Animal models: virology II Flashcards

1
Q

What is the percentage of results obtained in mice models that are somehow translated to humans?

A

~ 8%

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2
Q

Why do we want fewer animals?

A

3E’s –> Ethics, Economics and Efficacy

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3
Q

What is needed to replace animal models? (3)

A
  • Legislation
  • Mindset
  • Science
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4
Q

How can horizontal virus transmission between rodents occur? (3)

A
  • Saliva
  • Urine
  • Faeces
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5
Q

What are Orthohantavirus transmission risk factors? (3)

A
  • Virus in the environment
  • Human infection
  • Rodent infection
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6
Q

What are risk factors for orthohantaviruses to be in the environment? (3)

A
  • Rodent density
  • Host infection phase
  • Temperature, moisture, UV-radiation
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7
Q

What are the risk factors for rodent infection? (9)

A
  • Seasonality
  • Age
  • Sex
  • Maternal antibodies
  • Co-infections
  • Resistance genes
  • Rodent density
  • Species diversity
  • Habitat type
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8
Q

What are the risk factors for human infection? (5)

A
  • Rodent density
  • Occupation/recreation
  • Environmental conditions
  • Season
  • Resistance genes
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9
Q

Why does each strain of Orthohantavirus have a specific rodent host?

A

Strains have co-evolved with specific rodent species

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10
Q

Why is there a clear geographical distribution of Orthohantavirus strains?

A

Strains have co-evolved with specific rodent species –> live in certain location

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11
Q

Rodents carrying Orthohantaviruses are mostly symptomatic/asymptomatic

A

Asymptomatic

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12
Q

What symptoms do Orthohantaviruses often cause?

A

Haemorrhagic fever with renal syndrome (HFRS)

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13
Q

Which Orthohantaviruses are found in rodents in Europe? (4)

A
  • Puumala virus
  • Dobrava virus
  • Seoul virus
  • Tula virus
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14
Q

On what factors is a Orthohantavirus infection diagnosis based?

A

Clinical signs and serology

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15
Q

Serological assays to confirm Orthohantavirus infection are based on?

A

Cross-reactive antigens

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16
Q

What are the three different clinical manifestations of Orthohantavirus infections?

A
  • Haemorrhagic fever with renal syndrome (HFRS)
  • Nephropathia epidemics (NE)
  • Hantavirus cardiopulmonary syndrome (HCPS)
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17
Q

Which Orthohantavirus causes nephropathia epidemica (NE)?

A

Puumala virus

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18
Q

What are the structural proteins of Orthohantaviruses? (3)

A
  • Membrane glycoproteins Gn and Gc
  • Polymerase L
  • Nucleocapsid N
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19
Q

How do Orthohantaviruses attach?

A

Interaction with Gn/Gc (cell surface receptors)

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20
Q

Which process is initiated by the attachment of Gn/Gc proteins to integrin?

A

Endocytosis

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21
Q

Which integrins are bound by pathogenic orthohantaviruses?

A

B3-integrins

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22
Q

Which integrins are bound by NON-pathogenic orthohantaviruses?

A

B1-receptors

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23
Q

Integrins are present on which cell types?

A

Endothelial cells, macrophages and platelets

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24
Q

Integrins are involved in…(3)

A
  • Regulation of endothelial cell adhesion
  • Platelet aggregation
  • Extracellular matrix interactions
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25
Q

Which process plays a central role in the pathogenesis of Orthohantavirus infections?

A

Vascular barrier loss

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26
Q

Orthohantaviruses: what causes disruption of vascular integrity?

A

Binding of Orthohantavirus glycoproteins to B3-integrin

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27
Q

What encompasses the disruption of vascular integrity caused by Orthohantaviruses? (3)

A
  • Capillaries more permeable
  • Arteriole vasoconstriction & vasodilation disrupted
  • Binding to platelet receptor affects clotting and platelet function
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28
Q

Why do different Orthohantaviruses have different pathogenesis?

A

Different viruses bind endothelium in different locations

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29
Q

Where do HFRS bind to the endothelium?

A

Lungs, kidneys, spleen

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30
Q

Where do HCPS bind to the endothelium?

A

Lungs, liver, heart, spleen

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31
Q

Which immune response is mounted in humans to combat Orthohantaviruses?

A

Virus epitopes expressed on surface of host cells –> CD8+ T cell attack on host tissue

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32
Q

Why does the immune response against Orthohantaviruses that is mounted in human not occur in rodents?

A

Downregulation by Tregs

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33
Q

Describe the kinetics of Orthohantavirus infection in rodents (2)

A
  • After acute infection: virus disappears from blood
  • As infection continues: long-lasting antibody response are produced
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34
Q

Why do rodents not get sick from hantavirus infection

A

Local increases of Tregs, and decreases of CD8+ T cells at primary sites of replication

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35
Q

What does the term ‘neuro-invasion’ mean?

A

The ability of a virus enter either the PNS or CNS

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36
Q

How is neuro-invasion described in the case of respiratory viruses?

A

The ability to travel from the respiratory tract to the CNS

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37
Q

What are the routes of virus spread into the CNS? (4)

A
  • Peripheral nerves
  • BB barrier
  • Cranial nerves
  • Blood-CSF barrier
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38
Q

Virus spread: which kind of transport is used via the peripheral nerves?

A

Retrogade transport

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39
Q

Virus spread: which kind of transport is used via the BB barrier?

A

Intracellular (Trojan Horse)

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40
Q

What does the term neurotropism mean?

A

The ability of a virus to infect and replicate in cells of the nervous system

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41
Q

What does the term neurovirulence mean?

A

The ability of a virus infection to cause lesions in the CNS that contribute to the development of clinical disease of the nervous system independently of its neuroinvasiveness or neurotropism

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42
Q

Which route of neuro-invasion occurs in SARS-CoV2 infection?

A

Mostly olfactory nerve

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43
Q

Describe the preferred neurotropism in SARS-CoV2 infection (2)

A
  • Glomerular layer neural cells
  • Cortical neurons
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44
Q

Describe the neurovirulence in SARS-CoV2 infection (4)

A
  • Lasting immune activation -> cognitive impairments
  • Loss of oligodendrocytes -> memory impairments
  • Shrinkage of specific brain regions
  • Increased risk for depression, anxiety, dementia, psychosis
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45
Q

What categories of pathological changes due to virus infections exist? (5)

A
  • Necrosis
  • Inflammation
  • Hyperplasia/neoplasia
  • Hypoplasia
  • Atrophy
46
Q

What is the difference between hypoplasia and atrophy?

A

Hypoplasia = something is not formed which would usually be formed (lack of growth)
Atrophy = loss of tissue

47
Q

Name an example of necrosis caused by viral infection

A

Herpesviruses cause necrosis of epithelial cells

48
Q

Name an example of inflammation caused by viral infection

A

Pneumonia from H5N1 influenza –> infection of alveolar epithelial incited marker immune response from host

49
Q

Name an example of hyperplasia/neoplasia caused by viral infection

A

Pock from poxvirus –> hyperplasia of epidermal cells

50
Q

Name an example of hypoplasia caused by viral infection

A

Cerebellar hypoplasia from ZIKV

51
Q

Name an example of atrophy caused by viral infection

A

Villus atrophy from coronaviruses (in animals)

52
Q

What are the pathological mechanisms of viruses? (3)

A
  • Direct damage to host cell
  • Induction of host immune responses
  • Transformation of infected host cell
53
Q

Direct damage to infected host cell, can be through… (4)

A
  • Inhibition of host cell DNA/RNA/protein synthesis
  • Damage to host cell integrity
  • Lysis of host cells
  • Induction of apoptosis/necroptosis
54
Q

Why can inhibition of host cell DNA/RNA/protein synthesis cause direct damage to infected host cells?

A

Viruses take over the replication machinery to produce virus particles

55
Q

What causes damage to host cell integrity by viral infection?

A

Viral replication processes

56
Q

What is often the purpose of lysis of host cells caused by viral infection?

A

Release of viral particles

57
Q

Induction of apoptosis/necroptosis is often not a viral strategy, what is it?

A

Usually a side effect of cellular replication

58
Q

What are the main components of the innate immune response against viral infection? (5)

A
  • Infected cells
  • Sentinel cells
  • Effector cells
  • Cytokines
  • Complement
59
Q

What is the main effector cytokine during viral infection?

A

IFN –> triggers neighboring cells to prevent further infection

60
Q

Describe the events (in order) of the innate immune response against viral infections (3)

A
  • PRRs on sentinel cells recognize virus motifs
  • Infected/sentinel cells produce cytokines
  • Complement activation
61
Q

What are the typical presentations of innate immune responses against viral infections? (5)

A
  • Calor
  • Rubor
  • Tumor
  • Dolor
  • Functio laesa
62
Q

What is Calor?

A

Increased blood flow

63
Q

What is Rubor?

A

Increased blood flow (by cytokines) -> dilutes pathogen, extravasation of inflammatory cells

64
Q

What is Tumor?

A

Increased blood vessel permeability (oedema) -> dilutes pathogen, extravasation of inflammatory cells

65
Q

What is Dolor?

A

Inflammatory processes

66
Q

What are examples of lesions associated with the adaptive immune response? (B-cells) (2)

A
  • Virus-antibody complexes
  • Non-protective antibodies
67
Q

Name an example of virus-antibody complexes in viral infection

A

LCMV infection in mice, hepatitis B infection in humans

68
Q

Name an example of non-protective antibodies in viral infection

A

Dengue virus infections in humans

69
Q

Name two examples of viruses that have caused systemic inflammatory response syndrome

A
  • H5N1 influenza
  • SARS-CoV2
70
Q

What are the mechanisms of transformation of the infected host cell caused by viral infections? (3)

A
  • Expression virus-encoded oncogenes
  • Insertional mutagenesis
  • Anti-apoptotic
71
Q

Name an example of the expression of virus-encoded oncogenes

A

HPV production of E6 and E7

72
Q

Name an example of insertional mutagenesis

A

HPV viral genes inserted into host genome

73
Q

Name an example of anti-apoptotic measures

A

EBV -> immortalization and proliferation of B cells

74
Q

What are the possible evolutionary benefits/selective advantages causing disease due to viral infections? (3)

A
  • Virulence is coincidental byproduct
  • Virulence is beneficial for the virus
  • Co-evolution of virus and host
75
Q

What are the characteristics of ‘virulence is a coincidental byproduct’? (2)

A
  • No advantage to virus
  • Often in zoonotic viruses
76
Q

What are the two main hypotheses of ‘virulence is beneficial for the virus’?

A

A: intermediate virulence hypothesis
B: short-sighted evolution hypothesis

77
Q

Describe the intermediate virulence hypothesis

A
  • Virulence = cost to virus -> lower host survival
  • Virulence = benefit -> higher transmission
  • Intermediate level of virulence = favoured
78
Q

Describe the short-sighted evolution hypothesis

A
  • Within host competition among virus genotypes
  • Genotype with higher virulence favoured without enhanced transmission
79
Q

What are the three scenarios of co-evolution of virus and host?

A
  • A: Slow host evolution
  • B: Fast host evolution
  • C: Balance in evolution
80
Q

How does slow host evolution work?

A

Virus has upper hand -> outcome = high virulence

81
Q

How does fast host evolution work?

A

Host has upper hand -> outcome = low virulence

82
Q

How does balance in evolution of virus and host work?

A

Both host and virus evolve -> outcome = intermediate virulence

83
Q

What are possible complications in understanding evolution of host-virus interaction? (3)

A
  • Host usually infected by multiple pathogens
  • Some viruses can infect several host species
  • Environment involved
84
Q

Which Rabiesvirus structural proteins are involved in blocking of the interferon/antiviral response? What do they inhibit? (3)

A
  • P-protein -> IRF3/STAT1,2 signaling
  • N-protein -> RIG-I activation
  • M-protein -> NF-kB activation
85
Q

Which Rabiesvirus structural protein is involved in the blocking of inflammatory responses?

A

G-protein -> inhibition of inflammatory responses by macrophages

86
Q

How does the P-protein of Rabiesvirus inhibit IFR3 signaling?

A

Blocks phosphorylation of IRF3 -> no dimerization and translocation to nucleus

87
Q

How does the N-protein of Rabiesvirus inhibit RIG-I activation?

A

Inhibits RIG-I’s ability to recruit factors to phosphorylate IRF3

87
Q

How does Rabiesvirus block NF-kB activation?

A

No translocation to nucleus of p50-p65

88
Q

How does Rabiesvirus block interferon-stimulated genes (ISGs)?

A

P-protein blocks translocation to nucleus & binding of the complex to ISGs tf’s

89
Q

Dog bites normally recruit strong immune responses due to high bacteria numbers and tissue damage. Which pathway prevents this during Rabiesvirus infection?

A

Cholinergic anti-inflammatory pathway

90
Q

What are commonly used in vivo (invertebrate) models that replace animal models? (3)

A
  • Drosophila melanogaster
  • Nematode
  • Galleria mellonella = greater wax moth larvae
91
Q

What is a con of using G.Mellonella as a model?

A

Relatively new model -> no techniques to genetically manipulate this organism

92
Q

What are the pro’s of using G.Mellonella as a model? (5)

A
  • Drug discovery
  • Tissue recovery
  • Phagocytosis
  • Hyphae evaluation
  • Mammalian temperatures
93
Q

What kind of infection models are used using G.Mellonella as a model? (4)

A
  • Gram-negative bacteria
  • Gram-positive bacteria
  • Fungi
  • Virus
94
Q

Why is G.Mellonella a relatively cheap model?

A

Larvae can be brought or grown on site

95
Q

What can be said about the temperature at which you keep the G.Mellonella larvae? Why is this advantageous?

A

Can be kept at 37C -> not possible for some other invertebrate model systems

96
Q

What are types of experiments that can be performed using G.Mellonella? (5)

A
  • Survival/virulence
  • Fungal load
  • Histology
  • Host responses
  • Efficacy of treatments
97
Q

The immune system of G.Mellonella has similarities/no similarities to that of humans

A

Similarities

98
Q

How do you perform a survival experiment using G.Mellonella?

A

Infect larvae using injection in pro-legs -> monitor survival

99
Q

How do you monitor survival of G.Mellonella larvae?

A

Melanisation -> immune system uses melanin -> blacker larvae = sicker larvae

100
Q

How can you analyse the survival data of G.Mellonella larvae? (2)

A
  • Kaplan-Meier plots -> survival for various doses
  • LD50 -> concentration at which 50% of larvae die
101
Q

Which two techniques can be used to determine the fungal load in G.Mellonella larvae?

A
  • Harvest hemolymph and plate out
  • Homogenize complete larvae
102
Q

How do you perform a histological examination of infection of G.Mellonella larvae? (2)

A
  • Fix larvae in formalin
  • Process for histology
103
Q

G.Mellonella only has an innate immune system, consisting of two components… (2)

A
  • Cellular immune response: haemocytes
  • Humoral immune response
104
Q

What are haemocytes? Why?

A

They are like neutrophils -> receptors, signaling pathways and defense mechanisms are similar

105
Q

What encompasses the humoral immune response of G.Mellonella? (3)

A
  • Opsonins
  • Antimicrobial peptides
  • Melanisation
106
Q

How is the severity of the immune response measured in G.Mellonella?

A

Haemolymph melanisation -> measuring concentrations

107
Q

What types of experiments are possible using G.Mellonella as a drug discovery model? (3)

A
  • Toxicity
  • PK/PD
  • Survival analysis
108
Q

How do you perform a PK/PD experiment using G.Mellonella?

A

Harvesting hemolymph at different timepoints after injection -> how fast is compound broken down?

109
Q

What is a difficulty in antifungal therapy studies using G.Mellonella?

A

Consecutive injection impossible due to small larvae size (max. 3 injections)