The effects of agar depth, Inoculum size, cations & method of application on antibiotic inhibition zones and the effects of antibiotics on different strains of bacteria.
Introduction:
Agar is a growth medium that contains the essential mix of minerals and nutrients required for the optimum growth of bacteria. The agar can be changed to better support different types of bacteria and it provides the ideal surface to investigate the bacteriostatic capabilities of antibiotics.
In the presence of an antibiotic, bacteria will fail to grow and to determine how effective the antibiotic is it can be tested through agar plate diffusion. By allowing bacteria to grow on a plate treated with antibiotics we can observe the clear area around the antibiotic called the zone of inhibition. This size of this zone tells us how effective the antibiotic is, a large zone, sometimes larger than the plate, means the antibiotics is effective at low concentrations, whilst a small zone means a higher concentration is needed to have the same effect. From this we can compare the effects of different antibiotics on different bacteria.
Concentration of the antibiotic is not the only thing to affect the zone of inhibition; many factors like agar depth all have distinct effects on the antibiotic. In this practical we will be using the bacteria E.coli, P.aeruginosa & S.aureus and the antibiotics Streptomycin & ampicillin to investigate four factors being tested; (i) Agar depth by pouring out 10, 20, 30 ml of agar into plates, (ii) Inoculum size by growing a diluted and undiluted culture of E.coli, (iii) Cations, through mixing KCl and CaCl2 into the agar, the effects of the monovalent K+and divalent Ca2+ can be observed,(iv) Method of administering antibiotic, either by soaking the sterile discs or pipetting the antibiotics onto them.
Finally and gradient plate will be setup to compare the effects on an antibiotic on three strains of unknown bacteria.
Method: (as shown in module