Although HCV has hepatic tropism, the presence of the virus in extra-hepatic compartments has been well documented. Platelets have been described as carriers of the virus in the circulation and may be a natural reservoir for the virus. However, few studies have been performed to evaluate the levels of HCV RNA in plasma and platelets are equal or differ in some way. Therefore, the aim of this study was to perform a comparative evaluation of the stability of HCV RNA in plasma and isolated platelets. Four aliquots of whole plasma obtained from patients infected with HCV were incubated at 37 °C for 0, 48, 96 and 144 h. After incubation, the plasma and platelet pellet was obtained from each aliquot. Viral RNA in plasma and platelets was quantified by q-PCR. The results showed a decrease in HCV RNA levels in plasma with incubation time. However, platelet HCV RNA levels were stable up to 144 h incubation. The results of this study showed that HCV RNA in platelets, although at lower concentrations than in plasma, is preserved from degradation over time, suggesting that the virus may persist longer in the body when associated with platelets, which could have an impact on the efficiency of antiviral therapy.

Several reports showed outbreaks of histoplasmosis acquired while bat-inhabited caves were visited by tourists, miners or researchers. We evaluated the performance of double immunodifusion (DI) and immunoblotting (IB) assays, employed for the histoplasmosis outbreak elucidation occurred in Vale do Paraíba, São Paulo. The existence of epidemiologic link, four patients with clinical signs suggestive of histoplasmosis and mycological confirmation has made that all 35 individuals involved to the cave visit were subjected to serological evaluation. By DI, we observed reactivity against H. capsulatum antigen in a single serum examined nearly 20 days after exposure to fungal propagules. On the other hand, IB showed reactivity against H and M fractions in 50% of samples evaluated. The analysis of the second sample batch, collected two months after the exposure showed that 96.7% were reactive by DI with antibodies titers ranging from 1 to 16 and 100% of reactivity against H and M fractions, by IB, suggesting an acute infection. The analysis of the overall agreement between the methods showed to be reasonable (κ = 0.37). This study confirms the importance and efficacy of more sensitive methodologies, such as IB assay, to early elucidation of disease, especially in cases of patients without mycological information.

This case reports an immunocompetent 29-year-old woman with suspected pneumonia, suggestive of fungal infection. Immunoblotting analysis reactivity againstHistoplasma capsulatum and Paracoccidioides brasiliensis were observed. Nested-PCR in blood employing species-specific primers was positive for H. capsulatum andCryptococcus neoformans. The evaluation of paucisymptomatic patients with positive results for H. capsulatum and C. neoformans could be relevant for the prevention as well as the possible evaluation of the reactivated quiescent foci. In conclusion, the associated methodology may have contributed to the monitoring endogenous reactivation of these diseases.

O objetivo do estudo consistiu em detectar seqüências no ADNr e as suas regiões no Histoplasma capsulatum, que pudessem ser consideradas espécie-específicas e usadas como método molecular para o diagnóstico pela técnica da reação em cadeia da polimerase aninhada ("nested PCR") com seqüências específicas ("primers") para H. capsulatum: regiões 18S ADNr (HC18), 100kDa (HC100) e a seqüência 5.8 S ADNr-ITS (HC5.8). A "nested PCR" com as seqüências HC18, HC100 e HC5.8 resultaram em 100% de especificidade. Os ensaios moleculares podem aumentar a especificidade, sensibilidade e rapidez na diagnose da Histoplasmose.

The aim of the study was to detect the rDNA sequences and their regions in Histoplasma capsulatum, which could be considered species-specific and used as a molecular method for this diagnosis by the technique of nested polymerase chain reaction (nested PCR), employing specific sequences (primers) for H. capsulatum: 18S rDNA region (HC18), 100 kDa (HC100) and the sequence 5.8 S-ITS rDNA (HC5.8). The PCR sequences HC18, HC100 and HC5.8 resulted in a specificity of 100%. The molecular assays may increase the specificity, sensitivity and speed in the diagnosis of Histoplasmosis.

CONTEXTO:O câncer gástrico é uma das principais neoplasias que causam o óbito no Brasil e no mundo. Helicobacter pylori é um carcinógeno do tipo I relacionado à gastrite crônica. Diferenças no grau de virulência de suas cepas levam a maior risco de desenvolvimento de doenças gástricas. A metilação de ilhas CpGs está envolvida com o processo de tumorigênese em diferentes tipos de câncer. CDH1 é um gene supressor tumoral que, quando inativado, pode aumentar as chances de metástase. A metilação deste gene em estágios precoces da carcinogênese gástrica ainda não é totalmente compreendida. OBJETIVO: Investigar o padrão de metilação do gene CDH1 em amostras de gastrites crônicas e correlacionar com a presença do H. pylori. MÉTODOS: Foram usadas 60 biopsias de mucosas gástricas. A detecção de H. pylori foi realizada por PCR para o gene da urease C e a genotipagem com PCR para os genes cagA e vacA (região s e m). O padrão de metilação do gene CDH1 foi analisado usando a técnica de PCR e específica para a metilação e sequenciamento direto dos produtos de PCR. RESULTADOS: A bactéria H. pylori foi detectada em 90% das amostras de gastrites crônicas; destas, 33% portavam o gene cagA e 100% vacA s1. O genótipo vacA s2/m1 não foi detectado nas amostras analisadas. Metilação de CDH1 foi detectada em 63,3% das amostras de gastrites e 95% delas eram portadoras de H. pylori. CONCLUSÃO: Os resultados deste estudo sugerem que a metilação em CDH1 e a infecção pelo H. pylori são eventos frequentes em amostras de pacientes brasileiros com gastrite crônica e reforça a correlação entre infecção por H. pylori e inativação do gene CDH1 em estágios precoces da tumorigênese gástrica.

CONTEXT: Gastric cancer is one of the top list of cancer types that most leads to death in Brazil and worldwide. Helicobacter pylori(H. pylori) is a class I carcinogen and infect almost 90% of chronic gastritis patients. Some genotypes confer different virulent potential to H. pylori and can increase the risk of gastritis development. Methylation of CpG islands can inactivate tumor suppressor genes and therefore, it can be involved in the tumorigenic process. CDH1 is a tumor suppressor gene that encodes the E-cadherin protein, which is important in maintaining cell-cell contacts. The inactivation of this gene can increase the chance of metastasis. Promoter methylation of CDH1 at early steps of gastric carcinogenesis is not yet completely understood. OBJECTIVE: In this study, we investigated the methylation status of CDH1 in chronic gastritis samples and correlated it with the presence of H. pylori. METHODS: Sixty gastric mucosal biopsies were used in this study. The detection of H. pylori was performed with the PCR primers specific to urease C gene. H. pylori genotyping was performed by PCR to cagA and vacA (s and m region). The methylation status of these gene CDH1 was analyzed using methylation-specific polymerase chain reaction and direct sequencing of the PCR products was performed using primers methylated and unmethylated in both forward and reverse directions. RESULTS: H. pylori was detected in 90% of chronic gastritis samples; among these 33% were cagA positive and 100% vacA s1. The genotype vacA s2/m1 was not detected in any sample analyzed. Methylation of CDH1 was detected in 63.3% of chronic gastritis samples and 95% of them were also H. pylori-positive. CONCLUSION: This work suggests that CDH1 gene methylation and H. pylori infection are frequent events in samples from Brazilian patients with chronic gastritis and reinforces the correlation between H. pylori infection and CDH1 inactivation in early steps of gastric tumorigenesis.

INTRODUÇÃO: Apesar da alta freqüência de infecção por Helicobacter pylori na população, somente uma minoria de indivíduos desenvolve câncer gástrico. É provável que a colonização da mucosa por cepas patogênicas, levando a maior agressão e inflamação da mucosa seja um dos elos da cadeia de eventos da oncogênese gástrica. OBJETIVOS: Investigar a freqüência de cepas patogênicas cagA e vacA do H. pylori em pacientes com câncer gástrico. MATERIAL E MÉTODOS: Foram estudados retrospectivamente 42 pacientes com câncer gástrico. A infecção por H. pylori foi avaliada por exame histológico e pelo PCR para identificação dos genótipos cagA e vacA em amostras de material fixado em formalina e incluído em parafina. RESULTADOS: A análise histológica permitiu a visualização direta do H. pylori em 85,7% dos casos, e o método de PCR para o gene urease C demonstrou a presença de DNA da bactéria em 95% dos casos. O gene cagA foi detectado em amostras de 23 pacientes (54,7%) com câncer gástrico. O alelo s1 do gene vacA foi identificado em amostras de 24 pacientes (57,1%) e o alelo m1, em amostras de 26 pacientes (61,9%). Os alelos s1 e m1 foram identificados simultaneamente em 24 pacientes (57,1%). O alelo s2 foi identificado em amostras de quatro pacientes (9,5%), e o alelo m2, em amostras de três pacientes (7,1%). A freqüência de infecção pelo Helicobacter pylori foi similar em ambos os tipos histológicos de câncer gástrico (intestinal e difuso). CONCLUSÕES: Os resultados confirmam a relevância dos genótipos patogênicos cagA e vacA do H. pylori para lesões orgânicas significativas tais como o câncer gástrico, sugerindo a participação dessa bactéria na cadeia de eventos da oncogênese gástrica.

BACKGROUND: The rates of Helicobacter pylori infection are very high worldwide, but only a minority of infected patients develop gastric carcinoma. This might be related, among several factors, to the colonization of the human stomach by pathogenic Helicobacter pylori strains. OBJECTIVE: To investigate the distribution of cagA and vacA genotypes of Helicobacter pylori in paraffin-embedded gastric samples from patients with gastric cancer. MATERIAL AND METHODS: Paraffin-embedded samples from 42 patients with gastric cancer were histologically examined and evaluated by PCR for H. pylori cagA and vacA (s and m regions) genotypes. RESULTS: Histological analysis allowed direct visualization of H. pylori in 85.7% of cases and PCR for urease C gene detected H. pylori in 95% of cases. The presence of cagA gene was detected in 23 (54.7%) patients with gastric cancer. The s1 allele from vacA gene was found in samples from 24 (57.1%) patients and the m1 allele in 26 (61.9 %). The s1m1 genotype was detected in 24 (57.1%) patients with gastric cancer. The s2 allele was found in samples from four patients (9.5%) and the m2 allele in three (7.1%) patients. The distribution of H. pylori genotypes was similar in both intestinal and diffuse types of gastric carcinoma. CONCLUSION: Our results confirm the relevance of the pathogenic cagA and vacA H. pylori genotypes for significant organic lesions, such as gastric cancer, suggesting a possible role for H. pylori in the pathogenesis of gastric carcinoma.

O fator de Leiden é uma mutação genética que predispõe seus portadores ao tromboembolismo venoso. O objetivo do estudo foi investigar a distribuição dos alelos em 21 membros da família de três pacientes portadores de trombose com a presença da mutação do fator V de Leiden. A detecção da mutação no gene do fator V foi realizada entre portadores da mutação no estado heterozigoto. Este estudo foi realizado no Centro de Hematologia e Hemoterapia do Ceará - Hemoce. Observou-se a presença da mutação no estado heterozigoto na família 1 (83,3%), na família 2 (40%) e na família 3 (50%). No total de 24 membros (pacientes e familiares) analisados, 50% (12/24) apresentaram a mutação, todos no estado heterozigoto, 66,7% (8/12) não apresentaram trombose. A detecção do fator V de Leiden em pacientes portadores de eventos trombóticos é recomendado para esclarecimento das causas e para efetuar o rastreamento em membros de sua família, ainda sem o aparecimento de eventos trombóticos, de forma a avaliar os riscos associados e assim determinar um acompanhamento médico preventivo.

Factor V Leiden is a mutation that can cause venous thrombosis. When associated to other risk factors such as the use of contraceptives, important surgical intervention, pregnancy and malignant diseases, the risk for heterozygous carriers increases by three-fold to 10 times or even 80 times for two mutated alleles. The factor V Leiden is found in about 20% of the population with a history of venous thromboembolism. It is present in about 4 to 6% of general population but this percentage changes depending on the ethnicity. This study shows the distribution of alleles in family members of three carriers of factor V Leiden diagnosed with deep venous thrombosis. The mutation investigation of the factor V Leiden gene was performed in 21 family members of 3 heterozygous carriers. The study was performed in the Hematology and Hemotherapy Center from Ceará - Hemoce, Brazil. In this study the V Leiden mutation in its heterozygous form was observed in 83.3% of family 1, 40% of family 2 and 50% of family 3. From the total of 24 members analyzed 50% (12/24) showed the mutation and 66.7% (8/12) did not present the thrombotic disease. An investigation of the factor V Leiden in patients suffering thrombotic events is recommended to explain the etiology and effect of the disease as well as screening of family members so that preventative actions can be adopted.

Type 1 diabetes, as an autoimmune disease, presents several islet cell-specific autoantibodies such as islet cell antibody (ICA), anti-insulin, anti-glutamic acid decarboxylase (GAD) and the antibody (Ab) against tyrosine phosphatase (PTP)-like protein known as ICA-512 (IA-2). In order to determine the frequency of the anti-GAD and anti-IA-2 autoantibodies in Brazilian type 1 diabetes patients we studied 35 diabetes mellitus (DM) type 1 patients with recent-onset disease (<FONT FACE="Symbol">£</font>12 months) and 37 type 1 diabetes patients with long-duration diabetes (>12 months) who were compared to 12 children with normal fasting glucose. Anti-GAD65 and anti-IA-2 autoantibodies were detected with commercial immunoprecipitation assays. The frequency of positive results in recent-onset DM type 1 patients was 80.0% for GADAb, 62.9% for IA-2Ab and 82.9% for GADAb and/or IA-2Ab. The long-duration type 1 diabetes subjects presented frequencies of 54.1% for GADAb and IA-2Ab, and 67.5% for GAD and/or IA-2 antibodies. The control group showed no positive cases. Anti-GAD and IA-2 assays showed a high frequency of positivity in these Brazilian type 1 diabetes patients, who presented the same prevalence as a Caucasian population.

We have investigated the relationship between fetal hemoglobin (HbF) levels and metabolic control in subjects with insulin-dependent (N = 79) and non-insulin-dependent diabetes mellitus (N = 242). HbF and hemoglobin A1c (HbA1c) levels were increased in subjects with type 1 and type 2 diabetes as compared to levels in nondiabetic individuals (P<0.0001), and were significantly higher in type 1 than in type 2 diabetes subjects. Lower levels of HbA1c and HbF were observed in type 2 diabetes subjects treated by diet, intermediate levels in those treated with oral hypoglycemic agents, and higher levels in those treated with insulin. HbF and HbA1c levels were correlated in type 1 diabetes (R2 = 0.57, P<0.0001) and type 2 diabetes (R2 = 0.58, P<0.0001) subjects. Following intense treatment, twelve diabetic patients showed significant improvement both in HbA1c and HbF values. We conclude that increased HbF levels reflect poor metabolic control in subjects with diabetes mellitus.

Glucokinase (GCK) is an enzyme that regulates insulin secretion, keeping glucose levels within a narrow range. Mutations in the glucokinase gene cause a rare form of diabetes called maturity-onset diabetes of the young (MODY). An early onset (less than 25 years), autosomal dominant inheritance and low insulin secretion stimulated by glucose characterize MODY patients. Specific insulin and proinsulin were measured in serum by immunofluorimetric assays (IFMA) during a 75-g oral glucose tolerance test (OGTT). Two kindreds (SA and LZ) were studied and compared to non-diabetic unrelated individuals (control group 1) matched for age and body mass index (BMI). In one kindred, some of these subjects were also obese (BMI >26 kg/m2), and other family members also presented with obesity and/or late-onset NIDDM. The MODY patients were also compared to a group of five of their first-degree relatives with obesity and/or late-onset NIDDM. The proinsulin profile was different in members of the two MODY kindreds. Fasting proinsulin and the proinsulin/insulin ratio were similar in MODY members of kindred LZ and subjects from control group 1, but were significantly lower than in MODY members of kindred SA (P<0.02 and P<0.01, for proinsulin and proinsulin/insulin ratio, respectively). Moreover, MODY members of family SA had higher levels of proinsulin and proinsulin/insulin ratio, although not significantly different, when compared to their first-degree relatives and to subjects from control group 2. In conclusion, we observed variable degrees of proinsulin levels and proinsulin/insulin ratio in MODY members of two different kindreds. The higher values of these parameters found in MODY and non-MODY members of kindred SA is probably related to the obesity and late-onset NIDDM background present in this family.