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Chapter 2.
Bacterial Morphology and Structure
Methods to study bacterial morphology and
structure
Optical methods
• The light microscope 100-power objective lens with a 10-power ocular
lens magnifying the specimen 1000 times
• The electron microscopes
Transmission electron microscope (TEM)
can resolve particles with 0.001 μm in size
Scanning electron microscope (SEM)
is particularly providing three-dimensional images of
the surface structure of microscopic objects
• Phase microscope
observe living cells
Treponemes are recognized by their characteristic
corkscrew shape and deliberate forward and
backward movement with rotation about the
longitudinal axis.
Electron micrograph of Spirillum serpens, showing
lophotrichous flagellation (9000 x ).
Methods to study bacterial morphology and
structure
Staining methods
•Simple staining methods
•Differential staining methods (Gram stain, Acid-fast stain)
•Special staining methods
The spore staining method
The flagella staining method
The capsule staining
Negative staining
Size of bacteria
• Unit for measurement:
Micron or micrometer, μm:
1μm=10-3 mm
• Size:
Varies with kinds of bacteria, and
also related to their age and
external environment.
Shape of bacteria
• Spherical (Cocci, sing. Coccus )
• Rods (Bacilli, sing. Bacillus)
• Spiral (Spiral bacteria)
vibrio
spirillum
helicobacterium
Spherical bacteria
Different arrangements depending on the
plane of division
Diplococci: Pair of cells divide in one plane
Streptococci: Chain of cells formed by dividing
in one plane several times
Tetrad: Divide in two planes
Sarcinae: Divide in three planes
Staphylococci: Divide in many planes and
remain together as a cluster
Rod-shaped bacteria
Considerable variation in length and diameter: 0.51 um in width and 2-5 um in length.
Most of rod-shaped bacteria are single
arrangement.
Diplobacilli: Bacilli that remain in pairs after they
divide.
Streptobacilli: Bacilli that remain in chains after they
divide.
Coccobacilli: A short Bacilli that nearly looks like a
cocci.
Spiral-shaped bacteria
Divided into:
Vibrio: comma shaped
Spirillum: helical rigid (spirillum) or flexible (spirochete)
Helicobacterium: curved rod-shaped
Structure of Bacteria
Bacteria are prokaryotes
•Except ribosomes, lack membrane-bound
cytoplasmic organelles
•The plasma membrane performs many of the
functions carried out by membranous organelles in
eukaryotes
•Have a nucleoid (nuclear body) rather than an
enveloped nucleus
•Despite their lack of complexity compared to eukaryotes, a number
of bacterial structures may be defined:
Basic structure:
Cell wall
Cell membrane
Cytoplasm
Nucleoid
Specific structure:
Flagellum
Pilus
Capsules and slime layers
Endospores (spores)
•Only plasma membrane, which belonging to cell envelope, is the essential
component for any bacteria.
Important bacterial structures
Cell envelope
•The cell envelope may be defined as the cell membrane and
cell wall.
•Usually, bacterial cell envelopes fall into two major categories:
Gram positive and Gram negative.
•This is based on Gram staining characteristics that reflect major structural
differences between the two bacterial groups.
1884: Christian Gram: First publication for the Gram stain method
Editor's note: I would like to testify that I have found the Gram
method to be one of the best and for many cases the best method
which I have ever used for staining Schizomycetes.
Gram-positive cocci
Gram-negative bacilli
periplasmic space
Simplified diagram and electronic microscopy
pictures of the cell envelope of G+ and G- bacteria
Cell wall
Situation: outmost portion.
15-30 nm in thickness
10%-25% of dry weight.
The cell wall consists of the peptidoglycan (murein) layer
and attached structures.
peptidoglycan
•The peptidoglycan is a single bag-shaped, highly cross-linked
macromolecule that surrounds the bacterial cytoplasmic
membrane and provides rigidity (which decides the shape of a
bacterium) .
•It is huge (billions in molecular weight).
•Peptidoglycan is found in all eubacteria except Chlamydia and
Mycoplasma.
peptidoglycan
•Peptidoglycan consists of a glycan
(polysaccharide) backbone consisting
of alternatively residues of N-acetyl
muramic acid and N-acetyl
glucosamine connected by -1,4
linkage.
•with tetrapeptide side chains usually containing D- and L- amino acid residuals,
and in some instances containing diaminopimelic acid (DAP) residual.
•The side chains are cross-linked by peptide bridges. These peptide bridges vary in
structure among bacterial species (gram-negative bacteria have no peptide
bridges)
•Lysozyme can block the -1,4 linkage.
•So lysozyme can kill G+ and G- bacteria by destroying their glycan
backbone .
•Penicillin can block the linkage of tetrapeptide side chains and peptide bridges.
• So penicillin can kill G+ bacteria by inhibiting their peptidoglycan synthesis.
Cell wall: characteristics of gram-positive bacteria
•A gram-positive bacterium has a
thick cell wall (20-80 nm).
•The peptidoglycan layer of it is
thick (15-50 layers).
•It may also include special
components such as teichoic
acids and proteins.
•Teichoic acids are water-soluble, a polymer of polyphosphates.
•Wall teichoic acids are linked to the peptidoglyacan.
•Membrane teichoic acids are anchored in the cytoplasmic membrane.
Lipoteichoic acids
Cell wall: characteristics of gram-negative bacteria
•A gram-negative cell has a
thin (10-15 nm) cell wall but
it is more complex than
gram-positive cell walls, both
structurally and chemically.
•The peptidoglycan layer of
it is thin (1-2 layers).
•It has no any teichoic acids but has an unique outer membrane which is external
to the peptidoglycan layer.
•The space between cytoplasm and outer membrane is periplasmic space. It
contains transport, degradative, and cell wall synthesis proteins.
Outer membrane
Outer membrane of a G- bacterium contains lipopolysaccharide (LPS),
Phospholipids and lipoproteins.
A group of transmembrane proteins is known as porins which form channels to
allow passage of small hydrophilic nutrients (such as sugars, amino acids and
certain ions) through the outer membrane.
phospholipids
Lipopolysaccharide (LPS)
LPS is also called endotoxin because it is poisonous to mammal cells.
It consists of 3 regions: an external O antigen, a middle core, and
an inner lipid A.
• The O antigen (somatic antigen) is a specific
polysaccharide region composed of repeating
units by some specific monosaccharides.
Highly variable
•The core polysaccharide is similar within a
single genus.
•lipid A contains β hydroxy fatty acids
(uncommon in nature). This molecule
displays endotoxin activity.
What is the difference between
G+ cell wall and G- cell wall?
Property
Gram positive Gram negative
Number of layers in wall 1
2
Peptidoglycan content
>50%
10-20%
Teichoic acid
+
-
Outer membrane
-
+
lipopolysaccharide
-
+
Sensitive to penicillin
yes
Less sensitive
Digested by lysozyme
yes
weakly
Cell wall: function
• Maintaining a bacterial characteristic shape
• Possessing the resistance to osmotic pressure
• Providing a platform for surface appendages such as flagella
and pili.
• Providing a pathogenic function to adhere host cells
For G+ bacteria, the major adhering molecular is teichoic acids.
For G- bacteria, the major adhering molecular is the pili as well as
some of the outer membrane proteins).
Cell wall: function
• Providing attachment sites for bacteriophages
• Play an essential role in cell division
• Participating the bacterial material exchange
• Be the sites of major antigenic determinants of the cell surface.
For G+ bacteria, the major surface antigen is the teichoic acids and then
the polysaccharides and the peptidoglycan.
For G- bacteria, the major surface antigen is the LPS and then the outer
membrane proteins.
Wall-less forms of bacteria
• When bacteria are treated with 1) enzymes that are lytic for the cell wall
e.g. lysozyme or 2) antibiotics that interfere with biosynthesis of
peptidoglycan, wall-less bacteria are often produced.
• Usually these treatments generate non-viable organisms. Wall-less
bacteria that can not propagate are referred to as spheroplast (G-) or
protoplast ( G+).
• Occasionally, some wall-less bacteria can propagate (so called as L forms
of bacteria).
Bacterial L form
•L-forms of bacteria are artificial cell wall-less organisms (means they are
different from Mycoplasma) first found by the Lister institute in 1935.
•Can be produced from normal bacteria by damaging the cell wall (by
Penicillin, salt solutions, antisera etc.).
•L-form cells are difficult to cultivate and usually require a medium with a
right osmotic strength.
•They can reconvert to wild type. Rarely they can develop to stable L-forms.
•May cause chronic infections which are relatively resistant to antibiotic
treatment and difficulty to detectable using routing serological diagnosis
methods, because the target sites of some antibiotics (penicillin) and somatic
antigens (O antigen) are absent.
Electron micrograph of Staphylococcus
A: L-form
B: Reconvert to wild type
•Because of the lack of a rigid cell wall, L-form cells generally
show various shapes (such as filar) compared to the original
shapes and gram-negative staining.
Important bacterial structures
Cell membrane
Site of biosynthesis of DNA, cell wall polymers and membrane lipids.
Selective permeability and transport of solutes into cells
Electron transport and oxidative phosphorylation
Excretion of hydrolytic exoenzymes
Mesosomes
Mesosomes are specialized structures folded invaginations in
the plasma membrane of bacteria
divided into septal and lateral mesosome.
Functions of the cytoplasmic membrane
(1) selective permeability and transport of solutes;
(2) electron transport and oxidative phosphorylation, in aerobic
species;
(3) excretion of hydrolytic exoenzymes;
(4) bearing the enzymes and carrier molecules that function in the
biosynthesis of DNA, cell wall polymers, and membrane lipids;
(5) bearing the receptors and other proteins of the chemotactic and
other sensory transduction systems.
Important bacterial structures
Cytoplasm
•
Composed largely of water, together with proteins,
nucleic acid, lipids and small amount of sugars and salts
•
Ribosomes:
Plasmids: extrachromosomal genetic elements
capable of autonomous replication
• Inclusions: sources of stored energy, e,g volutin
•
Ribosomes
•15-20 nm in diameter with 70S, distributed throughout the
cytoplasm
•sensitive to streptomycin
and erythromycin
• a workbench for protein
synthesis, numerous
• the target of antimicrobial
agents:
Streptomycin
Tetracyline
Plasmids
•Plasmids are small, circular / line, extrachromosomal double-stranded DNA.
•Usually present in multiple copies and
are capable of self-replication.
•Often code for pathogenic factors and
antibiotic resistant factors.
Are not essential for bacterial survival
Inclusions
are aggregates of various compounds that are normally involved in storing
energy reserves or building blocks for the cell. Inclusions accumulate
when a cell is grown in the presence of excess nutrients and they are
often observed under laboratory conditions.
granulose
Inclusions of Bacteria
Important bacterial structures
caryoplasm
• Lacking nuclear membrane,
absence of nucleoli, hence
known as nucleic material or
nucleoid, one to several per
bacterium.
Special bacterial structures
Capsules and slime layers
•Are structures surrounding the outside of the cell envelope.
•Usually, slime layer is thinner than capsule.
•They are usually demonstrated by the negative staining or
“capsule stain” which gives color to the background.
Capsules and slime layers
•They are usually composed of polysaccharide, however, in certain
bacilli they are composed of a polypeptide.
• Some strains within a species can produce a capsule, whereas the
others can not.
•They are not essential to bacterial viability.
•Capsules are often lost during in vitro culture.
•The capsules contribute to the invasiveness (virulence) of
pathogenic bacteria by protecting them from phagocytosis by
phagocytes.
Special bacterial structures
Flagellum: general description
• a thin, threadlike appendage on many bacterial cells that is responsible for
their motility
• Flagellum consist of a number of proteins including flagellin
• Are embedded in the cell membrane, extend through the cell envelope and
project as a long strand.
• The diameter of a flagellum is thin (20 nm) and long with a length 10
times of the bacterial cell diameter. Due to their small diameter,
flagellum cannot be seen under light microscope unless a special stain is
applied.
Flagellum: types
• Three types of arrangement are known:
monotrichous
single polar flagellum
lophotrichous
in one plane with cell
peritrichous
flagella distributed over the entire cell
Electron graph
monotrichous
lophotrichous
peritrichous
Flagella stain
Light microscopy graph
Flagellum: composition
• Composition: protein
A bacterial flagellum is made up of several thousand molecules of a protein
subunit called as flagellin. They move the bacterial cell by rotating with a
propeller like action.
Flagellins are highly
antigenic (H antigens).
Flagella: function
•Motility of bacteria:
Flagella enable bacteria to taste their environment and respond to
specific chemical foodstuffs or toxic materials and move towards or away from
them (chemotaxis).
•Identification of bacteria:
According to the mobility and antigenicity of flagellins (H antigen).
•Possible Pathogenesis:
In the past, flagella were considered not to be relative to bacterial
pathogenicity. In the recent data, the flagella of some bacterial species have
adhering ability to host cells.
Special bacterial structures
Pilus
•Pili are hair-like appendages of many bacterial cells.
•shorter and thinner than flagella, only visible under electron
microscope.
•receptors for certain bacterial viruses. Chemical nature is pilin
Pilus
•Pili are composed of structural protein subunits termed pilins.
•Two types can be distinguished:
Common pili
•Shorter, thinner, numerous for one bacterium
•Relative to bacterial adhesion (adhering to host cells)
•Contribute to virulence of some pathogenic bacteria
Sex pili
•Longer, coarser, only 1-4 for one bacterium
•Relative to bacterial conjugation (a pattern of bacterial
genetic material exchanges)
•The recent data revealed that sex pili of some bacteria has
the ability to adhere host cells.
Common pili
Sex pili
Recipient
Donor bacterium
Electron graph of pili
Special bacterial structures
Endospores (spores)
Endospores (Spores)
• Under adverse conditions, such as nutrient/water depletion, some
bacteria form a thick wall inside the cytoplasmic membrane leading
to a resting stage known as spores.
Endospores (spores)
• Spore is an intracellular body which found in some species of
bacteria. One spore-forming bacterium can only produce one spore.
Spore can be seen after staining with dyes by heating the
preparation. Sometimes, it can also be seen as a colorless area by
using conventional bacterial staining methods.
Endospores (Spores)
•A highly resistant resting structure produced within a bacterium.
It enables the bacterium in soil to survive many years. It can
withstand heating, freezing, chemicals and radiation.
•Spore has no ability for propagation (multiplication).
•Under favorable conditions, one spore germinates into one
vegetative bacterial cell (propagation / multiplication). But spore
has no ability for proliferation.
•Spores are commonly found gram-positive bacilli in the genera
Bacillus and Clostridium.
•The different sizes, shapes and positions of spores will help us to
identify spore-forming bacteria.
Contents of endospore
Core
spore wall /core
Cortex
Coat
exosporium
Process of endospore formation:
Sporogenesis/sporulation
Classification of bacteria
•Since there is the diversity of organisms including bacteria,
it is necessary to group similar organisms together and
organize these groups in a non-overlapping hierarchical
arrangement.
•Taxonomy is the science of biological classification, which
includes
the
nomenclature.
two
disciplines
of
classification
and
Classification of bacteria
The bacteria are classified in a hierarchic system
based on phenotypic characteristics (morphological,
physiological, and chemical characteristics).
The basic unit is the species. Similar and related
species are classified in a single genus and related
genera are placed in a single family.
Classification of bacteria
•In formal terms, the prokaryotes are classified in phyla,
classes, orders, families, genera, and species, plus subtaxa
if any:
Family (familia)
Enterobacteriaceae
Genus
Escherichia
Species
E. coli
Variety or type
Serovar O157:H7
Strain
xyz
nomenclature of bacteria
•Scientifically, species is named by a Latin binominal. Each species
receives a name of two parts, and the first is the genus name and the
second is the species name.
•The genus name is always capitalized but the species name is not.
Both species and genus are usually in italics.
•The genus name may be abbreviated by just using its initial letter.
•While described in Chinese, the genus is followed by species name.
Staphylococcus aureus
S. aureus
Genus
species
Oxygen
• Obligate aerobe
an organism that grows only in the presence of oxygen
• Microaerophile
an organism that requires a low concentration of oxygen for
growth
• Facultative anaerobe
an organism that grows with or without oxygen
• Oblige anaerobe
an organism that grows only in the absence of oxygen
Temperature
• Psychrophile (15-20 ℃ )
an organism that grows best at cold temperatures
• Mesophile
an organism that grows best at moderate temperatures
• Thermophile (50-60 ℃ )
an organism that grows best at high temperatures
pH
• Acidophile
an organism that grows best at a pH below 6
• Neutrophile
an organism that grows best at a pH between 6 and 8
• Alkalophile
an organism that grows best at a pH above 8
Some other features that have been used
to classify bacteria
• Gram stain (G+/G-)
• Cell shape
(coccus/ bacillus/ spiral bacterium)
• Ability to form spores
(spore-forming/non-spore-forming clostridia)
Summary
Structure of bacteria include essential structures of cell wall, cell
membrane, cytoplasm, and nuclear material (nucleoid).
Some bacteria also have one or more of the particular structures
of capsule, flagella, pili, endospores.
The difference between G+ cell wall and G- cell wall
Medical importance of four special structures