1. Control of microorganisms Many bacteria cause disease and food spoilage Need exists to kill or inhibit the growth of these bacteria

2. Frequently used terms Sterilization - removal or destruction of all living cells, viable spores, viruses and viriods Disinfection - removal or destruction of pathogens (spores and some other microorganisms remain) Sanitization - reduction of microbial population to safe levels Antisepsis - prevention of infection (accomplished by antiseptics)

3. Frequently used terms Bactericide - substance that kills bacteria Bacteriostatic - substance that prevents growth of bacteria

4. Pattern of microbial death Microorganisms usually die logarithmically (i.e. the population will be reduced by the same fraction at regular intervals)

5. Pattern of microbial death Microorganisms usually die logarithmically (i.e. the population will be reduced by the same fraction at regular intervals)

6. Conditions influencing effectiveness of antimicrobial agents Population size Population composition Concentration or intensity of agent Duration of exposure to agent Temperature Local environment (e.g. pH, presence of organic material)

7. Physical methods of control Heat Low temperature Filtration Radiation

8. Measuring heat-killing efficiency Thermal death point (TDP) - lowest temperature at which all microorganisms in a suspension are killed in 10 minutes Decimal reduction time (D value) - time it takes to kill 90% of the cells or spores in a sample at a specific temperature

9. Decimal reduction time (D value)

10. Measuring heat-killing efficiency Z value - the increase in temperature required to reduce D to 1/10 its value F value - time in minutes at a specific temperature required to kill a population of spores or cells

11. Measuring heat-killing efficiency

12. Moist heat Effective against all types of microorganisms Degrades nucleic acids, denatures proteins, and disrupts membranes

13. Moist heat Autoclaves are used to kill endospores Uses steam under pressure to achieve temperatures above boiling

14. Pasteurization Controlled heating at temperatures below boiling Does not sterilize Kills pathogens and reduces levels of spoilage microorganisms First used to preserve wine, now used for milk, beer, juice and other beverages

15. Pasteurization Traditional method: 63 ºC for 30 minutes Flash pasteurization: 72 ºC for 15 seconds

16. Ultrahigh temperature (UHT) sterilization Sometimes used by the milk industry Milk heated at 140 to 150 ºC for 1 to 3 seconds Products can be stored at room temperature for 1 to 2 months

17. Dry heat sterilization Less effective, requiring higher temperatures and longer exposure times Does not corrode metal and glass instruments Oxidizes cell constituents and denatures proteins

18. Low temperature Refrigeration- storage at 4 ºC slows microbial growth (only used for short-term storage) Freezing- storage at - 20 ºC stops microbial growth (does not kill microorganisms) Freezing at -30 to -70 ºC used to preserve microbial samples

19. Filtration Can be used to sterilized or reduce the microbial population of heat-sensitive liquids Removes microorganisms rather than destroying them Solutions often forced through filters by pressure or a vacuum

20. Depth filters Thick layers of fibrous or granular material with small diameter channels Made of various materials (e.g. diatomaceous earth or asbestos)

21. Membrane filters Porous membrane about 0.1 mm thick Entrap microbes on their surface Pore size of 0.2 um diameter removes most cells but not viruses

22. Air filtration Surgical masks Cotton plugs on culture vessels

23. Air filtration Laminar flow biological safety cabinets Employ high efficiency particulate air (HEPA) filters Remove 99.97 % of particles larger than 0.3 um Air forced through filter forms a curtain of sterile air across the opening of the cabinet

24. Laminar flow biological safety cabinets

25. Radiation Ultraviolet (UV) radiation Ionizing radiation

26. Ultraviolet (UV) radiation Wavelength of about 260 nM Lethal but does not penetrate glass Used to sterilize air or exposed surfaces Can sterilize water in a thin layer Burns the skin

27. Ionizing radiation Penetrates deep into objects Not always effective against viruses Gamma radiation from Cobalt 60 often used Used to treat meat, fruits, vegetables and spices

28. Chemical agents Are most commonly used agents for disinfection and antisepsis

29. Phenolics Phenol first used by Lister Phenol and derivatives used as disinfectants in hospitals and labs Effective in the presence of organic material Can cause skin irritation

30. Phenolics Act by denaturing proteins and disrupting cell membranes

31. Alcohols Not effective against spores or lipid-containing viruses Ethanol and isopropanol most commonly used (at 70- 80 %) Act by denaturing proteins and possibly dissolving membrane lipids

32. Halogens Include fluorine, chlorine, bromine, iodine and astatine Iodine used as a skin disinfectant Chlorine used to disinfect water Both act by oxidizing cell material and iodinating or chlorinating molecules

33. Aldehydes Formaldehyde and gutaraldehyde are the most commonly used Are highly reactive molecules Inactivate proteins and DNA by cross-linking alkylating molecules

34. Quaternary ammonium compounds (QUATS) Are a class of detergents Consist of a positively charged quaternary nitrogen with a long hydrophobic chain Act by disrupting membranes and denaturing proteins

35. Sterilizing gases Ethylene oxide (EtO) is used to sterilize heat-sensitive material Kills by combining with proteins Rapidly penetrates packing material

36. Sterilizing gases Betapropiolactone (BPL) is occasionally used Less difficult to eliminate and kills more quickly Does not penetrate well and may be carcinogenic

37. Heavy metals Ions of mercury, silver, arsenic, zinc and copper were used for many years as germicides (less toxic and more effective compounds have generally replaced them) Silver nitrate can be used in the eyes of newborns (antibiotics more common now) Silver sulfadiazine used on burns Copper sulfate used as an algicide

38. Heavy metals Act by combining with proteins and inactivating or precipitating them

39. Evaluation of antimicrobial agent effectiveness The Environmental protection agency (EPA) regulates disinfectants The Food and Drug Administration (FDA) regulates agents used on humans and animals

40. Phenol coefficient Potency of disinfectant compared to phenol Coefficient greater than 1 indicates agent is more potent than phenol Not always indicative of potency during normal use

41. Other evaluation methods Use dilution test Measures rate at which selected bacteria are destroyed by various chemical agents In use test Testing done using conditions that approximate normal use of disinfectant