Amino acid Decarboxylase Test Principle, Procedure, Results, Uses

Decarboxylase Test Definition

The Decarboxylase Test is a biochemical method. It is used to determine an organism’s ability to produce decarboxylase enzymes. These enzymes remove the carboxyl group (–COOH) from specific amino acids.

This reaction results in the formation of amines and carbon dioxide (CO₂), leading to a rise in pH.

Objectives of Decarboxylase Test

• To detect the presence of Decarboxylase enzymes produced by bacteria.
• To differentiate bacterial species based on their ability to decarboxylate specific amino acids.
• To assist in the identification of Enterobacteriaceae and other gram-negative bacilli.

Principle of Decarboxylase Test

The test is based on the ability of bacteria to Decarboxylase amino acids, producing alkaline amines. These reactions occur in anaerobic conditions and are usually glucose-dependent. The pH indicator (e.g., bromocresol purple) helps detect the pH change from acid to alkaline.

Microorganisms Tested

• Members of Enterobacteriaceae (e.g., Klebsiella, Enterobacter, Salmonella)
• Some non-enteric gram-negative rods
• Differentiation within Proteus, Shigella, etc.

Media, Reagents, and Supplies Used

Media

• Moeller’s Decarboxylase Base Medium
• Specific amino acid supplements: lysine, ornithine, arginine

Reagents Used

• pH indicator (e.g., bromocresol purple)
• Mineral oil (to supply anaerobic environment)

Supplies Used

• Test tubes
• Inoculating loop
• Incubator (35–37°C)
• Sterile pipettes or droppers

Procedure of Amino Acid Decarboxylase Test

A. Preparation of the Media

1. Prepare Moeller’s base medium according to manufacturer’s instructions.
2. Add the specific amino acid (e.g., 1% L-lysine).
3. Dispense into test tubes and sterilize by autoclaving.

B. Decarboxylase Test

For Glucose-Fermenting Organisms

1. Inoculate the test organism into two tubes:
   • One containing only base medium (control)
   • One with the amino acid supplement
2. Overlay both with 1 cm of sterile mineral oil.
3. Incubate at 35–37°C for 18–48 hours.
4. Observe color change:
   • Yellow → acidic fermentation
   • Purple → alkaline reaction (positive decarboxylation)

Glucose-Nonfermenting Organisms

• May not acidify the medium adequately to induce decarboxylase enzyme.
• Extended incubation or use of different testing methods may be necessary.

Control Organisms

• Positive control: Enterobacter aerogenes (for lysine/ornithine)
• Negative control: Shigella dysenteriae

Result Interpretation of Amino Acid Decarboxylase Test

• Purple (alkaline): Positive – decarboxylation occurred
• Yellow (acidic): Negative – no decarboxylation
• No change: Invalid or inconclusive – check for growth

Uses of Decarboxylase Test

• Identification and differentiation of Enterobacteriaceae
• Differentiation of Proteus species
• Diagnostic support in clinical microbiology

Limitations of Decarboxylase Test

• False negatives with glucose-nonfermenters
• Requires optimal pH and anaerobic conditions
• Some organisms require prolonged incubation
• Media quality and amino acid concentration affect outcomes