Transcript Lecture 8_PPT - DrMinkovskyScienceWiki

Microbial
Nutrition,
Ecology, and
Growth
Chapter 7
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Learning Objectives:
• Classify microbes into five groups on the basis of
preferred temperature range.
• Identify how and why the pH of culture media is
controlled.
• Explain the importance of osmotic pressure to
microbial growth.
• Explain how microbes are classified on the basis
of oxygen requirements.
• Identify ways in which aerobes avoid damage by
toxic forms of oxygen
Learning Objectives:
• Define bacterial growth, including binary fission.
• Compare the phases of microbial growth and
describe their relation to generation time.
• Describe three direct methods for measuring
microbial growth.
• Differentiate between direct and indirect methods
for measuring cell growth.
• Explain three indirect methods of measuring cell
growth
Temperature Optima
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Psychrophile
Rate of Growth
Optimum
Psychrotroph
Thermophile
Mesophile
Extreme thermophile
Minimum
-1 5 -10 -5
0
Maximum
5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95 100 105110115120125 130
Temperature °C
Psychrotrophs
• Grow
between 0°C
and 20°C
• Cause food
spoilage
pH Optima
• Most bacteria grow between pH 6.5 and 7.5
• Molds and yeasts grow between pH 5 and 6
• Acidophiles grow in acidic environments
• Alcaliphiles grow in basic environments
pH 0
1
2
3
Acidic
4
[H+]
5
6
7
Neutral
8
9
[OH–]
10
11
12
Basic
(alkaline)
13
14
Plasmolysis
Osmotic pressure
•Hypertonic environments, increase salt or sugar, cause
plasmolysis
•Extreme or obligate halophiles require high osmotic pressure
•Facultative halophiles tolerate high osmotic pressure
Conclusions:
• On the basis of preferred temperature range, microbes are
classified as psychrophiles cold-loving), mesophiles
(moderate-temperature-loving), and thermophiles (heatloving).
• The minimum growth temperature is the lowest temperature
at which a species will grow, the optimum growth
temperature – at which it grows best, and the maximum
temperature – the highest at which growth is possible.
• Most bacteria grow best at a pH value between 6.5 and 7.5
• In a hypertonic solution, most microbes undergo
plasmolysis; halophiles can tolerate high salt concentrations
Oxygen Requirements
•
Oxygen is needed for aerobic respiration.
•
Oxygen is a powerful oxidizing agent (toxic to
cells)
•
Some microbes use oxygen and can detoxify it.
•
Some microbes do not use oxygen and cannot
detoxify it.
•
Some microbe do not use oxygen but can
detoxify it.
Toxic Oxygen Species
• Singlet oxygen: O2 boosted to a higher-energy
state
• Superoxide free radicals: O2–
• Peroxide anion: O22–
• Hydroxyl radical (OH)
Microbe Requirements for Growth
Obligate
Facultative
Aerobes
Anaerobes Anaerobes
Require
Does not
O2
require O2
Obligate
No O2
Aerotolerant
Micro-
Anaerobes
aerophiles
Can survive in
Require low
presence of O2
concentration of O2
Table 6.1
Oxygen Requirement in
Thioglycollate Broth
• Aerobes
• Microaerophiles
• Anaerobes
• Facultative anaerobes
• Aerotolerant anaerobe
Anaerobic
Culture
Methods
• Anaerobic jar
serves the
purpose of
chemically
removing oxygen
Figure 6.5
Capnophiles
Require High CO2
• Candle jar can be used
to grow Neissseria
meningitidis
• CO2-packet is used to
generate an environment
that contains more
carbon dioxide than
oxygen
Figure 6.7
Other Factors
• Barometric pressure - barophiles
• Dry - Xerophiles
Conclusions:
• On the basis of oxygen requirements, organisms
are classified as obligate aerobes, facultative
anaerobes, obligate anaerobes, aerotolerant
anaerobes, and microaerophiles.
• Aerobes, facultative anaerobes, and aerotolerant
anaerobes must have the enzymes superoxide
dismutase, and either catalase or peroxidase.
Bacterial Reproduction
• Binary fission
• Asexual
process
• Doubling time
(generation)
if 20 minutes,
then in 24
hours
Cell wall
Cell membrane
Chromosome 1
Chromosome 2
Ribosomes
1 A young cell.
Chromosome is
replicated and
2 new and old
chromosomes
move to different
sides of cell.
3 Protein band
forms in
center of cell.
4 Septum formation
begins.
• 1  4.7 x 1021
cells
• 5,100 tons
When septum is
complete, cells are
5 considered divided.
Some species will
separate completely
as shown here, while
others remain attached,
The Math of Bacterial Growth
• X = X0 * 2Y
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10
9
10,000,000,000
8
Number
of cells
7
(
Number 1
Number of
generations
Exponential
value
(a)
2
4
8
16
32
Log of
6
)
number
5
of cells
(
3
10,000
1,000
2
100
4
1
21
(2 × 1)
2
3
4
5
22
23
24
25
(2 × 2)
(2 × 2 × 2)
(2 × 2
(2 × 2
× 2 × 2) × 2 × 2 × 2)
0
(b)
Time
1
)
Bacterial Growth Curve
Implications for:
•Applying antimicrobial agents
•Treating infections
Conclusions:
• The normal reproductive method of bacteria is binary fission,
in which a single cell divides into two identical cells.
• The time required for a cell to divide or a population to double
is the generation time.
• Bacterial division occurs according to a logarithmic
progression.
• During the lag phase, there is little or no change in the
number of cells, but metabolic activity is high.
• During the log phase, the bacteria multiply at a fastest rate
possible under the conditions provided.
• During the stationary phase, there is an equillibrium between
cell division and death
• During the death phase, the number of deaths exceeds the
number of new cells formed.
Measuring Microbial Growth
Direct methods
Indirect methods
• Plate counts
• Turbidity
• Filtration
• Metabolic activity
• Direct microscopic count
• Dry weight
• Automated cell count
Plate Count
• After incubation, count colonies on plates that have
25-250 colonies (CFUs)
Plate Count
Method
• Inoculate Petri
plates from serial
dilutions using
either method
• Incubate plates
and count up the
number of
colonies
Figure 6.16
Direct Measurements of Microbial
Growth
• Filtration
Direct Cell Counts
• Cytometer
• Known volume
• Count total cells
• Both dead and live cells are counted
Direct Measurements of Microbial
Growth
• Direct microscopic count
Automated Cell Counting
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• Coulter counter
• Flow cytometer
• Can sort cells
• Requires tagging
Automatic
counter
Sample in
liquid
Bacterial
cell
Tube
Counting orifice
Electronic detector
Bacteria Scatter Light
• Spectrophotometer
• Transmittance
• Absorbance
• Red light scatters best
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Percentage of light
transmitted
High
(1)
Low
(a)
(2)
(b)
a: © Kathy Park Talaro/Visuals Unlimited.
Measure Cell Components
• We can measure growth by increase in mass…
• Measure dry weight of cells
• Assume that relative fraction of the cell
components is relatively stable
• Lipid content
• DNA content
• Protein determinations (most common)
Conclusions:
• A standard plate count reflects the number of viable
microbes and assumes that each bacterium grows
into a single colony; it is reported as the number of
colony forming units (CFU)
• In filtration, bacteria are retained on the surface of a
membrane filter and then transferred to a culture
medium to grow and to be counted.
• In a direct count, the microbes in a measured
volume of a bacterial suspension are counted with
the use of a specially designed slide.
Conclusions:
• A spectrophotometer is used to determine
turbidity by measuring the amount of light that
passes through a suspension of cells.
• An indirect way of estimating bacterial numbers
is measuring the metabolic activity of a
population (for example, acid production or
oxygen consumption)