22.6 Culturing Microorganims In The Lab

Question 1

Why do health and safety procedure need to be carried out when microoorganisms are cultured, even when they’re harmless?

Answer 1

• there is always the risk of a mutation taking place making the strain pathogenic
• there may be contamination with pathogenic microorganisms from the environment

Question 2

Basic technique used to culture microorganisms in the lab?

Answer 2

• on an agar plate (a sterile Petri dish containing agar jelly)
• nutrients can be added to the agar to improve the growing conditions
• microorganisms you use are likely to be in a liquid broth (a mixture of distilled water and nutrients)
• to culture the microorganisms, use a sterile wire inoculation loop to transfer some of the same to the plate
• plates need to be incubated to allow the microorganisms to grow

Question 3

Why is aseptic technique important?

Answer 3

To prevent contamination of cultures by unwanted microorganisms, which may affect the growth rate of the bacteria being cultured
- leads to imprecise result and may be a health hazard

• contamination on an industrial scale can be very costly as entire cultures are affected and would need to be thrown away

Question 4

Examples of aseptic technique

Answer 4

• disinfect work surfaces
• work near Bunsen flame, hot air removes micro-organisms away from your culture
• sterilise loop wire before and after each use (e.g. by passing it through Bunsen flame)
• briefly pass the neck of the broth container through a Bunsen flame just after its opened and just before its closed
• minimise the time to agar plate is open, keep lid on after each streak
• sterilise glassware (e.g. using autoclave)
• wear a lab coat

Question 5

To investigate factors that affect growth of microorganisms

Answer 5

Temp: place agar plates at diff temperatures (e..g one in the fridge and the other in an incubator), then count the the number of colonies that have formed

-pH: add buffers at diff pH levels to the broth (contains the sample of microorganisms)

Nutrient availability: use agars that have been prepared differently, which contain different nutrients

Question 6

What’s a closed culture

Answer 6

When growth has taken place in a vessel thats isolated from external environment (e.g. extra nutrients aren’t added and waste products aren’t removed from vessel during growth)

Question 7

Describe and explain the growth curve in a closed culture, e.g. batch fermentation?

Answer 7

Standard growth curve, has 4 phases

1) Lag phase: the population size increases slowly because the microorganisms have to make enzymes and other molecules before they can reproduce. Reproduction rate is slowly.

2) Exponential (log) phase: population size increases quickly because the culture conditions are most favourable for reproduction (lots of food and little competition). The number of microorganisms doubles at regular intervals

3) stationary phase: population size stays level because death rate = reproduction rate (by binary fission) so total growth rate is zero

4) decline phase: death rate is greater than reproduction rate. This because food is very scarce and waste products are at toxic levels

Question 8

Limiting factors which prevent exponential growth of culture of bacteria

Answer 8

• nutrient availability
• oxygen levels
• temperature (due to enzyme controlled reactions within microorganisms)
• build up of waste (toxic material may inhibit further growth or even kill the culture)
• change in pH (as CO2 produced by respiration of bacterial cells, pH falls which can affect enzyme activity)