1. Growth and Multiplication of Bacteria
Hugh B. Fackrell
Sept 1997
Filename: Growth.ppt

2. Requirements for Growth/Multiplication
ALL required nutrients
correct pH
temperature
salinity,
moisture
redox potential
atmosphere

3. Growth Liquid vs Solid Media
Liquid: clear >>turbid
Solid: individual colonies
each colony derived from a single cell

4. Growth Event Absorption of water & nutrients
Catabolism of carbon source
inorganic or organic
Biosynthesis of new cellular components
major energy consumption
Cell enlargement
Cell division ( Binary Fission)

5. Binary Fission DNA replication Plasma membrane invaginate
Cell wall deposited in invaginated space
Cross wall completed
Cells separate

6. Binary Fission
Light micrograph

7. Binary Fission

8. Consequences of Binary Fission
Very large number of cells very fast
Mathematical progressions
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

9. Logarithmic Plots Can plot very large Range of numbers
Phases of growth demonstrated
Generation time easily calculated

10. Cell Multiplication 1 20 0 l 2 21 log21 ll 4 22 log22 llll
log23 lllllllll
log24 lllllllllllllllll

11. Mathematics of bacterial growth
Cells
Generation # Log Log10

12. Growth Data #Generation #cells Log10 1 1 0 5 32 1.51 10 1,024 3.01 , ,
,048,

13. Growth curves for exponentially increasing population
Number of cells
Log number of cells
Time (hours)

14. Bacterial Growth Curve
Stationary phase
Death phase
Log phase
Lag phase 1 5 10
Time (hours)

15. Measurement of Growth Constants
G: Generation Time
K: Mean Growth Rate Constant
G := 1/K

16. G: Generation time
Time in minutes or hours for a population of bacteria to double in number

17. Calculation of Generation Time
Log Number of Bacteria
Log phase
Double # cells
Generation time 1 5 10
Time (hours)

18. Slope of Log phase proportional to generation time
Time (hours)
Log Number of bacteria
Fast
Medium
Doubling number
Slow

19. K: Mean Growth Rate Constant
K= n/t
K= (log10Nt - log10Nt0)/ 0.301t
N= number of cells
n=: number of generations
t = time (hr or min)
K = 1/slope ( semi log growth plot)
Therefore G = 1/K

20. Sample calculation for K & G
Population increase from 103 to 109 in 10hrs
K= (log log 103) / x 10
K= 9-3/3.01 = 2 generations/hours
G = 1/K = 1/2 = 0.5 hr/generation

21. Factors influencing lag phase
Age of culture inoculum
old culture -> long lag
young culture-> short lag
Size of inoculum
few cells -> long lag
many cells -> short lag
Environment
pH, temp, gases,salinity
sub optimum -> long lag
optimum-> short lag

22. Growth Responses: Temperature
Rate of Growth
Thermophile
Mesophile
Extreme Thermophile
Psychrotroph
Pyschrophile
-10 10 20 30 40 50 60 70 80 90
100
Temperature (o C)

23. Growth Responses: pH pH Rate of Growth Neutrophile Alkalophile
Acidophile 1 2 3 4 5 6 7 8 9 10 11 12 pH

24. Diauxic Growth Growth on two carbon sources Mixed sugars
Each sugar used separately
Glucose ALWAYS used first
Second sugar ONLY used when glucose GONE

25. Diauxic Growth: 2 carbon sources
[Sugar]
Arabinose
Glucose
Time (hr)

26. Synchronous Growth Filtration Temperature shock Starvation
Smaller cells
all same size
Temperature shock
Hot/cold brings cells to same metabolic state
Starvation
deplete medium of selected nutrient

27. Synchronous vs Asynchronous growth
Number of Cells
Asynchronous growth
Synchronous growth
Time (min)

28. Growth in Limited Nutrients
Limiting concentration of Required nutrient
YIELD
number of cells
Linear increase yield with nutrient conc
Yield = Mass of organisms formed
Mass of nutrients used

29. Growth in Limited Nutrients
Total Growth
Growth Rate
[Nutrient]

30. Applications of Limiting [Nutrient]
Chemostat (continuous culture)
Bio-Assay

31. Bio-Assay: Procedure Bacterium: CANNOT synthesize nutrient
Medium: all growth requirements except nutrient to be assayed
Add
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)

32. Bio-Assay Vitamin B-12 measurement in Green beans
Lactobacillus leichmanni
Microbial Growth
Growth of known [nutrient]
Growth of unknown
[Nutrient] in unknown
[Nutrient] mg/ml

33. Chemostat Description of Instrument Principle Steady State
Sample Results
Application

34. Chemostat: Description of Instrument

35. 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
Steady State
K = D
Flow rate
Vessel volume
D =

36. Chemostat: Sample Results
Measurement
Value
Cell density or biomass
Nutrient conc
Generation time
Dilution Rate of Nutrient

37. Chemostat: Applications
Growing large amounts of cells
Industrial production
vaccines
pharmaceuticals
hormones
Long term studies of specific growth phase
Selecting for specific mutants
Aquatic systems

38. Bacterial Growth in Natural Environments
Animal Tissues
Soil
Water- freshwater- marine
Plants

39. Bacterial Growth in Natural Environments
Active
Short bursts of growth & metabolism
usually low rates of growth
Quiescent
Viable cannot culture
Stressed
starvation semi viable

40. Biofilms: Body Catheter Prostheses Tampons IUD Foley: Intravenous:
latex silicone
Intravenous:
polyurethane S. epidermidis
Prostheses
Hip joints
Dental implants
voicebox
Tampons
IUD

41. Biofilms: Water Dental lines Spacecraft Drinking filters ALL surfacces
Biofilm in gut of a mollusc

42. Biofilms: Disease Cystic fibrosis Ulcers Dental caries
lung-alveolar surface
Ulcers
Helicobacter jejuni
Dental caries
Streptococccus spp