Butyrate Disk Test- Principle, Procedure and Result Interpretation

Purpose of Butyrate Disk Test

This is a rapid test to detect the enzyme butyrate esterase, to aid identification of Moraxella (Branhamella) catarrhalis.

Principle of Butyrate Disk Test

Organisms capable of producing butyrate esterase hydrolyze bromochlorindolyl butyrate. Hydrolysis of the substrate in the presence of butyrate esterase releases indoxyl, which in the presence of oxygen spontaneously forms indigo,
a blue to blue-violet color.

Procedure of Butyrate Disk Test

  1. Remove a disk from the vial and place on a glass microscope slide.
  2. Add 1 drop of reagent-grade water. This should leave a slight excess of water on the disk.
  3. Using a wooden applicator stick, rub a small amount of several colonies from an 18- to 24-hour pure culture onto the disk.
  4. Incubate at room temperature for up to 5 minutes.

Result Interpretation of Butyrate Disk Test

Positive: Development of a blue color during the 5-minute incubation period.
Negative: No color change.

Limitations of Butyrate Disk Test

Incubation longer than 5 minutes may result in a false-positive reaction. False-negative reactions may occur if the inoculum is too small. If the organism is negative, repeat with a larger inoculum and follow-up with additional methods.

Quality Control

Positive: Moraxella catarrhalis (ATCC25240)— formation of blue color
Negative: Neisseria gonorrhoeae (ATCC43069)—no color change