Chapter
3
Microscopy and Cell
Structure
Overview
Microscopes are the basic tools for
the study of microscopic form and structure. This chapter introduces
the various kinds of
microscopes and staining techniques used by microbiologists to study both
prokaryotic and eukaryotic microorganisms. The
prokaryotic cell is presented with special emphasis on
the cell wall,
external structures, cell membrane and internal components of the cell. The
eukaryotic cell
is also presented with
a focus on the components that are involved in microbial infection and
resistance.
The two kinds of cells
are compared and contrasted.
Learning
Objectives
After studying
the material in this chapter,
you should be able to:
Microscopy
1.
List and describe
the kinds of microscopes used in microbiology.
2.
Define and
contrast:
• Magnification
• Resolution
3.
Describe the preparation of a smear used in staining procedures.
4. List the two kinds of differential staining
procedures; give specific examples and uses of each type.
5. List the three kinds of special staining
procedures; give specific examples and uses of each type.
6.
Describe the Gram
stain procedure and explain the function of each step.
7.
Interpret a Gram
stain.
Prokaryotic Cells
1.
Describe the different morphological forms and arrangements of
prokaryotic cells and give specific
examples of each.
2. List the characteristics of prokaryotic
cells.
3.
Describe the structure of a prokaryotic cell and give the specific
function of each of the major
components.
4.
Describe the fluid mosaic model of the cytoplasmic membrane.
5.
Describe the major roles of the cytoplasmic membrane.
6. Describe the role of the cytoplasmic
membrane in energy transformation.
7. Describe the processes by which substances
are transported across cell membranes by:
• Passive transport mechanisms
• Active transport mechanisms
8. Define:
• Simple diffusion
• Osmosis
• Osmotic pressure
9.
Describe what will occur when a bacterial cell with a cell wall is place
into a:
• Hypertonic solution (higher concentration of
solute outside of the cell than within of the cell)
• Hypotonic solution (higher concentration of
solute inside of the cell than outside of the cell)
10. Define:
• Transport protein
• Facilitated diffusion
• Active transport
• Proton motive force
• ABC transport systems
• Group translocation
11. Describe the
structure of the bacterial cell wall. Differentiate between Gram-negative and
Gram-
positive cell walls.
12. Describe a typical
bacterial chromosome.
13. Describe a plasmid
and explain what it does.
14. Describe a
bacterial ribosome.
15. Give the function
of endospores (bacterial spores) and name the two genera in which endospores
are
generally found.
Key Concepts
1.
Microscopes are the basic tools that make it possible to observe the
microscopic world.
2. The
functions of microscopes are to enlarge (magnify) and to make visible as
separate objects
(resolve) two objects
that are close together.
3. The major differences between the different
types of microscopes are the types of lenses and how the
specimen is
illuminated.
4.
Staining techniques make it possible to study more clearly the structure
of both prokaryotic and
eukaryotic cells.
5. Prokaryotic cells are designed to carry on
the basic functions of life and they have the appropriate
structures to do so.
6. The structure of the bacterial cell wall is
an important determinant in many of the properties of
bacteria including
differential staining characteristics, susceptibility to antibiotics such as
penicillin
and susceptibility to
changes in extracellular osmotic concentrations.
7. The cytoplasmic membrane is the major
determinant as to what enters and leaves a cell.
8. Prokaryotic cells have developed cellular
modifications such as the glycocalyx, sheaths, flagella and
pili that enable them
to compete successfully.
9. The
bacterial chromosome consists of a single, double-stranded DNA molecule that
resides in the
cytoplasm of the cell
rather than in a nucleus.
10. Extrachromosomal DNA called plasmids are
sometimes found in prokaryotic cells and may be
advantageous, but not
essential, to the cell.
11. The structure of ribosomes differs in
prokaryotic and eukaryotic cells making the prokaryotic
ribosomes useful
targets for antimicrobial agents.
12. Endospores are dormant forms within the life
cycle of some bacterial species that enable the microbe
to withstand difficult
environmental conditions.
Summary Outline
I.
Microscopic techniques: the instruments
A. Light Microscopes
1. The bright-field
microscope: Visible light passes
through the specimen
2. Phase
contrast microscope: Amplifies differences in refraction
3. Dark-field
microscope: Directs light toward a
specimen at an angle
4. The fluorescent
microscope: used to observe cells
that have been stained with
fluorescent dyes
5. The confocal
scanning laser microscope: Used
to construct a three-dimensional image
of a thick structure
and to provide detailed sectional views of the interior of an intact cell
B. Electron
Microscopes
1. Use electromagnetic lenses, electrons, and phosphorus screens to
produce a magnified
image
a) Transmission
electron microscopes (TEM) transmit electrons through a
specimen that has been
prepared by thin-sectioning, freeze-fracturing, or freeze-etching
b) Scanning
electron microscopes scan a beam of electrons back and forth over the
surface of a specimen,
producing a three-dimensional effect
2.
Scanning probe microscopes: Maps the bumps and valleys of a surface on an atomic
scale
C. Dyes
and staining
1. Differential stains
a) The Gram
Stain
(1)
Gram-positive bacteria stain purple
(2)
Gram-negative bacteria stain pink
b) The acid-fast
stain: Stains organisms such as mycobacteria, which do not take
up stains readily;
acid-fast organisms stain pink and all other organisms stain
blue
2. Special stains to observe cell structures
a) Capsule
stain colors the background,
allowing the capsule to stand out as a halo
around an organism
b) Spore
stain stains endospores
c) Flagella
stain stains flagella
3. Fluorescent
dyes and tags: Some fluorescent dyes
bind compounds that characterize
all cells, others bind
to compounds specific to only certain cell types
D. Morphology of prokaryotic cells
1. Shapes
a) Cocci
b) Rods
c) Coccobacilli
d) Vibrios
e) Spirilla
f) Spirochetes
g) Pleomorphic
bacteria have variable shapes
2. Groupings: Cells adhering to one another
following division form a characteristic
arrangement that
depends on the plane in which the bacteria divide
3. Multicellular associations
a) Associations containing multiple cells,
such as myxobacteria
b) Biofilms
often alter their activities when a critical number of cells are present
II. The Structure of the Prokaryotic Cell
A. The cytoplasmic
membrane
1. Structure and chemistry of the
cytoplasmic membrane
a) Phospholipid
bilayer embedded with a variety of
different proteins
b) Differential
barrier between the cell and the
surrounding environment
c) Membrane proteins function in transport
or provide a mechanism by which
cells can sense and adjust to their surroundings
2. Permeability of the cytoplasmic membrane
a) Selectively
permeable
b) Inflow of water into the cell exerts more
osmotic pressure on the cytoplasmic
membrane than it can
generally withstand
3. The role of the cytoplasmic membrane is
involved in energy generation: Electron
transport chain within
the membrane expels protons, generating an electrochemical
gradient, which
contains a form of energy called proton motive force
B. Directed movement of materials across the
cytoplasmic membrane
1. Transport
systems
a) Facilitated
diffusion: Moves impermeable
compounds from one side of the
membrane to the .other
by exploiting the concentration gradient
b) Active transport mechanisms use energy to accumulate compounds against a
concentration gradient
c) Members of the major facilitator superfamily use proton motive force for energy
(1)
Symporters
(2) Antiporters
(3) Uniporters
d) ABC
transport systems require ATP for
energy
e) Group
translocation chemically modifies a
molecule during its passage through
the cytoplasmic
membrane
2. Secretion:
The general secretory pathway is the primary mechanism used to secrete
proteins
C. Cell Wall
1. Peptidoglycan
a) Found only in bacteria and provides
rigidity to the cell wall
b) Composed of peptidoglycan which contains
alternating subunits of N-
acetylmuramic acid (NAM) and N-acetylglucosamine (NAG)
interconnected
via the tetrapeptide
chains on NAM
2. The Gram-positive
cell wall
a) Relatively thick layer of peptidoglycan
b) Teichoic
acids and lipoteichoic acids stick
out of the peptidoglycan molecule
3. The Gram-negative cell wall
a) Thin
layer of peptidoglycan sandwiched
between the cytoplasmic membrane
and an outer membrane
b) Periplasm
contains a variety of proteins
c) The outer membrane contains
lipopolysaccharides. The Lipid A portion of the
lipopolysaccharide
molecule is toxic, which is why LPS is called endotoxin
d) Porins
form small channels that permit small molecules to pass through the outei
membrane
4. Antibacterial compounds that target
peptidoglycan
a) Penicillin binds to proteins involved in
cell wall synthesis
b) Lysozyme breaks the bond that links
alternating NAG and NAM molecules
5. Characteristics of bacteria that lack a
cell wall
a) Because mycoplasmas do not have a cell wall, they are extremely variable in
shape and are not
effected by lysozyme or penicillin
D. Surface layers external to the cell wall
1. Glycocalyx:
Enable bacteria to adhere to surfaces; some capsules allow disease-
causing microorganisms
to thwart the innate defense system
a) Capsule:
A distinct and gelatinous layer made of polysaccharide
b) Slime
layer: Diffase and irregular layer
of polysaccharide
2. Sheath:
A tube that holds a linear chain of cells; it is thought to protect the
enclosed
organisms from
disruption
E. Filamentous protein appendages
1. Flagella
a) Long protein structures responsible for
most types of bacterial motility
b) Chemotaxis
is the directed movement toward an attractant or away from a
repellant
2. Pili
a) Many types of pili (fimbriae) enable
attachment of cells to specific surfaces
b) Some pili play a role in specific types of
motility
c) Sex pili
are involved in conjugation, which enables DNA to be transferred from
one cell to another
F. Internal structures
1. Chromosome
a) The chromosome of prokaryotes resides in the nucleoid
b) The typical chromosome is a single,
double-stranded DNA molecule that
contains all the
genetic information required by a cell
2. Plasmids
a) Plasmids are circular, double-stranded
DNA molecules that typically encode
genetic information
that may be advantageous, but not required by the cell
b) Populations of cells can gain and lose
plasmids, depending on the relative
advantages
3. Ribosomes
a) Ribosomes facilitate the joining of amino
acids
b) The 70S bacterial ribosome is
composed of a 50S and a 30S subunit
4. Storage
granules: Dense accumulations of
high molecular weight polymers, which are synthesized from a nutrient that a
cell has in relative excess
5. Gas
vesicles are gas-permeable,
water-impermeable rigid structures that provide
buoyancy to aquatic
cells
6. Endospores
a) Dormant stage produced by members of the genera Bacillus and Clostridium,
they can germinate to
become a vegetative cell
b) Resistant
to conditions such as heat, desiccation, toxic chemicals, and UV
irradiation
c) Sporulation
is an eight-hour process initiated when cells are grown under
nutrient-limiting
conditions
d) Germination
is the process by which an endospore leaves its dormant state
Terms You Should
Know
Microscopy and Cell
Morphology
Bacillus
Binary
fission
Capsule
Coccobacillus
Coccus
Compound microscope
Condenser lens
Contrast
Decolorizing agent
Diplococcus
Electron photomicrograph
Endospore
Fluorescent dyes
Immunofluorescence
Mordant
Objective
lens
Ocular
lens
Pleomorphic
Refraction
Resolution
Spirillum
Spirochete
Prokaryotic Cell
Active
transport
Antiporter
Capsule
Cell wall
Chemotaxis
Chromosome
Cytoplasm
Cytoplasmic membrane
Electrochemical gradient
Endospore
Endotoxin
Facilitated diffusion
Flagella
Fluid mosaic model
Gas vesicles
Germination
Glycocalyx
Group translocation
Lipid A
Lipopolysaccharide layer (LPS)
N-acetylglucosamine (NAG)
N-acetylmuramic acid
(NAM)
Nucleoid
O
polysaccharide
Osmosis
Osmotic pressure
Passive transport
Peptidoglycan
Pili
(fimbriae)
Plasmid
Forms
Proton
motive force
Ribosome
Selectively permeable
Sex pili
Sheath
Simple
diffusion
Slime
layer
Sporulation
Storage
granules
Symporter
Teichoic acids
Transport proteins (permeases or carriers)
Uniporter