Long-term cryopreservation of basidiomycetes

Abstract

Basidiomycetes have several biotechnological and industrial applications such as enzyme production, bioremediation, pharmaceutical and functional food production. Due to climatic features, the preservation of several basidiomycetes is threatened, and to guarantee the preservation of this genetic resource, the development of long-term preservation techniques is necessary once there is no universal protocol for the cryopreservation of basidiomycetes. Cryopreservation is a technique in which microorganisms are submitted to ultralow temperatures. Therefore, this study aimed to collect information on the main conditions for long-term cryopreservation of basidiomycetes in the last 20 years. Scientific articles on cryopreservation of basidiomycetes published from 1997 to 2016, were researched, and only the studies on two intervals of cryopreservation were considered: from 1 to 2 years and for longer than 2 years. The analyzed conditions of basidiomycete cryopreservation were: most studied genera, cryopreservation temperature, substrate, cryoprotectant (and preservation substrate), cryopreservation period, thawing temperature and cultivation medium after thawing, physiological and genetic stability of basidiomycetes after thawing in cryopreservation. In this review, the viability of the main cryopreservation conditions of basidiomycetes studied in the last 20 years are presented and discussed.

Semisolid culture medium improves mycelial recovery of Agaricus subrufescens cryopreserved in cereal grains

“…cereal grains in mycelial viability cryopreservation, probably because grains provide an internal capillary net, besides protectant substances, which works as a physical protection to mycelia, confirming the reports of Tanaka et al [17] and Mata and Pérez-Merlo [26]. Another aspect is that glycerol addition to the substrate in order to obtain a Bpreservation substrate^ [22] was not effective to keep the mycelial viability in all treatments after 5-year cryopreservation. The association of the semisolid culture medium was effective to recover the mycelial viability only when wheat or rye grains were used for mycelial growth, without glycerol addition to the substrate, and with glycerol addition to the cryotube.…”

“…For the 5-year cryopreservation at − 70°C of A. subrufescens, the fungal growth in whole grains of hard endosperm wheat or rye is recommended, even though MEA and potato dextrose agar disks were broadly used in the basidiomycete cryopreservation [22]. Colauto et al [12] reported effective mycelial viability recovery of A. subrufescens grown in whole grains of rice with hulls and cryopreserved at − 86°C for 1 year with 10% glycerol.…”

“…Most basidiomycete cryopreservation studies evaluate the cryopreservation effect of temperature [12][13][14], substrates for fungal growth before cryopreservation [15][16][17], class and concentration of cryoprotectants [18,19], and maintenance period [18,[20][21][22]. However, few studies assess the effect of cereal grains for cryopreservation and the consistency of recovery culture medium on the mycelial viability after cryopreservation.…”

“…There are many studies about the initial and intermediate stages of the cryopreservation process but few studies on the recovery culture media after thawing [22]. Recovery cultivation media are usually the same ones used for fungal growth before freezing.…”

“…Recovery cultivation media are usually the same ones used for fungal growth before freezing. The most used ones are potato dextrose agar (PDA) and malt extract agar (MEA) [22], but they are not always the best choice. Fungal cryopreservation is a very stressful freezing process and our hypothesis is that the increase in free water using semisolid media after thawing could improve mycelial recovery viability.…”