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

ABSTRACT: 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.

SUBMITTER: Kar M

PROVIDER: S-EPMC6823047 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature

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

Kar Meenakshi M Nisheetha Amul A Kumar Anuj A Jagtap Suraj S Shinde Jitendra J Singla Mohit M M Saranya S Pandit Awadhesh A Chandele Anmol A Kabra Sushil K SK Krishna Sudhir S Roy Rahul R Lodha Rakesh R Pattabiraman Chitra C Medigeshi Guruprasad R GR

International journal of infectious diseases : IJID : official publication of the International Society for Infectious Diseases 20181207

<h4>Objective</h4>To characterize the in vitro replication fitness, viral diversity, and phylogeny of dengue viruses (DENV) isolated from Indian patients.<h4>Methods</h4>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.<h4>Results</h4 ...[more]

PMID: 30528666

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Project description:Nucleotide sequence of complete genome of a new isolate (KAN-6) of tomato leaf curl New Delhi virus (ToLCNDV) from Kanpur, Uttar Pradesh, India was determined. Sequence analysis indicated that it shared maximum identity to ToLCNDV isolates from pumpkin and ashgourd. Infectious clones of isolate KAN-6 along with two other ToLCNDV isolates (MOD-21 & FAI-19) obtained from potato fields of Modipuram and Faizabad, India were produced and used in symptom expression studies in N. benthamiana and potato plants through agro-inoculation. These isolates produced different symptoms both in N. benthamiana and potato. Severe symptoms of yellow mottling, downward curling and stunted growth were observed in N. benthamiana plants inoculated with KAN-6. MOD-21-inoculated plants also showed downward curling, stunted growth, but yellow mottling was observed only in older leaves whereas FAI-19-inoculated plants produced only downward curling symptoms. In case of potato, typical symptoms of apical leaf curl disease were observed in cultivar Kufri Pukhraj inoculated with MOD-21 and KAN-6 that are similar to those produced by virus-infected plants in the field. However, MOD-21 produced more prominent yellow mosaic symptoms as compared to KAN-6. FAI-19 produced only restricted yellow spots in Kufri Pukhraj. Only mild symptoms appeared in KAN-6 and no symptoms were observed in MOD-21- and FAI-19-inoculated Kufri Bahar plants which is known to show lowest seed degeneration under field conditions. Analysis of genomic components indicated that these isolates had 94.8-94.9% and 87.9-97.3% identity among them in DNA A and DNA B, respectively. The results of the study indicate the association of ToLCNDV isolates of different symptomatology with apical leaf curl disease of potato. This is also a first experimental demonstration of Koch's postulate for a begomovirus associated with apical leaf curl disease of potato.Author names: Please confirm if the author names (Swarup Kumar Chakrabarti) are presented accurately and in the correct sequence (given name, middle name/initial, family name).Yes. It is correct.Supplementary informationThe online version contains supplementary material available at 10.1007/s13205-021-02752-5.

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

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

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