Profiling of human body fluids in healthy and diseased states using gas chromatography and mass spectrometry,with special reference to organic acids

This review summarizes recent advances in the application of gas chromatography and mass spectrometry to the study of human diseases. Emphasis is placed upon the organic acid profiles of the various body fluids. Methods for sample work-up prior to separation and mass spectrometric analysis are reviewed, and artifacts and pitfalls are discussed. Organic acid profiles, obtained with packed or capillary columns attached to mass spectrometers with or without computer systems, have led to the discovery of new normal metabolites, new metabolic disorders, and to new knowledge about a number of other diseases. Stable isotopes and gas chromatography—mass spectrometry are suitable for quantitative analysis of many compounds in the body fluids, and well suited for investigation of metabolic pathways.     

Structural,Biochemical, and Computational Characterization of the Glycoside Hydrolase Family 7 Cellobiohydrolase of the Tree-killing Fungus Heterobasidion irregulare

Root rot fungi of the Heterobasidion annosum complex are the most damaging pathogens in temperate forests, and the recently sequenced Heterobasidion irregulare genome revealed over 280 carbohydrate-active enzymes. Here, H. irregulare was grown on biomass, and the most abundant protein in the culture filtrate was identified as the only family 7 glycoside hydrolase in the genome, which consists of a single catalytic domain, lacking a linker and carbohydrate-binding module. The enzyme, HirCel7A, was characterized biochemically to determine the optimal conditions for activity. HirCel7A was crystallized and the structure, refined at 1.7 Å resolution, confirms that HirCel7A is a cellobiohydrolase rather than an endoglucanase, with a cellulose-binding tunnel that is more closed than Phanerochaete chrysosporium Cel7D and more open than Hypocrea jecorina Cel7A, suggesting intermediate enzyme properties. Molecular simulations were conducted to ascertain differences in enzyme-ligand interactions, ligand solvation, and loop flexibility between the family 7 glycoside hydrolase cellobiohydrolases from H. irregulare, H. jecorina, and P. chrysosporium. The structural comparisons and simulations suggest significant differences in enzyme-ligand interactions at the tunnel entrance in the −7 to −4 binding sites and suggest that a tyrosine residue at the tunnel entrance of HirCel7A may serve as an additional ligand-binding site. Additionally, the loops over the active site in H. jecorina Cel7A are more closed than loops in the other two enzymes, which has implications for the degree of processivity, endo-initiation, and substrate dissociation. Overall, this study highlights molecular level features important to understanding this biologically and industrially important family of glycoside hydrolases.     

Molecular Genetic Analysis of Stomach Contents Reveals Wild Atlantic Cod Feeding on Piscine Reovirus (PRV) Infected Atlantic Salmon Originating from a Commercial Fish Farm

In March 2012, fishermen operating in a fjord in Northern Norway reported catching Atlantic cod, a native fish forming an economically important marine fishery in this region, with unusual prey in their stomachs. It was speculated that these could be Atlantic salmon, which is not typical prey for cod at this time of the year in the coastal zone. These observations were therefore reported to the Norwegian Directorate of Fisheries as a suspected interaction between a local fish farm and this commercial fishery. Statistical analyses of genetic data from 17 microsatellite markers genotyped on 36 partially-degraded prey, samples of salmon from a local fish farm, and samples from the nearest wild population permitted the following conclusions: 1. The prey were Atlantic salmon, 2. These salmon did not originate from the local wild population, and 3. The local farm was the most probable source of these prey. Additional tests demonstrated that 21 of the 36 prey were infected with piscine reovirus. While the potential link between piscine reovirus and the disease heart and skeletal muscle inflammation is still under scientific debate, this disease had caused mortality of large numbers of salmon in the farm in the month prior to the fishermen''s observations. These analyses provide new insights into interactions between domesticated and wild fish.     

Positive short-term effects of sheep grazing on the alpine avifauna

Grazing by large herbivores may negatively affect bird populations. This is of great conservation concern in areas with intensive sheep grazing. Sheep management varies substantially between regions, but no study has been performed in less intensively grazed systems. In a fully replicated, landscape scale experiment with three levels of sheep grazing, we tested whether the abundance and diversity of an assemblage of mountain birds were negatively affected by grazing or if grazing facilitated the bird assemblage. Density of birds was higher at high sheep density compared with low sheep density or no sheep by the fourth grazing season, while there was no clear effect on bird diversity. Thus, agricultural traditions and land use politics determining sheep density may change the density of avifauna in either positive or negative directions.     

Site-specific recombination in human cells catalyzed by phage lambda integrase mutants

Phage lambda Integrase (Int) is the prototype of the so-called integrase family of conservative site-specific recombinases, which includes Cre and FLP. The natural function of Int is to execute integration and excision of the phage into and out of the Escherichia coli genome, respectively. In contrast to Cre and FLP, however, wild-type Int requires accessory proteins and DNA supercoiling of target sites to catalyze recombination. Here, we show that two mutant Int proteins, Int-h (E174 K) and its derivative Int-h/218 (E174 K/E218 K), which do not require accessory factors, are proficient to perform intramolecular integrative and excisive recombination in co-transfection assays inside human cells. Intramolecular integrative recombination is also detectable by Southern analysis in human reporter cell lines harboring target sites attB and attP as stable genomic sequences. Recombination by wild-type Int, however, is not detectable by this method. The latter result implies that eukaryotic co-factors, which could functionally replace the prokaryotic ones normally required for wild-type Int, are most likely not present in human cells.     

ClusTrack: Feature Extraction and Similarity Measures for Clustering of Genome-Wide Data Sets

Clustering is a popular technique for explorative analysis of data, as it can reveal subgroupings and similarities between data in an unsupervised manner. While clustering is routinely applied to gene expression data, there is a lack of appropriate general methodology for clustering of sequence-level genomic and epigenomic data, e.g. ChIP-based data. We here introduce a general methodology for clustering data sets of coordinates relative to a genome assembly, i.e. genomic tracks. By defining appropriate feature extraction approaches and similarity measures, we allow biologically meaningful clustering to be performed for genomic tracks using standard clustering algorithms. An implementation of the methodology is provided through a tool, ClusTrack, which allows fine-tuned clustering analyses to be specified through a web-based interface. We apply our methods to the clustering of occupancy of the H3K4me1 histone modification in samples from a range of different cell types. The majority of samples form meaningful subclusters, confirming that the definitions of features and similarity capture biological, rather than technical, variation between the genomic tracks. Input data and results are available, and can be reproduced, through a Galaxy Pages document at http://hyperbrowser.uio.no/hb/u/hb-superuser/p/clustrack. The clustering functionality is available as a Galaxy tool, under the menu option "Specialized analyzis of tracks", and the submenu option "Cluster tracks based on genome level similarity", at the Genomic HyperBrowser server: http://hyperbrowser.uio.no/hb/.     

Pakistanis living in Oslo have lower serum 1,25-dihydroxyvitamin D levels but higher serum ionized calcium levels compared with ethnic Norwegians. The Oslo Health Study

Background

Clinical determination of mid-parental height is an important part of the assessment of a child's growth, however our clinical impression has been that parents cannot be relied upon to accurately report their own heights. Therefore, we conducted this study to assess the accuracy of parental height self-reporting and its effect on calculated mid-parental target height for children presenting to a pediatric endocrinology office.

Methods

All parents bringing their children for an initial evaluation to a pediatric endocrinology clinic over a period of nine months were questioned and then measured by a pediatric endocrinologist. Parents were blinded to the study. Mid-parental target heights, based on reported and actual height were compared.

Results

There were 241 families: 98 fathers and 217 mothers in our study. Mean measured paternal height was 173.2 cm, self reported 174.9 cm (p < 0.0001), partner reported 177 cm (p = 0.0004). Only 50% of fathers and 58% of mothers reported their height within ± 2 cm of their measured height, while 15% of fathers and 12% of mothers were inaccurate by more than 4 cm. Mean measured maternal height was 160.6 cm, self-reported 161.1 cm (NS), partner reported 161.7 cm (NS). Inaccuracy of height self-report had a small but significant effect on the mean MPTH (0.4 cm, p = 0.045). Analysis showed that only 70% of MPTH calculated by reported heights fell within ± 2 cm of MPTH calculated using measured heights, 24% being in ± 2–4 cm range, and 6% were inaccurate by more than 4 cm.

Conclusion

There is a significant difference in paternal measured versus reported heights with an overall trend for fathers to overestimate their own height. A large subset of parents makes a substantial error in their height self-report, which leads to erroneous MPTH. Inaccuracy is even greater when one parent reports the other parent's height. When a child's growth is in question, measured rather than reported parental heights should be obtained.