HiCrome™ MM Agar

Intended Use:

Recommended for identification and differentiation of Salmonella and non-salmonella like Citrobacter from water and clinical samples.

Composition**

Final pH (at 25°C) 7.6±0.2

**Formula adjusted, standardized to suit performance parameters

# Equivalent to Beef extract

Directions

Suspend 49.13 gram 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

HiCrome™ MM Agar was formulated by Miller and Mallison (1) for specific isolation and detection of Salmonellae. This medium is superior to XLT4 Agar in supporting growth of Salmonella due to the presence of appropriate proportion of four sugars. Most differential and selective media are formulated with one or more sugars and pH indicators respectively. The utilization of sugars by organisms results in pH-changes. This is used as a means of distinguishing Salmonella from competing bacteria on the basis of colony colour.

Salmonella usually are unable to ferment these sugars (2) that supports growth of competing bacteria. Thus other bacteria tend to overgrow Salmonellae, masking their presence. The inclusion of sugars like mannitol, cellobiose and trehalose stimulate the better initial growth of Salmonella cells. However, the low concentrations of these sugars do not interfere with the utilization of protein and H2S production. Presence of lactose suppresses H2S production by non-salmonellae like Citrobacter freundii. The chromogenic mixture, present in this medium helps to differentiate between lactose fermenters and nonfermenters. Lactose fermenters give bluish green coloured colonies, which would have been impossible to differentiate with an indicator based on pH change. Inclusion of tergitol 4 in the medium suppresses the presence of Proteus and Providencia colonies. Peptone and HM peptone B provide essential nitrogen compounds.

Type of specimen

Clinical samples - Faeces, urine, etc.; Food samples; Water samples

Specimen Collection and Handling:

For water samples, follow appropriate techniques for sample collection, processing as per guidelines and local standards (3).For food samples, follow appropriate techniques for sample collection and processing as per guidelines (4). For clinical samples follow appropriate techniques for handling specimens as per established guidelines (5,6). After use, contaminated materials must be sterilized by autoclaving before discarding.

Warning and Precautions :

In Vitro diagnostic Use. 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 :

1. Individual organisms differ in their growth requirement and may show variable growth patterns on the medium.
2. Though most of the Salmonella produce H2S certain non H2S producing Salmonella species may appear as colourless colonies.
3. Certain Salmonella species which are lactose fermenters may show as bluish green coloured colonies.
4. Further confirmation may be carried out on suspected colonies.

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 1.5% Agar gel

Colour and Clarity of prepared medium Light amber coloured, slightly opalescent gel forms in Petri plates

Reaction Reaction of 4.91% w/v aqueous solution at 25°C. pH: 7.6±0.2

pH 7.40-7.80

Cultural Response

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

Key: (*)Corresponding WDCM numbers.

Storage and Shelf Life

Store between 15-25°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 (5,6).

Reference

1. Miller R.G. and Mallison E.T., 2000, J. Food Protection, 63(10), 1443-46.
2. Miller R.G., Tate C.R., Mallinson E.T. and Scherrer J.A., 1991, Pault Sa 70:2429-32.
3. Lipps WC, Braun-Howland EB, Baxter TE,eds. Standard methods for the Examination of Water and Wastewater, 24th ed. Washington DC:APHA Press; 2023.
4. Salfinger Y., and Tortorello M.L., 2015, Compendium of Methods for the Microbiological Examination of Foods, 5th Ed., American Public Health Association, Washington, D.C.
5. Isenberg, H.D. Clinical Microbiology Procedures Handbook 2nd Edition.
6. 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.