Abstract
This study attempted to eradicate Apple stem pitting virus (ASPV) and Apple stem grooving virus (ASGV) from ‘Gala’ apple using shoot tips from of adventitious shoots derived from leaf segments. Leaf segments were excised from in vitro 4-week-old stock shoots and cultured on shoot regeneration medium containing 9.1 µM thidiazuron and 0.25 µM indole-3-butyric acid (IBA) to induce adventitious shoot formation. Shoot tips of different size and different developmental stage were excised from adventitious shoots and cultured on shoot tip culture medium containing 1.1 µM 6-benzyladenine and 0.05 µM IBA. Results showed size and developmental stage of shoot tips excised from adventitious shoots did not influence survival rate, but significantly affected shoot regrowth rate and ASPV-free frequency. Shoot regrowth rates increased from 10 to 15 % in 0.3 mm shoot tips containing two leaf primordia (LP) excised after 2–3 weeks of shoot regeneration, to 53–55 % in those containing three LP excised after 3–4 weeks. The highest shoot regrowth rate (82 %) was obtained in shoot tips of 0.4 mm shoot tips containing four LP excised after 4 weeks. ASPV-free frequencies (95–100 %) were high in 0.2–0.4 mm shoot tips containing two to three LP excised after 2–4 weeks, but low (20 %) in 0.4 mm shoot tips containing four LP excised after 4 weeks. None of the shoots regenerated from the shoot tips were ASGV-free, regardless of the size and developmental stage at which shoot tips were excised. Histological studies and virus localization provided explanations to the varying frequencies of the virus eradication using different size of shoot tips that were excised from adventitious shoots at different developmental stages. The protocol described here was characteristic of high frequency ASPV eradication and high efficiency of production of shoot tips that were used for virus eradication, and thus may have potential applications to virus eradication in other plant species.
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Abbreviations
- AD:
-
Apical dome
- ApMV:
-
Apple mosaic virus
- ASGV:
-
Apple stem grooving virus
- ASPV:
-
Apple stem pitting virus
- BA:
-
6-Benzyladenine
- CMV:
-
Cucumber mosaic virus
- GCLV:
-
Garlic common latent virus
- GLV:
-
Garlic latent virus
- IBA:
-
Indole-3-butyric acid
- ISSR:
-
Inter-simple sequence repeat
- LP:
-
Leaf primordia
- LSV:
-
Lily symptomless virus
- LYSV:
-
Leek yellow stripe virus
- MD:
-
Meristematic dome
- MS:
-
Murashige and Skoog medium
- NAA:
-
Naphthalene acetic acid
- OYDV:
-
Onion yellow dwarf virus
- RAPD:
-
Random amplified polymorphic DNA
- RT-PCR:
-
Reverse transcription-polymerase chain reaction
- SLV:
-
Shallot latent virus
- SRM:
-
Shoot regeneration medium
- STCM:
-
Shoot tip culture medium
- TDZ:
-
Thidiazuron
References
Aldwinckle H, Malnoy M (2009) Plant regeneration and transformation in the Rosaceae. Transgenic Plant J 3:1–39
Ayabe M, Sumi S (2001) A novel and efficient tissue culture method—“stem-disc dome culture”—for producing virus-free garlic (Allium sativum L.). Plant Cell Rep 20:503–507
Brischia R, Piccioni E, Standardi A (2002) Micropropagation and synthetic seed in M.26 apple rootstock (II): a new protocol for production of encapsulated differentiating propagules. Plant Cell Tissue Organ Cult 68:137–141
Brison M, Boucaud M-T, Pierronnet A, Dosba F (1997) Effect of cryopreservation on the sanitary state of a cv. Prunus rootstock experimentally contaminated with Plum Pox Potyvirus. Plant Sci 123:189–196
Caboni E, Lauri P, Damiano C, D’Angeli S (2000) Somaclonal variation induced by adventitious shoot regeneration in pear and apple. Acta Hortic 530:195–201
Cieslinska M, Rutkowski KP (2008) Effect of Apple chlorotic leaf spot virus on yield and quality of fruits from ‘Golden Delicious’ and ‘Sampion’ apple trees. Acta Hortic 781:119–124
Conejero A, Romero C, Cunill M, Mestre MA, Martínez-Calvo J, Badenes ML, Llácer G (2013) In vitro shoot-tip grafting for safe Prunus budwood exchange. Sci Hortic 150:365–370
Dobránszki J, Teixeira da Silva JA (2010) Micropropagation of apple—a review. Biotechnol Adv 28:462–488
EPPO (1998) Certification Schemes PM4/1-26 European and Mediterranean Plant Protection Organization, Paris
Faccioli G, Marani F (1998) Virus elimination by meristem tip culture and tip micrografting. In: Hadidi A, Khetarpal RK, Koganezawa H (eds) Plant virus disease control. APS Press, St Paul, pp 346–380
Feng C-H, Cui Z-H, Li B-Q, Chen L, Ma Y-L, Zhao Y-H, Wang QC (2013) Duration of sucrose preculture is critical for shoot regrowth of in vitro-grown apple shoot-tips cryopreserved by encapsulation-dehydration. Plant Cell Tissue Organ Cult 112:369–378
Gupta R, Modgil M, Chakrabarti SK (2009) Assessment of genetic fidelity of micropropagated apple rootstock plants, EMLA 111, using RAPD markers. Ind J Exp Bot 47:925–928
Hadidi A, Barba M (2011) Economic impact of pome and stone fruit viruses and viroids. In: Hadidi A, Barba M, Candresse W, Jelkman W (eds) Virus and virus-like diseases of pome and stone fruits. APS Press, St Paul, pp 1–7
Han D-S, Niimi Y, Kimura S (2006) Localization of lily symptomless virus and cucumber mosaic virus in anther- and filament-derived calluses and effect of callus culture duration on virus-free bulblet production in Lilium ‘Enchantment’. Plant Cell Tissue Organ Cult 87:211–217
Hansen J, Lane WD (1985) Elimination of Apple chlorotic leaf spot virus from apple shoot cultures by ribavirin. Plant Dis 69:134–135
Hu GJ, Hong N, Wang LP, Hu HJ, Wang GP (2012) Efficacy of virus elimination from in vitro-cultured sand pear (Pyrus pyrifolia) by chemotherapy combined with thermotherapy. Crop Prot 37:20–25
Hu GJ, Dong YF, Zhang ZP, Fan XD, Ren F, Zhou J (2015) Virus elimination from in vitro apple by thermotherapy combined with chemotherapy. Plant Cell Tissue Organ Cult 121:435–443
Huang SC, Millikan DF (1980) In vitro micrografting of apple shoot tips. HortScience 15:741–743
Hull R (2002) Matthews’ plant virology, 4th edn. Academic Press, London
James D, Trytten PA, Mackenzie DJ, Towers GHN, French CJ (1997) Elimination of Apple stem grooving virus by chemotherapy and development of an immunocapture RT-PCR for rapid sensitive screening. Ann Appl Biol 131:459–470
Jelkmann W, Paunovic S (2011) Apple stem pitting virus. In: Hadidi A, Barba M, Candresse W, Jelkman W (eds) Virus and virus-like diseases of pome and stone fruits. APS Press, St Paul, pp 35–40
Ji Z, Zhao X, Duan H, Hu T, Wang S, Wang Y, Cao K (2013) Multiplex RT-PCR detection and distribution of four apple viruses in China. Acta Virol 57:435–441
Kartha KK, Gamborg OL (1975) Elimination of cassava mosaic disease by meristem culture. Phytopathol 65:826–828
Kaushal N, Modgil M, Thakur M, Sharma DR (2005) In vitro clonal multiplication of an apple rootstock by culture of shoot apices and axillary buds. Ind J Exp Biol 43:561–565
Koike H, Makita H, Tsukahara T, Sukahara K (1993) Effect of an Apple-chlorotic-leaf-spot-virus free M.9 rootstock on the growth of apple trees. J Jpn Soc Hortic Sci 62:499–504
Laimer M, Barba M (2011) Elimination of systemic pathogens by thermotherapy, tissue culture, or in vitro micrografting. In: Hadidi A, Barba M, Candresse W, Jelkman W (eds) Virus and virus-like diseases of pome and stone fruits. APS Press, St Paul, pp 389–393
Li B-Q, Feng C-H, Hu L-Y, Wang M-R, Chen L, Wang Q-C (2014) Shoot regeneration and cryopreservation of shoot tips of apple (Malus) by encapsulation-dehydration. In Vitro Cell Dev Biol Plant 50:357–368
Li B-Q, Feng F-C, Hu L-Y, Wang M-R, Wang Q-C (2016) Shoot tip culture and cryopreservation for eradication of Apple stem pitting virus (ASPV) and Apple stem grooving virus (ASGV) from apple rootstocks ‘M9’ and ‘M26’. Ann Appl Biol 168:142–150
MacKenzie DJ, McLean MA, Mukerji S, Green M (1997) Improved RNA extraction from woody plants for the detection of viral pathogens by reverse transcription-polymerase chain reaction. Plant Dis 81:222–226
Magyar-Tábori K, Dobránszki J, Teixeira da Silva JA, Bulley SM, Hudák I (2010) The role of cytokinins in shoot organogenesis in apple. Plant Cell Tissue Organ Cult 101:251–267
Massart S, Jijakli MH, Kummert J (2011) Apple stem grooving virus. In: Hadidi A, Barba M, Candresse W, Jelkman W (eds) Virus and virus-like diseases of pome and stone fruits. APS Press, St Paul, pp 29–33
Menzel W, Jelkmann W, Maiss E (2002) Detection of four apple viruses by multiplex RT-PCR assays with co-amplification of plant mRNA as internal control. J Virol Methods 99:81–92
Mink GI, Wample R, Howell WE (1998) Heat treatment of perennial plants to eliminate phytoplasms, viruses and viroids while maintaining plant survival. In: Hadidi A, Khetarpal RK, Koganezawa H (eds) Plant virus disease control. APS Press, St Paul, pp 332–345
Modgil M, Mahajan K, Chakrabarti SK, Sharma DR, Sobti RC (2005) Molecular analysis of genetic stability in micropropagated apple rootstock MM106. Sci Hortic 104:151–160
Montecelli S, Gentile A, Damino C (2000) In vitro shoot regeneration of apple cultivar Gala. Acta Hortic 530:219–223
Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco cell cultures. Physiol Plant 15:473–497
O’Herlihy EA, Croke JT, Cassells AC (2003) Influence of in vitro factors on titre and elimination of model fruit tree viruses. Plant Cell Tissue Organ Cult 72:33–42
Paprstein F, Sedlak J, Polak J, Svobodova L, Hassan M, Bryxiova M (2008) Results of in vitro thermotherapy of apple cultivars. Plant Cell Tissue Organ Cult 94:347–352
Pathak H, Dhawan V (2010) Molecular analysis of micropropagated apple rootstock MM111 using ISSR markers for ascertaining clonal fidelity. Acta Hortic 865:73–80
Pathak H, Dhawan V (2012) ISSR assay for ascertaining genetic fidelity of micropropagated plants of apple rootstock Merton 793. In Vitro Cell Dev Biol Plant 48:137–143
Plopa C, Preda S (2013) Elimination of Apple mosaic virus by tissue culture of some infected apple cultivars. Acta Hortic 981:517–522
Sakai W (1973) Simple method for differential staining of paraffin embedded plant material using toluidine blue O. Stain Technol 48:247–249
Sedlak J, Paprstein F, Talacko L (2011) Elimination of Apple stem pitting virus from pear cultivars by in vitro chemotherapy. Acta Hortic 923:111–115
Tan RR, Wang LP, Hong N, Wang GP (2010) Enhanced efficiency of virus eradication following thermotherapy of shoot-tip cultures of pear. Plant Cell Tissue Organ Cult 101:229–235
Viršcek-Marn M, Javornik B, Štampar F, Bohanec B (1998) Assessment of genetic variation among regenerants from in vitro apple leaves using molecular markers. Acta Hortic 484:299–303
Wang Q-C, Mawassi M, Li P, Gafny R, Sela I, Tanne E (2003) Elimination of Grapevine virus A (GVA) by cryopreservation of in vitro-grown shoot tips of Vitis vinifera L. Plant Sci 165:321–327
Wang LP, Wang GP, Hong N, Tang RR, Deng XY (2006) Effect of thermotherapy on elimination of Apple stem grooving virus and Apple chlorotic leaf spot virus for in vitro-cultured pear shoot tips. HortSci 41:729–732
Wang LP, Hong H, Wang GP, Xu WX, Michelutti R, Wang AM (2010) Distribution of apple stem grooving virus and apple chlorotic leaf spot virus in infected in vitro pear shoots. Crop Prot 29:1447–1451
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The authors acknowledge financial support from the fund provided Department of Science and Technology of Shaanxi Province (2014KTCL02-05).
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Wang, MR., Li, BQ., Feng, CH. et al. Culture of shoot tips from adventitious shoots can eradicate Apple stem pitting virus but fails in Apple stem grooving virus . Plant Cell Tiss Organ Cult 125, 283–291 (2016). https://doi.org/10.1007/s11240-016-0948-y
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DOI: https://doi.org/10.1007/s11240-016-0948-y