Physical and Chemical Agents for Microbial Control

1. Physical and Chemical Agents for Microbial Control Chapter 11

2. Control of microbes Physical and chemical methods to destroy or reduce microbes in a given area

3. OVERVIEW 3

4. Why do we want to control microbes? • How do we do it?

5. Highest resistance Bacterial endospores Prions Moderate resistance Pseudomonas sp. Mycobacterium tuberculosis Staphylococcus aureus Protozoan cysts Least resistance most vegetative cells Fungal spores enveloped viruses Yeast Protozoan trophozoites Relative resistance of microbes

6. Methods that kill microbes • _______________ – a process that destroys all viable microbes, including viruses & endospores • Heat, sterilants • _______________– a process to destroy vegetative pathogens, not endospores • disinfectants or germicides – chemical - kills pathogenic microorganisms • _____________________ – destroy/inhibit vegetative pathogens on exposed body surfaces • Sepsis – infection/growth in body • Asepsis – preventing entry of pathogens and infection

7. Methods that reduce numbers • _______________– any cleansing technique that mechanically removes microbes and reduces numbers to safe levels • _______________– reduces the number of microbes on skin • ___________________________ – controls numbers by preventing growth (multipication)

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9. Microbial death • Permanent termination of an organism’s vital processes • microbiological definition: Involves permanent loss of reproductive capability, even under optimum growth conditions

10. Factors that influence microbial killing 10

11. How antimicrobial agents work: Cellular targets of control (mode of action) • Cell wall • Cell membrane • Cellular synthetic processes (DNA, RNA) • Proteins

12. mode of action Cell wall • Action: • Block synthesis • Digestion • Disrupt surface • w/o cell wall, bacteria will lyse (especially gram positive… remember?) • Examples: penicillin, detergents, alcohol

13. mode of action Surfactants disrupt

14. mode of action Affect on synthesis (DNA, RNA) • Proteins – have many functions in the cell! • Antimicrobials can block: DNA synthesis (master code), transcription, translation • Mutagens (radiation = permanent inactivation of DNA) • Antimicrobial therapy (drugs) • Chemicals – some destroy nucleic acids

15. mode of action Heat, pH, heavy metals can alter proteins

16. Practical concerns • Does the application require sterilization? • Is the item to be reused? (time, $$) • 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?

17. Types of Control: I. Methods of Physical Control • Heat • Cold temperatures • Desiccation • Radiation • Filtration

18. Physical Control 1. Heat Moist vs. dry • Moist heat = lower temp and shorter time • Causes coagulation/denaturation of protein • Dry heat = higher temp/longer time • Dehydrates cell, removes water, denatures proteins, oxidation (burning)

19. Physical Control

20. Physical Control 1. Heat – moist heat Moist heat uses hot water or steam • sterilization • _______________ 15 psi/121oC/10-40min (steam under pressure) • intermittent sterilization – 100oC 30-60 min for 3 days (unpressurized steam) • disinfection • Boiling at 100oC for 30 minutes to destroy non-spore-forming pathogens • _______________ kills Salmonella, Listeria & overall microbe count

21. Physical Control

22. Pasteurization Pasteurization – heat  kill potential agents of infection and spoilage without destroying the food flavor or value 63°C–66°C for 30 minutes (batch method) 71.6°C for 15 seconds (flash method) Not sterilization – kills non-spore-forming pathogens and lowers overall microbe count; does not kill endospores or many nonpathogenic microbes 22

23. Physical Control 1. Heat – dry heat Dry heat uses higher temperatures than moist heat, can also sterilize • incineration – 600-1200oC combusts & dehydrates cells • dry ovens – 150-180oC- coagulate proteins

24. Physical Control Thermal death Thermal death time (TDT) – shortest length of time required to kill all test microbes at a specified temperature Thermal death point (TDP) – lowest temperature required to kill all microbes in a sample in 10 minutes

25. Physical Control 2. Cold temperatures • _______________– slows the growth of microbes • refrigeration 0-15oC & freezing <0oC • used to ____________food, media and cultures

26. Physical Control 3. Desiccation • gradual removal of water from cells, leads to metabolic inhibition • not effective microbial control – many cells retain ability to grow when water is reintroduced • _______________= freeze-drying Note: cold and dessication ARE NOT good methods of disinfection or sterilization.

27. Physical Control 4. Radiation • _______________radiation – deep penetrating power, breaks DNA • gamma rays, X-rays, cathode rays • used to sterilize medical supplies & food products • _______________radiation – little penetrating power to sterilize air, water & solid surfaces • uv light creates thymine dimers, which interfere with replication

28. Ionizing radiation Physical Control

29. Physical Control Nonionizing radiation - UV

30. UV treatment of wastewater Physical Control

31. Physical Control Other “waves” • Sound (high frequency) • Can also be used to disrupt cells (vibrations) or generate heat • Ultrasonic devices are used clean dental, medical instruments before sterilization

32. Physical Control 5. Filtration • physical removal of microbes by passing a gas or liquid through filter • Pores of filter large enough for liquid but too small for microbe (<1 μm) • used to

33. OVERVIEW 33

34. Types of Control: II. Methods of Chemical Control Categories: • Halogens • Phenolics • Chlorhexidine • Alcohols • Hydrogen peroxide • Detergents & soaps • Heavy metals • Aldehydes • Uses: • Disinfectants • antiseptics • sterilants • degermers • preservatives

35. Chemical antimicrobials • 10,000 manufactured today • About 1,000 routinely used • Society is obsessed with “killing germs” – to the point of being excessive • Result: widespread overuse  resistance of pathogens, death of natural flora

36. Qualities of chemical antimicrobials • Rapid action, even in low concentration • Water/alcohol soluble, stabile • Broad spectrum w/o being toxic • Penetration, sustained action • Resitance to inactivation • Noncorrosive, nonstaining • Sanitizing and deodorizing • Inexpensive and available

37. Levels of activity

38. Factors that affect activity • Type of microorganism being treated • Material being treated • Amt of contamination • Exposure time • Strength/action of germicide • Appendix C shows procedures for testing effectiveness

39. Ways to express strength/concentration • Dilution (1:200 is one part chemical in 200 parts dilutant such as water) • ppm – parts per million • Percent – 70% alcohol, or mg/ml • _______________ solutions – have water as the solvent • _______________– are dissolved in alcohol

40. Chemical control - categories • Halogens • Phenolics • Chlorhexidine • Alcohols • Hydrogen peroxide • Detergents & soaps • Heavy metals • Aldehydes

41. 1. Halogens • Ionic (halide) or nonionic • Mostly _______________ • Germicidal and sporicidial with long exposure • Affect protein structure (bonds)

42. 1. Halogens • Chlorine – Cl2, hypochlorites (chlorine bleach - OCl), chloramines • In water – release hypochlorous acid (HOCl) • Denaturation of proteins by disrupting disulfide bonds • Can be sporicidal • 0.6-1 ppm Cl2 to clean water • Bleach – sanitization/disinfection

43. 1. Halogens • Iodine - I2, iodophors (betadine) • Denature proteins – similar to Chlorine but not affected by organic matter or pH • Broad spectrum microbicide, can be sporicidal • Milder medical & dental degerming agents, disinfectants, ointments, topical antiseptic • Betadine (iodophor) = iodine + neutral polymer; allows for slow release and increased penetration – used as antiseptic

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45. 2. Phenolics • Phenol ring (aromatic carbon ring) + groups • Disrupt cell membranes & precipitate (denature) proteins; bactericidal, fungicidal, virucidal, not sporicidal • _______________ • ____________antibacterial additive to soaps • Mouthwash! (thymol) • Can be VERY toxic! Not typically used as antiseptics.

47. _______________

48. 3. _______________ • Hibiclens, Hibitane • A surfactant & protein denaturant with broad microbicidal properties • Not sporicidal • Used as skin _________ agents for preoperative scrubs, skin cleaning & burns

49. 4. Alcohols • Ethyl, isopropyl in solutions of 50-90% (water needed for protein coagulation) • Act as surfactants dissolving membrane lipids and coagulating proteins of vegetative bacterial cells and fungi • _______________ • Isopropanol = rubbing alcohol, but vapors can be toxic

50. 5. Hydrogen peroxide • Weak (3%) to strong (25%) • _______________ agent (steals electrons) • Produce highly reactive hydroxyl free radicals that damage protein & DNA while also decomposing to O2 gas (bubbles) • toxic to _______________ , overwhelms catalase in aerobes • Strong solutions are sporicidal