Bacteria – Morphology &
Classification
Additional Organelles

Plasmid –
Extranuclear genetic elements consisting of DNA

Transmitted to daughter cells during binary fission

May be transferred from one bacterium to another

Not essential for life of the cell
1.
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Confer certain properties e.g. drug resistance,
toxicity
2.
Capsule & Slime layer –
 Viscous layer secreted around the cell wall.
 Polysaccharide / polypeptide in nature
a)
Capsule – sharply defined structure,
antigenic in nature
• Protects bacteria from lytic enzymes
• Inhibits phagocytosis
• Stained by negative staining using India Ink
• Can be demonstrated by Quellung reaction
(capsule swelling reaction)
b)
Slime layer – loose undemarcated secretion
Functions of capsule:
1.
2.
3.
4.
Antiphagocytic
Adherence
Resistance to dessication.
Excludes viruses and hydrophobic molecules
Capsule Demonstration
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Staining
Positive: crystal violet,
Hiss, Muir s
Negative : India ink
Electron microscopy
Serology- capsular antigens
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Quellung reaction
3. Flagella

Long (3 to 12 µm), filamentous surface appendages

Organs of locomotion

Chemically, composed of proteins called flagellins
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The number and distribution of flagella on the
bacterial surface are characteristic for a given species
- hence are useful in identifying and classifying
bacteria
Flagella may serve as antigenic determinants (e.g. the
H antigens of Gram-negative enteric bacteria)
Presence shown by motility e.g. hanging drop
preparation
Ultra structure of Flagella: Threadlike locomotor
appendages
Types of flagellar arrangement
Polar/ Monotrichous – single
flagellum at one pole ex: Vibrio
Lophotrichous – tuft of flagella at
one pole ex: Pseudomonas
Amphitrichous – flagella at both
poles. Ex: Spirillum
Peritrichous – flagella all over. ex :
E coli, Salmonella
Amphilophotrichous – tuft of
flagella at both ends
Demonstration:
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wet mount method,
Staining :
Kirkpatrick, Leifsons ,Loefflers etc.
Electron microscopy
Additional Organelles
Fimbriae/ Pili –
4.
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Thin, hairlike appendages on the surface of many
Gram-negative bacteria

10-20µ long, acts as organs of adhesion
(attachment) - allowing bacteria to colonize
environmental surfaces or cells and resist
flushing

Made up of proteins called pilins.
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Pili can be of two types –
Common pili – short & abundant
Sex pili - small number (one to six), very long
pili, helps in conjugation (process of transfer of
DNA)
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Demonstration: Electron microscopy.
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Function: Attachment to solid surfaces
Antigenic
Agglutination to RBCs
Additional Organelles
Spores –
5.

Highly resistant resting stages formed during
adverse environment (depletion of nutrients)

Formed inside the parent cell, hence called
Endospores
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Very resistant to heat, radiation and drying and
can remain dormant for hundreds of years.

Formed by bacteria like Clostridia, bacillus
The cycle of spore formation and germination
At the beginning of spore formation, a septum forms, separating the nascent spore from
the rest of the cell and all of the genetic material of the cell is copied into the newlyforming cell. The spore contents are dehydrated and the protective outer coatings are laid
down. Once the spore is matured it is released from the cell. On germination, the spore
contents rehydrate and a new bacterium emerges and multiplies.
Shape & position of bacterial spore
Oval central
Spherical central
Non bulging
Oval sub terminal
Oval sub terminal
Oval terminal
Spherical terminal
Free spore
Bulging
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Uses:
Bacillus stearothermophilus -spores
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Used for quality control of heat sterilization
equipment
Bacillus anthracis - spores
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Used in biological warfare
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Demonstration:
Unstained in Grams stain
Modified Zeihl Neelson, Malachite green,
Fuschin Nigrosin
Unstained wet preparation in phase contrast
microscopy
Pleomorphism & Involution forms
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Pleomorphism – great variation in shape & size of
individual cells e.g. Proteus species
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Involution forms – swollen & aberrant forms in
ageing cultures, especially in the presence of high
salt concentration e.g. plague bacillus
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Cause – defective cell wall synthesis
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THANK YOU
Microbial Growth 
refers to the increase in number of cells,
not the size of the cells
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Bacteria divide by binary fission
The interval of time between two cell divisions or the time
required by the bacterium to give rise to two daughter cells
under optimum conditions, is known as the generation time
or population doubling time.
Generation Time Under Optimal Conditions
Organism
Generation
Time
Bacillus cereus
28 min
Escherichia coli
12.5 min
Staphylococcus aureus (causes many types of infections)
27-30 min
Mycobacterium tuberculosis (agent of Tuberculosis)
18 – 24 hrs
Treponema pallidum (agent of Syphilis)
30 hrs
Lepra bacilli
20 days
• When
pathogenic bacteria multiply in host tissues, the
situation maybe intermediate between a batch culture and a
continuous culture; the source of nutrients maybe inexhaustible
but the parasite has to face the defence mechanisms of the
body.
•Bacteria growing on solid media form colonies.
•Each colony represents a clone of cells derived from a single
parent cell.
•In liquid media, growth is diffuse.
• When
a bacterium is seeded into a suitable liquid medium
and incubated, its growth follows a definite course.
•If bacterial counts are made at intervals after inoculation and
plotted in relation to time, a growth curve is obtained.
It shows 4 phases :
Lag,
Log
Exponential,
Stationary & phase of Decline.
Bacterial growth curve
1)
Lag phase:
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No cell division.
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The bacteria form the enzymes and molecules needed
for replication.
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Clinical significance: this phase = incubation period
of a disease.
2)
Logarithmic phase:

Rapid cell division occurs.

The number of living bacteria increases by time.

Clinical significance: this phase = symptoms and
signs of the disease.
Nutrients and conditions are not limiting
growth = 2n
or
X = 2nX0
Where X0 = initial number of cells
X = final number of cells
n = number of generations
3)
Stationary phase:

Nutrients are exhausted.

Waste products are accumulated.

The number of dying cells = number of new cells.

The number of living bacteria remains constant.
4) Decline phase:

Nutrients are more exhausted.

Waste products are more accumulated.

The number of dying cells > number of new cells.

The number of living bacteria decreases by time.

Clinical significance: this phase = recovery and
convalescence.
morphological and physiological alterations of the cells
in various stages of the growth curve
•Bacteria have the maximum cell size towards the end of the
lag phase.
•In the log phase, cells are smaller and stain uniformly.
•In the stationary phase, cells frequently are gram variable and
show irregular staining due to the presence of intracellular
storage granules.
•Sporulation occurs at this stage.
production of exotoxins & antibiotics
•Phase of Decline –involution forms(with ageing).
Nutritional Requirements
Growth of prokaryotes depends on nutritional factors as
well as physical environment
Main factors to be considered are:
Required elements
Growth factors
Energy sources
Major elements (CHONPS + K, Mg, Fe, Ca)
Carbon, oxygen, hydrogen, nitrogen, sulfur,
phosphorus, potassium, magnesium, iron, and calcium
Essential components for macromolecules
Trace elements (Co, Cu, Ni, Zn, Se, Mg)
Cobalt, zinc, copper, molybdenum and manganese
Required in minute amounts
Assist in enzyme function
Nutrient
Autotrophic bacteria sHeterotrophic bacteria
o Use simple inorganic
materials like CO2 as a source
of carbon and ammonium as a
source of nitrogen
o Require complex organic
materials
o They form complex organic
compounds from the simple
inorganic materials
o Can not form organic
compounds from the simple
inorganic materials
o Derive their energy:
From light (photosynthetic
bacteria)
Oxidation of inorganic
materials (chemolithotrophic
bacteria)
o Derive their energy:
Oxidation or fermentation of
organic compounds such as
glucose
Most bacteria of medical
importance are heterotrophic
bacteria
Growth factors
 These are organic compounds which bacteria must
contain to grow.
 But, they are unable to synthesize them
 So, they must be added ready formed to the culture
medium
 Examples: amino acids, vitamins, purines and
pyrimidines
Oxygen
Class
Obligate aerobe
Definition
Examples
Grow only in
Mycobacterium
the presence of
tuberculosis
O2
Obligate anaerobe
Can not grow in
Clostridia
the presence of
O2
Facultative
Can grow in the
Most bacteria of
anaerobe
presence or
medical
absence of O2
importance
Microaerophilic
Require low O2
Campylobacter
bacteria
tension
2H2O  2O2  2H2O2  O2
In the presence of oxygen, two toxic substances to the
bacteria are produced which are hydrogen peroxide and
superoxide anion.
In obligate aerobes and facultative anaerobes:
Catalase and peroxidase enzymes degrade hydrogen peroxide.
Superoxide dismutase enzyme degrades superoxide anion.
BU
T
In obligate anaerobes:
These enzymes are not present.
So, the presence of oxygen is toxic to them.
1.
Obligate Aerobes
2. Obligate Anaerobes
3. Facultative Aerobes
Facultative Anaerobes
Carbon dioxide

Most bacteria require CO2 in small concentration as that in
the air.

However, some bacteria require higher concentrations of
CO2.

These bacteria are called capnophilic bacteria.
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Example:

Neisseria requires 5-10% CO2
Temperature
Bacteria
Range of
temperature
Optimum
temperature
Mesophilic
18 - 42
37
Psychrophilic
5 - 30
15 - 20
Thermophilic
25 - 80
50 - 60
Psychrophile
0o to 18o C
Psychrotroph
20°C to 30°C
Important in food spoilage
Mesophile
25°C to 45°C
More common
Majority of human pathogens
Thermophiles
45°C to 70°C
Common in hot springs and hot water heaters.
Lipids in PM more saturated than mesophiles (higher melting points)
Hyperthermophiles
70°C to 110°C
Live at very high temperatures, high enough where water threatens to
become a gas
Usually members of Archaea
Found in hydrothermal vents
Most bacteria of medical
importance are mesophilic
bacteria
Hydrogen ion concentration
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Hydrogen ion concentration is called pH.
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Most bacteria grow within pH range 7.2-7.6
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However, some bacteria require different pH.
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Examples:

Vibrio cholera needs alkaline pH

Lactobacilli need acidic pH
pH is the negative logarithm of the hydrogen ion
concentration
Acidophiles grow best between pH 0
and 5.5
Neutrophiles grow best between pH
5.5 and 8.0
Alkalophiles grow best between pH
8.5 and 11.5
•
Alkalophiles grow best at pH 10.0 or higher
•
Despite wide variations in habitat pH, the internal pH
of most microorganisms is maintained near neutrality
either by proton/ion exchange or by internal buffering
•
Sudden pH changes can inactivate enzymes and
damage PMs
 Microbes that require a high water activity (near or at 1) are
termed nonhalophiles. (Halophile = salt-loving)
 Some bacteria require salt to grow and are called
halophiles. If a very high concentration of salt is required
(around saturation), 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.
Pressure:
Barotolerant organisms are adversely affected by
increased pressure, but not as severely as are
nontolerant organisms
Barophilic organisms require, or grow more rapidly in
the presence of, increased pressure
Radiation
Ultraviolet radiation damages cells by causing the
formation of thymine dimers in DNA
Photoreactivation repairs thymine dimers by direct
splitting when the cells are exposed to blue light
Dark reactivation repairs thymine dimers by excision
and replacement in the absence of light
Ionizing radiation such as X rays or gamma rays are
even more harmful to microorganisms than ultraviolet
radiation
Low levels produce mutations and may indirectly
result in death
High levels are directly lethal by direct damage to
cellular macromolecules or through the production
of oxygen free radicals
Measurement of
Microbial Growth
Growth in numbers can be studied by bacterial counts.
Two types of bacterial counts can be made- total count and
viable count.
The total count gives the total number of cells in the sample
irrespective of whether they are living or not.
It can be obtained by
1.Direct counting under the microscope using counting
chambers,
2.Counting in an electronic device as in the Coulter counter,
3.Direct counting using stained smears prepared by spreading a known
volume of the culture over a measured area of a slide,
4.Comparing relative numbers in smears of the culture mixed with known
numbers of other cells,
5. By opacity measurements using an absorptiometer or nephalometer,
6.By separating the cells by centrifugation or filtration and measuring their
wet or dry weight and
7. Chemical assay of cell components such as nitrogen.
• The
viable count measures the number of living cells, that is,
cells capable of multiplication.
• Viable counts are obtained by dilution or plating methods.
In the dilution method, the suspension is diluted to a point
beyond which unit quantities do not yield growth when
inoculated into suitable liquid media(extinction).
Several tubes are inoculated with varying dilutions and the
viable count calculated statistically from the number of tubes
showing growth.
DIRECT METHODS:
1) viable plate counts
2) membrane filtration
3) microscopic counts
4) electronic counters
5) Most Probable Number
Spectrophotometric determination
Light is scattered and is proportional to cell number
Linear relationship between absorbance and cell
density
Often written as % transmittance (as absorbance
increases, transmittance decreases)
Requires cultures to be ~107/ml and upwards (slight
turbidity)
Metabolic Activity