Microbial Enumeration Flashcards
Methods of Microbial Enumeration
- Direct
- Indirect
Direct Methods of Microbial Enumeration
- Viable counting
- Serial Dilution
- Spread Plate and Pour Plate
- Filtration
- Direct Microscopic Count
Viable count/Plate count Method
- Spread Plate
- Pour Plate
It is the most frequently used method of measuring bacterial populations
Viable count/Plate count
Advantage of Plate count
It measure the number of viable cells
Disadvantage of Plate count
It takes some time, usually 24 hours or more, for visible colonies to form
A colony often results, not from a single bacterium, but from short segments of a chain or from a bacterial clump. To reflect this reality, plate counts are often reported as
colony-forming units (CFU)
What causes inaccurancies in using viable count method?
- too many colonies are present
- some cells are overcrowded and do not develop
original inoculum that is diluted
several times
Serial Dilution
- put a 1 ml or 0.1 ml of dilution of the bacterial suspension into the petri dish
- the agar liquid is then poured into the plates which is then mixed into the medium by gentle agitation of the plate
Pour Plate
- a 0.1-ml inoculum is added to the surface of a prepoured, solidified agar medium
- the inoculum is then spread unformaly over the surface using a metal rod
- this method positions all the colonies on the surface and avoids contact between the cells and melted agar
Spread Plate
When the quantity of bacteria is very small, as in lakes or relatively pure streams, bacteria can be counted by
Filtration method
Filtration method is applied frequently to
detection and enumeration of coliform bacteria, which are indicators of fecal contamination of food or water
In this method a measured volume of a bacterial suspension is placed within a defined area on a microscope slide
Direct Microscopic Count
Direct Microscopic Count is often used to
count the number of bacteria in milk
A ______ sample is spread over a marked square centimeter of slide, _____ is added so that the bacteria can be seen, and the sample is viewed under the ___ _________ objective lens
- 0.01 ml
- stain
- oil immersion
- a specially designed slide
- used in direct microscopic count
Petroff-Hausser cell counter
Disadvantages of Direct Microscopic Count
- motile bacteria are difficult to count
- a rather high concentration of cells is required to be countable
Advantage of Direct Microscopic Count
no incubation time is required, usually reserved for applications in which time is the primary consideration
- automatically count the number of cells in a measured volume of liquid
- used in some research laboratories and hospitals
Electronic cell counters or Coulter counters
Indirect Methods of Mirobial Enumeration
- Most Probable Number (MPN)
- Turbidity
- Metabolic Activity
- Dry Weight
- a method for determining the number of bacteria in a sample
- statistical estimating technique is based on the fact that the greater the number of bacteria in a sample, the more dilution is needed to reduce the density to the point at which no bacteria are left to grow in the tubes in a dilution series
Most Probable Number (MPN)
When is the MPN most useful?
when the microbes being counted will not grow on solid media (such as the chemoautotrophic nitrifying bacteria)
MPN is also useful when
the growth of bacteria in a liquid differential medium is used to identify the microbes (such as coliform bacteria, which selectively ferment lactose to acid, in water testing)
