Growth curve of bacteria and factors effect bacterial growth curve

Growth curve
of Bacteria
Dr. Harinatha Reddy M.sc, Ph.D.
biohari14@gmail.com
Department of Microbiology
Sri Krishnadevaraya University
Anantapur, A.p. India
08/06/2017

Bacterial growth
• Growth may be defined as an increase in cellular components
followed by increase in the number of cells.
• Microorganisms reproduce by processes like binary fission.

Binary fission
1. Bacterial cells grow by
increasing in cell
number
2. Replication is by binary
fission, the splitting of
one cell into two
3. Therefore, bacterial
populations increase by
a factor of two (double)
every generation time.

Generation time
• The time required to for a population to double in number Or Time
require for the cell division.
• Ex. E. coli double every 20-30 minutes
• Ex. Mycobacterium tuberculosis double every 12 to 24 hours
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Bacterial growth curve:
• Graphical representation the pattern of bacterial growth, or
a graph of viable cells versus the incubation time is called
Bacterial growth curve.
• Bacterial growth curve obtained from Batch system OR
Closed system.
• Because limited nutrients and no fresh medium is provided
during incubation, nutrient concentrations decline and
concentrations of wastes increase.
• The shape of the Bacterial growth curve is ‘S” or Sigmoid
shape.

The bacterial growth curve has four distinct phases.
1.lag
2.log
3.Stationary phase
4.Death phase.

Lag phase:
• When microorganisms are introduced into fresh culture medium, usually no
immediate increase in cell number, therefore this period is called the lag phase.
• No cell division and there is no net increase in mass, But cells increase in size.
• The cell is synthesizing new components.
• Phase of cell enlargement or Preparation phase/cell adjustment phase.
• The lag phase varies considerably in length with the condition of the
microorganisms and the nature of the medium. This phase may be quite long if
the inoculum is from an old culture or one that has been refrigerated.

Log phase:
• Also known as exponential phase.
• microorganisms are growing and dividing at the maximal rate possible given
their genetic potential, the nature of the medium, and the conditions under
which they are growing.
• Their growth rate is constant during the log phase; that is, the
microorganisms are dividing and doubling in number at regular intervals.
• The population is most uniform in terms of chemical and physical properties
during this phase; therefore log phase cultures are usually used in
biochemical and physiological studies.
• Cells are small in size, and produce primary metabolites.
• Examples for primary metabolites?

• Log phase growth is balanced growth phase.
• All cellular components are manufactured at constant rates.
• The cells first construct new ribosomes to enhance their
capacity for protein synthesis. This is followed by increases in
DNA replication. Finally, the expected rise in reproductive rate
takes place.
• If nutrient levels or other environmental conditions change,
unbalanced growth results.
• Unbalanced growth also results when a bacterial population is
shifted down from a rich medium to a poor one.

Stationary Phase:
• This stationary phase usually is attained by bacteria at a
population level of around 109 cells per ml.
• Other microorganisms normally do not reach such high
population densities, protozoan and algal cultures often having
maximum concentrations of about 106 cells per ml.
• The final population size depends on nutrient availability and
other factors, as well as the type of microorganism being
cultured.

• In the stationary phase the total number of viable
microorganisms remains constant. This may result from a
balance between cell division and cell death.
• In this phase bacteria growth rate is decreases, Due to the,
deposition of toxic waste products and empty of nutrients.
• The secondary metabolites are produced in this phase.
• For example, streptococci can produce so much lactic acid and
other organic acids from sugar fermentation that their medium
becomes acidic and growth is inhibited.

• Bacteria in a batch culture may enter stationary phase in response to
starvation.
• Starvation can be a positive experience for bacteria.
• In starvation bacteria only decrease in overall size, by protoplast shrinkage
and nucleoid condensation.
• The more important changes are in gene expression. Starving bacteria
frequently produce a variety of starvation proteins, which make the cell
much more resistant.

• They increase peptidoglycan cross-linking and cell wall strength. The Dps
(DNA-binding protein from starved cells) protein protects DNA.
Chaperones prevent protein denaturation and renature damaged proteins.
• As a result of these and many other mechanisms, the starved cells become
harder to kill by temperature changes, oxidative and osmotic damage, and
toxic chemicals. These changes are so effective that some bacteria can
survive starvation for years.
• Salmonella typhimurium and some other bacterial pathogens become
more virulent during starvation.

Death Phase:
• The number of viable cells are decline death phase.
• The death of a microbial population, is usually logarithmic (that is,
a constant proportion of cells dies every hour).
• Environmental changes like nutrient deprivation and the buildup
of toxic wastes lead to the decline in the number of viable cells

1. Bacteria growing in batch culture produce a growth curve with up to
four distinct phases.
2. Batch cultures are grown in tubes or flasks and are closed systems where
no fresh nutrients are added or waste products removed.
3. Lag phase occurs when bacteria are adjusting to them medium. For
example, with a nutritionally poor medium, several anabolic pathways
need to be turned on, resulting in a lag before active growth begins.
4. In log the cells are growing as fast. During this phase, almost all cells are
alive, they are most nearly identical.
5. Due to nutrient depletion and/or accumulation of toxic end products,
replication stops and cells enter a stationary phase where there is no net
change in cell number.
6. Death phase occurs when cells can no longer maintain viability and
numbers decrease.
Growth in Batch Culture

Factors affecting growth
Dr.Harinatha Reddy, Aswartha
09/06/2017

The Influence of Environmental Factors on Growth:
• Microbes, even if they are supplied with all the necessary
nutritional requirements, still may not grow.
• Bacteria, and other microbes, can only survive and reproduce
within a certain range of environmental conditions like.
• Various factors affecting growth of bacteria:
 Solutes and Water Activity
 Radiation
 Temperature
 Oxygen requirement
 pH

Solutes and Water Activity
• Permeable plasma membrane separates microorganisms from their surroundings
environments.
• If a microorganism is placed in a hypotonic solution (solution with low solute con.), water
will enter the cell and cause it to burst.
• Moment of water from the outer medium in to the cell when cell placed in a hypotonic
solution is known as Endoosmosis.
• Most bacteria, algae, and fungi have thick cell walls that maintain the shape and integrity
of the cell.
• When microorganisms with thick cell walls are placed in a hypertonic (solution with high
solute con.) environment, water leaves and the plasma membrane shrinks away from the
wall, a process known as plasmolysis.
• This dehydrates the cell and may damage the plasma membrane; the cell usually becomes
metabolically inactive.

• Most prokaryotes increase their internal osmotic concentration in a
hypertonic environment through the synthesis or uptake of choline,
proline, glutamic acids and other amino acids; elevated levels of
potassium ions.
• A few prokaryotes like Halobacterium raise their osmotic concentration
with potassium ions.
• Halobacterium’s enzymes have been altered so that they actually require
high salt concentrations for normal activity, generally halophilenequires
high levels of sodium chloride.
• Osmotolerant: Able to grow over wide ranges of water activity or osmotic
concentration Staphylococcus aureus.

Solutes and Water Activity
 Some bacteria require salt to grow and are
called halophiles.
 If a very high concentration of salt is required
the organisms are termed extreme halophiles.
 A nonhalophile that can grows best with
almost no salt but can still grow with low
levels of salt (~ 7%) is called halotolerant.

radiation:
• Ionising gamma rays emitted by cobalt-60 and X-rays to inactivate microbial pathogens, particularly in
the food industry.
• Bacteria such as Deinococcus radiodurans are particularly resistant to radiation, but are not
pathogenic.
• Active microbes, such as Corynebacterium aquaticum, Pseudomonas putida, and Micrococcus diversus,
have been retrieved from spent nuclear fuel.
• These microbes were again exposed to controlled doses of radiation. All the species survived radiation
doses with little damage, while only the gram-positive species survived much larger doses.
• The spores of gram-positive bacteria contain storage proteins that bind tightly to DNA, possibly acting
as a protective barrier to radiation damage.

Temperature:
• Bacteria can maintain T.M.P stability by :
1. By maintaining plasma membrane integrity.
2. By controlling structural stability of protein.
3. By controlling enzyme activity.
But bacteria doesn’t have constant temperature.
Organisms which changes their body temperature according to
environment are called poikilotherms.

Cardinal range temperature: Temperature at which bacteria grow effectively.
Psychrophilic = Cold loving; optimum growth at 0 – 20 C
Mesophilic = middle living; optimum growth at 30 - 37 C
Thermophilic = heat loving; optimum growth at 50 - 60 C
• Psychrophiles are commonly isolated from cold waters and bottoms of lakes and oceans.
Many bacteria grow well at 0 to 4 C and some at -7 C.
• Mesophiles are the most common bacteria grow best around 30-40 C.
 Thermophiles were first found in hot springs and hot water heaters 50 - 60 C.

• Some species of microorganism can grow at temperatures as low as -10o C, and
others at temperatures as high as 100o C.
• At lower temperatures molecules move slowely, enzymes cannot mediate in
biochemical reactions, and cell activity decreases.
• As the temperature increases, molecules move faster, enzymes speed up
metabolism and cells rapidly increase in size.
• But, above a certain value all of these activities are proceeding at such high
rates, enzymes start to denature, Cellular growth decreases.

Temperature
• Psychrophile
0o to 18o C
• Psychrotroph
20°C to 30°C
• Important in food spoilage
• Mesophile
25°C to 45°C
• More common disease causing
• Thermophiles
– 45°C to 70°C
Common in hot springs and hot water heaters.
• Hyperthermophiles
– 70°C to 110°C
• ProKaryotes such as Bacillus infernus even seem able to live over 2 miles
below the Earth’s surface, without oxygen and at temperatures above 60°C.
Microorganisms that grow in such harsh conditions are often called
extremophiles.

pH:
• Most bacteria are neutrophiles, meaning they grow at neutral pH of 7. Most
familiar bacteria, like E. coli, Staphylococci, and Salmonella spp. are
neutrophiles.
• Microorganisms that grow optimally at pH less than 5.55 are called acidophiles.
• For example, the sulfur-oxidizing Sulfolobus spp. isolated from sulfur mud fields
and hot springs in Yellowstone National Park are extreme acidophiles.
• Ferroplasma live in acid mine drainage at pH values of 0–2.9.
• Lactobacillus bacteria, which are an important part of the normal microbiota of
humans can tolerate acidic environments at pH values 3.5–6.8.
• Alkaliphiles are , microorganisms that grow best at pH between 8.0 and
10.5. Vibrio cholerae, the pathogenic agent of cholera, grows best at the slightly
basic pH of 8.0; it can survive pH values of 11.0 but is inactivated by the acid of
the stomach.

 Extreme alkaliphiles have adapted to their harsh environment
through evolutionary modification of lipid and protein
structure and due to the proton motive force in an alkaline
environments.
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Oxygen:
• An organism able to grow in the presence of atmospheric O2 is an aerobe, whereas
one that can grow in its absence is an anaerobe. Almost all multicellular organisms
are completely dependent on atmospheric O2 for growth—that is, they are obligate
aerobes.
• Oxygen serves as the terminal electron acceptor for the electron- transport chain in
aerobic respiration.
• Facultative anaerobes do not require O2 for growth but do grow better in its
presence. In the presence of oxygen they will use aerobic respiration.
• Aerotolerant anaerobes such as Enterococcus faecalis simply ignore O2 and grow
equally well whether it is present or not.
• In contrastobligate anaerobes (e.g., Bacteroides, Fusobacterium, Clostridium
pasteurianum, Methanococcus) do not tolerate O2 at all and die in the presence of
oxygen..

• Oxygen accepts electrons and is readily reduced because its two outer
orbital electrons are unpaired. Flavoproteins several other cell
components, promote oxygen reduction.
• The result is usually some combination of the reduction products
superoxide radical, hydrogen peroxide, and hydroxyl radical.
O2 + e- → O2– (superoxide radical)
O2– + e- +2H → H2O2 (hydrogen peroxide)
H2O2 + e- + H+ → H2O +OH (hydroxyl radical)
• These products of oxygen reduction are extremely toxic because they are
powerful oxidizing agents and rapidly destroy cellular organells.

• Many microorganisms possess enzymes protect cells against O2 toxic
products. Obligate aerobes and facultative anaerobes usually contain
the enzymes superoxide dismutase (SOD) and catalase, which catalyse
the destruction of superoxide radical and hydrogen peroxide,
respectively.
2O2- +2H+ --- superoxide dismutase-- O2+H2O2
H2O2 ------Catalase----- 2H2O+ O2
• Aerotolerant microorganisms may lack catalase but almost always have
superoxide dismutase.
• All Obligate anaerobes lack both enzymes or have them in very low
concentrations and therefore cannot tolerate O2.

Growth of prokaryotes depends on nutritional requirements;
Major elements: (CHONSP + K, Mg, Fe, Ca)
Carbon, oxygen, hydrogen, nitrogen, sulfur, phosphorus, potassium,
magnesium, iron, and calcium.
Essential components for macromolecules.
Trace elements: ( Cu, Ni, Zn, Mo, Mn):
zinc, copper, molybdenum and manganese required in minute amounts, which
are Assist in enzyme function. (which are co factors for enzymes)

Element % dry wgt Source
Carbon 50 organic compounds or CO2
Oxygen 20
H2O, organic compounds, CO2, and
O2
Nitrogen 14 NH3, NO3, organic compounds, N2
Hydrogen 8 H2O, organic compounds, H2
Phosphorus 3 inorganic phosphates (PO4)
Sulfur 1
SO4, H2S, organic sulfur
compounds
Potassium 1 Potassium salts
Magnesium 0.5 Magnesium salts
Calcium 0.5 Calcium salts
Iron 0.2 Iron salts

Growth factors
• The medium without growth factors is known as
minimal medium.
• Organisms do not require growth factors is called
Prototrophs.
• Organisms which can’t synthesize essential
components from raw material form the medium is
called Auxotrophs.
• The most important vitamin for the growth of the
bacteria is B –Complex.

Culture Media
• Complex (contains undefined components)
• Chemically defined (all concentrations are known)
• Selective (favors the growth of a particular organism or group
of organisms)
• Differential (has reactions that give isolates different
appearance)
• Anaerobic (oxygen-free)
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Growth curve of bacteria and factors effect bacterial growth curve

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