Plot No. C40, Road No. 21Y, MIDC, Wagle Industrial Estate,
Thane(W)-400604, MH, India.
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Intended Use
Recommended for heterotrophic plate count of treated potable water using longer incubation periods.
Composition
| Ingredients | g/L |
|---|---|
| Casitose # | 0.500 |
| Yeast extract | 0.500 |
| Proteose peptone | 0.500 |
| Dextrose (Glucose) | 0.500 |
| Starch | 0.500 |
| Dipotassium hydrogen phosphate | 0.300 |
| Magnesium sulphate | 0.024 |
| Sodium pyruvate | 0.300 |
| Agar | 15.000 |
| Final pH (at 25°C) | 7.2±0.2 |
**Formula adjusted, standardized to suit performance parameters
# Equivalent to Casein Hydrolysate
Directions
R-2A Agar is a ready to use solid media in glass bottle. The medium is pre-sterilized, hence it does not need sterilization. Medium in the bottle can be melted either by using a pre-heated water bath or any other method. Slightly loosen the cap before melting. When complete melting of medium is observed dispense the medium in tubes as butts /slants or in plates as desired and allow to solidify. If on plate, either streak, inoculate or surface spread the test inoculum (50-100 CFU) aseptically.
Principle And Interpretation
The heterotrophic plate count (HPC), formerly known as the standard plate count is a procedure for estimating the number of live heterotrophic bacteria in water and measuring changes during water treatment, in distribution systems or in swimming pools. R-2A Agar is recommended by APHA (1,2) for estimating the heterotrophic plate count by the pour plate, spread plate or membrane filter procedure. R-2A Agar is formulated as per Reasoner and Geldreich (3). Stressed or injured organisms during water treatment are unable to grow on high nutrient media, since the faster growing organisms outgrow the former (4). Therefore the use of a low nutrient medium like R-2A Agar incubated for longer incubation periods allows these stressed organisms to grow well as per EP and BP (5,6).
Many bacteria from natural waters which contain limited nutrients at ambient temperature, grow best on the media with less nutrient levels. They grow better at the temperatures below the routine laboratory incubation temperatures of 35 to 37°C (4). Casitose, proteose peptone and yeast extract provide nitrogen, carbon compounds, vitamins, amino acids and minerals. Dextrose/ glucose serves as an energy source. Soluble starch aids in the recovery of injured organisms by absorbing toxic metabolic byproducts while sodium pyruvate increases the recovery of stressed cells. Magnesium sulphate is a source of divalent cations and sulphate. Dipotassium hydrogen phosphate is used to balance the pH of the medium. The number of colonies on a plate are reported as CFU (Colony Forming Units) per volume of sample.
Type of specimen
Water samples
Specimen Collection and Handling
For water samples, follow appropriate techniques for sample collection, processing as per guidelines and local standard (1). 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
- Longer incubation time other than specified is required for slow growing microorganisms.
- The media is intended for water samples for recovery of stressed or injured organisms.
- Further biochemical and serological testing is required 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: Sterile glass bottle containing slightly opalescent R-2A Agar.
Colour medium: Light amber coloured medium.
Quantity of medium: 500 ml of medium in glass bottle.
pH: 7.00-7.40
Sterility Check: Passes release criteria
Cultural Response: Cultural characteristics observed *by using standard ATCC cultures after an incubation at 35-37°C for 24-72 hours. (*-In case of water samples from fields it is suggested to incubate further for upto 7 days to ascertain the absence of organisms).
| Organism | Inoculum (CFU) |
Growth | Recovery |
|---|---|---|---|
| Candida albicans ATCC 10231 (00054*) | 50-100 | good-luxuriant | >=50% |
| Escherichia coli ATCC 25922 (00013*) | 50-100 | good-luxuriant | >=50% |
| Salmonella Enteritidis ATCC 13076 (00030*) | 50-100 | good-luxuriant | >=50% |
| ^Pseudomonas paraeruginosa ATCC 9027 (00026*) | 50-100 | good-luxuriant | >=50% |
| Staphylococcus aureus subsp. aureus ATCC 6538 (00032*) | 50-100 | good-luxuriant | >=50% |
| ** Bacillus spizizenii ATCC 6633 (00003*) | 50-100 | good-luxuriant | >=50% |
| # Aspergillus brasiliensis ATCC 16404(00053*) | 50-100 | good-luxuriant | >=50% |
| Enterococcus faecalis ATCC 29212 (00087*) | 50-100 | good-luxuriant | >=50% |
| Salmonella Typhi ATCC 6539 | 50-100 | good-luxuriant | >=50% |
Key: (*) Corresponding WDCM numbers.
^ Formerly known as Pseudomonas aeruginosa
**Formerly known as Bacillus subtilis subsp. spizizenii
# Formerly known as Aspergillus niger
Storage and Shelf life
On receipt store between 15-30°C Use before expiry date on the label. 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 (7,8).
| Product Name | R2A Agar |
|---|---|
| SKU | SMEB962D |
| Customized Product Available | No |
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