v Principle:-

This test is used to detect the enzymatic ability of an organism  to decarboxylase (hydrolyse) an amino acid to form an amine. Hydrolysis of an amino acid results in an alkaline pH change leading  to formation of pink color.

v Requirements:-

1.      Glucose non-fermenting organisms

2.      Glucose fermenting organisms

3.      Decarboxylase broaths (lysine, arginine, ornithine)

4.      Sterile mineral oil

5.      Suspensions of suspected organisms grown on 5% sheep blood agar (18-24 hours) in brain- heart infusion broth (BHIB).

v Procedure:-

A.    Glucose non-fermenting organisms

1.      Prepare a heavy suspension of the organisms in brain-heart infusion broth.

2.      Inoculate each of the three decarbxylase broths and one control broth  (without amino   acid).

3.      Add a 4 mm layer of sterile mineral oil in each tube.

4.      Incubate at 37° up to seven days.

5.      Observe color change.

  B.  Glucose fermenting organisms

1.      Inoculate each of the decarboxylase tubes with 1 drop of brain-heart infusion broth   culture.

2.      Add 4 mm layer of sterile mineral oil in each tube.

3.      Incubate at 37° up to seven days.

4.      Observe color change.

vObservations: By comparing with the color of the control tube 

      1   Positive:   Purple color (alkaline color change)

           Lysine:        Klebsiella pneumonia

          Arginine:     Enterobacteria cloacae

         Ornithine:    Enterobacteria cloacae

       Negative:     No color change or yellow color

         (Due to fermentation of glucose in BHIB).

v  Use following microorganisms to confirm the reliability of reagents:

1   Positive test:       Lysine:      Klebsiella pneumonia

                                  Arginine:   Enterobacter cloacae

                                 Ornithine:  Enterobacter cloacae

 2  Negative test:      Lysine:       Enterobacter cloaeae

                                   Arginine:    Klebsiella pneumonia

                                  Ornithine:   Klebsiella pneumonia

v Note

1         Reference Book Text book of medical laboratory Technology by Dr. Praful B. Godkar