control of microbial growth chapter 5. approaches to control control mechanisms either physical or...

Control of Microbial Growth

Chapter 5

Approaches to Control

Control mechanisms either physical or chemical May be a combination of both Physical methods

Heat Irradiation Filtration Mechanical removal

Chemical methods Use a variety of antimicrobial chemicals Chemical depends on circumstances and degree of

control required


Principles of control Sterilization

Removal of all microorganisms Sterile item is absolutely free of microbes, endospores and

viruses Can be achieved through filtration, heat, chemicals and

irradiation Disinfection

Eliminates most pathogens Some viable microbes may exist

Disinfectants = used on inanimate objects and surfaces Antiseptics = used on living tissues

Pasteurization Brief heat treatment used to reduce organisms that cause

food spoilage Surfaces can also be pasteurized

Principles of control Decontamination

Treatment to reduce pathogens to level considered safe Degerming

Mechanism uses to decrease number of microbes in an area

Particularly the skin

Sanitized Implies a substantially reduced microbial population

This is not a specific level of control

Preservation Process used to delay spoilage of perishable items

Often includes the addition of growth-inhibiting ingredients



Situational considerations Microbial control methods

are highly variable Depends on situation

and degree of control required

Daily life Hospital Microbiology laboratories Food and food

production facilities Water treatment

Daily life Washing a scrubbing with soaps and

detergents achieves routing control Hand washing single most important step to

achieving control Soap acts as wetting agent

Aids in mechanical removal of microorganisms Removes numerous organism from outer layer of

skin Normal flora usually unaffected because it

resides in deeper layers


Hospitals Minimizing microbial population very important

Due to danger of nosocomial infections Patients are more susceptible to infection Pathogens more likely found in hospital setting

Numerous organisms develop antimicrobial resistance due to high concentrations of antibiotics

Instruments must be sterilized to avoid introducing infection to deep tissues


Microbiology laboratories Use rigorous methods of control

To eliminate microbial contamination to experimental samples and environment

Aseptic technique and sterile media used for growth Eliminates unwanted organisms

Contaminated material treated for disposal Eliminate contamination of environment


Food and food production facilities Retention of food quality; enhanced through

prevention of microbial growth and contamination

Achieved through physical removal and chemical destroying organisms

Heat treatment most common and most reliable mechanism

Irradiation approved to treat certain foods Chemicals prevent spoilage

Risk of toxicity


Water treatment facilities Ensures drinking water is safe Chlorine generally used to disinfect water

Can react with naturally occurring chemicals Form disinfection by-products (DBP)

Some DBP linked to long-term health risks Some organism resistant to chemical

disinfectants


Selection of Antimicrobial Procedure

Selection of effective procedure is complicated Ideal method does not exist

Each has drawbacks and procedural parameters Choice of procedure depends on numerous factors

Type of microbe Extent of contamination

Number of organisms Environment Risk of infection Composition of infected item

Type of microorganism Most critical consideration

Is organism resistant or susceptible to generally accepted methods?

Resistant microbes include Bacterial endospores

Resistant to heat, drying and numerous chemicals Protozoan cysts and oocysts

Generally excreted in feces and cause diarrheal disease Mycobacterium species

Cell wall structure initiates resistance Pseudomonas species

Can grow in presence of many chemical disinfectants Naked viruses

Lack envelope and are more resistant to chemical killing



Number of organisms initially present Time it takes to kill it directly

affected by population size Large population = more time

Commercial effectiveness is gauged by decimal reduction time

A.k.a D value Time required to kill 90% of

population under specific conditions

Washing reduces time required to reach disinfection or sterilization

Environmental conditions Environmental conditions strongly influence

effectiveness pH, temperature and presence of organic

materials can increase or decrease effectiveness Most chemicals are more effective at higher

temperatures and lower pH Effectiveness can be hampered by the presence of

organism molecules Can interfere with penetration of antimicrobial

agent


Potential risk of infection Medical items categorized according to potential

risk of disease transmission Critical items = come in contact with body tissues

Needles and scalpels Semicritical instruments = contact mucous

membranes but do not penetrate body tissues Endoscope

Non-critical instruments = contact unbroken skin only Show little risk of transmission Stethoscope


Composition of the item Some sterilization and disinfection methods

inappropriate for certain items Heat inappropriate for plastics and other heat

sensitive items


Heat as Control

Heat treatment most useful for microbial control Relatively fast, reliable, safe and inexpensive

Heat can be used to sterilize or disinfect Methods include

Moist heat Dry heat

Heat as Control

Moist heat Destroys through irreversible coagulation of

proteins Moist heat includes

Boiling Pasteurization Pressurized steam

Boiling (100 C) Destroys most microorganisms and viruses Not effective means of sterilization

Does not destroy endospores Pasteurization

Pasteur developed to avoid spoilage of wine Does not sterilize but significantly reduces organisms Used to increase shelf life of food Most protocols employ HTST method

Heated to 72°C and held for 15 seconds Other protocol UHT

Heated to 140°C - 150°C, held for several seconds then rapidly cooled

Heat as Control

Heat as Control Pressurized steam

Autoclave used to sterilize using pressurized steam

Heated water steam increased pressure

Preferred method of sterilization

Achieves sterilization at 121°C and 15psi in 15 minutes

Effective against endospores

Flash autoclaving sterilizes at 135°C and 15psi in 3 minutes

Prions destroyed at 132°C and 15psi for 4.5 hours

Dry heat Not as effective as moist heat

Sterilization requires longer times and higher temperatures

200°C for 1.5 hours vs. 121°C for 15 minutes for moist Incineration method of dry heat sterilization

Oxidizes cell to ashes Used to destroy medical waste and animal carcasses Flaming laboratory inoculation loop incinerates

organism Results in sterile loop

Heat as Control

Other Physical Methods of Control Heat sensitive materials require other

methods of microbial control Filtration Irradiation High-pressure treatment

Other Physical Methods of Control

Filtration Membrane filtration used

to remove microbes from fluids and air

Liquid filtration Used for heat

sensitive fluids Membrane filters allow

liquids to flow through Traps microbes on

filter Depth filters trap

microbes using electrical charge

Filtration of air High efficiency particulate

air (HEPA) filter remove nearly all microbes from air

Filter has 0.3µm pores to trap organisms


Radiation Electromagnetic radiation

Energy released from waves

Based on wavelength and frequency

Shorter wavelength, higher frequency = more energy

Range of wavelength is electromagnetic spectrum

Radiation can be ionizing or non-ionizing

Ionizing radiation Radiation able to strip electrons from atoms Three sources

Gamma radiation X-rays Electron accelerators

Causes damage to DNA and potentially to plasma membrane

Used to sterilize heat resistant materials Medical equipment, surgical supplies, medications Some endospores can be resistant


Ultraviolet radiation Non-ionizing radiation

Only type to destroy microbes directly Damages DNA

Causes thymine dimers Used to destroy microbes in air, drinking water

and surfaces Limitation

Poor penetrating power Thin films or coverings can limit effect


High pressure processing Used in pasteurization of commercial foods

Does not use high temperatures Employs high pressure

Up to 130,000 psi Destroys microbes by denaturing proteins and

altering cell membrane permeability


Chemicals as Control

Chemicals can be used to disinfect and sterilize Called germicidal

chemicals Reacts with vital cell

sites Proteins DNA Cell membrane


Potency of chemicals Formulations generally

contain more than one antimicrobial agent

Regulated by FDA

Antiseptics EPA

Disinfectants

Germicidal agents grouped according to potency

Sterilants = Destroy all microorganisms

High-level disinfectants Destroys viruses and vegetative

cells, Not endospores

Intermediate-level disinfectants Kills vegetative cells fungi, most

viruses, Not endospores

Low-level disinfectants Removes fungi, vegetative

bacteria and enveloped viruses Not mycobacteria, naked

viruses or endospores

Selecting appropriate chemical Points to consider

Toxicity Benefits must be weighed against risk of use

Activity in presence of organic material Many germicides inactivated in presence of organic matter

Compatibility with material being treated Liquids cannot be used on electrical equipment

Residue Residues can be toxic or corrosive

Cost and availability Storage and stability

Concentrated stock relieves some storage issues Environmental risk

Is germicidal agent harmful to environment


Classes of chemicals Germicides represent a number or chemical families

Alcohols Aldehydes Biguanides Ethylene oxide Halogens Metals Ozone Peroxides Phenolics Quaternary ammonium compounds


Alcohols Solutions of 60% - 80% isopropyl or ethyl alcohol kill

vegetative bacteria and fungi Not effective against endospores and some naked

viruses Mode of action

Coagulation of proteins and essential enzymes Damage to lipid membranes

Commonly used as antiseptic and disinfectant Limitations

Evaporates quickly limiting contact time May damage material such as rubber and some plastics


Aldehydes Destroy organisms by inactivating proteins

and DNA 2% glutaraldehyde solution most widely used

liquid sterilant Orthophthalaldehyde studied as alternative

Formalin used to kill bacteria and inactivate viruses

Also used for specimen preservation Formalin is solution made from formaldehyde


Biguanides Most effective member of group is

chlorhexidine Extensively used in antiseptics Relative low toxicity Destroys wide range of organisms


Ethylene oxide Useful gaseous sterilant

Destroys microbes including endospores and viruses

Mode of action Reacts with proteins

Useful in sterilizing heat or moisture sensitive items

Limitations Mutagenic and potentially carcinogenic


Halogens Common disinfectants Mode of action

Oxidizing proteins and other cell components Includes chlorine and iodine Chlorine

Destroys all types of organisms and viruses Used as disinfectant

Caustic to skin and mucous membranes Chlorine dioxide replacing chlorine in many applications

Iodine Kills vegetative cells

Not reliable with endospores Used in tincture or iodophore on skin


Metal compounds Compounds combine with enzymes and

proteins Interfering with function

High concentrations of many metals toxic to human tissue

Silver still used as disinfectant Creams containing silver sulfadiazine used to

prevent secondary infections Also available on bandages for wound care


Ozone O3

Unstable form of oxygen Powerful oxidizing agent Used as alternative to chlorine

As disinfectant for drinking and waste water


Peroxygens Includes hydrogen peroxide and peracetic

acid Powerful oxidizing agents Readily biodegradable Less toxic than ethylene oxide and

glutaraldehyde


Hydrogen peroxide Effectiveness depends on surface being treated

Living tissue produce catalase enzyme Breaks down hydrogen peroxide to oxygen and water More effective on inanimate object

Useful as disinfectant Leaves no residue Doesn’t damage most materials

Hot solutions used in food industry Vapor-phase can be used as sterilant

Peracetic acid More potent then hydrogen peroxide Effective on organic material

Can be used on wide range of material


Phenolics A.k.a carbolic acid One of the earliest disinfectants

Now has limited use Active ingredient in Lysol Mode of action

Destroy plasma membrane Denature proteins

Kills most vegetative cells Can kill mycobacterium at high concentrations Not reliable on all groups of viruses

Triclosan and hexachlorophene phenols used in soaps and lotions


Quaternary ammonium compounds A.k.a Quats Cationic detergents Nontoxic

Used to disinfect food preparation surfaces Mode of action

Reduces surface tension Aids in removal of dirt and organic matter Facilitates mechanical removal of organisms

Positive charge attracts Quats to negative charge of cell surface

Reacts with membrane Destroys vegetative bacteria and enveloped viruses Not effective on endospores, mycobacteria and naked

viruses


Preservation of Perishable Products

Preservation extends shelf-life of many products Chemicals are often added to prevent or slow

growth of microbes Other methods include

Low temperature storage Freezing Reducing available water

Chemical preservatives Numerous chemicals are used as preservatives

Formaldehyde, Quats, and phenols Weak organic acids often used as food preservatives

Benzoic, ascorbic and propionic acids Used in bread, cheese and juice Mode of action

Alter cell membrane function Interfere with energy transformation

Nitrates and nitrites used in processed meats Inhibits germination of endospores and growth of

vegetative cells Have been shown to be potent carcinogen


Low temperature storage Microbial growth is temperature dependent

Low temperatures slow down or stop enzymatic reactions of mesophiles and thermophiles

Some psychrophiles still able to grow

Freezing means of food preservation Essentially stops microbial growth Irreversibly damages cell

Kills up to 50% of microbes Remaing cells still pose potential threat


Reducing water availability Decreasing water availability accomplished by salting or

drying food Addition of salt increases environmental solutes

Causes cellular plasmolysis Numerous bacteria can continue to grow in high salt

environments Staphylococcus aureus can survive in high salt concentrations

Desiccation or drying is often supplemented by other methods

Salting Lyophilization (freeze drying)

Widely used to preserve foods like coffee, milk and meats


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