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

OBJECTIVES: 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 isolation of DENV RNA from plasma samples of the same patients and processed for next generation sequencing.

RESULTS: We were able to isolate 64 clinical isolates, mostly DENV-2, of which 25 were further used for growth curve analysis in vitro which showed a wide range of replication kinetics. Highest viral titers were of isolates from dengue with warning signs and severe dengue cases. We obtained full genome sequences of 21 DENV isolates. 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 was not significantly different but the viral titers between the isolates varied by two orders of magnitude suggesting differences in replication fitness among the circulating DENV-2 isolates.