Microbiology

Classification

The three classification of bacteria you need to know.
Can also be classified by Gram staining:
-All bacteria has polysaccharide and polypeptide in there membrane, Gram negative bacteria also have lipopolysaccharide which means penicillin and lysozome don't affect it.
-Gram staining produces a violet colour for positive and red for negative.
-Gram negative do not retain the dye and only are shown by the counterstain (red).

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Nutrients - On a agar plate with the glucose and nitrogen containing material for growth.
Temperature - The range of 25-45 degrees provides a favourable temperature.
pH - Most bacteria prefer slightly alkali conditions, and fungi slightly acidic.
Oxygen - Many require O2 for metabolism, some prefer not to, and some cannot survive in it.
-Obligate aerobes: Respire aerobically and require O2.
-Facultative aerobes: Prefer O2, but can survive without it.
-Obligate anaerobes: Cannot survive in presence of O2.

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Sterilise all apparatus and media before intitial contamination.
Sterilise the work surface with work surface with disinfectant.
Hold the culture bottle in the little finger to prevent not putting it on the workplace.
Flame the mouth of the bottle for 2-3 seconds.
Inoculating loop requires flaming initially.
Lift the lid only slightly.
Secure the lid with a little tape, do not make anaerobic.
-This produces harmful anaerobes.
Incubate at 25 degrees C.
-If incubated at 37 degrees C then higher chance of human pathogens.
To sterilise afterwards use an autoclave, high temperatures and pressure kills any bacteria.

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Rough estimate can be made with measuring the diameter of the colony, finding the rate is called the dilution plating technique.
Serial dilution allows for a colony to be diluted into countable amounts.
-1cm3 of the solution to be added to added to 9cm3 of sterile saline, doing this several times gives a small sample which can be added to an agar sample and produces countable colonies.
More accurate method which determines total number of cells, but not just a viable count.
A colorometer measures the turbidity of a solution, which is then is used to work out total count of cells.

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Rapid production of cells, low temperature, and relatively cheap to do on a large scale.
The following describes a batch fermenter:
-A pure culture is needed in a sterile vessel.
-Must have suitable conditions and a sterile medium.
-Must be protected from contamination and constantly aerated.
-Mixing will improve contact with nutrients for growth.
-Temperature and pH monitors are needed, and a cooling water jacket.
-Air inlets use spargers to force aeration
In the production of penicillin:
-Fermenter is sterile and cultured with Penicillium; which then grows.
-Takes 30 hours for production of penicillin to occur as it is a secondary metabolite.
-The penicillin is extracted after 6 days.
-This is batch culture as all the nutrients are added at the begining with no input.

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