1. Control of Microbial Growth
Chapter 7
Control of Microbial Growth
Siti Sarah Jumali

2. Terminology Sepsis refers to microbial contamination.
Asepsis is the absence of significant contamination.
Antisepsis: Removal of pathogens from living tissue
Degerming: Removal of microbes from a limited area
Sanitization: Lower microbial counts on eating utensils

3. Terminology cont’d Decontaminated
- item that has been treated to reduce # of disease causing organisms
Preservation
- delaying spoilage of foods
Sterilization, Disinfection, Antiseptic, Bacteriocidal, Bacteriostatic

4. Terms used: Sterilization vs. Disinfection
Sterilization: destroying all forms of life
Disinfection: destroying pathogens or unwanted organisms
Disinfectant vs. Antiseptic
Disinfectant: antimicrobial agent used on inanimate objects
Antiseptic : antimicrobial agent used on living tissue

6. cidal vs. static Biocide/Germicide: Kills microbes
Bacteriostasis: Inhibiting, not killing, microbes
Examples:
- Bactericidal - kills bacteria
- Bacteriostasis (Bacteriostatic)- inhibits bacterial growth
- Fungicidal
- Fungistatic
- Algacidal
- Algastatic

7. Methods of control Physical or chemical?
physical control includes heat, irradiation, filtration and mechanical removal
Chemical control involves the use of microbial chemicals
Depends on the situation
degree of control required

8. Methods of control cont’d
Daily life
- Cooking
- refrigeration
- cleaning
- soap water
mechanical
Mechanical and chemical

9. Methods of control cont’d
Hospitals
Important to minimize nosocomial infection (hospital acquired infection) due to
- weakened patients’ condition
- breaching of intact skin
- high concentration of pathogens from patients and workers
Sterile condition

10. Methods of control cont’d
Microbiology lab utilizes
Sterile equipment
Aseptic technique
And possesses workers who takes care of the nature of of microbiologists (GLP)

11. Methods of control cont’d
Foods/food production industry
- physical removal
- adding chemicals
- may result in toxicity
- clean surface/ machinery

12. Selection of Control Method
Antimicrobial procedure used for control of microbial growth is based on
Types of microbe
Extent of contamination
Environmental conditions
Potential risk

13. Selection of Control Method cont’d
Types of microorganism
- some organisms are more resistant and require stronger measures for control
- endospores require chemical treatment for hours
- Mycobacterium’s waxy cells are resistant to chemicals
Mycobacterial cell wall: 1-outer lipids, 2-mycolic acid, 3-polysaccharides (arabinogalactan), 4-peptidoglycan, 5-plasma membrane, 6-lipoarabinomannan (LAM), 7-phosphatidylinositol mannoside, 8-cell wall skeleton

14. Selection of Control Method cont’d
Extent of microbial population
- larger population take more time to destroy
- usually 90% of the population is destroyed in
a given period
e.g if in 1st 3 minutes 90% of the population is destroyed, then 90% of the remaining population gets destroyed in the next 3 minutes and so on

16. Selection of Control Method cont’d
Environmental conditions
- pH, temperature
- presence of
- organics: blood
- dirt
- grease
- the potential risk of transmitting infectious agents
- critical items
- semicritical items
- non-critical items
Must be cleaned first, then controlled

17. Selection of Control cont’d
Critical items have
- indirect contact with body tissues
- needles, scalpels
Semicritical items have contact with
- mucous membranes but it does not penetrate
endoscopes, endotrachial tubes

18. Selection of Control cont’d
Non-critical items have
- indirect contact with unbroken skin
- countertops, stethoscopes

19. Methods to Control Microbial Growth
Physical Methods
Heat Sterilization
Dry Heat
Incineration
Moist Heat
Boiling
Autoclaving
Pasteurization
Filtration
Air
Liquid
Radiation
UV Light
Ionizing Radiation

20. Physical Microbial Controls: Heat
Heat as a microbial control
- fast, reliable, inexpensive
- does not introduce potential toxic substances
Types of heat control include
- moist heat
- pasteurization
- pressurized steam
- dry heat

21. Physical Microbial Controls: Heat cont’d
Moist heat
- causes irreversible coagulation of proteins found in microorganisms
- 10 minutes of boiling
- most microbes and viruses will be destroyed except endospores and few others which can survives hours of boiling

22. Physical Microbial Controls: Heat cont’d
Pasteurization
- reduces number of heat sensitive organisms
- widely used in milk and juices
increases shelf life and does not alter quality
- original pasteurization was 62ºC, 30 mins
- now: UHT-shorter time 72ºC, 15 secs

23. Physical Microbial Controls: Heat cont’d
Pressurized steam
- pressure cooker or autoclave
- higher air pressure increases the temperature
at which steam forms
- 15 psi (lbs/square inch) at 121ºC for 15 mins
-effective to kill endospores

24. The autoclave: Moist heat and pressure
15psi, 121ºC, 15 minutes
Thermal death point (TDP): Lowest temperature at which all cells in a culture get killed in 10 mins
Thermal death time (TDT): time to kill all cells in a culture
Decimal reduction time (DRT): Minutes to kill 90% of a population at a given temperature

25. The autoclaving machine

26. Temperature of steam and Pressure at sea level

27. The autoclaving machine

28. Physical Microbial Controls: Heat cont’d
Dry heat
- without moisture e.g. flaming
- burns cell constituents
- object is oxidized to ash
- irreversibly denatures proteins
- takes longer (200ºC, 1.5 hrs dry=121ºC, 15 min moist)
- advantages are for powders, does not corrode metals and blunt sharps
- e.g flasks, tubes, pipettes in microbiological laboratories.

29. Physical Microbial Controls: Filtration
Used for heat sensitive fluids
air

30. Physical Microbial Controls: Filtration cont’d
Fluids
solutions of antibiotics, vitamins, tissue extracts, animal serum, etc.
Depth filters
–able to retain microorganisms while allowing fluids to pass through
Membrane Filter
- The use of graded pore size µm

31. Physical Microbial Controls: Filtration cont’d
Air
- HEPA (High Efficient Particulate Air) filter and laminar air flow are commonly used
- filter incoming air and outgoing air respectively
- HEPA filter prevents the income of 0.3µm and large size particles to enter.

32. Physical Microbial Controls: Radiation
“Cold sterilization“ for disposable materials made up of plastics, wool, cotton, etc without altering the material.
Radiation damages DNA
Ionizing radiation (X-rays, gamma rays, electron beams)
Non-ionizing radiation
- UV
- Microwaves kill by heat not especially
antimicrobial

34. Physical Microbial Controls: Radiation cont’d
Gamma irradiation
- penetrate deeply
- for heat sensitive materials
- causes biological damage to microorganisms
- does not alter food flavor (meat)

35. Physical Microbial Controls: Radiation cont’d
UV light
damages the structure and function of nucleic acids
Penetrate poorly- cannot penetrate even into liquid.
Used to disinfect surfaces
Can cause damage to human cells
Germicidal lamps -kill or reduce the number of viable microorganisms to sterilize microbiological laboratories hospital operating rooms, and specific filling rooms in various industries

36. Physical Microbial Controls: Radiation cont’d
Microwave
Kills by heat
Does not affect microorganisms directly

37. Physical Methods used to control Microbial growth

42. Chemical Microbial Control
Chemical Method
Gas sterilants
Ethylene oxide
Antiseptics and disinfectants
Germicidal chemical

44. Chemical Microbial Control cont’d
Grouped according to potency
Sterilants
High-level
Intermediate level
Low level

45. Chemical Control sterilants
destroy microorganisms, endospores and viruses
used for critical equipment-scalpels

46. chemical control cont'd
high level
- destroy viruses and  vegetative microorganisms (no endospores)
- used for semicritical equipment: endoscopes
intermediate level
- destroy vegetative microorganisms, some viruses
- used for non-critical equipment: sthetoscopes
low level
-destroy fungi, vegetative microorganisms
- used for general purpose disinfectants

47. Selecting Germicidal Chemical
Germicide: An agent capable of killing pathogens and non-pathogens but not necessarily endospores
toxicity to human or environment?
- weigh the benefits vs the risks
presence of organic material
- hypochlotrite is inactivated by the presence of organic matter
compatibility
-electrical equipment with a liquid??
residue
- some have to be rinsed with sterile water
cost and availability

48. Selecting germicidal Chemical cont'd
storage and stability
- may come in concentrated form for ease in storing
- those have to be mixed
environmental risk
- is neutralization necessary before disposal?

49. Classes of Germicidal Chemicals
alcohols alcohols
aldehydes
biguanides
ethylene oxide 
halogens: oxidize proteins

50. Classes of Germicidal Chemicals
alcohols
- coagulated enzymes and proteins
- damage lipid membranes
- on-toxic
- inexpensive
- no residue

51. Classes of Germicidal Chemicals cont’d
aldehydes
- inactivate proteins and nucleic acids
- toxic to humans
Peroxygens
- oxidizing agents
- hydrogen peroxide
- leaves no residue

52. Classes of Germicidal Chemicals cont’d
biguanides
- extensive antiseptic use
- adheres and persists on skin, mucous membranes
- low toxicity
Phenolic compounds
Hitorically important
Irritant, unpleasant odor
Destroy cytoplasmic membrane and denatures protein

53. Classes of Germicidal Chemicals cont’d
ethylene oxide 
- reacts with proteins
- gas: penetrable 
- mutagenic
Metal compounds
Interfere protein function
Toxic
pollutants

54. Classes of Germicidal Chemicals cont’d
halogens: oxidize proteins
-chlorine
-irritating to skin
-organic compounds consume free chlorine
-iodine
-tincture
- Iodophore

55. Effect of germicidal activity on Microbes
Cytoplasmic membrane
Biguanides
Phenolics
Quats
DNA
Ethylene oxides
Aldehydes
Proteins
Alcohols
Halogens
Metals
Ozone
Peroxygens

56. Effect of germicidal activity on Microbes

57. Chemical methods of microbial control
Evaluating a disinfectant
- Disk diffusion method

58. Preservation of Perishables
Extends shelf life
Slow or halts microbial growth thus delaying spoilage

59. Preservation of Perishables
Chemical preservatives
Some chemical preservatives are used in non-food items
Food preservatives must be non-toxic to humans
Benzoic acid, propionic acids, nitrate are commonly used
Nitrate
Inhibits germination of C. botulinum endospores

60. Preservation of Perishables
Low temperature storage
Temperature dependent
most microorganisms do not reproduce in ordinary refrigerator (0-7ºC)
Freezing
ice crystals can cause irreversible damage to many microorganisms (kills up to 50% growth)
Freezing stops all growth, but may start to reproduce again once food is thawed

61. Preservation of Perishables
Reducing water
Salt/sugar
Draw water out of cell
Less available for microorganisms
Drying
Desiccation
Removing water such as milk powder
Salt cured meat

62. Preservation of Perishables
Lyophilization
Freeze drying
Freeze food first
followed by putting in vacuum

63. Factors that influence effectiveness
Number of microbes
Environmental influences
Time of exposure
Microbial characteristics

64. Microbial death curve

65. Limitations: Microbial Characteristics

66. Actions of Microbial Control Agents
Alteration of membrane permeability
Damage to proteins
Damage to nucleic acids

67. Questions????
Sand rich in salts n nitrates preserved mummy