Microbial Death and Control

Microbial Death and Control

• Focus of chapter 11 involves antiseptics and aseptic methods.
• Aseptic methods: preventing infection.
• Antisepsis: destroying/inhibiting vegetative pathogens.
• The standards for each environment differ.
• Controlling microbes requires physical, mechanical, and chemical strategies.

Death

• Microbes do not show obvious signs of life.
• Microbial death involves the permanent loss of reproductive capability, even under optimal growth conditions.
• Control can static (inhibit growth) or cidal (kill).

Definitions

• Germicide kills pathogens and many non-pathogens, but not necessarily endospores.
• Static agents inhibit microbial growth.
• Sterilization is the process to destroy all viable microbes, though often only a reduction of microbial numbers to a safe level are needed.
• Disinfection removes vegetative pathogens, but not bacterial endospores, typically on inanimate objects.
• Sterilization is the complete removal of all microorganisms, usually on inanimate objects.
• Antiseptics are chemicals used to body surfaces to destroy or inhibit vegetative pathogens.
• Chemotherapy involves chemicals used internally to kill or inhibit microorganism growth within host tissues.
• Decontamination reduces the number of undesirable microbes.
• Sepsis is the growth of microorganisms in the tissues.
• Asepsis are techniques to prevent the entry of microorganisms into sterile tissues.
• Sanitization cleanses techniques to remove microorganisms and debris from inanimate surfaces.
• Degermation cleanses living tissue to remove microorganisms and debris.

Factors Affecting Death

• The number of microbes affects microbial death.
• The nature of microbes in the population affects microbial death.
• Population death occurs exponentially.
• Decimal reduction time shows the measure of agent's killing efficiency, and the time to kill.
• Larger populations take longer to kill than smaller populations.
• Temperature and pH of the environment affect microbial death.
• Concentration or dosage of the agent affect microbial death.
• The Mode of action of the agent affects microbial death.
• Presence of solvents, organic matter, or inhibitors affects microbial death.
• Prions are the most resistant microbes.
• Enveloped viruses are the most susceptible microbes.

Practical Concerns

• The selection of method of control depends on circumstances.
• Circumstances include deciding whether the application requires sterilization, if the item can be reused and if the item can withstand heat, pressure, radiation, or chemicals.
• The method must be suitable, and will the agent penetrate to the necessary extent.
• Is the method cost- and labor-efficient and is it safe.

Cellular/Physical Targets

• The cell wall becomes fragile and lyses when impacted by some antimicrobial drugs, detergents, and alcohol.
• The cell membrane loses integrity when impacted by surfactants.
• Protein and nucleic acid synthesis prevention of replication, transcription, translation, and peptide bond formation.

Physical Methods of Control

• The majority of microbes can be readily controlled by abrupt changes in their environment.
• Some physical methods are heat, cold temperatures, desiccation, radiation and filtration.

Heat Control

• Mode of action of heat involves moist heat at lower temperatures and shorter exposure time, coagulation, and denaturation of proteins which halts metabolism.
• Mode of action of heat involves dry heat at moderate to high temperatures that dehydrates and alters protein's structures, accomplished by incineration.
• Thermal death time (TDT) is the shortest length of time required to kill all test microbes at a specified temperature.
• Thermal death point (TDP) is the lowest temperature required to kill all microbes in a sample in ten minutes.
• Moist heat, such as carried out above 100°C, requires saturated steam under pressure and uses an Autoclave.
• Quality control includes strips with Geobacillus stearothermophilus

Methods of Moist Heat Control

• Tyndallization is intermittent sterilization for substances that cannot withstand autoclaving.
• Items exposed to free-flowing steam for 30-60 minutes, incubated for 23-24 hours, and then subjected to steam again.
• Repeat this cycle for three days.
• Tyndallization is used for canning foods and lab media.
• Boiling water at 100°C for 30 minutes is used to inactivate non-spore-forming pathogens.

Pasteurization

• Pasteurization involves controlled heating at temperatures well below boiling.
• Pasteurization Process does not sterilize.
• The Pasteurization Process kills pathogens and slows spoilage by reducing the total load of organisms present.
• Flash method: At 71.6°C for 15 seconds is not sterilization since it only kills non-spore-forming pathogens, lowers overall microbe court, does not kill endospores or many non-pathogenic microbes.
• Ultrapasteurized (sterile) milk is processed (UHT) at 134°C for 2-5 seconds.

Dry Heat

• Incineration is applied by flame, electric heating coil, or infrared incinerators ignites and reduces microbes and other substances.
• Hot air ovens heated by circulated air at 150°C-180°C, 12min-4h coagulate proteins.

Cold and Desiccation Effects

• Refrigeration at 0-15°C and freezing at less than 0°C microbiostatic slows the growth of microbes, and is often used to preserve food, media, and cultures.
• It can be static or cidal in its effects.
• Desiccation involves the gradual removal of water from cells, which leads to metabolic inhibition.
• This process that is not effective for microbial - many cells retain ability to grow when water is reintroduced.
• Lyophilization is a form of freeze drying that allows preservation.

Radiation Methods

• Radiation involves energy emitted from atomic activities and dispersed at high velocity through matter or space.
• One control method consists of ionizing radiation, such as gamma rays, X rays, and cathode rays.
• Another radiation method is noniodizing or ultraviolet (UV) radiation.

Cellular Effects of Irradiation

• Ionizing radiation has deep penetrating power sufficient energy to cause electrons to leave their orbit.
• This includes gamma rays, X rays, and cathode rays which breaks DNA.
• It is used for sterilization and pasteurization of antibiotics, hormones, sutures, plastic disposable supplies, and food.
• Nonionizing radiation has little penetrating power.
• UV is the most common form of radiation when discussing nonionizing radation.
• UV light creates pyrimidine dimers and interferes with replication.
• It is limited to surface sterilization because it does not penetrate glass, dirt films, water, and other substances.

Filtration Method

• It physically removes of microbes by passing a gas or liquid through filter.
• Filtration is Used to sterilize heat sensitive liquids and air in hospital isolation units and industrial clean rooms.
• Filters in this method come in a variety of pore sizes.
• High-efficiency particulate air (HEPA) filters are used in laminar flow biological safety cabinets used in a laboratory.
• Surgical masks, N-95 Masks - 95% >0.3 uM, and cotton plugs on culture vessels can be considered rudimentary filtration devices.

Germicides

• Germicides are chemical agents in microbial control consist of disinfectants, antiseptics, sterilants, degermers, and preservatives.
• Desirable qualities of germicides that they are fastacting, soluble in water or alcohol, stable, broad spectrum, low toxicity, penetrating, noncorrosive and non-staining, affordable, and readily available.

Levels of Chemical Decontamination

• High-level germicides kill endospores and may be sterilants.
• Devices that use are not heat-sterilizable and intended to be used in sterile environments (body tissue).
• Intermediate-level use kills fungal spores (not endospores), tubercle bacillus, and viruses.
• Devices used are to disinfect that will come in contact with mucous membranes but are not invasive.
• Low-level chemicals used eliminate only vegetative bacteria, vegetative fungal cells, and some viruses.
• Low-level chemicals used clean surfaces that touch skin but not mucous membranes.

Factors That Affect Germicidal Activity of Chemicals

• The factors are nature of the material being treated; degree of contamination; time of exposure required; concentration of the chemical agent, strength, and chemical action of the germicide expressed in dilution factor.

Categories

• Halogens, Phenolics, Chlorhexidine, Alcohols, Hydrogen peroxide, Aldehydes,Gases Detergents and soaps, Heavy metals, Dyes Acids and Alkalis

Halogens

• Halogens are Chlorine – Cl₂, hypochlorites (chlorine bleach), chloramines used in germicidal preparations denaturate proteins by disrupting disulfide bonds.
• It Is intermediate level, unstable in sunlight, inactivated by organic matter and good to use on water, sewage, wastewater, inanimate objects.
• Iodines is most commonly 12 and iodophors.
• I2 interferes with disulfide bonds of proteins, has an intermediate level, a milder medical and dental degerming agent, disinfectant, ointments.

Phenol

• Phenol is a carbolic acid is an acrid, poisonous compound that disrupts cell walls and membranes and precipitate proteins
• Its low to intermediate level, bactericidal, fungicidal, virucidal
• Toxicity makes them a questionable choice as antiseptics, such as Lysol and Triclosan antibacterial additive to soaps.
• It is tuberculocidal, effective in presence of organic material, and long lasting

Chlorhexidine

• It is used for skin cleaning, burns, and preoperative scrubs.
• It is a surfactant and protein denaturant with broad microbicidal properties.
• Chlorhexidine contains Hibiclens and Hibitane and is low to intermediate in its efficiency.

Alcohols

• Ethyl and isopropyl are used for microbial agents.
• Alcohol concentrations of 50% can dissolve membrane lipids and coagulate proteins of vegetative bacterial cells and fungi.
• Alcohol has greater microbiocidal activity at 70% concentration than at 100%

Oxidizing Agents

• Oxidizing agents Produce highly reactive hydroxyl-free radicals that damage protein and DNA while also decomposing to O₂ as a gas.
• The agent can Antiseptic at low concentrations. Strong solutions are sporicidal.

Aldehydes

• Glutaraldehyde can be used to kill by alkylating protein and DNA. In 2% solution the high level can be used as sterilant for heat sensitive instruments.
• Formaldehyde can be used to kill by alkylating protein and DNA. Formalin – 37% aqueous solution with its Intermediate to high level can be used as disinfectant or preservative as long as toxicity limits are considered.

Gases

• It contains strong alkylating agents: Ethylene oxide (ETO), propylene oxide (PO), and chlorine dioxide.
• It had a Level: High level, used to sterilize and disinfect plastics and prepackaged devices, foods.

Soaps

• Detergents includes Polar molecules and surfactants is Quaternary ammonia compounds (quats), can be used that alter membrane permeability of some bacteria and fungi is very low
• Mechanically removes soil and grease containing microbes, but weak microbicides can destroy only highly sensitive forms (gonorrhea, meningitis, and syphilis).

Quaternary Ammonium

• They are amphipathic organic cleansing agents, like cationic detergents and are effective disinfectants that kill most bacteria, but not M. tuberculosis or endospores.
• Can be safely and easily used, as long as they are inactivated by hard water and soap.

Heavy Metals

• Metals are created with solutions of heavy metals and kill at incredibly low concentrations and have have antimicrobial effects.

Acids/Alkalis

• They have a generally low level of activity to disrupt spores or bacterial/fungal growth.
• They have ingredients like acetic acid, propionic acid, lactic acid, and benzoic/asorbic acid to disrupt microorganisms.

Agent Evaluation

• Complex test that must pass agencies like Environmental Protection Agency, Food and Drug Administration, and/or the CDC.
• Test must show effective death to microorganisms, but safe and easy to use.