Bacterial Anatomy

CHAPTER TWO
 Bacteriology
A branch of Science that deals with the
study of Bacteria
REPRODUCTION
▪ Binary fission – “ division in half ”
✔Asexual reproduction of bacteria by a separation of the body into
two new body
✔a simple division of one cell into two cells following DNA
replication and the formation of separating membrane and cell wall
Binary fission
BACTERIAL
CELL
STRUCTURE
 All bacteria are prokaryotes
□unicellular organism that lack a true nucleus and nuclear membrane
□cells are about 10 times smaller than eukaryotic cells
□very simple cells when compared to eukaryotes and yet they are able to carry on the
necessary process of life
□do not contain organelles
□ all functions take place in the cytoplasm or cytoplasmic membrane
of the cell
□has cell wall composed of peptidoglycan, cell wall is absent in
eukaryote
 PROCARYOTIC CELL STRUCTURES

CELL INTERIOR (CYTOPLASMIC STRUCTURE)

▪ structures and substances that are bounded internally by the cytoplasmic
membrane composes the cell interior which includes:
1. cytosol or cytoplasm
2. endospores
3. nucleoid
4. plasmids
 CYTOPLASMIC STRUCTURE
Cytosol or cytoplasm
▪ where other functions not
conducted by the cell membrane
occurs
▪ Contains a complex mixture of
materials required by the cell for its
metabolic functions
▪ has granular appearance caused by
the presence of ribosomes and
inclusions (storage reserve granules)
 CYTOPLASMIC STRUCTURE
Bacterial Ribosomes
▪ often occurs in clusters
polyribosomes or polysomes
✔ consist of RNA and protein
✔ attached to cytoplasmic membrane
✔ site of protein biosynthesis
✔ 70s in size and dissociate into two
subunits, 50s and 30s in
S= Svedberg units
size •sedimentation rates (unit of time) during high speed
centrifugation
•50s binds with 30s = 70s (loss of surface area)
 Cytoplasmic granules
▪ maybe stained by use of a suitable st ain,
and then identified microscopically
▪ granules are storage deposits which m ay
consist of polysaccharides such as:
✔Glycogen – storage form of glucose
✔lipids such as poly-β hydroxybutyrate
✔polyphosphate – storage form for inorgan ic
phosphates

✔ sulfur
✔ iron
✔ other stored substances
Cytoplasmic Granules
▪ Metachromatic or volutin granules indicate accumulation of food
reserve
1. Babes –Ernst granules of Corynebacterium diphtheriae
2. Much granules of Mycobacteria spp.
 CYTOPLASMIC STRUCTURE
Bacterial endospores or spores
▪ produced by:
□ aerobic genus Bacillus
□ anaerobic genus Clostridia
▪ small, dormant (inactive
state), asexual spores that develop
inside the bacterial cell as a means of
survival
▪ Becomes vegetative (active state)
when harsh conditions are removed
▪ Not a means of reproduction
▪ Differe
nt from reproductive spores of
fungi
 CYTOPLASMIC STRUCTURES
Bacterial Spores or Endospores
□spores survive all sorts of trauma including:
▪ lack of nutrients (starvation)
▪ lack of water (dehydration)
▪ extreme heat or cold (temperature change)
▪ exposure to most chemicals such as disinfectants
▪ UV and gamma radiation
▪ desication or drying
 CYTOPLASMIC STRUCTURE
Bacterial Spores or Endospores
▪ thick protein coat makes them
highly resistant
▪ Calcium dipicolinate
✔ Up to 20% of the endospore dry
weight
✔ Stabilize DNA
✔ Dipicolinic acid – heat resistance of
the spore
✔ Calcium – aid in resistance to
heat and oxidizing agents
 CYTOPLASMIC STRUCTURE
Bacterial Spores or Endospores
▪ Highly refractile bodies in the cell
▪ Unstained areas in a cell with use of
traditional bacterial stains
▪ Schaeffer-Fulton stain – most
commonly used; stain the spore green and
bacterial bodies red
▪ Moeller stain – endospore stains
red while the rest stains blue
▪ Dorner’s stain – endospore stains red
in colorless cytoplasm against a dark
background
 CYTOPLASMIC STRUCTURE

Bacterial Spores or Endospores

▪ Sporulation – process by which spores are formed
▪ Spores can survive for many years, even for centuries
▪ Spores are the ones responsible for the perpetuation (to last
indefinitely) of the species, but not for multiplication
 CYTOPLASMIC STRUCTURES
Bacterial Spores or Endospores
▪Size, shape, and interior location of the spore can
be used as identifying characteristic
a) Terminal spores – produced at the end of the
bacterial cells
Clostridium tetani – terminal endospore,
giving the organism a drumstick, tennis
racquet-shaped, or lollipop-shaped
appearance
b) Subterminal spores – produced within the cell
and whether or not it causes a swelling of the cell
serves as clues to the identity of the organism
ex. Clostridium sporogenes
c) Central – ex. Clostridium botulinum
 CYTOPLASMIC STRUCTURE
Nucleoid
▪Genome consists of a single circular
chromosomes
▪bacterial chromosomes exists as a
diffuse nucleoid or chromatin body
▪attached to mesosome, sac-like
structure in the cell membrane

▪no nuclear membrane and mitotic

apparatus
▪serves as the control center of the
bacterial cell
 CYTOPLASMIC STRUCTURE

Plasmid
▪ other genetic elements that exist
independently in the cytosol
▪ Smaller circular molecules of
extrachromosomal circular DNA
▪ replicate independently of the main
chromosomes
 CYTOPLASMIC STRUCTURE

Plasmid
• can be readily transferred between partners when bacteria
conjugate
• Plasmids provide the cell with genes for:
✔Resistance to antibiotics
✔Metabolism of unusual nutrients not present in the normal environment
✔Other special contingencies
 CYTOPLASMIC STRUCTURE
PLASMIDS
▪ Classification based on function:
✔ Fertility F-plasmids, which
contain tra genes - capable
of conjugation and result in the
expression of sex pili
✔ Resistance R plasmids - contain
genes that provide resistance against
antibiotics or poisons
✔ Virulence plasmids - turn the
bacterium into a pathogen
 CYTOPLASMIC STRUCTURE
Plasmids
▪ Classification based on function:
✔ Col plasmids - contain genes that
code for bacteriocins,
proteins that can kill other bacteria
✔ Degradative plasmids, which
enable the digestion of unusual
substances, e.g. toluene and
salicylic acid
 CELL ENVELOPE STRUCTURE

Layers that surround the prokaryotic cell

▪ It comprises:
i. an outer membrane - in gram (-) bacteria only
ii. a cell wall - composed of the peptidoglycan macromolecule (also
known as the murein layer)
iii. periplasm - in gram (-) negative bacteria only
iv. the cytoplasmic membrane –deepest layer of the cell envelope
 CELL ENVELOPE STRUCTURE
Cytoplasmic (inner) or cell or plasma membrane present in
both gram (-) and gram (+)bacteria

▪ deepest layer of the cell envelope

▪ phospholipid bilayer
▪ heavily embedded proteins that
envelope the cytoplasm, including
number of enzymes vital to cellular
metabolism
▪ functionally similar to several of the
eukaryotic cell’s organelles
 CELL ENVELOPE STRUCTURE
Cytoplasmic (inner) or cell or plasma membrane present in
both gram (-) and gram (+)bacteria
▪ does not contain sterols, in contrast to
eukaryotes except for Mycoplasma
▪ contain sterol-like molecules called
hopanoids
▪ acts as osmotic barrier
▪ location of the electron transport
chain
 CELL ENVELOPE STRUCTURE
Function of the cytoplasmic or cell or plasma membrane
▪ Transport solutes into and out of the cell
▪ Housing enzymes involved in outer membrane synthesis, cell wall
synthesis, and in the assembly and secretion of extra-cytoplasmic and
extracellular substances
▪ Generation of chemical energy (i.e., ATP)
▪ mediation of chromosomal segregation during replication

▪ housing molecular sensors that monitor chemical and physical changes in

the environment
 CELL ENVELOPE STRUCTURE
Cell wall (murein layer) - also referred to as the peptidoglycan

▪ an essential structure found in nearly

all clinically important bacteria
▪ a rigid structure that gives the bacterial cell
shape and strength to withstand changes in
environmental osmotic pressures that would
otherwise result in lysis
 CELL ENVELOPE STRUCTURE
Cell wall (murein layer) - also referred to as the peptidoglycan

▪ Protects against mechanical disruption

of the cell
▪ offers some barrier to the passage of
larger substances essential to the
survival of bacteria
▪ its synthesis and structure have been
the primary target for development and
design of several antimicrobial agents
 CELL ENVELOPE STRUCTURE
Cell wall (murein layer) - also referred to as the peptidoglycan

▪ bacteria are grouped into Gram

positive and Gram negative based on
their difference in the cell wall structure
▪ modified cell wall for mycobacteria
are called acid fast cell wall
▪ Mycoplasma have no cell wall
▪ Gram Stain - most valuable tools for
identifying bacteria
 CELL ENVELOPE STRUCTURE
Cell wall (Gram-positive bacteria)
▪ consist mainly of a very thick
peptidoglycan layer (0.02 – 0.06 nm)
□Glycan (polysaccharide) chains of
alternating N-acetyl-d-glucosamine
(NAG) and
□N-acetyl-d-muramic acid (NAM)
▪ antibiotics effective against gram (+)
organisms act by preventing synthesis of
peptidoglycan
 CELL ENVELOPE STRUCTURE
Cell wall (Gram – positive bacteria)
▪ peptidoglycan layer does not block
diffusion of low molecular weight
compounds, so substances that damage
the cytoplasmic membrane (such as
antibiotics, dyes, and detergents) can
pass through
 CELL ENVELOPE STRUCTURE

Cell wall (Gram – positive bacteria)

▪ contains teichoic acid –

anchored to the peptidoglycan
▪ lipoteichoic acid molecules
– anchors to the plasma membrane
▪ Antigenic polysaccharides
may be present on peptidoglycan
layer surface
 CELL ENVELOPE STRUCTURE
Cell wall (Gram - negative bacteria)
consists of two layers:
▪ inner peptidoglycan (0.01nm) –
much thinner than in gram positive
cell walls
▪ outer membrane - unique to gram
negative bacteria that consist of:
✔protein
✔phospholipids
✔lipopolysaccharide layers (LPS)
 CELL ENVELOPE STRUCTURE

Cell wall (Gram - negative bacteria)

✔lipopolysaccharide (LPS)
blocks the passage of
substances (such as antibiotics,
dyes, and detergents) to the
peptidoglycan layer and sensitive
inner cytoplasmic membrane
 CELL ENVELOPE STRUCTURE
Cell wall (Gram - negative bacteria)
LPS contains three regions: a.antigenic
O specific polysaccharide b.core
polysaccharide
c.Inner lipid A (also called endotoxin)
✔ responsible for producing fever and
shock in patients infected with
gram (-) bacteria
 CELL ENVELOPE STRUCTURE

Cellwall
Outer membrane – only found in
gram-negative bacteria
Functions as:
▪ cell’s initial barrier to the environment
▪ Acts a sieve through protein-lined
channels called porins
▪ Provides attachment sites that enhance
attachment to host cells
 CELL ENVELOPE STRUCTURE
Cell wall
Outer membrane – only found in
gram-negative bacteria
▪ Bi-layered structure composed of
lipopolysaccharide (LPS) that gives
the surface of gram-negative bacteria a
net negative charge
▪ plays a significant role in the ability
of certain bacteria to cause disease
▪ Provides attachment sites that
enhance attachment to host cells
 CELL ENVELOPE STRUCTURE
Cell wall
Periplasmic space – found only in
gram (-) bacteria
▪bounded by the internal surface of the
outer membrane and the external surface
of the cellular membrane
▪ Gel-like matrix containing
nutrient-binding proteins and
degradative and detoxifying enzymes
 CELL ENVELOPE STRUCTURE
Cellwall
Periplasmic space – found only in
gram (-) bacteria
•Contains several enzymes that degrade macromolecules and detoxify
environmental solutes, including antibiotics, that enter through the outer
membrane
• virulence factors such as collagenases, hyaluronidases and
proteases are compartmentalized in the periplasmic space
 CELL ENVELOPE STRUCTURE

Porin – protein structures that are scattered throughout the

lipopolysaccharide macromolecules
▪ water-filled structures that controls the passage of nutrients and other
solutes, including antibiotics, through the outer membrane
▪ In addition to porins, other proteins, murein lipoproteins facilitate
attachment of the outer membrane to the next deeper layer in the cell
envelope, the cell wall.
Differences Between Gram-Positive and Gram-Negative Bacteria

Gram-Positive Bacteria Gram-Negative Bacteria

Cell envelope has two layers: Cell envelope has three layers:
.Cytoplasmic membrane 1.Cytoplasmic membrane
.Outer peptidoglycan layer 2.Inner peptidoglycan layer (0.01 nm)
(0.02-0.06 nm) 3.Outer membrane with LPS

Low lipid content High lipid content

No endotoxin Endotoxin (LPS) – Lipid A moiety
No periplasmic space Periplasmic space has detoxifying
enzymes
No porin channel Porin channel is present
Vulnerable to lysozyme and penicillin Resistant to lysozyme and penicillin
since LPS blocks their passage
Cell Envelope Structures
▪ Cell Wall
▪ Acid-fast cell wall
✔ Certain genera, such as
Mycobacterium and Nocardia
✔ Gram-positive cell wall structure
but also contain a waxy layer of
glycolipids and fatty acids, called
mycolic acid, bound to the
exterior of the cell wall
✔ More than 60% of cell wall is
lipid, with mycolic acid as the
major lipid component
 CELL ENVELOPE STRUCTURE
Cell wall (Acid fast organism)
▪ Mycolic acid
✔ is strong “hydrophobic” molecule that
forms a lipid shell around the organism
and affects its permeability
✔ Make bacteria difficult to stain with
gram stain
✔ Best stained with an acid-fast stain
 CELL ENVELOPE STRUCTURE

Absence of cell wall

□ Prokaryotes that belong to the
Mycoplasma and Ureaplasma
genera lack a cell wall
□ Contains sterols in their cell
membrane
□ Seen in various shape
(pleomorphic) microscopically
due to lack of rigidity
 CELL ENVELOPE STRUCTURES
Absence of cell wall
▪ Gram-positive and gram-negative
bacteria may lose their cell walls
and grow as L-forms in media
supplemented with serum or sugar
▪ Serum or sugar prevents osmotic
rupture of the cell membrane
 CELL ENVELOPE STRUCTURES
Cellular appendages
▪ can play a role in causing infections and in laboratory identification
a. Surface polymers
✔ Capsule
✔ slime layers
✔ glycocalyx
b. Cell appendages
✔ pili
✔ fimbriae
✔ flagella
 CELL ENVELOPE STRUCTURES
Surface Polymers
Capsule / glycocalyx /slime layer
▪ Extracellular polymer synthesized by the
enzymes located at the surface of the
bacterial cell
✔ production depends on the
environment and growth conditions
surrounding the bacterial cell
✔ immediately exterior to the murein
layer of the gram (+) and outer
membrane of gram (-)
 CELL ENVELOPE STRUCTURES
Surface polymers
Capsule
✔when the polymer forms a condensed
well-defined layer closely surrounding
the cell
✔during ID by serologic typing, capsules
must be removed to detect the somatic
(cell wall) antigens present underneath
them
✔removal is accomplished by boiling
a suspension of the
microorganism
 CELL ENVELOPE STRUCTURES
Surface Polymers
Capsule acts as virulence factors:
✓ protect bacteria from attack by cells of
the human defense system
✓ facilitates and maintains bacterial
colonization of biologic surfaces (ex.
teeth) and surfaces of inanimate
objects such as implanted medical
devices (ex. prosthetic heart valves
through the formation of biofilms)
✓ anti-phagocytic function
✓ Capsules enable the bacteria to
adhere to their substrate
 CELL ENVELOPE STRUCTURE
Surface Polymers
Capsule
▪ Salmonella Typhi must have its
capsular (Vi) antigen removed to
observe agglutination with
Salmonella somatic (O) antisera
▪ Does not ordinarily stain with use
of common stains (Anthony
staining method)
▪ Appears as clear area (“halo-like”)
between the stained organism and
amorphous background material
 CELL ENVELOPE STRUCTURE

Surface Polymers
Slime Layer
▪ Similar to capsule but more diffuse
layers surrounding the bacterial cell
▪ Made of polysaccharides
▪ Serve either to inhibit phagocytosis
or, in some cases, aid in adherence
to host tissue or synthetic implants
▪ Example – Sarcina lutea
 CELL ENVELOPE STRUCTURE

Surface Polymers
Glycocalyx
✔when it forms a loose meshwork of fibrils extending outward from the
cell
 CELL ENVELOPE STRUCTURE
Cellular appendages
Pili (singular: pilus)
▪ Also known as conjugation pili
▪ Non-motile, long, hollow protein
tubes that connect to bacterial cells
and mediate DNA exchange
▪ most often seen on gram-negative
bacteria
▪ much thinner than flagella
▪ not associated with motility
▪ primary function is to mediate
adherence of bacterial cell to host
cells
 CELL ENVELOPE STRUCTURE
Cellular appendages 2
TYPES OF PILI
i.common pili
enables bacteria to adhere or attach to host surfaces, often as the first
step in establishing infection
ii. sex pili
enables transfer of genetic material from one bacterial cell to another
conjugation
 CELL ENVELOPE STRUCTURE
Cellular appendages
Fimbriae
▪ Singular fimbria
▪ Incorrectly called attachment pili
▪ Non-flagellar, sticky,
proteinaceous, hair-like appendages
that adhere some bacterial cells to one
another and to environmental surfaces
▪ Essential in biofilm formation
 CELL ENVELOPE STRUCTURE
Cellular appendages
Flagellum
▪ Organ of locomotion
▪ complex, threadlike, protein
appendages
▪ motility plays an important role in
survival and ability of some
bacteria to cause disease
▪ bacterial flagella consist of three,
four, or more threads of protein
(called flagellin) twisted like a rope
▪ Originate from cell membrane
 CELL ENVELOPE STRUCTURE
Cellular Appendages
Flagellum
▪ Special staining techniques for
flagella
✔ Gray’s method
✔ Leifson’s carbol fuchsin method
✔ Loeffler’s method
✔ Casares-Gil’s method
✔ Bailey’s method
 CELL ENVELOPE STRUCTURE
Cellular Appendages
Flagellar arrangement
Messea’s Classification
1. monotrichous – one flagella at one end
(polar)
2. lophotrichous – tuft of flagella at one or
both ends
3. amphitrichous – single flagella at each
end
4. peritrichous – flagella cover those entire
surface (perimeter)
5. Atrichous – no flagellum
 CELL ENVELOPE STRUCTURE
Cellular Appendages
Flagellum
▪ Proteus vulgaris – peritrichous
▪ Escherichia coli – peritrichous
▪ Vibrio cholerae – monotrichous
▪ Helicobacter pylori – lophotrichous
▪ Pseudomonas aeruginosa
– monotrichous
▪ Rhodospirillum rubrum
- amphitrichous
BACTERIAL CELL MORPHOLOGY
Many characteristics of bacteria are examined to provide data
for identification and classification. These characteristics
include:
1. Cell morphology (shape)
2. Staining reactions
3. Motility
4. Growth requirements
a) Physical requirements
b) Nutritional requirements
5. Colony morphology
6. Biochemical and metabolic activities
7. Specific enzymes that the organism produces
8. Pathogenicity
9. Genetic composition
 CELL MORPHOLOGY

easily observed in light microscope

(a) size
(b) shape
(c) cell arrangement
 CELL MORPHOLOGY
Size of bacteria
□Metric system is used to express the sizes of microorganisms
▪ 1 micrometer (μm) = 0.000001 = 10-6 mm
▪ 1 nanometer (nm) = 10-9 mm
▪ 1 Angstrom = 10-20 mm
□Most clinically relevant bacterial species range in the following sizes
▪ 0.25 to 1 μm in width
▪ 1 to 3 μm in length
□Average size varies from 0.4 – 2.0 μm
□Average coccus is 1 μm in diameter while average bacillus is 1 x 3 μm
 CELL MORPHOLOGY
Shape and Arrangement
□Based on shapes, bacteria may be grouped into three categories:
▪ Cocci – spherical
▪ Bacilli – rod-shaped
▪ Spiral – spirillum and spirochete
□Some would argue that a fourth category is necessary to classify some
bacteria appearing in different shapes (and sizes) due to the absence of a
cell wall or when grown into a pure culture
▪ Pleomorphic
 CELL MORPHOLOGY

▪ BASIC SHAPES OF BACTERIA

I COCCI – round, spherical or coccoid bacterium
- maybe oval, rounded, lancet-shaped or bean shaped
- arrangement depend on plane division
II BACILLI – often referred to as rods maybe
(a) short (coccobacilli) or long (filamentous)
(b) thick or thin
(c) pointed or with curved - blunt ends
(d) fusiform bacilli – tapered, pointed ends
 CELL MORPHOLOGY

▪ BASIC SHAPES OF BACTERIA

III CURVED AND SPIRAL FORMS
- often referred to as spirochetes
- possesses curved, spiral or cork- screw bodies

IV PLEOMORPHIC ORGANISM
- appears in different shape because they lack cell wall
 CELL MORPHOLOGY

▪ MORPHOLOGICAL ARRANGEMENT
following binary fission, the daughter cells may separate completely
from each other or may remain connected, forming various
morphological arrangements

depends on the particular species and the manner in which the cell
divides
 MORPHOLOGICAL ARRANGEMENT OF COCCI
1. Singly

2. in pairs (diplococci) – division in one plane

• lancet-shaped
(Streptococcus
pneumoniae)
• bean-shaped (Neisseria
gonorrhoeae)
 MORPHOLOGICAL ARRANGEMENT OF COCCI

Streptococcus pneumoniae Neisseria gonorrhoeae

(lancet shaped) (bean-shaped)
MORPHOLOGICAL ARRANGEMENT OF COCCI

3. in chains
(Streptococci)
•bead-like rows of cells
•division in one plane
4. in clusters
(Staphylococcus and
Micrococcus)
•grapelike clusters
•Division in random planes
MORPHOLOGICAL ARRANGEMENT OF COCCI
MORPHOLOGICAL ARRANGEMENT OF COCCI

5. in tetrads
(Peptococcus)
• packets of four
•division in two planes

6. in cube of eight – cuboidal

(sarcina)
•packets of eight
•division in three planes
Sarcina
 MORPHOLOGICAL ARRANGEMENT OF BACILLI

1. singly

2. in pairs (diplobacilli)

3.in chains
(streptobacilli)

4. coccobacilli
• very short rods that
resembles elongated
cocci
MORPHOLOGICAL ARRANGEMENT OF BACILLI
MORPHOLOGICAL ARRANGEMENT OF BACILLI

5. in long filaments
6. branched palisade arrangement (slipping)
• lined side by side like matchsticks
7. chinese character (snapping)
• organism tend to bend at the point of division Ex.
Corynebacterium diphtheriae
8. Curve forms
• single, comma- shaped/curve bacilli (Vibrio)
 Long filamentous Long filamentous bacilli
fusiform bacilli Legionella pneumophila
Fusobacterium nucleatum
MORPHOLOGICAL ARRANGEMENT OF BACILLI
 MORPHOLOGY OF SPIRAL FORMS

▪ Different species vary in their:

(a) size
(b) length
(c) rigidity
(d) number and amplitude of their coils
 MORPHOLOGY OF SPIRAL FORMS
A. SPIRILUM – rigid spirals
- actual spirals, like corkscrew with terminal
appendages
- spiral whose long axis is rigid in motion
B. SPIROCHETES – able to flex
and wiggle their bodies while moving
about
i. Treponema (tightly coiled)
ii. Leptospira
iii. Borrelia (less tightly coiled)
Spirochete – Treponema pallidum Spirochete – Treponema pallidum
Fontana Tribondeau Silver Stain Gram stain