BRUCELLA AGAR
CAT Nº: 1012
For the cultivation of Brucella in diverse clinical specimens, foods and other
materials of sanitary interest
FORMULA IN g/l
Meat peptone
10.00
Yeast Extract
2.00
Casein Peptone
10.00
Dextrose
1.00
Sodium Chloride
5.00
Sodium Bisulfite
0.10
Bacteriological Agar
15.00
Final pH 7.0 ± 0.2 at 25ºC
PREPARATION
Suspend 43.1 grams of the 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 121ºC for 15 minutes. Cool to 45-50ºC
and aseptically add 5% sterile sheep defibrinated blood. Be careful to avoid bubble formation when adding the blood to
the cooled medium and rotate the flask or bottle slowly to create a homogeneous solution. Homogenize gently and
dispense into Petri dishes. Once solidified, invert the plates to dry up any excess of moisture.
Brucella Agar can be made selective to yield higher numbers of positive isolations by aseptically adding two vials of the
Brucella Supplement (Cat. 6017) previously reconstituted in 10 ml of 1:1 solution of methanol / sterile distilled water.
The prepared medium should be stored at 8-15°C. The color is amber, slightly opalescent. The color with blood is cherry
red.
The dehydrated medium should be homogeneous, free-flowing and light beige in color. If there are any physical
changes, discard the medium.
Brucella Supplement (Cat. 6017)
(1 vial for 500 ml of the medium)
Nystatin…………………..50000 IU
Bacitracin……………….12500 IU
Polymyxin B……………….2500 IU
Cycloheximide………………..50 mg
Vancomycin……………………10 mg
Nalidixic Acid………………….2.5 mg
For better growth, Polyenrichment Supplement (Cat. 6011) may be added if required.
USES
BRUCELLA AGAR, being rich in nutrients and growth factors, is very suitable to grow and isolate fastidious
microorganisms.
It is used to successfully isolate Brucella from diverse specimens contaminated with microflora, both saprophytes and
commensals, in clinical samples as well as in foods. This medium is also used to produce clostridial toxins. It can also be
utilized in the isolation of many anaerobes both of human and animal origin. Food samples can be inoculated directly on
the plates of Brucella Agar, while clinical specimens are more convenient as suspensions or macerations in sterile saline
solutions.
The Meat peptone and Casein peptone provide nitrogen, vitamins, minerals and amino acids essential for growth. Yeast
extract is a source of vitamins, particularly of the B-group essential for bacterial growth. Sodium bisulfite is the reducing
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agent; Sodium chloride supplies essential electrolytes for transport and osmotic balance. Dextrose is the fermentable
carbohydrate providing carbon and energy. The addition of blood provides extra growth factors for fastidious
microorganisms. Bacteriological agar is the solidifying agent. The addition of the supplement enhances the medium’s
selectivity for the growth of Brucella.
Brucella species are level 3 pathogens and cause brucellosis, a zoonotic disease. It is usually transmitted through milk,
dairy products, meat and direct contact with infected animals.
Inoculations and incubation at 35 ± 2°C should be made in duplicate - one plate under normal conditions and one plate
under 5 -10% CO2. Observe after 24 - 72 hours.
Note: For an excellent medium for anaerobes, add 5 mg/ml of hemin and 10 mcg/ml of Vitamin K1 (phytomenadione)
to the basal medium, culture and incubate under anaerobic conditions.
MICROBIOLOGICAL TEST
The following results were obtained in the performance of the medium from type cultures, adding the supplements
Brucella Supplement (Cat. 6017), Polyenrichment Supplement (Cat. 6011), and 5% sterile sheep defibrinated blood after
incubation at a temperature of 35 ± 2°C, under 5 -10% CO2 atmosphere, and observed after 24 - 72 hours.
Growth
Inoculum
Brucella abortus ATCC 4315
Good
103-104
Brucella melitensis ATCC 4309
Good
103-104
Brucella suis ATCC 4314
Good
103-104
Microorganisms
BIBLIOGRAPHY
Kzudas and Morse, J. Bact. 66:502. 1953 Rennoux G. Ann. Inst. Pasteur, 87:325. 1954 Standard Methods for the Examination of Diary
Products. 10th Ed. APHA, Inc. New York, 1960
Smith Louis Ds. The Pathogenic Anaerobic Bacteria. C. Thomas Pub. Springfield, Il, 1975.
Koneman, Allen, Dowell, and Sommers. Color Atlas and Textbook of Diagnostic Microbiology, J.B. Lippincott, Philadelphia, 1979.
STORAGE
25ºC
Once opened keep powdered medium closed to avoid hydration.
2ºC
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LABORATORIOS CONDA, S.A.
www.condalab.com