ANTIBIOTIC SENSITIVITY TEST

 ANTIBIOTIC SENSITIVITY TEST

Aim

            To determine the sensitivity (sensitive or resistant) of Antibiotics against bacterial pathogens.

Principle

            An antibiotic sensitivity (or susceptibility) test is done to help choose the antibiotic that will be most effective against the specific types of bacteria infecting an individual person. One method that is used to determine antibiotic susceptibility is the sensitivity disk method of Kirby - Bauer (named after W. Kirby and A. W. Bauer in 1966). In this method, antibiotics are impregnated onto paper disks and then placed on a seeded Mueller-Hinton agar plate using a mechanical dispenser or sterile forceps. The plate is then incubated for 16 to 18 hours, and the diameter of the zone of inhibition around the disk is measured. The inhibition zone diameter that is produced will indicate the susceptibility or resistance of a bacterium to the antibiotic. Antibiotic susceptibility patterns are called Antibiograms. The Kirby-Bauer method is not restricted to antibiotics. It may also be used to measure the sensitivity of any microorganism to a variety of antimicrobial agents such as sulfonamides and synthetic chemotherapeutics.

Requirements

1. 24 Hours old culture of Bacteria.
2. Mueller Hinton agar plates
3. Antibiotic discs
4. Sterile cotton swabs
5. Bunsen burner
6. Glass markers
7. Ruler  

Procedure

a)     A sterile cotton swab is taken and dipped into a pure culture of bacteria was separately inoculate into Mueller Hinton Agar (MHA) plate by Swabing method first in horizontal direction and then is vertical direction.

b)     Agar surface was allowed to dry 5 minutes.

c)   Different antibiotic discs were placed on seeded Mueller Hinton agar surface using flamed sterile forceps.

d)     The plate was incubated at 37 °C for 24 hours.

e)     The plates were examined for the zone of inhibition around the disc (6 mm in diameter).

f)      Diameter of the zone of inhibition was measured as mm in diameter using the ruler.

Observation and Result

·       Sensitive – Zone of inhibition observed

·       Resistant – No zone of inhibition observed

(Zone formation – Sensitive; No zone formation – Resistance)

Antibiotic sensitivity test (Kirby – Bauer’s Disc method)