Abstract
Citrus tristeza virus (CTV) is the most economically important viral disease of citrus worldwide. Cultivars with improved CTV tolerance or resistance are needed to manage CTV-induced diseases. The citrus relatives Poncirus trifoliata (L.) Raf., Swinglea glutinosa (Blanco) Merr., and Severinia buxifolia (Poir) Ten. are potential sources of CTV resistance, but their resistance mechanisms are poorly characterized. As a first step to examine the mechanisms of resistance to CTV in these citrus relatives and selected Citrus × Poncirus hybrids, it was necessary to develop methods for protoplast isolation and viral inoculation to allow examination of CTV multiplication in this range of citrus varieties and relatives. Leaf and/or cultured cell protoplasts were isolated and inoculated with four biologically distinct CTV isolates. Northern-blot hybridization analyses for progeny RNAs and immuno-electron microscopy assays for newly produced virions showed that CTV replicated and produced infectious particles in protoplasts from all of the resistant plants tested. These results suggest that resistance to CTV observed at the plant level results from a lack of virus movement and/or some induced resistance response, rather than lack of viral multiplication at the cellular level.
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Albiach-Marti M.R., Mawassi M., Gowda S., Satyanarayana T., Hilf M.E., Shanker S., Almira E.C., Vives M.C., Lopez C., Guerri J., Flores R., Moreno P., Garnsey S.M., and Dawson W.O. 2000. Sequences of citrus tristeza virusseparated in time and space are essentially identical. J. Virol. 74: 6856–6865.
Barrett H.C. 1990. US119, an intergeneric hybrid citrus scion breeding line. HortScience 25: 1670–1671.
Bar-Joseph M., Marcus R. and Lee R.F. 1989. The continuous challenge of citrus tristeza virus control. Annu. Rev. Phytopathol. 27: 291–316.
Castle W.S. 1987. Citrus rootstocks, p. 361–399. In: Rom R.C. and Carlson R.F. (eds), Rootstocks for fruit crops. Wiley, New York, USA.
Dawson W.O. and Hilf M.E. 1992. Host-range determinants of plant viruses. Annu. Rev. Plant Physiol. Plant Mol. Biol. 43: 527–555.
Dawson T.E. and Mooney P.A. 2000. Evidence for trifoliate resistance breaking isolates of citrus tristeza virus in New Zealand, pp. 69–76. In: da Graca J.V., Lee R.F. and Yokomi R.K. (eds), Proc. 14th Conf. Int. Org. Citrus Virologist. IOCV-1998.
Deng Z., Huang S., Xiao S. and Gmitter F.G. Jr. 1997. Development and characterization of SCAR markers linked to the citrus tristeza virus resistance gene from Poncirus trifoliata. Genome 40: 697–704.
Deng A., Huang S., Ling P., Chen C., Yu C., Weber C.A., Moore G.A. and Gmitter F.G. Jr. 2000. Cloning and characterization of NBS-LRR class resistance gene candidate sequences in citrus. Theor. Appl. Genet. 101: 812–814.
Deng A., Huang S., Ling P., Yu C., Tao Q., Chen C., Wendell M.K., Zhang H.B. and Gmitter F.G. Jr. 2001. Fine genetic mapping and BAC containing development for the citrus tristeza virus resistance gene locus in Poncirus trifoliata(Raf.). Mol. Genet. Genomics 2265: 739–747.
Derrick K. and Brlansky R.H. 1976. Assay for viruses and mycoplasmas using serologically specific electron microscopy. Phytopathology 66: 815–820.
Fang D.Q., Federici C.T. and Roose M.L. 1998. A high-resolution linkage map of the citrus tristeza virus resistance gene region in Poncirus trifoliata(L.) Raf. Genetics 150: 883–890.
Fraser R.S.S. 1990. The genetics of resistance to plant viruses. Annu. Rev. Phytopathol. 28: 179–200.
Garnsey S.M., Barrett H.C. and Hutchison D.J. 1987. Identification of citrus tristeza virus resistance in citrus relatives and its potential applications. Phytophylactica 19: 187–191.
Garnsey S.M., Permar T.A., Cambra M. and Henderson C.T. 1993. Direct tissue blot immunoassay (DTBIA) for detection of citrus tristeza virus, pp. 39–50. In: Moreno P., da Graca J.V. and Timmer L.W. (eds), Proc. 12th Conf. Int. Org. Citrus Virologist (IOCV). IOCV-1993.
Garnsey S.M., Su H.J. and Tsai M.C. 1996. Differential susceptibility of pummelo and Swingle citrumelo to isolates of citrus tristeza virus, pp. 138–146. In: da Graca J.V., Moreno P. and Yokomi R.K. (eds), Proc. 13th Conf. Int. Org. Citrus Virologist. IOCV-1996.
Gmitter F.G. Jr., Xiao S.Y., Huang S., Hu X.L., Garnsey S.M. and Deng Z. 1996. A localized linkage map of the citrus tristeza virus resistance gene region. Theor. Appl. Genet. 92: 688–695.
Grosser J.W. and Gmitter F.G. Jr. 1990. Protoplast fusion and citrus improvement. Plant Breed Rev. 8: 339–374.
Grosser J.W. and Chandler J.L. 2000. Somatic hybridization of high yield, cold-hardy and disease resistant parents for citrus rootstock improvement. J. Hortic. Sci. Biotech. 75: 641–644.
Guerra M., dos Santos K.G., Barros E., Silva A.E. and Ehrendorfer F. 2000. Heterochromatin banding patterns in Rutaceae-Aurantioideae-a case of parallel chromosomal evolution. Am. J. Bot. 87: 735–747.
Herrero R., Asins M.J., Carbonell E.A. and Navarro L. 1996a. Genetic diversity in the orange subfamily Aurantioideae. I. Intraspecies and intragenus genetic variability. Theor. Appl. Genet. 92: 599–609.
Herrero R., Asins M.J., Pina J.A., Carbonell E.A. and Navarro L. 1996b. Genetic diversity in the orange subfamily Aurantioideae. II. Genetic relationships among genera and species. Theor. Appl. Genet. 93: 1327–1334.
Hilf M.E., Karasev A.V., Pappu H.R., Gumpf D.J., Niblett C.L. and Garnsey S.M. 1995. Characterization of citrus tristeza virus subgenomic RNAs in infected tissue. Virology 208: 576–582.
Hilf M.E., Karasev A.V., Albiach-Marti M.R., Dawson W.O. and Garnsey S.M. 1999. Two paths of sequence divergence in the citrus tristeza virus complex. Phytopathology 89: 336–342.
Hilf M.E. and Garnsey S.M. 2000. Characterization and classification of citrus tristeza virus isolates by amplification of multiple molecular markers, pp. 18–27. In: da Graca J.V., Lee R.F. and Yokomi R.K. (eds), Proc. 14th Conf. Int. Org. Citrus Virologist. IOCV-1998.
Iwamasa M. and Nito N. 1988. Cytogenetics and the evolution of modern cultivated citrus, pp. 265–275. In: Goren R. and Mendel K. (eds), Proc. 6th Int. Citrus Congress. Margraf Scientific Books, Weikersheim, Germany.
Lewandowski D.J. and Dawson W.O. 1998. Deletion of internal sequences results in tobacco mosaic virus defective RNAs that accumulate to high levels without interfering with replication of the helper virus. Virology 251: 427–437.
Mansky L.M. and Hill J.H. 1993. Molecular basis for virus disease resistance in plants. Arch. Virol. 131: 1–16.
Mawassi M., Mietkiewska E., Hilf M.E., Ashoulin L., Karasev A.V., Gafny R., Lee R.F., Garnsey S.M., Dawson W.O. and Bar-Joseph M. 1995. Multiple species of defective RNAs in plants infected with citrus tristeza virus. Virology 214: 264–268.
Mestre P.F., Asins M.J., Pina J.A., Carbonell E.A. and Navarro L. 1997a. Molecular markers flanking citrus tristeza virus resistance gene from Poncirus trifoliata(L.) Raf. Theor. Appl. Genet. 94: 458–464.
Mestre P.F., Asins M.J., Carbonell E.A. and Navarro L. 1997b. New gene(s) involved in the resistance of Poncirus trifoliata(L.) Raf. to citrus tristeza virus. Theor. Appl. Genet. 95: 691–695.
Mestre P.F., Asins M.J., Pina J.A. and Navarro L. 1997c. Efficient search for new resistant genotypes to the citrus tristeza virus in the orange subfamily Aurantioideae. Theor. Appl. Genet. 95: 1282–1288.
Moore G.A. 2001. Oranges and lemons: Clues to the taxonomy of citrus from molecular markers. Trends Genet. 17: 536–540.
Navas-Castillo J., Albiach-Marti M.R., Gowda S., Hilf M.E., Garnsey S.M., and Dawson W.O. 1997. Kinetics of accumulation of citrus tristeza virus RNAs. Virology 228: 92–97.
Nicolosi E., Deng Z.N., Gentile A., La Malfa S., Continella G. and Tribulato E. 2000. Citrus phylogeny and genetic origin of important species as investigated by molecular markers. Theor. Appl. Genet. 100: 1155–1166.
Olivares-Fuster O., Albiach-Marti M.R., Gowda S., Fleming G.H., Dawson W.O., and Grosser J.W. 2000. A co-transformation strategy for achieving RNA mediated virus resistance in citrus, (Abstr. P-3018). J. Soc. In Vitro Biol. 36: Part II.
Pappu H.R., Karasev A.V., Anderson E.J., Pappu S.S., Hilf M.E., Febres V.J., Eckloff R.M.G., McCaffery M., Boyko V., Gowda S., Dolja V.V., Koonin E.V., Gumpf D.J., Cline K., Garnsey S.M., Dawson W.O., Lee R.F. and Niblett C.L. 1994. Nucleotide sequence and organization of eight 3' open reading frames of citrus tristeza closterovirus genome. Virology 199: 35–46.
Satyanarayana T., Gowda S., Mawassi M., Albiach-Marti M.R., Ayllon M.A., Robertson M.A., Garnsey S.M., and Dawson W.O. 2000. Closterovirus encoded HSP70 homolog and p61 in addition to both coat proteins function in efficient virion assembly. Virology 278: 253–265.
Scora R.W. 1988. Biochemistry, taxonomy and evolution of modern cultivated citrus, pp. 277–289. In: Goren R. and Mendel K. (eds), Proc. 6th Int. Citrus Congress. Margraf Scientific Books, Weikersheim, Germany.
Yang Z.N., Ye X.R., Choi S., Molina J., Moonan F., Wing R.A., Roose M.L. and Mirkov T.E. 2001. Construction of a 1.2 Mb contig including the Citrus tristeza virus resistance gene locus using a bacterial artificial chromosome library of Poncirus trifoliata. Genome 44: 382–393.
Yoshida T. 1985. Inheritance of susceptibility to citrus tristeza virus in trifoliate orange. Bull. Fruit tree Res. Stn. Ser. B (Okitsu) 12: 17–25. (in Japanese with English summary)
Yoshida T. 1993. Inheritance of immunity to citrus tristeza virus of trifoliate orange in some citrus intergeneric hybrids. Bull. Fruit tree Res. Stn. Ser B (Okitsu) 25: 33–43. (in Japanese with English summary)
Yoshida T. 1996. Graft compatibility of Citrus with plants in the Aurantiodeaeand their susceptibility to citrus tristeza virus. Plant Dis. 80: 414–414.
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Albiach-Marti, M.R., Grosser, J.W., Gowda, S. et al. Citrus tristeza virus replicates and forms infectious virions in protoplasts of resistant citrus relatives. Molecular Breeding 14, 117–128 (2004). https://doi.org/10.1023/B:MOLB.0000038000.51218.a7
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DOI: https://doi.org/10.1023/B:MOLB.0000038000.51218.a7