Dehydrated Culture Medium
One-step MPN broth for faecal coliforms and E. coli in water, wastewater, seawater and shellfish
Cat. No. AS-110131.5 g/LpH 6.9 ± 0.2Selective brothMPN method
At a glance
- Target organismsFaecal coliforms, E. coli
- Incubation35 °C for 3 h, then 44.5 °C
- Time24 ± 2 h in total
- Sterilisation121 °C, 10 min
Manufactured in Melbourne. TDS and SDS available. Batch COA on request.
Overview
A-1 Medium is a selective lactose broth for the Most Probable Number (MPN) enumeration of faecal coliforms in water, wastewater, seawater, shellfish and foods. It gives a faecal coliform result in about 24 hours in a single step, without a separate presumptive test.
Tubes are incubated at 35 °C for 3 hours to let stressed cells recover, then at 44.5 °C for a further 21 ± 2 hours. Gas in the Durham tube indicates faecal coliforms.
Key features
- One-step faecal coliform result in about 24 hours
- Triton X-100 included in the powder: just suspend 31.5 g/L
- Double-strength preparation for 10 mL sample volumes
Principle of the Medium
| Component | Function |
|---|---|
| Enzymatic digest of casein | Nitrogen, amino acids and peptides |
| Lactose | Fermentable carbohydrate; fermentation produces gas |
| Salicin | Additional fermentable carbohydrate |
| Sodium chloride | Maintains osmotic balance |
| Triton X-100 | Non-ionic detergent that inhibits non-coliform organisms |
Autoclave for 10 minutes only. Longer heating can break down the lactose and affect gas production.
Typical Composition
| Ingredient | g/L |
|---|---|
| Enzymatic digest of casein | 20.00 |
| Lactose | 5.00 |
| Sodium chloride | 5.00 |
| Salicin | 0.50 |
| Triton X-100 | 1.00 |
| Total | 31.50 |
Final pH 6.9 ± 0.2 at 25 °C. Formula may be adjusted and/or supplemented to meet performance specifications.
Reading the Results
| Observation after 21 ± 2 h at 44.5 °C | Indicates |
|---|---|
| Gas in the Durham tube (any amount) | Positive: faecal coliforms present |
| No gas | Negative at that dilution |
Applications
| Field | Typical use |
|---|---|
| Water and wastewater | Faecal coliform MPN in source water, recreational water and treated wastewater |
| Seawater and shellfish | Faecal coliform MPN in shellfish growing waters and shellfish |
| Food microbiology | Faecal coliform and E. coli MPN in foods |
| Teaching laboratories | Demonstrating the MPN method |
Comparable Products
| Manufacturer | Product | Relationship |
|---|---|---|
| HiMedia Laboratories | M874 A-1 Broth | Same published formulation |
| Neogen | NCM0124 A-1 Medium | Same published formulation |
| Merck | 1.00415 A-1 Broth GranuCult® prime | Comparable published formulation (granulated) |
Frequently Asked Questions
Why is the autoclave time only 10 minutes?
Longer heating can break down the lactose and affect gas production. Standard Methods specifies 121 °C for 10 minutes for A-1 Medium.
Why incubate at 35 °C for 3 hours first?
Faecal coliforms in water and seawater are often stressed. Three hours at 35 °C lets them recover before the selective 44.5 °C step.
When do I use double-strength medium?
When you add 10 mL of sample to 10 mL of medium. Prepare it at 63 g/L so the final concentration is correct.
How is A-1 Medium different from the LTB and EC method?
The LTB and EC method takes two steps: a presumptive test in lauryl tryptose broth, then confirmation in EC medium, over 48–72 hours. A-1 Medium gives a faecal coliform result in one step in about 24 hours.
Intended use: For laboratory microbiological testing, quality control and research use only. Not intended for direct use in food, human or veterinary therapeutic applications, diagnosis, or consumption.
Mode of Action
The casein digest supplies nitrogen and amino acids. Lactose and salicin are fermented by coliforms with acid and gas, and the gas collects in the Durham tube. Triton X-100 inhibits non-coliform organisms. Incubation at 44.5 °C restricts gas production to thermotolerant (faecal) coliforms, mainly Escherichia coli.
Preparation
- Suspend 31.5 g of powder in 1 litre of purified or distilled water. For 10 mL sample volumes, prepare double-strength medium at 63 g/L.
- Mix until completely dissolved, warming gently if needed.
- Dispense into test tubes containing inverted Durham tubes, so that the Durham tubes are fully covered.
- Sterilise by autoclaving at 121 °C for 10 minutes.
- Cool to room temperature. Check that the Durham tubes contain no air bubbles before use.
Use double-strength medium when the sample volume equals the medium volume (for example 10 mL sample into 10 mL medium), so the final concentration is correct.
Incubation and Interpretation
| Parameter | Condition |
|---|---|
| Inoculation | Sample volumes and dilutions as specified in the MPN method used |
| Step 1 | 35 ± 0.5 °C for 3 hours |
| Step 2 | Transfer to a water bath at 44.5 ± 0.2 °C for 21 ± 2 hours, with the water level above the medium in the tubes |
| Positive result | Gas in the Durham tube |
Count positive tubes at each dilution and calculate the MPN from standard MPN tables. Confirm E. coli with further tests if required by your method.
Limitations
- Accurate temperature control is essential: use a circulating water bath at 44.5 ± 0.2 °C, not an air incubator.
- Some thermotolerant coliforms other than E. coli can produce gas at 44.5 °C.
- A positive tube is presumptive for E. coli; confirm if the method requires it.
- Do not autoclave longer than 10 minutes.
Literature and References
- Andrews WH, Presnell MW. Rapid recovery of Escherichia coli from estuarine water. Appl Microbiol. 1972;23:521–523.
- Andrews WH, Diggs CD, Wilson CR. Comparative validity of members of the total coliform and fecal coliform groups for indicating the presence of Salmonella in the Eastern oyster. Appl Microbiol. 1975;29:130–131.
- Standridge JH, Delfino JJ. A-1 medium: alternative technique for fecal coliform organism enumeration in chlorinated wastewaters. Appl Environ Microbiol. 1981;42:918–920.
- APHA, AWWA, WEF. Standard Methods for the Examination of Water and Wastewater, 23rd ed., Section 9221 E. APHA; 2017.
- US Food and Drug Administration. Bacteriological Analytical Manual, Media: A-1 Medium.
Hazard Identification
Not Dangerous Goods under the ADG Code. Contains Triton X-100 (about 3% w/w), which is an eye irritant: see the SDS for the full GHS classification.
Avoid contact with eyes. Dust may cause mild mechanical irritation of the respiratory tract. Contains casein-derived material: people with milk protein allergy should avoid exposure.
Composition
| Ingredient | CAS No. | % w/w |
|---|---|---|
| Enzymatic digest of casein | 91079-40-2 | 63.5 |
| Lactose | 63-42-3 | 15.9 |
| Sodium chloride | 7647-14-5 | 15.9 |
| Salicin | 138-52-3 | 1.6 |
| Triton X-100 (octylphenol ethoxylate) | 9002-93-1 | 3.2 |
First Aid
| Route | Action |
|---|---|
| Inhalation | Move to fresh air. Seek medical advice if symptoms persist. |
| Skin | Wash with soap and water. |
| Eyes | Rinse cautiously with water for at least 15 minutes. Remove contact lenses if easy to do. Seek medical advice. |
| Ingestion | Rinse mouth. Do not induce vomiting. Seek medical advice if unwell. |
Poisons Information Centre (Australia, 24 h): 13 11 26
Handling, Spills and Disposal
- Avoid generating and breathing dust, and avoid contact with eyes.
- Spills: sweep up or HEPA-vacuum without raising dust; wash the area with water.
- Unused product: dispose of in accordance with local regulations. Triton X-100 is harmful to aquatic life; do not discharge large quantities to drains.
- Inoculated medium: treat as biohazardous and decontaminate (for example by autoclaving) before disposal.
Quality Control Specification
Each batch is tested for growth performance and gas production. Incubation at 35 °C for 3 hours, then 44.5 °C for 21 ± 2 hours.
| QC organism | Strain | Inoculum (CFU) | Expected result |
|---|---|---|---|
| Escherichia coli | ATCC 25922 | 50–100 (≤ 100) | Good growth, gas |
| Klebsiella aerogenes | ATCC 13048 | 50–100 (≤ 100) | No gas at 44.5 °C |
| Bacillus spizizenii | ATCC 6633 | ≥ 103 | Inhibited |
Physical and Chemical Tests
| Test | Specification |
|---|---|
| Appearance, powder | Cream to light yellow, homogeneous |
| Solubility | Dissolves completely (31.5 g/L) |
| Appearance, prepared medium | Light amber, clear to slightly opalescent |
| pH, prepared medium (25 °C) | 6.70–7.10 |
The information on this page is provided in good faith as a guide to the properties and use of this product and does not constitute a warranty of fitness for a particular purpose. Users are responsible for confirming suitability for their application. Ausamics Pty Ltd · ABN 56 676 640 467 · 36/25 Trafalgar Road, Epping VIC 3076, Australia · support@ausamics.com