Microbial Growth

Requirements
for Growth

Culture Media

Obtaining
Pure Cultures

Growth of
Bacterial Cultures

Measurement of
Bacterial Growth
Objectives:

16. List and describe the methods for the measurement of bacterial growth generally used in the lab.

The Growth Of Bacterial Cultures

 

Direct Measurement of Microbial Growth

A standard plate count reflects the number of viable microbes and assumes that each bacterium grows into a single colony. Because it is impossible to say that each colony actually arose from an indivual cell (cells clump, fact of life) plate counts are reported as the number of colony-forming units (CFU) instead of the number of cells.

 

If the concentration of bacteria is too great the colonies will grow into each other and the plate will be uncountable.

 

To insure a countable plate a series of dilutions should be plated.  The serial dilutions should give at least one countable plate in the series (25-250 or 30-300, depending on preference of the individual lab).

 

 

A plate count may be done on plates prepared by either the pour plate method or the spread plate method.

 

 

Some samples may be very dilute to begin with, so a plate prepared with undiluted inoculum may still have too few colonies to be countable.   In this case the concentration of bacteria can be increased by filtering the sample.

 

In filtration, bacteria are retained on the surface of a membrane filter and then transferred to a culture medium to grow and subsequently be counted.

 

 

The most probable number (MPN) method can be used for microbes that will grow in a liquid medium; it is a statistical estimation.

 

 

In a direct microscopic count, the microbes in a measured volume of a bacterial suspension are counted with the use of a specially designed slide.

 

 

Estimating Bacterial Numbers by Indirect Methods

 

A spectrophotometer is used to determine turbidity ("cloudiness") by measuring the amount of light that passed through a suspension of cells.

 

More cells = more turbidity; more turbidity = less light passing through the suspension

 

%T is percent transmission - fewer cells present (less turbidity) will allow more light to pass through, the %T is higher when the cell number is lower.

 

Absorbance is the opposite of %T.  More light is absorbed when more cells are present - some people like this measure better because absorbance goes up as turbidity (or cell number) goes up.

 

 

Another indirect way of estimating bacterial numbers is measuring the metabolic activity of the population (for example, acid production or oxygen consumption).

 

For filamentous organisms such as fungi, measuring dry weight is a convenient method of growth measurement.

 

You won't know how the measurement of turbidity with a spectrophotometer or measurement of metabolic activity or any other indirect measurement correlates to cell number unless you do a standard curve.

 

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