Reverse Genetics System for Severe Fever with Thrombocytopenia Syndrome Virus

Brennan, Benjamin; Li, Ping; Zhang, Shuo; Li, Aqian; Liang, Mifang; Li, Dexin; Elliott, Richard M.

doi:10.1128/jvi.03432-14

Cited by 71 publications

(91 citation statements)

References 49 publications

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“…Using SFTSV generated by a reverse-genetic system, Brennan et al showed that the F330S mutation in the M segment was responsible for plaque formation and had a high growth rate in Vero E6 cells (15). The mutation in the GP was detected in the high plaque-forming strain HB29pp, and it was related to Vero E6 cell adaption.…”

Section: Discussionmentioning

confidence: 99%

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Establishment of Subclones of the Severe Fever with Thrombocytopenia Syndrome Virus YG1 Strain Selected Using Low pH-Dependent Cell Fusion Activity

et al. 2017

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Section: Discussionmentioning

confidence: 99%

“…Recently, SFTSV and other genetically related viruses that are pathogenic for humans have been reported in East Asia and North America (8)(9)(10)(11)(12)(13). However, their molecular biological characteristics and pathogenic mechanisms are not fully understood (5,14,15).…”

Section: Introductionmentioning

confidence: 99%

Establishment of Subclones of the Severe Fever with Thrombocytopenia Syndrome Virus YG1 Strain Selected Using Low pH-Dependent Cell Fusion Activity

et al. 2017

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“…In addition, we have shown that NSs is not essential for virus replication but that its absence results in a degree of growth attenuation in cell culture. Recently, we described a reverse-genetics system for pathogenic SFTSV (28). The developed reverse-genetics systems will be useful tools to elucidate the molecular determinants of virulence of the more pathogenic tick-borne phleboviruses.…”

Section: Discussionmentioning

confidence: 99%

“…Thus, UUKV would be a useful comparator to include in studies on the molecular basis of the pathogenesis of STSFV and HRTV in humans. Such investigations will be aided by the availability of reverse-genetics systems that allow specific manipulation of the viral genome, and we have recently established such a system for SFTSV (28). Here, we describe a reverse-genetics system for the recovery of infectious UUKV entirely from cDNA copies of the genome.…”

mentioning

confidence: 99%

Generation of Mutant Uukuniemi Viruses Lacking the Nonstructural Protein NSs by Reverse Genetics Indicates that NSs Is a Weak Interferon Antagonist

Rezelj

Överby

Elliott

2015

J Virol

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Uukuniemi virus (UUKV) is a tick-borne member of the Phlebovirus genus (family Bunyaviridae) and has been widely used as a safe laboratory model to study aspects of bunyavirus replication. Recently, a number of new tick-borne phleboviruses have been discovered, some of which, like severe fever with thrombocytopenia syndrome virus and Heartland virus, are highly pathogenic in humans. UUKV could now serve as a useful comparator to understand the molecular basis for the different pathogenicities of these related viruses. We established a reverse-genetics system to recover UUKV entirely from cDNA clones. We generated two recombinant viruses, one in which the nonstructural protein NSs open reading frame was deleted from the S segment and one in which the NSs gene was replaced with green fluorescent protein (GFP), allowing convenient visualization of viral infection. We show that the UUKV NSs protein acts as a weak interferon antagonist in human cells but that it is unable to completely counteract the interferon response, which could serve as an explanation for its inability to cause disease in humans. IMPORTANCE Uukuniemi virus (UUKV) is a tick-borne phlebovirus that is apathogenic for humans and has been used as a convenient model to investigate aspects of phlebovirus replication. Recently, new tick-borne phleboviruses have emerged, such as severe fever with thrombocytopenia syndrome virus in China and Heartland virus in the UnitedStates, that are highly pathogenic, and UUKV will now serve as a comparator to aid in the understanding of the molecular basis for the virulence of these new viruses. To help such investigations, we have developed a reverse-genetics system for UUKV that permits manipulation of the viral genome. We generated viruses lacking the nonstructural protein NSs and show that UUKV NSs is a weak interferon antagonist. In addition, we created a virus that expresses GFP and thus allows convenient monitoring of virus replication. These new tools represent a significant advance in the study of tick-borne phleboviruses. The Bunyaviridae are the largest grouping of RNA viruses, comprising Ͼ350 members. The family includes a wide range of enveloped, single-stranded RNA viruses with tripartite genomes of negative-sense or ambisense polarity. The Phlebovirus genus contains 70 viruses divided into two groups: the sandfly fever group, transmitted mainly by phlebotomines and mosquitoes, and the Uukuniemi-like virus group, transmitted by ticks. Uukuniemi virus (UUKV) itself was originally isolated in 1964 from an Ixodes ricinus tick in Finland (1). UUKV has served as a prototype tick-borne phlebovirus for many years and has aided in advances in bunyavirus structural and architectural determination (2-4), bunyavirus glycoprotein studies (5-9), and studies of bunyavirus entry into mammalian cells (10-12).Like other phleboviruses, the three UUKV genome segments, designated small (S), medium (M), and large (L), are found in the form of ribonucleoprotein (RNP) complexes in association with the nucleocapsid (N)...

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“…Generation of viral stocks was performed in Vero76 cells, with titers determined by plaque assay as previously described (18,25). A multiplicity of infection (MOI) of ϳ0.01 was used in all experiments involving virus infection, unless stated otherwise.…”

Section: Methodsmentioning

confidence: 99%

Extracellular Vesicles Mediate Receptor-Independent Transmission of Novel Tick-Borne Bunyavirus

Silvas

Popov

Paulucci-Holthauzen

et al. 2016

J Virol

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Severe fever with thrombocytopenia syndrome (SFTS) virus is a newly recognized member of the genus Phlebovirus in the family Bunyaviridae. The virus was isolated from patients presenting with hemorrhagic manifestations and an initial case fatality rate of 12 to 30% was reported. Due to the recent emergence of this pathogen, there is limited knowledge on the molecular virology of SFTS virus. Recently, we reported that the SFTS virus NSs protein inhibited the activation of the beta interferon (IFN-␤) promoter. Furthermore, we also found that SFTS virus NSs relocalizes key components of the IFN response into NSs-induced cytoplasmic structures. Due to the important role these structures play during SFTS virus replication, we conducted live cell imaging studies to gain further insight into the role and trafficking of these cytoplasmic structures during virus infection. We found that some of the SFTS virus NSs-positive cytoplasmic structures were secreted to the extracellular space and endocytosed by neighboring cells. We also found that these secreted structures isolated from NSs-expressing cells and SFTS virus-infected cells were positive for the viral protein NSs and the host protein CD63, a protein associated with extracellular vesicles. Electron microscopy studies also revealed that the isolated CD63-immunoprecipitated extracellular vesicles produced during SFTS virus infection contained virions. The virions harbored within these structures were efficiently delivered to uninfected cells and were able to sustain SFTS virus replication. Altogether, these results suggest that SFTS virus exploits extracellular vesicles to mediate virus receptor-independent transmission to host cells and open the avenue for novel therapeutic strategies against SFTS virus and related pathogens. IMPORTANCESFTS virus is novel bunyavirus associated with hemorrhagic fever illness. Currently, limited information is available about SFTS virus. In the present study, we demonstrated that extracellular vesicles produced by SFTS virus-infected cells harbor infectious virions. We sought to determine whether these "infectious" extracellular vesicles can mediate transmission of the virus and confirmed that the SFTS virions were efficiently transported by these secreted structures into uninfected cells and were able to sustain efficient replication of SFTS virus. These results have significant impact on our understanding of how the novel tick-borne phleboviruses hijack cellular machineries to establish infection and point toward a novel mechanism for virus replication among arthropod-borne viruses. The Bunyaviridae family comprises five genera, including Orthobunyavirus, Phlebovirus, Nairovirus, Hantavirus, and Tospovirus. The majority of viruses within this family (with the exception of Hantaviruses) are considered arthropod-borne viruses and are important causes of morbidity and mortality around the world. These viruses are associated with a range of clinical symptoms characterized by febrile illness and in the most severe cases fatal hepa...

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Reverse Genetics System for Severe Fever with Thrombocytopenia Syndrome Virus

Cited by 71 publications

References 49 publications

Establishment of Subclones of the Severe Fever with Thrombocytopenia Syndrome Virus YG1 Strain Selected Using Low pH-Dependent Cell Fusion Activity

Establishment of Subclones of the Severe Fever with Thrombocytopenia Syndrome Virus YG1 Strain Selected Using Low pH-Dependent Cell Fusion Activity

Generation of Mutant Uukuniemi Viruses Lacking the Nonstructural Protein NSs by Reverse Genetics Indicates that NSs Is a Weak Interferon Antagonist

Extracellular Vesicles Mediate Receptor-Independent Transmission of Novel Tick-Borne Bunyavirus

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