1. MICROBIAL
CELL
BIOLOGY
Biology 120 Lecture 3
Reference: Chapter 3
and 4 TORTORA
Tuesday, July 3, 2012
2. GETTING TO KNOW
YOUR BACTERIA
• Unit of measure
(micrometers or
nanometers)
• Microscopy = most
essential tool in
microbial cell biology
Tuesday, July 3, 2012
3. ESSENTIALS IN
MICROSCOPY
• Simple versus Compound
• number of lenses
• Brightfield versus Darkfield
• background
• Light versus Electron Microscopy
• light/ beam of electrons
• SEM versus TEM
• 3D surface/ 2D internal structures
Tuesday, July 3, 2012
10. PREPARATION OF SPECIMENS
FOR MICROSCOPY
• FIXATION
• Process by which the internal and external structures of
cells and microorganisms are preserved and fixed in
position
• Inactivates enzymes that might disrupt cell morphology
• Toughens cell structures to prevent changes during
staining and observation
• Usually microbes are killed when fixed
Tuesday, July 3, 2012
11. TYPES OF FIXATIONS
• preserves overall
morphology but not
structures within cells
• smear ready for staining!!!
• Penetrates
cells and
react with
cellular
components
Tuesday, July 3, 2012
12. STAINING
•STAINS
•salt of + or - ions, one of which is
colored (chromophore)
•+ basic dye (CV, MB, MG, Sf )
•- acidic dye (Ng)
•Types of Staining: SIMPLE,
DIFFERENTIAL, SPECIAL
Tuesday, July 3, 2012
14. STAINING
• SIMPLE • SPECIAL
• e.g. MB, CF, CV, Sf • e.g. negative,
endospore, flagella
• highlight shapes and staining
arrangements
• staining special
• DIFFERENTIAL structures
• e.g. Gram stain and
Acid fast stain
• differentiate and
distinguish one kind of
bacteria from another
Tuesday, July 3, 2012
26. SIZE, SHAPE &
ARRANGEMENT
OF BACTERIAL CELLS
Tuesday, July 3, 2012
27. WHAT ARE THE 3 BASIC
SHAPES OF BACTERIA?
•Measure: 0.2-2.0µm
(diameter) x 2-8µm
(length)
•Basic shapes: coccus,
bacillus, spiral
Tuesday, July 3, 2012
28. COCCI
• usually round, can be oval, elongated or
flattened on one side
• e.g. diplococci (remain in pairs)
• e.g. streptococci (remain in chains)
• e.g. tetrads (remain attached in cube-like
group of 4)
• e.g. sarcinae (remain attached in cube-like
group of 8)
• e.g. staphylococci (divide in multiple planes,
grape-like)
Tuesday, July 3, 2012
31. BACCILI
• divide only across their short axis; mostly
single rods
• e.g. diplobacilli (remain in pairs after
division)
• e.g streptobacilli (occurs in chains)
• e.g. coccobacilli (cocci-like)
Tuesday, July 3, 2012
34. SPIRALS &
CURVED
• have on or more twists
• never staright
• e.g. vibrios (curved rods(
• e.g. spirilla (helical, cork-screw, rigid)
• e.g spirochetes (helical but flexible)
Tuesday, July 3, 2012
37. THE
OTHERS
Stella sp. Haloarcula sp.
Tuesday, July 3, 2012
38. SHAPE = heredity
* Monomorphic, maintain a single shape
** Pleomorphic more than one shape
Tuesday, July 3, 2012
39. ORGANIZATION
IN A TYPICAL
PROKARYOTE
STRUCTURE:
1. Structures external
to the cell wall
2. the cell wall
3. structures internal
to the cell wall
Tuesday, July 3, 2012
40. STRUCTURES
EXTERNAL TO THE
CELL WALL
Tuesday, July 3, 2012
41. GLYCOCALYX or
SUGAR COAT
• secreted on prokaryotic surface
• viscous, sticky, gelatinous polymer
• composed of polysaccharide, polypeptide or
both
• made inside the cell and secreted outside
• CAPSULE: organized and firmly attached to
the cell wall
• SLIMY LAYER: unorganized and loosely
attached to the cell wall
Tuesday, July 3, 2012
42. FUNCTIONS:
• contributory to virulence (degree of
pathogenicity)
• protect pathogen from phagocytosis
• attachment to various surfaces for survival
• prevent cell from dehydration
• viscosity = inhibits movement of nutrients out
of the cell
• EPS (extracellular polysaccharide) = capsules
made up of sugars
Tuesday, July 3, 2012
44. OBSERVATION OF
GLYCOCALYX
•Bacillus anthracis
(anthrax)
•Streptococcus
pneumoniae
(bacterial pneumonia)
•Streptococcus mutans
(dental caries)
Tuesday, July 3, 2012
45. FLAGELLA
• long filamentous appendages that
propel bacteria
• 3 basic parts:
• filament = long outermost region
(flagellin)
• hook = where filament is attached
(various protein)
• basal body = anchors flagellum to
cell wall and plasma membrane
Tuesday, July 3, 2012
46. FLAGELLA
• atrichous = lacks flagellum
• monotrichous = single polar
• amphitrichous = tufts at both ends
• lophotrichous =two or more on
one or both ends
• peritrichous = distributed over the
entire cell
Tuesday, July 3, 2012
52. MOTILITY
• ability of an organism to
MOVE by itself
• RUN-TUMBLE-RUN
routine
• TAXIS: move away or
towards a stimuls
(chemotaxis, phototaxis)
Tuesday, July 3, 2012
54. FLAGELLAR
PROTEINS
• H antigen: useful for
distinguishing among
serovars of bacteria
• e.g. E. coli O157:H7
• NOTE: there are at least
50 fifferent H antigens
for E. coli
Tuesday, July 3, 2012
55. AXIAL
FILAMENTS
• locomotory structure for
spirochetes
• also called “endoflagella”
• bundles of fibrils that
arise at the ends of the
cell beneath an outer
sheath
• spirals around the cell
Tuesday, July 3, 2012
57. FIMBRIAE & PILI
• hair-like appendages
• shorter, straighter and thinner
than flagella
• used for attachment (F) and
transfer of DNA/conjugation (P)
• essential for colonization in
Neisseria (F)
• NOT for motility!
Tuesday, July 3, 2012
60. CELL WALL
• complex, semi-rigid structure
• gives the shape of the cell
• surrounds the plasma membrane and protects interior
from adverse changes in the outside environment
• prevents rupture of bacterial cells
• contributes to ability of some species to cause disease
• site of action of some antibiotics
• ALMOST ALL prokaryotes have cell walls
Tuesday, July 3, 2012
61. CELL WALL:
Composition & Characteristics
• peptidoglycan or
murein
• N-acetylglucosamine
(NAG) and
N-acetylmuramic acid
(NAM)
• Linked by
polypeptides
Tuesday, July 3, 2012
62. CELL WALL:
Composition & Characteristics
Tuesday, July 3, 2012
69. ATYPICAL CELL WALL
• e.g. Mycoplasma
• no walls or have little
wall materials
• plasma membrane have
sterols (help protect
from lysis vice the CW)
Tuesday, July 3, 2012
70. ATYPICAL CELL WALL
• e.g. Archaea
(Halobacterium sp)
• may lack walls or have
unusual walls composed of
polysaccharides and
proteins not peptidoglycan
(pseudomurein)
• PSEUDOMUREIN = lacks
the D-amino acids found in
bacterial CWs
• NOTE: Gram stain not
applicable
Tuesday, July 3, 2012
71. ATYPICAL CELL WALL
• e.g. Mycobacterium and
Nocardia
• high concentrations of
mycolic acids in CWs (60%)
• prevents the uptakes of
dyes
• Note: Gram stain will work
only if mycolic acids
removed
Tuesday, July 3, 2012
76. IMPORTANT STRUCTURES
• GLYCOPROTEINS & GLYCOLIPIDS
• help protect and lubricate the cell
• involved in cell-to-cell interactions
(e.g. pathogen binding in inlfuenza)
Tuesday, July 3, 2012
77. CM FUNCTIONS
• selective barrier
• breakdown of nutrients and
production of energy
• What happens when CM
destroyed?
• cell leakage
Tuesday, July 3, 2012
78. PHOTOSYNTHETIC STRUCTURES IN
THE CM
green sulfur bacteria
purple non-sulfur bacteria
PHOTOSYNTHETIC PIGMENTS IN
MEMBRANE FOLDINGS:
1. Chromatophores
2. Chlorosomes
3. Photosynthetic lamellae
purple sulfur bacteria
Tuesday, July 3, 2012
79. BACTERIA VS ARCHAEA CM
• EUBACTERIA
• Ester linkage
• Weaker linkage
• ARCHAEBACTERIA
• Ether linkage
• Stronger linkage
Tuesday, July 3, 2012
81. THE NUCLEAR AREA
• “NUCLEOID”
• contains the
bacterial
chromosome
• not surrounded by
a nuclear envelope
(membrane)
• do not include
histones
Tuesday, July 3, 2012
82. THE NUCLEAR AREA
• PLASMIDS
• extrachromosomal
genetic element
• replicate independently
• Gene: antibiotic
resistance, tolerance to
toxic metals, toxin
production and
synthesis of enzymes
• can be transferred from
one bacterium to
another via conjugation
Tuesday, July 3, 2012
83. BACTERIAL RIBOSOMES
• ALL PROKARYOTES & EUKARYOTES
HAVE RIBOSOMES!!!
• site of protein synthesis
• composed of two units:
• protein sub-unit
• ribosomal RNA subunit
• NOTE: differ from EUK ribosomes in the
number of proteins and rRNA molecules
they contain and they are less dense
Tuesday, July 3, 2012
84. BACTERIAL RIBOSOMES
• PROK = 70S ribosomes while EUK = 80S
ribosomes
• The 70S = 30S (1 rRNA molecule) + 50S
(2 rRNA molecules)
16S ribosomal DNA = prokaryotes
18S ribosomal DNA = eukaryotes
Tuesday, July 3, 2012
85. BACTERIAL RIBOSOMES
• Antimicrobials:
• streptomycin and
gentamicin = attach
to 30S and interfere
with protein synthesis
• erythromycin and
chloramphenicol =
attach to 50S and
interfere with protein
synthesis
• THUS only prokaryotes
are affected by these
antimicrobials
Tuesday, July 3, 2012
86. INCLUSIONS
• inclusions = reserve deposits used when
supply are deficient
• METACHROMATIC GRANULES
• POLYSACCHARIDE GRANULES
• LIPID INCLUSIONS
• SULFUR GRANULES
• CARBOXYSOMES
• GAS VACUOLES
• MAGNETOSOMES
Tuesday, July 3, 2012
87. METACHROMATIC
GRANULES
• “volutin”
• large inclusions
• stain red with certain blue dyes (e.g. MB)
• inorganic phosphate/polyphosphate
reserves
• used for ATP synthesis
• BACTERIA, ALGAE, FUNGI & PROTOZOA
• Corynebacterium diphtheriae (diagnostic)
Tuesday, July 3, 2012
88. METACHROMATIC
GRANULES
“chinese
characters”
diagnostic for
C. diptheriae
Tuesday, July 3, 2012
89. POLYSACCHARIDE
GRANULES
• consist of glycogen and
starch
• demonstrated when
iodine is applied to cells
• appear reddish brown
(Glycogen)
• appear blue (Starch)
Tuesday, July 3, 2012
90. LIPID INCLUSIONS
• Mycobacterium, Bacillus,
Azotobacter, Spirillum
etc
• e.g. poly-B-
hydroxybutyric acid
(PHBs)
• revealed using Sudan
dyes (fat soluble dye)
Tuesday, July 3, 2012
91. SULFUR GRANULES
• Thiobacillus spp,
Beggiatoa
Beggiatoa sp.
• they derive energy by
oxidizing sulfur and
sulfur-containing
compounds
• deposit sulfur
granules as energy
reserves
Tuesday, July 3, 2012
93. GAS VACUOLES
• hollow cavities in aquatic
prokaryotes
• cyanobacteria, anoxygenic
photosynthetic bacteria and
halobacteria
• maintain buoyancy
Tuesday, July 3, 2012
94. MAGNETOSOMES
• inclusion of iron oxide
• Magnetospirillum
magnetotacticum
• used to move downward
until they reacha suitable
attachment site (act like
magnets)
• can decompose hydrogen
peroxide (to protect cells
from its accumulation)
Tuesday, July 3, 2012
95. ENDOSPORES
• Clostridium, Bacillus (Bacteria)
• Thermoactinomyces vulgaris (Archaea)
• specialized resting cells
• resistant to adverse conditions (extreme
heat, lack of water, exposure to toxic
chemicals and radiation)
• Dipicolinic acid (DPA) with calcium ions
directly involved in spore heat resistance
Tuesday, July 3, 2012
96. SPORULATION/SPOROGENESIS
SPORULATION:
SPORE FORMATION
GERMINATION:
SPORE TO VEGETATIVE
CELL
Tuesday, July 3, 2012
101. EUKARYOTIC
FLAGELLA & CILIA
Tuesday, July 3, 2012
102. For cellular locomotion
• Flagella: projections that are few and long in relation
to the size of the cell (e.g. Euglena)
• Cilia: projections that are numerous and short in
relation to the size of the cell (e.g. Tetrahymena)
• Difference between prokaryotic flagella:
• PROK = rotates
• EUK = moves in a wave-like manner
Tuesday, July 3, 2012
107. CELL WALLS
• EUK have simpler cell walls
• Algae: cellulose
• most Fungi: chitin
• Yeasts: glucan and mannan
• Protozoa: DO NOT HAVE a typical cell
wall = pellicle (flexible outer protein
covering)
Tuesday, July 3, 2012
108. GLYCOCALYX
• strengthens the cell
surface
• helps attach cells
together
• involved in cell to cell
recognition
Tuesday, July 3, 2012
109. EUKARYOTIC
PLASMA MEMBRANE
Tuesday, July 3, 2012
110. PLASMA MEMBRANE
• similar in function and basic structure with
prokaryotes
• differences are the proteins found in the
membranes
• also contain carbohydrates which serves as
attachment sites for bacteria and as receptor sites
for cell-to-cell recognition
• contains sterols (resist lysis due to osmotic
pressure)
Tuesday, July 3, 2012
111. PLASMA MEMBRANE
• NOTE: group
translocation do not
occur in eukaryotic
membranes
• instead ENDOCYTOSIS
(e.g. pinocytosis and
phagocytosis)
Tuesday, July 3, 2012
112. CYTOPLASM
• substance inside
the plasma
membrane and
outside the
nucleus
• cytosol = fluid
portion of the
cytoplasm
Tuesday, July 3, 2012
113. CYTOPLASM
• Major difference:
• EUK have complex internal
structures (microfilaments,
intermediate filaments,
microtubules) which forms
the cytoskeleton (provides
support for cytoplasmic
streaming)
• many enzymes fund in
cytoplasmic fluid of PROK are
sequestered in the organelles
of EUK
Tuesday, July 3, 2012
114. EUKARYOTIC
RIBOSOMES &
ORGANELLES
Tuesday, July 3, 2012
115. RIBOSOMES
• same function as in PROK
• larger and denser than PROK (80S = 60S with 3
molecules of rRNA; and 40S with 1 molecule of rRNA)
Tuesday, July 3, 2012
116. RIBOSOMES
• free ribosomes: unattached, protein
synthesis used inside the cell
• membrane-bound ribosomes: attached
to nuclear membrane and ER, protein
synthesis for insertion in the plasma
membrane or for export from the cell
• polyribosome: located within
mitochondria, synthesis of
mitochondrial proteins (10-20
ribosomes joined together in a string-
like arrangement)
Tuesday, July 3, 2012
117. ORGANELLES
• organelle: structure with specific shapes
and specialized functions; absent in
prokaryotes
• Nucleus, ER, golgi complex, lysosomes,
vacuoles, mitochondria, chloroplasts,
peroxisomes and centrosomes
Tuesday, July 3, 2012
118. ORGANELLES
• Nucleus = houses the chromosome
• ER = transport and storage
• Golgi complex = membrane foration
and protein secretion
• Lysosomes = store digestive enzymes
• Vacuoles = storage and rigidity
Tuesday, July 3, 2012
119. ORGANELLES
• Vacuoles = storage and rigidity
• Mitochondria = site of ATP production
• Chloroplasts = contain chlorophyll and
enzymes for photosynthesis
• Peroxisomes = oxidation of organic
compiunds (e.g. catalase) destroying
hydrogen peroxide)
• Centrosomes = contains centrioles for
mitotic spindle formation
Tuesday, July 3, 2012
122. HOW THEY CAME
ABOUT...
• 3.5-4B years ago = simple organisms
(similar to prokaryotes)
• 2.5B years ago = eukaryotes from
prokaryotes
• Lyn Margulis: The Endosymbiotic Theory
• larger bacterial cells lost their CW and
engulfed smaller bacterial cells
• endosymbiosis = lives within another
Tuesday, July 3, 2012
123. ENDOSYMBIOTIC
THEORY
• ancestral EUK developed a rudimentary nucleus
when the plasma membrane folded around the
chromosome (NUCLEOPLASM)
• Nucleoplasm ingested aerobic bacteria and lived
inside it
• evolved into a symbiotic relationship (host supply
nutrients, while bacteria produce the energy from
supplied nutrients
Tuesday, July 3, 2012
124. ENDOSYMBIOTIC
THEORY
• CHLOROPLASTS =
descendants of
photosynthetic
prokaryotes ingested
by the nucleoplasm
• FLAGELLA & CILIA =
motile spiral bacteria/
spirochetes
Tuesday, July 3, 2012
125. NEXT MEETING:
JOURNAL REPORTING
Tuesday, July 3, 2012
