Chapter 7 Bacteria Separation Designs Paper Design a bacterial separation problem using the terms from Chapter 7. The problem should include 3 or more bact

Chapter 7 Bacteria Separation Designs Paper Design a bacterial separation problem using the terms from Chapter 7. The problem should include 3 or more bacteria in a mixed culture. Design two problems for 1 bonus point each, and answer 1 problem designed by a classmate for 1 bonus point. *** 3 Bonus points possible

Example Problem: You are given a mixed culture containing an aerobic thermophile, aerobic psychrophile that is also a halophile, and an anerobic mesophile bacteria. How will you separate these into pure cultures?

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Example answer:

1) Separate the culture into 3 new tubes for isolation conditions

2) To isolate the Thermophile: Expose tube 1 to high temperatures above 45 Degrees Celsius, all bacteria will die except the Thermophile.

3) To isolate the Psychrophile: Expose tube 2 to cold temperatures below -15 degrees Celisius, all bacteria will die except the Psychrophile.

4) To isolate the Anaerobic Mesophile: Expose tube 3 to anaerobic conditions without oxygen, both aerobes will die, only the Anaerobe will live.

Note: If two bacteria are too closely related for the above approach, you could use the streak plate to isolate colonies to obtain a pure culture. You can differentiate different bacteria by colony color, form, elevation, margin on a agar plate. Chapter 7
Microbial Nutrition,
Ecology, and Growth
Microbial Biofilms
• Biofilms result when organisms attach to a
substrate by some form of extracellular matrix
that binds them together in complex organized
layers
• Dominate the structure of most natural
environments on earth
• Communicate and cooperate in the formation
and function of biofilms – quorum sensing
See the Youtube video montage regarding Biofilms

2
Biofilm Formation and Quorum Sensing
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Chromosome
Quorum-dependent
proteins
Inducer
molecule
1
5
4
2
Matrix
3
1
Free-swimming cells settle on a surface and remain there.
2
Cells synthesize a sticky matrix that holds them tightly to
the substrate.
3
When biofilm grows to a certain density (quorum), the cells release
inducer molecules that can coordinate a response.
4
Enlargement of one cell to show genetic induction. Inducer molecule
stimulates expression of a particular gene and synthesis of a protein
product, such as an enzyme.
5
Cells secrete their enzymes in unison to digest food particles.
3
Interrelationships Between
Microbes and Humans
• Human body is a rich habitat
for symbiotic bacteria, fungi,
and a few protozoa – normal
microbial flora
• Consider how antibiotics alter
your normal flora. Do you
think you should take
antibiotics for bacterial
infections that your immune
system could clear naturally?
4
Ecological Associations Among
Microorganisms
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Microbial Associations
Symbiotic
Nonsymbiotic
Organisms live in close
nutritional relationships;
required by one or both members.
Organisms are free-living;
relationships not required
for survival .
Mutualism Commensalism Parasitism
Obligatory,
The commensal
Parasite is
dependent;
benefits;
dependent
both members
other host is
and benefits;
benefit.
not harmed.
host harmed.
Synergism
Antagonism
Members Some members
cooperate
are inhibited
and share
or destroyed
nutrients.
by others.
5
Ecological Associations
• Symbiotic – two organisms
live together in a close
partnership
– Mutualism – obligatory,
dependent; both
members benefit
– Commensalism –
commensal member
benefits, the host is not
harmed but does not
benefit
– Parasitism – parasite
is dependent and
benefits; host is harmed
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Staphylococcus
aureus
growth
Haemophilus
satellite
colonies
© Science VU/Fred Marsik/Visuals Unlimited
Courtesy Arthur Hauck (Germany)
6
Ecological Associations
Non-symbiotic – organisms are free-living; relationships not
required for survival
• Synergism –
members cooperate to
produce a result that
none of them could do
alone
• Antagonism – actions
of one organism affect
the success or
survival of others in
the same community
(competition)
Nutritional Requirements: Carbon
• Carbon sources
• Heterotroph – must obtain
carbon in an organic form
made by other living
organisms such as proteins,
carbohydrates, lipids, and
nucleic acids
• Autotroph – an organism
that uses CO2, an inorganic
gas as its carbon source
– Not nutritionally
dependent on other living
things
8
Nutritional Requirements: Energy
Chemotroph – gain energy
from chemical compounds
• Phototrophs – gain energy
through photosynthesis
9
Growth Factors:
Essential Organic Nutrients
• Organic compounds that cannot
be synthesized by an organism
because they lack the genetic
and metabolic mechanisms to
synthesize them
• Growth factors must be
provided as a nutrient
– Essential amino acids,
vitamins
10
Heterotrophs and Their Energy Sources
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Digestion in Bacteria and Fungi
• Majority are chemoheterotrophs
– Aerobic respiration
• Two categories
– Saprobes: free-living
microorganisms that feed on
organic detritus from dead
organisms
• Opportunistic pathogen
• Facultative parasite
– Parasites: derive nutrients from
host
• Pathogens
• Some are obligate parasites
Organic debris
(a)
Walled cell is a barrier.
Enzymes
(b)
Enzymes are transported outside the wall.
(c)
Enzymes hydrolyze the bonds on nutrients.
11
(d)
Smaller molecules are transported across the
wall and cell membrane into the cytoplasm.
Concept Check:
If an organism is degrading large organic molecules
to get both carbon and energy, it would be best
described as a
A. Photoheterotroph
B. Photoautotroph
C. Chemoheterotroph
D. Chemoautotroph
Transport: Movement of Chemicals
Across the Cell Membrane
• Passive transport – does not require energy;
substances exist in a gradient and move from
areas of higher concentration toward areas of
lower concentration
– Diffusion
– Osmosis – diffusion of water
– Facilitated diffusion – requires a carrier
• Active transport – requires energy and carrier
proteins; gradient independent
– Examples are:
• endocytosis, exocytosis, pinocytosis
13
Osmosis – Diffusion of Water (Passive Transport)
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Membrane sac
with solution
Glass
tube
Solute
Water
Container
with
water
Pore
a. Inset shows a close-up of the osmotic process.
The gradient goes from the outer container
(higher concentration of H2O) to the sac (lower
concentration of H2O). Some water will diffuse
the opposite direction but the net gradient
favors osmosis into the sac.
b. As the H2O diffuses into the sac, the volume
increases and forces the excess solution into
the tube, which will rise continually.
c. Even as the solution becomes diluted, there
will still be osmosis into the sac. Equilibrium
will not occur because the solutions can never
become equal. (Why?)
15
Response to solutions of different osmotic content
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Cells with
Cell Wall
Isotonic Solution
Hypotonic Solution
Hypertonic Solution
Cell wall
Cell
membrane
Cell membrane
Water concentration is equal inside and
outside the cell, thus rates of diffusion
are equal in both directions.
Cells Lacking
Cell Wall
Net diffusion of water is into the cell; this
swells the protoplast and pushes it tightly
against the wall. Wall usually prevents
cell from bursting.
Early
Water diffuses out of the cell and
shrinks the cell membrane away from
the cell wall; process is known as
plasmolysis.
Early
Cell membrane
Late
(osmolysis)
Late
Rates of diffusion are equal
in both directions.
Direction of net water movement.
Diffusion of water into the cell causes
it to swell, and may burst it if no
mechanism exists to remove the water.
Water diffusing out of the cell causes
it to shrink and become distorted.
16
Facilitated Diffusion (Passive Transport)
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Outside
cell
Inside
cell
Outside
cell
Inside
cell
17
(a)
(b)
Carrier Mediated Active Transport
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Membrane
Membrane
Membrane
Protein
Protein
Protein
Protein
Protein
Protein
Extracellular
Intracellular
Extracellular
Intracellular
Extracellular
Intracellular
(a) Carrier-mediated active transport. The membrane proteins (permeases) have attachment sites for essential nutrient molecules. As these
molecules bind to the permease, energy from ATP pumps them into the cell’s interior through special membrane protein channels. Microbes
have these systems for transporting various ions (sodium, iron) and small organic molecules.
Group Translocation
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Extracellular
Membrane
Membrane
Protein
Protein
Protein
Protein
Intracellular
Extracellular
19
Intracellular
(b) In group translocation, the molecule is actively captured, but along the route of transport, it is chemically altered. By coupling transport
with synthesis, the cell conserves energy.
Example problem 1:
• Cell A has 60% glucose and is placed in a
solution with 15% glucose and the cell
membrane is permeable to the solvent.
• Based on this background answer the following
questions:
1. What type of problem is this?
A.
B.
C.
D.
Osmosis
Diffusion
Facilitated diffusion
Active transport
20
Example problem 1:
• Cell A has 60% glucose and is placed in a
solution with 15% glucose and the cell
membrane is permeable to the solvent.
• Based on this background answer the following
questions:
60% glucose
40% H20
2. Which way will water move?
A. Into the cell
B. Out of the cell
C. Neither into or out of the cell
15% glucose
85% H20
21
Example problem 1:
• Cell A has 60% glucose and is placed in a
solution with 15% glucose and the cell
membrane is permeable to the solvent.
• Based on this background answer the following
60% glucose
questions:
40% H20
3. The solution is ___ to the cell.
A. Hypertonic
B. Hypotonic
C. Isotonic
15% glucose
85% H20
22
Example problem 2:
• Cell A has 20% glucose and cell B has 30% glucose and
the protein carrier in the cell membrane is pumping
solute across.
• Based on this background answer the following
questions:
1. What type of problem is this?
A.
B.
C.
D.
Osmosis
Diffusion
Facilitated diffusion
Active transport
20% glucose 30% glucose
80% H20
70% H20
A
B
23
Example problem 2:
• Cell A has 20% glucose and cell B has 30% glucose and
the protein carrier in the cell membrane is pumping
solute across.
• Based on this background answer the following
questions:
2. Which way is the solute moving?
A.
B.
C.
From Cell A to B
From Cell B to A
No movement
20% glucose 30% glucose
80% H20
70% H20
A
B
24
Example problem 2:
• Cell A has 20% glucose and cell B has 30% glucose and
the protein carrier in the cell membrane is pumping
solute across.
• Based on this background answer the following
questions:
3. Cell B is ____ to cell A.
A.
B.
Hypertonic
Hypotonic
C.
Isotonic
20% glucose 30% glucose
80% H20
70% H20
A
B
25
Endocytosis: Eating and Drinking by Cells
• Endocytosis: bringing substances into the cell
through a vesicle or phagosome
– Phagocytosis ingests substances or cells
– Pinocytosis ingests liquids
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Phagocytosis
Pinocytosis
4
Pseudopods
Microvilli
3
Liquid enclosed
by microvilli
Oil droplet
2
Vacuoles
1
Vesicle with liquid
26
Summary of Transport Processes
27
Concept Check:
If a cell is in a concentrated glucose solution and the
glucose is moving into the cell through a carrier protein,
this would be an example of
A. Diffusion
B. Facilitated Diffusion
C. Active Transport
D. Endocytosis
E. Pinocytosis
Environmental Factors That
Influence Microbes
• Niche: totality of adaptations organisms make to
their habitat
• Environmental factors affect the function of
metabolic enzymes
• Factors include:





Temperature
Oxygen requirements
pH
Osmotic pressure
Barometric pressure
29
3 Cardinal Temperatures
• Minimum temperature – lowest temperature
that permits a microbe’s growth and metabolism
• Maximum temperature – highest temperature
that permits a microbe’s growth and metabolism
• Optimum temperature – promotes the fastest
rate of growth and metabolism
30
3 Temperature Adaptation Groups
Psychrophiles – optimum temperature below 15oC; capable of
growth at 0oC
Mesophiles – optimum temperature 20o-40oC; most human
pathogens
Thermophiles – optimum temperature greater than 45oC
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Optimum
Minimum
Psychrophile
Mesophile
Thermophile
Maximum
-15-10 -5 0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90
Temperature °C
31
Gas Requirements
Oxygen
• As oxygen is utilized it is transformed into
several toxic products:
– Singlet oxygen (1O2), superoxide ion (O2-), peroxide
(H2O2), and hydroxyl radicals (OH-)
• Most cells have developed enzymes that
neutralize these chemicals:
– Superoxide dismutase, catalase
• If a microbe is not capable of dealing with toxic
oxygen, it is forced to live in oxygen free habitats
32
Categories of Oxygen Requirement
• Aerobe – utilizes oxygen and can detoxify it
• Obligate aerobe – cannot grow without oxygen
• Facultative anaerobe – utilizes oxygen but can
also grow in its absence
• Microaerophilic – requires only a small amount
of oxygen
• Anaerobe – does not utilize oxygen
• Obligate anaerobe – lacks the enzymes to
detoxify oxygen so cannot survive in an oxygen
environment
• Aerotolerant anaerobes – do not utilize oxygen
but can survive and grow in its presence
33
Culturing by Oxygen Requirement
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Photo by Keith Weller, USDA/ARS
© Terese M. Barta, Ph.D.
Carbon Dioxide Requirement
All microbes require some carbon dioxide in their
metabolism
• Capnophile – grows best at higher CO2 tensions
than normally present in the atmosphere
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
35
Courtesy and © Becton, Dickinson and Company
Effects of pH
• Majority of microorganisms grow at a pH between
6 and 8 (neutrophiles)
• Acidophiles – grow at extreme acid pH
• Alkalinophiles – grow at extreme alkaline pH
36
Osmotic Pressure
• Most microbes exist under hypotonic or isotonic
conditions
• Halophiles – require a high concentration of salt
• Osmotolerant – do not require high
concentration of solute but can tolerate it when it
occurs
37
Other Environmental Factors
• Barophiles – can survive under extreme
pressure and will rupture if exposed to normal
atmospheric pressure
• Ex: Deep ocean prokaryotes
38
Concept Check:
Chlamydomonas nivalis grows on Alaskan glaciers and it’s
photosynthetic pigments give the snow a red crust. This
organism would be best described as a
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
A. Psychrophile
B. Alkalinophile
C. Microaerophile
D. Osmotolerant
E. Barophile
Image courtesy Nozomu Takeuchi
Image courtesy Nozomu Takeuchi
(a)
(b)
Rate of Population Growth
• Time required for a complete fission cycle is called
the generation, or doubling time
• Each new fission cycle increases the population
by a factor of 2 – exponential growth
• Generation times vary from minutes to days
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
4500*
*12
4000
3500
3000
Log of
(
Number
2500 of cells (
2000
) number
of cells
using the 11
power
of 2
10
1500
1000
500
9
0
(b)
Number
of cells
Number of
generations
Exponential
value
(a)
1
2
4
8
16
32
1
2
3
4
5
24
25
21
22
(21)
(22)
23
(222)
(2222) (22222)
)
0
Time
40
Concept Check:
Escherichia coli has a
doubling time of 20 minutes.
If there are 5 cells at the
beginning of the experiment,
how many will there be in 3
hours?
Time
Colony Forming Units
(CFU)
0
5
20
10
40
20
60
40
80
80
100
160
120
320
140
640
160
1280
180
2560
Viable Plate Count
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Flask inoculated
Samples taken at equally spaced intervals
(0.1 ml)
60 min
500 ml
120 min
180 min
240 min
300 min
360 min
420 min
480 min
540 min
600 min
135
230
0.1
ml
Sample is
diluted in
liquid agar
medium
and poured
or spread
over surface
of solidified
medium
Plates are
incubated,
colonies
are counted
None
Number of
colonies (CFU)
per 0.1 ml

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