3- Bacterial Pathogens II

Question 1

what is an endotoxin?

Answer 1

a structural toxic component of gram-negative bacterial cell walls - often lipopolysaccharides

Question 2

what is the structure of lipopolysaccharides (LPS)?

Answer 2

three components: lipid A, core polysaccharide, O-side chain polysaccharide

Question 3

describe the lipid A component of LPS

Answer 3

• lipid component, composed of phosphorylated glucosamines and variable amounts of long fatty acid chains between species
• hydrophobic
• anchors LPS to outer membrane of gram-negative bacterial cell wall
• responsible for LPS endotoxic activity

Question 4

describe the core polysaccharide

Answer 4

• conserved amongst different species of gram-neg bacteria
• contains KDO (ketodeoxyoctanoic acid) and heptose sugar
• hydrophilic

Question 5

describe the O-side chain polysaccharide component of LPS

Answer 5

• highly variable repeat units of tri/tetra/penta-saccharide sugars between different bacterial strains and species
• hydrophilic
• responsible for antigenic diversity amongst gram-neg bacteria

Question 6

what are the variable components of LPS?

Answer 6

lipid A part, O-side chain polysaccharide

variability in the lipid A component with number and length of fatty acid chains

most variability in the O-side chain polysaccharide

Question 7

describe the endotoxin characteristics of the LPS components

Answer 7

lipid A has a high, active inflammatory role but isn’t immunogenic = hard for the immune system to produce antibodies against it

O-antigen is highly variable and immunogenic = easier to produce antibodies against the different bacterial species and strains based on their varying O-antigens

Question 8

describe the endotoxin characteristics of LPS as a whole

Answer 8

heat stable

not converted to toxoids = harder to produce antibodies against the molecule or make it non-toxic

diseases associated with LPS are severe - e.g. sepsis = leads to systemic inflammation and organ dysfunction

LPS molecules are made stable by non-covalent cross-bridges with Ca2+ and Mg2+ ions = forms a barrier that restricts the passage of hydrophobic molecules

Question 9

what is sepsis?

Answer 9

life-threatening organ dysfunction caused by a dysregulated host response to infection

caused by an overreactive and uncontrolled immune response triggered by infection - trigger being gram-negative endotoxins like LPS

Question 10

describe the mechanism of sepsis and how it’s detrimental

Answer 10

LPS endotoxin detected by resident immune cells, triggers their hyperactivation

resident immune cells detect DAMPs (e.g. LPS) and PAMPs (from damaged host cells) through cell membrane receptors like TLRs and C-type lectin receptors & intracellular cytosol receptors like NOD-like & RIG-1 like receptors

high levels of pro-inflammatory cytokines (e.g. TNF-a, IL-1 & 6) produced, and inflammasomes are activated through intrac. mechanisms to produce IL-1b & 18 – promotes apoptosis & extensive inflammation from cytokine release

extensive inflammation causes tissue damage, leads to more leukocyte recruitment and more cytokines produced, creates a cytokine storm

tissue damage breeches barriers, allows bacteria & endotoxins to enter the blood = colonises many tissues, causes a systemic infection

systemic infection leads to vasoconstriction = loss of blood flow = organ damage

Question 11

list resident immune cells

Answer 11

macrophages
monocytes
granulocytes
natural killer cells
dendritic cells

Question 12

what are DAMPs?

Answer 12

damage associate molecular patterns - endogenous molecules released/expressed by stressed or dying cells

e.g. ATP, heat shock proteins

Question 13

what are PAMPs?

Answer 13

pathogen associated molecular patterns - associated with microbial pathogens, recognised by pattern recognition receptors like toll-like receptors on immune cells

e.g. LPS

Question 14

compare gram-positive and gram-negative bacterial infections

Answer 14

gram-positive:
- produce superantigens = activate T cells by bridging MHC class II molecules with T-cell receptors, leading to polyclonal T cell activation and cytokine release
- associated with toxic shock syndrome = can lead to organ dysfunction and systemic inflammation

gram-negative:
- produce endotoxins - e.g. LPS = components on the outer membrane, which activate resident immune cells leading to release of pro-inflammatory cytokines
- associated with sepsis, infection evolves to colonise other tissues, cause systemic inflammation and organ dysfunction

Question 15

describe the TLR4-LPS signal transduction pathway that initiates an immune response and helps manage a gram-negative bacterial infectionn

Answer 15

gram negative bacteria produce LPS specific to it

MD2 detects and binds to lipid A. TLR4 on resident immune cell/ macrophage detects and binds to O-side chain polysaccharide

TLR4 complex dimerises with another TLR4 receptor – not necessarily LPS complexed

CD14 interacts with dimerised complex = initiates a downstream signalling pathway mediated by adaptor molecule Myd88 which triggers TRAF6 activity

leads to activation and transcription of transcription factor NF-kB

NF-kB translocates from cytoplasm to nucleus, sits on specific gene promoters to increase transcription of pro-inflammatory cytokines

pro-inflammatory cytokines produced by macrophages = initiates an immune response, mediate inflammation, help manage gram-neg bacterial infection

Question 16

how does dysregulation contribute to sepsis?

Answer 16

when adaptive immune responses fail to control infections and inflammation = leads to systemic injury and potential sepsis

Question 17

list components of dysregulation in sepsis?

Answer 17

uncontrolled ROS production

dysregulated complement activation via C5a

widespread immunothrombosis leading to disseminated intravascular coagulation

mitochondrial damage

Question 18

what is the role of ROS in sepsis?

Answer 18

generated by immune cells to destroy pathogens within phagolysosomes

dysregulated production can cause bystander tissue damage and impair mitochondrial function

Question 19

how does dysregulated complement activation contribute to sepsis?

Answer 19

dysregulated complement activation, especially via C5a leads to:
- increased ROS production
- release of granulocyte enzymes
- increased endothelial permeability
- increased tissue factor expression

Question 20

what is the consequence of widespread immunothrombosis in sepsis?

Answer 20

leads to disseminated intravascular coagulation

impaired microvascular function
organ dysfunction
reduced blood flow to organs
organ failure

Question 21

What role does mitochondrial damage play in sepsis?

Answer 21

systemic mitochondrial damage occurs in later stages of sepsis - leads to:
- worse organ dysfunction and cell death
- decreased intracellular ATP levels = cells go into hibernation

Question 22

what is tissue factor?

Answer 22

protein important in the initiation of the coagulation cascade, expressed by endothelial cells and monocytes

initiates extrinsic pathway of coagulation by interacting with factor VII, leads to thrombin generation and clot formation

Question 23

under what natural conditions can sepsis resolve?

Answer 23

it can resolve if the infection isn’t severe enough to cause irreversible damage

body undergoes an active process of resolution = counteracts dysregulated inflammation

Question 24

list three ways for how can sepsis resolve under natural conditions

Answer 24

anti-inflammatory cytokines
autophagy of DAMPs and PAMPs
apoptosis of damaged cells, cleared by macrophages

Question 25

what role do anti-inflammatory cytokines play in resolving sepsis?

Answer 25

control the dysregulated immune response in sepsis and return it to a regulated state by:
- suppressing the production of pro-inflammatory cytokines
- stimulating the production of soluble TNF receptors and IL-1 receptor antagonists

Question 26

how does autophagy contribute to the resolution of sepsis?

Answer 26

removes PAMPs and DAMPs = triggers for sepsis

clearance of cellular debris and infectious agents helps resolve the inflammatory response

Question 27

what happens to damaged cells in the resolution of sepsis?

Answer 27

damaged cells undergo apoptosis and are engulfed and cleared by macrophages

this helps regulate inflammation and remove damaged tissue

Question 28

what is meningococcal sepsis caused by?

Answer 28

Neisseria meningitidis, also called meningococcus

gram negative diplococcus, has various serotyped associated with different diseases and areas - e.g. serotype A associated with Africa, serotype B with the UK

Question 29

what makes meningococcus effective in causing sepsis?

Answer 29

production of LOS/ lipo-oligosaccharide
- acts as an endotoxin and triggers a fast and aggressive immune response
- it’s similar to LPS but has a shorter core and lacks an O-side chain

release of blebs = small vesicles containing LOS and other bacterial components
- allows blebs to carry out effects as antibodies will target the bacterial cell surface, bacteria are able to evade antibodies and keep infecting

Question 30

what are the clinical manifestations of meningococcal sepsis?

Answer 30

areas of red rash
non-bouncing skin oedema
disseminated rash
severe fatigue and headaches