Differential short-term and long-term metabolic and cytokine responses to infection of severe fever with thrombocytopenia syndrome virus.

INTRODUCTION: Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease caused by the SFTS virus (SFTSV), which has a wide geographic distribution. The primary clinical manifestations of SFTS are fever and thrombocytopenia, with multiorgan failure being the leading cause of death. While most patients recover with treatment, little is known about the potential long-term metabolic effects of SFTSV infection. OBJECTIVES: This study aimed to shed light on dysregulated metabolic pathways and cytokine responses following SFTSV infection, which pose significant risks to the short-term and long-term health of affected individuals. METHODS: Fourteen laboratory-confirmed clinical SFTS cases and thirty-eight healthy controls including 18 SFTSV IgG-positive and 20 IgG-negative individuals were recruited from Taizhou city of Zhejiang province, Eastern China. Inclusion criteria of healthy controls included residing in the study area for at least one year, absence of fever or other symptoms in the past two weeks, and no history of SFTS diagnosis. Ultrahigh-performance liquid chromatography-mass spectrometry (UHPLC-MS) was used to obtain the relative abundance of plasma metabolites. Short-term metabolites refer to transient alterations present only during SFTSV infection, while long-term metabolites persistently deviate from normal levels even after recovery from SFTSV infection. Additionally, the concentrations of 12 cytokines were quantified through fluorescence intensity measurements. Differential metabolites were screened using orthogonal projections to latent structures discriminant analysis (OPLS-DA) and the Wilcoxon rank test. Metabolic pathway analysis was performed using MetaboAnalyst. Between-group differences of metabolites and cytokines were examined using the Wilcoxon rank test. Correlation matrices between identified metabolites and cytokines were analyzed using Spearman's method. RESULTS AND CONCLUSIONS: We screened 122 long-term metabolites and 108 short-term metabolites by analytical comparisons and analyzed their correlations with 12 cytokines. Glycerophospholipid metabolism (GPL) was identified as a significant short-term metabolic pathway suggesting that the activation of GPL might be linked to the self-replication of SFTSV, whereas pentose phosphate pathway and alanine, aspartate, and glutamate metabolism were indicated as significant long-term metabolic pathways playing a role in combating long-standing oxidative stress in the patients. Furthermore, our study suggests a new perspective that α-ketoglutarate could serve as a dietary supplement to protect recovering SFTS patients.

The Phlebovirus Ribonucleoprotein: An Overview.

In negative strand RNA viruses, ribonucleoproteins, not naked RNA, constitute the template used by the large protein endowed with polymerase activity for replicating and transcribing the viral genome. Here we give an overview of the structures and functions of the ribonucleoprotein from phleboviruses. The nucleocapsid monomer, which constitutes the basic structural unit, possesses a flexible arm allowing for a conformational switch between a closed monomeric state and the formation of a polymeric filamentous structure competent for viral RNA binding and encapsidation in the open state of N. The modes of N-N oligomerization as well as interactions with vRNA are described. Finally, recent advances in tomography open exciting perspectives for a more complete understanding of N-L interactions and the design of specific antiviral compounds.

The Effect of Tryptophan-to-Tyrosine Mutation at Position 61 of the Nonstructural Protein of Severe Fever with Thrombocytopenia Syndrome Virus on Viral Replication through Autophagosome Modulation.

In our prior investigations, we elucidated the role of the tryptophan-to-tyrosine substitution at the 61st position in the nonstructural protein NSsW61Y in diminishing the interaction between nonstructural proteins (NSs) and nucleoprotein (NP), impeding viral replication. In this study, we focused on the involvement of NSs in replication via the modulation of autophagosomes. Initially, we examined the impact of NP expression levels, a marker for replication, upon the infection of HeLa cells with severe fever thrombocytopenia syndrome virus (SFTSV), with or without the inhibition of NP binding. Western blot analysis revealed a reduction in NP levels in NSsW61Y-expressing conditions. Furthermore, the expression levels of the canonical autophagosome markers p62 and LC3 decreased in HeLa cells expressing NSsW61Y, revealing the involvement of individual viral proteins on autophagy. Subsequent experiments confirmed that NSsW61Y perturbs autophagy flux, as evidenced by reduced levels of LC3B and p62 upon treatment with chloroquine, an inhibitor of autophagosome-lysosome fusion. LysoTracker staining demonstrated a decrease in lysosomes in cells expressing the NS mutant compared to those expressing wild-type NS. We further explored the mTOR-associated regulatory pathway, a key regulator affected by NS mutant expression. The observed inhibition of replication could be linked to conformational changes in the NSs, impairing their binding to NP and altering mTOR regulation, a crucial upstream signaling component in autophagy. These findings illuminate the intricate interplay between NSsW61Y and the suppression of host autophagy machinery, which is crucial for the generation of autophagosomes to facilitate viral replication.

Gn protein expressed in plants for diagnosis of severe fever with thrombocytopenia syndrome virus.

Severe fever with thrombocytopenia syndrome virus (SFTSV) causes the highly fatal disease in humans. To facilitate diagnosis, the native form of subunit glycoprotein (Gn), a prime target for potential vaccines and therapies, was produced in Nicotiana benthamiana using a Bamboo mosaic virus-based vector system. By fusion with secretory signal tags, SSExt, derived from the extension protein, and the (SP)10 motif, the yield of the recombinant Gn (rGn) was remarkably increased to approximately 7 mg/kg infiltrated leaves. Ultimately, an rGn-based ELISA was successfully established for the detection of SFTSV-specific antibodies in serum samples from naturally infected monkeys. As validated with the reference method, the specificity and sensitivity of rGn-ELISA were 94% and 96%, respectively. In conclusion, utilizing well-suited fusion tags facilitates rGn production and purification in substantial quantities while preserving its antigenic properties. The rGn-ELISA, characterized by its commendable sensitivity and specificity could serve as a viable alternative diagnostic method for assessing SFTSV seroprevalence. KEY POINTS: • SFTSV Gn, fused with secretory signal tags, was expressed by the BaMV-based vector. • The plant fusion tags increased expression levels and eased the purification of rGn. • The rGn-ELISA was established and validated; its specificity and sensitivity > 94%.

CCR2 is a host entry receptor for severe fever with thrombocytopenia syndrome virus.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is an emerging tick-borne bunyavirus causing a high fatality rate of up to 30%. To date, the receptor mediating SFTSV entry remained uncharacterized, hindering the understanding of disease pathogenesis. Here, C-C motif chemokine receptor 2 (CCR2) was identified as a host receptor for SFTSV based on a genome-wide CRISPR-Cas9 screen. Knockout of CCR2 substantially reduced viral binding and infection. CCR2 enhanced SFTSV binding through direct binding to SFTSV glycoprotein N (Gn), which is mediated by its N-terminal extracellular domain. Depletion of CCR2 in C57BL/6J mouse model attenuated SFTSV replication and pathogenesis. The peripheral blood primary monocytes from elderly individuals or subjects with underlying diabetes mellitus showed higher CCR2 surface expression and supported stronger binding and replication of SFTSV. Together, these data indicate that CCR2 is a host entry receptor for SFTSV infection and a novel target for developing anti-SFTSV therapeutics.

The SFTSV Nonstructural Proteins Induce Autophagy to Promote Viral Replication via Interaction with Vimentin.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is a newly identified phlebovirus associated with severe hemorrhagic fever in humans. Studies have shown that SFTSV nucleoprotein (N) induces BECN1-dependent autophagy to promote viral assembly and release. However, the function of other SFTSV proteins in regulating autophagy has not been reported. In this study, we identify SFTSV NSs, a nonstructural protein that forms viroplasm-like structures in the cytoplasm of infected cells as the virus component mediating SFTSV-induced autophagy. We found that SFTSV NSs-induced autophagy was inclusion body independent, and most phenuivirus NSs had autophagy-inducing effects. Unlike N protein-induced autophagy, SFTSV NSs was key in regulating autophagy by interacting with the host's vimentin in an inclusion body-independent manner. NSs interacted with vimentin and induced vimentin degradation through the K48-linked ubiquitin-proteasome pathway. This negatively regulating Beclin1-vimentin complex formed and promoted autophagy. Furthermore, we identified the NSs-binding domain of vimentin and found that overexpression of wild-type vimentin antagonized the induced effect of NSs on autophagy and inhibited viral replication, suggesting that vimentin is a potential antiviral target. The present study shows a novel mechanism through which SFTSV nonstructural protein activates autophagy, which provides new insights into the role of NSs in SFTSV infection and pathogenesis. IMPORTANCE Severe fever with thrombocytopenia syndrome virus (SFTSV) is a newly emerging tick-borne pathogen that causes multifunctional organ failure and even death in humans. As a housekeeping mechanism for cells to maintain steady state, autophagy plays a dual role in viral infection and the host's immune response. However, the relationship between SFTSV infection and autophagy has not been described in detail yet. Here, we demonstrated that SFTSV infection induced complete autophagic flux and facilitated viral proliferation. We also identified a key mechanism underlying NSs-induced autophagy, in which NSs interacted with vimentin to inhibit the formation of the Beclin1-vimentin complex and induced vimentin degradation through K48-linked ubiquitination modification. These findings may help us understand the new functions and mechanisms of NSs and may aid in the identification of new antiviral targets.

Development and evaluation of a mouse model susceptible to severe fever with thrombocytopenia syndrome virus by rAAV-based exogenous human DC-SIGN expression.

Experimental animal model is indispensable to evaluate the prophylactic and therapeutic candidates against severe fever with thrombocytopenia syndrome virus (SFTSV). To develop a suitable mouse model for SFTSV infection, we delivered human dendritic cell-specific ICAM-3-grabbing non-integrin (hDC-SIGN) by adeno-associated virus (AAV2) and validated its susceptibility for SFTSV infection. Western blot and RT-PCR assays confirmed the expression of hDC-SIGN in transduced cell lines and a significantly increased viral infectivity was observed in cells expressing hDC-SIGN. The C57BL/6 mice transduced with AAV2 exhibited a stable hDC-SIGN expression in the organs for 7 days. Upon SFTSV challenge with 1 × 105 FAID50, the mice transduced with rAAV-hDC-SIGN showed a 12.5% mortality and reduced platelet and white blood cell count in accordance with higher viral titer than control group. Liver and spleen samples collected from the transduced mice had pathological signs similar to the IFNAR-/- mice with severe SFTSV infection. Collectively, the rAAV-hDC-SIGN transduced mouse model can be used as an accessible and promising tool for studying the SFTSV pathogenesis and pre-clinical evaluation of vaccines and therapeutics against the SFTSV infection.

Development of functional anti-Gn nanobodies specific for SFTSV based on next-generation sequencing and proteomics.

Severe fever with thrombocytopenia syndrome (SFTS) is an acute infectious disease caused by novel bunyavirus (SFTSV), with a mortality rate of 6.3% ~ 30%. To date, there is no specific treatment for SFTS. Previously, we demonstrated that SFTSV surface glycoprotein (Glycoprotein N, Gn) was a potential target for the development of SFTS vaccine or therapeutic antibodies, and anti-Gn neutralizing antibodies played a protective role in SFTS infection. Compared with traditional antibodies, nanobodies from camelids have various advantages, including small molecular weight, high affinity, low immunogenicity, convenient production by gene engineering, etc. In this study, we combined next-generation sequencing (NGS) with proteomics technology based on affinity purification-mass spectrometry (AP-MS) and bioinformatics analysis to high-throughput screen monoclonal anti-Gn nanobodies from camel immunized with Gn protein. We identified 19 anti-Gn monoclonal nanobody sequences, of which six sequences were selected for recombinant protein expression and purification. Among these six anti-Gn nanobodies, nanobody 57,493 was validated to be highly specific for Gn. The innovative high-throughput technical route developed in this study could also be expanded to the production of nanobodies specific for other viruses like SARS-CoV-2.

In silico study on miRNA regulation and NSs protein interactome characterization of the SFTS virus.

Severe fever with thrombocytopenia syndrome causing virus i.e. SFTS virus has increased in the last few years. The underlying cause and mechanism of disease progression and development of symptoms is not well known. Many viruses including Hepatitis B, Hepatitis C, HIV-1, Herpes virus, Dengue virus and many others have been seen to regulate their functions at the miRNA level. This study aimed to find out those cellular miRNAs, which can be mimicked or antagonized by the viral genome and analyze the effect of these miRNAs on various gene functions. Investigations in this study suggest a correlation between miRNA regulation with the disease symptoms and progression. By exhaustive literature survey we have tried to identify the interacting partners of the Non Structural S (NSs) protein and characterized the protein-protein interactions. The binding interface that can serve as target for therapeutic studies involving the interfacial residues was analyzed. This study would serve as an avenue to design therapeutics making use of not only protein-protein interactions but also miRNA based regulation as well.

A Viral NOD to Encephalitis.

In this issue, Gao and colleagues (J Virol 96:e00167-22, https://doi.org/10.1128/JVI.00167-22) dissect innate immune signaling in a microglial cell line infected with severe fever with thrombocytopenia syndrome virus (SFTSV). This virus has been designated a priority pathogen by the World Health Organization due to its capacity to induce a fatal cytokine storm. The study's findings attribute the pathogenesis to induction of the host inflammasome response by the SFTSV nonstructural protein.

Nonstructural Protein NSs Activates Inflammasome and Pyroptosis through Interaction with NLRP3 in Human Microglial Cells Infected with Severe Fever with Thrombocytopenia Syndrome Bandavirus.

Severe fever with thrombocytopenia syndrome (SFTS) is a tick-borne febrile disease caused by SFTS virus (SFTSV), or Dabie bandavirus, in the Phenuiviridae family. Clinically neurological disorders in SFTS have been commonly reported, but their neuropathogenesis has rarely been studied. Microglia are a type of neuroglia accounting for 10 to 12% of all cells in the brain. As resident immune cells, microglial cells are the first line of immune defense present in the central nervous system (CNS). Here, we report that SFTSV was able to infect microglial cells and stimulate interleukin 1ß (IL-1ß) secretion in the brains of infected neonatal BALB/c mice. We characterized the cell death induced in infected human microglial HMC3 cells, also susceptible to SFTSV, and found that the NOD-like receptor protein 3 (NLRP3) inflammasome was activated, leading to secretion of IL-1ß and pyroptosis. Knockdown of NLRP3 or inhibition of the NLRP3 inflammasome activation suppressed the viral replication, suggesting that the activation of the NLRP3 inflammasome may support SFTSV replication in microglial cells. Viral nonstructural protein NSs, a known modulator of immune responses, interacted and colocalized with NLRP3 for the inflammasome activation. It appeared that the N-terminal fragment, amino acids 1 to 66, of NSs was critical to promote the assembly of the inflammasome complex by interacting with NLRP3 for its activation in microglial cells. Our findings provide evidence that SFTSV may cause neurological disorders through infecting microglia and activating the inflammasome through its nonstructural protein NSs for neural cell death and inflammation. This study may have revealed a novel mechanism of SFTSV NSs in dysregulating host response. IMPORTANCE Encephalitis or encephalopathy during severe fever with thrombocytopenia syndrome (SFTS) is considered a critical risk factor leading to high mortality, but there have been no studies to date on the pathogenesis of encephalitis or encephalopathy caused by SFTS virus. Here, we report that SFTSV infection can active the NLRP3 inflammasome and induce IL-1ß secretion in the brains of infected newborn mice. In infected human HMC3 microglia, SFTSV activated the NLRP3 inflammasome via the viral nonstructural protein NSs through interaction with its N-terminal fragment. Notably, our findings suggest that the activation of the NLRP3 inflammasome may promote SFTSV replication in infected microglial cells. This study may reveal a novel mechanism by SFTSV to dysregulate host responses through its nonstructural protein, which could help us understand viral neuropathogenesis in SFTS patients.

A bioactive phlebovirus-like envelope protein in a hookworm endogenous virus.

Endogenous viral elements (EVEs), accounting for 15% of our genome, serve as a genetic reservoir from which new genes can emerge. Nematode EVEs are particularly diverse and informative of virus evolution. We identify Atlas virus-an intact retrovirus-like EVE in the human hookworm Ancylostoma ceylanicum, with an envelope protein genetically related to GN-GC glycoproteins from the family Phenuiviridae. A cryo-EM structure of Atlas GC reveals a class II viral membrane fusion protein fold not previously seen in retroviruses. Atlas GC has the structural hallmarks of an active fusogen. Atlas GC trimers insert into membranes with endosomal lipid compositions and low pH. When expressed on the plasma membrane, Atlas GC has cell-cell fusion activity. With its preserved biological activities, Atlas GC has the potential to acquire a cellular function. Our work reveals structural plasticity in reverse-transcribing RNA viruses.

The AST/ALT Ratio (De Ritis Ratio) Represents an Unfavorable Prognosis in Patients in Early-Stage SFTS: An Observational Cohort Study.

BACKGROUND: Severe fever with thrombocytopenia syndrome (SFTS), a widely prevalent infectious disease caused by severe fever with thrombocytopenia syndrome virus (SFTSV) that carries with it a high mortality rate, has emerged to be a public health concern. This study aimed to investigate the epidemiological and clinical characteristics of patients infected with SFTSV, seeking novel prognostic risk factors for SFTS. METHODS: In this retrospective and cross-sectional study, confirmed SFTS patients from the First Affiliated Hospital of Anhui Medical University were enrolled from September 1, 2019, to December 12, 2020. Cases were analyzed for epidemiological, demographic, clinical, and laboratory data. Logistic regression models were used to assess the association between predictors and outcome variables. A generalized additive mixed model (GAMM) was conducted to analyze the trending shift of aspartate aminotransferase/alanine transaminase-ratio (AST/ALT-ratio) and platelet (PLT) in SFTS patients treated with ribavirin. p values ≤ 0.05 were considered statistically significant. RESULTS: Clinical and laboratory results of 107 hospitalized patients with SFTSV infection were retrospectively described. The mean age at onset of disease was 60.38 ± 11.29 years old and the ratio between male and female was 1:1.2. Fever and thrombocytopenia are hallmark features of SFTS. Furthermore, multiple cases also experienced neurological complications, gastrointestinal/skeletal muscle symptoms together with other non-specific clinical manifestations; laboratory dataset outcomes reported dysregulated levels for routine blood biomarkers, coagulation function, and biochemistry. Overall, 107 patients were segregated into two groups according to patient condition at the clinical endpoint (survivors/non-survivors). SFTS survivors had a higher level of PLT- counts, total protein (TP), and estimated glomerular filtration rate (eGFR), while levels of activated partial thromboplastin time (APTT), thrombin time (TT), D-dimer (D-D), fibrinogen degradation products (FDP), ALT, AST, AST/ALT-ratio, creatinine (Cr), creatine phosphokinase (CK) and procalcitonin (PCT) was higher in non-survivors. Results from univariate Cox regression revealed that elevated levels of FDP, TT, AST/ALT-ratio, PCT, as well as decreased eGFR level and presence of central nervous system symptoms (CNS), were significant predictors for SFTS prognostic, results from multivariate logistic regression analysis in three adjusted models showed AST/ALT-ratio and PCT were independent risk factors for the prognosis of SFTS patients. Kaplan-Meier survival analysis showed that SFTS patients with AST/ALT-ratio >2.683 were associated with a shorter futime (means survival time), therefore indicating an unfavorable prognosis. Treatment with ribavirin could increase PLT count while decreasing AST/ALT-ratio within SFTS patients. CONCLUSION: SFTS is an emerging infectious disease, possibly leading to multiple-organ injury; AST/ALT-ratio was an independent risk factor for the prognosis of SFTS patients. Further investigation should be performed in order to gain more knowledge on this disease and guide clinical management.

Bunyavirus SFTSV exploits autophagic flux for viral assembly and egress.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is an emerging negatively stranded enveloped RNA bunyavirus that causes SFTS with a high case fatality rate of up to 30%. Macroautophagy/autophagy is an evolutionarily conserved process involved in the maintenance of host homeostasis, which exhibits anti-viral or pro-viral responses in reaction to different viral challenges. However, the interaction between the bunyavirus SFTSV and the autophagic process is still largely unclear. By establishing various autophagy-deficient cell lines, we found that SFTSV triggered RB1CC1/FIP200-BECN1-ATG5-dependent classical autophagy flux. SFTSV nucleoprotein induced BECN1-dependent autophagy by disrupting the BECN1-BCL2 association. Importantly, SFTSV utilized autophagy for the viral life cycle, which not only assembled in autophagosomes derived from the ERGIC and Golgi complex, but also utilized autophagic vesicles for exocytosis. Taken together, our results suggest a novel virus-autophagy interaction model in which bunyavirus SFTSV induces classical autophagy flux for viral assembly and egress processes, suggesting that autophagy inhibition may be a novel therapy for treating or releasing SFTS.

Comparison of RT-PCR, RT-nested PCRs, and real-time PCR for diagnosis of severe fever with thrombocytopenia syndrome: a prospective study.

We designed a highly sensitive reverse transcription nested polymerase chain reaction targeting the M-segment (NPCR-M) of severe fever with thrombocytopenia syndrome (SFTS) virus. NPCR-M was performed in parallel with three other referenced PCR assays QPCR-S, PCR-M, and NPCR-S to assess their clinical usefulness as routine diagnostic techniques for SFTS. In this multi-centered prospective study, 122 blood samples from 38 laboratory-confirmed SFTS patients and 85 control samples were used. The results demonstrated that QPCR-S and NPCR-S had better sensitivity rate up to 21 days after symptom onset however, the PCR-M showed poor sensitivity after 7 days of symptom onset. Our designed NPCR-M had a higher detection rate up to 40 days from symptom onset and revealed the persistence of SFTSV RNA in the early convalescent phase. No false-positive results were seen for the control samples. Additionally, NPCR-M showed positive results for a sample that initially showed negative results from other PCRs and for many other samples collected in the convalescent phase of SFTS. Our designed nested PCR is suitable for SFTSV detection in patients' blood collected in the acute and early convalescent phase of SFTS, and shows better sensitivity and high specificity even up to 40 days after symptom onset.

Nonstructural Protein NSs Hampers Cellular Antiviral Response through LSm14A during Severe Fever with Thrombocytopenia Syndrome Virus Infection.

The nonstructural protein (NSs) of severe fever with thrombocytopenia syndrome virus (SFTSV) plays multiple functions in the virus life cycle. Proteomic screening for host proteins interacting with NSs identified the cellular protein LSm14A. LSm14A, a member of the LSm family involved in RNA processing in the processing bodies, binds to viral RNA or synthetic homolog and mediates IFN regulatory factor 3 activation and IFN-ß induction. NSs interacted with and colocalized with LSm14A, and this interaction effectively inhibited downstream phosphorylation and dimerization of IFN regulatory factor 3, resulting in the suppression of antiviral signaling and IFN induction in several cell types of human origin. Knockdown of NSs resulted in the suppression of SFTSV replication in host cells. Viral RNA bound to LSm14A-NSs protein complex during the interaction. A newly discovered LRRD motif of NSs functioned to interact with LSm14A. Altogether, our data demonstrated a mechanism used by SFTSV to inhibit host innate immune response.

Clinical effect and antiviral mechanism of T-705 in treating severe fever with thrombocytopenia syndrome.

Severe fever with thrombocytopenia syndrome (SFTS) virus (SFTSV) is an emerging tick-borne virus with high fatality and an expanding endemic. Currently, effective anti-SFTSV intervention remains unavailable. Favipiravir (T-705) was recently reported to show in vitro and in animal model antiviral efficacy against SFTSV. Here, we conducted a single-blind, randomized controlled trial to assess the efficacy and safety of T-705 in treating SFTS (Chinese Clinical Trial Registry website, number ChiCTR1900023350). From May to August 2018, laboratory-confirmed SFTS patients were recruited from a designated hospital and randomly assigned to receive oral T-705 in combination with supportive care or supportive care only. Fatal outcome occurred in 9.5% (7/74) of T-705 treated patients and 18.3% (13/71) of controls (odds ratio, 0.466, 95% CI, 0.174-1.247). Cox regression showed a significant reduction in case fatality rate (CFR) with an adjusted hazard ratio of 0.366 (95% CI, 0.142-0.944). Among the low-viral load subgroup (RT-PCR cycle threshold ≥26), T-705 treatment significantly reduced CFR from 11.5 to 1.6% (P = 0.029), while no between-arm difference was observed in the high-viral load subgroup (RT-PCR cycle threshold <26). The T-705-treated group showed shorter viral clearance, lower incidence of hemorrhagic signs, and faster recovery of laboratory abnormities compared with the controls. The in vitro and animal experiments demonstrated that the antiviral efficacies of T-705 were proportionally induced by SFTSV mutation rates, particularly from two transition mutation types. The mutation analyses on T-705-treated serum samples disclosed a partially consistent mutagenesis pattern as those of the in vitro or animal experiments in reducing the SFTSV viral loads, further supporting the anti-SFTSV effect of T-705, especially for the low-viral loads.

Diagnostic system for the detection of severe fever with thrombocytopenia syndrome virus RNA from suspected infected animals.

BACKGROUND: Severe fever with thrombocytopenia syndrome virus (SFTSV) causes severe hemorrhagic fever in humans and cats. Clinical symptoms of SFTS-infected cats resemble those of SFTS patients, whereas SFTS-contracted cats have high levels of viral RNA loads in the serum and body fluids. Due to the risk of direct infection from SFTS-infected cats to human, it is important to diagnose SFTS-suspected animals. In this study, a reverse transcription polymerase chain reaction (RT-PCR) was newly developed to diagnose SFTS-suspected animals without non-specific reactions. METHODOLOGY/PRINCIPLE FINDINGS: Four primer sets were newly designed from consensus sequences constructed from 108 strains of SFTSV. A RT-PCR with these four primer sets successfully and specifically detected four clades of SFTSV. Their limits of detection are 1-10 copies/reaction. Using this RT-PCR, 5 cat cases among 56 SFTS-suspected animal cases were diagnosed as SFTS. From these cats, IgM or IgG against SFTSV were detected by enzyme-linked immunosorbent assay (ELISA), but not neutralizing antibodies by plaque reduction neutralization titer (PRNT) test. This phenomenon is similar to those of fatal SFTS patients. CONCLUSION/SIGNIFICANCE: This newly developed RT-PCR could detect SFTSV RNA of several clades and from SFTS-suspected animals. In addition to ELISA and PRNT test, the useful laboratory diagnosis systems of SFTS-suspected animals has been made in this study.

Structure of severe fever with thrombocytopenia syndrome virus L protein elucidates the mechanisms of viral transcription initiation.

Segmented negative-sense RNA viruses (sNSRVs) encode a single-polypeptide polymerase (L protein) or a heterotrimeric polymerase complex to cannibalize host messenger RNA cap structures serving as primers of transcription, and catalyse RNA synthesis. Here, we report the full-length structure of the severe fever with thrombocytopaenia syndrome virus (SFTSV) L protein, as determined by cryogenic electron microscopy at 3.4 Å, leading to an atomic model harbouring three functional parts (an endonuclease, an RNA-dependent RNA polymerase and a cap-binding domain) and two structural domains (an arm domain with a blocker motif and a carboxy-terminal lariat domain). The SFTSV L protein has a compact architecture in which its cap-binding pocket is surprisingly occupied by an Arg finger of the blocker motif, and the endonuclease active centre faces back towards the cap-binding pocket, suggesting that domain rearrangements are necessary to acquire the pre-initiation state of the active site. Our results provide insight into the complete architecture of sNSRV-encoded L protein and further the understanding of sNSRV transcription initiation.

Zwiesel bat banyangvirus, a potentially zoonotic Huaiyangshan banyangvirus (Formerly known as SFTS)-like banyangvirus in Northern bats from Germany.

Bats are reservoir hosts for several emerging and re-emerging viral pathogens causing morbidity and mortality in wildlife, animal stocks and humans. Various viruses within the family Phenuiviridae have been detected in bats, including the highly pathogenic Rift Valley fever virus and Malsoor virus, a novel Banyangvirus with close genetic relation to Huaiyangshan banyangvirus (BHAV)(former known as Severe fever with thrombocytopenia syndrome virus, SFTSV) and Heartland virus (HRTV), both of which have caused severe disease with fatal casualties in humans. In this study we present the whole genome of a novel Banyangvirus, named Zwiesel bat banyangvirus, revealed through deep sequencing of the Eptesicus nilssonii bat virome. The detection of the novel bat banyangvirus, which is in close phylogenetic relationship with the pathogenic HRTV and BHAV, underlines the possible impact of emerging phenuiviruses on public health.