Cetrimide is a quaternary ammonium, cationic detergent that is toxic to most bacteria, except Pseudomonas aeruginosa and a few other bacteria. Nitrogen and phosphorus are released from the bacterial cell and growth is inhibited when they come in contact with cetrimide. However, P. aeruginosa and a few other pseudomonads able to withstand this germicidal activity and hence growth is observed on the inoculated slant of agar medium containing cetrimide for a positive result. Magnesium chloride and potassium sulfate in the medium enhance the production of pyocyanin and pyoverdin (fluorescein) by P. aeruginosa. If the colonies of non-glucose-fermentative, gram-negative rods are isolated from the media containing cetrimide, the suggestive organism is P. aeruginosa.

Requirements:

i. Cetrimide agar: composition

• Gelatin          —————————– 20.0 g/ltr
• K2SO4                         ——————————-              10.0 g/ltr
• MgCl2            ——————————               1.4 g/ltr
• cetyltrimethylammonium bromide —-               0.3 g
• Agar              ————————-        13.6 g
• Glycerin       ——————–                             10.0 ml
• Four milliliters of a 22.5% solution of cetrimide (0.9 g of hexadecyltrimethylammonium) can substitute for the cetyltrimethylammonium bromide.
• Final pH- 7.2
• Sterile inoculating loops or sticks

ii. UV light (360 nm)

iii. Positive control: P. aeruginosa ATCC 27853= growth; yellow-green to blue pigment

iv. Negative control:  Escherichia coli ATCC 25922= inhibited

Procedure:

1. Prepare a slanted cetrimide agar media in a test tube
2. Pick a well isolated colony from cultured agar plate and streak the slant back and forth with inoculum loosely place the cap on the tube
3. Incubate aerobically at 37 °C for up to 7 days.
4. Observe for growth and pigment.
5. If no pigment is visible, examine growth under UV light for the presence of fluorescein.
6. If pigment is still not visible in UV light then incubate for 24 hrs in the dark to enhance pigment production.

Result interpretation:

• Positive result:
  • Growth is observed with a yellow-green (fluorescein) to dark blue-green (pyocyanin) colored fluorescein
  • P. aeruginosa grows on cetrimide agar and produces a blue-green (pyocyanin) pigment.
• Negative result: No growth is observed

Cetrimide test: Principle, Requirements, Procedure and Result interpretation

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