Many pandemics have been attributed to the ability of some RNA viruses to change their host range to include humans. Here, we review the mechanisms of disease emergence that are related to the host-range specificity of selected mosquito-borne alphaviruses and flaviviruses. We discuss viruses of medical importance, including Venezuelan equine and Japanese encephalitis viruses, dengue viruses and West Nile viruses.

Since it emerged in Japan in the 1870s, Japanese encephalitis has spread across Asia and has become the most important cause of epidemic encephalitis worldwide. Four genotypes of Japanese encephalitis virus (JEV) are presently recognized (representatives of genotypes I to III have been fully sequenced), but its origin is not known. We have determined the complete nucleotide and amino acid sequence of a genotype IV Indonesian isolate (JKT6468) which represents the oldest lineage, compared it with other fully sequenced genomes, and examined the geographical distribution of all known isolates. JKT6468 was the least similar, with nucleotide divergence ranging from 17.4 to 19.6% and amino acid divergence ranging from 4.7 to 6.5%. It included an unusual series of amino acids at the carboxy terminus of the core protein unlike that seen in other JEV strains. Three signature amino acids in the envelope protein (including E327 Leu3Thr/Ser on the exposed lateral surface of the putative receptor binding domain) distinguished genotype IV strains from more recent genotypes. Analysis of all 290 JEV isolates for which sequence data are available showed that the IndonesiaMalaysia region has all genotypes of JEV circulating, whereas only more recent genotypes circulate in other areas (P < 0.0001). These results suggest that JEV originated from its ancestral virus in the IndonesiaMalaysia region and evolved there into the different genotypes which then spread across Asia. Our data, together with recent evidence on the origins of other emerging viruses, including dengue virus and Nipah virus, imply that tropical southeast Asia may be an important zone for emerging pathogens.

Summary The Asian lineage of Zika virus (ZIKV) has recently caused epidemics and severe disease. Unraveling the mechanisms causing increased viral transmissibility and disease severity requires experimental systems. We report an infectious cDNA clone of ZIKV that was generated using a clinical isolate of the Asian lineage. The cDNA clone-derived RNA is infectious in cells, generating recombinant ZIKV. The recombinant virus is virulent in established ZIKV mouse models, leading to neurological signs relevant to human disease. Additionally, recombinant ZIKV is infectious for Aedes aegypti and thus provides a means to examine virus transmission. The infectious cDNA clone was further used to generate a luciferase ZIKV that exhibited sensitivity to a pan-flavivirus inhibitor, highlighting its potential utility for antiviral screening. This ZIKV reverse genetic system, together with mouse and mosquito infection models, may help identify viral determinants of human virulence and mosquito transmission, as well as inform vaccine and therapeutic strategies.

Neutralization of flaviviruses in vivo correlates with the development of an antibody response against the viral envelope (E) protein. Previous studies demonstrated that monoclonal antibodies (MAbs) against an epitope on the lateral ridge of domain III (DIII) of the West Nile virus (WNV) E protein strongly protect against infection in animals.Based on X-ray crystallography and sequence analysis, an analogous type-specific neutralizing epitope for individual serotypes of the related flavivirus dengue virus (DENV) was hypothesized. Using yeast surface display of DIII variants, we defined contact residues of a panel of type-specific, subcomplex-specific, and cross-reactive MAbs that recognize DIII of DENV type 2 (DENV-2) and have different neutralizing potentials. Type-specific MAbs with neutralizing activity against DENV-2 localized to a sequence-unique epitope on the lateral ridge of DIII, centered at the FG loop near residues E383 and P384, analogous in position to that observed with WNV-specific strongly neutralizing MAbs. Subcomplex-specific MAbs that bound some but not all DENV serotypes and neutralized DENV-2 infection recognized an adjacent epitope centered on the connecting A strand of DIII at residues K305, K307, and K310. In contrast, several MAbs that had poor neutralizing activity against DENV-2 and cross-reacted with all DENV serotypes and other flaviviruses recognized an epitope with residues in the AB loop of DIII, a conserved region that is predicted to have limited accessibility on the mature virion. Overall, our experiments define adjacent and structurally distinct epitopes on DIII of DENV-2 which elicit type-specific, subcomplex-specific, and cross-reactive antibodies with different neutralizing potentials.Dengue fever (DF), the most prevalent arthropod-borne viral illness in humans, is caused by dengue virus (DENV). The four serotypes of DENV are transmitted to humans primarily by the mosquitoes Aedes aegypti and Aedes albopictus. DENV is a member of the Flaviviridae family and is related to the viruses that cause yellow fever and the Japanese, St. Louis, and West Nile encephalitides (8). Infection by DENV causes a spectrum of clinical disease, ranging from an acute, debilitating, selflimited febrile illness (DF) to a life-threatening hemorrhagic and capillary leak syndrome (dengue hemorrhagic fever/dengue shock syndrome). At present, no approved antiviral treatment or vaccine is available, and therapy is supportive in nature. DENV causes an estimated 25 to 100 million cases of DF and 250,000 cases of dengue hemorrhagic fever per year worldwide, with 2.5 billion people at risk for infection (27,48).DENV is an enveloped virus with a single-stranded, positivesense RNA genome (11). The 10.7-kilobase genome is translated as a single polyprotein, which is then cleaved into three structural proteins (C, prM/M, and E) and seven nonstructural (NS) proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5) by virus-and host-encoded proteases. The 500-Å DENV mature virion has a well-organized outer protein she...

Endemic/epidemic dengue viruses (DEN) that are transmitted among humans by the mosquito vectors Aedes aegypti and Aedes albopictus are hypothesized to have evolved from sylvatic DEN strains that are transmitted among nonhuman primates in West Africa and Malaysia by other Aedes mosquitoes. We tested this hypothesis with phylogenetic studies using envelope protein gene sequences of both endemic/epidemic and sylvatic strains. The basal position of sylvatic lineages of DEN-1, -2, and -4 suggested that the endemic/epidemic lineages of these three DEN serotypes evolved independently from sylvatic progenitors. Time estimates for evolution of the endemic/epidemic forms ranged from 100 to 1,500 years ago, and the evolution of endemic/epidemic forms represents relatively recent events in the history of DEN evolution. Analysis of envelope protein amino acid changes predicted to have accompanied endemic/epidemic emergence suggested a role for domain III in adaptation to new mosquito and/or human hosts.Dengue viruses (DEN) (Flaviviridae: Flavivirus) are serious human pathogens that occur nearly throughout the tropics, with ca. 100 million cases annually (16). DEN comprise four serotypes (DEN-1 to DEN-4); although epidemiologically similar, they are genetically and antigenically distinct. Infection with one serotype leads to lifelong protection against homologous reinfection but only brief protection against heterologous challenge (21, 38).DEN cause dengue fever, a self-limited febrile illness lasting 2 to 10 days that has been known in the medical literature for over 200 years. Infrequent epidemics of dengue fever occurred in tropical areas until the 1950s. After War World II, this pattern of disease was disrupted by the emergence of dengue hemorrhagic fever and dengue shock syndrome, more severe diseases characterized by thrombocytopenia, hemorrhage, and excessive plasma leakage (16,28). Two principal hypotheses have been proposed to explain the hemorrhagic form of disease: (i) the immune enhancement theory maintains that hemorrhage occurs in secondary infections when DEN-specific antibodies and memory T cells resulting from primary infection with another serotype enhance the binding of virus-immunoglobulin G complexes to FcY receptors on monocytic cells, and (ii) certain phenotypes of DEN are more virulent than others. Recent phylogenetic studies suggest that an Asian genotype of DEN-2 recently introduced into the New World may be associated with increased risk for hemorrhagic fever and shock in the presence of heterologous antibody (32). DEN strains of reduced virulence have also been described; endemic transmission on the South Pacific islands of Tonga, involving vectors other than Aedes aegypti, may result in less severe disease because of the lesser selection for high viremia imposed by more susceptible vectors, like Aedes polynesiensis (13). Two distinct DEN transmission cycles occur: (i) endemic and epidemic DEN involving human hosts and transmission byA. aegypti, with Aedes albopictus and other Aedes mosquitoes serv...