Abstract
Multiplication of certified pathogen-free stock plants in vitro makes an important contribution to the production of disease-free planting material for vegetatively propagated crops. Meristem culture is extensively used to eliminate pathogens and contaminants from microbially contaminated plants prior to micropropagation. The approach to pathogen and contamination management differs. It is essential to avoid the release of pathogen-contaminated microplants and to ensure this the plant pathogen-testing guidelines and protocols issued by the Food and Agriculture Organisation (FAO) and its regional representative organizations should be followed. Where in vitro methods are used to eliminate pathogens, the progeny plants should be established in vivo under quarantine conditions and tested under FAO guidelines before being used as stock plants for in vitro multiplication. At establishment of microplants in vitro (stage 1), cultures should be culture-indexed for the presence of microbial contaminants. If pathogenand contaminant-free cultures are established, then the risk is that of managing laboratory contamination by common environmental microorganisms based on Hazard Analysis Critical Control Points (HACCP) principles.
International plant health certification organizations are conservative and rely on established pathogen indexing protocols. They are reluctant to accept DNA-based tests and do not accept testing of in vitro cultures. Given that in time both of these restrictions may be relaxed, micropropagators may look forward to availing more of diagnostic service providers using polymerase chain reaction-based multiplex assays for pathogen-indexing and advances in diagnostic kits for environmental microorganisms in support of laboratory contamination management; with the caveat that molecular tests for pathogens may continue to require confirmation by inoculation of indicator plant species.
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References
Cassells, A. C. and Tahmatsidou, V. (1996) The influence of local plant growth conditions on non-fastidious bacterial contamination of meristem-tips of Hydrangea cultured in vitro. Plant Cell Tiss. Org. Cult. 47, 15–26.
Leifert, C., Morris, C., and Waites W. M. (1994) Ecology of microbial saprophytes and pathogens in tissue cultured and field grown plants. CRC Crit. Rev. Plant Sci. 13, 139–183.
Leifert, C., Nicholas, J. R., and Waites, W. M. (1990) Yeast contaminants of micropropagated plant cultures. J. Appl. Bacteriol. 69, 471–476.
Pype, J., Everaert, K., and Debergh, P. C. (1997) Contamination by microarthropods, in Pathogen and Microbial Contamination Management in Micropropagation (Cassells, A. C., ed.), Kluwer Academic Publishers, Dordrecht, pp. 259–266.
Krczal, G. (1998) Virus certification of ornamental plants—the European strategy, in Plant Virus Disease Control APS Press, (Hadidi, A., Khetarpal, R. K., and Koganezawa, H., eds.), St. Paul, pp. 277–287.
Leifert, C. and Cassells, A. C. (2001) Microbial hazards in plant tissue and cell cultures. In Vitro Cell Dev. Biol. 37, 133–138.
Lelliott, R. A. and Stead, D. E. (1987) Methods for the Diagnosis of Bacterial Diseases of Plants, Blackwell Scientific Publications, Oxford.
Schaad, N. W., Jones, J. B., and Chun, W. (2001) Laboratory Guide for Identification of Plant Pathogenic Bacteria, APS Press, St. Louis.
Agrios, G.N. (1997) Plant Pathology, Academic Press, London.
Larone, D. H. (1987) Medically Important Fungi. A Guide to Identification, Elsevier Publishers, New York.
Krantz, G. W. (1978) A Manual of Acarolgy, Oregon State University, Corvallis.
Hull, R. (2001) Matthews Plant Virology, Academic Press, New York.
Hari, V. and Das, P. (1998). Ultra microscopic detection of plant viruses and their gene products, in Plant Virus Disease Control (Hadidi, A., Khetarpal, R. K., and Koganezawa, H., eds.), APS Press, St. Paul.
Cassells, A. C. (1991) Problems in tissue culture: culture contamination, in Micropropagation: Technology and Applications (Debergh, P. C. and Zimmerman, R. H. eds.). Kluwer Dordrecht, pp. 31–44.
Stead, D. E., Elphinstone, J. G., Weller, S., Smith, N., and Hennessy, J. (2000) Modern methods for characterizing, identifying and detecting bacteria associated with plants. Acta Hort. 530, 45–60.
O’Herlihy, E. A. and Cassells, A. C. (2003) Influence of in vitro factors on titre and elimination of model fruit tree viruses. Plant Cell Tiss. Org. Cult. 72, 33–42.
Dijkstra, J. and de Jager, C. P. (1998) Practical Plant Virology: Protocols and Exercises, Springer Verlag, Berlin.
Lee, B. H. and Nagamune, T. (2004) Protein microarrays and their application. Biotechnol. Bioprocess. Eng. 9, 69–75.
Schaad, N. W. and Frederick, R.D (2002) Real-time PCR and its application for rapid plant disease diagnostics. Can. J. Plant Pathol. 24, 250–258.
Mishra, P. K., Fox, R. T. V., and Culham, A. (2003) Development of a PCR-based assay for rapid and reliable identification of pathogenic Fusaria. FEMS Microbiol. Letts. 218, 329–332.
Singh, K. K., Mathew, R., Masih, I. E., and Paul, B. (2003) ITS region of the rDNA of Pythium rhizosaccharum sp. Nov. isolated from sugarcane roots: taxonomy and comparison with related species. FEMS Microbiol. Letts. 221, 233–236.
Klerks, M. M., Leone, G. O. M., Verbeek, M., van den Heuvel, J. F., and Schoen, C. D. (2001) Development of a multiplex AmpliDet RNA for the simultaneous detection of Potato Leafroll Virus and Potato Virus Y in potato tubers. J. Virol. Meth. 93, 115–125.
Bailey, A. M., Mitchell, D. J., Manjunath, K. L., Nolasco, G., and Niblett, C. L. (2002) Identification to the species level of the plant pathogens Phytophthora and Pythium by using unique sequences of the ITS1 region of ribosomal DNA as capture probes for PCR ELISA. FEMS Microbiol. Letts. 207, 153–158.
Gosalvez, B., Navarro, J. A., Lorca, A., Botella, F., Sanchez-Pina, M. A., and Pallas, V. (2003). Detection of melon necrotic spot virus in water samples and melon plants by molecular methods. J. Virol. Meth. 113, 87–93.
Rigotti, S., Gindro, K., Richter, H., and Viret, O. (2002). Characterization of molecular markers for specific and sensitive detection of Botrytis cinerea Pers.: Fr. in Strawberry (Fragaria x ananassa Duch.) using PCR. FEMS Microbiol. Letts. 209, 169–174.
Filion, M., St. Arnaud, M., and Jabaji-Hare, S. H. (2003) Direct quantification of fungal DNA from soil substrate using real-time PCR. J. Microbiol. Meth. 53, 67–76.
Langrell, S.R.H. and Barbara, D.J. (2001) Magnetic capture hybridisation for improved PCR detection of Nectria galligena from lignified apple extracts. Plant Molec. Biol. Rep. 19, 5–11.
Ozakman, M. and Schaad, N. W. (2003) A real-time BIO-PCR assay for the detection of Ralstonia solanacearum race 3, biovar 2, in asymptomatic potato tubers. Can. J. Plant Pathol. 25, 232–239.
Watanabe, K., Kodama, Y., and Harayama, S. (2001) Design and evaluation of PCR primers to amplify bacterial 16S ribosomal DNA fragments used for community fingerprinting. J. Microbiol. Meth. 44, 253–262.
Lay, J. O. (2001) MALDI-TOF mass spectrometry of bacteria. Mass Spectrom. Revs. 20, 172–194.
Wang, Z. P., Dunlop, K., Long, S. R., and Li, L. (2002) Mass spectrometric methods for generation of protein mass database used for bacterial identification. Anal. Chem. 74, 3174–3182.
Nedelkov, D., Rasooly, A., and Nelson, R. W. (2000) Multitoxin buiosensor-mass spectrometry analysis: a new approach for rapid, real-time, sensitive analysis of staphylococcal toxins in food. Int. J. Food Microbiol. 60, 1–13.
Wenning, M., Seiler, H., and Scherer, S. (2002) Fourier-transformed infrared microscopy, a novel and rapid toot for identification of yeasts. Appl. Environ. Microbiol. 68, 4717–4721.
Bull, A. T., Goodfellow, M., and Slater, J. H. (1992) Biodiversity as a source of innovation in biotechnology. Ann. Rev. Microbiol. 46, 219–252.
George, E. F. (1996) Plant Propagation by Tissue Culture, Exegetics, Basingstoke.
Cassells, A. C. (1997) Pathogen and Microbial Contamination Management in Micropropagation, Kluwer Academic Publishers, Dordrecht.
Menard, D., Coumans, M., and Gaspar, T. H. (1985) Micropropagation du Pelargonium a partir de meristems. Meded. Fac. Landbouwett. Rijksuniv. Gent. 50, 327–331.
Barrett, C. and Cassells, A. C. (1994) An evaluation of antibiotics for the elimination of Xanthomonas campestris pv. Pelargonii (Brown) from Pelargonium x domesticum cv. Grand Slam explants in vitro. Plant Cell Tiss. Org. Cult. 36, 169–175.
Hoffman, P. N., Death, J. E., and Coates, D. (1981) The stability of sodium hypochlorite solutions, in Disinfectants: Their Use and Evaluation of Effectiveness (Collins, C. H., Allwood, M. C., Bloomfield, S. J. and Fox, A., eds.), Academic Press, London, pp. 77–83.
Gregory, P. H. (1973) The Microbiology of Atmosphere, Leonard Hill Books, Aylesbury.
Weller, R. and Leifert, C. (1996) Transmission of Trichophyton interdigitale via an intermediate plant host. Brit. J. Dermatol. 135, 656–657.
Saar, D. E., Polans, N. O., Sorensen, P. D., and Duvall, M. R. (2001) Angiosperm DNA contamination by endophytic fungi: detection and methods of avoidance. Plant Mol. Biol. Rep., 19, 249–260.
Barnett, H. L. and Hunter, B. B. (1998) Illustrated Genera of Imperfect Fungi, 4th ed., APS Press, St. Louis.
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Cassells, A.C., Doyle, B.M. (2006). Pathogen and Biological Contamination Management. In: Loyola-Vargas, V.M., Vázquez-Flota, F. (eds) Plant Cell Culture Protocols. Methods in Molecular Biology™, vol 318. Humana Press. https://doi.org/10.1385/1-59259-959-1:035
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DOI: https://doi.org/10.1385/1-59259-959-1:035
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