Thiosulfate-citrate-bile salts-sucrose agar
The TCBS agar is a nutrient medium for the selective isolation and cultivation of bacteria of the genus Vibrio, such as the causative agents of cholera - generate and other vibrios, the vomiting and diarrhea disease ( gastroenteritis), such as Vibrio parahaemolyticus, or the so - Vibrio cholerae called NAG vibrios ( non- agglutinating vibrios ), where it is non -O1 strains of Vibrio cholerae.
TCBS Agar Thiosulfate Citrate Bile stands for ( Salts ) sucrose agar and refers to the most important components of the nutrient medium: sodium thiosulfate, sodium citrate, bile salts ( in English bile for bile) and sucrose ( sucrose in English ). The original (1962 ) published the recipe of Japanese Y. Nakanishi was modified a year later by T. Kobayashi.
Applications
This medium is recommended by the American Public Health Association for the microbiological examination of foods. Furthermore, used in food microbiology test methods for Vibrio cholerae and other Vibrio species by the ISO 21872 and in the U.S. by the Bacteriological Analytical Manual ( BAM ) of the Food and Drug Administration (FDA ) - the U.S. Agency for Food and pharmaceutical drug - prescribed, and through them the use of TCBS agar is also recommended.
For samples such as water or food, first a accumulation of vibrios in alkaline peptone water. This broth has a high concentration of sodium chloride and an alkaline pH - value of pH 8.5, the growth of a number of other bacteria is inhibited by these two parameters. If the cultivation is carried out in this medium at 42 ° C, the enhancement is even more selective, since other mesophilic bacteria are growth inhibited by the high temperature. For the isolation of Vibrio species a small volume of nutrient broth on TCBS agar was plated after enrichment. In clinical samples, such as Stool samples or vomit can they be shared with a swab on the nutrient medium. In addition to the selective medium TCBS agar also a little selectively acting nutrient medium should be inoculated.
Operation
TCBS Agar Thiosulfate Citrate Bile stands for ( Salts ) sucrose agar and refers to the most important components of the nutrient medium: High concentrations of sodium thiosulfate and sodium citrate largely inhibit the growth of Gram-negative Enterobacteriaceae, which are also present for example in stool samples. Ox bile ( bile in English ) and sodium cholate ( the sodium salt of cholic acid, a bile acid ) inhibits the growth of gram- positive microorganisms, v. a enterococci. Sucrose ( sucrose, in English ) is the only carbohydrate in TCBS agar and restricts the growth of bacteria, which are unable to utilize sucrose. Together with the accompanying pH indicators thymol blue and bromothymol the degradation of carbohydrate by Vibrio species on the acid formation can be visualized, while a color change of the pH indicator from blue ( alkaline pH) to yellow occurs (acidic pH).
Uninoculated, the medium to an alkaline pH ( pH 8.6 ), also takes place through the inhibition of growth of other bacteria. Through the accompanying pH indicators it is colored blue-green. The one with the TCBS agar to be isolated vibrios are halophilic ( " salt- loving" ), that can be cultured in or on nutrient media with increased salt concentration, so that the high concentration does not impair sodium chloride and other salts in the medium to grow.
Evaluation
The Petri dishes containing the inoculated TCBS agar is incubated for 18 to 24 hours at 35 ° C under aerobic conditions. Vibrio cholerae forms on the nutrient medium flat, yellow, 2-3 mm large colonies, Vibrio alginolyticus large yellow colonies, and also the medium around the colonies is colored yellow by acid formation. V. parahaemolyticus, however, is smaller colonies with a blue-green center.
In addition to the vibrios, there are only a few Proteus strains that can utilize sucrose with acid formation and similar form colonies such as Vibrio species. However, they are, like other bacteria from the family Enterobacteriaceae inhibited by other ingredients in the nutrient medium and therefore show only weak or no growth. Bacteria of the genus Pseudomonas only grow and can be inhibited also by the blue colony color of the vibrios differ because they are unable to utilize sucrose with acid formation. On TCBS agar grown colonies need to be further investigated for confirmation and differentiation of Vibrio species, for example, by biochemical tests of a "colorful series", the assignment to the serotypes of Vibrio cholerae can be done by means of an agglutination test.
Typical Composition
The soil mostly consists of ( in grams per liter):
- Peptone 10.0
- Yeast extract 5.0
- Sodium thiosulfate 10.0
- Sodium citrate 10.0
- Oxgall 5.0
- Sodium cholate 3.0
- Sucrose 20.0
- Sodium chloride 10.0
- Iron (III) citrate 1.0
- Agar- agar 14.0