Sensitivity of Antibiotics in Bacteria

In this article we will discuss about the Sensitivity of Antibiotics in Bacteria.

Principle:

Antibiotics are chemical substances produced, due to metabolism, by some microorganisms, which in small concentrations are capable of inhibiting the growth of other microorganisms. Antibiotics differ chemically and in their mode of action from one ano­ther. The sensitivity to any antibiotic depends on the type of microorganism against which it is used and the age of the culture.

Some microbes are very sensitive to a particular antibiotic while others are not and the sensitive ones are again resistant to some other antibiotic. To find out the sensitivity of one microorganism against different antibiotics, the test organism is spread on an agar plate and different anti­biotics of high conc. (100 µgm./ml) are applied in two ways (paper disc and cup) at the centre of a marked quadrant.

After incubation for 24 hrs. The zone of inhibition around the cup or disc is measured. The diameter of the zone of inhibition is directly pro­portional to the strength of the antibiotic as well as to the sensitivity of the microbe. Zone of inhibition is the zone on agar plate at which growth of the test organisms docs not occur.

Materials:

(i) 24 hours broth culture of:

(a) Escherichia coli,

(b) Staphylococcus aureus,

(c) Serratia marcescens.

(ii) Antibiotics

(a) Benzyl ponicillin,

(b) Streptomycin,

(c) Chloramphenicol,

(d) Oxytcfracyclin,

(iii) Dry nutrient agar plates

(iv) Cork borer

(v) Paper discs

(vi) Pipettes and petridishes (sterilised)

(vii) Sterilised distilled water

(viii) Spreader, forceps etc.

Procedure:

Nutrient agar medium is prepared, sterilised and then poured into sterilised petridishes each containing 20 ml. After the plates have solidified, a quadrant is marked on the lower surface of the petridish. 0.2 ml. of test organism is uniformly spread all over the plate with a glass spreader.

A. Cup Method:

A cork borer is sterilised by dipping into alcohol and flaming and 4 cups are cut in the centre of the 4 quadrants of the plate spread with the test organism.

10 µgm. of each of the above named antibiotics are dissolved in 10 ml. of sterilised distilled water to make 100 µgm./ml. solution.

With sterilised pipettes, the 4 cups of each plate are filled with 4 different antibiotic solutions. The plates are incubated at 37°C for 24 hrs. and zone of inhibition noted with its diameter being measured.

B. Paper Disc Method:

In the paper disc method, paper discs of filter paper are sterilised. A few of them are soaked in antibiotic solution for 5 minutes in sterilised petridishes and then are transferred to another sterilised petridish for drying.

With the help of a sterilised forceps, the different antibiotic soaked paper discs are placed in the centre of different quadrants of the plate with test organism. The plates are then incubated in an inverted position at 37°C for 24 hours and the zone of inhibition measured. In the cup method, the plates with cups of antibiotics are incubated in normal position without any disturbance.

Results:

Conclusion:

From the diameter of zone of inhibition, it is evident that E. coli is resistant to penicillin and most sensitive to oxy-tetracyclin.

The diameter of zone of inhibition is maximum for penicillin (3 cm), in case of S. aureus. Hence it is most sensitive to penicillin amongst the antibiotics tested.

S. marcescens is most sensitive to chloram­phenicol and resistant to penicillin. Thus, the sensitivity of an organism to an antibiotic varies.