Microbial growth requirements:
Chemical growth factors:
What are the chemical growth factors required for isolation of
microbes in vitro?
1-Carbon: organic: Glucose.
Inorganic: CO2
2- Hydrogen occurs in organic hydrocarbon molecules and in
inorganic molecules such as water.
3-Oxygen or gaseous requirements of microbes.
4-Nitrogen is used in protein/ amino acid synthesis and nucleic acid
polymerization.
5- Phosphorus is essential for nucleic acid synthesis and formation
of phospholipids.
6- Sulfur present in certain amino acids such as cysteine and
methionine .
Physical growth factors:
1-Acidity (pH requirements):
Pathogenic bacteria grow best at neutral or biological pH which is
typically between pH 6.8 to pH 7.4.
Fungi such as yeasts and molds prefer slightly more acidic
conditions and grow best between pH 5 to pH 6.
The pathogen Helicobacter pylori is able to survive with pH of the
stomach by producing urease. So it is acidophilic bacterium.
On the other extreme, bacteria that prefer alkaline (basic) conditions
are known as alkaliphiles. Example: Vibrio cholerae (prefers pH 9).
a
2- Acids, Bases, Salts concentration:
An ACID can be described as a hydrogen donor.
A BASE can be described as a hydrogen acceptor.
A SALT is an ionic compound that dissociates in water into anions
and cations.
3- Temperature requirements:
Mesophiles: Grow at optima between 25 C and 40 C.
Normal microbiota and most pathogens grow at or near human
body temp of 37 C.
Thermophiles:
heat-loving microbes of Optima between 50 and 60 C .
4- Gaseous requirements and Humidity :
According to oxygen requirements, microbes can be classified into:
1- Strict ,Obligate Aerobe: O2 must be present during
cultivation of microbe. Example: Pseudomonas aeruginosa.
2- Strict or obligate Anaerobes:
Molecular oxygen is toxic to these organisms. Lack enzymes
catalase and superoxide dismutase (SOD) to neutralize
hydrogen peroxide and free radicals, respectively.
Example: Bacteriodes and Clostridium.
3- Facultative anaerobes:
Adaptable organisms that use oxygen when present but can
switch to anaerobic pathways in its absence.
Example: Escherichia coli.
4- Microaerophilic:
Also known as "Capnophilic" or carbon-dioxide loving. Only use low
concentrations of oxygen. Example : Helicobacter pylori.
Humidity: The percentage of water vapor in the cultivation atmosphere.
The Bacterial Growth Curve:
Bacterial growth is the division of one bacterium into two daughter
cells in a process called binary fission.
During lag phase, bacteria adapt themselves to growth conditions.
Exponential phase (sometimes called the log phase): is a period
characterized by cell doubling.
For this type of exponential growth, plotting the natural logarithm of cell
number against time produces a straight line.
a
During stationary phase, the growth rate slows as a result of nutrient
depletion and accumulation of toxic products.
This phase is a constant value as the rate of bacterial growth is equal
to the rate of bacterial death.
At death phase, bacteria run out of nutrients and die.
Bacterial culture media:
An undefined medium (also known as a basal or complex medium). It
is a medium that contains:
1-A carbon source such as glucose for bacterial growth
2-water
3-various salts needed for bacterial growth.
Defined media (also known as chemically defined media or synthetic
media).
Classification of media:
Media can be classified into:
1-Minimal media ( simple medium): nutrient broths and agar media
2- Selective media:
Selective media are used for the growth of only selective microbes.
It contains antibiotics, dye, or specific chemicals that inhibits the
growth of most types of microbe and stimulate the isolation of one
type.
Mannitol salt agar (MSA)
which is selective for
Gram-positive bacteria.
Blood-free, charcoal-based selective
medium agar (CSM) for isolation of
Campylobacter.
3-Differential media:
Differential media or indicator media distinguish one microorganism type
from another growing on the same media.
Indicators (such as neutral red, phenol red, eosin y, or methylene blue) could
be used.
Examples of differential media include:
Eosin methylene blue (EMB), which is differential for lactose and sucrose
fermentation.
MacConkey (MCK), which is differential for lactose fermentation.
4- Enriched media:
Enriched media contain the nutrients required to support the growth
of a wide variety of organisms, including some of the more
fastidious ones.
Blood agar is an enriched medium in which nutritionally rich whole
blood supplements the basic nutrients. Chocolate agar is enriched
with heat-treated blood (40-45°C).
5- Transport media:
Transport medium is a simple organic medium that Maintain the
viability of all organisms in the specimen without altering their
concentration.
This type of medium mainly used for Temporary storage of
specimens being transported to the laboratory for cultivation.
Examples of transport media include:
Thioglycolate broth for strict anaerobes.
Antibiotics:
An antibacterial agent is a compound or substance that kills or slows
down the growth of bacteria.
Antibiotic(s) has come to include a broader range
of antimicrobial compounds, including anti-fungal and other
compounds.
These include, for example, the beta-lactam antibacterials, which
include the penicillins (produced by fungi in the genus
Penicillium),and the cephalosporins.
Compounds that are still isolated from living organisms are the
aminoglycosides, whereas other antibacterial agents—for example,
the sulfonamides,and the quinolones, are produced by chemical
synthesis.
Classification of Antibiotics:
According to agent action:
In this classification antibacterial agents are divided into two broad
groups according to their biological effect on
microorganisms: bactericidal agents kill bacteria, and bacteriostatic
agents slow down or stall bacterial growth.
Bactericidal antibiotics:
Antibiotics that inhibit cell wall synthesis: the Beta-lactam antibiotics
(penicillin derivatives, and cephalosporins).
Aminoglycosidic antibiotics are usually considered bactericidal,
although they may be bacteriostatic with some organisms.
a
Bacteriostatic antibiotics limit the growth of bacteria by interfering
with bacterial protein production, DNA replication, or other aspects
of bacterial cellular metabolism.
This group includes:
Tetracyclines, sulphonamides , trimethoprim ,chloramphenicol,
and macrolides.
The Beta-lactam: Penicillin.
The Tetracycline.
a
The Erythromycin.
The Trimethoprim
The chloramphenicol
Antibiotic sensitivity test:
Antibiotic sensitivity is a term used to describe the susceptibility
of bacteria to antibiotics.
Antibiotic susceptibility testing (AST) is usually carried out to
determine which antibiotic will be most successful in treating a
bacterial infection in vivo.
Testing for antibiotic sensitivity is often done by the Kirby-Bauer
method ( Disc-diffusion method).
Other methods to test antimicrobial susceptibility include the Etest (also based on antibiotic diffusion).
Agar and Broth dilution methods for Minimum Inhibitory
Concentration determination.
Antibiotic sensitivity test:
Antibiotic sensitivity Test : antibiotic have been placed on an agar
plate growing bacteria. Bacteria are not able to grow around
antibiotics to which they are sensitive.