Practical 2 - Nutritional & physical requirements of microbes

Question 1

What are the two major categories of media used for routine cultivation of bacteria?

Answer 1

Chemically defined media and Complex media

Question 2

What is chemically defined media?

Answer 2

Media that is composed of known quantities of chemically pure, specific organic/inorganic compounds.

Question 3

The use of chemically defined media requires a microbiologist to know an organisms

Answer 3

Specific nutritional needs

Question 4

What is complex media?

Answer 4

Media where the exact chemical composition is unknown.

Question 5

What is complex media made of?

Answer 5

Extracts of plant and animal tissues - variable in their chemical composition. Most contain amino acids, sugars, vitamins, minerals but quantities are unknown

Question 6

What are the three types of special-purpose media?

Answer 6

1. Selective media
2. Differential/selective media
3. Enriched media

Question 7

How does selective media work?

Answer 7

By selecting a specific group of bacteria. They incorporate chemical substances that inhibit the growth of one type of bacteria while permitting the growth of another.

Question 8

Give three examples of selective media

Answer 8

1. Phenylethyl alcohol agar
2. Crystal violet agar
3. 7.5% sodium chloride agar

Question 9

How does phenylethyl alcohol agar work as a selective medium?

Answer 9

Isolates most gram positive bacteria. The phenylethyl alcohol is partially inhibitory to gram negative organisms

Question 10

How does crystal violet agar work?

Answer 10

This is selective for most gram negative microbes

Question 11

How does 7.5% sodium chloride agar work?

Answer 11

Inhibitory to most organisms other than halophilic organisms. Useful in detection of members of the genus staphylococcus

Question 12

What does differential/selective media distinguish between?

Answer 12

These can distinguish among morphologically and biochemically related groups of organisms.

Question 13

Give three examples of differential/selective media

Answer 13

1. Mannitol salt agar MSA
2. MacConkey agar MAC
3. Eosin Methylene Blue agar EMB

Question 14

What does mannitol salt agar contain and how does it work at selectivity?

Answer 14

Contains a high salt concentration which in inhibitory to most bacteria other than staphylococci (SELECTIVITY)

Question 15

Mannitol salt agar contains mannitol as the carbon source, how does this differentiate between staphylococci strains?

Answer 15

Only certain staphylococci can metabolise mannitol

Question 16

When mannitol is metabolised what happens?

Answer 16

An acid is produced which turns the indicator in the medium from red to yellow

Question 17

What does MacConkey agar contain and how does it work at selectivity?

Answer 17

MacConkey agar contains crystal violet
Inhibits the growth of gram +ve organisms

Also contains bile salts and these inhibit many bacteria except enteric bacteria - those that live in the human gut

Question 18

What are enteric bacteria?

Answer 18

Ones that live in the human gut

Question 19

How does MacConkey agar work at differentiation?

Answer 19

Carbohydrate source is lactose when this is fermented acid is produced which results in yellow colour in medium.
Lactose fermenters will take up neutral red dye from medium to develop a pink colour
Non-lactose - colourless

Question 20

How does Eosin Methylene Blue agar work at selectivity?

Answer 20

Partially inhibitory to gram positive bacteria

Question 21

How does Eosin Methylene Blue agar work at differentiating?

Answer 21

Differentiates between enteric lactose fermenters and non-fermenters. E.Coli particularly distinctive - green sheen

Question 22

What is enriched media?

Answer 22

Media that have been supplemented with highly nutritious materials such as blood, serum or yeast extract.

Question 23

An example of enriched media is

Answer 23

Blood agar

Question 24

What does blood agar contain?

Answer 24

Blood and other nutrients

Question 25

What is a fastidious organism?

Answer 25

A fastidious organism is any organism that has a complex nutritional requirement. In other words, a fastidious organism will only grow when specific nutrients are included in its diet.

Question 26

What are the three haemolytic activities?

Answer 26

1. Gamma haemolysis
2. Alpha haemolysis
3. Beta haemolysis

Question 27

What happens in gamma haemolysis?

Answer 27

No lysis of red blood cells - no significant change in appearance surrounding colony

Question 28

What happens in alpha haemolysis?

Answer 28

Incomplete lysis of RBCs
Reduction of haemoglobin to methomeglobin
Results in greenish halo around the bacterial growth

Question 29

What happens in beta haemolysis?

Answer 29

Lysis of the RBCs

Clearing zone around the colony

Question 30

Organisms whose growth range falls between 5ºC and 20ºC with an optimum of 11ºC are called

Answer 30

Psychrophiles

Question 31

Bacterial species that will grown within 20ºC to 45ºC - many grow at 37ºC are called

Answer 31

Mesophiles

Question 32

Bacterial species that have an optimum growth temperature of 45ºC to 65ºC are called

Answer 32

Thermophiles

Question 33

Organisms with an optimum temperature at about 90ºC are called

Answer 33

Extreme thermophiles

Question 34

What is special about endospore forming organisms in terms of temperature?

Answer 34

Fairly narrow temperature limits in vegetative state

Spores can survive in a dormant stage over a very wide temperature range

Question 35

Having a pH optima between 6.5 and 7.5

Answer 35

Neutrophiles

Question 36

Bacteria able to grow below pH6 are

Answer 36

Acidophiles

Question 37

Bacteria able to grow above 8.5 are

Answer 37

Alkalophiles