Dengue Diagnostics

Dengue virus (four serotypes, DENV-1 to DENV-4) is transmitted by Aedes mosquitoes. Diagnosis combines timing from onset with the right test. In the first 5 to 7 days (febrile phase) the virus, its RNA and non-structural protein 1 (NS1) antigen are detectable. From about day 5, IgM antibody appears and IgG follows.

Tests include RT-PCR, NS1 antigen (laboratory or rapid), IgM and IgG serology, and serotyping. Alongside these, the full blood count (haematocrit and platelets) and liver enzymes track severity and warn of plasma leakage.

Dengue affects an estimated 100 to 400 million people each year, and over 5 million cases were reported in 2023. About 1 in 20 develop severe disease, which includes plasma leakage with shock, severe bleeding and organ impairment. Risk is highest with a second infection by a different serotype, in infants born to immune mothers and in people with comorbidity. Case fatality in severe dengue falls under 1% with good fluid management.

Indications. Acute fever of 2 to 7 days with headache, retro orbital pain, myalgia, arthralgia, rash, nausea or leucopenia in an endemic area or after travel in the previous 14 days. Any warning sign (abdominal pain, persistent vomiting, mucosal bleeding, lethargy, liver enlargement, rising haematocrit with falling platelets). Outbreak investigation. Blood donors and organ donors from outbreak areas.

Specimen. Serum or plasma (no haemolysis) 2 to 3 mL, kept at 2 to 8 °C and frozen at minus 20 °C or lower for RT-PCR if delayed beyond 48 hours. Whole blood in EDTA for the full blood count.

Choice of test by day of illness.

NS1 antigen. Detects a secreted viral glycoprotein from day 1 to day 9. Sensitivity is higher in primary than secondary infection (often 80 to 90% versus 60 to 70%), because pre-existing antibody complexes the antigen. Specificity is over 95%. A positive result confirms infection. A negative result does not exclude it, particularly after day 5 or in secondary infection. Rapid lateral flow tests have lower sensitivity (50 to 80%).

RT-PCR. Detects viral RNA and gives serotype, which is of public health importance. A positive result confirms infection. Viraemia falls rapidly after defervescence.

IgM antibody capture ELISA (MAC-ELISA). A positive IgM after day 5 suggests recent infection. Cross-reactivity with other flaviviruses (Zika, Japanese encephalitis, yellow fever, West Nile) is frequent, and vaccination against yellow fever or Japanese encephalitis can produce false positives. Paired sera (acute and convalescent, 7 to 14 days apart) showing a fourfold rise in IgG or IgM seroconversion confirm infection.

IgG. In primary infection IgG appears after day 10 and persists for life. In secondary infection IgG rises within the first days to high titres (over 1 in 2560), which identifies a secondary immune response (a risk factor for severe disease).

Classification (2009). Dengue with or without warning signs, and severe dengue (plasma leakage with shock or respiratory distress, severe bleeding, or organ involvement with AST or ALT over 1000 U/L, impaired consciousness or cardiac involvement).

Supporting laboratory findings.

• Leucopenia (total white cells under 5000 /microL) is common early and precedes the platelet fall.
• Thrombocytopenia under 100 000 /microL in most patients. Platelets under 20 000 /microL with bleeding need urgent action.
• Haematocrit rise of 20% or more over baseline signals plasma leakage. The baseline is the earliest measured value or the expected value for age and sex.
• AST and ALT raised in most patients, with AST typically higher. Hypoalbuminaemia, hyponatraemia and raised lactate in severe disease.
• Prolonged APTT, low fibrinogen and raised D-dimer in severe cases.
• Ultrasound or chest radiograph showing pleural effusion, ascites or gallbladder wall thickening confirms plasma leakage.

Critical values and findings needing immediate action.

• Rising haematocrit with falling platelets at the time of defervescence (day 3 to 7): critical phase, with intravenous fluids and close monitoring.
• Shock (narrow pulse pressure under 20 mmHg, cold extremities, weak pulse, capillary refill over 2 seconds): immediate isotonic crystalloid resuscitation.
• Falling haematocrit with shock: occult haemorrhage, and blood transfusion is needed.
• Severe bleeding or platelets under 20 000 /microL with bleeding.
• AST or ALT over 1000 U/L, altered consciousness or myocarditis.
• Positive result in a blood donor.

Interferences and pitfalls.

• Testing outside the right window gives false negatives. A negative NS1 or PCR on day 6 or later is repeated with IgM.
• Serological cross-reaction with other flaviviruses and Zika can confuse diagnosis, and neutralisation tests (plaque reduction) are the reference method for specificity.
• Rheumatoid factor and some other conditions can cause false positive IgM.
• Aspirin and non-steroidal drugs worsen bleeding and are avoided.
• Dengue should be distinguished from malaria, chikungunya, Zika, leptospirosis, typhoid, influenza, rickettsial infection, early meningococcaemia and viral haemorrhagic fevers. Co-infection is possible.
• Fluid overload in the recovery phase causes pulmonary oedema, and haematocrit falls with convalescent reabsorption.
• Platelet count correlates poorly with bleeding risk, and prophylactic platelet transfusion is not routine.

Special groups.

• Pregnancy: risk of severe disease, vertical transmission, preterm birth and haemorrhage at delivery.
• Infants: mothers' antibody wanes between 4 and 12 months, with risk of severe disease on primary infection.
• Older adults and those with diabetes, hypertension, renal disease or obesity: higher risk of severe dengue.
• Regions: periodic epidemics in South and South-East Asia, Latin America and increasing numbers in Africa and the Pacific.

Follow up tests. Haematocrit and platelet count every 6 to 12 hours in the critical phase (more often in shock), liver enzymes, creatinine, electrolytes, albumin, coagulation tests, lactate, glucose, ultrasound for effusions, blood group and cross-match, and repeat serology for paired sera.