Molecular analysis of the dengue virus type 1 and 2 in Brazil based on sequences of the genomic envelope-nonstructural protein 1 junction region

The genomic sequences of the Envelope-Non-Structural protein 1 junction region (E/NS1) of 84 DEN-1 and 22 DEN-2 isolates from Brazil were determined. Most of these strains were isolated in the period from 1995 to 2001 in endemic and regions of recent dengue transmission in São Paulo State. Sequence data for DEN-1 and DEN-2 utilized in phylogenetic and split decomposition analyses also include sequences deposited in GenBank from different regions of Brazil and of the world. Phylogenetic analyses were done using both maximum likelihood and Bayesian approaches. Results for both DEN-1 and DEN-2 data are ambiguous, and support for most tree bipartitions are generally poor, suggesting that E/NS1 region does not contain enough information for recovering phylogenetic relationships among DEN-1 and DEN-2 sequences used in this study. The network graph generated in the split decomposition analysis of DEN-1 does not show evidence of grouping sequences according to country, region and clades. While the network for DEN-2 also shows ambiguities among DEN-2 sequences, it suggests that Brazilian sequences may belong to distinct subtypes of genotype III.

Analysis of HIV- type 1 protease and reverse transcriptase in Brazilian children failing highly active antiretroviral therapy (HAART)

The aim of this study was to evaluate the genotypic resistance profiles of HIV-1 in children failing highly active antiretroviral therapy (HAART). Forty-one children (median age = 67 months) receiving HAART were submitted to genotypic testing when virological failure was detected. cDNA was extracted from PBMCs and amplified by nested PCR for the reverse transcriptase and protease regions of the pol gene. Drug resistance genotypes were determined from DNA sequencing. According to the genotypic analysis, 12/36 (33.3%) and 6/36 (16.6%) children showed resistance and possible resistance, respectively, to ZDV; 5/36 (14%) and 4/36 (11.1%), respectively, showed resistance and possible resistance to ddI; 4/36 (11.1%) showed resistance to 3TC and D4T; and 3/36 (8.3%) showed resistance to Abacavir. A high percentage (54%) of children exhibited mutations conferring resistance to NNRTI class drugs. Respective rates of resistance and possible resistance to PIs were: RTV (12.2%, 7.3%); APV (2.4%, 12.1%); SQV(0%, 12.1%); IDV (14.6%, 4.9%), NFV (22%, 4.9%), LPV/RTV (2.4%, 12.1%). Overall, 37/41 (90%) children exhibited virus with mutations related to drug resistance, while 9% exhibited resistance to all three antiretroviral drug classes.

T CD4+ cells count among patients co-infected with human immunodeficiency virus type 1 (HIV-1) and human T-cell leukemia virus type 1 (HTLV-1): high prevalence of tropical spastic paraparesis/HTLV-1-associated myelopathy (TSP/HAM)

INTRODUCTION: HIV positive patients co-infected with HTLV-1 may have an increase in their T CD4+ cell counts, thus rendering this parameter useless as an AIDS-defining event. OBJECTIVE: To study the effects induced by the co-infection of HIV-1 and HTLV-1 upon CD4+ cells. MATERIAL AND METHODS: Since 1997, our group has been following a cohort of HTLV-1-infected patients, in order to study the interaction of HTLV-1 with HIV and/or with hepatitis C virus (HCV), as well as HTLV-1-only infected asymptomatic carriers and those with tropical spastic paraparesis/HTLV-1 associated myelopathy (TSP/HAM). One hundred and fifty HTLV-1-infected subjects have been referred to our clinic at the Institute of Infectious Diseases "Emílio Ribas", São Paulo. Twenty-seven of them were also infected with HIV-1 and HTLV-1-infection using two ELISAs and confirmed and typed by Western Blot (WB) or polymerase chain reaction (PCR). All subjects were evaluated by two neurologists, blinded to the patient's HTLV status, and the TSP/HAM diagnostic was based on the World Health Organization (WHO) classification. AIDS-defining events were in accordance with the Centers for Disease Control (CDC) classification of 1988. The first T CD4+ cells count available before starting anti-retroviral therapy are shown compared to the HIV-1-infected subjects at the moment of AIDS defining event. RESULTS: A total of 27 HIV-1/HTLV-1 co-infected subjects were identified in this cohort; 15 already had AIDS and 12 remained free of AIDS. The median of T CD4+ cell counts was 189 (98-688) cells/mm³ and 89 (53-196) cells/mm³ for co-infected subjects who had an AIDS-defining event, and HIV-only infected individuals, respectively (p = 0.036). Eight of 27 co-infected subjects (30%) were diagnosed as having a TSP/HAM simile diagnosis, and three of them had opportunistic infections but high T CD4+ cell counts at the time of their AIDS- defining event. DISCUSSION: Our results indicate that higher T CD4+ cells count among HIV-1/HTLV-1-coinfected subjects was found in 12% of the patients who presented an AIDS-defining event. These subjects also showed a TSP/HAM simile picture when it was the first manifestation of disease; this incidence is 20 times higher than that for HTLV-1-only infected subjects in endemic areas.

Evaluation of the human immunodeficiency virus type 1 and 2 antibodies detection in dried whole blood spots (dbs) samples

Human Immunodeficiency Vírus Type 1 and 2 antibodies detection was performed in 457 dried whole blood spots samples (S&S 903). Q-Preven HIV 1+2 was the screening test used. The results were compared with the gold standard serum tests by ELISA (Cobas Core e Axsym HIV1/2 gO) and imunofluorescence was the definitive confirmatory test. The samples were obtained from the Hospital Nossa Senhora da Conceição in Porto Alegre, RS - Brazil, through whole blood transfer to filter paper card and sent to Caxias do Sul, RS - Brazil where the tests were performed. The dried whole blood spot stability was evaluated with two different panels. The first one was composed of five negative and five positive samples stored at room temperature, 4 ºC, -20 ºC and -70 ºC, while the second was composed of two negative and three positive samples stored at 37 ºC (humidity <50%). Each sample was screened every week for six weeks. These measurement results didn't show variation during the study period. The detected sensibility was 100%, specificity was 99.6%, the positive predictive value was 99.5% and negative predictive values were 100%. The results demonstrated high performance characteristics, opening a new perspective of dried whole blood spot utilization in HIV screening diagnosis.