MICROBIAL GROWTH

MICROBIAL GROWTH
 A. Increase in cell number
 -Binary fission
 B. Increase in cell mass
- Doubling size
- Anabolic process
-

 Binary Fission
 Procaryotic cell division:


• Binary fission is the process by which

most prokaryotes replicate.
•
• Binary fission generally involves the
separation of a single cell into two
more or less identical daughter cells,
each containing, among other things,
at least one copy of the parental
DNA.

 Generation time
[Doubling Time]
• Interval time between successive
binary fissions of a cell.
• It helps determine the amount of
time that passes before disease
symptoms appear in an infected
individual.
– Staphylococcus aureus 30 mins
– Mycobacterium tuberculosis 5 hrs.
– Treponema pallidum (syphilis) 33
hrs
– E. Coli 20 mins.
 Standard Bacterial
Growth Curve
– Describes various stages of growth a
pure culture of bacteria will go
through, beginning with the addition
of cells to sterile media and ending
with the death of all of the cells
present.
– Shows the growth of cells over time
– To study population growth
– Plot of logarithm of cell versus
incubation time
–


  The phases of growth typically
observed include:
• lag phase
• exponential (log,
logarithmic) phase
• stationary phase
• death phase (exponential or
logarithmic decline)
•

• Lag Phase
– Increase in mass but has no change in
number.
• Log Phase
– Number double with each generation
time.
– Cells are most metabolically active.
– Applications:
• Human’s diseases: symptoms usually
develop cause by tissue damage.
• Broth tubes- become turbid
• Solid medium- microorganisms
become more visible

• Stationary Phase
•
• Period of equilibrium
• Number of microbial death balances
the number of new cells
• Population stabilizes
•

• Death phase [logarithmic
decline, exponential
decline]
•

– Death phase is a physiological point

at which cell deaths exceed cell
births.
– More specifically, viable count
declines.
–
• During the decline phase, many cells
undergo involution---that is, they
assume a variety of unusual
shapes, which makes them difficult
• Cell Death
– the point at which reinitiating of
division is no longer possible.

Growth Curve:
Factors Affecting Microbial
Growth
 Factors Affecting Microbial
Growth
• Temperature
• pH
• Oxygen
• Light
• Moisture
• Nutrient Content

 TEMPERATURE
• Bacterial species are poikilothermic
• Each bacterial species grows at particular
minimum, optimum, and maximum
temperatures


– MINIMUM GROWTH TEMPERATURE-

lowest temperature at which the
species will grow
– OPTIMUM GROWTH TEMPERATURE-
temp. at which the species will grow
best
– MAXIMUM GROWTH TEMPERATURE-
 O PTM U M
G ro w th R a te

M IN IM U M M A X IM U
M
Te m p e ra tu
re
• Psychrophiles
 min – 0°C
 opt – 15°C
 max – 20°C


 Can contaminate and grow in

food and dairy products when
refrigerated.
 Ex: Vibrio marinus
• Psychrotrophs or facultative
psychrotrophs
 min – 0°C
 opt – 20-30 °C
 max – 35°C

• Mesophiles – moderate
temperature
 min – 15-20°C
 opt – 20-45 °C
 max – 45°C or lower


 (most human pathogens

 are mesophiles).
 E.g. Corynbacterium
dephtherius, Nesseria
gonorrhoea
 E. Coli (opt.-37°C)
• Thermophiles – heat
loving
 min – 15-20°C
 opt – 55-65 °C
 max – 100°C


 (not pathogenic to human)

 e.g. Thurmus aquaticus.
• Hyperthermophiles / Extreme
thermophiles


 opt – 70-110 °C


- Achaea
- Highest recorded: 113 °C
 pH
• Bacteria: 6.5 – 7.5
• Molds and Yeasts: pH
4
• Neutrophiles: 5.5-8
• Alkalophiles: 8.5-11.5
• Acidophiles:acid
tolerant bacteria.


• Human Blood pH:

• 7.2-7.4
Acidophiles
• Lactobacillus
• Streptococcus
MILK
•
•
 pH
• Effects of acids on organisms
– Energy required to maintain cell’s
internal pH
– Enzyme activity affected
– Proteins, DNA, other molecules
denatured
– Longer lag, less rapid growth
 Oxygen
• AEROBE –depends on plentiful
supply of oxygen to be its final
electron acceptor to make cellular
energy.
• MICROAEROPHILES – concentration
of low oxygen
– Treponema pallidum (syphilis)
– Body Parts: oral cavity urinary tract,
gastrointestinal tract
 Oxygen
• AEROTOLERANT – insensitive to
oxygen and will grow in
environment with or w/o oxygen.
• OBLIGATE ANAEROBE- die if the
oxygen is present.
– e.g. Mycobacterium
– Clostrridium botulism –expired canned
goods
– Costridium tetanii- tetanus
– Result to the putrid odors of marshes,
landfills, and swamps.
 Oxygen
• FACULTATIVE ANAEROBES
– grow in the absence of oxygen, but
more slowly in than if oxygen were
presesnt
– e.g. Saccharomyces (yeast); E. Coli,
Streptococcus, Staphylococcus, and
Bacillus.
– FACULTATIVE AEROBE –prefers
anaerobic conditions but grows
aerobically.
– FACULTATIVE ANAEROBE- prefers
• CAPNOPHILIC – require an
atmosphere low in oxygen but rich
in carbon dioxide.


– Genera of Neisseria and

streptococcus
 Light
• Essential for photoautotroph
• As well as its intensity its wavelength
may be significant.
Moisture (Water Availability)
• Microbes must
have a supply of
water available
• Bacteria most
restricted, then
yeasts, then
moulds
• Organisms tolerant
of low water
levels:
– Halophilic
bacteria
– Osmophilic
yeasts
• WATER ACTIVITY (Aw) - depends
upon its presence in the
atmosphere (relative humidity) or
its presence in solution or a
substance.
• Microorganisms live over a range of
Aw from 1.0 to 0.7.

 
(Aw)
 100% H2O 1.0
 human blood 0.99
 Seawater 0.98
 Maple syrup 0.90
 Great Salt Lake 0.75.
 Agricultural soils 0.9 - 1.0.

• HALOPHILES-require some NaCl for
growth
– MILD HALOPHILES (1-6% salt)
– MODERATE HALOPHILES (6-15%
salt)
– EXTREME HALOPHILES (15-30%
NaCl)
– HALOTOLERANT (w/ and w/o the
presence of NaCl)
– OSMOPHILES (organisms that are
able to live in environments
high in sugar)
• XEROPHILES (Organisms which live
Effects of low water levels:

– Longer lag,
slower
growth
– Impaired
transport
– Loss of
membrane
permeability
 Nutritional Requirement
• chemicals and elements of this
environment that are utilized for
bacterial growth


• Require by all organisms: water,

carbon. Nitrogen, minerals


• Organic growth factors needed to

varying degrees
– Gram + > Gram - > yeast > moulds
 Major elements, their sources and
functions in bacterial cells.
Element % of dry weight Source Function
Carbon 50 organic compounds or Main constituent of cellular
CO2 material

Oxygen 20 H2O, organic compounds, Constituent of cell material

CO2, and O2 and cell water; O2 is
electron acceptor in aerobic
respiration

Nitrogen 14 NH3, NO3, organic Constituent of amino acids,

compounds, N2 nucleic acids nucleotides,
and coenzymes

Hydrogen 8 H2O, organic compounds, Main constituent of organic

H2 compounds and cell water
Element % of dry weight Source Function

Phosphorus 3 inorganic phosphates (PO4) Constituent of nucleic acids,

nucleotides, phospholipids, LPS,
teichoic acids

Sulfur 1 SO4, H2S, So, organic Constituent of cysteine, methionine,

sulfur compounds glutathione, several coenzymes

Potassium 1 Potassium salts Main cellular inorganic cation and

cofactor for certain enzymes

Magnesium 0.5 Magnesium salts Inorganic cellular cation, cofactor for

certain enzymatic reactions
Element % of dry Source Function
weight
Calcium 0.5 Calcium salts Inorganic cellular cation,
Iron 0.2 Iron salts cofactor
Componentfor of
certain
cytochromes
enzymes and
and certain a component
nonheme iron-
of endospores
proteins and a cofactor for
some enzymatic reactions
Trace Elements
• Metal ions required by certain cells in such small
amounts that it is difficult to detect (measure)
them, and it is not necessary to add them to
culture media as nutrients.
• Required in such small amounts that they are
present as "contaminants" of the water or other
media components.
• As metal ions, the trace elements usually act as
cofactors for essential enzymatic reactions in
the cell.
• One organism's trace element may be another's
required element and vice-versa.
• Usual cations that qualify as trace elements in
bacterial nutrition are Mn, Co, Zn, Cu, and Mo.
 Major Nutritional Types of
Procaryotes
Nutritional Type Energy Source Carbon Source Examples
Photoautotrophs Light CO2 Cyanobacteria,
Photoheterotrophs Light Organic some
Some Purple
Purple and
and
Chemoautotrophs Inorganic compounds
CO2 Green
A few Bacteria
Green Bacteria
Bacteria
or Lithotrophs
Chemoheterotrophs compounds,
Organic e.g. Organic and
Mostmany
Bacteria,
(Lithoautotrophs)
or Heterotrophs H 2, NH3, NO2,
compounds compounds Archaea
some Archaea
H2 S
 Growth Factor -required in small
amounts by cells because they fulfill
specific roles in metabolism.
 The need for a growth factor results

from either a blocked or missing

metabolic pathway in the cells.


 Growth factors are organized into three

categories:


 purines and pyrimidines: required for

synthesis of nucleic acids (DNA
and RNA)
 amino acids: required for the synthesis of
proteins
 CULTURE MEDIA
• Provide all the essential nutrients in
solution for bacterial growth.
• Employed in the isolation and
maintenance of pure cultures of
bacteria and are also used for
identification of bacteria according
to their biochemical and
physiological properties.
• Liquid (broth) vs. Solid media
– Liquid medium
• Components are dissolved in water
and sterilized
– Solid medium
• A medium to which has been added a
gelling agent
• Agar (most commonly used)
• Gelatin
• Silica gel (used when a non-organic
gelling agent is required)
 Uses of Liquid Media vs
Solid Media
• Liquid media- growth of pure/batch
cultures
• Solidified media - isolation of pure
cultures
– for estimating viable bacterial
populations, and a variety of other
purposes.
–

 AGAR -hydrocolloid derived from red

algae.
 - melts at 100°C and remains liquid
until cooled to 40°C the temperature at
which it gels)
 - it cannot be metabolized by most
Microbiological Media
– Chemically Defined Media
• The exact chemical composition is
known
• e.g. minimal media used in bacterial
genetics experiments
– Complex Media
• Exact chemical composition is not
known
• Often consist of plant or animal
extracts, such as soybean meal,
milk protein, etc.
• Include most routine laboratory
media,
 Microbiological Media
• Selective media
– Contain agents that inhibit the
growth of certain bacteria while
permitting the growth of others
– Frequently used to isolate specific
organisms from a large population
of contaminants
• Differential media
– Contain indicators that react
differently with different organisms
(for example, producing colonies
with different colors)
 Isolation of Pure
cultures
• Spread Plates


• Streak Plates


• Pour Plates

COLONIES – accumulation of cells large enough to

be seen without a microscope