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BIO 208 Unit 2 – Microbial Growth and Control
Unit Two – Microbial Growth and Control
In Lecture 7 we will be considering how microbes grow, what they need to be able to grow, and
how they get what they need.
We will be reviewing several figures from your text. It may be helpful to have the text with you
and open to Ch. 6 so you can mark those figures.
I. Microbial Growth (Chapter 6)
A. Basics
“Growth” – increase in number of cells, increase in size of population.
1. Bacterial Division (Fig. 6.12, 6.14)
 binary fission
2. Generation Time = time required for a cell to divide (or a population to double).
 varies greatly with species
 cell number = 2n, n = # of divisions (or generations)
Ex. start with 5 cells and they each divide 9 times
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BIO 208 Unit 2 – Microbial Growth and Control
B. Equation for Cell Growth
NUTRIENTS + INFORMATION + ENERGY 
POLYMERS  MACROMOLECULES  NEW CELL
 source of nutrients  source of information –
 source of energy 1. Energy (E):
a. E from chemicals –
1) organic (-C-C-) chemicals - chemo organo troph
Ex. C6H12O6 + 6O2 6CO2 + 6H2O
2) inorganic chemicals – chemo litho troph
Ex. 4FeS2 + 15O2 + 14H2O  4Fe(OH)3 + 8H2SO4
Thiobacillus ferrooxidans
b. E from light -
Ex. Chlamydomonas nivalis
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BIO 208 Unit 2 – Microbial Growth and Control
2. Nutrients
a. Macronutrients: need in large quantities: Carbon, Hydrogen, Nitrogen, Oxygen,
Phosphorous, Sulfur (CHNOPS)
1). Carbon
sources:
 CO2 – auto trophs
 organic (-C-C-) chemicals – hetero trophs
*note that organic chemicals can serve as both energy and carbon sources
2) Hydrogen source –
3) Nitrogen
sources:
 amino groups (-NH2)
 ammonia (NH3)
 nitrate (NO3-)
 atmospheric nitrogen (N2)
4) Oxygen source –
5) Phosphorous
sources:
 phosphate (PO43-)
 organic molecules
6) Sulfur
sources:
 sulfate (SO42-)
 sulfide (S2-)
b. Micronutrients: K, Mg, Ca, Fe
 required in small but sig. amts.
 act as cofactors for many enzymes (enz)
 are important in cell structures
c. Trace elements: Co, Zn, Cu
 required by a small number of enz.
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BIO 208 Unit 2 – Microbial Growth and Control
C. Growth of Bacteria in Culture
Assume cultivation in:
 liquid medium
 a closed system
affected by:
o nutrient shortages
o waste accumulation
Phases of Growth -4(Fig. 6.15)
1. lag phase -
2. log phase -
3. stationary phase -
4. death phase -
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BIO 208 Unit 2 – Microbial Growth and Control
D Growth of Bacteria “in the Wild”
 Affected by:
1.Biofilm - A multilayered bacterial population embedded in a polysaccharide matrix and
attached to some surface.
Also see Fig. 6.5
2. Cell to cell communication a. quorum sensing –
AHL (aka HSL) – acylhomoserine lactone
b. regulating cellular processes
Biofilm lifestyle allows these structurally simple yet physiologically diverse microbes to
coexist in an environment --and the activity and growth of the community exceeds what is
possible for an individual.
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BIO 208 Unit 2 – Microbial Growth and Control
E. Measurement of Microbial Growth – most of this will be covered in lab, so we won’t cover
much in lecture
1. Cell Numbers
a. Direct counts


advantages - quick, cheap, cell size & morph
disadvantages - pop. needs to be large, can’t tell living from dead
*b. Plate counts
Diluted sample dispersed over solid agar surface - each microbe grows into 1 colony
Original # viable cells in a sample can be back calculated
Ex. dilute 1 ml sample into 100 ml water (a 1/100 dilution)  plate incubate
count 150 colonies
# colonies
x
inverse of dilution
= # original per ml
150
x
100
= 15,000 CFU/ml


advantages - simple, sensitive
disadvantages - have to know how to culture the microbe
Modification - counts from membrane filters
2. Cell Mass
a. turbidity – cells interfere with transmission of light, looks cloudy. The lower the light
transmission, the greater the mass of cells.
b. metabolic activity – e.g., ATPase activity.
c. dry weight
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BIO 208 Unit 2 – Microbial Growth and Control
In Lecture 8 we will look at the effects of the physical environment on the growth of microbes.
F. What Affects Microbial Growth?
1. Nutrients
a. presence in the environment
b. ability to transport across plasma membrane
2. Physical environment: water, pH, temp., oxygen – for each physical parameter, microbes
will display a range of tolerances (too low, optimum, too high), which will vary from
species to species.
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BIO 208 Unit 2 – Microbial Growth and Control
a. Water
1) effects of water imbalance
 hypotonic  hypertonic -
2) adapted to hypertonic environments
 Halotolerant –
habitat Ex. Staphylococcus aureus

Facultative halophile habitat -

Extreme halophile habitat Ex. Halobacterium (Archaea)
b. pH
 internal pH of cell is
protozoa & most bact. prefer
fungi & algae prefer
1) effects of being in the wrong pH
 disrupts plasma membrane
 inhibits transport
 inactivates enz
2). pH classifications

Neutrophile: 5.5 - 8
most microbes

Acidophile: 0 - 5.5
some molds, some bact.
habitats - ore mines, bogs, stomach
Ex. Helicobacter pylori
uses for humans – enzymes to function in low pH industrial environments

Alkalophile: 8.5 - 11.5
habitats - soda lakes
uses – enzymes to perform in alkaline laundry detergent
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BIO 208 Unit 2 – Microbial Growth and Control
c. Temperature
1) effects of being at the wrong temperature
 influences membrane fluidity
 affects enzyme function
o minimum –
o optimum –
o maximum -
2) temperature classifications

Psychrophile –
optimum Ex. Chlamydomonas nivalis

Psychrotroph –
optimum Ex. Listeria monocytogenes

Mesophile optimum Ex.

Thermophile –
optimum Ex. Thermoplasma (Archaea)

Hyperthermophile –
optimum Ex. Pyrolobus fumarii
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BIO 208 Unit 2 – Microbial Growth and Control
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d. Oxygen
1) evolution of Earth’s atmosphere
4.8 billion
2.25 billion
2 billion
Today
no O2 (reducing = anoxic)
oxygenic photosynthesis  O2
1% O2
21% O2
2) why is oxygen bad?
O2 - accepts e- and becomes reduced to H2O
O2 + e-  O2O2- + e- + 2H+  H2O2
H2O2 + e- + H+  H2O + OH
pull e- off of other molecules (DNA, plasma membranes)
3) what protects O2 respiring cells from bad effects of O2? superoxide dismutase (SOD)
2O2 + 2H+
O2 + H2O2
catalase (cat)
2H2O2
2H2O + O2
4) Oxygen tolerance classification –we will not go over in lecture but you will need to
know these terms
 Obligate anaerobe – does not require O2 for growth, will not survive exposure
to oxygen, has neither catalase (cat) nor superoxide dismutase (SOD).
 Aerotolerant anaerobe – does not require O2 for growth; will survive exposure,
has SOD only.
 Microaerophile – needs a little O2 for growth, but less than amount present in air.
 Facultative anaerobe – can grow with or without O2, has both cat and SOD.
 Aerobe – requires O2 for growth, has both cat and SOD.
Assignments
Ch. 6 – Read entire chapter
Review – 1- 3, 5-8
MC – all
CT – 1, 4
CA – all
BIO 208 Unit 2 – Microbial Growth and Control
In Lecture 9 we will explore control of microbial growth, including the use of antibiotics.
II. Control of Microbial Growth (Chapter 7 and pp. 554-558)
A. Terminology
1. sterilization/sterilize - destroy all viable cells, spores, viruses
2. disinfection/disinfectant - kill pathogens on inanimate surfaces
3. antisepsis/antiseptic - kill pathogens on living tissue
4. de-germ – mechanical removal
5. sanitization/sanitize - lower # of pathogens to acceptable levels
B. How do we kill microbes?
1. Nonspecific – work against almost all microbes in the same way
a. Physical methods
b. Chemical methods
1) phenols - denature proteins, disrupt membranes
Ex.
2) halogens - oxidation of cellular material
Exs.
3) alcohols - denature proteins, dissolve lipid membranes Ex.
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BIO 208 Unit 2 – Microbial Growth and Control
2. Specific – specifically kill some types of microbes, others are left unharmed.
Antibiotics (pp. 554-558)
Antibiotic –natural substance produced by one microorganism that inhibits the growth of
another
a. How were antibiotics discovered?
Alexander Fleming (1928)
Penicillium notatum (Eukarya – fungi)
b. How do antibiotics work?
Bactericidal - kill
Bactriostatic – inhibit
*Selective toxicity – no harm to host
**
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BIO 208 Unit 2 – Microbial Growth and Control
c. Cellular Target Sites of Antibiotics – 4 - Important
Fig. 20.2
1) cell wall synthesis
 prevent synthesis of new peptidoglycan
 work only on growing cells
 selective - how?  least toxic
Exs. penicillin, methacilllin, cephalosporin
2) plasma membrane integrity and/or function
 alter permeability
 selective - how? Eukarya –
Bacteria –
Exs. polymyxin B, nystatin
3) nucleic acid synthesis
 interfere with enzymes gyrase and polymerase
 selective - how? Ex. rifampin, quinolones like ciprofloxacin
4) protein synthesis
 target 70S ribosome
 greater toxicity – why?
Exs. tetracycline, chloramphenicol, erythromycin
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BIO 208 Unit 2 – Microbial Growth and Control
3. Antibiotic Resistance
a. history
1940s 1969 1980s 1980s 1990s and on -
b. how did we get in to this predicament?
amount manufactured number of prescriptions -
Antibiotic resistance can develop extremely rapidly - even in a patient receiving
treatment in a hospital
Notes from clinical case:
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BIO 208 Unit 2 – Microbial Growth and Control
15
Assignment
Read Chapter 7 and pages 554-558
Review 1, 2, 5, 7- 9
MC 1,9,10
CT 1-3
CA 3
FYI
Q. How do you know if an antibiotic is going to work?
A. You should feel better within 24-48 hours of starting antibiotic treatment.
If you do not feel better:
1. You have a viral infection and not a bacterial infection OR
2. You have a bacterial infection but the antibiotic prescribed is not effective against the bacteria you
have OR
3. You have a bacterial infection but bacteria are resistant to the antibiotic that was prescribed
Then you should contact your doctor and let her/him know that the antibiotic is not working.
What can you do to reduce the likelihood that bacteria will become antibiotic resistant?
1. Take the correct dosage of your antibiotic and always take the entire prescription. If you don’t,
infectious bacteria that have not yet been killed off may survive, reproduce, and cause a more severe
relapse, one that may not be treatable.
2. Ask the doctor to tailor the prescription to fit your schedule so that you don’t miss a dose.
3. Ask if you can take the antibiotic for the shortest amount of time possible.
4. Ask the doctor to prescribe a narrow-spectrum antibiotic, one that works specifically against a few
strains of bacteria, rather than a broad-spectrum antibiotic that targets more strains. The more bacteria
exposed to antibiotics, the greater the chance that a strain will develop antibiotic resistance.
5. Use the antibiotic only for the prescribed illness. Never take antibiotics that you have left over
from a previous illness. Never take antibiotics that were prescribed for someone else (not even
your mom).
BIO 208 Unit 2 – Microbial Growth and Control
Review - Important Concepts for Lectures over Metabolism
I know that you have had an introduction to the basics of metabolism in BIO 110. The
metabolism you learned was focused on the types of metabolism that animal and plant cells carry
out --aerobic respiration. The microorganisms are tremendously more diverse and complex in
metabolic patterns than are Eukarya and I want to spend our time emphasizing what microbes can
do, not just covering what you have already had in other courses.
My focus in metabolism is that as microbes create and store and use energy for transporting
nutrients, making their cellular components, growing, and moving in their environment.
Importantly, as a consequence of their metabolism, they can profoundly change that environment!
So, if you do not remember the basics of metabolism you will need to review. The following
pages should serve as a reminder. If it doesn’t all come back to you then read Chapter 5 in the
text. If you have not had chemistry you will also need to read Chapter 2.
Review of nutritional patterns:
Source of energy
Source of carbon
Chemicals = chemotroph
make it (CO2) = autotrophs
Organic = chemoorgano
eat it (organic molecules (-C-C-C-)) = heterotrophs
inorganic = chemolitho – you were not exposed to this concept in BIO 110
Light = phototroph
Most common combinations of Energy gaining strategy plus Carbon gaining strategy – this
terminology was not used in BIO 110, but you were exposed to the concepts behind the terms
“chemoorgano heterotroph” and “photo autotroph”)
Chemoorgano heterotrophs
Chemolitho autotrophs
Photo autotrophs
Photo heterotrophs
You should also know definitions of metabolism, anabolism, and catabolism
You should know what ATP is and does. ATP (Adenosine Tri Phosphate) connects reactions
that produce energy with reactions that use energy. It is made to store energy for later use – it is
the energy “currency” for the cell.
During catabolism – ATP  ADP + Pi + energy
During anabolism – ADP + Pi + energy  ATP
(Pi = inorganic phosphate; ADP stands for adenosine diphosphate)
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BIO 208 Unit 2 – Microbial Growth and Control
ATP can be formed in 3 ways:
1. by substrate level phosphorylation – the simplest, oldest, and least-evolved way to make ATP a high energy phosphate is removed from a substrate and is added to ADP to make ATP.
Ex. C-C-C~P + ADP  C-C-C + ATP
2. by oxidative phosphorylation, aka electron transport phosphorylation – electrons are transferred
from organic compounds to electron carrier molecules and then to final electron acceptor
molecules. The transfer of electrons releases energy that is used to convert ADP  ATP.
3. by photophosporylation – occurs in photosynthetic cells only. Light energy is converted to ATP.
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BIO 208 Unit 2 – Microbial Growth and Control
18
You should understand basics of energy production
All molecules have energy that is associated with the electrons that form bonds between atoms.
The electrons can move around in a cell from molecule to molecule, transferring energy as they
move. The molecules are changed as they either gain or lose electrons.
Oxidation – Reduction Reactions (redox)
In biological systems:
H

Hydrogen
atom
H+
+
proton
eelectron
Oxidation = a loss of an e- (and in biological systems, usually a loss of the H+ as well)
Ex.
H2O - 2e- - 2H+

H2 + ½O2
+
Ex.
NO2 - 2e - 2H

NO3Reduction = a gain of an e- (and in biological systems, usually a gain of the H+ as well)
Ex.
½O2 + 2e- + 2H+

H2O
Ex.
NO3-+ 2e- + 2H+

NO2-+ H2O
Remember as LEO the lion says GER (Lose of Electron is Oxidation, Gain of Electron is Reduction)
Redox reactions are always balanced.
e- donor = reducing agent – causes its partner molecule to become reduced, to gain ee- cannot exist free in a cell, it must go somewhere. So if are e- removed from one molecule
they are added to another.
e- acceptor = oxidizing agent – causes its partner molecule to become oxidized, to lose e-
In biological molecules it is usually the entire H atom (electron and proton) that is lost or gained,
but not always. Sometimes the electrons are separated from the proton and only the electrons are
lost or gained; and sometimes it may be one H atom + 1 electron (from a second H atom) that are
lost or gained.
Ex.
C3H4O3
pyruvate
C3H6O3
lactic acid
oxidized
pair one
reduced
2e- + H+
NADH + H  NAD+
reduced
pair two
oxidized
NADH passes 2e- and 1 H+ to C3H4O3, as soon as C3H4O3 accepts the e-s and H+, it becomes
C3H6O3 and NADH + H becomes NAD+
BIO 208 Unit 2 – Microbial Growth and Control
In any pair of molecules you can distinguish which molecule is in the oxidized state (has lost an
e-) and which molecule is in the reduced state (has gained an e-):
Oxidized state
Reduced state:
Contains more oxygen atoms OR
fewer hydrogen atoms AND
therefore has fewer electrons and is
less negative or more positive
Contains fewer oxygen atoms OR
more hydrogen atoms AND
therefore has more electrons and is
more negative or less positive
Example pairs:
Glucose
C6H12O6
Pyruvate
C3H4O3
NAD+
NADH
Sulfate
SO4
Hydrogen sulfide
H2S
End Review
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BIO 208 Unit 2 – Microbial Growth and Control
In Lectures 10-13 we will explore how microbes create, store, and use energy. Through these
processes microbes can profoundly change their environment.
While we will be covering the topics outlined in Chapter 5 of your text, we will be doing it in a
very different manner from how it is presented in the text. Please be prepared to take careful notes
in class.
III. Patterns of Metabolism in the Microbial World (a.k.a. how do microbes make a living –
and why should we care?)
A. The Basics: quick review of basics from BIO 110
Metabolism = sum of all chem. rxns occurring within a living organism
All cells need a source of energy for:
Catabolism- breaking bonds in molecules –
Ex. glucose to carbon dioxide and water
Anabolism – creating bonds -
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BIO 208 Unit 2 – Microbial Growth and Control
21
B. Patterns of Energy Production Among Living Organisms
1. Basic information
Patterns among Eukarya:
 alcohol fermentation (yeast)
 lactic acid fermentation (muscle cells, neutrophils)
 aerobic respiration (mold, protozoa, animals)
 oxygenic photosynthesis (algae, plants)
Bacteria and Archeae do all the above plus:
 anaerobic respiration: uses inorganic molecules other than 02 as a final electron
acceptor
 lithotrophy: use of inorganic substances as sources of energy
 photoheterotrophy: use of organic compounds as a carbon source during bacterial
photosynthesis
 anoxygenic photosynthesis: photophosphorylation in the absence of O2
 methanogenesis: uses H2 as an energy source and produces methane
 light-driven nonphotosynthetic photophosphorylation: converts light energy into chemical
energy
We will explore only a few of these
There are 2 initial sources of usable energy:
1. sunlight –
2. chemical bonds of molecules Heterotrophs - energy is created by breaking bonds in a molecule and harvesting the
electrons released from the H atoms in:
Organic molecules Inorganic molecules –
The more electrons a molecule has, the more energy it is capable of releasing. This initial
molecule is called the ____________________________________________.
Ex. glucose (C6H12O6) has a lot of H atoms (12) and therefore a lot of electrons, the oxidation
of glucose will release a lot of electrons. **Glucose is a high energy electron donor.
BIO 208 Unit 2 – Microbial Growth and Control
22
The electrons released from a molecule such as glucose have to go somewhere - they get passed from
the initial donor of released electrons (electron donor) to intermediate electron carrier molecules.
Example intermediate electron carrier - NAD (Nicotinamide Adenine Dinucleotide) -
accepts 2e- (and 1 proton) and becomes reduced to
+
NAD
NADH
PROBLEM: NADH can’t accept anymore electrons. If energy production is going to continue,
the NADH must be converted back to NAD+, which means NADH must transfer the electrons
somewhere.
We will examine some of the solutions to this problem by seeing what chemoorgano
heterotrophs do during carbohydrate catabolism (beginning next page)
BIO 208 Unit 2 – Microbial Growth and Control
Many types of molecules can undergo catabolism to release energy:
Proteins  amino acids
Lipids  glycerol + fatty acids
Carbohydrates  sugars
2. One Example – generation of energy via carbohydrate catabolism – specifically the
carbohydrate glucose – by a chemoorgano heterotroph
 watch movement of e- and regeneration of NAD+
 watch for formation of ATP (energy storage molecule)
Glycolysis (via Embden-Meyerhof Parnas (EMP) pathway) occurs in the cytoplasm – the initial
electron donor is glucose
Glucose + 2 ATP
C6H12O6
initial e- donor
2 Pyruvate + 4 ATP
2 C3H4O3
net ATP production =
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BIO 208 Unit 2 – Microbial Growth and Control
At end of glycolysis Need NAD+; NADH needs to get rid of e1. First strategy:
Fermentation –pass e- from NADH to an organic molecule, NADH becomes NAD+ fermentation reactions occur in the cytoplasm of the cell.
organic molecule
(pyruvate)
2 C3H4O3
2 C3H6O3
(lactic acid)
End of fermentation -
Inefficient process–
2 NADH + 2H+
2 NAD+
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BIO 208 Unit 2 – Microbial Growth and Control
25
2. Alternative (to fermentation) strategy:
Respiration – pass e- from NADH (becomes NAD+) along a series of intermediate electron
carrier molecules, ultimately to a final (or terminal) electron acceptor molecule.
Occurs in 2 steps:
Step 1 – Tricarboxylic acid (TCA) cycle (also known as citric acid cycle or Krebs cycle) – occurs
in the cytoplasm - harvests the energy still within the bonds of pyruvate, but transfers even more eto NAD+ (so more NAD+ is converted to NADH). Doesn’t solve the shortage of NAD+ problem.
At end of TCA:
For each pyruvate (C3H4O3) 
since we get 2 pyruvate per glucose…
For each glucose 
(FAD is another intermediate electron carrier that functions so much like NAD that for the
purposes of this course we will consider them equal)
BIO 208 Unit 2 – Microbial Growth and Control
Step 2 – Electron Transport Chain – the soluble NADH and FADH2 carry e- from the cytoplasm
(where glycolysis took place) to the cytoplasmic membrane and pass them off to a series of
membrane associated proteins (when NADH passes off the e- it becomes NAD+). These proteins
function as intermediate electron acceptors, accepting e- and becoming reduced, then passing the
e- off to the next protein in the chain, becoming oxidized again, ending with the final or terminal
electron acceptor (which accepts the e- and becomes reduced).
This final electron acceptor may be oxygen –
Final e- acceptor (oxidized state)
accepts e- and becomes reduced to
aerobic respiration
1 molecule of C6H12O6 oxidized completely to CO2 coupled to reduction of oxygen to water
(aerobic respiration) can yield up to a max of 38 ATP.
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BIO 208 Unit 2 – Microbial Growth and Control
OR
The final electron acceptor is an inorganic molecule other than oxygen –
anaerobic respiration
Examples of final e- acceptors for anaerobic respiration:
Final e- acceptor (oxidized state)
becomes reduced to:
Fe3+
ferric iron
Fe2+
Iron respiration
NO3nitrate
NO2-, N2O, N2
Nitrate respiration
SO42sulfate
HS-
Sulfate respiration
CO2
carbon dioxide
CH4
methane
Methanogenesis
S0
sulfur
HS-
Sulfur respiration
Yield of ATP by cells undergoing anaerobic respiration is greater than the 2 ATP produced by
glycolysis (and maintained in fermentation), but fewer than the 38 ATP produced by
aerobic respiration.
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BIO 208 Unit 2 – Microbial Growth and Control
C. Some Exciting Implications of Microbial Activity:
1. Metabolism of the Human Intestinal Microbial Community
a. Where does your gut microbial community come from?
At birth
Progression of your gut community if you were a breast-fed baby
Day 1 - First colonizer was Escherichia coli
 facultative anaerobe
 chemoorgano heterotroph (gets both C and E from organic molecules)
Where did E. coli come from?
What organic compound does E. coli use as a C and E source?
How does E. coli get C and E from
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BIO 208 Unit 2 – Microbial Growth and Control
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BIO 208 Unit 2 – Microbial Growth and Control
Day 3 – 2 more bacteria joined you gut community
Enterococcus
obligate but aerotolerant anaerobes
chemoorgano heterotrophs
obligate fermentative metabolism
Bifidobacterium
Lactic acid bacteria (named from their final fermentation end product)
e1 Glucose
NAD+
2 Pyruvate
eNADH
2 Lactic acid
NAD+
glycolysis
2 ATP
fermentation
Soon after added:
Enterobacter- facultative anaerobe
Clostridium - obligate (aerotolerant) anaerobe
Butanediol fermenters
e1 Glucose
NAD+
2 Pyruvate
eNADH
glycolysis
2 ATP
fermentation
ethanol
acetic acid
Acetoin
lactic acid
CO2 + H2
succinic acid
2,3-Butanediol + CO2
NAD+
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BIO 208 Unit 2 – Microbial Growth and Control
Microbial gut community from 1 week to ~ 3.5 months if you were a breast-fed infant:
E. coli
Enterococcus
Bifidobacterium
Enterobacter
Clostridium
3.5 months to weaning
99% Bifidobacterium infantis –
When meat was introduced to your diet:
Gram-negative anaerobes:
Bifidobacterium
Clostridium
Fusobacterium
Eubacterium
Ruminococcus
Peptococcus
Peptostreptococcus
Bacteroides – 30% of total adult community
Bacteroides
obligate anaerobe
extremely oxygen sensitive
fermentative metabolism
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BIO 208 Unit 2 – Microbial Growth and Control
32
b. What are the benefits of a stable, mature gut microbial community?
1) Nutritional
2) Prevents colonization by pathogens
2) and 3) in supplemental info at the end of this unit, p.
3) Trains the immune system
38. We do not have time to cover them in this class 
1) Nutrition
a) Gut microbe metabolism converts complex polysaccharides to volatile fatty acids (vfa) – good
Bacteroides is the key player in this process
Host and dietary carbohydrates – complex carbs, starch, cellulose
saccharolases
hydrolases
fermentation
by gut community
short-chain volatile fatty acids*
acetic acid
butyric acid
propionic acid
reabsorbed through the large intestine
used by you as an energy source
provide a significant proportion of your daily energy requirement (540 kcal)
* These products in brown are good for your health metabolic by-products
BIO 208 Unit 2 – Microbial Growth and Control
b) Gut microbes can metabolize dietary fats too – not so good
Bacteroides is the key player in this process also
Dietary fats
Liver
Bile acids
Absorbed by small intestine
----------------------------------------------------------------------------------------------------
If fats and bile acids are not reabsorbed by small intestine but make it to colon
deconjugated
deoxycholic acid
lithocholic acid
intermediate products
Bacteroides thetaiotomicron
ethyl ester
*These products in purple are mutagenic, carcinogenic products; they can induce cancer –
bad for your health products
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BIO 208 Unit 2 – Microbial Growth and Control
34
c) Gut microbes can also metabolize dietary protein – can be bad
By the combined activity of the colonic microbial community
Dietary protein
Peptides
Amino acids
Absorbed by small intestine
---------------------------------------------------------------------------------------------------If peptides are not reabsorbed by small intestine but make it to colon
Amino acids
R
+
H3N – C – C – OH–O
deamination
decarboxylation
aromatic
amino acids
sulfur
amino acids
reduced to
phenolic
compounds
SO4
H2S gas
anaerobic respiration by
sulfate-reducing bacteria
fermentation
by many microbes
ammonia
H2
branched chain
fatty acids
volatile
fatty acids
CO2
reduced to
CH4 gas
anaerobic respiration by Methanogens (which are Archaea)
brown and purple as explained before. red are final electron acceptors in anaerobic respiration
BIO 208 Unit 2 – Microbial Growth and Control
Autotrophy - Background
All microbes need:
a source of energy (electrons = ATP)
a source of C to build macromolecules (-C-C-)
Heterotrophs get C to make –C-C- by recycling the C contained in organic molecules. (all
heterotrophs get energy [e-, ATP] to make –C-C- from breaking chemical bonds)
**Autotrophs – get C to make –C-C- from CO2
But there are 2 sources of energy that can be used to turn CO2  -C-C- and the source defines 2
groups of autotrophs:
1. Photo autotrophs
energy from sunlight (C from CO2) (I’ll leave this for Botany, but lots of microbes do this too)
2. Chemolitho autotrophs
energy is generated from inorganic chemicals (C from CO2)
Many different inorganic chemicals can serve as electron donors to provide the energy for
microorganisms via aerobic respiration (notice the presence of O2 as electron acceptor in all the
equations following – therefore all chemolitho autotrophs are obligate aerobes):
a. Hydrogen gas as an electron donor
e- donor
H2
+
e- acceptor
1/2 O2

donor reduced to
H2O
Hydrogen bacteria
Ex. Alcaligenes faecalis (from Lab 8)
b. Sulfur compounds as electron donors
e- donor
2S + 2H2O
+
e- acceptor
3O2

donor reduced to
2H2SO4
Ex. Sulfur bacteria like Thiomargareta namibiensis or the bacteria that form snot-tites in
caves.
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BIO 208 Unit 2 – Microbial Growth and Control
c. Nitrogen compounds as electron donors
Nitrifying Bacteria
2 groups of Nitrifying Bacteria:
e- donor
1) 2NH3
+
e- acceptor
3O2

donor reduced to
NO2 + 2H2O + 2H+
e- acceptor
2O2

donor reduced to
2NO3 + 2H+
Ex. Nitrosomonas
e- donor
2) 2NO2
+
Ex. Nitrobacter
d. iron as an electron donor
e- donor
Fe2+
+
e- acceptor
1/2 O2 + 2H+ 
donor reduced to
Fe3+ + H2O
Ex. Iron bacteria like Ferroplasma
Two scenarios where chemolitho autotrophs are very important:
Metabolism of Wastewater Treatment
How do we go from toilet water to treated water? (stay tuned, we will discuss this in Unit 4  )
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BIO 208 Unit 2 – Microbial Growth and Control
2. Metabolism of the Deep - What? No photosynthesis???
Deep sea hydrothermal vents – provide all the necessary inorganic chemicals
 Black smokers – vent hydrogen, sulfur, iron (electron donors for energy), and CO2 (for
carbon) from the Earth’s core
 Sea water contains dissolved oxygen (electron acceptor for aerobic respiration)
 Everything that is needed for chemolitho autotrophs to grow.
Chemolitho autotrophic metabolism turns CO2 and inorganic chemicals into bacterial biomass,
with excess energy to spare!
Animals (chemoorgano heterotrophs)
Giant tube worms
with endosymbiotic chemolitho autotrophs
Giant mussels
Brittle stars
Limpets
Worms
Crabs
Vent fish
Sharks
Assignment
Read Chapter 5
Review 4, 5b,c, 6,7,9
MC 1,4,6
CT 1,3,5
This ends the lecture material for Test 2.
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BIO 208 Unit 2 – Microbial Growth and Control
Supplemental Information – If we have any extra time I may cover some of these topics
Aside - How can 2 people eat the same foods, 1 person gains weight and the other stays lean?
colonization of gut by microbes increases
glucose uptake in the intestine
↓
microbial fermentation
↓
resulting in substantial elevations in serum
glucose and insulin
results in production of short-chain fatty acids
both
stimulate lipogenesis in the liver
↓
triglycerides into the circulation
↓
taken up by adipocytes (fat cells)
The composition and operation of your gut microbiota influences your energy balance.
Relatively high-efficiency gut microbial communities would promote energy storage (weight
gain), whereas lower efficiency communities would promote weight loss.
Small but long-term differences between energy intake and expenditure can, in principle, produce
major changes in body composition.
Ex. if energy intake exceeds energy expenditure by +12 kcal/day, >1 lb of fat could be gained in a
year; this is the average annual weight gain experienced by Americans between ages 25 and 55.
1. b. Metabolism of Human Intestinal Microbial Community continued from p. 32
2). Mature gut microbial community prevents colonization by pathogens – pathogens like
Salmonella, Shigella, Campylobacter, the pathogenic strains of E. coli, etc. that cause intestinal
disease.
a. competition for attachment sites – the gut epithelium is so densely colonized by normal
microbiota, nowhere for pathogens to attach.
b. competition for nutrients – if pathogens do attach, they have to fight normal microbiota for a
share of nutrients
c. antimicrobial chemicals – and then the normal microbiota secrete antimicrobial chemicals
that kill pathogens.
Ex. E. coli – produces a chemical called colicin
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BIO 208 Unit 2 – Microbial Growth and Control
3) Mature gut microbial community trains the immune system
The primary barrier between the outside world and you is a single layer (1 cell thick) of gut
epithelium. This barrier is tight, but not impenetrable.
Microvilli – where adsorption takes place
Epithelium
Submucosa
Muscle
The surface of the intestinal epithelium is protected by your immune system – the antibody
IgA, and white blood cells called T and B lymphocytes, and phagocytic macrophages.
The gut epithelium tests the contents of the
gut lumen (open cavity) and can directly sense
the antigens of microbes using “pattern
recognition receptors” (PRRs) – the
epithelium recognizes conserved structures of
bacteria and viruses and then alerts the host to
the potential of infection.
Normal microbiota of the gut and dietary
antigens in food are tolerated (should not
stimulate an immune response).
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BIO 208 Unit 2 – Microbial Growth and Control
40
1. continued.
c. How does what you eat influence your gut community and in turn your health? –So very cool!!
1). Over stimulation of microbial growth and metabolism
Ex. Lactose intolerance
In babies, the enzyme human lactase is secreted by the small intestine and will break milk
lactose into glucose and galactose. By the age of weaning, humans stop secreting human
lactase.
After the age of weaning if lactose is consumed in dairy, it will pass undigested to the large
intestine. In the large intestine E. coli will secrete the enzyme -galactosidase, which will
now break lactose in to glucose and galactose. The E. coli will use the glucose as a carbon
and energy source to support rapid population growth.
As a result of their fermentative metabolism on this bounty of glucose, E. coli will produce a
lot of 3 carbon fermentation end products, and a lot of CO2 gas. The 3 C end products
increase the osmotic pressure in the large intestine, which combined with the CO2 will
results in the symptoms of bloating and diarrhea
Adult lactose intolerance is the normal state for humans. People who as adults can tolerate
lactose had ancestors that acquired a mutation that allows them to continue to secrete human
lactase in to adulthood.
2). Diet can upset immune system training
The gut immune system has the challenge of responding to disease-causing microbes but not
responding to food antigens and the normal gut microbial community.
In developed countries like the U.S., this discriminatory ability appears to be breaking down.
High-fat, high-sugar, low-fiber diet changes gut community composition, which upsets
immune training resulting in allergies and/or chronic inflammation
Ex.1. Allergies
Children w/ allergies have a higher chance of having bad Clostridium difficile and
Staphylococcus aureus and lower prevalence of good Bacteroides and Bifidobacteria in their
gut.
Ex. 2. Chronic inflammation
Crohn's disease and ulcerative colitis (UC)
? breakdown in tolerance to Bacteroides initiates an autoimmune reaction?
Experimental txtt - whipworms
BIO 208 Unit 2 – Microbial Growth and Control
3). Diet can promote abnormal cell growth – i.e., cancer
Examples of suggested links between microbial metabolism and cancer:
1. High fat diet – go back and look at diagram of what happens to fat in the gut
 conjugated secondary bile acids – are carcinogens
2. High protein diet – go back and look at protein diagram again
 protein fermentations may be sources of systemic toxins
 Heterocyclic amines (HCA) are converted into carcinogens.
 phenolics from aromatic amino acids may enhance production of mutagens.
 reduced sulfur compounds (like H2S) may be toxic to the colonic epithelium.
3. Alcohol consumption
 acetaldehyde toxicity
Look again at diagram of lactose utilization by E. coli. See where ethanol is produced by
mixed acid fermentation? An intermediate molecule in the pathway Acetyl CoA  ethanol is
a toxin called acetaldehyde
Acetyl CoA  acetaldehyde  ethanol
Part of this pathway also runs in the reverse direction:
oxidation
mitochondria in the liver cells
ethanol

acetaldehyde (bad)

alcohol dehydrogenase
aldehyde dehydrogenase
acetic acid (good)
If there is a lot of ethanol being converted to acetaldehyde, the hepatic mitochondrial enzyme
aldehyde dehydrogenase cannot keep up, and acetaldehyde levels build in the liver and blood.
This causes symptoms of hangover in the short term, in the long term the acetaldehyde causes
mutations in DNA that can lead to cancer.
Prebiotics are complex carbohydrates that you cannot digest, such as fructo oligosaccharides
(FOS). They pass to the intestines where they stimulate the growth and activity of intestinal
bacteria that secrete beneficial metabolic end products. Fruits and vegetables contain
oligosaccharides; bananas and artichokes are especially high.
Probiotics are living bacteria from genera that produce favorable end products, such as
Bifidobacterium and Lactobacillus.
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BIO 208 Unit 2 – Microbial Growth and Control
Review – Metabolism Basics
All cells need:
1. A source of carbon for making cellular molecules.
There are two strategies for obtaining carbon:
a. recycle the C already present in some organic (-C-C-) molecule
b. use CO2 from the atmosphere
2. A source of energy for performing all cellular work (building molecules, transport across the
plasma membrane, locomotion, etc.)
Energy is created by harvesting the electrons (e-) present in:
a. Organic
molecules.
(specifically the e- in the H
atoms in the molecules)
Hydrogen – showing the
proton and electron
like a sugar or an
amino acid
OR
b. Inorganic
molecules.
e- in molecules
like
ammonia
hydrogen sulfide
The more electrons a molecule has, the more energy the molecule is capable of yielding – so
look at glucose compared to hydrogen sulfide – which molecule should yield the most energy?
(glucose has 12 H vs. 2 in H2S)
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BIO 208 Unit 2 – Microbial Growth and Control
The electrons that are
released when bonds are
broken have to go
somewhere, so they get
passed from the donor (the
molecule that you started
with that had all the
electrons) to intermediate
electron carriers.
NAD+ is a soluble carrier
present in the cytoplasm. It
is lacking 1 electron (1 H)
and so it can accept 1
electron (1 H). As it
accepts the electron, it is
reduced to NADH.
Oxidized state
fewer H, fewer emore positive (NAD+)
Reduced state
more H, more emore negative (NADH)
NAD+ is in limiting quantities in the cell and it must be converted back to NADH if energy
production is to continue.
There are 2 ways convert NADH back to NAD+:
1. NADH passes the electron to an organic
molecule like pyruvate – this process is
called fermentation - as NADH loses the
electron it becomes oxidized to NAD+
again. As pyruvate accepts the electron it
becomes reduced to acetic acid or to
ethanol, etc., which are excreted from the
cell, carrying waste electrons with them.
Acetic acid, ethanol, etc. still have
electrons, so potential energy is lost in the
fermentation strategy.
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BIO 208 Unit 2 – Microbial Growth and Control
2. NADH travels to the
cytoplasmic membrane
and passes the electron
off to the electron
transport chain. This
process is called
respiration.
electrochemical gradient - energy
Fig. 5.16
(NADH then becomes
NAD+ )
pH 8.5
The electrons are passed along the chain, generating two types of usable energy along the way –
electrochemical gradient and ATP - until they reach a final electron acceptor, an inorganic
molecule which can be:
a. oxygen (aerobic respiration)
As oxygen accepts e- it will
become reduced to H2O
OR
b. some other inorganic molecule
(anaerobic respiration)
like nitrate
or sulfate
becomes reduced to
nitrite (NO2)
becomes reduced to hydrogen
sulfide (H2S)
Note – fermentation is NOT anaerobic respiration. By definition respiration requires both an
electron transport chain and an inorganic terminal electron acceptor. Fermentation does not
employ an electron transport chain and the terminal electron acceptor is an organic molecule.
Fermentation takes place in the absence of oxygen, it can occur in anoxic (no oxygen but has
nitrate) and anaerobic (no oxygen, no nitrate) environments, but it is not respiration!
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BIO 208 Unit 2 – Microbial Growth and Control
Comparison of Respiration vs Fermentation in Chemoorganotrophs
Initial electron donor:
examples:
Intermediary electron
carrier(s):
Final electron acceptor
examples:
final electron acceptor
reduced to:
example organisms
Potential net ATP yield:
Respiration
organic molecule
carbohydrates, amino acids, lipids
NADH, FADH2, carriers in the
electron transport chain
inorganic molecule
O2
CO2, NO3, SO4
H2O
CH4, NO2, H2S
Mitochondria,
E. coli,
Pseudomonas,
S. aureus
Methanogens, E.
coli,
Pseudomonas,
Sulfate-reducing
bacteria
as many as 38 if starting with 1
glucose by aerobic respiration with
an electron transport chain
containing all the cytochromes – but
often far fewer than 38 - but still
more than 2.
Fermentation
organic molecule
carbohydrates, amino
acids, lipids
NADH
organic molecule
pyruvate
lactic acid, acetic acid,
ethanol, etc.
Bifidobacterium,
Lactobacillus, E. coli,
Clostridium, Bacteroides
2
Comparison of Respiration in Chemoorganotrophs vs Chemolithotrophs
Initial electron donor:
examples:
Electron donor oxidized
to:
Final electron acceptor
examples:
electron acceptor
reduced to:
example organisms
Chemoorganotroph
organic (-C-C-) molecule
carbohydrates, amino acids, lipids
CO2
inorganic molecule
CO2, NO3, SO4
O2 (aerobic
(anaerobic
respiration)
respiration)
Chemolithotroph
inorganic molecule
hydrogen gas, ammonia,
nitrate, hydrogen sulfide
water, nitrate, nitrite,
sulfuric acid
inorganic molecule
O2 (aerobic respiration)
H2O
CH4, NO2, H2S
H2O
Mitochondria,
E. coli,
Pseudomonas,
S. aureus
Methanogens,
E. coli,
Pseudomonas,
Sulfate-reducing
bacteria
Alcaligenes,
Nitrosomonas,
Nitrobacter,
Thiomargarita
45