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Micro Bio
6-C Methods for Measuring Microbial Growth
18
Microbiology
Professional
10/21/2011

Additional Microbiology Flashcards

 


 

Cards

Term

Growth of Bacterial Populations

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

Term

Generation time

Definition

 

o   Time to double

o   Factors

§  Temperature

§  pH

§  Nutrients

§  Properties of species

  • Most bcteria generation time is between 20-120 minutes
Term

 

Bacterial Growth Curve

 

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

Term

    Lag Phase

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

Term

 

Log Phase

 

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

Term

 

Stationary Phase

 

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

Term

Death Phase

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

Term

 

Synchronous Growth

 

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

Term

 

Continuous growth

 

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

Term

 

Measuring microbial growth

 

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 methodsmass of population

§  Turbidity

§  Dry weight of culture

§  Metabolic product of culture

Term

Standard (Viable) Plate counts

 

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

Term

Membrane filtration counts

 

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

Term

 

Microscopic counts

 

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

Term

 

Electronic particles count

 

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

Term

 

   Most Probable Number Method

 

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

Term

 

Turbidity

 

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

 

 

Term

Dry weight

 

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

Term

Measuring metabolic product

 

Definition

 

o   Nitrogen or protein content or oxygen uptake measured indirectly

o   Tedious to carry out

 

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