Dengue Diagnosis

World Health Organization (WHO) Case Definition of Dengue Fever (DF)

Probable DF refers to an acute febrile illness with ≥2 of the following:

• Headache, retro-orbital pain
• Myalgia, arthralgia
• Rash
• Hemorrhagic manifestations
• Leukopenia, thrombocytopenia, hematocrit rise of 5-10%
• Supportive serology
  • Hemagglutination-inhibition (HI) Ab titer of ≥1:1280
  • Comparable immunoglobulin G (IgG) titer with enzyme-linked immunosorbent assay (ELISA)
  • Positive immunoglobulin M antibody (IgM Ab) test on a late acute or convalescent-phase serum specimen
• Occurrence at the same location and time as other confirmed cases of DF

Confirmed DF is a probable case confirmed by at least 1 of the following laboratory criteria:

• Isolation of the dengue virus from serum, cerebrospinal fluid (CSF), or autopsy samples
• Demonstration of a ≥4-fold rise in serum IgG or IgM antibody titers particular to dengue virus
• Demonstration of dengue virus Ag in autopsy tissue, serum or CSF samples by immunohistochemistry, immunofluorescence or ELISA
• Detection of dengue virus genomic sequences through reverse transcriptase-polymerase chain reaction (RT-PCR)

Any probable or confirmed case of dengue should be reported

WHO Case Definition of Dengue Hemorrhagic Fever (DHF)

All of the following must be present:

• Fever, or history of acute fever, lasting 2-7 days, occasionally biphasic
• Hemorrhagic tendencies, evidenced by at least 1 of the following:
  • Positive tourniquet test
  • Petechiae, ecchymoses or purpura
  • Bleeding from the mucosa, gastrointestinal (GI) tract, injection sites or other locations
  • Hematemesis or melena
• Thrombocytopenia (≤100,000/mm³)
• Evidence of plasma leakage due to increased vascular permeability, manifested by at least 1 of the following:
  • A rise in the hematocrit (Hct) ≥20% above average for the corresponding age, sex and population
  • A drop in the hematocrit following volume-replacement treatment ≥20%
  • Signs of plasma leakage eg pleural effusion, ascites and hypoproteinemia

WHO Case Definition of Dengue Shock Syndrome (DSS)

All of the 4 criteria for DHF must be present, plus evidence of circulatory failure manifested by:

• Rapid and weak pulse, tachycardia
• Narrow pulse pressure (<20 mmHg) with increased diastolic blood pressure (DBP)
• Hypotension for age
• Cold clammy skin, restlessness, lethargy

Grading Severity of Dengue Hemorrhagic Fever (DHF) [Based on World Health Organization (WHO)]

• DHF is classified into 4 grades of severity, where grades III and IV are considered to be dengue shock syndrome (DSS). The presence of thrombocytopenia with concurrent hemoconcentration differentiates grades I and II from dengue fever (DF)
  
• Grade I: Fever accompanied by nonspecific constitutional symptoms; hemorrhagic manifestation is a positive tourniquet test, evidence of plasma leakage
  
• Grade II: Spontaneous bleeding in addition to the manifestations of Grade I patients, usually in the form of skin or other hemorrhages
  
• Grade III: Circulatory failure manifested by a rapid weak pulse and narrowing of pulse pressure or hypotension, with the presence of cold clammy skin and restlessness
  
• Grade IV: Profound shock with undetectable blood pressure (BP) and/or pulse

Criteria for Hospitalization

General Condition

• Continuous fever ≥3 days
• Lethargy, restlessness, irritability, excessive tiredness, drowsiness
• Generalized flushing
• Shortness of breath
• No improvement or worsening of condition

Dehydration

• Unable to tolerate oral fluid intake
• Vomiting, diarrhea

Abdominal Discomfort

• Epigastric pain, tender hepatomegaly

Hemorrhagic Manifestations

• Positive tourniquet test
• Petechiae, ecchymoses or purpura
• Spontaneous mucosal bleeding, epistaxis
• GI bleeding, excessive menstrual bleeding, hematuria
• Thrombocytopenia (platelet count: ≤100,000/mm³)
• Patients with active bleeding regardless of platelet count
• Patients without bleeding tendency but platelet count on a rapid downward trend

Plasma Leakage

• Rapidly rising hematocrit
• Hematocrit ≥20% of baseline
• Serositis eg pleural effusion, ascites
• Suspected plasma leakage in:
  • Males with hematocrit >47%
  • Females with hematocrit >40%

Circulatory Failure/Shock

• Rapid and weak pulse
• Narrowing of the pulse pressure to <20 mmHg
• Hypotension
• Cool, mottled or pale skin; cold extremities with circumoral cyanosis
• Changes in mental status, restlessness, lethargy
• Oliguria
• Tachypnea due to metabolic acidosis

Clinical Predictors for Bleeding

• Hypotension
• Narrowing of pulse pressure to <20 mmHg
• Hepatosplenomegaly or hepatomegaly
• Severe thrombocytopenia (platelet count <50,000/mm³)
• Leukopenia
• Elevated serum alanine aminotransferase (ALT) (>3x compared to normal value)
• Presence of vomiting, abdominal pain, restlessness, serosal effusion, or rashes

Criteria for Discharging Hospitalized Patients

• Absence of fever for at least 24 hours without the use of antipyretics
• Rising platelet count of >50,000/mm³
• Stable hematocrit
• Visible clinical improvement
• Return of appetite
• Good urine output
• No respiratory distress
• At least 2-3 days have passed after recovering from shock
  
  
  
  

Dengue Fever (DF)

• Blood pressure (BP)
• Hydration status, capillary refill time
• Tourniquet test which is performed as follows:
  • Inflate a BP cuff on the upper arm to a point midway between the systolic and diastolic pressures for 5 minutes
  • Test is positive when ≥20 petechiae/square inch are observed

Dengue Hemorrhagic Fever (DHF)

• Positive tourniquet test
  • Discrete fine petechiae scattered in the extremities, axillae, face and soft palate seen during the early febrile phase
  • Increase in capillary fragility reflected by positive tourniquet test and easy bruising
• BP is decreased as an effect of plasma leakage into the extravascular compartment following an acute increase in vascular permeability

Dengue Shock Syndrome (DSS)

• Rapid weak pulse with narrowing of the pulse pressure (<20 mmHg)
• Pleural effusion and ascites detected by physical examination or radiography

Other Lab Tests for Dengue

Complete Blood Count (CBC)

• Normal total white blood cell (WBC) but leukopenia may develop throughout the febrile period
• Mild increase in hematocrit may be due to dehydration
• Platelet counts and factors of blood clotting mechanism can be normal
  • Thrombocytopenia may occur

Albumin

• Usually low

Liver Function Tests (LFTs)

• Elevation of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels

Urinalysis

• Check for hematuria

Dengue Hemorrhagic Fever (DHF)

Radiography

• Evidence of pleural effusion and ascites due to acute increase in vascular permeability

Dengue Fever (DF)

Detection of Antigens

• Developments in enzyme-linked immunosorbent assay (ELISA) and dot blot assays directed to the envelope/membrane (E/M) antigen and the non-structural protein (NS1) showed that high concentrations of these antigens (immune complexes) could be detected in patients during the early period of the disease and in patients with primary and secondary dengue infections up to 6 days after the start of illness
• Non-structural protein 1 (NS1) ELISA
  • Has been shown to be a useful diagnostic tool in acute dengue infections
  • Produced by all flaviviruses and is secreted from mammalian cells
  • Dengue NS1 antigen has been detected in the serum of dengue virus-infected patients as early as 1 day post-symptom onset (PSO) and up to 18 days post onset (DPO)
  • May also be useful for differential diagnosis between flaviviruses because of its specificity
  • NS1 antigen detection kits are commercially available and can be used by laboratories with limited equipment and yield results within a few hours

Serology

• Hemagglutination inhibition (HI) test
  • Simple, sensitive and reproducible
  • Requires paired sera: Soon following hospital admission (acute) and 10-14 days after discharge (convalescent)
  • Titer of ≥1:1280 in an acute or convalescent phase serum sample is considered presumptive evidence of current dengue
  • Negative in early acute blood specimen and a repeat specimen should be tested before confirming or excluding dengue infection
  • Limited by inability to discriminate between infections by closely related flaviviruses

• Immunoglobulin M (IgM) antibody capture enzyme-linked immunosorbent assay (MAC-ELISA)
  • Serological test of choice because ELISA IgM detected is specific for flavivirus
  • Rapid, simple, requires little sophisticated equipment
  • A single, properly timed blood sample may be adequate
  • High increase in molar fraction of IgM indicates a primary acute flavivirus infection and low increase in molar fraction of IgM indicates a secondary acute flavivirus infection
  • High IgM antibody (Ab) response indicates recent primary flavivirus infection and low IgM Ab response indicates recent secondary flavivirus infection
  • Positive dengue IgM result indicates acute or recent past infections (up to 90 days); a single positive dengue IgM may not be indicative of a present dengue infection
  • May also be used to detect cerebrospinal fluid (CSF) IgM
  • However, MAC-ELISA cannot be used to identify infecting virus serotype
  • Positive results do not necessarily denote a current dengue infection
    • Caution must be exercised when making decisions about patient management

• IgG ELISA
  • Comparable to the HI test and can be used to differentiate primary and secondary infections

• Neutralization test (NT)
  • Most specific and sensitive serologic test for dengue virus
  • Not commonly used because it is expensive, requires a long time for performance and is technically difficult
  • Serum dilution plaque reduction NT is the most common protocol

• Complement fixation (CF) test
  • Least sensitive serological assay
  • CF appears later than IgM or HI Ab and is usually less specific
  • Has greater specificity for primary infections but is not specific in secondary infections

Other Diagnostic Tests

• Virus isolation
• Molecular techniques including reverse transcriptase-polymerase chain reaction (RT-PCR)
  • Specificity and sensitivity are better than virus isolation with a faster turnaround time

Dengue Hemorrhagic Fever (DHF)

Hematology Tests

• Evidence of disseminated intravascular coagulation
  • Thrombocytopenia on complete blood count (CBC)
  • Prolonged prothrombin time and partial thromboplastin time (PTT)
  • Decreased fibrinogen level and increased level of fibrinogen degradation products
• Leukopenia, hemoconcentration (rising hematocrit)

Dengue Shock Syndrome (DSS)

• The finding of a continuing drop in the platelet count concurrent with a rise in the hematocrit is an important indication of DSS