EVOLUTIONARY CONSEQUENCES OF DENGUE VIRUS EMERGENCE

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Sylvatic dengue viruses circulate in an unknown reservoir in the forest canopies of Africa and Asia and do not contribute to human dengue disease, which is caused by endemic dengue viruses that circulate only between humans and human-biting mosquitoes. Because the barriers that prevent sylvatic viruses have not been identified, the risk that sylvatic viruses pose for emergence of new dengue strains cannot be assessed. Three hypotheses to explain the absence of sylvatic viruses in humans include: 1. Sylvatic viruses are less fit in humans than endemic viruses 2. Sylvatic viruses are less fit in human-biting mosquitoes than endemic viruses 3. Sylvatic viruses and endemic viruses are equally fit in both humans and human-biting mosquitoes, but occupy separate spatial niches (the forest canopy versus the forest floor). To distinguish among these hypotheses, my lab has compared the whole genome sequence of two sylvatic dengue type 2 viruses with eight endemic dengue type 2 viruses and identified conserved genetic differences between the two groups in each of the ten genes and two untranslated regions that compose the dengue virus genome. Next the kinetics and magnitude of replication, which represents one component of fitness, of a panel of four sylvatic dengue type 2 viruses and four endemic dengue type 2 viruses will be measured in human liver (HuH-7) cells and mosquito (C6/36) cells. If hypotheses 1 or 2 is correct then sylvatic viruses should show significantly lower levels of replication than endemic viruses in human or mosquito cells, respectively, whereas a lack of difference between the two groups would support hypothesis 3. Sylvatic viruses showing low replication will be serially passaged on that cell type to allow for adaptation and then sequenced to identify candidate mutations that could be associated with adaptations for human infection.