Evaluation of commercial rapid diagnostic test kits for dengue infections in India

Dengue is acommon acute viral febrile illness in the tropics. Dengue fever (DF), caused bya mosquito-borne virus of the flaviviridaefamily, is endemic to India and can present with a wide range of clinical manifestations.It comprises four distinct serotypes (DEN-1, DEN-2, DEN-3 and DEN-4).Typical DF is characterized by high fever, severe headache, myalgia,arthralgia, retro-orbital pain and maculopapular rash. Some patients developpetechiae, bruising or thrombocytopenia. TheNational Vector Born Disease Control Programme(NVBDCP) in India reported 111,880 dengue cases and 227 deaths due to dengue inyear 2016. The overall case fatality rate (CFR) of dengue fever is lessthan 1%. However CFR of patients who progressto develop dengue haemorragic fever isbetween 3–5%. Althoughresulting in an estimated 372 (210–520) DALYs per million inhabitants inSoutheast Asia , approximately 90% of patients with dengue infection remainasymptomatic. It is, however, likelythat many of the patients who do present with mild-moderate symptoms of dengueare inappropriately prescribed antibiotics.

Recognitionof acute dengue infection may be challenging since the symptoms arenon-specific and resemble several other causes of acute undifferentiatedfebrile illness, particularly in areas where there are other vector-borne diseases. The World HealthOrganization (WHO) syndromic case definition for dengue can help in identifyingdengue cases in endemic areas. However, the syndromic approach alone may beinadequate in the diagnosis or management of severe dengue cases.  The clinical presentation of acute dengueinfection is non-specific, but aproportion of patients progress to severe dengue haemorrhagic fever/dengue shock syndrome (DHF/DSS). Therefore, early and accuratediagnosis of dengue infection and initiation of appropriate treatment are thekey components in the management of severe dengue infection.

Thecharacteristics of an ideal dengue diagnostic test depend on the purpose forwhich the test will be used. The ideal test for early diagnosis of dengueinfection should be able to distinguish it from other diseases of similarclinical spectrum (such as malaria,leptospirosis, typhoid, typhus and chikungunya). Thetest needs to be highly sensitive during the acute stage of infection, rapid,inexpensive, easily performable and can be utilisedat temperatures above 30 °C.

Thetarget product profile (TPP) of an ideal test for use for the epidemiologicalsurveillance and outbreak investigation of dengue should include givingpositive results as soon as possible after onset of symptoms to provide earlywarning. Unlike the TPP needed for routine use in disease control programmes, these types of test needs to behighly specific and should be able to determine the dengue virus serotypes. Thesecond priorities of an ideal test for epidemiological surveillance andoutbreak investigations should be its ability to distinguish between primaryand secondary infection, its high throughput capacity and long shelf life.

Current WHO recommendations for the diagnosis ofdengue include enzyme-linked immunosorbent assay (ELISA)-baseddetection of dengue-specific Immunoglobulin M (IgM) antibodies or a ≥4-fold increase in the titre of total antibodies to dengue virus in paired acute andconvalescent sera, or detection of dengue virus by reversetranscription–polymerase chain reaction (RT-PCR). In the Indiancontext, the government of India’s National Vector Borne Disease Control Programme(NVBDCP) guidelines from 2015 recommend the use of an ELISA-based antigendetection test (NS1) for diagnosing the cases from day 1 to day 5 of illness,and the antibody detection test IgM Capture ELISA (MAC ELISA) for diagnosingthe cases after the 5th day of onset of disease for confirmation of dengueinfection.

However,these tests are expensive, time-consuming,arduous, technologically demanding and are not always available in most resource-limited settings and during epidemics.Hence, a large fraction of medical practitioners depend on readily availablerapid diagnostic test (RDT) kits which are cheaper and have less turn-aroundtime for the early diagnosis of dengue infection. Most of the commercially available RDTs are based onimmuno-chromatographic or ELISA methodsto detect IgM and Immunoglobulin G (IgG)antibodies- with or without NS1 antigen- in the serum of the patients. Manydifferent RDT kits are manufactured in India (and used internationally) and other RDTsthat are not produced in India but are registered in the country are available commercially. The latter are widely used forthe diagnosis of dengue fever in secondary and tertiary care centres across India as these give results withinan hour and are technically less demanding.

Theissue, however, lies with the reliability and performance of these tests, whichare yet to be independently evaluated against an acceptable reference standardin India.  A WHO/TDR/PDVI laboratory networkin 2009 evaluated selected commercial ELISAs and first-generation rapiddiagnostic tests, finding that ELISAs generally performed better than rapidtests. They, however,did not look at NS1 antigen based rapid tests or combination tests, and none ofthe study sites were within India. There is limited evidence of evaluation ofsuch RDTs against clinician based diagnosis. However,comparison with a laboratory-confirmed diagnosis based on ELISA is needed toensure the robustness of the results. Therefore, in this study, we propose to  evaluate readily available commercial rapiddiagnostic test kits for the diagnosis of dengue fever.

Inthis laboratory-based phase 2 diagnostic evaluation study using archived serumsamples, we will evaluate the sensitivity and  specificity of six commonly used rapid diagnostictest kits in India manufactured by five different manufacturers:Panbio Diagnostics,  Australia; Standard Diagnostics, Korea;  J. Mitra, India;  Zephry Biomedicals,India; and MP diagnostics,USA.