Microbial Growth Control Methods

• Exercise 8 focuses on controlling microbial growth on inanimate objects.

Controlling Microorganisms

• Controlling microbial populations on inanimate objects is important.
• Removing all microorganisms from an object is sometimes necessary
• Methods for controlling microbial growth depend on the material type, volume, and size.

Sterilization vs. Disinfection

• Sterilization kills all microorganisms, including spores, viruses, and viroids in a material
• Sterility is absolute, meaning an object is either sterile or nonsterile
• Sterilization ensures highly resistant bacterial endospores and fungal spores are killed when done properly
• Disinfection reduces the quantity of pathogenic microorganisms through killing, inhibiting, or removal
• Disinfectants are used on inanimate objects and do not necessarily sterilize an object

Microbial Growth Control Methods:

• Physical techniques include osmotic pressure changes, heat (pasteurization, dry/moist heat sterilization, tyndallisation), and radiation.

Learning Objectives:

• Demonstrate the effect of dry and moist heat on bacterial growth.
• Demonstrate the effect of osmotic pressure on bacterial growth using different salt concentrations.
• Demonstrate the effect of radiation on the pigment production of bacteria.

Materials for Experiments:

• Sterile nutrient agar plates, nutrient broth tubes, wire loop, yeast culture, unsterilized nutrient agar flask, Serratia marcescens broth culture, UV lamp, E. coli, S. aureus, M. luteus broth culture, sodium chloride, and yeast and Rhizopus .

Direct Heat Experiment

• Streak an unflamed wire loop on a sterile agar plate and label it A
• Flame the loop, cool it, then streak on another sterile agar plate and mark it as B
• Incubate the plates for 24-48 hours at ambient temperature.
• Unheated loop shows more bacterial growth than heated loop streaked on a different agar plate

Moist Heat under Pressure

• Sterilize three nutrient agar flasks each containing 50 ml of NA at 15 psi, for 15/10/5 minutes
• An unsterilized nutrient agar flask will serve as the control
• Observe and score growth in each flask using plus (+) signs

Osmotic Pressure Experiment

• Use five nutrient broth tubes with different concentrations of sodium chloride.
• Inoculate each tube with a loopful of bacterial culture to test osmotic pressure effects
• Prepare two more sets of tubes and inoculate them with yeast culture.
• Observe the culture for growth after 2 and 7 days and score the growth rate

Ultraviolet Radiation Experiment

• Obtain a 24-hour broth culture of Serratia marcescens and five nutrient agar plates
• Label the plates A, B, C, D, and E
• Spread 0.01 ml of the culture on each plate, immediately cover plate A with foil and incubate at 30°C for 24-48 hours (your control which has no UV exposure)
• Remove the top dish and expose plates B-E to UV radiation at varying time intervals (10, 30, 60 and 120 seconds) using a UV lamp under laminar hood and/or mini isolation chamber. Position the plates 4 inches from the lamp
• Cover the plates with foil after exposure, incubate as with plate A, and compare to control and record observations
• UV tends to inhibit bacterial growth and gives a pinkish colouration in the colonies compared to red on the irradiated plate