OF BASAL MEDIUM (HUGH AND LEIFSON)
CAT Nº: 1500
For the identification of non-fermenting bacilli
of medical and sanitary importance
FORMULA IN g/l
Sodium Chloride
5.00
Bromothymol Blue
0.03
Casein Peptone
2.00
Bacteriological Agar
2.50
Dipotassium Phosphate
0.30
Final pH 7.1 ± 0.2 at 25ºC
PREPARATION
Suspend 9.8 grams of the medium in one litre of distilled water. Heat with frequent agitation until dissolved. Sterilize in
autoclave at 121°C for 15 minutes. Add 10 ml of 10% glucose (or any suitable sugar) solution, sterilized by filtration, to
100 ml of liquid medium. Mix and aseptically dispense 5 ml per tube. If preferred, add 1.0 grams of carbohydrate directly
to 100 ml. of medium and sterilize in the autoclave at 118°C for 10 minutes to avoid the degradation of the sugar. The
prepared medium should be stored at 2-8°C. The color is green-bluish.
The dehydrated medium should be homogeneous, free-flowing and beige with a greenish tint in color. If there are any
physical changes, discard the medium.
USES
OF BASAL MEDIUM is a semisolid medium, prepared according to Hugh and Leifson’s formula, and is used to determine
the metabolism (oxidation, fermentation) of Gram-negative bacteria. It is useful for Pseudomonas, Salmonella, Shigella
and Alcaligenes.
Casein peptone provides nitrogen, vitamins, minerals and amino acids essential for growth. Bromothymol blue is the pH
indicator. Sodium chloride supplies essential electrolytes for transport and osmotic balance. Dipotassium phosphate acts
as a buffer system. Bacteriological agar is the solidifying agent.
Inoculate 2 fresh tubes by stabbing with a fresh culture of the organism in study. If the medium has been prepared and
stored, remelt in a water bath to expel the dissolved gases.
After inoculation, add a layer of 4 to 5 mm of paraffin oil to one of the tubes. It is not recommended to use mineral oil.
Incubate both tubes at 35°C for 48 hours or more, up to 7 days with the caps loose. To facilitate the identification of
Gram-negative non-fermenting bacilli, also use Indole Nitrate Medium (Cat. 1504).
Results:
1 Fermentation: Yellow color in both tubes with or without gas.
2 Oxidation: Yellow color only in tube without oil.
3 No oxidation/fermentation: No change in the color of the tubes. The carbohydrates have not been
fermented or oxidized. Inert microorganisms, e.g. Alcaligenes faecalis.
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 48 - 72 hours.
1
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Microorganisms
Alcaligenes faecalis ATCC 8750
Escherichia coli ATCC 25922
Pseudomonas aeruginosa ATCC 27853
Salmonella enteritidis ATCC 13076
Shigella flexneri ATCC 12022
Without sugar
O
K
K
K
K
K
O
K
K
K
K
K
With glucose
O
K
AG
A
AG
A
O
K
AG
K
AG
A
With lactose
O
K
AG
K
K
K
With Sucrose
O
O
K
AG
K
K
K
K
K
K
K
K
O
K
K
K
K
K
 = Opened
 = Closed
K = Alkaline, green (without change)
A = Acid, yellow
G = Gas, sometimes perceptible
BIBLIOGRAPHY
Hugh, R. and Leifson, E.J. Bact. 66:24-26. 1953. Lisenko J. Gen. Microbiol., 35:379, 1961. Edwards y Ewing Identification of
Enterobacteriaceae. Burguess Publ. Co. Minneapolis, Minn., 1972.
STORAGE
25ºC
Once opened keep powdered medium closed to avoid hydration.
2ºC
2
LABORATORIOS CONDA, S.A.
www.condalab.com