Molecular cloning and functional expression of a novel Helicobacter pylori α-1,4 fucosyltransferase

Helicobacter pylori is an important human pathogen which causes both gastric and duodenal ulcers and is associated with gastric cancer and lymphoma. This microorganism synthesizes fucosylated oligosaccharides, predominantly the Galb-1,4GlcNAc (Type II) blood group antigens Lewis X and Y, whereas a small population also expresses the Galb-1,3GlcNAc (Type I) blood group antigens Lewis A and B. These carbohydrate structures are known to mimic host cell antigens and permit the bacteria to escape from the host immune response. Here, we report the cloning and characterization of a novel H. pylori α-1,4 fucosyltransferase (FucT). In contrast to the family members characterized to date, this enzyme shows exclusively Type I acceptor substrate specificity. The enzyme consisting of 432 amino acids (MW 50,502 Da) was cloned using a polymerase chain reaction (PCR)-based approach. It exhibits a high degree of identity (75-87%) and similar structural features, for example, in the heptamer repeat pattern, with other H. pylori FucTs. The kinetic characterization revealed a very efficient transferase (kcat/Km = 229 mM21s21) for the Type I acceptor substrate (Gal)-1,3 GlcNAc-Lem (1). Additionally, the enzyme possesses a broad tolerance toward nonnatural Type I acceptor substrate analogs and therefore represents a valuable tool for the chemoenzymatic synthesis of Lewis A, sialyl Lewis A as well as mimetics thereo

The Soil Microbiome of GLORIA Mountain Summits in the Swiss Alps

While vegetation has intensively been surveyed on mountain summits, limited knowledge exists about the diversity and community structure of soil biota. Here, we study how climatic variables, vegetation, parent material, soil properties, and slope aspect affect the soil microbiome on 10 GLORIA (Global Observation Research Initiative in Alpine environments) mountain summits ranging from the lower alpine to the nival zone in Switzerland. At these summits we sampled soils from all four aspects and examined how the bacterial and fungal communities vary by using Illumina MiSeq sequencing. We found that mountain summit soils contain highly diverse microbial communities with a total of 10,406 bacterial and 6,291 fungal taxa. Bacterial α-diversity increased with increasing soil pH and decreased with increasing elevation, whereas fungal α-diversity did not change significantly. Soil pH was the strongest predictor for microbial β-diversity. Bacterial and fungal community structures exhibited a significant positive relationship with plant communities, indicating that summits with a more distinct plant composition also revealed more distinct microbial communities. The influence of elevation was stronger than aspect on the soil microbiome. Several microbial taxa responded to elevation and soil pH. Chloroflexi and Mucoromycota were significantly more abundant on summits at higher elevations, whereas the relative abundance of Basidiomycota and Agaricomycetes decreased with elevation. Most bacterial OTUs belonging to the phylum Acidobacteria were indicators for siliceous parent material and several OTUs belonging to the phylum Planctomycetes were associated with calcareous soils. The trends for fungi were less clear. Indicator OTUs belonging to the genera Mortierella and Naganishia showed a mixed response to parent material, demonstrating their ubiquitous and opportunistic behaviour in soils. Overall, fungal communities responded weakly to abiotic and biotic factors. In contrast, bacterial communities were strongly influenced by environmental changes suggesting they will be strongly affected by future climate change and associated temperature increase and an upward migration of vegetation. Our results provide the first insights into the soil microbiome of mountain summits in the European Alps that are shaped as a result of highly variable local environmental conditions and may help to predict responses of the soil biota to global climate change

Diagnostic and prognostic accuracy of clinical and laboratory parameters in community-acquired pneumonia

BACKGROUND: Community-acquired pneumonia (CAP) is the most frequent infection-related cause of death. The reference standard to diagnose CAP is a new infiltrate on chest radiograph in the presence of recently acquired respiratory signs and symptoms. This study aims to evaluate the diagnostic and prognostic accuracy of clinical signs and symptoms and laboratory biomarkers for CAP. METHODS: 545 patients with suspected lower respiratory tract infection, admitted to the emergency department of a university hospital were included in a pre-planned post-hoc analysis of two controlled intervention trials. Baseline assessment included history, clinical examination, radiography and measurements of procalcitonin (PCT), highly sensitive C-reactive protein (hsCRP) and leukocyte count. RESULTS: Of the 545 patients, 373 had CAP, 132 other respiratory tract infections, and 40 other final diagnoses. The AUC of a clinical model including standard clinical signs and symptoms (i.e. fever, cough, sputum production, abnormal chest auscultation and dyspnea) to diagnose CAP was 0.79 [95% CI, 0.75–0.83]. This AUC was significantly improved by including PCT and hsCRP (0.92 [0.89–0.94]; p < 0.001). PCT had a higher diagnostic accuracy (AUC, 0.88 [0.84–0.93]) in differentiating CAP from other diagnoses, as compared to hsCRP (AUC, 0.76 [0.69–0.83]; p < 0.001) and total leukocyte count (AUC, 0.69 [0.62–0.77]; p < 0.001). To predict bacteremia, PCT had a higher AUC (0.85 [0.80–0.91]) as compared to hsCRP (p = 0.01), leukocyte count (p = 0.002) and elevated body temperature (p < 0.001). PCT, in contrast to hsCRP and leukocyte count, increased with increasing severity of CAP, as assessed by the pneumonia severity index (p < 0.001). CONCLUSION: PCT, and to a lesser degree hsCRP, improve the accuracy of currently recommended approaches for the diagnosis of CAP, thereby complementing clinical signs and symptoms. PCT is useful in the severity assessment of CAP

Label-Free FimH Protein Interaction Analysis Using Silicon Nanoribbon BioFETs

The detection of biomarkers at very low concentration and low cost is increasingly important for clinical diagnosis. Moreover, monitoring affinities for receptor-antagonist interactions by time-resolved measurements is crucial for drug discovery and development. Biosensors based on ion-sensitive field-effect transistors (BioFETs) are promising candidates for being integrated into CMOS structures and cost-effective production. The detection of DNA and proteins with silicon nanowires has been successfully demonstrated using high affinity systems such as the biotin-streptavidin interaction. Here, we show the time-resolved label-free detection of the interaction of the bacterial FimH lectin with an immobilized mannose ligand on gold-coated silicon nanoribbon BioFETs. By comparing our results with a commercial state of the art surface plasmon resonance system, additional surface effects become visible when using this charge based detection method. Furthermore, we demonstrate the effect of sensor area on signal-to-noise ratio and estimate the theoretical limit of detection

Intraspecific trait variation in alpine plants relates to their elevational distribution

Climate warming is shifting the distributions of mountain plant species to higher elevations. Cold-adapted plant species are under increasing pressure from novel competitors that are encroaching from lower elevations. Plant capacity to adjust to these pressures may be measurable as variation in trait values within a species. In particular, the strength and patterns of intraspecific trait variation along abiotic and biotic gradients can inform us whether and how species can adjust their anatomy and morphology to persist in a changing environment. Here, we tested whether species specialized to high elevations or with narrow elevational ranges show more conservative (i.e. less variable) trait responses across their elevational distribution, or in response to neighbours, than species from lower elevations or with wider elevational ranges. We did so by studying intraspecific trait variation of 66 species along 40 elevational gradients in four countries in both hemispheres. As an indication of potential neighbour interactions that could drive trait variation, we also analysed plant species' height ratio, its height relative to its nearest neighbour. Variation in alpine plant trait values over elevation differed depending on a species' median elevation and the breadth of its elevational range, with species with lower median elevations and larger elevational range sizes showing greater trait variation, i.e. a steeper slope in trait values, over their elevational distributions. These effects were evidenced by significant interactions between species' elevation and their elevational preference or range for several traits: vegetative height, generative height, specific leaf area and patch area. The height ratio of focal alpine species and their neighbours decreased in the lower part of their distribution because neighbours became relatively taller at lower elevations. In contrast, species with lower elevational optima maintained a similar height ratio with neighbours throughout their range. Synthesis. We provide evidence that species from lower elevations and those with larger range sizes show greater intraspecific trait variation, which may indicate a greater ability to respond to environmental changes. Also, larger trait variation of species from lower elevations may indicate stronger competitive ability of upslope shifting species, posing one further threat to species from higher ranges