DENGUE VIRUS

Case-study
17 year-old female admitted into Princeton Plainsborough
Hospital. The patient present the following symptoms:

High Fever
Severe nausea
Mild nosebleeds
Joint pain
New appearance of skin rashes

Case-study

BACKGROUND INFORMATION
Word origin may be derived from Swahili or Spanish
3 forms of dengue:
Dengue fever (DF), dengue haemorrhagic fever (DHF), and
dengue shock syndrome (DSS)
Arboviral infection transmitted by the Aedes aegypti mosquito
Originates in African forests independent of humans breeds
in water storage containers slave and commerce trade
brought it to South-East
Asia & New World in 17th-19th centuries 1800 in global
tropical coasts

(Comprehensive Guidelines for Prevention

and Control of Dengue and Dengue
Haemorrhagic Fever, 2011)

GLOBAL DISTRIBUTION
(Comprehensive Guidelines for Prevention and Control of Dengue and Dengue Haemorrhagic Fever, 2011)

Found in tropical and subtropical regions of the world, 2.5 billion at risk
Endemic in more than 100 countries in the WHO regions:
Africa, Americas, Eastern Mediterranean, South-East Asia, and Western Pacific
South-East Asia and Western Pacific regions are the most seriously affected

EPIDEMIOLOGY
~ 50-100 million infections occur worldwide, 0.5 million hospitalized for DHF

~ 90% are children under 5 years

Classical DF more common in adults

frequency of epidemics observed, infection rates of the previously unexposed is 40% - 50%,
can reach 80% - 90%
Needs coincidence of many vector mosquitoes, many people with no immunity to virus
types, and opportunity for contact
Infection with any 1 of 4 serotypes lifelong immunity to that 1 serotype
Provides 1-3 years of cross-protection against other 3 serotypes
Reoccurring infection with different serotypes severe dengue
(DHF/DSS)
(CDC, 2014)

EPIDEMIOLOGY CONTD
Ae. aegypti strongly attracted to humans, domesticated, and is a
nervous feeder
Bites more than one host to complete blood meal and gonotropic cycle
Breeds in safe clean water, feeds at dusk and dawn
Results in multiple cases in cities
Transmission usually occurs in rainy seasons when humidity
and temperature are conducive for survival and breeding
transport, human contact, urbanization,
drinking water supply in rural areas brought DF to
urban and rural areas globally

(Comprehensive Guidelines for Prevention and Control of Dengue and

Dengue Haemorrhagic Fever, 2011)

PREVENTION
Key is disease surveillance to detect epidemics
Dengue-endemic regions should be educated about the virus,
recognize symptoms, and prevent transmission:
Regularly remove sources of stagnant water to prevent breeding
Use mosquito repellent, coils, and nets
Wear long, loose clothing in the daytime
Use nets and coils on those with DF to prevent mosquitoes
transmitting infections
Stay in AC or well-screened housing
Infected mosquitoes like to live in/around homes
with clean water

EVOLUTION
Genus: Flavivirus
Diverse
4 serotypes
1970: Central
America and Africa
(DEN 1 & 2)
Southeast Asia
(Den1-4)
2004: Worldwide
(Den1-4)
65% genome shared
Clinical

(Holmes,E & Twiddy, S, 2003)

VIRUS STRUCTURE & COMPONENTS

Dengue
Class IV: Positive Sense Single
Stranded RNA Virus

DNA Components
Internal Structure: 10 genes (3
structural and 7 non-structural )

Molecular Structure
External Structure : icosahedral &
50 nm in diameter

(Kuhn, R.J. et al. , 2002)

TRANSMISSION
Mosquito (female mosquito)
Possibly through blood
transfusions
Human incubation period ~ 4
days
Viremia lasts ~5 days
During viremia biting mosquitos
are susceptible to infection
Mosquitos incubation period ~812 days, which then it is
infectious for life
Virus resides in salivary glands
of infected mosquito (anticlotting factors, ect)

(Nishiura, H., & Halstead, 2007; Rodenhuis-Zybert, 201

DENGUE LIFE CYCLE

1.

1. Receptor-Mediated Endocytosis

2.

2. Fusion

3.

3. Nucleocapsid Release

4.

4. RNA Replication

5.

5. Translation

6.

6. Budding

7.

7. Furin Cleavage

8.

8. Progeny Release

Envelope Proteins
Lipid Bilayer
Capsid
RNA Genome

THE DENGUE VIRUS

(Martina, B. et al, 2009)

DENGUE LIFE CYCLE

Infected Mosquito Biting

DENGUE LIFE CYCLE

Extracellular
Matrix

DENGUE LIFE CYCLE

Extracellular Space
Dengue virus

Receptor
Endocytosis
Binding

Cognate receptor

Cell Membrane
Cytoplasm

DENGUE LIFE CYCLE

Fusion
Proton Pump

H+

H+

Endosome

Lowered pH Change
Conformational

Cytoplasm

DENGUE LIFE CYCLE

Proton Pump

H+

H+

Endosome

Fusion
Conformational
Change

Cytoplasm

DENGUE LIFE CYCLE

Fusion
Nucleocapsid
Release

Cytoplasm

DENGUE LIFE CYCLE

Nucleocapsid Release

Cytoplasm

DENGUE LIFE CYCLE

Cytoplasm

Viral RNA
Ribosomes

Rough Endoplasmic Reticulum

Viral RNA Translation

DENGUE LIFE CYCLE

Cytoplasm

Viral RNA
Ribosomes

Rough Endoplasmic Reticulum

Viral RNA Translation

DENGUE LIFE CYCLE

Cytoplasm

Capsid Proteins

RNA dependent
RNA Polymerase

Envelope Proteins

Rough Endoplasmic Reticulum

Viral RNA Translation

DENGUE LIFE CYCLE

Cytoplasm

Capsid Proteins

RNA dependent
RNA
Polymerase
RdRp

Proteases

Cell Membrane

Peptidases
Envelope Proteins

Viral RNA Translation

Rough Endoplasmic Reticulum

DENGUE LIFE CYCLE

Cytoplasm

Doublestranded
strandedRNA
RNA
Single
RdRp
RNA dependent RNA Polymerase

RNA Replication

DENGUE LIFE CYCLE

Cytoplasm

Double stranded RNA

RdRp

RNA Transcription

DENGUE LIFE CYCLE

Cytoplasm

Viral RNA
Ribosomes

Rough Endoplasmic Reticulum

Viral RNA Translation

DENGUE LIFE CYCLE

Cytoplasm

Viral RNA
Ribosomes

Rough Endoplasmic Reticulum

Viral RNA Translation

DENGUE LIFE CYCLE

Cytoplasm

RdRp
Capsid Proteins
R

Envelope Proteins

Rough Endoplasmic Reticulum

Viral RNA Translation

DENGUE LIFE CYCLE

Cytoplasm

Cell Membrane

Budding
Viral RNA
Translation
Rough Endoplasmic Reticulum

DENGUE LIFE CYCLE

RER

Golgi Apparatus

DENGUE LIFE CYCLE

Furin

Furin Cleavage
Golgi Apparatus

Golgi Apparatus

Cytoplasm

Molecular Structure
External Structure : icosahedral

Cognate receptor

Progeny Release

Extracellular Space

PATHOPHYSIOLOGY

Complement
Activation

T-cell
Immunology

ADE of
virus

Plasma
Leakage

Bleeding
Diathesis

Hypovolemic
Shock

(Nishiura, H., & Halstead, 2007; Halstead & Orourke,

1977) ).

PATHOPHYSIOLOGY- ADE Activation

Main Result: Increase in virus replication and
numbers

Heterotypic
antibodies
- Low concentration
- Partial neutralization
- Ie. prM

(Halstead et al. 2010)

PATHOPHYSIOLOGY- ADE Activation

Main Result: Increase in virus replication and
numbers

Heterotypic
antibodies
- Low concentration
- Partial neutralization
- Ie. prM

Virus uptake
- Delivery to Fc
receptor cells
(Halstead et al. 2010)

PATHOPHYSIOLOGY- ADE Activation

Main Result: Increase in virus replication and
numbers

Heterotypic
antibodies
- Low concentration
- Partial neutralization
- Ie. prM

Virus uptake
- Delivery to Fc
receptor cells

Replication
- Increased viral
replication compared
to virus alone
(Halstead et al. 2010)

PATHOPHYSIOLOGY- Complement
Activation

NS1- Glycoprotein secreted by dengue

infected cells

Activation of innate immune system on

endothelial cell surface
1) Classical complement
2) Alternative complement

Result:
1) Apoptosis of endothelial cell
2) Inflammation = CYTOKINES!

Anaphylatoxin
s
(Guzman & Kouri, 2002)

PATHOPHYSIOLOGY- T-Lymphocyte Pathogenesis

Primary
Infection

DENV-1

T- cell
selection

(Dejnirattisai et al., 2010)

PATHOPHYSIOLOGY- T-Lymphocyte Pathogenesis

Primary
Infection

DENV-1

T- cell
selection

Secondary
Infection

DENV-2

Clonal
Expansion
Cross-Reaction

(Dejnirattisai et al., 2010)

PATHOPHYSIOLOGY- T-Lymphocyte Pathogenesis

Primary
Infection

DENV-1

T- cell
selection

CYTOKINE
STORM

Secondary
Infection
IL-2
TNF

DENV-2

Clonal
Expansion
Cross-Reaction

IFN

(Dejnirattisai et
al., 2010)

ENDOTHELIAL CELL ACTIVATION

(Manson et al. 2003)

Cytokines induce changes

in endothelial
morphology:
Loss of vascular integrity
Increased adhesion molecules
Increased cytokine production
http://www.scielo.br/img/revistas/rb/v43n6/en_a13fig01.jpg

IMPLICATIONS: Plasma
leakage

Edema

Hypovolemic
shock

Thrombocytopeni
a

COAGULOPATHY

(Sellahewa, K, 2012)

SYMPTOMS
Dengue Fever and Dengue Hemorrhagic Fever
Febrile:
Hallmark trait is fever
Convulsions may occur due to high fever
(DHF)
Thrombocytopenia, leukopenia (levels more
drastic with DHF)
Critical
Plasma leakage into pleural cavities, ascites
(DHF)
Subnormal temperatures, defervescence
Varying degrees of hemorrhage (worsened
for DHF)
Recovery
Reabsorption of accumulated fluids
Improved vital signs
Important to monitor
(CDC, 2010)

Case-study
Back at Princeton Plainsborough, House has ordered his team to perform an
endoscopy to check for internal hemorrhaging

New symptom: Vomiting blood, excessive bleeding in gastrointestinal tract

Complications: Excessive loss of fluid, increased lymphatic return, overcompensation of
cardiovascular/adrenal/renal mechanisms, anoxia = DENGUE SHOCK SYNDROME

DIAGNOSIS

Clinical symptoms
Often misdiagnosed with influenza,
malaria, Typhoid fever,
Leptospirosis
Travel history
Viral markers IgG, IgM, and NS1
Laboratory confirmation:
IgM Capture ELISA
NS-1 specific assays
Lateral flow test to detect IgM & IgG
antibody, NS1 antigen

(CDC, 2010; Mayo Clinic, 2013)

ANTIVIRAL THERAPY
AG129 Mouse Model
deficient for interferon-// receptors
one of only models that permit infection by all 4
serotypes
used to target virus entry, membrane fusion, RNA
genome replication, assembly, & release from
DENVax
infected cell
targets protein E to interfere with viral
N-nonyl7-deaza-2-Creplication

methyl-adenosine

deoxynojirimycin

6-O-butanoyl
castanospermine

treatment tested in viremia mouse model (AG129 mice) during acute

phase
block viral replication
(Schul et al., 2007; Wilder-Smith et al.,
2010)

LIVE-ATTENUATED VACCINATION

DENVax

DENVax

based on the PDK53 DENV-2 backbone

containing pre-membrane & E genes of
serotypes 1-4
tested in cynomolgus macaques
Immunogenicity & efficacy results

LATV
contains 30 nucleotide deletion; tetravalent
tested in flavivirus-nave adults
no significant difference in adverse events
between vaccines and placebo-recepients
Race factor influencing infectivity of LATV virus

(Durbin et al., 2013; Osorio et al., 2011)

CHIMERIC LIVE-ATTENUATED VACCINATION

ChimeriVax Vaccine (Sanofi Pasteur)
uses 17D yellow fever vaccine virus
Phase I, II, III
shows 57% overall efficacy
reduction of hospitalization by 80%
89% reduction of dengue haemorrhagic fever
DENVax

Mahidol
PDK-53 DEN-2 backbone
Phase 1 trial (U.S. & Columbia)
protective when administered in monkeys and
mice
(Fink & Shi, 2014; Wilder-Smith et al.,
2010)

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