Anaerobic bacteria infection in cystic fibrosis airway disease

Depletion of the periciliary liquid in "Cystic Fibrosis" airway disease results in reduced mucociliary transport, persistent mucus hypersecretion and consequently increased height of the luminal mucus layer, so hypoxic gradients in the mucus plugs are developed. Because of anaerobic lung zones, it is highly probable that anaerobic bacteria not detected by routine bacteriologic culture methods also reside within the mucus. Notwithstanding this evidence, microbiology laboratories working in the cystic fibrosis field do not generally use strict anaerobic bacteriologic cultures to determine the presence of anaerobic bacteria in the Cystic Fibrosis lung. The aim of this review is to focus on the published data regarding the finding of anaerobic bacteria in cystic fibrosis airway disease. Therefore, microbiology, diagnosis, antimicrobial susceptibility and possible impact on clinical management of anaerobic bacteria lung infection in cystic fibrosis are described.     

Proteomic analysis in the identification of allergenic molecules

Conventional and innovative strategies can be exploited to identify and characterize new allergenic proteins. With the aim of obtaining suggestions for future improvements, this article describes our attempt to understand and describe some of the advantages and pitfalls of the methodologies and procedures often used in this field. The analysis includes the protein extract preparation, starting from the allergenic source, the separation of the proteins contained in a mixture and the detection, identification and characterization of IgE-binding molecules. Classic and emerging proteomic technologies, including mass spectrometry-based methodologies, Edman degradation procedure, microarray-based techniques and bioinformatics search strategies, have been explored. A comparative analysis of biochemistry-based proteomics and molecular biology strategies has also been given.     

Glutamine-binding protein from Escherichia coli specifically binds a wheat gliadin peptide. 2. Resonance energy transfer studies suggest a new sensing approach for an easy detection of wheat gliadin

In this work is presented the first attempt to develop a fluorescence assay for detection of traces of gluten in food by utilizing the recombinant glutamine-binding protein (GlnBP) from E. coli. We found that GlnBP specifically binds the sequence of amino acids present both in gliadin and other prolamines classified as toxic for celiac patients. Affinity chromatography experiments together with mass spectrometry experiments demonstrated that GlnBP can bind the following amino acid sequence XXQPQPQQQQQQQQQQQQL. Sequence alignment experiments pointed out that this sequence is exclusively representative of the gliadin and the other prolamines considered toxic for celiac patients. These findings suggest the development of a competitive resonance energy transfer (RET) assay for an easy and rapid detection of this sequence in raw and cooked food.     

The extended multidomain solution structures of the complement protein Crry and its chimeric conjugate Crry-Ig by scattering,analytical ultracentrifugation and constrained modelling: implications for function and therapy

Complement receptor-related gene/protein y (Crry) is a cell membrane-bound regulator of complement activation found in mouse and rat. Crry contains only short complement/consensus repeat (SCR) domains. X-ray and neutron scattering was performed on recombinant rat Crry containing the first five SCR domains (rCrry) and mouse Crry with five SCR domains conjugated to the Fc fragment of mouse IgG1 (mCrry-Ig) in order to determine their solution structures at medium resolution. The radius of gyration R(G) of rCrry was determined to be 4.9-5.0 nm, and the R(G) of the cross-section was 1.2-1.5 nm as determined by X-ray and neutron scattering. The R(G) of mCrry-Ig was 6.6-6.7 nm, and the R(G) of the cross-section were 2.3-2.4 nm and 1.3 nm. The maximum dimension of rCrry was 18 nm and that for mCrry-Ig was 26 nm. The neutron data indicated that rCrry and mCrry-Ig have molecular mass values of 45,000 Da and 140,000 Da, respectively, in agreement with their sequences, and sedimentation equilibrium data supported these determinations. Time-derivative velocity experiments gave sedimentation coefficients of 2.4S for rCrry and 5.4S for mCrry-Ig. A medium-resolution model of rCrry was determined using homology models that were constructed for the first five SCR domains of Crry from known crystal and NMR structures, and linked by randomly generated linker peptide conformations. These trial-and-error calculations revealed a small family of extended rCrry structures that best accounted for the scattering and ultracentrifugation data. These were shorter than the most extended rCrry models as the result of minor bends in the inter-SCR orientations. The mCrry-Ig solution data were modelled starting from a fixed structure for rCrry and the crystal structure of mouse IgG1, and was based on conformational searches of the hinge peptide joining the mCrry and Fc fragments. The best-fit models showed that the two mCrry antennae in mCrry-Ig were extended from the Fc fragment. No preferred orientation of the antennae was identified, and this indicated that the accessibility of the antennae for the molecular targets C4b and C3b was not affected by the covalent link to Fc. A structural comparison between Crry and complement receptor type 1 indicated that the domain arrangement of Crry SCR 1-3 is as extended as that of the CR1 SCR 15-17 NMR structure.     

THE ELEMENTAL COMPOSITION OF SOME MARINE PHYTOPLANKTON1

We analyzed the cellular content of C, N, P, S, K, Mg, Ca, Sr, Fe, Mn, Zn, Cu, Co, Cd, and Mo in 15 marine eukaryotic phytoplankton species in culture representing the major marine phyla. All the organisms were grown under identical culture conditions, in a medium designed to allow rapid growth while minimizing precipitation of iron hydroxide. The cellular concentrations of all metals, phosphorus, and sulfur were determined by high‐resolution inductively coupled plasma mass spectrometry (HR‐ICPMS) and those of carbon and nitrogen by a carbon hydrogen nitrogen analyzer. Accuracy of the HR‐ICPMS method was validated by comparison with data obtained with ⁵⁵Fe radioactive tracer and by a planktonic reference material. The cellular quotas (normalized to P) of trace metals and major cations in the biomass varied by a factor of about 20 among species (except for Cd, which varied over two orders of magnitude) compared with factors of 5 to 10 for major nutrients. Green algae had generally higher C, N, Fe, Zn, and Cu quotas and lower S, K, Ca, Sr, Mn, Co, and Cd quotas than coccolithophores and diatoms. Co and Cd quotas were also lower in diatoms than in coccolithophores. Although trace element quotas are influenced by a variety of growth conditions, a comparison of our results with published data suggests that the measured compositions reflect chiefly the intrinsic (i.e. genetically encoded) trace element physiology of the individual species. Published field data on the composition of the planktonic biomass fall within the range of laboratory values and are generally close to the approximate extended Redfield formula given by the average stoichiometry of our model species (excluding the hard parts): While clearly this elemental stoichiometry varies between species and, potentially, in response to changes in the chemistry of seawater, it provides a basis for examining how phytoplankton influence the relative distributions of the ensemble of major and trace elements in the ocean.     

Effect of solid meal on gastric emptying of,and glycemic and cardiovascular responses to,liquid glucose in older subjects

Gastric emptying is a determinant of the postprandial glycemic and cardiovascular responses to oral carbohydrate. We evaluated the effects of a solid meal on gastric emptying and the glycemic and cardiovascular responses to oral glucose in healthy older subjects. Ten subjects aged 72.1 +/- 1.9 yr were studied. Each subject had measurements of gastric emptying, blood glucose, serum insulin, blood pressure, and heart rate after ingestion of a 50-g glucose drink (300 ml) with (mixed meal) or without (liquid only) a solid meal (300 g ground beef). Gastric emptying of liquid was initially slightly more rapid (P < 0.05) after the mixed meal compared with liquid only at 5 min (92.0 +/- 1.5 vs. 96.0 +/- 1.3%) and much slower (P < 0.05) after 120 min. The time to peak blood glucose was less (39.0 +/- 4.0 vs. 67.5 +/- 10.3 min; P < 0.01) and blood glucose subsequently lower (P < 0.01) after the mixed meal. The increase in serum insulin was greater (P < 0.001) after the mixed meal. Blood pressure fell (P < 0.05) in the first 30 min, with no difference between the two meals. Increase in heart rate after both meals (P < 0.005), was greater (P < 0.05) after the mixed meal. The presence of a noncarbohydrate solid meal had discrepant effects on early and subsequent emptying of a nutrient liquid, which affects postprandial glycemia and increased heart rate.     

Functional expression and characterization of EryA,the erythritol kinase of Brucella abortus,and enzymatic synthesis of L-erythritol-4-phosphate

The eryA gene of the bacterial pathogen Brucella abortus has been functionally expressed in Escherichia coli. The resultant EryA was shown to catalyze the ATP-dependent conversion of erythritol to L-erythritol-4-phosphate (L-E4P). The steady state kinetic parameters of this reaction were determined and the enzyme was used to prepare L-E4P which was shown to be a weak inhibitor of 2-C-methyl-D-erythritol-4-phosphate cytidyltransferase (YgbP).     

Atypical antiinflammatory activation of microglia induced by apoptotic neurons

In the central nervous system (CNS), apoptosis plays an important role during development and is a primary pathogenic mechanism in several adult neurodegenerative diseases. A main feature of apoptotic cell death is the efficient and fast removal of dying cells by macrophages and nonprofessional phagocytes, without eliciting inflammation in the surrounding tissue. Apoptotic cells undergo several membrane changes, including the externalization of so-called "eat me" signals whose cognate receptors are present on professional phagocytes. Among these signals, the aminophospholipid phosphatidylserine (PS) appears to have a crucial and unique role in preventing the classical pro-inflammatory activation of macrophages, thus ensuring the silent and safe removal of apoptotic cells. Although extensively studied in the peripheral organs, the process of recognition and removal of apoptotic cells in the brain has only recently begun to be unraveled. Here, we summarize the evidence suggesting that upon interaction with PS-expressing apoptotic neurons, microglia may no longer promote the inflammatory cascade, but rather facilitate the elimination of damaged neurons through antiinflammatory and neuroprotective functions. We propose that the anti-inflammatory microglial phenotype induced through the activation of the specific PS receptor (PtdSerR), expressed by resting and activated microglial cells, could be relevant to the final outcome of neurodegenerative diseases, in which apoptosis seems to play a crucial role.