R2A Broth

Intended Use

R2A Broth (DM1403) is recommended for cultivation and maintenance of heterotrophic bacteria from potable water.

Product Summary and Explanation

R2A media (Reasoner´s 2A) was developed for culturing heterotrophic bacteria that inhabit treated potable water. Nutritionally rich media, such as Plate Count Agar, support the growth of fast-growing bacteria, but may suppress the slow growing or stressed bacteria found in treated water.⁽¹⁾ For this reason a nutritionally reduced medium was described. R2A Agar is a modification of this medium.^(2,3) R2A is a low nutrient medium that in combination with a lower incubation temperature and longer incubation time will stimulate the growth of stressed and chlorine-tolerant bacteria. R2A Agar is also recommended by APHA⁽⁴⁾ for pour plate, spread plate and membrane filter technique. R2A Broth is similar to R2A Agar except agar. Total count recommended for the bacterial examination of potable waters gives an estimate of the aerobic and facultative anaerobic bacteria, which grow best at 35°C in a rich medium.⁽³⁾ R2A Broth enables better recovery of these bacteria from treated waters under different incubation conditions. Many bacteria from natural waters, which contain limited nutrients at ambient temperature, grow best on the media with less nutrient levels and at the temperatures below the routine laboratory incubation temperatures.⁽³⁾

Principles of the Procedure

R2A Broth contains casein acid hydrolysate, proteose peptone serves as source of nitrogen, vitamins, amino acids, carbon and minerals and essential growth factors required for metabolism of the bacteria. Yeast extract provides a source of trace elements and vitamins. Dextrose is the carbon and energy source. Starch acts as a neutralizer that neutralizes any toxic metabolites, if present. Potassium phosphate is used to balance the pH and provide a source of phosphate. Sodium pyruvate supplies additional nutrition and increases the recovery of stressed cells. Magnesium sulphate serves as a source of ions. Due to the presence of the above mentioned ingredients these media allow the growth of stressed and chlorine tolerant bacteria present in treated waters.

Formula / Liter

Precautions

1. For Laboratory Use only.
2. IRRITANT. Irritating to eyes, respiratory system, and skin.

Directions

1. Suspend 3.12 grams of the medium in one liter of distilled water.
2. Heat if necessary to dissolve the medium completely.
3. Dispense into tubes.
4. Autoclave at 121°C, 15 psi pressure, for 15 minutes / validated cycle.
5. DO NOT OVERHEAT.

Quality Control Specifications

Expected 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)

The organisms listed are the minimum that should be used for quality control testing.

Test Procedure

Refer to appropriate references for standard test procedures.

Results

Refer to appropriate references and test procedures for interpretation of results.

Storage

Store the sealed bottle containing the dehydrated medium at 10 – 30°C. Once opened and recapped, place container in a low humidity environment at the same storage temperature. Protect from moisture and light.

Expiration

Refer to the expiration date stamped on the container. The dehydrated medium should be discarded if not free flowing, or if the appearance has changed from the original color. Expiry applies to medium in its intact container when stored as directed.

Limi tations of the Procedure

1. For identification, organisms must be in pure culture. Morphological, biochemical and/or serological tests should be performed for final identification.
2. Consult appropriate texts for detailed information and recommended procedures.