Quantitative Measurement Of Bacterial Growth (Total & Viable Count)

Quantitative Measurement of Bacterial Growth (Total & Viable Count)

In the field of microbiology, the quantitative measurement of bacterial growth is a fundamental practice that provides critical insights into microbial populations. Whether you’re studying the effectiveness of antibiotics or monitoring bacterial contamination in pharmaceuticals, understanding how to accurately measure bacterial growth is essential. This article delves into the methods used to quantify bacterial growth, focusing on both total and viable counts, and highlights their importance in various applications.

Table of Contents

Total Count Methods for Quantitative Measurement of Bacterial Growth

Total count methods aim to quantify the entire bacterial population in a sample, including both living and dead cells. Here are the primary techniques used:

Direct Microscopic Count

Description: This method involves counting bacteria directly under a microscope using a specialized counting chamber, such as a Petroff-Hausser counting chamber.

Procedure:

A small volume of the bacterial suspension is placed on the counting chamber.
The chamber has a grid etched on its surface, which helps in counting the cells.
The sample is observed under a microscope, and the number of bacteria in several grid squares is counted.
The average count is then used to calculate the total number of bacteria per unit volume.

Advantages:

Quick and straightforward.
Provides an immediate count of bacteria.

Disadvantages:

Cannot distinguish between live and dead cells.
Requires a high concentration of bacteria for accurate counting.
Potential for human error and subjectivity.

Electronic Enumeration (Coulter Counter)

Description: This method uses an electronic device, such as a Coulter Counter, to count cells as they pass through a small orifice.

Procedure:

The bacterial suspension is passed through a small aperture.
As each cell passes through the aperture, it disrupts an electrical current.
These disruptions are counted electronically, providing a total count of cells.

Advantages:

Automated and rapid.
Reduces human error and subjectivity.

Disadvantages:

Expensive equipment.
Cannot differentiate between live and dead cells.
May require sample preparation to avoid clogging the aperture.

Turbidity Measurement

Description: This indirect method measures the cloudiness (turbidity) of a bacterial suspension using a spectrophotometer.

Procedure:

The bacterial suspension is placed in a cuvette.
The spectrophotometer measures the amount of light that passes through the suspension.
The turbidity, which correlates with the bacterial population, is recorded as optical density (OD).
A standard curve is used to convert OD values to bacterial counts.

Advantages:

Quick and easy.
Suitable for monitoring bacterial growth over time.

Disadvantages:

Indirect method; requires calibration with a direct counting method.
Cannot distinguish between live and dead cells.
Less accurate at very high or low bacterial

Viable Count Methods for Quantitative Measurement of Bacterial Growth

Plate Count Method

Description: This is one of the most commonly used methods to determine the number of viable bacteria in a sample. It involves spreading a diluted bacterial sample on an agar plate and counting the colonies that form after incubation.

Procedure:

Prepare a series of dilutions of the bacterial sample.
Spread a measured volume of each dilution onto the surface of an agar plate.
Incubate the plates at an appropriate temperature for a specified period.
Count the number of colonies that form on each plate.
Calculate the number of viable bacteria in the original sample based on the dilution factor.

Advantages:

Provides a direct count of viable bacteria.
Can be used to isolate and identify different bacterial species.

Disadvantages:

Time-consuming due to the incubation period.
Only counts bacteria that can grow under the specific conditions provided.

Most Probable Number (MPN) Method

Description: The MPN method is a statistical technique used to estimate the number of viable bacteria in a sample. It is particularly useful for samples with low bacterial counts or when the bacteria cannot be easily cultured on solid media.

Procedure:

Prepare a series of dilutions of the bacterial sample.
Inoculate multiple tubes of liquid media with each dilution.
Incubate the tubes and observe for bacterial growth (e.g., turbidity or gas production).
Use statistical tables to estimate the number of viable bacteria based on the pattern of positive and negative tubes.

Advantages:

Useful for samples with low bacterial counts.
Can be used for bacteria that are difficult to culture on solid media.

Disadvantages:

Less precise than plate counts.
Requires multiple tubes and media, making it more labor-intensive.

Membrane Filtration

Description: This method involves filtering a known volume of the bacterial sample through a membrane that traps bacteria. The membrane is then placed on an agar plate, and the colonies that form are counted.

Procedure:

Filter a measured volume of the bacterial sample through a membrane filter.
Place the membrane on the surface of an agar plate.
Incubate the plate at an appropriate temperature for a specified period.
Count the number of colonies that form on the membrane.

Advantages:

Suitable for samples with low bacterial counts.
Can filter large volumes of liquid, concentrating the bacteria for easier counting.

Disadvantages:

Requires specialized equipment (membrane filters and filtration apparatus).
Only counts bacteria that can grow on the specific agar used.

Conclusion

Quantitative measurement of bacterial growth is crucial in microbiology. By using total count methods to count all bacteria and viable count methods to count only the living ones, we gain valuable insights into bacterial populations. These techniques are essential for various fields, including medicine, pharmaceuticals, and environmental studies. Understanding these methods helps students and professionals make informed decisions and improve their work in microbiology.

Quantitative Measurement of Bacterial Growth (Total & Viable count)