Increased Cleavage of Japanese Encephalitis Virus prM Protein Promotes Viral Replication but Attenuates Virulence.

In flavivirus, the furin-mediated cleavage of prM is mandatory to produce infectious particles, and the immature particles containing uncleaved prM cannot undergo membrane fusion and release to the extracellular environment. However, the detailed relationship between viral replication or pathogenicity and furin in Japanese encephalitis virus (JEV) hasn't been clarified. Here, JEV with the mutations in furin cleavage sites and its nearby were constructed. Compared with WT virus, the mutant virus showed enhanced cleavage efficiency of prM protein and increased replication ability. Furthermore, we found that the mutations mainly promote genomic replication and assembly of JEV. However, the mutant formed smaller plaques than WT virus in plaque forming assay, indicating the lower cytopathogenicity of mutant virus. To assess the virulence of JEV mutant, an in vivo assay was performed using a mouse model. A higher survival rate and attenuated neuroinflammation were observed in JEV mutant-infected mice than those of WT-infected mice, suggesting the cleavage of prM by furin was closely related to viral virulence. These findings will provide new understanding on JEV pathogenesis and contribute to the development of novel JEV vaccines. IMPORTANCE Japanese encephalitis virus (JEV) is the leading cause of viral encephalitis epidemics in Southeast Asia, affecting mostly children, with high morbidity and mortality. During the viral maturation process, prM is cleaved into M by the cellular endoprotease furin in the acidic secretory system. After cleavage of the prM protein, mature virions are exocytosed. Here, the mutant in furin cleavage sites and its nearby was constructed, and the results showed that the mutant virus with enhanced replication mainly occurred in the process of genomic replication and assembly. Meanwhile, the mutant showed an attenuated virulence than WT virus in vivo. Our study contributes to understanding the function of prM and M proteins and provides new clues for live vaccine designation for JEV.

Effects of biochar on soil microbial community and functional genes of a landfill cover three years after ecological restoration.

Landfills, as the most common approach to disposing of municipal solid waste worldwide, disturb native ecosystems and create a need for ecological restoration. The restoration of landfill cover with biochar has shown immediate potential to improve soil microbial functions within one year. However, such characteristics could change after a longer period of time. Here, soil properties, microbial communities, and microbial functional genes related to nutrient cycling were investigated three years after the biochar amendment of the topsoil of a subtropical landfill cover. The results showed that the levels of soil organic matter, water content, total carbon (C), total nitrogen (N) and total phosphorus (P) of biochar-amended soils were higher than those of control soil. Different soil microbial community structures were observed in the biochar-amended and control soils. Nine phyla, including Proteobacteria and Acidobacteria, but not Actinobacteria or Chloroflexi, were enriched in the biochar-amended soil. Although the impact of biochar on shaping microbial communities increased after a longer period of restoration, no differences were observed in soils that were amended using different biochar:soil ratios. The abundances of functional genes related to C and N cycling decreased, whereas those of genes related to P cycling were higher in soils that received biochar amendment. This finding suggests that compared with the control soil, biochar-amended soils were less active in processes involved in C and N cycling but enhanced in processes related to P cycling. This study can serve as a reference for future ecological restoration of degraded lands using biochar.

Synergistic Effect and Characterization of Graphene/Carbon Nanotubes/Polyvinyl Alcohol/Sodium Alginate Nanofibrous Membranes Formed Using Continuous Needleless Dynamic Linear Electrospinning.

In this study, a self-made continuous needleless dynamic linear electrospinning technique is employed to fabricate large-scale graphene (Gr)/carbon nanotubes (CNT)/polyvinyl alcohol (PVA)/sodium alginate (SA) nanofibrous membranes. The synergistic effect of Gr and CNT fillers in the PVA/SA membrane is explored in depth by changing the volume ratio (v/v) of Gr and CNT as 10:0, 8:2, 6:4, 4:6, 2:8, and 0:10. Microstructure, functional group, conductivity, and hydrophilicity of PVA/SA/Gr/CNT membranes was characterized. Results show that the linear electrode needleless electrospinning technique can be spun into 200-nm diameter fibers. The PVA/SA/Gr/CNT fibrous membrane has good hydrophilicity and thermal stability. A Gr/CN ratio of 6:4 possessed the optimal synergistic effect, which showed the lowest surface resistivity of 2.53 × 103 Ω/m2. This study will provide a reference for the large-scale preparation of nanofibrous membrane used as a artificial nerve conduit in the future.

Characterization and Microstructure of Linear Electrode-Electrospun Graphene-Filled Polyvinyl Alcohol Nanofiber Films.

With the aim of achieving controllable mass production of electrospun nanofiber films, this study proposes and investigates the feasibility of using a custom-made linear electrode- electrospun device to produce conductive graphene (GR)-filled polyvinyl alcohol (PVA) nanofibers. The film morphology and diameter of nanofibers are observed and measured to examine the effects of viscosity and conductivity of the PVA/GR mixtures. Likewise, the influence of the content of graphene on the hydrophilicity, electrical conductivity, electromagnetic interference shielding effectiveness (EMSE), and thermal stability of the PVA/GR nanofiber films is investigated. The test results show that the PVA/GR mixture has greater viscosity and electric conductivity than pure PVA solution and can be electrospun into PVA/GR nanofiber films that have good morphology and diameter distribution. The diameter of the nanofibers is 100 nm and the yield is 2.24 g/h, suggesting that the process qualifies for use in large-scale production. Increasing the content of graphene yields finer nanofibers, a smaller surface contact angle, and higher hydrophilicity of the nanofiber films. The presence of graphene is proven to improve the thermal stability and strengthens the EMSE by 20 dB at 150⁻1500 MHz. Mass production is proven to be feasible by the test results showing that PVA/GR nanofiber films can be used in the medical hygiene field.

Mass-Production and Characterizations of Polyvinyl Alcohol/Sodium Alginate/Graphene Porous Nanofiber Membranes Using Needleless Dynamic Linear Electrospinning.

The aim of this study was to investigate the feasibility of large-scale preparation of porous polyvinyl alcohol/sodium alginate/graphene (Gr) (Gr-AP) nanofiber membranes using a copper wire needleless dynamic linear electrode electrospinning machine. Furthermore, the effects of Gr concentrations (0, 0.0375, 0.075, 0.25, 0.5, and 0.75 wt.%) on the morphology, electrical, hydrophilicity and thermal properties were evaluated. Results indicate that the dynamic linear electrospun Gr-AP membranes have a high yield of 1.25 g/h and are composed of porous finer nanofibers with a diameter of 141 ± 31 nm. Gr improved the morphology, homogeneity, hydrophobicity and thermal stability of Gr-AP nanofiber membranes. The critical conductive threshold is 0.075 wt.% for Gr, which provides the nanofiber membranes with an even distribution of diameter, an optimal conductivity, good hydrophilicity, appropriate specific surface area and optimal thermal stability. Therefore, needleless dynamic linear electrospinning is beneficial to produce high quality Gr-AP porous nanofiber membranes, and the optimal parameters can be used in artificial nerve conduits and serve as a valuable reference for mass production of nanofiber membranes.