Term
Growth of Bacterial Populations |
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Definition
o Unicellular
o Reproduce by Binary fission
§ Cell increases size
§ Replicates DNA
§ Divides into genetically identical daughter cells of same size
o Daughter cells grow and divide at same rate
o Based off 2x
o Exponential growth
o Plotted on a semi-log scale à straight line |
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Term
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Definition
o Time to double
o Factors
§ Temperature
§ pH
§ Nutrients
§ Properties of species
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Term
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Definition
o Bacterial growth vs time
§ Sterile broth culture inoculated
§ Growth measured by turbidity with a spectrophotometer at time intervals
§ Measured turbidities converted to log and plotted as log10turbidity vs time
o Four growth phases
§ Lag phase
§ Log phase
§ Stationary phase
§ Death phase
§ Figure 6c-7 shows phases |
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Term
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Definition
o Do not multiply immediately upon inoculation
o Factors
§ Properties of bacteria species
§ Age and size of inoculum
§ Nutrients and pH of new medium
§ Nutrients and pH of old medium
§ Whether essential enzymes or other molecules need to be synthesizes before growth |
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Term
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Definition
o After lag à log phase
o Cell number doubles each generation
o Positively sloped straight line
o Generation time calculated by determining doubling time during log phase |
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Term
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Definition
o Culture cannot continue log phase forever
§ Exhaustion of nutrients
§ Changes in ph
§ Insufficient O2
§ Accumulation of inhibitory waste products
o Stop growing, remain viable
o After 24 hrs enter stationary phase
§ Deaths = new cells |
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Term
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Definition
o Death rate exceeds new cell formation
o Some die quickly in spend medium, others take months
o Old cells assume unusual shapes and may not Gram stain properly |
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Term
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Definition
o All cells of population in same stage
o Normally any stage contains same number of cells due to random nature
o Special techniques to induce synchronous growth
o Useful to study division process and cell cycle |
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Term
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Definition
o All cells in log phase
o Maintained by
§ Adding fresh medium
§ Maintaining optimal oxygen and pH levels
§ Continually removing some cells and spend medium
o Carried out in chemostat |
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Term
Measuring microbial growth
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Definition
o Direct methods - Determine number of cells
§ Standard (viable) plate counts
§ Membrane filtration counts
§ Microscopic counts
§ Electronic counter counts
§ The most probable number method
o Indirect methods – mass of population
§ Turbidity
§ Dry weight of culture
§ Metabolic product of culture |
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Term
Standard (Viable) Plate counts
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Definition
· Most frequently used method to count cells
· Measures viable cells only
· Two assumptions
o Each live cell will grow on agar
o Each visible colony derived from single cell
· Make serial dilutions
· Plate each on an agar medium
· Too many bacteria à too hard to count
· Too few bacteria à not statistically significant
· Serial 1/10 dilutions will give you one level that is countable at least
· Find plate, count, multiply by reciprocal of dilution to estimate full population
· Two methods
o Pour plate
§ Pour bacteria, pour agar, swirl
o Spread plate
Pour agar, solidify, spread bacteria |
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Term
Membrane filtration counts
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Definition
o Determine microbial cells in air or water
o Only works on few microbes per volume
o 50ml to 1 L passed through .2um filter, then cultured by placing it right on agar |
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Term
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Definition
o Special ruled glass slide – Petroff-Hauser Counter
§ Allows .02 mm3 in a ruled 1 mm2 area
o Countable range
§ 107 -108 cells per ml
§ Tedious to count and measure viable and dead |
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Term
Electronic particles count
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Definition
o Coulter counters
o Count cells in sample
o Give poor results for prokaryotic cells
o Better results with larger cells like white blood cells |
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Term
Most Probable Number Method
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Definition
o Estimate coliform bacteria in a water sample
o Coliforms – gram-negative, facultatively anaerobic rods that ferment lactose to acids and gas
§ Majority of those found in large intestine and feces
§ Found in water à contaminated with feces
o MPN uses statistics to estimate number of cells using serial dilutions
o 3 concentrations tested in quintuplet
o Number of tubes that turn yellow counted
o Statistics estimates level of cells |
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Term
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Definition
o Optical density (turbidity) is the Simplest and most frequently used indirect method for liquid cultures
o Resuspended in culture medium
o Sample taken with spectrophotometer
o Accurately measured between .01 to 2.0 O.D. units
§ 108 – 1011 cells per ml
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Term
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Definition
o Indirectly measured by calculating dry weight of microbial cells in known volume of culture medium
o Samples pulled out at regular intervals
§ Centrifuged out of solution
§ Washed
§ Centrifuged
§ Resuspended
§ Frozen
§ Lyophilized (freeze-dried)
§ Weighed
o Tedious to carry out
§ Used for fungi and filamentous organisms that grow on surface of liquid media |
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Term
Measuring metabolic product
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Definition
o Nitrogen or protein content or oxygen uptake measured indirectly
o Tedious to carry out
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