Preservation Techniques for Industrially Important Microorganisms

Question: What are the preservation techniques for maintenance of the industrially important microorganisms ?

Answer:

For industrially important microorganisms, maintenance of the desirable strains is very much important. Preservation of these strains ensure optimal long time viability, genetic stability and purity of the microorganism. Several  preservation methods have been developed and they are as follows,

1. Periodic Transfer to Fresh Media: Strains can be maintained by periodically preparing a fresh subculture from previous stock culture. The culture medium, the storage temperature and the time interval at which the transfer undergoes , varies depending upon the species. The temperature and the growth medium should  support a slow growth rather than a rapid one in order to make the time interval between transfers as long as possible.
   Disadvantages:
   * Repeated sub-culturing is time consuming.
   * Cannot prevent the development of mutants. 
2. Refrigeration: pure cultures can be stored at 4°C for a comparatively short period of time (2-3 weeks) either in refrigerator or on cold rooms. Because of thee low temperature, the metabolic activity of the microorganisms are greatly slowed down but not stopped. They continue to consume nutrients and grow slowly.
   Disadvantages:
   * Need subculturing after some time.
   * Cannot be used for long time strain preservation. 
3. Culture Covered With Mineral Oil: Sterile liquid paraffin is poured over the slant of culture and stored upright at room temperature. The layer of paraffin endures anaerobic conditions and prevents dehydration of the medium. This helps the microorganisms of the pure culture to live in a dormant state, and therefore the culture can be preserved for months to years (depending upon the species). When needed we can take some of the growth under he oil with a transfer needle or a micro-pipette and inoculate a fresh medium while preserving the original culture. It is very simple, effective and economical process of  culture preservation.
   Disadvantage:
   * Mutant strains may develop.  
4. Cryopreservation: The cultures are grown in a culture tube. Then 10-30% cryoprotective agent ( glycerol or Dimethyle Sulfoxide ) is added into the tube so that it could protect thee culture from cold shock. The culture is dispensed into ampules, sealed and kept under liquid Nitrogen at -176°C to -196°C . Most species can remain viable under these conditions for 10-30 years without undergoing any change in their characteristics.
   Disadvantage:
   * Very much expensive process.
5. Lyophilization (Freeze drying): Freeze drying is a process in which water and other solvents are removed from a frozen product via sublimation ( i.e. conversion of a frozen liquid directly into a gaseous state without entering the liquid state ).
   
   
       In this method the microbial suspension is put in small ampules and frozen through drying under vacuum, resulting in sublimation of cell water. Then the ampules are sealed in vacuum and stored in dark 4°C. Under these conditions the microbial cells become dehydrated and their metabolic activity stops ; as a result microbes become dormant and retain there viability for years, and remain unchanged.
   Disadvantage:
   * This is an expensive process.
   * Not every kind of microorganism can be lyophilized. 
   
6. Storage Under Glycerol: 20% glycerol (sterile) is poured onto a slant of culture and the cells are resuspended in the glycerol aseptically. Then it can be stored at 4°C for months. When needed, the culture is quickly thawed under running water, a few drops inoculated is a growth medium and the rest is preserved. 
7. Storage in Saline Suspension: At high concentration salts act as bacterio-static. Hence bacterial culture is preserved in 1% salt solution in capped tubes to prevent evaporation and stored at room temperature. 
8. Immobilization of Microbial Cells:  This method involves adsorption of cells to glass beads (2 mm diameter) that may be placed in frozen storage, from which individual beads may be removed when needed without thawing the whole sample. 
9. Drying in Vacuum: The cultures are dried over CaCl2 in a vacuum, then stored in a refrigerator. At such conditions organisms survive for longer period than air dried cultures. 
10. Preservation in Sterile Soil: Fresh soil is taken, sterilized and then mixed with microbes under aseptic condition. Moisture is removed by maintaining reduced pressure over a drying agent. Then the tubes are sealed and stored.