Optochin sensitivity test Principle, procedure (streptococcus pneumonia)

An optochin sensitivity test is used for the alpha-hemolytic streptococcus pneumoniae biochemical test.

With the help of this test, we diagnose streptococcus pneumonia another alpha-hemolytic streptococcus.

Optochin sensitivity test principle

Optochin is a chemical (ethylhydrocupreine hydrochloride). It is an antibiotic. It interferes with ATP synthesizing enzyme called ATPase in bacteria. It causes the change in the fragility of the cell membrane of particular bacteria resulting in membrane rapture due to surface tension.

In the optochin sensitivity test, the optochin-impregnated disk is placed in the middle of the test organisms on the blood agar plate.

This antibiotic diffuses in the media. It inhibits the growth of susceptible organisms around itself and creates a clear zone.

Note: 14 mm clear zone is an indication of optochin-sensitive organisms.

The procedure of the optochin sensitivity test

1. Select and pick the isolated colony of alpha-hemolytic organisms with the help of a sterile loop.
2. Streak the organism on the sheep blood agar plate.
3. Now pick the optochin disk with sterile forceps and place it on between the test organisms on the agar plate.
4. Gently press the disk, so that it may adhere to the agar surface.
5. Incubate the plate at 37 C for 24 hours, at 5% CO₂ environments.
6. Observe the plate after 24 hours and note the clear zone around the disk.

NOTE: If the clear area is equal to 15 mm or more than 15 mm, then the organism is optochin test sensitive. But, if the clear area is less than 15 mm then it will be an optochin-resistant organism.

Which bacterias are Optochin sensitive?

Only streptococcus Pneumonia is Optochin sensitive organism. The clear area will be more than 14 mm around the optochin disk.

What bacteria is Optochin resistant?

Alpha hemolytic organism other than Streptococcus pneumoniae is optochin resistant. There will be a small clear zone around the optochin disk.

Optochin sensitivity test with no clear zone

When there is no clear zone around the optochin disk after 24 hours, it will be concluded that the test organism is other than Streptococcus pneumonia. Therefore other biochemical tests are necessary for further identification.

What are the limitations of the Optochin test?

1. The Optochin susceptibility test is only used to identify the Alpha hemolytic organism, not beta-hemolytic organisms.
2. We can not differentiate other alpha-hemolytic organisms on the base small clear zone, for this purpose we need to do more biochemical tests bile solubility tests, and more.
3. For this test, we need a special chamber with 5-10 % CO₂ for bacteria growth.