DDEENNGGUUEE

Introduction

Dengue fever is an acute febrile infectious disease, caused by all four serotypes (DEN 1, 2, 3 or 4) of a virus from genus Flavivirus, called dengue virus. It’s the most prevalent flavivirus infection of humans, with a worldwide distribution in the tropics and warm areas of the temperate zone corresponding to that of the principal vector, Aedes aegypti (A. aegypti) When simultaneous

or sequential introduction of two or more serotypes occurs in the same area, there may be an increased number of cases with worse clinical presentation (dengue hemorrhagic fever). The term ‘hemorrhagic’ is imprecise, because what characterizes this form of the disease is not the presence of hemorrhagic manifestations, but the abrupt increase of capillary permeability, with diffuse capillary leakage of plasma, hemoconcentration and, in some cases, with non-hemorrhagic hypovolemic shock (dengue shock syndrome).

Epidemiology

The highest incidence of dengue is in Southeast Asia, India and the American tropics, where A. aegypti can be found. In the 1980s, dengue emerged in explosive epidemics in Rio de Janeiro (1986 - serotype 1 and 1990 - serotype 2 was isolated in Niterói city), São Paulo and in many other towns and cities in Brazil. In areas such as southeast of Asia, where all four dengue virus types are hyper endemic, children are almost exclusively affected, and seroprevalence approaches 100% by young adulthood.

Transmission occurs by the bite of Aedes aegypti female mosquitoes - the same vector of urban yellow fever - a day-active species with low fly-autonomy that is abundant in and around human habitations. In Brazil and other countries Aedes albopictus may also be responsible for transmission. Viremic humans (till the fifth day of disease) serve as the source of virus for mosquito infection; there is no person-to-person transmission. Movement of viremic humans provides the principal means of spread, and rapid air travel is a factor in most recent epidemic emergences.

Although homotypic immunity (immunity to a particular strain of the virus) is complete and lifelong, cross-protection between virus of the same group is incomplete and transient. As mentioned before, there is a strict association between reinfection by

another serotype and the occurrence of dengue hemorrhagic fever (DHF), due to immunopathologic processes, such as immune enhancement and immune clearance. Outbreaks usually occur in summer, when ambiental conditions are ideal for vectors’ proliferation.

Problem statement:

• Since last 20 years, there is global increase in frequency of Dengue infection.

• 2.5 – 3.0 billion people are currently at risk worldwide. • Worldwide 50-100 million cases are being reported annually. • Each year approximately 500,000 cases of Dengue

hemorrhagic fever (DHF) require hospitalization, including 90% children less than 15 years of age. Mortality of Dengue hemorrhagic fever is approximately 5% with 25,000 deaths reported each year.

• Usually it is considered an urban disease but now spreading its tentacles in rural areas also.

Indian perspective:

India, Sri Lanka and Myanmar are considered hyper endemic for Dengue fever because of significant increase in the proportions of dengue cases with severe disease. According to WHO, in India, Dengue Hemorrhagic fever is an emergent disease in view of frequent cyclical epidemics and circulation of multiple virus serotypes at present.

Transmission of Dengue

Dengue viruses are transmitted by humans through the bite of infected Ades mosquito, principally Ades aegypti, and are therefore considered to be arboviruses (arthropod – borne

viruses). Once infected the mosquito remains infected for life, transmitting the virus to susceptible individuals during probing and feeding. Humans are the main amplifying host of the virus. The virus circulates in the blood of the infected humans at approximately the time that they have fever, and the uninfected mosquito may acquire the virus if they feed on an individual when he or she is viraemic. The virus then develops in the mosquito for a period of 8- 10 days before it can be transmitted to the other humans during subsequent probing or feeding. The length of time required for this extrinsic incubation depends in part on environmental conditions, especially ambient temperatures.

The Virus

Dengue belongs to the family Flaviviridae. The four serotypes of dengue virus (i.e. DEN1, DEN2, DEN3 & DEN4) can be distinguished by serological methods. Infection in humans by one serotype produces life long immunity against reinfection, by that same serotype, but only temporary and partial protection against the other serotypes. Dengue viruses share many characteristics with the other flaviviruses, having a single- stranded RNA genome surrounded by icosahedral Nucleocapsid and covered by a lipid envelope. The virion is approximately 50nm in diameter.

The Vector

Ades aegypti is one of the most efficient mosquito vectors for arboviruses, because it is highly anthropophilic and thrives in close proximity to humans and often lives indoors. Dengue outbreaks have also been attributed to A. albopiticus, A. polyneniensis, and several species of the A. scutellaris complex. A factor complicating the eradication of the vector is that A. aegypti eggs can withstand long periods of desiccation, sometimes for more than a year.

The Host

Man is probably the main reservoir of the disease. There is transovarial transmission of dengue virus in the mosquito and this can be repeated for a few generations without the adults feeding on an infected host.

Clinical Manifestations

Dengue virus infections may be asymptomatic or may lead to undifferentiated fever, dengue fever (DF) or dengue haemorrhagic fever (DHF) with plasma leakage that may lead to hypovalaemic shock. (Dengue shock syndrome, DSS) Dengue Fever: - The Clinical features of DF frequently depend on the age of the patient. Infants and young children may have an undifferentiated febrile disease, often with a rash (accompanied with discoloration of skin). Older children and adults may have either mild fibril syndrome or the classic incapacitating disease with high fever of abrupt onset, severe headache, pain behind the eyes, muscle and bone joint pains, nausea and vomiting, and rash. Leucopenia (loss in number of leucocytes) is usually seen. In some epidemics, DF may be accompanied by bleeding complications. Though case fatality in DF, however, is less than 1%.

Dengue Haemorrhagic fever:- Typical cases of DHF are characterized by four major clinical manifestations: high fever, haemorrhagic phenomena, and often, hepatomelagy and circulatory failure. Moderate to marked thrombocytopenia with concurrent haemoconcentration is a distinctive clinical finding of DHF. The major pathophysiological change that determines the severity of disease in DHF- and differentiates it from DF- is the leakage of plasma, as manifested by an elevated haematocrit (haemoconcentration), a serous effusion.

(Picture exhibiting DHF symptoms.)

The most common Haemorrhagic phenomenon is a positive TOURNIQUET TEST, easy bruising and bleeding at venepuncture site. The severity of the disease can be modified by the early diagnosis and replacement of plasma loss. Thrombocytopenia and haemoconcentration are usually detectable before the subsidence of fever and onset of shock. Dengue Shock Syndrome:- The conditions of the patients who progress to the shock deteriorate after a fever of 2-7 days duration. There are the typical signs of circulatory failure: the skin becomes cool, blotchy, and congested; circumoral cyanosis is frequently observed; the pulse becomes rapid. Patients may initially be lethargic then becomes restless and rapidly enter a critical stage of shock. If not attended to the patient may pass into stage of profound shock, with the blood pressure or pulse becoming imperceptible. The duration of shock is short; typically the patient dies within 12-24 hours, or recovers rapidly following apt. volume replacement therapy. DDHHFF rreesseemmbblleess DDFF dduurriinngg iinniittiiaall pphhaassee ooff iillllnneessss.. HHoowweevveerr,, tthhrroommbbooccyyttooppeenniiaa aaccccoommppaanniieedd bbyy // ffoolllloowweedd bbyy aa rriissiinngg hhaaeemmaattooccrriitt ddiissttiinngguuiisshheess DDHHFF ffrroomm DDFF aanndd ootthheerr ddiisseeaasseess.. IItt iiss nnoott cclleeaarrllyy kknnoowwnn wwiitthh cceerrttaaiinnttyy wwhhyy ddooeess DDHHFF ooccccuurr iinn ssoommee ccaasseess wwhheenn tthhee mmoosstt hhaavvee oonnllyy ddeenngguuee ffeevveerr tthhoouugghh iitt iiss ssuuggggeesstteedd tthhaatt wwhheenn tthheerree aarree cciirrccuullaattiinngg aannttiibbooddiieess aaggaaiinnsstt oonnee sseerroottyyppee dduuee ttoo eeaarrlliieerr iinnffeeccttiioonn aanndd llaatteerr tthheerree iiss iinnffeeccttiioonn wwiitthh aannootthheerr sseerroottyyppee,, iitt mmaayy rreessuulltt iinn ddeenngguuee hhaaeemmoorrrrhhaaggiicc ffeevveerr // ddeenngguuee sshhoocckk ssyynnddrroommee.. TThhee rriisskk ooff DDHHFF iiss 00..22%% dduurriinngg

tthhee ffiirrsstt ddeenngguuee iinnffeeccttiioonn bbuutt iitt iinnccrreeaasseess 1100--ffoolldd dduurriinngg iinnffeeccttiioonn wwiitthh aa sseeccoonndd sseerroottyyppee ooff ddeenngguuee vviirruuss.. HHoowweevveerr,, tthhee cciirrccuullaattiioonn ooff mmuullttiippllee ddeenngguuee sseerroottyyppeess ddooeess nnoott aallwwaayyss pprroodduuccee DDHHFF..

Transmission Cycle

• Cycle begins with a dengue-infected person

• This person will have virus circulating in the blood—a

viremia that lasts for about five days.

• During the viremic period, an uninfected female Aedes aegypti mosquito bites the person and ingests blood that contains dengue virus

• Then, within the mosquito, the virus replicates during an

extrinsic incubation period of eight to twelve days.

• The mosquito then bites a susceptible person and transmits the virus to him or her, as well as to every other susceptible person the mosquito bites for the rest of its lifetime

• The virus then replicates in the second person and produces

symptoms. The symptoms begin to appear an average of four to seven days after the mosquito bite—this is the intrinsic incubation period, within humans

• While the intrinsic incubation period averages from four to

seven days.

• The viremia begins slightly before the onset of symptoms

• Symptoms caused by dengue infection may last three to 10 days, with an average of five days, after the onset of symptoms—so the illness persists several days after the viremia has ended.

Serology:-

• In primary infection, circulating IgM antibody to the viral coat proteins is detected 5-6 days after the onset of illness, and gradually decreases within 1-2 months of onset. IgG antibody to dengue virus is detected approximately 14 days after onset in primary infections, and by day 2 in secondary infections.

• In secondary infection, IgM antibody may reappear but gradually diminishes, while IgG antibody persists, often at high titer.

• These patterns of dengue antibody development permit serological differentiation of primary and secondary infections.

• Characteristically, acute patients with primary infections have a higher IgM:IgG ratio than are found in secondary infections. Patients with secondary infections generally have higher IgG levels.

• Acute or recent infections are identified by a rise in antibody titer as well as high IgM levels.

Diagnosis:-

Clinical Diagnosis:

• Pancythemia may occur after 3-4 days of illness, characterized by leucopoenia lesser than 2000, mild acidosis, and hypoproteinemia.

• ECG changes may include bradycardia, flattened T waves, and prolongation of the P-R interval.

• Additional or common hematologic changes in DHF and DSS include: haemconcentrations of greater than 20% in hematocrit, thrombocytopenia, prolonged bleeding time; and decreased PT seldom below than 40% of the control.

• Chest x-ray may reveal pleural effusions (greater on the left side as compared to right).

• Elevated hepatic transaminase levels have been reported in over 80% of dengue cases.

• The tourniquet test is frequently positive. A tourniquet test is used to assess if capillaries can withstand increased pressure, which is important to assess as the vasculopathy that leads to DSS and DHF, which is often first, visualized in the form of petechiae.

The tourniquet test is assessed by inflating a blood pressure cuff on an arm to above venous pressure (70 mm Hg) for 5 minutes and then releases the pressure. Then extremity for petechiae is observed. If there are more than 2 petechiae then the test is positive. This test does not have high specificity.

Laboratory Diagnosis

1. Virus Isolation

Mosquito inoculation is the most sensitive method for dengue virus isolation. Isolation rates of up to 100% of

serologically confirmed dengue infections are not uncommon, and this is the only method sensitive enough for routine successful virologic confirmation of fatal DHF and DSS cases. Moreover, there are many endemic dengue virus strains that can be recovered only by this method.

Virus detection in the mosquito is generally performed by the direct fluorescent-antibody DFA test.

The mosquito inoculation technique has the disadvantages of being labor-intensive and requiring an insectaries to produce large numbers of mosquitoes for inoculation. Also, unless strict safety precautions are maintained, the chance of laboratory infections

increases.

2. Serological Tests:

Haemagglutination Inhibition Test:

If a virus has a surface protein that can attach to the RBC, it can agglutinate RBCs. If the concentration of virus in a sample is high, when the sample is mixed with RBCs, a lattice of viruses and RBCs will be formed. This phenomenon is called hemagglutination.

If antibodies against the hemagglutinins are present, hemagglutination will be prevented. During the

hemagglutination inhibition test, serial dilutions of serum are mixed with a known amount of virus. After incubation, RBCs are added, and the mixture is left to sit for several hours. If hemagglutination is inhibited, a pellet of RBCs forms at the bottom of the tube. If hemagglutination is not inhibited, a thin film is formed.

Easy, simple and ideal for seroepidemiological studies. Haemagglutination inhibitor antibodies appear by day 5-6 of illness but this test lacks specificity.

• Complement fixation Test:

The test consists of two antigen-antibody reactions, one of which is the indicator system. The first reaction, between a known virus antigen and a specific antibody takes place in the presence of a predetermined amount of complement. The complement is removed or "fixed" by the antigen-antibody complex. The second antigen-antibody reaction consists of reacting sheep RBc with haemolysin. When this indicator system is added to the reactants, the sensitized RBC’s will only lyse in the presence of free complement.

Difficult to perform. Not specific for secondary infections.

• Neutralization Test:

Neutralization of a virus is defined as the loss of infectivity through reaction of the virus with specific antibody. Virus and serum are mixed under appropriate condition and then inoculated into cell culture. The presence of unneutralized virus may be detected by reactions such as haemagglutination. The loss of infectivity is bought about by interference by the bound Ab’s of any one of the steps leading to the release of the viral genome into the host cells.

Most specific and sensitive but technically difficult and time consuming.

ELISAS

• IgM-capture enzyme -linked immunosorbent assay (MAC-ELISA):

MAC-ELISA has become the most widely used serologic test for dengue diagnosis in the past few years. Anti-dengue IgM antibody develops a little faster than IgG antibody.

• Nearly all patients tested between 10 and 20 days had detectable IgM antibody.

• IgM antibody titers in primary infections are significantly higher than in secondary infections.

• Widely used, simple and rapidly defects Dengue specific IgM antibodies by capturing them out of solution using antihuman IgM previously bound to the solid phase.

Indirect IgG-ELISA

This Elisa can be used to differentiate primary and secondary dengue infections. The test is simple and easy to perform and is thus useful for high-volume testing.

Can be used to differentiate primary and secondary Dengue infection. But non-specific and cross-reacts with other flaviviruses. Does not give serotype diagnosis.

RT-PCR:

Useful diagnostic test for confirmation of Dengue virus infection (primary or secondary).

Rapids: Easy to use rapids are another option in the detection of Dengue infection. Such as our -------à

DDIIAAGGNNOOSS DDEENNGGUUEE CCAARRDD.. INTENDED USE DIAGNOS DENGUE CARD is a rapid solid phase immuno-chromatographic assay for the qualitative and differential detection of IgG and IgM antibodies to dengue virus in human serum / plasma. This test is for in vitro diagnostic use only and is intended as an aid in the presumptive diagnosis between primary and secondary dengue infection.

PRINCIPLE

Serum / plasma samples may be used with this test. When a specimen is added to the test, IgG and IgM antibodies in the specimen sample react with red particles coated with dengue proteins. As this specimen/ particle mixture migrates along the length of the test, the anti-dengue IgG or IgM antibody particle complex is captured by the relevant IgG and /or IgM test bands located in the test device window causing a pale to dark red band to form at the IgG or IgM region of the test device window. The intensity of the bands will vary depending upon the amount of antibody present in the sample. The appearance of any colour in a specific test region (IgG or IgM) should be considered as positive for that particular antibody

SALIENT FEATURES

• High Specificity and sensitivity. • No equipments required. • Easy to read colour bands. • This tests provides for the detection of both Primary and Secondary infections in Dengue.

• See- through devices for easy interpretation.

• Built in control line. • Results within 15 minutes. • Shelf life 12 months at 2-8 0C. • Available is convenient pack sizes.

Treatment There is no specific treatment for dengue fever. However, careful clinical management by experienced physicians and nurses frequently saves the lives of DHF patients. With appropriate intensive supportive therapy, mortality may be reduced to less than 1%. Maintenance of the circulating fluid volume is the central feature of DHF case management.

Immunization Vaccine development for dengue and DHF is difficult because any of four different viruses may cause disease, and because protection against only one or two dengue viruses could actually

increase the risk of more serious disease. Nonetheless, progress is being made in the development of vaccines that may protect against all four dengue viruses. Such products may become available for public health use within several years.

Prevention and control At present, the only method of controlling or preventing dengue and DHF is to combat the vector mosquitoes. The rapid geographic spread of this species has been largely attributed to the international trade in used tyres. Vector control is implemented using environmental management and chemical methods. Proper solid waste disposal and improved water storage practices, including covering containers to prevent access by egg laying female mosquitoes are among methods that are encouraged through community-based programmes. The application of appropriate insecticides to larval habitats, particularly those which are considered useful by the householders, e.g. water storage vessels, prevent mosquito breeding for several weeks but must be re-applied periodically. Small, mosquito-eating fish and copepods (tiny crustaceans) have also been used with some. Regular monitoring of the vectors' susceptibility to the most widely used insecticides is necessary to ensure the appropriate choice of chemicals. Active monitoring and surveillance of the natural mosquito population should accompany control efforts in order to determine the impact of the program.

Glossary Ø Hemorrhagic:- Relating or belonging to the escape of blood from any part of the vascular system. Ø Hypovolemic:- Conditions In which the volume

of the blood in the body is diminished. Ø Haemoconcentration/ Haemocrit:- Formation of a

clump of blood corpuscles. Ø Viraemic:- A virus in the blood. Ø Febrile:- Feverish; having the character of or

belonging to a fever; characterized by the presence of fever.

Ø Leucopenia:- Decrease in the number of leucocytes. Ø Thrombocytopenia:- thrombosis produced by an

abnormal accumulation or agglutination of Blood platelets.

Ø Maculpapular Rash:- A spot which has the characteristics between those of a macula (Change in colour of skin without change in any other property of the skin, Also k/as Petechiae) and a papule (pimple).

Ø Hepatomelagy:- Enlargement of Liver. Ø Hypoproteinaemia:- Decrease in the protein content

in blood. Ø Circumoral Cyanosis:- Blue appearance of the skin

and mucous membrane around the mouth. Ø Blotchy:- Discolored/ pustule/ blister. Ø Pancythemia/ pancytopenia:- Decrease in all

elements of blood (viz. RBC’s, WBC’s & Platelets). Ø Haemadsorption:- Adherence of RBC’s to the surface

of something (as a virus or cell).

Ø Bradycardia:- Relatively slow heart action whether physiological or pathological.

Ø Transovarial:- Transmission within the virus and it’s progeny.

Reference:-

Ø Centers for Disease Control and Prevention. 1600 Clifton Rd., NE, Atlanta, GA 30333. (800) 311-3435, (404) 639-3311. http://www.cdc.gov.

Ø Farley, Dixie. "Treating Tropical Diseases." FDA Consumer (Jan./Feb. 1997): 26+.

Ø Carlini, M.E. & Shandera, W.X. (2001). Infectious diseases: Viral and rickettsial. In L.M. Tierney, S.J. McPhee, & M.A. Papadakis (Eds.), Current Medical Diagnosis & Treatment (40th ed.) (pp.1330-1331). Stamford Connecticut: Appleton & Lange.

Ø Chin, J. (Ed.) (2000). Control of communicable diseases manual (17th ed.). Washington, DC: American Public Health Association.

Ø Chuansumrit,A., Phimolthares, V., Tardtong, P., Tapaneya-Olam, C., Tapaneya-Olarn, W., Kowsathit, P., & Chantarojsiri, T. (2000). Transfusion Requirements in Patients with Dengue Hemorragic Fever. The Southern Asian Journal of Tropical Medicine and Public Health, 31(1), 10-14.

Ø Gubler, D.J. (1998). Dengue and dengue hemorrhagic fever. Clinical Microbial Review,11(3), 480-496.

Ø World Health Organization (1998). The World Report, 1998. Geneva: World Health Organization.

Ø WHO Dengue and dengue haemorrhagic fever.htm Ø ANGEL BALMASEDA, SAMANTHA N. HAMMOND, LEONEL PÉREZ,

YOLANDA TELLEZ, SAIRA INDIRA SABORÍO, JUAN CARLOS MERCADO, RICARDO CUADRA, JULIO ROCHA, MARIA ANGELES PÉREZ, SHEYLA SILVA, CRISANTA ROCHA, AND EVA HARRIS*.

“SEROTYPE-SPECIFIC DIFFERENCES IN CLINICAL MANIFESTATIONS OF DENGUE”.

Ø Antoine Talarmin, Bhety Labeau, Josiane Lelarge, and Jean-Louis Sarthou. “Immunoglobulin A-Specific Capture Enzyme-Linked Immunosorbent Assay for Diagnosis of Dengue Fever.”

Ø Zhonghua Min Guo Wei Sheng Wu Ji Mian Yi Xue Za Zhi. 1989 Nov;22(4):278-85 MAC-ELISA for the detection of IgM antibodies to dengue type I virus (rapid diagnosis of dengue type I virus infection).

This Manual is meant for Internal circulation only.