Long-term preservation of potato leafroll virus, potato virus S, and potato spindle tuber viroid in cryopreserved shoot tips
Availability of and easy access to diverse plant viruses and viroids is a prerequisite in applied and basic studies related to viruses and viroids. Long-term preservation of viruses and viroids is difficult. A protocol was described for long-term preservation of potato leafroll virus (PLRV), potato virus S (PVS), and potato spindle tuber viroid (PSTVd) in cryopreserved shoot tips of potato cv. Zihuabai. Shoot regrowth levels following cryopreservation were higher in 1.5 mm-shoot tips (58–60%) than in 0.5-mm-ones (30–38%). All shoots recovered from 0.5-mm-shoot tips were PVS- and PSTVd-preserved, but none of them were PLRV-preserved. Cryopreservation of 1.5-mm-shoot tips resulted in 35% and 100% of PLRV- and PVS- and PSTVd-preserved shoots. Studies on cell survival patterns and virus localization provided explanations to the varying PLRV-preservation frequencies produced by cryopreservation of the two sizes of shoot tips. Although micropropagation efficiencies were low after 12 weeks of subculture following cryopreservation, similar efficiencies were obtained after 16 weeks of subculture in pathogen-preserved shoots recovered from cryopreservation, compared with the diseased in vitro stock shoots (the control). Pathogen concentrations in the three pathogens-preserved shoots analyzed by qRT-PCR were similar to those in micropropagated shoots. The three pathogens cryopreserved in shoot tips were readily transmitted by grafting and mechanical inoculation to potato plants. PLRV, PVS, and PSTVd represent a diverse range of plant viruses and viroid in terms of taxonomy and infectious ability. Therefore, shoot tip cryopreservation opens a new avenue for long-term preservation of the virus and viroid.
KeywordsCryopreservation Potato Shoot tips Virus Viroid
Author contribution statements
J-W Li: performance of experiments, data collection and analysis, and preparation of manuscript; M-R Wang and Z-H Cui: assistance to the performance of experiments, data collection, and analysis; H-Y Chen: assistance to the performance of experiments; Z Zhang: valuable discussions; D-R Blystad: valuable discussions; L Zhao: financial support, experimental design, and preparation of manuscript; Q-C Wang: chief scientist of the project, financial supports, experimental design, and preparation of manuscript.
This research was funded by the National Natural Science Foundation of China (No. 31701761), Chinese Universities Scientific Fund (No. 2452017061), and Department of Science and Technology of Shaanxi Province, China (2014KTCL02–05).
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
This article does not contain any studies with human or animals performed by any of the authors.
- Bai J, Chen X, Lu X, Guo H, Xin X, Zhang Z (2012) Can cryopreservation eliminate the Potato virus X (PVX) and Potato spindle tuber viroid (PSTVd)? Biosci Meth 3:34–40Google Scholar
- Di Serio F, Navarro B, Flores R (2017) Origin and evolution of viroids. In: Hadidi A, Flores R, Randles JW, Palukaitis P (eds) Viroids and satellites. Academic Press, London, UK. pp. 125–134Google Scholar
- Fan NTF, Chen LF, Zhou YC, Shapiro B, Stiller M, Heintzman PD, Varsani A, Kondov NO, Wong W, Deng X, Andrews TD, Moorman BJ, Meulendyk T, MacKay G, Gilbertson RL, Delwart E (2014) Preservation of viral genomes in 700-y-old caribou feces from a subarctic ice patch. Proc Natl Acad Sci U S A 111:16842–16847CrossRefGoogle Scholar
- Helliot B, Panis B, Poumay Y, Swenen R, Lepoivre P, Frison E (2002) Cryopreservation for the elimination of cucumber mosaic and banana streak viruses from banana (Musa spp.). Plant Cell Rep 20:1117–22Google Scholar
- Hull R (2002) Mathews’ plant virology. Academic Press, London, UKGoogle Scholar
- Kim HH, Popova E, Shin DJ, Yi JY, Kim CH, Lee JS (2012) Cryobanking of Korean Allium germplasm collections: results from 10 year experience. CryoLett 33:45–57Google Scholar
- Kushnarenko S, Romadanova N, Aralbayeva M, Zholamanova S, Alexandrova A, Karpova O (2017) Combined ribavirin treatment and cryotherapy for efficient Potato virus M and Potato virus S eradication in potato (Solanum tuberosum L.) in vitro shoots. In Vitro Cell Dev Biol-Plant 53:425–4322CrossRefGoogle Scholar
- Li J-W, Chen H-Y, Li J, Zhang Z, Blystad D-R, Wang Q-C (2018) Growth, microtuber production and physiological metabolism in virus-free and virus-infected potato in vitro plantlets grown under NaCl-induced salt stress. Eur J Plant Path 152:417–432. https://doi.org/10.1007/s10658-018-1485-9 CrossRefGoogle Scholar
- Lomonossoff GP, Evans DJ (2011) Applications of plant viruses in bionanotechnology. Curr Top Microbiol 375:61–87Google Scholar
- Mckinney HH, Silver G, Greeley LW (1965) Longevity of some plant viruses stored in chemically dehydrated tissues. Phytopath 55:1043–1044Google Scholar
- Owens RA, Flores R, Di Serio F, Li SF, Pallas V, Randles JW (2012) Viroids. In: King AMQ, Adams MJ, Carstens EB, Lefkowitz EJ (eds), Virus taxonomy, ninth reports of the international committee on taxonomy of viruses. Elsevier/Academic Press, London, UK, pp. 1221–1234Google Scholar
- Sakai A (1960) Survival of the twigs of woody plants at −196°C. Nature 185:392–394Google Scholar
- Sudarshana MR, Roy G, Falk BW (2007) Methods for engineering resistance to plant viruses. In: Ronald PC (ed) Plant-pathogen interactions, methods in molecular biology, vol 354. Humana Press, New York, pp. 183–195Google Scholar
- Wang QC, Valkonen JPT (2008) Eradication of two synegistically interacting viruses from sweetpotato using shoot tip culture and cryotherapy of shoot tips. J Virol Meth 154:135–45Google Scholar
- Wang B, Yin Z-F, Feng C-H, Shi X, Li Y-P, Wang QC (2009a) Cryopreservation of potato shoot tips. In: Benkeblia, N. and Tennant, P. (eds), Potato I. Fruit, Vegetable and Cereal Science and Biotechnology 2 (Special Issue 1), Global Science Book, London, pp. 46–53Google Scholar
- Wang B, Zhang Z, Wang R-R, Ma L-Y, Wang Q-C (2013) Development of three vitrification-based cryopreservations of shoot tips for China’s potato. CryoLett 34:369–380Google Scholar
- Zhang Z, Haugslien S, Clark Liu JH, Spetz C, Lee Y, Sivertsen A, Skjeseth G, Blystad D-R, Wang Q-C (2014) Cryotherapy could not eradicate Chrysanthemum stunt viroid from infected Argyranthemum maderense ‘Yellow Empire. Acta Hortic (1039):201–208Google Scholar