Nanobodies with cross-neutralizing activity provide prominent therapeutic efficacy in mild and severe COVID-19 rodent models.

The weakened protective efficacy of COVID-19 vaccines and antibodies caused by SARS-CoV-2 variants presents a global health emergency, which underscores the urgent need for universal therapeutic antibody intervention for clinical patients. Here, we screened three alpacas-derived nanobodies (Nbs) with neutralizing activity from twenty RBD-specific Nbs. The three Nbs were fused with the Fc domain of human IgG, namely aVHH-11-Fc, aVHH-13-Fc and aVHH-14-Fc, which could specifically bind RBD protein and competitively inhibit the binding of ACE2 receptor to RBD. They effectively neutralized SARS-CoV-2 pseudoviruses D614G, Alpha, Beta, Gamma, Delta, and Omicron sub-lineages BA.1, BA.2, BA.4, and BA.5 and authentic SARS-CoV-2 prototype, Delta, and Omicron BA.1, BA.2 strains. In mice-adapted COVID-19 severe model, intranasal administration of aVHH-11-Fc, aVHH-13-Fc and aVHH-14-Fc effectively protected mice from lethal challenges and reduced viral loads in both the upper and lower respiratory tracts. In the COVID-19 mild model, aVHH-13-Fc, which represents the optimal neutralizing activity among the above three Nbs, effectively protected hamsters from the challenge of SARS-CoV-2 prototype, Delta, Omicron BA.1 and BA.2 by significantly reducing viral replication and pathological alterations in the lungs. In structural modeling of aVHH-13 and RBD, aVHH-13 binds to the receptor-binding motif region of RBD and interacts with some highly conserved epitopes. Taken together, our study illustrated that alpaca-derived Nbs offered a therapeutic countermeasure against SARS-CoV-2, including those Delta and Omicron variants which have evolved into global pandemic strains.

GEM-PA-Based Subunit Vaccines of Crimean Congo Hemorrhagic Fever Induces Systemic Immune Responses in Mice.

The Crimean Congo Hemorrhagic Fever Virus (CCHFV) is a tick-borne bunyavirus of the Narovirus genus, which is the causative agent of Crimean Congo Hemorrhagic Fever (CCHF). CCHF is endemic in Africa, the Middle East, Eastern Europe and Asia, with a high case-fatality rate of up to 50% in humans. Currently, there are no approved vaccines or effective therapies available for CCHF. The GEM-PA is a safe, versatile and effective carrier system, which offers a cost-efficient, high-throughput platform for recovery and purification of subunit proteins for vaccines. In the present study, based on a GEM-PA surface display system, a GEM-PA based vaccine expressing three subunit vaccine candidates (G-GP, including G-eGN, G-eGC and G-NAb) of CCHFV was developed, displaying the ectodomains of the structural glycoproteins eGN, eGC and NAb, respectively. According to the immunological assays including indirect-ELISA, a micro-neutralization test of pseudo-virus and ELISpot, 5 µg GPBLP3 combined with Montanide ISA 201VG plus Poly (I:C) adjuvant (A-G-GP-5 µg) elicited GP-specific humoral and cellular immunity in BALB/c mice after three vaccinations via subcutaneous injection (s.c.). The consistent data between IgG subtype and cytokine detection, ELISpot and cytokine detection indicated balanced Th1 and Th2 responses, of which G-eGN vaccines could elicit a stronger T-cell response post-vaccination, respectively. Moreover, all three vaccine candidates elicited high TNF-α, IL-6, and IL-10 cytokine levels in the supernatant of stimulated splenocytes in vitro. However, the neutralizing antibody (nAb) was only detected in A-G-eGC and A-G-eGC vaccination groups with the highest neutralizing titer of 128, suggesting that G-eGC could elicit a stronger humoral immune response. In conclusion, the GEM-PA surface display system could provide an efficient and convenient purification method for CCHFV subunit antigens, and the G-GP subunit vaccine candidates will be promising against CCHFV infections with excellent immunogenicity.

Can we use mine waste as substrate in constructed wetlands to intensify nutrient removal? A critical assessment of key removal mechanisms and long-term environmental risks.

The utilization of natural ores and/or mine waste as substrate in constructed wetlands (CWs) to enhance nutrient removal performance has been gaining high popularity recently. However, the knowledge regarding the long-term feasibility and key removal mechanisms, particularly the potential negative environmental effects of contaminants leached from mine waste is far insufficient. This study, for the first time, performed a critical assessment by using different CWs with three mine waste (coal gangue, iron ore and manganese ore) as substrates in a 385-day experiment treating wastewater with varying nutrient loadings. The results showed that the addition of mine waste in CWs increased removal of total phosphorus (TP) by 17-34%, and total nitrogen (TN) by 11-51%. The higher removal of TP is mainly attributed to the strong binding mechanism of phosphate with the oxides and hydroxides of Mn, Fe and/or Al, which are leached out of mine waste. Moreover, integration of mine waste in CWs also significantly stimulated biofilm establishment and enriched the relative abundance of key functional genes related to the nitrogen cycle, supporting the observed high-rate nitrogen removal. However, leaching of heavy metals (Fe, Mn, Cu and Cr) from the beded mine waste in the experimented CWs was monitored, which further influenced cytoplasmic enzymes and created oxidative stress damage to plants, resulting in a decline of nutrient uptake by plants.

Viral and Host Transcriptomes in SARS-CoV-2-Infected Human Lung Cells.

Coronaviruses are commonly characterized by a unique discontinuous RNA transcriptional synthesis strategy guided by transcription-regulating sequences (TRSs). However, the details of RNA synthesis in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have not been fully elucidated. Here, we present a time-scaled, gene-comparable transcriptome of SARS-CoV-2, demonstrating that ACGAAC functions as a core TRS guiding the discontinuous RNA synthesis of SARS-CoV-2 from a holistic perspective. During infection, viral transcription, rather than genome replication, dominates all viral RNA synthesis activities. The most highly expressed viral gene is the nucleocapsid gene, followed by ORF7 and ORF3 genes, while the envelope gene shows the lowest expression. Host transcription dysregulation keeps exacerbating after viral RNA synthesis reaches a maximum. The most enriched host pathways are metabolism related. Two of them (cholesterol and valine metabolism) affect viral replication in reverse. Furthermore, the activation of numerous cytokines emerges before large-scale viral RNA synthesis. IMPORTANCE SARS-CoV-2 is responsible for the current severe global health emergency that began at the end of 2019. Although the universal transcriptional strategies of coronaviruses are preliminarily understood, the details of RNA synthesis, especially the time-matched transcription level of each SARS-CoV-2 gene and the principles of subgenomic mRNA synthesis, are not clear. The coterminal subgenomic mRNAs of SARS-CoV-2 present obstacles in identifying the expression of most genes by PCR-based methods, which are exacerbated by the lack of related antibodies. Moreover, SARS-CoV-2-related metabolic imbalance and cytokine storm are receiving increasing attention from both clinical and mechanistic perspectives. Our transcriptomic research provides information on both viral RNA synthesis and host responses, in which the transcription-regulating sequences and transcription levels of viral genes are demonstrated, and the metabolic dysregulation and cytokine levels identified at the host cellular level support the development of novel medical treatment strategies.

Improvement of wheat seed vitality by dielectric barrier discharge plasma treatment.

Influences of discharge voltage on wheat seed vitality were investigated in a dielectric barrier discharge (DBD) plasma system at atmospheric pressure and temperature. Six different treatments were designed, and their discharge voltages were 0.0, 9.0, 11.0, 13.0, 15.0, and 17.0 kV, respectively. Fifty seeds were exposed to the DBD plasma atmosphere with an air flow rate of 1.5 L min-1 for 4 min in each treatment, and then the DBD plasma-treated seeds were prepared for germination in several Petri dishes. Each treatment was repeated three times. Germination indexes, growth indexes, surface topography, water uptake, permeability, and α-amylase activity were measured. DBD plasma treatment at appropriate energy levels had positive effects on wheat seed germination and seedling growth. The germination potential, germination index, and vigor index significantly increased by 31.4%, 13.9%, and 54.6% after DBD treatment at 11.0 kV, respectively, in comparison to the control. Shoot length, root length, dry weight, and fresh weight also significantly increased after the DBD plasma treatment. The seed coat was softened and cracks were observed, systematization of the protein was strengthened, and amount of free starch grain increased after the DBD plasma treatment. Water uptake, relative electroconductivity, soluble protein, and α-amylase activity of the wheat seed were also significantly improved after the DBD plasma treatment. Roles of active species and ultraviolet radiation generated in the DBD plasma process in wheat seed germination and seedling growth are proposed. Bioelectromagnetics. 39:120-131, 2018. © 2017 Wiley Periodicals, Inc.

Alleviation of adverse effects of drought stress on wheat seed germination using atmospheric dielectric barrier discharge plasma treatment.

Atmospheric dielectric barrier discharge (DBD) was attempted to improve the resistance of wheat seed to drought stress. Effects of DBD plasma on wheat seed germination, seedling growth, osmotic-adjustment products, lipid peroxidation, reactive oxygen species (ROS), antioxidant enzyme activity, abscisic acid, and drought resistant related genes expression under drought stress were investigated. The changes of the wheat seed coat before and after the DBD plasma treatment were explored. Experimental results showed that the DBD plasma treatment could alleviate the adverse effects of drought stress on wheat seed germination and seedling growth; the germination potential and germination rate increased by 27.2% and 27.6%, and the root length and shoot length of the wheat seedlings also increased. Proline and soluble sugar levels under drought stress were improved after the DBD plasma treatment, whereas the malondialdehyde content decreased. ROS contents under drought stress were reduced after the DBD plasma treatment, whereas the activities of superoxide dismutase, catalase, and peroxidase were promoted. DBD plasma treatment promoted abscisic acid generation in wheat seedlings, and it also regulated functional gene LEA1 and stimulated regulation genes SnRK2 and P5CS to resist drought stress. Etching effect and surface modification occurred on the seed coat after the DBD plasma treatment.

Enhanced degradation of azo dye in wastewater by pulsed discharge plasma coupled with MWCNTs-TiO2/γ-Al2O3 composite photocatalyst.

In order to improve the photocatalytic performance of TiO2 in pulsed discharge plasma systems, easily recycled multi-walled carbon nanotubes (MWCNTs)-TiO2 supported on γ-Al2O3 (MWCNTs-TiO2/γ-Al2O3) composite photocatalyst were prepared. The morphology and physicochemical properties of the prepared catalysts were investigated using XRD, SEM, FTIR and UV-vis spectroscopy. The photocatalytic activity was evaluated by degradation of azo dye acid orange II (AO7) in wastewater under pulsed discharge plasma. The results indicate that the MWCNTs-TiO2/γ-Al2O3 composite catalyst possesses enhanced photocatalytic activity facilitating the decomposition of AO7 compared with TiO2/γ-Al2O3 composite in pulsed discharge plasma systems. Under pulsed discharge plasma, almost 100% AO7 is degraded by the MWCNTs-TiO2/γ-Al2O3 composite after 60 min at optimal conditions. The degradation efficiency of AO7 is also affected by the dosage of the composite catalyst and pulsed discharge peak voltage. As the amount of MWCNTs-TiO2/γ-Al2O3 composite and pulsed discharge peak voltage increases, the degradation efficiency of AO7 increases. The photocatalyst was implemented for 6 cycles and the degradation efficiency of AO7 remains higher than 85% under pulsed discharge plasma. Results indicate that the catalyst displays easy separation and minimal deactivation after several uses. Possible decomposition mechanisms were also investigated. MWCNTs are capable of improving the photocatalytic activity of TiO2/γ-Al2O3 composite in pulsed discharge plasma systems primarily due to the photo-induced-electron absorption effect and the electron trap effect of MWCNTs. The results of this study establish the feasibility and potential implementation of MWCNTs-TiO2/γ-Al2O3 composites in pulsed discharge plasma systems for the degradation of dye wastewater.

Research on dye wastewater decoloration by pulse discharge plasma combined with charcoal derived from spent tea leaves.

Pulsed discharge plasma (PDP) combined with charcoal (PDP-charcoal) was employed to treat dye wastewater, with methyl orange (MO) as the model pollutant. The charcoal was prepared using spent tea leaves and was characterized by scanning electron microscopy, Fourier-transform infrared spectroscopy, and Boehm titration to investigate the adsorption and catalytic characteristics before and after adsorption and PDP treatment. The prepared charcoal exhibited a high MO adsorption capacity, and the adsorption process followed the pseudo-second-order kinetic model and the Freundlich model. The MO decoloration efficiency reached 69.8 % within 7.5 min of treatment in the PDP-charcoal system, whereas values of 29.2 and 25.9 % were achieved in individual PDP and charcoal systems, respectively. The addition of n-butanol and H2PO4 (-) presented inhibitive effects on MO decoloration in the PDP system. However, these effects were much weaker in the PDP-charcoal system. In addition, the effects of charcoal on O3 and H2O2 formation were evaluated, and the results showed that both the O3 and H2O2 concentrations decreased in the presence of charcoal. The MO decomposition intermediates were analyzed using UV-Vis spectrometry and GC-MS. 1,4-Benzoquinone, 4-nitrophenol, 4-hydroxyaniline, and N,N'-dimethylaniline were detected. A possible pathway for MO decomposition in this system was proposed.