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0:00

Overview

1:15

Principles of Bacterial Growth Curve Analysis

3:46

Collection of Bacterial Culture Aliquots

5:09

Serial Dilution Plating to Quantify Bacteria

7:19

Counting Colonies and Calculating Mean Generation Time

9:31

Applications

11:50

Summary

Bacterial Growth Curve Analysis and its Environmental Applications

Source: Laboratories of Dr. Ian Pepper and Dr. Charles Gerba - The University of Arizona
Demonstrating Author: Luisa Ikner

Bacteria are among the most abundant life forms on Earth. They are found in every ecosystem and are vital for everyday life. For example, bacteria affect what people eat, drink, and breathe, and there are actually more bacterial cells within a person’s body than mammalian cells. Because of the importance of bacteria, it is preferable to study particular species of bacteria in the laboratory. To do this, bacteria are grown under controlled conditions in pure culture, meaning that only one type of bacterium is under consideration. Bacteria grow quickly in pure culture, and cell numbers increase dramatically in a short period of time. By measuring the rate of cell population increase over time, a “growth curve” to be developed. This is important when aiming to utilize or inoculate known numbers of the bacterial isolate, for example to enhance plant growth, increase biodegradation of toxic organics, or produce antibiotics or other natural products at an industrial scale.

1. Collection of Bacterial Culture Aliquots

One day before collection of growth time points, inoculate 20 mL of trypticase soy broth (TSB) medium in a 50-mL flask with E. coli.
Incubate overnight at 27 °C. This relatively long incubation period results in a stationary phase population of wild type E. coli of approximately 10⁹ CFU/mL.
On the following day, use 100 µL of the prepared culture to inoculate 250 mL of TSB (in a 500-mL flask). Mix thoroughly

Following a serial dilution plating experiment, the following data was obtained. At the beginning of exponential growth designated here as time t = 0, the initial concentration of bacterial cells is 1,000 CFU/mL. At time t = 6 h, the concentration of cells is 16,000 CFU/mL.

Now, X = 2ⁿ x X₀

Where: X₀ = initial concentration of cells = 1,000 CFU/mL
X = concentration of cells after time t = 16,000 CFU/mL
n

Knowledge of bacterial growth kinetics and bacterial numbers in a culture medium is important from both a research and commercial point of view. In research, it is often critical to know the number of bacteria in a sample, so the experiment can be replicated, if need be, with the exact same numbers. For example, during experiments in which bacterial inoculants are added to a plot of soil, a minimum of 10⁴ CFU needs to be added per gram of soil to get the desired effect, such as enhanced biodegradation of toxic

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