Identification of novel orthonairoviruses from rodents and shrews in Gabon, Central Africa.

In Africa, several emerging zoonotic viruses have been transmitted from small mammals such as rodents and shrews to humans. Although no clinical cases of small mammal-borne viral diseases have been reported in Central Africa, potential zoonotic viruses have been identified in rodents in the region. Therefore, we hypothesized that there may be unrecognized zoonotic viruses circulating in small mammals in Central Africa. Here, we investigated viruses that have been maintained among wild small mammals in Gabon to understand their potential risks to humans. We identified novel orthonairoviruses in 24.6 % of captured rodents and shrews from their kidney total RNA samples. Phylogenetic analysis revealed that the novel viruses, Lamusara virus (LMSV) and Lamgora virus, were closely related to Erve virus, which was previously identified in shrews of the genus Crocidura and has been suspected to cause neuropathogenic diseases in humans. Moreover, we show that the LMSV ovarian tumour domain protease, one of the virulence determination factors of orthonairoviruses, suppressed interferon signalling in human cells, suggesting the possible human pathogenicity of this virus. Taken together, our study demonstrates the presence of novel orthonairoviruses that may pose unrecognized risks of viral disease transmission in Gabon.

5-amino levulinic acid inhibits SARS-CoV-2 infection in vitro.

The current COVID-19 pandemic requires urgent development of effective therapeutics. 5-amino levulinic acid (5-ALA) is a naturally synthesized amino acid and has been used for multiple purposes including as an anticancer therapy and as a dietary supplement due to its high bioavailability. In this study, we demonstrated that 5-ALA treatment potently inhibited infection of SARS-CoV-2, a causative agent of COVID-19, in cell culture. The antiviral effects could be detected in both human and non-human cells, without significant cytotoxicity. Therefore, 5-ALA is worth to be further investigated as an antiviral drug candidate for COVID-19.

Different effects of two mutations on the infectivity of Ebola virus glycoprotein in nine mammalian species.

Ebola virus (EBOV), which belongs to the genus Ebolavirus, causes a severe and often fatal infection in primates, including humans, whereas Reston virus (RESTV) only causes lethal disease in non-human primates. Two amino acids (aa) at positions 82 and 544 of the EBOV glycoprotein (GP) are involved in determining viral infectivity. However, it remains unclear how these two aa residues affect the infectivity of Ebolavirus species in various hosts. Here we performed viral pseudotyping experiments with EBOV and RESTV GP derivatives in 10 cell lines from 9 mammalian species. We demonstrated that isoleucine at position 544/545 increases viral infectivity in all host species, whereas valine at position 82/83 modulates viral infectivity, depending on the viral and host species. Structural modelling suggested that the former residue affects viral fusion, whereas the latter residue influences the interaction with the viral entry receptor, Niemann-Pick C1.

Functional mutations in spike glycoprotein of Zaire ebolavirus associated with an increase in infection efficiency.

Ebola virus (EBOV) is extremely virulent, and its glycoprotein is necessary for viral entry. EBOV may adapt to its new host humans during outbreaks by acquiring mutations especially in glycoprotein, which allows EBOV to spread more efficiently. To identify these evolutionary selected mutations and examine their effects on viral infectivity, we used experimental-phylogenetic-structural interdisciplinary approaches. In evolutionary analysis of all available Zaire ebolavirus glycoprotein sequences, we detected two codon sites under positive selection, which are located near/within the region critical for the host-viral membrane fusion, namely alanine-to-valine and threonine-to-isoleucine mutations at 82 (A82V) and 544 (T544I), respectively. The fine-scale transmission dynamics of EBOV Makona variants that caused the 2014-2015 outbreak showed that A82V mutant was fixed in the population, whereas T544I was not. Furthermore, pseudotype assays for the Makona glycoprotein showed that the A82V mutation caused a small increase in viral infectivity compared with the T544I mutation. These findings suggest that mutation fixation in EBOV glycoprotein may be associated with their increased infectivity levels; the mutant with a moderate increase in infectivity will fix. Our findings showed that a driving force for Ebola virus evolution via glycoprotein may be a balance between costs and benefits of its virulence.

A loop-mediated isothermal amplification assay for rapid and sensitive detection of bovine papular stomatitis virus.

Bovine papular stomatitis virus (BPSV) causes pustular cutaneous disease in cattle worldwide. This paper describes the development of a specific loop-mediated isothermal amplification (LAMP) assay to detect BPSV which did not cross-react with other parapoxviruses. To assess analytical sensitivity of this LAMP assay, DNA was extracted from serially diluted BPSV from which the infectious titer was determined by a novel assay based on calf kidney epithelial cells. The LAMP assay had equivalent analytical sensitivity to quantitative PCR, and could detect as few as 86 copies of viral DNA per reaction. These results suggest that the assay is a specific and sensitive technique to rapidly diagnose bovine papular stomatitis in domestic animals.

Tofla virus: A newly identified Nairovirus of the Crimean-Congo hemorrhagic fever group isolated from ticks in Japan.

Ixodid ticks transmit several important viral pathogens. We isolated a new virus (Tofla virus: TFLV) from Heamaphysalis flava and Heamaphysalis formsensis in Japan. The full-genome sequences revealed that TFLV belonged to the genus Nairovirus, family Bunyaviridae. Phylogenetic analyses and neutralization tests suggested that TFLV is closely related to the Hazara virus and that it is classified into the Crimean-Congo hemorrhagic fever group. TFLV caused lethal infection in IFNAR KO mice. The TFLV-infected mice exhibited a gastrointestinal disorder, and positron emission tomography-computed tomography images showed a significant uptake of (18)F-fluorodeoxyglucose in the intestinal tract. TFLV was able to infect and propagate in cultured cells of African green monkey-derived Vero E6 cells and human-derived SK-N-SH, T98-G and HEK-293 cells. Although TFLV infections in humans and animals are currently unknown, our findings may provide clues to understand the potential infectivity and to develop of pre-emptive countermeasures against this new tick-borne Nairovirus.

Enhanced replication of human T-cell leukemia virus type 1 in T cells from transgenic rats expressing human CRM1 that is regulated in a natural manner.

Human T-cell leukemia virus type 1 (HTLV-1) is the etiologic agent of adult T-cell leukemia (ATL). To develop a better animal model for the investigation of HTLV-1 infection, we established a transgenic (Tg) rat carrying the human CRM1 (hCRM1) gene, which encodes a viral RNA transporter that is a species-specific restriction factor. At first we found that CRM1 expression is elaborately regulated through a pathway involving protein kinase C during lymphocyte activation, initially by posttranscriptional and subsequently by transcriptional mechanisms. This fact led us to use an hCRM1-containing bacterial artificial chromosome clone, which would harbor the entire regulatory and coding regions of the CRM1 gene. The Tg rats expressed hCRM1 protein in a manner similar to expression of intrinsic rat CRM1 in various organs. HTLV-1-infected T-cell lines derived from these Tg rats produced 100- to 10,000-fold more HTLV-1 than did T cells from wild-type rats, and the absolute levels of HTLV-1 were similar to those produced by human T cells. We also observed enhancement of the dissemination of HTLV-1 to the thymus in the Tg rats after intraperitoneal inoculation, although the proviral loads were low in both wild-type and Tg rats. These results support the essential role of hCRM1 in proper HTLV-1 replication and suggest the importance of this Tg rat as an animal model for HTLV-1.