Chapter 8 Control of Microorganisms by Physical and

Chapter 8 Control of Microorganisms by Physical and Chemical Agents 1

Definition of Frequently Used Terms • Sterilization – – Removes all viable microorganisms including viruses Material is said to be sterile Usually for inanimate objects Mostly performed with heat Autoclave use saturated steam under pressure to reach temperatures (above boiling) 2

Definitions…. • Disinfection – disinfectants • agents, usually chemical, used for disinfection • 5% bleach solution • The use of a physical process or chemical agent (disinfectant) to destroy vegetative pathogens • Does not destroy bacterial endospores • Usually used only on inanimate objects • Also removes toxins

Decontamination & Sanitization • Any cleansing technique that mechanically removes microorganisms to reduce contamination to safe levels (reduction of microbial population to levels deemed safe (based on public health standards) • Sanitizer: compound such as soap or detergent that sanitizes • Sanitary: may not be free from microbes but are safe for normal use 4

Antisepsis • Antisepsis – prevention of infection of living tissue by microorganisms • Antiseptics chemical agents that kill or inhibit growth of microorganisms when applied directly to exposed body surfaces to destroy or inhibit vegetative pathogens • Sepsis: the growth of microorganisms in the blood and other tissues • Asepsis: any practice that prevents the entry of infectious agents into sterile tissues

Figure 8. 1

Antimicrobial agents • –cide: to kill – – – Bactericide: chemical that destroys bacteria (not endospores) Fungicide: a chemical that can kill fungal spores, hyphae, and yeasts Virucide: a chemical that inactivates viruses Sporicide: can destroy bacterial endospores Germicide and microbicide: chemical agents that kill microorganisms • -static: to stand still (inhibit) – Bacteristatic: prevent the growth of bacteria – Fungistatic: inhibit fungal growth – Microbistatic: materials used to control microorganisms in the body, for example

The Pattern of Microbial Death • microorganisms are not killed instantly • population death usually occurs exponentially • microorganisms are considered to be dead when they are unable to reproduce in conditions that normally support their reproduction 8

Conditions Influencing the Effectiveness of Antimicrobial Agent Activity • population size – larger populations take longer to kill than smaller populations • population composition – microorganisms differ markedly in their sensitivity to antimicrobial agents • Concentration of an antimicrobial agent – usually higher concentrations or intensities kill more rapidly – relationship is not linear Copyright © The Mc. Graw-Hill Companies, Inc. Permission required for reproduction or 9

More conditions… • duration of exposure to agents longer exposure more organisms killed • temperature – higher temperatures usually increase amount of killing • local environment – Many other factors such as p. H, viscosity and concentration of organic matter can profoundly impact effectiveness Copyright © The Mc. Graw-Hill Companies, Inc. Permission required for reproduction or 10

The Use of Physical Methods in Control • • heat low temperatures filtration radiation

Heat • moist heat – effective against all types of microorganisms – degrades nucleic acids, denatures proteins, and disrupts membranes • dry heat sterilization – less effective, requiring higher temperatures and longer exposure times – oxidizes cell constituents and denatures proteins Copyright © The Mc. Graw-Hill Companies, Inc. Permission required for reproduction or 12

Measuring heat-killing efficiency • thermal death time (TDT) – shortest time needed to kill all microorganisms in a suspension at a specific temperature and under defined conditions • decimal reduction time (D or D value) – time required to kill 90% of microorganisms or spores in a sample at a specific temperature Copyright © The Mc. Graw-Hill Companies, Inc. Permission required for reproduction or 13

Figure 8. 3 14

Moist heat…. 15

Figure 8. 4 16

Moist heat… Pasteurization – controlled heating at temperatures well below boiling – reduces total microbial population and thereby increases shelf life of treated material • flash pasteurization – high temperature short-term – HTST – 72°C for 15 seconds then rapid cooling • ultrahigh-temperature (UHT) sterilization – 140 to 150°C for 1 to 3 seconds 17

Dry Heat (Oven) • Usually an electric oven • Coils radiate heat within an enclosed compartment • Exposure to 150°C to 180°C for 2 to 4 hours • Used for heat-resistant items that do not sterilize well with moist heat

Low Temperatures Ø To slow growth of cultures and microbes in food during processing and storage Ø Cold does not kill most microbes; freezing can actually preserve cultures • freezing – stops microbial reproduction due to lack of liquid water – some microorganisms killed by ice crystal disruption of cell membranes • refrigeration – slows microbial growth and reproduction Lyophilization: a combination of freezing and drying; used to preserve microorganisms and other cells in a viable state for many years 19

Filtration • Reduces microbial population or sterilizes solutions of heatsensitive materials by removing microorganisms • also used to reduce microbial populations in air • depth filters – thick fibrous or granular filters that remove microorganisms by physical screening, entrapment, and/or adsorption • membrane filters – porous membranes with defined pore sizes that remove microorganisms primarily by physical screening – Filters are usually thin membranes of cellulose acetate, polycarbonate, and a variety of plastic materials 20

Figure 9. 9 Filtration equipment-overview

Filtering air • surgical masks • cotton plugs on culture vessels • high-efficiency particulate air (HEPA) filters – used in laminar flow biological safety cabinets Figure 7. 6 a 22

Figure 7. 6 b Copyright © The Mc. Graw-Hill Companies, Inc. Permission required for

Radiation • ultraviolet (UV) radiation (non ionizing) – limited to surface sterilization because UV radiation does not penetrate glass, dirt films, water, and other substances – Germicidal lamp: 254 nm – not as penetrating as ionizing radiation • ionizing radiation (Gamma Rays, X Rays) – penetrates deep into objects – destroys bacterial endospores; not always effective against viruses – used for sterilization and pasteurization of medical products and food 24

Phenolics • commonly used as laboratory and hospital disinfectants • act by denaturing proteins and disrupting cell membranes • tuberculocidal, effective in presence of organic material, and long lasting • disagreeable odor and can cause skin irritation 26

Alcohols • bactericidal, fungicidal, but not sporicidal • inactivate some viruses • denature proteins and possibly dissolve membrane lipids 27

Halogens Iodine – skin antiseptic – Microbicidal and sporicidal with longer exposure – oxidizes cell constituents and iodinates proteins – at high concentrations may kill spores – skin damage, staining, and allergies can be a problem – iodophore • iodine complexed with organic carrier

Halogens… Chlorine – oxidizes cell constituents – important in disinfection of water supplies and swimming pools, used in dairy and food industries, effective household disinfectant – destroys vegetative bacteria and fungi, but not spores – can react with organic matter to form carcinogenic compounds 29

Heavy Metals • Ions of mercury, silver, arsenic, zinc, and copper • effective but usually toxic • combine with and inactivate proteins; may also precipitate proteins 30

Quaternary Ammonium Compounds Detergents – organic molecules with hydrophilic and hydrophobic ends – act as wetting agents and emulsifiers • cationic detergents are effective disinfectants – kill most bacteria, but not Mycobacterium tuberculosis or endospores – safe and easy to use, but inactivated by hard water and soap 31

Aldehydes • highly reactive molecules • sporicidal and can be used as chemical sterilants • combine with and inactivate nucleic acids and proteins 32

Sterilizing Gases • used to sterilize heat-sensitive materials • microbicidal and sporicidal • combine with and inactivate proteins 33

Phenol coefficient test 34