10. Dengue virus immunity and vaccination 1 Flashcards
1
Q
What is the global burden of dengue?
A
- It is the most abundant and rapidly spreading arboviral infection.
- Estimated 390 million infections with 96 million clinically apparent.
- 500,000 cases of dengue hemorrhagic fever and dengue shock syndrome. These are more severe forms of dengue.
- Causes 20,000 deaths annually.
- Mostly exists in tropical and sub-tropical areas but it is spreading.
2
Q
What type of virus is dengue virus?
A
- An orthoflavivirus
- ssRNA virus
3
Q
What are examples of orthoflaviviruses?
A
- Dengue virus
- Zika
- Yellow fever
- West Nile Virus
4
Q
How is dengue virus transmitted?
A
- By the Aedes mosquitoes.
- Aedes aegypti is the main vector
- Aedes Albopictus is the secondary vector
5
Q
How many dengue serotypes are there?
A
- 4 infectious serotypes
- They cocirculate.
- This provides a challenge for vaccine design.
6
Q
How is Dengue fever treated?
A
- There are no specific therapies.
- Only general pain killers and fluids can be given
7
Q
What vaccines are available for dengue virus?
A
- 2 partially protective vaccines
- Dengvaxia by Sanofi-Pasteur
- Qdengue by Takeda
- Dengvaxia has some serious problems.
8
Q
Are cases of dengue fever increasing or decreasing?
A
Increasing
9
Q
Where is dengue spreading to geographically?
A
- Places it shouldn’t and hasn’t been.
- Coastal regions of Europe like Italy and Spain.
- South America
10
Q
Why is dengue spreading out of tropical regions?
A
- The dengue vector Aedes Albopictus is now endemic in Europe due to rising temperatures.
- Travellers return from dengue endemic regions with the infection and can pass the infection to the vectors.
- This then causes local transmission and spread of the virus in Europe and other places.
- The winters are still too cold for the vector so infection drop but climate change increases this risk.
- South and Central America is mostly effected by this.
11
Q
What are the 4 clinical stages of Dengue Virus?
A
- Asymptomatic
- Self-limiting dengue fever
- Severe dengue: Dengue hemorrhagic fever and dengue shock syndrome
12
Q
What is the normal course of dengue infection?
A
- mosquito bite
- Around 5 days of incubation
- Then viremia starts to develop alongside fever.
- After 5 days is the biggest risk of developing severe dengue.
- around 10% of cases become severe dengue
13
Q
When does severe dengue manifest?
A
- Around day 5 of infection
- Symptoms start to appear
- This is later in infection, so the viral load is decreasing, but the immune response is increasing.
14
Q
What are the forms of severe dengue?
A
- Dengue hemorrhagic fever
- Dengue shock syndrome
- These occur in the minority of cases, but the effects can be debilitating and last a long time.
15
Q
What is non-severe dengue?
A
- A normal dengue infection.
- symptoms include vomiting, rashes and aches.
- Some symptoms act as warning signs for severe dengue.
- This includes abdominal pain, fluid accumulation or liver enlargement.
16
Q
What are the symptoms of severe dengue?
A
- Severe plasma leakage due to increased vascular permeability.
- This leads to hypervolemic shock, severe bleeding and organ impairment.
- This is caused by a massive immune mediated cytokine storm.
- It is not clear what part of immunity mediates severe dengue
17
Q
What are the host risk factors for severe dengue?
A
- Secondary dengue infection
- Age - very young or old
- Co morbidities like hypertension and diabetes.
- Obesity
- Genetic polymorphisms, SNPs.
18
Q
How does secondary dengue infection increase the risk for severe dengue?
A
- There are 4 different stereotypes of dengue circulating
- Infection with a different serotype at the same time or later. (heterologous infection)
- This risk of severe dengue increases for the 2nd exposure massively.
- After two exposures, the risk decreases again, suggesting immunity develops to protect from subsequent infections.
19
Q
How does SNPs increase the risk for severe dengue?
A
- Some SNPs in genes important with T cell responses and cytokines can increase the risk of severe dengue.
- MICB is the main one. It was found through a GWAS study.
- Also PLCE1, TNFa, IL-10, HLA class 1 and 2
20
Q
What is heterologous dengue infection?
A
When someone is infected with a different serotype of dengue then they were previously infected with.
21
Q
What are the key features of the adaptive immune response?
A
- Consists of T and B lymphocytes and their secreted products.
- It is a sophisticated antigen-specific defence system.
- Immunological memory
22
Q
How do T cell recognise antigens?
A
- MHC/HLA molecules.
- MHC1 presents cytosolic proteins via the TAP pathway to CD8+ T cells.
- MHC2 presents extracellular proteins and activate CD4+ T cells.
23
Q
How do T cells recognise viral antigens?
A
- Viruses are intracellular pathogens so their proteins are found in the cytosol.
- This means they are leaded via the TAP pathway into MHC1.
- These are then presented to CD8+ T cells to activate them.
24
Q
What are the 3 phases of a primary T cell response?
A
- Expansion
- Contraction
- Memory
25
What happens in the contraction phase of the T-cell response?
1. Around 5-10% of T cell from the main response survive and become memory cells.
2. IL-7Ra (CD127) is important for this persistence without antigen stimulation.
3. This allows the T cells to respond to omostatic cytokines, which allows persistence.
26
When does the T cell response peak?
1. After the peak of viral infection
2. After this peak most T cells die.
27
How many naive T cells are there for any given antigen?
1. Around 1 in 100,000 T cells.
2. This is very low frequency
28
How many memory T cells are there for any given antigen?
1. Memory T cell precursors are much more common than naive T cell precursors.
2. They also have a lower activation threshold
3. This is an advantage of immune memory.
4. The frequency of memory cells depends on the pathogen.
29
What are the main characteristics of T memory cells?
1. They clonally expand from naive precursors.
2. They have a lower activation threshold then naive cells.
3. They mount secondary responses and confer immediate protection. This response is bigger then a naive response.
4. Memory cells persist for a lifetime in the absence of antigen.
5. Heterogeneous for effector functions and migratory capacities. There are lots of different subsets.
30
What does the immune response to primary dengue infection look like?
1. Bite from mosquito and infection.
2. 5 day incubation that leads to viremia
3. Day 0 is when symptoms appear.
4. Peak of viremia is around day 2/3.
5. Peak of T cell response is around day 8/9. About 6 days later.
6. Antibodies also develop in patients.
7. IgM response starts around day 1.
8. IgG response appears around day 5/6.
31
What does the immune response to secondary dengue infection look like?
1. Bite from mosquito and infection.
2. Viremia peaks higher and earlier than in primary infection. Around Day 0
3. Viremia is shorter due to pre-existing antibodies that can clear the infection.
4. Risk point for developing severe dengue is around day 5.
5. The immune response is increasing as viremia decreases.
6. Vascular permeability increases and peaks when the risk is highest for severe dengue.
32
What is severe dengue associated with?
1. A higher dengue virus load early in infection.
2. This is independent of dengue serotype or host immune status.
33
What is the structure of the dengue virus proteome?
1. It is an ssRNA virus that is encoded by 1 polyprotein.
2. There are 3 structural proteins. The capsid, membrane and envelope proteins.
3. There are 7 non-structural proteins: NS1, NS2a, NS2b, NS3, NS4a, NS4b, NS5.
4. The non-structural proteins encode enzymes important for viral lifecycle and replication.
34
What is the structure of the dengue virus particle?
1. Mature dengue has a smooth surface made by repetitive E protein pattern.
2. Immature dengue has a spikey surface made by the prM precursor peptide.
3. This immature structure prevents premature membrane fusion.
4. The cleavage of prM depends on pH and is done by host or viral proteases.
35
How were dengue T cell epitopes determined?
1. Using ELISpot assays.
2. These are used to detect and measure immune cells that produce antibodies or cytokines.
3. It is good as it is high throughput.
4. The limitation is that you don’t know which type of T cell is causing the response.
36
What dengue epitopes were recognised by T cells?
1. All dengue proteins were recognised to some degree.
2. The T cell response to non-structural proteins was bigger especially to NS3 and NS5.
3. Structural proteins still also trigger an immune response.
37
Does dengue cause a CD4 or CD8 T cell response?
1. Using flow cytometry and intracellular cytokine staining you can characterise the T cell response.
2. The majority of the response is a CD8+ response to non structural proteins.
3. NS3 and NS5 produce the largest immune response.
4. There is still a CD4+ T cell response that mainly targets the Capsid and envelope proteins.
38
What regions in the dengue virus polyprotein trigger the immune response?
1. The T cell response can be mapped to the dengue proteome and map how frequently a peptide region is recognised.
2. 50% of the T cell responses to dengue target 25 antigenic regions across NS3 and NS5.
3. This shows the T cell response is broad but also targeted.
4. This is important to consider for vaccine design.
39
How does dengue activate the immune response?
1. The virus enters through the skin.
2. The dengue antigens are picked up by Langerhans cells which present antigens to T cells.
3. The T cells recognise the antigen presented by the Langerhans cell which also imprint skin-homing signals so the T cells go back to the skin.
4. This is an evolutionary response to increase the chance of antigen re-encounter.
5. This homing signal is CLA and all dengue specific T cells express CLA.
6. Later in infection they start to express CXCR6 which homes to the liver and other tissues.
7. This aids dengue spreading through the blood to the liver, and T cells migrate to these secondary sites.
40
What are Langerhans cells?
Skin resident dendritic cells
41
What other immune cell is involved in dengue infection?
1. Mast cells
2. They degranulate to limit infection
3. They can have a role in vascular permeability changes in severe dengue.
4. They also produce cytokines to recruit T cells.
42
How do we know adaptive immunity has a role in dengue immunopathology?
The fact that heterologous dengue infection is a big risk factor for severe dengue.
43
What are the 2 ways pre-existing antibodies can act in secondary dengue infection?
1. They can be cross-reactive and strongly neutralising and efficient at clearing dengue virus.
2. Not specific enough to neutralise dengue so non-neutralising antibodies.
44
How do non neutralising antibodies cause antibody dependent enhancement of dengue infection?
1. Non-neutralising antibodies don't coat the viral particle properly but can still bind to dengue.
2. The Fc part of the antibody binds to FcR and then the dengue virus can be internalised into phagocytes and replicate.
3. This leads to higher viral loads.
4. This is Fc receptor mediated viral entry and causes ADE.
45
What are the 3 types of antibodies that cause antibody-dependent enhancement (ADE)?
1. Cross-reactive low affinity antibodies.
2. Low antibody concentrations.
3. Antibodies targeting specific epitopes.
46
What is the main antibody epitope involved in ADE?
1. Anti prM antibodies.
2. prM is the membrane precursor protein for dengue.
3. These particles are not infectious but if antibodies bind the immature virus prM they can then enter cells through Fc-FcR binding.
4. This makes the immature virus particles infectious so they can replicate.
47
What are the 2 types of ADE?
1. Extrinsic ADE
2. Intrinsic ADE
48
What is Extrinsic ADE in dengue?
The increase viral entry into cells due to Fc-FcR binding and non-neutralising antibodies.
49
What is intrinsic ADE in dengue?
1. This is the modulation of the immune response once the antibody-virus complex is internalised.
2. This is the blocking of the type 1 interferon response.
3. This creates a more favourable environment for viral replication.
50
How does intrinsic ADE modulate the immune response to dengue?
1. The antibody virus complex binds to activation Fc receptors
2. This includes FcyR1, FcyR2a, FcyR3.
3. These should activate the anti-viral response but they don't.
4. It causes a drop in Type 1 IFN responses and leads to ADE.
5. There is decreased type 1 interferon signalling and production of interferon stimulated genes.
51
How does dengue binding to LILRB1 inhibit the FcyR induced anti-viral response?
1. When the virus is not properly coated by antibodies some viral receptors are left open.
2. These bind to LILR-B1 which triggers Syk dephosphorylation.
3. This prevents signalling for type 1 interferon production and interferon stimulated genes.
4. This increases viral replication inside cells and triggers ADE.
52
Do pre-existing antibodies increase the risk of severe dengue or ADE?
1. A paediatric cohort was followed for 12 years and blood samples were collected every time they were sick.
2. If these kids had antibody levels within a specific window, that put them at risk of severe dengue.
3. The risk decreases against in really high antibody levels.
53
What else can increase the risk of severe dengue?
increased levels of afucosylated IgG
54
What is afucosylated IgG?
1. 94% of circulating IgG is fucosylated.
2. Fucosylation is a post translational modification.
55
Why does afucosylated IgG increase the risk of severe dengue?
1. It has an increase affinity for FcyR3a/b.
2. We don’t know why but patients with severe dengue have more afucosylated IgG of specific dengue antibodies or total antibodies.
56
What is antigenic sin in dengue infection?
1. In primary infection, T cells develop to the specific dengue serotype.
2. On secondary infection with another serotype, T memory cells can be activated due to the lower activation threshold and higher frequency.
3. Dengue memory cells for other serotypes can recognise, clear and lyse infected cells in secondary infection.
4. OR these T memory cells can be sub-optimally activated as the TCR has lower affinity for the dengue antigens.
5. This virus is not cleared, and the cells are not lysed. This can cause cytokine storms and plasma leakage.
57
What are the differing roles of T cells during dengue infection?
1. T cell could cause immunopathology and poor viral clearance and cytokine storms.
2. They can also cause immune protection and viral clearance.
3. This depends on the situation and T cell affinity.
58
What are T cell response like in severe dengue?
1. T cell responses to all the dengue proteins is bigger in severe dengue patients.
2. Particularly NS3.
59
How does T cell function change in primary and secondary dengue infections?
1. CD107a shows degranulation capacity hasn’t changed.
2. But in secondary infection patients the T cells how more cytokine production and less cytotoxic functions.
3. Dengue could cause impairments of cytotoxic functions.
4. It could also skew the T cell response towards cytokine production.
60
How does MICB SNPs increase the risk for severe dengue?
1. MICB is a stress ligand expressed during dengue infection and other stress.
2. SNPs in MICB increase the risk of developing severe dengue.
3. MICB binds to NKG2D which activated CD8 and NK cells and is important for cytotoxic functions.
4. Dengue could be compromising these cytotoxic functions.
61
What is antigenic sin?
1. When the epitope of an infection is similar to the epitope of a previous similar infection, the immune system relies on the memory of the previous infection rather than mounting a new primary response.
2. This can result in an inadequate immune response to the infection as the immune system is not adapting to the new infection.
3. This is antigenic sin
4. This can apply to vaccines where the vaccine can act as this first infection.
5. This can cause antibody dependent enhancement of disease
