Microbial Growth
Growth Curve
Growth can be defined as :
1. increase in cellular constituents. Cells growing
longer and larger.
2. increase the number of cells in a population
( growth in size of population) .
Population growth is studied by analyzing the
growth curve of a microbial culture.
The growth curve has four distinct phases:
1. lag phase 2. exponential ( log) phase ,( growth
phase)
3. Stationary phase
4. death phase
How did biologists determine the presence
of such phases??
By growing bacteria in batch culture.
Batch culture: bacteria grown in liquid medium
and incubated in closed culture vessel with a
single batch of medium : no fresh medium is
provided during incubation so
1. Nutrient concentration decrease
2. wastes concentration increase
• Under such conditions the growth of a
population of microbes reproducing by binary
fission can be blotted as the logarithm of the
number of viable cells versus the incubation
time .
1.Lag phase
• Cells are synthesizing new components, no
increase in cell number occurs .
• This phase is necessary because :
1. Cells may be old and depleted of : ATP ,
essential cofactor , and ribosome‘s that must
be synthesized before growth can begin.
2. microorganisms may transferred from
medium to another so new enzymes needed
to use different nutrients.
3. Possibly the microorganisms have been
injured and require time to recover.
In this phase cells begin to replicate DNA, and
increase in mass.
2. Exponential phase ( log phase )
• Microorganism are growing and dividing at the
maximal rate possible . this affected by :
1.Genetical potential
2. Nature of the medium
3. Environmental condition
• Cells doubling in number at regular intervals ( the
rate of growth is constant)
This phase used in biochemical and physiological
studies because the population is most uniform in
terms of chemical and physiological properties
Balanced and unbalanced growth
Balanced growth :
Cellular constituents are manufactured at a constant
rates relative to each other .
In exponential phase growth is balanced.
Unbalanced growth :
the rates of synthesis of cell components vary
relative to one another until a new balanced state is
reached.
Unbalanced observed in two types of
experiments
1. shift up : a culture is transferred from a
nutritionally poor medium to a rich one >>>
there is a lag while the cells first construct new
ribosomes to enhance their capacity for protein
synthesis.
2. Shift down : a culture is transferred from a rich
medium to poor one >>> there is a lag in growth
because cells need time to make the enzymes
required for the biosynthesis of a available
nutrients.
• This show that microbial growth is under
coordinated control and respond quickly to a
change in environmental conditions
• once the cell are able to grow again , balanced
growth is resumed and the culture enters the
log phase.
3. Stationary phase
• Population growth eventually cease and the growth
curve becomes horizontal .
• In stationary phase , the total number of viable
microorganisms remain constant. This result from :
1.A balance between cell division and cell death or
2. Population may cease to divide but remain
metabolically active.
• Microbial population enter the stationary phase
because several factors operating in concert:
1. Nutrient limitation
e.g Aerobic organisms often limited by O2
availability.
2. Accumulation of toxic waste products
e.g Streptococci ferment sugar ( in absence of
O2) >> produce much lactic acid and other
organic acid. >>> medium become acidic
>>growth inhibited
3. Growth may cease when critical population
level is reached.
stationary phase is attained by most bacteria
at a population level of around 109 cells per
milliliter.
Protest cultures often have maximum
concentration of about 106 cells per milliliter.
4. Senescence and Death
• Death phase: in this phase , the number of viable
cells often declines at an exponential rate.
Two hypothesis to interpret such decline:
1. Cells enter temporarily phase under which they
can not grow at least under the laboratory
conditions used. This called viable but
nonculturable ( VBNC)
( become dormant as some types form spores in
response to unfavorable conditions
• When temperature change or the microorganisms
pass to animals >>> microbes resume growth.
• This phenomenon is important as many assays that
test for food and drinking water safety are culture
based.
• 2. programmed cell death >> predict that a fraction
of the microbial population is genetically
programmed to die after growth ceases.
In such case the nutrients leak from dead cells
enable the eventual growth of cells did not initiate
cell death.
Some microbes have a very gradual decline in
the number of culturable cells >> this can last
months to years >> during this time bacterial
population evolve so actively, reproducing
cells are those :
1.Best able to use the nutrients released from
dead cells.
2. Best able to tolerate the accumulated
toxins.
Mechanisms of growth
During the exponential phase, each
microorganism is dividing at constant intervals >>>
the population double in number during a specific
length of time called generation ( doubling )time .
e,g culture tube inoculated with one cell that divides
every 20 minutes , after 20 minutes will be 2 cells
After 40 minutes will be 4 cells and so …..
So the increase in population is always 2n where n
is the number of generations
Influences of Environmental factors on
growth
An understanding of environmental influences aids
in :
1. The control of microbial growth
2. The study of the ecological distribution of
microorganisms
Most important environmental factors :
1. Solutes and water activity
2. PH
3. Temperature 4. oxygen level 5. pressure
6. radiation
1. Solutes and water activity
• A selectively permeable plasma membrane
separates microorganisms from their
environment can be affected by change in the
osmotic concentration of their surroundings
• According to their osmotic pressure solution
can be :
1.Hypotonic solution : one with lower
osmotic concentration ( low solutes)
• 2. hypertonic solution : one with higher
osmotic concentration ( high solutes)
In osmosis water will move from hypotonic
solution to the hypertonic .
In hypotonic solution Water will enter the cell
and cause it to burst unless something is done
prevent the influx or inhibit plasma
membrane expansion
• In hypertonic solution water will flow out of
the cell . In microbes with cell wall, plasma
membrane shrinks away from the cell wall =
plasmolysis.
Dehydration in hypertonic medium may
damage the cell membrane and cause the cell
to become metabolically inactive.
Halobacterium and other extremely
halophilic archaea accumulate enormous
quantity of ions to remain hypertonic to their
environments as:
1. chloride ions.
which is also needed to stabilize the plasma
membrane and cell wall of halobacterium
When sodium ions decrease plasma
membrane and cell wall disintegrate
• Halophiles adapted successfully to extreme
environmental condition but lost ecological
flexibility.
Water activity (aw) : a low water activity
indicates that most of the water is chemically
or structurally bound to other components in
the medium >> it is a quantitatively the
degree of water availability .
Microorganisms must expend extra effort to
grow in a habitat with low aw because it must
maintain a high internal solute concentration
to retain water.
Osmotolerant
Grow over wide range of water activity
e. g1. Staphyllococcus aureus are halotolerant
>>>>> cultured in media containing sodium
chloride up to 3M.
(58.44 g of NaCl is needed to make 1M)
E,g 2. yeast Saccharomyces rouxii grow in sugar
solutions with aw as low as 0,6.
How microorganisms respond to change in the
osmotic concentration of their environment??
1. In hypotonic environment:
Microbes can reduce the osmotic concentration of
their cytoplasm by several mechanisms
A. in bacteria: some bacteria have
mechanosensitive (MS) channels in their plasma
membrane >>> in hypotonic environment water
will enter the cell >> increase hydrostatic pressure
and cellular swelling >> Ms channels open and
allow solutes to leave .
B. In protist : many protist do not have cell wall so
they use contractile vaculoes to expel exess
water.
2 In hybertonic environment
Compatible solutes
Many microorganisms keep the osmotic
concentration of their cytoplasm above that the
surrounding environment by use compatible
solutes >> they are a chemicals do not interfering
with metabolism and growth but used to
increase the internal osmotic concentration.
Most bacteria and archea use choline( watersoluble vitamen- like essential nutrients ,it is a
constituent of lecithin ….) beatine, aminoacids as
proline and glutamic acid and elevated levels of
potassium and chloride ions
Protists and fungi use : sucrose and polyols( is an
organic compound containing mutiple hydroxyl
groups): arabitol ( is a sugar alcohol) , glycerol,
and mannitol ( type of a sugar alcohl )as
compatible solutes.
Halophiles : microbes adapted to extreme
hypertonic environments need high level of
sodium chloride to grow ( between 2M and
saturation about 6,2M.
• The archaeon halobacterium can be isolated
from the Dead sea and other aquatic habitats
with salt concentration approach saturation.
2. PH
PH is a measure of the relative acidity of a
solution .
Each species has :
1.PH growth range
PH growth optimum
• Acidophiles : their growth optimum between
0 and 5.5
• Neutrophiles: their growth optimum between
5.5 and 8.0
• Alkalophiles: their growth optimum between
8.0 and 11.5
• Extreme alkalophiles: >>growth optima at PH
10 or higher .
• Most known bacteria and protists are neutrophiles.
• Most fungi prefer more acidic surroundings about
4 to 6.
Photosynthetic protists seem to favor slight acidity .
• Many archaea are acidophiles
e.g the archaeon Sulfolobus acidocaldarius :
-It is a common inhabitant of acidic hot springs
-It grows well from PH 1 to 3 and at high temperatur
• Alkalophiles are distributed among all three
domains of life :
• Bacteria : genera Bacillus, micrococcus ,
pseudomonas and streptomyces
• Yeast and filamentous fungi
• Numerous archaea
• alkaliphilic.
• When H+ concentration in out side much
grater than inside ( low PH) H+ move into the
cytoplasm and lower PH .
• Drastic variation in cytoplasmic PH can harm
Microorganisms ( affect plasma membrane,
enzymes, transport protein).
• Mechanisms to maintain cytoplasmic PH:
1. Neutrophiles : exchange potassium to
protons
2. Internal buffering may contribute .
• If external PH become too acidic >>> below about
5.5 to 6.0 Salmonella enterica ,Serovar
typhimurium and E.coli show acidic tolerance
response >>>synthesis new proteins .
- A proton translocating ATPase enzyme
contribute by pumping protons out of the cell.
• If PH decrease to 4.5 or lower acid shock proteins
and heat shock proteins are synthesized >> these
prevent the denaturation of proteins and aid in
the refolding of denatured proteins in acidic
condition.
Microbes at PH extremes
• Extremes alkalophiles as Bacillus alcalophiles
maintain internal PH close to neutrality by
exchange internal sodium ions for external
protons.
Acidophiles maintain suitable internal PH by:
1. Transport of cations (e.g pottasium ions) into
the cell. Thus decreasing the movement of H+
into the cell.
2.Proton transporters that pumps H+ out if they
get in .
3. Highly impermeable membranes.
3.Temperature
• Microorganisms susceptible to external temperature
because they can not regulate their internal
temperature.
• Change in temperature is important as temperature
some enzyme-catalyzed reactions are temperature
sensitive.
• Each enzyme has a temperature at which it function
optimally , below or above this temperature the
activity of enzymes affected.
• High temperature denature enzymes.
Temperature has significant effect on microbial
membrane:
1. At very low temperature , membrane
solidify
2. At very high temperature , the lipid bilayer
simply melts and disintegrates.
When organisms above or below their optimum
temperature both function and cell structure
affected.
• Each microbes has minimum, optimum,
maximum growth temperature.
• The temperature optimum closer to the
maximum than to the minimum.
• Microorganisms are placed in different classes
based on their temperature ranges for growth.
1. A- psychrophiles : grow well at zero C , optimum
growth 150C , maximum around 20 oC
Habitats isolated from Arctic and antractica
habitats
• Oceans constitutes an enormous habitat for
psychrophiles because 90% of ocean water is 5oC or
colder.
• In psychrophiles enzymes, transport system, protein
synthetic machinery function well at low
temperature.
• Their cell membrane have high level of unsaturated
fatty acids and remain semi fluid when cold.
• Psychrotrophs ( psychrotolerants)
grow at o or higher , maximum about 35oC
Psychrophilic bacteria and fungi are the major
causes of refrigerated food spoilage.
Mesophiles
• Most microorganisms probably fall within this
category.
• Optima around 20- 45 .
• Minimum 15 to 20 oC.
• Maximum about 45 oC.
• Almost all human pathogens are mesophiles,
as might be expected because the human
body temperature is 37oc.
Thermophiles
•
•
•
•
Grow between 55 to 85oC.
Minimum around 45oC
Optima between 55 and 65oC.
The vast majority are bacteria and archaea,
few photosynthetic protists and fungi.
• Habitats : hot water lines and hot springs.
•
•
•
•
Hyperthermophiles
Optima between 85 and 113oC
Do not grow below 55oC.
Thermophiles and hyperthermophiles differ
from mesophiles in many ways.
1. They have heat- stable enzymes, and protein
synthesis systems that function properly at
high temperatures
2. Have large quantities of aminoacids such as
proline also make polypeptide chains flexible
and more heat stable.
3. Presence of chaperone proteins that stabilize
and aided in folding of proteins.
4. Nucleoid- associated proteins stabilize the
DNA of thermophilic bacteria.
5. Membrane lipids are also quite temperature
stable>> it is more saturated, more branched
of higher molecular weight >>increasing
melting points.
4. Oxygen concentration
• Aerobes: An organisms able to grow in the
presence of atmospheric O2.
• Anaerobe: an organisms can grow in absence
of oxygen.
• Obligate aerobes: organisms completely
dependent on atmospheric O2 for growth
( almost all multicellular organisms).
• Microaerophiles: damaged by the normal
atmospheric level of O2 ( 20%) and require O2
level in the range of 2 to 10% for growth.
• Facultative anaerobes: do not require O2 for
growth but grow better in its presence. In the
presence of oxygen they use O2 as the
terminal electron transport acceptor during
aerobic respiration .
• Aerotolernat anaerobes: grow equally well
whether O2 is present or not.
• Obligate anaerobes: killed by presence of O2.
5. Pressure
• Organisms live on land or surface of water subjected
to pressure of 1 atmosphere ( atm) and are never
affected significantly by pressure.
• Organisms including many bacteria and archaea live
in the deep sea ( ocean depths of 1000 m or more ),
where hydrostatic pressure can reach 600 to 1100
atm and the temperature is about 2 to 3oC.
• These high hydrostatic pressures affect membrane
fluidity and membrane- associated function.
• Barotolerant: found at great ocean depths.
Increased pressure adversely affects them but not as
much as it does nontolerant.
• Piezophilic: grow rapidly at high pressure, defined as:
an organism that has maximal growth rate at
pressures grater than 1 atm but less than about 950
atm.
• Hyper piezophiles: having growth rate maxima grater
than 590 atm.
• An important adaptation observed in piezophiles Is
>>> they change their membrane lipids in response
to increasing pressure.
• Some bacteria piezophiles increase the
amount of unsaturated fatty acids and shorten
the length of their fatty acids
• Piezophiles are thought to play important
roles in nutrient recycling in the deep sea.
6. Radiation
• Radiation behaves as if it were composed of waves
moving through space like waves traveling on the
surface of water.
• Te distance between two crests or troughs is the
wave length .
• As the wavelength of electromagnetic radiation
decrease , the energy of the radiation increases.
• Sun light is the major source of radiation on earth
, it includes:
1. Visible light 2. ultraviolet ( UV) radiation
3. Infrared rays 4. radio waves
•
Many forms of electromagnetic radiation is very
harmful to microorganisms.
• Ionizing radiation ( cause atoms to lose
electron);
1. X rays: produced artificially
2. Gamma rays : emitted during radioisotops decay
• Low level of ionizing radiation may produce
mutations and may indirectly result in death,
whereas higher level are directly lethal.
• Ionizing radiation can be used to sterilize
items.
• Some bacteria and bacterial endospore are
extremely resistant to large doses of ionizing
radiation.
• Ultraviolet ( UV) radiation from 10 to 400 can
kill microorganisms ( short wavelength+ high
energy .
• The most lethal UV radiation has a wavelength
of 260 nm , the wave length most effectively
absorbed by DNA.
• The primary mechanisms of UV damage is the
formation of thymine dimers in DNA, which
inhibit DNA replication and function.
• Thymine dimers are formed when two
adjacent thymins in the same DNA strands are
covalently joined.
• Even visible light when present in sufficient
intensity can damage or kill microbial cells.