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Role of duck plague virus glycoprotein C in viral adsorption:Absence of specific interactions with cell surface heparan sulfate

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Journal of Integrative Agriculture 2017年第5期16卷 1145-1152页

作者： JING Yan-chun WU Ying SUN Kun-feng wang ming-shu CHENG An-chun CHEN shun JIA Ren-yong ZHU De-kang LIU Ma-feng YANG Qiao JING Bo CHEN Xiao-yue Institute of Preventive Veterinary Medicine Sichuan Agricultural UniversityChengdu 611130P.R.China Key Laboratory of Animal Diseases and Human Health of Sichuan Province Chengdu 611130P.R.China Avian Diseases Research Center College of Veterinary MedicineSichuan Agricultural UniversityChengdu 611130P.R.China

Many mammalian herpes viruses utilize heparan sulfate （HS） moieties present on cell surface proteoglycans as receptors for cell entry, and this process also requires viral glycoprotein C （gC） homologues. However, our understanding of the role of gC in facilitating attachment of other alpha-herpes viruses such as the duck plague virus （DPV） remains preliminary. To study the role of gC during DPV infection, we used a gC-deleted mutant virus （DPV-AgC-EGFP）. Examination of the viral copy number by real-time PCR, as well as time course studies of viral adsorption and proliferation revealed that gC was involved in the viral binding to the cell surface. The affinity of viral glycoproteins （gB-DPV, gC-DPV, and gE-DPV） to HS was assessed using a prokaryotic expression system and HJTrapTM HeparJn HP column chromatography. In addition, to confirm that gC played a role in the interaction between DPV and HS, viruses were treated with the HS analogue heparin and host cells were treated with its inhibitors heparinase prior to exposure to DPV-△gC-EGFP or wild-type strain Chinese virulent duck plague virus （DPV-CHv）. The effects of heparin and heparinase on virus infectivity demonstrated that function of gC on Viral adsorption is independent of interactions between gC and heparin sulfate on cell surface. All in all, this study demonstrated that the gC of DPV can mediate viral adsorption in an HS-independent manner, which distinguish it from the gC of some other alpha-herpes viruses. Future studies will be required to identify the receptors involved in gC protein binding to cells. This work provides us a foundation for further studies of examining the roles of gC in the adsorption during duck plague virus infection.

关键词： duck plague virus （DPV） glycoprotein C （gC） heparan sulfate （HS） viral adsorption

Development and optimization of a double antibody sandwich ELISA for the detection of goose T cell surface CD8α molecule

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Journal of Integrative Agriculture 2016年第10期15卷 2363-2368页

作者： ZHANG Wei CHENG Bei-bei CHEN shun wang ming-shu JIA Ren-yong ZHU De-kang LIU Ma-feng LIU Fei SUN Kun-feng YANG Qiao WU Ying CHEN Xiao-yue CHENG An-chun Institute of Preventive Veterinary Medicine Sichuan Agricultural University Avian Disease Research Center College of Veterinary Medicine Sichuan Agricultural University Key Laboratory of Animal Disease and Human Health of Sichuan Province Sichuan Agricultural University

CD8, a glycoprotein on the surface of T cells, is involved in the defense against viral infection and plays significant roles in antigen presentation and in the antiviral immune response. CD8 is composed of two chains. Of these, the CD8α chain was chosen for the detection because it involved in both the CD8αα homodimer and the CD8αβ heterodimer. Here, we established a double antibody sandwich enzyme-linked immunosorbent assay（DAS-ELISA） for specific detection of goose CD8α（go CD8α）. The results showed that the optimal coated antibody and antigen dilutions were 1：50（the antibody titer was 1：12 800） and 1：32（0.3 ng m L^–1）, respectively, while the optimal capture antibody and horseradish peroxidase（HRP）-labelled goat anti-rabbit Ig G dilutions were 1：50（the antibody titer was 1：51 200） and 1：4 000（the antibody titer was 1：5 000）, respectively. The optimal blocking buffer was 5% bovine serum albumin（BSA）. The best incubating condition was overnight at 4℃, the best blocking time was 120 min and the best anti-capture antibody working time was 150 min. In addition, the minimum dose detectable by DAS-ELISA was 5×10^–3 ng m L^–1. Most importantly, go CD8α expression levels in goose spleen mononuclear cells（MNCs） post-Goose parvoviruse（GPV） infection were found to be significantly up-regulated using the DAS-ELISA method, which was consistent with previous results obtained using real-time quantitative PCR. In conclusion, the DAS-ELISA method reported here is a novel, specific technique for the clinical detection of go CD8α.

关键词： T cells goose CD8α polyclonal antibody double antibody sandwich ELISA

The codon-optimized capsid gene of duck circovirus can be highly expressed in yeast and self-assemble into virus-like particles

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Journal of Integrative Agriculture 2017年第7期16卷 1601-1608页

作者： YANG Cui XU Yu JIA Ren-yong LIU Si-yang wang ming-shu ZHU De-kang CHEN shun LIU Ma-feng ZHAO Xin-xin SUN Kun-feng JING Bo YIN Zhong-qiong CHENG An-chun Avian Disease Research Centre Sichuan Agricultural UniversityChengdu 611130P.R.China Institute of Preventive Veterinary Medicine Sichuan Agricultural UniversityChengdu 611130P.R.China Key Laboratory of Animal Disease and Human Health of Sichuan Province Chengdu 611130P.R.China

The capsid （Cap） protein, which is the only structural protein of duck circovirus （DuCV）, is the most important antigen for the development of vaccines against DuCV and the virus＇s serological diagnostic methods. In order to use yeast expression system to produce a large quantities of DuCVCap protein which is close to its natural form to display the antigen peptides perfectly, the Cap gene was optimized into the codon-optimized capsid （Opt-Cap） gene towards the preference of yeast firstly. Then, the genes of Cap and Opt-Cap were separately cloned into pPIC9K plasmid and transformed into Picha pas- toris GSl15. The strains that displayed the phenotype of Mut~ and contained multiple inserts of expression cassette were selected from those colonies. After the induction expression, the secretory type of Cap protein, which was about 43 kDa, was best expressed under 0.5% （v/v） methanol and sorbitol induction. Compared with the Cap gene, the expression level of Opt-Cap gene was much higher. What＇s more, the purified Cap protein had a good reactivity to its specific polyclone antibody and DuCV-positive serum, and it was able to self-assemble into virus-like particles （VLPs）. These VLPs, with a diameter of 15-20 nm and without a nucleic acid structure, showed a high level of similarity to DuCV particles in size and shape. All of the resultsdemonstrated that, based on the codon-optimization, it is suitable to use the P. pastoris expression system to produce DuCV VLPs on a large scale. It is the first time that a large amounts of DuCV VLPs were produced successfully in P. pastoris, which might be particularly useful for the further studies of serological diagnosis and vaccines of DuCV.

关键词： capsid gene codon＇optimization duck circovirus virus-like particles

Immunogenicity and protective efficacy of DHBV DNA vaccines expressing envelope and capsid fusion proteins in ducks delivered by attenuated Salmonella typhimurium

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Journal of Integrative Agriculture 2018年第4期17卷 928-939页

作者： LIU Si-yang JIA Ren-yong LI Qing-qing FENG Dai-shen SHEN Hao-yue YANG Cui wang ming-shu ZHU De-kang CHEN shun LIU Ma-feng ZHAO Xin-xin YIN Zhong-qiong JING Bo CHENG An-chun Avian Disease Research Center Sichuan Agricultural University Chengdu 611130 P.R. China Institute of Preventive Veterinary Medicine Sichuan Agricultural University Chengdu 611130 P.R. China Key Laboratory of Animal Disease and Human Health of Sichuan Province Chengdu 611130 P.R.China

Duck hepatitis B virus(DHBV) shares many basic characteristics with hepatitis B virus(HBV) and is an attractive model for vaccine development. In this study, DHBV DNA vaccines were designed to express envelope and capsid fusion proteins to enhance the breadth of immune response in ducks. Attenuated Salmonella typhimurium(SL7207) was used as a carrier and adjuvant to boost the magnitude of immune response. Based on this strategy, novel DNA vaccines(SL7207-p VAX1-LC and SL7207-p VAX1-SC) were generated. Growth kinetics, genetic stabilities and relative transcription levels of the L, S and C genes introduced by these vaccine strains were measured before inoculation to guarantee safety and efficacy. The relative transcript levels of the CD4 and CD8 T genes and the antibody levels(Ig Y) in ducks receiving the vaccines were higher than those in single gene delivered groups. Additionally, the copy number of covalently closed circular DNA in hepatocytes after DHBV challenge also provided evidence that our fusion vaccines could enhance the protective efficiency against DHBV infection in ducks.

关键词： fusion DNA genes attenuated Salmonella typhimurium(SL7207) hepatitis B virus(HBV) duck hepatitis B virus(DHBV) oral immunization

Developing a diagnostic model for predicting prostate cancer: a retrospective study based on Chinese multicenter clinical data

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Asian Journal of Andrology 2024年第1期26卷 34-40页

作者： Chang-ming wang Lei Yuan Xue-Han Liu shu-Qiu Chen Hai-Feng wang Qi-Fei Dong Bin Zhang ming-shuo Huang Zhi-Yong Zhang Jun Xiao Tao Tao Department of Urology The First Affiliated Hospital of USTCDivision of Life Sciences and MedicineUniversity of Science and Technology of ChinaHefei 230001China Department of Radiology The First Affiliated Hospital of USTCDivision of Life Sciences and MedicineUniversity of Science and Technology of ChinaHefei 230001China Core Facility Center for Medical Sciences The First Affiliated Hospital of USTcDivision of Life Sciences and MedicineUniversity of Science and Technology of ChinaHefei 230001China Department of Urology Affiliated Zhongda Hospital of Southeast UniversityNanjing 210009China Department of Urology Shanghai East HospitalTongji University School of MedicineShanghai 200120China Department of Urology Shanghai Changhai HospitalSecond Military Medical UniversityShanghai 200000China Department of Urology Affiliated Anhui Provincial Hospital of Anhui Medical UniversityHefei 230001China

The overdiagnosis of prostate cancer(PCa)caused by nonspecific elevation serum prostate-specific antigen(PSA)and the overtreatment of indolent PCa have become a global problem that needs to be solved urgently.We aimed to construct a prediction model and provide a risk stratification system to reduce unnecessary biopsies.In this retrospective study,clinical data of 1807 patients from three Chinese hospitals were used.The final model was built using stepwise logistic regression analysis.The apparent performance of the model was assessed by receiver operating characteristic curves,calibration plots,and decision curve analysis.Finally,a risk stratification system of clinically significant prostate cancer(csPCa)was created,and diagnosis-free survival analyses were performed.Following multivariable screening and evaluation of the diagnostic performances,a final diagnostic model comprised of the PSA density and Prostate Imaging-Reporting and Data System(PI-RADS)score was established.Model validation in the development cohort and two external cohorts showed excellent discrimination and calibration.Finally,we created a risk stratification system using risk thresholds of 0.05 and 0.60 as the cut-off values.The follow-up results indicated that the diagnosis-free survival rate for csPCa at 12 months and 24 months postoperatively was 99.7%and 99.4%,respectively,for patients with a risk threshold below O.05 after the initial negative prostate biopsy,which was significantly better than patients with higher risk.Our diagnostic model and risk stratification system can achieve a personalized risk calculation of csPCa.It provides a standardized tool for Chinese patients and physicians when considering thenecessity of prostatebiopsy.

关键词： nomogram prostate biopsy prostate cancer Prostate Imaging-Reporting and Data System prostate-specific antigen density

利用高密度电阻率法进行多年冻土探测的研究与应用

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地质与资源 2024年第3期33卷 339-348页

作者：杜海双王书明宋庆元周传芳殷启春张天恩孙桐中国地质调查局哈尔滨自然资源综合调查中心黑龙江哈尔滨150000 中国地质大学地球物理与空间信息学院湖北武汉430074

多年冻土退化后,融区周期性冻结和融化作用会严重损害输油管道的稳定性.为查明原油管道埋设区多年冻土分布、发育现状,计划分别采用二维、三维高密度电法进行探测.为了进一步研究高密度电法不同探测方式的有效性和冻土的电阻率响应特征... 详细信息

多年冻土退化后,融区周期性冻结和融化作用会严重损害输油管道的稳定性.为查明原油管道埋设区多年冻土分布、发育现状,计划分别采用二维、三维高密度电法进行探测.为了进一步研究高密度电法不同探测方式的有效性和冻土的电阻率响应特征,首先在管道下部空间建立未融化、半融化、贯穿融化3种模型,采用EarthImager 2D和EarthImager 3D软件进行有限单元法正演和圆滑约束的最小二乘法反演;随后根据数值模拟结果,对比研究3种模型电阻率响应特征及反演结果的差异性,总结二维、三维高密度电法勘探效果;最后利用勘探实例进一步验证.结果表明,高密度电法在多年冻土探测领域是一种行之有效的方法:二维高密度电法对未融化、贯穿融化探测效果优于半融化状态;三维高密度电法采集数据更加丰富,目标体形态展示更加直观精细,反演结果更加接近理论模型,适用各种冻土融化状态.野外实例进一步验证了数值模拟结果,三维高密度电法的应用也对多年冻土精细探测领域提供了新思路.

关键词：高密度电阻率法二维模型三维模型多年冻土原油管道

Alphaherpesvirus-vectored vaccines against animal diseases: Current progress

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Journal of Integrative Agriculture 2020年第8期19卷 1928-1940页

作者： HU Yang wang ming-shu CHENG An-chun JIA Ren-yong YANG Qiao WU Ying LIU Ma-feng ZHAO Xin-xin ZHU De-kang CHEN shun ZHANG Sha-qiu wang Yin GAO Qun OU Xu-min MAO Sai WEN Xing-jian XU Zhi-wen CHEN Zheng-li ZHU Ling LUO Qi-hui TIAN Bin PAN Lei-chang Mujeeb Ur REHMAN LIU Yun-ya YU Yan-ling ZHANG Ling CHEN Xiao-yue Institute of Preventive Veterinary Medicine Sichuan Agricultural UniversityChengdu 611130P.R.China Key Laboratory of Animal Disease and Human Health of Sichuan Province Sichuan Agricultural UniversityChengdu 611130P.R.China Avian Disease Research Center College of Veterinary MedicineSichuan Agricultural UniversityChengdu 611130P.R.China

Recombinant virus-vectored vaccines are novel agents that can effectively activate specific and nonspecific immunity,are multivalent and multieffective,and have high safety ratings.Animal alphaherpesviruses have a large genome,contain multiple nonessential regions that do not affect viral replication and are capable of accepting the insertion of an exogenous gene and expressing the antigen protein.Furthermore,animal alphaherpesviruses have a wide host spectrum,can replicate in the host and continuously stimulate the animal to produce immunity to the corresponding pathogen,thus making them ideal carriers for recombinant virus-vectored vaccines.With the development of gene-editing technology,recombinant viruses capable of expressing foreign genes can be constructed by various methods.Currently,studies on recombinant virusvectored vaccines constructed based on animal alphaherpesviruses have involved poultry,pigs,cattle,sheep,and companion animals.Studies have shown that the construction of recombinant animal alphaherpesviruses enables the acquisition of immunity to multiple diseases.This article mainly summarizes the current progress on animal alphaherpesvirus-vectored vaccines,aiming to provide reference for the development of new animal alphaherpesvirus-vectored vaccines.

关键词： recombinant vectored-vaccine recombinant alphaherpesviruses immunity gene-editing exogenous gene

Mitogenome-based phylogeny of mosquitoes(Diptera:Culicidae)

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Insect Science 2024年第2期31卷 599-612页

作者： De-Hong Chen shu-Lin He Wen-Bo Fu Zhen-Tian Yan Yun-Jian Hu Huan Yuan ming-Bin wang Bin Chen Chongqing Key Laboratory of Vector Insects Institute of Entomology and Molecular BiologyChongqing Normal UniversityChongqingChina

Mosquitoes are of great medical significance as vectors of many deadly diseases.Mitogenomes have been widely used in phylogenetic studies,but mitogenome knowledge within the family Culicidae is limited,and Culicidae phylogeny is far from resolved.In this study,we surveyed the mitogenomes of 149 Culicidae species,including 7 newly sequenced species.Comparative analysis of 149 mosquito mitogenomes shows gene composition and order to be identical to that of an ancestral insect,and the AT bias,length variation,and codon usage are all consistent with that of other reported Dipteran mitogenomes.Phylogenetic analyses based on the DNA sequences of the 13 protein-coding genes from the 149 species robustly support the monophyly of the subfamily Anophelinae and the tribes Aedini,Culicini,Mansonini,Sabethini,and Toxorhynchitini.To resolve ambiguous relationships between clades within the subfamily Culicinae,we performed topological tests and show that Aedini is a sister to Culicini and that Uranotaenini is a sister to(Mansonini)+(Toxorhynchitini+Sabethini).In addition,we estimated divergence times using a Bayesian relaxation clock based on the sequence data and 3 fossil calibration points.The results show mosquitoes diverged during the Early Jurassic with massive Culicinae radiations during the Cretaceous,coincident with the emergence of angiosperms and the burst of mammals and birds.Overall,this study,which uses the largest number of Culicidae mitogenomes sequenced to date,comprehensively reveals the mitogenome characteristics and mitogenome-based phylogeny and divergence times of Culicidae,providing information for further studies on the mitogenome,phylogeny,evolution,and taxonomic revision of Culicidae.

关键词： characterization Culicidae evolution mitogenomes mosquitoes phylogeny

Amino acids modified nanoscale zero-valent iron:Density functional theory calculations,experimental synthesis and application in the Fenton-like degradation of organic solvents

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Journal of Environmental Sciences 2024年第1期135卷 296-309页

作者： Xingchen Yang Fucheng ming Jianlong wang Lejin Xu Department of Nuclear Engineering and Technology School of Energy and Power EngineeringHuazhong University of Science and TechnologyWuhan 430074China China-EU Institute for Clean and Renewable Energy Huazhong University of Science and TechnologyWuhan 430074China Institute of Nuclear and New Energy Technology Key Laboratory of Advanced Reactor Engineering and Safety of Ministry of EducationTsinghua UniversityBeijing 100084China Beijing Key Laboratory of Radioactive Wastes Treatment Tsinghua UniversityBeijing 100084China

To improve the adsorption and catalytic performance of heterogeneous Fenton-like catalysts for oil wastes,amino acids were used to modify nanoscale zero-valent iron(AA@Fe^(0)),which were applied in the Fenton-like degradation of organic solvents(tributyl phosphate and n-dodecane,named TBP and DD).Twelve amino acids,i.e.,glycine(Gly),alanine(Ala),leucine(Leu),proline(Pro),phenylalanine(Phe),methionine(Met),cysteine(Cys),asparagine(Asn),serine(Ser),glutamic acid(Glu),lysine(Lys)and arginine(Arg),were selected and calculated by density functional theory(DFT).The optimized structure,charge distribution,the highest occupied molecular orbital(HOMO),the lowest unoccupied molecular orbital(LUMO),interaction region indicator(IRI)isosurface map and adsorption energy of AA@Fe^(0),AA@Fe^(0)-TBP and AA@Fe^(0)-DD were studied,which indicated that Fe is more likely to approach and charge transfer with-COO and-NH_(3) on theα-carbon of amino acids.There is strong attraction between Fe and–COO,and Van der Waals force between Fe and-NH_(3),respectively.In the interaction of AA@Fe^(0)with TBP and DD,Van der Waal force plays an important role.AA@Fe^(0)was synthesized in laboratory and characterized to investigate physicochemical properties.In Fenton-like degradation of organic solvents,the change of COD in water phase during the degradation process as well as the volume of the organic phase after the reaction were investigated.The results of calculations combined with experiments showed that Ser-modified Fe^(0)performed the best in these amino acids,with 98%removal of organic solvents.A possible catalytic mechanism was proposed in which amino acids acted a linking role between Fe and organic solvents,activating H_(2)O_(2)to generate hydroxyl radicals for the degradation of organic solvents.

关键词： Amino acids Nanoscale zero-valent iron Density functional theory Organic solvents Fenton-like degradation