Dengue Virus (DEN1, DEN2, DEN3, DEN4)

Characteristics
Morphology	Dengue Virus is an arbovirus of the Flaviviridae family and Flavivirus genus. Dengue is an enveloped virus, 40-60 nm in size, with an isometric nucleocapsid of 25-30 nm and a ~10.7 kb, linear, positive-sense RNA genome. Dengue virus exists as four serotypes (Dengue 1–4) and is genetically related to other flaviviruses such as yellow fever and tick-borne encephalitis viruses.
Growth Conditions	Cell culture (both mammalian and mosquito cell lines)

Health Hazards
Host Range	Humans, Simians, and Mosquitoes
Modes of Transmission	Dengue viruses are spread to people through the bite of an infected Aedes species (Ae. aegypti or Ae. albopictus) mosquito or through contaminated blood transfusion. Peak mosquito biting times are 2-3 hours after dawn and several hours before dusk
Signs and Symptoms	1st Exposure: Sudden onset of fever for 3-5 days with an intense headache, myalgia, arthralgia, retro-orbital pain, anorexia, and rash. Symptoms of dengue typically last 2–7 days. Most people will recover after about a week. 2nd Exposure: Dengue hemorrhagic fever is characterized by abnormal vascular permeability, hypovolemia, and abnormal clotting mechanisms. Fatality rate is 40-50% and warning signs usually begin in the 24–48 hours after your fever has gone away.
Infectious Dose	Human LD50 is <10 PFU. Fewer than 10 PFU led to infection in 50% of volunteers treated with an attenuated Dengue virus vaccine candidate.
Incubation Period	Ranges from 3-15 days but is typically 4-7 days.

Medical Precautions/Treatment
Prophylaxis	None publically available
Vaccines	None publically available
Treatment	There is no specific medicine to treat dengue. Treat the symptoms by taking non-aspirin pain relievers and drink plenty of water. Seek medical attention if they become severe.
Surveillance	Monitor for symptoms. Real-time PCR assay can be used to quantitatively measure RNA number as an indicator of viral load in infection. Induced seroconversion can be detected using ELISA
GWU Requirements	Report all incidents to the Office of Risk Management as well as the Office of Research Safety (ORS) IMMEDIATELY FOLLOWING THE OCCURRENCE.

Containment
BSL-2	Risk Group 2 classification is applied to dengue. Containment Level 2 facilities, equipment, and operational practices for work involving infectious or potentially infectious material. All procedures that may produce aerosols, or involve high concentrations or large volumes should be conducted in a biological safety cabinet (BSC). The use of needles, syringes, and other sharp objects should be strictly limited. Additional precautions should be considered with work involving animals or large scale activities. Enhanced containment practices should be taken when working with samples obtained from patients (due to presence of etiologic agents).
ACL-2	Specifically for in vivo work involving infectious mosquitoes

Containment

BSL-2

Risk Group 2 classification is applied to dengue. Containment Level 2 facilities, equipment, and operational practices for work involving infectious or potentially infectious material. All procedures that may produce aerosols, or involve high concentrations or large volumes should be conducted in a biological safety cabinet (BSC). The use of needles, syringes, and other sharp objects should be strictly limited. Additional precautions should be considered with work involving animals or large scale activities. Enhanced containment practices should be taken when working with samples obtained from patients (due to presence of etiologic agents).

ACL-2

Specifically for in vivo work involving infectious mosquitoes

Spill Procedures
Small Spills (<1 liter)	If the spill occurred inside a biological safety cabinet, close the sash and allow the cabinet to operate for 15 minutes before continuing with the spill cleanup. While dengue exposure via aerosol transmission is uncommon, extra caution should still be taken during a spill outside of a BSC. Leave the room immediately and allow the aerosols to dissipate for 15 minutes. Notify others working in the lab. Don appropriate PPE. Cover area of the spill with paper towels or any absorbent material and apply an EPA registered disinfectant effective against enveloped viruses (alcohols, bleach, and quaternary ammonium compounds), working from the perimeter towards the center. Allow 30 minutes of contact time before disposal and cleanup of spill materials.
Large Spills	Alert lab personnel in the laboratory to the spill and keep people out of the area to prevent spread of the contamination. Check if you have been contaminated or if any of your PPE has been breached. If so follow exposure procedures. Remove any contaminated clothing and place it the biohazard waste. Wash your hands and post a sign on the door. Notify your supervisor of the incident and call ORS (4-8258) for assistance. If the situation involves an imminently life-threatening injury or has catastrophic potential, call 911.

Spill Procedures

Small Spills (<1 liter)

If the spill occurred inside a biological safety cabinet, close the sash and allow the cabinet to operate for 15 minutes before continuing with the spill cleanup.

While dengue exposure via aerosol transmission is uncommon, extra caution should still be taken during a spill outside of a BSC. Leave the room immediately and allow the aerosols to dissipate for 15 minutes. Notify others working in the lab. Don appropriate PPE. Cover area of the spill with paper towels or any absorbent material and apply an EPA registered disinfectant effective against enveloped viruses (alcohols, bleach, and quaternary ammonium compounds), working from the perimeter towards the center. Allow 30 minutes of contact time before disposal and cleanup of spill materials.

Large Spills

Alert lab personnel in the laboratory to the spill and keep people out of the area to prevent spread of the contamination. Check if you have been contaminated or if any of your PPE has been breached. If so follow exposure procedures. Remove any contaminated clothing and place it the biohazard waste. Wash your hands and post a sign on the door. Notify your supervisor of the incident and call ORS (4-8258) for assistance. If the situation involves an imminently life-threatening injury or has catastrophic potential, call 911.

Exposure Procedures
Mucus Membrane	Flush eyes, mouth or nose for 15 minutes at eyewash station.
Other Exposure	Wash area with soap and water for 15 minutes.
Reporting	Report ALL injuries to the PI immediately and reported to the Office of Risk Management at [email protected] IMMEDIATELY FOLLOWING THE OCCURRENCE. Exposures that involve a bloodborne-pathogen or recombinant DNA also need to be reported to the Office of Research Safety at [email protected]. If the injury requires immediate medical attention, call GWPD at 202-994-6111 or call 911.
Medical Monitoring	Seek immediate medical evaluation, treatment, and post exposure follow-up at the Employee Health Office at GWU Hospital (900 23rd St., NW, Suite G-1090, Phone: 202-715-4275). Students should go to the Students Health Office at Marvin Center. After hours treatment can be received at the GWU hospital emergency room.

Stability
Disinfection	Susceptible to 70% Ethanol, 10% bleach, and 2% glutaraldehyde. Quaternary ammonium compounds can be used as well
Inactivation	Viruses are sensitive to moist heat (121°C for at least 15 min), dry heat (160-170°C for at least 1 hour), low temperature sterilization (i.e. Ethylene oxide or plasma sterilization), and low pH
Survival Outside Host	The virus is stable in dried blood for up to 9 weeks at room temperature

Personal Protective Equipment (PPE)
Minimum PPE Requirements	At minimum, personnel are required to don gloves, closed toed shoes, lab coat, and appropriate face and eye protection prior to working with dengue. Additional PPE may be required depending on lab specific SOPs.
Additional Precautions	Additional protection may be worn over laboratory clothing when infectious materials are directly handled, such as solid-front gowns with tight fitting wrists, gloves, and respiratory protection.

References
CDC (https://www.cdc.gov/dengue/) Infection of Human Cells by Dengue Virus Is Modulated by Different Cell Types and Viral Strains (https://journals.asm.org/doi/10.1128/JVI.74.17.7814-7823.2000) Canadian PHAC (https://www.canada.ca/en/public-health/services/laboratory-biosafety-bi…)