VMC 211: General Veterinary Microbiology (1+1)

MOIST HEAT

• Moist heat kills microorganisms primarily by coagulation of protein caused by breakage of hydrogen bonds that hold the proteins in three dimensional structure.
• It is used for sterilization of media and other liquids required to retain their content of water.
• It is not applicable to water proof materials like oils and dry materials in sealed containers.

Boiling at 100^oC

• Boiling at 100C for 10 minutes is sufficient to kill all vegetative bacteria and some spores.
• All spores are not killed. Metal, glass and rubber items are held on a removable tray are immersed in boiling water in a metal bath.
• After boiling for ten minutes the articles are removed using a long sterile forceps and held to dry it by evaporation.

Steaming at 100ºC

• The articles are exposed to steam. The temperature of the steam is 100C.
• Exposure at this temperature for 60 minutes kills all organisms except slow viruses and spores of most resistant mesophilic bacteria and thermophilic bacteria. This method is more convenient for bottles, flasks and tubes of culture media and fluids as they may crack if heated directly to boil their contents.

Tyndallization

• The media are exposed to steam for a period of 20-45 minutes on three successive days. The spores that survive the initial steaming will germinate when held at room temperature.
• The resulting vegetative bacteria will be killed when heated again. This process will not kill the bacteria whose spores cannot germinate in the particular medium or under the conditions of storage in between treatment. So the method cannot be used for non nutrient media. This method was used in the past for sterilization of media containing heat labile sugars. Now for this purpose filtration is used.

Autoclaving (steam under pressure)

• Steam under pressure is one of the efficient methods for sterilization. When the pressure of the steam is increased to 15 lbs/sq.inch the temperature of the steam is 121C. At this temperature both the vegetative cells and spores are killed.
• The standard for efficient sterilization is 121C for 15 minutes at 15 lbs/sq.inch. the equipment used for this purpose is called Autoclave. An autoclave has two chambers, a sterilization chamber and steam generation chamber. In the steam generation chamber water is heated continuously and steam is produced.
• The materials to be sterilized are placed in the sterilization chamber. Bacteriological media, clothings , heat resistant plastics and rubber items are sterilized in autoclave.
• The glasswares kept in the autoclave should be made of Corning or Pyrex or Borosil glass.
• The mouth of the flask containing the medium should be plugged to prevent escape of medium during heating.
• When petridishes are sterilized the base and lid have to be separated.
• When heat resistant plastics are sterilized they should be placed sideways for easy entry of steam.