This document outlines a lecture on controlling microbial growth. It defines terms like sterilization, disinfection, and antisepsis. It discusses conditions that influence the effectiveness of antimicrobial agents like population size, composition, concentration, and duration of exposure. Physical methods of control like heat and filtration are described. Heat can kill microbes through degradation of nucleic acids and denaturation of proteins. The use of heat to sterilize foods is discussed.
The Zero-ETL Approach: Enhancing Data Agility and Insight
Biology 120 lecture 5 2011 2012
1. CONTROL OF
MICROORGANISMS
Lecture 5
Thursday, September 15, 2011
2. LECTURE OUTLINE
Definition of Terms in Microbial Control
Pattern of Microbial Death
Conditions Influencing the Effectiveness of
Antimicrobial Agents
Use of Physical Agents
Use of Chemical Agents
Thursday, September 15, 2011
4. The Control of
Microbial Growth
SEPSIS
microbial contamination
ASEPSIS
absence of significant
contamination
Thursday, September 15, 2011
5. The Control of
Microbial Growth
SEPSIS
microbial contamination
ASEPSIS
absence of significant
contamination
Thursday, September 15, 2011
6. The Control of
Microbial Growth
STERILIZATION
Removal of all microbial life
COMMERCIAL STERILIZATION
Killing C. botulinum endospores
Thursday, September 15, 2011
7. The Control of
Microbial Growth
STERILIZATION
Removal of all microbial life
COMMERCIAL STERILIZATION
Killing C. botulinum endospores
Thursday, September 15, 2011
8. The Control of
Microbial Growth
DISINFECTION
Removal of pathogens
ANTISEPSIS
from living tissue
DEGERMING
from a limited area
Thursday, September 15, 2011
9. The Control of
Microbial Growth
DISINFECTION
Removal of pathogens
ANTISEPSIS
from living tissue
DEGERMING
from a limited area
Thursday, September 15, 2011
10. The Control of
Microbial Growth
SANITATION
Lower microbial counts on eating
utensils
BIOCIDE/GERMICIDE: Kills microbes
BACTERIOSTATS: Inhibiting, not
killing, microbes
Thursday, September 15, 2011
11. The Control of
Microbial Growth
SANITATION
Lower microbial counts on eating
utensils
BIOCIDE/GERMICIDE: Kills microbes
BACTERIOSTATS: Inhibiting, not
killing, microbes
Thursday, September 15, 2011
12. PATTERN OF MICROBIAL
DEATH
Bacterial
populations
die at a
constant
logarithmic
rate
Thursday, September 15, 2011
13. PATTERN OF MICROBIAL
DEATH
Bacterial
populations
die at a
constant
logarithmic
rate
Thursday, September 15, 2011
14. The Control of
Microbial Growth
HOW DO WE DECIDE
WHETHER THEY ARE
ACTUALLY DEAD?
Thursday, September 15, 2011
15. The Control of
Microbial Growth
HOW DO WE DECIDE
WHETHER THEY ARE
ACTUALLY DEAD?
Thursday, September 15, 2011
16. The Control of
Microbial Growth
HOW DO WE DECIDE
WHETHER THEY ARE
ACTUALLY DEAD?
“a microbe is defined DEAD if it does not
grow when inoculated into culture medium
that would normally support its growth”
Thursday, September 15, 2011
17. Conditions Influencing
Effectiveness of
Antimicrobial Agent Activity
1. Population size
Larger population requires a longer time to die
2. Population composition
Microorganisms vary markedly on
susceptibility
Vegetative versus Spores
Young versus Mature cells
Thursday, September 15, 2011
18. Conditions Influencing
Effectiveness of
Antimicrobial Agent Activity
1. Population size
Larger population requires a longer time to die
2. Population composition
Microorganisms vary markedly on
susceptibility
Vegetative versus Spores
Young versus Mature cells
Thursday, September 15, 2011
19. Conditions Influencing
Effectiveness of
Antimicrobial Agent Activity
3. Concentration or Intensity of an Antimicrobial
Agent
The more concentrated an agent the more
rapidly microbes can be destroyed
BUT sometimes an agent may be more
effective at lower concentrations (e.g. 70%
alcohol)
4. Duration of Exposure
The longer the exposure to an agent the more
they will be killed
Thursday, September 15, 2011
20. Conditions Influencing
Effectiveness of
Antimicrobial Agent Activity
3. Concentration or Intensity of an Antimicrobial
Agent
The more concentrated an agent the more
rapidly microbes can be destroyed
BUT sometimes an agent may be more
effective at lower concentrations (e.g. 70%
alcohol)
4. Duration of Exposure
The longer the exposure to an agent the more
they will be killed
Thursday, September 15, 2011
21. Conditions Influencing
Effectiveness of
Antimicrobial Agent Activity
5. Temperature
An increase in temperature at which a
chemical acts often enhances it activity
Example: acids used in high T = more
effective
6. Local environment
pH, organic matter, etc
Controls or Protects the pathogen
Thursday, September 15, 2011
22. Conditions Influencing
Effectiveness of
Antimicrobial Agent Activity
5. Temperature
An increase in temperature at which a
chemical acts often enhances it activity
Example: acids used in high T = more
effective
6. Local environment
pH, organic matter, etc
Controls or Protects the pathogen
Thursday, September 15, 2011
24. PHYSICAL METHODS:
HEAT
Fire and boiling
Sufficient to destroy vegetative
cells (10 minutes)
Not high for killing endospores
Disinfection but not
sterilization!
Thursday, September 15, 2011
25. PHYSICAL METHODS:
HEAT
Fire and boiling
Sufficient to destroy vegetative
cells (10 minutes)
Not high for killing endospores
Disinfection but not
sterilization!
Thursday, September 15, 2011
26. PHYSICAL METHODS:
HEAT
Thermal Death Point (TDP)
The lowest temperature at which a
microbial suspension in killed in 10 minutes
Thermal Death Time (TDT)
The shortest time needed to kill all
organisms in a microbial suspension at a
specific temperature and under defined
conditions
Thursday, September 15, 2011
27. PHYSICAL METHODS:
HEAT
Thermal Death Point (TDP)
The lowest temperature at which a
microbial suspension in killed in 10 minutes
Thermal Death Time (TDT)
The shortest time needed to kill all
organisms in a microbial suspension at a
specific temperature and under defined
conditions
Thursday, September 15, 2011
28. However, such a destruction is
logarithmic and it is theoretically NOT
POSSIBLE to “completely destroy”
microbes in a sample
Thursday, September 15, 2011
29. However, such a destruction is
logarithmic and it is theoretically NOT
POSSIBLE to “completely destroy”
microbes in a sample
Thursday, September 15, 2011
30. However, such a destruction is
logarithmic and it is theoretically NOT
POSSIBLE to “completely destroy”
microbes in a sample
Decimal Reduction Time or D value
Time required to kill 90% of the
microorganisms or spores in a sample at
a specified temperature
Time required for the line to drop by one
log cycle or tenfold
Used to estimate the relative resistance
of a microbe to different temperatures
Thursday, September 15, 2011
33. PHYSICAL METHODS:
HEAT
z Value
The increase in temperature required to
reduce D to 1/10 its value or to reduce it by
one log cycle
Thursday, September 15, 2011
34. PHYSICAL METHODS:
HEAT
z Value
The increase in temperature required to
reduce D to 1/10 its value or to reduce it by
one log cycle
F value
Time in minutes at a specific temperature
needed to kill a population of cells or spores
Usually 121°C
Thursday, September 15, 2011
37. APPLICATION: FOOD
INDUSTRY
After a food have been canned, it must be
heated to eliminate the risk of botulism arising
from the presence of Clostridium spores
Thursday, September 15, 2011
38. APPLICATION: FOOD
INDUSTRY
After a food have been canned, it must be
heated to eliminate the risk of botulism arising
from the presence of Clostridium spores
Example: (1012 to 100 spores)
IF the D value = 0.204 minutes
It would take 12D or 2.5 minutes to
reduce the spore number by heating at the
specified temperature
Thursday, September 15, 2011
41. APPLICATION: FOOD
INDUSTRY
If the z value for Clostridium spores is 10°C
Thursday, September 15, 2011
42. APPLICATION: FOOD
INDUSTRY
If the z value for Clostridium spores is 10°C
It takes a 10°C change in temperature to
alter the D value tenfold
Thursday, September 15, 2011
43. APPLICATION: FOOD
INDUSTRY
If the z value for Clostridium spores is 10°C
It takes a 10°C change in temperature to
alter the D value tenfold
Thursday, September 15, 2011
44. APPLICATION: FOOD
INDUSTRY
If the z value for Clostridium spores is 10°C
It takes a 10°C change in temperature to
alter the D value tenfold
THUS: if the cans are to be processed at
111°C rather than 121°C, the D value would
increase by tenfold t 2.04 minutes
Thursday, September 15, 2011
45. APPLICATION: FOOD
INDUSTRY
If the z value for Clostridium spores is 10°C
It takes a 10°C change in temperature to
alter the D value tenfold
THUS: if the cans are to be processed at
111°C rather than 121°C, the D value would
increase by tenfold t 2.04 minutes
The 12D value = 24.5 minutes
Thursday, September 15, 2011
55. HOW DOES HEAT KILL
MICROBES
Thursday, September 15, 2011
56. HOW DOES HEAT KILL
MICROBES
Thursday, September 15, 2011
57. HOW DOES HEAT KILL
MICROBES
MOIST HEAT
Kill effectively by degradation of nucleic
acids and by denaturation of enzymes and
other essential proteins
May also disrupt cell membranes
Thursday, September 15, 2011
58. HOW DOES HEAT KILL
MICROBES
MOIST HEAT
Kill effectively by degradation of nucleic
acids and by denaturation of enzymes and
other essential proteins
May also disrupt cell membranes
Thursday, September 15, 2011
59. HOW DOES HEAT KILL
MICROBES
MOIST HEAT
Kill effectively by degradation of nucleic
acids and by denaturation of enzymes and
other essential proteins
May also disrupt cell membranes
DRY HEAT
Microbial death results from the oxidation of
cell constituents and denaturation of proteins
Thursday, September 15, 2011
60. PHYSICAL METHODS:
FILTRATION
Applicable for heat-sensitive materials that needs
sterilization
Types of filters
Depth filters: consist of fibrous or granular
materials that have been bonded into a thick
layer filled with twisting channels of small
diameter
Membrane filters: porous membranes; 0.2 µm
pore sizes
Thursday, September 15, 2011
61. PHYSICAL METHODS:
FILTRATION
Applicable for heat-sensitive materials that needs
sterilization
Types of filters
Depth filters: consist of fibrous or granular
materials that have been bonded into a thick
layer filled with twisting channels of small
diameter
Membrane filters: porous membranes; 0.2 µm
pore sizes
Thursday, September 15, 2011
62. PHYSICAL METHODS:
FILTRATION
Laminar flow hood versus
biological safety cabinets (HEPA
filters)
High Efficiency Particulate Air
Remove 99.97% particles
0.02 µm
for sterilizing AIR
Thursday, September 15, 2011
63. PHYSICAL METHODS:
FILTRATION
Laminar flow hood versus
biological safety cabinets (HEPA
filters)
High Efficiency Particulate Air
Remove 99.97% particles
0.02 µm
for sterilizing AIR
Thursday, September 15, 2011
64. BIOSAFETY
CABINETS
Class1 (from room=outside)
protection: person and environment
Class 2 (Type A and B) )
Type A and B: product, person and
environment
difference: type A air is recirculated back
to room, type B exhausted outside the
building
Class 3: contained facility, higher level of
protection and containment
Thursday, September 15, 2011
65. BIOSAFETY
CABINETS
Class1 (from room=outside)
protection: person and environment
Class 2 (Type A and B) )
Type A and B: product, person and
environment
difference: type A air is recirculated back
to room, type B exhausted outside the
building
Class 3: contained facility, higher level of
protection and containment
Thursday, September 15, 2011
66. PHYSICAL METHODS:
RADIATION
IONIZING RADIATION
X rays, gamma rays, electron beams
Excellent as a sterilizing agent and penetrates
deep into objects
NON-IONIZING RADIATION
UV (about 260nm)
Quite lethal but does not penetrate glass, dirt
films, water and other substances very effectively
Microwaves: kill by heat not usually antimicrobial
Thursday, September 15, 2011
67. PHYSICAL METHODS:
RADIATION
IONIZING RADIATION
X rays, gamma rays, electron beams
Excellent as a sterilizing agent and penetrates
deep into objects
NON-IONIZING RADIATION
UV (about 260nm)
Quite lethal but does not penetrate glass, dirt
films, water and other substances very effectively
Microwaves: kill by heat not usually antimicrobial
Thursday, September 15, 2011
70. CHEMICAL
METHODS
PHENOLICS QUATERNARY
AMMONIUM
ALCOHOLS COMPOUNDS
ALDEHYDES
HALOGENS
STERILIZING
HEAVY METALS
GASES
Thursday, September 15, 2011
71. CHEMICAL
METHODS
PHENOLICS QUATERNARY
AMMONIUM
ALCOHOLS COMPOUNDS
ALDEHYDES
HALOGENS
STERILIZING
HEAVY METALS
GASES
Thursday, September 15, 2011
72. CHEMICAL
METHODS
Chemical agent Effectiveness against
Endospores Mycobacteria
Phenolics Poor Good
Quats None None
Chlorines Fair Fair
Alcohols Poor Good
Glutaraldehyde Fair Good
Thursday, September 15, 2011
73. CHEMICAL
METHODS
Chemical agent Effectiveness against
Endospores Mycobacteria
Phenolics Poor Good
Quats None None
Chlorines Fair Fair
Alcohols Poor Good
Glutaraldehyde Fair Good
Thursday, September 15, 2011
74. CHEMICAL
METHODS: Phenolics
First widely used antiseptic and disinfectant
Joseph Lister (1867): reduced the risk of
infection during operations
Example: LYSOLR
Act by denaturing proteins and
disrupting cell membranes
Thursday, September 15, 2011
75. CHEMICAL
METHODS: Phenolics
First widely used antiseptic and disinfectant
Joseph Lister (1867): reduced the risk of
infection during operations
Example: LYSOLR
Act by denaturing proteins and
disrupting cell membranes
Thursday, September 15, 2011
76. Disruption of
Cell Membranes
Thursday, September 15, 2011
77. CHEMICAL
METHODS: Phenolics
ADVANTAGES: effective in the
presence of organic material and remain
active on surfaces long after application
DISADVANTAGE: disagreeable odor
and can cause skin irritation and in
some instances brain damage
(hexachlorophene)
Thursday, September 15, 2011
78. CHEMICAL
METHODS: Phenolics
ADVANTAGES: effective in the
presence of organic material and remain
active on surfaces long after application
DISADVANTAGE: disagreeable odor
and can cause skin irritation and in
some instances brain damage
(hexachlorophene)
Thursday, September 15, 2011
79. CHEMICAL
METHODS: Alcohols
Widely used disinfectant and antiseptics
Bactericidal and fungicidal but not
sporicidal
May not destroy lipid-containing viruses
Thursday, September 15, 2011
80. CHEMICAL
METHODS: Alcohols
Widely used disinfectant and antiseptics
Bactericidal and fungicidal but not
sporicidal
May not destroy lipid-containing viruses
Thursday, September 15, 2011
81. DENATURES
PROTEINS,
DISSOLVES
LIPIDS
Thursday, September 15, 2011
82. CHEMICAL
METHODS: Alcohols
Example: ethanol and isopropanol (70-80%
concentration)
Act by denaturing proteins and possibly by
dissolving membrane lipids
10-15 soaking in alcohol is sufficient to
disinfect thermometers and small instruments
Thursday, September 15, 2011
83. CHEMICAL
METHODS: Alcohols
Example: ethanol and isopropanol (70-80%
concentration)
Act by denaturing proteins and possibly by
dissolving membrane lipids
10-15 soaking in alcohol is sufficient to
disinfect thermometers and small instruments
Thursday, September 15, 2011
84. CHEMICAL METHODS:
Halogens
Iodine
Kills by oxidizing cell constituents
and iodinating cell proteins
Kill spores at high concentrations
DISADVANTAGE: a stain may be left
(answer = iodophor)
Thursday, September 15, 2011
85. CHEMICAL METHODS:
Halogens
Iodine
Kills by oxidizing cell constituents
and iodinating cell proteins
Kill spores at high concentrations
DISADVANTAGE: a stain may be left
(answer = iodophor)
Thursday, September 15, 2011
86. CHEMICAL METHODS:
Halogens
Chlorine
Usually for water supply
Kills by oxidation of cellular materials and
destruction of vegetative bacteria, fungi
Will not kill spores
Death within 30 minutes
Thursday, September 15, 2011
87. CHEMICAL METHODS:
Halogens
Chlorine
Usually for water supply
Kills by oxidation of cellular materials and
destruction of vegetative bacteria, fungi
Will not kill spores
Death within 30 minutes
Thursday, September 15, 2011
88. CHEMICAL METHODS:
Heavy Metals
Mercury, Arsenic, Zinc, Copper
Used as germicides
How do they Kill:
Heavy metals combine with proteins,
often with their sulfhydryl groups and
inactivate them
May also precipitate cell proteins
Thursday, September 15, 2011
89. CHEMICAL METHODS:
Heavy Metals
Mercury, Arsenic, Zinc, Copper
Used as germicides
How do they Kill:
Heavy metals combine with proteins,
often with their sulfhydryl groups and
inactivate them
May also precipitate cell proteins
Thursday, September 15, 2011
90. CHEMICAL METHODS:
Quats
DETERGENTS
Amphipathic (both polar and non-polar
ends)
Kill by disrupting microbial membranes and
denature proteins
ADVANTAGE: stable, non-toxic
DISADVANTAGE: inactivated by hard
Thursday, September 15, 2011
91. CHEMICAL METHODS:
Quats
DETERGENTS
Amphipathic (both polar and non-polar
ends)
Kill by disrupting microbial membranes and
denature proteins
ADVANTAGE: stable, non-toxic
DISADVANTAGE: inactivated by hard
Thursday, September 15, 2011
93. CHEMICAL METHODS:
Aldehydes
FORMALDEHYDES
Very reactive molecules that
combine with proteins and inactivate
them
Sporicidal and can be used as
sterilants
Thursday, September 15, 2011
94. CHEMICAL METHODS:
Aldehydes
FORMALDEHYDES
Very reactive molecules that
combine with proteins and inactivate
them
Sporicidal and can be used as
sterilants
Thursday, September 15, 2011
95. EVALUATION OF
ANTIMICROBIAL AGENT
EFFECTIVENESS
PHENOL COEFFICIENT TEST
Best-known disinfectant screening test
Potency of a disinfectant is compared
with that of phenol
The highest dilution that killed bacteria
after a 10 minutes exposure are used to
calculate phenol coefficient
Thursday, September 15, 2011
96. EVALUATION OF
ANTIMICROBIAL AGENT
EFFECTIVENESS
PHENOL COEFFICIENT TEST
Best-known disinfectant screening test
Potency of a disinfectant is compared
with that of phenol
The highest dilution that killed bacteria
after a 10 minutes exposure are used to
calculate phenol coefficient
Thursday, September 15, 2011
97. CALCULATING PHENOL
COEFFICIENTS
The reciprocal of the appropriate test
disinfectant dilution is divided by that for phenol
to obtain the coefficient
Example: phenol dilution = 1/90 and the
maximum effective dilution for disinfectant X
= 1/450
Phenol coefficient = 5
Thursday, September 15, 2011
98. CALCULATING PHENOL
COEFFICIENTS
The reciprocal of the appropriate test
disinfectant dilution is divided by that for phenol
to obtain the coefficient
Example: phenol dilution = 1/90 and the
maximum effective dilution for disinfectant X
= 1/450
Phenol coefficient = 5
Thursday, September 15, 2011
99. CALCULATING PHENOL
COEFFICIENTS
The higher the phenol
coefficient value, the more
effective the disinfectant
under this conditions
Thursday, September 15, 2011
100. CALCULATING PHENOL
COEFFICIENTS
The higher the phenol
coefficient value, the more
effective the disinfectant
under this conditions
Thursday, September 15, 2011
101. DILUTION TESTS
Metal rings dipped in test bacteria are dried
Dried cultures placed in disinfectant for 10
min at 20°C
Rings transferred to culture media to
determine whether bacteria survived
treatment
Thursday, September 15, 2011
102. DILUTION TESTS
Metal rings dipped in test bacteria are dried
Dried cultures placed in disinfectant for 10
min at 20°C
Rings transferred to culture media to
determine whether bacteria survived
treatment
Thursday, September 15, 2011
107. CHEMOTHERAPEUTIC
AGENTS
Antibiotics are medicines used to treat infections
caused by bacteria only
Infections are usually caused by bacteria or viruses
Antibiotics, therefore, do not cure all infections
Many infections like the common cold, flu, mild sore
throat or diarrhea are caused by viruses
Thursday, September 15, 2011
108. CHEMOTHERAPEUTIC
AGENTS
Antibiotics are medicines used to treat infections
caused by bacteria only
Infections are usually caused by bacteria or viruses
Antibiotics, therefore, do not cure all infections
Many infections like the common cold, flu, mild sore
throat or diarrhea are caused by viruses
Thursday, September 15, 2011
110. WHAT IF ANTIBIOTICS WERE
USED INCORRECTLY?
No healing effect - If antibiotics are used for
viral infections, there will be no effect on the
illness
Antibiotic resistance - This occurs when one
antibiotic no longer works on a specific type of
bacteria
A stronger antibiotic will be needed to treat the
infection caused by this resistant strain of bacteria
Thursday, September 15, 2011
111. WHAT IF ANTIBIOTICS WERE
USED INCORRECTLY?
No healing effect - If antibiotics are used for
viral infections, there will be no effect on the
illness
Antibiotic resistance - This occurs when one
antibiotic no longer works on a specific type of
bacteria
A stronger antibiotic will be needed to treat the
infection caused by this resistant strain of bacteria
Thursday, September 15, 2011
112. ANTIBIOTIC
MECHANISMS
Thursday, September 15, 2011
113. ANTIBIOTIC
MECHANISMS
Thursday, September 15, 2011
122. DO YOU CONTRIBUTE
TO RESISTANCE?
Another factor that contributes to resistance is
that when patients are prescribed antibiotics for a
just cause, many do not finish their medication
This allows resistant bacteria to survive more
easily
The practice of saving unused medication to
treat themselves or others at a later date can
also lead to resistant strains
Thursday, September 15, 2011
125. DO YOU CONTRIBUTE
TO RESISTANCE?
Also contributing to antibiotic resistance is the
widespread use of antibiotics to promote weight
gain and to control disease in cattle, pigs, and
chickens
Forty to fifty percent of antibiotics produced are
used in livestock feed
This leads to an increase of resistant bacteria
in these animals, which is then spread to
humans
Thursday, September 15, 2011
126. ANY
QUESTIONS???
Thursday, September 15, 2011
127. NEXT MEETING:
INTERACTIVE LECTURE/
QUIZ ON METABOLISM
Thursday, September 15, 2011