A highly effective ferritin-based divalent nanoparticle vaccine shields Syrian hamsters against lethal Nipah virus.

The Nipah virus (NiV), a highly deadly bat-borne paramyxovirus, poses a substantial threat due to recurrent outbreaks in specific regions, causing severe respiratory and neurological diseases with high morbidity. Two distinct strains, NiV-Malaysia (NiV-M) and NiV-Bangladesh (NiV-B), contribute to outbreaks in different geographical areas. Currently, there are no commercially licensed vaccines or drugs available for prevention or treatment. In response to this urgent need for protection against NiV and related henipaviruses infections, we developed a novel homotypic virus-like nanoparticle (VLP) vaccine co-displaying NiV attachment glycoproteins (G) from both strains, utilizing the self-assembling properties of ferritin protein. In comparison to the NiV G subunit vaccine, our nanoparticle vaccine elicited significantly higher levels of neutralizing antibodies and provided complete protection against a lethal challenge with NiV infection in Syrian hamsters. Remarkably, the nanoparticle vaccine stimulated the production of antibodies that exhibited superior cross-reactivity to homologous or heterologous henipavirus. These findings underscore the potential utility of ferritin-based nanoparticle vaccines in providing both broad-spectrum and long-term protection against NiV and emerging zoonotic henipaviruses challenges.

Novel antiviral discoveries for Japanese encephalitis virus infections through reporter virus-based high-throughput screening.

Japanese encephalitis (JE) caused by JE virus (JEV), remains a global public health concern. Currently, there is no specific antiviral drug approved for the treatment of JE. While vaccines are available for prevention, they may not cover all at-risk populations. This underscores the urgent need for prophylaxis and potent anti-JEV drugs. In this context, a high-content JEV reporter system expressing Nanoluciferase (Nluc) was developed and utilized for a high-throughput screening (HTS) of a commercial antiviral library to identify potential JEV drug candidates. Remarkably, this screening process led to the discovery of five drugs with outstanding antiviral activity. Further mechanism of action analysis revealed that cepharanthine, an old clinically approved drug, directly inhibited virus replication by blocking GTP binding to the JEV RNA-dependent RNA polymerase. Additionally, treatment with cepharanthine in mice models alleviated JEV infection. These findings warrant further investigation into the potential anti-JEV activity of cepharanthine as a new therapeutic approach for the treatment of JEV infection. The HTS method employed here proves to be an accurate and convenient approach that facilitates the rapid development of antiviral drugs.

Vaccines based on the fusion protein consensus sequence protect Syrian hamsters from Nipah virus infection.

Nipah virus (NiV), a bat-borne paramyxovirus, results in neurological and respiratory diseases with high mortality in humans and animals. Developing vaccines is crucial for fighting these diseases. Previously, only a few studies focused on the fusion (F) protein alone as the immunogen. Numerous NiV strains have been identified, including 2 representative strains from Malaysia (NiV-M) and Bangladesh (NiV-B), which differ significantly from each other. In this study, an F protein sequence with the potential to prevent different NiV strain infections was designed by bioinformatics analysis after an in-depth study of NiV sequences in GenBank. Then, a chimpanzee adenoviral vector vaccine and a DNA vaccine were developed. High levels of immune responses were detected after AdC68-F, pVAX1-F, and a prime-boost strategy (pVAX1-F/AdC68-F) in mice. After high titers of humoral responses were induced, the hamsters were challenged by the lethal NiV-M and NiV-B strains separately. The vaccinated hamsters did not show any clinical signs and survived 21 days after infection with either strain of NiV, and no virus was detected in different tissues. These results indicate that the vaccines provided complete protection against representative strains of NiV infection and have the potential to be developed as a broad-spectrum vaccine for human use.

Both chimpanzee adenovirus-vectored and DNA vaccines induced long-term immunity against Nipah virus infection.

Nipah virus (NiV) is a highly lethal zoonotic paramyxovirus that poses a severe threat to humans due to its high morbidity and the lack of viable countermeasures. Vaccines are the most crucial defense against NiV infections. Here, a recombinant chimpanzee adenovirus-based vaccine (AdC68-G) and a DNA vaccine (DNA-G) were developed by expressing the codon-optimized full-length glycoprotein (G) of NiV. Strong and sustained neutralizing antibody production, accompanied by an effective T-cell response, was induced in BALB/c mice by intranasal or intramuscular administration of one or two doses of AdC68-G, as well as by priming with DNA-G and boosting with intramuscularly administered AdC68-G. Importantly, the neutralizing antibody titers were maintained for up to 68 weeks in the mice that received intramuscularly administered AdC68-G and the prime DNA-G/boost AdC68-G regimen, without a significant decline. Additionally, Syrian golden hamsters immunized with AdC68-G and DNA-G via homologous or heterologous prime/boost immunization were completely protected against a lethal NiV virus challenge, without any apparent weight loss, clinical signs, or pathological tissue damage. There was a significant reduction in but not a complete absence of the viral load and number of infectious particles in the lungs and spleen tissue following NiV challenge. These findings suggest that the AdC68-G and DNA-G vaccines against NiV infection are promising candidates for further development.

ASPP2 binds to hepatitis C virus NS5A protein via an SH3 domain/PxxP motif-mediated interaction and potentiates infection.

Hepatitis C virus (HCV) infects millions of people worldwide and is a leading cause of liver disease. Despite recent advances in antiviral therapies, viral resistance can limit drug efficacy and understanding the mechanisms that confer viral escape is important. We employ an unbiased interactome analysis to discover host binding partners of the HCV non-structural protein 5A (NS5A), a key player in viral replication and assembly. We identify ASPP2, apoptosis-stimulating protein of p53, as a new host co-factor that binds NS5A via its SH3 domain. Importantly, silencing ASPP2 reduces viral replication and spread. Our study uncovers a previously unknown role for ASPP2 to potentiate HCV RNA replication.

Effect of standardized nursing cooperation on intravenous thrombolysis with recombinant tissue plasminogen activator in acute ischemic stroke.

OBJECTIVE: This study explored the impact of standardized nursing cooperation on intravenous thrombolysis with recombinant tissue plasminogen activator (rt-PA) in acute ischemic stroke (AIS). METHODS: From June 2019 to June 2020, a total of 235 AIS patients that received rt-PA intravenous thrombolysis were enrolled as the research subjects. Among them, there were 101 patients who were admitted between June 2019 and December 2019 and were placed into control-group and received traditional routine nursing collaboration procedures; and the remaining 134 subjects admitted between January 2020 to June 2020 were classified into the observation-group and received standardized care collaboration procedures. The time spent (from admission to CT examination, from completion of CT to medication and from admission to medication), the thrombolysis within 1 h, 1-2 h, 2-3 h and 3-4.5 h, the degree of damage of neurological function before and after nursing intervention, the occurrence of complications and satisfaction with nursing care were compared between the two groups. RESULTS: The time spent in each procedure of thrombolytic therapy in the observation group was remarkably less than that in control group (P<0.05). The distribution of thrombolysis in the observation group was superior to that in control group (P<0.05). NIHSS score of subjects in observation group after intervention was obviously lower than that in the control group, with statistically significant difference [(3.34±0.87) points, (4.82±0.93) points, t=12.5318, P=0.0000]. The incidence of complications in the observation group was 5.97%, and that in the control group was 24.75%, with a statistically significant difference (X2 =16.8317, P=0.0000). The nursing satisfaction of the observation group was 91.04%, which was significantly higher than 73.27% in the control group, and the difference was statistically significant (X2 =13.1496, P=0.0003). CONCLUSION: The standardized nursing cooperation for AIS patients with rt-PA intravenous thrombolysis is beneficial for effectively reducing the treatment delay and the incidence of complications, and improving the neurological function and satisfaction of nursing care, and as such it which is worthy of clinical promotion.

Long-range assembly of sequences helps to unravel the genome structure and small variation of the wheat-Haynaldia villosa translocated chromosome 6VS.6AL.

Genomics studies in wild species of wheat have been limited due to the lack of references; however, new technologies and bioinformatics tools have much potential to promote genomic research. The wheat-Haynaldia villosa translocation line T6VS·6AL has been widely used as a backbone parent of wheat breeding in China. Therefore, revealing the genome structure of translocation chromosome 6VS·6AL will clarify how this chromosome formed and will help to determine how it affects agronomic traits. In this study, chromosome flow sorting, NGS sequencing and Chicago long-range linkage assembly were innovatively used to produce the assembled sequences of 6VS·6AL, and gene prediction and genome structure characterization at the molecular level were effectively performed. The analysis discovered that the short arm of 6VS·6AL was actually composed of a large distal segment of 6VS, a small proximal segment of 6AS and the centromere of 6A, while the collinear region in 6VS corresponding to 230-260 Mb of 6AS-Ta was deleted when the recombination between 6VS and 6AS occurred. In addition to the molecular mechanism of the increased grain weight and enhanced spike length produced by the translocation chromosome, it may be correlated with missing GW2-V and an evolved NRT-V cluster. Moreover, a fine physical bin map of 6VS was constructed by the high-throughput developed 6VS-specific InDel markers and a series of newly identified small fragment translocation lines involving 6VS. This study will provide essential information for mining of new alien genes carried by the 6VS·6AL translocation chromosome.

Roles of Symbiotic Microorganisms in Arboviral Infection of Arthropod Vectors.

Arthropod vectors serve as native reservoirs and transmitters of hundreds of arboviruses. In arthropod vectors, symbiotic microorganisms residing in the gut lumen and/or hemocoelic tissues maintain complicated relationships with their host and influence multiple aspects of vector physiology. Recently, accumulating evidence has established an important role for symbiotic microorganisms in vector-virus interactions which could potentially be used to control viral transmission. Herein, we review recent progress on symbiotic microbe-arbovirus interactions and summarize the molecular mechanisms by which commensal microbes act on hosts and arboviruses. Understanding the sophisticated interactions among arthropod vectors, microbiota, and arboviruses may offer new strategies for the prevention of arboviral diseases in the future.

Phenotypic analysis of mutations at residue 146 provides insights into the relationship between NS5A hyperphosphorylation and hepatitis C virus genome replication.

The hepatitis C virus genotype 2a isolate, JFH-1, exhibits much more efficient genome replication than other isolates. Although basic replication mechanisms must be conserved, this raises the question of whether the regulation of replication might exhibit isolate- and/or genotype-specific characteristics. Exemplifying this, the phenotype of NS5A hyperphosphorylation is genotype-dependent; in genotype 1b a loss of hyperphosphorylation correlates with an enhancement of replication. In contrast, the replication of JFH-1 is not regulated by hyperphosphorylation. We previously identified a novel phosphorylation site in JFH-1 NS5A: S146. A phosphomimetic substitution (S146D) had no effect on replication but correlated with a loss of hyperphosphorylation. In genotype 1b, residue 146 is alanine and we therefore investigated whether the substitution of A146 with a phosphorylatable (S), or phosphomimetic, residue would recapitulate the JFH-1 phenotype, decoupling hyperphosphorylation from replication. This was not the case, as A146D exhibited both a loss of hyperphosphorylation and a reduction in replication, accompanied by a perinuclear restriction of replication complexes, reductions in lipid droplet and PI4P lipid accumulation, and a disruption of NS5A dimerization. In contrast, the S232I culture-adaptive mutation in the low-complexity sequence I (LCSI) also exhibited a loss of hyperphosphorylation, but was associated with an increase in replication. Taken together, these data imply that hyperphosphorylation does not directly regulate replication. In contrast, the loss of hyperphosphorylation is a consequence of perturbing genome replication and NS5A function. Furthermore, we show that mutations in either domain I or LCSI of NS5A can disrupt hyperphosphorylation, demonstrating that multiple parameters influence the phosphorylation status of NS5A.

A role for domain I of the hepatitis C virus NS5A protein in virus assembly.

The NS5A protein of hepatitis C virus (HCV) plays roles in both virus genome replication and assembly. NS5A comprises three domains, of these domain I is believed to be involved exclusively in genome replication. In contrast, domains II and III are required for the production of infectious virus particles and are largely dispensable for genome replication. Domain I is highly conserved between HCV and related hepaciviruses, and is highly structured, exhibiting different dimeric conformations. To investigate the functions of domain I in more detail, we conducted a mutagenic study of 12 absolutely conserved and surface-exposed residues within the context of a JFH-1-derived sub-genomic replicon and infectious virus. Whilst most of these abrogated genome replication, three mutants (P35A, V67A and P145A) retained the ability to replicate but showed defects in virus assembly. P35A exhibited a modest reduction in infectivity, however V67A and P145A produced no infectious virus. Using a combination of density gradient fractionation, biochemical analysis and high resolution confocal microscopy we demonstrate that V67A and P145A disrupted the localisation of NS5A to lipid droplets. In addition, the localisation and size of lipid droplets in cells infected with these two mutants were perturbed compared to wildtype HCV. Biophysical analysis revealed that V67A and P145A abrogated the ability of purified domain I to dimerize and resulted in an increased affinity of binding to HCV 3'UTR RNA. Taken together, we propose that domain I of NS5A plays multiple roles in assembly, binding nascent genomic RNA and transporting it to lipid droplets where it is transferred to Core. Domain I also contributes to a change in lipid droplet morphology, increasing their size. This study reveals novel functions of NS5A domain I in assembly of infectious HCV and provides new perspectives on the virus lifecycle.

First isolation of reticuloendotheliosis virus from mallards in China.

Reticuloendotheliosis virus (REV) causes an oncogenic, immunosuppressive and runting syndrome in many avian hosts worldwide. REV infection has never been reported in mallard ducks, however. To identify REV infection in mallards, we collected 40 mallard duck samples from Jilin Province of China. In this study, the REV strain, DBYR1102, was first isolated from a mallard in China and identified by PCR, indirect immunofluorescence assay and electron microscopy. The gp90 gene and complete LTR of DBYR1102 were amplified and sequenced. Phylogenetic analysis based on gp90 genes of REV indicated that the REV strain DBYR1102 is closely related to strain HLJR0901 from northeastern China, the prairie chicken isolate APC-566, and REV subtype III, represented by chick syncytial virus. This new strain is distantly related to two other subtypes of REV, 170A and SNV. Phylogenetic analysis based on the LTR yielded information similar to that obtained with the gp90 genes. The results of this study not only expand our epidemiological understanding of REV in the wild birds of China but also demonstrate the potential role of wild waterfowl in REV transmission.

Antigenic analysis of monoclonal antibodies against different epitopes of σB protein of avian reovirus.

BACKGROUND: Avian reovirus (ARV) causes arthritis, tenosynovitis, runting-stunting syndrome (RSS), malabsorption syndrome (MAS) and immunosuppression in chickens. σB is one of the major structural proteins of ARV, which is able to induce group-specific antibodies against the virus. METHODS AND RESULTS: The present study described the identification of two linear B-cell epitopes in ARV σB through expressing a set of partially overlapping and consecutive truncated peptides spanning σB screened with two monoclonal antibodies (mAbs) 1F4 and 1H3-1.The data indicated that (21)KTPACW(26) (epitope A) and (32)WDTVTFH(38) (epitope B) were minimal determinants of the linear B cell epitopes. Antibodies present in the serum of ARV-positive chickens recognized the minimal linear epitopes in Western blot analyses. By sequence alignment analysis, we determined that the epitopes A and B were not conserved among ARV, duck reovirus (DRV) and turkey reovirus (TRV) strains. Western blot assays, confirmed that epitopes A and B were ARV-specific epitopes, and they could not react with the corresponding peptides of DRV and TRV. CONCLUSIONS AND SIGNIFICANCE: We identified (21)KTPACW(26) and (32)WDTVTFH(38) as σB -specific epitopes recognized by mAbs 1F4 and 1H3-1, respectively. The results in this study may have potential applications in development of diagnostic techniques and epitope-based marker vaccines against ARV groups.

Identification of a linear B-Cell epitope on avian reovirus protein sigmaC.

SigmaC (σC) protein, which mediates virus attachment to target cells, is the most variable proteins of avian reovirus (ARV). It is responsible for inducing protective antibody immune responses in animals. To understand the antigenic determinants of σC protein, a set of partially overlapping and consecutive peptides spanning σC were expressed and then screened with the monoclonal antibody (mAb) 2B5 directed against σC. The mAb 2B5 recognized peptides with the σC motif (45)ELLHRSISDISTTV(58). Further identification of the displayed B-cell epitope was conducted with a set of truncated peptides expressed as GST fusion proteins. The Western blot and ELISA results indicated that (45)ELLHRSISDI(54) was the minimal determinant of the linear B-cell epitope. Using sequences analysis, we found that this epitope was not a common motif shared among the other members of the ARV and DRV groups. Furthermore, cross reactivity analysis showed that the associated coding motif of other ARV and DRV groups was not recognized by 2B5. These data suggested that (45)ELLHRSISDI(54) was a type-specific linear B-cell epitope of avian reovirus. The results in this study may have potential applications in the development of diagnostic techniques and epitope-based marker vaccines against ARV, which is prevalent in China.

Molecular characterization and phylogenetic analysis of the reticuloendotheliosis virus isolated from wild birds in Northeast China.

To analyze the status of reticuloendotheliosis (RE) infection of wild birds in China, 585 samples from wild birds collected in Liaoning, Jilin and Heilongjiang provinces China were investigated and analyzed. The sampled birds represent 3 orders and more than 40 species. Virus isolation and PCR amplification showed that some of the wild birds were infected with REV, and 10 REV strains were isolated. The gp90 gene from each of the 10 REV strains was amplified, cloned, and sequenced. Sequence analysis indicated that the gp90 genes of the 10 REV strains isolated in this study were more similar at the nucleotide level with the northeast Chinese strains HLJR0901 and HLJR0801 and some REV strains found in the US and Taiwan than with the early Chinese REV isolate HA9901. Furthermore, phylogenetic analysis indicated that the gp90 genes of the 10 REV strains were more similar to the REV subtype III-representing strain (CSV) than to strains 170A (subtype I) or SNV (subtype II). This is the first study to investigate the status of wild birds infected with REV. The results of this paper will not only provide necessary information for further understanding the evolution of REV, but they also identify the potential role of wild birds in REV transmission and furthers our understanding of the ecology of REV in wild bird species.

A Disaster Response and Management Competency Mapping of Community Nurses in China.

BACKGROUND: It is widely accepted in many parts of the world thatcommunity nurses are of vital importance in various phases of disaster response and management. In China, however, it is not clear whether the Chinese community nurses are able to assume disaster-related duties due to the lack of a systematic assessment. METHODS: A pre-designed and well-tested questionnaire was employed to evaluate the competency in disaster response and management among 205 valid registered Chinese community nurses between September and October 2009. Statistical analyses were performed with SPSS Version 13.0 using one way ANOVA, Least Significant Difference (LSD) and multiple stepwise regression analysis. RESULTS: This group of Chinese community nurses scored at an intermediate level of competency (a score of 3.68 (SD 0.48) out of a perfect score of 5) in disaster response and management, suggesting that they have the basic ability to participate in disaster-related nursing. Four factors, namely, Experiences in Disaster Relief, Participation in Disaster Training, the Age and Duration in Job, were identified to be the predominant factors contributing significantly to the integrated competency in disaster response and management of an individual. CONCLUSION: Most of the Chinese community nurses have basic qualifications and competencies to undertake the responsibilities of disaster response and management. However, more targeted disaster training including virtual-reality based drills should be provided in order to improve their competency.

Development and application of real-time PCR for detection of subgroup J avian leukosis virus.

Subgroup J avian leukosis virus (ALV-J) is an avian retrovirus that causes severe economic losses in the poultry industry. The early identification and removal of virus-shedding birds are important to reduce the spread of congenital and contact infections. In this study, a TaqMan-based real-time PCR method for the rapid detection and quantification of ALV-J with proviral DNA was developed. This method exhibited a high specificity for ALV-J. Moreover, the detection limit was as low as 10 viral DNA copies. The coefficients of variation (CVs) of both interassay and intra-assay reproducibility were less than 1%. The growth curves of ALV-J in DF-1 cells were measured by real-time PCR, yielding a trend line similar to those determined by 50% tissue culture infective dose (TCID(50)) and p27 antigen detection. Tissue samples suspected of ALV infection were evaluated using real-time PCR, virus isolation, and routine PCR, and the positivity rates were 60.1%, 41.6% and 44.5%, respectively. Our data indicated that the real-time PCR method provides a sensitive, specific, and reproducible diagnostic tool for the identification and quantification of ALV-J for clinical diagnosis and in laboratory research.

Heterojunction topology versus fill factor correlations in novel hybrid small-molecular/polymeric solar cells.

The authors present organic photovoltaic (OPV) devices comprising a small molecule electron acceptor based on 2-vinyl-4,5-dicyanoimidazole (Vinazene) and a soluble poly(p-phenylenevinylene) derivative as the electron donor. A strong dependence of the fill factor (FF) and the external quantum efficiency [incident photons converted to electrons (IPCE)] on the heterojunction topology is observed. As-prepared blends provided relatively low FF and IPCE values of 26% and 4.5%, respectively, which are attributed to significant recombination of geminate pairs and free carriers in a highly intermixed blend morphology. Going to an all-solution processed bilayer device, the FF and IPCE dramatically increased to 43% and 27%, respectively. The FF increases further to 57% in devices comprising thermally deposited Vinazene layers where there is virtually no interpenetration at the donor/acceptor interface. This very high FF is comparable to values reported for OPV using fullerenes as the electron acceptor. Furthermore, the rather low electron affinity of Vinazene compound near 3.5 eV enabled a technologically important open circuit voltage (V(oc)) of 1.0 V.

[Isolation and analysis of the drug resistance of the flavobacterium and its production of beta-lactamases].

OBJECTIVE: To investigate the drug resistance of flavobacterium and its ability to produce BLA (beta-lactamases) and ESBLs (Extended-spectrum beta-lactamases). METHODS: The production of BLA and ESBLs from 6 clinical isolated flavobacterium strains was determined by nitrocefin disc test and double-disc synergy method, respectively. The antibiotic susceptibilities of the strains were determined by Kirby-Bauer disc diffusion test and the agar dilution method and the MIC was assessed. RESULTS: All the six flavobacteria were BLA-producing strains and more than 80% of them were ESBLs-producing, and they were highly resistant to beta-lactamase antibiotics (MIC 32 - 256 mg/L), but susceptible to fluoroquinolones and cephalosporin with beta-lactamase inhibitors (MIC 0.125 - 8 mg/L). CONCLUSION: Most of the flavobacteria in nosocomial infections were beta-lactamase-producing and were highly resistant to beta-lactamase antibiotics. Fluoroquinolones and beta-lactamase antibiotics with lactamase inhibitors should be the first choice for the management of infection caused by flavobacterium.