BRILLIANT GREEN AGAR EUROPEAN PHARMACOPEIA, USP
CAT Nº: 1078
Highly selective medium for the isolation of Salmonella,
other than Salmonella typhi, from foods, faeces and dairy products
FORMULA IN g/l
Peptone (Meat and Casein)
10.00
Yeast Extract
3.00
Lactose Monohydrate
10.00
Phenol Red
0.08
Sucrose
10.00
Brilliant Green
0.0125
Sodium Chloride
5.00
Bacteriological Agar
20.00
Final pH 6.9 ± 0.2 at 25ºC
PREPARATION
Suspend 58.1 grams of medium in one liter of distilled water. Mix well and dissolve by heating with frequent agitation.
Boil for one minute until complete dissolution. Sterilize in autoclave at 121º C for 15 minutes. Cool to 45-50ºC, mix well
and dispense into plates. If necessary, allow to dry about for approximately 2 hours with the covers partially removed.
The prepared medium should be stored at 8-15 ºC. The color is dusky green.
The dehydrated medium should be homogeneous, free−flowing and pink in color. If there are any physical changes,
discard the medium.
USES
BRILLIANT GREEN AGAR, as recommended by the European Pharmacopoeia, is used for the selective isolation of
Salmonella spp other than S. typhi in foods and clinical specimens, via Lactose/Sucrose fermentation.
The Peptone mixture provides nitrogen, vitamins, minerals and amino acids essential for growth. Yeast extract is a
source of vitamins, particularly the B-group. Sucrose and Lactose are fermentable carbohydrates providing carbon and
energy. Phenol red is the pH indicator, turning the medium a yellow color with the formation of acid as a result of
Lactose/Sucrose fermentation. Brilliant green inhibits Gram positive bacteria and most Gram negative bacilli other than
Salmonella spp Lactose/Sucrose fermenters are usually inhibited. Sodium chloride supplies essential electrolytes for
transport and osmotic balance. Bacteriological agar is the solidifying agent.
When there is a suspicion that the material under study contains low concentrations of Salmonellae, it is necessary to
initially inoculate the sample in Brilliant Green Tetrathionate Broth (Cat. 1253) or Selenite Cystine Broth (Cat. 1220) as a
preenrichment step.
As this medium is very inhibitory it allows for moderately heavy inocula, which should be evenly distributed on the
surface of the medium.
Inoculate and incubate at 35 ± 2 ºC for 18 – 24 hours. At the same time, inoculate other selective media that are less
inhibitory, such as Desoxycholate Agar (Cat. 1020), Salmonella Shigella Agar (Cat. 1064), XLD Agar (Cat. 1274),
MacConkey Agar (Cat. 1052), EMB Agar (Cat. 1050) or Hektoen Enteric Agar (Cat. 1030).
The medium, which has a coffee color at the beginning, changes to red during incubation at 35-37ºC. A probable
presence of Salmonellae is indicated by small, transparent, either colorless or pink or opaque-white colonies, often
surrounded by a pink or red zone. Some of the uninhibited Gram negative, Lactose/Sucrose fermenting organisms
present opaque green-yellow colonies, surrounded by a yellow halo. Other lactose negative microorganisms, such as
Proteus spp., form colonies of a pale pink or red color, transparent and surrounded by a brilliant red halo.
1
LABORATORIOS CONDA, S.A.
www.condalab.com
Confirmation of suspect samples is carried out by transferring a few of the suspect colonies to TSI (Cat. 1046).
The European Pharmacopoeia recommends in the Paragraph 2.6.13 “Microbiological examination of non-Sterile
products”:
Subculture in this medium after incubation in Brilliant Green Tetrathionate Bile Broth (Cat. 1253), at 41-43ºC for 18-24
hours and incubate at 35-37 ºC for 18-72 h. The probable presence of Salmonellae is indicated by the growth of cultures
having the following appearance in this medium: small, transparent, colourless or pink or opaque-white colonies often
surrounded by pink or red zone. Precise confirmation may be carried out by appropriate biochemical and serological test.
The product passes the test if colonies of the type described do not appear or if the confirmatory biochemical and
serological test are negative.
MICROBIOLOGICAL TEST
The following results were obtained in the performance of the medium from type cultures after incubation at a temperature
of 35-± 2°C and observed after 18-24 hours.
Microorganisms
Escherichia coli ATCC 25922
Salmonella enteritidis ATCC 13076
Growth
Inhibited-Moderate
Good
Staphylococcus aureus ATCC 25923
Salmonella typhi ATCC 19430
*Salmonella typhimurium ATCC 14028
Inhibited
Inhibited – Moderate
Good
*According European Pharmacopeia Incubate at 35-37ºC for 18-72 hours
Colony color
Yellow-green
Pink-white
Red
Pink - white
According ISO 11133 24-48h/37ºC (Productivity)
Microorganisms
Salmonella typhimurium ATCC 14028
Salmonella enteritidis ATCC 13076
Inoculum
(cfu/ml)
103 -104
103 -104
Productivity
Qualitative
Good growth
Good growth
BIBLIOGRAPHY
European Pharmacopoeia. 6th Ed. 2007.
American Public Health Association. Standard Methods for the Examination of Water and Waster water, 11th Edition APHA, New
York, 1960. American Public Health Association. Recommended Methods for the Microbiological Examination of Foods, APHA, Inc.
New York, 1958.
EP
USP
STORAGE
25ºC
Once opened keep powdered medium closed to avoid hydration.
2ºC
2
LABORATORIOS CONDA, S.A.
www.condalab.com