Bacterial Cultivation & Growth
• Culturing Microorganisms
– Binary fission & sporulation
– What determines growth?
– Media Types
– Batch vs Continuous Culture Growth
Binary Fission in Prokaryotes
Dividing vegetative cells
But what happens when stressed or starved?
Endospores:
• Resting stage during “lean
or stressful times”.
• Resistant protein coat!
• Develop in different
locations of vegetative cell:
free;
sub-terminal;
central;
terminal
•Schaeffer – Fulton Stain:
Young (24 h)
Old (96 h)
Endospores
& Sporulation
(cortex)
Spore coat
proteins resist
toxic chemicals.
Dipicolinic Acid &
Calcium protects
DNA from heat.
exosporium
Spore Germination:
activation; germination; outgrowth
Culturing Microorganisms
• Tolerance to All Environmental Factors (Shelford’s
Law of Tolerance)
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Temperature
Solute Concentration / Water Activity
pH (acidity versus alkalinity)
Oxygen Concentration
Barometric Pressure
Electromagnetic Radiation
• Growth Limiting Resource (Liebig’s Law of the
Minimum): Nutrient in least supply relative to
bacterial needs will cap growth yield.
Closed (“batch”) Culture Systems
“growth curve”
Unbalanced
Growth
Balanced Growth:
Rates of RNA =
Protein = DNA =
binary fission
Exponential Growth Phase
Nt = No + 2n
Number of generations (n) = (log Nt – log No) / log 2
Growth Rate Constant (k) = n/t
It is expressed in units of generations per hours (h-1)
Generation time (g) = 1/k; it is expressed in units of hours (h).
k = (log Nt – log No) / 0.301 t
Generation time (g) = doubling time
Time (hours)
Nutrient Concentration Effects in Batch Cultures:
• Total growth will increase until limiting nutrients are exhausted
(included oxygen for aerobes) or metabolic byproducts accumulate
that change environmental conditions to inhibit growth (toxicity).
• Growth rate will also increase with increasing nutrient concentration
up to a some maximum value, beyond which there is no effect
(transporters are saturated with there substrate.
Open (“continuous”) Culture Systems
Chemostat: growth rate = dilution rate (D = f/V);
constant dilution rate with nutrient limiting growth.
Turbidostat: dilution rates varies to maintain constant
turbidity (cell density); no limiting nutrient.
Flow (f)
Volume (V)
Wash
out!