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Molecular characterization of an outbreak-involved Bacillus anthracis strain confirms the spillover of anthrax from West Africa

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Infectious Diseases of Poverty 2024年第1期13卷 93-94页

作者： Shuchao Wang Roland Suluku Mohamed B.Jalloh Ahmed F.Samba Baogui Jiang Yubiao Xie Doris Harding Mengyao Zhang Foday Sahr Mahmud E.Sesay James S.Squire Mohamed A.Vandi Moinina N.Kallon Shoufeng Zhang rongliang hu Yuee Zhao Zhiqiang Mi Changchun Veterinary Research Institute Chinese Academy of Agricultural Sciences Changchun China Department of Animal Sciences School of Agriculture and Food Sciences Njala University Njala Sierra Leone Department of Microbiology College of Medicine and Allied Health Sciences University of Sierra Leone Freetown Sierra Leone Ministry of Agriculture and Food Sciences Freetown Sierra Leone Beijing Institute of Microbiology and Epidemiology Beijing China Ministry of Health and Sanitation Freetown Sierra Leone Beijing Institute of Biotechnology Beijing China

Background Anthrax,a zoonotic disease caused by the spore-forming bacterium Bacillus anthracis,remains a major global public health concern,especially in countries with limited resources.Sierra Leone,a West African country historically plagued by anthrax,has almost been out of report on this disease in recent decades.In this study,we described a large-scale anthrax outbreak affecting both animals and humans and attempted to characterize the path-ogenusingmoleculartechniques.Methods The causative agent of the animal outbreak in Port Loko District,Sierra Leone,between March and May 2022 was identified using the nanopore sequencing technique.A nationwide active surveillance was implemented from May 2022 to June 2023 to monitor the occurrence of anthrax-specific symptoms in humans.Suspected cases were subsequently verified using quantitative polymerase chain reaction.Full-genome sequencing was accomplished by combining long-read and short-read sequencing methods.Subsequent phylogenetic analysis was performed based on the full-chromosome single nucleotide polymorphisms.Results The outbreak in Port Loko District,Sierra Leone,led to the death of 233 animals between March 26th and May 16th,2022.We ruled out the initial suspicion of Anaplasma species and successfully identified B.anthracis as the causative agent of the outbreak.As a result of the government's prompt response,out of the 49 suspected human cases identifed during the one-year active surveillance,only 6 human cases tested positive,all within the frst month after the official declaration of the outbreak.The phylogenetic analysis indicated that the BaSL2022 isolate responsible for the outbreak was positioned in the A.Br.153 clade within the TransEuroAsian group of B.anthracis.Conclusions We successfully identifed a large-scale anthrax outbreak in Sierra Leone.The causative isolate of B.anthracis,BaSL2022,phylogenetically bridged other lineages in A.Br.153 clade and neighboring genetic groups,A.Br.144 and A.Br.148,eventually confirming the spillover of anthrax f

关键词： Molecular identification Anthrax Bacillus anthracis Sierra Leone Phylogeny Single nucleotide polymorphism

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A Semi-quantitative Serological Method to Assess the Potency of Inactivated Rabies Vaccine for Veterinary Use

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Virologica Sinica 2012年第4期27卷 259-264页

作者： Ye Liu Shoufeng Zhang Fei Zhang rongliang hu Laboratory of Epidemiology Military Veterinary Research InstituteAcademy of Military Medical SciencesChangchun 130122China

Potency is one of the most important indexes of inactivated vaccines.A number of methods have been established to assay the potency,of which the NIH test and single-dose mouse protection test are the "prescribed methods".Here,we report a method to semi-quantitatively assay the potency of an inactivated rabies vaccine,which uses fewer animals and takes less time to complete.Depending on the quality requirements of a vaccine(e.***.minimum potency),a rabies reference vaccine is,for example,diluted to the minimum potency,and 50 μL of the dilution is taken to inoculate 10 mice.The same amount of the test rabies vaccine is inoculated into another 10 mice.After two weeks,all mice are bled and serum samples are assayed for viral neutralizing antibody by the fluorescent antibody virus neutralization(FAVN) test.By comparing the median and interquartile range of antibody titers of the reference vaccine with those of the test vaccine,the test vaccine potency can be semi-quantitatively judged as to whether it is in accord with the required quality.The reliability of this method was also confirmed in dogs.The procedure can be recommended for batch potency testing during inactivated rabies vaccine production.

关键词：狂犬病疫苗半定量检测灭活疫苗血清学方法效价美国国立卫生研究院病毒中和试验保护测试

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Immunogenicity of multi-epitope-based vaccine candidates administered with the adjuvant Gp96 against rabies

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Virologica Sinica 2016年第2期31卷 168-175页

作者： Yange Niu Ye Liu Limin Yang Hongren Qu Jingyi Zhao rongliang hu Jing Li Wenjun Liu Key Laboratory of Pathogenic Microbiology and Immunology Institute of Microbiology ChineseAcademy of Sciences Beijing 100101 China University of Chinese Academy of Sciences Beijing 100049 China Laboratory of Epidemiology and Key Laboratory of Jilin Province for Zoonosis Prevention and Control Veterinary Research Institute Academy of Military Medical Sciences Changchun 130122 China Beijing Center for Animal Disease Control and Prevention Chinese Veterinary Medical Association of Animal Clinic Branch Beijing 102629 China State Key Laboratory of Veterinary Biotechnology Harbin Veterinary Research Institute Chinese Academy of Agricultural Sciences Harbin 150001 China

Rabies, a zoonotic disease, causes > 55,000 human deaths globally and results in at least 500 million dollars in losses every year. The currently available rabies vaccines are mainly inactivated and attenuated vaccines, which have been linked with clinical diseases in animals. Thus, a rabies vaccine with high safety and efficacy is urgently needed. Peptide vaccines are known for their low cost, simple production procedures and high safety. Therefore, in this study, we examined the efficacy of multi-epitope-based vaccine candidates against rabies virus. The ability of various peptides to induce epitope-specific responses was examined, and the two peptides that possessed the highest antigenicity and conservation, i.e., AR16 and h PAB, were coated with adjuvant canineGp96 and used to prepare vaccines. The peptides were prepared as an emulsion of oil in water(O/W) to create three batches of bivalent vaccine products. The vaccine candidates possessed high safety. Virus neutralizing antibodies were detected on the day 14 after the first immunization in mice and beagles, reaching 5–6 IU/m L in mice and 7–9 IU/m L in beagles by day 28. The protective efficacy of the vaccine candidates was about 70%–80% in mice challenged by a virulent strain of rabies virus. Thus, a novel multi-epitope-based rabies vaccine with Gp96 as an adjuvant was developed and validated in mice and dogs. Our results suggest that synthetic peptides hold promise for the development of novel vaccines against rabies.

关键词：狂犬病疫苗 gp96 免疫原性多表位疫苗注射佐剂狂犬病病毒候选疫苗

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Epidemic and Maintenance of Rabies in Chinese Ferret Badgers (Melogale moschata) indicated by Epidemiology and the Molecular Signatures of Rabies Viruses

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Virologica Sinica 2013年第3期28卷 146-151页

作者： Shoufeng Zhang Ye Liu Yanli Hou Jinghui Zhao Fei Zhang Ying Wang rongliang hu Key Laboratory of Jilin Province for Zoonosis Prevention and Control Laboratory of Epidemiology Veterinary Research Institute Academy of Military Medical Sciences Centre for Computer Fundamental Education Jilin University

An epidemic of Chinese ferret badger-associated human rabies was investigated in Wuyuan county, Jiangxi province and rabies viruses isolates from ferret badgers in different districts in Jiangxi and Zhejiang provinces were sequenced with their nucleotides and amino acids and aligned for epidemiological analysis. The results showed that the human rabies in Wuyuan are only associated with ferret badger bites; the rabies virus can be isolated in a high percentage of ferret badgers in the epidemic areas in Jiangxi and Zhejiang provinces; the isolates share the same molecular features in nucleotides and have characteristic amino acid signatures, i.e., 2 sites in the nucleoprotein and 3 sites in the glycoprotein, that are distinct from virus isolates from dogs in the same region. We conclude that rabies in Chinese ferret badgers has formed an independent transmission cycle and ferret badgers may serve as another important rabies reservoir independent of dog rabies in China.

关键词：流行病学分析狂犬病毒分子特征鼬獾中国签名南瓜维护

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RNA疫苗研究进展及狂犬病RNA疫苗

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中国兽医学报 2024年第5期44卷 1078-1086页

作者：汪芋缪发明张守峰么乃全扈荣良吉林农业大学动物医学院吉林长春130118 中国农业科学院长春兽医研究所吉林长春130122

狂犬病(rabies)是由狂犬病病毒(rabies virus, RABV)引起的一种严重人畜共患的急性接触性传染病,其致死率为100%,每年全球约6万人死于该病,其中亚洲和非洲发生最多。RABV是狂犬病的致病病原体,由结构蛋白和负链RNA构成。狂犬病一旦发病... 详细信息

狂犬病(rabies)是由狂犬病病毒(rabies virus, RABV)引起的一种严重人畜共患的急性接触性传染病,其致死率为100%,每年全球约6万人死于该病,其中亚洲和非洲发生最多。RABV是狂犬病的致病病原体,由结构蛋白和负链RNA构成。狂犬病一旦发病致死率为100%,目前疫苗接种仍是预防和控制狂犬病最为有效的策略。RNA疫苗是新一代核酸疫苗,其安全性好,有效性高,设计灵活,研发周期短,且借助高效递送系统,可刺激机体快速产生强烈免疫应答。RNA疫苗这一特点对于潜伏期短且病死率高的狂犬病的防控具有重要应用价值。现对RNA疫苗及狂犬病RNA疫苗的研究进行系统梳理,以阐明其研究现状及应用前景。

关键词： RNA疫苗狂犬病免疫应答递送系统

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Fox- and raccoon-dog–associated rabies outbreaks in northern China

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Virologica Sinica 2014年第5期29卷 308-310页

作者： Ye Liu Shoufeng Zhang Jinghui Zhao Fei Zhang Nan Li Hai Lian Wurengege Shiyu Guo rongliang hu Key Laboratory of Jinlin Provincial Zoonosis Laboratory of Epidemiology Veterinary Research Institute Academy of Military Medical Sciences Department of Physiology Hetao College Department of Physiology School of Medicine Showa University

Dear Editor,Rabies is a generally fatal disease caused by the rabies virus(RABV),and is transmitted mainly by Carnivora and Chiroptera(Fooks A R,et al.,2014;Tao X,et al.,2013).In China,stray dogs and some wild animals(e.g.,Chinese ferret badgers,foxes,and raccoon dogs)are the principal reservoirs for RABV(hu R L,et al.,2009).Historically,rabies in wild foxes and raccoon dogs(Nyctereutes procyonoides)was recorded in the

关键词： rabies Fox reservoirs fatal transmitted isolate cattle Changchun Heilongjiang surveillance

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Nanopore sequencing of African swine fever virus

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Science China(Life Sciences) 2020年第1期63卷 160-164页

作者： Lijia Jia Mengwei Jiang Ke Wu Juefu hu Yang Wang Weipeng Quan Mengchan Hao Haizhou Liu Hongping Wei Wenhui Fan Wenjun Liu rongliang hu Depeng Wang Jing Li Jianjun Chen Di Liu CAS Key Laboratory of Special Pathogens and Biosafety Wuhan Institute of VirologyChinese Academy of SciencesWuhan 430071China Computational Virology Group Center for Bacteria and Viruses Resources and BioinformationWuhan Institute of VirologyChinese Academy of SciencesWuhan 430071China African Swine Fever Regional Laboratory of China Wuhan 430071China University of Chinese Academy of Sciences Beijing 100049China GrandOmics Biosciences Beijing 101312China NextOmics Biosciences Wuhan 430074China CAS Key Laboratory of Pathogenic Microbiology and Immunology Institute of MicrobiologyChinese Academy of SciencesBeijing 100101China Center for Biosafety Mega-Science Chinese Academy of SciencesWuhan 430071China Institute of Military Veterinary Medicine Academy of Military Medical ScienceChangchun 130122China

Dear Editor,African swine fever(ASF)is one of the most pathogenic viral diseases in pigs caused by African swine fever virus(ASFV).The fatality rate is almost 100%,which brings huge economic losses to the hog industry in countries with epiepidemics(Galindo and Alonso,2017).China was the first Asian country to have an ASF epidemic,and it spread quickly across the country after the first epidemic was re・ported in August 2018(Ge et al.,2018).After that,Mongolia,Vietnam,Cambodia and North Korea also reported on the ASF epidemic in succession(OIE,2019).

关键词： swine fever African

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A SYBR-Green I quantitative Real-time Reverse Transcription-PCR assay for rabies viruses with different virulence

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Virologica Sinica 2014年第2期29卷 131-132页

作者： Lindong Wang Ye Liu Shoufeng Zhang Ying Wang Jinghui Zhao Fuchun Miao rongliang hu Key Laboratory of Jilin Province for Zoonosis Prevention and Control Laboratory of Epidemiology Veterinary Research InstituteAcademy of Military Medical Sciences

Dear Editor,To establish a more specific diagnostic assay than thecurrent methods for rabies virus(RABV)infection in ca-nines and wildlife species,we developed a SYBR-GreenI quantitative real-time reverse transcription-PCR(RT-PCR).Three primers were designed and synthesized:oneof which,located within the leader sequences and target-ed for the whole viral genome amplification to obtain

关键词：狂犬病病毒逆转录PCR 实时定量 SYBR PCR检测 RT-PCR方法核蛋白基因毒力

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非洲猪瘟病毒p54间接ELISA抗体检测方法的建立及试剂盒研制

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中国兽医学报 2024年第5期44卷 907-913页

作者：岳慧贤李其炫惠丽丽赵心馨陈腾张艳艳王述超缪发明张菲张守峰扈荣良中国农业科学院长春兽医研究所吉林长春130122

非洲猪瘟是对全球养猪业危害最严重的病毒性传染病,目前尚无有效疫苗。p54蛋白是非洲猪瘟病毒(ASFV)的重要结构蛋白之一,参与病毒的吸附和侵入,是非洲猪瘟血清学诊断的重要靶标之一。本研究采用无标签的重组p54蛋白作为包被抗原,建立了A... 详细信息

非洲猪瘟是对全球养猪业危害最严重的病毒性传染病,目前尚无有效疫苗。p54蛋白是非洲猪瘟病毒(ASFV)的重要结构蛋白之一,参与病毒的吸附和侵入,是非洲猪瘟血清学诊断的重要靶标之一。本研究采用无标签的重组p54蛋白作为包被抗原,建立了ASFV p54间接ELISA抗体检测方法,优化了检测条件,组装成了试剂盒。根据对374份阴性猪血清的检测,确定了本试剂盒检测的临界值,并利用不同猪病原阳性质控血清进行了敏感性、特异性和稳定性研究。结果显示,本方法检测阴/阳性血清的临界值为0.250;敏感性血清质控品的最小检出限为1∶1 600倍,与ASFV阳性血清、猪伪狂犬病病毒阳性血清、猪繁殖与呼吸病毒综合征病毒阳性血清、猪圆环病毒2型阳性血清、猪细小病毒阳性血清和猪抗大肠杆菌阳性血清等6种特异性质控血清均无交叉反应;试制的3批试剂盒批间和批内变异系数均小于10%。试剂盒在2~8℃贮存9个月,特异性、敏感性保持不变。以上试验结果表明,本试剂盒具有良好的敏感性、特异性和稳定性,与OIE推荐的p72阻断ELISA抗体检测试剂盒具有较高符合率,为非洲猪瘟临床血清学诊断和抗体监测提供了良好的技术支持。

关键词：非洲猪瘟病毒 p54蛋白间接ELISA 试剂盒

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非洲猪瘟病毒p30蛋白在本氏烟草中的表达及其抗体间接ELISA检测方法的建立

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中国兽医学报 2024年第1期44卷 12-19,28页

作者：刘悦魏天张春雨张建峰王成宇张芳毓朱日宁鞠安琪王思月曲磊张守峰扈荣良梁静王永志吉林农业大学生命科学学院吉林长春130118 吉林省农业科学院畜牧兽医研究所吉林长春130124 中国农业科学院长春兽医研究所吉林长春130122

为建立一种非洲猪瘟病毒(African swine fever virus,ASFV)抗体间接ELISA检测方法,利用马铃薯Y病毒(potato virus Y,PVY)介导的植物生物反应器在本氏烟草中表达ASFV p30蛋白,并以表达的蛋白作为包被抗原,对间接ELISA各反应条件进行优化... 详细信息

为建立一种非洲猪瘟病毒(African swine fever virus,ASFV)抗体间接ELISA检测方法,利用马铃薯Y病毒(potato virus Y,PVY)介导的植物生物反应器在本氏烟草中表达ASFV p30蛋白,并以表达的蛋白作为包被抗原,对间接ELISA各反应条件进行优化。依据ASFV SY-18株的CP204L(p30)基因序列,按照本氏烟草密码子的偏好性,对p30基因(231~536 bp)的核苷酸序列进行优化、人工合成及克隆,通过In-Fusion连接技术将扩增的p30基因插入到PVY基因组的P1和HC-Pro基因之间,构建PVY-p30重组病毒质粒,以摩擦的方式将质粒接种本氏烟草叶片,14 d后,通过RT-PCR和Western blot分析目的基因的转录和重组蛋白的表达情况,并利用Ni-NTA-琼脂糖亲和层析柱纯化本氏烟草中表达的p30重组蛋白,Western blot分析p30重组蛋白的纯化效果和反应原性,将纯化后的p30重组蛋白作为包被抗原,建立ASFV抗体间接ELISA检测方法。结果显示,构建的PVY-p30重组病毒能够系统性侵染本氏烟草;p30重组蛋白能够在本氏烟草叶片中表达,且具有良好的反应原性;纯化的p30重组蛋白效果较好,能够被p30蛋白单克隆抗体识别。建立的ELISA检测方法最佳反应条件为抗原质量浓度0.5 mg/L,37℃包被1 h;待检血清1∶400倍稀释,37℃孵育0.5 h;HRP标记二抗37℃孵育0.5 h;TMB室温避光显色10 min;阴阳性临界值为0.461。当ASFV阳性血清稀释度为1∶2560时,仍检测为阳性,具有较高的敏感性;该方法与其他常见病阳性血清不发生交叉反应,能够特异性地检测ASFV抗体;重复性试验显示批内变异系数小于5%,批间变异系数小于10%;该方法与ASFV商品化检测试剂盒相比,总体符合率为85.0%。结果表明,本研究构建的PVY-p30重组病毒载体能介导p30重组蛋白在本氏烟草中表达,以植物表达的p30重组蛋白为抗原建立的ASFV抗体间接ELISA检测方法可用于疾病的临床检测及疫情监测,同时为可饲疫苗的研发提供了基础材料。

关键词：间接ELISA 非洲猪瘟病毒马铃薯Y病毒植物生物反应器 p30蛋白

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