Transcript Growth and Multiplication of Bacteria

Growth and Multiplication of
Bacteria
Hugh B. Fackrell
Sept 1997
Filename: Growth.ppt
Requirements for
Growth/Multiplication
 ALL required nutrients
 correct
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pH
temperature
salinity,
moisture
redox potential
atmosphere
Growth
Liquid vs Solid Media
 Liquid: clear >>turbid
 Solid: individual colonies
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each colony derived from a single cell
Growth Event
 Absorption of water & nutrients
 Catabolism of carbon source
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inorganic or organic
 Biosynthesis of new cellular components
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major energy consumption
 Cell enlargement
 Cell division ( Binary Fission)
Binary Fission
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DNA replication
Plasma membrane invaginate
Cell wall deposited in invaginated space
Cross wall completed
Cells separate
Binary Fission
 Light micrograph
Binary Fission
Consequences of Binary Fission
 Very large number of cells very fast
 Mathematical progressions
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arithmetic (1>2>4>6>8>10>12>14>16)
geometric(1>2>4>8>16)
• exponential expression (20 > 21 > 22 >23>24)
• logarithmic expression(0 >log21>log22>log23>log24
Logarithmic Plots
 Can plot very large Range of numbers
 Phases of growth demonstrated
 Generation time easily calculated
Cell Multiplication
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0
1 2 0
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2 21 log21 
4 22 log22 
8 23 log23 
16 24 log24 
Mathematics of bacterial growth
Cells
Generation #
Log 2 Log10
0
1
0
0.000
1
2
1
0.301
2
4
2
0.602
3
8
3
0.903
4
16
4
1.204
5
32
5
1.505
6
64
6
1.806
7
128
7
2.107
8
256
8
2.408
Growth Data
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#Generation
1
5
10
15
20
#cells
1
32
1,024
32,768
1,048,576
Log10
0
1.51
3.01
4.50
6.02
Growth curves for exponentially
increasing population
Number
of cells
Log number
of cells
Time (hours)
Bacterial Growth Curve
Stationary phase
Death
phase
Log phase
Lag phase
1
5
Time (hours)
10
Measurement of
Growth Constants
 G: Generation Time
 K: Mean Growth Rate Constant
G := 1/K
G: Generation time
Time in minutes or hours for a
population of bacteria to double in
number
Calculation of Generation Time
Log Number
of Bacteria
Double
# cells
Log phase
Generation time
1
5
Time (hours)
10
Slope of Log phase proportional
to generation time
Fast
Log
Number of
bacteria
Medium
Doubling number
Slow
Time (hours)
K: Mean Growth Rate Constant
 K= n/t
 K= (log10Nt - log10Nt0)/ 0.301t
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N= number of cells
n=: number of generations
t = time (hr or min)
K = 1/slope ( semi log growth plot)
Therefore G = 1/K
Sample calculation for K & G
 Population increase from 103 to 109 in 10hrs
 K= (log 109 - log 103) / 0.301 x 10
 K= 9-3/3.01 = 2 generations/hours
 G = 1/K = 1/2 = 0.5 hr/generation
Factors influencing lag phase
 Age of culture inoculum
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old culture -> long lag
young culture-> short lag
 Size of inoculum
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few cells -> long lag
many cells -> short lag
 Environment
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pH, temp, gases,salinity
sub optimum -> long lag
optimum-> short lag
Growth Responses: Temperature
Thermophile
Mesophile
Rate of
Psychrotroph
Growth Pyschrophile
-10 0 10
Extreme
Thermophile
20 30 40 50 60 70 80
Temperature (o C)
90 100
Growth Responses: pH
Neutrophile Alkalophile
Rate of
Growth
Acidophile
1
2
3
4
5
6
7
pH
8
9
10
11 12
Diauxic Growth
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Growth on two carbon sources
Mixed sugars
Each sugar used separately
Glucose ALWAYS used first
Second sugar ONLY used when glucose
GONE
Diauxic Growth: 2 carbon sources
Growth
[Sugar]
Arabinose
Glucose
Time (hr)
Synchronous Growth
 Filtration
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Smaller cells
all same size
 Temperature shock
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Hot/cold brings cells to same metabolic state
 Starvation
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deplete medium of selected nutrient
Synchronous vs Asynchronous
growth
Number
of Cells
Synchronous
growth
Asynchronous growth
Time (min)
Growth in Limited Nutrients
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Limiting concentration of Required nutrient
YIELD
number of cells
Linear increase yield with nutrient conc
Yield = Mass of organisms formed
Mass of nutrients used
Growth in Limited Nutrients
Growth
Rate
Total
Growth
[Nutrient]
Applications of
Limiting [Nutrient]
 Chemostat (continuous culture)
 Bio-Assay
Bio-Assay: Procedure
 Bacterium: CANNOT synthesize nutrient
 Medium: all growth requirements except
nutrient to be assayed
 Add
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equal amounts of medium to each tube
equal numbers of bacteria to each tube
increasing amounts of the nutrient to be assayed
 [Unknown]
 Incubate
 Measure growth (turbidity or viable count)
Bio-Assay
 Vitamin B-12 measurement in Green beans
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Lactobacillus leichmanni
Growth of known [nutrient]
Microbial
Growth
Growth of unknown
0
0
0
0
0
[Nutrient] in
unknown
[Nutrient] mg/ml
0
Chemostat
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Description of Instrument
Principle
Steady State
Sample Results
Application
Chemostat: Description of Instrument
Chemostat: Principle
 Essential nutrient is limited
 Growth rate(K) controlled by supply rate of
nutrient
 Yield controlled by concentration of nutrient
 Dilution rate (D): speed of nutrient flow into the
culture vessel
D = Flow rate
Vessel volume
Steady State
K=D
Chemostat: Sample Results
Cell density or biomass
Measurement
Value
Generation time
Nutrient conc
Dilution Rate of Nutrient
Chemostat: Applications
 Growing large amounts of cells
 Industrial production
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vaccines
pharmaceuticals
hormones
 Long term studies of specific growth phase
 Selecting for specific mutants
 Aquatic systems
Bacterial Growth in
Natural Environments
 Natural Environments
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Animal Tissues
Soil
Water- freshwater- marine
Plants
Bacterial Growth in
Natural Environments
 Active
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Short bursts of growth & metabolism
usually low rates of growth
 Quiescent
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Viable cannot culture
 Stressed
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starvation semi viable
Biofilms: Body
 Catheter
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Foley:
• latex silicone
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Intravenous:
• polyurethane S. epidermidis
 Prostheses
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Hip joints
Dental implants
voicebox
 Tampons
 IUD
Biofilms: Water
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Dental lines
Spacecraft
Drinking filters
ALL surfacces
Biofilm in gut of a mollusc
Biofilms: Disease
 Cystic fibrosis
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lung-alveolar surface
 Ulcers
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Helicobacter jejuni
 Dental caries
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Streptococccus spp