M-Aeromonas Selective Agar Base (Havelaar)

Intended Use:

Recommended for the detection of Aeromonas species in water and other liquid samples by membrane filter technique.

Composition**

Final pH (at 25°C): 7.5±0.2

**Formula adjusted, standardized to suit performance parameters

Directions

Suspend 36.74 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 and aseptically add rehydrated contents of 1 vial of Amp Selective Supplement (FD082). Mix well and pour into sterile Petri plates.

Principle And Interpretation

Aeromonas species are natural inhabitants of aquatic environments worldwide. Procedures to isolate, enumerate and identify Aeromonas from water and wastewater sources is of significance, because of their role in causing human and animal disease, their ability to colonize treatment plants and distribution systems and their presence and distribution as an alternative indicator of the trophic state of water (1). M-Aeromonas Selective Agar Base is recommended for the detection of Aeromonas species in water sample by the membrane filter technique. This medium was formulated by Havelaar et al (2,3) and also complies with the recommendations of USEPA Approved Method 1605 (2001) which describes Ampicillin Dextrin Agar with Vancomycin (4). Aeromonas utilize Dextrin in the medium to form acids which are detected by the pH indicator bromothymol blue by changing the colour from blue to yellow. Tryptose and yeast extract provide nitrogenous compounds along with other essential nutrients for growth of Aeromonas. Sodium chloride maintains the osmotic balance of the medium. Aeromonas forms acid from dextrin, which is indicated by change in colour from blue to yellow. Selectivity of the medium is increased by the addition of ampicillin. The effectiveness of ampicillin as a selective agent has been reported by several workers (5-8). Membrane filters through which water samples have been passed are aseptically placed on M-Aeromonas Selective Agar Base plates. After an incubation at 35-37°C for 24 hours Aeromonas species appear as large, yellow colonies with a purple periphery.

Type of specimen

Water samples.

Specimen Collection and Handling:

For water samples, follow appropriate techniques for sample collection, processing as per guidelines and local standards.(1) After use, contaminated materials must be sterilized by autoclaving before discarding.

Warning and Precautions

Read the label before opening the container. The media should be handled by trained personnel only. 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: Light yellow to greenish yellow homogeneous free flowing powder

Gelling: Firm, comparable with 1.3% Agar gel.

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

Reaction: Reaction of 3.67% w/v aqueous solution at 25°C. pH: 7.5±0.2

pH: 7.30-7.70

Cultural Response: Cultural characteristics observed after an incubation at 35-37°C for 24 hours with added Amp Selective Supplement (FD082).

Key: * Corresponding WDCM numbers

Storage and Shelf Life

Store between 10-30°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 sample must be decontaminated and disposed of in accordance with current laboratory techniques (9,10).

Reference

1. Lipps WC, Braun-Howland EB, Baxter TE,eds. Standard methods for the Examination of Water and Wastewater, 24th ed. Washington DC:APHA Press; 2023.
2. Havelaar A. H., During M. and Versteegh J. F. M., 1987, J. Appl. Micro. 62:279-287.
3. Havelaar A. H., Vonk M., 1988, Lett. Appl. Microbiol. 7:169
4. United States Environmental Protection Agency (USEPA), Method 1605: Aeromonas in Finished Water by Membrane Filtration using Ampicillin Dextrin Agar with Vancomycin (ADA-V) October 2001.
5. Atkinson M., 1986 Culture; Vol. 7, No. 2.
6. Moulsdale M. T., 1983, The Lancet, 1:351.
7. Richardson C. J., Robinson J. O., Wagerer L. B., Burker V. J., 1982, Antimicrob., Chemother., 9:267.
8. Rogol M., Sechter 1., Grenber L., Gerichter Ch. B., 1979, J. Med. Microbiol., 12:229.
9. Isenberg, H.D. Clinical Microbiology Procedures Handbook 2nd Edition.
10. 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.