Bacterial evolution

Question 1

Outline Lamarckian evolution

Answer 1

First person to suggest: “life isn’t fixed”
Idea: if organisms uses something more frequently then increases, if not used then it will shrink.

e.g. giraffe necks will grow if used to reach higher fruit etc.

Question 2

Outline Darwinian evolution

Answer 2

Idea: Change is spontaneous ->
Natural selection = survival of the fittest.

e.g. some giraffes with long neck can reach leaves + survive = traits are conserved.

Question 3

What were the 2 opposing exceptions to Darwins idea?

Answer 3

1. Lysenkoism -> soviet propaganda
2. Prokaryotic evolution -> thought to have a different type of evolution.

Question 4

Outline the observation of the Luria-Delbrück experiment (1943)

Answer 4

Trying to explain if add toxic agent e.g. antibiotic or phage to culture the whole culture will become resistant.

Question 5

What was the interpretation associated with the Luria-Delbrück experiment?

Answer 5

The toxic agent added = causes all cells to become resistant

Question 6

What was the conclusion from the Luria-Delbrück experiment was testing?

Answer 6

Bacteria follow Lamarckian evolution

i.e. as there is a change in environment there is a change in bacteria.

Question 7

What were the 2 hypotheses tested in the Luria-Delbrück experiment?

Answer 7

1. Follows Darwinian:
   - Random mutations appear randomly
   - Mutations appear before adding agent
   - Adding of agent = provides conditions for natural selection between wild and mutants
2. Follows Lamarckian:
   - Directed change i.e. mutations only occur after addition of the toxic agent.

Question 8

What were the predictions for the Luria-Delbrück experiment if following the Lamarckian model?

Answer 8

• The culture will divide + grow exponentially.
• Only after toxic T1 added should mutations appear.
• Mutation rate should be constant in every culture -> no variability in no of bacteria w mutation.

Question 9

What does T1 do to the bacteria?

Answer 9

Kills them off

Question 10

What were the predictions for the Luria-Delbrück experiment if following the Darwinian model?

Answer 10

• Mutations happen at anytime are random in each culture
• Daughter cells will get mutations throughout randomly in each culture and will pass on to subsequent offspring.
• Once toxic T1 added -> some of the population will be resistant if resistance present previously.
• Huge variability.

Question 11

Steps for experiment

Answer 11

• Grow multiple cultures of EC in parallel and plate small amount of each onto separate plates.
• Add toxic agent
• Count no of bacteria resistant to T1 in each culture

Question 12

How to work out if Darwinian or Lamarckian

Answer 12

Out of the bacteria counted:
- Huge variability in no of resistants = Darwinian
- Small variability in no of resistants = Lamarckian

Question 13

What was trying to be concluded from the Luria-Delbrück experiment?

Answer 13

Is there a difference in the variability of the number of resistant bacteria in each culture compared with that of the control/baseline calculated number?

Question 14

What is the phenotype for the T1 phage?

Answer 14

Ton ^S = sensitive to T1
Ton ^R = resistant to T1

Question 15

What was the control present in the Luria-Delbrück experiment ?

Answer 15

One culture of Ton S plated onto separate plates. -> no to very little variation as all from one EC culture

Question 16

What was concluded from the Luria-Delbrück experiment?

Answer 16

• Big mutation in no of mutations vs control.
  = mutations had to occur before adding T1
  ∴ Bacteria evolve due to random mutation (only important upon addition of natural selection pressure). DARWINIAN EVOLUTION

Question 17

How can the old observation “once toxic added, whole population is resistant” be explained?

Answer 17

• Mutant is fitter than others around it -> so not acc resistant whole culture, just non-mutants cannot survive and only mutants continue to grow.

Question 18

What was the point of the Newcombe experiment (1949) ?

Answer 18

• To support the conclusion -> bacterial evolution is darwinian.

Question 19

7 Steps of Newcombe experiment

Answer 19

1. Culture of phage sensitive bacteria
2. Plate onto 2 plates of media
3. Incubate for few hours
4. Leave one plate (B) to grow normally
5. Re-spread other plate (A) + incubate A.
6. Spray both w phage.
7. Count visible colonies

Question 20

What are the results from the Newcombe experiment?

Answer 20

• There will be more colonies on the re-spread plate A.

Question 21

What was the Lederberg and Lederberg experiment (1952) aim?

Answer 21

Use replica plating to further back up idea of Darwinian evolution.

Question 21

What is the interpretation of Newcombe experiment results?

Answer 21

• On each plate = some mutants.
• If re-spreading one plate = mutants spread around.
• Not re-spread plate = mutants in one area.

i.e. Mutants survive -> more colonies if spread out via re-spreading after spraying of toxic agent.

Question 22

What is replica plating?

Answer 22

1. Take a master plate
2. Dip a piece of cloth onto master plate.
3. Then dip this onto a new plate.
4. Two plates will be identical (replicas) w same colonies in the same places on both.

Some bacteria onto cloth, some stay on plate.

Question 23

What were the results and conclusion of Lederberg + Lederberg experiment?

Answer 23

Results:
- Each replica plate = colonies in same position as master plate.

Conclusion:
- Mutation happened before plating as all colonies in the same place.

Question 23

Outline the steps of the Lederberg + Lederberg experiment

Answer 23

1. Master plate with EC sensitive to T1 phage.
2. Multiple replica plates
3. Each replica sprayed with phage.

Question 24

How to apply this to antibiotic resistance in bacteria?

Answer 24

• Antibiotic selects pre-existing mutations
• With higher conc of antibiotic = selection pressure for mutated bacteria present.