Vaccine candidates derived from a novel infectious cDNA clone of an American genotype dengue virus type 2

A full-length cDNA clone was generated from the DEN-2 virus and used to produce recombinant DEN-2 (rDEN-2) and rDEN2Δ30. Viruses were evaluated for replication in SCID mice transplanted with human hepatoma cells (SCID-HuH-7 mice), in mosquitoes, and in rhesus monkeys. Neutralizing antibody induction and protective efficacy were also assessed in rhesus monkeys.The rDEN2Δ30 virus was ten-fold reduced in replication in SCID-HuH-7 mice when compared to the parent virus. The rDEN-2 viruses were not infectious for Aedes mosquitoes, but both readily infected Toxorynchites mosquitoes. In rhesus monkeys, rDEN2Δ30 appeared to be slightly attenuated when compared to the parent virus as measured by duration and peak of viremia and neutralizing antibody induction. A derivative of rDEN2Δ30, designated rDEN2Δ30-4995, was generated by incorporation of a point mutation previously identified in the NS3 gene of DEN-4 and was found to be more attenuated than rDEN2Δ30 in SCID-HuH-7 mice.The rDEN2Δ30 and rDEN2Δ30-4995 viruses can be considered for evaluation in humans and for inclusion in a tetravalent dengue vaccine.The increased prevalence of disease caused by the mosquito-borne dengue (DEN) viruses (four serotypes; DEN-1 – DEN-4) has intensified the effort to generate a vaccine that would both confer protection and be economically feasible for use in countries with limited resources for healthcare [1]. Dengue fever and dengue hemorrhagic fever and shock (DHF/DSS) are a severe disease burden for tropical and semitropical countries inhabited by more than 2.5 billion people [2]. Risk factors for the more severe disease, DHF/DSS, include the strain of virus, age and genetic background of the host, and secondary infection by a DEN serotype different from that which caused the primary infection [2]. Increased risk associated with secondary infection by a different DEN serotype is believed to be caused both by increased virus replication resulting from antibody-dependent enhancement and by a