Culturing microorganisms and measuring growthEdexcel International A Level Biology: Subtopic test

1. 1
   A technician is preparing nutrient agar plates and inoculating them with a culture of a non-pathogenic strain of Escherichia coli for a practical class. She works near a lit Bunsen burner and uses a metal inoculating loop. The plates will be incubated for 48 hours. The technician must prevent the culture from being contaminated by other microorganisms and must prevent the culture from escaping into the laboratory.

   (a)

   The technician passes the neck of the culture bottle through the Bunsen flame after removing the lid. What is the main purpose of this?

   [1 mark]

   • ATo warm the culture to its optimum temperature
   • BTo sterilise the neck of the bottle and create an upward air current that carries airborne microorganisms away
   • CTo kill the bacteria in the culture so it is safe to handle
   • DTo increase the oxygen concentration in the bottle

   (b)

   School laboratories normally incubate cultures at 25 °C or below, not at 37 °C. What is the reason?

   [1 mark]

   • ABacteria cannot grow at 37 °C
   • BAgar melts at 37 °C
   • CEnzymes in bacteria are denatured at 25 °C
   • DHuman pathogens grow best near 37 °C, so a lower temperature reduces the chance of growing them

   (c)

   Explain two aseptic techniques that the technician should use when inoculating the plates.

   [2 marks]

   Total for question 1: 4 marks

2. 2
   A food scientist wants to find the number of viable bacteria in a sample of milk. She makes a serial dilution by transferring 1 cm³ of the milk into 9 cm³ of sterile water, mixing, and repeating the transfer with 1 cm³ of each dilution to give dilutions of 10⁻¹, 10⁻², 10⁻³ and 10⁻⁴. She spreads 0.1 cm³ of the 10⁻³ and of the 10⁻⁴ dilutions onto separate nutrient agar plates and incubates them. The 10⁻³ plate has too many colonies to count and the 10⁻⁴ plate has 42 colonies.

   (a)

   What is the dilution factor of each step in this serial dilution?

   [1 mark]

   • A10
   • B9
   • C100
   • D0.1

   (b)

   The count from the agar plates is described as a viable count. What does a viable count measure?

   [1 mark]

   • AThe total number of cells, living and dead
   • BThe mass of the cells
   • COnly the living cells that can divide and form a colony
   • DThe number of different species present

   (c)

   Calculate the number of viable bacteria in 1 cm³ of the original milk.

   [2 marks]

   Total for question 2: 4 marks

3. 3
   A flask of nutrient broth was inoculated with bacteria and incubated at 37 °C. Samples were taken at intervals and the viable count was found by dilution plating. The count was 5.0 × 10⁴ cells cm⁻³ at 2 hours, when the culture had entered its exponential phase, and 3.2 × 10⁶ cells cm⁻³ at 8 hours, when it was still in this phase. The count then stayed at about 5 × 10⁸ cells cm⁻³ between 14 and 20 hours and fell steadily after 20 hours.

   (a)

   Explain the changes in the viable count between 14 and 20 hours and after 20 hours.

   [3 marks]

   (b)

   Calculate the growth rate constant, k, for the exponential phase between 2 and 8 hours, and explain why this value could not be used to predict the count at 20 hours.

   [4 marks]

   Total for question 3: 7 marks

4. 4
   A student investigates the growth of yeast in a liquid culture of glucose broth at 30 °C in a shaking incubator, over 8 hours. She wants to monitor growth throughout this time. She has access to a colorimeter, a haemocytometer and microscope, sterile dilution and agar plating equipment, and a drying oven and balance.

   (a)

   Describe how the student could set up the culture and monitor its growth over 8 hours using a colorimeter, so that her results are valid.

   [6 marks]

   (b)

   Evaluate the use of turbidity, direct cell counts, dilution plating and dry mass for measuring the growth of the yeast culture.

   [6 marks]

   Total for question 4: 12 marks

End of questions