RESUMO
Dengue is hyperendemic in Colombia, where a cyclic behavior of serotype replacement leading to periodic epidemics has been observed for decades. This level of endemicity favors accumulation of dengue virus genetic diversity and could be linked to disease outcome. To assess the genetic diversity of dengue virus type 2 in Colombia, we sequenced the envelope gene of 24 virus isolates from acute cases of dengue or severe dengue fever during the period 2013-2016. The phylogenetic analysis revealed the circulation of the Asian-American genotype of dengue virus type 2 in Colombia during that period, the intra-genotype variability leading to divergence in two recently circulating lineages with differential geographic distribution, as well as the presence of nonsynonymous substitutions accompanying their emergence and diversification.
Assuntos
Vírus da Dengue/genética , Dengue/virologia , Variação Genética , Genótipo , RNA Viral/sangue , Adolescente , Adulto , Bancos de Espécimes Biológicos , Criança , Pré-Escolar , Colômbia/epidemiologia , Dengue/epidemiologia , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Pessoa de Meia-Idade , Filogenia , RNA Viral/genética , Estudos Retrospectivos , Sorogrupo , Proteínas do Envelope Viral/genética , Adulto JovemRESUMO
Dengue viruses (DENV) have become the most important arthropod-borne viruses, causing dengue and severe dengue fever in at least 50-100 million cases each year, mainly in tropical and subtropical countries. During recent years, important advances in the molecular biology concerning the life cycle of these viruses have allowed the manipulation and generation of recombinant viruses and replicons with multiple applications, mainly in viral biology and the screening of antiviral compounds. In the present study, we describe the construction of an enhanced green fluorescent protein-bearing DENV replicon under the control of the cytomegalovirus immediate early promoter. Following a rational in silico design and cloning by standard molecular biology techniques, a reporter DENV-2 replicon and a replication-deficient mutant were constructed, and characterized by confocal microscopy and real-time RT-PCR. The results showed successful transcription, translation, and autonomous viral RNA replication of the DENV replicon from its DNA clone. This novel DENV replicon will allow the study of viral replication and testing of antiviral candidates without the need for in vitro transcription.