A new method for the preservation of fungus stock cultures by deep-freezing

Kitamoto, Yutaka; Suzuki, Akira; Shimada, Sanae; Yamanaka, Katsuji

doi:10.1007/s102670200021

Cited by 57 publications

(31 citation statements)

References 4 publications

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“…Após esse período, ocorre a sua degeneração natural, que perde a capacidade de formar novos descendentes (Kitamoto et al, 2002). Cita-se como exemplo o fungo C. cladosporioides, que tem como temperatura mínima de crescimento -4ºC, e temperatura ótima, 22ºC.…”

Section: Discussionunclassified

Imobilização de fungos filamentosos com potencial para uso agroindustrial

Elizei

Chalfoun

Botelho³

et al. 2014

Arq. Inst. Biol.

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Cellular immobilization represents an alternativefor the bioprocess conduction, in which the cells are kept in a matrix and can be used over long periods. The objective of this work was to test a new fungi immobilization methodology with reduced cost to evaluate the viability of these fungi when submitted to the new encapsulation method, and to determine the ideal temperature to store the immobilized fungi. The mycelium of the fungi Aspergillus niger, Cladosporium cladosporioides and Penicillium solitum were mixed with 15 g of titrated rice and 3 g of sodium alginate, which was dripped in a 0.25 M calcium chloride solution for the formation of pellets. After drying in an oven at 26ºC, the granules were stored at three temperatures: room, refrigerator and freezer. The platings were carried out every 15 days in culture medium. The evaluations of the colony size and sporulation were carried out 7, 14 and 12 days after plating, for 195 days for A. niger, 225 days for C. cladosporioides, and 210 days for P. solitum. Storage temperature did not affect the mycelial development of A. niger and P. solitum. However, sporulation was reduced for the granules stored in the freezer. The mycelial development of C. cladosporioides was influenced by temperature. The granules conserved at room temperature had lower viability than those stored in the refrigerator and freezer. In the Scanning Electronic Microscopy analysis, it was observed that the immobilization is a safe method in which the fungus mycelium remains inside the granule, facilitating transport, storage and application of micro-organisms. KEYWORDS: cellular immobilization; encapsulation of fungi; sodium alginate; granulated formulation. RESUMO:A imobilização celular representa uma alternativa para a condução de bioprocessos. As células ficam retidas em matrizes e podem ser utilizadas por longos períodos. O objetivo deste trabalho foi testar uma nova metodologia de imobilização de fungos com custo reduzido, avaliar a viabilidade e segurança dos fungos submetidos ao novo método de encapsulamento e determinar a temperatura ideal para armazenar os fungos imobilizados. Os micé-lios dos fungos Aspergillus niger, Cladosporium cladosporioides e Penicillium solitum foram misturados com 15 g de arroz triturado e 3 g de alginato de sódio, que gotejava em uma solução de cloreto de cálcio a 0,25 M para a formação dos grânulos. Após a secagem em estufa a 26ºC, os grânulos foram armazenados em temperaturas ambiente, geladeira e freezer. Os plaqueamentos foram realizados a cada 15 dias em meio de cultura. As avaliações do tamanho das colônias e esporulação foram realizadas 7, 14 e 21 dias após o plaqueamento, durante 195 dias para A. niger, 225 dias para C. cladosporioides e 210 dias para P. solitum. A temperatura de armazenamento não afetou o desenvolvimento micelial de A. niger e P. solitum. Porém, a esporulação foi reduzida para os grânulos armazenados no freezer. O desenvolvimento micelial de C. cladosporioides foi influenciado pela temperatura. Os grânulos conservados ...

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Section: Discussionunclassified

Imobilização de fungos filamentosos com potencial para uso agroindustrial

Elizei

Chalfoun

Botelho³

et al. 2014

Arq. Inst. Biol.

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“…Kitamoto et al (2002) grew cultures over SDM (65% moisture W/W) with 10% glycerol as the cryoprotectant and then rapidly froze them at 285uC and tested viability by inoculating on PDA; several zygomycete cultures remained viable for 20 mo. These and other methods are discussed in detail by Fennell (1960), Jong and Birmingham (2001), and Nakasone et al (2004).…”

Section: Maintenancementioning

confidence: 99%

Methods Used by Dr. R. K. Benjamin, and Other Mycologists, to Isolate Zygomycetes

Benny¹

2008

aliso

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The methods that Dr. Richard K. Benjamin used to isolate Zygomycetes are discussed. These processes involved the following five steps: (1) collection, (2) plating, (3) isolation, (4) culture, and (5) maintenance. Additional methods, materials and modifications used to isolate Zygomycetes are summarized. The author considers the flattening of the aerial hyphae onto the substrate of the faster-and higher-growing Mucorales for several consecutive days to be the critical step in isolating species of Coemansia, Piptocephalis, Syncephalis, and Dimargaritales. The methods used by other scholars to isolate, culture, and study many taxa in Zygomycetes also are discussed.

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“…Cryopreservation at -70°C is more effective than at -20°C but it requires expensive equipment. Kitamoto et al [11] report a long-term cryopreservation protocol for some species of basidiomycetes using ultrafreezers at -80°C. However, Ryan et al [3] reported that no method can be universally applied to all fungi and that while some species are difficult to preserve, others can be preserved by almost any method.…”

Section: Introductionmentioning

confidence: 99%

Cryopreservation at −20 and −70 °C of Pleurotus ostreatus on Grains

et al. 2012

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Alternative substrates for cryopreservation at -20°C have been little explored for basidiomycetes and could bring new possibilities of lower cost cryopreservation. Nevertheless, freezing temperatures between -15 and -60°C are very challenging because they frequently result in cryoinjuries. The objective of this study was to evaluate substrates associated to cryoprotective agents for Pleurotus ostreatus cryopreservation at -20 or -70°C in order to develop alternative techniques for basidiomycete cryopreservation. P. ostreatus was grown on potato dextrose agar or whole grains of oat, wheat, rice or millet and transferred to cryovials with cryoprotective solution with 1 % dimethyl sulfoxide, 5 % glycerol, 10 % saccharose, 4 % glucose, 6 % polyethylene glycol-6000 or 5 % malt extract. The mycelium in the cryovials were cryopreserved at -20 or -70°C and recovered for evaluation of the mycelial growth viability after 1 and 3 years. Both substrates and cryoprotectants affect the viability of the mycelial growth cryopreserved at -20 or -70°C; wheat grains combined with cryoprotectants such as saccharose or glucose are effective for keeping mycelium viable after cryopreservation at -20°C for 1 or 3 years; for cryopreservation at -70°C after 1 or 3 years, any substrate combined with any cryoprotectant is effective for preserving the mycelium viable, except for millet grains with polyethylene glycol after 3 years; semi-permeable cryoprotective agents such as saccharose and glucose are the most effective for cryopreservation at -20 or -70°C for at least 3 years.

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A new method for the preservation of fungus stock cultures by deep-freezing

Cited by 57 publications

References 4 publications

Imobilização de fungos filamentosos com potencial para uso agroindustrial

Imobilização de fungos filamentosos com potencial para uso agroindustrial

Methods Used by Dr. R. K. Benjamin, and Other Mycologists, to Isolate Zygomycetes

Cryopreservation at −20 and −70 °C of Pleurotus ostreatus on Grains

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