Transcript 2. Cell Wall - Belle Vernon Area School District

Techniques & Tools
I. Types of Microscopes
1. Light Microscopes
A.
Simple –
B.
B. Complex –
C.
Visual Light.
 3 objectives, low, high, oil
immersion.
 Magnification –
 Resolving Power
Light Microscopes
Review
 Why
is a light microscope called a light
microscope?
 Name
the two lenses in a compound
microscope.
2. U.V.



light for illumination
Magnification Resolving
.
.
3. Fluorescent

light with specimens
that’s been stained with



Magnification Resolving -
4. Dark-field


of visible
light, light is refracted
by specimen as light
enters the objective.
Mag
Resolving -
.
.
5. Phase Contrast
Microscope




Observe
matter, unstained.
Detects changes
(phases) in visible
light as it
through a specimen.
Mag Resolving -
6. Transmission Electron
Microscope (TEM)




Beam of
Thin specimen that’s been
stained with electrons
opaques heavy metal.
Travels
specimen.
Viewed on
screen.

Mag -

Resolving -
7. Scanning Electron
Microscope (SEM)
 Beam of
.

surface & produces
a
image of
specimen.

Mag -

Resolving
Scanning Tunneling Electron
Micrograph
Review
 What
is the difference between
magnification and resolution?
 Which
microscope gives the greatest
magnification?
 What
is the magnification limit most light
microscopes?
II. Techniques for Microscopic
Staining
1. Living Microscopic
Organisms.
A. Some Bacteria are
mobile, others are not
but appear to move.
1.
movement due to
air or water
molecules hitting
the
microorganism.
B. Non Staining
Techniques - hard to see.
Ex.
2. Stained Microorganisms
Colored by a chemical dye to make visible.
General Techniques for all staining.
1. Smear - slide out the drop with the bacteria in it.
2. Stained - reveal size, shape, arrangement & presence of
some internal structures.
Types of Staining
1. Simple
A. Smear.
B. Single stain is used (Sudan Black).
2. Differential Staining
A. Gram
1.
2.
3.
4. .
5. .
- decolorizes
after alcohol wash, need a counter
stain (red, safranin).
stain.
- retains blue
B. Acid Fast - Staining
Used for the genus
.
Retain carbolfuchsin (red)when
washed with acidic alcohol.
Counter with
.
Gram Staining
Review
 What
 Why
is a differential stain?
is a Gram stain so useful?
 What
structure of the cell determines its
Gram staining properties?
III. Bacterial Morphology
Shapes are recognized, but
others exist.
1.
- round
A. pairs B. chains C. cubes • Depending when they divide
and then adhere to each other.
2. - rod or cylinder shape.
3.
Spiral Forms twisted rods or cylinders
A.
- actual spirals
(corkscrew); rigid.
B.
- flexible.
Acid Fast Stain
Carbol Fuchsin - 5minutes
 Wash with tap water.
 Acid – 2 minutes.
 Wash with tap water.
 Methylene Blue – 2 minutes.
 Wash with tap water.
 Blot dry.
 Acid Fast bacteria do no decolorize with
acid alcohol. They appear red against a
blue background.

4.
- Extensions
on their surface which gives
them a star appearance.
With age bacteria morphology
may change - swell or show rudimentary
branching.
IV. Reproduction
(
reproduction) - one cell
divides into two identical
daughter cells.
Each cells lives
on their own after replication
even though they may not be
Prokaryotes
Surface Layers - Capsule
Review
 How
do bacteria mainly reproduce? What
is the advantage of that type of
reproduction? What is the disadvantage
of that type of reproduction?
2. Cell Wall
•Function is to hold the cell together.
Composed of sub-units found nowhere else in nature.
Produces symptoms of diseases.
Site of action of some of the most effective antibiotics.
- Common structural component made up of
sugar & amino acids that makes the cell wall rigid.
Components, two different sugars.
A.
B.
Differences in
staining properties.
determines the gram-
– effective against cell walls of bacteria.
V. Bacterial Structure
A.
- Collectively known as the
1.
- material secreted by bacteria that
adheres to the exterior of the bacterial cell.
Functions
Antigenic
Possible
Waste products
A.
- organized, thickened material around
each cell or pairs of cells.
Ex. - Streptococcus pneumoniae - can not survive in a
host unless it can synthesize a capsule, because the
host will quickly destroys it.
B.
- Unorganized loosely attached
polysaccharide.
C.
- oral bacteria that secrete glycocalyx.
Review
 What
is the function of the cell wall?
 What
are the affects of penicillin on
bacterial cell walls?
 What
are the cell walls made of? Where is
the only place on earth that this
component is found at?
A. Gram Positive
(
)
1.Numerous layers of
peptidoglycan (up
to
).
2.
associated in the
peptidoglycan
layer.
B. Gram Negative (
)
1. Outer layer consists
of a
that consists of several
other molecules.
2. Lipopolysaccharides
(LPS),
,&
Porin proteins.
3. Cell Membrane or Plasma membrane.
Located inside the cell wall.
Composition is 60% protein & 40% Lipids.
Functions
1. Osmotic regulator.
2. Enzymes necessary for the synthesis & transport of
peptidoglycan, teichoic acid & other membrane
components.
3. Secretes extracellular hydrolytic enzymes.
4. Ensures segregation of nuclear material during cell
division.
5. Transport of electrons & protons that are released
during aerobic oxidation & turns it into chemical
energy that can be used by the cell.
6. Barrier to the entry of most molecules into the cell
(nutrients entering & wastes leaving).
A. Semi-permeable allows some things to pass, but not others.
1. Diffusion - do not use energy.
a. Passive or Simple Diffusion -molecules flow freely in
& out of the cell (Small).
b. Facilitated Diffusion - Use membrane transport
proteins (Permeases) for molecules to move in & out of
the cell (Large).
2. Active transport - uses energy to cross the membrane.
Several proteins are required for this.
Surface binding proteins.
Membrane transport proteins.
B.
-structures
that extend from the
surface.
1.
- long
slender , protein used for
locomotion.
Bacteria can move
lengths /minute (6ft
man running 82 mph).
A. Composed of three
parts.
1.
2.
3.
B. Types of flagella
arrangements.
C.
movement toward food
substances or away from
harmful substances.
Toward- straight line Away - tumble away -
Due to the
of
the flagella movement (
)
Possible proteins on the cytoplasm
D.
- responds
to various amount of
light.
E.
- responds
to various amount of
oxygen.
F.
- reacts to
the earth’s magnetic
field due to a row of
magnetic particles.
Line up north-south
direction.
Chemotaxsis
2.
- shorter & thinner than flagella.
Functions
1. Movement of
reproduction.
2.
during
to surfaces.
sexual
C. Cytoplasm
1.Cytoplasm water, other 20% Nucleic
Acids, Proteins, Carbohydrates, Lipids.
Primary site of synthetic synthesis of proteins
2.
- singular circular double
strand of DNA that is supercoiled.
that the Bacteria.
, but histonelike structures.
No
or useless DNA.
3.
- Small circular pieces of double
stranded DNA that contain genetic information to
resistance of particular antibiotics.
4.
- Site of protein synthesis
Composed of 60% RNA & 40% Protein.
5.
6.
- Not integral parts of the cell
structure
A.
- reserve source of
phosphate & energy (Volutin).
B. Granulose C. Others - Sulfur, lipids, glycose.
- special membranes
systems found in certain photosynthetic bacteria &
cyanobacteria that contain pigment concerned with
photosynthesis.
D. Special Structures
1.
Minute highly durable body
formed with in the cell &
capable of development into
a new vegetative organism.
adverse conditions
Outer layers
Cortex layers
to
Lack many enzymes; but may
have some in small amounts.
amounts of
,high calcium, protein &
Polysaccharide antigens.
Endospore Formation
Capsule & Endospore Staining
Directions

CAPSULE

Crystal Violet 2-5 minutes

Drain extra crystal violet
in sink.


Wash with copper sulfate
for 30 seconds.
Dry.

Endospore
 Stain with toluidine
blue for 5-15 minutes.

Rinse with tap water.

Dry.
VI. Preparation of a Pure Culture
Culture is a medium in which microorganisms can grow on.
Pure Cultural Techniques
2 Techniques are commonly used.
1.
2.
-General preparation.
.
Inoculating a dilution of the mixed cultures in to a
melted agar.
Poured onto a sterile petri dish.
Bacteria grow sedately (isolated) in a solid
medium. Help ensure pure culture.
VII. Types of Culture Media
pH and temperature must be controlled in all media
1.
2.
– Common
Boiling ground meat with water and filtering off the solid
material-leaves a clear liquid infusion.
Make solid by adding agar to it.
Artificial or Complex Medium - beef extracts, yeast, blood
Synthetic or Defined Medium - can write the chemical
formula for.
broth.
- Various sugars are added to the nutrient
3. Selective & Differential Media
A.
- Chemicals are added to the medium
to inhibit the growth of some bacteria while letting others
grow.
B.
- Acid indicator is added so that many
colonies that formed acid will turn colors.
4.
(Liquid) - Help the growth of certain bacteria
and not others.
VII. Oxygen Requirements
1.
2.
3.
4.
grow.
- will not grow in the
presence of free oxygen,
may even be killed.
oxygen.
-prefers the presence of low
lives in the presence of both.
a.
b.
5.
- require free oxygen to
- produce lactic acids.
- carbon dioxide.
- will grow in the
presence of oxygen but do not posse an
oxidative metabolism.
-
VIII. Sterilization Methods
No living organisms are in the media
when inoculated.
1. Autoclave-steam under pressure
15lbs/in2.
2. Filter.