Sporulation

Question 1

How is an endospore formed

Answer 1

1. DNA duplication
2. Unequal divisions of cytoplasm to create septum
3. Smaller compartment engulfed to make a double membrane
4. Peptidoglycan deposited between the two
5. Protein coat deposited
6. Cell is lysed releasing endospore

Question 2

What are the key processes during sporulation

Answer 2

1. Formation of cortex
2. Accumulation in spore specific components (Ca and Dipicolinic acid (DPA))
3. Formation of coat
4. Small acid soluble proteins

Question 3

Where is the cortex and it’s composition

Answer 3

• formed between inner and outer forespore membrane

Composed of:

• peptidoglycan with no teichoic acid
• thick cell wall like layer
• lightly cross linked so can expand and contract
• Cortex development correlates with dehydration

Question 4

Calcium content in endospore

Answer 4

Low levels in vegative cells
2% dry weight
accumulated due to facilitated diffusion

Question 5

DPA contents and ratios in spores

Answer 5

Made in mother cell

1:1 ratio with calcium

Leads to low free calcium in spore core

The ratio reduces water availability

Question 6

How is the coat formed on a spore

Answer 6

Synthesised by the mother

Contains stable disulphides rich proteins

Resistant the chemicals and UV as of absorbent pigments

Question 7

What are small acid soluble proteins (SASPs)

Answer 7

• provides resistance to UV and dry heat
• alpha/beta SASPS
• made in the development forespore stages
• contains 3-8% of spore proteins
• dsDNA binding proteins change DNA to B to A form making it more compact which protects pyramidine dimers causing resistance to heat and UV

Question 8

List structures of bacterial spore from inner to outer

Answer 8

Core
Inner membrane 
Germ cell wall 
Outer membrane 
Coat 
Exosporium

Question 9

bacillus anthracis

Answer 9

92% mortality
Often caused by agriculture with animals
Weaponised in 2001 in letter case

Question 10

Clostridium tetani

Answer 10

Causes tenitus
Deep wound causes anaerobic conditions
Vaccine developed

Question 11

Clostridium difficile

Answer 11

Antibiotic resistant associated diarrhoea

Leads to pseudomenbronous colitis

Resistant

Question 12

Clostridium botulinum

Answer 12

Low O2 levels

Treated with antitoxin

Question 13

When are spores released

Answer 13

In starvation / harsh conditions

Question 14

Sporulation initiation

Answer 14

Five histidine kinases Kin A-E acts as sensors and phosphorylates SpoOF which phosphorylates SpoOB

SpoOB phosphorylates SpoOA causing transcription of sporulation genes

Question 15

What is each Kinase used for in sporulation initiation

Answer 15

KinA/B add phosphates to SpoOF

KinC phosphorylates SpoOA

KinD works against SpoOA

Question 16

Sporulation phosphorelay

Answer 16

• KinA is major kinase
• over expression of KinA causes sporulation
• Contains PAS domain to sense stimuli
• no single PAS domain essential to inducing sporulation

Question 17

What’s SpoOA

Answer 17

Transcription regulator that interacts with target DNA on SpoOA box.

Regulates 121 genes 40 positively regulated and 82 negatively.

Key genes activated spoIIE, spoIIG

400 genes indirectly regulated

Question 18

Morphogenesis and gene regulation during spore formation

Answer 18

• activation of SpoOF/A and sigma H leads to asymmetric division

Sigma factor - gives promotor recognition to RNA polymerase core enzyme

Question 19

Spore germination stages

Answer 19

Stage 1.

• Cation released
• calcium / DPA release
• partial core hydration and loss of some resistance

Stage 2.

• cortex hydrolysis
• further core hydration
• loss of resistance and dormancy

Stage 3. Overgrown

• metabolism
• SASP degradation
• macromolecule synthesis and escape from coat.

Stage 4 germination

• activation
• can have resistant spore to sensitive mea