Bacterial lawn

1. Bacterial lawn Learning objectives • To be able to describe how to measure the concentration or potency of a substance by its effect on living cells or tissues (bioassay) • To be able to describe the use of a bacterial lawn as a bioassay technique to determine the effectiveness of antibiotics and disinfectants on inhibition of bacterial growth.

2. A bacterial lawn is a layer of bacteria growing on the surface of an agar plate. • This is used to test the effectiveness of antimicrobial substances such as antiseptics or antibiotics, using a disk diffusion test There are two ways of making a bacterial lawn.

3. Pour plate method • In the pour plate method a universal bottle of warm, sterile, molten agar is inoculated with bacteria, stirred, and then the agar is poured into a plate, where it sets. The bacteria will grow uniformly on the surface.

4. Spread plate method • In the spread plate method an agar plate is inoculated with a small volume of broth culture, which is then spread even over the surface using a glass spreader or a cotton bud.

5. Advantages and disadvantages • The pour plate method • Advantage: • gives a more even lawn, • Disadvantage: • requires universal bottles of agar at just the right temperature. (Why?) • The spread plate method • Advantage: • is easier to do • Disadvantage: • may give an uneven lawn.

6. Testing • Testing may be achieved either by either of 2 methods: • placing antibiotic liquid into wells or ditches which have been cut into the agar, or - applying discs containing measured amounts of antibiotics, which will diffuse out.

9. Testing bacteria for sensitivity to antibiotics • Individual bacterial strains can be tested against a variety of antibiotics (or vice versa) by growing the bacteria as "lawns" on agar in the presence of different concentrations of a single antibiotic, or several different antibiotics may be tested at the same time.

10. Under what circumstances might individual bacterial strains be tested against a variety of antibiotics? • To see if a bacterium causing an infection can be controlled by a specific antibiotic/find the best one (for a particular patient) • Under what circumstances might individual antibiotics be tested against a variety of bacterial strains? • To see if an antibiotic under development by a drug company is likely to work in a given circumstance - i.e. control a certain disease

11. The size of the zone of inhibition - in which bacteria will not grow - gives an indication of the sensitivity of the strain involved, i.e. how easily the bacterial strain will be controlled by the particular antibiotics.

12. In a medical context, the prescription by a doctor of an appropriate antibiotic or dose rate for a particular patient may thus be confirmed from these laboratory tests. How long would these laboratory tests take? Explain why. About 24 hours (or at least overnight) - to give the culture a chance to grow (or be killed!)

13. What is the difference between a disinfectant and a chemotherapeutic agent? What is meant by the following? Therapeutic dose Toxic dose Therapeutic index Broad spectrum antibiotic Narrow spectrum antibiotic Minimal inhibitory concentration Minimum lethal concentration