Élcio Leal

Estrogen receptor alpha polymorphism and susceptibility to uterine leiomyoma

Uterine leiomyoma is the most frequent pelvic tumor found in female genital tract. Some studies have suggested an association between single nucleotide polymorphisms (SNPs) in estrogen receptors genes with susceptibility in developing uterine leiomyoma. In this work, we estimated the frequency of two SNPs: one located in the intron 1 (rs9322331) and other in the exon 1 (rs17847075) of the estrogen receptor alpha (ESR1) gene in 125 women with uterine leiomyoma and 125 healthy women. To do this we used a PCR-RFLP method with MspI and HaeIII restriction enzymes to respectively detect C/T SNPs in the intron 1 and in the exon 1 of ESR1. To our knowledge this is the first study aimed to investigate the association of ESR1 SNPs with the risk of developing uterine leiomyoma in Brazilian women. Our results showed that the allele frequencies of the exon 1 and the intron 1 of the ESR1 gene did not differ between cases and controls (P = 0.325 and 0.175, respectively). Furthermore, our findings provided little support for the association of these SNPs on ESR1 with leiomyoma. However, we found that the SNP in the intron 1 of the ESR1 gene was underrepresented in the Brazilian female population.

Phylogenetic Detection of Recombination with a Bayesian Prior on the Distance between Trees

Genomic regions participating in recombination events may support distinct topologies, and phylogenetic analyses should incorporate this heterogeneity. Existing phylogenetic methods for recombination detection are challenged by the enormous number of possible topologies, even for a moderate number of taxa. If, however, the detection analysis is conducted independently between each putative recombinant sequence and a set of reference parentals, potential recombinations between the recombinants are neglected. In this context, a recombination hotspot can be inferred in phylogenetic analyses if we observe several consecutive breakpoints. We developed a distance measure between unrooted topologies that closely resembles the number of recombinations. By introducing a prior distribution on these recombination distances, a Bayesian hierarchical model was devised to detect phylogenetic inconsistencies occurring due to recombinations. This model relaxes the assumption of known parental sequences, still common in HIV analysis, allowing the entire dataset to be analyzed at once. On simulated datasets with up to 16 taxa, our method correctly detected recombination breakpoints and the number of recombination events for each breakpoint. The procedure is robust to rate and transition∶transversion heterogeneities for simulations with and without recombination. This recombination distance is related to recombination hotspots. Applying this procedure to a genomic HIV-1 dataset, we found evidence for hotspots and de novo recombination.

Diversity of HIV-1 Subtype B: Implications to the Origin of BF Recombinants

Background The HIV-1 subtype B epidemic in Brazil is peculiar because of the high frequency of isolates having the GWGR tetramer at V3 loop region. It has been suggested that GWGR is a distinct variant and less pathogenic than other subtype B isolates. Methodology/Principal Findings Ninety-four percent of the HIV-1 subtype B worldwide sequences (7689/8131) obtained from the Los Alamos HIV database contain proline at the tetramer of the V3 loop of the env gene (GPGR) and only 0.74% (60/8131) have tryptophan (GWGR). By contrast, 48.4% (161/333) of subtype B isolates from Brazil have proline, 30.6% (102/333) contain tryptophan and 10.5% (35/333) have phenylalanine (F) at the second position of the V3 loop tip. The proportion of tryptophan and phenylalanine in Brazilian isolates is much higher than in worldwide subtype B sequences (chi-square test, p = 0.0001). The combined proportion of proline, tryptophan and phenylalanine (GPGR+GWGR+GFGR) of Brazilian isolates corresponds to 89% of all amino acids in the V3 loop. Phylogenetic analysis revealed that almost all subtype B isolates in Brazil have a common origin regardless of their motif (GWGR, GPGR, GGGR, etc.) at the V3 tetramer. This shared ancestral origin was also observed in CRF28_BF and CRF29_BF in a genome region (free of recombination) derived from parental subtype B. These results imply that tryptophan substitution (e.g., GWGR-to-GxGR), which was previously associated with the change in the coreceptor usage within the host, also occurs at the population level. Conclusions/Significance Based on the current findings and previous study showing that tryptophan and phenylalanine in the V3 loop are related with coreceptor usage, we propose that tryptophan and phenylalanine in subtype B isolates in Brazil are kept by selective mechanisms due to the distinct coreceptor preferences in target cells of GWGR, GFGR and GFGR viruses.

Rhinovirus species and their clinical presentation among different risk groups of non-hospitalized patients†

Infections caused by Human Rhinoviruses (HRVs) account for 25–50% of respiratory illnesses among individuals presenting influenza‐like illness (ILI). HRVs could be classified in at least three species: HRV‐A, HRV‐B, and HRV‐C. The HRV‐C species has frequently been described among children and has led to severe illness resulting in hospitalization; however, the occurrence among adults is unknown. The aim of this study was to assess the clinical presentation and species distribution of HRV infections in different populations during 2001–2008. A total of 770 samples were collected. Subjects consisted of 136 adults from the general community and 207 health‐care workers (2001–2003), 232 renal‐transplanted outpatients (2002–2004), 70 children with congenital heart disease (2005) and 125 children from a day‐care center (2008). Amplification of HRV genes was performed by reverse transcriptase‐polymerase chain reaction (RT‐PCR) and followed by sequencing and phylogenetic analysis. HRV was detected in 27.4% of samples (211/770), with 72 children (36.9%) and 139 adults infected (24.2%). A total of 89.61% (138/154) unknown HRV strains were sequenced, and 79.22% (122/138) were analyzed. We identified 74 isolates (60.7%) of the HRV A species, 21 (17.2%) of the HRV B species and 27 isolates (22.1%) of the HRV C species. HRV species A and B caused ILI among adult patients, whereas HRV‐C did not. The dynamics of infection among different species deserve further analysis. J. Med. Virol. 82:2110–2115, 2010.

Selective regimes and evolutionary rates of HIV-1 subtype B V3 variants in the Brazilian epidemic

Half of subtype B Brazilian HIV-1 harbors the V3 tip GWGR instead of the GPGR. To investigate the evolution of GW variants, we analyzed 81 env sequences and 5 full-length GW genomes from antiretroviral-naive individuals sampled between 1983 and 1999. Phylogenetic analysis indicated that GW strains intermingle in the tree with other subtype B sequences. The mean d(N)/d(S) values of GW strains were proximal to those of the other sequences, regardless of sampling years or clinical status. In sequences from patients with CD4+ T cell counts >or=200 cells/microL, the mean d(N)/d(S) ratio was greater than one, suggesting a positive selection. The prevalence of GW variants was lower among individuals in whom disease progressed. This is probably attributable to the fact that tryptophan is replaced by other amino acids over time, whereas the GP motif does not evolve as rapidly.

Loci Polymorphisms of the APOBEC3G Gene in HIV Type 1-Infected Brazilians

The human APOBEC3G (A3G) protein activity obstructs retrovirus infection by inducing mutations of guanosines to adenosines (G → A) in the viral DNA. These G → A mutations may disrupt the reading frames of the viral genes. It has been observed that A3G polymorphisms can affect the degree of G → A mutations and the disease progression. For example, one study showed that the nonsynonymous substitution H186R was linked to AIDS progression in African Americans. Other studies, however, found no association between A3G polymorphisms and progression to AIDS in Europeans or in Asians. The genetic structure of a host population likely affects the dynamics of HIV-1 infection. The AIDS infection in Brazil is unique because of the high incidence of isolates with an unusual motif (GWGR) at the V3 region of the env gene. Since the Brazilian population is a mix of Portuguese, native Amerindians, and Africans we aimed to explore the influence of A3G polymorphisms in HIV-1 infection in this heterogeneous host population. We analyzed seven loci polymorphisms of A3G in 400 HIV-1-infected individuals naive to drug therapy. Our findings indicated no significant influence of A3G polymorphisms on disease status. The exception was the SNP -571 (rs5757463) in which heterozygous individuals (C/G) and homozygous individuals (G/G) presented lower CD4(+) T cell counts compared to homozygous (C/C) individuals (Mann-Whitney test p-value = 0.0076). Furthermore, the loci diversity of A3G in Brazilians was similar to that of Europeans. Consequently, if there is any host factor that could be used to explain the peculiar subtype B HIV-1 infection in Brazil it is not associated with the innate immunity of the A3G gene.

Molecular and structural characterization of HIV-1 subtype B Brazilian isolates with GWGR tetramer at the tip of the V3-loop

One of most intriguing features of the HIV-1 subtype B epidemic in Brazil is the high frequency of isolates exhibiting tryptophan (W) in the tetramer (GWGR) at the tip of the V3 loop. We observed that the frequencies of glutamic and aspartic acids at site 25 of the V3 loop are quite distinct in GWGR isolates compared with viruses with other tetramers. The basic amino acids at sites 11 and 25 of V3 are strongly linked with CCR5-to-CXCR4 coreceptor shift. We therefore predicted phenotype usage and found that GWGR isolates are exclusively CCR5-using. Further evidence of this came from intrahost sequences, where basic amino acid substitutions at sites 11 and 25 emerged only in isolates presenting a tryptophan-to-glycine replacement at the tetramer of the V3. In addition, modeled 3D-structures of the V3 loop of GWGR and GGGR in intrahost viruses differ essentially in the binding region of the coreceptor.

Viral diseases and human evolution

The interaction of man with viral agents was possibly a key factor shaping human evolution, culture and civilization from its outset. Evidence of the effect of disease, since the early stages of human speciation, through pre-historical times to the present suggest that the types of viruses associated with man changed in time. As human populations progressed technologically, they grew in numbers and density. As a consequence different viruses found suitable conditions to thrive and establish long-lasting associations with man. Although not all viral agents cause disease and some may in fact be considered beneficial, the present situation of overpopulation, poverty and ecological inbalance may have devastating effects on human progress. Recently emerged diseases causing massive pandemics (e.g., HIV-1 and HCV, dengue, etc.) are becoming formidable challenges, which may have a direct impact on the fate of our species.

Frequency of human rhinovirus species in outpatient children with acute respiratory infections at primary care level in Brazil.

This study assessed the occurrence of human rhinovirus (HRV) species in outpatient children attending day-care in Sao Paulo, Brazil. HRV reverse transcriptase polymerase chain reaction and amplicon sequencing were done in 120 samples collected in 2008. HRV was detected in 27.5% of samples. HRV C was detected in 60.7% of wheezers, a frequency not different from that observed in nonwheezers (69.6%).

Viral etiology among the elderly presenting acute respiratory infection during the influenza season

INTRODUCTION Acute respiratory tract infections are the most common illness in all individuals. Rhinoviruses have been reported as the etiology of more than 50% of respiratory tract infections worldwide. The study prospectively evaluated 47 elderly individuals from a group of 384 randomly assigned for acute respiratory viral infections (cold or flu) and assessed the occurrence of human rhinovirus (HRV), influenza A and B, respiratory syncytial virus and metapneumovirus (hMPV) in Botucatu, State of São Paulo, Brazil. METHODS Forty-nine nasal swabs collected from 47 elderly individuals following inclusion visits from 2002 to 2003 were tested by GenScan RT-PCR. HRV-positive samples were sequenced for phylogenetic analysis. RESULTS No sample was positive for influenza A/B or RSV. HRV was detected in 28.6% (14/47) and hMPV in 2% (1/47). Of 14 positive samples, 9 isolates were successfully sequenced, showing the follow group distribution: 6 group A, 1 group B and 2 group C HRVs. CONCLUSIONS The high incidence of HRV during the months of the influenza season requires further study regarding HRV infection impact on respiratory complications among this population. Infection caused by HRV is very frequent and may contribute to increasing the already high demand for healthcare during the influenza season.