Tryptone Salt Agar, w/1% NaCl

Intended Use

Recommended for differentiation of El Tor and Classical biotypes of Vibrio in accordance with FDA BAM, 1998.

Composition

**Formula adjusted, standardized to suit performance parameters

Directions

Suspend 40.0 grams in 1000 ml purified / distilled water. Heat to boiling to dissolve the medium completely. For slants, dispense into tubes. Sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes. Cool to 45-50°C. Solidify tubes as slants or mix well and pour into sterile Petri plates.

Principle And Interpretation

Members of the genus Vibrio are defined as Gram-negative, asporogenous rods that are straight or have a single, rigid curve. They are motile; most have a single polar flagellum, when grown in liquid medium. Different methods used for the confirmation of Vibrio species include physical, biochemical and serological assays (5). Tryptone salt agar w/ 1% NaCl is used for the growth of Vibrio sp. in accordance with FDA BAM (1) for differentiation of El Tor and Classical biotypes.

Blend the food sample to be analysed with Alkaline peptone water (APW) in appropriate ratio and incubate as per the recommendation by FDA BAM. Pure cultures can be isolated from APW by plating a loopful of the inoculum into TCBS agar. For biochemical and serological identification of Vibrio, colonies from crowded plates must be streaked to Tryptone salt agar, w/1% NaCl for purity. Incubate overnight at 35 ±2° C and proceed with identification using a single isolated colony for differentiation of Classic and El Tor biotypes. Further biochemical tests can also be done using colonies from this medium.

Type of specimen

Food samples - Raw, undercooked, contaminated, or recontaminated seafood.

Specimen Collection and Handling

For food samples, follow appropriate techniques for sample collection and processing as per guidelines (4).

After use, contaminated materials must be sterilized by autoclaving before discarding.

Warning and Precautions

Read the label before opening the container. Wear protective gloves/protective clothing/eye protection/ face protection. Follow good microbiological lab practices while handling specimens and culture. Standard precautions as per established guidelines should be followed while handling specimens. Safety guidelines may be referred in individual safety data sheets.

Limitations

1. Further biochemical and serological tests must be carried out for complete identification.

Performance and Evaluation

Performance of the medium is expected when used as per the direction on the label within the expiry period when stored at recommended temperature.

Quality Control

Appearance: Cream to yellow homogeneous free flowing powder

Gelling: Firm, comparable with 2.0% Agar gel

Colour and Clarity of prepared medium: Light yellow coloured clear to slightly opalescent gel forms in Petri plates

Cultural Response: Cultural characteristics observed after an incubation at 35-37°C for 18-48 hours.

Key : (*) Corresponding WDCM numbers.

Storage and Shelf Life

Store between 10-30°C in a tightly closed container and the prepared medium at 20-30°C. Use before expiry date on the label. On opening, product should be properly stored dry, after tightly capping the bottle in order to prevent lump formation due to the hygroscopic nature of the product. Improper storage of the product may lead to lump formation. Store in dry ventilated area protected from extremes of temperature and sources of ignition. Seal the container tightly after use. Product performance is best if used within stated expiry period.

Disposal

User must ensure safe disposal by autoclaving and/or incineration of used or unusable preparations of this product. Follow established laboratory procedures in disposing of infectious materials and material that comes into contact with sample must be decontaminated and disposed of in accordance with current laboratory techniques (2,3).

Reference

1. FDA, U.S. 1998. Bacteriological Analytical Manual. 8 ed. Gaithersburg, Md. : AOAC International.
2. Isenberg, H.D. Clinical Microbiology Procedures Handbook. 2nd Edition.
3. Jorgensen, J.H., Pfaller, M.A., Carroll, K.C., Funke, G., Landry, M.L., Richter, S.S and Warnock., D.W. (2015) Manual of Clinical Microbiology, 11th Edition. Vol. 1.
4. Salfinger Y., and Tortorello M.L. Fifth (Ed.), 2015, Compendium of Methods for the Microbiological Examination of Foods, 5th Ed., American Public Health Association, Washington, D.C.
5. Vera. 1944. J. Bact., 47.