Control of
Microorganisms
Microbiology
Effected in two basic ways:
1. Biocidal action: Killing Microorganisms
2. Biostatic action :By inhibiting the Growth of
Microorganisms
Usually involves the use of:
1. Physical Agents that Kill or Prevent Growth
2. Chemical Agents that Kill or Prevent Growth
3. Biological Agents: Bacteria, Fungi, Phage
etc.
Control of Microbial Growth
Level of Resistance
1.
2.
3.
4.
5.
5.
6.
5.
Endospores (Most)
Parasites
Mycobacteria
Fungal Spores
Small Non-enveloped Viruses
- Polio, Rotavirus, Rabies
Vegetative Fungal Cells
Enveloped Viruses
- Herpes, Hepatitis B & C, HIV
Vegetative Bacteria (Least)
Principles of Microbial
Control
Prevention / Control of Growth to Prevent
Infection and Spoilage, Asepsis
 Sterilization / Complete Destruction
- Real sterilization is All or Nothing)

 Commercial Sterilization
The best we can do
Reduce numbers to safe level
- Utilizes
a. Heat
b. Radiation
c. Chemicals
d. Physical Removal
e. Hyperbaric (High pressure)






Commercial Sterilization / Heat Treatment of
Canned Foods
Disinfection / Reducing Growth Nonliving
Surfaces
Antisepsis / Reducing Growth Living Tissue
Antibiotics ( “magic bullets”) Internal use
Preservatives
Sanitize – Subject to High Temperature
(70oC) Washing (Dishwashers) or Hypo
Principles of Microbial Control
Cide – Suffix Meaning to Kill
 Stat – Suffix Meaning to Inhibit
 Sepsis – Bacterial Contamination
 Asepsis – Lack of Bacterial Contamination

Principles of Microbial Control
Rate of Microbial Death


Bacteria Usually Die At a Constant Rate
Plotted Logarithmically This Will Give a Straight
Line
Time to Kill in Proportion to
the Population Size
Large Numbers Require Greater Time
 Small Numbers Require Less Time

Susceptibilities Vary
Endospores are Difficult to Kill
 Organic Matter May Interfere with
Heat Treatments and Chemical
Control Agents
 Must clean first



Alteration of Membrane Permeability
- Susceptibility of membrane is due to its
lipid and protein composition
- Control Agents can alter permeability
Damage to Proteins and Nucleic Acids
- Break hydrogen and covalent bonds in
proteins
- Interfere with DNA, RNA, Protein
Synthesis
Control Agents Act By:
Physical Methods of
Microbial Control






Heat
Filtration
Low Temperatures
Desiccation
Osmotic Pressure
Radiation
Physical Methods
Most Frequent and Widely Used.
 Always Consider
1. Type of Heat
2. Time of Application
3. Temperature
 Endospores are the most heat resistant of
all cells.

Heat





Moist Heat / Denatures Enzymes
Thermal Death Point (TDP) / Lowest
Temp to Kill All the Bacteria in a Broth in 10
Minutes
Thermal Death Time (TDT) / Time Span
Required to Kill All the Bacteria in a Broth at
a Given Temperature
Decimal Reduction Time (DRT) / Length
of Time in Which 90% of a Bacterial
Population will be Killed at a given
Temperature
Boiling / Kills Many Vegetative Cells and
Inactivates Viruses Within 10 Minutes
Moist Heat Sterilization
Boiling
 Autoclaving

Boiling
• 100° C for 30 Minutes
• Kills Everything Except Some Endospores
• Intermittent boiling can kills spores
Boil, incubate, boil,
incubate, boil
Does not always work
Autoclaving
Steam Under
Pressure
 121° C for 15
Minutes at
15 lb/in2
 Heat-labile
Substances
will be
Denatured
 Steam Must
Contact the
Material

 thermal
death time (TDT)
◦ shortest time needed to kill all
microorganisms in a suspension at a
specific temperature and under defined
conditions
 decimal
reduction time (D or D value)
◦ time required to kill 90% of
microorganisms or spores in a sample at
a specific temperature
Measuring heat-killing efficiency
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18
Figure 7.1
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19
Other measures…

Z value
◦ increase in
temperature
required to
reduce D by
1/10

F value
◦ time in minutes
at a specific
temperature
needed to kill a
population of
cells or spores
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required for reproduction or
display.
Figure 7.2
20
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reproduction or display.
21
Moist heat
Autoclaves
– used to kill endospores efficiently
– use saturated steam under pressure
to reach temperatures
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above boiling
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22

pasteurization
◦ controlled heating at temperatures well below
boiling
◦ reduces total microbial population and thereby
increases shelf life of treated material
Moist heat…
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reproduction or display.
23

flash pasteurization (high temperature
short-term – HTST)
◦ 72°C for 15 seconds then rapid cooling

ultrahigh-temperature (UHT) sterilization
◦ 140 to 150°C for 1 to 3 seconds
Pasteurization of milk
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24
Cook Chill F values
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Cook Chill
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26
Dry Heat Sterilization
Direct Flaming
 Incineration
 Hot-Air Sterilization
(Oven)

Flaming the Loop
 Flaming the loop
helps to prevent
contamination of
the bacteria.
 When flaming the
loop, make sure
that all of the wire
has been heated to
redness.
Incineration
• Burns and Physically Destroys Organisms
• Used for
a. Needles
b. Inoculating Wires
c. Glassware
d. Body Parts?
Dry Heat (Hot Air Oven)
• 160° C for 2 Hours or 170° C for 1 hour
• Used for
a. Glassware
b. Metal
c. Objects That Won’t Melt
Pasteurization
A High Temperature
Is Used For a Short
Time
 Batch Method (LTLT)
63 °C for 30 Minutes
 Flash Method (HTST)
72 °C for 15 Seconds
 Ultra-HighTemperature is 140 °C
for 3 seconds

Filtration
The passage of a liquid or gas through a filter
with pores small enough to retain microbes.
 Especially important to sterilize solutions
which would be denatured by heat (antibiotics,
injectable drugs, amino acids, vitamins.)


High-Efficiency Particulate Air Filters
1. Operating Rooms
2. Burn Units
3. Fume Hoods
HEPA Filters
Decreasing Temperature
Decreases Chemical Activity
 Low Temps are Not Bactericidal
 Refrigeration <50 C

◦ Listeria still grows but slow
◦ Spoilage organisms still grow

Freezing -180 C
◦ Spoilage due to enzymes
Low Temperatures
Disrupts Microbial Metabolism
 Stops Growth / Still Viable
 Freeze-drying / Dehydration

◦ Used to preserve bacteria

Viruses and Endospores
Can Resist Desiccation
Desiccation
Plasmolysis
 Sugar Curing / Salting
 May Still Get Some Mold or Yeast Growth

◦ Staphylococcus and Listeria grows in 10% salt
Osmotic Pressure
Radiation
Acts By Destroying DNA or Damaging
It.
 Dependent on the Wavelength,
Intensity, and Duration


Ionizing
Radiation
1. High Degree of
Penetration
2. Examples
- Gamma Rays
- X-rays
- High Energy
Electron
Beams
Radiation

Ultraviolet Radiation
1. Nonionizing
2. Low Degree of
Penetration
3. Low Penetration
4. Harmful / Skin / Eyes
5. Cell Damage /
Thymine
6. Germicidal / 260 nm

Kill Microbes Indirectly with Heat
Microwaves









Phenols and Phenolics
Halogens
Alcohols (70% ethanol) (iso-propanal)
Heavy Metals and Their Compounds
Surface-Active Agents
Quaternary Ammonium Compounds
Chemical Food Preservatives
Aldehydes
Antibiotics
Chemical Control Methods
Two Conditions Influence the
Effectiveness of Chemical
Disinfectants
Type of Microbe
- G+ More Susceptible to
Disinfectants
- Pseudomonands Can Grow in
Disinfectants and Antiseptics
- M. tuberculosis is Resistant to Many
Disinfectants
- Endospores Most Resistant
 Environment
- Organic Matter and Increased Temp

Evaluating a Disinfectant

Old Standard is the Phenol Coefficient
Test
(FYI -- The phenol coefficient is the value
obtained by dividing the highest dilution
of the test solution by the highest dilution
of phenol that sterilizes the given culture
of bacteria under standard conditions of
time and temperature.)
In Lab We Use Soaked Filter
Papers
Antibiotic Sensitivities
Types of Disinfectants

Phenol and Phenolics
- Another Name for Carbolic Acid /
Lysol
- Joseph Lister
- Exert Influence By
1. Injuring Plasma membranes
2. Inactivating Enzymes
3. Denaturing Proteins
- Long Lasting, Good for Blood and
Body
Fluids, No Effect on Spores
Types of Disinfectants

Halogens
- Can be Used Alone or in Solution
- Chlorine -- Purifies Drinking Water
a. 2-4 Drops of Chlorine per Liter / 30
Min
b. Forms an Acid Which is Bactericidal
c. Disinfectant in Gaseous Form or in
Solution as Calcium Hypochlorite
- Iodine – combines with Amino Acids
a. Inactivates Enzymes
b. Tincture / Alcohol
c. Iodophor / Organic Molecule /
Betadine

Alcohols
- Denature Proteins
- Dissolve Lipids
- Tinctures
- Wet Disinfectants
a. Aqueous Ethanol (60% 95%)
b. Isopropal Alcohol
Types of Disinfectants
Types of Disinfectants

Heavy Metals and Their Compounds
- Used for Burn Treatment
- Prevents Neonatal Gonorrheal
Opthalmia
Silver nitrate
- Denature Proteins
Types of Disinfectants

Surface-Active Agents
- Decrease Molecular Surface
Tension
- Include Soaps and Detergents
- Soaps Have Limited Germicidal
Action but
Assist in the Removal of
Organisms by
Scrubbing
- Acid-Anionic Detergents / Dairy
Types of Disinfectants

Quaternary Ammonium
Compounds (QUATS)
- Cationic Detergents Attached to
NH4+1
- Disrupt Plasma Membranes
- Most Effective on Gram-Positive
Bacteria
- Mouthwashes and Sore Throat
Remedies

Chemical Food Preservatives
- Sorbic Acid
- Benzoic Acid
InhibitFungus
- Propionic Acid
- Nitrate and Nitrite Salts / Meats /
To Prevent Germination of
Clostridium
botulinum endospores
Types of Disinfectants

Aldehydes
- Formaldehyde
- Glutaraldehyde
- Most Effective of all Chemical
Disinfectants
- Carcinogenic
- Oxidize Molecules Inside Cells
Types of Disinfectants
Types of Disinfectants

Gases
◦
◦
◦
◦
◦
◦
◦
◦
Ethylene oxide
Ozone
Sulfur dioxide,
chlorine,
glutaraldehyde,
methylene chloride,
formaldehyde,
ammonia
Types of Disinfectants
Antibiotics
- Used to Preserve
Cheese
- Tomato products pH
> 4.5
- Nisin
 Stock feeds

 Promote faster growth
Growing Problem
 Indiscriminant and Inappropriate Use
 Super Bugs
1. Methicillin Resistant S. aureus
2. Vancomycin Resistant Enterococcus
3. Multidrug Resistant M. tuberculosis
 Important to test Sensitivities

Antibiotic Resistance