TBX CHROMOGENIC AGAR
(TRYPTONE BILE X-GLUCURONIDE) ISO 16649-2,3
CAT Nº: 1151
Selective medium for the detection and enumeration of
Escherichia coli in foods
FORMULA IN g/l
Casein Peptone
20.00
X-β-Glucuronide
0.075
Bile Salts Nº3
1.50
Bacteriological Agar
15.00
Final pH 7.2 ± 0.2 at 25ºC
Escherichia coli
ATCC 25922
PREPARATION
Suspend 36.6 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,
mix well and dispense into plates. The prepared medium should be stored at 8-15°C. The color is amber, slightly
opalescent.
The dehydrated medium should be homogeneous, free-flowing and beige in color. If there are any physical changes,
discard the medium.
USES
TBX CHROMOGENIC AGAR is based on Tryptone Bile Salts Agar medium, used to detect and enumerate E. coli in foods,
with the addition of a chromogenic agent, x-ß-D-Glucuronide, to detect the presence of the enzyme glucuronidase,
which is highly specific for E. coli.
The released chromophore in TBX Agar is colored and target colonies are easily identified. E. coli absorbs the
chromogenic agent x-ß-D-glucuronide, and the intracellular glucuronidase enzyme activity breaks the bond between the
chromophore and the glucuronide. The released chromophore is colored and builds up within the cells, causing the E.
coli colonies to be blue-green colored.
Casein peptone provides nitrogen, vitamins, minerals and amino acids essential for growth. Bile Salts are inhibitors to
other Gram-positive organisms and suppress coliform bacteria. Bacteriological agar is the solidifying agent.
ISO 16649 specifies a horizontal method for the enumeration of ß-glucuronidase-positive E. coli in products intended for
human consumption or for the feeding of animals. In part 2, it uses a colony count technique at 44°C on a solid medium
containing a chromogenic ingredient for the enzyme ß-glucuronidase. Inoculate and incubate at a temperature of 44ºC
for 18-24 hours.
ISO 16649-3 uses the most probable number technique. The most probable number of β-glucuronidase-positive E.coli
are determined according to the number of tubes of Minerals Modified Glutamate Broth (Cat. 1365) whose subcultures
have produced blue or blue-green colonies on tryptone bile glucuronide agar. Inoculate and incubate at a temperature of
44°C for 20 – 24 hours.
1
LABORATORIOS CONDA, S.A.
www.condalab.com
Note: The negative b-ß-glucuronidase-negative E. coli colonies are colorless, e.g. E. coli O157:H7. The high
temperatures (44°C) inhibit the growth of E. coli O157:H7.
MICROBIOLOGICAL TEST
The following results were obtained from type cultures in the performance of the medium after incubation at a
temperature of 37ºC and 44°C and observed after 18-24 hours.
Microorganisms
Growth
44ºC 37ºC
Colony Color
Good
Good
Blue-Green
Salmonella typhimurium ATCC 14028
Inhibited
Inhibited
Colorless
Enterococcus faecalis ATCC 19433
Inhibited
Inhibited
-
Klebsiella pneumoniae ATCC 13883
Inhibited
Inhibited
Colorless
Escherichia coli ATCC 25922
According to 11133(24 h/44ºC) Productivity and Selectivity
Microorganisms
Escherichia coli ATCC 25922
Enterococcus faecalis ATCC 29212
Inoculum
(cfu/ml)
102
10 4/ 106
Productivity
Quantitative
pr ≥ 0.9
Selectiviity
Qualitative
Specificity
Qualitative
Blue
Inhibited
Reference Media Productivity: TSA
BIBLIOGRAPHY
International standard ISO 16649-2 Microbiology of food animal feeding stuffs. Horizontal method for the enumeration of presumptive
ß-glucuronidase –positive. Part 2:Colony-count technique at 44°C using 5-bromo-4-chloro-3-indoly ß-D-glucuronide. Part 3: Most
probable number technique using 5-bromo-4-chloro-3-indoly ß-D-glucuronide
ISO
STORAGE
25ºC
Once opened keep powdered medium closed to avoid hydration.
2ºC
2
LABORATORIOS CONDA, S.A.
www.condalab.com