Construction and Characterization of Severe Fever with Thrombocytopenia Syndrome Virus with a Fluorescent Reporter for Antiviral Drug Screening.

Severe fever with thrombocytopenia syndrome (SFTS) caused by a novel bunyavirus (SFTSV) is an emerging infectious disease with up to 30% case fatality. Currently, there are no specific antiviral drugs or vaccines for SFTS. Here, we constructed a reporter SFTSV in which the virulent factor nonstructural protein (NSs) was replaced by eGFP for drug screening. First, we developed a reverse genetics system based on the SFTSV HBMC5 strain. Then, the reporter virus SFTSV-delNSs-eGFP was constructed, rescued, and characterized in vitro. SFTSV-delNSs-eGFP showed similar growth kinetics with the wild-type virus in Vero cells. We further detected the antiviral efficacy of favipiravir and chloroquine against wild-type and recombinant SFTSV by the quantification of viral RNA, and compared the results with that of fluorescent assay using high-content screening. The results showed that SFTSV-delNSs-eGFP could be used as a reporter virus for antiviral drug screening in vitro. In addition, we analyzed the pathogenesis of SFTSV-delNSs-eGFP in interferon receptor-deficient (IFNAR-/-) C57BL/6J mice and found that unlike the fatal infection of the wild-type virus, no obvious pathological change or viral replication were observed in SFTSV-delNSs-eGFP-infected mice. Taken together, the green fluorescence and attenuated pathogenicity make SFTSV-delNSs-eGFP a potent tool for the future high-throughput screening of antiviral drugs.

Accuracy of reverse-transcription polymerase chain reaction and loop-mediated isothermal amplification in diagnosing severe fever with thrombocytopenia syndrome: A systematic review and meta-analysis.

Nucleic acid molecular diagnostic technology plays an important role in the detection of severe fever with thrombocytopenia syndrome (SFTS). However, no relevant reports have been published on the accuracy of reverse-transcription polymerase chain reaction (RT-PCR) and reverse-transcription loop-mediated isothermal amplification (RT-LAMP) in the diagnosis of SFTS. Thus, we conducted a meta-analysis and systematic review to evaluate the accuracy of the two methods. On June 19, 2022, we comprehensively searched the PubMed, Embase, Cochrane Library, Web of Science, Scoups, Ovid, Proquest, China National Knowledge Infrastructure Database, Wan Fang Data, Traditional Chinese Medicine Database (Sinomed), VIP Database, and Reading Showing Database for articles on nucleic acid diagnostic techniques, such as RT-PCR and RT-LAMP, used to diagnose SFTS. Statistical analysis was performed using STATA 14.0 and Meta-Disc 1.4. Sixteen articles involving 2942 clinical blood samples were included in the analysis. RT-PCR and RT-LAMP were used as index tests, whereas RT-PCR or other detection methods were used as reference standards. The pooled values for the sensitivity, specificity, positive and negative likelihood ratios of the RT-PCR test were 0.97 (95% confidence interval [CI]: 0.92-0.99), 1.00 (95% CI: 0.98-1.00), 483.87 (95% CI: 58.04-4033.76), and 0.03 (95% CI:0.01-0.08), respectively. Those for the RT-LAMP test were 0.95 (95% CI: 0.91-0.97), 0.99 (95% CI: 0.93-1.00), 111.18 (95% CI: 13.96-885.27), and 0.05 (95% CI: 0.03-0.09), respectively. Both RT-PCR and RT-LAMP have high diagnostic value in SFTS and can be applied in different scenarios for laboratory confirmation or on-site screening.

The role of selenium in severe fever with thrombocytopenia syndrome: an integrative analysis of surveillance data and clinical data.

OBJECTIVES: Selenium deficiency can be associated with increased susceptibility to some viral infections and even more severe diseases. In this study, we aimed to examine whether this association applies to severe fever with thrombocytopenia syndrome (SFTS). METHOD: An observational study was conducted based on the data of 13,305 human SFTS cases reported in mainland China from 2010 to 2020. The associations among incidence, case fatality rate of SFTS, and crop selenium concentration at the county level were explored. The selenium level in a cohort of patients with SFTS was tested, and its relationship with clinical outcomes was evaluated. RESULTS: The association between selenium-deficient crops and the incidence rate of SFTS was confirmed by multivariate Poisson analysis, with an estimated incidence rate ratio (IRR, 95% confidence interval [CI]) of 4.549 (4.215-4.916) for moderate selenium-deficient counties and 16.002 (14.706-17.431) for severe selenium-deficient counties. In addition, a higher mortality rate was also observed in severe selenium-deficient counties with an IRR of 1.409 (95% CI: 1.061-1.909). A clinical study on 120 patients with SFTS showed an association between serum selenium deficiency and severe SFTS (odds ratio, OR: 2.94; 95% CI: 1.00-8.67) or fatal SFTS (OR: 7.55; 95% CI: 1.14-50.16). CONCLUSION: Selenium deficiency is associated with increased susceptibility to SFTS and poor clinical outcomes.

M Segment-Based Minigenome System of Severe Fever with Thrombocytopenia Syndrome Virus as a Tool for Antiviral Drug Screening.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is an emerging tick-borne bunyavirus that causes severe disease in humans with case fatality rates of approximately 30%. There are few treatment options for SFTSV infection. SFTSV RNA synthesis is conducted using a virus-encoded complex with RNA-dependent RNA polymerase activity that is required for viral propagation. This complex and its activities are, therefore, potential antiviral targets. A library of small molecule compounds was processed using a high-throughput screening (HTS) based on an SFTSV minigenome assay (MGA) in a 96-well microplate format to identify potential lead inhibitors of SFTSV RNA synthesis. The assay confirmed inhibitory activities of previously reported SFTSV inhibitors, favipiravir and ribavirin. A small-scale screening using MGA identified four candidate inhibitors that inhibited SFTSV minigenome activity by more than 80% while exhibiting less than 20% cell cytotoxicity with selectivity index (SI) values of more than 100. These included mycophenolate mofetil, methotrexate, clofarabine, and bleomycin. Overall, these data demonstrate that the SFTSV MGA is useful for anti-SFTSV drug development research.

Several catechins and flavonols from green tea inhibit severe fever with thrombocytopenia syndrome virus infection in vitro.

INTRODUCTION: Severe fever with thrombocytopenia syndrome (SFTS) is an emerging tick-borne hemorrhagic fever caused by SFTS virus (SFTSV). The mortality rate of SFTS is pretty high, but no vaccines and antiviral drugs are currently available. METHODS: The antiviral effects of six green tea-related polyphenols, including four catechins and two flavonols, on SFTSV were evaluated to identify natural antiviral compounds. RESULTS: Pretreatment with all polyphenols inhibited SFTSV infection in a concentration-dependent manner. The half-maximal inhibitory concentrations of (-)-epigallocatechin gallate (EGCg) and (-)-epigallocatechin (EGC) were 1.7-1.9 and 11-39 µM, respectively. The selectivity indices of EGCg and EGC were larger than those of the other polyphenols. Furthermore, pretreatment with EGCg and EGC dose-dependently decreased viral attachment to the host cells. Additionally, the treatment of infected cells with EGCg and EGC inhibited infection more significantly at a lower multiplicity of infection (MOI) than at a higher MOI, and this effect was less effective than that of pretreatment. Pyrogallol, a trihydroxybenzene that is the structural backbone of both EGCg and EGC, also inhibited SFTSV infection, as did gallic acid. CONCLUSIONS: Our study revealed that green tea-related polyphenols, especially EGCg and EGC, are useful as candidate anti-SFTSV drugs. Furthermore, the structural basis of their antiviral activity was identified, which should enable investigations of more active drugs in the future.