Unit 6- Microbial Techniques Flashcards

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1
Q

What are the different types of mixtures the microbes are cultured on?

A
  • agar plate (made of agar and a nutrient broth)

- selective media

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2
Q

What is the agar plate?

A

A solid medium of agar mixed with a nutrient broth in a Petri dish

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3
Q

What is a a selective media?

A

A medium made of a specific recipe to allow the growth of only one group of microbes, it may include antiobiotics if the wanted bacteria is antibiotic resistant

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4
Q

How is equipment sterilised?

A

Using ultra violet
autoclave (large pressure cooker)
Using virkon, bleach or ethanol
Flaming

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5
Q

What conditions does a culture provide?

A
  • nutrients
  • suitable temp and pH
  • usually access to O2
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6
Q

Aseptic techniques are used to avoid contamination, give some examples of aseptic techniques:

A
  • all equipment should be sterile
  • flame the appropriate equipment in Bunsen burner before and after
  • replace lids as quickly as possible
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7
Q

What are the problems with culturing microbes?

A
  • pathogens may enter the culture and grow risking disease
  • safe organisms may mutate to dangerous strains
  • contamination can spoil an investigation
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8
Q

What precautions can be taken to reduce the risk of culturing microbes?

A
  • adequate O2 supply as pathogens prefer to grow in anaerobic conditions
  • culture the medium at about 20 degrees for a few days as body temperature encourages the growth of potentially dangerous human pathogens
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9
Q

What 4 different techniques can be used to measure microbial growth?

A
  • cell counting with haemocytometer
  • turbidity with a colorimeter
  • dilution plating until the microbes can be spread on an agar plate and counted
  • weighing dry mass of a known volume
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10
Q

How can we measure the number of microbes in a culture broth by dilution plating?

A
  • dilute the culture until the number of cultures can be counted when spread on an agar plate
    (Dilution plating is the same as serial dilution)
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11
Q

How can you count the number of bacteria in a culture using a haemocytometer?

A
  • Pipette the culture onto the haemocytometer
  • the volume of the haemocytometer is 0.1mm
  • count the number of cells in a 0.2mm box
  • calculate the number of cells per mm^3
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12
Q

What is the convention when counting using a haemocytometer

A

Count the cells overlapping the north west boundary and discard the cells over the south east boundary

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13
Q

How do you only count living cells using a haemocytometer?

A
  • add a vital stain (trypan blue) which only stains dead cells blue
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14
Q

What does turbidity mean?

A

Cloudiness

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15
Q

How can we measure cell density using a colorimeter?

A
  • dilute the culture broth until it has an absorbable of below 0.5
  • the greater the concentration of cells the more turbid the dilution giving a higher absorbance
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16
Q

What is an issue of using a colorimeter?

A
  • must be calibrated regularly

- reading must be taken quick before cells settle at bottom of the curettes

17
Q

How can we count cells using dilution plating?

A
  • take a small sample from a culture broth using aseptic techniques and a sterile pipette
  • spread the sample evenly over the agar plate using a sterile spreader
  • the agar plate is incubated at 30degrees for a day, each living cell will form a colony
  • count the number of individual colonies, and this is the number of colonies in the dilution
18
Q

What are the 4 stages of a bacterial growth curve?

A
  • lag phase
  • log phase
  • stationary phase
  • death phase
19
Q

What occurs in the lag phase?

A
  • cell division and cell death rates are low so there is little change
  • the microbes are preparing to use the nutrients and adjusting to the new conditions
  • switching on genes and mostly synthesising proteins
20
Q

What happens during the log phase?

A

There are no limiting factors of growth
Growth is exponential due to rapid doubling
Scale is log so graph appears linear

21
Q

What occurs during stationary phase?

A

Cell division rate slows down and cell death rate increased
Until the rates are equal
Due to a build up of metabolic waste, lack of space or nutrients

22
Q

What occurs during the death rate?

A

Population of living cells falls due to less nutrients, space available and more toxic metabolic waste