DNA Encoding an HIV-1 Gag/Human Lysosome-Associated Membrane Protein-1 Chimera Elicits a Broad Cellular and Humoral Immune Response in Rhesus Macaques

Previous studies of HIV-1 p55Gag immunization of mice have demonstrated the usefulness of targeting antigens to the cellular compartment containing the major histocompatibility complex type II (MHC II) complex molecules by use of a DNA antigen formulation encoding Gag as a chimera with the mouse lysosome-associated membrane protein (mLAMP/gag). In the present study, we have analyzed the magnitude and breadth of Gag-specific T-lymphocyte and antibody responses elicited in Rhesus macaques after immunization with DNA encoding a human LAMP/gag (hLAMP/gag) chimera. ELISPOT analyses indicated that the average Gag-specific IFN-γ response elicited by the hLAMP/gag chimera was detectable after only two or three naked DNA immunizations in all five immunized macaques and reached an average of 1000 spot-forming cells (SFC)/10(6) PBMCs. High IFN-γ ELISPOT responses were detected in CD8(+)-depleted cells, indicating that CD4(+) T-cells play a major role in these responses. The T-cell responses of four of the macaques were also tested by use of ELISPOT to 12 overlapping 15-amino acids (aa) peptide pools containing ten peptides each, encompassing the complete Gag protein sequence. The two Mamu 08 immunized macaques responded to eight and twelve of the pools, the Mamu B01 to six, and the other macaque to five pools indicating that the hLAMP/gag DNA antigen formulation elicits a broad T-cell response against Gag. Additionally, there was a strong HIV-1-specific IgG response. The IgG antibody titers increased after each DNA injection, indicating a strong amnestic B-cell response, and were highly elevated in all the macaques after three immunizations. Moreover, the serum of each macaque recognized 13 of the 49 peptides of a 20-aa peptide library covering the complete Gag amino acid sequence. In addition, HIV-1-specific IgA antibodies were present in the plasma and external secretions, including nasal washes. These data support the findings of increased immunogenicity of genetic vaccines encoded as LAMP chimeras, including the response to DNA vaccines by non-human primates

Early molecular markers predictive of dengue hemorrhagic fever

The management of acute dengue patients during outbreaks is a challenging problem. Most of the dengue fever cases are benign, but some cases develop into a severe and possibly lethal vasculopathy, known as dengue hemorrhagic fever. Early symptoms of dengue and hemorrhagic fever are very similar. An early differential diagnosis is needed to predict which of these two clinical presentations is crucial to proper patient care and public health management. This study evaluates the predictive potential of specific mRNA expression markers of dengue hemorrhagic fever using quantitative real-time PCR assays. Six candidate "dengue hemorrhagic fever specific signature genes" were evaluated and all showed good correlation among their transcription levels at early days of infection and the later development of severe vasculopathy. The markers selected were able to indicate, at early stages of infection, the evolution of a dengue-infected patient to the severe form of the illness. Despite the fact that these results grant further validation studies, the panel of candidate prognostic markers obtained demonstrated the potential to be useful for clinical use in the form of a fast assay based in blood samples

Identification of a Zika NS2B Epitope for Which Absence of IgG Response Is Associated with Severe Neurological Symptoms and the Design of a Biomarker Capable of Discriminatory Diagnostics Between Severe and Non4 Severe Clinical Phenotypes

In this manuscript we describe the engineering of a biomarker for the diagnosis and prognosis of Zika-associated neurological disease. Although the causal association between congenital Zika virus (ZIKV) infection and neurological manifestations has been well documented in the recent years, biomarkers for proper diagnostic and disease outcome still remain to be defined. Combining high-density peptide array and multivariate analysis, we have identified an ZIKV epitope that is associated to a lack of IgG antibody response in patients with severe neurological symptoms. An engineered chimera was developed to discriminate between mild and severe clinical forms of the disease. </div

Complement factor H gene (CFH) polymorphisms C-257T, G257A and haplotypes are associated with protection against severe dengue phenotype, possible related with high CFH expression

Four genetic polymorphisms located at the promoter (C-257T) and coding regions of CFH gene (exon 2 G257A, exon 14 A2089G and exon 19 G2881T) were investigated in 121 dengue patients (DENV-3) in order to assess the relationship between allele/haplotypes variants and clinical outcomes. A statistical value was found between the CFH-257T allele (TT/TC genotypes) and reduced susceptibility to severe dengue (SD). Statistical associations indicate that individuals bearing a T allele presented significantly higher protein levels in plasma. The 257T variant is located within a NF-jB binding site, suggesting that this variant might have effect on the ability of the CFH gene to respond to signals via the NF-jB pathway. The G257A allelic variant showed significant protection against severe dengue. When CFH haplotypes effect was considered, the ancestral CG/CG promoter-exon 2 SNP genotype showed significant risk to SD either in a general comparison (ancestral all variant genotypes), as well as in individual genotypes comparison (ancestral each variant genotype), where the most prevalent effect was observed in the CG/ CG CA/TG comparison. These findings support the involvement of 257T, 257A allele variants and haplotypes on severe dengue phenotype protection, related with high basal CFH expression