Dengue Viral Infections: A Major Public Health Issue

*1Nadeem Sajjad Raja,  2Nishi Nihar Singh,  3Tahir Mehmood, 4Habib Ur Rehman Sethi,  5Najam Ul Haq Raja , 6Khalid Abbas Janjua.

 

1Department of Medical Microbiology, Norfolk and Norwich University Hospital, Norwich

NR2 3TX, UK.

 

2 Cardiff School of Biosciences, Cardiff University, Museum Avenue, Cardiff,  UK.

 

3Medici Medical Centre (General Practice), Luton UK.

 

4Army Medical Corps, Rawalpindi, Pakistan.

 

5Fauji Foundation Medical Centre, Rawalpindi, Pakistan.

 

6Chemical Laboratory, Rawalpindi General Hospital, Rawalpindi, Pakistan.

 

*To whom correspondence should be addressed:

Dr Nadeem Sajjad Raja

Department of Medical Microbiology,

Norfolk and Norwich University Hospital,

Norwich

NR2 3TX, UK.

Email: rajanadeem@doctors.net.uk

 

 

 

Abstract

Dengue virus infection has emerged as a major public health problem worldwide in the recent decades. The incidence of this infection has increased in tropics and subtropics. It’s estimated that approximately one third of the world population are at risk of acquiring dengue infection with 100 million case of dengue fever (DF) and half million dengue haemorrhagic fever (DHF) and dengue shock syndrome (DSS) every year.  Dengue is a mosquito borne infection caused by four serotypes (type 1-4) of dengue virus. Severe forms (DHF/DSS) of dengue infection are under-recognised and the lack of awareness of the clinical features especially plasma leakage can lead to delayed recognition of DHF and DSS. No antiviral agents are available to treat dengue infection at the moment. Nevertheless, attempt to control dengue vector and introduction of safe and effective vaccine remain the important preventive measure. In this review article, we describe the epidemiology, virology, risk factors, pathogenesis and pathology, clinical features and management of DF, DHF and DSS, this review is also an update of diagnostics methods such as virus isolation, serology and molecular methods employed in clinical virology laboratory for the diagnosis of dengue infection.

 

Keywords: Dengue, Dengue Virus, Dengue haemorrhagic fever, Dengue shock syndrome

 

INTRODUCTION

Dengue is a widespread mosquito-borne illness in humans of the tropics and subtropics of the world (1, 2). It is thought that about 100 million cases of dengue fever, 500,000 cases of dengue hemorrhagic fever (DHF) and 12000 deaths occur worldwide annually (3, 4,5,6, 7). Aedes aegypti (A. aegypti), the mosquito vector of dengue virus is present in almost all tropics and sub-tropic of the world and it poses the greatest threat to one third of the worlds population. This infection has been expanding its endemic areas for several years. The current epidemiologic trend underscores the importance of dengue infections and there is need for appropriate understanding of presentation patterns, diagnosis, treatment and long-term improvements for disease control and surveillance of dengue infections. This article reviews the epidemiology, virology, clinical manifestations including pathogenesis and pathology, laboratory diagnosis, treatment and prevention of dengue infections.

 

EPIDEMIOLOGY

Dengue infections have largely been endemic in Asia (Pakistan, India, Sri Lanka, Maldives, Bangladesh and China), Southeast Asia, several southern and central Pacific countries and Americas. No dengue epidemic has yet been reported from Africa however periodic cases have occurred (8, 9). In Pakistan, cases of dengue infections have previously been reported in the medical literature from various regions of which, 3 dengue outbreaks have recently been documented in 1994, 2003 and 2005 (10, 11, 12). Ali and co-workers reported that 72 patients with suspected dengue fever (DF) were investigated of which 58% were found to be IgM positive. Only one fatal outcome was recorded in this case series (11). Another study reported ten confirmed cases of dengue with four deaths from the north eastern city of Pakistan in 2003 (12). Several dengue outbreaks have also been reported from India and Sri Lanka (13, 14). Dengue has emerged as an infectious disease in Southeast Asia especially in the Philippines, Indonesia, Malaysia and Thailand. Dengue infection poses a great threat to the local inhabitants of Malaysia due to rapid increase in population and industrialisation in the past few decades. Significant outbreaks of dengue infections have occurred frequently in 1974, 1978, 1982, 1986 and 1990. It is estimated that incidence of dengue infection remained high since 1990 therefore it remains an important cause of morbidity and morbidity (15, 16). A large study from Dong Nai Paediatric Centre, Vietnam revealed  of the 1136 children who were admitted in hospital; 712 had acute dengue infection, 339 suffered from moderate dengue and 85 were confirmed as not having dengue. Of the 712 children with acute dengue, 312 (44%) had severe dengue fever, 319 (45%) were classified as dengue haemorrhagic fever. No death was noted down in this case series (17). Dengue infection has increasingly been reported from Brazil, Cuba and Americas in recent years (18, 19).

 

VIROLOGY

Dengue infection (also called “break bone fever”) is an important infectious disease that is caused by the dengue virus, which belongs to family flaviviridae of the genus flavivirus (3). The flaviviruses are small surrounded by a spherical lipid envelope. The Dengue virus is a single stranded RNA virus, approximately 11 kilo bases with an icosahedral nucleocapsid covered by a lipid envelope (4). The dengue virus has four closely related but distinct serotypes, DEN1 to DEN4; within which are several genotypes. The virion is composed of 3 structural proteins (known as core, membrane and envelope) and 7 non-structural (NS1, NS2a, NS2b, NS3, NS4a, NS4b AND NS5) proteins (20). As the infection with 1 dengue virus provides life long immunity, there is no cross protective immunity to the other dengue viruses, therefore all dengue virus types may infect a person living in an endemic area. Dengue is understood to be an urban disease. These viruses maintain cycle of infection that uses the mosquito, the A. aegypti as a vector to infect the human host, who in turn serves as sources of viral amplification. The A. aegypti is a small highly domesticated, black and white tropical insect that prefers to feed on humans during the daytime. There are two peaks of biting activity; early morning for 2 to 3 hours and in the afternoon for several hours before dark. It breads in artificial containers in and around homes. Female A. aegypti feeds on several persons and may transmit dengue virus to many persons in short course of time (6, 21, 8).

 

CLINICAL FEATURES

Humans happen to infected with dengue virus after an infected mosquito prods on the vulnerable human host. There are several other possible routes of transmission without mosquito vector such as mucocutaneous transmission by blood of infected patient with dengue, needle stick injuries, bone marrow transplant, blood transfusion, intrapartum and vertical transmission (22). The incubation period of dengue infections is from 3 to 14 days (average 4-7 days). Serotype and virulence of dengue virus, sex, age, genetic background and immune status of the host are the established risk factors in dengue infections. For example, the elderly patients are more susceptible than younger children however the risk for severity of dengue infections does however fluctuate by age (9, 18, 23).  The clinical presentation of dengue infection depicts a wide spectrum of findings from asymptomatic or mild undifferentiated illness with maculopapular rash or mild self-limiting infection of DF to DHF and dengue shock syndrome (DSS) (3, 21, 24). DF is an acute febrile illness that appears after an incubation period of 4-7 days. In older patients the disease may be mild or severe with sudden onset of high-grade fever, chills, frontal headache, myalgia, arthralgia and a rash, vomiting and loss of appetite (6, 25). This febrile phase lasts for 2-7 days. Although it is a serious debilitating condition it is usually not fatal (25).  Clinical presentation of dengue infections in children is different and the major symptoms include are seizure, rash, coryza, and diarrhoea and less common symptoms are vomiting, abdominal pain and headache (26). Patients with DF also describe blurred vision after seven days of onset of illness while other clinical manifestations are disappearing. Focal retinal haemorrhage, Roth like spots, retinal oedema, cotton wool spots, vasculitis, and optic neuritis are the most common ocular manifestations (27).

 

DHF occurs less frequently than DF. Although it is primarily a disease of young children, adults may also suffer from DHF. Its clinical presentations are dramatic with abrupt onset of high-grade fever that subsides in approximately 2-7 days. However fever in DHF gives biphasic or saddleback curve. The signs of circulatory failure appear before or about 24 hours after the temperature hits normal or below (28). Blood tests usually illustrate thrombocytopenia and hemoconcentration as evidence of vascular leak syndrome. Skin haemorrhages like purpuric lesions, petechiae and ecchymosis are obvious hemorrhagic manifestation in DHF. The tourniquet test, which indicates that patient has increased capillary fragility, may be diagnostically helpful to the physicians (8). Scattered petechiae appear on the extremities, trunk, face and other parts of the body in patients with severe DSS. More severely ill patients may have gastrointestinal bleed. Shock in dengue infection is usually caused by plasma leakage. Von willebrand factor (VWF) antigen is a macromolecular antigen that is secreted predominantly by endothelial cells and smaller amount from platelet cell. Endothelial activation or injury leads to the release of VWF antigen from endothelial cells. This injury is manifested by a spectrum of clinical effects from hypotension to lung injury.  One study revealed that high levels of plasma VWF were associated with hospital deaths and a longer duration of mechanical ventilation (29). It is also thought that endothelial injury may contribute to the widespread formation of platelet micro thrombi leading to tissue ischemia and multiorgan dysfunction. The potential value of VWF antigen as a predictor of the development of acute respiratory distress has been established (5, 28, 29). 

 

In dengue infection, viremia usually hits the peak at the time or shortly after the onset of illness and remains detectable during illness period ranging 2 to 12 days. The number of cells infected with the virus determines the severity of dengue disease and the number of cells infected is related to antibody dependent enhancement (ADE) infection of peripheral leukocytes in secondary infections.  It is also hypothesised that there is an association between dengue viremia early in the course of illness and disease outcome in patients with secondary dengue virus 1 and dengue virus 2 infections. However no association has been found in patients with primary dengue virus 1 infection. It has been established that dengue virus 3 viremia (measured as dengue virus 3 genome equivalent levels) and subsequent immune activation were of greater magnitude in more severe clinical disease. The magnitude of plasma leakage was primarily linked to the magnitude of viremia. Sensitive and reproducible quantitative RT-PCR assays have been reliably used to find associations between higher and maximum viremia levels and increasing disease severity (30, 31). The clinical and laboratory parameters are summarised in Table1.

 

Complications in dengue infections are rare though, the reports of complications are on the rise. However severe dengue infections may give rise to complications (Table2) for example, liver failure, myocarditis, acute renal failure, haemolytic ureaemic syndrome, acute transverse myelitis, and encephalopathy and disseminated intravascular coagulation (DIC) (26, 32). Patients with persistent uncorrected shock may progress to acute respiratory distress syndrome (ARDS), abdominal compartment syndrome (ACS), neurological symptoms, diastolic dysfunction contributing to refractory shock and acute disseminated encephalomyelitis. ACS was described as abdominal distension with intra abdominal pressure >15 mmHg, and any of two signs from the following oliguria or anuria or metabolic acidosis or hypotension shock or respiratory distress (33).

 

 

 

PATHOGENESIS

The pathogenesis of DF, DHF and DSS is less understood and controversial. Dengue virus is a complex interplay of host and viral factors. Several risk factors for severe disease have been established such as age (8, 23), viral serotype (8), genotype (14) and genetic background (34). The ability to severe illness in primary infection varies between DV serotypes but secondary infection is by a heterologous serotype is the paramount risk factor for DHF and DSS (34, 35). Several studies have indicated that certain DEN2 and DEN3 genotypes are associated with DF and DHF (14, 36).  Watts et. al. (1999) showed that phylogenetic analysis of American genotype of DEN2 appeared to be more associated with DF while Asian genotypes were more linked with DHF (36).  Another study confirmed that Asian genotypes replicated to higher levels in human monocytes derived macrophages and dentdritic cells (37).

 

There are two common theories to explain the pathogenesis of dengue infection. The “immune enhancement hypothesis” is most commonly regarded as reliable. This hypothesis elaborates that patients in second infection with a heterologus dengue virus serotype have a profound higher risk for developing DHF or DSS (38). Already present heterologus dengue antibody (from previous infection) recognises the infecting virus and establishes an antigen-antibody complex, which is bound and internalise by immunoglobulin Fc receptor on the cell membrane of leukocytes. In such, circumstances the antibody is heterologus; the virus is not neutralized and is free to replicate inside macrophages. This is known as antibody dependent enhancement (ADE) and this phenomenon boosts the infectious process and replication of the virus in the cells of the mononuclear lineage (5, 30, 38).  Anti-DENV antibodies have demonstrated that they cross-react with platelets, clotting factors, and endothelial cells in humans (39). Lin et al (2005) described that anti-NS1 antibodies bind and induce apoptosis in endothelial cells and increase vascular permeability in DHF and DSS by the secretion of proinflammatory cytokines and chemokines (40). These cells are responsible for the increased vascular permeability that eventually leads to hypovolumia and shock.

 

The second theory claims that dengue virus does vary and mutates as a result of selection pressure as they replicate in human and or mosquitoes. There are also some virus strains that have greater epidemic potential (31). Phenotypic genetic changes in the virus genome may contribute in virus replication and viremia, severity of the disease and epidemic potential. Avirutnan et. al. (2006) revealed a strong association between peak levels of viremia and progression to DHF for circulation serotypes in Thailand. High levels of NS1 protein were found in the serum of patients with DHF therefore, NS1 could be useful prognostic indicator in DHF (41). Other viral proteins play less efficient roles in the severity of dengue disease.

 

PATHOLOGY

The pathology of DHF and DSS has been studied extensively. Pathological studies of tissues taken at autopsies have shown diffused petechial haemorrhages of most organs, as well as serous effusion in the pericardial, pleural and peritoneal cavities. Microscopically perivascular oedema and loss of integrity of endothelial junctions are found. There is no prominent damage to the endothelial cells or blood vessels. Midzonal necrosis and Councilman bodies are frequently found in the liver. Recent isolation of dengue virus from brain, cerebrospinal fluid and intrathecal antibody production suggest that dengue virus crosses the blood-brain barrier. There is increased proliferation of reticuloendothelial cells in the bone marrow, spleen, lymph nodes and lungs (24, 42). In a recent cohort study, Jessie and co workers revealed presence of dengue viral antigen in many tissues like liver, spleen, thymus, lung, lymph nodes, kidney and mononuclear phagocytic cells. This study also shows that B and T lymphocytes, endothelial and fibroblast cells could be probable source of infection and replication of virus. It has been established that viral replication occurs in vascular endothelial cells and it contributes to DSS (43).  

 

LABORATORY DIAGNOSIS

The diagnosis of dengue infections is difficult on the basis of clinical grounds only because more than half of patients are either asymptomatic or have diffuse picture of fever. This diffuse picture can be attributed to other similar infection such as malaria, typhoid fever, measles, leptospirosis, Epstein-Barr virus, cytomegalovirus, rubella, rickettsial infection and HIV seroconversion illness. Leucopoenia, thrombocytopenia, raised liver function tests, and hyponateremia are important laboratories features in dengue infections (12, 44, 45). A definitive diagnosis of dengue infection can be made only in the virology laboratories. Several types of methods are currently being employed in virology labs for the definitive diagnosis of dengue infections. The methods include virus isolation, detection of viral antigen and or antibodies and genomic sequencing by nucleic acid amplification assays (46, 47).  The dengue virus can be found in serum or plasma, circulating blood cells tissues from immune system between 2-7 days of onset of infection (47). Laboratory diagnostic methods are summarised in Table3.

 

Virus isolation

Virus isolation is considered as gold standard methods in the establishment of dengue disease. It is useful in further virological studies and detection of serotype of dengue virus by immunofluorescence staining method. The rate of virus isolation can be improved by collection of blood sample within first six days of disease as viral load is high during first week of illness (48). There are currently four isolation systems used for the isolation of dengue viruses; intracerebral inoculation of 1-3 days old baby mice, the use of mammalian cell cultures (primarily LLC-MK2 Cells), intrathoracic inoculation of adult mosquito and use of mosquito cell structure (3, 25). All DV serotypes are isolated by intracerebellar inoculation of new born mice. This method has not been popular in recent years because of low sensitivity, long isolation time and high cost (49). On the other hand mosquito inoculation and mosquito cell cultures are the most sensitive virological methods. The former method is rarely used for virus isolation now adays. The main disadvantages are extraordinary precautions required to prevent the release of infected mosquito and hard work needed to produce large number of mosquitoes for this method. Four mosquito species such as Aedes aegypti, Aedes albopticus, Tooxorhynchities amboinensis and Tooxorhynchities splendens are used for virus isolation. Serotyping of all DV is done on the brain, salivary gland tissues of mosquito by the immunofluorescence assay (IFA) after one week of isolation at suitable temperature (50, 51). IFA is used for identification of viruses. It is a more reliable and rapid method. The different types of mosquito cell culture for example C6/36 (A. alpopictus), AP61 (A. pseudoscutellaris) and TRA 284 (Toxorhynchities amboinensis) are used for virus isolation and then serotyping. The diluted serum is introduced to the mosquito cell culture monolayer on screw cap tubes, dishes of flasks. Cytopathic effect (CPE) is often detected in within 11 days after inoculation at suitable temperature. IFA stain with all four different serotype specific monocloncal antibodies is used to determine the serotype of DV (48, 50, 51). The cell line C/636 is considered as the method of choice for routine dengue virus isolation. Mammalian cell culture like the Vero cell culture, primarily LLC-MK2 cells are not recommended for routine dengue virus isolation in the clinical laboratories. DV needs multiple passages in this cell culture in order to induce CPE.

 

Serological diagnosis

Five basic serological techniques have been routinely used for the diagnosis infection; hemagglutination inhibition (HI), complement fixation test (CF), neutralization test (NT), immunoglobulin IgM capture enzyme linked immunosorbent assay (ELISA) and indirect IgG ELISA (1, 3, 6). Regardless of test used, serological diagnosis depends upon a significant 4-fold rise in the titre of specific antibodies between acute and convalescent serum samples. The dengue antibodies are best detected around the day 5 of the illness. It is understood that on the whole, serological techniques used in clinical laboratories have several limitations. The most important drawback is the high cross reactivity between antigens of flaviviruses including all four DV, yellow fever virus, Japanese encephalitis virus or St. Louis encephalitis virus. It also remains difficult to establish the serodiagnosis of past, recent and present dengue infection due to long standing persistence of immunoglobulin G. It is a well established factor that the diagnosis of dengue virus infection is more complicated than the diagnosis of other viral infections because the patient may have been infected with more than one serotype of DV. Infection with one serotype does not confer immunity to other three serotypes of the DV (20, 52, 53).

 

HI has remained the standard serological diagnostic technique in dengue viral infections due to its high sensitivity rate, reproducibility and easy of execution. It is also used to differentiate primary infection from secondary infection. In primary dengue, antibodies are detected on 5 or 6 day of onset of acute period of disease and antibodies titres are more than 1:10. The antibody titre in convalescent sera is often less than 1:640. On the contrary, the antibodies in secondary dengue can be detected soon after the appearance of clinical illness. Rapid rise in antibodies titres in early days of illness is an important diagnostic feature. In secondary infection seroconvalescent antibodies titres are higher than or equal to 1:2560. In majority of patients, the antibodies levels remain for 2 to 3 months then start to decline. HI is also used for seroepidemiological studies in several parts of the world. The disadvantages include inability to differentiate different serotypes of DV, lack of specificity and require paired samples (46, 54, 55). CF is rarely used for routine dengue diagnosis in the virology laboratories as it requires highly qualified and trained technical staff and is difficult to perform to attain reliable results (53, 54). Although NT is an expensive and tedious technique, is has high specificity for dengue virus diagnosis and it can be used to identify the infecting serotype in primary dengue infection.

 

Among all serological diagnostic techniques, capture IgM and or IgG ELISA is the most routinely used method for the daily diagnosis of DV infections in clinical laboratories around the world. It has high sensitivity and specificity, easy to perform, no need for sophisticated equipments as well as it provides evidence of recent infection. It also has the ability to distinguish primary dengue infection from secondary dengue infection by capturing IgM or IgG. However, it has a few drawbacks such as the existence of rheumatoid factor in patient’s serum. This may interfere with the specificity of ELISA test in IgM detection and difficulty in detecting DV specific antibody due to cross reactivity between different serotypes of dengue virus. The test is also used to detect antigens (46, 47, 56, 57). IgM appears firstly in the serum and is detectable form day 3 to 5 of illness in patients. Several studies have shown anti-dengue IgM levels reach peak levels in approximately 2 weeks time; they start disappearing to undetectable levels in 60-90 days. IgG appears shortly after the disappearance of IgM IgM titres are markedly high in primary infection as compared to secondary infection (46, 53, 54). IgG ELISA is as sensitive as HI and is used in seroepidemiological studies. It can be used for the differentiation between primary and secondary DV infection. This test is simple to perform although it is not specific, cross reacts with other flaviviruses and unable to differentiate between dengue serotypes (58).

 

New diagnostic techniques

In the past few years, several new modern and latest methods have been introduced for dengue infection diagnosis. These include polymerase chain reaction (PCR), hybridization probes and immunohistochemistry. These methods have proven useful in the diagnosis of virus infection (1-4, 21, 25).

 

TREATMENT

There is no specific therapy and uncomplicated dengue infections are usually resolved spontaneously. Nevertheless, dengue viral infections with life threatening complications should be managed in hospital with purely supportive management (44, 45).

 

Dengue fever can easily be managed at home by administering analgesics and antipyretic or tepid sponging as it is a mild self limiting illness. Paracetamol (60 mg/kg/day) is the drug of choice in febrile patients. Aspirin and diclofenac sodium should be avoided because these agents pose a risk of gastric irritation and may cause bleeding. All patients should be observed closely by their general practitioner for the early detection of DHF and DSS. It is highly recommended that platelet counts and packed cells volume should be checked on daily basis. The low levels of the cells indicate the plasma leakage in dengue disease (59, 33). Antiemetic should be administered in patients with vomiting. Hospital admission is necessary if patient develops acute abdominal pain, decreased level of consciousness, cold extremities, bleeding, restless, laboratory evidence of DHF, urine output, and haemoconcentration. Patients with underlying risk factors such as age <1 years, heart disease, anaemia, massive bleeding, comatose, overweight/obese should be monitored closely (26, 60).  Patients with probable diagnosis of DHF should be hospitalised in rehydration ward. As mentioned above, antipyretic therapy is given and blood tests are done daily. The fluid and its volume should be determined to the degree of dehydration and electrolyte.  It is imperative to monitor the patients for the early signs of DSS. Shock usually occurs after the third day during transition from pyrexia to defervescence period. These patients have low platelet count or have significant loss of blood may require platelet transfusions (61). Patients with probable diagnosis DSS should be provided intensive unit therapy. Vital signs, haemoatocrit and platelet counts, urine output, level of consciousness and haemorrhagic manifestations need to be monitored regularly. Fluid replacement therapy plays a crucial rule in the reversal of DSS. Ringers lactate or 5% glucose at a rate of 10-20 ml/kg/hour is sufficient for infusion. It can be infused rapidly in severe shock. Dose can be raised to 20-30 ml/kg/hour. In case of persistent shock, plasma expander (10-20 ml/kg/hour) can be added (61). Fluid overload should be avoided in order to prevent pulmonary oedema, myocardial oedema and pleural effusion (62, 63). Diastolic dysfunction is an established complication in patients with persistent shock and should be investigated by echocardiography. ACS can be relieved by lowering intra abdominal pressure; hence it improves cardiopulmonary symptoms (33). Prothrombin and partial thromoboplastin times should be measures in patients with DIC and profuse bleeding. In these patients, fresh frozen plasma, platelet concentrate or cryoprecipitate is suggested (26). Management of dengue virus infection is summarised in Table4.

 

A good prognosis depends on the early diagnosis of the dengue infection, monitoring of the clinical condition of the patient like blood pressure, pulse, urine output, conscious level and good nursing care. The onset of plasma leakage in DHF and DSS progress rapidly and hemotocrit rises abruptly. If it is not managed quickly it may then lead to tissue hypoperfusion, tissue anoxia, metabolic acidosis and organ failure. Fluid replacement has significant role in the desirable outcome in the most dengue infections.

 

PREVENTION AND CONTROL

At the moment the tools available to prevent dengue infection is limited. There are no vaccines available in the market and options for mosquito control are also disappointing. In these circumstances the emphasis should be on disease prevention if the trend of emergent disease is to be reversed.

 

Active surveillance

Active surveillance remains a fundamental point of dengue prevention program. The main aim of this program should be to provide an early warning or predictive capability for epidemic transmission. Thus the epidemic can be prevented by emergency mosquito control (5).

 

Mosquito control

Prevention and control of dengue infections is based on controlling the mosquito vector in and around the houses, where most transmission occurs. The most effective way to control the mosquitoes is the reduction of larva by eliminating or cleaning of water holding containers that serve as the larval habitat for A. aegypti. Public involvement is necessary in order to implement mosquito control program. It can be achieved by public education and law enforcement (64). Massive media campaign against dengue should be launched in the country. All sections of the society such as school teachers, health workers, religious leaders, community leaders and school children should be encouraged to participate in dengue control program. Government should take necessary step to make people aware of dengue disease and its prevention.

 

Vaccine

An effective tetravalent vaccine remains a significant challenge. Two live attenuated dengue virus vaccines, attenuated by passing several times I non-human cells, have been developed. Trial of a tetravalent vaccine showed significant seroconversion rates (89%) against all 4 serotypes of DV after the third dose. However, two doses of this vaccine confer 80-90% protection in children (65). Other vaccine, prepared by Walter Reed Army Institute of Research, produced similar seroconversion rates in adult volunteer (60). Several risk factors are associated with live attenuated RNA vaccine such as reversal to a virulent phenotype, and short life.

 

CONCLUSION

Dengue virus infections are a major and emerging global health problem in present era. Since these infections are on the continuous rise, it remains important to describe and categorise the common manifestation of dengue infections, employ laboratory tests for its diagnosis and utilise all possible sources for the management and prevention of the disease.

 

Table1: The clinical and laboratory parameters and differential diagnosis of dengue infections 9, 12, 26, 44, 45.

Category

Clinical features

Laboratory features

Differential diagnosis

DF

Sudden onset, high grade fever, flu like illness, myalgia, retro-bulbar pain, maculopapular rash, cervical and occipital lymphadenopathy, ocular manifestations

Low white blood cells count, Low platelet count

Influenza, measles, rubella, leptospirosis, infectious mononucleosis, chickengunya, coxsackie,  parovirus and rickettsial infections,

DHF

Similar as above, intermittent high grade fever, flushing, abdominal pain, vomiting, anorexia, bleeding tendencies such as petechiae, bruises, haematemesis and meleana, epistaxis, signs of circulatory failure

Low platelet count, Low white blood cells count, increased haematocrit concentration, low albumin, disturbed liver function tests and electrolytes

Viral haemorrhagic disease such as yellow fever,  leptospirosis, meningococcemia, acute abdomen, hanta viral infection, chickengunya infection

DSS

As above as DF and DHF, cold extremities, signs of shock such oliguria, as low pulse, tachycardia, and hypotension, abdominal tenderness, impaired mental status, encephalopathy and coma

increased haematocrit concentration, metabolic acidosis,

Other viral haemorrhagic fevers, Septicaemia, meningococcemia,

 DF, Dengue fever; DHF,  Dengue haemorrhagic fever; DS,  Dengue shock syndrome

 

Table 2: Complications of the dengue infections.26,32,33

Complications

liver failure, myocarditis, acute renal failure, haemolytic ureaemic syndrome, acute transverse myelitis, encephalopathy, disseminated intravascular coagulation, acute respiratory distress syndrome, abdominal compartment syndrome, neurological symptoms, diastolic dysfunction contributing to refractory shock, acute disseminated encephalomyelitis

 

Table 3: Methods used in clinical laboratory for the diagnosis tests of dengue infection. 1-4,6,25

Laboratory tests

Virus isolation:

Intracerebral inoculation in baby mice

Mammalian cell culture

Mosquito cell culture

Adult mosquito inoculation techniques

Serological diagnosis:

Haemagglutination inhibition test

Enzyme linked immunosorbent assay

Indirect IgG Enzyme linked immunosorbent assay

Complement fixation tests

Neutralisation test

Molecular methods

Reverse-Transcriptase Polymerase Chain Reaction

Hybridization probes

Immunohistochemistry.

 

Table 4: Management of patients with dengue infection28,46, 63

Category

Management

DF

Home/outpatient treatment,

Analgesic and antipyretic (Paracetamol, aspirin is contraindicated)

Recognise early signs of DHF/DSS

DHF

Hospitalise the patient, Analgesic and antipyretic

Monitor vital signs, urine output and conscious level

Daily blood biochemistry (liver function tests, platelet, white blood cells, hematocrit concentration) for early diagnosis DHF and DSS

Increase fluid intake

Intravenous fluids to correct dehydration (Hartman’s solution, 5% dextrose saline)

Observe for haemorrhagic manifestations

DSS

·         Admit in Intensive care unit

·         Management as above

·         Oxygen

·         nursing care

·         Ringers’ acetate at a rate of 10-20 ml/kg body weight/hour, if no improvement increase 20-30 ml/kg body weight/hour and add plasma expander 10-20 ml/kg body weight/hour.

 

 

 

REFERENCES:

1.      Yamada KI, Tokasaki T, Nawa M, Yabe S, Kurane I.  Antibody response for Japanese dengue fever patients by neutralization and hemagglutination inhibition assays demonstrate cross reactivity between dengue and Japanese encephalitis virus. Clin Diagn Lab Iummunol 2003; 10:  725-28.

 

2.      Libraty DH, Young PR, Pickering D Endy TP, Kalayanarooj S, Green S, Vaughn DW, Nisalak A, Ennis FA, Rothman AL.. High circulating levels of the dengue virus nonstructural protein NS1 early in dengue illness correlate with the development of dengue hemorrhagic fever. J Clin Infect 2002; 186: 1165-8.

 

3.      Wang WK, Sung TL, Tsai YC Kao CL, Chang SM, King CC..  Detection of dengue virus replication in peripheral blood mononuclear cells from dengue virus type 2 infected patients by Reverse Transcription-Real-time PCR assay. J of Clinic Micrbiol 2002; 40: 4472-4478.

 

4.      Callahan JD, Wu SJ, Dion-Schultz A Mangold B,  Peruski LF, Watts DM, Porter KR, Murphy GR, Suharyono W, King CC, Hayes CG, Temenak JJ.  Development and evaluation of serotype- and group-specific fluorogenic reverse transcriptase PCR (TaqMan) assays for dengue virus. J Clin Micribiol 2001; 39: 4119-24.

 

5.      Lam SK. Dengue haemorrhagic fever. Rev in Med Microbiol 1995; 6: 39-48

 

6.      Koraka P, Suharti C, Setiati T, Mairuhu AT, Van Gorp E, Hack CE, Juffrie M, Sutaryo J, Van Der Meer GM, Groen J, Osterhaus AD..  Kinetics of dengue virus specific serum immunoglobulin classes and subclasses correlate with clinical outcome of infection. J Clin Microbiol 2001; 39: 4332-38.

 

7.      Wang WK, Lee CN, Kao CL, Lin YL, King CC. Quantitative competitive reverse transcription-PCR for quantification of dengue virus RNA. J Clin Microbiol 2000; 38: 3306-10.

 

8.      Gubler DJ. Dengue and Dengue Hemorrhagic Fever. Clin Microbiol Rev 1998; 11: 480-96.

 

9.      Raja NS, Devi S. The incidence Dengue disease in a University teaching hospital in Malaysia in 2003 and 2004.  Infect Dis J 2006; 15:99-102.

 

10.  Ansari JK, Siddique M, HussainT, Baig I, Tariq W.  Karachi outbreak of Dengue hemorrhagic fever in Karachi. Pak Armed Forces Med J  2001; 5: 94-8.

 

11.  Naseem Salahuddin, Farheen Ali, Muhammad Ali and Fauzia Rashid. Dengue fever outbreak in Karachi, 2005—a clinical experience. Infect Dis J 2005; 14: 115-7.

 

12.  Ali N,  Nadeem A, Anwar M, Tariq WU,  Chotani RA. Dengue fever in malaria endemic areas. J Coll Physicians Surg Pak  2006; 16: 340-2.

 

13.  Dar L,  Broor S, Sengupta S, Xess I,  Seth P. The first major outbreak of dengue hemorrhagic fever in Delhi, India. Emerg Infect Dis 1999; 5: 589-90.

 

14.  Messer WB, Gubler DJ, Harris E, et al. Emergence and global spread of a dengue serotype 3, subtype III virus. Emerg Infect Dis 2003; 9: 800–9.

 

15.  Fong, MY, Koh CL, Lam SK.  Molecular epidemiology of Malaysian dengue 2 viruses isolated over twenty five years (1968-1993). Res Virol 1998; 149: 457-64.

 

16.  Porter KR, Beckett CG, Kosasih H, Tan RI, Alisjahbanan B, Rudiman PF et al.  Epidemiology of dengue and dengue hemorrhagic fever in a cohort of adults living in Bandung, West Java, Indonesia. Am J Trop Med Hyg 2005; 72: 60-6.

 

17.   Phuong CX, Nhan NT, Kneen R, Thuy PT, van Thien C, Nga NT, Thuy TT, Solomon T, Stepniewska K, Wills B; Dong Nai Study Group. Clinical diagnosis and assessment of severity of confirmed dengue infections in Vietnamese children: is the world health organization classification system helpful? Am J Trop Med Hyg 2004; 70: 172-9.

 

18.  Vaughn DW. Deng ue Lessons from Cuba. Am J Epidemiol 2000; 152: 800–3.

 

19.  Siqueira JB Jr. Martelli CM, Coelho GE, SImplicio CR, Hatch DL. Dengue and dengue hemorrhagic fever, Brazil, 1981-2002. Emerg Infect Dis 2005; 11: 48-53.

 

20.  Guzman MG, Kouri G. Advances in dengue diagnosis. Clin Diagn Lab Immunol 1996; 3: 621–7.

 

21.  Hahn CS, French OG, Foley P, Martin EN, Taylor RP. Bispecific monoclonal antibodies mediate binding of dengue virus to erythrocytes in a monkey model of passive viremia.J of Immunol 2001; 166: 1057-65.

22.  Chen LH, Wilson ME. Transmission of dengue virus without a mosquito vector:   nosocomial mucocutaneous transmission and other routes of transmission. Clin Infect Dis 2004; 39: 56-60.

 

23.  Egger JR, Coleman PG. age and clinical dengue illness. Emerg Infect Dis 2007; 13: 924-5.

 

24.  Lam SK.  Dengue infection with central nervous system infections. Neurol J Southeast Asia 1996; 1: 3-6.

 

25.  Messer WB, Vitarana UT, Sivananthan K, Elvtigala J, Preethimala LD, Ramesh R, Withana N, Gubler DJ, De Silva AM. Epidemiology of dengue in Sri Lanka before and after the emergence of epidemic dengue hemorrhagic fever. Am J Trop Med Hyg 2002; 66: 765-773.

 

26.  Malavige GN,  Fernando S,  Fernando DJ, Seneviratne SL. Dengue viral infections.  Postgrad  Med  J  2004; 80; 588-601.

 

27.  Chia A, Luu CD, Mathur R, Cheng B, Chee SP. Electrophysiological findings in patients with dengue-related maculopathy. Arch Opthalmol 2006; 124: 1421-26.

 

28.  Anonymous. Dengue hemorrhagic fever, diagnosis, treatment and control. World health Organization. Geneva Switzerland, 1986.

 

29.  Ware LB, Conner ER, Matthay MA. von Willebrand factor antigen is an independent marker of poor outcome in patients with early acute lung injury. Crit Care Med 2001; 29: 2325-2331.

 

30.  Halstead SB, O'Rourke EJ.  Antibody enhanced dengue virus infection in primate leukocytes. Nature 1977;  265: 739-741.

 

31.  Gubler DJ, Reed D, Rosen L, Hitchcock JR Jr. Epidemiological, clinical and virological observation on Dengue in the Kingdom of Tonga. Am J Trop Med Hyg 1978; 27: 581-89.

32.  Seet RC, Lim EC, Wilder-Smith EP. Acute transverse myelitis following dengue virus infection. J Clin Virol  2006; 35: 310-2.

 

33.  Kamath SR,  Ranjit S. Clinical features, complications and atypical manifestation of children with severe forms of dengue haemorrhagic fever in South India. Indian J Paediatr 2006; 73: 889-95.

 

34.  Guzman MG, Kouri G, Valdes L, Bravo J, Vazquez  S, Halstead SB. enhanced severity of secondary dengue-2 infections: death rates in 1981 and 1997 Cuban outbreaks. Rev Panam Salud Publica 2002; 11: 223-7.

 

35.  Vaughn DW, Green S, Kalayanarooj S, Innis BL, Nimmannitya S,  et al. Dengue viremia titre, antibody response pattern and virus serotypes correlate with disease severity. J Infect Dis 2000; 181: 2-9.

 

36.  Watts DM, Porter KR, Putvatana P, Vasquez B, Calampa C, Hayes CG, Halstead SB. Failure of secondary infection with American genotype dengue 2 to cause dengue haemorrhagic fever. Lancet 1999; 354: 1431-4.

 

37.  Cologna R, Rico-Hesse R. American genotype structure decrease dengue virus output from human virus output from human monocytes and dentdritic cells. J Virol 2003; 77: 3929-38.

 

38.  Halstead SB.  Pathogenesis of dengue: challenges to molecular biology. Science 1988; 239: 476-481.

 

39.  Saito M, Oishi K, Inoue S, Dimanno EM, Alera MT,  Robles Am et al. association of increased platelet-associated immunoglobulins with thromobocytopenia and the severity of disease in secondary virus infection. Clin Exp Immunol 2004; 138: 299-303.

 

40.  Lin CF, Chiu SC, Hsiao YL, Wan SW, Lei HY, Shiau AL, Liu HS, Yeh TM, Chen SH, Liu CC, Lin YS. Expression of cytokine, chemokine, and adhesion molecules during endothelial cell activation induced by antibodies against dengue virus nonstructural protein 1. J Immunol 2005; 174: 395-403.

 

41.  Avirutnan P, Punyadee N, Noisakran S, Komoltri C, Thiemmeca S, Auethavornanan K, et al. Vascular leakage in severe dengue virus infections: a potential role for the nonstructural viral protein NS1 and complement.J Infect Dis 2006; 193: 1078-88.

 

42.  Bhamarapravati N. Haemostatic defects in dengue hemorrhagic fever. J Infect Dis 1989;  2 : S826-829.

43.  Jessie K, Fong MY, Devi S, Lam SK, Wong KT. Localization of dengue in naturally infected human tissues, by immunohistochemistry and in situ hybridization. J Infect Dis 2004 ; 189: 1411-18.

44.  Senanayake S. Dengue fever and dengue haemorrhagic fever. A diagnostic challenge. Aust Fam Phy 2006; 35: 609-12.

45.  Leggat PA.  Assessment of febrile illness in the returned traveller. Aust Fam Phys 2007; 38: 328-33.

 

46.  World Health Organisation. Dengue haemorrhagic fever. Diagnosis, treatment prevention and control, 2nd ed. World Health Organisation Geneva, Switzerland 1997.

 

47.  Shu PY, Huang JH. Current advances in dengue diagnosis. Clin Diagn Lab Immunol 2004; 11: 642-50.

 

48.  Kao CL, King CC, Chao DY, Wu HL, Chang GJ. Laboratory diagnosis of dengue virus infection: current and future prospective in clinical diagnosis and public health. J Microbiol Immunol Infect 2005; 38: 5-16.

 

49.  Thongcharoen P, Wasi C, Puthavathana P. Dengue viruses mono on dengue/dengue haemorrhagic fever. Ed., Prasert Thongcharoen. World Health Organisation, New Delhi, India 1993.

 

50.  Gubler DJ, Suharyono W, Sumarmo, Wulur H, Jahja E, Sulianti Saroso J.  Virological surveillance for dengue haemorrhagic fever in Indonesia using the mosquito inoculation technique. Bull World Health Organ 1979; 57: 931-6.

 

51.  Rosen L, Gubler D. The use of mosquitoes to detect and propagate dengue viruses. Am J Trop Med Hyg  1974; 23: 1153-60.

 

52.  Gubler DJ. Serological diagnosis of dengue/dengue haemorrhagic fever. Dengue Bull 1996; 20: 20-3.

 

53.  Innis BL, Nisalak A, Nimmannitya S, Kusalerdchariya S, Chongswasdi S, Suntayakorn S, Puttisri P, Hoke CH. An enzyme-linked immunosorbent assay to characterize dengue infections where dengue and Japanese encephalitis co-circulate. Am J Trop Med Hyg 1989; 40: 418-27.

54.  Vodam V, Kuno G. Laboratory diagnosis of dengue virus infections. In DJ Guber and G Kuno (ed). Dengue and dengue haemorrhagic fever, cab international, London, United Kingdom 1997: pp313-34.

55.  Shu PY, Chen LK, Chang SF, Yueh YY, Chow L, Chien LJ, Chin C, Lin TH, Huang JH. Comparison of capture immunoglobulin M (IgM) and IgG enzyme-linked immunosorbent assay (ELISA) and nonstructural protein NS1 serotype-specific IgG ELISA for differentiation of primary and secondary dengue virus infections. Clin Diagn Lab Immunol 2003; 10: 622-30.

 

56.  Jelinek T, Wastlhuber J, Pröll S, Schattenkirchner M, Löscher T Influence of rheumatoid factor on the specificity of a rapid immunochromatographic test for diagnosing dengue infection. Eur J Clin Microbiol Infect Dis.  2000; 19: 555-6.

 

57.  Ludolfs D, Schilling S, Altenschmidt J, Schmitz H. Serological differentiation of infections with dengue virus serotypes 1 to 4 by using recombinant antigens. J Clin Microbiol 2002; 40: 4317-20.

 

58.  De Paula S O,  da Fonseca BA. Dengue: A review of the laboratory tests a clinician must know to achieve a correct diagnosis. The Brazilian J Infect Dis 2004; 8: 390-8.

 

59.  World Health Organisation. Guidelines for treatment dengue fever/ Dengue haemorrhagic fever in small hospitals. Geneva: WHO; 1999.

 

60.  Singhi S, Kissoon N, Bansal A. Dengue and Dengue haemorrhagic fever: management issues in an intensive care unit. J Pediatr (Rio J) 2007; 83: s22-35.

 

61.  Chuansumrit A, Phimolthares V, Tardtong P, Tapaneya-Olarn C, Tapaneya-Olarn W, Kowsathit P, Chantarojsiri T. Transfusion requirements in patients with dengue haemorrhagic fever. Southeast Asian J Trop Med Public Health 2000; 31: 10-4.

 

62.  Graham, TP. Disorders of the circulation. In Fuhrman BP, Zimmerman JJ eds. Pediatric critical care 2nd edition, St Louis Mosby, 1998; 261-71.

 

63.  Ranjit S, Kissoon N, Jayakumar I.. Aggressive management of dengue shock syndrome may decrease mortality rate: a suggested protocol. Paediatr Crit Care Med 2005; 6: 412-9.

 

64.  Ooi EE, Goh KT, Gubler DJ. Dengue prevention and 35 years of vector control in Singapore. Emerg Infect Dis 2006; 12: 887-93.

 

65.  Monath T, McCarthy K, Bedford P, Johnson CT, Nicholas R, Yoksan S, et al. Clinical proof of principal for Chimerivax TM: recombinant live, attenuated vaccine against Flavivirus infections. Vaccine 2002; 20: 1004-18.