Vocabulary
• Bactericidal: Bacteria-killing
• Bacteriostasis: Bacterial growth inhibiting
• Disinfection: Destruction of vegetative pathogens
• Antisepsis: Destruction of vegetative pathogens on living tissue
• Aseptic: Absence of a pathogen from an object or an area

Vocabulary Cont.
• Sanitation: Reduction of pathogens on eating utensils, etc. to public health standards
• Antiseptic: chemical used for disinfection of skin or mucous membranes
• Disinfectant: Chemical used on inanimate objects to kill or inhibit bacterial growth
• Sterile: Without life, free of microorganisms

Measuring the effectiveness of bacterial control methods
• Not all genera of bacteria are equally susceptible to a given method of control.
– Example:
• E. coli bacteria are highly susceptible to heat and die quickly when they reach boiling temperatures.
• Pathogens that create endospores on the other hand, can survive long periods of boiling.
• How do we determine how hot and how long we need to keep a microbe at to kill it?

Three acceptable ways to measure microbial tolerance to heat

1. Decimal reduction time (DRT): time in which
90% of the population of bacteria at a given temperature will die.
2. Thermal death time (TDT): Time in which all cells in a suspension are killed at a given temperature.
3. Thermal death point (TDP): The lowest temperature in which all microorganisms will be killed in 10 minutes.

Microbes die at a constant, logarithmic rate when treated.

What is the DRT for the above graph?
What is the TDT?

• More effective than dry heat
• Kills by coagulation of proteins
• Three methods: boiling, autoclaving, pasteurization

• 100 C for 10 min: kills vegetative bacteria, most viruses, and fungal spores.
– Endospores and some viruses (hepatitis) can survive for long boiling periods.
• Good for making drinking water and food safe for consumption.
– Unreliable for sterilization of surgical equipment
(endospores survive without stomach acid to kill them). Free flowing unpressurized steam is equivalent to boiling.

• Pressurizing of steam to make it hotter. 1 atm temp = 121 C kills all organisms and their endospores. Most effective and most preferred method of sterilization. Steam must contact all solid surfaces since solids don’t have convection currents like liquids do.

• Mildly heating food products to kill particular spoilage microorganisms or pathogens. 72 C for 15 seconds with refrigeration needed afterwards. Many heat resistant bacteria survive pasteurization but these are unlikely to cause disease.
• Ultra high temperature pasteurization 140 C for 3 seconds sterilizes milk.

• Kills and sterilizes by oxidation effect.
• Direct flaming or incineration
• Hot air sterilization (170 C for 2 hours)

• Filters are made of cellulose or plastic polymers with pores of about .2 micrometers.
Viruses can still get through.
• There are filters with pore size as small as .01 micrometers for viruses but filtration is very slow.
• Filters are important when you can’t heat something up because you’ll destroy what you’re trying to keep.

• Cold is bacteriostatic
• Slow freezing is more harmful as crystals can set up better.
• Deep freezing is quick freezing at temp between -50 and -95 degrees. This is usually done to preserve the specimen.

• Desiccation is the absence of water. Microbes need water in order to grow and reproduce.
However, microbes in a desiccated state can be viable for years. When water is added, they can resume growth.
• Desiccation is bacteriostatic, though the ability to survive varies by species.

• A suspension of microbes is quickly frozen at temp between -54 and -72 C and water is removed by high vacuum. Microbes can be stored for years in this state.

• Different strains of microbes have different levels of susceptibility to different chemotherapeutic agents.
• Susceptibility of a microorganism can change over the course of a treatment.
• Physicians need to know the sensitivities of the pathogen before treatment can be started. Various tests are employed to give
Physicians the knowledge they need.

• The American Official Analytical Chemist’s
Use-dilution method (Use-Dilution Method)
• The Disk-diffusion Method
• Three bacteria, Salmonella, Staphylococcus, and Pseudomonas, are used as standards to examine the effectiveness of a given disinfectant.

• Small stainless steel cylinders are dipped and coated with these bacteria and than allowed to dry.
• The cylinder is than dipped into one of several dilutions of the chemical agent to be tested for 10 minutes, removed, rinsed with water, and placed into a tube of nutrient broth.
• Chemicals that prevent growth at the greatest dilutions are considered to be the most effective.

Disinfecting Agents
Dilution Ratio Agent 1 Agent 2
1:1 -
1:10
1:50
1:500
+
+
+
-
-
-
+ indicates growth
- indicates no growth
Agent 3
-
-
+
+

Disk-diffusion (Filter Paper) Method
• Most common testing method.
• Small disc of filter paper is soaked with chemical and placed on surface of agar plate that has been previously inoculated with the test organism.

• The chemical is concentrated in one area, and as it diffuses out, it is less concentrated. The effectiveness is based on how big of a circle
“kill-zone” or zone of inhibition is around where the chemical is introduced.
• Multiple chemicals can be tested against one bacteria at a time with this method.

• Can kill in three ways:
– Damage of Plasma Membrane
– Inactivate Enzymes
– Denature Proteins

• Phenol: First used by Joseph Lister in an operating room. It proved too toxic for skin and had a terrible odor.
• Phenolic: Adapted version of phenol that is less potent and more useful.
• Kills in all three ways.

• Kills by inactivating enzymes by attaching to the amino acid tyrosine.
• Effective against bacteria, endospore, fungi, and some viruses.
• Iodine is available as tincture solution (in solution with alcohol). Tincture solutions often stain and patients may develop hypersensitivity to it.
• Iodine can come in Iodophor (combined with an organic molecule for slower releasing).

• Chlorine is mostly just a disinfectant. Chlorine turns to hyochlorous acid, its active form, when it’s added to water.
• Kills by deactivating enzymes. The acid oxidizes cellular molecules into inactive forms.

• Alcohols work in two ways: by denaturing proteins and dissolving lipids (plasma membrane).
• Effective against bacteria and fungi but not endospores and non-enveloped viruses.
Alcohol is most effective when it is diluted by water. (ethanol 70%, isopropyl 90%). Nonwatered alcohol doesn’t carry into cells very well.

Don’t drink isopropyl alcohol.
That is all.

• Gold, silver, copper and zinc are germicidal.
Silver nitrate is used in infants’ eyes.
• Selenium and zinc in shampoos control fungi that cause dandruff.
• Heavy metals combing with –SH groups to denature proteins.