Growth Control          

Chapter 4

The Control of Microbial Growth

n    Sepsis refers to microbial contamination.

n    Asepsis is the absence of significant contamination.

n    Aseptic surgery techniques prevent microbial contamination of wounds.

TERMINOLOGY

TERMINOLOGY

n    Biocide/Germicide

–  Kills microorganisms

n    Bacteriostatic

–  Inhibits (not kill) growth of microorganisms

n    Bacterial populations die at a constant logarithmic rate.

Effectiveness of Antimicrobial Treatment

n     Depends on:

–   Number of containing microbes

–   Environment influence (organic matter, pH, temperature, biofilms)

–   Length or time of exposure

–   Microbial characteristics

Actions of Microbial Control Agents

n    Alteration of plasma membrane

–   Loss of permeability barrier

n    Damage to proteins (enzymes)

–   Metabolism and transport disrupted

n    Damage to nucleic acids (DNA)

–   Cannot produce enzymes or make new membrane

n    Cell Wall

–   Lysis

Physical Methods of Microbial Control

n    Heat

–  Thermal death point (TDP): Lowest temperature at which all cells in a culture are killed in 10 minutes

–  Thermal death time (TDT): Time to kill all cells in a culture at a given temperature

Physical Methods of Control:  Heat

n    Bacteriocidal

n    Heat resistance by a population varies

n    Factors that effect sterilization by heat:

–   Time

–   Organism

–   Temperature

–   Concentration

–   Presence of organic material

–   pH

–   Spores or no spores

–   Type of heat

–   Osmotic pressure

Moist Heat

n     Denatures enzymes

–   Coagulation of proteins

n     Boiling (100°C water)

–   Usually 10 minutes for vegetative cells

–   Virus and spore problems

Moist Heat

n      Autoclave

–       Steam under pressure

–       Raising the pressure in the chamber raises the temperature of the steam

–       121ºC at 15 psi

–       Can kill endospores and hepatitis

n      15 minutes       

–       Volume versus time

–       Most popular sterilization technique in laboratory

Moist Heat

n    Pasteurization

–   Reduces spoilage organisms and pathogens

–   Killing food pathogens without affecting flavor

–   Different times for different consistencies (fat content)

–   Phosphatase test

–   Equivalent treatments

n  63°C for 30 min

n  High temperature, short time: 72°C for 15 sec

n  Ultra high temperature: 140°C for <1 sec

Physical Methods of Microbial Control

n    Dry heat sterilization kills by oxidation

–   Flaming

n  Used during aseptic technique

–   Hot-air sterilization

n  Heat not transferred well in air

n  More time than moist heat

–   170 °C air for 2 hours

n  Not used very often in labs

Other Methods of Control

n     Filtration

}  Filter out or removes microbes

}  Sterilize heat sensitive materials; used to make heat sensitive media

}  HEPA removes microbes >0.3 µm

}  Membrane filtration removes microbes >0.22 µm

n     Low temperatures inhibits microbial growth

–    Refrigeration

–    Deep freezing

n     Desiccation

–    Prevents metabolism

n     Osmotic pressure

–    Causes plasmolysis

–    Curing

–    Salt or sugar

Radiation

n     Damages DNA

n     Two types

–    Ionizing (gamma rays, X rays, high energy electron beams)

n  Creates hydroxyl radicals from wateràdamages organic molecules (especially DNA) by stealing electrons

n  Sterilize medical supplies, some pharmaceuticals and food preservation

–    Nonionizing (UV)

n  Thymine dimersàDNA mutations and inhibition of replication

n  Not penetratingàdirect exposure required

n  Vaccine disinfections and germicidal lamps in hospitals

 }  Microwaves kill by heat; not especially antimicrobial

Chemical Control Methods

n     Factors affecting efficiency of chemicals

–   Initial load

–   Environmental:  Heat and presence of organic matter

–   Time

–   Organism

n  Spores versus vegetative cells

n  Mycobacterium and lipid rich cell wall

n  Gram negative organisms and outer membrane

Resistance to Biocides

n    Gram negative

–   Outer membrane with selective porins

n    Mycobacteria

–   Lipid rich cell wall

n    Endospores

–   Thick spore coat protection