Pseudomonas Agar (For Fluorescein)

Pseudomonas Agar (For Fluorescein)

Intended Use:

Recommended for detection of fluorescein production by Pseudomonas species.

Composition**

**Formula adjusted, standardized to suit performance parameters

Directions

Suspend 38 grams in 1000 ml purified / distilled water containing 10 ml glycerol. 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

Pseudomonas Agar (For Fluorescein) is based on the formula described by King et al (3) and as modified in the U.S. Pharmacopeia (5) for the detection of fluorescein production a water soluble, chloroform insoluble fluorescent pigment by Pseudomonas species (4). The medium enhances the elaboration of fluorescein by Pseudomonas and inhibits the pyocyanin formation. The fluorescein pigment diffuses from the colonies of Pseudomonas into the agar and shows yellow fluorescent colouration. Some Pseudomonas strains produce small amounts of pyocyanin resulting in a yellow-green colouration.

Tryptone and proteose peptone provide the essential nitrogenous nutrients, carbon, sulphur and trace elements for the growth of Pseudomonas. Dipotassium hydrogen phosphate buffers the medium while magnesium sulphate provides necessary cations for the activation of fluorescein production. Salt concentration exceeding 2% affects pigment production. UV illumination may be bactericidal, so make sure that there is good growth before placing culture under UV light (4).

A pyocyanin-producing Pseudomonas strain will usually also produce fluorescein. It must, therefore, be differentiated from other simple fluorescent Pseudomonads by other means. Temperature can be a determining factor as most other fluorescent strains will not grow at 35°C. Rather, they grow at 25-30°C (4).

Type of specimen

Pharmaceutical samples

Specimen Collection and Handling:

For pharmaceutical samples follow appropriate techniques for handling specimens as per established 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.This medium is general purpose medium and may not support the growth of fastidious organisms.

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

Yellow coloured clear to slightly opalescent gel forms in Petri plates

Reaction

Reaction of 3.8% w/v aqueous solution (containing 1% v/v glycerol) at 25°C. pH : 7.0±0.2

pH

6.80-7.20

Cultural Response

Cultural characteristics observed with added 1% glycerol after an incubation at 35-37°C for 18-24 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 (1,2).

Reference

1. Isenberg, H.D. Clinical Microbiology Procedures Handbook 2^nd Edition.
2. 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, 11^th Edition. Vol. 1.
3. King, Ward and Raney, 1954, J. Lab. Clin. Med., 44: 301.
4. MacFaddin J., 1985, Media for Isolation-Cultivation-Identification-Maintenance of Medical Bacteria, Vol. I, Williams and Wilkins, Baltimore.
5. The United States Pharmacopoeia, 2006, USP29/NF24, The United States Pharmacopeial Convention, Rockville, MD.