Immunogenicity in Mice Immunized with Recombinant Adenoviruses Expressing Varicella-Zoster Virus Envelope Glycoprotein E.

Herpes zoster (HZ) is a disease caused by the reactivation of latent varicella-zoster virus (VZV). The subunit vaccine, Shingrix®, and live attenuated vaccine, Zostavax®, could be used as an HZ vaccine that prevents HZ from being developed due to the reactivation of latent VZV in the sensory ganglia due to aging, stress or immunosuppression. In this study, the recombinant adenoviruses rChAd63/gE expressing glycoprotein E (gE) of VZV based on chimpanzee adenovirus serotype 63 (ChAd63) were constructed and investigated for the immunogenicity of different immune pathways in C57BL/6 mice. The results showed similar CD4+ T and CD8+ T cell responses to Shingrix® were induced in mice vaccinated using rChAd63/gE via different immune pathways. This study elucidates that recombinant adenoviruses expressing VZV gE could be appropriate for further development as a new HZ vaccine candidate via different immune pathways.

Comparative analysis of human respiratory syncytial virus evolutionary patterns during the COVID-19 pandemic and pre-pandemic periods.

The COVID-19 pandemic has resulted in the implementation of strict mitigation measures that have impacted the transmission dynamics of human respiratory syncytial virus (HRSV). The measures also have the potential to influence the evolutionary patterns of the virus. In this study, we conducted a comprehensive analysis comparing genomic variations and evolving characteristics of its neutralizing antigens, specifically F and G proteins, before and during the COVID-19 pandemic. Our findings showed that both HRSV A and B exhibited an overall chronological evolutionary pattern. For the sequences obtained during the pandemic period (2019-2022), we observed that the HRSV A distributed in A23 genotype, but formed into three subclusters; whereas the HRSV B sequences were relatively concentrated within genotype B6. Additionally, multiple positively selected sites were detected on F and G proteins but none were located at neutralizing antigenic sites of the F protein. Notably, amino acids within antigenic site III, IV, and V of F protein remained strictly conserved, while some substitutions occurred over time on antigenic site Ø, I, II and VIII; substitution S389P on antigenic site I of HRSV B occurred during the pandemic period with nearly 50% frequency. However, further analysis revealed no substitutions have altered the structural conformations of the antigenic sites, the vial antigenicity has not been changed. We inferred that the intensive public health interventions during the COVID-19 pandemic did not affect the evolutionary mode of HRSV.

Tracking of Mutational Signature of SARS-CoV-2 Omicron on Distinct Continents and Little Difference was Found.

The Omicron variant is currently ravaging the world, raising serious concern globally. Monitoring genomic variations and determining their influence on biological features are critical for tracing its ongoing transmission and facilitating effective measures. Based on large-scale sequences from different continents, this study found that: (i) The genetic diversity of Omicron is much lower than that of the Delta variant. Still, eight deletions (Del 1-8) and 1 insertion, as well as 130 SNPs, were detected on the Omicron genomes, with two deletions (Del 3 and 4) and 38 SNPs commonly detected on all continents and exhibiting high-occurring frequencies. (ii) Four groups of tightly linked SNPs (linkage I-IV) were detected, among which linkage I, containing 38 SNPs, with 6 located in the RBD, increased its occurring frequency remarkably over time. (iii) The third codons of the Omicron shouldered the most mutation pressures, while the second codons presented the least flexibility. (iv) Four major mutants with amino acid substitutions in the RBD were detected, and further structural analysis suggested that the substitutions did not alter the viral receptor binding ability greatly. It was inferred that though the Omicron genome harbored great changes in antigenicity and remarkable ability to evade immunity, it was immune-pressure selected. This study tracked mutational signatures of Omicron variant and the potential biological significance of the SNPs, and the linkages await further functional verification.

The protective immunity induced by intranasally inoculated serotype 63 chimpanzee adenovirus vector expressing human respiratory syncytial virus prefusion fusion glycoprotein in BALB/c mice.

Human respiratory syncytial virus (RSV) is a ubiquitous pediatric pathogen causing serious lower respiratory tract disease worldwide. No licensed vaccine is currently available. In this work, the coding gene for mDS-Dav1, the full-length and prefusion conformation RSV fusion glycoprotein (F), was designed by introducing the stabilized prefusion F (preF) mutations from DS-Cav1 into the encoding gene of wild-type RSV (wtRSV) F protein. The recombinant adenovirus encoding mDS-Cav1, rChAd63-mDS-Cav1, was constructed based on serotype 63 chimpanzee adenovirus vector and characterized in vitro. After immunizing mice via intranasal route, the rChAd63-mDS-Cav1 induced enhanced neutralizing antibody and F-specific CD8+ T cell responses as well as good immune protection against RSV challenge with the absence of enhanced RSV disease (ERD) in BALB/c mice. The results indicate that rChAd63-mDS-Cav1 is a promising mucosal vaccine candidate against RSV infection and warrants further development.

Evolutionary Pattern Comparisons of the SARS-CoV-2 Delta Variant in Countries/Regions with High and Low Vaccine Coverage.

It has been argued that vaccine-breakthrough infections of SARS-CoV-2 would likely accelerate the emergence of novel variants with immune evasion. This study explored the evolutionary patterns of the Delta variant in countries/regions with relatively high and low vaccine coverage based on large-scale sequences. Our results showed that (i) the sequences were grouped into two clusters (L and R); the R cluster was dominant, its proportion increased over time and was higher in the high-vaccine-coverage areas; (ii) genetic diversities in the countries/regions with low vaccine coverage were higher than those in the ones with high vaccine coverage; (iii) unique mutations and co-mutations were detected in different countries/regions; in particular, common co-mutations were exhibited in highly occurring frequencies in the areas with high vaccine coverage and presented in increasing frequencies over time in the areas with low vaccine coverage; (iv) five sites on the S protein were under strong positive selection in different countries/regions, with three in non-C to U sites (I95T, G142D and T950N), and the occurring frequencies of I95T in high vaccine coverage areas were higher, while G142D and T950N were potentially immune-pressure-selected sites; and (v) mutation at the N6-methyladenosine site 4 on ORF7a (C27527T, P45L) was detected and might be caused by immune pressure. Our study suggested that certain variation differences existed between countries/regions with high and low vaccine coverage, but they were not likely caused by host immune pressure. We inferred that no extra immune pressures on SARS-CoV-2 were generated with high vaccine coverage, and we suggest promoting and strengthening the uptake of the COVID-19 vaccine worldwide, especially in less developed areas.

The mechanism and pharmacodynamics of 2-((1H-indol-3-yl)thio/sulfinyl)-N-pheny acetamide derivative as a novel inhibitor against human respiratory syncytial virus.

Human respiratory syncytial virus (RSV) is a leading cause of lower respiratory tract infection worldwide. Until now, there are no licenced vaccines or effective antiviral drugs against RSV infections. In our previous work, we found 2-((1H-indol-3-yl)thio/sulfinyl)-N-pheny acetamide derivatives (4-49 C and 1-HB-63) being a novel inhibitor against RSV in vitro. Here, we explored the underlying mechanism of 2-((1H-indol-3-yl)thio/sulfinyl)-N-pheny acetamide derivatives to inhibit RSV replication in vitro and disclosed that 4-49 C worked as the inhibitor of membrane fusion and 1-HB-63 functioned at the stage of RSV genome replication/transcription. Yet, both of them could not inhibit RSV infection of BALB/c mice by using RSV-Luc, in vivo imaging and RT-qPCR analyses, for which it may be due to the fast metabolism in vivo. Our work suggests that further structural modification and optimisation of 2-((1H-indol-3-yl) thio/sulfinyl)-N-pheny acetamide derivative are needed to obtain drug candidates with effective anti-RSV activities in vivo.

What is the Potential Cause for the Predominance of GII.2[P16] Norovirus in Acute Gastroenteritis Outbreaks in China?

GII.2[P16] noroviruses (NoV) reemerged and rapidly became the main epidemic strain in acute gastroenteritis (AGE) outbreaks in Asian countries since 2016. The current GII.2 [P16] NoV showed the same antigenicity to the ones before 2016, but several unique amino acid substitutions existed in the RNA dependent RNA polymerase (RdRp) and other non-structural proteins, and the viral load of the current GII.2[P16] NoV was higher than those of other genotypes, it was estimated that the viral replication ability may have improved. However, other genotypes, such as GII.1 and GII.3, also had recombination with the novel RdRp, were not prevalent in AGE-outbreaks; thus, it was inferred that the capsid proteins also played an important role in the enhanced replication process. The viral infection could also be affected by other factors, such as the population genetic background, the climate and environment, and people's lifestyles. Continued surveillance on genetic diversity and evolutionary pattern for the GII.2[P16] NoV is necessary.

Evolutionary analyses of emerging GII.2[P16] and GII.4 Sydney [P16] noroviruses.

GII.2[P16] and GII.4 Sydney [P16] are currently the two predominant norovirus genotypes. This study sought to clarify their evolutionary patterns by analyzing the major capsid VP1 and RNA-dependent RNA polymerase (RdRp) genes. Sequence diversities were analyzed at both nucleotide and amino acid levels. Selective pressures were evaluated with the Hyphy package in different models. Phylogenetic trees were constructed by the maximum likelihood method from full VP1 sequences, and evolutionary rates were estimated by the Bayesian Markov Chain Monte Carlo approach. The results showed that (1) several groups of tightly linked mutations between the RdRp and VP1 genes were detected in the GII.2[P16] and GII.4[P16] noroviruses, and most of these mutations were synonymous, which may lead to a better viral fitness to the host; (2) although the pattern of having new GII.4 variants every 2-4 years has been broken, both the pre- and the post-2015 Sydney VP1 had comparable evolutionary rates to previously epidemic GII.4 variants, and half of the major antigenic sites on GII.4 Sydney had residue substitutions and several caused obvious changes in the carbohydrate-binding surface that may potentially alter the property of the virus; and (3) GII.4 Sydney variants during 2018-21 showed geographical specificity in East Asia, South Asia, and North America; the antigenic sites of GII.2 are strictly conserved, but the GII.2 VP1 chronologically evolved into nine different sublineages over time, with sublineage IX being the most prevalent one since 2018. This study suggested that both VP1 and RdRp of the GII.2[P16] and GII.4 Sydney [P16] noroviruses exhibited different evolutionary directions. GII.4[P16] is likely to generate potential novel epidemic variants by accumulating mutations in the P2 domain, similar to previously epidemic GII.4 variants, while GII.2[P16] has conserved predicted antigenicity and may evolve by changing the properties of nonstructural proteins, such as polymerase replicational fidelity and efficiency. This study expands the understanding of the evolutionary dynamics of GII.2[P16] and GII.4[P16] noroviruses and may predict the emergence of new variants.

Biological Significance of the Genomic Variation and Structural Dynamics of SARS-CoV-2 B.1.617.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants have been emerging and circulating globally since the start of the COVID-19 pandemic, of which B.1.617 lineage that was first reported in India at the end of 2020, soon became predominant. Tracing genomic variations and understanding their impact on the viral properties are the foundations for the vaccine and drug development and for the mitigation measures to be taken or lifted. In this study, 1,051 near-complete genomes and 1,559 spike (S) sequences belonging to the B.1.617 were analyzed. A genome-wide spread of single nucleotide polymorphisms (SNPs) was identified. Of the high frequency mutations identified, 61% (11/18) involved structural proteins, despite two third of the viral genome encoding nonstructural proteins. There were 22 positive selection sites, mostly distributed across the S protein, of which 16 were led by non-C to U transition and should be of a special attention. Haplotype network revealed that a large number of daughter haplotypes were continually derived throughout the pandemic, of which H177, H181 H219 and H286 from the ancestor haplotype H176 of B.1.617.2 were widely prevalent. Besides the well known substitutions of L452R, P681R and deletions of E156 and F157, as well as the potential biological significance, structural analysis in this study still indicated that new amino acid changes in B.1.617, such as E484Q and N501Y, had reshaped the viral bonding network, and increasingly sequenced N501Y mutant with a potential enhanced binding ability was detected in many other countries in the follow-up monitoring. Although we can't conclude the properties of all the mutants including N501Y thoroughly, it merits focusing on their spread epidemically and biologically.

Genetic diversity and molecular evolution of human respiratory syncytial virus A and B.

Human respiratory syncytial viruses (RSVs) are classified into two major groups (A and B) based on antigenic differences in the G glycoprotein. To investigate circulating characteristics and phylodynamic history of RSV, we analyzed the genetic variability and evolutionary pattern of RSVs from 1977 to 2019 in this study. The results revealed that there was no recombination event of intergroup. Single nucleotide polymorphisms (SNPs) were observed through the genome with the highest occurrence rate in the G gene. Five and six sites in G protein of RSV-A and RSV-B, respectively, were further identified with a strong positive selection. The mean evolutionary rates for RSV-A and -B were estimated to be 1.48 × 10-3 and 1.92 × 10-3 nucleotide substitutions/site/year, respectively. The Bayesian skyline plot showed a constant population size of RSV-A and a sharp expansion of population size of RSV-B since 2005, and an obvious decrease 5 years later, then became stable again. The total population size of RSVs showed a similar tendency to that of RSV-B. Time-scaled phylogeny suggested a temporal specificity of the RSV-genotypes. Monitoring nucleotide changes and analyzing evolution pattern for RSVs could give valuable insights for vaccine and therapy strategies against RSV infection.

Dimethylglyoxime Clathrate as Ligand Derived Nitrogen-Doped Carbon-Supported Nano-Metal Particles as Catalysts for Oxygen Reduction Reaction.

Nitrogen-doped carbon-supported metal nano-particles show great promise as high-performance catalysts for novel energies, organic synthesis, environmental protection, and other fields. The synergistic effect between nitrogen-doped carbon and metal nano-particles enhances the catalytic properties. Thus, how to effectively combine nitrogen-doped carbon with metal nano-particles is a crucial factor for the synthesis of novel catalysts. In this paper, we report on a facile method to prepare nitrogen-doped carbon-supported metal nano-particles by using dimethylgly-oxime as ligand. The nano-particles of Pd, Ni, Cu, and Fe were successfully prepared by the pyrolysis of the corresponding clathrate of ions and dimethylglyoxime. The ligand of dimethylglyoxime is adopted as the source for the nitrogen-doped carbon. The nano-structure of the prepared Pd, Ni, Cu, and Fe particles are confirmed by X-ray diffraction, scanning electron microscopy, and trans-mission electron microscopy tests. The catalytic performances of the obtained metal nano-particles for oxygen reduction reaction (ORR) are investigated by cyclic voltammetry, Tafel, linear sweeping voltammetry, rotating disc electrode, rotating ring disc electrode, and other technologies. Results show that the nitrogen-doped carbon-supported metal nano-particles can be highly efficient catalysts for ORR. The results of the paper exhibit a facile methodology to prepare nitrogen-doped carbon-supported metal nano-particles.

Evaluation of the Safety and Immune Efficacy of Recombinant Human Respiratory Syncytial Virus Strain Long Live Attenuated Vaccine Candidates.

Human respiratory syncytial virus (RSV) infection is the leading cause of lower respiratory tract illness (LRTI), and no vaccine against LRTI has proven to be safe and effective in infants. Our study assessed attenuated recombinant RSVs as vaccine candidates to prevent RSV infection in mice. The constructed recombinant plasmids harbored (5' to 3') a T7 promoter, hammerhead ribozyme, RSV Long strain antigenomic cDNA with cold-passaged (cp) mutations or cp combined with temperature-sensitive attenuated mutations from the A2 strain (A2cpts) or further combined with SH gene deletion (A2cptsΔSH), HDV ribozyme (δ), and a T7 terminator. These vectors were subsequently co-transfected with four helper plasmids encoding N, P, L, and M2-1 viral proteins into BHK/T7-9 cells, and the recovered viruses were then passaged in Vero cells. The rescued recombinant RSVs (rRSVs) were named rRSV-Long/A2cp, rRSV-Long/A2cpts, and rRSV-Long/A2cptsΔSH, respectively, and stably passaged in vitro, without reversion to wild type (wt) at sites containing introduced mutations or deletion. Although rRSV-Long/A2cpts and rRSV-Long/A2cptsΔSH displayed  temperature-sensitive (ts) phenotype in vitro and in vivo, all rRSVs were significantly attenuated in vivo. Furthermore, BALB/c mice immunized with rRSVs produced Th1-biased immune response, resisted wtRSV infection, and were free from enhanced respiratory disease. We showed that the combination of ΔSH with attenuation (att) mutations of cpts contributed to improving att phenotype, efficacy, and gene stability of rRSV. By successfully introducing att mutations and SH gene deletion into the RSV Long parent and producing three rRSV strains, we have laid an important foundation for the development of RSV live attenuated vaccines.

Complete genome of a novel recombinant human astrovirus and its quasispecies in patients following hematopoietic stem cell transplantation.

Human astroviruses (HAstVs) were first identified in 1975 and can be classified into three clades: classic HAstVs (HAstV 1-8), MLB (MLB1-3) and VA (VA1-5), with MLB and VA were newly identified. Recombination and a high mutation rate make HAstV as one of the rapidly evolving infectious agents. This study reported a novel identified recombinant human astrovirus (Y/1-CHN) and its long existence in two immunocompromised patients with diarrhea following allogeneic hematopoietic stem cell transplantation (allo-HSCT). The identified Yu/1-CHN genome contains 6801 base pairs encoding three open reading frames, with ORF1a best hit to the HAstV1 (Pune strain, 97 % nucleotide identity), while ORF1b and ORF2 best hit to HAstV-5 (DL30 strain, 99 % nucleotide identity). Possible recombination breakpoint was predicted to be located in the boundary of ORF1a and ORF1b. Different quasispecies were found in the host, and the dN/dS ratios of the S and P domains were determined to be 1.189 and 1.444, respectively, suggesting a positive selection existed. Fecal samples collected in different clinical phases from the two patients were all positive for Yu/1-CHN, suggesting a long existence of the virus in the host. It was indicated that immunocompromised patients may a reservoir for astrovirus, their excreta should be monitored even after discharge from hospital.

Intra-Host Evolution of Norovirus GII.4 in a Chronic Infected Patient With Hematopoietic Stem Cell Transplantation.

Human noroviruses (NVs) are the leading cause of acute gastroenteritis outbreaks worldwide. The majority of outbreaks are caused by genogroup II.4 (GII.4), with new variants emerging every 2 to 4 years. Immunocompromised patients are hypothesized to be important reservoirs where new NV variants emerge. Here, we examined intra-host NV variants and assessed immune-driven NV evolution in chronically infected immunocompromised hosts. Three NV GII.4-positive samples were collected from the same patient in different clinical phases following allogeneic hematopoietic stem cell transplantation, and had viral RNA concentrations of 2.46 × 106, 1.47 × 106, and 2.26 × 106 genome copies/mL. The non-synonymous (dN) and synonymous (dS) substitution ratio of the sequences in the partial P domain were >1, indicating strong positive selection in the patient. Both the number and the frequency of the single nucleotide variants increased over time in the patient. Also, the majority of capsid amino acid changes were located at blocking epitopes and histo-blood group antigen (HBGA)-binding sites, and 11 positive selection sites were found in the capsid region, of which 8 sites were presented in blocking epitopes or HBGA-binding sites. Homodimeric P-domain capsid models also suggested a structural change in the epitopes and HBGA-binding sites. The results suggested that novel variants of NV GII.4 with HBGA and antigenic site changes were produced in the immunocompromised patient. Further functional and epidemiological studies are needed to determine whether the new variants are a risk to public health.

Analysis of Continuous Mutation and Evolution on Circulating SARS-CoV-2.

Monitoring the mutation and evolution of the virus is important for tracing its ongoing transmission and facilitating effective vaccine development. A total of 342 complete genomic sequences of SARS-CoV-2 were analyzed in this study. Compared to the reference genome reported in December 2019, 465 mutations were found, among which, 347 occurred in only 1 sequence, while 26 occurred in more than 5 sequences. For these 26 further identified as SNPs, 14 were closely linked and were grouped into 5 profiles. Phylogenetic analysis revealed the sequences formed 2 major groups. Most of the sequences in late period (March and April) constituted the Cluster II, while the sequences before March in this study and the reported S/L and A/B/C types in previous studies were all in Cluster I. The distributions of some mutations were specific geographically or temporally, the potential effect of which on the transmission and pathogenicity of SARS-CoV-2 deserves further evaluation and monitoring. Two mutations were found in the receptor-binding domain (RBD) but outside the receptor-binding motif (RBM), indicating that mutations may only have marginal biological effects but merit further attention. The observed novel sequence divergence is of great significance to the study of the transmission, pathogenicity, and development of an effective vaccine for SARS-CoV-2.

Diversified Variants of Astrovirus MLB2 in Patients Following Hematopoietic Stem Cell Transplantation and the Evolutionary Rates and Patterns of the Virus.

We assessed the quasispecies heterogeneity of a human astrovirus MLB2 (HAstV-MLB2-YJMGK) in immunocompromised patients following hematopoietic stem cell transplantation and performed genetic and evolutionary analyses of HAstV isolates circulating worldwide. The result showed that the virus had diversified variants and a strong positive selection in the patient, indicating that such patients may be a reservoir for astrovirus. The time to the most recent common ancestor of MLB2 and classic HAstVs was around 1800 years, and it has a decline in effective population size of HAstVs in the late 100 years.

A novel astrovirus identified in wild rhesus monkey feces in China.

The discovery and analysis of pathogens carried by non-human primates are important for understanding zoonotic infections in humans. We identified a highly divergent astrovirus (AstV) from fecal matter from a rhesus monkey in China, which has been tentatively named "monkey-feces-associated AstV" (MkAstV). The full-length genome of MkAstV was determined to be 7377 nt in length. It exhibits the standard genomic AstV organization of three open reading frames (ORFs) and is most closely related to duck AstV (28%, 49%, and 35% amino acid sequence identity in ORF1a, ORF1b, and ORF2, respectively). Coincidentally, while this report was being prepared, an astrovirus sequence from Hainan black-spectacled toad became available in the GenBank database, showing 95%, 94% and 92% aa sequence identity in ORF1a, ORF1b and ORF2, respectively, to the corresponding ORFs of MkAstV. Phylogenetic analysis of ORF1a, ORF1b, and ORF2 indicated that MkAstV and the amphibian-related astroviruses formed an independent cluster in the genus Avastrovirus. The host of MkAstV remains unknown. Epidemiological and serological studies of this novel virus should be undertaken in primates, including humans.

A Hybrid Material Combined Copper Oxide with Graphene for an Oxygen Reduction Reaction in an Alkaline Medium.

In this work, an electrode material based on CuO nanoparticles (NPs)/graphene (G) is developed for ORR in alkaline medium. According to the characterization of scanning electron microscope and transmission electron microscope, CuO NPs are uniformly distributed on the wrinkled G sheets. The X-ray diffraction test reveals that the phase of CuO is monoclinic. The CuO/G hybrid electrode exhibits a positive onset potential (0.8 V), high cathodic current density (3.79 × 10-5 mA/cm²) and high electron transfer number (four-electron from O2 to H2O) for ORR in alkaline media. Compared with commercial Pt/C electrocatalyst, the CuO/G electrode also shows superior fuel durability. The high electrocatalytic activity and durability are attribute to the strong coupling between CuO NPs and G nanosheets.

Synthesis and Characterizations of Zinc Oxide on Reduced Graphene Oxide for High Performance Electrocatalytic Reduction of Oxygen.

Electrocatalysts for the oxygen reduction (ORR) reaction play an important role in renewable energy technologies, including fuel cells and metal-air batteries. However, development of cost effective catalyst with high activity remains a great challenge. In this feature article, a hybrid material combining ZnO nanoparticles (NPs) with reduced graphene oxide (rGO) is applied as an efficient oxygen reduction electrocatalyst. It is fabricated through a facile one-step hydrothermal method, in which the formation of ZnO NPs and the reduction of graphene oxide are accomplished simultaneously. Transmission electron microscopy and scanning electron microscopy profiles reveal the uniform distribution of ZnO NPs on rGO sheets. Cyclic voltammograms, rotating disk electrode and rotating ring disk electrode measurements demonstrate that the hierarchical ZnO/rGO hybrid nanomaterial exhibits excellent electrocatalytic activity for ORR in alkaline medium, due to the high cathodic current density (9.21 × 10-5 mA/cm²), positive onset potential (-0.22 V), low H2O2 yield (less than 3%), and high electron transfer numbers (4e from O2 to H2O). The proposed catalyst is also compared with commercial Pt/C catalyst, comparable catalytic performance and better stability are obtained. It is expected that the ZnO/rGO hybrid could be used as promising non-precious metal cathode in alkaline fuel cells.