CP05 - Microbial Cultures

Question 1

What antiseptic techniques can be used to avoid contamination?

Answer 1

● Spray working area with disinfectant and wipe dry
● Wash hands with antibacterial wash
● Flame the neck of the culture bottle
● Lift the lid of the agar plate at an angle
● Dip spreader in ethanol and pass through Bunsen flame before spreading bacteria
● Use forceps to place antibiotic discs

Question 2

Describe how you could investigate the effect of antibiotics on bacterial growth using zones of inhibition

Answer 2

● Divide agar plate with bacteria into three segments
● Use forceps to place filter paper disc with antiseptic in each zone
● Loosely tape lid onto agar plate to allow oxygen to reach bacteria
● Incubate at 25°C for 48 hours
● Measure the diameter of clear zones (zones of inhibition) using a ruler
from two opposite directions - calculate mean of measurements
● Calculate area of clear zones

Question 3

How would you measure the zone of inhibition?

Answer 3

Use a ruler to measure from a point on one side to a point directly opposite (with lid still in place)

Measure again at 90° to first diameter measurement in order to calculate a mean

Question 4

Why should you not completely seal the agar plate?

Answer 4

To allow oxygen to enter the agar plate, preventing the growth of harmful anaerobic bacteria

Question 5

Why is it necessary to measure the diameter of the zone of inhibition twice?

Answer 5

Clear zones are not always uniform - taking more than one measurement allows a mean diameter to be calculated

Question 6

What equation is used to calculate the area of clear zones?

Answer 6

Area = πr^2

Question 7

What is a source of error in this practical?

Answer 7

The area of the clear zone may be irregular and it is difficult to determine a suitable width to be recorded.