Sel’skokhozyaistvennaya Biologiya [Agricultural Biology], 2017, Vol. 52, № 1, p. 95-104.
(how to cite this article)

doi: 10.15389/agrobiology.2017.1.95eng

UDC 635.21:573.6.086.83:[632.935.11+632.934.3

Acknowledgements:
The paper was prepared with the support from the Basic Research Program performed at state academies for 2013-2020. The final stage of the work was partially supported by the Special Complex Program of the Russian Federation (2016-2025) on improving the competitiveness of agricultural products (priority for potato research).

 

ERADICATION OF VIRUSES IN MICROPLANTS OF THREE
CULTIVATED POTATO SPECIES (Solanum tuberosum L.,
S. phureja Juz. & Buk., S. stenotomum Juz. & Buk.) USING COMBINED
THERMO-CHEMOTHERAPY METHOD

O.Yu. Antonova, O.V. Apalikova, Yu.V. Ukhatova, E.A. Krylova,
O.Yu. Shuvalov, A.R. Shuvalova, T.A. Gavrilenko

Federal Research Center the N.I. Vavilov All-Russian Institute of Plant Genetic Resources, Federal Agency of Scientific Organizations, 42-44, ul. Bol’shaya Morskaya, St. Petersburg, 190000 Russia,
e-mail tatjana9972@yandex.ru

ORCID:
Antonova O.Yu. orcid.org/0000-0001-8334-8069
Apalikova O.V. orcid.org/0000-0002-1188-2446
Ukhatova Yu.V. orcid.org/0000-0001-9366-0216
Gavrilenko T.A. orcid.org/0000-0002-2605-6569

Received October 17, 2016

 

A modified method of combined chemo-thermotherapy for eradication the most damaging viruses in potato plants is proposed. The materials for study included 91 clones of 85 accessions of three South-American cultivated species Solanum phureja (18 accessions), S. stenotomum (26 accessions), S. tuberosum (S. tuberosum ssp. andigenum and S. tuberosum ssp. tuberosum) (41 accessions) from the VIR field collection (N.I. Vavilov All-Russian Institute of Plant Genetic Resources). The results of ELISA for 91 clones from the field collection and RT-PCR analysis of the corresponding microplants, when introducing clones into in vitro culture, have shown that all material was infected by viruses (potato viruses Y, X, S, M and potato leafroll virus — PVY, PVX, PVS, PVM and PLRV) to varying degrees. Single infections were detected in 25.3 % of the 91 clones, and the remaining clones were multi-infected and contained several viruses in various combinations. Based on the termo- and chemotherapy protocols as well as complex therapy used in leading potato genebanks (Leibniz Institute of Plant Genetics and Crop Plant Research — IPK, Germany; International Potato Centre — CIP, Peru), we have developed a modified method of combined therapy comprising three successive cycles of in vitro plant cultivation on Murashige and Skoog basal medium (MS) supplemented with ribavirin (30 mg/l) at 37 °С during 4 weeks. Between the cycles, the microplants were incubated for 2 days at 26 °С on the same medium with ribavirin. Each clone was tested for the presence of viruses by RT (revers transcription) PCR before and after the combined thermo-chemotherapy. In RT-PCR, 42 clones (46 %) were completely free from tested viruses, 44 % were free from some viruses, and 10 % of the clones were unable to improve. On the whole of 91 clones, PLRV was eliminated from 72.7 %, PVY from 71.4 %, PVM from 63.9 % and PVS from 57.4 % of microplants. Differences in elimination frequency between the viruses were not significant (р > 0.05). Also, there were no significant differences in elimination of different viruses between the accessions of different cultivated species. The difference of this modification from IPK protocols (Germany) lays in simultaneous effect of high temperature (37 °C) and ribavirin (instead of sequential application), and, unlike the CIP (Peru) scheme, it comprises less steps.

Keywords: potatoes, Solanum tuberosum, Solanum phureja, Solanum stenotomum, microplants, PVY, PVX, PVS, PVM, PLRV, virus eradication, RT-PCR, thermotherapy, chemotherapy.

 

Full article (Rus)

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REFERENCES

  1. Polevaya kollektsiya kartofelya VIR [VIR Potato field collection]. Available http://vir.nw.ru/otd_r.htm#tuber. Accessed Ocober 4, 2016 (in Russ.).
  2. Jeffries C.J. Potato. FAO/IPGRI technical guidelines for the safe movement of germplasm. FAO and IPGRI, Rome, 1998.
  3. Virus and virus-like diseases of potatoes and production of seed potatoes. G. Loenbenstein, P.H. Berger, A.A. Brunt, R.H. Lawson (eds.). Springer Netherlands, The Netherlands, 2001 CrossRef
  4. Vlasov Yu.I., Larina E.I., Truskinov E.V. Sel'skokhozyaistvennaya fitovirusologiya [Agricultural phytovirology]. St. Petersburg, 2016 (in Russ.).
  5. Anisimov B.V., Belov G.L., Varitsev Yu.A., Elanskii S.N., Zhurom-
    skii G.K., Zavriev S.K., Zeiruk V.N., Ivanyuk V.G., Kuznetsova M.A., Plyakhnevich M.P., Pshechenkov K.A., Simakov E.A., Sklyarova N.P., Uskov A.I., Yashina I.M. Zashchita kartofelya ot boleznei, vreditelei i sornyakov [Poteto protection from diseases, pests and weeds]. Moscow, 2009 (in Russ.).
  6. Morel G.T., Martin C. Guerison de dahlias atteints dune maladie a virus. Comptes Rendus de l'Académie des Sciences, Ser. III, 1952, 235: 1324-1325.
  7. Truskinov E.V. Sel'skokhozyaistvennaya biologiya [Agricultural Biology], 1976, 11(2): 250-255 (in Russ.).
  8. Wang Q.C., Liu Y., Xie Y.H., You M. Cryotherapy of potato shoot tips for efficient elimination of Potato leaf roll virus (PLRV) and Potato virus Y (PVY). Potato Res., 2006, 49(2): 119-129 CrossRef
  9. Badarau C.L., Chiru N. Effect of some therapies on potato plantlets infected with potato virus X (PVX). Journal of EcoAgriTourism, 2014, 10(1/28): 11-17.
  10. Mahmoud S.Y.M., Hosseny M.H., Abdel-Ghaffar M.H. Evaluation of some therapies to eliminate Potato Y potivirus from potato plants. Int. J. Virol., 2009, 5(2): 64-76 CrossRef
  11. Griffiths H.M., Slack S.A., Dodds J.H. Effect of chemical and heat therapy on virus concentration in in vitro plantlets. Can. J. Bot., 1990, 68: 1515-1521.
  12. Cassells A.C., Long R.D. The elimination of potato viruses X, Y, S and M in meristem and explant cultures of potato in the presence of Virazole. Potato Res., 1982, 25: 165-173.
  13. Faccioli G., Colalongo M.C. Eradication of Potato virus Y and Potato leafroll virus by chemotherapy of infected potato stem cuttings. Phytopathol. Mediterr., 2002, 41: 76-78.
  14. Truskinov E.V., Rogozina E.V. Fiziologiya rastenii, 1997, 43: 432-439 (in Russ.).
  15. Atabekov I.G., Tal'yanskii M.E. V sbornike: Dostizheniya sel'skokhozyaistvennoi nauki [In: Progress in agricultural science]. Moscow, 1987: 121-136 (in Russ.).
  16. Nascimento L.C., Pio-Ribeiro G., Willadino L., Andrade G.P. Stock indexing and potato virus Yelimination from potato plants cultivated in vitro. Scientia Agricola, 2003, 60(3): 525-530 CrossRef
  17. Dhital S.P., Sakha B.M., Lim H.T.Utilization of shoot cuttings for elimination of PLRV and PVY by thermotherapy and chemotherapy from potato (Solanum tuberosum L.). Nepal Journal of Science and Technology, 2006, 7: 1-6.
  18. Techniques in plant virology. Section 4.2. Virus eradication: tissue culture of meristems, thermotherapy, and chemotherapy. L. Salazar, U. Jayasinghe (eds.). International Potato Center, Lima, Peru, 1999.
  19. Panta A., Tay D., Gomez R., Zea B., Rojas E., Simon R., Roca W. Status and impact of the in vitro conservation of root and tubers at the International Potato Centre (CIP). Proc. 15th Triennial Symposium of the International Society for Tropical Root Crops. Lima, Peru, 2009: 15-24.
  20. Operational Genebank manual of IPK. Gatersleben, 2011.
  21. Lizarraga R., Panta A., Jayasinghe U., Dodds J. Tissue culture for elimination of pathogens. CIP Research Guide 3. International Potato Center, Lima, Peru, 1991.
  22. Jianming B., Xiaoling C., Xinxiong L., Huachun G., Xia X., Zhie Z. Can cryopreservation eliminate the Potato virus X (PVX) and Potato spindle tuber viroid (PSTVd)? Bioscience Methods, 2012, 3(5): 34-40 CrossRef.
  23. Faccioli G. Control of potato viruses using meristem and stem-cutting cultures, thermotherapy and chemotherapy. In: Virus and virus-like diseases of potatoes and production of seed-potatoes. G. Loebenstein, P.H. Berger, A.A Brunt, R.H. Lawson (eds.). Springer Netherlands, 2001: 365-390 CrossRef
  24. Heldak J. Detection of potato virus S by RT-PCR in potato regenerants derived from in vitro heat-treated shoot tips. Acta fytotechnica et zootechnica, 2001, 4: 275-277.
  25. Klein R.E., Livingston C.H. Eradication of potato viruses X and S from potato shoot-tip cultures with ribavirin. Phytopathology, 1983, 73(7): 1049-1050 CrossRef
  26. Yang L., Nie B., Liu J., Song B. A reexamination of the effectiveness of ribavirin on eradication of viruses in potato plantlets in vitro using ELISA and quantitative RT-PCR. Am. J. Potato Res., 2014, 91(3): 304-311 CrossRef
  27. Sanchez G.E., Slack S.A., Dodds J.H. Response of selected Solanum species to virus eradication therapy. Am. Potato J., 1991, 68: 299-315 CrossRef
  28. Hawkes J.G. The potato. Evolution, biodiversity, genetic resources. Belhaven Press, London, 1990.
  29. Bukasov S.M. Trudy po prikladnoi botanike, genetike i selektsii (Leningrad), 1978, 62(1): 3-35 (in Russ.).
  30. Dunaeva S.E., Pendinen G.I., Antonova O.Yu., Shvachko N.A., Volkova N.N., Gavrilenko T.A. Sokhranenie vegetativno razmnozhaemykh kul'tur v in vitro i kriokollektsiyakh /Pod redaktsiei T.A. Gavrilenko [Preservation of vegetatively propagated crops in vitro and in cryo collections. T.A. Gavrilenko (ed.)]. St. Petersburg, 2011 (in Russ.).
  31. Nabor «Vydelenie total'noi RNK na magnitnykh chastitsakh, pokrytykh SiO2» [Kit for total RNA isolation on magnetic particles coated with SiO2]. Aailable http://www.sileks.com/ru/production.php?folder=189. Accessed September 25, 2016 (in Russ.).
  32. Nabor «Sintez pervoi tsepi kDNK (rendom)» firmy Sileks [Kit for first cDNA shain random synthesis. Sileks company]. Available http://www.sileks.com/ru/production.php?folder=65. Accessed September 25, 2016 (in Russ.).
  33. Singh R.P. Development of the molecular methods for potato virus and viroid detection and prevention. Genome, 1999, 42: 592-604.
  34. Xu H., D’Aubin J., Nie J. Genomic variability in Potato virus M and the development of RT-PCR and RFLP procedures for the detection of this virus in seed potatoes. Virol. J., 2010, 7: 25 CrossRef
  35. Bostan H., Peker P.K. The feasibility of tetraplex RT-PCR in the determination of PVS, PLRV, PVX and PVY from dormant potato tubers. Afr. J. Biotechnol., 2009, 8(17): 4043-4047.
  36. Nicot N., Hausman J.-F., Hoffmann L., Evers D. Housekeeping gene selection for real-time RT-PCR normalization in potato during biotic and abiotic stress. J. Exp. Bot., 2005, 56(421): 2907-2914 CrossRef
  37. Nasir I.A., Tabassum B., Latif Z., Javed M.A., Haider M.S., Javed M.A., Husnain T. Strategies to control potato virus y under in vitroconditions. Pak. J. Phytopathol., 2010, 22(1): 63-70.
  38. Zapata C., Miller J.C., Jr., Smith R.H. An in vitro procedure to eradicate potato viruses X, Y, and S from Russet Norkotah and two of its strains. In Vitro Cellular & Developmental Biology — Plant, 1995, 31(3): 153-159 CrossRef
  39. Morozova Z.R. Biologicheskie nauki, 1992, 338(2): 128-134 (in Russ.).
  40. Leonova N.S., Salganik N.I. Sibirskii vestnik sel'skokhozyaistvennoi nauki, 1991, 5: 25-28 (in Russ.).
  41. Lopez-Delgado H., Mora-Herrera M.E., Zavaleta-Mancera H.A., Cadena-Hinojosa M., Scott I.M. Salicylic acid enhances heat tolerance and potato virus X (PVX) elimination during thermotherapy of potato microplants. Am. J. Potato Res., 2004, 81(3): 171-176 CrossRef
  42. Paet C.N., Zamora A.B. Efficacy of thermotherapy and group culture of isolated potato meristems for the elimination of single and mixed infections of Potato Virus Y, Potato Virus S and Potato Leaf Roll Virus. Philipp. J. Crop. Sci., 1990, 15(2): 113-118.

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