Direct- Requiring Incubation:
Serial Dilution and viable plate count- used if number of cells in a sample is TOO GREAT TO COUNT. Dilution of a liquid culture, then count the colonies on plates with 25-250 visible colonies and then multiply that number by the reciprocal dilution to estimate the number of bacteria in the original culture.
Membrane Filtration: USED IF POPULATION DENSITY IS VERY SMALL. Large sample is poured and filtered through a membrane. Then the membrane is transferred to a solid medium and colonies present after incubation are counted.
Most Probably Number (MPN)- Statistical estimation based on the fact that the more bacteria there are in a solution, the more dilutions are required to reduce their numbers. Typically it starts at 5 tubes and goes to 15 to dilute. This method works well with microorganisms that do not grow in solid media, when bacteria require counts routinely, and when samples are too few to do a plate count. Also used with algal cells.
Direct- NOT Requiring Incubation:
Microscopic Counts- Used with stained prokaryotes and larger eukaryotes. A cell counter is used in conjunction with a microscope. Several squares are counted and then multiplied by remaining square numbers to estimate.
Electronic Counters: Coulter counter device directly counts cells as they interrupt an electrical current. Counts larger cells of yeast, unicellular algae and protozoa. Flow cytometer is another counter that uses light sensitivity. It is used to distinguish among cells that have differential fluorescent stains. They can count bacteria in a solution.
It is not always necessary to count microorganisms to estimate population or density. Indirect measures other variables.
Metabolic Activity: Under standard temperature conditions, the rate at which a population of cells uses nutrients and produces wastes is dependent on their number. Once we know their metabolic rate, we can measure the rest to calculate population.
Dry Weight: Used particularly with filamentous microorganisms. Organisms are filtered from their culture medium, dried and weighed. It is suitable for broth cultures.
Turbidity (cloudy): The greater the population, the cloudier the broth. Measures changes in turbidity using a spectrophometer that measures the amount of light transmitted through a culture. This method is easy and fast. It is only useful is the number of cells exceeds 1 million per mL. If a pedicle or sediment form, the number will be underestimated. It does not distinguish between living and dead cells.
Genetic Methods: Isolating of unique DNA sequences representing unculturable prokaryotic species using genetic techniques.
9th EditionCain, Jackson, Minorsky, Reece, Urry, Wasserman
11th EditionLisa A. Urry, Michael L. Cain, Peter V Minorsky, Steven A. Wasserman
5th EditionCharlotte W. Pratt, Donald Voet, Judith G. Voet
1st EditionDay, Stewart