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. 2016 Aug 24:4:e2326.
doi: 10.7717/peerj.2326. eCollection 2016.

Analysis of genotype diversity and evolution of Dengue virus serotype 2 using complete genomes

Vaishali P Waman  1 , Pandurang Kolekar  1 , Mukund R Ramtirthkar  2 , Mohan M Kale  2 , Urmila Kulkarni-Kale  1
Affiliations

Analysis of genotype diversity and evolution of Dengue virus serotype 2 using complete genomes

Vaishali P Waman et al. PeerJ. .

Abstract

Background: Dengue is one of the most common arboviral diseases prevalent worldwide and is caused by Dengue viruses (genus Flavivirus, family Flaviviridae). There are four serotypes of Dengue Virus (DENV-1 to DENV-4), each of which is further subdivided into distinct genotypes. DENV-2 is frequently associated with severe dengue infections and epidemics. DENV-2 consists of six genotypes such as Asian/American, Asian I, Asian II, Cosmopolitan, American and sylvatic. Comparative genomic study was carried out to infer population structure of DENV-2 and to analyze the role of evolutionary and spatiotemporal factors in emergence of diversifying lineages.

Methods: Complete genome sequences of 990 strains of DENV-2 were analyzed using Bayesian-based population genetics and phylogenetic approaches to infer genetically distinct lineages. The role of spatiotemporal factors, genetic recombination and selection pressure in the evolution of DENV-2 is examined using the sequence-based bioinformatics approaches.

Results: DENV-2 genetic structure is complex and consists of fifteen subpopulations/lineages. The Asian/American genotype is observed to be diversified into seven lineages. The Asian I, Cosmopolitan and sylvatic genotypes were found to be subdivided into two lineages, each. The populations of American and Asian II genotypes were observed to be homogeneous. Significant evidence of episodic positive selection was observed in all the genes, except NS4A. Positive selection operational on a few codons in envelope gene confers antigenic and lineage diversity in the American strains of Asian/American genotype. Selection on codons of non-structural genes was observed to impact diversification of lineages in Asian I, cosmopolitan and sylvatic genotypes. Evidence of intra/inter-genotype recombination was obtained and the uncertainty in classification of recombinant strains was resolved using the population genetics approach.

Discussion: Complete genome-based analysis revealed that the worldwide population of DENV-2 strains is subdivided into fifteen lineages. The population structure of DENV-2 is spatiotemporal and is shaped by episodic positive selection and recombination. Intra-genotype diversity was observed in four genotypes (Asian/American, Asian I, cosmopolitan and sylvatic). Episodic positive selection on envelope and non-structural genes translates into antigenic diversity and appears to be responsible for emergence of strains/lineages in DENV-2 genotypes. Understanding of the genotype diversity and emerging lineages will be useful to design strategies for epidemiological surveillance and vaccine design.

Keywords: Bioinformatics; Comparative genomics; Dengue virus serotype 2; Evolution; Genetic structure; Genotype/genetic diversity; Molecular phylogeny; Population genetics; Recombination; Selection pressure.

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Conflict of interest statement

The authors declare there are no competing interests.

Figures

Figure 1
Figure 1. Phylogenetic tree of DENV-2 strains obtained using Neighbor-joining (NJ) method in MEGA.
Complete genomes of 990 strains of DENV-2 with 1,000 bootstrap replicates were used to reconstruct phylogenetic tree using NJ method. The fifteen lineages, which are also obtained using STRUCTURE program, are depicted in the tree using color codes as indicated. The bootstrap value (%) associated with each lineage is indicated. There are two lineages (S1 and S2) of Sylvatic genotype, two lineages (C1 and C2) of the cosmopolitan genotype, two lineages (AI-1 and AI-2) of Asian I genotype and a total of seven lineages (AA1–AA7) of Asian/American genotype. The American (AM) and Asian II (AII) genotypes formed independent clusters. AA4* indicates the clade of admixed strains that were found to belong to the AA4 lineage by the STRUCTURE program.
Figure 2
Figure 2. Genetic structure of Asian/American genotype obtained using the STRUCTURE program using admixture model.
Sublevel cluster analysis of the dataset of 552 strains of Asian/American genotype was carried out using STRUCTURE program. The presence of seven distinct lineages are depicted as a bar plot. The seven lineages are AA1 (green), AA2 (red), AA3 (orange), AA4 (pink), AA5 (yellow), AA6 (cyan) and AA7 (blue). These lineages correspond to the clusters obtained using the NJ-based tree (Fig. 1) and have been colour-coded. The STRUCTURE program resolved the cluster assignments of admixed strains of all the seven lineages in general and that of AA4 lineage, in particular. The admixed strain of AA4 cluster, denoted as AA4*, are found to group with the strains of AA3 lineage in the NJ tree, have >0.50 membership for AA4 (indicated in pink) subpopulation and ∼0.35 membership for AA3 (orange) subpopulation.
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Grants and funding

This work received infrastructural and financial support provided by the Center of Excellence (CoE) grant from the Department of Biotechnology (DBT), Government of India, New Delhi, India and the Savitribai Phule Pune University, Pune. UKK received financial assistance from DBT CoE. VPW received a DBT fellowship. MR received support from the Department of Science & Technology (DST), New Delhi, India for an internship under the National Network for Mathematical and Computational Biology (NNMCB) scheme. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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