CHAPTER 3: MICROBIAL GROWTH AND CONTROL bacterium into two daughter cells, in a

process called binary fission. Providing

INTRODUCTION
no mutational event occurs, the
Early civilization practiced salting. Smoking, resulting daughter cells are genetically
pickling, drying, and exposure of food and identical to the original cell
clothing to sunlight to control microbial growth.
Use of spices in cooking was to mask taste of
spoiled food. Some spices prevented spoilage.
In mid 1800s Semmelweis and Lister helped
MICROBIAL GROWTH
developed aseptic techniques to prevent
contamination of surgical wounds
● Increase in number of cells rather an
BEFORE THEN increase in size

● Nosocomial infections caused death in ● Understanding the requirements for

10% of surgeries microbial growth

● Up to 25% mothers delivering in the ● Allows us determine how to control the

hospitals died due to infection growth of microbes

● Specially of those microbes that cause
CONTROL OF MICROBIAL GROWTH
disease and food spoilage
● Means to inhibit or prevent growth of BACTERIAL GROWTH
microorganisms
● This control is affected in two basic ● What is growth rate of bacteria?

ways ● The growth rate of bacteria formula is

1. By killing microorganisms defined as the rate of exponential
growth of a bacterial culture which is
2. By inhibiting the growth of microorganism expressed as generation time, also the
CONTROLLING MICROBIAL GROWTH doubling time of bacterial population
and represented as G = T/n or
● Is important in the medical field, generation time = Time/No. of
generation
pharmaceutical and biotechnology
industries, academic research, and food BACTERIAL GROWTH RATE
industry
1.Lag Phase
● Chemical agents that can eliminate or
suppress microbial life are separated in ● Phase when bacteria absorb nutrients,
different groups based on their use and synthesize enzyme and prepare for
reproduction.
CONTROL OF MICROORGANISMS
2. Logarithmic Growth Phase or Exponential Growth
● Is essential in order to prevent the Phase
transmission of diseases and infection,
stop decomposition and spoilage, and ● Phase when bacteria multiply so rapidly
prevent unwanted microbial that the population number doubles
contamination with each generation time
● Microorganisms are controlled by Generation Time - time between the formation of a
means of physical agents and chemical new bacterium and its division into two daughter cells.
magnets Growth rate is greatest during the logarithmic phase.
CONTROL GROWTH 3. Stationary Phase

● Usually involves the use of physical or ● Phase when rate of division slows and
chemical agents which either kill or the number of bacteria dividing equals
prevent the growth of microorganisms the number of dying.
● Agents which kill cells are called cidal ● It is during this phase that its culture
agents maintains its greatest population
● Agents which inhibits the growth of density

cells (without killing them) are referred 4. Death Phase or Decline Phase
to as static agents
● microorganism die at rapid rate
● Example: Bacterial growth is the asexual
because of overriding, the toxic waste
reproduction, or cell division, of a
 products increase and nutrients and is the destruction or removal of infections or
oxygen supply decreases harmful microorganism from non-living things
by physical or chemical means
CONTROL OF MICROBIAL GROWTH: Definitions
example: Pasteurization – use of disinfectants
Sterilization: and use of antiseptic
Note: Disinfection is not sterilization because
Killing or removing all forms of microbial life (including not all microbes are destroyed in disinfection.
endospores) in a material or an object. Heating is the
most commonly used method of sterilization. Disinfectants

Heating -most commonly used method of sterilization. chemicals used to disinfect inanimate objects
like bedside and OR equipment
Commercial Sterilization: example: Cydex, chlorox, phenol, carbolic acid,
Heat treatment that kills endospores of Clostridium eresol, xylenols and orthophenylphenol
botulinum the causative agent of botulism, botulism (in
canned food). Sepsis:
Does not kill endospores of thermophiles, which
are not pathogens and may grow temperature ● Comes from Greek for decay or putric.
above 45oC
● Indicates bacterial contamination
Does not kill just preventing

CONTROL OF MICROBIAL GROWTH Asepsis:

1. STERILIZATION ● Absence of significant contamination

The complete destruction of all living organisms ● Septic techniques are used to prevent
including cells, viable spore and viruses
contamination of surgical instruments, medical
Sterilization can be accomplished by:
personnel, and the patient during surgery
1. Heat ● Septic techniques are also used to prevent
2. Heat and steam under pressure bacterial contamination in food industry
3. Gas (ethylene acide) Bacteriostatic Agents:
4. Chemicals (formaldehyde)
⮚ An agent that inhibits the growth of bacteria,
5. Radiation of the ultraviolet rays or gamma rays but does not necessarily kill them. Suffix stasis:
to stop or steady. Germicide: an agent that kills
certain microorganisms
Disinfection:
Bactericide:
Reducing the number of pathogenic
microorganisms so they can no longer cause ⮚ An agent that kills bacteria, most do not kill
diseases. Usually involves the removal of endospores
vegetative ornonendospore forming pathogens
Viricide:
MAY USE PHYSICAL OR CHEMICAL METHODS:
⮚ An agent that inactivates viruses
Disinfectants

chemicals used to disinfect inanimate objects Fungicide:

Applied to inanimate objects
⮚ An agent that kills fungi
Antiseptic:
Sporocide:
● Applied to living tissue (antisepsis).
⮚ An agent that kills bacterial endospores of
Degerming:
fungal spores
Mechanical removal of most microbes in a
Rate of Microbial Death When bacterial populations are
limited area
heated or treated antimicrobial chemicals, they usually
Example: alcohol swab on skin
die at a constant rate
Sanitation

Use of chemical agent on food-handling

Several factors influence the effectiveness of
equipment meet public health standards and
antimicrobial treatment:
minimize chances of disease transmission.
Example: Hot soap & water
⮚ 1. Number of microbes:
Disinfection o The more microbes present, the
more time it takes to eliminate
population
 Moist Heat (Continued):
⮚ 2. Type of Microbe
o Endospores are very difficult to ● Reliable sterilization with moist heat requires
destroy. Vegetative pathogens
temperatures above that of boiling water
vary widely in susceptibility to
different methods of microbial Autoclave:
control
● Chamber which is filled with hot steam under
⮚ 3. Environmental influences: pressure. Preferred method of sterilization,
o Presence of organic material unless material is damaged by heat, moisture,
(blood, feces, saliva) tends to or high pressure
inhibit antimicrobials, Ph. ● Autoclave: Closed chamber with high
temperature and pressure
● Temperature of steam reaches 121oC at twice
⮚ 4. Time of Exposure:
atmospheric pressure
o Chemical antimicrobials and
radiation treatments are more ● Most effective when organisms contact steam
effective at longer times. In directly or are contained in a small volume of
heat treatments, longer liquid
exposure compensates for
● All organisms and endospores are killed within
lower temperatures.
15 minutes
PHYSICAL METHODS OF MICROBIAL CONTROL:
Pasteurization:
* Heat:
● Developed by Louis Pasteur to prevent the
● Kills microorganisms by denaturing their
spoilage of beverages. Used to reduce microbes
enzymes and other proteins. Heat resistance responsible for spoilage of beer, milk, wine,
varies widely among microbes. juices, etc.
* Thermal Death Point (TDP): Classic Method of Pasteurization:

● Lowest temperature at which all of the ● Milk was exposed to 65oC for 30 minutes
microbes in a liquid suspension will be killed in
ten minutes. High Temperature Short Time Pasteurization (HTST):

* Thermal Death Time (TDT): ● Used today milk is exposed to 72oC for 15
seconds
● Minimal length of time in which all bacteria will
be killed at a given temperature. Ultra-High Temperature Pasteurization (UHT):

* Decimal Reduction Time (DRT): ● Milk is treated at 1400oC for 3 seconds and
then cooled very quickly in a vacuum chamber
● Time in minutes at which 90% of bacteria at a
● Advantage: milk can be stored several months
given temperature will be killed. Used in
canning industry at room temperature for months

* Moist Heat: Dry Heat:

● Kills microorganisms by coagulating their ● Kills by oxidation effects

proteins. In general, moist heat is much more
Direct Flaming:
effective than dry heat.

* Boiling: ● Used to sterilized noculating loops and needles

● Heat metal until it has a red glow

● Heat to 100 dc or more at sea level. Kills
vegetative forms of bacterial pathogens, almost Incineration:
all viruses, and fungi and their spores within 10
minutes or less. Endospores and some viruses ● Effective way to sterilize disposable items
are not destroyed this quickly. However brief (paper cups, dressings) and biological waste
boiling will kill most pathogens.
Hot Air Sterilization:
● Hepatitis virus: Can survive up to 30
minutes of boiling. ● Place objects in an oven, requires 2 hours at
● Endospores: Can survive up to 20 hours or 1700oC for sterilization.
more of boiling.
 ● Dry heat is transfers heat less effectively to a ● Clostridium spp., and bacillus spp.: may survive
cool body, than moist heat for decades

Filtration: Osmotic Pressure:

● Removal of microbes by passage of a liquid or ● using high concentrations of salts and sugars in
gas through a screen like material with small foods is used to increase the osmotic pressure
pores. and create a hypertonic environment.
● Used to sterilize heat sensitive materials like Plasmolysis:
vaccines, enzymes, antibiotics, and some
culture media ● As water leaves the cell, plasma membrane
shrinks away from cell wall. Cell may not die,
High Efficiency Particulate Air (HEPA):
but usually stops growing in plants
● Used in operating rooms and burn units to ● Yeast and molds: more resistant to high osmotic
remove bacteria form air pressures

Membrane Filters: ● Staphylococci spp. That live on skin are fairly

resistant to high osmotic pressure
● Uniform pore size, used in industry and
A. Radiation: Three types of radiation kill microbes:
research
1. Ionizing Radiation:
● Different sizes; 0.22 and 0.45µm pores

● Used to filter most bacteria ● gamma rays, X rays, electron beams, or higher
energy rays
Low Temperature:
● Have short wavelength (less than 1 nanometer)
● Effect depends on microbes and treatment
● Dislodge electrons from atoms and form ions.
applied
Cause mutations in DNA and produce peroxides
Refrigeration: ● Used sterilize pharmaceuticals and disposable
medical supplies
● Temperatures from 0 to 7oC, Bacteriostatic
effect. Reduces metabolic rate of most ● Food industry is interested in using ionizing
microbes so they cannot reproduce or produce radiation
toxin. U freezing temperatures below 0Oc ● Disadvantages: Penetrates human tissues. May
Flash Freezing: cause genetic mutations in humans

2. Ultraviolet light (Non-ionizing Radiation):

● Does not kill most microbes, slow freezing,
more harmful because ice crystal disrupt cell ● Wavelength is longer than 1 nanometer.
structure
Damages DNA by producing thymine dimers,
● Over a third of vegetative bacteria may survive which cause mutations
1 year ● Used to disinfect operating rooms, nurseries,
● Most parasites are killed by a few days of cafeterias
freezing ● Disadvantages: Damages skin, eyes. Doesn’t
Desiccation: penetrate paper, glass, and cloth

3. Microwave Radiation:
● In the absence of water, microbes, cannot grow
or reproduce, but some may remain viable for ● Wavelength ranges from 1 millimeter to 1
years
meter
● After water becomes available, they start
● Heat is absorbed by water molecules. May kill
growing again
vegetative cells in moist foods
● Susceptibility to desiccation varies widely:
● Bacterial endospores, which do not contain
● Neisseria gonorrhea: Only survives about one water, are not damaged by microwave radiation
hour ● Solid foods are unevenly penetrated by
● Mycobacterium tuberculosis: may survive microwaves
several months
B. CHEMICAL METHODS OF MICROBIAL CONTROL
● Viruses are fairly resistant to desiccation
TYPES OF DISINFECTANT’S
1. Phenols and Phenolics:
● Evaporate, leaving no residue

● Phenol (carbolic acid) was first used by Lister as ● Used to mechanically wipe microbes off skin
a disinfectant. before injections or blood drawing
● Rarely used today because it is a skin irritant ● Not good for open wounds, because cause
and has strong odor protein to coagulate
● Used in some throat sprays and lozenges ● Ethanol: drinking alcohol. Optimum

● Acts as local anesthetic concentration is 70%

● Isopropanol: Rubbing alcohol. Better
● 1.2 Phenolics are chemical derivatives of phenol
disinfectant than ethanol. Also cheaper and less
● 1.2 1 Cresols: Derived from coal tar (Lysol) volatile

● 1.2 2 Biphenols (pHisoHex): Effective against 4. Heavy Metals:

gram-positive staphylococci. Used in nurseries.
● Include copper, selenium, mercury, silver,
Excessive use in infants may cause neurological
damage. Destroy plasma membrane and and zinc.
denature proteins. ● Oligodynamic action: Very tiny amounts are
● Advantages: stable, persist for long times after effective
applied, and remain active in the presence of
organic compounds ● A.) Silver:

2. Halogens: ● 1% silver nitrate used to protect infants

against gonorrheal eye infections until
● Effective alone or in compounds. recently
● B.) Mercury:
● A. Iodine:
● Organic mercury compounds like
● Tincture of iodine (alcohol solution) was one of
Merthiolate the mercurochrome are used
first antiseptics used to disinfect skin wounds
● Combines with amino acid tyrosine in proteins ● C.) Copper:
and denatures proteins
● Copper sulfate is used to kill algae in pools
● Stains skin and clothes, somewhat irritating
and fish tanks
● Iodophors: Compounds with iodine that are ● D.) Selenium:
slow releasing, take several minutes to act.
Used to skin antiseptic in surgery. Not effective ● Kills fungi and their spores. Used for fungal
against bacterial endospores. Example: infections. Also used in dandruff shampoos
Betadine or Lodine
● E.) Zinc:
● B. Chlorine: ● Zinc chloride is used in mouthwashes
● When mixed in water forms hypochlorous acid: ● Zinc oxide is used as antifungal agent in
Cl2 + H2O 🡪 H+ + Cl- + HOCI paints

5. Quaternary Ammonium Compounds (Quats):

● Hypochlorous acid used to disinfect drinking
water, pools, and sewage ● Widely used surface active agents
● Chlorine is easily inactivated by organic
● Cationic (positively charge) detergents
materials
● Sodium hypochlorite (Na OCI): Is active ● Effective against gram positive bacteria, less

ingredient of bleach effective against gram-negative bacteria

● Chloramines: Consist of chlorine and ammonia. ● Also destroy fungi, amoebas, and enveloped

Less effective as germicides viruses:

● Zephiran, Cepacol, also found in our lab spray
3. Alcohols:
bottles
● Kill bacteria, fungi, but not endospores or naked ● Pseudomonas strains that are resistant and can
viruses grow in presence of Quats are a big concern in
● Act by denaturing proteins and disrupting cell hospitals
membranes
● Advantages:
 ● Strong antimicrobial action, colorless, odorless, ● Oxidize cellular components of treated
tasteless, stable, and non-toxic microbes.
● Disrupt membranes and proteins
● Disadvantages:

● Form foam ● A.) Ozone:

● Organic matter interferes with effectiveness. ● Used along with chlorine to disinfect water
Neutralized by soaps andanionic detergents
● Helps neutralize unpleasant tastes and odors
6. Aldehydes:
● More effective killing agent than chlorine, but
● Include some of the most effective less stable and more expensive
antimicrobials. ● Highly reactive from of oxygen
● Inactive proteins by forming covalent cross links ● Made by exposing oxygen to electricity or UV
with several functional groups light

● A.) Formaldehyde Gas: ● B.) Hydrogen peroxide:

● Excellent disinfectant ● Used as an antiseptic
● Commonly used as formalin, a 37% aqueous ● Not good for open wounds because quickly
solution. broken down by catalase present in human cells
● Formalin was used extensively to preserve ● Effective in disinfection of inanimate objects
biological specimens and inactivate viruses and
bacteria in vaccines ● Sporicidal at higher temperature

● Irritates mucous membranes, strong odor ● Used by food industry and to disinfect contact
lenses
● Also used in mortuaries for embalming
● C.) Benzoyl Peroxide:
● B.) Glutaraldehyde:
● Used in acne medications
● Less irritating and more effective than
formaldehyde C. CHEMOTHERAPEUTIC AGENTS
● One of the few chemical disinfectants that is a
● chemical substances used in treating diseases
sterilizing agent
● Antibiotics
● A 2% solution of glutaraldehyde (Cidex) is:
● - special type of chemotherapeutic agents used
● Bactericidal, tuberculocidal, and viricidal in 10
in treating infectious diseases
minutes. Sporicidal in 3 to 10 hours
● - usually obtained from living microorganisms
● Commonly used to disinfect hospital
instruments
● Also used in mortuaries for embalming CHEMICAL ANTIMICROBIAL AGENTS MODE OF ACTION

7. Gaseous Sterilizers: ● 1. Damage to cell wall or inhibition of cell wall

● Denature proteins, by replacing functional synthesis

groups with alkyl groups. ● 2. Alteration of the cytoplasm membrane

● Chemicals that sterilize in a chamber similar to permeability

an autoclave ● 3. Alteration of the physical and chemical state

of proteins and nucleic acid
● A.) Ethylene Oxide:
● 4. Inhibition of enzymes function
● Kills all microbes and endospores, but requires
● 5. Inhibition of protein and nucleic acid
exposure of 4 to 18 hours
synthesis
● Toxic and explosive in pure form
FACTORS AFFECTING THE MICROBIAL ACTIVITY
● Highly penetrating
● 1. Concentration of the agent
● Most hospitals have ethylene oxide chambers
to sterilize mattresses and large equipment ● 2. Temperature (increase temperature,
increase reaction)
8. Peroxygens (Oxidizing Agents):
 ● 3. Contact time

● 4. Ph

● 5. Nature of microorganisms (species)

● 6. Growth phase of microorganisms

● 7. Presence of extraneous materials (blood,

pus)

QUALITIES OF GOOD DISINFECTANT

● 1. High co-efficient of disinfection

● 2. Stable

● 3. Water soluble

● 4. Non-toxic, non-corrosive

● 5. Easy to use

● 6. Cheap