1. Microbiology: A Systems Approach, 2nd ed.
Chapter 11: Physical and Chemical Control of Microbes

2. 11.1 Controlling Microorganisms
General Considerations in Microbial Control
Sterilization: the destruction of all microbial life
Disinfection: destroys most microbial life, reducing contamination on inanimate surfaces
Antisepsis: destroys most microbial life, reducing contamination on a living surface
Decontamination: the mechanical removal of most microbes from an animate or inanimate surface

3. Figure 11.1

4. Relative Resistance of Microbial Forms
Primary targets of microbial control: microorganisms that can cause infection or spoilage that are constantly present in the external environment
Contaminants that need to be controlled
Bacterial vegetative cells and endospores (so resistant, the goal is sterilization)
Fungal hyphae and spores
Yeasts
Protozoan trophozoites and cysts
Worms
Viruses
Prions

6. Terminology and Methods of Microbial Control
Sterilization
Removes all viable microorganisms including viruses
Material is said to be sterile
Usually reserved for inanimate objects
Mostly performed with heat
Sometimes chemicals called sterilants are used

7. Disinfection
The use of a physical process or chemical agent (disinfectant) to destroy vegetative pathogens
Does not destroy bacterial endospores
Usually used only on inanimate objects
Also removes toxins
5% bleach solution

8. Antisepsis
Antiseptics applied directly to exposed body surfaces to destroy or inhibit vegetative pathogens
Sepsis: the growth of microorganisms in the blood and other tissues
Asepsis: any practice that prevents the entry of infectious agents into sterile tissues

9. The Agents Versus the Processes
–cide: to kill
Bactericide: chemical that destroys bacteria (not endospores)
Fungicide: a chemical that can kill fungal spores, hyphae, and yeasts
Virucide: a chemical that inactivates viruses
Sporicide: can destroy bacterial endospores
Germicide and microbicide: chemical agents that kill microorganisms
Stasis and static: to stand still
Bacteristatic: prevent the growth of bacteria
Fungistatic: inhibit fungal growth
Microbistatic: materials used to control microorganisms in the body, for example

10. Decontamination
Used when actual sterilization isn’t needed but need to decrease the risk of infection or spoilage (ex. food industry)
Sanitization: any cleansing technique that mechanically removes microorganisms to reduce contamination to safe levels
Sanitizer: compound such as soap or detergent that sanitizes
Sanitary: may not be free from microbes but are safe for normal use
Degermation: reduces the numbers of microbes on the human skin (ex. alcohol wipes)

11. Practical Concerns in Microbial Control
Does the application require sterilization, or is disinfection adequate?
Is the item to be reused or permanently discarded?
If it will be reused, can it withstand heat, pressure, radiation, or chemicals?
Is the control method suitable for a given application?
Will the agent penetrate to the necessary extent?
Is the method cost- and labor-efficient, and is it safe?

12. What is Microbial Death?
When various cell structures become dysfunctional and the entire cell sustains irreversible damage
If a cell can no longer reproduce under ideal environmental conditions
Death begins when a certain threshold of microbicidal agent is met, and continues in a logarithmic manner

13. Figure 11.2

14. Factors that Influence the Action of Antimicrobial Agents
The number of microorganisms
The nature of the microorganisms in the population
The temperature and pH of the environment
The concentration of the agent
The mode of action of the agent
The presence of solvents, interfering organic matter, and inhibitors

15. How Antimicrobial Agents Work: Their Modes of Action
The Cell Wall
Block its synthesis
Digest it
Break down its surface
The cell becomes fragile and is lysed easily
The Cell Membrane

16. Figure 11.3

17. Protein and Nucleic Acid Synthesis
Binding to ribosomes to stop translation
Bind irreversibly to DNA preventing transcription and translation
Mutagenic agents

18. Protein Function
Figure 11.4

19. 11.2 Methods of Physical Control
Heat as an Agent of Microbial Control
Generally, elevated temperatures are microbicidal and lower temperatures are microbistatic
Can use moist heat or dry heat

21. Heat Resistance and Thermal Death of Spores and Vegetative Cells

22. Practical Concerns in the Use of Heat: Thermal Death Measurements
Temperature and length of exposure must be considered
Higher temperatures generally allow shorter exposure times; lower temperatures generally require longer exposure times
Thermal death time (TDT): the shortest length of time required to kill all test microbes at a specified temperature
Thermal death point (TDP): the lowest temperature required to kill all microbes in a sample in 10 minutes

23. Common Methods of Moist Heat Control
Steam under pressure
Pressure raises the temperature of steam
Autoclave is used
Most efficient pressure-temperature combination for sterilization: 15 psi which yields 121°C

24. Figure 11.5

25. Nonpressurized Steam Intermittent sterilization or tyndallization
Expose to free-flowing steam for minutes, incubate for hours, treat again; repeat for 3 days in a row

26. Pasteurization Used to disinfect beverages
Heat is applied to liquids to kill potential agents of infection and spoilage, while retaining the liquid’s flavor and food value
Special heat exchangers
Flash method: expose to 71.6°C for 15 seconds
Batch method: expose to 63°C to 66°C for 30 minutes
Does not kill endospores or thermoduric microbes

27. Boiling Water For disinfection and not sterilization
Expose materials to boiling water for 30 minutes

28. Dry Heat: Hot Air and Incineration
Ignites and reduces microbes to ashes and gas
Common practice in microbiology lab- incineration on inoculating loops and needles using a Bunsen burner
Can also use tabletop infrared incinerators

29. Figure 11.6

30. Dry Oven Usually an electric oven
Coils radiate heat within an enclosed compartment
Exposure to 150°C to 180°C for 2 to 4 hours
Used for heat-resistant items that do not sterilize well with moist heat

31. The Effects of Cold and Desiccation
To slow growth of cultures and microbes in food during processing and storage
Cold does not kill most microbes; freezing can actually preserve cultures
Desiccation: dehydration of vegetative cells when directly exposed to normal room air
Lyophilization: a combination of freezing and drying; used to preserve microorganisms and other cells in a viable state for many years

32. Radiation as a Microbial Control Agent
Radiation: energy emitted from atomic activities and dispersed at high velocity through matter or space
For microbial control:
Gamma rays
X rays
Ultraviolet radiation

33. Modes of Action of Ionizing Versus Nonionizing Radiation
Ionizing radiation: if the radiation ejects orbital electrons from an atom causing ions to form
Nonionizing radiation: excites atoms by raising them to a higher energy state but does not ionize them

34. Figure 11.7

35. Ionizing Radiation: Gamma Rays, X Rays, and Cathode Rays
Cold sterilization
Dosage of radiation- measured in Grays
Exposure ranges from 5 to 50 kiloGrays
Gamma rays, most penetrating; X rays, intermediate; cathode rays, least penetrating

36. Applications of Ionizing Radiation
Food products
Medical products

37. Nonionizing Radiation: Ultraviolet Rays
Wavelength approximately 100 nm to 400 nm
Germicidal lamp: 254 nm
Not as penetrating as ionizing radiation
Powerful tool for destroying fungal cells and spores, bacterial vegetative cells, protozoa, and viruses

38. Figure 11.9

39. Applications of Ultraviolet Radiation
Usually disinfection rather than sterilization
Hospital rooms, operating rooms, schools, food prep areas, dental offices
Treat drinking water or purify liquids

40. Figure 11.10

41. Decontamination by Filtration: Techniques for Removing Microbes
Effective for removing microbes from air and liquids
Fluid strained through a filter with openings large enough for fluid but too small for microorganisms
Filters are usually thin membranes of cellulose acetate, polycarbonate, and a variety of plastic materials
Pore size can be controlled and standardizes

42. Figure 11.11

43. Applications of Filtration
Prepare liquids that can’t withstand heat
Can decontaminate beverages without altering their flavor
Water purification
Removing airborne contaminants (HEPA filters)

44. 11.3 Chemical Agents in Microbial Control
Approximately 10,000 different antimicrobial chemical agents are manufactured
Approximately 1,000 used routinely in health care and the home
Occur in liquid, gaseous, or solid state
Tinctures: solutions dissolved in pure alcohol or water-alcohol mixtures

45. Choosing a Microbicidal Chemical
Rapid action even in low concentrations
Solubility in water or alcohol and long-term stability
Broad-spectrum microbicidal action without being toxic to human and animal tissues
Penetration of inanimate surfaces to sustain a cumulative or persistent action
Resistance to becoming inactivated by organic matter
Noncorrosive or nonstaining properties
Sanitizing and deodorizing properties
Affordability and ready availability

46. Factors that Affect the Germicidal Activity of Chemicals
Nature of microorganisms being treated
Nature of the material being treated
Degree of contamination
Time of exposure
Strength and chemical action of the germicide

48. Germicidal Categories According to Chemical Group
Halogen Antimicrobial Chemicals
Fluorine, bromine, chlorine, and iodine
Microbicidal and sporicidal with longer exposure
Chlorine compounds: liquid and gaseous chlorine, hypochlorites, chloramines
Kills bacteria and endospores
Also kills fungi and viruses
Example: Household bleach
Iodine compounds: free iodine and iodophors
Topical antiseptic
Disinfectant

49. Phenol and its Derivatives
Phenol coefficient: compares a chemical’s antimicrobic properties to those of phenol
High concentrations: cellular poisons
Lower concentrations: inactivate certain critical enzyme systems

50. Chlorhexidine
Complex organic base containing chlorine and two phenolic rings
Targets cell membranes and protein structure
At moderate to high concentrations, it is bactericidal for both gram-positive and gram-negative bacteria but inactive against spores
Mild, low toxicity, rapid action

51. Alcohols as Antimicrobial Agents
Only ethyl and isopropyl alcohols are suitable for microbial control
Mechanism of action depends in part upon its concentration
50% and higher dissolve membrane lipids, disrupt cell surface tension, and compromise membrane integrity
50% to 90% denatures proteins through coagulation; but higher concentration does not increase microbicidal activity
100% (absolute alcohol) dehydrates cells and inhibits their growth
Does not destroy bacterial spores at room temperature but can destroy resistant vegetative forms
More effective in inactivating enveloped viruses than nonenveloped viruses

52. Hydrogen Peroxide and Related Germicides
Germicidal effects are due to the direct and indirect actions of oxygen
Oxygen forms hydroxyl free radicals which are highly toxic and reactive to cells
Bactericidal, virucidal, and fungicidal
In higher concentrations is sporicidal

53. Chemicals with Surface Action: Detergents
Act as surfactants
Anionic detergents have limited microbicidal power
Cationic detergents are more effective because the positively charged end binds well with the predominantly negatively charged bacterial surface proteins
Soaps are weak microbicides but gain germicidal value when mixed with agents such as chlorhexidine or iodine

54. Figure 11.14

55. Figure 11.15

56. Heavy Metal Compounds Hg, Ag, Au, Cu, As, and Zn have been used
Only Hg and Ag still have significance as germicides
Oligodynamic action: having antimicrobial effects in exceedingly small amounts
Bind onto functional groups of proteins and inactivating them
Drawbacks to using metals in microbial control:
Can be very toxic to humans
Often cause allergic reactions
Large quantities of biological fluids and wastes neutralize their actions
Microbes can develop resistance to them

57. Figure 11.16

58. Aldehydes as Germicides
–CHO functional group on the terminal carbon
Glutaraldehyde and formaldehyde (formalin- aqueous solution)- most often used in microbial control

59. Figure 11.17

60. Gaseous Sterilants and Disinfectants
Ethylene oxide (ETO)
Propylene oxide
Chlorine dioxide

61. Dyes as Antimicrobial Agents
Primary source of certain drugs used in chemotherapy
Aniline dyes (crystal violet and malachite green) are very active against gram-positive species of bacteria and various fungi
Yellow acridine dyes (acriflavine and proflavine) sometimes used for antisepsis and wound treatment
Limited applications because they stain and have a narrow spectrum of activity

62. Acids and Alkalis
Very low or high pH can destroy or inhibit microbial cells
Limited in applications due to their corrosive, caustic, and hazardous nature