Background: SARS-CoV-2 is a novel coronavirus that emerged in 2019 and is now classified in the genus Coronavirus with closely related SARS-CoV. SARS-CoV-2 is highly pathogenic in humans and is classified as a biosafety level (BSL)-3 pathogen, which makes manipulating it relatively difficult due to its infectious nature. Methods: To circumvent the need for BSL-3 laboratories, an alternative assay was developed that avoids live virus and instead uses a recombinant VSV expressing luciferase and possesses the full length or truncated spike proteins of SARS-CoV-2. Furthermore, to measure SARS-CoV-2 neutralizing antibodies under BSL2 conditions, a chemiluminescence reduction neutralization test (CRNT) for SARS-CoV-2 was developed. The neutralization values of the serum samples collected from hospitalized patients with COVID-19 or SARS-CoV-2 PCR-negative donors against the pseudotyped virus infection evaluated by the CRNT were compared with antibody titers determined from an enzyme-linked immunosorbent assay (ELISA) or an immunofluorescence assay (IFA). Results: The CRNT, which used whole blood collected from hospitalized patients with COVID-19, was also examined. As a result, the inhibition of pseudotyped virus infection was specifically observed in both serum and whole blood and was also correlated with the results of the IFA. Conclusions: In conclusion, the CRNT for COVID-19 is a convenient assay system that can be performed in a BSL-2 laboratory with high specificity and sensitivity for evaluating the occurrence of neutralizing antibodies against SARS-CoV-2.

Severe fever with thrombocytopenia syndrome (SFTS), caused by SFTS virus (SFTSV), is a viral hemorrhagic fever with a high case fatality rate. Favipiravir was reported to be effective in the treatment of SFTSV infection in vivo in type I interferon receptor knockout (IFNAR^−/−) mice at treatment dosages of both 60 mg/kg/day and 300 mg/kg/day for a duration of 5 days. In this study, the efficacy of favipiravir at dosages of 120 mg/kg/day and 200 mg/kg/day against SFTSV infection in an IFNAR^−/− mouse infection model was investigated. IFNAR^−/− mice were subcutaneously infected with SFTSV at a 1.0 × 10⁶ 50% tissue culture infectious dose followed by twice daily administration of favipiravir, comprising a total dose of either 120 mg/kg/day or 200 mg/kg/day. The treatment was initiated either immediately post infection or at predesignated time points post infection. Neutralizing antibodies in the convalescent-phase mouse sera was examined by the pseudotyped VSV system. All mice treated with favipiravir at dosages of 120 mg/kg/day or 200 mg/kg/day survived when the treatment was initiated at no later than 4 days post infection. A decrease in body weight of mice was observed when the treatment was initiated at 3–4 days post infection. Furthermore, all control mice died. The body weight of mice did not decrease when treatment with favipiravir was initiated immediately post infection at dosages of 120 mg/kg/day and 200 mg/kg/day. Neutralizing antibodies were detected in the convalescent-phase mouse sera. Similar to the literature-reported peritoneal administration of favipiravir at 300 mg/kg/day, the oral administration of favipiravir at dosages of 120 mg/kg/day and 200 mg/kg/day to IFNAR^−/− mice infected with SFTSV was effective.

This study provides a diagnostic evidence of test validity, which can lead to vaccine efficacy and proof of recovery after COVID-19. It is not easy to know neutralization against SARS-CoV-2 in the clinical laboratory because of technical and biohazard issues.

Heartland bandavirus (HRTV) is an emerging tick-borne virus that is distributed in the United States and that causes febrile illness with thrombocytopenia and leukocytopenia. It is genetically close to Dabie bandavirus, which is well known as severe fever with thrombocytopenia syndrome (SFTS) virus (SFTSV). The mortality rate of human HRTV infection is approximately 10%; however, neither approved anti-HRTV agents nor vaccines exist. An appropriate animal model should be developed to evaluate the efficacy of antiviral agents and vaccines against HRTV. The susceptibility of IFNAR^−/− mice with HRTV infection was evaluated using subcutaneous, intraperitoneal, and retro-orbital inoculation routes. IFNAR^−/− mice intraperitoneally infected with HRTV showed the most severe clinical signs, and the 50% lethal dose was 3.2 × 10⁶ TCID₅₀. Furthermore, to evaluate the utility of a novel lethal IFNAR^−/− mice model, IFNAR^−/− mice were orally administered favipiravir, ribavirin, or a solvent for 5 days immediately after a lethal dose of HRTV inoculation. The survival rates of the favipiravir-, ribavirin-, and solvent-administered mice were 100, 33, and 0%, respectively. The changes in bodyweights and HRTV RNA loads in the blood of favipiravir-treated IFNAR^−/− mice were the lowest among the three groups, which suggests that favipiravir is a promising drug candidate for the treatment of patients with HRTV infection.

Since mRNA vaccines utilize wild-type SARS-CoV-2 spike protein as an antigen, there are potential concerns about acquiring immunity to variants of this virus. The neutralizing activity in BNT162b2-vaccinated individuals was higher against the wild-type virus than against its variants; this effect was more apparent in older age groups.

Severe fever with thrombocytopenia syndrome (SFTS) is an infectious disease with a high case fatality rate caused by the SFTS virus, and currently there are no approved specific treatments. Neutralizing monoclonal antibodies (mAbs) against the virus could be a therapeutic agent in SFTS treatment, but their development has not sufficiently been carried out. In the present study, mouse and human mAbs exposed to the viral envelope proteins Gn and Gc (16 clones each) were characterized in vitro and in vivo by using recombinant proteins, cell culture with viruses, and an SFTS animal model with IFNAR^-/- mice. Neutralization activities against the recombinant vesicular stomatitis virus bearing SFTS virus Gn/Gc as envelope proteins were observed with three anti-Gn and six anti-Gc mAbs. Therapeutic activities were observed among anti-Gn, but not anti-Gc mAbs with neutralizing activities. These results propose an effective strategy to obtain promising therapeutic mAb candidates for SFTS treatment, and a necessity to reveal precise roles of the SFTS virus Gn/Gc proteins.