Bacterial Pathogens and Diseases II (Endotoxins)

Question 1

what are endotoxins a part of?

Answer 1

the bacterial cell wall

Question 2

what do all bacteria have?

Answer 2

an inner membrane

Question 3

what do gram positive bacteria have?

Answer 3

an outer peptidoglycan layer

Question 4

what do gram negative bacteria have?

Answer 4

extra membrane on the outside – they have an inner and outer membrane, and in between this (the periplasmic space) you have the peptidoglycan layer

Question 5

what is a big component of gram negative bacteria?

Answer 5

lipopolysaccharide – the endotoxin

-causes symptoms

Question 6

describe the polysaccharide structure:

Answer 6

split into 3 parts:

LIPID A COMPONENT

• Main toxigenic component of the endotoxin
• Toxic lipid chain that anchors into the membrane
• Hydrophobic
• No. and type of fatty acid vary by species

CORE POLYSACCHARIDE

• Relatively constant between gram negative species
• unique sugars
• Hydrophilic

O – SIDE CHAIN

• repeats units of sugars tri, tetra or pentasaccharide sugars in different combinations
• highly variable between species AND between strains of the same species
• hydrophilic

Question 7

variability in lipids are responsible for what?

Answer 7

the differences in virulence that the lipid A component projects

Question 8

how do strains differ in terms of structure, and how will this affect pathogenicity?

Answer 8

bacteria can acquire genes that vary their genetic makeup, for example certain coli strains can make diff combinations of polysaccharides on the hydrophilic o-side chain so they differ antigenically, so you might have an antibody against 1 side chain that doesn’t recognise another

eg. difference in o side chain that means complement can no longer bind and assemble on the surface

Question 9

what are LPS molecules non covalently cross bridged by?

Answer 9

Ca and Magnesium ions

-provides a barrier to molecules including bile salts

Question 10

what exactly is the “endotoxin”?

Answer 10

the lipopolysaccharide (LPS)

• Found only in gram negative bacteria.
• Heat stable
• Not converted to toxoids.
• Major initiator of the sepsis pathway.

Question 11

can you make an immune response against all the parts of the LPS?

Answer 11

No,

the lipid A is the active component and not immunogenic, ie. you cannot make an immune response against it or a vaccine

but, the o antigen is highly immunogenic and immune specific.

Question 12

Sepsis – What is it?

Answer 12

Life threatening organ dysfunction caused by a dysregulated host response to infection.

Question 13

what is sepsis primarily driven by?

Answer 13

the innate immune system response:

• macrophages
• monocytes
• granulocytes
• natural killer cells
• dendritic cells

Question 14

in sepsis, the cells of the innate immune system detect what?

Answer 14

PAMPs and DAMPs

Question 15

what are DAMPs?

Answer 15

damage associated molecular patterns from damaged host cells

Eg if a bacteria attaches to surface of an epithelial cell and releases toxins that damage the endothelial cell – the components of the damaged cells will be recognised by dendritic cells or macrophages etc.

Question 16

what are PAMPs?

Answer 16

pathogen associated molecular patterns eg. endotoxin

Question 17

how are PAMPs and DAMPs detected?

Answer 17

cell membrane receptors (TLR’s, C-type lectin receptors)

cytosol receptors (NOD-like receptors, RIG-I-like receptors)

Question 18

what is the effect of PAMPs and DAMPs being detected by cell membrane and cytosol receptors?

Answer 18

Production of pro-inflammatory cytokines TNFα, IL-1, IL-6

via inflammasomes to produce IL-1β and IL-18 that cause rapid programmed cell death

Question 19

what are the effects of pro-inflammatory cytokines?

Answer 19

Increase number, lifespan and activation state of innate immune cells.

Increase adhesion molecule and chemokine expression by endothelial cells

Increase acute phase protein such as complement , fibrinogen and CRP

Cause fever.

Causes neutrophils to release extra-cellular traps (NETs) made of DNA and antimicrobial proteins that forms a scaffold for platelet activation

Cause release of microparticles by activated platelets

Increase tissue factor expression by blood monocytes

Question 20

consequences of sepsis?

Answer 20

increased vascular permeability

hypotension leading to hypovolaemic shock

DIC (disseminated intravascular coagulation)

multiple organ failure

Question 21

bacteria vs septicaemia?

Answer 21

Bacteraemia – you can find bacteria in the blood, but these are phagocytosed and killed quickly. But, fi those bacteria cannot be cleared, you get symptoms of immune response – septic shock and septicaemia.

Question 22

Sepsis – dysregulation

Answer 22

Production of ROS damages cellular proteins, DNA and lipids and impairs mitochondria.

Complement activation (esp. C5a) – increase ROS, granulocyte enzyme release, endothelial permeability and tissue factor expression

Widespread immunothrombosis leading to DIC with impaired microvasculature function and organ dysfunction.

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

Question 23

resolution of sepsis?

Answer 23

Autophagy of PAMP’s and DAMP’s – removal

Anti-inflammatory IL-10 produced early in process

Damaged cells – undergo apoptosis and engulfment by macrophages.

Question 24

Meningococcal Sepsis - what causes it?

Answer 24

Neisseria meningitidis, a gram negative diplococcus

- has a potent LPS that drives sepsis

Question 25

Neisseria meningitidis - how do the strains differ and what is the consequence?

Answer 25

the strains all have different antigenic natures – the LPS o-side chain all vary from each other. This means if you have antibodies against strain A, you will NOT have protective immunity against strain C because you aren’t recognising the same structure

Question 26

What makes meningococcus so effective in sepsis?

Answer 26

meningococcus LPS is different to the normal standard LPS. Lacks o-antigen on the surface, and has only got short o-antigen rather than a long one – called an oligosaccharide rather than a polysaccharide, but it has the same very potent immunostimulating effect.

Lipid blobs of the membrane hat it keeps releasing, full of LPS. Once inside the body it starts producing lots of these blebs, causing an overwhelming endotoxin response – not all gram negatives to do this, but nisserium does.

Question 27

clinical presentation of meningococcus septacaemia

Answer 27

Tumbler test (for someone with purpurik rash, gram negative septacaemia)

• 30% of people with septacaemia get pin prick rashes on their skin which get wider and larger and start to become purpurik and then they coallesce together
• large haemorrhages in the skin which wont move due to the thrombotic effect, and all the platelets having been used up
• oedema due to the inflammatory process.