1.1.7 Culturing Microorganisms

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Microorganisms are very small, so in order for scientists to study them they need to grow many of them in the lab using nutrients (culturing them).The culture medium contains carbohydrates for energy, minerals, proteins and vitamins.

There are two ways to grow microorganisms in the lab:

In nutrient broth solution- involves making a suspension of bacteria to be grown and mixing with sterile nutrient broth (the culture medium), stoppering the flask with cotton wool to prevent air from contaminating it and shaking regularly to provide oxygen for the growing bacteria.
On an agar gel plate- the agar acts as the culture medium, and bacteria grown on it form colonies on the surface.

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Making the plate:

Hot sterilised agar jelly is poured into a sterilised Petri dish, which is left to cool and set
Wire loops called inoculating loops are dipped in a solution of the microorganism and spread over the agar evenly
A lid is taped on and the plate is incubated for a few days so the microorganisms can grow (stored upside down)

The reasons why we follow certain steps in this procedure need to be understood.

Step	Why ?
Petri dishes and culture media must be sterilised before use, often done by an autoclave (an oven) or UV light.	If this step does not take place, they are likely to be contaminated with other microorganisms. These could be harmless but will compete with the desired bacteria for nutrients and space, or they could be harmful (for example through a mutation taking place), potentially producing a new pathogen.
Inoculating loops must be sterilised by passing them through a flame.	This kills unwanted microorganisms, which is needed for reasons above.
The lid of the Petri dish should be sealed (but not completely) with tape.	Sealing stops airborne microorganisms from contaminating the culture, but it should not be sealed all the way around as this would result in harmful anaerobic bacteria growing (due to no oxygen entering).
The Petri dish should be stored upside down.	This is to prevent condensation from the lid landing on the agar surface and disrupting growth.
The culture should be incubated at 25 degrees.	If it were incubated at a higher temperature, nearer 37 degrees (human body temperature), it would be more likely that bacteria that could be harmful to humans would be able to grow as this is their optimum temperature. At lower temperatures, colonies of such bacteria would not be able to grow.

If they have a supply of nutrients and a suitable temperature, bacteria can multiply by binary fission (one splitting into two) as fast as every 20 minutes. You can calculate the number of bacteria in a population after a certain time if given the mean division time.

The formula is: bacteria at beginning x 2 number of divisions = bacteria at end

To calculate the number of divisions, divide the time the population is left for by the mean division time for that bacteria
The number of bacteria at the end of the growth period can be very large, so it is common for it to be left in standard form (see above in 'microscopy' for more on standard form)

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If the microorganisms are bacteria, they can be used to test the effects that different antibiotics (or disinfectants) have on their growth. The investigation involves soaking paper discs in different antibiotics, which are placed on an agar plate with bacteria. After leaving the plate, the size of the clear area around the discs shows how many bacteria have died, indicating therefore how effective the antibiotic is.

Soak the paper discs in different types/concentrations of antibiotics and place on an agar plate evenly spread with bacteria. One disc should be a control, soaked in sterile water. There should be no death of bacteria with this disc- showing only the type of antibiotic affects the size of the inhibition zone (the clear area left when they die).
If the bacteria are resistant to the antibiotic they will not die, but non-resistant will die, leaving an inhibition zone.
Leave the plate at 25 degrees for 2 days.
The zone of inhibition can be measured- the bigger it is, the more bacteria are killed and therefore the more effective the antibiotic is.

In both investigations- growing bacteria and testing the effectiveness of antibiotics- you need to calculate cross-sectional areas (of colonies or inhibition zones). This involves using the formula $πr^2$ , where r is the radius of the circle.

Culturing Microorganisms Simplified Revision Notes for GCSE AQA Biology

1.1.7 Culturing Microorganisms

There are two ways to grow microorganisms in the lab:

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