Smith D, Coulson G E, Morris G J
J Gen Microbiol. 1986 Jul;132(7):2013-21. doi: 10.1099/00221287-132-7-2013.
The changes in morphology of Penicillium expansum Link and Phytophthora nicotianae Van Breda de Haan during freezing and thawing in a growth medium with and without the cryoprotective additive glycerol were examined with a light microscope fitted with a temperature-controlled stage. Viability of 0.5-1.0 mm diameter colonies of both fungi was determined after equivalent rates of cooling to -196 degrees C in the presence or absence of glycerol. In P. expansum shrinkage occurred in all hyphae at rates of cooling of less than 15 degrees C min-1; at faster rates intracellular ice nucleation occurred. The addition of glycerol increased the rate of cooling at which 50% of the hyphae formed intracellular ice from 18 degrees C min-1 to 55 degrees C min-1. This species was particularly resistant to freezing injury and recovery was greater than 60% at all rates of cooling examined. At rapid rates of cooling recovery occurred in hyphae in which intracellular ice had nucleated. In contrast, during the cooling of Ph. nicotianae in the growth medium, shrinkage occurred and no samples survived on thawing from -196 degrees C. However, on the addition of glycerol, shrinkage during freezing decreased and viable hyphae were recovered upon thawing; at rates of cooling over 10 degrees C min-1 the loss of viability was related to glycerol-induced osmotic shrinkage during cooling rather than to the nucleation of intracellular ice.
使用配备温控载物台的光学显微镜，研究了扩展青霉（Penicillium expansum Link）和烟草疫霉（Phytophthora nicotianae Van Breda de Haan）在含有和不含有冷冻保护添加剂甘油的生长培养基中冷冻和解冻过程中的形态变化。在有或没有甘油存在的情况下，将两种真菌直径为0.5 - 1.0毫米的菌落以相同的冷却速率冷却至-196℃后，测定其活力。在扩展青霉中，当冷却速率低于15℃/分钟时，所有菌丝都会发生收缩；当冷却速率更快时，细胞内会发生冰核形成。添加甘油后，50%的菌丝形成细胞内冰的冷却速率从18℃/分钟提高到了55℃/分钟。该物种对冷冻损伤具有特别的抗性，在所研究的所有冷却速率下，恢复率均大于60%。在快速冷却时，细胞内已形成冰核的菌丝仍能恢复。相比之下，在生长培养基中冷却烟草疫霉时，会发生收缩，从-196℃解冻后没有样品存活。然而，添加甘油后，冷冻期间的收缩减少，解冻后可恢复有活力的菌丝；当冷却速率超过10℃/分钟时，活力的丧失与冷却期间甘油诱导的渗透收缩有关，而不是与细胞内冰的成核有关。
