Development of a Dot-Blot Assay for the Detection of Mould-Specific IgE in the Belgian Population

Background

Data on mould sensitization in the general population are scarce and mostly on Aspergillus fumigatus, Alternaria alternata and Cladosporium herbarum.

Objectives

To validate a dot-blot assay for the detection of specific IgE and evaluate the prevalence of mould sensitization in a healthy population.

Methods

The dot-blot assay was validated against the CAP test. Sensitization rate to ten common indoor and outdoor mould species in 344 serum samples was calculated. For each serum with more than one reactivity, the “major sensitization” defined as the strongest response against a single mould species was calculated.

Results

Intra- and inter-assay variations were both below 20%, and the positivity threshold of the test was of 0.418 kU/L for A. fumigatus. Correlation with CAP results was strong. The overall prevalence of sensitization was 32.8%, and the commonest sensitizations were against A. alternaria, A. flavus and A. niger (around 15%). The most frequent “major reactivities” were against A. niger and A. alternata (20–30%). In 25.1% of the samples, “major reactivities” were directed against a group of moulds commonly found indoor (Penicillium spp., Aspergillus versicolor, Cladosporium sphaerospermum and Cladosporium cladosporioides).

Conclusions

The dot-blot assay was validated for the detection of mould-specific IgE. In the general population, sensitization to indoor species was common and accounted for 25% of overall mould sensitizations.

     

Biochemical characterization of metabolism‐based atrazine resistance in Amaranthus tuberculatus and identification of an expressed GST associated with resistance

Rapid detoxification of atrazine in naturally tolerant crops such as maize (Zea mays) and grain sorghum (Sorghum bicolor) results from glutathione S‐transferase (GST) activity. In previous research, two atrazine‐resistant waterhemp (Amaranthus tuberculatus) populations from Illinois, U.S.A. (designated ACR and MCR), displayed rapid formation of atrazine‐glutathione (GSH) conjugates, implicating elevated rates of metabolism as the resistance mechanism. Our main objective was to utilize protein purification combined with qualitative proteomics to investigate the hypothesis that enhanced atrazine detoxification, catalysed by distinct GSTs, confers resistance in ACR and MCR. Additionally, candidate AtuGST expression was analysed in an F₂ population segregating for atrazine resistance. ACR and MCR showed higher specific activities towards atrazine in partially purified ammonium sulphate and GSH affinity‐purified fractions compared to an atrazine‐sensitive population (WCS). One‐dimensional electrophoresis of these fractions displayed an approximate 26‐kDa band, typical of GST subunits. Several phi‐ and tau‐class GSTs were identified by LC‐MS/MS from each population, based on peptide similarity with GSTs from Arabidopsis. Elevated constitutive expression of one phi‐class GST, named AtuGSTF2, correlated strongly with atrazine resistance in ACR and MCR and segregating F₂ population. These results indicate that AtuGSTF2 may be linked to a metabolic mechanism that confers atrazine resistance in ACR and MCR.     

Polymer:Nonfullerene Bulk Heterojunction Solar Cells with Exceptionally Low Recombination Rates

Organic semiconductors are in general known to have an inherently lower charge carrier mobility compared to their inorganic counterparts. Bimolecular recombination of holes and electrons is an important loss mechanism and can often be described by the Langevin recombination model. Here, the device physics of bulk heterojunction solar cells based on a nonfullerene acceptor (IDTBR) in combination with poly(3‐hexylthiophene) (P3HT) are elucidated, showing an unprecedentedly low bimolecular recombination rate. The high fill factor observed (above 65%) is attributed to non‐Langevin behavior with a Langevin prefactor (β/β_L) of 1.9 × 10^?4. The absence of parasitic recombination and high charge carrier lifetimes in P3HT:IDTBR solar cells inform an almost ideal bimolecular recombination behavior. This exceptional recombination behavior is explored to fabricate devices with layer thicknesses up to 450 nm without significant performance losses. The determination of the photoexcited carrier mobility by time‐of‐flight measurements reveals a long‐lived and nonthermalized carrier transport as the origin for the exceptional transport physics. The crystalline microstructure arrangement of both components is suggested to be decisive for this slow recombination dynamics. Further, the thickness‐independent power conversion efficiency is of utmost technological relevance for upscaling production and reiterates the importance of understanding material design in the context of low bimolecular recombination.     

Appointment of New Features Editor

Transporters are essential players in bacterial growth and survival, since they are key for uptake of nutrients on the one hand, and for defence against endogenous and environmental stresses on the other hand. Remarkably, in addition to their primary role in substrate translocation, it has become clear that some transporters have acquired a secondary function as sensors and information processors in signalling pathways. In this review, we describe recent advances in our understanding of the role of transporters in such signalling cascades, and discuss some of the emergent dynamic behaviour found in hallmark examples. A particular focus is placed on new insights into mechanistic details of information transfer between transporters and regulatory proteins. Quantitative considerations reveal that these signalling complexes can implement a remarkable diversity of regulatory logic functions, where the transporter can act as activity switch, as positive or negative reporter of transport flux, or as a signalling hub for the integration of multiple inputs. Such a dual use of transport proteins not only enables efficient substrate translocation but is also an elegant strategy to integrate important information about the cell's external conditions with its current physiological state.     

Isolation and characterization of iron chelators from turmeric (<Emphasis Type="Italic">Curcuma longa</Emphasis>): selective metal binding by curcuminoids

Iron overload disorders may be treated by chelation therapy. This study describes a novel method for isolating iron chelators from complex mixtures including plant extracts. We demonstrate the one-step isolation of curcuminoids from turmeric, the medicinal food spice derived from Curcuma longa. The method uses iron-nitrilotriacetic acid (NTA)-agarose, to which curcumin binds rapidly, specifically, and reversibly. Curcumin, demethoxycurcumin, and bisdemethoxycurcumin each bound iron-NTA-agarose with comparable affinities and a stoichiometry near 1. Analyses of binding efficiencies and purity demonstrated that curcuminoids comprise the primary iron binding compounds recovered from a crude turmeric extract. Competition of curcuminoid binding to the iron resin was used to characterize the metal binding site on curcumin and to detect iron binding by added chelators. Curcumin-Iron-NTA-agarose binding was inhibited by other metals with relative potency: (>90% inhibition) Cu²⁺ ~ Al³⁺ > Zn²⁺ ≥ Ca²⁺ ~ Mg²⁺ ~ Mn²⁺ (<20% inhibition). Binding was also inhibited by pharmaceutical iron chelators (desferoxamine or EDTA) or by higher concentrations of weak iron chelators (citrate or silibinin). Investigation of the physiological effects of iron binding by curcumin revealed that curcumin uptake by cultured cells was reduced >80% by addition of iron to the media; uptake was completely restored by desferoxamine. Ranking of metals by relative potencies for blocking curcumin uptake agreed with their relative potencies in blocking curcumin binding to iron-NTA-agarose. We conclude that curcumin can selectively bind toxic metals including iron in a physiological setting, and propose inhibition of curcumin binding to iron-NTA-agarose for iron chelator screening.     

A mechanistic model of snakebite as a zoonosis: Envenoming incidence is driven by snake ecology,socioeconomics and its impacts on snakes

Snakebite is the only WHO-listed, not infectious neglected tropical disease (NTD), although its eco-epidemiology is similar to that of zoonotic infections: envenoming occurs after a vertebrate host contacts a human. Accordingly, snakebite risk represents the interaction between snake and human factors, but their quantification has been limited by data availability. Models of infectious disease transmission are instrumental for the mitigation of NTDs and zoonoses. Here, we represented snake-human interactions with disease transmission models to approximate geospatial estimates of snakebite incidence in Sri Lanka, a global hotspot. Snakebites and envenomings are described by the product of snake and human abundance, mirroring directly transmitted zoonoses. We found that human-snake contact rates vary according to land cover (surrogate of occupation and socioeconomic status), the impacts of humans and climate on snake abundance, and by snake species. Our findings show that modelling snakebite as zoonosis provides a mechanistic eco-epidemiological basis to understand snakebites, and the possible implications of global environmental and demographic change for the burden of snakebite.     

The colonic pathogen Entamoeba histolytica activates caspase-4/1 that cleaves the pore-forming protein gasdermin D to regulate IL-1β secretion

A hallmark of Entamoeba histolytica (Eh) invasion in the gut is acute inflammation dominated by the secretion of pro-inflammatory cytokines TNF-α and IL-1β. This is initiated when Eh in contact with macrophages in the lamina propria activates caspase-1 by recruiting the NLRP3 inflammasome complex in a Gal-lectin and EhCP-A5-dependent manner resulting in the maturation and secretion of IL-1β and IL-18. Here, we interrogated the requirements and mechanisms for Eh-induced caspase-4/1 activation in the cleavage of gasdermin D (GSDMD) to regulate bioactive IL-1β release in the absence of cell death in human macrophages. Unlike caspase-1, caspase-4 activation occurred as early as 10 min that was dependent on Eh Gal-lectin and EhCP-A5 binding to macrophages. By utilizing CRISPR-Cas9 gene edited CASP4/1, NLRP3 KO and ASC-def cells, caspase-4 activation was found to be independent of the canonical NLRP3 inflammasomes. In CRISPR-Cas9 gene edited CASP1 macrophages, caspase-4 activation was significantly up regulated that enhanced the enzymatic cleavage of GSDMD at the same cleavage site as caspase-1 to induce GSDMD pore formation and sustained bioactive IL-1β secretion. Eh-induced IL-1β secretion was independent of pyroptosis as revealed by pharmacological blockade of GSDMD pore formation and in CRISPR-Cas9 gene edited GSDMD KO macrophages. This was in marked contrast to the potent positive control, lipopolysaccharide + Nigericin that induced high expression of predominantly caspase-1 that efficiently cleaved GSDMD with high IL-1β secretion/release associated with massive cell pyroptosis. These results reveal that Eh triggered “hyperactivated macrophages” allowed caspase-4 dependent cleavage of GSDMD and IL-1β secretion to occur in the absence of pyroptosis that may play an important role in disease pathogenesis.     

Influence of human model resolution on computed currents induced in organs by 60-Hz magnetic fields

The effects of human body model resolution on computed electric fields induced by 60 Hz uniform magnetic fields are investigated. A recently-developed scalar potential finite difference code for low-frequency electromagnetic computations is used to model induction in two anatomically realistic human body models. The first model consists of 204 290 cubic voxels with 7.2-mm edges, while the second comprises 1 639 146 cubic voxels with 3.6-mm edges. Calculations on the lower-resolution model using, for example, the finite difference time domain or impedance methods, push the capabilities of workstations. The scalar method, in contrast, can handle the higher-resolution model using comparable resources. The results are given in terms of average and maximum electric field intensities and current density magnitudes in selected tissues and organs. Although the lower-resolution model provides generally acceptable results, there are important differences that make the added computational burden of the higher-resolution calculations worthwhile. In particular, the higher-resolution modelling generally predicts peak electric fields intensities and current density magnitudes that are slightly higher than those computed using the lower-resolution modelling. The differences can be quite large for small organs such as glands. Bioelectromagnetics 18:478–490, 1997. © 1997 Wiley-Liss, Inc.     

The tandem PDZ protein Syntenin interacts with the aminoacyl tRNA synthetase complex in a lysyl-tRNA synthetase-dependent manner

Syntenin-1 is a tandem PDZ protein that binds a diverse array of signaling molecules that are often associated with cell adhesion and intracellular trafficking. With the use of a MS-based functional proteomics approach, we identified several members of the aminoacyl-tRNA synthetase macromolecular (ARS) complex in a syntenin-1 pull down assay. Interaction of these proteins with syntenin-1 was confirmed by co-immunoprecipitation from cultured cells. We demonstrate a direct interaction of syntenin-1 with lysyl-tRNA synthetase (KRS), which contains a PDZ binding motif at its C-terminus. This motif is important for the interaction of the entire complex with syntenin-1. A point mutation in the PDZ2 domain of syntenin-1 abrogates interaction with KRS. As a result, other components of the ARS complex no longer co-immunoprecipitate with syntenin-1. We further show that syntenin-1 regulates KRS activity. These findings suggest that syntenin-1 is an adaptor modulating the activity of KRS.