2. INTRODUCTION
โข Growth in a biological system is an orderly increase in the
quantity of cellular constituents and which depends upon the
ability of the cell to new protoplasm from nutrients available.
โข But, in bacteria growth involves increase in cell mass and no.
of ribosomes ,duplication of bacterial chromosomes,septum
formation and cell divisions.
โข Method of growth measurements involve both direct and
indirect techniques:-
A. Direct method
B. Indirect methods
3. DIRECT METHODS
1. Cell count
โข Haemocytometer
It is a simplest, most convenient and
cheapest method of determining the
no. of cell in a sample is to use a
haemocytometer and microscope. It
is a special slide that has counting
chambers with a known volume of
liquid. The engraved grid on the
surface of counting chambers ensures
the no. of particles in a defined
volume of liquid. The
haemocytometer is placed on the
microscope stage and cell suspension
is counted.
4. โข Viable plate count
Viable plate count is used to estimate
the no. of viable cells that are present
in a sample.
This method relies on bacteria
growing a colony on a nutrient
medium.
The colony become visible to the
naked eye and the no. of colonies on
a plate can be counted.
To be effective , on average between
30 and 300 colonies of the target
bacterium are grown.
5. โข Membrane filter count
Using the membrane filter technique,
sample is passed through the
membrane using a filter funnel and
vaccum system.
Any organism in sample is
concentrated on the surface of the
membrane.
The membrane is then placed in a
special plate with appropriate
medium.
The passage of nutrients through the
filter facilitates the growth of
microorganisms in the form of
colonies on the upper surface of
membrane.
Now, the discrete colonies can easily
be counted.
6. โข Direct microscopic count
In the direct microscopic count, a
counting chamber consists of ruled
slide and coverslip.
Number of bacteria in small known
volume is directly counted
microscopically and the no. of
bacteria in original sample is
determined by extrapolation.
Petroff-Hausser counting chamber
is commonly used in dairy
industry.
7. 2. Cell mass
โข Wet weight
A known volume of culture sample is centrifuged, then , the wet
weight of pellet is measured by using pre weighed filter paper.
โข Dry weight
A known volume of culture sample is centrifuged , pellet is dried by
removing water and then dry weight of pellet is measured by
using pre weighed filter paper.
8. โข Total nitrogen content
Major constituent of cell material is protein and since nitrogen is a
characteristic part of protein , bacterial population can be
measured in terms of bacterial nitrogen.
Bacteria average approx. 14% N2 on a day weight basis .
To measure growth by this technique, first harvest the cells and
wash them free of medium and perform a quantitative chemical
analysis of nitrogen.
9. โข Turbidity
When bacteria growing in a liquid
medium are mixed , the culture
appears turbid.
This is because a bacterial
culture act as a colloidal
suspension that block and
reflects light passing through
culture.
Light absorbed by the bacterial
suspension will be directly
proportional to the concentration
of the cells in the culture.
By measuring the amount of light
absorbed one can estimate the
number of bacteria present.
The instrument used to measure
turbidity is spectrophotometer.
10. INDIRECT METHODS
The bacterial growth can be indirectly estimated by detecting
specific changes caused in growth medium as a result of activity
and multiplication of bacterial cells. It includes detecting activity
cell products such as acid and gas production.
11. โข OXYGEN CONSUMPTION
The dye reduction tests such as methylene blue and resazurin
reduction tests is based on the fact that the color imparted to milk
by the addition of a dye such as methylene blue will disappear
more or less quickly.
The removal of the oxygen from milk and the formation of reducing
substances during bacterial metabolism cause the color to
disappear.
The agencies responsible for the oxygen consumption are the
bacteria.
Thus, the time of reduction is taken as a measure of the number of
organisms in milk although actually it is likely that it is more truly a
measure of the total metabolic reactions proceeding at the cell
surface of the bacteria.
12. โข CO2 PRODUCTION
Gas production by bacteria is
another major activity which
can be taken up as an index of
bacterial growth.
Detection of gas production
using Durham tube and change
in color of the growth medium
due to reduction of pH
sensitive ingredients present in
medium are commonly used
for detection of acid and gas
producing coliforms and
yeasts.
