The principle of the disc diffusion method is that when a filter disc is impregnated with a chemical that is placed on the agar, the chemical in the agar diffuses around the disc. The solubility of the chemical and its molecular size determines the size of the area if chemical infiltration around the disc. When an organism is placed on the agar around the disc, if it is susceptible to the chemical then there would be no growth around the area where the chemical is diffused and this area is called as the "zone o the inhibition".
This inhibition can be measured and compared with a standard control strain and normal tables, Stokes and Kirby-Bauer method respectively. The factors that affect in this method are the concentration of the bacterial inoculums, the depth and types of agar, the incubation conditions and the time of incubation. All this factors should be always taken into account while performing the test. An alternative test that can be performed is the determination of the amount of antibiotic required to either inhibit the growth of organism or to kill the organism which is done by incubating a fixed concentration of the organism in increasing concentrations of antibiotics and checking for growth after 24h of incubation.
The as3. Methods.
The assigned culture was swabbed onto the agar plate in three directions for maximal coverage. The plates were allowed to settle for 10 mins, followed by impregnating the antibiotic discs by pushing the dispenser over the agar. The antibiotic and the disc code were noted and the plates
were kept in incubator at 37C.
4. Results. The results obtained from Test 1 and Test 2 are displayed in Table 1 and 2 respectively.
RESULTS: TEST 1
RESULTS: TEST 2
5. Discussion: Antibiotic susceptibility of given strain was determined by Kirby-Bauer disc diffusion method. Disc diffusion method is widely used across the laboratory to determine the effect of any anti- microbial agents like antiseptics, antibiotics, bactericidal or bacteriostatic compound etc. To avoid any lab to lab variation standardized protocol was developed that was recommended by National Committee for Clinical laboratory standards like Meuller-Hilton Agar is used as growth medium; plates will be incubated at 37C for 18-24 hr, while turbidity of bacterial suspension will be set at 0.5 McFarland method or 1.0 OD by spectrophotometer. The biggest advantages of disc diffusion method over other methods are that it is rapid, efficient, cost effective and reliable. While there are certain limitation of this methods like any variation in cell number, incubation time, diffusibility of antibacterial compound, media etc, which leads to very high variation in final results. Similarly, it gives only quantitative idea and determination of bactericidal concentration or MIC need to be back calculated. For pathological determination serum to antibiotic ratio must be taken in to consideration to evaluate effective dose. In this practical, we evaluated three cultures for its Antibiotics susceptibility against various antibiotics. Among these Pseudomonas