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PowerPoint to accompany. Foundations in Microbiology. Fifth Edition. Talaro. Chapter 11. Copyright The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Physical and Chemical Control of Microbes. Chapter 11. Control of microbes.

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Foundations in microbiology l.jpg

PowerPoint to accompany

Foundations in Microbiology

Fifth Edition




Copyright The McGraw-Hill Companies, Inc. Permission required for reproduction or display.

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Control of microbes

Physical and chemical methods to destroy or reduce microbes in a given area

Physical – heat and radiation

Chemical – disinfectants and antiseptics

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Relative resistance of microbes

  • Highest resistance

    • Bacterial endospores

  • Moderate resistance

    • Bacteria with resistant vegetative cells:

      • Pseudomonas sp.

      • Mycobacterium tuberculosis

      • Staphylococcus aureus

    • Protozoan cysts

    • Some viruses (naked more resistant than enveloped)

  • Least resistance

    • most bacterial vegetative cells

    • Fungal spores

    • Enveloped viruses

    • Yeast

    • Protozoan trophozoites

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  • Decontamination - The destruction, removal, or reduction in number of undesirable microbes

  • Sepsis - The growth of microorganisms in the tissues

  • Asepsis - Techniques that prevent the entry of microorganisms into sterile tissues

  • Antiseptic - Chemicals applied to body surfaces to destroy or inhibit vegetative pathogens

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  • Sterilization – a process that destroys all viable microbes, including viruses & endospores (autoclave and radiation)

  • Disinfection – a physical or chemical agent that destroys vegetative pathogens, not endospores, on inanimate objects (bleach, boiling water)

  • Sanitization – any cleansing technique that mechanically removes microbes (soaps and detergents)

  • Degermation – reduces the number of microbes from living tissue (surgical hand scrub)

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Microbial death

  • Involves permanent loss of reproductive capability, even under optimum growth conditions

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Factors that influence action of antimicrobial agents:

  • Number of microbes

  • Nature of microbes in the population (young cells die more quickly)

  • Temperature & pH of environment

  • Concentration or dosage of agent

  • Mode of action of the agent

  • Presence of solvents, organic matter, or inhibitors

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Cellular targets of control by antimicrobial agent

  • Cell wall

  • Cell membrane

  • Cellular synthetic processes (DNA, RNA)

  • Proteins

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Surfactants inserting into the lipid bilayer, alter permeability, cause leakage both in and out of the cell

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a) The native state is maintained by bonds that create active sites to fit the substrate. Agents denature by breaking 2o and 3o bonds

c) Random bonding and incorrect folding

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Selecting a workable method of microbial control active sites to fit the substrate. Agents denature by breaking 2

Practical concerns

  • Does the application require sterilization? Or is disinfection adequate?

  • Is the item to be reused?

  • Can the item withstand heat, pressure, radiation, or chemicals?

  • Is the method suitable?

  • Will the agent penetrate to the necessary extent?

  • Is the method cost- and labor-efficient & is it safe?

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Methods of Physical Control active sites to fit the substrate. Agents denature by breaking 2

  • Heat

  • Cold temperatures

  • Desiccation

  • Radiation

  • Filtration

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1. Heat active sites to fit the substrate. Agents denature by breaking 2

Moist heat – use of hot water or steam

  • Mode of action – denaturation of proteins, destruction of membranes & DNA

  • sterilization

  • autoclave 15 psi/121oC/10-40min (raise the temp of steam by increasing the pressure)

  • intermittent sterilization – unpressurized steam at 100oC 30-60 min for 3 days

  • disinfection

  • Pasteurization <100oC for seconds; kills Salmonella, Listeria & overall microbe count

  • Boiling at 100oC for 30 minutes to destroy non-spore-forming pathogens

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Sterilization using steam under pressure active sites to fit the substrate. Agents denature by breaking 2

The Autoclave

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Dry heat active sites to fit the substrate. Agents denature by breaking 2 using higher temperatures than moist heat, can also sterilize, less efficient that moist heat

  • incineration – 600-1200oC combusts & dehydrates cells (limited to metals, ex. loop)

  • dry ovens – 150-180oC- coagulate proteins, not suitable for plastics/paper

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Dry heat- Incineration active sites to fit the substrate. Agents denature by breaking 2

Shield prevents splattering of microbial samples during flaming

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2. Cold temperatures active sites to fit the substrate. Agents denature by breaking 2

  • Microbistatic – slows the growth of microbes

  • refrigeration 0-15oC & freezing <0oC

  • Pathogens can survive several months

  • used to preserve food, media and cultures

  • -70oC to -135oC can preserve cultures of bacteria and viruses for long periods of time

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3. Desiccation active sites to fit the substrate. Agents denature by breaking 2

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

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

  • A valuable way to preserve foods

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4. Radiation active sites to fit the substrate. Agents denature by breaking 2

Defined as energy emitted from atomic activities and dispersed at high velocity through matter or space

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4. Radiation active sites to fit the substrate. Agents denature by breaking 2

Ionizing radiation – ejects orbital electron from an atom causing ions to form

  • Deep penetrating power, breaks DNA

  • gamma rays, X-rays, cathode rays

  • used to sterilize medical supplies & food products

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4. Radiation active sites to fit the substrate. Agents denature by breaking 2

Nonionizing radiation: Ultraviolet Rays – excites atoms by raising them to a higher energy state, leads to the formation of abnormal bonds

  • Little penetrating power to sterilize air, water and solid surfaces

  • UV light creates thymine pyrimidines dimers, which interfere with replication

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Dislodges electron from molecules active sites to fit the substrate. Agents denature by breaking 2

Excites electron-abnormal bonds form

A solid barrier cannot be penetrated by nonionizing radiation

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Formation of pyrimidine dimers by the action of UV radiation active sites to fit the substrate. Agents denature by breaking 2

Pyrimidine bases=

Thymine and cystosine

Dimers prevent that segment from correctly replicating or transcribing

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5. Filtration water

  • physical removal of microbes by passing a gas or liquid through filter

  • used to sterilize heat sensitive liquids & air in hospital isolation units & industrial clean rooms

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Filtration water

Scanning electron micrograph of filter showing relative size of pores and bacteria trapped on surface (8um to 0.2um)

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Chemical control water

  • Halogens

  • Phenolics

  • Chlorhexidine

  • Alcohols

  • Hydrogen peroxide

  • Detergents & soaps

  • Heavy metals

  • Aldehydes

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Halogens water(used for disinfection for 200 yrs)

  • Chlorine – Cl2, hypochlorites (chlorine bleach), chloramines

    • Denaturation of proteins by disrupting disulfide bonds

    • Can be sporicidal

  • Iodine - I2, iodophors (Betadine)

    • Denature proteins

    • Can be sporicidal

    • Milder medical & dental degerming agents, disinfectants, ointments

    • Less susceptible to inactivation by organic material

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2. Phenolics water

  • Phenolics-one or more aromatic carbon rings with additional functional groups

  • Disrupt cell membranes & precipitating proteins; bactericidal, fungicidal, virucidal, not sporicidal

    • Lysol

    • triclosan- antibacterial additive to soaps

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3. Chlorhexidine additional compounds

  • Hibiclens, Hibitane

  • A surfactant & protein denaturant with broad microbicidal properties

  • Not sporicidal

  • Used as skin degerming agents for preoperative scrubs, skin cleaning & burns

  • Mildness, low toxicity and rapid action

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4. Alcohols additional compounds

  • Colorless hydrocarbons with one or more -OH functional groups

  • Ethyl or isopropyl alcohol in solutions of 50-95%. At 50% or greater concentration dissolves cell membrane lipids

    • 70% proteins coagulate

    • 100% dehydrates

  • Act as surfactants dissolving membrane lipids and coagulating proteins of vegetative bacterial cells, fungi, and enveloped viruses

  • Skin degerming agent. Not sporicidal

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5. Hydrogen peroxide (H additional compounds2O2)

  • A colorless, caustic liquid that decomposes in the presence of light into water and oxygen gas

  • Weak (3%) to strong (25%)

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

  • Strong solutions are sporicidal

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6. Detergents & soaps additional compounds

  • Quaternary ammonia cpds act as surfactants that alter membrane permeability of some bacteria & fungi

    • Positively charged detergent

    • Not sporicidal

  • Soaps- mechanically remove soil and grease containing microbes

    • Alkaline cmpds made by combining the fatty acids in oils with sodium or potassium salts

    • Weak microbicides

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Detergents are polar molecules with charged head and uncharged tail

A common quarternary ammonium detergent

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7. Heavy metals- uncharged tail(relatively high atomic weight: mercury, silver, and gold)

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

  • Oligodynamic action-the property of having antimicrobial effects in exceedingly sm. amts

  • Not sporicidal

  • Bind to functional groups of proteins and inactivate them

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A pour plate inoculated with saliva uncharged tail

Contains silver and mercury

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8. Aldehydes uncharged tail

  • Organic substances bearing a reducing functional group on terminal carbon

  • Glutaraldehyde & formaldehyde kill by alkylating protein & DNA

  • glutaraldehyde in 2% solution (Cidex) used as sterilant for heat sensitive instruments, high-level disinfectant

  • formaldehyde - disinfectant, preservative, toxicity limits use

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Gases & aerosols uncharged tail

  • Ethylene oxide, propylene oxide, betapropiolactone & chlorine dioxide

  • Strong alkylating agents, sporicidal

  • Sterilizing inanimates

  • Ethylene oxide is the gas used in chemiclaves

  • Reactions with functional groups of DNA and proteins, therefore, blocks both DNA replication and enzymatic actions

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Sterilization Using Gas uncharged tail

An automatic ethylene oxide (ETO) sterilizer. Machine equipped with gas canisters containing ETO and carbon dioxide, chamber to hold items, and ways to evacuate and introduce air

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Table 11.7 uncharged tail

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Fig. 11.1 uncharged tail