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Physical and Chemical Control of Microorganisms I. Terms II. Factors which determine the effectiveness of control methods III. Methods of physical control IV. Chemical agents Terms 1) “Control” -- Limiting exposure to _______________________ agents Microorganisms are part of our environment Infection can be controlled by limiting exposure to infectious agents 2) _______________ -- Removal or destruction of all _________________ microorganisms. “sterile” 3) Disinfection -- Removal or destruction of pathogens from ________________________ areas. “disinfectant” 4) Antisepsis -- Removal or destruction of vegetative forms from _______ _____________. “antiseptic” Note: Antiseptics and disinfectants do the same thing, but in different environments Terms (continued) 5) Sanitization -- any cleansing technique that mechanically removes microorganisms. Usually soaps More terms 6) “-cide” refers to ______________. “Bacteriocide” kills bacteria, “fungicide” kills fungi, etc. 7) “-static” means ‘to stand still’. Refers to an agent which prevents _______________ but doesn’t kill. “Bacteriostatic” or “fungistatic” or “virustatic” agent Factors which determine the effectiveness of a control method 1. The number of microorganisms “Death curve” 107 106 # of 105 ________ 104 cells Note log scale on y axis (each increment is 10X reduction in numbers) 103 102 101 100 1 2 3 4 5 Time (min.) 6 7 8 107 106 105 104 103 102 101 100 1 2 3 4 5 Time (min.) 6 7 8 Decimal reduction time (DRT) -- Time for a _____ reduction in population size. Here DRT = ______ If the goal is to reduce numbers to 102 then the size of the initial population will determine how long it will take, in this case, ____________ minutes. Factors which determine the effectiveness of a control method (cont.) Spore formers vs. non-spore formers 107 106 # of 105 viable 4 cells 10 103 102 101 100 __________________ Vegetative cells 1 2 3 4 5 Time (min.) 6 7 8 Factors which determine effectiveness of a control agent (cont.) • Temperature and _______ of the environment • ___________________ of the agent • Mode of action of the agent • Microbistatic vs. microbicidal Cellular target Presence of ________________ compounds or inhibitors of the control agent Methods of Physical Control: Heat _________ heat Hot water, boiling water, steam 60° - 135° C Denatures proteins, nucleic acids __________ heat Ovens 160° - >1000° C Denatures, oxidizes At lower temperatures, moist heat is __________ effective than dry heat Moist heat methods I. Boiling water -- ________ min. boiling kills all vegetative cells II. Pasteurization -- Heating to temp. below boiling to kill specific pathogens and increase shelf-time Batch method -- 63°-66° for 30 min. ___________ method -- 71.6° for 15 sec. Results in killing of most viruses and 97-99% reduction in veg. stages of bacteria and fungi UHT (Ultra High Temp) -- 134° 1-2 ___________. Milk can last up to 3 months unrefrigerated Primary pathogen targets of pasteurization: ______________, Listeria, Mycobacterium, Campylobacter, Brucella Moist heat methods (cont.) III. Steam under pressure Highest temp. of moist heat at sea level -- _________° C If ____________ increased, higher temps. can be reached Autoclaving Commonly: _______° C at 15 PSI ________ min. at this temp. is very effective Effects on cellular proteins Text, Fig. 11.4 Physical methods: radiation Physical methods: radiation ____________ -- knocks electrons off atoms --> ions e.g. gamma rays, x-rays Damages DNA and proteins by breaking bonds -- Fig. 11.8 Exposure hazard to humans Penetrates ___________ and liquids Used in fruits & vegetables, now meats Non-ionizing -- excites atoms but doesn’t ionize e.g. ____________, sunlight Damages DNA by creating T-T dimers -Fig. 11.8, 11.10 Relatively safe to use Doesn’t penetrate solids or liquids well ______________ sterilization, water treatment Text, Fig. 11.8 Text, Fig. 11.10 Other physical control methods Sonication -- Disruption using ___________ waves Filtration Text, Fig. 11.13 Chemical control agents I. Many kinds but most function by: Disrupting Altering __________________ ________________ and/or nucleic acid structure II. Effectiveness is determined by: Concentration Contact time Major groups • • • • • • ________________ Phenol and its derivatives Alcohols Hydrogen peroxide Detergents Heavy ___________ Halogens • Mainly _______________, also iodine and fluorine • Iodine “iodophors” most common iodine compound used in hospitals: Betadine, Povidone, etc. Less ____________ than free iodine due to slow release of free iodine. • 3 Chlorine forms: Cl2 (chlorine gas) OCl (hypochlorite -- common ‘bleach’) NH2Cl (chloramines) In solution, these compounds combine with water to form hypochlorous acid (HOCl). This is the active form which destroys by ____________ Effective against: bacteria, fungi, spores, viruses Limitations: ineffective at _____________ pH ______________: light and O2 cause breakdown Phenols p-cresol (lysol) phenol OH OH CH3 Hexachlorophene (phisohex) OH Cl OH CH2 Cl Cl Cl Cl Cl Phenolics affect protein function and/or disrupt membranes Alcohols • Ethanol -- 70% to 95% • Isopropanol -- more microbicidal but also more __________ Destroy cell membranes, can coagulate proteins Hydrogen peroxide (H2O2) Forms _______________ free radicals (e.g. superoxides, hydroxyl radicals: see last lecture) which are toxic to all cells Also breaks down to H2O and O2, therefore is strongly effective against _____________. Detergents All solubilize membranes and disrupt proteins Text, Fig. 11.3 Detergents (cont.) Two types: ___________ (charged) and non-ionic Non-ionic and anionic detergents (like soaps) are not very microbicidal, although they may be very effective in sanitization. _____________ detergents (like benzalkonium chloride) are the most effective ionic detergents. Heavy metals Mainly only mercury and silver preparations are used now. Merthiolate, mercurochrome, silver nitrate, etc. Form ions which complex with cell components, stopping growth Disadvantages: _________________ _________________ Microbes can develop ___________________