Single nucleotide polymorphisms in candidate genes and dengue severity in children: A case–control, functional and meta-analysis study Caroline Xavier-Carvalho a , Gerusa Gibson b , Patrícia Brasil c , Ralph X. Ferreira d , Reinaldo de Souza Santos b , Oswaldo Gonçalves Cruz e , Solange Artimos de Oliveira d , Marília de Sá Carvalho e , Antonio G. Pacheco e , Claire F. Kubelka f,⇑,1 , Milton O. Moraes a,⇑,1 a Laboratório de Hanseníase, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, FIOCRUZ, Avenida Brasil 4365, Manguinhos, 21040-360 Rio de Janeiro, Brazil b Departamento de Endemias Samuel Pessoa, Escola Nacional de Saúde Pública (ENSP), FIOCRUZ, Rio de Janeiro, Brazil c Instituto de Pesquisa Clínica Evandro Chagas (IPEC) FIOCRUZ, Rio de Janeiro, Brazil d Departamento de Medicina Clínica, Disciplina de Doenças Infecciosas e Parasitárias, Hospital Universitário Antonio Pedro, UFF, Niterói, Rio de Janeiro, Brazil e Programa de Computação Científica (PROCC) – Prédio da Antiga Residência Oficial FIOCRUZ, Rio de Janeiro, Brazil f Laboratório de Imunologia Viral, Instituto Oswaldo Cruz – FIOCRUZ, Rio de Janeiro, Brazil article info Article history: Received 8 April 2013 Received in revised form 13 August 2013 Accepted 21 August 2013 Available online 7 September 2013 Keywords: CLEC5A DC-SIGN TNF Cytokines Dengue DHF abstract Dengue is an arthropod-borne emerging viral disease with high morbidity and mortality risk in tropical coun- tries like Brazil. Clinical manifestations are vast, ranging from asymptomatic to most severe forms of dengue such as shock. Previous data have shown that host genetics play a role in disease susceptibility and severity. Herein, we have tested the association of single nucleotide polymorphisms (SNPs) at TNF, IL10, MIF, DCSIGN, CLEC5A, NOD2, CCR5 and MRC1 as candidate genes using a matched case–control study design including 88 severe children cases of dengue patients and 335 healthy unrelated subjects that was also separated in IgG + and IgG  controls. We demonstrated that the TT genotype of CLEC5A SNP (rs1285933 C>T) is associated with dengue severity (OR = 2.25; p = 0.03) and that GG genotype of 336G>A DCSIGN (CD209) SNP is associ- ated with protection to severe dengue (OR = 0.12; p = 0.04). Both comparisons were borderline significant when cases were compared with IgG + controls subgroup. Nevertheless, genotype–phenotype correlation was also assessed using serum levels of TNF from infected patients at the onset of dengue fever, and CT/TT carriers in CLEC5A secreted higher levels of TNF than CC individuals in 5–7 days of infection. No significant difference was observed in TNF levels between genotypes GG versus AG/AA at DCSIGN promoter. Next, we performed a meta-analysis retrieving results from the literature for 336G>A DCSIGN and 308G>A TNF SNPs demonstrating that the consensus estimates of these SNPs indicated no association with dengue severity (when compared to Dengue fever) in the overall analysis. But, a subgroup analysis in the 336G>A DCSIGN, the G allele was associated with severe dengue susceptibility in Asians (OR allele = 2.77; p = 0.0001; OR carri- ers = 2.99; p = 0.0001) and protection in Brazilians (OR allele =0.66; p=0.013). In summary, our results suggest that genetic variations at CLEC5A increase the risk and regulate TNF secretion in dengue severity among Bra- zilians. Also, combined data of the literature suggest population-specific effect of the 336 DCSIGN SNP more prominent in Asians and in a different direction than Brazilians. Ó 2013 Elsevier B.V. All rights reserved. 1. Introduction During the last decades dengue became the most important arthropod-borne emerging viral disease with high morbidity and mortality risk and is considered a neglected disease by the World Health Organization (WHO/TDR, 2009). It is caused by the Dengue Virus (DENV), which has four circulating serotypes, DENV-1 to DENV-4 (Guzman et al., 2010; Wilder-Smith et al., 2010). In Brazil, after the introduction of DENV-3 in 2002, the disease has been presenting increasing numbers of reported cases and deaths. It recently became alarming when DENV-2 re-emerged in Rio de Janeiro during 2008 (Rodriguez-Barraquer et al., 2011; Teixeira et al., 2009) and, more recently, by the DENV-4 introduction in 2010 raising serious concerns about the increase of severe cases of the disease in the next few years (Nogueira and Eppinghaus, 2011; Temporao et al., 2011). 1567-1348/$ - see front matter Ó 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.meegid.2013.08.017 ⇑ Corresponding authors. Address: Laboratório de Imunologia Viral, Instituto Oswaldo Cruz – FIOCRUZ, Rio de Janeiro Av. Brasil 4365, Manguinhos, Rio de Janeiro – RJ, CEP 21040-360, Brazil (C.F. Kubelka), Leprosy Laboratory – FIOCRUZ, Av. Brasil 4365, Manguinhos, Rio de Janeiro – RJ, CEP 21040-360, Brazil. Tel.: +21 25621556 (M.O. Moraes). E-mail addresses: claire@ioc.fiocruz.br (C.F. Kubelka), mmoraes@fiocruz.br (M.O. Moraes). 1 These authors share senior authorship in this work. Infection, Genetics and Evolution 20 (2013) 197–205 Contents lists available at ScienceDirect Infection, Genetics and Evolution journal homepage: www.elsevier.com/locate/meegid