Generation of Wild-Type Dengue Viruses for Use in Rhesus Monkey Infection Studies

Dengue virus is a positive-sense RNA virus belonging to the Flavivirus genus of the family Flaviviridae. Dengue virus is widely distributed throughout the tropical and semitropical regions of the world and is transmitted to humans by mosquito vectors. Dengue virus is a leading cause of hospitalization and death in children in at least eight tropical Asian countries. There are four serotypes of dengue virus (DEN-1, DEN-2, DEN-3, and DEN-4) that annually cause an estimated 50-100 million cases of dengue fever and 500,000 cases of the more severe form of dengue virus infection known as dengue hemorrhagic fever/dengue shock syndrome (DHFIDSS). This latter disease is seen predominately in children and adults experiencing a second dengue virus infection with a serotype different than that of their first dengue virus infection and in primary infection of infants who still have circulating dengue-specific maternal antibody. A vaccine is needed to lessen the disease burden caused by dengue virus, but none is licensed.

Because of the association of more severe disease with secondary dengue virus infection, a successful vaccine must induce immunity to all four serotypes. Immunity is primarily mediated by neutralizing antibody directed against the envelope (E) glycoprotein, a virion structural protein. Infection with one serotype induces long-lived homotypic immunity and a short-lived heterotypic immunity. Therefore, the goal of immunization is to induce a long-lived neutralizing antibody response against DEN-1, DEN-2, DEN-3, and DEN-4, which can best be achieved economically using live attenuated virus vaccines. This is a reasonable goal since a live attenuated vaccine has already been developed for the related yellow fever virus, another mosquito-borne flavivirus present in tropical and semitropical regions of the world.

The evaluation of live attenuated dengue vaccine candidates in rhesus monkeys requires wild type control viruses for each of the four dengue serotypes. These control viruses are used for comparison to the attenuated strains and post-vaccination challenge to assess vaccine efficacy. As such, these viruses need to be well characterized and sufficiently pure to ensure that they will replicate to consistent levels in rhesus monkeys. Characterization generally includes sequence analysis, titration, and evaluation in monkeys. The following viruses have been characterized: (1) DEN1 WP (2) DEN1 Puerto Rico/94 (3) DEN2 NGC prototype (4) DEN2 Tonga/74 (5) DEN3 Sleman/78 and (6) DEN4 Dominica/81.