Immune profile and responses of a novel dengue DNA vaccine encoding an EDIII-NS1 consensus design based on Indo-African sequences.

The ongoing COVID-19 pandemic highlights the need to tackle viral variants, expand the number of antigens, and assess diverse delivery systems for vaccines against emerging viruses. In the present study, a DNA vaccine candidate was generated by combining in tandem envelope protein domain III (EDIII) of dengue virus serotypes 1-4 and a dengue virus (DENV)-2 non-structural protein 1 (NS1) protein-coding region. Each domain was designed as a serotype-specific consensus coding sequence derived from different genotypes based on the whole genome sequencing of clinical isolates in India and complemented with data from Africa. This sequence was further optimized for protein expression. In silico structural analysis of the EDIII consensus sequence revealed that epitopes are structurally conserved and immunogenic. The vaccination of mice with this construct induced pan-serotype neutralizing antibodies and antigen-specific T cell responses. Assaying intracellular interferon (IFN)-γ staining, immunoglobulin IgG2(a/c)/IgG1 ratios, and immune gene profiling suggests a strong Th1-dominant immune response. Finally, the passive transfer of immune sera protected AG129 mice challenged with a virulent, non-mouse-adapted DENV-2 strain. Our findings collectively suggest an alternative strategy for dengue vaccine design by offering a novel vaccine candidate with a possible broad-spectrum protection and a successful clinical translation either as a stand alone or in a mix and match strategy.

Isolation and molecular characterization of dengue virus clinical isolates from pediatric patients in New Delhi.

OBJECTIVE: To characterize the in vitro replication fitness, viral diversity, and phylogeny of dengue viruses (DENV) isolated from Indian patients. METHODS: DENV was isolated from whole blood collected from patients by passaging in cell culture. Passage 3 viruses were used for growth kinetics in C6/36 mosquito cells. Parallel efforts also focused on the isolation of DENV RNA from plasma samples of the same patients, which were processed for next-generation sequencing. RESULTS: It was possible to isolate 64 clinical isolates of DENV, mostly DENV-2. Twenty-five of these were further used for growth curve analysis in vitro, which showed a wide range of replication kinetics. The highest viral titers were associated with isolates from patients with dengue with warning signs and severe dengue cases. Full genome sequences of 21 DENV isolates were obtained. Genome analysis mapped the circulating DENV-2 strains to the Cosmopolitan genotype. CONCLUSIONS: The replication kinetics of isolates from patients with mild or severe infection did not differ significantly, but the viral titers varied by two orders of magnitude between the isolates, suggesting differences in replication fitness among the circulating DENV-2.