Ion-channel function and cross-species determinants in viral assembly of nonprimate hepacivirus p7

Nonprimate hepacivirus (NPHV), the closest homolog of hepatitis C virus (HCV) described to date, has recently been discovered in horses. Even though the two viruses share a similar genomic organization, conservation of the encoded hepaciviral proteins remains undetermined. The HCV p7 protein is localized within endoplasmic reticulum (ER) membranes and is important for the production of infectious particles. In this study, we analyzed the structural and functional features of NPHV p7 in addition to its role during virus assembly. Three-dimensional homology models for NPHV p7 using various nuclear magnetic resonance spectroscopy (NMR) structures were generated, highlighting the conserved residues important for ion channel function. By applying a liposome permeability assay, we observed that NPHV p7 exhibited liposome permeability features similar to those of HCV p7, indicative of similar ion channel activity. Next, we characterized the viral protein using a p7-based trans-complementation approach. A similar subcellular localization pattern at the ER membrane was observed, although production of infectious particles was likely hindered by genetic incompatibilities with HCV proteins. To further characterize these cross-species constraints, chimeric viruses were constructed by substituting different regions of HCV p7 with NPHV p7. The N terminus and transmembrane domains were nonexchangeable and therefore constitute a cross-species barrier in hepaciviral assembly. In contrast, the basic loop and the C terminus of NPHV p7 were readily exchangeable, allowing production of infectious trans-complemented viral particles. In conclusion, comparison of NPHV and HCV p7 revealed structural and functional homology of these proteins, including liposome permeability, and broadly acting determinants that modulate hepaciviral virion assembly and contribute to the host-species barrier were identified

Comparison of different in situ hybridization techniques for the detection of various RNA and DNA viruses

In situ hybridization (ISH) is a technique to determine potential correlations between viruses and lesions. The aim of the study was to compare ISH techniques for the detection of various viruses in different tissues. Tested RNA viruses include atypical porcine pestivirus (APPV) in the cerebellum of pigs, equine and bovine hepacivirus (EqHV, BovHepV) in the liver of horses and cattle, respectively, and Schmallenberg virus (SBV) in the cerebrum of goats. Examined DNA viruses comprise canine bocavirus 2 (CBoV-2) in the intestine of dogs, porcine bocavirus (PBoV) in the spinal cord of pigs and porcine circovirus 2 (PCV-2) in cerebrum, lymph node, and lung of pigs. ISH with self-designed digoxigenin-labelled RNA probe

Status, sources and contamination levels of organochlorine pesticide residues in urban and agricultural areas: a preliminary review in central–southern Italian soils

Organochlorine pesticides (OCPs) are synthetic chemicals commonly used in agricultural activities to kill pests and are persistent organic pollutants (POPs). They can be detected in different environmental media, but soil is considered an important reservoir due to its retention capacity. Many different types of OCPs exist, which can have different origins and pathways in the environment. It is therefore important to study their distribution and behaviour in the environment, starting to build a picture of the potential human health risk in different contexts. This study aimed at investigating the regional distribution, possible sources and contamination levels of 24 OCP compounds in urban and rural soils from central and southern Italy. One hundred and forty-eight topsoil samples (0–20 cm top layer) from 78 urban and 70 rural areas in 11 administrative regions were collected and analysed by gas chromatography–electron capture detector (GC–ECD). Total OCP residues in soils ranged from nd (no detected) to 1043 ng/g with a mean of 29.91 ng/g and from nd to 1914 ng/g with a mean of 60.16 ng/g in urban and rural area, respectively. Endosulfan was the prevailing OCP in urban areas, followed by DDTs, Drins, Methoxychlor, HCHs, Chlordane-related compounds and HCB. In rural areas, the order of concentrations was Drins > DDTs > Methoxychlor > Endosulfans > HCHs > Chlordanes > HCB. Diagnostic ratios and robust multivariate analyses revealed that DDT in soils could be related to historical application, whilst (illegal) use of technical DDT or dicofol may still occur in some urban areas. HCH residues could be related to both historical use and recent application, whilst there was evidence that modest (yet significant) application of commercial technical HCH may still be happening in urban areas. Drins and Chlordane compounds appeared to be mostly related to historical application, whilst Endosulfan presented a complex mix of results, indicating mainly historical origin in rural areas as well as potential recent applications on urban areas. Contamination levels were quantified by Soil Quality Index (SoQI), identifying high levels in rural areas of Campania and Apulia, possibly due to the intensive nature of some agricultural practices in those regions (e.g., vineyards and olive plantations). The results from this study (which is in progress in the remaining regions of Italy) will provide an invaluable baseline for OCP distribution in Italy and a powerful argument for follow-up studies in contaminated areas. It is also hoped that similar studies will eventually constitute enough evidence to push towards an institutional response for more adequate regulation as well as a full ratification of the Stockholm Convention

Vaccine-associated enhanced respiratory pathology in COVID-19 hamsters after T(H)2-biased immunization

Vaccine-associated enhanced respiratory disease (VAERD) is a severe complication for some respiratory infections. To investigate the potential for VAERD induction in coronavirus disease 2019 (COVID-19), we evaluate two vaccine leads utilizing a severe hamster infection model: a T helper type 1 (T(H)1)-biased measles vaccine-derived candidate and a T(H)2-biased alum-adjuvanted, non-stabilized spike protein. The measles virus (MeV)-derived vaccine protects the animals, but the protein lead induces VAERD, which can be alleviated by dexamethasone treatment. Bulk transcriptomic analysis reveals that our protein vaccine prepares enhanced host gene dysregulation in the lung, exclusively up-regulating mRNAs encoding the eosinophil attractant CCL-11, T(H)2-driving interleukin (IL)-19, or T(H)2 cytokines IL-4, IL-5, and IL-13. Single-cell RNA sequencing (scRNA-seq) identifies lung macrophages or lymphoid cells as sources, respectively. Our findings imply that VAERD is caused by the concerted action of hyperstimulated macrophages and T(H)2 cytokine-secreting lymphoid cells and potentially links VAERD to antibody-dependent enhancement (ADE). In summary, we identify the cytokine drivers and cellular contributors that mediate VAERD after T(H)2-biased vaccination

Über die Inhaltsstoffe des grünen Knollenblätterpilzes, XXXVII. Die absoluten Konfigurationen der in den Phytotoxinen enthaltenen γ-Hydroxyaminosäuren und der γ-Hydroxynorvaline

Durch Analyse der NMR-Spektren von teils aus den Phytotoxinen isolierten, teils synthetisch erhaltenen α-Amino-γ-lactonhydrochloriden ließen sich die absoluten Konfigurationen bestimmen. Der im Phalloidin vorkommende Baustein ist (2S,4R)-4.5-Dihydroxy-leucin (Lacton 3), der des γ-Amanitins (2S,4R)-4-Hydroxy-isoleucin (Lacton 6), der des α-Amanitins (2S,4S)-4.5-Dihydroxy-isoleucin (Lacton 8). Von den beiden diastereomeren Lactonen des γ-Hydroxynorvalins, die sich dünnschichtchromatographisch trennen lassen, leitet sich das langsamer wandernde 9 vom (2S,4R)-4-Hydroxy-norvalin (oder vom Antipoden) ab. Die aus Lathyrus odoratus von L. Fowden isolierte Aminosäure (S. 210) hat die (2S,4S)-Konfiguration. - Die in saurer wäßr. Lösung vorliegenden Gleichgewichte von Lacton und offener Säure wurden für einige γ-Hydroxyaminosäuren NMR-spektroskopisch ermittelt (Tab. 1, S. 212)

Experimental and calculated conformational characteristics of the bicyclic heptapeptide phalloidin

Several conformations generated from approximate potential energy calculations are presented for the bicyclic heptapeptide phalloidin which are consistent with the conformation-dependent information obtained from proton nuclear magnetic resonance measurements performed on phalloidin in dimethylsulfoxide solution. In each conformation, the cysteine amide proton is intramolecularly hydrogen bonded, the tryptophan amide is internally buried and the methyl group of the alanine residue preceding tryptophan is shielded by the tryptophan ring. Thus, phalloidin appears to be a relatively rigid molecule in solution

Spectral study of Na source with a smartphone and a diffraction grating

A home-made spectrometer, based on the use of inexpensive diffraction transmission gratings coupled with a smartphone camera, can be assembled and employed to obtain quantitative spectra of different sources with surprisingly high accuracy to the point of being able to observe the separation of the Na-doublet (the wavelength difference between the two emission D-lines of the sodium spectrum). We describe the set up and data analysis required to acquire the emission spectra of a low-pressure sodium discharge lamp and perform such measurement. The experiment addresses a relevant and engaging topic in physics, namely optical spectroscopy, and is suitable for the undergraduate laboratory and for working with high school students