Bacterial Pathogens and Diseases II (Endotoxins)

Question 1

Describe the endotoxin in the gram negative bacteria.

Answer 1

The endotoxin (the lipopolysaccharide) is an integral part of the gram negative bacterial cell wall.
It sits in the outer membrane of the bacteria; it is not secreted or released.

Question 2

What are the three main areas of a lipopolysaccharide?

Answer 2

LIPID A.:

• Phosphorylated glucosamines attached to long chain fatty acids.
• The number and type of fatty acid vary by species.
• Hydrophobic

POLYSACCHARIDE CORE:

• Ketodeoxyoctanoic acid (KDO) and heptose. - Relatively constant between species.
• Hydrophilic

O – SIDE CHAIN:

• Repeat units of tri, tetra or pentasaccharide sugars. - Highly variable between species
• Hydrophilic

Question 3

List some characterstics of endotoxins.

Answer 3

• an endotoxin is lipopolysaccharide (LPS)
• Lipid A is the active component. – not immunogenic.
• O antigen is highly immunogenic and immune specific.
• it’s found only in gram negative bacteria.
• it’s heat stable
• it’s not converted to toxoids.
• it’s a major initiator of the sepsis pathway

Question 4

What is sepsis?

Answer 4

It is life threatening organ dysfunction caused by a dysregulated host response to infection.

Question 5

Describe the immune response that sepsis is primarily driven with.

Answer 5

Sepsis primarily driven by the innate immune system response. This includes macrophages, monocytes, granulocytes, natural killer cells and dendritic cells.

These cells detect:

• pathogen associated molecular patterns (PAMP’s) such as endotoxin
• damage associated molecular patterns (DAMP’s) from damaged host cells

This detection is mediated by:

• cell membrane receptors – toll-like receptors (TLR) and C-type lectin receptors
• cytosol receptors - NOD-like receptors, RIG-I-like receptors.

These can have an effect via:

• production of pro- inflammatory cytokines TNFα, IL-1, IL-6
• using inflammasomes to produce IL-1β and IL-18 that cause rapid programmed cell death

Question 6

What is the mechanism of sepsis?

Answer 6

The endotoxin form the gram negative bacteria goes and binds to CD14 receptors on macrophages.

The macrophages then pass the endotoxin to TLRs, which then dimerise. This dimerisation activates a cascade event that ends in activation of NF-κB and the production of TNF-α and other cytokines.

Question 7

What are some effects of pro-inflammatory cytokines?

Answer 7

1. Increase number, lifespan and activation state of innate immune cells.
2. Increase adhesion molecule and chemokine expression by endothelial cells.
3. Increase acute phase protein such as complement , fibrinogen and CRP.
4. Cause fever.
5. Causes neutrophils to release extra-cellular traps (NETs) made of DNA and antimicrobial
   proteins that forms a scaffold for platelet activation.
6. Cause release of microparticles by activated platelets
7. Increase tissue factor expression by blood monocytes

5,6 and 7 cause the formation of a thrombus (immunothrombosis); microbes get trapped within this, which attracts and activates further leucocytes.

Question 8

How can sepsis get dysregulated?

Answer 8

The process described achieves rapid control of localised and minor infections.

However, the process may pass a threshold, causing systemic injury.

It’s basically when the process has gone out of control.

Question 9

Describe the dysregulation of sepsis.

Answer 9

There is production of reactive oxygen species (ROS) – such as hydroxyl and nitric oxide – which damages cellular proteins, DNA and lipids and impairs mitochondria.

The complement activation (esp. C5a) – increases ROS, granulocyte enzyme release, endothelial permeability and tissue factor expression.

There is widespread immunothrombosis leading to disseminated intravascular coagulation (DIC) with impaired microvasculature function and organ dysfunction.

The mitochondrial damage leads to decreased intracellular ATP and cells enter state of hibernation – exacerbates organ dysfunction.

Question 10

Describe the process of resolution in sepsis.

Answer 10

It’s an active process – not passive.

Anti-inflammatory IL- 10 is produced early in process. it has helpful effects:

• supresses production of IL-6 and γ-interferon
• stimulates production of soluble TNF receptor and IL-1 receptor antagonist

There is the autophagy of PAMP’s and DAMP’s, causing their removal.

Damaged cells undergo apoptosis and engulfment by macrophages.

Question 11

Describe Meningococcal Sepsis.

Answer 11

It is caused by Neisseria meningitidis. It’s a gram negative diplococcus.

There are multiple serotypes like A,B,C, Y, W135.

Serotype A is associated with large outbreaks in Sahel region of Africa – thus called the Meningitis belt.
Serotype B,C and W135 are found in UK – but have declined since the introduction of MenC and now MenB vaccine.

It can cause disease ranging from meningitis to life threatening meningococcal sepsis.

Question 12

What makes meningococcus so effective in sepsis?

Answer 12

It has six fatty acid chains, which makes it highly effective as a toxin.

It’s carbohydrates are very similar to the ones expressed on human cells, and so the immune response to it is not very effective.

It sheds a lot of polysaccharides constantly in blebs, so produces huge amounts of toxin.