Using Chemicals to Destroy Microorganisms and Viruses 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

Approaches to Control Principles of control Sterilization Disinfection 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

Approaches to Control Principles of control Pasteurization Brief heat treatment used to reduce organisms that cause food spoilage Surfaces can also be pasteurized Decontamination Treatment to reduce pathogens to level considered safe to handle Degerming Mechanism uses to decrease number of microbes in an area Particularly the skin (antiseptics)

Approaches to Control Principles of control Sanitized Preservation 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

Approaches to Control 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

Approaches to Control Daily life Washing and 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 organisms from outer layer of skin Normal flora usually unaffected because it resides in deeper layers

Approaches to Control 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

Approaches to Control 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 Eliminates contamination of environment

Approaches to Control Food and food production facilities Retention of 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

Approaches to Control 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 organisms 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

Selection of Antimicrobial Procedure 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

Bacterial endospores Protozoans Clostridium botulinum – causes botulism, resists boiling, but autoclaving kills Protozoans Giardia lamblia and Cryptosporidium parvum Cause digestive problems

Selection of Antimicrobial Procedure 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

Selection of Antimicrobial Procedure 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

Selection of Antimicrobial Procedure 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

Selection of Antimicrobial Procedure 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

Heat as Control Boiling (100° C) Pasteurization 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 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

Heat as Control 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 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

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 removes nearly all microbes from air Filter has 0.3µm pores to trap organisms

Other Physical Methods of Control 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

Other Physical Methods of Control 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

Other Physical Methods of Control 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

Other Physical Methods of Control 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

Chemicals as Control Potency of chemicals Sterilants = 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 Destroy viruses and vegetative cells, Not endospores Intermediate-level disinfectants Kill vegetative cells fungi, most viruses, Low-level disinfectants Removes fungi, vegetative bacteria and enveloped viruses, Not mycobacteria, naked viruses or endospores

Chemicals as Control 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

Chemicals as Control Selecting appropriate chemical Points to consider 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

Chemicals as Control Classes of chemicals Germicides represent a number or chemical families Alcohols Aldehydes Biguanides Ethylene oxide Halogens Metals Ozone Peroxides Phenolics Quaternary ammonium compounds

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

Chemicals as Control 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

Chemicals as Control 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 as means of food preservation Essentially stops microbial growth Irreversibly damages cell Kills up to 50% of microbes Remaining cells still pose potential threat

Chemicals as Control 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