pre

Thiosulfate–citrate–bile
salts–sucrose agar

Thiosulfate–citrate–bile salts–sucrose agar (TCBS agar) is a selective agar

medium used for the isolation of Vibrio cholerae and other enteric pathogens. In
this presentation, we will explore the composition, preparation, and uses of
TCBS agar.

Presenters:
Salman ,Kainat bb
,Irshadullah, Asim khan
,Abbas Ahmed
Introduction
What is TCBS agar?
TCBS agar is a culture medium used to
isolate Vibrio cholerae bacteria.
History of TCBS agar
Developed by Kobayashi et al. in 1963,
TCBS agar was first used to select for
Vibrio cholerae in water samples.
How does TCBS agar work?
TCBS agar contains bile salts, which
inhibit the growth of most Gram-positive
bacteria, and sucrose, which fosters the
growth of TCBS-fermenting bacteria.
Why is TCBS agar important?
TCBS agar is an essential tool used in the
detection and diagnosis of Vibrio cholerae,
particularly in developing countries where
cholera outbreaks are prevalent.
Purpose
Isolating Vibrio
cholerae
The primary purpose of TCBS
agar is to isolate Vibrio
cholerae bacteria from
environmental samples or
human stools.
Counting colony-
forming units (CFUs)
TCBS agar is also used to
calculate the number of Vibrio
cholerae colony-forming units
in a given sample.
Preventing the spread of
cholera
TCBS agar plays a critical
role in cholera prevention
efforts by enabling the rapid
identification and tracking of
Vibrio cholerae outbreaks.
Composition

1 Peptone
Provides nitrogen and other
essential nutrients for bacterial
TCBS Selective 2 growth
Supplements
Include bile salts, and sodium
thiosulfate, which inhibit the growth
of most bacteria and enrich the 3 Sucrose
growth of vibrios respectively Provides a fermentable carbohydrate
source for the detection of TCBS-
fermenting bacteria
Sodium chloride and pH 4
indicator
Help maintain the pH The pH of
TCBS Agar is set at 8.6 and indicate
the fermentation of sucrose by
changing color from green to yellow
Uses
1 Diagnosing cholera
TCBS agar is an essential tool in the
diagnosis of cholera, particularly in
developing countries.
3 Epidemiological investigations
TCBS agar is used to identify the source
and transmission routes of outbreaks and
to study the ecology and diversity of
Vibrio species
2 Environmental monitoring
TCBS agar is used to detect and isolate
Vibrio cholerae from water, seafood, and
other environmental samples.
4 Research and development
TCBS agar is used in vaccine and drug
development, as well as in the field of
metagenomics to understand the
microbiome ecology of coastal
environments.
Preparation and incubation
Preparation Incubation
1. Add 22 grams of TCBS agar powder to • Inoculate the petri dish with a sample
1000 ml of distilled water in a flask. using a sterile loop or swab.

2. Boil until the powder has dissolved. • Incubate the plate at 37°C for 24-48 hours.
3. Autoclave is not essential ,it will • Look for yellow colonies with a black
degrade some ingredients center as an indication of Vibrio

4. Pour the sterilized agar into petri cholerae growth.

dishes and let it cool to room

temperature.
Limitations
False-positive results Low sensitivity
Unusual bacteria or contaminants can grow TCBS agar is not ideal for detecting low
on TCBS agar, leading to false-positive levels of Vibrio cholerae in fecal samples.
results and misidentification of Vibrio
cholerae.

Environmental biases Difficult to interpret

TCBS agar is selective only for vibrios and TCBS agar can be challenging to interpret
may not be sensitive to other pathogens due to the wide range of Vibrio species and
present in the sample. morphological variations in colonies.
 TCBS Agar Principle:
TCBS is a selective and differential medium used to isolate and identify bacteria like Vibrio
cholerae. It contains thiosulfate, citrate, bile salts, and sucrose which help in the growth of
Vibrio species. It's commonly used in clinical and research laboratories.

• The high concentration of thiosulphate and citrate and higher alkalinity of this medium
largely inhibits the Enterobacteriaceae growth.
• Bile and sodium citrate inhibit gram-positive bacteria and coliforms
• Yeast extract and peptones provide essential growth nutrients like nitrogen, vitamins,
and amino acids for growth of bacteria.
• When sucrose is fermented, acid is produced, and pH drops which with the help of
bromothymol blue and thymol blue Turns the media yellow from blue.
• NaCl provides optimum growth and metabolic activity of halophilic Vibrio species.
• Agar is solidifying Agent
Conclusion
Thiosulfate–citrate–bile salts–sucrose agar is a vital tool in the detection,
diagnosis, and prevention of cholera. Although it has some limitations, TCBS
agar remains the gold standard for isolating and identifying Vibrio cholerae from
environmental samples and human stools. Understanding the composition,
preparation, and uses of TCBS agar is essential for anyone involved in public
health or microbiology.
Quality Control on TCBS Agar
The stringent quality control measures on TCBS Agar ensure accurate results and promote reliability in identifying
specific microbial species.

Positive Control = Vibrio parahaemolyticus ATCC 17802 (Growth,

blue green)

Negative Control = Enterococcus faecalis ATCC 29212 (No Growth)