ALBERT

Description: Background: Crescentic glomerulonephritis is a rare condition in children and is typically associated with renal insufficiency. Dysfunction of the alternative complement pathway is an unusual aetiology with an unknown mechanism.Case presentationWe report a case of a previously healthy 12-year-old Caucasian girl who was examined on emergency owing to an asymptomatic gross haematuria. An active urinary sediment and nephrotic-range proteinuria were identified, and serologic examination showed a decreased serum C3 concentration not associated with any immunologic or infectious cause. Oedema, hypertension, and renal insufficiency were not observed. A renal biopsy was performed, and crescentic glomerulonephritis associated with C3 glomerulonephritis was diagnosed. Prompt treatment with intravenous steroids resulted in complete resolution of the gross haematuria. Further examination did not detect any underlying acquired cause. A combination of oral steroids and cyclophosphamide, followed by mycophenolate mofetil, was maintained and resulted in clinical remission during an 8-month follow-up. Conclusion: The presence of severe injury such as crescentic glomerulonephritis secondary to C3 glomerulonephritis is extremely unusual in children. This is the first known case of paediatric crescentic glomerulonephritis secondary to C3 glomerulonephritis that presented with gross haematuria and was treated early and effectively with immunosuppressive therapy based on its severe histologic features.

Description: Background: The rearrangements in the 22q11.2 chromosomal region, responsible for the 22q11.2 deletion and microduplication syndromes, are frequently associated with congenital heart disease (CHD). The present work aimed to identify the genetic basis of CHD in 87 patients from the S?o Miguel Island, Azores, through the detection of copy number variants (CNVs) in the 22q11.2 region. These structural variants were searched using multiplex ligation-dependent probe amplification (MLPA). In patients with CNVs, we additionally performed fluorescent in situ hybridization (FISH) for the assessment of the exact number of 22q11.2 copies among each chromosome, and array comparative genomic hybridization (array-CGH) for the determination of the exact length of CNVs. Results: We found that four patients (4.6%; A to D) carried CNVs. Patients A and D, both affected with a ventricular septal defect, carried a de novo 2.5?Mb deletion of the 22q11.2 region, which was probably originated by inter-chromosomal (inter-chromatid) non-allelic homologous recombination (NAHR) events in the regions containing low-copy repeats (LCRs). Patient C, with an atrial septal defect, carried a de novo 2.5?Mb duplication of 22q11.2 region, which could have been probably generated during gametogenesis by NAHR or by unequal crossing-over; additionally, this patient presented a benign 288 Kb duplication, which included the TOP3B gene inherited from her healthy mother. Finally, patient B showed a 3?Mb triplication associated with dysmorphic facial features, cognitive deficit and heart defects, a clinical feature not reported in the only case described so far in the literature. The evaluation of patient B?s parents revealed a 2.5?Mb duplication in her father, suggesting a paternal inheritance with an extra copy. Conclusions: This report allowed the identification of rare deletion and microduplication syndromes in Azorean CHD patients. Moreover, we report the second patient with a 22q11.2 triplication, and we suggest that patients with triplications of chromosome 22q11.2, although they share some characteristic features with the deletion and microduplication syndromes, present a more severe phenotype probably due to the major dosage of implicated genes.

Description: Background: Innate immune system is the first line of research when studying immune response to diverse infections and autoimmune/inflammatory diseases. This immune response has been reported to be genetically diverse, due to polymorphisms coded by different genes. For this reason, our purpose was to develop a multiplex assay that allows the genotyping of candidate single nucleotide polymorphisms (SNPs) in innate immune genes.FindingsWe developed three multiplex PCR panels coupled with the minisequencing (SNaPshot) technique (multiplex PCR, multiplex primer extension, and capillary electrophoresis). The panels were tested in a sample set composed of 100 anonymous DNAs from healthy blood donors living in Sao Miguel Island (Azores, Portugal). Sixteen relevant SNPs among nine genes of the innate immune system -- IL1alpha, IL1beta, IL6, IL10, IL12RB1, TLR2, TLR4, TLR9 and CD14 -- were genotyped and validated by direct sequencing, with the exception of one that was undetected by minisequencing. We suggest that these panels can be used in future studies for detection of risk gene variants in several populations and/or diseases. Conclusions: In summary, we propose a multiplex assay that is able to identify the most frequent candidate SNPs in innate immune genes, using a medium scale genotyping platform. The assays can be used to evaluate the risk gene variants in populations of various geographic origins.

Description: Background: Restriction site-associated DNA sequencing (RAD-Seq) is a genome complexity reduction technique that facilitates large-scale marker discovery and genotyping by sequencing. Recent applications of RAD-Seq have included linkage and QTL mapping with a particular focus on non-model species. In the current study, we have applied RAD-Seq to two Atlantic salmon families from a commercial breeding program. The offspring from these families were classified into resistant or susceptible based on survival/mortality in an Infectious Pancreatic Necrosis (IPN) challenge experiment, and putative homozygous resistant or susceptible genotype at a major IPN-resistance QTL. From each family, the genomic DNA of the two heterozygous parents and seven offspring of each IPN phenotype and genotype was digested with the SbfI enzyme and sequenced in multiplexed pools. Results: Sequence was obtained from approximately 70,000 RAD loci in both families and a filtered set of 6,712 segregating SNPs were identified. Analyses of genome-wide RAD marker segregation patterns in the two families suggested SNP discovery on all 29 Atlantic salmon chromosome pairs, and highlighted the dearth of male recombination. The use of pedigreed samples allowed us to distinguish segregating SNPs from putative paralogous sequence variants resulting from the relatively recent genome duplication of salmonid species. Of the segregating SNPs, 50 were linked to the QTL. A subset of these QTL-linked SNPs were converted to a high-throughput assay and genotyped across large commercial populations of IPNV-challenged salmon fry. Several SNPs showed highly significant linkage and association with resistance to IPN, and population linkage-disequilibrium-based SNP tests for resistance were identified. Conclusions: We used RAD-Seq to successfully identify and characterise high-density genetic markers in pedigreed aquaculture Atlantic salmon. These results underline the effectiveness of RAD-Seq as a tool for rapid and efficient generation of QTL-targeted and genome-wide marker data in a large complex genome, and its possible utility in farmed animal selection programs.

Description: Background: Keeping up-to-date with bioscience literature is becoming increasingly challenging. Several recent methods help meet this challenge by allowing literature search to be launched based on lists of abstracts that the user judges to be 'interesting'. Some methods go further by allowing the user to provide a second input set of 'uninteresting' abstracts; these two input sets are then used to search and rank literature by relevance. In this work we present the service 'Caipirini' (http://caipirini.org) that also allows two input sets, but takes the novel approach of allowing ranking of literature based on one or more sets of genes. Results: To evaluate the usefulness of Caipirini, we used two test cases, one related to the human cell cycle, and a second related to disease defense mechanisms in Arabidopsis thaliana. In both cases, the new method achieved high precision in finding literature related to the biological mechanisms underlying the input data sets. Conclusions: To our knowledge Caipirini is the first service enabling literature search directly based on biological relevance to gene sets; thus, Caipirini gives the research community a new way to unlock hidden knowledge from gene sets derived via high-throughput experiments.

Description: Background: Crescentic glomerulonephritis is a rare condition in children and is typically associated with renal insufficiency. Dysfunction of the alternative complement pathway is an unusual aetiology with an unknown mechanism.Case presentationWe report a case of a previously healthy 12-year-old Caucasian girl who was examined on emergency owing to an asymptomatic gross haematuria. An active urinary sediment and nephrotic-range proteinuria were identified, and serologic examination showed a decreased serum C3 concentration not associated with any immunologic or infectious cause. Oedema, hypertension, and renal insufficiency were not observed. A renal biopsy was performed, and crescentic glomerulonephritis associated with C3 glomerulonephritis was diagnosed. Prompt treatment with intravenous steroids resulted in complete resolution of the gross haematuria. Further examination did not detect any underlying acquired cause. A combination of oral steroids and cyclophosphamide, followed by mycophenolate mofetil, was maintained and resulted in clinical remission during an 8-month follow-up. Conclusion: The presence of severe injury such as crescentic glomerulonephritis secondary to C3 glomerulonephritis is extremely unusual in children. This is the first known case of paediatric crescentic glomerulonephritis secondary to C3 glomerulonephritis that presented with gross haematuria and was treated early and effectively with immunosuppressive therapy based on its severe histologic features.

Description: Background: The fucoid brown algae (Heterokontophyta, Phaeophyceae) are increasingly the focus of ecological genetics, biodiversity, biogeography and speciation research. The molecular genetics underlying mating system variation, where repeated dioecious -- hermaphrodite switches during evolution are recognized, and the molecular evolution of sex-related genes are key questions currently hampered by a lack of genomic information. We therefore undertook a comparative analysis of male and female reproductive tissue transcriptomes against a vegetative background during natural reproductive cycles in Fucus vesiculosus. Results: Over 300 k reads were assembled and annotated against public protein databases including a brown alga. Compared with the vegetative tissue, photosynthetic and carbohydrate metabolism pathways were under-expressed, particularly in male tissue, while several pathways involved in genetic information processing and replication were over-expressed. Estimates of sex-biased gene (SBG) expression were higher for male (14% of annotated orthologues) than female tissue (9%) relative to the vegetative background. Mean expression levels and variance were also greater in male- than female-biased genes. Major female-biased genes were carbohydrate-modifying enzymes with likely roles in zygote cell wall biogenesis and/or modification. Male-biased genes reflected distinct sperm development and function, and orthologues for signal perception (a phototropin), transduction (several kinases), and putatively flagella-localized proteins (including candidate gamete-recognition proteins) were uniquely expressed in males. Overall, the results suggest constraint on female-biased genes (possible pleiotropy), and less constrained male-biased genes, mostly associated with sperm-specific functions. Conclusions: Our results support the growing contention that males possess a large array of genes regulating male fitness, broadly supporting findings in evolutionarily distant heterogametic animal models. This work identifies an annotated set of F. vesiculosus gene products that potentially regulate sexual reproduction and may contribute to prezygotic isolation, one essential step towards developing tools for a functional understanding of species isolation and differentiation.

Description: Background: Nonalcoholic fatty liver disease (NAFLD) related to obesity has been rising in the last decades, though the morphological and metabolic determinants are remain unclear in children. The aim of this study was to analyze the morphological determinants and metabolic abnormalities in obese children and adolescents, classified either as with (P-NAFLD) or without (N-NAFLD). The sample comprised 190 individuals, aged 6 to 16 years-old, assigned into one of 4 groups according to sex and presence or absence of NAFLD. Obesity was obtained according to body mass index (BMI) cut-points. Body composition variables was estimated by Dual-Energy X-ray Absorptiometry (DEXA). Total cholesterol (TC), triglycerides (TG), HDL-cholesterol (HDL-C), LDL-cholesterol (LDL-C), insulin, fasting glucose (FG) and blood pressure were also analyzed. The diagnosis of NAFLD, as well as the measurement of intra-abdominal fat tissue thickness (IAF) and subcutaneous abdominal fat tissue thickness (SCF), was carried-out by ultrasound. Results: Males and females belonging to P-NAFLD group showed, respectively, higher TFM and IAF. When data were adjusted for sex, age and total fat mass, those in P-NAFLD showed statistically higher IAF, TFM and TG. Conclusion: Our study showed that obese youngsters who were assigned to P-NAFLD group were twice as likely to present higher concentration of triglycerides, higher levels of trunk fat, as well as intra-abdominal fat compared to their N-NAFLD counterparts even after adjustments for sex, age, pubertal stage and total body fat mass.

Description: Background: Nonalcoholic fatty liver disease (NAFLD) related to obesity has been rising in the last decades, though the morphological and metabolic determinants are remain unclear in children. The aim of this study was to analyze the morphological determinants and metabolic abnormalities in obese children and adolescents, classified either as with (P-NAFLD) or without (N-NAFLD). The sample comprised 190 individuals, aged 6 to 16 years-old, assigned into one of 4 groups according to sex and presence or absence of NAFLD. Obesity was obtained according to body mass index (BMI) cut-points. Body composition variables was estimated by Dual-Energy X-ray Absorptiometry (DEXA). Total cholesterol (TC), triglycerides (TG), HDL-cholesterol (HDL-C), LDL-cholesterol (LDL-C), insulin, fasting glucose (FG) and blood pressure were also analyzed. The diagnosis of NAFLD, as well as the measurement of intra-abdominal fat tissue thickness (IAF) and subcutaneous abdominal fat tissue thickness (SCF), was carried-out by ultrasound. Results: Males and females belonging to P-NAFLD group showed, respectively, higher TFM and IAF. When data were adjusted for sex, age and total fat mass, those in P-NAFLD showed statistically higher IAF, TFM and TG. Conclusion: Our study showed that obese youngsters who were assigned to P-NAFLD group were twice as likely to present higher concentration of triglycerides, higher levels of trunk fat, as well as intra-abdominal fat compared to their N-NAFLD counterparts even after adjustments for sex, age, pubertal stage and total body fat mass.

Description: Background: Chlamydia trachomatis is an obligate intracellular human pathogen causing ocular and urogenital infections that are a significant clinical and public health concern. This bacterium uses a type III secretion (T3S) system to manipulate host cells, through the delivery of effector proteins into their cytosol, membranes, and nucleus. In this work, we aimed to find previously unidentified C. trachomatis T3S substrates. Results: We first analyzed the genome of C. trachomatis L2/434 strain for genes encoding mostly uncharacterized proteins that did not appear to possess a signal of the general secretory pathway and which had not been previously experimentally shown to be T3S substrates. We selected several genes with these characteristics and analyzed T3S of the encoding proteins using Yersinia enterocolitica as a heterologous system. We identified 23C. trachomatis proteins whose first 20 amino acids were sufficient to drive T3S of the mature form of beta-lactamase TEM-1 by Y. enterocolitica. We found that 10 of these 23 proteins were also type III secreted in their full-length versions by Y. enterocolitica, providing additional support that they are T3S substrates. Seven of these 10 likely T3S substrates of C. trachomatis were delivered by Y. enterocolitica into host cells, further suggesting that they could be effectors. Finally, real-time quantitative PCR analysis of expression of genes encoding the 10 likely T3S substrates of C. trachomatis showed that 9 of them were clearly expressed during infection of host cells. Conclusions: Using Y. enterocolitica as a heterologous system, we identified 10 likely T3S substrates of C. trachomatis (CT053, CT105, CT142, CT143, CT144, CT161, CT338, CT429, CT656, and CT849) and could detect translocation into host cells of CT053, CT105, CT142, CT143, CT161, CT338, and CT429. Therefore, we revealed several C. trachomatis proteins that could be effectors subverting host cell processes.