Polymorphisms of CDKN1A gene and risk of retinoblastoma.

UNLABELLED: Retinoblastoma (RB) is a malignant neoplasia that occurs mostly in children under 5 years. Recently, CDKN1A gene has been shown to be up-regulated in a context of loss of function of pRb. This gene encodes the p21 protein, which is the bona fide effector of p53. We hypothesized whether two putatively functional single nucleotide polymorphisms (SNPs) of CDKN1A (rs1801270 C>A and rs1059234 C>T) may influence the risk and/or survival of RB patients. We genotyped both SNPs in 141 RB patients and 120 unrelated healthy individuals. Statistical analyses consisted of chi-square (χ(2)), odds ratio (OR) and survival curves by Kaplan-Meier method. We found that patients who carry the genotype CA for rs1801270 and CT for rs1059234 were associated to an increased risk of RB [OR = 2.5, 95% confidence interval (CI) = 1.38-4.53], whereas patients with CC for both polymorphisms were associated to a lower risk of developing RB (OR = 0.43, 95% CI = 0.25-0.74). On the other hand, Kaplan-Meier curves did not show statistically significant differences in survival among the studied polymorphisms. We conclude that the minor alleles of rs1801270 and rs1059234 polymorphisms may act as risk factors for the development of RB in our sample. SUMMARY: The minor alleles of polymorphisms rs1801270 C>A and rs1059234 C>T in CDKN1A (p21) gene may act as risk factors for the development of RB; however, they do not seem to influence overall survival.

Sequence analysis of NAT2 gene in Brazilians: identification of undescribed single nucleotide polymorphisms and molecular modeling of the N-acetyltransferase 2 protein structure.

N-Acetyltransferase 2 (NAT2) metabolizes a variety of xenobiotics that includes many drugs, chemicals and carcinogens. This enzyme is genetically variable in human populations and polymorphisms in the NAT2 gene have been associated with drug toxicity and efficacy as well as cancer susceptibility. Here, we have focused on the identification of NAT2 variants in Brazilian individuals from two different regions, Rio de Janeiro and Goiás, by direct sequencing, and on the characterization of new haplotypes after cloning and re-sequencing. Upon analysis of DNA samples from 404 individuals, six new SNPs (c.29T>C, c.152G>T, c.203G>A, c.228C>T, c.458C>T and c.600A>G) and seven new NAT2 alleles were identified with different frequencies in Rio de Janeiro and Goiás. All new SNPs were found as singletons (observed only once in 808 genes) and were confirmed by three independent technical replicates. Molecular modeling and structural analysis suggested that p.Gly51Val variant may have an important effect on substrate recognition by NAT2. We also observed that amino acid change p.Cys68Tyr would affect acetylating activity due to the resulting geometric restrictions and incompatibility of the functional group in the Tyr side chain with the admitted chemical mechanism for catalysis by NATs. Moreover, other variants, such like p.Thr153Ile, p.Thr193Met, p.Pro228Leu and p.Val280Met, may lead to the presence of hydrophobic residues on NAT2 surface involved in protein aggregation and/or targeted degradation. Finally, the new alleles NAT2*6H and NAT2*5N, which showed the highest frequency in the Brazilian populations considered in this study, may code for a slow activity. Functional studies are needed to clarify the mechanisms by which new SNPs interfere with acetylation.

Analysis of the DMPK gene CTG repeat in healthy Brazilians

Myotonic dystrophy (DM) is a neuromuscular disorder caused by the expansion of the cytosine-thymine-guanine (CTG) repeat of the myotonic dystrophy protein kinase gene (DMPK). This repeat is highly polymorphic in healthy individuals [(CTG)5-37], and it has been proposed that expanded CTG alleles originated from larger sized normal alleles [(CTG)19-37]. According to this hypothesis, a positive correlation should be expected between the frequency of these large-sized normal alleles and the prevalence of the disorder in a population. We determined the distribution of CTG alleles of the DMPK gene in 156 healthy Brazilians from Rio de Janeiro city. Our analyses of 312 chromosomes detected 20 different alleles ranging in size from 5 to 27 CTG repeats, with 24 alleles having more than 18 repeats (7.69 percent). This frequency of (CTG)3(19) alleles observed in our population suggests that the prevalence of DM in Rio de Janeiro should not be different from the prevalence in European populations.

Polymorphic markers suggest a gene flow of CFTR gene from Sub-Saharan/Arabian and Mediterranean to Brazilian Population.

The analysis of 2 diallelic loci (M470V and T854T) and a microsatellite IVS8(T)n of the cystic fibrosis transmembrane conductance regulator (CFTR) gene has shown different haplotype distribution in Brazilian cystic fibrosis (CF) chromosomes carrying different CF mutations. The DeltaF508 mutation was in absolute linkage disequilibrium with 1-1 haplotype (M470V-T854T). Most of DeltaF508 chromosomes (84%) were found to carry the IVS8-9T. The most frequent haplotypes IVS8-7T and 2-1 (M470V-T854T) were found associated with Non-DeltaF508 mutations. Although there is a remarkable linkage disequilibrium between these markers with CFTR locus, the mutations R334W (7T-1-2 and 7T-2-1) and the 3120 + 1G --> A (7T-1-2 and 9T-1-2) are associated with two different haplotypes probably introduced in the Brazilian population by migration. These findings suggest that recombination events from the original haplotype and gene flow among different ethnic groups (sub-Saharan and Mediterranean) might have resulted in CF mutations associated with different haplotypes by independent introductions.

Preliminary data on the performance of Aedes aegypti and Aedes albopictus immatures developing in water-filled tires in Rio de Janeiro

A monthly survey of Aedes aegypti and Aedes albopictus immatures in discarded tires at a site in metropolitan Rio de Janeiro showed that Ae. albopictus was much more abundant in the rainy season, but Ae. aegypti abundance showed a less clear seasonal pattern. Pupal masses for Ae. albopictus showed a seasonal trend. In contrast, Ae. aegypti pupae did not show any clear trend in weight. Large Ae. albopictus pupae were found in the warmer months, when water volume was higher, pH lower, and larval abundance lower. Further studies should be carried out to assess how seasonal variations in body size may impact vector competence of these species in Brazil.

Molecular analysis of 23 exons of the CFTR gene in Brazilian patients leads to the finding of rare cystic fibrosis mutations.

To define mutations present in 23 exons and flanking intronic sequences of the cystic fibrosis transmembrane conductance regulator (CFTR) gene in 95 patients from Rio de Janeiro, Brazil, we carried out single-strand conformation polymorphism analysis and automated direct sequencing. Mutation detection was achieved in 45% of the alleles presented, and complete genotyping (two mutated alleles) was accomplished in 34.7% of the patients. Twenty patients (21.1%) were found to carry only one mutation, whereas mutated alleles could not be observed in 42 patients (44.2%). Eleven mutations were found, of which four were characterized as rare mutations: P205S (1.05%), Y1092X (0.53%), S549R (0.53%), and S4X (0.53%). The DF508 mutation in this population sample showed a frequency of 28.42%. The low number of individuals (10 of 95; 10.5%) with compound heterozygous (DF508/non-DF508) genotypes could indicate the presence of another severe mutation leading to the premature death of these individuals. In 4 of the aforementioned 10 individuals with compound heterozygous genotypes, the D-7-2-1-2 (XV2c-KM19-IVS6a-TUB9-M470-T854) haplotype was defined.

Haplotype distribution of and linkage disequilibrium between four polymorphic markers near the CFTR locus in Brazilian cystic fibrosis patients.

To contribute to a better understanding of the origin and distribution of CFTR mutations in the Brazilian population, we have investigated the linkage between four polymorphic markers (XV2c, KM19, GATT, and TUB9) within or near the CFTR locus. The distribution of alleles for each polymorphism for both parental and cystic fibrosis (CF) chromosomes from Rio de Janeiro CF families were ascertained using a maximum-likelihood method. This same method was applied to study the distribution of the haplotypes defined by these markers. There was no significant association between the XV2c and KM19 loci on the parental and CF chromosomes. On the other hand, a strong association between GATT and TUB9 loci was observed on both CF and parental chromosomes, and striking linkage disequilibrium between the GATT-TUB9 pair and deltaF508 was observed (chi2 = 26.48, p < 0.0001). Remarkable linkage disequilibrium between the GATT-TUB9 marker pair and non-deltaF508 was also found (chi2 = 17.05, p < 0.0001). Our finding of a linkage disequilibrium between GATT-TUB9 and the CFTR locus could suggest that gene flow between different ethnic groups, mainly sub-Saharan and Mediterranean populations, with Brazilian populations could have resulted in some CF mutations originating on chromosomes that carried the GATT-TUB9 marker haplotype 7-2 (OR = 1.34 < 2.83 < 6.00; p = 0.0066).

Rastreamento da fibrose cística usando-se a análise combinada do teste de IRT neonatal e o estudo molecular da mutaçäo deltaF508 / Cystic fibrosis screening by two-tiered newborn IRT assay and deltaF508 mutation molecular analysis

Um total de 117 cartöes de rastreamento neonatal foi selecionado anonimamente para a avaliaçäo de fibrose cística (FC) pela análise da mutaçäo deltaF508 usando-se a técnica da reaçäo em cadeia da polimerase (PCR), seguida de eletroforese em gel de poliacrilamida (Page) e pela quantificaçäo da imunotripsina reativa (IRT, Delfia). Uma concentraçäo de IRT menor que 140ng/ml foi encontrada em 116 recém-nascidos. Entre estes foi detectado um heterozigoto deltaF508 com uma concentraçäo de IRT de 4,44ng/ml. Um dos 117 recém-nascidos era homozigoto näo-deltaF508 e apresentava uma IRT anormal de 410,7ng/ml. A média da concentraçäo da IRT diferia significativamente conforme este recém-nascido com IRT alterada era incluído ou excluído da amostragem populacional (n = 117, média = 8,207 ± 38,101; n = 116, média = 4,737 ± 6,597, respectivamente). Outra amostra de oito recém-nascidos, previamente rastreados pelo teste de IRT e com níveis elevados do analito, foi testada para a mutaçäo deltaF508. Em cinco deles a mutaçäo deltaF508 foi encontrada em um ou em ambos os cromossomos, correspondendo a 62,25 por cento dos recém-nascidos. De acordo com os resultados obtidos com triagem neonatal pela análise combinada IRT/DNA, pode-se concluir que o método só será eficiente se: a) forem excluídos os fatores que determinam os falso-positivos ou falso-negativos; e b) a detecçäo de outras mutações forem incluídas na análise dos resultados duvidosos

Seroprevalence of Hepatitis B virus infection among an afro-descendant community in Brazil

Furnas dos Dionísios is an Afro-Brazilian black community whose descendants were mainly fugitive slaves that established themselves in the State of Mato Grosso do Sul (MS), Brazil. The population is comprised mainly of low socioeconomic individuals who are engaged in agricultural activities. The objective of this study was to investigate the prevalence of hepatitis B (HB) and its correlation with epidemiological data obtained from the community. The studied population totaled 260 individuals with ages varying from 1 to 79 years (median 20). One hundred thirty-three (51.2 percent) were females and 127 (48.8 percent) were males. A high prevalence for anti-HBc was observed (42.7 percent), with present infection detected in 9.2 percent of the subjects who were also HB surface antigens (HBs Ag) positive; 27.3 percent were anti-HBc and anti-HBs reactive, and 6.2 percent had anti-HBc as only marker. The prevalence for anti-HBc was proportional to age, reaching its highest peak in age categories greater than 50. No serological marker was detected in children under the age of 2 years, however anti-HBc was present in 12 subjects with ages between 2 and 14 years, of these 8 (7.4 percent) were HBsAg positive. Among individuals over the age of 15 years, 99 were anti-HBc reactive, of these 16 (10.5 percent) were also HBsAg positive, thus suggesting an increased prevalence of HBV carriers among children and adolescents. The risk factors observed in this community that were significantly associated with anti-HBc positivity were age (over 20 years) and having an anti-HBc positive mother. Both HBeAg and anti-HBe were detected in 44.4 percent of the samples tested. HBsAg subtypes found in the studied population were adw2 (77.7 percent) and ayw2 (23.3 percent). While intrafamilial transmission was most likely responsible for HBV infection among children, other routes such as sexual contact might be considered for individuals with ages over 15 years

Role of tumor necrosis factor-alpha and interleukin-10 promoter gene polymorphisms in leprosy.

Single-nucleotide polymorphisms within the genes coding for tumor necrosis factor (TNF)-alpha and interleukin (IL)-10 have been associated with several infectious diseases. To determine whether such polymorphisms are associated with leprosy, genotyping was performed at the -308 and -238 positions of the promoter of the TNF-alpha gene in 210 and 191 patients with multibacillary (MB) leprosy, respectively; 90 and 79 patients with paucibacillary (PB) leprosy; and 92 control subjects. For the -592 and -819 positions within the promoter of the IL-10 gene, 143 patients with MB leprosy, 79 patients with PB leprosy, and 62 control subjects were included in the analysis. TNF2 allele frequency was significantly higher among control subjects than among all patients with leprosy or in the MB group (P<.05 and P<.01). For the IL-10 gene, the frequency of the homozygous -819TT genotype was significantly higher among patients than among control subjects. These data indicate that a relationship exists between TNF-alpha and IL-10 promoter polymorphisms and the development of PB leprosy.

Studies on populations of Lutzomyia longipalpis (Lutz & Neiva, 1912) (Diptera: Psychodidae: Phlebotominae) in Brazil

Studies were performed on five Brazilian populations of Lutzomyia longipalpis: Salvaterra (PA), São José do Ribamar (MA), Canindé (CE), Natal (RN) and Gruta da Lapinha, Lagoa Santa (MG). No morphological differences were observed that could distinguish between these populations. Homogeneity tests showed that the allopatric populations display a certain heterogeneity and that the sympatric populations, with different patterns of spots, are homogeneous. The Student-Newman-Keuls test, represented by Euler-Venn diagrams, showed a disjunction between the populations from the north/northeast and the one from Gruta da Lapinha. Genetic distances between the four populations (excluding the Canindé population) were within the range of intrapopulational differences. The Gruta da Lapinha population displayed a heterozygotic deficiency that could be a consequence of high levels of inbreeding due to cryptic habits of living in a small cave. These results do not favor the hypothesis of a L. longipalpis species complex in Brazil, and the species should be considered high polymorphic.