showed higher similarity of foreign counties. In some cases, they were similar to isolates of the neighbour countries such as Slovakia and Austria. It could be supposed that mainly the exchange of virus infected propagation materials caused the dissemination of GLRaV isolates.

Keywords

Grapevine; Virus; GLRaV-1; GLRaV-3; HSP70; RT-PCR; Hungary

Conflicts of Interest

The authors declare no conflicts of interest.

References

[1]	M. Kolber, J. Lázár, G. Farkas and J. Lehoczky, “Szolo Le-Vélsodródását Okozó Closterovírusok Kimutathatósága. 40th. Plant Protection Scientific Days. Budapest. Hungary (in Hungarian),” 1994, p. 86.
[2]	G. P. Martelli, “Graft Transmissible Diseases of Grapevines,” FAO Press, Rome, 1993, p. 263.
[3]	R. Sforza, E. Boudon-Padieu and C. Greif, “New Mealybug Species Vectoring Grapevine leafroll-associated viruses-1 and 3 (GLRaV-1 and -3),” European Journal of Plant Pathology, Vol. 209, pp. 975-981.
[4]	P. Komínek, M. Glasa and M. Bryxiová, “Analysis of the Molecular Variability of Grapevine leafroll-associated virus 1 Reveals the Presence of Two Distinct Virus Groups and Their Mixed Occurrence in Grapevines,” Virus Genes, Vol. 31, 2005, pp. 247-255.
[5]	C. Cabailero and A. Segura, “Field Transmission of Grapevine Leafroll Associated Virus 3 (GLRaV-3) by the Mealybug Planococcus citri,” Plant Disease, Vol. 81, 1997, pp. 283-286.
[6]	M. Cid, S. Pereira, C. Cabaleiro, F. Faoro and A. Segura, “Presence of Grapevine leafroll-associated virus 3 in Primary Salivary Glands of the Mealybug Vector Planococcus citri Suggests a Circular Transmission Mechanism,” European Journal of Plant Pathology, Vol. 118, 2007, pp. 23-30.
[7]	C. Cabaleiro, C. Couceiro, S. Pereira, M. Cid, M. Barrasa and A. Segura, “Spatial Analysis of Epidemics of Grapevine Leafroll Associated Virus-3,” European Journal of Plant Pathology, Vol. 121, 2008, pp. 121-130.
[8]	C. W. Tsai, J. Chau, L. Fernandez, D. Bosco, K. M. Daane and R. P. P. Almeida, “Transmission of Grapevine leafroll-associated virus 3 by the Vine Mealybug (Planococcus ficus),” Plant Disease, Vol. 98, 2008, pp. 1093-1098.
[9]	C. L. Petersen and J. G. Charles, “Transmission of Grapevine Leafroll-Associated Closteroviruses by Pseudococcus longispinosus and P. calceolariae,” Plant Pathology, Vol. 46, 1997, pp. 509-515. http://dx.doi.org/10.1046/j.1365-3059.1997.d01-44.x
[10]	D. A. Golino, S. T. Sim, R. Gill and A. Rowhani, “California Mealybugs Can Spread Grapevine Leafroll Disease,” California Agriculture, Vol. 56, No. 6, 2002, pp. 196-201. http://dx.doi.org/10.3733/ca.v056n06p196
[11]	C. Cabaleiro and A. Segura, “Temporal Analysis of grapevine leafroll associated virus 3 Epidemics,” European Journal of Plant Pathology, Vol. 114, No. 4, 2006, pp. 441-446. http://dx.doi.org/10.1007/s10658-006-0006-4
[12]	A. Little and M. A. Rezaian, “Improved Detection of Grapevine Leafroll-Associated Virus 1 by Magnetic Capture Hybridisation RT-PCR on a Conserved Region of Viral RNA,” Archives of Virology, Vol. 151, No. 4, 2006, pp. 753-761. http://dx.doi.org/10.1007/s00705-005-0649-z
[13]	C. Turturo, P. Saldarelli, D. Yafeng, M. Digiaro, A. Minafra, V. Savino and G. P. Martelli, “Genetic Variability and Population Structure of Grapevine leafroll-associated virus 3 Isolates,” Journal of General Virology, Vol. 86, No. 1, 2005, pp. 217-224. http://dx.doi.org/10.1099/vir.0.80395-0
[14]	A. A. Agranovsky, V. P. Boyko, A. V. Karasev, N. A. Lunina, E. V. Koonin and V. V. Dolja, “Nucleotide Sequence of the 3’-Terminal Half of Beet Yellows Closterovirus RNA Genome: Unique Arrangement of Eight Virus Genes,” Journal of General Virology, Vol. 72, No. 1, 1991, pp. 15-23. http://dx.doi.org/10.1099/0022-1317-72-1-15
[15]	K. M. Sefc, W. Leonhardt and H. Steinkellner, “Partial Sequence Identification of Grapevine-leafroll-associated virus-1 and Development of a Highly Sensitive IC-RT-PCR Detection Method,” Journal of Virological Methods, Vol. 86, No. 1, 2000, pp. 101-106. http://dx.doi.org/10.1016/S0166-0934(00)00135-X
[16]	V. I. Maliogka, C. I. Dovas and N. I. Katis, “Evolutionary Relationships of Virus Species Belonging to a Distinct Lineage within the Ampelovirus Genus,” Virus Research, Vol. 135, No. 1, 2008, pp. 125-135. http://dx.doi.org/10.1016/j.virusres.2008.02.015
[17]	M. F. Clark and A. N. Adams, “Characteristics of the Microplate Method of Enzyme-Linked Immunosorbent Assay for the Detection of Plant Viruses,” Journal of General Virology, Vol. 34, No. 3, 1977, pp. 475-483. http://dx.doi.org/10.1099/0022-1317-34-3-475
[18]	J. Sambrook, E. F. Fritsch and T. Maniatis, “Molecular Cloning: A Laboratory Manual,” 2nd Edition, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, 1989, p. 1659.
[19]	M. Fuchs, T. E. Martinson, G. M. Loeb and H. C. Hoch, “Survey for the Three Major Leafroll Disease-Associated Viruses in Finger Lakes Vineyards in New York,” Plant Disease, Vol. 93, No. 4, 2009, pp. 395-401. http://dx.doi.org/10.1094/PDIS-93-4-0395
[20]	A. E. C. Jooste and D. E. Goszczynski, “Single-Strand Conformation Polymorphism (SSCP), Cloning and Sequencing Revelas Two Major Groups of Divergent Molecular Variants of Grapevine leafroll-associated virus 3 (GLRaV3),” Vitis, Vol. 44, 2005, pp. 39-43.
[21]	A. E. C. Jooste, H. J. Maree, D. U. Bellstedt, D. E. Goszczynski, G. Pietersen and J. T. Burger, “Three Genetic Grapevine leafroll-associated virus 3 Variants Identified from South African Vineyards Show High Variability in Their 5’UTR,” Archives of Virology, Vol. 155, No. 2, 2010, pp. 1997-2006. http://dx.doi.org/10.1007/s00705-010-0793-y
[22]	H. J. Maree, M. J. Freeborough and J. T. Burger, “Complete Nucleotide Sequence of a South African Isolate of Grapevine leafroll-associated virus 3 Reveals a 5’UTR of 737 Nucleotides,” Archives of Virology, Vol. 153, No. 4, 2008, pp. 755-757. http://dx.doi.org/10.1007/s00705-008-0040-y