Control of Microbial Growth

A few terms

• Bacteriostatic: inhibits bacterial growth• Bactericidal: something capable of killing

bacteria• Antiseptic: an agent that is used to inhibit/kill

bacterial growth on skin and mucus membranes• Disinfectant: an agent that is used to inhibit/kill

bacterial growth on inanimate objects

What factors influence the success of microbial control?

• What type of organism(s) are you targeting?

• What type of environment are you treating?

Are all microbes equally sensitive?

What parts of a bacterial cell are sensitive to physical treatments

and chemicals?

• Plasma membrane

• DNA and proteins

**

Physical Methods

• Heat– Dry: kills by oxidation, ex: incineration– Moist: promotes coagulation of proteins

• Boiling (100oC)• Autoclave(121oC, 15lbs/sq inch)• Pasteurization

– Classic-63oC for 30 minutes– HTST-72oC for 15 seconds– UHT-140oC for 3 seconds

Autoclave

Autoclaves work due to steam under pressure

**Indicators used in autoclaving**

What does a color change on the autoclave tape indicate? Why would it be necessary to also place an indicator in the center of the pack? What type of biological indicator is used for quality assurance in an autoclave monitoring program? Why? Where would the biological indicator be placed during the autoclaving procedure?

What if the substance is heat sensitive?

• Filtration is the best choice

• Pore sizes can be either .45um or .22um

Physical Methods

• Low temperature: freezing does not kill bacteria, most cultures are stored at -80oC

• Dessication: remove the water and bacteria can remain viable

Physical Methods

• Radiation– Ionizing radiation (gamma rays, x-rays)– Non-ionizing radiation (uv rays)– Microwaves

Electromagnetic Spectrum

Chemical Methods

• Most are only able to reduce the numbers of organisms, not achieve sterility

• Types of chemicals– Phenol and phenolics– Halogens– Alcohols– Heavy metals– Soaps– Quaternary Ammonium compounds– Biguanides

As a general rule: if you need to achieve sterility then you should be thinking about physical, not chemical methods of microbial control.

Evaluation of chemicals: disc diffusion method

Chemical control: choosing the right germicidal chemical

• Goal: sterility vs. disinfection; level of disinfection required dictates potency of chemical required

• Toxicity: risk benefit analysis• Activity in presence of organic material: most are

diminished or inactivated• Sensitivity of the material to be treated• Residue: toxic or corrosive vs residual desired

antimicrobial effect• Cost and availability• Storage and stability: concentrate vs stock solution• Environmental risk: antimicrobials in the environment

Chemical control

• PhenolicsDestroy cytoplasmic membranes, denature proteins (binds

to tyrosine, alters cell membranes)• Triclosan• Hexachlorophene

Target: vegetative bacteria +/- mycobacteria; not sporocidal; inadequate if viricidal activity is required; leave an active antimicrobial residue

Chemical control

• Halogens1. Iodine (binds to tyrosine, alters cell membranes)

• Tincture• Iodophore

Target: all bacteria, fungi, most endospores, and some viruses

2. Chlorine (strong oxidizing agent)

Target: all types of microbes and viruses; inactivated by organic matter

Chemical Control• Alcohols

– Mechanism of action is protein denaturation, can also dissolve lipids

– Most common are ethanol and isopropanolTarget: kill vegetative cells and fungi, do not kill endospores and

some naked viruses

Chemical control

• Heavy metals exert “oligodynamic action” = ability of very low concentrations of metals to kill microbes: interfere with function of enzymes by binding to –SH

• Silver (and formerly, copper, mercury, lead, arsenic)

Chemical control

• Surfactants (soaps)– Little value as an antiseptic– Acid-anionic surface-active sanitizers are

important in cleaning of dairy equipment and utensils

Chemical control

• Quaternary Ammonium Compounds (Quats)– Group of positively charged detergentsAction: react with the cell membrane → reduced surface

tension, membrane damage

Target: kill many vegetative bacteria and enveloped viruses, fungi; do NOT kill endospores, Mycobacteria, or naked viruses

Chemical control

– Biguanides (chlorhexidine)• Used in antiseptic products• Adheres to skin and mucus membranes, low toxicity• Action: membrane disruption• Target: wide range of bacteria, fungi, and some

enveloped viruses; does not kill endospores; will not destroy naked viruses

Which antiseptic works the best?