Sequence and Structural Analysis of the 5 ` Noncoding Region of Hepatitis C Virus in Patients With Chronic Infection

Hepatitis C virus (HCV), exhibits considerable genetic diversity, but presents a relatively well conserved 5 ` noncoding region (5 ` NCR) among all genotypes. In this study, the structural features and translational efficiency of the HCV 5 ` NCR sequences were analyzed using the programs RNAfold, RNAshapes and RNApdist and with a bicistronic dual luciferase expression system, respectively. RNA structure prediction software indicated that base substitutions will alter potentially the 5 ` NCR structure. The heterogeneous sequence observed on 5 ` NCR led to important changes in their translation efficiency in different cell culture lines. Interactions of the viral RNA with cellular transacting factors may vary according to the cell type and viral genome polymorphisms that may result in the translational efficiency observed. J. Med. Virol. 81: 1212-1219, 2009. (C) 2009 Wiley-Liss, Inc.

Modulation of tumor necrosis factor receptors 1 and 2 in chronic hepatitis B and C: The differences and implications in pathogenesis

Tumor necrosis factor (TNF) plays a role in the pathogenesis of chronic hepatitis B (CHB) and chronic hepatitis C (CHC). The difference in the cytokine responses between hepatitis B virus (HBV) and hepatitis C virus (HCV) infections may have implications in the pathogenesis of these diseases. We performed a comparative study to examine the possible differences in the TNF-TNF receptor (TNFR) response between CHB and CHC. We studied the cytokine levels of 38 patients with CHB, 40 patients with CHC and 9 patients with dual hepatitis B and C, and compared them with the baseline levels of 12 healthy controls. The plasma levels of TNF-, interferon-, interleukin (IL)-2, IL-4, IL-10 and soluble TNFR-1 and 2 (sTNFR-1 and 2) were quantified by enzyme-linked immunosorbent assays. The expression of TNFR-1 and 2 in liver tissues was examined in 30 cases of CHB and 15 cases of CHC by semiquantitative reverse transcription polymerase chain reaction. The results showed that sTNFR-1 levels correlated with liver inflammation in all patients, whereas this correlation was not found with sTNFR-2 or other cytokines. Liver inflammation indicators were higher in HCV RNA+ than in HCV RNA– CHC. Most significantly, sTNFR-1 levels correlated with liver inflammation in CHB, but not in CHC. However, the expression of TNFR-1 and 2 in liver was similar between CHB and CHC. These findings suggest that the TNFR signal transduction pathway is modulated differently in HBV and HCV infection.

Effect of bronchoalveolar lavage-directed therapy on pseudomonas aeruginosa infection and structural lung injury in children with cystic fibrosis

Context Early pulmonary infection in children with cystic fibrosis leads to increased morbidity and mortality. Despite wide use of oropharyngeal cultures to identify pulmonary infection, concerns remain over their diagnostic accuracy. While bronchoalveolar lavage (BAL) is an alternative diagnostic tool, evidence for its clinical benefit is lacking.

Objective To determine if BAL-directed therapy for pulmonary exacerbations during the first 5 years of life provides better outcomes than current standard practice relying on clinical features and oropharyngeal cultures.

Design, Setting, and Participants The Australasian Cystic Fibrosis Bronchoalveolar Lavage (ACFBAL) randomized controlled trial, recruiting infants diagnosed with cystic fibrosis through newborn screening programs in 8 Australasian cystic fibrosis centers. Recruitment occurred between June 1, 1999, and April 30, 2005, with the study ending on December 31, 2009.

Interventions BAL-directed (n=84) or standard (n=86) therapy until age 5 years. The BAL-directed therapy group underwent BAL before age 6 months when well, when hospitalized for pulmonary exacerbations, if Pseudomonas aeruginosa was detected in oropharyngeal specimens, and after P aeruginosa eradication therapy. Treatment was prescribed according to BAL or oropharyngeal culture results.

Main Outcome Measures Primary outcomes at age 5 years were prevalence of P aeruginosa on BAL cultures and total cystic fibrosis computed tomography (CF-CT) score (as a percentage of the maximum score) on high-resolution chest CT scan.

Results Of 267 infants diagnosed with cystic fibrosis following newborn screening, 170 were enrolled and randomized, and 157 completed the study. At age 5 years, 8 of 79 children (10%) in the BAL-directed therapy group and 9 of 76 (12%) in the standard therapy group had P aeruginosa in final BAL cultures (risk difference, −1.7% [95% confidence interval, −11.6% to 8.1%]; P=.73). Mean total CF-CT scores for the BAL-directed therapy and standard therapy groups were 3.0% and 2.8%, respectively (mean difference, 0.19% [95% confidence interval, −0.94% to 1.33%]; P=.74).

Conclusion Among infants diagnosed with cystic fibrosis, BAL-directed therapy did not result in a lower prevalence of P aeruginosa infection or lower total CF-CT score when compared with standard therapy at age 5 years.

Trial Registration anzctr.org.au Identifier: ACTRN12605000665639

Influenza A virus infection impairs mycobacteria-specific T cell responses and mycobacterial clearance in the lung during pulmonary coinfection.

Individuals infected with mycobacteria are likely to experience episodes of concurrent infections with unrelated respiratory pathogens, including the seasonal or pandemic circulating influenza A virus strains. We analyzed the impact of influenza A virus and mycobacterial respiratory coinfection on the development of CD8 T cell responses to each pathogen. Coinfected mice exhibited reduced frequency and numbers of CD8 T cells specific to Mycobacterium bovis bacille Calmette-Guérin (BCG) in the lungs, and the IFN-γ CD8 T cell response to BCG-encoded OVA was decreased in the lungs of coinfected mice, when compared with mice infected with BCG alone. Moreover, after 2 wk of infection, mice coinfected with both pathogens showed a significant increase in the number of mycobacteria present in the lung compared with mice infected with BCG only. Following adoptive transfer into coinfected mice, transgenic CD8 T cells specific for OVA257–264 failed to proliferate as extensively in the mediastinal lymph nodes as in mice infected only with BCG-OVA. Also noted was a reduction in the proliferation of BCG-specific CD4 transgenic T cells in mice coinfected with influenza compared with mice infected with BCG alone. Furthermore, phenotypic analysis of CD11c+ dendritic cells from mediastinal lymph nodes of the infected mice showed that coinfection was associated with decreased surface expression of MHC class II and class I. Thus, concurrent pulmonary infection with influenza A virus is associated with decreased MHC expression on dendritic cells, reduced activation of BCG-specific CD4 and CD8 T cells, and impaired clearance of mycobacteria.

Entecavir for the treatment of chronic hepatitis B infection

This paper presents a summary of the evidence review group (ERG) report into the clinical and cost-effectiveness of entecavir for the treatment of chronic hepatitis B (CHB) in adults based upon a review of the manufacturer's submission to the National Institute for Health and Clinical Excellence (NICE) as part of the single technology appraisal (STA) process. The submission's evidence came from five randomised controlled trials (RCTs), of good methodological quality and measuring a range of clinically relevant outcomes, comparing entecavir with lamivudine. After 1 year of treatment entecavir was statistically superior to lamivudine in terms of the proportion of patients achieving hepatitis B virus (HBV) DNA suppression, alanine aminotransferase (ALT) normalisation and histological improvement, but not in terms of the proportion of patients achieving hepatitis B e antigen (HBeAg) seroconversion. The incidence of adverse or serious adverse events was similar for both treatments. The results of the manufacturer's mixed treatment comparison (MTC) model to compare entecavir with the comparator drugs in nucleoside-naive patients were considered to be uncertain because of concerns over its conduct and reporting. For the economic evaluation the manufacturer constructed two Markov state transition models, one in HBeAg-positive and one in HBeAg-negative patients. The modelling approach was considered reasonable subject to some uncertainties and concerns over some of the structural assumptions. In HBeAg-positive patients the base-case incremental cost-effectiveness ratios (ICER) for entecavir compared with lamivudine and pegylated interferon alpha-2a were 14,329 pounds and 8403 pounds per quality-adjusted life-year (QALY) respectively. Entecavir was dominated by telbivudine. In HBeAg-negative patients the base-case ICERs for entecavir compared with lamivudine, pegylated interferon alpha-2a and telbivudine were 13,208 pounds, 7511 pounds and 6907 pounds per QALY respectively. In HBeAg-positive lamivudine-refractory patients entecavir dominated adefovir added to lamivudine. In one-way deterministic sensitivity analysis on all key input parameters for entecavir compared with lamivudine in nucleoside-naive patients, ICERs generally remained under 30,000 pounds per QALY. In probabilistic sensitivity analysis in nucleoside-naive HBeAg-positive patients the probability of the ICER for entecavir being below 20,000 pounds per QALY was 57%, 82% and 45% compared with lamivudine, pegylated interferon alpha-2a and telbivudine respectively. In nucleoside-naive HBeAg-negative patients the probabilities were 90%, 100% and 96% respectively. The manufacturer's lifetime treatment scenario for HBeAg-negative patients and the ERG's 20-year treatment scenario for HBeAg-positive patients increased the ICERs, particularly in the latter case. Amending the HBeAg-negative model so that patients with compensated cirrhosis would also receive lifetime treatment gave probabilities of entecavir being cost-effective at a willingness to pay of 20,000 pounds and 30,000 pounds of 4% and 40% respectively. The NICE guidance issued in August 2008 as a result of the STA states that entecavir is recommended as an option for the treatment of people with chronic HBeAg-positive or HBeAg-negative hepatitis B in whom antiviral treatment is indicated.

Acute phase proteins: a potential approach for diagnosing chronic infection by Trypanosoma vivax

O objetivo do presente estudo foi verificar possíveis alterações nas proteínas de fase aguda em ovinos infectados experimentalmente com Trypanosoma vivax. Para tanto, foram utilizados oito ovinos machos, sendo quatro usados como controle e quatro infectados com 10(5) tripomastigotas de T. vivax. Colheram-se amostras de sangue em dois tempos antes da infecção e, posteriormente, aos 5, 7, 9, 11, 13, 15, 20, 30, 45, 60, 75, 90, 105 e 120 dias após a infecção (dpi); após centrifugação e aliquotização das amostras. As proteínas de fase aguda foram separadas por eletroforese em gel de acrilamida, contendo dodecil sulfato de sódio, e suas concentrações foram determinadas através de densitometria computadorizada. A dosagem de proteína total foi realizada pelo método colorimétrico do biureto. A contagem dos tripanossomas foi realizada diariamente, utilizando-se uma alíquota de 5 µL de sangue disperso em lâmina de microscopia, sob lamínula de 22 × 22 mm, contando-se os parasitos em 100 campos microscópicos, com objetiva de 40×, multiplicados pelo fator de correção do microscópio, e o resultado expresso em parasitos por mL de sangue. Para a análise estatística, empregou-se o teste de Wilcoxon a 5% de probabilidade. Foi observada a diminuição de diversas proteínas de fase aguda e aumento de antitripsina e transferrina que podem ser utilizadas para auxiliar no diagnóstico da infecção por T. vivax, principalmente na fase crônica da infecção.

Astrocytic and microglial response and histopathological changes in the brain of horses with experimental chronic Trypanosoma evansi infection

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Immunohistochemical characterization of mononuclear cells and MHC II expression in the brain of horses with experimental chronic Trypanosoma evansi infection

Este estudo objetivou caracterizar a resposta imune celular no sistema nervoso central (SNC) de eqüinos com infecção crônica experimental por Trypanosoma evansi. Para este propósito, foram utilizados os métodos histoquímicos (HE) e imunoistoquímicos do complexo avidina-biotina peroxidase (ABC). O fenótipo do infiltrado celular foi caracterizado com o auxílio de anticorpos anti - CD3, para linfócitos T e antiBLA36 para linfócitos B. Os macrófagos foram marcados com anticorpo antiantígenos da linhagem mielóide/histiócitos (Clone Mac387). A lesão no sistema nervoso central (SNC) dos eqüinos infectados com T. evansi foi caracterizada como meningoencefalite e meningomielite não supurativa. A gravidade das lesões variou em diferentes segmentos do SNC, refletindo distribuição irregular das alterações vasculares. A distribuição de células T e B e antígenos do complexo maior de histocompatibilidade classe II foram avaliados dentro do SNC de eqüinos cronicamente infectados com T. evansi. O infiltrado perivascular e meníngeo eram constituídos predominantemente por células T e B. Macrófagos foram raramente visualizados. T.evansi não foi identificado no parênquima do SNC dos eqüinos.

Influence of Helicobacter pylori infection on the expression of MLH1 and MGMT in patients with chronic gastritis and gastric cancer

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Effect of Helicobacter pylori infection on IL-8, IL-1 and COX-2 expression in patients with chronic gastritis and gastric cancer

Objective. Helicobacter pylori infection is related to gastric cancer development, and chronic inflammation is presumed to be the main cause. The aim of the present study was to evaluate the influence of H. pylori cagA, vacA, iceA, and babA genotypes on COX-2, IL-1, and IL-8 expression. Material and methods. of the 217 patients included in the study, 26 were uninfected, 127 had chronic gastritis and were H. pylori-positive, and 64 had gastric cancer. Bacterial genotypes were evaluated by polymerase chain reaction (PCR), and the expression values were determined by quantitative real-time PCR and immunohistochemistry. Results. An association was found between the infection with cagA, vacA s1m1 strains and gastric cancer development. Regarding the 3' region of the cagA gene, we also found an association between the infection with cagA EPIYA-ABCCC strains and clinical outcome. Higher levels of IL-8, IL-1, and COX-2 were detected in gastric mucosa from infected patients with chronic gastritis, and they were also associated with the infection by cagA, vacA s1m1 strains. The IL-8 and IL-1 levels decrease significantly from chronic gastritis to gastric cancer, while the relative expression remained unaltered when COX-2 expression was analyzed among patients with gastritis and cancer. Conclusions. Since inflammatory response to H. pylori infection plays an important role in cellular proliferation and gastric mucosal damage, the up-regulation of IL-1, IL-8, and COX-2 in patients with chronic gastritis has an important clinical implication in gastric carcinogenesis.

Effects of Helicobacter pylori Infection on the Expressions of Bax and Bcl-2 in Patients with Chronic Gastritis and Gastric Cancer

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Immunization protected well nourished mice but not undernourished ones from lung injury in Methicillin-resistant Staphylococcus aureus (MRSA) infection

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Brain and lung cryptococcoma and concurrent corynebacterium pseudotuberculosis infection in a goat: a case report

A four-year-old male goat with a history of neurological disorder was euthanized. It presented uncommon nodules in the brain and lungs associated with multiple abscesses, predominantly in the spleen and liver. Histological examination of brain and lung sections revealed yeast forms confirmed to be Cryptococcus gattii after a combination of isolation and polymerase chain reaction (PCR) procedures. Moreover, Corynebacterium pseudotuberculosis infection was diagnosed by PCR of samples from the lung, spleen and liver. The present report highlights the rare concurrent infection of C. gatti and C. pseudotuberculosis in an adult goat from São Paulo state, Brazil, and indicates the necessity of surveillance in the treatment of goats with atypical pulmonary infections associated with neurological disorders.