1. The Chemostat
Continuous culture devices are a means of maintaining cell populations in exponential growth for long periods.
In a chemostat, the rate at which the culture is diluted governs the growth rate and growth yield.

2. Microbial Growth in a Chemostat

3. Microbial growth: measurement and influence of environmental factors

4. Measurement of Microbial Growth
Can measure changes in number of cells in a population
Direct cell counts
-counting chambers
-on membrane filters
Viable cell counts
-plating methods
-membrane filtration methods
Can measure changes in mass of population
-dry weight
-quantity of a particular cell constituent
-turbidometric measures

5. Counting chambers easy, inexpensive, and quick
cannot distinguish living from dead cells
examples: Petroff-Hauser or hemocytometers
Figure 6.12

6. Direct counts on membrane filters
Cells filtered through special membrane that provides dark background for observing cells
Cells are stained with fluorescent dyes
Useful for counting bacteria
With certain dyes, can distinguish living from dead cells

7. Membrane filtration method
especially useful for analyzing aquatic samples
Figure 6.13

8. Measurement of Cell Mass
Dry weight
time consuming and not very sensitive
Quantity of a particular cell constituent
protein, DNA, ATP, or chlorophyll
Turbidometric measures (light scattering)
quick, easy, and sensitive

9. more cells  more light scattered less light detected Figure 6.15
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more cells 
more light
scattered
less light
detected
Figure 6.15

10. Environmental Factors on Growth
Most organisms grow in fairly moderate environmental conditions
Extremophiles
grow under harsh conditions that would kill most other organisms

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Table 6.3

12. Water Activity (aw) and osmosis
amount of water available to organisms
reduced by interaction with solute molecules (osmotic effect)
higher [solute]  lower aw

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Table 6.4

14. Halophilic and halotolerant microorganisms
Halophilic microorganisms
Absolute requirement of salt for growth
Accumulate K+ (primarily in archaea)
Accumulate organic compounds (compatible solutes) (primarily in bacteria)
Halotolerant microorganisms
No absolute requirement of salt for growth
grow over wide ranges of salinity
many use compatible solutes

15. halophiles extreme halophiles grow optimally at > 0.2 M
require > 2 M
Figure 6.18

16. pH acidophiles neutrophiles alkalophiles
growth optimum between pH 0 and pH 5.5
neutrophiles
growth optimum between pH 5.5 and pH 7
alkalophiles
growth optimum between pH 8.5 and pH 11.5

17. pH
Most acidophiles and alkalophiles maintain an internal pH near neutrality
The plasma membrane is impermeable to protons
Symport, antiport systems can be used to maintain pH closer to neutrality
Synthesize proteins that provide protection
e.g., acid-shock proteins
Many microorganisms change pH of their habitat by producing acidic or basic waste products
most media contain buffers to prevent growth inhibition

18. Temperature Figure 6.20 Greatly effects enzyme activities.
Organisms exhibit distinct cardinal growth temperatures
minimal
maximal
optimal
Figure 6.20

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Figure 6.21

21. Hyperthermophiles in Hot Springs

22. Adaptations of thermophiles
Protein structure stabilized by a variety of means
more H bonds
more proline
chaperones
Histone-like proteins stabilize DNA
Membrane stabilized by variety of means
more saturated, more branched and higher molecular weight lipids, lipid monolayers
e.g., ether linkages (archaeal membranes)

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Table 6.5

24. Oxygen Concentration need oxygen ignore oxygen < 2 – 10% oxygen
prefer
oxygen
oxygen is
toxic
Figure 6.22

25. Oxygen toxicity Some enzymes are extremely sensitive to oxygen.
oxygen easily reduced to toxic products
superoxide radical
hydrogen peroxide
hydroxyl radical
aerobes produce protective enzymes
superoxide dismutase (SOD)
catalase

28. Figure 6.24

29. Pressure barotolerant barophilic organisms
adversely affected by increased pressure, but not as severely as nontolerant organisms
barophilic organisms
require or grow more rapidly in the presence of increased pressure

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Radiation
Figure 6.25

31. Radiation Damage Ionizing radiation X-rays and gamma rays
mutations  death
disrupts chemical structure of DNA
damage may be repaired by DNA repair mechanisms

32. Radiation Damage… Non-Ionization radiation -Ultraviolet (UV) radiation
mutations  death
causes formation of thymine dimers in DNA
DNA damage can be repaired by several repair mechanisms

33. Radiation damage… Visible light
at high intensities generates singlet oxygen (1O2)
powerful oxidizing agent
carotenoid pigments protect many light-exposed microorganisms from photooxidation