Chapter 11: Physical and chemical Agents for Microbial Control

what does Burning wood release?

formaldehyde

what do herbs, perfume, and vinegar contain

mild antimicrobial substances

Decontamination

physical, chemical, and mechanical methods to destroy or reduce undesirable microbes in a given area

Microorganism capable of causing infection or spoilage

• Vegetative bacterial cells and endospores

• Fungal hyphae and spores, yeast

• Protozoan trophozoites and cysts

• Worms

• Viruses

• Prions

Antisepsis

Chemical agents (antiseptics) applied to body surfaces, wounds, and surgical incisions to destroy or inhibit vegetative pathogens

Disinfection

use of a physical process or a chemical agent (disinfectant) to destroy or remove vegetative pathogens but not bacterial endospores from inanimate objects or materials

Sterilization

The complete removal or destruction of all viable microorganisms. Used on inanimate objects

High resistance microbes

Prions, bacterial endospores (bacillus, Clostridium)

Moderate resistance microbes

• protozoan cysts; naked viruses

• bacteria with no endospores but resistant walls: Mycobacterium, Pseudomonas, Acinetobacter, other gram(-). Staphylococcus (most heat- and chemical-resistant bacteria)

Least resistance microbes

• Most bacterial vegetative cells

• Fungal spores and hyphae

• Yeasts

• Enveloped viruses

• Protozoan trophozoites

Microbicidal agents (germicides)

antimicrobial agent aimed at destroying a certain group of microorganisms (bactericide, fungicide, virucide, sporicidal)

• Disinfection

• Antisepsis

Agents that cause microbistasis

antimicrobial agent aimed at temporarily prevent microbes from multiplying

Methods that reduce the number of microorganisms

• sanitation

• degermination

Sanitation

any cleansing technique that removes microorganisms from inanimate surfaces to reduce the potential for infection and spoilage

Degermation

reduction of microbial load from living tissue by mechanical means

Microbial death

permanent loss of reproductive capability, even under optimum growth conditions

hard to detect, microbes often reveal no conspicuous vital signs to begin with

Factors that affect microbial death

• number of microbes

• nature of microbes in the population

• temperature and pH of environment

• Concentration or dosage of agent

• mode of action of the agent

• presence of solvents, organic matter, or inhibitors

Cellular targets of physical and chemical agents

• Cell wall

• cell membrane

• protein and nucleic acid synthesis

• protein function

effects of physical and chemical agents on cell wall

becomes fragile and cell lyses (some antimicrobial drugs, detergents, and alcohol)

effects of physical and chemical agents on cell membrane

loses integrity (surfactants)

effects of physical and chemical agents on protein and nucleic acid synthesis

prevention of replication, transcription, translation, peptide bond formation, protein synthesis (chloramphenicol, ultraviolet radiation, formaldehyde)

effects of physical and chemical agents on protein function

disrupts or denature proteins (alcohols, phenols, acids, heat)

what are majority of microbes readily controlled by

abrupt changes in their environment

Physical method of control

• heat (moist and dry)

• cold temperatures

• desiccation

• radiation

• filtration

Moist heat

lower temperatures and shorter exposure time; coagulation and denaturation of proteins, which halts cellular metabolism

Dry heat

moderate to high temperatures; dehydration, alters protein structure; incineration

uses higher temperatures than moist heat

• incineration

• hot air (dry) ovens

Thermal death resistance

Bacterial endospores are most resistant, while vegetative states of bacterial the least

Thermal death time (TDT)

shortest length of time required to kill all test microbes at a specified temperature

Thermal death point (TDP)

lowest temperature required to kill all microbes in a sample in 10 minutes

Methods of Moist heat control

1. Sterilization with steam under pressure

2. nonpressurized steam

3. Boiling water: Disinfection

4. Pasteurization: Disinfection of Beverages

Sterilization with Steam under pressure

Autoclave- (15 psi/121^oC/10-40min) Pressure increases steam temperature

• steam must reach surface of item being sterilized

• item must not be heat or moisture sensitive

• produces denaturation of proteins, destruction of membranes and DNA

Nonpressurized steam

tydallization- intermittent sterilization for substances that cannot withstand autoclaving

• items exposed to free-flowing steam for 30-60 minutes, incubated for 23-24 hours and them subjected to steam again

• repeat cycle for 3 days

• used for some canned foods and laboratory media

Boiling water: Disinfection

boiling water at 100^o C for 30 minutes to destroy non-spore-forming pathogens

Pasteurization: Disinfection of Beverages

Heat is applied to kill potential agents of infection and spoilage without destroying the food flavor or value

• flash method

• Ultra pasteurized (sterile)

Flash method

71.6^o C, 15 seconds. Not sterilization- kills non-spore-forming pathogens and lowers overall microbe count; does not kill endospores or many nonpathogenic microbes

Ultapasteurized (sterile)

milk is processed (UHT)- 134^o C, 2-5 seconds

Incineration

flame, electric heating coil, infrared incinerators

• ignites and reduces microbes and other substances

• very common practice in the microbiology lab

Hot air (Dry) ovens

heated circulated air (150^o C - 180^o C, 12min - 4hr)

coagulate proteins

Cold

• Microbiostatic – slows the growth of microbes

• Refrigeration 0–15^oC and freezing < 0^oC

• Used to preserve food, media, and cultures

Desiccation

• Gradual removal of water from cells, leads to metabolic inhibition

• Not effective microbial control – many cells retain ability to grow when water is reintroduced

• Lyophilization – freeze drying; preservation

Radiation

Energy emitted from atomic activities and dispersed at high velocity through matter or space

Types of radiation suitable for microbial control

1. Ionizing radiation

2. Nonionizing radiation

Ionizing radiation

deep penetrating power sufficient energy to cause electrons to leave the orbit

• Gamma rays, X rays, cathode rays

• breaks DNA

Nonionizing radiation

little penetrating power

• Ultraviolet (UV) radiation

• UV light created pyrimidine dimers

• Interferes with replication

Filtration

Physical removal of microbes by passing a gas or liquid through filter

Used to sterilize heat sensitive liquids and air in hospital isolation units and industrial clean rooms

Chemical agents in microbial control

Disinfectants, antiseptics, sterilants, degermers, and preservatives

Some desirable qualities of germicides

• Rapid action in low concentration

• Solubility in water or alcohol, stable

• Broad spectrum, low toxicity

• Penetrating

• Noncorrosive and nonstaining

• Affordable and readily available

High-level germicides

kill endospores; may be sterilants

• Devices that are not heat-sterilizable and intended to be used in sterile environments (body tissue)

Intermediate-level

kill fungal spores (not endospores), tubercle bacillus, and viruses

• Used to disinfect devices that will come in contact with mucous membranes but are not invasive

Low-level

eliminate only vegetative bacteria, vegetative fungal cells, and some viruses

• Clean surfaces that touch skin but not mucous membranes

Levels of Chemical

1. High-level germicides

2. Intermediate-level

3. Low-level

Factors that Affect Germicidal Activity of Chemicals

1. Nature of the material being treated

2. Degree of contamination

3. Time of exposure required

4. Concentration of the chemical agent. Strength and chemical action of the germicide expressed in various ways (dilution factor (ppm), dilution percentage)

Halogens used in germicidal preparations

• Chlorine

• Iodine

Chlorine

Cl₂ , hypochlorites (chlorine bleach), chloramines

• Denature proteins by disrupting disulfide bonds

• Intermediate level

• unstable in sunlight, inactivated by organic matter

• water, sewage, wastewater, inanimate objects

Iodine

I₂ , iodophors (betadine)

• interferes with disulfide bonds of proteins

• Intermediate level

• Milder medical and dental degerming agents, disinfectants, ointments

Phenol (carbolic acid)

is an acrid, poisonous compound. Disrupt cell walls and membranes and precipitate proteins

• Low to intermediate level

• Bactericidal, fungicidal, virucidal (not sporicial)

• Lysol and Triclosan- antibacterial additive to soaps

• Toxicity of phenolics makes then a questionable choice as antiseptics

Chlorhexidine

A surfactant and protein denaturant with broad microbial properties

• low to intermediate level

• used as skin degerming agents foe preoperative scrubs, skin cleaning, and burns

Alcohols

Only ethyl and isopropyl are suitable for microbial control

Intermediate level

• Concentrations of 50% and higher dissolve membrane lipids and coagulate proteins of vegetative bacterial cells and fungi

• Water is needed for proteins to coagulate, thus alcohol shows a greater microbicidal activity at 70% concentration than at 100%

Hydrogen peroxide

Produce highly reactive hydroxyl-free radicals that damage protein and DNA while also decomposing to O2 gas – toxic to anaerobes

Antiseptic at low concentrations; strong solutions are sporicidal

Aldehydes

Kill by alkylating protein and DNA

• Glutaraldehyde

• Formaldehyde

Glutaraldehyde

• In 2% solution (Cidex)

• high level

• used as sterilant for heat sensitive instruments

Formaldehyde

• Formalin- 37% aqueous solution

• Intermediate to high level

• Disinfectant, preservative, toxicity limits use

Gases and Aerosols

• Strong alkylating agents: Ethylene oxide (ETO), propylene oxide (PO), and chlorine dioxide

• High level

• Sterilize and disinfect plastics and prepackaged devices, foods

Detergents

Polar molecules, surfactant

• Quaternary ammonia compounds (quats) act as surfactants that alter membrane permeability of some bacteria and fungi

• Very low level

Soaps

alkaline compounds

• Mechanically remove soil and grease containing microbes

• Weak microbicides, destroy only highly sensitive forms (gonorrhea, meningitis, and syphilis)

Heavy metals

• Solutions of silver and mercury kill vegetative cells in low concentrations by inactivating proteins

• Oligodynamic action (having antimicrobial effects in exceedingly small amounts)

• Low level

• Merthiolate, silver nitrate, silver

Acids and Alkalis

Low level of activity

• Organic acids prevent spore germination and bacterial and fungal growth

• Acetic acid inhibits bacterial growth

• propionic acid retards molds

• lactic acid prevents anaerobic bacterial growth

• Benzoic and sorbic acid inhibits yeast