Epidemiology and Mutational Analysis of Global Strains of Crimean-Congo Haemorrhagic Fever Virus

Crimean-Congo hemorrhagic fever (CCHF) is a severe illness with high fatality.Cases are reported in several countries in Africa,Europe,the Middle East,and Asia.Phylogenetic analyses based on the virus S (nucleocapsid),M (glycoprotein),and L (polymerase) genome segments sequences indicate distinct geographic lineages exist but their specific genetic characteristics require elucidation.In this work we collected all full length S segment sequences and generated a phylogenetic tree based on the alignment of these 62 samples.We then analyzed the alignment using entries from AAIndex,the Amino Acid Index database,to identify amino acid mutations that performed significant changes in charge,pka,hydropathy and side chain volume.Finally,we mapped these changes back to the tree and alignment to identify correlated mutations or sites that characterized a specific lineage.Based on this analysis we are able to propose a number of sites that appear to be important for virus function and which would be good candidates for experimental mutational analysis studies.

Production of CCHF Virus-Like Particle by a Baculovirus-Insect Cell Expression System

Crimean-Congo Haemorrhagic Fever Virus (CCHFV) is a tick-born virus of the Nairovirus genus within the Bunyaviridae family,which is widespread and causes high fatality.The nucleocapsid of CCHFV is comprised of N proteins that are encoded by the S segment.In this research,the N protein of CCHFV was expressed in insect cells using a recombinant baculovirus.Under an electron microscope,Virus-Like Particles (VLPs) with various size and morphology were observed in cytoplasmic vesicles in the infected cells.Sucrose-gradient purification of the cell lysate indicated that the VLPs were mainly located in the upper fraction after ultracentrifugation,which was confirmed by Western blot analysis and immuno-electron microscopy (IEM).

Crimean-Congo hemorrhagic fever from the immunopathogenesis, clinical, diagnostic, and therapeutic perspective: A scoping review

Crimean-Congo hemorrhagic fever virus(CCHFV) is responsible for widespread tick-borne zoonotic viral disease CCHF in African, Middle Eastern, Asian, and European countries. CCHFV can be spread to humans through tick bites or contact with infected animals or humans, and it often progresses from asymptomatic to severe/lethal illness, with fatality rates ranging from 10% to 40% in humans. Today, CCHF is growing into a significant public health concern due to its very high prevalence, severity of the condition, and lack of available vaccines and specific treatments. Recent research has been drawn towards a more accurate study of CCHFV characteristics, including the structure, genetic diversity, mechanisms involved in pathogenesis and immunopathogenesis, and clinical features. In addition, the use of animal models(mouse and non-human primates) and advanced diagnostic tools in recent years has resulted in a significant advance in CCHF related studies. In this context, we summarized the latest findings about CCHF research, its health complications, animal models, current diagnosis, vaccination, and CCHF treatments, and therapeutic strategies. Furthermore, we discussed existing deficiencies and problems in CCHFV analysis, as well as areas that still need to yield conclusive answers.

Prevalence and Phylogenetic Analysis of Crimean-Congo Hemorrhagic Fever Virus in Ticks from Different Ecosystems in Xinjiang,China

The Crimean-Congo hemorrhagic fever virus(CCHFV),a member of the genus Orthonairovirus and family Nairoviridae,is transmitted by ticks and causes severe hemorrhagic disease in humans.To study the epidemiology of CCHFV in different ecosystems in Xinjiang,China,a total of 58,932 ticks were collected from Tarim Basin,Junggar Basin,Tianshan Mountain,and Altai Mountain from 2014 to 2017.Hyalomma asiaticum asiaticum was the dominant tick species in Tarim and Junggar basins,whereas Dermacentor nuttalli and Hyalomma detritum were found in Tianshan Mountain and Altai Mountain,respectively.Reverse transcription-polymerase chain reaction of the CCHFV small(S)genome segment was used for the molecular detection.The CCHFV-positive percentage was 5.26%,6.85%,1.94%,and 5.56% in Tarim Basin,Junggar Basin,Tianshan Mountain,and Altai Mountain,respectively.Sequences of the S segment were used for phylogenetic analysis and the results showed that the newly identified CCHFV strains belonged to two clades.Our study confirms that H.asiaticum asiaticum is the major vector of CCHFV in desert habitats which is consistent with previous studies,and also suggests that H.detritum and D.nuttalli are emerging vectors for CCHFV in Xinjiang.Moreover,this study reports the presence of CCHFV in the mountain habitat of Xinjiang for the first time,suggesting that future surveillance of CCHFV should also include mountainous areas.

A new strain of Crimean-Congo hemorrhagic fever virus isolated from Xinjiang, China

Crimean-Congo hemorrhagic fever virus(CCHFV) is a highly pathogenic tick-borne virus with a fatality rate of up to 50% in humans. CCHFV is widely distributed in countries around the world.Outbreaks of CCHFV infection in humans have occurred in prior years in Xinjiang Province, China.Epidemiological surveys have detected CCHFV RNA in ticks and animals; however, few isolates were identified. In this study, we identified and isolated a new CCHFV strain from Hyalomma asiaticum asiaticum ticks collected from north of Tarim Basin in Xinjiang, China. A preliminary investigation of infection and antigens expression of CCHFV was performed in newborn mice. The target tissues for CCHFV replication in newborn mice were identified. The analysis of the phylogenetic relationships with other Chinese strains suggested that diverse genotypes of CCHFV have circulated in Xinjiang for years. These findings provide important insights into our understanding of CCHFV infection and evolution as well as disease prevention and control for local residents.

Isolation,Characterization,and Phylogenetic Analysis of Two New Crimean-Congo Hemorrhagic Fever Virus Strains from the Northern Region of Xinjiang Province,China

Crimean-Congo hemorrhagic fever(CCHF)caused by the CCHF virus(CCHFV)is a tick-borne natural focal disease with a mortality rate of approximately 50%.CCHFV is widely prevalent in Africa,southern Asia,the Middle East,and southeast Europe.CCHF outbreaks have been reported previously in Xinjiang province,China,especially in its southern region.Epidemiological surveys conducted on ticks and animals have revealed the presence of CCHFV strains in ticks,rodents,and infected individuals from cities and counties in southern Xinjiang.Phylogenetic analyses revealed that the Chinese CCHFV strains belong to one genotype,based on complete sequences of the S segments of its negative-stranded RNA genome.The present study reports two new CCHFV strains isolated from Hyalomma asiaticum asiaticum ticks collected from Fukang City and Wujiaqu City in the northern region of Xinjiang.Viral characteristics and their evolutionary relationships were analyzed through metagenomic and reverse-transcription PCR analyses;these analyses indicated that the genotype of both strains was different from that of other Chinese strains.Furthermore,previous reports of CCHFV in Xinjiang were reviewed and phylogenetic analyses were performed.CCHFV was found to prevail in Fukang City in Junggar Basin for more than 20 years,and that Fukang City and Wujiaqu City are considered natural reservoirs of different genotypes of CCHFV strains.Our findings facilitate the understanding of CCHFV distribution in Xinjiang province and provide insights into the evolutionary relationships among Chinese CCHFV strains.

Cryo-EM structure of glycoprotein C from Crimean-Congo hemorrhagic fever virus

Crimean-Congo hemorrhagic fever virus(CCHFV)is a causative agent of serious hemorrhagic diseases in humans with high mortality rates.CCHFV glycoprotein Gc plays critical roles in mediating virus-host membrane fusion and has been studied extensively as an immunogen.However,the molecular mechanisms involved in membrane fusion and Gc-specific antibody-antigen interactions remain unresolved largely because structural information of this glycoprotein is missing.We designed a trimeric protein including most of the ectodomain region of Gc from the prototype CCHFV strain,Ib Ar10200,which enabled the cryo-electron microscopy structure to be solved at a resolution of 2.8Å.The structure confirms that CCHFV Gc is a class Ⅱ fusion protein.Unexpectedly,structural comparisons with other solved Gc trimers in the postfusion conformation revealed that CCHFV Gc adopted hybrid architectural features of the fusion loops from hantaviruses and domain Ⅲ from phenuiviruses,suggesting a complex evolutionary pathway among these bunyaviruses.Antigenic sites on CCHFV Gc that protective neutralizing antibodies target were mapped onto the CCHFV Gc structure,providing valuable information that improved our understanding of potential neutralization mechanisms of various antibodies.

Crimean-Congo hemorrhagic fever virus in Central,Eastern,and South-eastern Asia

Crimean-Congo hemorrhagic fever(CCHF),caused by Crimean-Congo hemorrhagic fever virus(CCHFV),is endemic in Africa,Asia,and Europe,but CCHF epidemiology and epizootiology is only rudimentarily defined for most regions.Here we summarize what is known about CCHF in Central,Eastern,and South-eastern Asia.Searching multiple international and country-specific databases using a One Health approach,we defined disease risk and burden through identification of CCHF cases,anti-CCHFV antibody prevalence,and CCHFV isolation from vector ticks.We identified 2313 CCHF cases that occurred in 1944–2021 in the three examined regions.Central Asian countries reported the majority of cases(2,026).In Eastern Asia,China was the only country that reported CCHF cases(287).In South-eastern Asia,no cases were reported.Next,we leveraged our previously established classification scheme to assign countries to five CCHF evidence levels.Six countries(China,Kazakhstan,Kyrgyzstan,Tajikistan,Turkmenistan,and Uzbekistan)were assigned to level 1 or level 2 based on CCHF case reports and the maturity of the countries’surveillance systems.Two countries(Mongolia and Myanmar)were assigned to level 3 due to evidence of CCHFV circulation in the absence of reported CCHF cases.Thirteen countries in Eastern and South-eastern Asia were categorized in levels 4 and 5 based on prevalence of CCHFV vector ticks.Collectively,this paper describes the past and present status of CCHF reporting to inform international and local public-health agencies to strengthen or establish CCHFV surveillance systems and address shortcomings.

Transcriptome profiling highlights regulated biological processes and type III interferon antiviral responses upon Crimean-Congo hemorrhagic fever virus infection

Crimean-Congo hemorrhagic fever virus(CCHFV)is a biosafety level-4(BSL-4)pathogen that causes Crimean-Congo hemorrhagic fever(CCHF)characterized by hemorrhagic manifestation,multiple organ failure and high mortality rate,posing great threat to public health.Despite the recently increasing research efforts on CCHFV,host cell responses associated with CCHFV infection remain to be further characterized.Here,to better understand the cellular response to CCHFV infection,we performed a transcriptomic analysis in human kidney HEK293 cells by high-throughput RNA sequencing(RNA-seq)technology.In total,496 differentially expressed genes(DEGs),including 361 up-regulated and 135 down-regulated genes,were identified in CCHFV-infected cells.These regulated genes were mainly involved in host processes including defense response to virus,response to stress,regulation of viral process,immune response,metabolism,stimulus,apoptosis and protein catabolic process.Therein,a significant up-regulation of type III interferon(IFN)signaling pathway as well as endoplasmic reticulum(ER)stress response was especially remarkable.Subsequently,representative DEGs from these processes were well validated by RT-qPCR,confirming the RNA-seq results and the typical regulation of IFN responses and ER stress by CCHFV.Furthermore,we demonstrate that not only type I but also type III IFNs(even at low dosages)have substantial anti-CCHFV activities.Collectively,the data may provide new and comprehensive insights into the virus-host interactions and particularly highlights the potential role of type III IFNs in restricting CCHFV,which may help inform further mechanistic delineation of the viral infection and development of anti-CCHFV strategies.

Serial Expression of the Truncated Fragments of the Nucleocapsid Protein of CCHFV and Identification of the Epitope Region

The Crimean-congo hemorrhagic fever virus(CCHFV)is a geographically widespread fatal pathogen. Identification of the epitope regions of the virus is important for the diagnosis and epidemiological studies of CCHFV infections.In this study,expression vectors carrying series truncated fragments of the NP(nucleocapsid protein)gene from the S fragment of CCHFV strain YL04057 were constructed.The recombinant proteins were expressed in E.coli and purified for detection.The antigenic of the truncated fragments of NP was detected with a polyclonal serum(rabbit)and 2 monoclonal(mAbs)(14B7 and 43E5)against CCHFV by Western-blot analyses. The results showed that the three expressed constructs,which all contained the region 235AA to 305AA could be detected by mAbs polyclonal serum.The results suggest that region 235-305 aa of NP is a highly antigenic region and is highly conserved in the NP protein.

Mapping the viruses belonging to the order Bunyavirales in China

Background:Viral pathogens belonging to the orderBunyavirales pose a continuous background threat to global health,but the fact remains that they are usually neglected and their distribution is still ambiguously known.We aim to map the geographical distribution ofBunyavirales viruses and assess the environmental suitability and transmission risk of majorBunyavirales viruses in China.Methods:We assembled data on allBunyavirales viruses detected in humans,animals and vectors from multiple sources,to update distribution maps of them across China.In addition,we predicted environmental suitability at the 10 km×10 km pixel level by applying boosted regression tree models for two importantBunyavirales viruses,including Crimean-Congo hemorrhagic fever virus(CCHFV)and Rift Valley fever virus(RVFV).Based on model-projected risks and air travel volume,the imported risk of RVFV was also estimated from its endemic areas to the cities in China.Results:Here we mapped all 89 species ofBunyavirales viruses in China from January 1951 to June 2021.Nineteen viruses were shown to infect humans,including ten species first reported as human infections.A total of 447,848 cases infected withBunyavirales viruses were reported,and hantaviruses,Dabie bandavirus and Crimean-Congo hemorrhagic fever virus(CCHFV)had the severest disease burden.Model-predicted maps showed that Xinjiang and southwestern Yunnan had the highest environmental suitability for CCHFV occurrence,mainly related toHyalomma asiaticum presence,while southern China had the highest environmental suitability for Rift Valley fever virus(RVFV)transmission all year round,mainly driven by livestock density,mean precipitation in the previous month.We further identified three cities including Guangzhou,Beijing and Shanghai,with the highest imported risk of RVFV potentially from Egypt,South Africa,Saudi Arabia and Kenya.Conclusions:A variety ofBunyavirales viruses are widely distributed in China,and the two major neglectedBunyavirales viruses including CCHFV and RVFV,both have the potential for outbreaks in local areas of China.Our study can help to promote the understanding of risk distribution and disease burden ofBunyavirales viruses in China,and the risk maps of CCHFV and RVFV occurrence are crucial to the targeted surveillance and control,especially in seasons and locations at high risk.

Prevalence of Crimean-Congo hemorrhagic fever in Pakistan and its new research progress

Crimean-Congo hemorrhagic fever is a deadly and life-threatening viral sickness spreading throughout the world with high mortality rate of 10%–40%.The causative agents are ticks which show diversity in their strains and thus it is difficult to make vaccine,however some strains are conserved which is the positive point in vaccines development.From 2012 to 2015,a total of 161 cases were confirmed for Crimean-Congo hemorrhagic fever virus in Pakistan.And from 2012 to July 2014,45 deaths were reported in the country.It is spreading sporadically in Pakistan.Mapping of endemic areas and cross-border veterinary surveillance should be developed in high risk areas and workshops should be arranged to increase public awareness about the disease.Control measures must be taken to avoid spreading of the disease to new areas.

新疆地区克里米亚刚果出血热病毒M片段的遗传分析

目的为进一步了解克里米亚刚果出血热病毒(CCHFV)M基因的特征,对新疆地区4株克里米亚刚果出血热病毒分离株亚东璃眼蜱的部分M片段核苷酸序列进行测定及分析。方法从感染CCHF病毒的鼠脑中提取RNA,应用逆转录聚合酶链反应扩增,测定CCHF病毒的部分M片段核苷酸序列,与GenBank中的已登录的部分CCHF病毒M基因的同源序列进行比较分析和种系进化分析。结果经测序得到了4株病毒的3962～4385 nt位点的424 bp核苷酸序列(位点依据IbAr10200的M基因序列),该序列共编码139个氨基酸。前3株病毒序列有相当高的核苷酸同源性(99.5%～99.8%),将所得到的序列与已发表的中国、尼日利亚、巴基斯坦、乌兹别克斯坦、塔吉克斯坦和俄罗斯病毒株相比,中国株之间的核苷酸同源性为78.1%～99.8%,而非中国株则为42.9%～94.8%。但它们所编码的氨基酸序列变异不大,同源性非常高。系统发生树表明,所有的毒株被分为5个进化支(所比较的M基因长度为3962～4385 nt核苷酸),来自新疆南部和东部的YT05099、YLD4041和YL05035共处于同一分支。结论相比于新疆其他分离株,YT05099、YL04041和YL05035的部分M基因特征有极高的相似性。新疆株CCHF病毒M基因与中东和远东病毒株之间有较近的亲缘关系。