Flashcard Answers on Micro Bio: Microbiology
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Unlock answersGrowth of Bacterial Populations |
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 |
Generation time |
o Time to double o Factors § Temperature § pH § Nutrients § Properties of species
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Bacterial Growth Curve
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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 |
Lag Phase |
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 |
Log Phase
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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 |
Stationary Phase
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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 |
Death Phase |
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 |
Synchronous Growth
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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 |
Continuous growth
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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 |
Measuring microbial growth
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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 |
Standard (Viable) Plate counts
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· 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 |
Membrane filtration counts
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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 |
Microscopic counts
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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 |
Electronic particles count
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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 |
Most Probable Number Method
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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 |
Turbidity
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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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Dry weight
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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 |
Measuring metabolic product
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o Nitrogen or protein content or oxygen uptake measured indirectly o Tedious to carry out
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