A nearly complete genome sequence of a dengue virus serotype 2 strain detected in the serum of a patient in 2019 during the largest outbreak of dengue fever in Bangladesh is reported

Hasanuzzaman, Md

Hooda, Yogesh

Malaker, Apurba R.

Malaker, Roly

Rahman, Hafizur

Saha, Samir K.

Saha, Senjuti

Sajib, Mohammad S. I.

PubMed

Enlighten

Genome Sequence of a Dengue Virus Serotype 2 StrainIdentified during the 2019 Outbreak in BangladeshRoly Malaker,a Mohammad S. I. Sajib,a Apurba R. Malaker,a Hafizur Rahman,a Yogesh Hooda,a,b Md Hasanuzzaman,aSamir K. Saha,a,c,d Senjuti SahaaaChild Health Research Foundation, Dhaka, BangladeshbMRC Laboratory of Molecular Biology, Cambridge, United KingdomcDhaka Shishu (Children) Hospital, Dhaka, BangladeshdBangladesh Institute of Child Health, Dhaka, BangladeshABSTRACT A nearly complete genome sequence of a dengue virus serotype 2 straindetected in the serum of a patient in 2019 during the largest outbreak of denguefever in Bangladesh is reported.Dengue is a mosquito-borne viral disease caused by dengue virus, a single-strandedpositive-sense RNA virus of the family Flaviviridae (1). Here, we report the genomesequence of a dengue virus detected in a clinically symptomatic patient during the2019 dengue outbreak in Bangladesh (2).All protocols were approved by the ethical review board of the Bangladesh Instituteof Child Health. Samples were collected for clinical care and diagnostic testing at thediscretion of the attending physicians, and written consent was obtained from thecaregiver for additional laboratory analysis.For this study, a subset of dengue virus-positive serum specimens, tested using theNS1 antigen kit (product no. 09DEN10D; SD Biosensor, South Korea), were retestedusing reverse transcription-quantitative PCR (RT-qPCR) to estimate the dengue viralload (3). One specimen collected on 27 July 2019 in Dhaka, Bangladesh, had the high-est viral load, with a cycle threshold value of 24, and was selected for next-generationsequencing. For library preparation, viral nucleic acid was extracted from a 200-ml se-rum sample using the Quick-DNA/RNA microprep extraction kit (product no. D7005;Zymo, CA, USA) according to the manufacturer’s protocol. The NEBNext Ultra II RNAlibrary preparation kit (product no. E7770; New England Biolabs, MA, USA) was used toprepare the sequencing library. External RNA Controls Consortium Collection spike-incontrol mix (product no. 4456740; Thermo Fisher Scientific, MA, USA) was used as amarker of potential library preparation errors. The final library was sequenced on anIllumina iSeq100 sequencer using 150-nucleotide paired-end sequencing.The library generated 7,688,000 reads, and 2,467,976 reads passed the default filterof BaseSpace (Illumina). Raw fastq files were uploaded to the IDseq portal to identifythe pathogenic organisms, as described elsewhere (4, 5). In brief, the human readswere filtered out from the fastq file by alignment with a reference database using theSpliced Transcripts Alignment to a Reference (STAR) algorithm. Next, a series of qualitycontrol steps were performed to exclude low-quality reads, duplicates, and low-complexity reads with the help of the Paired-Read Iterative Contig Extension (PRICE)package, the CD-HIT-DUP tool (v4.6.8), and the Lempel-Ziv-Welch (LZW) algorithm.Another round of host filtering was executed by applying the Bowtie2 package. Theremaining reads were queried against the most recent version of the NCBI nucleotideand nonredundant protein databases using Genomic Short-read Nucleotide AlignmentProgram (GSNAP) (v2018-10-26) and RAPSearch 2 (v2.24), respectively. Finally, theCitationMalaker R, Sajib MSI, Malaker AR,Rahman H, Hooda Y, Hasanuzzaman M, SahaSK, Saha S. 2021. Genome sequence of adengue virus serotype 2 strain identifiedduring the 2019 outbreak in Bangladesh.Microbiol Resour Announc 10:e01246-20.https://doi.org/10.1128/MRA.01246-20.Editor Kenneth M. Stedman, Portland StateUniversityCopyright © 2021 Malaker et al. This is anopen-access article distributed under the termsof the Creative Commons Attribution 4.0International license.Address correspondence to Senjuti Saha,senjutisaha@gmail.com.Received 29 October 2020Accepted 25 November 2020Published 7 January 2021January 2021 Volume 10 Issue 1 e01246-20 mra.asm.org 1GENOME SEQUENCESDownloaded from https://journals.asm.org/journal/mra on 09 July 2021 by 2.98.192.99.matched reads and their relative abundance were calculated and mapped at the genuslevel and visualized.The final nearly complete genome of the dengue virus consisted of 10,699 bases,with a GC content of 45.8% and an average coverage depth of 237. It had 92.5% nu-cleotide and 97.5% amino acid sequence identity to the reference dengue virus sero-type 2 genome (GenBank accession no. NC_001474.2). The genome was submitted tothe NCBI GenBank database (accession no. MN328061) and subsequently contextual-ized in the dengue virus serotype 2 build of Nextstrain (https://nextstrain.org/dengue/FIG 1 Phylogenetic trees of dengue virus serotype 2 (DENV2) strains rendered using Nextstrain. (A) Phylogenetic tree ofCHRF_DenV002 (GenBank accession no. MN328061) contextualized with 1,202 dengue virus serotype 2 genomes available onNextstrain. The circle bordered in red represents the position of CHRF_DenV002 in the tree. (B) Phylogenetic tree of CHRF_DenV002contextualized with the closest genomes sequenced from Southeast and East Asian countries (access date, 18 October 2020).Malaker et al.January 2021 Volume 10 Issue 1 e01246-20 mra.asm.org 2Downloaded from https://journals.asm.org/journal/mra on 09 July 2021 by 2.98.192.99.denv2) among 1,202 available dengue virus serotype 2 genomes sampled between1944 and 2019 (6) (Fig. 1A). The sequenced genome is closely related to dengueviruses seen in other Southeast and East Asian countries, specifically, Singapore,Malaysia, Indonesia, South Korea, and China (Fig. 1B).Data availability. The dengue viral genome from Bangladesh has been depositedin GenBank (accession no. MN328061). The raw reads have been deposited in the NCBISequence Read Archive (SRA accession no. SRR12901070). The BioProject and BioSampleaccession numbers are PRJNA672124 and SAMN16557024, respectively.ACKNOWLEDGMENTSWe are grateful to the Chan Zuckerberg Biohub and the Chan Zuckerberg Initiativefor their intellectual support and guidance. We specifically thank Madeline Y. Maydayand Lillian M. Khan for their support with library preparation and Joseph L. DeRisi,Cristina M. Tato, and Rebacca Eggar for overall supervision and guidance with genomeanalysis.This project was funded by the Bill and Melinda Gates Foundation (grantOPP1198769).REFERENCES1. Kautner I, Robinson MJ, Kuhnle U. 1997. Dengue virus infection: epidemiol-ogy, pathogenesis, clinical presentation, diagnosis, and prevention. JPediatr 131:516–524. https://doi.org/10.1016/s0022-3476(97)70054-4.2. Hsan K, Hossain MM, Sarwar MS, Wilder-Smith A, Gozal D. 2019. Unprece-dented rise in dengue outbreaks in Bangladesh. Lancet Infect Dis 19:1287.https://doi.org/10.1016/S1473-3099(19)30616-4.3. Santiago GA, Vergne E, Quiles Y, Cosme J, Vazquez J, Medina JF, Medina F,Colon C, Margolis H, Munoz-Jordan JL. 2013. Analytical and clinical per-formance of the CDC real time RT-PCR assay for detection and typing ofdengue virus. PLoS Negl Trop Dis 7:e2311. https://doi.org/10.1371/journal.pntd.0002311.4. Kalantar KL, Carvalho T, de Bourcy CFA, Dimitrov B, Dingle G, Egger R, HanJ, Holmes OB, Juan Y-F, King R, Kislyuk A, Lin MF, Mariano M, Morse T,Reynoso LV, Cruz DR, Sheu J, Tang J, Wang J, Zhang MA, Zhong E, AhyongV, Lay S, Chea S, Bohl JA, Manning JE, Tato CM, DeRisi JL. 2020. IDseq: anopen source cloud-based pipeline and analysis service for metagenomicpathogen detection and monitoring. Gigascience 9:giaa111. https://doi.org/10.1093/gigascience/giaa111.5. Saha S, Ramesh A, Kalantar K, Malaker R, Hasanuzzaman M, Khan LM,Mayday MY, Sajib MSI, Li LM, Langelier C, Rahman H, Crawford ED, TatoCM, Islam M, Juan YF, de Bourcy C, Dimitrov B, Wang J, Tang J, Sheu J,Egger R, De Carvalho TR, Wilson MR, Saha SK, DeRisi JL. 2019. Unbiasedmetagenomic sequencing for pediatric meningitis in Bangladesh revealsneuroinvasive chikungunya virus outbreak and other unrealized patho-gens. mBio 10:e02877-19. https://doi.org/10.1128/mBio.02877-19.6. Hadfield J, Megill C, Bell SM, Huddleston J, Potter B, Callender C,Sagulenko P, Bedford T, Neher RA. 2018. Nextstrain: real-time tracking ofpathogen evolution. Bioinformatics 34:4121–4123. https://doi.org/10.1093/bioinformatics/bty407.Microbiology Resource AnnouncementJanuary 2021 Volume 10 Issue 1 e01246-20 mra.asm.org 3Downloaded from https://journals.asm.org/journal/mra on 09 July 2021 by 2.98.192.99.

Genome sequence of a dengue virus serotype 2 strain identified during the 2019 outbreak in Bangladesh

Genome sequence of a dengue virus serotype 2 strain identified during the 2019 outbreak in Bangladesh

Abstract

Similar works

Full text

Available Versions

Enlighten