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PowerPoint to accompany Microbiology: A Systems Approach Cowan/Talaro Chapter 11 Physical and Chemical Control of Microbes Copyright The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Chapter 11 Topics - Controlling Microorganisms - Physical Control - Chemical control 2 An overview of the microbial control methods. Fig. 11.1 Microbial control methods 3 Controlling Microorganisms • • • • Microbial agents Sanitation Effectiveness Mode of action 4 Microbial agents • • • • • • -static agents -cidal agents Resistance Terms Effectiveness Mode of action 5 -static agents • Temporarily prevent the growth of microbes – Bacteriostatic – Fungistatic 6 -cidal • Kill or destroy a microorganism – Germicide – Bactericide 7 Resistance • Highest resistance - bacterial spores and prions • Moderate resistance - some bacteria, protozoan cysts, fungal sexual spores, naked viruses • Least resistance - most bacteria, fungal nonsexual spores and hyphae, enveloped viruses, yeast, protozoan trophozoites 8 Comparing the resistance between endospores and vegetative cells to control agents. 9 Table 11.1 Relative resistance of bacterial endospors and vegetative cells Terms • • • • • Sterilization Disinfection Antisepsis Sanitation Degermination 10 Sterilization • A process that destroys or removes all viable microorganisms, including spores and viruses • Physical or chemical agents • Inanimate objects – Surgical instruments, commercially packaged foods 11 Disinfection • Use of physical process or chemical agent (disinfectant) to destroy vegetative pathogens. • Removes toxins • Won’t destroy bacterial endospores 12 Antisepsis • Chemical agents (antiseptics) destroy or inhibit vegetative pathogens • Agent is used on skin and mucous membranes 13 Sanitation • Reducing the number of microorganisms to a safe public health standard • Physical and chemical agents • Dishwashers, sanitizers 14 Degermination • On Human skin - reducing the number of microorganisms by mechanical means – Wiping the microbes from the skin surface • Physical and chemical agents can be used 15 Factors that influence Effectiveness • Number of microorganisms • Target population (bacteria, fungi, spores, viruses) • Temperature and pH • Concentration of agent • Mode of action • Interfering agents present (solvents, debris, saliva, blood, feces) 16 Factors that influence the effectiveness of antimicrobial agents. Fig. 11.2 Factors that influence the rate at which microbes are killed by antimicrobial agents 17 Mode of action How Antimicrobial Agents Work • Targets – – – – – Cell wall Cell membrane Nucleic acid synthesis Protein synthesis Protein function 18 Target: Cell Wall • Bacteria and fungi have cell walls – Block synthesis of cell wall – Degrade cellular components of cell wall – Destroy or reduce stability of cell wall • Agents – Penicillin, detergents, alcohols 19 Target: Cell membrane • All microbes have a cell membrane • Enveloped viruses have an envelope (an outer bilayer of phospholipids) • Agents that destroy cell membranes bind and penetrate lipids – Membrane loses selective permeability (leakage) • Agents – Surfactants 20 The effect of surfactants on the cell membrane. Fig. 11.3 Mode of action of surfactants 21 Nucleic acid synthesis • Some agents irreversibly bind to DNA – Stop transcription and translation – Can cause mutations • Agents – Chemical agent – formaldehyde – Physical agent – radiation 22 Protein synthesis • Agents that bind to ribosomes – Stop translation – Prevent peptide bonds • Agent – Chloramphenicol • Remember: bacteria have 70S ribosomes; eukaryotes have 80S ribosomes but have 70S ribosomes in the mitochondria 23 Protein function • • • • Block protein active sites Prevent binding to substrate Denature protein Agent – Physical – Heat, pH change – Chemical – alcohols, acids, phenolics, metallic ions 24 The effects of heat, pH, and blocking agents on the function of proteins. Fig. 11.4 Modes of action affecting protein function 25 Physical Control • Heat • Radiation • Filtration 26 HEAT Mode of action • Moist heat – Coagulation of proteins – Denaturation of proteins • Dry heat – Dehydration – Denaturation – Oxidation (burning to ashes) • Thermal death time 27 Moist Heat Methods • Boiling water • Steam and pressure • Pasteurization 28 Boiling water • Boiling water at 100 ˚C for 30 minutes – Decontaminates but doesn’t sterilize • Kills most non-spore forming pathogens • Won’t destroy endospores • Examples: home sanitizing and disinfecting, disinfecting unsafe water 29 Steam and pressure • Increasing pressure above normal atmospheric pressure will result in water boiling temperatures above 100˚C • 15 psi / 121˚C / 15-30 min. • Effectively destroys spores • Sterilizes inanimate objects (glassware) – Material must be able to withstand both heat and moisture • Ex. Autoclave and home pressure cooker 30 A diagram of an autoclave. 31 Fig. 11.5 Steam sterilization with the autoclave Pasteurization • Disinfection of beverages • Exposes beverages to 71.6 ˚C for 15 seconds – Stops fermentation • Prevents the transmission of milk-borne diseases – Salmonella, Campylobacter, Listeria, Mycobacteria • Examples: Milk industry, wineries, breweries 32 Dry heat • Hot air ( e.g., oven) • Incineration • Not as efficient as using moist heat – Temperature and /or time of exposure is greater than moist heat 33 Hot air • Hot air – Oven – Effective at 150˚C to 180˚C for 2-4 hrs – Effective for inanimate objects and oils 34 Incineration • Destroys microbes to ashes or gas – Flame - 1870˚C – Furnace - 800˚C to 6500˚C 35 An infrared incinerator uses flame to burn or oxidize materials into ashes. Fig. 11.6 Dry heat incineration 36 Effects of cold and dessication • Cold temperatures reduce the activity of some microbes, not psychrophiles – Not a disinfection or sterilization method • Dessication or dehydration kill some microorganisms – Lyophilization – freezing and drying method used to preserve microbes 37 Radiation • Types of radiation • Modes of action • Applications 38 Types of radiation • Ionizing – Very High Energy – Gamma rays (Highest ionizing energy) – X-rays (Intermediate ionizing energy) – Cathode rays (least ionizing energy) • Nonionizing – Ultraviolet 39 Mode of actions • Ionizing radiation ejects electrons from an atom – High energy • Penetrates liquids and solids effectively • Nonionizing radiation raises atoms to a higher energy state – Lower energy than ionizing • Less penetration capability • Pyrimidine dimers form in DNA 40 The effects of ionizing and nonionizing radiation on DNA. 41 Fig. 11.7 Cellular effects of irradiation Ultraviolet (UV) radiation can cause the formation of pyrimidine dimers on DNA. Fig. 11.9 Formation of pyrimidine dimers by the action of UV radiation. 42 Applications • Ionizing radiation – Alternative sterilization method – Materials sensitive to heat or chemicals – Some foods (fruits, vegetables, meats) • Nonionizing radiation – Alternative disinfectant – Germicidal lamp in hospitals, schools, food preparation areas (inanimate objects, air, water) 43 A gramma radiation machine (ionizing radiation) used to sterilize fruits, vegetables, meats, fish, and spices. Fig. 11.8 Sterilization with Ionizing Radiation 44 A UV treatment system can be used to disinfect water. 45 Fig. 11.10 A UV treatment system for disinfection of water Filtration • Removes microbes and spores from liquids and air • Perforated membrane – Pore sizes vary • Applications – Liquids that are sensitive to heat • Serum, vaccines, media 46 An example of a filtration system. Fig. 11.11 Membrane filtration 47 Chemical control • Widely used agents • Applications 48 Example of chemical agents, their target microbe, level of activity, and toxicity. Table 11.5 Qualities of chemical agents used in health care 49 Applications • • • • • • • • • Halogens Phenolics Surfactants Hydrogen peroxide Detergents and soaps Heavy metals Aldehydes Gases Dyes, acids, and alkalis 50 Halogens • Chlorine – Disinfectant and antiseptic • Disrupt sulfhydryl groups in amino acids • Iodine – Topical antiseptic • Disruption is similar to chlorines 51 Phenolics • Vary based on functional groups attached to the aromatic ring • Examples: Hexachlorophene, Triclorsan – Microcidal – Ingredient in soaps to kitty litter • Disrupts cell walls and membranes, 52 Phenolics contain a basic phenolic aromatic ring with different functional groups. Fig. 11.12 Some phenolics 53 Alcohols • Ethyl alcohol, isopropyl (rubbing alcohol) – 70% concentration dissolve membrane lipids, disrupt cell surface tension, denatures proteins • Germicidal and skin degerming 54 Hydrogen peroxide • Colorless and caustic liquid • Form hydroxyl free radicals – Effective against anaerobes • Skin and wound cleaner • Quick method for sterilizing medical equipment 55 Examples of different devices used to sterilize medical equipment. Fig. 11.13 Sterile processing of invasive equipment protects patients. 56 Detergents and soaps • Quaternary ammonium – NH4 (quats) – Cationic – Bind and disrupt cell membrane – Low-level disinfectant in the clinical setting • Soaps – Fatty acids, oils, sodium or potassium salts – Cleaning agents – More effective if mixed with germicides 57 For detergents, the positive charge region binds bacteria and the uncharged region integrates into the cell membrane. Fig. 11.14 The structure of detergents 58 Comparison between nongermicidal and germicidal soaps. Fig. 11.15 Graph showing effects of handscrubbing 59 Heavy metals • Mercury, silver, – Inactivate proteins – Preservatives in cosmetics and ophthalmic solutions 60 Demonstration of the effects silver and gold have on microbial growth. Fig. 11.16 demonstration of the oligodynamic action of heavy metals. 61 Aldehydes • Glutaraldehyde – Crosslink with proteins on the cell surface – Disinfectant for surgical instruments • Formaldehyde – Formalin is used as a tissue preservative 62 Representation of the action of glutaraldehyde. Fig. 11.17 Actions of glutaraldehyde 63 Gases • Ethylene oxide – Reacts with functional groups of DNA and proteins – Sterilizes and disinfects plastic materials 64 Examples of different devices that use gas to sterilize equipment. Fig. 11.18 Sterilization using gas 65 Dyes • Crystal violet • Effective against Gram positive bacteria • Ointments 66 Acids and alkalis • Acetic acid • Ammonium hydroxide • Prevents spore germination and vegetative growth • Food preservative 67