Survival of thermophilic fungi in various preservation methods: A comparative study
Introduction
Several fungi are sources of many high-value compounds that are useful to all living beings. Some of the most important products that use fungi in their production are acetic acid, antibiotics, food and beverages, enzymes, fungal secondary metabolites, biocontrol agents, etc. Among the fungi, only a few species have the ability to thrive at temperatures between 45 and 55 °C. Thermophilic fungi have a growth temperature minimum at or above 20 °C and a growth temperature maximum at or above 50 °C [1]. They have the ability to produce biotechnologically important novel metabolites or enzymes because of their thermotolerance [5].
The isolation and purification of fungi are time consuming methods; thus, it is important to maintain the characteristic obtained as not to repeat those procedures once again. The selection of a preservation method for specific fungi such as thermophilic fungi depends on the characteristics of the technique and microorganism, stocking costs, importance of the collection etc. Therefore, a preservation method must be built for the purpose of maintaining fungal culture and stocks, while fungal characteristics, like morphological features, thermal stability, spore production and viability, are kept well preserved.
Fungal preservation is important for experimental ends, either for routine works or comparative studies that emphasize the biology, etiology, epidemiology, and production of biotechnological important substances [12]. The basic preservation techniques can be divided into two categories. The first ones are simple and cheap methods, such as constant sub-culturing, storage under mineral oil, under water, in soil, and in silica gel. The second ones are complex and expensive methods, such as lyophilization and cryopreserved in liquid nitrogen [9,12,15].
It's known that thermophilic fungi have the sensitivity to subminimum temperatures. However, further investigation for the effects of subminimum temperatures and rewarming on thermophilic fungi is needed, it should not necessarily be assumed that mycelial cultures of thermophilic fungi can be stored under refrigeration or at subminimum temperatures without the loss of viability [5]. Isolation of these fungi from nature is difficult as well as it is difficult to store the isolated species for use in later studies or production processes due to their low-temperature resistance. Depend on their highest growth temperature and sensitivity to subminimum temperatures that affect the survival of them, preservation of several thermophilic species are very difficult for long periods.
However, the literature is very limited as to the effects of different methods of preservation on morphology and viability of thermophilic fungi. So, there is no standard method to preserve thermophilic fungi for long-term maintenance. The aim of this work was to determine the best method of thermophilic fungi preservation among seven different preservation techniques. In accordance with this aim, we want to detect the feasibility of different preservation methods on thermophilic fungi in the way of the continued viability, purity of the cultures, and the influence of these conservation methods on the micro and macro morphological characteristics of the cultures. The long-term effects of seven different storage methods on 25 thermophilic fungi were tried to be determined for the first time.
Section snippets
Microorganisms
Twenty five thermophilic fungi previously isolated from hot spring waters in Turkey and identified by molecular techniques that performed using sequencing of the standard gene regions; internal transcribed spacer (ITS) regions and D1/D2 region of the rDNA genes were used in this study. The list of fungal strains is given in Table 1. Firstly, fungal isolates were grown in Emerson's YpSs agar or Sabouraud dextrose agar plates at 45 °C. After adequate growth, fresh cultures are used for further
Results
The thermophilic fungi isolates used for the present study belonged to the divisions of Ascomycota and Zygomycota. It also contains 11 different species (Table 1). Viability tests were performed at different periods for each species by 7 preservation techniques and the results obtained are given in Table 1. After different storage conditions for diverse periods, all viable thermophilic strains preserved their species-specific physiological abilities that recorded at the fungal diagnostic
Discussion
Considering the long-term storage methods of fungi, it is very difficult to talk about one successful method for all fungi. Because when it comes to long-term preservation, big differences occur even between sporulating and non-sporulating cultures. In order to determine a successful preservation method in the long term, a fungal class, family, and even species level research is required. The choice of storage method for fungi depends not only on the suitability of the organism and success of
Conclusions
The present research was particularly designed to compare various techniques and determine the most useful method for thermophilic fungi, because there are no reports regarding the preservation and maintenance of solely these extremophilic fungi. This study was carried out with 25 thermophilic fungi containing 11 different species. Considering that thermophilic fungi are represented by 30 species in the kingdom fungi [8], this is a sample number that will not be underestimated at all. Based on
Funding
This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.
Declaration of competing interest
Authors declare no conflicts of interest.
Acknowledgments
Author thanks Prof. Dr. Merih Kıvanç for her constructive feedback and all kinds of support to preserve the fungal cultures with all seven methods for more than 5 years.
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