Dengue early detection helps cure

Dr Ranil S Dassanayake and Dr Nilmini Silva Gunawardene

Dengue viral fever is one of the most serious medical problems in Sri Lanka and at present a faction of the nation is at risk. This year alone up to now, dengue virus has infected over 24,000 persons in Sri Lanka and caused over 175 deaths; and the highest incidence have been reported to occur in the five to nine year age-group

Attempts to eradicate Aedes aegypti, the most efficient mosquito vector of the dengue virus, have not been successful in countries where dengue viral fever is endemic. The control of dengue therefore, may be possible once an efficient vaccine has been developed.

No dengue vaccine has been licensed. The development of an efficient dengue vaccine is difficult because the vaccine must be tetravalent so that it could provide protection against all four dengue serotypes.

Dengue virus enters the human bloodstream after being bitten by the mosquitoes and then spreads throughout the body. Symptoms appear in about eight to 10 days after a bite from an infected mosquito. Symptoms are flu-like and can include high fever, nausea, vomiting, body aches, and headache.

Although dengue fever may lead to a considerable debilitating condition, it is not fatal. A small proportion of patients infected with dengue viruses develop dengue haemorrhagic fever (DHF) and dengue shock syndrome (DSS) which may be fatal. Although the pathogenesis of DHF is not clearly understood. It is known that previous exposure to dengue infections increases risk in developing severe disease.

A diagnosis of dengue fever can easily be missed or delayed as the symptoms such as fever and body aches, closely resemble symptoms of influenza of even infected individuals can be asymptomatic. If symptoms are mild, they may be overlooked or assumed to be due to a mild influenza infection.

Because of the risk of serious complications, such as dengue haemorrhagic fever and dengue shock syndrome, it is always vital to detect early and differentiate dengue virus infection in the acute phase to provide timely clinical treatment and etiologic investigation and disease control. This lowers the chances of the risk of developing severe disease.

Diagnostic tests

There are many diagnostic tests available for the detection of dengue infection such as serological diagnosis of viral antigen and antibodies produced by human body against the virus, and viral genetic material-based tests.

Antibody against the virus and the viral antigen to appear in blood circulation generally after five days of dengue fever and they can only be detected when the titer of these molecules is significantly high in the blood circulation.

Diagnostic test methods based on dengue genetic materials is very robust, accurate and specific, and these tests are designed to analyse the unique regions of genetic material of dengue viruses, so that these assays only detect the genetic material of Dengue virus.

The genetic material of the virus is made up of RNA and like any other organism the genetic material of dengue virus is made of nucleotides, and genome has 11000 such nucleotides.

The genome of dengue virus codes for three structural proteins (C, prM, E) and seven nonstructural proteins (NS1, NS2a, NS2b, NS3, NS4a, NS4b, NS5; and short non-coding regions on both the 5’ and 3’ ends). In the genetic testing of the virus targets one of the regions mentioned above of the dengue virus genome, that is unique to the latter virus.

According to the genetic information dengue virus could be mainly divided into four genetically distinct serotypes, serotypes one to four. Studies of genome between the four serologically distinct dengue viruses have shown that each serotype can further be subdivided into several major genotypes.

Primary infection by a particular serotype induces a life-long protective immunity to the homologous serotype and however infection by other serotypes can produce a wide spectrum of disease severity and it has generally been accepted that secondary infection or infection with secondary or multiple infections with various dengue virus serotypes is a major risk factor for DHF-DSS due to antibody-dependent enhancement.

It has been known that the serotypes three are more virulent compared to other serotypes, secondary infection with such serotype has high tendency to develop into DHF-DSS.

These dengue serotypes could be identified on the very first day of fever using viral genetic material-based tests. Therefore, the detection is important as this information will help the clinicians to better manage patients.

The widely used genetic material-based test to identify dengue virus is RT-PCR, which is a laboratory technique commonly used in molecular biology to generate many copies of a DNA sequence, a process termed “amplification”.

The genetic material-based assay could be designed to identify only dengue genetic material, in the presence of many other different genetic material and also in the presence of a few dengue viral particles in the blood to be analyzed, with greatest accuracy. These assays also can produce results within four to six hours following the arrival of the samples to the laboratory.

A tool to fore warn

The constant mosquito surveillance to identify whether the mosquitoes carry the viruses is also important as information derived from such surveys could be used to develop predictive tools to fore warn of an impending dengue outbreak. These studies can often be carried out using tools in molecular biology.

In addition to RT-PCR, the genome sequencing of circulating virus and subsequent analyses of these sequences with the previously sequenced genome sequences using tools in bioinformatics will tell us important information such as the origin of the virus (the virus has migrated from another country), whether circulating virus is a virus that was identified during the past epidemic, the virulent attributes of the circulating virus, whether circulating virus is a new virus that has evolved from the virus that was identified during the past epidemics.

This information will undoubtedly help health authorities to take action to prevent dengue outbreaks.

Dengue diagnostics using tools in molecular biology such as RT-PCR, Real Time PCR, genome sequencing is considered to be faster and more reliable than other methods such as immunodiagnostics due to the high sensitivity and specificity and also rapid turnaround time.

When Dengue viral infection is raging on, we are indeed very fortunate that these facilities are now available in Sri Lanka; both in universities as well as leading private hospitals in Colombo.

The writers are Senior Lecturers in Biochemistry and Molecular Biology and Molecular Medicine of the University of Colombo Chemistry Department and Molecular Medicine Unit, Medicine Faculty, Kelaniya University