Molecular characterisation of dengue virus type 1 reveals lineage replacement during circulation in Brazilian territory

Abstract

Dengue fever is the most important arbovirus infection found in tropical regions around the world. Dispersal of the vector and an increase in migratory flow between countries have led to large epidemics and severe clinical outcomes, such as dengue haemorrhagic fever and dengue shock syndrome. This study analysed the genetic variability of the dengue virus serotype 1 (DENV-1) in Brazil with regard to the full-length structural genes C/prM/M/E among 34 strains isolated during epidemics that occurred in the country between 1994-2011. Virus phylogeny and time of divergence were also evaluated with only the E gene of the strains isolated from 1994-2008. An analysis of amino acid differences between these strains and the French Guiana strain (FGA/89) revealed the presence of important nonsynonymous substitutions in the amino acid sequences, including residues E297 (Met→Thr) and E338 (Ser→Leu). A phylogenetic analysis of E proteins comparing the studied isolates and other strains selected from the GenBank database showed that the Brazilian DENV-1 strains since 1982 belonged to genotype V. This analysis also showed that different introductions of strains from the 1990s represented lineage replacement, with the identification of three lineages that cluster all isolates from the Americas. An analysis of the divergence time of DENV-1 indicated that the lineage circulating in Brazil emerged from an ancestral lineage that originated approximately 44.35 years ago.Key words: dengue virus type 1 -phylogeny analysis -divergence time The aim of the present study was to analyse the genetic variability at the level of the structural C/prM/ M/E genes of the DENV-1 isolates obtained over the course of epidemics that occurred in several Brazilian states between 1994-2011. Furthermore, we performed phylogenetic studies of the time of divergence of the DENV-1 strains to provide a more thorough understanding of the epidemiology profile of strains circulating in Brazil and their sources. MATERIALS AND METHODS Viral strains Real-time polymerase chain reaction (RT-PCR)and nucleotide sequencing -The C/prM/M/E genes of DENV-1 were fully amplified by RT-PCR according to a method adapted from Lanciotti et al. (1992) using four primer pairs specific for each gene (Supplementary data). The amplicons were purified using the PureLink Quick Gel extraction kit (Invitrogen, California, USA) according to the manufacturer's instructions. The purified cDNA was directly sequenced in an automated capillary ABI 3130 sequencer (Applied Biosystems, USA) using the ABI PRISM BigDye Terminator Cycle Sequencing Ready Reaction 3.1V kit (Applied Biosystems, USA) by the dideoxyribonucleotide chain termination method (Sanger et al. 1977). The amplicons were sequenced in both directions with the same primers used for RT-PCR.Nucleotide and amino acid sequence analysis -The nucleotide sequences were assembled, aligned and analysed with the SeqMan, Editseq and Megalign v.5.0 programs (LaserGene DNA Star ® Package, USA). To analyse the diversity of the genes, C/prM/E re...

Insights of the genetic diversity of DENV-1 detected in Brazil in 25years: Analysis of the envelope domain III allows lineages characterization

“…Aa substitutions were observed in all sequences analyzed, mainly in the domain III of the E gene and corroborate previous observations (Carneiro et al, 2012;Drumond et al, 2012). Substitutions on domain III may alter the viral particle immunogenicity, may influence pathogenicity and determine the receptor specificity, type of vector, host and cell tropism (Chen et al, 1996;dos Santos et al 2002;Gonçalvez et al, 2002;Drumond et al, 2012;Carneiro et al, 2012). As some regions in domain III are target for neutralizing antibodies in humans, mutations in this region could have important functional consequences (Thai et al, 2012).…”

“…In 2009, DENV-1 reemerged and caused a major DENV-1 epidemic in 2010 and, for the first time, distinct viral lineages were reported (dos.DENV are highly diverse(Wang et al, 2002;Aaskov et al, 2006;Thai et al, 2012) and this genetic diversity has been described in different genotypes within each serotype(Rico-Hesse, 1990;2003;Weaver & Vasilakis, 2009). The genotype V of DENV-1 has been previously investigated and the circulation and replacements of lineages have been reported in Brazil and in other American countries(Mendez et al, 2010;Yamanaka et al, 2011;Carneiro et al, 2012;Drumond et al, 2012; Muñoz- Jórdan et al, 2013; Añez & Rivers, 2013;Dash et al, 2015)…”

Molecular surveillance of dengue in Minas Gerais provides insights on dengue virus 1 and 4 circulation in Brazil

“…After the introduction of DENV‐2 into Brazil around 1988/89 [Drumond et al, ] and DENV‐3 at the end of the 1990s [De Araujo et al, ], those DENVs predominated in Brazil in the 1990s and 2000s, respectively [Nogueira et al, ]. Despite shifts in the predominant DENV types circulating in the country, DENV‐1, genotype V has been causing infections in humans throughout this time in Brazil [dos Santos et al, ; Carneiro et al, ; Drumond et al, ; de Bruycker‐Nogueira et al, ]. Different studies have been demonstrating the circulation of different lineages of DENV‐1, genotype V [Carneiro et al ; Drumond et al, ; de Bruycker‐Nogueira et al ; Cunha et al ] with different terminology for those lineages.…”

“…Despite shifts in the predominant DENV types circulating in the country, DENV‐1, genotype V has been causing infections in humans throughout this time in Brazil [dos Santos et al, ; Carneiro et al, ; Drumond et al, ; de Bruycker‐Nogueira et al, ]. Different studies have been demonstrating the circulation of different lineages of DENV‐1, genotype V [Carneiro et al ; Drumond et al, ; de Bruycker‐Nogueira et al ; Cunha et al ] with different terminology for those lineages. But the results presented here demonstrated that in fact, five DENV‐1 lineages were introduced into Brazil.…”

High prevalence of clinically unsuspected dengue disease among children in Ribeirao Preto city, Brazil

“…After the introduction of DENV‐4 in Boa Vista, Roraima State, in 2010, all four serotypes are now circulating in the Brazil [Nogueira et al, ; Amarilla et al, ; Aquino et al, ; Temporao et al, ; Alfonso et al, ; Carneiro et al, ; Drumond et al, , ]. Dengue occurs mainly in young adults [Siqueira et al, ; Teixeira et al, ; San Martín et al, ].…”