Ribosomal Tag Pyrosequencing of DNA and RNA Reveals “Rare” Taxa with High Protein Synthesis Potential in the Sediment of a Hypersaline Lake in Western Australia

Little is known about the potential activity of microbial communities in hypersaline sediment ecosystems. Ribosomal tag libraries of DNA and RNA extracted from the sediment of Lake Strawbridge (Western Australia) revealed bacterial and archaeal operational taxonomic units (OTUs) with high RNA/DNA ratios providing evidence for the presence of ‘rare’ but potentially “active” taxa. Among the ‘rare’ bacterial taxa Halomonas, Salinivibrio and Idiomarina showed the highest protein synthesis potential. Rare but ‘active’ archaeal OTUs were related to the KTK 4A cluster and the Marine-Benthic-Groups B and D. We present the first molecular analysis of the microbial diversity and protein synthesis potential of rare microbial taxa in a hypersaline sediment ecosystem.     

Regulation of Gag- and Env-Specific CD8+ T Cell Responses in ART-Na?ve HIV-Infected Patients: Potential Implications for Individualized Immunotherapy

Strategies to develop a functional cure for HIV infection will likely require boosting of effector T cell responses to eliminate reactivated, latently infected cells. We have recently explored an assay for assessing antigen-specific regulation of T cell proliferation, which was related to clinical progression in untreated patients and to vaccine efficacy in two trials of therapeutic Gag-based vaccines. We here expand the same assay to further investigate regulation mediated by various inhibitory pathways. Peripheral blood mononuclear cells from 26 asymptomatic HIV-infected, antiretroviral therapy-naïve patients were stimulated with Gag and Env overlapping peptide panels for 5 days. Monoclonal antibodies (mAbs) blocking inhibitory mediators interleukin (IL) 10, transforming growth factor (TGF) β, programmed death ligand (PD–L) 1 and herpes virus entry mediator (HVEM) were added to parallel cultures. Functional T cell regulation (FTR) was defined as the difference in proliferation between stimulated cultures with and without blocking mAbs. FTR was detected in 54% of patients. Blockade of IL-10/PD-L1 and IL10/TGF-β detected all cases with Gag- and Env-associated FTR, respectively. In accordance with previous findings, isolated Env FTR was associated with higher plasma HIV RNA and lower CD4 counts, while patients with both Gag and Env FTR also had higher Gag- and Env-specific proliferative CD8⁺ T cell responses. There was no association between FTR and frequencies of activated regulatory T cells. In conclusion, we observed substantial heterogeneity in FTR between patients, inhibitory pathways and HIV antigens. FTR may help to individualize immunomodulation and warrants further assessment in clinical immunotherapy trials.     

An infrared sensor analysing label‐free the secondary structure of the Abeta peptide in presence of complex fluids

The secondary structure change of the Abeta peptide to beta‐sheet was proposed as an early event in Alzheimer's disease. The transition may be used for diagnostics of this disease in an early state. We present an Attenuated Total Reflection (ATR) sensor modified with a specific antibody to extract minute amounts of Abeta peptide out of a complex fluid. Thereby, the Abeta peptide secondary structure was determined in its physiological aqueous environment by FTIR‐difference‐spectroscopy. The presented results open the door for label‐free Alzheimer diagnostics in cerebrospinal fluid or blood. It can be extended to further neurodegenerative diseases.

An immunologic ATR‐FTIR sensor for Abeta peptide secondary structure analysis in complex fluids is presented.     

Consistent performance of invasive plant species within and among islands of the Mediterranean basin

Since the success of an invasive species depends not only upon its intrinsic traits but also on particular characteristics of the recipient habitat, assessing the performance of an invader across habitats provides a more realistic analysis of risk. Such an analysis will not only provide insights into the traits related to invasiveness, but also the habitat characteristics that underpin vulnerability to invasion that, taken together, will facilitate the selection of management strategies to mitigate the invader’s effect. In the present study, we considered the Mediterranean basin islands as an excellent study region to test how the same invasive species perform in different habitats within a single island, and to scale up differences among islands with similar climate. We tested how the performance of three widespread plant invaders with clonal growth but contrasting life-history traits, a deciduous tree Ailanthus altissima, a succulent subshrub Carpobrotus spp., and an annual geophyte Oxalis pes-caprae, varied depending upon the species identity, habitat, and invaded island. The environmental parameters considered were habitat type, elevation, species diversity in the invaded plot, and several soil traits (% C, % N, C/N, pH, and relative humidity). The study documents that the performance of these three important and widespread plant invaders is dependent mainly on species identity, and less upon the invaded island’s general features. Likewise, differences in performance among habitats were only significant in the case of Ailanthus, whereas Carpobrotus and Oxalis appear to perform equally well in different environments. Ailanthus thus appears to have a broader spectrum of invasiveness, being able to invade a larger number of habitat types. On the contrary, Carpobrotus spp. have not yet invaded habitats different from those where the species have been originally introduced and where they are still commonly spread by humans. Oxalis distribution is mainly related to agricultural activities and disturbed sites, and the total area infested by this geophyte may be more reflection of the extent of suitable habitats than of invasiveness or ecological impact. Our results confirm the potential for these species to significantly alter the functioning of ecosystems in the Mediterranean islands and highlight the risk to other islands not yet invaded.     

Characterisation of the tryptophan synthase alpha subunit in maize

Background  

In bacteria, such as Salmonella typhimurium, tryptophan is synthesized from indole-3-glycerole phosphate (IGP) by a tryptophan synthase αββα heterotetramer. Plants have evolved multiple α (TSA) and β (TSB) homologs, which have probably diverged in biological function and their ability of subunit interaction. There is some evidence for a tryptophan synthase (TS) complex in Arabidopsis. On the other hand maize (Zea mays) expresses the TSA-homologs BX1 and IGL that efficiently cleave IGP, independent of interaction with TSB.     

Key regulatory molecules of cartilage destruction in rheumatoid arthritis: an <Emphasis Type="Italic">in vitro</Emphasis>study

Background  

Rheumatoid arthritis (RA) is a chronic, inflammatory and systemic autoimmune disease that leads to progressive cartilage destruction. Advances in the treatment of RA-related destruction of cartilage require profound insights into the molecular mechanisms involved in cartilage degradation. Until now, comprehensive data about the molecular RA-related dysfunction of chondrocytes have been limited. Hence, the objective of this study was to establish a standardized in vitro model to profile the key regulatory molecules of RA-related destruction of cartilage that are expressed by human chondrocytes.     

Geographical and technological differences in life cycle inventories shown by the use of process models for waste incinerators part I. technological and geographical differences

Goal and Background  Geographical and technological differences in Life Cycle Inventory data are an important source for uncertainty in the result of Life Cycle Assessments. Knowledge on their impact on the result of an LCA is scarce, and also knowledge on how to manage them in an LCA case study. Objective  Goal of this paper is to explore these differences for municipal solid waste incinerator plants, and to develop recommendations for managing technological and geographical differences. Methodology  The paper provides a definition of technological and geographical differences, and analyses their possible impacts. In a case study, the differences are caused intentionally in ‘games’, by virtually transplanting incineration plants to a different location and by changing parameters such as the composition of the waste input incinerated. The games are performed by using a modular model for municipal solid waste incinerator plants. In each case, an LCA including an Impact Assessment is calculated to trace the impact of these changes, and the results are compared. Conclusions  The conclusions of the paper are two-fold: (1) reduce the differences in inventory data where their impact on the result is high; where it is possible reducing them to a great extent, and the effort for performing the change acceptable; in the case of incineration plants: Adapt the flue gas treatment, especially a possible DeNOx step, to the real conditions; (2) make use of modular process models that allow adapting plant parameters to better meet real conditions, but be aware of possible modelling errors. The paper invites the scientific community to validate the model used for a waste incinerator plant, and suggest putting up similar models for other processes, preferably those of similar relevance for Life Cycle Inventories.     

PvuII-endonuclease induces structural alterations at the scissile phosphate group of its cognate DNA

We investigated the PvuII endonuclease with its cognate DNA by means of molecular dynamics simulations. Comparing the complexed DNA with a reference simulation of free DNA, we saw structural changes at the scissile phosphodiester bond. At this GpC step, the enzyme induces the highest twist and axial rise, inclination is increased and the minor groove widened. The distance between the scissile phosphate group and the phosphate group of the following thymine base is shortened significantly, indicating a substrate-assisted catalysis. A feasible reason for this vicinity is the catalytically important amino acid residue lysine 70, which bridges the free oxygen atoms of the successive phosphate groups. Due to this geometry, a compact reaction pocket is formed where a water molecule can be held, thus bringing the reaction partners for hydrolysis into contact. The O1-P-O2 angle of the scissile nucleotide is decreased, probably due to a complexation of the negative oxygen atoms through protein and solvent contacts.     

Identification of oxidized methionine residues in peptides containing two methionine residues by derivatization and matrix-assisted laser desorption/ionization mass spectrometry

Oxidation of methionine residues in peptides and proteins occurs in vivo or may be an artifact resulting from purification steps. We present a three step method for the localization of methionine sulfoxides in peptides with two methionine residues. In the first step, the N-terminus as well as other reactive side chain functions are blocked by acetylation. The resulting protected peptides are cleaved by cyanogen bromide. The cleavage does not occur at methionine sulfoxide but only at reduced methionine residues forming new amino termini. The newly formed amino group is then derivatized with a bromine containing compound in the last step of the procedure. The resulting peptide can easily be identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry using both the characteristic isotope pattern of the halogen and the metastable loss of methanesulfenic acid from oxidized residues. This procedure allows the unequivocal localization of oxidized methionines even in complex peptide mixtures.