Hi-Sensitivity™ Test Agar

Intended use

Hi-Sensitivity Test Agar is used for antimicrobial susceptibility tests.

Composition**

Final pH (at 25°C): 7.4±0.2

**Formula adjusted, standardized to suit performance parameters

Directions

Suspend 31.4 grams in 1000 ml purified/distilled water. Heat to boiling to dissolve the medium completely. Sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes. Cool to 45-50°C. Mix well and pour into sterile Petri plates.

Principle And Interpretation

The goal of an antimicrobial susceptibility test is to predict through an in vitro assessment the likelihood of successfully treating an infection with a particular antimicrobial agent. There are several continual or novel methods for performing antibacterial susceptibility testing. These include the disk diffusion test, broth microdilution, agar gradient and rapid automated instrument methods (1). Hi-Sensitivity Test Agar, which is used for antimicrobial susceptibility tests, is a semi-defined medium in which the mineral contents have been stabilized to give reproducible results. The thiamine and thymidine content is very low thus making it most suitable for testing antimicrobial activity of sulfonamide. However some mutant strains which are totally dependent on thiamine and thymidine for their growth, will not grow on Hi- Sensitivity Test Agar, due to very low levels of these compounds in the media as they are the naturally occurring antagonist of trimethoprim. These strains should be carefully recognized (2,3,4).

Hi-Sensitivity Test Agar has been so designed to overcome the problems occurring in Mueller-Hinton Media that are as

Type of specimen

Clinical samples -Isolated Microorganisms from urine, stool etc.

Specimen Collection and Handling

For clinical samples follow appropriate techniques for handling specimens as per established guidelines (1,14).

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

Warning and Precautions

In vitro diagnostic use only. For professional use only. 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 clinical specimens. Safety guidelines may be referred in individual safety data sheets.

Limitations

• This medium is recommended for susceptibility testing of pure cultures only.
• Inoculum density may affect the zone size. Heavy inoculum may result in smaller zones or too less inoculum may result in bigger zones.
• Fastidious organisms may not grow on this medium and may require supplementation of blood.
• However some mutant strains which are totally dependent on thiamine and thymidine for their growth, will not grow on Hi-Sensitivity Test Agar, due to very low levels of thiamine and thymidine these compounds in the media as they are the naturally occurring antagonist of trimethoprim. These strains should be carefully recognized (2,3,4).
• As antimicrobial susceptibility is carried with antibiotic disc, proper storage of the disc is desired which may affect the potency of the disc.
• Under certain circumstances, the in vitro results of antibiotic susceptibility may not show the same in vivo.

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 0.8% Agar gel.

Colour and Clarity of prepared medium

Basal medium: Light amber coloured, clear to slightly opalescent gel. After addition of 5%v/v laked blood : Red to chocolate coloured, opaque gel forms in Petri plates.

Reaction

Reaction of 3.14% w/v aqueous solution at 25°C. pH: 7.4±0.2

pH: 7.20-7.60

Cultural Response

Cultural characteristics observed with added 5%w/v laked blood, after an incubation at 35 - 37°C for 18 - 24 hours .

Key*- Corresponding WDCM numbers **Formerly known as Bacillus subtilis subsp. spizizenii * Formerly known as Bacteroides vulgatus

Storage and Shelf Life

Store between 2-8°C in a tightly closed container and the prepared medium at 2-8°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 clinical sample must be decontaminated and disposed of in accordance with current laboratory techniques (1,14).

Reference

1. 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.
2. Barker J., Healing D., and Hutchinson J. G. P., 1972, J. Clin. Path., 25:1086
3. Barker J., Healing D., and Hutchinson J. G. P., 1972, J. Clin. Path., 25:1086
4. Yourassowsky E., Vanderlinden M. P. and Schoutens E., 1974, J. Clin. Path., 27:897.
5. Bridson E.Y., 1976, Arztl. Lab., 22:373.
6. Duncan I. B. R., 1974, Antimicrob. Agents Chemother., 5:9.
7. Ericsson H. M. and Sherris J. C., 1971, Acta. Pathol. Microbiol Scand Suppl., 217:1.
8. Garrod L. P. and Waterworth P. M., 1971, J. Clin. Path., 24:779.
9. Neussil H., 1976, Chemother., Vol. 2:33.
10. Reller L. B., Schoenknecbt F. D., Kenny M. A. and Sherris J. C., 1974, J. Infect. Dis., 130:454.
11. Thomas M. and Bond L., 1973, Med. Lab. Technol., 30:277
12. Acar J. F., 1980, Antibiotics in Laboratory Medicine, Lorian V. (Ed.), Williams and Wilkins, Baltimore, USA, 48-51.
13. Hartzen S.H., Anderson L.P., Bremmelgaard A. et.al, 1997, Antimicrob. Agents Chemother., 41.2634-2639.
14. Isenberg, H.D. Clinical Microbiology procedures Handbook. 2nd Edition.