The Agent

Aedes aegypti mosquitoes after a blood meal (320x240)
credit: Rico-Hesse LabAedes aegypti mosquitoes after a blood meal

The dengue viruses, a group of four closely related viruses, are transmitted to humans through the bite of an infected mosquito. They cause a disease known as dengue fever, but they can also cause a more severe form of illness known as dengue hemorrhagic fever (or severe dengue), which potentially can be fatal. The incidence of both forms of the disease has been increasing dramatically in recent years.

Dengue fever is characterized by high fever, severe headache, pain behind the eyes, joint and bone pain, rash, and mild bleeding. In addition to these symptoms, the more severe form of the disease can lead to breathing difficulty, severe bleeding, shock, and possibly death. However, when recognized early and treated by maintaining body fluid volumes, mortality rates can be reduced to less than one percent.

Dengue infection occurs predominantly in tropical and subtropical climates around the world (see map), mostly in urban and semi-urban areas, and usually during the rainy season – the time when these regions are inhabited by large numbers of the particular type of mosquitoes that carry the virus.

The mosquito Aedes aegypti is the primary vector, or transmitter, of dengue virus. This is the same type of mosquito that transmits Zika virus. It is originally from Africa but now lives in urban habitats in tropical and subtropical regions worldwide. It breeds mostly in man-made containers that hold water, such as tires and flower pots. Another mosquito vector, Aedes albopictus, has spread from Asia to North America and Europe and was responsible for a dengue outbreak in Hawaii in 2001. The Aedes mosquitoes are common in the Gulf coast and mid-Atlantic regions of the United States and in over 100 countries around the world, including Europe.

The incidence of dengue infection in the United States is low, but it is on the rise. Most people in the United States who acquire a dengue virus infection do so while traveling abroad to an area where dengue viruses - and the mosquitoes that harbor them - are common. However, since 1980, locally acquired cases have been reported in Texas and more recently, in southern Florida.

A person becomes infected with dengue virus as a result of a bite from an infected female Aedes mosquito. Symptoms usually begin three to five days following the mosquito bite. Mosquitoes acquire dengue virus through a blood meal when they bite a dengue-infected human during the seven-day period when large amounts of virus are present in the blood. The infection of the mosquito can occur even if the person does not have symptoms of infection. The mosquito will then remain infected and capable of transmitting dengue virus to other people for the remainder of its life (up to a few weeks). If there is sufficient contact between Aedes mosquitoes and dengue-infected people in a particular area, then transmission of dengue is sustained.

There are four types of dengue virus - DENV1, DENV2, DENV3, and DENV4. Dengue remained a relatively minor and geographically restricted disease until World War II, when Aedes mosquitoes harboring the viruses are thought to have been transported around the world in cargo, and when returning soldiers carried the virus in their blood.

Once infected with one type of dengue virus, a person will become immune to infection by that particular dengue virus. However, infection with one dengue virus does not provide protection against the three other dengue viruses, and in fact sequential infections by different dengue virus types can lead to an increased risk of developing a more severe form of the disease.

Dengue viruses belong to a family of viruses known as Flaviviruses. Flaviviruses are enveloped viruses that contain genomes which consist of nonsegmented single-stranded positive-sense RNA. Viruses in this family, which are commonly transmitted by mosquitoes or ticks, include yellow fever virus (for which the family is named; flavus means yellow in Latin), Zika virus, West Nile virus, and hepatitis C virus.

The Problem

Dengue is the most common vector-borne viral disease in the world, causing an estimated 50 to 100 million infections globally each year and 25,000 deaths. In the tropics and subtropics, it is a leading cause of illness and death.

The incidence of dengue has jumped sharply in recent decades as the disease has spread into new geographic regions. Whereas before 1970, only nine countries experienced dengue epidemics, the viruses are currently transmitted in more than 100 countries. Over 2.5 billion people - or over 40% of the world’s population - are now at risk from dengue.

Not only are dengue epidemics occurring with greater frequency, the number of cases of the severe form of the disease, dengue hemorrhagic fever, is also increasing. In 2010, 1.6 million cases of dengue were reported in the Americas alone, with DHF accounting for nearly 50,000 of these cases. Although dengue has not been widespread in the United States, there is a threat that dengue could become continuously transmitted in the United States, since the mosquitoes occur throughout the southeast. There has also been a steady increase in the number of dengue infections in travelers returning to the United States from abroad.

The emergence of dengue can be attributed to increased urbanization and travel. The growth of urban centers provides a greater number of manmade containers, which are potential breeding sites for the Aedes mosquitoes, and a higher concentration of people in proximity to the mosquitoes. Air travel allows the disease to spread rapidly and into new geographic regions, especially in instances where people return home from popular tourist destinations where the disease is endemic, such as Latin America and Southeast Asia.

There is no specific treatment for dengue fever, nor is there a vaccine to protect against infection. Preventing and controlling outbreaks is very challenging, because it is impossible to eradicate the Aedes mosquitoes. They are very adaptable and resilient to environmental changes and human intervention strategies.

The Research

Structure and transmission of dengue virus

Dr. Rebeca Rico-Hesse’s laboratory is studying the structural (RNA and/or protein) and transmission (replication in Aedes aegypti) characteristics of dengue type 2 and type 3 virus variants that have produced dengue hemorrhagic fever in humans throughout the world.

The high mutability of RNA viruses (such as Flaviviruses) leads to a wide range of their genetic variants being transmitted in nature. Using rapid nucleotide sequencing techniques, they have obtained phylogenetic or family trees of the genetic relationships of many different viruses. These trees can then be interpreted to: (1) detect virus transmission pathways during epidemics and around the world, (2) determine which virus variants may be attenuated or less prone to producing disease, or conversely, which may be more virulent or cause more epidemics, (3) pick the best genetic and/or antigenic representative, to serve as a vaccine strain, and (4) detect and identify emerging viruses, to pinpoint their geographic origin and determine the best way to interrupt their transmission. For example, these studies have shown that the dengue viruses that are currently being transmitted on the border with Mexico (2005) are the most virulent type, having been imported from Southeast Asia a long time ago. These viruses displaced the less virulent variants that existed on this continent until 1995.

Dr. Rico-Hesse and her group use molecular clones of some of the viruses to determine which genetic/structural components are involved in pathogenesis and increased transmission, so that they can design new vaccines to control these exotic diseases at their origin.

Humanized mouse model for studying dengue virus

In a separate line of research, Rico-Hesse and colleagues have worked to overcome one of the main limitations for studying dengue fever, which is that dengue virus only causes the disease in humans, so no other animals can be used as models of the condition to develop preventive and therapeutic measures.

To overcome this challenge, they have been working with a mouse model of the human immune system, or ‘humanized mice’ which were developed by other research groups from mice naturally born without their own immune system. These severely immunodeficient mice received human stem cells that gave rise to many of the components of the human immune system, creating a living humanized animal model in which Rico-Hesse and her colleagues could study factors that may affect the development of dengue fever.

In 2012 they found that in these humanized mice, disease development was significantly different depending on whether the dengue virus was delivered through a mosquito bite or through a needle injection. The mice subjected to mosquito-bite delivery of the virus developed a more human-like disease with more of a rash, more fever and other characteristics that mimic the disease presentation in humans than did the needle-injection mice.

These observations led to the idea that mosquitoes are not just acting like ‘syringes,’ merely injecting viruses into the animals they feed on, but that their saliva seems to contribute significantly to the development of the disease. 

To test the effect of virus-free mosquito saliva on the immune response of humanized mice, the researchers held a vial containing mosquitoes against a footpad of anesthetized humanized mice, allowing a total of four mosquitoes to feed on both footpads. The researchers then took blood and a number of other tissue samples six hours, 24 hours and seven days after the mosquitoes bit the mice, and determined the levels of cytokines, as well as the number and activity of different types of immune cells using highly sensitive techniques. They compared these results with those obtained from humanized mice that had not been bitten by mosquitoes.

To their surprise, they found that mosquito-delivered saliva induced a varied and complex immune response. Both the immune cell responses and the cytokine levels were affected. They saw activation of T helper cells 1, which generally contribute to antiviral immunity, as well as activation of T helper cells 2, which have been linked to allergic responses. At various time points, the levels and activities of other types of immune cells also increased as others decreased.

Overall, the researchers found evidence that mosquito saliva alone can trigger long-lasting immune responses – up to seven days post-bite – in multiple tissue types, including blood, skin and bone marrow.

The researchers will continue this study by investigating which of the more than 100 proteins in mosquito saliva are mediating the effects on the immune system, or may help the virus become more infectious. Identifying these proteins could help in the design of strategies to fight transmission of dengue fever, as well as other viral diseases transmitted by Aedes aegypti, such as Zika virus and chikungunya virus.