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General laboratory information drawn from manufacturer instructions for use; not medical advice and not a procedure.
Short answer
Litmus milk is a liquid microbiology medium made from skim milk and the pH and redox indicator litmus. It is used to see what bacteria do to milk: ferment lactose to acid, form a clot, digest casein, make gas, or reduce the litmus to a colourless form. Manufacturers state the reactions are not specific on their own, so a result narrows an identification but does not complete one.
Definition and scope
Remel (Thermo Fisher) describes Litmus Milk Medium as a liquid medium for qualitative procedures that determine the action of bacteria on milk and that maintain lactic acid bacteria. It differentiates organisms on lactose fermentation, casein hydrolysis and coagulation, gas production and reduction of litmus. The manufacturer singles out two uses: species differentiation within the genus Clostridium, and separating Streptococcus bovis (no growth) from Streptococcus equinus (growth). HiMedia’s Litmus SM Broth is described for maintaining lactobacilli and for determining the action of bacteria on milk.
This page covers what the medium is and how readings are interpreted. It is one of several differential biochemical media; for related readouts see the guides on phenol red broth, the methyl red test and identifying bacteria.
How the reactions are read
Per the Remel instructions, litmus indicates both pH and oxidation-reduction potential. It is red in acid and blue in alkaline conditions. Lactose fermentation produces lactic acid and turns the medium pinkish-red. Organisms that attack nitrogenous substances release ammonia and turn it purplish-blue. Some organisms reduce litmus to a white leuco base through reductase activity. Proteolytic enzymes can coagulate milk protein into a clot, and casease can peptonize it, seen as the medium clearing to a watery appearance. Carbon dioxide and hydrogen gas may form; abundant gas breaks up an acid clot, a reaction called stormy fermentation that the manufacturer notes may occur with certain anaerobic Clostridium species.
Remel lists incubation at 35-37 C for 18-24 hours and states that longer periods, up to 14 days, may be necessary. It also says not to attempt to differentiate between a clot and a curd formation.
Reaction types: acid, alkaline and redox
The table combines the Remel code key with the HiMedia reaction classes. Every row is stated by one or both manufacturers.
| Appearance | Reading | Stated cause |
|---|---|---|
| Pinkish-red | Acid (Remel code A) | Lactose fermentation to lactic acid |
| Blue | Alkaline (ALK) | Ammonia or amines from nitrogenous substrates |
| Purplish-blue | No fermentation (NF) | No reaction on the substrate |
| Solid mass | Clot or curd (C) | Coagulation of milk protein; acid clots in acid, soft blue clots under alkaline coagulation (HiMedia: paracasein by rennin) |
| Clearing, clot dissolved | Digestion or peptonization (D) | Casein hydrolysis |
| Bubbles, clot broken | Gas (G) or stormy fermentation (SF) | Abundant gas disrupting an acid clot |
| White or decolourized | Reduction of litmus (RED) | Reductase activity, forming leucolitmus, typically in the depths of the tube |
What labs compare when evaluating and storing the medium
Two manufacturers publish different formulas, so results and lot comparisons should be read against the formula actually in use. Remel states its classical formula is adjusted to meet performance standards.
| Item | Remel Litmus Milk Medium (IFU 61274, rev. 2013) | HiMedia Litmus SM Broth (M609) |
|---|---|---|
| Skim milk component | Skim milk 100.0 g per litre | SM powder 100.0 g per litre |
| Litmus | 0.75 g per litre | 0.5 g per litre |
| Sodium sulphite | Not listed | 0.5 g per litre |
| Final pH | 6.8 +/- 0.2 at 25 C | 6.8 +/- 0.2 |
Quality-control organisms the manufacturers name
- Remel (up to 5 days at 33-37 C): Escherichia coli ATCC 25922, ambient air, gives an acid reaction; Clostridium innocuum ATCC 14501 is also listed as a control; Clostridium perfringens ATCC 13124, anaerobic, gives acid, clotting and digestion, including stormy fermentation.
- HiMedia (35-37 C, up to 14 days): C. perfringens ATCC 13124 gives stormy fermentation; Lactobacillus johnsonii ATCC 11506 gives an acid clot; Pseudomonas aeruginosa ATCC 27853 gives peptonization.
Storage and handling statements
HiMedia states that the powder is stored tightly closed at 10-30 C and the prepared medium at 15-30 C, that the powder is hygroscopic and lumps if stored improperly, that overheating during sterilization should be avoided, and that contaminated material is discarded by autoclaving. It describes the prepared medium as a light purple, opaque, milky solution. It lists dairy and water samples as specimen types.
Common misunderstandings
- A result identifies the organism. HiMedia states that reactions in this medium are not specific, that further tests must be carried out, and that further biochemical and serological testing is required for complete identification.
- Stormy fermentation proves a Clostridium. Remel says it may occur with certain anaerobic Clostridium species; it does not say it is exclusive to them.
- Clot and curd are separable readings. Remel instructs users not to attempt to differentiate between a clot and a curd formation.
- Every strain grows well. HiMedia notes that some strains may show poor growth because of nutritional variation.
- An 18-24 hour read is always enough. Remel notes that periods of up to 14 days may be necessary.
Other differential media answer different questions, for example the gelatin hydrolysis test for gelatin liquefaction and the urease test.
FAQ
What is litmus milk used for?
Remel describes it as a liquid medium for qualitative procedures that determine the action of bacteria on milk and that maintain lactic acid bacteria. It differentiates organisms on lactose fermentation, casein hydrolysis and coagulation, gas production and reduction of litmus.
Does a litmus milk result identify a bacterium?
No. HiMedia states that reactions in this medium are not specific and that further biochemical and serological testing is required for complete identification.
What is stormy fermentation?
Remel defines it as abundant gas breaking up an acid clot, which may occur with certain anaerobic Clostridium species.
How long can incubation take?
Remel lists 18-24 hours at 35-37 C and states that longer periods, up to 14 days, may be necessary.
Sources
- Thermo Fisher Scientific (Remel), Litmus Milk Medium instructions for use, IFU61274, revised April 4, 2013: documents.thermofisher.com/TFS-Assets/MBD/Instructions/IFU61274.pdf. Checked 2026-10-31.
- HiMedia Laboratories, Technical Data M609, Litmus SM Broth: himedialabs.com/media/TD/M609.pdf. Checked 2026-10-31.
