Dengue Virus: Dengue is a flu-like viral disease spread by the bite of Aedes infected mosquitoes. Dengue positive-strand RNA viruses of the genus Flavivirus, family FlaviviridaeDengue hemorrhagic fever is a severe, often fatal, and leads to several complications. Dengue occurs in most tropical areas of the world. There is no specific treatment for dengue. Dengue virus (DENV) infection can be divided into two phases: acute and convalescent.  The acute phase begins with the onset of symptoms and lasts approximately 5 days.  The convalescent phase begins once the fever subsides and lasts 4 to 7 days. Patients with dengue usually have symptoms when DENV is present in the blood.  Virus levels in the blood are usually higher during the first 5 days of the illness. Real Time RT-PCR assay detects DENV in a significantly high proportion of cases during this time, very early after the onset of symptoms of dengue.  This is important because an early laboratory confirmation of a dengue diagnosis helps physicians managing the case. Dengue fever is the most prevalent mosquito-borne viral disease in humans. Dengue fever is caused by 1 of 4 DV serotypes (dengue 1-4) Reinfection with a heterologous serotype of DV enhances the infection, resulting in the most severe clinical manifestations.

·         Dengue hemorrhagic fever (DHF)

·         Dengue shock syndrome (DSS)

Transmission occurs principally from the bite of an infected Aedes aegypti (and, less commonly, Ae. albopictus or Ae. polynesiensis) mosquito. Female mosquitoes acquire DENV by biting viremic humans and become infective after an extrinsic incubation period of 8–12 days. The infected mosquito can then transmit DENV for the rest of its life (the mosquito lifespan is approximately 1 month).Less common modes of DENV transmission include through exposure to DENV-infected blood, organs, or other tissues via blood transfusion, solid organ or bone marrow transplantation, and nosocomial injury (needlestick or mucous membrane contact with spilled blood). DENV can be vertically transmitted from an infected woman to her fetus in utero or to the infant during childbirth.

Prevention centers on avoiding mosquito bites in areas where dengue occurs or might occur and eliminating breeding sites. Dengue fever usually starts suddenly with a high fever, rash, severe headache, pain behind the eyes, and muscle and joint pain. The severity of the joint pain has given dengue the name “breakbone fever.” Nausea, vomiting, and loss of appetite are common.

 

 

Methodology: Taqman Real time PCR assay

Clinical Use:

  • RT-PCR method detects DENV very early after the onset of symptoms of dengue
  • he identification of dengue virus serotypes 1, 2, 3 or 4 from viral RNA in serum or plasma collected from human patients with dengue
  • Assess viral measured by changes in the Dengue virus RNA levels
  • To provide epidemiologic information for surveillance of circulating dengue viruses. Assess prognosis and early diagnosis for clinical management of patient and better cure
  • Confirm active Dengue virus infection In patient

 

 

 

Screening:

  • residing in areas where extended community outbreaks exist
  • international travelers to regions of the world where Dengue virus  is endemic
  • Vaccinated patient
  • During pregnancy
Performed:  Every day
Reported: 2-3 days

Specimen Required: Blood, serum, plasma, Collect in: Lavender (EDTA), pink (K2EDTA), or serum separator tube. Cerebrospinal fluid (CSF) Stability collection to initiation of testing On Cells: Ambient: 4 hours; after separation from cells: Refrigerated: 48 hours; Frozen at -20°C: 72 hours; Frozen at -70°C: 4 months. Do not thaw avoid repeated freezing and thawing.

NOTE: Collect CSF specimen in sterile screw capped bottles under all aseptic precautions for attempting isolation of virus minimum 0.5ml of CSF is required for acceptance of standard diagnosis.

Specimen Preparation: CSF, Separate serum or plasma from cells within 24 hours.

 

Storage/Transport Temperature: Frozen-20 0C. Refrigerate specimens at 2°C-4°C.

Unacceptable Conditions:  Heparinized specimens, Hemolysis sample, Quantity not sufficient for analysis, specimen grossly contaminated, specimen too old, frozen whole blood specimen, specimen leaky or tube broken.

 

Interpretation: This test can quantitate/detect Dengue Virus RNA Virus over the linear range 70-107 copies/mL. However this does not mean that lower copies or higher copies cannot be detected. The lower copies can be detected in some cases. This is a limitation of the currently available extraction systems. A negative result does not preclude the presence of Dengue Virus infection because results depend on adequate/proper patient sample storage and transportation as RNA is fragile and thermo labile, absence of inhibitors and sufficient RNA to be detected .Dengue Virus are a viral disease, and as such, antibiotics are of no value in the treatment of the infection. This assay detects DENV serotypes 1, 2, 3 or 4 from human serum or plasma collected from human patients with signs and symptoms consistent with dengue infection. The assay is intended to be used as a diagnostic test in patients and is not approved for blood bank screening. Real-Time RT-PCR Assay should not be performed unless the patient meets clinical and/or epidemiologic criteria for testing suspect dengue cases.

The result of this test must always be correlated with clinical status and history of the patient and other relevant data and should not be used alone for the interpretation.   

Note: The test is intended for use in conjunction with clinical presentation and other laboratory markers as an indicator of disease prognosis.

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