Bacterial Toxins

Question 1

What categories do we put toxin types into

Answer 1

Toxins can be broadly classified as either Endotoxins or Exotoxins

Question 2

What characteristics do endotoxins have

Answer 2

ENDOTOXIN
Innate part of cell wall (LPS) 
Low site specificity
Becomes available for action following cell death/lysis
Heat resistant usually

Question 3

What characteristics do exotoxins have

Answer 3

EXOTOXIN
Secondary metabolites
Typically exhibit high site specificity
High potency

Question 4

What are endotoxins an integral part of

Answer 4

The gram negative cell wall.

Question 5

What are endotoxins made of

Answer 5

Lipopolysaccharides - O-polysaccharide
Core polysaccharide
Lipid A - this stimulates the immune system

The saccharide protects against phagocystosis

Question 6

What are the three categories of exotoxins

Answer 6

1. Pore forming toxins (e.g.) pneumolysin
2. Toxins with enzymatic activity (e.g.) AB toxins
3. Superantigens (e.g.) toxic shock syndrome toxin

(however there is no clear systematic nomenclature)

Question 7

Give 4 examples of pore forming toxins

Answer 7

Streptococcus pneumoniae, group A streptococci, Escherichia coli, Staphylococcus aureus

Question 8

How do pore forming toxins work to damage eukaryotic cells

Answer 8

The toxin is a water soluble molecule which will bind to a receptor on the host cell membrane. This undergoes a conformational change which allows the toxin to create an aqueous pore in the membrane. This pore is usually formed of a beta barrel or alpha helix of various sizes. It results in the permeability of calcium, loss of potassium, halting of protein formation etc. The main effect, however, is cytolitic - the cell will die.

Question 9

Which bacteria causes pneumolysin and what are its characteristics

Answer 9

Streptococcus pneumoniae

Pneumolysin is a very common virulence factor utilised by pathogenic bacteria

Gram positive coccus
> 90 capsular types reported 
Non motile
Cytolytic
Cholesterol dependant toxin

Question 10

Which diseases can be caused by pneumolysin

Answer 10

Pneumolysin is one of numerous exotoxins produced by Strep. pneumo. when causing invasive disease:
Community acquired pneumonia (CAP)
Pneumococcal meningitis

Question 11

How does pneumolysin cause disease in the host

Answer 11

Oligomerises in host membrane to make a pore.

Significant involvement in pathogenesis of CAP

• This exotoxin is involved in the colonisation of the lung
• This exotoxin is involved in the histopathological changes in lung which are involved in the onset of symptoms

Question 12

What are the two phases pneumolysin is involved in in the development of cap by a patient

Answer 12

1. Immunosuppressive (early)
   Inhibition of mucociliary pathway (innate immune defence)
   Alveola macrophage apoptosis
2. Proinflammatory (late - higher conc. of toxin)
   Characterised by influx of neutrophils - higher conc of pneumolysin leads to neutrophil recruitment (as it activates complement and tissue damage leading to immune cell influx)
   Complement activation, Reactive Oxygen Species production
   All induces inflammatory mediated tissue damage

Question 13

What are the signs and symptoms of CAP

Answer 13

Fever, malaise

Dyspnoea (shortness of breath/breathlessness)

Productive cough (purulent discharge- contains white blood cells)

Focal chest signs (crackles, wheeze?)

May progress to acute respiratory failure or death (if untreated). Mortality in up to 18% individuals

Question 14

Which bacteria produces the dyptheria toxin and what are its characteristics

Answer 14

Producing agent:
Corynebacterium diphtheriae

Gram-positive
Non-motile
Clubbed morphology

Question 15

What are the effects of the diptheria toxin

Answer 15

Pseudomembrane formation (throat)
Difficulty swallowing
Systemic effects – heart complications, coma, death

Question 16

What is the diptheria toxin mode of action

Answer 16

1. Inhalation of aerosols (viable pathogen)
2. Colonisation of throat making toxins upon binding to the host membrane
3. Produces single chained AB toxin (DT)
4. AB toxin targets and inhibits Elongation Factor 2 (EF2) in eukaryotic cells. EF2 is invloved in moving trna in ribosomes during protein translocation. The inactivation leads to protein synthesis inhibition

Net effect is the pseudomembrane formation in throat

Death in 3 to 5% children

Question 17

Which vaccine is diptheria protection included in and how do we engineer vaccines to act against bacterial toxins

Answer 17

Diphtheria is currently covered in the UK vaccination schedule as part of the 6 in 1 vaccine (toxoid component)

The diptheria toxin is inactivated (a toxoid) and introduced into the host

Question 18

Which bacteria causes the botulinum toxin and what are its characteristics

Answer 18

Producing agent
Clostridium botulinum

Gram positive, motile rod
Obligate anaerobe
Sub-terminal spores

Question 19

How does the botulinum toxin cause botulism

Answer 19

Toxin causes botulism
Botulism is a toxin mediated disease that causes flaccid paralysis through toxin mediated effect on peripheral nervous system. Can cause respiratory failure. The muscles are no longer acting as they should

Rare (UK) due to public health interventions but fatal in 5-10% cases

Question 20

What is the mode of action of the botulism toxin (or any AB toxin)

Answer 20

FROM WIKI : Botulinum toxin exerts its effect by cleaving key proteins required for nerve activation. First, the toxin binds specifically to nerves that use the neurotransmitter acetylcholine. Once bound to the nerve terminal, the neuron takes up the toxin into a vesicle by receptor-mediated endocytosis. As the vesicle moves farther into the cell, it acidifies, activating a portion of the toxin that triggers it to push across the vesicle membrane and into the cell cytoplasm. Once inside the cytoplasm, the toxin cleaves SNARE proteins (proteins that mediate vesicle fusion, with their target membrane bound compartments) meaning that the acetylcholine vesicles cannot bind to the intracellular cell membrane, preventing the cell from releasing vesicles of neurotransmitter (acetylcholine). This stops nerve signaling, leading to paralysis.

Question 21

Detail the 3 categories of the pathenogenosis of botulism

Answer 21

Food bourne - toxin ingested in food - toxin enters bloodstream - flaccid paralysis

Infant botulism - spores ingested - colonisation of the GI tract - toxin enters bloodstream - flaccid paralysis. Why you cant give babies honey

Wound botulism - contamination of wound with spores toxin enters bloodstream - flaccid paralysis

Question 22

What are the signs and symptoms of botulism poisoning

Answer 22

Blurred vision, difficulty speaking due to muscle paralysis
Diarrhoea and vomiting
progression to paralysis
Recovery period takes months for the nerves to recover

Question 23

As a summary, give a sentance on how diptheria toxin and botulinum toxin affect host cells

Answer 23

DT targets protein translocation causing the production of pseudomembranes

BT targets the PNS and causes loss of excitory pathways leading to flaccid paralysis

Question 24

Give two examples of superantigens

Answer 24

Examples:

Toxic shock syndrome toxin (TSST)

Streptococcal pyrogenic exotoxins

Question 25

How does t-cell activation work

Answer 25

T cell activation typically involves a complicated cascade of events… First an antigen presenting cell comes into contact with an antigen The cell reconises and phagocytoses the micro-organism Microorganism is kept within a phagosome which is then fused with a lysosome to degrade the antigen into peptides. The t cell then creates MHC-II molecules within the endoplasmic reticulum Eventually the cell ends up with MHC-II molecules with a peptide epitope bound onto it. These compounds can be presented on the cells surface to activate a very specific T cells

Question 26

How do superantigens work to alter the t-cell activation process

Answer 26

The superantigens bind to MHC-II molecules (bypassing the antigen presenting cell process as they are able to bind the antigen presenting cell to t-cells that arent specific matches) This will activate significant numbers of T-cells Activation of the t-cells results in cytokine release (IL-2) - a cytokine storm 1 in 5 T-cells activated (as opposed to one in ten thousand under usual circumstances)

Question 27

What are the characteristics of toxic shock syndroms (signs and symptoms) and which toxin can cause it

Answer 27

Toxic Shock Syndrome Toxin causes toxic shock syndrome in humans Exotoxin production by Staphylococcus aureus Involves multiple organ systems BP drop, Fever, hypotension (which can lead to multiple organ failure) Rash Desquamation (peeling) of the skin on soles/palms

Question 28

Can bacteria utilise more than one toxin

Answer 28

Yes, in many cases, bacteria may utilise more than one type of toxin to achieve an infection (example - whooping cough)

Question 29

Which bacteria causes pertussis and what are its charactristics (whooping cough)

Answer 29

Causative agent Bordetella pertussis Gram-negative Aerobic cocco-bacillus Confined to humans

Question 30

What are the symptoms and progression of whooping cough

Answer 30

Paroxysms of cough (whoop - uncontrollable fits) Secondary pneumonia Highest mortality in < 4m yo

Question 31

How do the 3 toxins which cause whooping cough cause disease

Answer 31

``` 1. Pertussis toxin (toxin with enzymatic activity) Immune inhibition (reduced neutrophil recruitment makes them more receptive to increased bacterial load) ``` 2. Tracheal cytotoxin (endotoxin) Causes mucociliary pathway inhibition (prevents the scilia from beating the viable bacteria away from the lungs in the sputum) 3. Adenylate cyclase toxin (Pore forming and enzyme activity) Causes immune inhibition (has an affect on T cells and phagocytosis)

Question 32

Why can bacteria using toxins as a virulence factor be a benefitial to us

Answer 32

We can exploit their use in vaccines by inactivating the toxin, make it into a toxoid which allows us to develop an immune response Example - 6 in 1 vaccine: diptheria, tetanus, pertussis, hep B, polio, haemophilus influenza B

Question 33

How do we manage and prevent toxin mediated disease (as a general overview)

Answer 33

Vaccinate where possible (e.g. Diphtheria) Relevant antibiotic management to reduce the bacterial load hence bacterial toxin load Anti-toxins may be utilised: Post exposure prophylaxis. Used in tetanus Neutralise effects of toxin present in blood Can only be used before the toxin enters the cns