1. Dynamics of Prokaryotic Growth
Chapter 4

2. Preview Principles of bacteria growth. Bacteria growth in nature.
Bacteria growth in laboratory.
Factors affect bacteria growth.
Detecting bacteria growth

3. Principles of Bacterial Growth
Prokaryotic cells divide by binary fission
One cell divides into two
Cell growth is exponential
population double with each cell division.
Cell divide at constant pace
Generation time
Time it takes for population to double
A.k.a doubling time
Varies among species

4. Practical Problem 100 E. coli in potato salad.
How many bacteria are there in salad 2 hrs later (if the double time is 20 min)?
Why do we need to store the salad in cooler?

5. Principles of Bacterial Growth
Growth can be calculated
Nt = N0 x 2n
(Nt ) number of cells in population
(N0 ) original number of cells in the population
(n) number of divisions
Example
N0 = 10 cells in original population
n = 12
4 hours assuming 20 minute generation time
Nt = 10 x 212
Nt = 10 x 4,096
Nt = 40,960

6. Bacterial Growth in Nature
Conditions in nature have profound effect on microbial growth
Synthesize compounds useful for growth
produce multicellular associations to increase survivability
Biofilms: a community formed by a group of bacteria and their secreted slimes.
Biofilm layer

7. Biofilm Can cause disease Can be beneficial Difficult to kill biofilm
Architecture resist immune response and antimicrobial drugs
Can be beneficial
biofilm

8. Bacterial Growth in Nature
Prokaryotes live in mixed communities
Many interactions are cooperative
Some cells compete for nutrient

9. Bacteria growth in Laboratory
Culture media
broth media
Solid media is broth media with addition of agar
Agar marine algae extract
Liquefies above 95°C
Solidifies at 45°C
Remains solid at room temperature and body temperature
Bacteria grow in colonies on solid media surface
Single colony

10. Laboratory Cultivation
Special types of culture media
These are used to detect or isolate particular organisms
Are divided into selective and differential media

11. Laboratory Cultivation
Selective media
Inhibits the growth of unwanted organisms
Allows only sought after organism to grow
Example
Thayer-Martin agar (multiple antimicrobial)
For isolation of Neisseria gonorrhoeae
MacConkey agar (antimicrobial+bile salt)
For isolation of Gram-negative intestinal bacteria

12. Laboratory Cultivation
Differential media
Contains substance that bacteria change in recognizable way
Example
Blood agar
Test for hemolysis
MacConkey agar
pH indicator

13. Obtaining Pure Culture
Pure culture is defined as population of cells derived from single cell
All cells are genetically identical
to study functions of specific species
Obtain pure culture
Aseptic technique

14. Obtaining Pure Culture
Streak-plate method
Simplest and most commonly used in bacterial isolation
Object is to reduce number of cells being spread

15. Bacterial Growth in Laboratory Conditions
Cells in laboratory grown in closed or batch system
Population of cells increase in predictable fashion
Follows a pattern called growth curve 15

16. Bacterial Growth in Laboratory Conditions
The Growth Curve
Characterized by five distinct stages
Lag stage
Exponential or log stage
Stationary stage
Death stage
Phase of prolonged decline 16

17. Bacterial Growth in Laboratory Conditions
Lag phase
synthesis of cell components and prepare for division
Log phase
exponential growth
Cell divide at constant rate
Produce primary metabolites
Compounds required for growth
Cells enter late log phase
Cell wall and cell membrane component changes
Synthesize secondary metabolites
Used to enhance survival
Antibiotics 17

18. Bacterial Growth in Labortory Conditions
Stationary phase
Overall population remains relatively stable
Cells exhausted nutrients and build up toxic waste
Cell growth = cell death
Death phase
Total number of viable cells decreases
99% of cells die at constant rate
Death is exponential
Much slower rate than growth 18

19. Bacterial Growth in Laboratory Conditions
Phase of prolonged decline
Marked by very gradual decrease in viable population
Phase may last months or years
Most fit cells survive
Each new cell more fit that previous 19

20. Bacterial Growth in Laboratory Conditions
Continuous culture
Bacterial culture can be maintained
Continuous exponential growth can be sustained by use of chemostat 20

21. Bacterial Growth in Laboratory Conditions
Colony growth on solid medium
Position within colony determines resource availability
Cells on edge of colony have little competition and significant oxygen stores
Cells in the middle of colony have high cell density
Leads to increased competition and decreased availability of oxygen

22. Questions What is biofilm? What is pure culture?
What are the different stages of bacterial growth?
Selective and differential medium.

23. Environmental Factors on Growth
Major conditions that influence growth
Temperature
Oxygen pH
Water availability

24. Environmental Factors on Growth
Psychrophile
Optimum temperature -5°C to 15°C
Found in Arctic and Antarctic regions
Psychrotroph
20°C to 30°C
Important in food spoilage
Mesophile
25°C to 45°C
More common
Disease causing
Thermophiles
45°C to 70°C
Common in hot springs
Hyperthermophiles
70°C to 110°C
Usually members of Archaea
Found in hydrothermal vents

25. Environmental Factors on Growth
Temperature and food storage
4C can slow down bacteria growth
Freezing can stop bacteria growth
Temperature and disease
Different pathogen can only grow in different part of body.
Hansen’s disease
Syphilis disease

26. Environmental Factors on Growth
Oxygen
Prokaryotes divided based on oxygen requirements
Obligate aerobes
Absolute requirement for oxygen
Use for energy production
Micrococcus
Obligate anaerobes
No multiplication in presence of oxygen
May cause death
Clostridium

27. Environmental Factors on Growth
Facultative anaerobes
Grow better with oxygen
Use fermentation in absence of oxygen
E coil
Microaerophiles
Require oxygen in lower concentrations
Higher concentration inhibitory
Helicobacter pylori
Aerotolerant anaerobes
Indifferent to oxygen, grow with or without
Does not use oxygen to produce energy
Streptococcus

28. Environmental Factors - O2 availability
Decreasing O2

29. Environmental Factors - O2 availability

30. Environmental Factors on GrowthpH
Bacteria survive within pH range
Neutrophiles
Multiply between pH of 5 to 8
Maintain optimum near neutral
Acidophiles
Thrive at pH below 5.5
Maintains neutral internal pH pumping out protons (H+)
Alkalophiles
Grow at pH above 8.5
Maintain neutral internal pH through sodium ion exchange
Exchange sodium ion for external H+

31. Environmental Factors on Growth
Water availability
All microorganisms require water for growth
Water not available in all environments
In high salt environments
Bacteria increase internal solute concentration
Synthesize small organic molecules
Osmotolerant bacteria tolerate high salt environments
Bacteria that require high salt for cell growth termed halophiles

32. Nutritional Factors on Growth
Growth of prokaryotes depends on nutritional factors as well as physical environment
Main factors to be considered are:
Required elements
Energy sources
Growth factors

33. Nutritional Factors on Growth
Required elements
Major elements
Carbon, oxygen, hydrogen, nitrogen, sulfur, phosphorus, potassium, magnesium, calcium and iron
Essential components for macromolecules
Organisms classified based on carbon usage
Heterotrophs
Use organic carbon as carbon source
Autotrophs
Use inorganic carbon (CO2) as carbon source
Trace elements
Cobalt, zinc, copper, molybdenum and manganese
Required in minute amounts

34. Nutritional Factors on Growth
Energy Sources
Organisms derive energy from sunlight or chemical compounds
Phototrophs
Derive energy from sunlight
Chemotrophs
Derive energy from chemical compounds
Organisms often grouped according to energy source

35. Nutritional Factors on Growth
Nutritional Diversity
Organisms thrive due to their ability to use diverse sources of carbon and energy
Photoautotrouphs
Use sunlight and atmospheric carbon (CO2) as carbon source
Called primary producers (Plants)
Chemolithoautotrophs
A.k.a chemoautotrophs or chemolitotrophs
Use inorganic carbon for energy and use CO2 as carbon source
Photoheterotrophs
Energy from sunlight, carbon from organic compounds
Chemoorganoheterotrophs
a.k.a chemoheterotrophs or chemoorganotrophs
Use organic compounds for energy and carbon source
Most common among humans and other animals

36. Nutritional Factors on Growth
Growth factors
Some bacteria cannot synthesize some cell constituents
These must be added to growth environment
Referred to as growth factors
Organisms can display wide variety of factor requirements
Some need very few while others require many
These termed fastidious

37. Questions Major factors that affect bacteria growth Growth factor
Carbon source and energy source of chemoheterotroph

38. Detecting Bacterial Growth
Variety of techniques to determine growth
Number of cells
Total mass
Detection of cellular products

39. Detecting Bacterial Growth
Direct cell count
Plate count

40. Detecting Bacterial Growth
Direct microscopic count
Number is measured in a know volume
Liquid dispensed in specialized slide
Counting chamber
Viewed under microscope
Cells counted
Limitation
Must have at least 10 million cells per ml to gain accurate estimate

41. Detecting Bacterial Growth
Plate counts
Measures viable cells growing on solid culture media
Count based on assumption the one cell gives rise to one colony
Number of colonies = number of cells in sample
Ideal number to count
Between 30 and 300 colonies
Sample normally diluted in 10-fold increments
Plate count methods
pour-plates
Spread-plates methods