Microbial Growth Monitoring

The micro-organisms count can be performed by different methods: (a) serial dilutions; (b) Most Probable Number method – MPN; (c) Direct Microscope observation by traditional microscope- DM;  (d) Electronic cell count; (e) Optical Density – OD; (f) Metabolic activity; (g) Methylene blue dye (Reduction Test).

(a) Serial Dilutions

The sample is diluted out in a series of dilution tubes (9 ml) / bottles (90 or 99 ml) which are then plated out. The number of colonies (CFU) on the agar Petri dishes are counted and corrected for the dilution to calculate the number of micro-organisms in the original sample / inoculum.

(b) Most Probable Number – MPN

Several dilutions in test tubes are made with different initial volume of the sample / inoculum / specimen.

A statistical method is applied to estimate the most probable number of bacteria present in the test  tube dilutions. The results are recorded as a series of positive growth in the test tubes which can then be calculated to give a Most Probable Number in the sample.

(c) Direct Microscope observation (by traditional microscope)

The known amount of sample / specimen is filled inside a well of a special microscope slide with etched grid. The average number of micro-organisms in the bacterial suspension of each square grid is counted under the microscope and then multiplied by a factor to obtain the results per millilitre.

Selective staining is used to differentiate bacteria from non-living material.

(d) Electronic cell count

The electronic cell counter automatically counts the number of cells in a measured volume of liquid.

(e) Optical Density – OD. (Turbidity)

The turbidity of a liquid medium increases as bacteria multiply and can be measured on a spectrophotometer. The absorbency (optical density) of the sample is dependent on the number of cells, their shape and size.

(f) Metabolic activity

The amount of a metabolic product (e.g.: carbon dioxide, organic acids)   is measured and assumed to be proportional to the number of micro-organisms present in the sample / inoculum / specimen.

(g) Methylene blue dye (Reduction test)

The reduction test is based by the reduction of oxygen uptake using the methylene blue dye.

The blue dye changes colour from blue (presence of oxygen) to colourless (absence of oxygen).