Homework 2
Please use this document to take notes/draft your answers. Submit your final answers for grading to
the “Homework 2 Final Answers” file to be graded.
All answers must be in your own words. Answers copied directly from the textbook, PowerPoint, or an
online source will not be given credit.
Each question is worth 2 points.
1. What characteristics do all living organisms have? Do you agree that viruses are not living? Why
or why not?
2. How can fimbriae make a bacterial cell more virulent? How can a capsule make a bacterial cell
more virulent?
3. What is a biofilm and where might you find one?
4. How does a Gram Stain work? Describe each of the 4 reagents used and how they act on a
Gram-positive cell vs. a Gram-negative cell.
5. How does Penicillin work? What type of bacteria would it be most effective against?
6. Compare and contrast prokaryotic and eukaryotic cells. Describe at least two ways they are
similar and two ways they differ.
Topic 3 PPT
Cells – Prokaryotes in
Particular
Outline
• Characteristics of life
• Two types of cells
• Size, Shape, and Arrangements of
Prokaryotic Cells
• Parts of a Prokaryotic Cell
– External
– Cell Membrane
– Internal
Characteristics of life?
• Made of cells
– Has DNA for storage and transmission of genetic information
• Metabolism
– Takes in nutrients, uses enzymes, creates energy (ATP), and
excretes waste
• Reproduction
– Asexually or sexually
• Growth and development
• Movement and/or irritability
– respond to internal/external stimuli; self-propulsion of many
organisms
• Cell support, protection, and storage mechanisms
– cell walls, vacuoles, granules and inclusions
Characteristics of Life
4
Cells
• Cell Theory of Life
– “All life is composed of cells and all cells can carry
out the basic life processes (metabolism and
reproduction).”
• Two types of Cells:
• Prokaryotic Cells
– lack nucleus and other membrane-bound structures
(organelles)
– Domains of Archaea and Bacteria are both
prokarytotes
• Eukaryotic Cells
– have a nucleus and organelles
– Domain of Eukarya or eukaryotes
– Includes protozoa/protists, fungi, helminthes, and
algae
Before moving on…
• Brainstorm what you know about
prokaryotes and eukaryotes.
– What is the same?
– What is different?
• Can you diagram it? See the next slide
for an example.
Anything that is unique to eukaryotes would go in the
left circle. Anything unique to prokaryotes in the right
circle. Any structures, processes, or traits they share
would go in the portion that intersects in the middle.
Similarities between Prokaryotic and Eukaryotic Cells
(these would go in the middle portion of your circles)
• DNA
• the genetic material, made up of A, T, C, and G, and arranged in a double helix
• Plasma membrane
• a phospholipid bilayer with proteins that separates the cell from the surrounding
environment and functions as a selective barrier for the import and export of
materials
• Cytoplasm
• the rest of the material of the cell within the plasma membrane, excluding the
nucleoid region or nucleus, that consists of a fluid portion called the cytosol and
the organelles and other particulates suspended in it
• Ribosomes
• the organelles on which protein synthesis takes place
• Reproduction
• Both make exact copies of themselves
• Metabolism
• Both obtain nutrients, make energy, and expel waste
• Ability to maintain internal homeostasis
Differences Between Prokaryotic & Eukaryotic Cells (these
would go in one circle or the other)
• DNA location
– Prokaryotes have a central region called the nuclear region where genetic
material is concentrated, but not bound by a membrane
– Eukaryotes have a membrane bound organelle called the nucleus that contains
the genetic material
• DNA structure
– Prokaryotes have a singular, circular chromosome
– Eukaryotes have multiple, linear, paired chromosomes
• DNA storage (histone proteins)
– Present in Eukaryotic cells due to the massive amount of genetic material
present. Histones help condense it so it fits in the nucleus.
– Not present in Prokaryotes
• Extrachromosomal DNA
– Present in the mitochondria and chloroplasts of Eukaryotic cells.
– Present in plasmids in prokaryotic cells.
More Differences Between
Prokaryotic & Eukaryotic Cells
• Cell Walls
• Found in Prokaryotic cells and made of peptidoglycan. Helps
the cell maintain shape and avoid lysis via osmosis.
• Found in some Eukaryotic cells like plants (cellulose) and fungi
(chitin). Absent in animal cells.
• Locomotion
• Some Prokaryotes are mobile via one or more flagella. The
flagella are thin and rotate like a propeller.
• Some Eukaryotes are mobile via flagella. The flagella are thick
and move side to side. Some eukaryotes have cilia.
• Reproduction
• Prokaryotes use binary fission.
• Eukaryotes use mitosis and meiosis.
How many similarities and
differences did you come up with?
• Did you miss some that are on the list?
– Add them to your diagram.
• Did you come up with some that are not
on the list?
– Nice job!
•
For many students, reading the information alone (in the form of a
textbook or PPT) is not enough for them to learn the information. Try to
organize the information in a way that lets you see the connections (like
the activity you just completed) to help you better understand the
material.
Size of Prokaryotes
•
prokaryotes are the smallest of all organisms
• they typically range 0.5 – 2.0 µm in diameter and 1.0 – 60 µm in
length (Note: 1 µm = 1 millionth of a meter)
•
•
•
In Microbiology there is often
an exception to the general
rule!
Most prokaryotes are very
small, but Epulopiscium
fishelsoni is huge in
comparison. (Compare its
size to the E. coli cell
pictured.)
It is a bacterial symbiont of
sturgeon fish (80 µm in
diameter and 600 µm in
length.
Relative Size of Living Things
Magnification
Why are prokaryotes so small?
SA/V=
6
SA/V=3
SA/V = 1.5
Small size is an advantage!
A larger Surface Area (SA) to Volume (V) ratio = easier diffusion of nutrients
in to any part of the cell and quick diffusion of waste products out of the cell.
Bacterial Shapes
– Coccus – spherical
– Bacillus – rod
– Coccobacillus – very short and plump (too
long to be a cocci, too short to be a bacilli)
– Vibrio – gently curved, comma shaped
– Spirillum – helical, twisted rod, rigid
– Spirochete – spring-like, flexible
16
Common bacterial shapes
17
Pleomorphism
• Variation in cell shape
and size within a
single species
• Bacteria of the genus
Mycoplasma are
noted for their
pleomorphism
– they lack a cell wall to
hold a single shape
18
Bacterial Arrangements
•
•
Arrangements happen when the cell walls of
bacteria remain attached following binary fission
(reproduction).
There is no exchange of cytoplasm or other
materials between the connected cells.
• Cocci:
•
•
•
•
•
•
Singles – no arrangement
Pairs - diplococci
Groups of four - tetrads
Irregular cluster - staphylococci
Chains - streptococci
Cubical packets - sarcina
– Bacilli:
• Singles – no arrangement
• Pairs - diplobacilli
• Chains - streptobacilli
19
What arrangement is this? (1)
• Neisseria (22,578X)
• Notice how the cells
are still attached via
the cell wall (yellow)
and how there is no
sharing of cytoplasm
(blue.)
Streptococcus (9605X)
What arrangement is this? (2)
What arrangement is this? (3)
What arrangement is this? (4)
• Bacilli divide only on the
axis of the narrow ends
• Sometimes they
separate and you have
random arrangement of
single bacilli.
• Sometimes they remain
attached in chains. Bacillus megaterum (10,000X)
• They never divide along
*Rods in chains
the wide sides –
or Streptobacilli
therefore you will never
see a cluster of rods.
Medically Important Bacteria
Structure of a bacterial cell
An Overview of Structure of
bacterial cells
1. External Structures: Capsules, slime layers,
flagella, fimbriae, and pili
2. Cell envelope: cell wall and cell membrane
3. Internal structures: Cytoplasm with
ribosomes, nuclear region, and in some cases
granules and/or vesicles
External Structures
• Appendages
• Those used for motility
– flagella
– axial filaments (periplasmic flagella)
• Those used for attachment or as channels
– fimbriae
– pili
• Glycocalyx – surface coating
• Capsules
• Slime layers
27
Flagella
• How to classify them based on number
and location?
• What are they made of?
• How do they make a bacterial cell
move?
• Can a bacterial cell actually “steer” itself
towards something it wants?
Naming Flagellar Arrangements
Monotrichous –
single flagellum
at one end
Lophotrichous –
small bunches
emerging from
the same site
Amphitrichous –
flagella at both
ends of cell
Peritrichous –
flagella
dispersed over
surface of cell
29
1. One polar flagellum located at one end or pole
Monotrichous
flagella
= monotrichous flagellum
Pseudomonas (3,300X)
Amphitrichous flagella
Spirillum (694X)
3. Two or more flaggela at one or both ends =
Lophotrichous
flagella
Lophotrichous flagella
4. Flagella all over the surface =
Peritrichous
flagella (1)
Peritrichous
flagella
Salmonella (1200X)
Peritrichous flagella (2)
Atrichous
Parts of the Flagella
• 3 parts:
– Filament – long, thin, helical structure composed of
protein flagellin
– Hook – curved sheath
– Basal body – stack of rings firmly anchored in cell wall
36
Flagella movement (1)
• Bacterial flagella do not move side to side like
eukaryotic flagella do instead they rotate like
a spinning propeller
• When it rotates counter clockwise the
bacterium moves in a straight line referred to
as a “run”
• When it rotates clockwise it causes the
bacteria to “tumble” and change
diretionrandomly
• Usually bacteria move in a random run and
tumble pattern
• Very fast, about 10 body lengths/sec!
Flagella movement (2)
Flagella movement (3)
• Sometimes flagellar movement is nonrandom
movement towards a chemical (Chemotaxis) or
towards light (Phototaxis)
• Substances often exist in gradients: areas of
higher to lower concentration
• Bacteria can sense these gradients by “counting”
the number of a particular substance that hit
receptors on the cell surface
Flagella movement (4)
• If the substance is desirable such as
nutrient it is called an attractant and the
bacterium will move towards it.
• If the substance is harmful, it is
considered a repellent and the
bacterium will move away from it.
• The bacterium cannot steer itself in one
direction or another. Rather it continues
to “run” if it is going in a favorable
direction or initiates a “tumble” if it is
going in an unfavorable direction.
Flagella movement (5)
• What do you
notice about the
bacterium in
each of the three
conditions?
– Is there net
movement to one
side or the other?
– When does the
bacteria “run” for
a longer period of
time?
– What makes it
“tumble?”
Axial Filaments or Periplasmic
Flagella
• Found in spirochetes
• Attached to cell membrane at each end
of the cytoplasmic cylinder that forms
the body between the cell wall and an
outer sheath
• Twisting action causes the rigid body to
rotate like a corkscrew
Axial filaments or Endoflagella
Leptospira interrogans (50,000X)
Cross section of Axial Filaments
• Fine, proteinaceous,
hair-like bristles
emerging from the cell
surface
• Function in adhesion
to other cells and
surfaces
• Makes bacteria more
dangerous as they are
able to adhere and
colonize a surface
instead of being
washed away by
mucous, urine, etc.
Fimbriae
45
E. coli (14,300X)
Conjugation
Pili
• Attach two cells and form
a pathway for the transfer
of genetic material
(conjugation)
• Found only in gramnegative cells
• Leads to genetic variety
• Problematic for us
because this is one of the
ways antibiotic resistance
genes are spread among
bacterial populations
Glycocalyx
•
•
Coating of molecules external to the cell wall,
made of sugars and/or proteins
Two types:
1. Slime layer - loosely organized and attached
2. Capsule - highly organized, tightly attached
47
Capsule
• Complex polysaccharide molecules arranged in a
loose gel
• Secreted by cell wall
• Only certain bacteria form capsules and may not
do so under all conditions
– I.e. outside a host Bacillus anthracis does not form a
capsule and inside a host it will
– Why???
– Because encapsulated bacteria are able to evade host
defense mechanisms (phagocytosis) because the
capsule covers up all the bacterial structures the
immune system would recognize as foreign
Slime Layer
• Less tightly bound to the cell wall and is
usually thinner than a capsule
• Protects the cell against drying, traps
nutrients and binds cells together
(biofilm)
• Allows for adherence to rock surfaces or
root hairs of plants to keep them near
nutrients or oxygen
• Even allows for adherence to tooth
surfaces to cause plaque
Biofilms
• The slime layer helps
Biofilms form. This
picture is of a biofilm
of bacteria growing on
tooth enamel.
• What is a biofilm?
How does the ability
to form a biofilm
makes a bacterial
species more virulent
(able to cause disease.)
The Cell Envelope
• Under the capsule and the slime layer, external
covering outside the cytoplasm
• Composed of two basic layers:
– Cell wall and cell membrane
• Maintains cell integrity
• Two different groups of bacteria demonstrated by
Gram stain:
– Gram-positive bacteria: thick cell wall composed
primarily of peptidoglycan and cell membrane
– Gram-negative bacteria: outer cell membrane, thin
peptidoglycan layer, and cell membrane
51
Prokaryotic Cell Wall
• Main functions
– Maintains characteristic shape of the cell
• I.e. if the cell wall is removed the cell will take on a spherical shape
– Prevents the cell from bursting from osmotic shock
• Osmotic shock is a sudden change in the solute concentration around a cell,
causing a rapid change in the movement of water across its cell membrane.
– Under conditions of high concentrations of either salts, substrates ,or any solute in the environment
surrounding the cell, water is drawn out of the cells through osmosis. This also inhibits the transport
of substrates and cofactors into the cell thus “shocking” the cell. Alternatively, at low concentrations
of solutes, water enters the cell in large amounts, causing it to swell and either burst or undergo
apoptosis.
– Does not regulate entry of materials into and out of the cell (job of
cell membrane)
• is rigid yet extremely porous
– Differences in the properties of cell walls result in different appearances
when stained and the ability to distinguish between G-, G+, and acid-fast
bacteria.
Structure of
Cell Walls
• Peptidoglycan is the
primary component:
– Unique
macromolecule
composed of a
repeating framework
of long glycan chains
cross-linked by short
peptide fragments
– The Antibiotic penicillin is
effective because it interferes
with synthesis of peptidoglycan
components
53
Gram-Positive Cell Wall
– thick peptidoglycan layer
– Includes teichoic acid and
lipoteichoic acid: function in
cell wall maintenance and
enlargement during cell division;
move cations across the cell
envelope; stimulate a specific
immune response
– Some cells have a periplasmic
space, between the cell
membrane and cell wall
54
Gram-Negative Cell Wall
• Inner and outer membranes and
periplasmic space between
them contains a thin
peptidoglycan layer
• Outer membrane contains
lipopolysaccharides (LPS)
• Lipid portion (endotoxin) may
become toxic when released
during infections
• G- are less susceptible to
penicillin because this outer
membrane prevents against its
entry into the cell
• Contain porin proteins in upper
layer – regulate molecules
entering and leaving cell
55
Structures of Gram-Positive and
Gram-Negative Bacterial Cell Walls
56
Another view of Gram-Positive and
Gram-Negative Bacterial Cell Walls
Comparison of Gram-Positive
and Gram-Negative Cell Walls
58
What does the cell wall tell us
about bacteria?
• Differential stains (I.e. the Gram Stain or
the acid fast stain) allow us to
differentiate between types of bacteria
based on differences in their structures
The Gram Stain (1)
The Gram Stain (2)
How does the Gram Stain work?
• G+ bacteria have very thick peptidoglycan and they retain
the crystal violet = purple
• G- bacteria have very thin walls of peptidoglycan lose the
crystal violet easily during the decolorization stage and are
counter-stained with safranin = red
• Very good stain…been around for 150 years!
• But some bacteria, gram-variable and acid-fast must be
stained using other methods
Acid-fast bacteria
• Mycobacteria
– Have a different cell wall composition
• Thick like G+ bacteria
• 60% lipid (mycolic acid) and contains much less peptidoglycan
• These lipids make the bacteria highly impermeable to most stains as
well as protect them from acidic and alkali solutions
– Slow growers because this relatively impermiable membrane also
impedes entry of nutrients into the cell as well
– Acid fast staining uses carbolfuchsin which binds to the cytoplasm
then resists removal upon addition of an acidic solution
– Why is it so important to have a difinitive way to identify these
organisms?
• Mycobacterium tuberculosis and Mycobacterium leprae are both acid-fast
bacteria that cause serious disease in humans.
Acid Fast Cell Wall
Wall-Deficient Organisms
• Bacteria that belong to the genus Mycoplasma
have no cell walls
• Protected from osmotic swelling and bursting by
a strengthened cell membrane that contains
sterols
• One example of a sterol is cholesterol. Animal
cells also use this molecule to strengthen their
cell membranes.
Practice
• Draw the cell membrane and all parts of
the cell wall of the following bacterial
types:
– Gram-positive
– Gram-negative
– Acid-fast
Selective Toxicity
• Antibiotics can have selective toxicity – meaning
they are harmful to micorbes without causing damage
to the host. Typically this is due to them affecting a
structure that is present in microbes and absent in
the host.
– Penicillin inhibits peptidoglycan synthesis and therefore
prevents dividing bacteria from making a functional cell wall
and they die from osmotic shock.
– Can penicillin be harmful to humans? Do you consider this
drug to have good selective toxicity?
Prokaryotic Cell Membrane
• Phospholipid bilayer
with embedded proteins
• Represented by the
fluid mosaic model
• Functions in:
– Providing site for
energy reactions,
nutrient processing,
and synthesis
– Passage of nutrients
into the cell and
discharge of wastes
– Cell membrane is
selectively permeable
68
• Phospholipids (keeps
anything besides
gasses and water from
getting into or out of
the cell)
• Hydrophilic (waterloving), polar, phosphate
head group
• Hydrophobic (waterfearing), non-polar fatty
acid chains as tails
• Proteins serve as
gates to allow nutrients
in and wastes out
Selective
Permeability
Inside a Prokaryotic Cell
• Cytoplasm
• Ribosomes
• Nucleoid
– Chromosome
– Plasmids
• Storage Structures
• Cytoskeleton
• Endospores
Cytoplasm
• Semi-fluid substance inside the plasma
membrane
• 4/5 water
• 1/5 materials dissolved or suspended
•
•
•
•
Enzymes and other proteins
Carbohydrates
Lipids
Inorganic ions
• Site of many chemical reactions
Bacterial Ribosome
– Site of protein synthesis
– Found in all cells
– Made of 60% ribosomal RNA
and 40% protein
– Consist of two subunits: large
and small
– Prokaryotic ribosomes differ
from eukaryotic ribosomes in
size and number of proteins
• Some antibiotics, streptomycin
and erythromycin, disrupt bacterial
protein synthesis by binding to the
70S prokaryotic ribosomes but
leaving the 80S eukaryotic
ribosomes unaffected.
Nucleoid
• Chromosome
– Single, circular, doublestranded DNA molecule that
contains all the genetic
information required by a cell
• Plasmids
– Free small circular, doublestranded DNA
– Not essential to bacterial
growth and metabolism
– Used in genetic engineering readily manipulated and
transferred from cell to cell
Bacterial Internal Storage (1)
• Inclusions and granules
– Intracellular storage bodies
– Vary in size, number, and content
– Bacterial cell can use them when environmental
sources are depleted
74
Bacterial Internal Storage (2)
– 1. Granules
• Not membrane-bound
• Materials so densely compacted that they do not easily
dissolve in cytoplasm
• I.e. glycogen, phosphate
– 2. Inclusions
• Specialized membrane enclosed structures
• I.e. gas vacuole used by aquatic organisms to regulate
depth at which they float and therefore intensity of light
that reaches them
• I.e. containing the lipid p-B-hydroxygutyrate which is
used as an energy store
• I.e. containing iron (magnetosomes) enable the bacteria
to respond to the earth’s magnetic fields
Spores -Sporulation cycle
76
Endospores (1)
• Dehydrated, metabolically inactive
• Spore-formation is induced following nutrient
depletion
– Can also be found to occur in normal conditions and may
be a way that the bacteria hold troops in reserve so as
not to be wiped out by sudden onset of poor conditions
• Resistant to ordinary cleaning methods and boiling
– Pressurized steam at 120oC for 20-30 minutes will
destroy
• Some common spore forming bacteria are Bacillus
species and Clostridium species
• Are you familiar with Bacillus anthracis, Clostridium tetani,
Clostridium botulinum, and Clostridium difficile?
77
• How does the ability to form spores make these organisms more
dangerous to humans?
Endospores (2)
• NOTE: bacterial endospores are not the
same as fungal spores which are means of
reproduction, endospores are a means of
survival
Activity:
Can you name each structure? Can you briefly
describe its function? You may want to organize the
information in a table for study purposes.
How are prokaryotes
and eukaryotes related?
•
•
The first prokaryote appeared approximately 3.5 billion years ago
The first eukaryote appeared approximately 1.7-2.2 billion years ago
• What led to this increase in cellular complexity???
– Evidence suggests that large prokaryotes engulfed smaller
prokaryotes, lived together with them in symbiosis creating
a larger, more complex eukaryotic cell.
– This is called the Endosymbiotic Theory and was proposed
by Lynn Margulis in the 1960s. (Read 5.1 Making
80
Connections in your textbook on page 124)
The Endosymbiotic Theory
•
The term "endosymbiosis" means "to cooperate inside".
–
–
•
The smaller cells were likely what we know of as mitochondria and chloroplasts today.
–
–
–
•
•
Advantage to the smaller cell is…protection. No other cells will engulf (and digest!) them.
Advantage to the larger cells is…the smaller cells produce energy which can be used by the larger
cells.
Both of these organelles have their own DNA and their own ribosomes. The DNA is in a single
circular chromosome and the ribosomes are 70S. (Remind you of prokaryotic DNA and ribosomes?)
In fact, the DNA in the chloroplast is very similar to photosynthetic bacteria called cyanobacteria.
The DNA in the mitochondria is most like that of the bacteria that causes typhus.
Both of these organelles divide inside the eukaryotic cell independently of the cell cycle in a process
that looks very similar to binary fission. (Remind you of prokaryotic cell division?)
When the larger cell that had engulfed the smaller cells went to divide, copies of the smaller
prokaryotes inside were made and passed down to the daughter cells. Eventually, the
smaller prokaryotes that had been engulfed adapted and evolved into some of the
organelles we know of today in eukaryotic cells like the mitochondria and chloroplasts.
Other organelles eventually arose from these first organelles, including the nucleus where
the DNA in a eukaryote is housed, the endoplasmic reticulum and the Golgi Apparatus. In
the modern eukaryotic cell, these parts are known as membrane bound organelles. They
still do not appear in prokaryotic cells like bacteria and archaea, but are present in all
organisms classified under the Eukarya domain.
What do you think?
• Is the
endosymbiotic
theory a
plausible
explanation for
the appearance
of eukaryotic
cells on earth?
Topic1 – Main Themes in
Microbiology
Overall Course Goals
▪ To share knowledge from different sources to give
you a healthy appreciation of the microscopic
world around you.
▪ To introduce or reintroduce you to scientific
concepts that you will encounter in future science
courses and/or as a healthcare professional.
▪ To help you distinguish between a good source
and a poor source of scientific information. Help
you become a good “consumer” of science.
Outline
▪ What is microbiology?
▪ Why study microbiology?
▪ Nomenclature (scientific names) of bacteria as
well as other living organisms
▪ Classification of living organisms
▪ Historical contributions to the development of
microbiology
▪ Fields in microbiology
What is Microbiology?
▪ Microbiology = study of microorganisms or
microbes
▪ Microorganisms = organisms too small to be seen
with the naked eye
▪ Can you give an example of a(n)…
▪
▪
▪
▪
▪
▪
Bacteria
Algae
Fungi
Protozoa
Helminthe
Virus
Can you give an example of…
▪ Microbes that are harmful to humans?
– Disease causing microbes
– Microbes that cause food to spoil
▪ Microbes that are helpful to humans?
– Normal microbiota
– Microbes used to produce food/beverages
As you go through the following slides, you can
use this table to help you condense what you
know about the 7 classes of microorganisms.
Class of
Microorganism
Prokaryotic Single or
/Eukaryotic Multicellular?
or Neither?
Nutrition
type?
Examples?
Other info?
Bacteria
▪ Very small, relatively simple,
single-celled
▪ Prokaryotic cells
– Lacks a nucleus and other
membrane bound organelles
▪ Ubiquitous in Nature
▪ Absorb nutrients from their
environment
▪ Diverse metabolic
capabilities
▪ Reproduce by Binary Fission
▪ Many are pathogens, but
despite the large amount of
attention devoted to diseasecausing bacteria less than
1% of all known bacteria are
harmful!
Archaea
▪
▪
▪
Very small, simple, exist as single
cells
Prokaryotic
When first discovered, they got a
reputation as extremophiles
because they live in what we would
consider unlivable conditions
–
▪
▪
Even more metabolically diverse
than bacteria
Not known to be pathogenic, but
many are part of our normal flora
–
▪
Bottom of the ocean, in lakes full of
sulfuric acid deep in caves, hot springs
created by volcanoes, etc.
Normal flora are all of the microbes that
live in an on our bodies
More closely related to eukaryotic
cells like us than other prokaryotic
cells like bacteria!
Where do Archaea fit in?
▪ Eukaryotic cells
Algae
– Unlike prokaryotic bacteria
eukaryotic cells have a nucleus and
membrane bound organelles
▪ Most are microscopic, singlecelled organisms
▪ Some are large, multicellular
organisms
▪ Autotrophs (self feeders) vs. all
the other classes of microbes
that are Heterotrophs (other
feeders)
– Photosynthetic: important oxygen
producers in the ecosystem
▪ Found in fresh and salt water
▪ Few are harmful
– Red Tide
What is Red Tide?
Red tide is a result of population
explosion or algal bloom
▪ Organisms implicated include (protists)
dinoflagellate species Gymnodnium and
Gonyaulax
▪ The dinoflagellates produce a powerful neurotoxin
that affects vertebrates
▪ The shellfish that eat them are not affected but the
fish, seals, or even humans that eat the shellfish
get sick
Red Tide Continued
▪ Does sufficient cooking of shellfish which have fed
on Gonyaulax (the protist dinoflagellate that
produces red tide) protect us from getting sick?
▪ Unfortunately, no.
▪ There is no way to visually tell if your shellfish is affected. It
doesn’t look or taste differently. We are protected because the
shellfish we buy at the store or eat in a restaurant is regularly
tested for toxin levels.
▪ You would have to be more vigilant if you were to go out and
harvest your own wild shellfish.
Fungi
▪ Eukaryotic organisms (nucleus)
▪ Can be single-cellular like yeast or multicellular
like mushrooms
▪ Cell walls – most have chitin
▪ Saprophytic (feeds off dead or decaying organic
matter)
▪ Found everywhere – especially soil & water
▪ A few species are harmful to humans – i.e. yeast,
ringworm, maduromycosis.
Fruiting bodies, budding yeast,
candiasis of oral cavity,
& mandura foot
Protozoa
▪ Eukaryotic cells –
unicellular
▪ Ingest or engulf food from
surroundings
▪ Many can resist drying
out/death by going into a
dormant state called a
cyst
▪ Many known as pathogens: cause
malaria, amoebic dysentery, African
sleeping sickness, toxoplasmosis
Helminthes (Worms)
▪ Eukaryotic cells
▪ Multicellular organisms
▪ But many have
microscopic lifestyles,
such as dog
heartworm, swimmer’s
itch, tapeworms
Viruses
▪
▪
▪
▪
Very small
Noncellular (acellular)
No metabolism
The ultimate parasite must use a host to
reproduce
▪ All groups of organisms
are infected by viruses
▪ Many are harmful – HIV,
herpes, polio, common
cold, measles, rabies,
hepatitis, Ebola, etc., etc.,
etc.
Microbial Diversity: 6 of the 7 Types of
Microbes
Why aren’t archaea always included
in the classes of microbes?
▪ Depending on your source you may or may not
see archaea included in the types or classes of
microbes.
▪ Archaea were only discovered as being distinct
from bacteria in the late 70s.
▪ There are no known archaea that are human
pathogens.
▪ There are recently discovered archaea found to
live in the human gut!
▪ For the purposes of this course, we will consider
archaea the 7th class of microbes.
Microbial Structure
• Two cell lines
– Prokaryote – microscopic, unicellular organisms, lack
nuclei and membrane-bound organelles
– Eukaryote – unicellular (microscopic) and multicellular,
nucleus and membrane-bound organelles
• Viruses - Acellular, parasitic particles composed of a nucleic
acid and protein
Top Microbial Causes of Death
Why Study Microbiology(1)?
Helpful….
– Microbes cover the human body inside and out and are
an important part of human health
– They outnumber us 10 bacterial cells to 1 human cell!
– Important in protecting us from pathogen colonization
▪ Take up space in the body
▪ Compete for nutrients
▪ Even produce antimicrobial compounds
Harmful….
– On the other hand, pathogens are microbes too
– And even the “helpful” microbes can become harmful if
they find a way into an otherwise sterile part of the body
(opportunistic pathogens)
Why study Microbiology (2)?
▪ Understanding gained from studying complex life processes like
metabolism in simple single-celled organisms can be applied to
higher organisms
▪ Microbes play an important role in the ecosystem both as a part of
many cycles as well as in recycling dead material.
N Cycle
Why study Microbiology(3)?
▪ Microbes are used in research and medicine
– Much simpler than maintaining mice, rats, etc.
– Great numbers in little time (E. coli can double
in 20 minutes)
– Microbes are used to produce large amounts of
antibiotics, insulin, etc. at low cost
What is life?
All life is…
▪ Composed of cells
▪ Carries out similar functions like metabolism
– Obtaining nutrients and eliminating wastes
▪ Able to grow and reproduce
How do we name living
organisms (Taxonomy?)
▪ Carolus Linnaeus (1707-1778) devised the Binomial
Nomenclature System
– Each distinct species is given a scientific name by scientists that is
standard around the world in place of common names that may differ
and cause confusion.
– Scientific name consists of Genus & species
▪ Escherichia coli, Escherichia is the genus and coli is the
species.
▪ The Genus is always capitalized and the species is always lower
case.
▪ It is set apart from the rest of the text by putting it in italics if typed
or underlined if handwritten.
▪ It must be spelled out in its entirety the first time used.
Subsequent uses can be abbreviated with just the first letter of the
Genus and the entire species. (E. coli)
Sample Taxonomy
How do we classify living
organisms?
We can use the Five-kingdom classification
system or the Three-domain classification
system.
Imagine that you are a scientist
living in the 1700s…
▪ You identify a new organism.
▪ How would you classify it?
▪ What tools would you have available to you?
Five-Kingdoms
▪ The answer…not many!
You would make most
of your observations
with your eyes alone.
You’d note similarities
and differences that
you could see –
otherwise known as
morphology.
▪ You would probably
group anything green
that thrives in sunlight
in the “plants” branch.
Anything that moved
and ate other things in
the “animals” branch.
Five-Kingdoms Continued
▪ For the small, single celled
organisms, anything with a
nucleus was placed in
“Protists.” While anything
without a nucleus was placd
in “Monera.”
▪ Monera includes eubacteria
(most of the bacteria
relevant to human health),
cyanobacteria which used to
be called blue-green algae
because of their
photosynthetic nature, as
well as archaeobacteria (the
extremists) which live in
seemingly uninhabitable
environments (salty, hot,
acidic, cold, under great
pressure)
New inventions
▪ In 1977, Carl Woese looked at the genetic makeup
of living things and found that plants, fungi,
animals, and protists were remarkably similar .
▪ On the other hand, there were two very distinct
groups of prokaryotes lumped into Monera.
▪ He devised the Three Domain system of
classification to better represent how closely
related one organism is to another genetically.
Three Domains
Bacteria - true
bacteria
Archaea - odd
bacteria that live in
extreme
environments, high
salt, heat, etc.
Eukarya - have a
nucleus and
organelles
Three Domains Continued
▪ The closer two organism are to
one another = the greater their
genetic similarity.
▪ So if you look at the end of the
Eukarya branch (shown in
blue) you can see how closely
humans(animals) are related
to fungi!
▪ On the other hand, if you look
at E. coli (found on the green,
Bacteria, branch), you can see
how distantly related it is to
Methanogens (found on the
orange, Archaea, branch.)
Antonie van Leeuwenhoek (1632-1723)
▪
Dutch linen merchant
▪
First to observe living
microbes
▪
Single-lens magnified
up to 300X
▪
Made detailed drawings
of microbes he
observed in drops of
water and most
famously from scrapings
taken from teeth (see
his quote on the next
slide)
Anton van Leeuwenhoek Quote
“For my part I judge,
from myself
(howbeit I clean my
mouth like I’ve
already said), that
all the people living
in our United
Netherlands are not
as many as the
animals that I carry
in my own mouth
this very day.”
Leewenhoek’s “little animals”
History of Microbiological Thought
▪ Two opposing schools of thought:
– Spontaneous Generation is an early belief that some
forms of life could arise from vital forces present in
nonliving or decomposing matter
▪ The theory of Spontaneous generation would argue that microbes arise in the
body as a RESULT of disease state instead of as the CAUSE of the disease
state
– Theory of Biogenesis - the idea that living things can
only arise from other living things
▪ This theory states that the microbe invades which CAUSES the
disease state.
Francesco Redi
▪ Experimental
Design
– Three jars with a
piece of meat in the
bottom
– One left uncovered,
one corked, and
the last covered
with gauze
– Where are
maggots present?
– Did this experiment
confirm or refute
spontaneous
generation?
Redi’s Experiment Continued
▪ Redi’s experiment refuted spontaneous
generation.
– There were maggots present on the meat in the
container left open to the air (and the flies that lay egg
which develop into maggots.)
– There were no maggots found on the meat in the sealed
jar.
– There were maggots on top of the gauze in the third jar
(flies were fooled by being able to sense the meat.)
– If spontaneous generation were true there would be
maggots on all samples of meat.
Louis Pasteur
▪ Louis Pasteur – boiled infusions
in flasks and then drew glass
tops into swan-necks
▪ No microbes grow in broth
▪ Why did this work?
▪ Microbes can fall but not
fly…therefore they get trapped in
the dip of the swan neck and
never reach the broth to grow
▪ If the swan neck is broken off
Microbes will grow
Others tried to repeat Pasteur’s
work – but failed.
Were they wrong? Lack of skill? Or something else?
Spore forming microbes contaminated many of these flasks.
Pasteur did not have these in his laboratory.
Spore forming bacteria concentrate their genetic material into a hard
calcium shell that can resist desiccation (drying out), nutrient deprivation,
and heat treatments such as boiling or even some autoclaves.
(Do not confuse bacterial spores with fungal spores which are used for
reproduction. Bacterial spores are merely used for survival in an
unfavorable environment. So does that mean that bacterial spores are
similar to protozoa cysts?)
Once put into a favorable environment bacterial spores can regain their
vegetative state and reproduce
Pasteur Continued
▪
▪
▪
▪
Portrait painter
Teacher
Professor of Chemistry
Wine and silkworm industries
▪ Found yeast produced good wine and bacteria-contaminate wine was sour
▪ Identified three different pathogens of silkworms and associated each with a
specific disease
▪ Pasteurization of wine
▪ Heating in the absence of oxygen
▪ Any other food products pasteurized?
▪ Development of vaccines – rabies in particular
▪ Used the dried spinal cord from rabbits infected with rabies
Robert Koch
▪ Identified bacterium causing anthrax: Bacillus
anthracis; tuberculosis: Mycobacterium
tubuculosis; cholera: Vibrio cholerae.
▪ Recognized active versus dormant cells
(spores)
▪ Developed techniques for studying cells in vitro
▪ Pure cultures – found a way to grow specific
bacteria separated from other bacteria by
streaking (Help from Angelina and Walther
Hesse)
▪ Koch’s Postulates – most outstanding
achievement
▪ Research on malaria, typhoid fever, sleeping
sickness, and more.
Koch’s Postulates:
One organism-one Disease
▪ 1. The specific causative agent must be found in every
case of the disease.
▪ 2. The disease organism must be isolated in pure culture.
▪ 3. Inoculation of a sample of the culture into a healthy,
susceptible animal must produce the same disease.
▪ 4. The disease organism must be recovered from the
inoculated animal.
Koch’s Postulates Continued
Angelina & Walther Hesse
▪ Walther was a
colleague of Koch.
▪ Angelina suggested that
Koch add agar, a
cooking thickener, to
broth.
▪ Use of agar was a
turning point in studies
of bacteria because it
allowed for isolation of
pure culture necessary
to identify one organism
to one disease in
Koch’s Postulates.
Hospital Sanitation
▪ Ignaz Philipp Semmelweis
– encouraged sanitary
practices in hospital
▪ Joseph Lister – Father of
antiseptic surgery which
reduced the incidence of
infections
Early Immunology
▪ Immunology – The science of immunology deals with the
host’s response to microbial invasion. This science led to
the development of vaccines
▪ Jenner – discovered that cowpox protects against
smallpox. The word vaccine comes from the Latin
vacca for cow.
▪ Pasteur – developed vaccines for rabies and cholera.
Using the Scientific Method to
Eradicate Small Pox (1)
Using the Scientific Method to
Eradicate Small Pox (2)
Early Virology
Virology – special techniques were
necessary for isolating, propagating,
and analyzing
▪ Porcelain filters originally developed to
remove bacteria from water. Used to
isolate viruses!
▪ Beijerinck determined that filtrate could be
infectious and used the term virus (from
the Latin word for poison).
▪ Electron microscopes were developed in
the 1930s and first virus seen in 1939.
▪ Hershey and Chase showed that viruses
contained DNA and that only the DNA had
to go into a host cell (bacteria) to cause
more viruses to be formed.
▪ DNA is the genetic information
Early Chemotherapy
▪ Chemotherapy - development of drugs to treat
disease began at least by the first century AD with
herbal medicines
▪ Discovery: quinine from tree bark could treat malarial
infections.
▪ Paul Ehrlich – first serious researcher in this area
who discovered the first successful drug treatment
for syphilis. Coined term “chemotherapy”
▪ Fleming discovered penicillin in 1928. “Others”
purified and mass produced during WW II.
Antibiotics-the wonder drugs
▪ Along with improved working conditions, better sanitation,
better hygiene, immunizations, and less crowding, antibiotics
can take credit for increasing life expectancy from 47 in the
1900s to 68 in the 1950s
▪ Many thought we had conquered disease and that further
research was not imperative
▪ World focus switches to Cancer in the 1970s
▪ In 1981 the emergence of AIDS was the wake up call
▪ More recently we are dealing with the increase in antibiotic
resistant strains of bacteria (another wake up call)
Early Molecular Biology
▪ Molecular Biology – youngest
branch of microbiology.
▪ Watson and Crick discovered the
double helix structure of DNA in 1953.
▪ Microbial genetics led to many
important genetic discoveries including
the three domain classification system
Areas of Study: Fields of Microbiology
▪ Organismal groups: bacteriology, phycology (studies algae),
mycology (studies fungi), protozoology, parsitology, virology.
▪ Processes or functions: microbial metabolism, microbial genetics,
microbial ecology.
▪ Health Related Fields: immunology, epidemiolgy, etiology,
infection control (nosocomial infections), chemotherapy.
▪ Applications: disease related; food & beverage technology,
environmental; industrial, pharmaceutical, genetic engineering.
Tools of the Microbiology Lab
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The Microscope
Key characteristics of
a reliable
microscope are:
• Magnification –
ability to enlarge
objects
• Resolving power or
resolution– ability to
show detail
2
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Magnification
Magnification is the
enlargement of an
image due to an
interaction between
visible light waves and
the curvature of a lens.
3
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Magnification in Two Phases
– The objective lens is
found on the “nose”
of the microscope.
Our lab has
microscopes with 4X,
10X, 40X, and 100X
objectives.
– The ocular lens is
found in the
eyepiece of the
microscope. Our
oculars have a
magnification of 10X.
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Total Magnification
• Total magnification of
the final image is a
product of the separate
magnifying powers of
the two lenses
objective ocular
total
x
=
power
power magnification
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Resolution
• Resolution is the
capacity to distinguish or
separate two adjacent
objects
• The level of detail we can see
• depends on the wavelength of
light that forms the image
along with characteristics of
the objectives
• In the picture on the right, you
can see image B is better
resolved because the separate
fingers are visible
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The Purpose of Oil is to increase the resolving
power of a microscope by. In our lab it is used
with the 100X objective.
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Bright-field Microscopy
• most widely
used
• specimen is
darker than
surrounding
field
• used for live
and preserved
stained
specimens
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Electron Microscopy
• Forms an image with a beam of electrons (instead
of light) that can be made to travel in wavelike
patterns when accelerated to high speeds
• Electron waves are 100,000 times shorter than the
waves of visible light
• Electrons have tremendous power to resolve
minute structures because resolving power is a
function of wavelength
• Magnification between 5,000X and 1,000,000X
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Thermal worm – found in deep sea vents…smaller than a bacterial
cell and only visible with an electron microscope. What else is out
there that we haven’t seen yet?!?!
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Specimen Preparation for
Optical Microscopes
• Wet mounts and hanging drop mounts –
allow examination of characteristics of live
cells: size, motility, shape, and arrangement
• Fixed mounts are made by drying and
heating a film of specimen. This smear is
stained using dyes to permit visualization of
cells or cell parts.
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Staining
• Dyes are used to create contrast by imparting
color to either the microbe (positive stain) or to
the background (negative stain)
– Simple stains – one dye is used; reveals shape,
size, and arrangement
– Differential stains – use a primary stain and a
counterstain to distinguish cell types or parts
(examples: Gram stain, acid-fast stain, and
endospore stain)
– Structural stains – reveal certain cell parts not
revealed by conventional methods: capsule and
flagellar stains
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Staining Examples
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The 6 I’s of Culturing Microbes (1)
Inoculation – introduction of a sample into a
container of media to produce a culture of
observable growth
Isolation – separating one species from another
Incubation – under conditions that allow growth
Inspection
Information gathering
Identification
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The 6 I’s of Culturing Microbes (2)
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Isolation
• If an individual bacterial cell is separated from other
cells and has space on a nutrient surface, it will grow
into a mound of cells— a colony. A colony consists of
one species.
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Isolation Techniques
– Streak plate
technique
– Pour plate
technique
– Spread plate
technique
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Inspection
• If a single species is growing in the container, you have
a pure culture but if there are multiple species than
you have a mixed culture.
• Check for contaminants (unknown or unwanted
microbes) in the culture.
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Ways to Identify a Microbe:
• Cell and colony
morphology or
staining
characteristics
• DNA sequence
• Biochemical tests to
determine an
organism’s chemical
and metabolic
characteristics
• Immunological tests
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Media: Providing Nutrients in
the Laboratory
Media can be classified according to three properties:
1. Physical state – liquid, semisolid, and solid
2. Chemical composition – synthetic (chemically
defined) and complex
3. Functional type – general purpose, enriched,
selective, differential, anaerobic, transport, assay,
enumeration
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Physical States of Media
Liquid – broth; does not
solidify
Semisolid – contains
solidifying agent
Solid – firm surface for
colony formation
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Agar
– The most commonly used
solidifying agent
– Solid at room temperature,
liquefies at boiling (100oC),
does not re-solidify until it
cools to 42oC
– Provides framework to hold
moisture and nutrients
– Not digestible for most
microbes
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Chemical Content of Media
• Synthetic – contains pure organic and
inorganic compounds in an exact chemical
formula
• Complex or nonsynthetic – contains at least
one ingredient that is not chemically definable
• General purpose media – grows a broad
range of microbes, usually nonsynthetic
• Enriched media – contains complex organic
substances such as blood, serum,
hemoglobin, or special growth factors
required by fastidious microbes
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Examples of Enriched Media
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Selective & Differential Media
Differential
Selective media: contains one or more agents
media:that
allows
inhibit growth of some microbes and encourage
growth of several
growth of the desired microbes
types of microbes
and displays
visible differences
among those
microbes
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Some media can be both
Selective & Differential
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