Second partCONTROLLING MICROBIAL

GROWTH IN THE ENVIRONMENT

Topic Outline

1. Basic Principles of Microbial Control

2. The Selections of Microbial Control Methods

3. Physical Methods of Microbial Control

4. Chemical Methods of Microbial Control

Basic Principles of Microbial Control1. Terminology of Microbial Control

• Sterilization – The removal or destruction of all microbes, including viruses and bacterial endospores, in or on an object.

• Aseptic – An environment or procedure that is free of contamination by pathogens.

• Disinfection – The use of physical or chemical agents known as disinfectants.

• Degerming – The removal of microbes from a surface by scrubbing.

• Sanitization – The process of disinfecting places and utensils used by the public to reduce the number of pathogenic microbes to meet accepted public health standards.

• Pasteurization – The use of heat to kill pathogens and reduce the number of spoilage micoorganisms in food and beverages.

2. Microbial Death Rates

• Definition – The permanent loss of reproductive ability under ideal environmental conditions.

• Technique for evaluating the efficacy of an antimicrobial agent.

3. Action of Antimicrobial Agents• Modes of action fall into two basic catagories.

i. Alteration of cell walls (fungi) and membranes (virus).

- Give damage to a membrane’s proteins or phospholipids and therefore allows the cellular contents to leak out and causes death.

- Give damage to viral envelope that interrupts the reproduction.

ii. Damage to proteins and nucleic acids.

- By breaking hydrogen and disulfide bonds in three dimensional shape and resulting in proteins denaturation. Proteins cease to function.

- Genes of a cell or virus are composed of nucleid acids. Disruption of nucleic acid can produce fatal mutation.

The Selection of Microbial Control Methods1. Factors Affecting the Efficacy of Antimicrobial Methods

i. Site to be treated

- Harsh chemicals and extreme heat cannot be used on human, animals and fragile objects.

- To sterilized the utensils to be used on the body to prevent infections.

ii. Relative susceptibility of microorganisms- Often to select a method to kill the hardiest

microorganisms present, assuming that method will kill more fragile microbes as well.

- Germicides can be classified as high, intermediate or low depending on their effectiveness on inactivating or destroying microorganisms.

- a. High-level kill all pathogens, including endospores.

b. Intermediate-level kill fungal spores, protozoan cysts, viruses and pathogenic bacteria.

c. Low-level kill vegetative bacteria, fungi, protozoa and some viruses.

iii. Environmental conditions- Temperature: warm disinfectants work

better than cool ones. - pH: some disinfectants more effective at

low pH.- To clean objects before sterilization.

2. Methods for Evaluating Disinfectants and Antiseptics

i. Phenol coefficient- The first method used.- If >1.0 ; the agent is more effective than

phenol.- The larger the ratio, the greater the

effectiveness.ii. Use-Dilution Test- The current standard test- The most effective agent is the one that

entirely prevents microbial growth at the highest dilution.

iii. In-use test

- A more realistic method.

- Swabs are taken from actual objects before and after application of disinfectant.

- More accurate determination of a given disinfection agent for each specific situation.

PHYSICAL METHODS OF MIROBIAL CONTROL

1. Heat Related Methods- High temperatures denature proteins, interfere

with the integrity of cytoplasmic membranes and cell walls and disrupt the function and structure of nucleic acids.

- Thermal death point: lowest temperature that that kills all cells in a broth in 10 minutes.

- Thermal death time: the time it takes to completely sterilize a particular volume of liquid at a set temperature.

- Decimal reduction time (D): time required to destroy 90% of the microbes in a sample.

i. Moist heat- To disinfect, sanitize, sterilize and pasteurize

by denaturing proteins and destroying cytoplasmic membranes.

- More effective than dry heat because water is better conductor of heat than air.

- Methods: Boiling, Autoclaving, Pasteurization and Ultrahigh-Temperature Sterilization.

ii. Dry heat- Denatures proteins and fosters the oxidation of

metabolic and structural chemicals.- For substances cannot be sterilized by boiling

or steam or materials can be damaged by repeated exposure to steam.

• endospores are very heat resistance.They are to survive heat that would rapidly kill vegetative cells of the same species.

• a major factor in heat resistance is the amount and state of water within the endospores.

• during endospores formation, the protoplasma is reduced to a minimum volume as a result of the accumulation of Ca2+-dipicolinic acid complexes.

• also the protection of spore DNA by small acid-soluble proteins (SASP).

• this mixture forms a gel in the cytoplasm.• a thick cortex then forms around the

protoplast core.• contraction of the cortex results in a

shrunken, dehydrated protoplast with a water content of only 10-30% of a vegetative cells.

• high concentration of SASPs and low water content will give high heat resistance.

- A routine part of standard aseptic in microbiology laboratory procedure .

- Requires higher temperatures for longer times than moist heat.

2. Refrigeration and Freezing- Tempt. is between 0oC and 7oC for

refrigeration and below 0oC for freezing.- Will decrease microbial metabolism,

growth and reproduction.- Because slow in chemical reactions and

unavailable of liquid.

3. Desiccation And Lyophilization- Dessication or drying inhibits microbial

growth because the absent of water.- Lyophilization- technique combining

freezing and drying to preserve microbes and cells.

4. Filtration- The passage of a fluid (liquid or gas)

through a sieve to trap and separate particles (cells or viruses) from the fluid.

- To sterilize heat-sensitive materials (antibiotics, vaccines, enzymes etc.).

5. Osmotic Pressure- Cells in hypertonic solution (concentrated

salt or sugar) will lose water and therefore inhibits cellular metabolism.

6. Radiation.- Particulate radiation : consists high-

speed subatomic particles that have been freed from their atoms.

- Electromagnetic radiation: energy without mass traveling in waves in the speed of lights.

i. Ionizing Radiation

- Wavelengths shorter than 1 nm.

- Electron beams, gamma rays, and X rays.

ii. Nonionizing Radiation

- Wavelength greater than 1 nm.

- UV light, visible light, infrared radiation and radio waves.

CHEMICAL METHODS OF MICROBIAL CONTROL

i. Phenol and phenolics.- Phenolics: compounds derived from phenol

molecules that have been chemically modified by the addition of halogens or organic functional groups.

- Commonly used in health care settings, laboratories and households.

- -ve: disagreeable odor and possible side effect (skin irritation, brain damage in infants).

ii. Alcohols

- intermediate-level disinfectants.

- Commonly used are isopropanol and ethanol.

- Denature proteins and disrupt cytoplasmic membranes.

- Pure alcohol is not an effective as 70% and 90% because no water.

• in microbial control, alcohols acts on proteins and disrupt cytoplasmic membranes.

• however, water is needed for the coagulation reactions of proteins.

• this also called denaturation of proteins. • 100% ethanol contained no water to carry on the

coagulation process and therefore not effective in microbial control.

• moreover, 70% alcohol-water mixture penetrates more deeply than pure alcohol into most materials to be disinfected.

iii. Halogens (I-, Cl-, Br-, Fr-)

- Used both alone and combined with other elements in organic and inorganic compounds

- Iodine: well-known antiseptic for water.

- Iodophor (iodine-containing organic compound): used in medical institution.

- Chlorine: treat drinking water, swimming pools and waste water.

- Sodium/calcium hypochlorite: disinfectant.

iv. Oxidizing Agents- peroxides, ozone and peracetic acid.- High-level disinfectants and antiseptics.- Used by health care workers to kill anaerobes in

deep puncture wounds.

v. Surfactants- To reduce the surface tension of solvents

(water) by decreasing the attraction among molecules that the solvent becomes more effective at dissolving solute molecules (exp. Oils) and any bateria they harbor – are more easily wash away.

vi. Heavy metals- Such as arsenic, zinc, mercury, silver and

copper.- Can combine with sulfur atoms in molecules of

cystein, an amino acid.- Low-level bacteriostatic and fungistatic agents.

vii. Aldehydes- Compounds containing terminal –CHO groups.- Usually used by hospital personnel for

disinfecting medical and dental equipments.- Formadehyde irritates mucous membranes and

is carcinogenic.

viii. Gaseous Agents

- Suitable for large or bulky items.

- Sterilized within a closed chamber containing highly reactive microbicidal and sporicidal gases such as ethylene oxide, propylene oxide, and beta-propiolactone.

- Can be extremely hazardous to the people using them.

ix. Antimicrobials Drugs

- antibiotics, semisynthetics and synthetics.

- Antibiotics: chemicals produced naturally by microorganisms.

i. from fungi

ii from bacteria

- Semisynthetic: antibiotic undergo modification.

- Typically used for treatment of disease.