USE OF TRIPLE SUGAR IRON AGAR (AS PER BIS)

TRIPLE SUGAR IRON AGAR (AS PER BIS) for confirmation of gram negative enteric bacilli on behalf of dextrose, lactose and sucrose fermentation and H2S production.

Composition 

Ingredients	Gms/Ltr
Peptone	20.000
Yeast extract	3.000
Meat extract	3.000
Lactose	10.000
Sucrose	10.000
Dextrose	1.000
Sodium chloride	5.000
Ferrous sulphate, heptahydrate	0.200
Sodium thiosulphate, pentahydrate	0.300
Phenol red	0.024
Agar	12.000

* Dehydrated powder, hygroscopic in nature, store in a dry place, in tightly-sealed containers below 25°C and protect from direct Sunlight.

Instructions for Use

Suspend 64.32 grams in 1000 ml refined water. Gently heat to boiling with gently swirling to dissolve the medium completely. Sterilize by autoclaving at 15 psi pressure (121°C) for 15 mins. Allow the medium to cool at room temperature for preparing the slants.

Appearance: Pinkish - red colour, clear to slightly opalescent gel in slant position in screw cap tubes

pH (at 25°C) : 7.4 ± 0.2.

Principle

Triple Sugar Iron Agar was originally proposed by Sulkin and Willet and modified by Hajna for identifying Enterobacteriaceae. ISO Committee and BIS has recommended a slight modification for the identification of Salmonellae. BIS has recommended the medium for detection of Escherichia coli and Vibrios.

Triple Sugar Iron Agar is recommended for confirmation of gram negative bacilli on basis of dextrose, lactose and sucrose fermentation and H2S production. The ingredients included in the medium such as peptone, yeast extract, meat extract provide the nitrogen, carbon, and vitamins required for organism growth. Triple sugar Iron Agar Consists of three carbohydrates-dextrose, Lactose and Sucrose. When the carbohydrate are fermented, acid production is detected by the phenol red pH indicator. Sodium thiosulphate is reduced to hydrogen sulphide, and hydrogen sulphide reacts with an iron salt yielding the typical black iron sulphide. Ferric ammonium citrate is the hydrogen sulphide (H2S) indicator. Sodium chloride maintains the osmotic balance of the medium. Agar is used as a solidifying agent.

Interpretation

Cultural characteristics observed after an incubation at 35-37°C for 18-24 hours.

Test Strains	ATCC	Inoculum (cfu/ml)	Growth	Butt/Slants/Gas/H2S
Salmonella Paratyphi A	9150	103	Luxuriant	Yellow /Red/+/-
Salmonella Typhi	6539	103	Luxuriant	Yellow /Red/-/+
Salmonella Typhimurium	14028	103	Luxuriant	Yellow /Red/+/+
Vibrio cholerae	15748	103	Luxuriant	Yellow /Red/-/-

Reference

1. Sulkin E.S. and Willett J.C., 1940, J. Lab. Clin. Med.

2. Hajna A.A., 1945, J. Bacteriol, 49:516.

3. Finegold and Baron, 1986, Bailey and Scotts Diagnostic Microbiology, 7th ed., The C.V. Mosby      Co., St. Louis.

4. Greenberg A. E., Trussell R. R. and Clesceri L. S. (Eds.), 1985, Standard Methods for the Examination of Water and Wastewater, 16th ed., APHA, Washington, D.C.

5. MacFaddin J., 1985, Media for Isolation-Cultivation-Identification-Maintenance of Medical Bacteria, Vol. 1, Williams and Wilkins, Baltimore.

Type of Culture Media

Culture media is of essential significance for most microbiological tests: to get purecultures, to develop and to check microbial cells, and to develop and choose microorganisms.Without top notch media then the likelihood of accomplishing exact, reproducible andrepeatable microbiological test outcomes is decreased. A microbiological culture medium is asubstance which energizes the development, backing and survival of smaller scale living beings. Culture mediacontains supplements, development advancing components, vitality sources, cradle salts, minerals, metals andgelling operators (for strong media). Culture media has been utilized by microbiologists since thenineteenth century. Indeed, even with the expanded utilization of quick strategies most of techniquesfound in the pharmaceutical quality control research facility require development media. For theassessment of culture media, nobody conclusive standard exists. In light of this, this articlepresents a few contemplations for a testing system.

TM Media Culture Media Types

Potato Dextrose Agar (PDA) contains dextrose and potato infusion which provides a nutrient base for luxuriant growth of most fungi such as Aspergillus niger_ ATCC 16404 which show typically heavy black spores appressed to mycelium after incubation at 25°C for 5-7 days, To Know more click here

Growth on Sheep Blood agar is observed on the bases of hemolysis. It can be alpha, beta or gamma.

Luxuriant growth of Staphylococcus Aureus (ATCC 25923) shows Beta- hemolysis after incubation at 30-35 °C for 18-48 hours.

In Beta-hemolysis, the red blood cells are fully lysed in the middle and around the colonies. The area appears lighter (yellow) and transparent. Know More click here

Sabouraud Dextrose Agar is a conventional Culturemedium. 

Spergillus niger grows in white Mycelium with Black Spores, Where in conidia matures from center of colony at room temperature (25ºC), the media be incubated for 72 hrs. Get to know more Click here

Get precise results with TM Media VIOLET RED BILE AGAR of coli-aerogenes in water, milk and other dairy food products. ReadMore 

How to Use Thayer Martin Medium Base

Thayer Martin Medium Base

Carpenter and Morton reported an improved medium to isolate Gonococci in 24 hours. Later on the efficiency of GC medium supplemented with haemoglobin and yeast concentrate was demonstrated for isolating gonococci. Subsequently Thayer Martin Medium Base was developed for the primary isolation of Neisseria gonorrhoeae and Neisseria meningitidis from specimens containing mixed flora collected from throat, vagina, rectum an d urethra. Thayer and Martin used Vancomycin, Colistin and Nystatin. Martin and Lester used an additional antibiotic, Trimethoprim to make the medium selective.

PRINCIPLE

Special peptone provides nutrients to the organisms while starch neutralizes the toxic fatty acids if present in the agar. Haemoglobin provides the X factor whereas the V factor (N.A.D.) is provided by the added supplement. Starch neutralizes the toxic fatty acids if present in the agar. Supplement (TS 022) also supplies vitamins, amino acids, coenzymes etc. which enhances the growth of pathogenic Neisseria. Vancomycin and colistin inhibits gram-positive and gram-negative bacteria respectively. Nystatin inhibits fungi. This medium may inhibit Haemophilus species. Some strains of Capnocytophaga species may grow on this medium when inoculated with oropharyngeal specimens.

INSTRUCTION FOR USE

1. Dissolve 42.0 grams in 900 ml distilled water.
2. Heat to boiling to dissolve the medium completely.
3. Sterilize by autoclaving at 15 psi pressure (121°C) for 15 minutes.
4. Cool to 45-50°C.
5. Aseptically add 100 ml of sterile lysed blood and rehydrated contents of one vial of Vitamins     Amino Growth Supplement (Vitamins & Amino Acids Mixture) (TS 022) and V.C.N Supplement (TS 038) or V.C.N.T Supplement (TS 039).
6.  If desired GC Supplement with Antibiotics (TS 036) can be used as a single supplement.
7. Mix well before pouring into sterile Petri plates.
8. If Hemoglobin powder (TS 021) is used suspend 42.0 grams of Thayer Martin Medium Base in 500 ml distilled water.
9. Heat to boiling to dissolve the medium completely.
10. Prepare 500 ml of 2% hemoglobin solution.
11. Sterilize separately by autoclaving at 15 psi pressure (121°C) for 15 minutes.
12. Cool to 45°C.
13. Mix both and add the supplements as above. 

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