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Chapter 5
Bacteria structure and physiology
Objectives: After reading Chapter Five, you should understand…
• The enormous span of time that bacteria have spent on Earth as well as their contribution
to it.
• The various forms of bacteria and the structures of the cell.
• How bacteria reproduce and with what frequency.
• The environments (natural and artificial) in which bacteria live.
How old is the Earth?
What were conditions like on early Earth?
Where did energy come from?
Were the biochemical subunits we discussed previously present on early Earth?
What other factors might have been important for life?
How long did it take all of these parameters to fall into place such that microbial life
could begin?
From 3.5 billion years ago until about 1.5 billion years ago, bacteria (prokaryotes) were
the only life forms on Earth.
Stromatolites in a tidal pool.
Electron microscopy image of a stromatolite (100,000 x
magnification).
Ancient cyanobacteria began building up stromatolites about three-billion years
ago.
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Bacteria begin producing oxygen
Eukaryotes then evolved, and they required:
More energy, more nutrients, more space and “nicer” conditions.
Bacteria had an advantage…what was it?
Bacteria are still the dominant life form today, as they occupy
every conceivable niche on Earth.
Bacteria structure and physiology
General morphology
Three major forms
Bacilli (bacillus) – rod-shaped – Escherichia coli
Cocci (coccus) – spherical – Staphylococcus aureus
Spirilla (spirillum) – flexible – Vibrio cholera
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Escherichia coli
Staphylococcus aureus
Vibrio cholera
These configurations can be useful for identifying clinical bacteria.
How do we visualize bacteria?
Since bacteria generally do not exhibit coloration in their natural state, staining of
bacteria is often necessary in order to visualize them with a microscope.
Bacteria cytoplasm (the “insides”) and polar heads of the membrane are
negatively charged and will therefore attract a positively-charged stain, like
crystal violet or methylene blue.
Staining types:
Simple staining – easy, one step.
Corynebacterium diptheriae
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Gram staining – developed by Christian Gram (1880s)
Performed in multiple steps and separates bacteria into two groups.
Gram positive (G+) and Gram negative (G-)
G+ bacteria have a thick cell wall (peptidoglycan) that absorbs the stains,
while G- bacteria have thinner cell walls that don’t absorb as effectively.
Gram stain of E.
Streptococcus spp. (G+)
coli
(G-)
and
Question: Why do we do the last “safranin” step? Aren’t the bacteria
differentiated after the alcohol wash?
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Surface structures of bacteria
Know: Cell wall, cell membrane, glycocalyx, pilus, flagellum, as well as
associated structures.
Almost all bacteria are encased in a CELL WALL.
Contains a tough network of polysaccharide and protein called peptidoglycan.
Strength and rigidity – can withstand over 350 pounds of pressure per
square inch!
Peptidoglycan is found only in bacteria (not fungi, viruses or protists).
Gram positive – thick peptidoglycan layer
Gram negative – thin peptidoglycan layer (dual membrane layer)
The essential functions of peptidoglycan and its confinement to bacteria make it a
perfect target for antibiotics meant to rid the body of infections.
Certain antibiotics attack infecting organisms by targeting the synthesis of
peptidoglycan.
e.g. β-lactam antibiotics, such as penicillin, ampicillin, amoxicillin
and methicillin.
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Death of a bacterium after
treatment with an antibiotic
targeting peptidoglycan synthesis.
?
Which will be more susceptible to β-lactam antibiotics, Gram (-) or Gram (+)
bacteria?
The CELL MEMBRANE is internal to the cell wall and contains a double layer of
phospholipids (bilayer).
The membrane is the most important boundary between the interior of the
cell and the external environment.
The cell membrane encloses the CYTOPLASM.
The membrane will allow small molecules to pass through, but big
molecules cannot pass.
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Proteins are embedded in the layers and sometimes span the entire thickness of
the membrane.
These proteins serve two general functions (know the difference):
1. Enzymes that catalyze chemical reactions
2. Porins (membrane-spanning proteins) can carry molecules (e.g.
nutrients, antibiotics) across the membrane.
Small carbohydrates are found on the outside of the membrane and can serve as
recognition sites for host immune defenses (antibodies)
e.g. in E. coli O157:H7, the “O157” refers to a membrane surface
carbohydrate.
A GLYCOCALYX (or “capsule” if rigid and highly structured) can protect the cell
from environmental stresses.
External mesh of polysaccharides that coat the cell.
India Ink capsule stain of Klebsiella pneumoniae
showing white capsules (glycocalyx)
surrounding purple cells.
From: biosci.sierracollege.edu/.../capsule_stain.html
1 µm
The saccharides in the glycocalyx are sticky.
Why would bacteria want to stick to things?
What types of bacteria might benefit from a glycocalyx?
Hint: a glycocalyx can help a bacterium evade host defenses.
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Attachment can also be achieved through a PILUS (plural PILI).
Cylindrical rod of helical protein about 1 µm in length and 7 nm thick.
E. coli attaching to the lumen of a mouse.
From: http://www.pnas.org/content/97/16/F1.medium.gif
Pili allow attachment of infectious bacteria to host cells…
Example, some therapeutic drugs, such as the antibiotics ampicillin and
streptomycin, act to inhibit the formation of the pilus.
…and to each other.
1.
Pili are important for the exchange of
genetic material between two cells.
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2.
Streptococci and other oral bacteria can bind to small pockets between
teeth and gums, leading to dental caries.
Bacteria will break down sugars, releasing acids that eat away at the
tooth enamel = cavity.
As tough as tooth enamel is, it’s not indestructible. Acids from foods and bacteria
can eat away at it, causing erosion and cavities
Where did these sugars come from?
Why do the bacteria break them down?
Bacteria that penetrate the soft inner tissue of the tooth produce gases that
press against nerve ending in the tooth, resulting in a toothache.
Do bacteria move?
While most bacteria movement is passive, some bacteria can be propelled through
liquid media by the action of a FLAGELLUM.
Rigid, rotating filament of protein about 20 nm in thickness.
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Bacteria can have single, tufted or several flagella over the cell surface.
A-Monotrichous
B-Lophotrichous
C-Amphitrichous
D-Peritrichous
An E. coli flagellum can propel the bacterium about 2000 times its body
length in an hour.
Equivalent to a person walking 2.25 miles per hour.
Schematic of the structure of a flagellum. From: http://www.sedin.org/pics/flglm-lg.jpg
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Cytoplasmic structures (these are the ones inside the cell)
Know: Chromosome, plasmid, ribosome, endospores and associated structures.
The cytoplasm is essentially a soup of various compounds such as carbohydrates, lipids
and nucleotides.
CHROMOSOME – closed loop of DNA
Only one chromosome is present in bacteria
Contains thousands of genes (4000)
How many pairs of chromosomes do humans have? How many
genes?
What is the function of the chromosome?
Some bacteria contain tiny loops of DNA that are separate from the chromosome
called PLASMIDS.
Contains genes that encode nonessential (under normal conditions) cell
functions.
1. Antibiotic resistance
2. Contaminant degradation
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Plasmids can be swapped between bacteria. What structure can mediate
this?
This is called gene exchange.
RIBOSOMES are made of proteins/nucleic acids and link amino acids together
to make polymers called peptides.
Eventually, the peptide chains become proteins when they fold in specific
configurations.
Conceptual drawing of the shape of
the ribosome.
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mRNA
Multiple ribosomes translating a
mRNA strand (arrow) to make
proteins.
Can you tell which
direction the ribosomes are moving?
Growing peptide chain
that wil eventually
become a protein.
Ribosome…you can’t see it.
ENDOSPORES (spores) are structures that can confer a high degree of stress
tolerance.
Members of the genera Bacillus and Clostridium can produce endospores, but not
all bacteria have this ability (not E. coli)
Endospores
A stained preparation of
Bacillus subtilis showing
endospores (green) and the
vegetative cell (red). From:
Wikipedia.
Vegetative cells
Spores contain a chromosome (DNA), two cell membranes, cortex, spore coat,
and a surrounding cell wall (exosporium).
The cortex gives the spore strength and rigidity, what do you think it is made
from?
Spores are storage units (like “seeds”) of bacteria.
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Spore extremes:
Spores are likely the most resistant forms of life known.
1995 - Researchers isolated intact spores from a 25
million year-old fossilized bee in a piece of
amber.
2001 – Researchers revived Bacillus spores from
within salt crystals that were 250 million
years old.
Some spores can be boiled for two hours or left in
alcohol for 20 years and still remain viable.
The ability of bacteria to form spores provides a basis for certain forms of bioterrorism.
Bacillus anthracis, the bacteria that causes anthrax, is a popular bacterium for
these purposes.
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Bacillus anthracis (vegetative)
B. anthracis spores
Once in an environment that is suitable for growth, the spores will germinate,
become vegetative cells, and potentially infect.
Spores enter the body through:
Ingestion - Progresses to sepsis (25 – 65% mortality if not treated)
Inhalation - Macrophages engulf the inhaled endospores and transport
them to lymph nodes. Progresses to meningitis (100%)
Cutaneous (skin) contact – Ulcer at site of infection. Rapidly progresses to
necrosis. (20%).
What route of exposure might terrorist groups choose to use when
weaponizing B. anthracis?
Growing B. anthracis is easy, but weaponizing it is difficult.
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Aerosolization is a difficult process. What might make aerosolization difficult?
U.S. and Soviet bioweapons
specialists discovered that
adding silica particles to
germ powders made them
easier
to
disperse.
Illustration by C. Cain,
adapted from S. Jacobsen.
By alleviating the clumping, aerosol dissemination would be much easier
to achieve.
Eliminating clumping is likely the main obstacle preventing more
widespread use of B. anthracis as a bioterrorism weapon.
If released, it would only take kilogram quantities of spores to cover a 100 square
km are and cause 50% mortality.
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In 1979, an unintentional release of anthrax spores occurred in the former Soviet Union at
a biological weapons facility. It was reported that 94 cases of anthrax occurred among
citizens living near the facility resulting in 64 deaths. It was estimated that less than one
gram of Bacillus anthracis spores were released during this accident. For a detailed
report, see http://www.anthrax.osd.mil/documents/library/sverdlovsk.pdf
Bacteria reproduction (just a few words, more later)
Bacteria reproduce by binary fission. (How do eukaryotic organisms reproduce?)
Binary fission results in clones of cells (genetically identical).
This process can take as little as 20 minutes for E. coli under good
conditions.
Bacteria are cultivated in the laboratory on materials called culture media.
A culture medium is a solution of nutrients that encourage the growth of
microorganisms
What nutrients might be contained in
culture media?
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Two types of media exist:
Broth – liquid form
Gel form – contains agar
Agar is a solidifying agent
Growth media
1. Many nutrient media are general (nonselective), i.e., they support the growth
of many types of bacteria.
…but sometimes, one might wish to only grow certain species of bacteria.
2. Selective media contains compounds that inhibit the growth of some bacteria,
while promoting the growth of others.
E. coli growing on eosin
methylene blue agar.
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…but some bacteria are especially difficult to grow – fastidious bacteria.
3. Special nutrients are added to the medium to support the growth of fastidious
bacteria – enriched medium.
e.g. blood agar – general medium supplemented with red blood
cells.
Encourages the growth of streptococci (Streptococcus
spp.)
Hemolysis
4. Differential media
Contains compounds that allow one to visually differentiate between
bacteria.
e.g. E. coli vs. coliform bacteria.
Both types of bacteria live in the guts of warm-blooded
animals.
Diagnostic ability of the media is based on the activity of
two enzymes:
galactosidase – blue color when it breaks down
galactose.
glucuronidase – purple color when it breaks down
glucuronide.
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E. coli (purple)
Coliform (blue)
Prokaryotic diversity and spectrum
The spectrum of prokaryotes
How well do we understand the diversity and function of prokaryotes?
The vast majority remain unknown.
How many do you think have been identified?
Photosynthetic bacteria (bacteria, not archaea)
How does photosynthesis work and is it an advantage to the bacteria itself? Others?
Photosynthesizers are autotrophic – they synthesize their own food (fix carbon) from
CO2, and when they die, release it for other organisms (heterotrophs) to use.
Autotrophs, such as photosynthetic bacteria, are considered primary producers and are
important as the bottom members of the food chain.
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Cyanobacteria contain green pigments and often become dense in lakes, oceans and
swimming pools during cyanobacterial blooms.
Satellite photo of a cyanobacteria plume in western Lake Erie, 2003 (left).
Glass of lake water containing Microcystis spp. (right).
Photosynthetic bacteria are some of the most independent organisms on Earth. Why?
They can take carbon (from CO2) and nitrogen (from N2) in the air we breathe and
convert it to structural C and N.
Why are C and N important for bacteria?
…also usable by other organisms when the cyanobacteria die.
This unique behavior allows them to inhabit harsh environments.
Endolithic cyanobacteria.
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Archaea
Look like bacteria, but are actually not bacteria in the traditional sense.
1. Archaeal cell walls do not contain peptidoglycan
2. Archaeal cell membranes contain unusual lipids.
But more interesting…Archaea tend to live in harsh environments –
extremophiles
1. An example is the thermoacidophiles, which live under acidic and hot conditions.
Sulfolobus acidocaldarius
85o C, pH 1.0 (this is the pH of sulfuric acid)
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2. The most heat resistant Archaea known is Pyrolobus fumarii.
These live near in thermal vents (underwater volcanoes) in the ocean bottom.
Distribution of thermal vents in the world’s oceans.
Grow well at temperatures up to 113o C, but anything below 90o C is too cold.
What makes these Archaea so heat resistant?
The key to thermal stability is keeping proteins and membranes operating
efficiently.
Waxy substances present in membranes and a high proportion of sulfurcontaining proteins help the Archaea to resist the heat.
Certain amino acids contain sulfur and these sulfur atoms make strong
bonds that keep the proteins from denaturing.
Methionine
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3. Methanogens are a specific type of Archaea.
They produce ????
Live in anaerobic environments…remember when we talked about landfills?
Methanogens are autotrophs (they don’t use organic carbon). Instead they use
hydrogen (H2) and CO2 to gain energy and produce CH4.
4. Another example of Archaea is the extreme halophiles, which live under high salt
concentrations such as those in the Great Salt Lake (UT).
Many extreme halophiles actually require high concentrations of salt to grow and
are not viable in low-salt medium.
At least ~9% NaCl with an optimum between 12-23% NaCl
Maximum at 32% NaCl.
What environments will facilitate such a high salt concentration?
Freshwater lake = 0.05%
Ocean = 3.5%
What conditions will achieve 9% salinity?
Hint: the Dead Sea is ~15% salinity
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Images of the Dead Sea
Why does high salinity represent such an extreme environment?
Water will move to equilibrate a chemical gradient.
In a high salt environment, osmotic forces will pull water out of the cell,
dehydrating it if nothing is done to prevent it.
…then how do extreme halophiles adapt to such harsh conditions?
Extreme halophiles accumulate inorganic ions inside the cell at
concentrations that match the salt concentration outside the cell.
Sodium, calcium, potassium.
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In Halobacterium salinarum, a pump in the cell membrane transports high
amounts of potassium inside the cell so that the concentrations of potassium is
inside and outside of the cell are equal. In this case, potassium serves as the
compatible solute.
Outside
K+
What’s this?
Inside
What material is this pump made from?
With these adaptations, the extreme halophiles have evolved to live in
environments where most other organisms die. How does this benefit the
halophile?
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