Download Decarboxylase Broth Base - Thermo Fisher Scientific

DECARBOXYLASE BROTH (MOELLER)
BASE CONTROL, ARGININE, LYSINE, and ORNITHINE
INTENDED USE
Remel Decarboxylase Broths (Moeller) are liquid media recommended for use in qualitative procedures to measure the enzymatic ability of an
organism to decarboxylate an amino acid to aid in the differentiation of gram-negative bacilli.
SUMMARY AND EXPLANATION
In 1955, Moeller developed the amino acid decarboxylase media to detect the production of lysine and ornithine decarboxylase and arginine
dihydrolase.1 These media are used primarily to differentiate bacterial groups within the Enterobacteriaceae.2 However, arginine dihydrolase
testing may also be used to aid in the identification of pseudomonads.3 Ewing, Davis, and Edwards compared Moeller medium to Falkow
decarboxylase medium. They reported Moeller’s formulation is more reliable for differentiating Klebsiella and Enterobacter as compared to
other media tested.4 The Moeller method is considered the standard or reference method for determining the decarboxylase reactions of the
Enterobacteriaceae.
PRINCIPLE
The basal medium contains meat peptone and beef extract which supply nitrogenous nutrients to support bacterial growth. Dextrose is an
energy source and fermentable carbohydrate. Bromcresol purple and cresol red are pH indicators. Pyridoxal is an enzyme cofactor which
enhances decarboxylase activity.5 Arginine, lysine, and ornithine are amino acids that are added to the basal medium individually to detect the
production of enzymes which decarboxylate these substrates. When an organism ferments dextrose, acids are produced which lower the pH,
resulting in a color change from purple to yellow. If decarboxylation of the amino acid occurs in response to the acid pH, alkaline end-products
(amines) are formed which result in the medium reverting to its original color (purple). Decarboxylation of lysine yields cadaverine,
decarboxylation of ornithine yields putrescine, and arginine is first hydrolyzed to form ornithine which is then decarboxylated to form
putrescine.
REAGENTS (CLASSICAL FORMULAE)*
Base Control Medium:
Beef Extract ..................................................................... 5.0 g
Meat Peptone .................................................................. 5.0 g
Dextrose .......................................................................... 0.5 g
Bromcresol Purple.........................................................10.0 mg
Cresol Red ..................................................................... 5.0 mg
Pyridoxal ........................................................................ 5.0 mg
Demineralized Water .................................................1000.0 ml
pH 6.0 ± 0.2 @ 25°C
The following optional ingredients are added per liter of medium:
L-Arginine ...................................................................... 10.0 g
L-Lysine......................................................................... 10.0 g
L-Ornithine .................................................................... 10.0 g
*Adjusted as required to meet performance standards.
PROCEDURE
Note: For each test isolate, inoculate a Decarboxylase Broth Base Control in addition to selected Decarboxylase Broth(s) containing an
amino acid (i.e., Arginine, Lysine, and Ornithine).
1. Inoculate Decarboxylase Broth from a pure, 18-24 hour culture growing on an agar plate. Do not inoculate from a broth culture. Using a
sterile inoculating loop or needle, select 1 or 2 colonies of the test isolate and transfer to the Decarboxylase Broth.
2. Mix to distribute the inoculum throughout the medium.
3. Overlay the medium in each tube with 1 ml of sterile mineral oil to protect from air.
4. Incubate, with caps tightened, at 33-37°C for up to 10 days.
5. Examine tubes daily for a purple color change. The control tube contains glucose which is fermented forming a yellow color. If the
control becomes alkaline (purple) the test is invalid.
INTERPRETATION OF THE TEST
Positive Test - A turbid purple to faded out yellow-purple color
Negative Test - A bright, clear yellow color
QUALITY CONTROL
All lot numbers of Decarboxylase Broth (Moeller) Base Control, Arginine, Lysine, and Ornithine have been tested using the following quality
control organisms and have been found to be acceptable. Testing of control organisms should be performed in accordance with established
laboratory quality control procedures. If aberrant quality control results are noted, patient results should not be reported.
CONTROL
INCUBATION
RESULTS
Decarboxylase Broth Base Control:
Klebsiella pneumoniae ATCC® 27736
Enterobacter aerogenes ATCC® 13048
Ambient, 18-24 h @ 33-37°C
Ambient, 18-24 h @ 33-37°C
Negative (yellow)
Negative (yellow)
Decarboxylase Broth Arginine:
Pseudomonas aeruginosa ATCC® 27853
Escherichia coli ATCC® 25922
Ambient, 18-24 h @ 33-37°C
Ambient, 18-24 h @ 33-37°C
Positive (purple)
Negative (yellow)
Decarboxylase Broth Lysine:
Salmonella enterica serovar Typhimurium ATCC® 14028
Citrobacter freundii ATCC® 33128
Ambient, 18-24 h @ 33-37°C
Ambient, 18-24 h @ 33-37°C
Positive (purple)
Negative (yellow)
Decarboxylase Broth Ornithine:
Enterobacter aerogenes ATCC® 13048
Proteus vulgaris ATCC® 6380
Ambient, 18-24 h @ 33-37°C
Ambient, 18-24 h @ 33-37°C
Positive (purple)
Negative (yellow)
(Continued on back)
LIMITATIONS
1.
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4.
Decarboxylase tests can only be performed with microorganisms that can metabolize glucose to produce acid. An acid environment is
required for decarboxylase enzyme activity to occur.2
After standing during incubation, a decarboxylase test may show a yellow layer and a purple layer. Shake the tube gently before
interpreting the result.2
Any trace of purple color should be interpreted as a positive test. The test must be incubated for at least 24 hours to prevent a falsenegative result. Prolonged incubation from 6-10 days or longer may be required to demonstrate weak reactions due to an organism’s
poor decarboxylase activity.2
To obtain the appropriate reactions, the inoculated tubes must be protected from air with a layer of sterile mineral oil. Exposure to air
may cause alkalinization at the surface of the medium due to oxidation and deamination of peptones, causing a decarboxylase-negative
organism to appear positive.2
BIBLIOGRAPHY
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Moeller, V. 1955. Acta. Pathol. Microbiol. Scand. 36:158-172.
MacFaddin, J.F. 1985. Media for Isolation-Cultivation-Identification-Maintenance of Medical Bacteria. Vol. 1. Williams & Wilkins,
Baltimore, MD.
MacFaddin, J.F. 1980. Biochemical Tests for Identification of Medical Bacteria. 2nd ed. Williams & Wilkins, Baltimore, MD.
Ewing, W.H. 1986. Edwards and Ewing’s Identification of Enterobacteriaceae. 4th ed. Elsevier, New York, NY.
Gale, E.F. 1946. Adv. Enzymol. 6:1.
Refer to the front of Remel Technical Manual of Microbiological Media for General Information regarding precautions, product storage and
deterioration, specimen collection, storage and transportation, materials required, quality control, and limitations.
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