Abstract
Long-term maintenance of arbuscular mycorrhizal fungi (AMF) by in vitro or in vivo subcultivation is often expensive and time-consuming and could present the risk of contaminations and possibly morphological, physiological, and genetic variations over time. Recently, in vitro produced AMF isolates belonging to the genus Rhizophagus were successfully cryopreserved at −130 °C following encapsulation-drying. Here, this method was tested on 12 single species cultures belonging to six different genera (i.e., Rhizophagus, Glomus, Claroideoglomus, Septoglomus, Paraglomus, and Gigaspora) produced in vitro or in vivo. Their viability was estimated, after 1 month of cryopreservation at −130 °C, by the percentage of potentially infective beads (i.e., the percentage of beads that contained at least one germinated propagule) for the in vitro produced species and the percentage of infective beads (i.e., the percentage of beads that contained at least one propagule able to colonize a new host plant in pot culture) for the in vivo produced species. With the exception of Gigaspora sp. MUCL 52331 and Septoglomus constrictus PER 7.2, no significant differences were observed in the viability of the single species cultures before and after cryopreservation. These results, thus, demonstrated the suitability of the cryopreservation method by encapsulation-drying for AMF species belonging to different genera and produced in vitro or in vivo. This method opens the door to the long-term preservation at ultra-low temperature of a large number of AMF species and for the preservation of species that are still recalcitrant to in vitro cultivation.
References
Dalpé Y, de Souza FA, Declerck S (2005) The monoxenic culture of arbuscular mycorrhizal fungi as a tool for systematics and biodiversity. In: Declerck S, Strullu DG, Fortin JA (eds) In vitro culture of mycorrhizas. Springer, Berlin, pp 31–48
de Souza FA, Declerck S (2003) Mycelium development and architecture, and spore production of Scutellospora reticulata in monoxenic culture with Ri T-DNA transformed carrot roots. Mycologia 95:1004–1012
Declerck S, Strullu DG, Plenchette C, Guillemette T (1996) Entrapment of in vitro produced spores of Glomus versiforme in alginate beads: in vitro and in vivo inoculum potentials. J Biotechnol 48:51–57
Declerck S, Strullu DG, Plenchette C (1998) Monoxenic culture of the intraradical forms of Glomus sp. isolated from a tropical ecosystem: a proposed methodology for germplasm collection. Mycologia 90:579–585
Declerck S, Van Coppenolle MG (2000) Cryopreservation of entrapped monoxenically produced spores of an arbuscular mycorrhizal fungus. New Phytol 148:169–176
Declerck S, D’Or D, Cranenbrouck S, Le Boulengé E (2001) Modelling the sporulation dynamics of arbuscular mycorrhizal fungi in monoxenic culture. Mycorrhiza 11:225–230
Declerck S, D’Or D, Bivort C, de Souza FA (2004) Development of extraradical mycelium of Scutellospora reticulata under root organ culture: spore production and function of auxiliary cells. Mycol Res 108:84–92
Douds DD, Schenck NC (1990) Cryopreservation of spores of vesicular arbuscular mycorrhizal fungi. New Phytol 115:667–674
Karandashov VE, Kuzourina IN, George E, Marschner H (1999) Monoxenic culture of arbuscular fungi and plant hairy roots. Russ J Plant Physiol 46:87–92
Krüger M, Kruger C, Walker C, Stockinger H, Schüßler A (2012) Phylogenetic reference data for systematics and phylotaxonomy of arbuscular mycorrhizal fungi from phylum to species level. New Phytol 193:970–984
Kuszala C, Gianinazzi S, Gianinazzi-Pearson V (2001) Storage conditions for the long-term survival of AM fungal propagules in wet sieved soil fractions. Symbiosis 30:287–299
Lalaymia I, Cranenbrouck S, Draye X, Declerck S (2012) Preservation at ultra-low temperature of in vitro cultured arbuscular mycorrhizal fungi via encapsulation-drying. Fungal Biol 116:1032–1041
Lalaymia I, Declerck S, Cranenbrouck S (2013) Cryopreservation of in vitro-produced Rhizophagus species has minor effects on their morphology, physiology, and genetic stability. Mycorrhiza. doi:10.1007/s00572-013-0506-y
Plenchette C, Declerck S, Diop TA, Strullu DG (1996) Infectivity of monoxenic subcultures of the arbuscular mycorrhizal fungus Glomus versiforme associated with Ri T-DNA transformed carrot root. Appl Microbiol Biotechnol 46:545–548
Schüßler A, Walker C (2010) The Glomeromycota. A species list with new families and new genera. Published in Libraries at the Royal Botanic Garden Edinburgh, the Royal Botanic Garden Kew, Botanische Staatssammlung Munich, and Oregon State University.
Walker C (2005) A simple blue staining technique for arbuscular mycorrhizal and other root-inhabiting fungi. Inoculum 56:68–69
Young JPW (2012) A molecular guide to the taxonomy of arbuscular mycorrhizal fungi. New Phytol 193:823–826
Acknowledgments
This work was supported by the European Community’s Seventh Framework Program FP7/2007–2013 under grant agreement no. 227522, entitled “Valorizing Andean microbial diversity through sustainable intensification of potato-based farming systems.” The authors wish to thank Valentine Potten, Ph.D., student at the Université catholique de Louvain (Belgium), for the isolation of the in vivo produced AMF species and Dr. Christopher Walker from the Royal Botanic Garden Edinburgh (UK) and Carolina Senés from the University of Munich (Germany) for the taxonomic and molecular identification, respectively, of the in vivo produced species.
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Lalaymia, I., Declerck, S., Naveau, F. et al. Cryopreservation of arbuscular mycorrhizal fungi from root organ and plant cultures. Mycorrhiza 24, 233–237 (2014). https://doi.org/10.1007/s00572-013-0525-8
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DOI: https://doi.org/10.1007/s00572-013-0525-8