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Mother nature accomplishes efficient ammonia synthesis via cascade N 2 oxidation by lightning strikes followed with enzyme-catalyzed nitrogen oxyanion (NO x - , x = 2,3) reduction. The protein environment of enzymatic centers for NO x - -to-NH 4 + process greatly inspires the design of glutathione-capped (GSH) quantum dots (QDs) for ammonia synthesis under visible light (440 nm) in tandem with plasma-enabled N 2 oxidation. Mechanistic studies reveal that GSH induces positive shift of surface charge to strengthen the interaction between NO x - and QDs. Upon visible light irradiation of QDs, the balanced and rapid hole and electron transfer furnish GS·radicals for 2e - /2H + alcohol oxidation and H·for 8e - /10H + NO 3 - -to-NH 4 + reduction simultaneously. For the first time, mmol-scale ammonia synthesis is realized with apparent quantum yields of 5.45% ± 0.64%, and gram-scale synthesis of value-added acetophenone and NH 4 Cl proceeds with 1:4 stoichiometry and stability, demonstrating promising multielectron and multiproton ammonia synthesis efficiency and sustainability with nature-inspired artificial photocatalysts., (© 2024 Wiley‐VCH GmbH.)

As SARS-CoV-2 continues to evolve and COVID-19 cases rapidly increase among children and adults, there is an urgent need for a safe and effective vaccine that can elicit systemic and mucosal humoral immunity to limit the emergence of new variants. Using the Chinese Hu191 measles virus (MeV-hu191) vaccine strain as a backbone, we developed MeV chimeras stably expressing the prefusion forms of either membrane-anchored, full-length spike (rMeV-preFS), or its soluble secreted spike trimers with the help of the SP-D trimerization tag (rMeV-S+SPD) of SARS-CoV-2 Omicron BA.2. The two vaccine candidates were administrated in golden Syrian hamsters through the intranasal or subcutaneous routes to determine the optimal immunization route for challenge. The intranasal delivery of rMeV-S+SPD induced a more robust mucosal IgA antibody response than the subcutaneous route. The mucosal IgA antibody induced by rMeV-preFS through the intranasal routine was slightly higher than the subcutaneous route, but there was no significant difference. The rMeV-preFS vaccine stimulated higher mucosal IgA than the rMeV-S+SPD vaccine through intranasal or subcutaneous administration. In hamsters, intranasal administration of the rMeV-preFS vaccine elicited high levels of NAbs, protecting against the SARS-CoV-2 Omicron BA.2 variant challenge by reducing virus loads and diminishing pathological changes in vaccinated animals. Encouragingly, sera collected from the rMeV-preFS group consistently showed robust and significantly high neutralizing titers against the latest variant XBB.1.16. These data suggest that rMeV-preFS is a highly promising COVID-19 candidate vaccine that has great potential to be developed into bivalent vaccines (MeV/SARS-CoV-2).

Background: Large-scale vaccine production requires downstream processing that focuses on robustness, efficiency, and cost-effectiveness., Methods: To assess the robustness of the current vaccine production process, three batches of COVID-19 Omicron BA.1 strain hydrolytic concentrated solutions were selected. Four gel filtration chromatography media (Chromstar 6FF, Singarose FF, Bestarose 6B, and Focurose 6FF) and four ion exchange chromatography media (Maxtar Q, Q Singarose, Diamond Q, and Q Focurose) were used to evaluate their impact on vaccine purification. The quality of the vaccine was assessed by analyzing total protein content, antigen content, residual Vero cell DNA, residual Vero cell protein, and residual bovine serum albumin (BSA). Antigen recovery rate and specific activity were also calculated. Statistical analysis was conducted to evaluate process robustness and the purification effects of the chromatography media., Results: The statistical analysis revealed no significant differences in antigen recovery ( p = 0.10), Vero HCP residue ( p = 0.59), Vero DNA residue ( p = 0.28), and BSA residue ( p = 0.97) among the three batches of hydrolytic concentrated solutions processed according to the current method. However, a significant difference ( p < 0.001) was observed in antigen content., Conclusions: The study demonstrated the remarkable robustness of the current downstream process for producing WIBP-CorV vaccines. This process can adapt to different batches of hydrolytic concentrated solutions and various chromatography media. The research is crucial for the production of inactivated SARS-CoV-2 vaccines and provides a potential template for purifying other viruses.

The extensive use of carbamate pesticides has led to a range of environmental and health problems, such as surface and groundwater contamination, and endocrine disorders in organisms. In this study, we focused on examining the effects of toxic exposure to the carbamate pesticide methomyl on the hatching, morphology, immunity and developmental gene expression levels in zebrafish embryos. Four concentrations of methomyl (0, 2, 20, and 200 μg/L) were administered to zebrafish embryos for a period of 96 h. The study found that exposure to methomyl accelerated the hatching process of zebrafish embryos, with the strongest effect recorded at the concentration of 2 μg/L. Methomyl exposure also trigged significantly reductions in heart rate and caused abnormalities in larvae morphology, and it also stimulated the synthesis and release of several inflammatory factors such as IL-1β, IL-6, TNF-α and INF-α, lowered the IgM contents, ultimately enhancing inflammatory response and interfering with immune function. All of these showed the significant effects on exposure time, concentration and their interaction (Time × Concentration). Furthermore, the body length of zebrafish exposed to methomyl for 96 h was significantly shorter, particularly at higher concentrations (200 μg/L). Methomyl also affected the expression levels of genes associated with development (down-regulated igf1, bmp2b, vasa, dazl and piwi genes), demonstrating strong developmental toxicity and disruption of the endocrine system, with the most observed at the concentration of 200 μg/L and 96 h exposure to methomyl. The results of this study provide valuable reference information on the potential damage of methomyl concentrations in the environment on fish embryo development, while also supplementing present research on the immunotoxicity of methomyl., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest of this work., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Coxsackievirus A10 (CV-A10) has become one of the major pathogens of hand, foot and mouth disease (HFMD), and studies on the vaccine and animal model of CV-A10 are still far from complete. Our study used a mouse-adapted CV-A10 strain, which was lethal for 14-day-old mice, to develop an infected mouse model. Then this model was employed to establish an actively immunized-challenged mouse model to evaluate the efficacy of a formaldehyde-inactivated CV-A10 vaccine, which was prepared from a Vero cell-adapted strain. CV-A10 vaccine at a dose of 0.5 or 2.0 μg was inoculated intraperitoneally in neonatal Kunming mice on the third and ninth day. Then the mice were challenged on day 14. The survival rate of mice immunized with 0.5 or 2.0 μg vaccine were 90% and 100%, respectively, while all Alum-inoculated mice died. Compared to those in the two vaccinated groups, the Alum-inoculated mice showed severe pathological damage, strong viral protein expression and high viral loads. The antisera from vaccinated mice showed high level of neutralizing antibodies against CV-A10. Meanwhile, three potential T cell epitopes located at the carboxyl-terminal regions of the VP1 and VP3 were identified and exhibited CV-A10 serotype-specific. The humoral and cellular immunogenicity analysis showed that immunization with two doses of the vaccine elicited CV-A10 specific neutralizing antibody and T cell response in BALB/c mice. Collectively, these findings indicated that this actively immunized-challenged mouse model will be invaluable in future studies on CV-A10 pathogenesis and evaluation of vaccine candidates.

Rabies, caused by the rabies virus (RABV), is the most fatal zoonotic disease. It is a neglected tropical disease which remains a major public health problem, causing approximately 59,000 deaths worldwide annually. Despite the existence of effective vaccines, the high incidence of human rabies is mainly linked to tedious vaccine immunisation procedures and the overall high cost of post-exposure prophylaxis. Therefore, it is necessary to develop an effective vaccine that has a simple procedure and is affordable to prevent rabies infection in humans. RABV belongs to the genus Lyssavirus and family Rhabdoviridae. Previous phylogenetic analyses have identified seven major clades of RABV in China (China I-VII), confirmed by analysing nucleotide sequences from both the G and N proteins. This study evaluated the immunogenicity and protective capacity of SYS6008, an mRNA rabies vaccine expressing rabies virus glycoprotein, in mice and cynomolgus macaques. We demonstrated that SYS6008 induced sufficient levels of rabies neutralising antibody (RVNA) in mice. In addition, SYS6008 elicited strong and durable RVNA responses in vaccinated cynomolgus macaques. In the pre-exposure prophylaxis murine model, one or two injections of SYS6008 at 1/10 or 1/30 of dosage provided protection against a challenge with a 30-fold LD 50 of rabies virus (China I and II clades). We also demonstrated that in the post-exposure prophylaxis murine model, which was exposed to lethal rabies virus (China I-VII clades) before vaccination, one or two injections of SYS6008 at both 1/10 and 1/30 dosages provided better protection against rabies virus challenge than the immunization by five injections of commercial vaccines at the same dosage. In addition, we proved that SYS6008-induced RVNAs could neutralise RABV from the China I-VII clades. Finally, 1/10 of the dosage of SYS6008 was able to stimulate significant RABV-G specificity in the T cell response. Furthermore, we found that SYS6008 induced high cellular immunity, including RABV-G-specific T cell responses and memory B cells. Our results imply that the SYS6008 rabies vaccine, with a much simpler vaccination procedure, better immunogenicity, and enhanced protective capacity, could be a candidate vaccine for post-exposure prophylaxis of rabies infections., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Coxsackievirus A10 (CV-A10) is a prevailing causative agent of hand-foot-mouth disease, necessitating the isolation and adaptation of appropriate strains in cells allowed for human vaccine development. In this study, amino acid sequences of CV-A10 strains with different cell tropism on RD and Vero cells were compared. Various amino acids on the structural and non-structural proteins related to cell tropism were identified. The reverse genetic systems of several CV-A10 strains with RD + /Vero - and RD + /Vero + cell tropism were developed, and a set of CV-A10 recombinants were produced. The binding, entry, uncoating, and proliferation steps in the life cycle of these viruses were evaluated. P1 replacement of CV-A10 strains with different cell tropism revealed the pivotal role of the structural proteins in cell tropism. Further, seven amino acid substitutions in VP2 and VP1 were introduced to further investigate their roles played in cell tropism. These mutations cooperated in the growth of CV-A10 in Vero cells. Particularly, the valine to isoleucine mutation at the position VP1-236 (V1236I) was found to significantly restrict viral uncoating in Vero cells. Co-immunoprecipitation assays showed that the release of viral RNA from the KREMEN1 receptor-binding virions was restricted in r0195-V1236I compared with the parental strain r0195 (a RD + /Vero + strain). Overall, this study highlights the dominant effect of structural proteins in CV-A10 adaption in Vero cells and the importance of V1236 in viral uncoating, providing a foundation for the mechanism study of CV-A10 cell tropism, and facilitating the development of vaccine candidates.

Coxsackievirus A5 (CV-A5) is a re-emerging enterovirus that causes hand, foot, and mouth disease in children under five years of age. CV-A5-M14-611 is a mouse-adapted strain that can infect orally and lead to the death of 14-day-old mice. Here, recombinants based on CV-A5-M14-611 were constructed carrying three reporter genes in different lengths. Smaller fluorescent marker proteins, light, oxygen, voltage sensing (iLOV), and nano luciferase (Nluc) were proven to be able to express efficiently in vitro. However, the recombinant with the largest insertion of the red fluorescence protein gene (DsRed) was not rescued. The construction strategy of reporter viruses was to insert the foreign genes between the C-terminus of VP1 and the N-terminus of 2A genes and to add a 2A protease cleavage domain at both ends of the insertions. The iLOV-tagged or Nluc-tagged recombinants, CV-A5-iLOV or CV-A5-Nluc, exhibited a high capacity for viral replication, genetic stability in cells and pathogenicity in mice. They were used to establish a rapid, inexpensive and convenient neutralizing antibody assay and greatly facilitated virus neutralizing antibody titration. Living imaging was performed on mice with CV-A5-Nluc, which exhibited specific bioluminescence in virus-disseminated organs, while fluorescence induced by CV-A5-iLOV was weakly detected. The reporter-gene-tagged CV-A5 can be used to study the infection and mechanisms of CV-A5 pathogenicity in a mouse model. They can also be used to establish rapid and sensitive assays for detecting neutralizing antibodies.

Previous studies demonstrated Y chromosome haplogroup C2a-M48-SK1061 is the only founding paternal lineage of all Tungusic-speaking populations. To infer the differentiation history of these populations, we studied more sequences and constructed downstream structure of haplogroup C2a-M48-SK1061 with better resolution. In this study, we generated 100 new sequences and co-analyzed 140 sequences of C2a-M48-SK1061 to reconstruct a highly revised phylogenetic tree with age estimates. We also performed the analysis of the geographical distribution and spatial autocorrelation of sub-branches. Dozens of new sub-branches were discovered, many sub-branches were nearly unique for Ewenki, Evens, Oroqen, Xibe, Manchu, Daur, and Mongolian. The topology of these unique sub-branches is the key evidence for understanding the complex evolutionary relationship between different Tungusic-speaking populations. The revised phylogeny provided a clear pattern for the differentiation history of haplogroup C2a-M48-SK1061 in the past 2,000 years. This study showed that the divergence pattern of founder lineage is essential to understanding the differentiation history of populations., Competing Interests: Author K-JL was employed by the company Chengdu 23Mofang Biotechnology Co., Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Yu, Ao, Zhang, Liu, Wang, Meng, Li, Wei and Man.)

Outbreaks of hand, foot and mouth disease (HFMD) have occurred frequently in the Asian-Pacific region over the last two decades, caused mainly by the serotypes in Enterovirus A species. High-quality monoclonal antibodies (mAbs) are needed to improve the accuracy and efficiency of the diagnosis of enteroviruses associated HFMD. In this study, a mAb 1A11 was generated using full particles of CV-A5 as an immunogen. In indirect immunofluorescence and Western blotting assays, 1A11 bound to the viral proteins of CV-A2, CV-A4, CV-A5, CV-A6, CV-A10, CV-A16, and EV-A71 of the Enterovirus A and targeted VP3. It has no cross-reactivity to strains of Enterovirus B and C . By mapping with over-lapped and truncated peptides, a minimal and linear epitope 23 PILPGF 28 was identified, located at the N-terminus of the VP3. A BLAST sequence search of the epitope in the NCBI genus Enterovirus (taxid: 12059) protein database indicates that the epitope sequence is highly conserved among the Enterovirus A species, but not among the other enterovirus species, first reported by us. By mutagenesis analysis, critical residues for 1A11 binding were identified for most serotypes of Enterovirus A . It may be useful for the development of a cost-effective and pan- Enterovirus A antigen detection for surveillance, early diagnosis and differentiation of infections caused by the Enterovirus A species.

Hand, foot and mouth disease (HFMD) is caused by a variety of serotypes in species A of the Enterovirus genus, including recently re-emerged Coxsackievirus A2 (CV-A2), CV-A4 and CV-A5. For development of diagnostic reagents, for surveillance, and the development of multivalent vaccines against HFMD, the antigenicity of HFMD-associated enteroviruses warrants investigation. The purified virions of CV-A4 were inoculated into Balb/c mice and hybridomas were obtained secreting monoclonal antibodies (mAbs) directed against CV-A4 and cross-reacting with other closely related species A enteroviruses. The mAbs were characterized by ELISA, Western blotting and in vitro neutralizing assays. The majority of mAbs was non-neutralizing, with only 2% of the mAbs neutralizing CV-A4 specifically. Most of mAbs bound to linear VP1 epitopes of CV-A4. Interestingly, four types of mAbs were obtained which bound specifically to CV-A4 or were broadly to CV-A4/-A2, CV-A4/-A5 and CV-A4/-A2/-A5, respectively. Mapping with overlapping or single-amino-acid mutant peptides revealed that the four types of mAbs all bound to the first 15 amino acids at the N-terminus of the VP1. This region of picornaviruses is functionally important as it is involved in uncoating and releasing of viral RNA into the cytosol. The binding footprints of four type mAbs are composed of conserved and variable residues and are different from each other. The newly discovered broadly cross-reactive mAbs reflect the high homology of CV-A4/ CV-A2/CV-A5. The results also demonstrate that it is possible and beneficial to develop the diagnostic reagents to detect rapidly the main pathogens of enteroviruses associated with HFMD cause by CV-A4/CV-A2/CV-A5., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

The odyssey of photochemistry is accompanied by the journey to manipulate "electrons" and "protons" in time, in space, and in energy. Over the past decades, single-electron (1e - ) photochemical transformations have brought marvelous achievements. However, as each photon absorption typically generates only one exciton pair, it is exponentially challenging to accomplish multielectron and proton photochemical transformations. The multistep differences in thermodynamics and kinetics urgently require us to optimize light harvesting, expedite consecutive electron transfer, manipulate the interaction of catalysts with substrates, and coordinate proton transfer kinetics to furnish selective bond formations. Tandem catalysis enables orchestrating different photochemical events and catalytic transformations from subpicoseconds to seconds, which facilitates multielectron redox chemistries and brings consecutive, value-added reactivities. Joint efforts in molecular and material design, mechanistic understanding, and theoretical modeling will bring multielectron and proton synthetic opportunities for fuels, fertilizers, and chemicals with enhanced versatility, efficiency, selectivity, and scalability, thus taking better advantage of photons (i.e., sunlight) for our sustainable society.

Background: In recent years, Coxsackievirus A2 (CV-A2) has become one of the main serotypes of enterovirus species A associated with hand, foot and mouth disease (HFMD) in China. It has also caused HFMD epidemics in many countries all over the world. Currently, there are no effective, preventive vaccines against it., Methods: A CV-A2 strain was isolated in RD cells and then adapted to grow in Vero cells. This is in compliance with guidelines for cell substrates allowed for human vaccines by the Chinese regulatory authority. Groups of newborn Kunming mice were inoculated on day 3 and day 9 using two formulations of candidate vaccines, empty particles and full particles. They were then challenged on day 14 at a lethal dose with a mouse-adapted strain., Results: The mice in the control group all died within 14 days post-challenge whereas most of the mice in the candidate vaccine groups survived. It was found that the titers of neutralizing antibodies was dose-dependent in sera of immunized mice. The results also showed that the vaccine candidates stimulated a strong humoral immune response and protected the mice from disease and death. The virus loads in tissues or organs were significantly reduced and pathological changes were either weak or not observed in the immunized groups compared with those in Al(OH) 3 control group. Preliminary mapping of the nucleotide and amino acid residues potentially related to cell tropism of the vaccine strain and virulence of the challenge strain was performed., Conclusion: The results showed that the RD cell-isolated and Vero cell-adapted CV-A2 strain is a promising vaccine candidate. This active immunization-challenge mouse model mimics the vaccination and then exposure to wildtype viruses, compared with passive immunization-challenge model, and is invaluable for efficacy evaluation in studies on multivalent vaccines containing CV-A2 against HFMD., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

A bis(pyridyl)amine-bipyridine-iron(II) framework (Fe(BPAbipy)) of complexes 1-3 is reported to shed light on the multistep nature of CO 2 reduction. Herein, photocatalytic conversion of CO 2 to CO even at low CO 2 concentration (1 %), together with detailed mechanistic study and DFT calculations, reveal that 1 first undergoes two sequential one-electron transfer affording an intermediate with electron density on both Fe and ligand for CO 2 binding over proton. The following 2 H + -assisted Fe-CO formation is rate-determining for selective CO 2 -to-CO reduction. A pendant, proton-shuttling α-OH group (2) initiates PCET for predominant H 2 evolution, while an α-OMe group (3) cancels the selectivity control for either CO or H 2 . The near-unity selectivity of 1 and 2 enables self-sorting syngas production at flexible CO/H 2 ratios. The unprecedented results from one kind of molecular catalyst skeleton encourage insight into the beauty of advanced multi-electron and multi-proton transfer processes for robust CO 2 RR by photocatalysis., (© 2021 Wiley-VCH GmbH.)

Microorganisms are sensitive to changes in the external environment and are often used as indicators to monitor and reflect water quality. Using Illumina MiSeq sequencing, the characteristics of the microbial community in Shihou Lake water at different time points were analyzed and the key environmental factors affecting the bacterial community were identified. The microbial community diversity in Shihou Lake water was rich and showed significant differences over time. The main bacterial phyla were the Cyanobacteria , Proteobacteria , Actinobacteria , Verrucomicrobia , Bacteroidetes , Chloroflexi , Planctomycetes , Firmicutes , Chlorobi , WS6 and Saccharibacteria . The relative abundance of these major phyla in the sample accounted for 97.83%-99.07% of the total abundance; Cyanobacteria had the highest relative abundance, accounting for 13.07%-44.61% of the total, and the abundance of each dominant phylum was significantly different at different time points. The Shannon and Simpson indexes showed that the diversity of each month was as follows: August > October > July > September > November. The Chao1 and Ace indexes indicated that the order of richness was: November > October > July > August > September. Beta diversity analysis found significant differences in the samples from month to month. Environmental factors such as temperature, total nitrogen, chlorophyll- a , permanganate index, nitrite, pH and ammonia nitrogen had significant effects on microbial community structure., Competing Interests: Conflict of interestThere is no conflict of interest here in this article., (© Association of Microbiologists of India 2021.)

Coxsackievirus A6 (CV-A6) has been emerging as a major pathogen of hand, foot and mouth disease (HFMD). Study on the pathogenesis of CV-A6 infection and development of vaccines is hindered by a lack of appropriate animal models. Here, we report an actively immunized-challenged mouse model to evaluate the efficacy of a Vero-cell-based, inactivated CV-A6 vaccine candidate. The neonatal Kunming mice were inoculated with a purified, formaldehyde-inactivated CV-A6 vaccine on days 3 and 9, followed by challenging on day 14 with a naturally selected virulent strain at a lethal dose. Within 14 days postchallenge, all mice in the immunized groups survived, while 100% of the Alum-only inoculated mice died. Neutralizing antibodies (NtAbs) were detected in the serum of immunized suckling mice, and the NtAb levels correlated with the survival rate of the challenged mice. The virus loads in organs were reduced, and pathological changes and viral protein expression were weak in the immunized mice compared with those in Alum-only inoculated control mice. Elevated levels of interleukin-4, 6, interferon γ and tumour necrosis factor α were also observed in Alum-only control mice compared with immunized mice. Importantly, the virulent CV-A6 challenge strain was selected quickly and conveniently from a RD cell virus stock characterized with the natural multi-genotypes. The virulent determinants were mapped to V124M and I242 V at VP1. Together, our results indicated that this actively immunized mouse model is invaluable for future studies to develop multivalent vaccines containing the major component of CV-A6 against HFMD.

Objectives: Haplogroup C2a-M48 is the predominant paternal lineage of Tungusic-speaking populations, one of the largest population groups in Siberia. Up until now, the origins and dispersal of Tungusic-speaking populations have remained unclear. In this study, the demographic history of Tungusic-speaking populations was explored using the phylogenetic analysis of haplogroup C2a-M86, the major subbranch of C2a-M48., Materials and Methods: In total, 18 newly generated Y chromosome sequences from C2a-M48 males and 20 previously available Y-chromosome sequences from this haplogroup were analyzed. A highly revised phylogenetic tree of haplogroup C2a-M86 with age estimates was reconstructed. Frequencies of this lineage in the literature were collected and a comprehensive analysis of this lineage in 13 022 individuals from 245 populations in Eurasia was performed., Results: The distribution map of C2a-M48 indicated the most probable area of origin and diffusion route of this paternal lineage in North Eurasia. Most C2a-M86 samples from Tungusic-speaking populations belonged to the sublineage C2a-F5484, which emerged about 3300 years ago. We identified six unique sublineages corresponding to the Manchu, Evenks, Evens, Oroqen, and Daurpopulations; these sublineages diverged gradually over the past 1900 years. Notably, we observed a clear north-south dichotomous structure for sublineages derived from C2a-F5484, consistent with the internal north-south divergence of Tungusic languages and ethnic groups., Conclusions: We identified the important founding paternal haplogroup, C2a-F5484, for Tungusic-speaking populations as well as numerous unique subgroups of this haplogroup. We propose that the timeframe for the divergence of C2a-F5484 corresponds with the early differentiation of ancestral Tungusic-speaking populations., (© 2020 Wiley Periodicals LLC.)

Coxsackievirus A5 (CV-A5) has recently emerged as a main hand, foot, and mouth disease (HFMD) pathogen. Following a large-scale vaccination campaign against enterovirus 71 (EV-71) in China, the number of HFMD-associated cases with EV-71 was reduced, especially severe and fatal cases. However, the total number of HFMD cases remains high, as HFMD is also caused by other enterovirus serotypes. A multivalent HFMD vaccine containing 4 or 6 antigens of enterovirus serotypes is urgently needed. A formaldehyde-inactivated CV-A5 vaccine derived from Vero cells was used to inoculate newborn Kunming mice on days 3 and 10. The mice were challenged on day 14 with a mouse-adapted CV-A5 strain at a dose that was lethal for 14-day-old suckling mice. Within 14 days postchallenge, groups of mice immunized with three formulations, empty particles (EPs), full particles (FPs), and a mixture of the EP and FP vaccine candidates, all survived, while 100% of the mock-immunized mice died. Neutralizing antibodies (NtAbs) were detected in the sera of immunized mice, and the NtAb levels were correlated with the survival rate of the challenged mice. The virus loads in organs were reduced, and pathological changes and viral protein expression were weak or not observed in the immunized mice compared with those in alum-inoculated control mice. Another interesting finding was the identification of CV-A5 dense particles (DPs), facilitating morphogenesis study. These results demonstrated that the Vero cell-adapted CV-A5 strain is a promising vaccine candidate and could be used as a multivalent HFMD vaccine component in the future. IMPORTANCE The vaccine candidate strain CV-A5 was produced with a high infectivity titer and a high viral particle yield. Three particle forms, empty particles (EPs), full particles (FPs), and dense particles (DPs), were obtained and characterized after purification. The immunogenicities of EP, FP, and the EP and FP mixture were evaluated in mice. Mouse-adapted CV-A5 was generated as a challenge strain to infect 14-day-old mice. An active immunization challenge mouse model was established to evaluate the efficacy of the inactivated vaccine candidate. This animal model mimics vaccination, similar to immune responses of the vaccinated. The animal model also tests protective efficacy in response to the vaccine against the disease. This work is important for the preparation of multivalent vaccines against HFMD caused by different emerging strains., (Copyright © 2021 American Society for Microbiology.)

In the face of COVID-19 pandemic caused by the newly emerged SARS-CoV-2, an inactivated, Vero cell-based, whole virion vaccine candidate has been developed and entered into phase III clinical trials within six months. Biochemical and immunogenic characterization of structural proteins and their post-translational modifications in virions, the end-products of the vaccine candidate, would be essential for the quality control and process development of vaccine products and for studying the immunogenicity and pathogenesis of SARS-CoV-2. By using a panel of rabbit antisera against virions and five structural proteins together with a convalescent serum, the spike (S) glycoprotein was shown to be N-linked glycosylated, PNGase F-sensitive, endoglycosidase H-resistant and cleaved by Furin-like proteases into S1 and S2 subunits. The full-length S and S1/S2 subunits could form homodimers/trimers. The membrane (M) protein was partially N-linked glycosylated; the accessory protein 3a existed in three different forms, indicative of cleavage and dimerization. Furthermore, analysis of the antigenicity of these proteins and their post-translationally modified forms demonstrated that S protein induced the strongest antibody response in both convalescent and immunized animal sera. Interestingly, immunization with the inactivated vaccine did not elicit antibody response against the S2 subunit, whereas strong antibody response against both S1 and S2 subunits was detected in the convalescent serum. Moreover, vaccination stimulated stronger antibody response against S multimers than did the natural infection. This study revealed that the native S glycoprotein stimulated neutralizing antibodies, while bacterially-expressed S fragments did not. The study on S modifications would facilitate design of S-based anti-SARS-CoV-2 vaccines.

The ongoing COVID-19 pandemic is causing huge impact on health, life, and global economy, which is characterized by rapid spreading of SARS-CoV-2, high number of confirmed cases and a fatality/case rate worldwide reported by WHO. The most effective intervention measure will be to develop safe and effective vaccines to protect the population from the disease and limit the spread of the virus. An inactivated, whole virus vaccine candidate of SARS-CoV-2 has been developed by Wuhan Institute of Biological Products and Wuhan Institute of Virology. The low toxicity, immunogenicity, and immune persistence were investigated in preclinical studies using seven different species of animals. The results showed that the vaccine candidate was well tolerated and stimulated high levels of specific IgG and neutralizing antibodies. Low or no toxicity in three species of animals was also demonstrated in preclinical study of the vaccine candidate. Biochemical analysis of structural proteins and purity analysis were performed. The inactivated, whole virion vaccine was characterized with safe double-inactivation, no use of DNases and high purity. Dosages, boosting times, adjuvants, and immunization schedules were shown to be important for stimulating a strong humoral immune response in animals tested. Preliminary observation in ongoing phase I and II clinical trials of the vaccine candidate in Wuzhi County, Henan Province, showed that the vaccine is well tolerant. The results were characterized by very low proportion and low degree of side effects, high levels of neutralizing antibodies, and seroconversion. These results consistent with the results obtained from preclinical data on the safety.

Inspired by the natural [NiFe]-H 2 ase, we designed mimic 1, (dppe)Ni(μ-pdt)(μ-Cl)Ru(CO) 2 Cl to realize effective H 2 evolution under photocatalytic conditions. However, a new species 2 was captured in the course of photo-, electro-, and chemo- one-electron reduction. Experimental studies of in situ IR spectroscopy, EPR, NMR, X-ray absorption spectroscopy, and DFT calculations corroborated a dimeric structure of 2 as a closed-shell, symmetric structure with a Ru I center. The isolated dimer 2 showed the real catalytic role in photocatalysis with a benchmark turnover frequency (TOF) of 1936 h -1 for H 2 evolution, while mimic 1 worked as a pre-catalyst and evolved H 2 only after being reduced to 2. The remarkably catalytic activity and unique dimer structure of 2 operated in photocatalysis unveiled a broad research prospect in hydrogenases mimics for advanced H 2 evolution., (© 2020 Wiley-VCH GmbH.)

Human Y-chromosome haplogroup C2b-F1067 is one of the dominant paternal lineages of populations in Eastern Eurasia. In order to explore the origin, diversification, and expansion of this haplogroup, we generated 206 new Y-chromosome sequences from C2b-F1067 males and coanalyzed 220 Y-chromosome sequences of this haplogroup. BEAST software was used to reconstruct a revised phylogenetic tree of haplogroup C2b-F1067 with age estimates. The revised phylogeny of C2b-F1067 included 155 sublineages, 1986 non-private variants, and >6000 private variants. The age estimation suggested that the initial splitting of C2b-F1067 happened at about 32.8 thousand years ago (kya) and the major sublineages of this haplgroup experienced continuous expansion in the most recent 10,000 years. We identified numerous sublineages that were nearly specific for Korean, Mongolian, Chinese, and other ethnic minorities in China. In particular, we evaluated the candidate-specific lineage for the Dayan Khan family and the Confucius family, the descendants of the ruling family of the Chinese Shang dynasty. These findings suggest that ancient populations with varied C2b-F1067 sublineages played an important role during the formation of most modern populations in Eastern Eurasia, and thus eventually became the founding paternal lineages of these populations.

Tilapia were exposed to 0, 0.2, 2, 20, 200 µg/L methomyl for 30 days, and then transferred to methomyl-free water for 18 days. Caspase-8 in serum, apoptosis rate, microstructure and ultra-microstructure of testis were checked after methomyl exposure and at 18 days after transferring to methomyl-free water. There were no significant changes in Caspase-8 activity, apoptosis rate, and tissue structure in testis exposed to 0.2 and 2 µg/L compared with control. However, when tilapia exposed to 20 and 200 µg/L, the Caspase-8 activity and apoptosis rate were induced significantly, and tissue damage happened compared with the control. Thus it would appear 2 µg/L methomyl might be considered as the no observed adverse effect level. Recovery data showed that the effects produced by lower concentration of 20 µg/L were reversible but not at the higher 200 µg/L concentration.

The consumption of herbal teas has become popular in recent years due to their attractive flavors and outstanding antioxidant properties. The Five-Golden-Flowers tea is a herbal tea consisting of five famous edible flowers. The effects of microwave-assisted extraction parameters on the antioxidant activity of Five-Golden-Flowers tea were studied by single-factor experiments, and further investigated using response surface methodology. Under the optimal parameters (53.04 mL/g of solvent/material ratio, 65.52 °C, 30.89 min, and 500 W), the ferric-reducing antioxidant power, Trolox equivalent antioxidant capacity, and total phenolic content of the herbal tea were 862.90 ± 2.44 µmol Fe 2+ /g dry weight (DW), 474.37 ± 1.92 µmol Trolox/g DW, and 65.50 ± 1.26 mg gallic acid equivalent (GAE)/g DW, respectively. The in vivo antioxidant activity of the herbal tea was evaluated on alcohol-induced acute liver injury in mice. The herbal tea significantly decreased the levels of aspartate aminotransferase, total bilirubin, and malonaldehyde at different doses (200, 400, and 800 mg/kg); improved the levels of liver index, serum triacylglycerol, and catalase at dose of 800 mg/kg. These results indicated its role in alleviating hepatic oxidative injury. Besides, rutin, chlorogenic acid, epicatechin, gallic acid, and p -coumaric acid were identified and quantified by high performance liquid chromatography (HPLC), which could contribute to the antioxidant activity of the herbal tea.

The discovery of aerobic and anammox bacteria capable of generating methane in bio-filters in freshwater aquaculture systems is generating interest in studies to understand the activity, diversity, distribution and roles of these environmental bacteria. In this study, we used microbial enrichment of bio-filters to assess their effect on water quality. Profiles of ammonia-oxidizing bacterial communities generated using nested PCR methods and DGGE were used to assess the expression of 16S rRNA genes using DNA sequencing. Five dominant ammonia-oxidizing bacterial strains-clones; KB.13, KB.15, KB.16, KB.17 and KB.18-were isolated and identified by phylogenetic analysis as environmental samples closely related to genera Methylobacillus, Stanieria, Nitrosomonas, and Heliorestis. The methyl ammonia-oxidizing microbes thereby found suggest a biochemical pathway involving electron donors and carbon sources, and all strains were functional in freshwater aquaculture systems. Environmental parameters including TN (2.69-20.43); COD (9.34-31.47); NH 4 + -N (0.44-11.78); NO 2 - N (0.00-3.67); NO 3 - N (0.05-1.82), mg/L and DO (1.47-10.31 µg/L) assessed varied in the ranges in the different tanks. Principal component analysis revealed that these water quality parameters significantly influenced the ammonia oxidizing microbial community composition. Temperature rises to about 40 °C significantly affected environmental characteristics-especially DO, TN and NH 4 + -N-and directly or indirectly affected the microbial communities. Although the nested PCR design was preferred due to its high sensitivity for amplifying specific DNA regions, a more concise method is recommended, as an equimolar mixture of degenerate PCR primer pairs, CTO189f-GC and CTO654r, never amplified only 16S rRNA of ammonia-oxidizing bacteria.

Owing to promoted electron-hole separation, the catalytic activity of semiconducting quantum dots (QDs) towards solar hydrogen (H2) production has been significantly enhanced by forming self-assembled clusters with ZnSe QDs made ex situ. Taking advantage of the favored interparticle hole transfer to ZnSe QDs, the rate of solar H2 evolution of CdSe QDs can be increased to ∼30 000 μmol h-1 g-1 with ascorbic acid as the sacrificial reagent, ∼150-fold higher than that of bare CdSe QDs clusters under the same conditions.

Tilapia were exposed to sub-lethal concentrations of 0, 0.2, 2, 20 or 200 μg/L for 30 days, then transferred to methomyl-free water for 18 days. E 2 , T, 11-KTand VTG in serum were examined. There were no significant changes in all the parameters in serum of tilapia exposed to 0.2 μg/L and 2 μg/L methomyl compared to the control. However, 20 μg/L and 200 μg/L have the potential to disrupt the endocrine system of male tilapia, as shown by its ability to increase VTG and E 2 and decrease T and 11-KT in serum. Thus it would appear the no observed adverse effect level for sexual steroid hormones of methomyl is lower than 2 μg/L. Recovery data showed that the effects produced by 20μg/L were reversible but not at 200μg/L. Furthermore, the sensitivity of above parameters to methomyl followed the order of VTG>E 2 >11-KT>T>GSI, suggesting VTG being the better biomarkers., (© 2017 Wiley Periodicals, Inc.)

Tilapia were exposed to sublethal methomyl concentrations of 0, 0.2, 2, 20 or 200 μg/L for 30 days, and then transferred to methomyl-free water for 18 days. The sexual steroid hormones 17β-estradiol (E2), testosterone (T), and 11-ketotestosterone (11-KT) in tilapia testes were examined at 0, 6, 12, 18, 24 and 30 days after methomyl exposure, and at 18 days after fish were transferred to methomyl-free water. There were no significant changes in the hormone parameters in testes of tilapia exposed to low concentration 0.2 and 2 μg/L methomyl compared with the controls. However, high concentration 20 and 200 μg/L methomyl had the potential to disrupt the endocrine system of male tilapia, as shown by an increase in E 2 and a decrease in T and 11-KT in the testes. Thus, it would appear that the 2 μg/L methomyl might be considered the no-observed-adverse-effect level. Recovery data showed that the effects produced by the lower concentration of 20 μg/L were reversible but the effects were not reversible at the higher concentration of 200 μg/L.

The present study clearly showed that chronic exposure to polychlorinated biphenyls (PCBs) at environmentally relevant concentrations can damage juvenile tilapia livers by modulating antioxidant enzyme activities and gene transcription, which affects toxic bioaccumulation and histological congestion. The results suggest that PCBs caused a decrease in the activity of some hepatic antioxidative and biotransformation enzymes (SOD, CAT, GST, T-GSH, and MDA) in tilapia at 7 days, as well as transcriptional changes (sod, cat, and gst). Except for some antioxidant parameters (T-GSH, GSH/GSSG, T-AOC, and MDA), significant declines and increases occurred at 14 and 21 days, respectively. Most of the antioxidant enzymatic signatures and genotoxicity significantly increased at 14 and 21 days. This study presented evidence that PCBs could result in hepatic toxicity through oxidative stress in the early growth stages of tilapia, and we speculated that oxidative stress plays an important role in embryonic developmental toxicity induced by PCBs., Competing Interests: The authors declare no conflict of interest.

Objective: Ubiquitin-specific protease 4 (USP4) is a deubiquitinating enzyme with key roles in the regulation of TGF-β1 signaling, suggesting its importance in tumorigenesis. However, the underlying mechanisms causing this are not entirely clear. In the present study, we investigated the effect of USP4 on invasion and tumorigenesis of breast cancer cells, and explored its mechanism., Materials and Methods: Effects of USP4 overexpression or USP4 silencing by small interfering RNA (USP4 siRNA) on invasion of breast cancer MDA-MB-231 and T47D cells in vitro was detected. Using siRNAs and inhibitors to examine the USP4 signaling pathway., Results: The migration and invasion assays showed that USP4 promotes human breast cancer cell migration and invasion by USP4 overexpression, and knockdown of USP4 by siRNA inhibits human breast cancer cell migration and invasion. Treatment with RLX siRNAs, TGF-β1 siRNAs, Smad2 siRNAs or BB94 (MMPs inhibitor) to USP4-overexpressing breast cancer cells revealed that USP4- induced RLX via TGF-β1 pathway promotes the cell migration and invasion. Further studies demonstrated that USP4-mediated TGF-β1 activation not only enhances the phosphorylation of Smad2 through TGF-β, but also directly upregulate matrix metalloproteinase (MMP)-9-mediated cell migration and invasion of breast cancer cells., Conclusions: Therapies targeting the USP4 inhibits invasion of breast cancer cells via Relaxin/TGF-β1/Smad2/MMP-9 signal. These results indicate that USP4 is an attractive target for breast cancer therapy.

The changes of microbial community in aquaculture systems under the effects of stocking densities and seasonality were investigated in tilapia ponds. Total DNAs were extracted from the water samples, 16S rRNA gene was amplified and the bacterial community analyzed by Illumina high-throughput sequencing obtaining 3486 OTUs, from a total read of 715,842 sequences. Basing on the analysis of bacterial compositions, richness, diversity, bacterial 16S rRNA gene abundance, water sample comparisons and existence of specific bacterial taxa within three fish ponds in a 4 months period, the study conclusively observed that the dominant phylum in all water samples were similar, and they included; Proteobacteria, Cyanobacteria, Bacteroidetes, Actinobacteria, Planctomycetes and Chlorobi, distributed in different proportions in the different months and ponds. The seasonal changes had a more pronounced effect on the bacterioplankton community than the stocking densities; however some differences between the ponds were more likely caused by feed coefficient than by stocking densities. At the same time, most bacterial communities were affected by the nutrient input except phylum Cyanobacteria that was also affected by the feed control of tilapia.

Objective: PRAME (Preferentially Expressed Antigen in Melanoma) is a tumor-associated antigen recognized by immunocytes, and it induces cytotoxic T cell-mediated responses in melanoma. PRAME is expressed in a wide variety of tumors, but in contrast with most other tumor-associated antigens, it is also expressed in leukemias. The physiologic role of PRAME remains elusive. Recently, it has found PRAME could be involved in the regulation of cell death in leukemias, but the mechanism of the function is unclear. Here, we confirm that PRAME induces leukemias cell death by regulation of S100A4/p53 signaling., Materials and Methods: The pCDNA3-PRAME plasmid and its control were transfected with the KG-1 cells. The pCDNA3-PRAME transfected KG-1 cells were then transiently transfected with S100A4 cDNA or wt-p53 siRNA. The PRAME siRNA and its control were transfected with the K562 cells. The PRAME siRNA transfected K562 cells were then transiently transfected with S100A4 siRNA or pGMp53-Lu. PRAME, S100A4 and P53 were detected by Western blot assay in different time point. Annexin V/propidium iodide and MTT methods were used to detect apoptosis and cell survival rate., Results: KG-1 cells overexpressing the PRAME gene significantly induces apoptosis and decreases proliferation in vitro, followed by down-regulation of S100A4 and up-regulation of p53. Up-regulation of S100A4 by S100A4 transfection inhibits PRAME-induced p53 up-regulation. Furthermore, up-regulation of S100A4 by S100A4 transfection or down-regulation of p53 by p53 siRNA transfection reduces apoptosis and increases proliferation in vitro. Knockdown of PRAME in K562 cells significantly increases proliferation in vitro, followed by up-regulation of S100A4 and down-regulation of p53. The downregulation of S100A4 by S100A4 siRNA transfection increased p53 expression. Furthermore, downregulation of S100A4 by S100A4 siRNA transfection or up-regulation of p53 by p53 transfection decreases proliferation in vitro., Conclusions: Our results suggest that the leukemias expressing high levels of PRAME has a favorable prognosis. PRAME promotes in vitro leukemia cells death by regulating S100A4/p53 signaling.

Tilapia were exposed to sublethal concentrations of 0, 0.2, 2, 20, or 200 μg/L for 30 days, and then transferred to methomyl-free water for 18 days. GST, GPx, GR, GSH, and GSSG in tilapia serum were examined at 0, 6, 12, 18, 24, and 30 days after methomyl exposure and at 18 days after transferring to methomyl-free water. There were no significant changes in antioxidants activities and contents in serum of tilapia exposed to 0.2 μg/L. Significant increases in GST, GR, GPx, and GSSG accompanied by a decrease in GSH were observed following methomyl exposure to 2, 20, or 200 μg/L, suggesting the presence of oxidative stress. Thus, it would appear the 0.2 μg/L methomyl might be considered the no observed adverse effect level. Recovery data showed that the effects produced by lower concentration of 20 μg/L were reversible but not at the higher 200 μg/L concentration., (Copyright © 2013 Wiley Periodicals, Inc.)

Objective: Chemoresistance is the principal reason for poor survival and disease recurrence in osteosarcoma patients. Survivin, a family member of the inhibitor of apoptosis proteins, plays an important role in inhibition of apoptosis. Survivin is expressed in a vast majority of human cancers, which is often correlated with poor prognosis in a wide variety of cancer patients. Furthermore, survivin expression is often related with chemoresistance in cancer cells, including osteosarcoma (OS). Here, we evaluated the therapeutic potential of YM155, a selective survivin suppressant alone and in combination with cisplatin using human OS models., Materials and Methods: U-2 OS, SW1353, MG-63 cells were treated with YM155, and/or cisplatin, and cell viability, apoptosis, survivin protein expression levels were then evaluated. Furthermore, the efficacy of YM155 combined with cisplatin was further examined in established xenograft models., Results: YM155 was sufficient to induce spontaneous apoptosis of OS cells. Combination with YM155 significantly augmented the cytotoxicity of cisplatin in OS cells. Combination treatment of YM155 and cisplatin showed antiproliferative effects and induced a greater rate of apoptosis than the sum of the single-treatment rates and promoted tumor regression in established OS xenograft models., Conclusions: Our findings provide evidence that YM155 could act as a survivin inhibitor on OS cells. Chemotherapeutic approaches using YM155 might enhance the benefit of the cisplatin in the treatment of OS cells. YM155 could be further developed as a potential therapeutic agent for the treatment of OS.

Hepatic antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT) of Nile tilapia in response to pesticide methomyl and recovery pattern were researched by exposing tilapia to sub-lethal methomyl concentrations of 0, 0.2, 2, 20 and 200 μg/L for 30 days, and then transferred to methomyl-free water for 18 days. Hepatic SOD and CAT were measured at 10 min (day 0), 6, 12, 18, 24 and 30 days after starting the experiment and at 18 days after transferring to methomyl-free water. The results showed hepatic SOD and CAT activities in 2, 20 and 200 μg/L groups were affected significantly, however, that in 0.2 μg/L group didn't change significantly compared to control during 30-day exposure period. Thus it would appear the 0.2 μg/L methomyl might be considered the no observed adverse effect level. Recovery data showed that, for SOD, the effects produced by lower concentration of methomyl 2 μg/L were reversible but not at concentrations higher than 20 μg/L, however, for CAT, the effects produced by all the concentrations were reversible.

The chronic effect of methomyl on the antioxidant system in tilapia (Oreochromis niloticus) was investigated. Fish were exposed to sub-lethal concentrations of 0.2, 2, 20 and 200μgL(-1) for 30 days, and then transferred to methomyl-free water for 18 days. Hepatic antioxidant parameters, including Glutathione-S-transferase (GST), Glutathione peroxidase (GPx), Glutathione reductase (GR), Reduced glutathione (GSH) and oxidized glutathione (GSSG), were measured at 10min (day 0), 6, 12, 18, 24 and 30 days after starting the experiment and at 18 days after transferring to methomyl-free water. There were no significant changes in enzymatic activity and content of antioxidants in liver of tilapia exposed to 0.2μgL(-1) methomyl compared to controls. However, the results showed significant increases in activities of GST, GR, GPx and levels of GSSG accompanied by a decrease in GSH levels following methomyl exposure in tilapia to 2, 20 or 200μgL(-1) over the 30-day exposure period and the highest induction rates in GST, GR, GPx and GSSG were 150.87%, 163.21%, 189.76%, and 179.56% of the control respectively, and the highest inhibition rate in GSH was 50.67% of the control, suggesting the presence of oxidative stress. Thus it would appear that the 0.2μgL(-1) methomyl might be considered as the no observed adverse effect level (NOAEL). Recovery data showed that the effects produced by lower concentration of methomyl 20μgL(-1) were reversible but not at the higher 200μgL(-1) concentration., (© 2013 Published by Elsevier Inc.)

Tilapia were exposed to sublethal methomyl concentrations of 0, 0.2, 2, 20, or 200 μg/L for 30 d, and then were transferred to methomyl-free water for 18 d. Renal antioxidant parameters, including catalase (CAT), superoxide dismutase (SOD), glutathione S-transferase (GST), glutathione peroxidase (GPx) , glutathione reductase (GR), total glutathione (GSH), and reduced glutathione (GSSG), were examined in tilapia at d 0, 6, 12, 18, 24, and 30 after starting the experiment and at 18 d after transferring to methomyl-free water. There were no significant changes in enzymatic activity and content of antioxidants in kidney of tilapia exposed to 0.2 μg/L methomyl compared to controls. The results showed significant increases in SOD, CAT, GST, GR, GPx, and level of GSSG accompanied by a decrease in GSH levels following methomyl exposure in tilapia to 2, 20, or 200 μg/L over the 30-d exposure period, suggesting the presence of oxidative stress. Thus, it would appear the 0.2 μg/L methomyl might be considered the no-observed-adverse-effect level (NOAEL). Recovery data showed that the effects produced by lower concentration of methomyl at 20 μg/L were reversible but not at the higher 200 μg/L concentration.

Rabies virus (RABV) causes severe neurological disease and death. As an important mechanism for generating genetic diversity in viruses, homologous recombination can lead to the emergence of novel virus strains with increased virulence and changed host tropism. However, it is still unclear whether recombination plays a role in the evolution of RABV. In this study, we isolated and sequenced four circulating RABV strains in China. Phylogenetic analyses identified a novel lineage of hybrid origin that comprises two different strains, J and CQ92. Analyses revealed that the virus 3' untranslated region (UTR) and part of the N gene (approximate 500 nt in length) were likely derived from Chinese lineage I while the other part of the genomic sequence was homologous to Chinese lineage II. Our findings reveal that homologous recombination can occur naturally in the field and shape the genetic structure of RABV populations.

In order to study phylogeography, population dynamics and molecular evolution of rabies viruses (RABVs) isolates from China, especially spatio-temporal dynamics, the timescale of RABVs evolution and its pattern of migration, we performed an extensive comparative analysis of RABV N gene sequence data, representing 167 isolates sampled from 20 provinces in a 78-year period (from 1931 through 2009). The available Chinese isolates could be divided into two distinct clades:Phylogroup clades I comprised Chinese group 1-4; Phylogroup clades II contained Chinese group 5-8. We found no evidence for positive selection (dN/dS>1) acting at any codon and found strong selective constraints for N gene. Bayesian Markov Chain Monte Carlo (MCMC) analysis suggested that the Chinese rabies viruses originated within the last 2000 years and the mean rates of nucleotide substitution for the N gene were approximately 4 x 10(-4) substitutions per site per year. The analyses of the spatial and spatio-temporal evolution indicated that RABV isolates from China migrated among different Provinces.

Objective: Based on sequencing the genomes of glycoprotein (GP) gene of rabies viruses isolated in Zhejiang, we analyzed the properties of rabies viruses genetic variation in molecular level, and to compare with those of other representative vaccine strains and street virus strains, get the information about rabies viruses variation., Methods: Suckling mice against rabies virus were selected. Overlapped fragments were amplified by RT-PCR and full-length genomes were assembled to analyze the nucleotide and deduced protein similarities and phylogenetic analyses of the GP genes., Results: The fourteen full-length genomes were completely sequenced and they had the same genetic structure with 1575 nts and deduced protein with 524 aa. Genetic analysis revealed that the nucleotide and amino acid homologies of GP gene from Zhejiang strains and other vaccine strains or street virus strains were 82.3% - 99.9% and 85.1% - 99.8%. The fourteen strains were genotype 1 according to the phylogenetic analyses. The GP amino acids of Zhejiang strains rabies virus strains without any recombination occurred in GP and no larger variation appeared in the major antigenic sites., Conclusion: The comprehensive analysis based on the first-level structure of GP demonstrated that it was possible that some advantageous antigenic epitopes existed in certain areas and potential antigenic determinants. It was evident that the GP gene of Zhejiang strains appear to be stable and their sequence similarity with the representative strains of street virus in China were higher than those of other vaccine strains. Some differences showed in the genetic structure and evolution relationship among Zhejiang strains, other street strains in other regions and vaccine strains.

Based on sequencing the full-length genomes of four Chinese Ferret-Badger and dog, we analyze the properties of rabies viruses genetic variation in molecular level, get the information about rabies viruses prevalence and variation in Zhejiang, and enrich the genome database of rabies viruses street strains isolated from China. Rabies viruses in suckling mice were isolated, overlapped fragments were amplified by RT-PCR and full-length genomes were assembled to analyze the nucleotide and deduced protein similarities and phylogenetic analyses from Chinese Ferret-Badger, dog, sika deer, vole, used vaccine strain were determined. The four full-length genomes were sequenced completely and had the same genetic structure with the length of 11, 923 nts or 11, 925 nts including 58 nts-Leader, 1353 nts-NP, 894 nts-PP, 609 nts-MP, 1575 nts-GP, 6386 nts-LP, and 2, 5, 5 nts- intergenic regions(IGRs), 423 nts-Pseudogene-like sequence (psi), 70 nts-Trailer. The four full-length genomes were in accordance with the properties of Rhabdoviridae Lyssa virus by BLAST and multi-sequence alignment. The nucleotide and amino acid sequences among Chinese strains had the highest similarity, especially among animals of the same species. Of the four full-length genomes, the similarity in amino acid level was dramatically higher than that in nucleotide level, so the nucleotide mutations happened in these four genomes were most synonymous mutations. Compared with the reference rabies viruses, the lengths of the five protein coding regions had no change, no recombination, only with a few point mutations. It was evident that the five proteins appeared to be stable. The variation sites and types of the four genomes were similar to the reference vaccine or street strains. And the four strains were genotype 1 according to the multi-sequence and phylogenetic analyses, which possessed the distinct district characteristics of China. Therefore, these four rabies viruses are likely to be street viruses already existing in the natural world.

A group of 31 rabies viruses (RABVs), recovered primarily from dogs, one deer and one human case, were collected from various areas in China between 1989 and 2006. Complete G gene sequences determined for these isolates indicated identities of nucleotide and amino acid sequences of >or=87% and 93.8%, respectively. Phylogenetic analysis of these and some additional Chinese isolates clearly supported the placement of all Chinese viruses in Lyssavirus genotype 1 and divided all Chinese isolates between four distinct groups (I-IV). Several variants identified within the most commonly encountered group I were distributed according to their geographical origins. A comparison of representative Chinese viruses with other isolates retrieved world-wide indicated a close evolutionary relationship between China group I and II viruses and those of Indonesia while China group III viruses formed an outlying branch to variants from Malaysia and Thailand. China group IV viruses were closely related to several vaccine strains. The predicted glycoprotein sequences of these RABVs variants are presented and discussed with respect to the utility of the anti-rabies biologicals currently employed in China.

The selective extraction of yttrium from heavy lanthanide by liquid-liquid extraction using CA-100 in the presence of the complexing agent, such as EDTA, DTPA, and HEDTA was investigated. The extraction of heavy lanthanide in the present of the complexing agent was suppressed when compared to that of Y because of the masking effect, but the selective extraction of Y was enhanced. All complexing agents formed 1:1 complex with rare earth elements (RE), and only free rare earth ions could take part in the extraction. The condition for separation was obtained by exploring the effects of the complexing agent concentration, the extractant concentration, pH and the equilibration time on the extraction of the heavy rare earth elements.