Introduction of Tellurite Blood Agar (TBA)
TBA stands for Tellurite Blood Agar and it is applied for the selective isolation and cultivation of Corynebacterium species and they are Gram-positive, facultatively anaerobic, non-motile bacteria that exhibit a fermentative metabolism (carbohydrates to lactic acid) under certain conditions. Most species are normal flora of humans present virtually at all anatomic sites. Tellurite Blood Agar is a selective medium used for the isolation and cultivation of Corynebacterium species. It is selective due to the presence of inhibitors and differential by means of the ability of organisms to reduce potassium tellurite. Corynebacterium diptheriae is the causative agent of Diphtheria and another pathogenic etiological agent of this genus is are-C. pseudotuberculosis (also known as Corynebacterium ovis), Corynebacterium pyogenes, Corynebacterium aquaticum, C pseudodiphtheriticum (also known as Corynebacterium hofmannii), etc.
Principle of Tellurite Blood Agar (TBA)
Constituent of the medium, Biopeptone provides nitrogenous compounds whereas sodium chloride maintains the osmotic equilibrium of the medium. Dipotassium hydrogen phosphate buffers the medium. Corn starch is the toxic metabolites neutralizer while Haemoglobin and Vitamin Growth Supplement promote good growth of Corynebacterium species. Agar acts as a solidifying agent. Potassium tellurite acts as a selective agent and it inhibits most Gram-positive and Gram-negative bacteria except Corynebacterium species. Corynebacterium diphtheriae (causative agent of diphtheria) reduces potassium tellurite to tellurium and thereby produces gray-black colored colonies.
Composition of TBA
- Biopeptone: 10.0
- Sodium chloride: 5.0
- Dipotassium hydrogen phosphate: 4.0
- Corn starch: 1.0
- Monopotassium phosphate: 1.0
- Agar: 10.0
- Final pH (at 25°C) 7.2 ± 0.2
- Haemoglobin powder
- One vial of Vitamin Growth Supplement and Potassium Tellurite 1 %
Preparation of Tellurite Blood Agar(TBA)
- Suspend 31.5 grams of the dehydrated medium in 500 ml purified/distilled water.
- Heat to boiling to dissolve the medium completely.
- Sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes i.e. validated cycle.
- Cool to 45-50°C.
- Aseptically add extra ingredients provided by the vendor i.e. sterile prepared Haemoglobin powder solution (10 g in 490 mL distilled water) and sterile reconstituted contents of one vial of Vitamin Growth Supplement and Potassium Tellurite 1 %.
- Mix well before pouring into sterile Petri plates.
- Leave for drying.
Storage and Shelf life of TBA
- Store at 2-8ºC and away from direct light.
- Media should not be used if there are any signs of deterioration (shrinking, cracking, or discoloration), contamination.
- The product is light and temperature-sensitive; protects from light, excessive heat, moisture, and freezing.
Test Requirements for TBA
- Test specimen
- Tellurite Blood Agar
- Inoculating loop/ dropper
- Bunsen burner
- Control strains
- Waste bin
Procedure of Tellurite Blood Agar
- Allow the plates to warm at 37°C or to room temperature, and the agar surface to dry before inoculating.
- Inoculate and streak the specimen as soon as possible after collection.
- If the specimen to be cultured is on a swab, roll the swab over a small area of the agar surface.
- Streak for isolation with a sterile loop.
- Incubate plates aerobically at 35-37°C for 48 hours (or more).
- Examine colonial characteristics.
Colony Morphology of TBA
Organism Growth and Colour of the colony
Corynebacterium diphtheriae ATCC 11913: Good Grey-black
Escherichia coli ATCC 25922: Inhibited
Uses of Tellurite Blood Agar
- Tellurite Blood Agar is a selective and differential medium for Corynebacterium diphtheriae.
- It is also a selective medium for the cultivation and isolation of Corynebacterium species.
- It is applicable for both clinical and non-clinical specimens to recover Corynebacterium species.
- It is recommended that both selective (Tellurite Blood Agar) and non-selective (Loeffler’s serum slope) media should be used parallelly during suspected specimens of Corynebacterium diphtheriae inoculation and it is performed so for enhanced recovery.
Limitations of TBA
- Colony characteristics only provide presumptive identification and thus biochemical, immunological, molecular, or mass spectrometry testing be performed on colonies from pure culture for final identification.
- Serial inoculation may be required to assure adequate isolation of mixed flora samples.
Keynotes on Tellurite Blood Agar
- Throat or nasal swab is directly inoculated and streaked on this agar medium.
- A modification of Neill’s medium is preferred for the isolation and differentiation of Corynebacterium diphtheriae types.
- C. diphtheriae, also called as Klebs-Loeffler bacillus is a gram-positive, non-sporulated, non-encapsulated, non-motile facultative anaerobe.
- Diphtheria is an infectious disease caused by the etiological agent, C. diphtheriae and characterized by an inflammatory lesion and membranous exudates on the mucosa of the upper respiratory tract.
- Loffler serum slope restores virulence and other identifying properties (microscopic and colonial) after they have been lost due to prolonged incubation or repeated subculturing and it is also used for the demonstration of pigmentation and ascospores.
- Downie’s blood tellurite agar (BTA), Hoyle medium (modification of Neill’s medium), Tinsdale, and Loeffler’s serum slope are the most common media used for the cultural isolation and differentiation of Corynebacterium diphtheriae.
Further Readings on TBA
- Scott T. J., 1981, Microbiological Media, A Manual of Products and Procedures, Fieskeville, TI: Scott Laboratories, Inc.
- MacFaddin J. F., 1985, Media for Isolation-Cultivation-Identification-Maintenance of Medical Bacteria. I, Williams and Wilkins, Baltimore.
- Data on file: Microxpress®, A Division of Tulip Diagnostics (P) Ltd.