Required practical 2 - Microbiology

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 with bacteria into 3 segments.
• Use forceps to place filter paper disc with antiseptic into 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 the clear zones (zones of inhibition) using a ruler from two opposite direction - calculate a mean of measurements.
• Calculate area of clear zones.

Question 3

How would you measure 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 zone of inhibition twice?

Answer 5

Clear zones are not always uniform - taking more that 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 = πr²