Preservation of Thermophilic Fungi: Laboratory Protocols for Short- and Long-Term Storage.

Fungal preservation is critical for safeguarding genetic stability and viability for research and industrial applications. Methods are generally categorized into short- and long-term storage. Short-term maintenance relies on constant sub-culturing, which is technologically simple but labor-intensive and carries risks of phenotypic changes, mutations, or contamination. Alternative intermediate methods include storage under mineral oil, distilled water stasis (Castellani method), and silica gel. For permanent preservation, lyophilization (freeze-drying) and cryopreservation at ultra-low temperatures (liquid nitrogen or -80°C freezers) are preferred as they effectively halt cellular metabolism. Thermophilic fungi present a unique challenge due to their extreme sensitivity to subminimum temperatures below 20°C. Recent research indicates that the freezing block method at -20°C and lyophilization are the most effective strategies for these isolates, maintaining viability for up to 5 years. The use of cryoprotectants, such as glycerol, skim milk, or trehalose, is essential in these protocols to minimize cellular damage caused by ice crystal formation. Ultimately, since no universal protocol exists, the choice of method must be tailored to the specific fungal species and available laboratory resources.