Lecture 6

Question 1

Types of virulence factors

Answer 1

Offensive e.g. toxins, Tissue degrading enzymes, invasins, adhesins

Non-specific - extracellular enzymes, siderophores

Defensive - Capsules, IgA proteases, Superoxide dismutase

Question 2

What occurs after docking in biofilm development?

Answer 2

-Pathogenic bacteria divide and wait

• QS used to count themselves vs others
• Count others with generic autoinducer AI-2
• Quorum reached - virulence factors expressed/upregulated
• Bacterium becomes nasty

Question 3

How many pairs of LuxI/LuxR homologues are there in P. aeruginosa?

Answer 3

2

Question 4

First LuxI/LuxR gene in P. aeurginosa

Answer 4

LasI - LasR

Autoinducer - OdDHL (OC12)

Phenotype: Elastase, Alkaline protease, Exotoxin A

Question 5

Second LuxI/LuxR homologue

Answer 5

RhII - RhIR

Autoinducer: BHL (C4)

Example phenotypes: Cytotoxic lectin, procyanin, hemolysin ramnolipid

Question 6

Explain pathogen colonisation

Answer 6

• Pathogen attaches to antigen either non-specifically (matrix) or specifically (receptor)
• Secondary attachment to host adhesion proteins - immediate if no pre-existing biofilm, extracellular enzymes required to dissolve matrix otherwise
• Adhesins and extracellular enzymes virulence factors
• After docking, pathogen causes infection
• Cell division leads to a quorum - upregulates QS controlled virulence factors
• Disease caused

Question 7

What are invasins

Answer 7

QS controlled extracellular proteins which allow bacteria to penetrate cells

Host cells break down in immediate vinicity of bacterial growth

Question 8

Examples of invasins

Answer 8

Hyaluronidase - Produced by streptococci, staphylococci, clostridia

Neuraminidase - Produced by Vibrio cholerae, Shigella dysenteriae

Streptokinase and Staphylokinase - convert inactive plasminogen to plasmin to digest fibrin and prevent blood clotting - More rapid diffusion of bacteria

Collagenase - Produced by Clostridium perfringens and Clostridium histolyticum - Breaks down collagen

Coagulase - Produced by S. aureus - Makes more collagen to localize pathogen

Question 9

Bacterial toxins types

Answer 9

Exotoxins and Endotoxins

Question 10

What do Endotoxins do?

Answer 10

Formed from LPS and bound to cell

Release when bacteria lyse

Causes host cells to release endogenous pyrogens

Affects hypothalamus

Action is indirect - activates host systems which cause damage - Fever, shock, inflammation, rarely fatal

Question 11

What do Exotoxins do?

Answer 11

Secrete proteins

Act on specific targets

Question 12

Are endotoxins QS-controlled?

Answer 12

No

Question 13

Structure of endotoxins

Answer 13

3 covalently linked subunits (O-specific polysaccharide, Lipid A, core polysaccharide)

Polysaccharide fraction (O-antigen and Core) makes it water soluble and immunogenic

Lipid A toxic

LD50 is 200-400 micrograms/animal

Weaker than exotoxins

LD50 botulinum toxin is 25pg/animal

Question 14

How can endotoxins cause problems in the pharmaceutical industry?

Answer 14

• Drugs contaminated with endotoxins cause complications
• Acceptable limit 25pg/mL or 0.25 endotoxin units

Question 15

How is endotoxin removed from drugs

Answer 15

In vitro LAL assay

detects level of coagulation by mixing products with cells

250 degrees Celsius for 30 mins

Filtration

Question 16

Cytolytic toxins

Answer 16

Exotoxin

Membrane disruption by insertion to cytoplasmic membrane and pole formation, or phospholipid attack by enzymes

Question 17

How are cytolytic toxins most easily detected?

Answer 17

Breakdown of red blood cells (Haemolysins)

Also attack Leukocidins

Leukocins: a toxin (Staph), Staph and Strep (especially S. pneumoniae)

Question 18

Phospholipases/Lecithinase

Answer 18

a toxin - Clostridium perfringens

• S. aureus, P. aeruginosa, Listeria monocytogenes

Question 19

What are A-B toxins?

Answer 19

2 components

B-subunit binds glycan

Conformational change - pore forms in membrane

• A-subunit enters cytosol - B-subunit leaves receptor
• A-subunit interferes with normal cell functioning
• Protein synthesis inhibited

Question 20

Corynebacterium diptheriae

Answer 20

• Gram-positive Actinobacterium
• Upper respiratory tract infection - sore throat, fever
• Transmitted from person-to-person 5-10% cases lead to death
• Cause Diptheria by producing Diptheria A-B exotoxin encoded in temperate phage DNA

Beta-phage encodes tox gene

DtxR controls tox gene expression

Lysogenic bacterial cells produce toxin

Question 21

Clostridium botulinium

Answer 21

• Produces Botulinum neurotoxin (types A to G) - MOST lethal toxin

Sodium cyanide:Botulinium toxin type A is 1:150,000

0.8 micrograms will kill human

Question 22

Botulinum neurotoxin

Answer 22

• Binds glycan on pre-synaptic membrane on termini of motor neurons
• Causes paralysis then death

Question 23

What bacteria produce A-5B exotoxins?

Answer 23

E. coli - Gastroenteritis

V. cholerae - Cholera