1. GROWTH AND CULTURING OF BACTERIA
CHAPTER 6

2. Growth and Cell Division
Growth Definition
Binary Fission
Budding

3. Bacterial Growth Curve
Lag phase
Adjustment to nutrients
Enzyme synthesis
DNA synthesis
Log phase
Exponential cell division
Stationary phase
Nutrient depletion
Waste build-up
Death phase
More die than are produced
Always a few survivors

4. Counting Large Populations
Serial Dilutions/ Standard plate counts
Very large numbers may be counted by this method
Pour plate or spread plate techniques are used

5. Direct Counts
Colony Counts
300 upper limit
Hemocytometer

6. Counting Small Populations
Most Probable Number
Filtration

7. Other Measurements
Spectrophotometer
Tube turbidity

8. Physical Influences (pH)
Bacteria
Prefer pH 5.4 – 8.0
Most grow at pH 7.0 – 7.4
Rare acidophiles and alkaliphiles exist
Lactobacillus
Vibrio cholerae
Fungi
Can tolerate pH 5-6
Often the only cause of acidic food spoilage
Fruit and vegetable juices/products

9. Physical Influences (Temperature)
Psychrophiles 0oC – 20oC
Refrigerator spoilage
Listeria monocytogenes
Mesophiles 10oC – 45oC
Food / Disease
Thermophiles 40oC – 70oC
Soil composters
Extreme Thermophiles 70oC – 100oC
Archaea

10. Oxygen’s Influence on Growth
OBLIGATE AEROBE (Micrococcus)
Must use oxygen
OBLIGATE ANAEROBE (Clostridium)
Must have anaerobic environment
Why?
Lack enzymes to degrade oxygen radicals
Superoxide dismutase (O2-)
Catalase (H2O2)
MICROAEROPHILE (Campylobacter)
Low oxygen, high carbon dioxide environment
Respiration pathway may contain unique electron carriers
FACULTATIVE ANAEROBE (Escherichia coli, Staphylococcus)
Use oxygen when it is present
Can also grow anaerobically
AEROTOLERANT ANAEROBE (Lactobacillus)
Tolerate oxygen
Do not use oxygen

11. Growth positions in Thioglycollate medium

12. Physical Influences on Growth
Moisture
Endospores
Fungal preferences
Hydrostatic pressure
Barophiles
Undersea vents
Osmotic pressure
High salt/sugar used as food preservatives
Halophiles – salt tolerant
Radiation (Xrays, UV, γrays)
Deinococcus radiodurans
DNA repair systems

13. Nutritional Factors Affecting Bacterial Growth
C, N, S, P availability
Organic versus inorganic
Trace elements
Minerals for enzyme activity
Vitamins
Organics not synthesized by the cell
Nutritional complexity
Determined by the enzyme capabilities of the microbe
Fewer enzymes = more nutritional needs (or complexity)

14. Bacterial Interactions
Quorum sensing
Biofilms
Antibiotic resistances
Diseases
Tuberculosis
Cystic Fibrosis
Diabetic Foot Ulcers
Toxin production to limit competitor “groups”
Sociomicrobiology

15. Culturing – Plate techniques
Streak Plates
¼ “ spacing minimum for well isolated colonies

16. Culturing Plate Techniques
Pour Plates

17. Culture Media ? Types Defined Precise concentrations
Research into new microbes
Complex
Nutritionally sufficient
Unknown molecular composition
? Beef extract

18. Culture Media Purposes
Selective media
Suppress unwanted
Differential media
Distinguishes
Enrichment media
Favors fastidious microbes
Propagation media
General growth

19. Controlling Oxygen Content
Brewer jar
Anaerobic environment
Chemicals absorb oxygen
Candle jar
Burning a candle creates low O2, high CO2 environment

20. Applications of Culturing
Maintaining cultures
Preserved cultures
Diagnostic tests
Limitations on Culture