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Dr KAMRAN AFZAL
Asst Prof Microbiology
Dengue Virus
   Genus: Flavivirus
   Family: Flaviviridae
   Single-stranded RNA arbovirus
   40–50 nm, spherical, lipid env
   Genome is approximately 11 Kb
   4 serotypes:
       DEN-1, DEN-2, DEN-3 and DEN-4
       Infection with one dengue serotype provides lifelong immunity
        to that serotype, but there is no cross-protective immunity to
        the other serotypes
Dengue Virus Particles
Aedes aegypti
Aedes Mosquito - Characteristics
Distinct physical feature
   Black and white stripes

       on its body and legs
Flying time: dusk to dawn

Bites during the day

Short flight: house to house

Imperceptible bite
   Lays its eggs in clean, stagnant water
      Pots, pans and utensils
Life cycle of Aedes aegypti




                 4. Adult




3. Pupae                      1. Eggs




                2. Larvae
Did you know…

   The female mosquito lays 30–150 eggs every 2-3 days
   Only the female Aedes mosquitoes feed on blood
       Female mosquitoes need the protein found in blood to produce eggs
       Human blood is preferred with the ankle area as a favored feeding
        site
   Male mosquitoes feed on plant and fruit nectar
   After 8-12 days of acquiring the virus, mosquitoes become
    infective, remain so for whole life span (1-3 months)
   The eggs of A. aegypti can resist desiccation for up to 1 yr
Epidemiology
   Endemic in more than 100 countries
   A leading cause of hospitalization and death among
    children in Asia
   2.5 billion people or two fifths of the world population
    are now at risk
   50 million cases of DF worldwide every year
   0.5 million cases of DHF require hospitalization each
    year, a very large proportion of whom are children,
    ~ 5% die
   Without proper treatment, DHF fatality rates can
    exceed 20%
Worldwide Prevalence
World at Risk
Worldwide Resurgence
Transmission Cycle
   Transmission from human to human requires
    the same female mosquito to bite a viremic
    human and then bite a susceptible human at
    an interval of around 8-12 days
Pathogenesis of DHF - 1
   Homologous Antibodies form non-infectious complexes
Pathogenesis of DHF - 2
   Heterologous Antibodies form infectious complexes
Pathogenesis of DHF - 3
   Heterologous complexes enter monocytes, where
    virus replicates
Pathogenesis of DHF - 4
   Infected monocytes → vasoactive
    mediators → increased vascular
    permeability and hemorrhagic
    manifestations (DHF and DSS)
   Activation of memory T cells →
    Inflammatory cytokines, including TNF-α
    and interleukins (IL-2, IL-6, and IL-8) →
    Increased vascular endothelial permeability
    or trigger death of target cells through
    apoptosis
Pathology of DHF and DSS - 1
DF
 Rash in DF show a lymphocytic vasculitis in the dermis

DHF and DSS
 Most of the morphologic abnormalities result from DIC
  and shock
 Hemorrhage and congestion are seen in many organs

 Hemorrhagic effusions in the pleural, pericardial and
  abdominal cavities
Pathology of DHF and DSS - 2
   Liver and spleen are the target organs
       DEN virus antigen detected in hepatocytes
   Liver - Variable morphology
       Severe, diffuse hepatitis with mid-zonal necrosis
       Focal areas of necrosis
       No PMN cells, and no lymphocytes in the liver lesions
       Kupffer cells destroyed in cases of focal or severe necrosis
       Councilman hyaline body is present as an apoptotic cell
• Spleen - Enlarged and hemorrhagic
Dengue Clinical Syndromes

   Classic dengue fever (DF)
   Dengue hemorrhagic fever (DHF)
   Dengue shock syndrome (DSS)
       DSS is actually a severe form of DHF
Case Definition: Dengue Fever
   An acute febrile viral disease frequently presenting with
    headaches, bone or joint pain, muscular pains, rash and
    thrombocytopenia (Breakbone fever)
   Incubation period
       3-7 days (range 2-15 days)
   Temperature
       Bimodal peak (saddle-back form)
   Rash
       Maculo-papular, lasts for 3-4 days
Case Definition: DHF
4-Necessary Criteria (W.H.O.)
   Fever, or recent history of acute fever
   Hemorrhagic manifestations
   Low platelet count (100,000/mm3 or less)
   Objective evidence of “leaky capillaries”:
       Elevated hematocrit (20% or more over baseline)
       A positive ‘Tourniquet Test’
       Pleural or other effusions
Case Definition: DSS
   4-N criteria for DHF
       +
   Evidence of circulatory failure
       Rapid and weak pulse
       Narrow pulse pressure (< 20 mm Hg) OR
        hypotension for age
       Cold, clammy skin and altered mental status
   Frank shock is direct evidence of circulatory failure
   DEN-2
   Mortality rate 1-10% (up to 40%)
Hemorrhagic Manifestations

   Skin hemorrhages
       Petechiae, purpura, ecchymoses
   Gingival bleeding
   Nasal bleeding
   Gastrointestinal bleeding
       Hematemesis, melena
   Hematuria
   Increased menstrual flow
Tourniquet Test
   Inflate blood pressure cuff to
    a point midway between
    systolic and diastolic pressure
    for 5 minutes
   Positive test:
       20 or more petechiae per 1 inch²
        (6.25 cm²)
Differential Diagnosis
   Influenza
   Measles
   Rubella
   Malaria
   Typhoid fever
   Leptospirosis
   Meningococcemia
   Rickettsial infections
   Bacterial sepsis
   Other viral hemorrhagic fevers
Common Misconceptions - DHF
   Dengue fever + bleeding = DHF
       Apply 4 WHO criteria and assess capillary permeability
   DHF kills only by haemorrhage
       Patient dies as a result of shock
   Poor management turns dengue into DHF
       Poorly managed dengue can be more severe, but DHF is a
        distinct condition, which even well-treated patients may develop
   Positive tourniquet test = DHF
       Tourniquet test is a nonspecific indicator of capillary fragility
Treatment
   Monitor blood pressure, hematocrit, platelet
    count, level of consciousness and urine output
   Fluids
       The volume of fluid needed is similar to the treatment
        of diarrhea with mild to moderate isotonic dehydration
   Antipyretics
       Avoid aspirin and NSAIDS
   Blood and components therapy
   Continue monitoring after defervescence
       Patients develop DHF after defervescence
Laboratory Diagnosis
   Non-specific lab tests
       CBC - WBCs, platelets, hematocrit
       Urine - check for microscopic hematuria
   Dengue-specific lab tests
       Serology
            IgM ELISA / ICT
            IgG ELISA / ICT
            NS1 Antigen ELISA / ICT
       Virus isolation
            PCR to determine serotype
            Cell culture
            Mosquito Inoculation
ELISA Test for Serologic Diagnosis
Virus Isolation: Cell Culture/
 Fluorescent Antibody Test
Virus Isolation: Mosquito Inoculation
Prevention and Control
Mosquito Repellants
   Insecticide sprays
       Knock down and residual
   Repellant oils
   Repellant lotions
   Electric lamps
   Mats
   Coils
   High frequency sound
   Use a larvicide
       Temephos
Mosquito Barriers
   Keep patient in screened sickroom or
    under a mosquito net
   To prevent Aedes aegypti mosquitoes
    from biting infected patients and
    acquiring the virus
Outdoor Fogging
Indoor Fogging
Aerial Spraying
Media Campaigns
Health Ministry Campaign
CDC Handout
CDC Investigation Report
Dengue Vaccine?

   No licensed vaccine at present
   Effective vaccine must be ‘tetravalent’
   Clinical trials of an attenuated tetravalent
    vaccine currently underway
   Effective, safe and affordable vaccine will not
    be available in the immediate future
Role of Pak Army in Dengue
Role of Pak Army in Dengue
Future Outlook
   Continued vigilance and surveillance is very important
   True and active community participation is the key
   It is likely to be few years before the vaccine can be
    considered for widespread use in populations in endemic
    areas
   The only control method is:
           ‘ Efficient vector control ’
Aedes aegypti




 Thank you

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Dengue

  • 1. Dr KAMRAN AFZAL Asst Prof Microbiology
  • 2. Dengue Virus  Genus: Flavivirus  Family: Flaviviridae  Single-stranded RNA arbovirus  40–50 nm, spherical, lipid env  Genome is approximately 11 Kb  4 serotypes:  DEN-1, DEN-2, DEN-3 and DEN-4  Infection with one dengue serotype provides lifelong immunity to that serotype, but there is no cross-protective immunity to the other serotypes
  • 5. Aedes Mosquito - Characteristics Distinct physical feature Black and white stripes on its body and legs Flying time: dusk to dawn Bites during the day Short flight: house to house Imperceptible bite
  • 6. Lays its eggs in clean, stagnant water  Pots, pans and utensils
  • 7. Life cycle of Aedes aegypti 4. Adult 3. Pupae 1. Eggs 2. Larvae
  • 8. Did you know…  The female mosquito lays 30–150 eggs every 2-3 days  Only the female Aedes mosquitoes feed on blood  Female mosquitoes need the protein found in blood to produce eggs  Human blood is preferred with the ankle area as a favored feeding site  Male mosquitoes feed on plant and fruit nectar  After 8-12 days of acquiring the virus, mosquitoes become infective, remain so for whole life span (1-3 months)  The eggs of A. aegypti can resist desiccation for up to 1 yr
  • 9. Epidemiology  Endemic in more than 100 countries  A leading cause of hospitalization and death among children in Asia  2.5 billion people or two fifths of the world population are now at risk  50 million cases of DF worldwide every year  0.5 million cases of DHF require hospitalization each year, a very large proportion of whom are children, ~ 5% die  Without proper treatment, DHF fatality rates can exceed 20%
  • 13. Transmission Cycle  Transmission from human to human requires the same female mosquito to bite a viremic human and then bite a susceptible human at an interval of around 8-12 days
  • 14. Pathogenesis of DHF - 1  Homologous Antibodies form non-infectious complexes
  • 15. Pathogenesis of DHF - 2  Heterologous Antibodies form infectious complexes
  • 16. Pathogenesis of DHF - 3  Heterologous complexes enter monocytes, where virus replicates
  • 17. Pathogenesis of DHF - 4  Infected monocytes → vasoactive mediators → increased vascular permeability and hemorrhagic manifestations (DHF and DSS)  Activation of memory T cells → Inflammatory cytokines, including TNF-α and interleukins (IL-2, IL-6, and IL-8) → Increased vascular endothelial permeability or trigger death of target cells through apoptosis
  • 18. Pathology of DHF and DSS - 1 DF  Rash in DF show a lymphocytic vasculitis in the dermis DHF and DSS  Most of the morphologic abnormalities result from DIC and shock  Hemorrhage and congestion are seen in many organs  Hemorrhagic effusions in the pleural, pericardial and abdominal cavities
  • 19. Pathology of DHF and DSS - 2  Liver and spleen are the target organs  DEN virus antigen detected in hepatocytes  Liver - Variable morphology  Severe, diffuse hepatitis with mid-zonal necrosis  Focal areas of necrosis  No PMN cells, and no lymphocytes in the liver lesions  Kupffer cells destroyed in cases of focal or severe necrosis  Councilman hyaline body is present as an apoptotic cell
  • 20. • Spleen - Enlarged and hemorrhagic
  • 21. Dengue Clinical Syndromes  Classic dengue fever (DF)  Dengue hemorrhagic fever (DHF)  Dengue shock syndrome (DSS)  DSS is actually a severe form of DHF
  • 22. Case Definition: Dengue Fever  An acute febrile viral disease frequently presenting with headaches, bone or joint pain, muscular pains, rash and thrombocytopenia (Breakbone fever)  Incubation period  3-7 days (range 2-15 days)  Temperature  Bimodal peak (saddle-back form)  Rash  Maculo-papular, lasts for 3-4 days
  • 23. Case Definition: DHF 4-Necessary Criteria (W.H.O.)  Fever, or recent history of acute fever  Hemorrhagic manifestations  Low platelet count (100,000/mm3 or less)  Objective evidence of “leaky capillaries”:  Elevated hematocrit (20% or more over baseline)  A positive ‘Tourniquet Test’  Pleural or other effusions
  • 24. Case Definition: DSS  4-N criteria for DHF +  Evidence of circulatory failure  Rapid and weak pulse  Narrow pulse pressure (< 20 mm Hg) OR hypotension for age  Cold, clammy skin and altered mental status  Frank shock is direct evidence of circulatory failure  DEN-2  Mortality rate 1-10% (up to 40%)
  • 25. Hemorrhagic Manifestations  Skin hemorrhages  Petechiae, purpura, ecchymoses  Gingival bleeding  Nasal bleeding  Gastrointestinal bleeding  Hematemesis, melena  Hematuria  Increased menstrual flow
  • 26. Tourniquet Test  Inflate blood pressure cuff to a point midway between systolic and diastolic pressure for 5 minutes  Positive test:  20 or more petechiae per 1 inch² (6.25 cm²)
  • 27. Differential Diagnosis  Influenza  Measles  Rubella  Malaria  Typhoid fever  Leptospirosis  Meningococcemia  Rickettsial infections  Bacterial sepsis  Other viral hemorrhagic fevers
  • 28. Common Misconceptions - DHF  Dengue fever + bleeding = DHF  Apply 4 WHO criteria and assess capillary permeability  DHF kills only by haemorrhage  Patient dies as a result of shock  Poor management turns dengue into DHF  Poorly managed dengue can be more severe, but DHF is a distinct condition, which even well-treated patients may develop  Positive tourniquet test = DHF  Tourniquet test is a nonspecific indicator of capillary fragility
  • 29. Treatment  Monitor blood pressure, hematocrit, platelet count, level of consciousness and urine output  Fluids  The volume of fluid needed is similar to the treatment of diarrhea with mild to moderate isotonic dehydration  Antipyretics  Avoid aspirin and NSAIDS  Blood and components therapy  Continue monitoring after defervescence  Patients develop DHF after defervescence
  • 30. Laboratory Diagnosis  Non-specific lab tests  CBC - WBCs, platelets, hematocrit  Urine - check for microscopic hematuria  Dengue-specific lab tests  Serology  IgM ELISA / ICT  IgG ELISA / ICT  NS1 Antigen ELISA / ICT  Virus isolation  PCR to determine serotype  Cell culture  Mosquito Inoculation
  • 31. ELISA Test for Serologic Diagnosis
  • 32. Virus Isolation: Cell Culture/ Fluorescent Antibody Test
  • 35. Mosquito Repellants  Insecticide sprays  Knock down and residual  Repellant oils  Repellant lotions  Electric lamps  Mats  Coils  High frequency sound  Use a larvicide  Temephos
  • 36. Mosquito Barriers  Keep patient in screened sickroom or under a mosquito net  To prevent Aedes aegypti mosquitoes from biting infected patients and acquiring the virus
  • 44. Dengue Vaccine?  No licensed vaccine at present  Effective vaccine must be ‘tetravalent’  Clinical trials of an attenuated tetravalent vaccine currently underway  Effective, safe and affordable vaccine will not be available in the immediate future
  • 45. Role of Pak Army in Dengue
  • 46. Role of Pak Army in Dengue
  • 47. Future Outlook  Continued vigilance and surveillance is very important  True and active community participation is the key  It is likely to be few years before the vaccine can be considered for widespread use in populations in endemic areas  The only control method is: ‘ Efficient vector control ’