B: Microorganisms and Fermenters

Question 1

What are microorganisms?

Answer 1

organisms that are too small to see without magnification

Question 2

Give 3 examples of microorganisms.

Answer 2

bacteria, fungi, and some protoctista

Question 3

How can microorganisms also be referred to?

Answer 3

referred to as ‘microbes’

Question 4

Why are microorganisms widely used in industry?

Answer 4

• metabolically diverse, therefore possible to find a type to carry out many different reactions
• very small so large numbers can be grown
• fast growth rate

Question 5

What inhibits the growth of bacteria?

Answer 5

biocides

Question 6

Describe an experiment that demonstrates the inhibition of bacteria growth using biocides.

Answer 6

• sterilise Petri dishes
• put nutrient agar in Petri dish
• spread a pure culture of bacterium over surface of agar
• place paper discs soaked in biocide on agar surface
• incubate dishes at optimum temperature for bacterium
• examine after 36hours
• clear areas (where bacteria died) show zones of inhibition of bacterial growth

Question 7

How could you find out which antibiotics kill the bacteria causing a patient’s disease?

Answer 7

do the ‘experiment that demonstrates the inhibition of bacteria growth using biocides’
- just test lots of different biocides

Question 8

What determines differences in bacterial resistance to biocides?

Answer 8

• metabolism of bacterium

- or structure of the cell wall of bacterium

Question 9

What are the two main types of wall structure in bacterium?

Answer 9

Gram-positive and Gram-negative

Question 10

What do differences in metabolism or structure of the cell wall determine?

Answer 10

resistance (or not) to a particular biocide

Question 11

How can you deduce which type of wall structure a particular bacterium has?

Answer 11

do a Gram staining experiment

Question 12

Outline the procedure for Gram staining.

Answer 12

• smear small sample of a pure bacterial culture on a microscope slide with an inoculating loop
• pass through a flame to fix the bacteria to the slide
• stain with crystal violet for 30 seconds
• treat with Gram’s iodine for 30 seconds (to bind crystal violet to the outer surface of the bacteria)
• decolorize with alcohol for 20 seconds (to dissolve the outer membrane of Gram-negative bacteria and remove the crystal violet staining)
• counterstain with safranin (which is red) for 30 seconds, then rinse and blot dry

Under the microscope Gram-negative bacteria will be red or pink. Gram-positive bacteria will be violet

Question 13

In the process of Gram staining: why does one treat the bacteria with Gram’s iodine?

Answer 13

to bind crystal violet to the outer surface of the bacteria

Question 14

In the process of Gram staining: why does one decolorize the bacteria?

Answer 14

to dissolve the outer membrane of Gram-negative bacteria and remove the crystal violet staining

Question 15

Draw a diagram of the wall structure of Gram-positive Eubacteria. (p158)

Answer 15

1. thick layer of peptidoglycan

2. plasma membrane of phospholipids and proteins

Question 16

Draw a diagram of the wall structure of Gram-negative Eubacteria. (p158)

Answer 16

1. thin layer of peptidoglycan
2. plasma membrane of phospholipids and proteins
3. outer membrane of lipopolysaccharide and protein

Question 17

What is required in the large-scale production of useful substances by microbes?

Answer 17

fermenters

Question 18

What are fermenters usually made of? Why?

Answer 18

• stainless steel

- to make sterilization easy

Question 19

In the large-scale production of useful substances by microbes, what is the fermenter filled with?

Answer 19

• sterile nutrient medium

- which is inoculated with a chosen microbe

Question 20

What growing conditions are relevant in the large-scale production of useful substances by microbes? Describe the conditions in a fermenter. How are the conditions monitored?

Answer 20

• pH and temperature
• conditions are maintained at optimal levels for the growth of the microbe
• monitored using probes and levels are adjusted if they move too far from the optimum

Question 21

Why does a cooling jacket surround the fermenter?

Answer 21

to remove the heat energy generated as a waste product of metabolism and maintain optimum temperature conditions inside the fermenter

Question 22

How does the heat jacket surrounding the fermenter work?

Answer 22

• cool water flows through the heat jacket and takes away the heat energy from the fermenter

Question 23

Why is sedimentation of microbes prevented?

Answer 23

sedimentation (matter settling at the bottom of a liquid) would mean some microbes, the microbes on the bottom, would not get access to enough nutrients. This might kill them, which would be inefficient

Question 24

How is sedimentation of microbes prevented?

Answer 24

by an impeller (a rotating set of paddles)

Question 25

Why might sterile air be bubbled through the fermenter?

Answer 25

if the desired metabolic process is aerobic

Question 26

What does a pressure gauge do in a fermenter?

Answer 26

measure the gas build up and allows waste gases to escape

Question 27

Apart from too much/little temperature, pH and gas, what else can limit fermentation?*

*badly phrased, deal with it

Answer 27

other* waste products can build up and eventually limit the fermentation

*no specific ones stated

Question 28

How many main types of fermentation are there?

Answer 28

2

Question 29

What are the main types of fermentation?

Answer 29

• batch fermentation

- continuous culture

Question 30

When are nutrients added in batch fermentation? When is the product harvested?

Answer 30

• once at the beginning

- when the yield has reached a maximum the fermenter is drained (to harvest the product)

Question 31

When are nutrients added in continuous culture? When is the product harvested?

Answer 31

• nutrients added during the fermentation, so they do not run out
• product harvested during fermentation

Question 32

If you’re a bit sad: Draw and label a fermenter

Answer 32

p158

Question 33

What is the ‘useful product’ of microbes grown in fermenters called?

Answer 33

the metabolite of interest

Question 34

What does detailed knowledge and analysis of metabolic pathways allow scientists to do? What is this called?

Answer 34

• change conditions at multiple points to improve the yield of the metabolite of interest
• called pathway engineering

Question 35

What is pathway engineering?

Answer 35

change conditions in a fermenter at multiple points to improve the yield of the metabolite of interest

Question 36

Give some examples of changing conditions in pathway engineering.

Answer 36

• extra substrate may be added
• by-products that slow down the pathway may be removed
• range of products may be extended

Question 37

How is genetic engineering used in pathway engineering?

Answer 37

to introduce extra genes or change how the expression of existing microbial genes is regulated