Bacteria

Morphology and fine structure of bacteria.

Cell wall structure in details.

Cultivation of bacteria.

Reproduction and growth.

Growth Kinetics.

Isolation and preservation.

 Basic unit of all living things is the cell

Unicellular Multicellular

Bacteria Fungi
Protozoa Algae
Some algae, fungi Higher living forms

Differentiation by size, structural complexity and

organization of genetic material

Procaryotic
Eucaryotic
Similarities
• Cell contents bounded by a plasma membrane
• Genetic information encoded on DNA
• Ribosomes act as site of protein synthesis

Differences
Eubacteria
Archaebacteria
Metabolic properties
Pathogenicity
Nutritional requirements
Staining reactions
Antigenic properties
Bacteria are the smallest and most numerous organisms.

Bacterial cell structure is more complex than commonly

supposed.

Bacteria exhibit considerable diversity in both structure

and metabolism.

Bacteria are responsible for many diseases but also make

important contributions to ecosystems.
 Why are bacteria so small?
Size affects ‘surface to volume’ ratio

Advantages of large S/V:

diffusion rates
metabolic rates
reproductive rates

Limits to size reduction?

‘defective’ bacteria

Really big bacteria?

-- Epulopiscium fishelsoni

Epulopiscium web site

http://www.micro.cornell.edu/cals/micro/research/labs/
angert-lab/epulopiscium.cfm
Characteristics of Bacteria

Size and Shape

Genetic Information
Circular DNA, Single chromosome, Replication,
Plasmid

Cell structure
Cell wall, Peptidoglycan, EPS, Flagella, Pili
 The diversity of bacteria
Pseudomonas aeruginosa, Streptococcus. The spherical
a rod-shaped, flagellated individual bacteria (cocci)
bacterium (bacillus). adhere in chains in the
members of this genus
Spirillum volutans, one of
the spirilla. This large
bacterium, which occurs
in stagnant fresh water,
has a tuft of flagella at
each end
Bacterial Morphologies
 Other Cell Shapes

Hyphomicrobium,
electron
micrograph with
negative staining.

Myxococcus stipitatus
fruiting body composed of
thousands of cells.
 Bacterial Structures
• Flagella
• Pili
• Fimbriae
• Capsule
• Plasma Membrane
• Cytoplasm
• Cell Wall
• Lipopolysaccharides
• Teichoic Acids
• Inclusions
• Spores
 Cytoplasm
• 80% Water {20% Salts-Proteins)
– Osmotic Shock important
• DNA is circular, Haploid
– More efficient; grows quicker
– Mutations allow adaptation to environment
quicker
• Plasmids; extra circular DNA
– Antibiotic Resistance
• No organelles (Mitochondria, Golgi, etc.)
Genetic Material
A closed circle of double stranded DNA, associated with certain
bacterial proteins (not histones).

Escherichia coli is around 3–4 μm in length, but contains a DNA

molecule some 1400 μm in length!)

Plasmid: Additional DNA in the form of small, self-replicating

extra-chromosomal elements called plasmids.

These do not carry any genes essential for growth and

reproduction, and thus the cell may survive without them.

They include genes encoding toxins or resistance to antibiotics,

and can be passed from cell to cell.
Ribosomes

composed of a complex of protein and RNA, and are the site of

protein synthesis in the cell.
Ribosomes
Ribosomes
Many ribosomes may simultaneously be attached to a single
mRNA molecule, forming a threadlike polysome.
Inclusion bodies

Within the cytoplasm of certain bacteria may be found granular

structures known as inclusion bodies.

These act as food reserves, and may contain organic compounds such
as starch, glycogen or lipid.

In addition, sulphur and polyphosphate can be stored as inclusion

bodies, the latter being known as volutin or metachromatic granules.

Eg: Magnetosomes, which contain a form of iron oxide, help some

types of bacteria to orientate themselves downwards into favourable
conditions,

whilst gas vacuoles maintain bouyancy of the cell in blue greens and
some halobacteria.
 Functions

1) They are an essential structure for viability.

2) They are composed of unique components.

3) They are one of the most important sites for attack by antibiotics. 

4) They provide ligands for adherence and receptor sites for drugs or
viruses. 

5) They cause symptoms of disease in animals. 

6) They provide for immunological distinction and immunological

variation among strains of bacteria.
 Gram positive
Peptidoglycan-Thicker (20 to 80 nanometers)
forms around 90% of the dry weight of Gram-
positive bacteria.

Polysachharides –
Teichoic acids-
polymer of glycerol
and ribitol
phosphates. 
Also known as murein , is a polymer consisting of sugars and amino acids that
forms a mesh-like layer outside the plasma membrane of bacteria forming the cell
wall.

Which consist of three parts back bone,

N-acetylglucosamine and

N-acetylmuramic acid

in alternating chains interconnected by tri, tetra or penta pedtide chains.

b 1-4
O
 alanine

glutamic acid

diaminopimelic acid

alanine
Gram positive
 Gram positive

a glycine pentapeptide
(gly-gly-gly-gly-gly-)

 Staphylococcus aureus 
Gram negative

Transmembrane protein: green, yellow, purple

Gram negative
Gram negative
 Lipopolysaccharide (LPS)
• Endotoxin or Pyrogen
– Fever causing
– Toxin nomenclature
• Endo- part of bacteria
• Exo- excreted into environment
• Structure
– Lipid A
– Polysaccharide
• O Antigen of E. coli, Salmonella
• G- bacteria only
– Alcohol/Acetone removes
 LPS (cont’d)
• Functions
– Toxic; kills mice, pigs, humans
• G- septicemia; death due to LPS
– Pyrogen; causes fever
• DPT vaccination always causes fevers
– Adjuvant; stimulates immunity
• Heat Resistant; hard to remove
• Detection (all topical & IV products)
– Rabbits (measure fever)
– Horse shoe crab (Amoebocytes Lyse in
presence of LPS)
 LPS (cont’d.)
• Appearance of Colonies
– Mucoid = Smooth (lots of LPS or capsule)
– Dry = Rough (little LPS or capsule)
• O Antigen of Salmonella spp. and E. coli
– 2,000 different O Ags of Salmonella spp.
– 100’s different O Ags of E. coli
• E. coli O157
• O Ags differ in Sugars, not Lipid A
 Cell Membrane
• Bilayer Phospholipid
• Water can penetrate
• Flexible
• Not strong, ruptures easily
– Osmotic Pressure created by cytoplasm
Flagella
 Flagella
• A large number of bacteria are motile.

• Most possess one or more flagella on their surface that allow them to swim.

• Bacterial flagella are tiny hair like organelles of locomotion.

• The straight line movement is called a run and the turn is called a tumble.

• Motility of eukaryotic flagella is dependent upon ATPase activity .

• Their fine protein structure requires special staining techniques for

demonstrating them with the light microscope.

• The pattern of flagellation is an important feature in identification of motile

bacteria.
Flagella: complex organ of motility, a “motor”
A = monotrichous B = amphitrichous C = lophotrichous D = peritrichous

Example ????
 Flagella
• Protein-70-81%
• Lipid-13-23%
• Carbohydrate-1-6%

• The ultra structure of the flagellum of E. coli is illustrated.

• About 50 genes are required for flagella synthesis and function.

• The flagella apparatus consists of several distinct proteins.

Bacterial flagella are polymers approximately 15 nm in
diameter and as much as 15 μm in length, composed of a
protein called flagellin.

Each flagellum has a hollow core.

At its base, each flagellum has a rotory

motor ( 200-1000 rpm).

Three parts: Basal body, hook and filament

Basal body: M and S rings, located in the plasma
membrane, comprise the rotor apparatus
The outermost rings, the P and L rings, located in the
periplasm and the outer membrane respectively, function as
bushings to support the rod.
where it is joined to the hook of the filament on the cell
surface.
Deriving forces of flagellar motion

ATP is not used for flagellar motion.

The necessary free energy is derived from either the proton

gradient or sodium ion gradient that exists across the plasma
membrane.

Hence, either proton motive force or sodium motive force

facilitates flagellar motion.

Flagella do not rotate at a constant speed but instead can increase

or decrease their rotational speed in relation to the strength of
the proton motive force.
Proton motive force

The bacterial flagellum is driven by a rotary engine.

The engine is powered by proton motive force, i.e., by the flow

of protons (hydrogen ions) across the bacterial cell membrane due to
a concentration gradient set up by the cell's metabolism.
Sodium motive force

In some bacteria especially marine bacteria, ion channels

eject sodium ions from the cell.

This leads to formation of sodium ion gradient. This gradient

can also be used as intermediate energy storage for flagellar
rotation.

This gradient is a form of sodium motive force which assists

in flagellar rotation.
Flagellar Motor

Five components crucial to motor function have been identified.

MotA and MotB are membrane proteins.

Approximately 11 MotA - MotB pairs form a ring around the

base of the flagellum.

The proteins FliG, FliM, and FliN are part of a disc-like structure
called the MS (membrane and supramembrane) ring, with
approximately 30 FliG subunits coming together to form the ring.
Flagellar Motion

The MotA - MotB pair and FliG combine to create a proton

channel that drives rotation of the flagellum.

FliG serves as the rotating proton carrier

A proton from the periplasmic space passes into the outer half
channel and is transferred to an FliG subunit.

The MS ring rotates, rotating the flagellum with it and allowing

the proton to pass into the inner half-channel and into the cell.
 Filament
Hook

Motor
 Lab technique: Plate assay

Motile : Swarming Non- Motile : No Swarming

Lab technique:

Tube assay Hanging drop

Non- Motile Motile

Sensory Role of Flagella: Bacterial Chemotaxis

In the presence of a gradient of certain substances such as

glucose, bacteria swim preferentially toward the direction of the
higher concentration of the substance.

Such compounds are referred to as chemoattractants (positive

chemotaxis).

Bacteria also swim preferentially away from potentially harmful

compounds such as phenol, a chemorepellant (negative
chemotaxis).

The process of moving in specific directions in response to

environmental cues is called chemotaxis.
Spore
Spore
• Resistant structure
– Heat, irradiation, cold
– Boiling >1 hr still viable
• Takes time and energy to make spores
• Location important in classification
– Central, Subterminal, Terminal
• -spores
– Used for Bacillus stearothermophilus quality
control of heat sterilization equipment
• Bacillus anthracis - spores
– Used in biological warfare
 Spores Structure
• Spore Core – contains DNA and
cytoplasm

• Core Membrane – similar to cell

membrane

• Cortex – thickest spore layer,

made of loose peptidoglycan

• Inner/Outer Membranes (Spore

Coat) – made of protein,
impermeable, protects spore from
chemicals

• Exosporium Basal Layer – made

of glycoprotein and lipoproteins
Protein:
1) RecA is a 38 kilodalton protein essential for the repair and
maintenance of DNA : surrounds whole DNA

2) Div IVA leads to the initiation of cell division: polar side and
stretch DNA towards pole.
 Some spore forming bacteria are
capable of causing disease
• Clostridium botulinum – botulism
• Clostridium perfingens – gas gangrene
• Clostridium tetani – tetanus
• Bacillus anthracis – Woolsorter’s Disease and
wound infections
• The Schaeffer-Fulton Stain Procedure is used to
differentiate between endospores and vegetative
cells
Fimbriae and pili
Fimbriae and pili

They are used for attachment and transfer

of DNA rather than for motility.

These structures, which consist of a

protein called pilin arranged helically
around a central core.
Fimbriae and pili

The main difference between pili and

fibriae is,

pili are found only in gram negative

bacteria

but fibriae is found in both, gram negative

and gram positive bacteria.
Fimbriae (singular: fimbria) help cells adhere to surfaces.

They can number anywhere from a few to several hundred per cell

They are involved in forming biofilms - Fimbriae can also help bacteria adhere to
epithelial surfaces in the body.
E. coli 0157 - Pili (s ingular: pilus) are involved in twitching and gliding motility.

also, involved in DNA transfer, such pili are called conjugation (sex) pili.

Pili are usually longer than fimbriae and number only one or two per cell.
Fimbriae or
adhesin
 Mesosome
In prokaryotic cell division, called binary
fission, a membrane attachment mechanism is
used to allocate chromosome copies to the
two daughter cells.

Attachment of bacterial chromosomes,

indicating the possible role of the mesosome
(an inward fold of the cell membrane) in
ensuring the distribution of the
"chromosomes" in a dividing cell.

Upon attachment to the plasma membrane,

the DNA replicates and reattaches at separate
points. Continued growth of the cell gradually
separates the chromosomes.
Capsule
 Capsule
Capsule is a 
Gelatinous layer covering the entire bacterium. Composed of polysaccharide (i.e. poly:
many, saccharide: sugar). These polymers are composed of repeating oligosaccharide
units of two to four monosaccharides.

Virulence determinants:

Saving engulfed bacteria from the action of neutrophil:

Identification of bacteria:

Development of Vaccines: Polyvalent (23 serotypes) polysaccharide vaccine

of Streptococcus pneumoniae capsule. Polyvalent (4 serotypes) vaccine of Neisseria
meningitidis capsule.

Initiation of infection:
 Capsule
Capsule is a 
Gelatinous layer covering the entire bacterium. Composed of polysaccharide (i.e. poly:
many, saccharide: sugar). These polymers are composed of repeating oligosaccharide
units of two to four monosaccharides.

Examples of Capsulated bacteria

Mnemonics to remember capsulated bacteria–

Some Killers Have Pretty Nice Capsule

Streptococcus pneumoniae
Klebsiella pneumoniae
Haemophilus influenzae
Pseudomonas aeruginosa
Neisseria meningitidis
Cryptococcus neoformans
 Bacterial Cytoskeleton
Shape determining proteins

Eukaryotes possess three cytoskeletal elements:

• actin filaments,

• microtubules, and

• intermediate filaments

Homologues of all three types of eukaryotic

proteins have been identified in bacteria
 Bacterial Cytoskeleton

FtsZ protein in a chain of Bacillus subtilis cells; FtsZ-green fl uorescent (GFP)

fusion protein viewed by fl uorescence microscopy.
 Bacterial Cytoskeleton

The MreB-like cytoskeletal protein (Mbl) of B. subtilis; Mbl-GFP in live cells was
examined by fl uorescence microscopy. With this method, Mbl seems to form
helices (arrows).
 Bacterial Cytoskeleton

CreS (crescentin), in red, of Caulobacter crescentus. The DNA in the cells was stained
blue with DAPI.
 Classification Based on Nutrition

1. Autotrophs – can synthesize all their organic compounds by

utilizing atmospheric CO2 & N2.

2. Heterotrophs – unable to synthesize their own metabolites &

depend on preformed organic compounds.
Growth Factors

Some bacteria require certain organic compounds in

minute quantities – Growth Factors OR Bacterial Vitamins.

• It can be :

1. Essential – growth does not occur in their absence.

2. Accessory – they enhance growth, without being

absolutely necessary for it.
Bacterial Nutrition

• Water constitutes 80% of the total weight of bacterial

cells.

• Proteins, polysaccharides, lipids, nucleic acids,

mucopeptides & low molecular weight compounds make up
the remaining 20%.

• For growth & multiplication, the minimum nutritional

requirements are water, a source of carbon, a source of
nitrogen & some inorganic salts
Based on Nutritional Requirement

• Phototrophs – Bacteria which derive their energy from

sunlight. Eg. ?

• Chemotrophs – Bacteria which derive energy from

chemical reactions. Eg. ?

• Organotrophs : require organic sources of hydrogen. Eg. ?

• Lithotrophs : require inorganic sources of hydrogen like

NH3, H2S. Eg. ?
The Effect of Oxygen on the Growth of Various Types of Bacteria
The Effect of Oxygen on the Growth of Various Types of Bacteria
The Effect of Oxygen on the Growth of Various Types of Bacteria
The Effect of Oxygen on the Growth of Various Types of Bacteria
The Effect of Oxygen on the Growth of Various Types of Bacteria
Example ???