Monitoring of Group C Rotavirus in Children With Acute Gastroenteritis in Brazil: An Emergent Epidemiological Issue After Rotavirus Vaccine?

Group C rotavirus (GpCRV) has a worldwide distribution; however, its epidemiology and ecology are still unclear. Evidence for a possible zoonotic role has been postulated recently for Brazilian children strains. The aim of this study was to monitor GpCRV in children <= 15 years with acute gastroenteritis during the 2007-2010 national Brazilian rotavirus surveillance, and to undertake the molecular characterization of the major VP6 capsid protein. A total of 3,019 fecal samples were first screened for Group A rotavirus (GpARV). A total of 2,205 GpARV ELISA negative samples were tested further for the presence of GpCRV by SDS-PAGE, electronic microscopy, and RT-PCR for the VP6 gene. The genetic diversity of GpCRV was carried out by sequencing the VP6 gene. GpARV and GpCRV infections were detected in 24.6% (742/3,019) and 0.3% (8/3,019), respectively. The GpCRV detection rate increased from 0.2% (1/422) in 2007 to 1% (7/708) in 2008, and GpCRV cases were not detected in 2009 and 2010. The phylogenetic analysis indicated that the strains belonged to the human lineage, and showed a genetic relationship with the GpCRV strain from Japan isolated in 2009. None of the study sequences was related closely to animal GpCRV strains. This study provides further evidence that GpCRV is a minor cause of acute childhood gastroenteritis in Brazil, and does not suggest that GpCRV may assume epidemiological importance in the future, even after the introduction of a GpARV vaccine. In addition, the molecular analyses of the GpCRV samples in this study do not support the zoonotic hypothesis. J. Med. Virol. 83: 1631-1636, 2011. (C) 2011 Wiley-Liss, Inc.

Verbal fluency in the detection of mild cognitive impairment and Alzheimer`s disease among Brazilian Portuguese speakers: the influence of education

Background: Verbal fluency (VF) tasks are simple and efficient clinical tools to detect executive dysfunction and lexico-semantic impairment. VF tasks are widely used in patients with suspected dementia, but their accuracy for detection of mild cognitive impairment (MCI) is still under investigation. Schooling in particular may influence the subject`s performance. The aim of this study was to compare the accuracy of two semantic categories (animals and fruits) in discriminating controls, MCI patients and Alzheimer`s disease (AD) patients. Methods: 178 subjects, comprising 70 controls (CG), 70 MCI patients and 38 AD patients, were tested on two semantic VF tasks. The sample was divided into two schooling groups: those with 4-8 years of education and those with 9 or more years. Results: Both VF tasks - animal fluency (VFa) and fruits fluency (VFf) - adequately discriminated CG from AD in the total sample (AUC = 0.88 +/- 0.03, p < 0.0001) and in both education groups, and high educated MCI from AD (VFa: AUC = 0.82 +/- 0.05, p < 0.0001; VFf: AUC = 0.85 +/- 0.05, p < 0.0001). Both tasks were moderately accurate in discriminating CG from MCI (VFa: AUC = 0.68 +/- 0.04, p < 0.0001 - VFf:AUC = 0.73 +/- 0.04, p < 0.0001) regardless of the schooling level, and MCI from AD in the total sample (VFa: AUC = 0.74 +/- 0.05, p < 0.0001; VFf: AUC = 0.76 +/- 0.05, p < 0.0001). Neither of the two tasks differentiated low educated MCI from AD. In the total sample, fruits fluency best discriminated CG from MCI and MCI from AD; a combination of the two improved the discrimination between CG and AD. Conclusions: Both categories were similar in discriminating CG from AD; the combination of both categories improved the accuracy for this distinction. Both tasks were less accurate in discriminating CG from MCI, and MCI from AD.

Conservation of hepatitis C virus nonstructural protein 3 amino acid sequence in viral isolates during liver transplantation

As a consequence of selective pressure exerted by the immune response during hepatitis C virus (HCV) infection, a high rate of nucleotide mutations in the viral genome is observed which leads to the emergence of viral escape mutants. The aim of this study was to evaluate the evolution of the amino acid (aa) sequence of the HCV nonstructural protein 3 (NS3) in viral isolates after liver transplantation. Six patients with HCV-induced liver disease undergoing liver transplantation (LT) were followed up for sequence analysis. Hepatitis C recurrence was observed in all patients after LT. The rate of synonymous (dS) nucleotide substitutions was much higher than that of nonsynonymous (dN) ones in the NS3 encoding region. The high values of the dS/dN ratios suggest no sustained adaptive evolution selection pressure and, therefore, absence of specific NS3 viral populations. Clinical genotype assignments were supported by phylogenetic analysis. Serial samples from each patient showed lower mean nucleotide genetic distance when compared with samples of the same HCV genotype and subtype. The NS3 samples studied had an N-terminal aa sequence with several differences as compared with reference ones, mainly in genotype 1b-infected patients. After LT, as compared with the sequences before, a few reverted aa substitutions and several established aa substitutions were observed at the N-terminal of NS3. Sites described to be involved in important functions of NS3, notably those of the catalytic triad and zinc binding, remained unaltered in terms of aa sequence. Rare or frequent aa substitutions occurred indiscriminately in different positions. Several cytotoxic T lymphocyte epitopes described for HCV were present in our 1b samples. Nevertheless, the deduced secondary structure of the NS3 protease showed a few alterations in samples from genotype 3a patients, but none were seen in 1b cases. Our data, obtained from patients under important selective pressure during LT, show that the NS3 protease remains well conserved, mainly in HCV 3a patients. It reinforces its potential use as an antigenic candidate for further studies aiming at the development of a protective immune response.

The potential role of C-reactive protein in distinguishing cytomegalovirus from tuberculosis and bacterial infections in renal transplant recipients

Introduction: The delay in the diagnosis of infections can be deleterious in renal transplant recipients. Thus, laboratory tests leading to an earlier diagnosis are very useful for these patients. Purpose: To assess the behavior of C-reactive protein (CRP) in renal transplant recipients with a diagnosis of cytomegalovirus (CMV) infection, tuberculosis (TB) and bacterial infection (BI). Methods: A retrospective analysis of 129 patients admitted at our hospital, from 2006 to 2008 because of CMV, TB or BI, was carried out. Appropriate statistical analysis was done and values were expressed as medians, range. Results: When CRP levels were compared among the groups with CMV disease, TB or BI, the group with CMV disease presented lower levels of CRP (18.4 mg/L, 0.28-44 mg/L) than the TB and BI (p < 0.05) groups. The area under the receiver-operating characteristics curve, distinguishing CMV disease from TB/BI, was 0.96 (p < 0.0001), resulting in 100% sensitivity and 90.63% specificity to detect CMV disease when CRP < 44.5 mg/L. The subgroup analysis of CMV infection showed increasing levels of CRP (0.28, 16 and 29.5 mg/L) in the asymptomatic, symptomatic and invasive disease subgroups, respectively (p < 0.05). Conclusion: The measurement of CRP levels may be a useful tool for differentiating CMV infection from the other types (bacterial or TB) of infection in kidney transplant recipients.

Protein disulfide isomerase overexpression in vascular smooth muscle cells induces spontaneous preemptive NADPH oxidase activation and Nox1 mRNA expression: Effects of nitrosothiol exposure

Mechanisms regulating NADPH oxidase remain open and include the redox chaperone protein disulfide isomerase (PDI). Here, we further investigated PDI effects on vascular NADPH oxidase. VSMC transfected with wild-type PDI (wt-PDI) OF PDI mutated in all four redox cysteines (mut-PDI) enhanced (2.5-fold) basal cellular ROS production and membrane NADPH oxidase activity, with 3-fold increase in Nox1, but not Nox4 mRNA. However, further ROS production, NADPH oxidase activity and Nox1 mRNA increase triggered by angiotensin-II (AngII) were totally lost with PDI overexpression, suggesting preemptive Nox1 activation in such cells. PDI overexpression increased Nox4 mRNA after AngII stimulus, although without parallel ROS increase. We also show that Nox inhibition by the nitric oxide donor GSNO is independent of PDI. PDI silencing decreased specifically Nox1 mRNA and protein, confirming that PDI may regulate Nox1 at transcriptional level in VSMC. Such data further strengthen the role of PDI as novel NADPH oxidase regulator. (C) 2009 Elsevier Inc. All rights reserved.

Molecular Characterization of Patients with X-linked Hyper-IgM Syndrome: Description of Two Novel CD40L Mutations

Type 1, X-linked Hyper-IgM syndrome (HIGM1) is caused by mutations in the gene encoding the CD154 protein, also known as CD40 ligand (CD40LG). CD40L is expressed in activated T cells and interacts with CD40 receptor expressed on B lymphocytes and dendritic cells. Affected patients present cellular and humoral immune defects, with infections by intracellular, opportunistic and extracellular pathogens. In the present study we investigated the molecular defects underlying disease in four patients with HIGM1. We identified four distinct CD40L mutations, two of them which have not been previously described. P1 harboured the novel p.G227X mutation which abolished CD40L expression. P2 had a previously described frame shift deletion in exon 2 (p.I53fsX65) which also prevented protein expression. P3 demonstrated the previously known p.V126D change in exon 4, affecting the TNF homology (TNFH) domain. Finally, P4 evidenced the novel p.F229L mutation also located in the TNFH domain. In silico analysis of F229L predicted the change to be pathological, affecting the many hydrophobic interactions of this residue. Precise molecular diagnosis in HIGM syndrome allows reliable detection of carriers, making genetic counselling and prenatal diagnosis possible.

Epstein-Barr virus detection in invasive and pre-invasive lesions of the uterine cervix

In the present study, our aim was to investigate whether EBV DNA could be found in association with invasive and pre-invasive cervical cancer lesions. We hypothesize that EBV is not merely a commensal agent when present in malignant cervical lesions. DNA was extracted from cervical scrapings followed by nested PCR-based amplification. The patients were 66 women with high grade cervical intraepithelial neoplasia and 14 women with invasive cervical cancer. The control group consisted of 89 women with a normal Pap smear and colposcopy as well as a negative HPV DNA test. Analysis of our results, in conjunction with the work of other authors, leads us to propose that EBV is not merely a commensal agent when present in malignant cervical lesions. The presence of DNA from EBV is significantly associated with cervical cancer.

Influence of Single-Nucleotide Polymorphisms on C-Reactive Protein Levels in Chronic Kidney Disease Before and After Kidney Transplantation

Introduction. We sought to evaluate 2 sing] e-nucleotide polymorphisms (SNPs) in the C-reactive protein (CRP) gene promoter region for their effects on CRP levels in chronic kidney disease (CKD) patients before and after a successful kidney transplantation. Methods. Fifty CKD patients were evaluated before and at the first and second years after the graft. Two SNPs were studied, a bi-allelic (G -> A) at the -409 and a tri-allelic (C -> T -> A) variation at the -390 position in the CRP gene. Results. All patients presented the -409GG genotype. At the -390 position, the ""A"" allele was not found; there were 15 ""CC"" patients, 11 ""TT"" patients, and 24 ""CT"" patients. CRP levels were different among patients with various genotypes (P < .019). Also the presence of the allele ""T"" was sufficient to determine differences in CRP levels both in pretransplantation (P = .045) and at 1 year posttransplantation (P = .011), but not at the second year (P = .448). Conclusion. SNPs at the -390 position of the CRP gene promoter region influence CRP basal levels in such a way that the ""C"" allele correlated with the lowest and the ""T"" with the highest. We did not observe this influence in our patients at the second year posttransplantation.

New SMS mutation leads to a striking reduction in spermine synthase protein function and a severe form of Snyder-Robinson X-linked recessive mental retardation syndrome

We report the identification of a novel mutation at a highly conserved residue within the N-terminal region of spermine synthase (SMS) in a second family with Snyder-Robinson X-linked mental retardation syndrome ( OMIM 309583). This missense mutation, p.G56S, greatly reduces SMS activity and leads to severe epilepsy and cognitive impairment. Our findings contribute to a better delineation and expansion of the clinical spectrum of Snyder-Robinson syndrome, support the important role of the N-terminus in the function of the SMS protein, and provide further evidence for the importance of SMS activity in the development of intellectual processing and other aspects of human development.

Effects of exercise on leukocyte death: prevention by hydrolyzed whey protein enriched with glutamine dipeptide

Lymphocyte and neutrophil death induced by exercise and the role of hydrolyzed whey protein enriched with glutamine dipeptide (Gln) supplementation was investigated. Nine triathletes performed two exhaustive exercise trials with a 1-week interval in a randomized, double blind, crossover protocol. Thirty minutes before treadmill exhaustive exercise at variable speeds in an inclination of 1% the subjects ingested 50 g of maltodextrin (placebo) or 50 g of maltodextrin plus 4 tablets of 700 mg of hydrolyzed whey protein enriched with 175 mg of glutamine dipeptide dissolved in 250 mL water. Cell viability, DNA fragmentation, mitochondrial transmembrane potential and production of reactive oxygen species (ROS) were determined in lymphocytes and neutrophils. Exhaustive exercise decreased viable lymphocytes but had no effect on neutrophils. A 2.2-fold increase in the proportion of lymphocytes and neutrophils with depolarized mitochondria was observed after exhaustive exercise. Supplementation of maltodextrin plus Gln (MGln) prevented the loss of lymphocyte membrane integrity and the mitochondrial membrane depolarization induced by exercise. Exercise caused an increase in ROS production by neutrophils, whereas supplementation of MGln had no additional effect. MGln supplementation partially prevented lymphocyte apoptosis induced by exhaustive exercise possibly by a protective effect on mitochondrial function.

Topical interleukin-1 receptor antagonist inhibits inflammatory cell infiltration into the cornea

Interleukin (IL)-1 alpha and beta are important modulators of many functions of corneal epithelial and stromal cells that occur following injury to the cornea, including the influx of bone marrow-derived inflammatory cells into the stroma attracted by chemokines released from the stroma and epithelium. In this study, we examined the effect of topical soluble IL-1 receptor antagonist on bone marrow-derived cell influx following corneal epithelial scrape injury in a mouse model. C57BL/6 mice underwent corneal epithelial scrape followed by application of IL-1 receptor antagonist (Amgen, Thousand Oaks, CA) at a concentration of 20 mg/ml or vehicle for 24 h prior to immunocytochemical detection of marker CD11b-positive cells into the stroma. In two experiments, topical IL-1 receptor antagonist had a marked effect in blocking cell influx. For example, in experiment 1, topical IL-1 receptor antagonist markedly reduced detectible CD11b-positive cells into the corneal stroma at 24 It after epithelial injury compared with the vehicle control (3.5 +/- 0.5 (standard error of the mean) cells/400x field and 13.9 +/- 1.2 cells/400x field, respectively, p < 0.01). A second experiment with a different observer performing cell counting had the same result. Thus, the data demonstrate conclusively that topical IL-1 receptor antagonist markedly down-regulates CD-11b-positive monocytic cell appearance in the corneal stroma. Topical IL-1 receptor antagonist could be an effective adjuvant for clinical treatment of corneal conditions in which unwanted inflammation has a role in the pathophysiology of the disorder. (c) 2008 Elsevier Ltd. All rights reserved.

Scanning laser polarimetry with enhanced corneal compensation for detection of axonal loss in band atrophy of the optic nerve

PURPOSE: To compare the abilities of scanning laser polarimetry (SLP) with enhanced corneal compensation (ECC) and variable corneal compensation (VCC) modes for detection of retinal nerve fiber layer (RNFL) loss in eyes with band atrophy (BA) of the optic nerve. DESIGN. Cross-sectional study. METHODS: Thirty-seven eyes from 37 patients with BA and temporal visual field defect from chiasmal compression and 40 eyes from 40 healthy subjects were studied. Subjects underwent standard automated perimetry and RNFL measurements using an SLP device equipped with VCC and ECC. Receiver operating characteristic (ROC) curves were calculated for each parameter. Pearson correlation coefficients were obtained to evaluate the relationship between RNFL thickness parameters and severity of visual field loss, as assessed by the temporal mean defect. RESULTS: All RNFL thickness parameters were significantly lower in eyes with BA compared with normal eyes with both compensation modes. However, no statistically significant differences were observed in the areas under the ROC curves for the different parameters between GDx VCC and ECC (Carl Zeiss Meditec, Inc, Dublin, California, USA). Structure-function relationships also were similar for both compensation modes. CONCLUSIONS: No significant differences were found between the diagnostic accuracy of GDx ECC and that of VCC for detection of BA of the optic nerve. The use of GDx ECC does not seem to provide a better evaluation of RNFL loss on the temporal and nasal sectors of the peripapillary retina in subjects with BA of the optic nerve.

Cellular prion protein (PrP(C)) and superoxide dismutase (SOD) in vascular cells under oxidative stress

The PrP(C) is expressed in several cell types but its physiological function is unknown. Some studies associate the PrP(C) with copper metabolism and the antioxidant activity of SOD. Our hypothesis was that changes in PrP(C) expression lead to abnormal copper regulation and induce SOD downregulation in the vascular wall. Objectives: to study whether the PrP(C) expression undergoes induction by agents that trigger endoplasmic reticulum stress (ERS) and, in this context, to evaluate the SOD activity. Methods: To trigger ERS, in vitro, rabbit aortic smooth muscle cells were challenged for 4, 8 and 18 hours, with angiotensin-II, tunicamycin and 7-ketocholesterol. For in vivo studies rabbit aortic arteries were subjected to injury by balloon catheter. Results: In vitro baseline SOD activity, determined through inhibition of cytochrome-c reduction, was 13.9 +/- 1.2 U/mg protein, angiotensin-II exposed for 8 hours produced an increase in SOD activity, and cellular copper concentration was about 9 times greater only under these conditions. Western blotting analysis for SOD isoenzymes showed an expression profile that was not correlated with the enzymatic activity. PrP(C) expression decreased after exposure to all agents after different incubation periods. RT-PCR assay showed increased mRNA expression for PrP(C) only in cells stimulated for 8 hours with the different stressors. The PrP(C) mRNA expression in rabbit aortic artery fragments, subjected to balloon catheter injury, showed a pronounced increase immediately after overdistension. The results obtained indicated a PrP(C) protection factor during the early part of the ERS exposure period, but did not demonstrate a SOD-like profile for the PrP(C). (C) 2009 Elsevier GmbH. All rights reserved.

A low-protein, high-carbohydrate diet increases the adipose lipid content without increasing the glycerol-3-phosphate or fatty acid content in growing rats

Le taux de triacylglycerol (TAG) qui s`accumule dans le tissu adipeux depend de 2 mecanismes opposes : la lipogenese et la lipolyse. Nous avons montre anterieurement que le poids des lipides du tissu adipeux de l`epididyme (EPI) de meme que leur taux augmentent chez les rats en croissance soumis a une diete hypoproteique hyperglucidique (HPHG) pendant 15 jours. La presente etude a eu pour but d`examiner les voies impliquees dans la lipogenese et la lipolyse qui regulent l`accumulation des lipides dans le tissu. On a evalue in vivo la synthese de novo des acides gras, qui s`est revelee similaire chez les rats soumis a la diete HPHG ou a une diete temoin; toutefois, chez les rats soumis a la diete HPHG, une diminution de l`activite de la lipoproteine lipase dans le tissus adipeux de l`EPI a ete observee, ce qui laisse croire a une diminution de la capture des acides gras des lipoproteines circulantes. La diete HPHG n`a eu aucun effet sur la synthese du glycerol-3-phosphate (G3P) par la glycolyse ou la glyceroneogenese. L`activite de la glycerokinase, c.-a-d. la phosphorylation du glycerol issu de l`hydrolyse du TAG endogene pour former le GP3, n`a pas ete modifiee non plus par la diete HPHG. A l`oppose, les adipocytes des rats HPHG stimules par la norepinephrine ont eu une plus faible reponse lipolytique, meme si le taux lipolytique basal des adipocytes a ete similaire chez les 2 groupes. Ainsi, les resultats donnent a penser que la diminution de l`activite lipolytique stimulee par la norepinephrine joue un role essentiel dans l`augmentation du TAG observee dans le tissu adipeux de l`EPI des animaux HPHG, probablement en perturbant le processus d`activation de la lipolyse.