The increasing burden of tick-borne orthonairovirus infections, such as Crimean-Congo hemorrhagic fever, is becoming a global concern for public health. In the present study, we identify a novel orthonairovirus, designated Yezo virus (YEZV), from two patients showing acute febrile illness with thrombocytopenia and leukopenia after tick bite in Hokkaido, Japan, in 2019 and 2020, respectively. YEZV is phylogenetically grouped with Sulina virus detected in Ixodes ricinus ticks in Romania. YEZV infection has been confirmed in seven patients from 2014–2020, four of whom were co-infected with Borrelia spp. Antibodies to YEZV are found in wild deer and raccoons, and YEZV RNAs have been detected in ticks from Hokkaido. In this work, we demonstrate that YEZV is highly likely to be the causative pathogen of febrile illness, representing the first report of an endemic infection associated with an orthonairovirus potentially transmitted by ticks in Japan.

Classical swine fever virus (CSFV) is the causative agent of classical swine fever (CSF), a highly contagious disease of pigs. There are numerous CSFV strains that differ in virulence, resulting in clinical disease with different degrees of severity. Low-virulent and moderately virulent isolates cause a mild and often chronic disease, while highly virulent isolates cause an acute and mostly lethal hemorrhagic fever. The live attenuated vaccine strain GPE ⁻ was produced by multiple passages of the virulent ALD strain in cells of swine, bovine, and guinea pig origin. With the aim of identifying the determinants responsible for the attenuation, the GPE ⁻ vaccine virus was readapted to pigs by serial passages of infected tonsil homogenates until prolonged viremia and typical signs of CSF were observed. The GPE ⁻ /P-11 virus isolated from the tonsils after the 11th passage in vivo had acquired 3 amino acid substitutions in E2 (T830A) and NS4B (V2475A and A2563V) compared with the virus before passages. Experimental infection of pigs with the mutants reconstructed by reverse genetics confirmed that these amino acid substitutions were responsible for the acquisition of pathogenicity. Studies in vitro indicated that the substitution in E2 influenced virus spreading and that the changes in NS4B enhanced the viral RNA replication. In conclusion, the present study identified residues in E2 and NS4B of CSFV that can act synergistically to influence virus replication efficiency in vitro and pathogenicity in pigs.

Classical swine fever (CSF) caused by CSF virus (CSFV) is a highly contagious disease of pigs. The viral protein N^pro of CSFV interferes with alpha- and beta-interferon (IFN-α/β) induction by promoting the degradation of interferon regulatory factor 3 (IRF3). During the establishment of the live attenuated CSF vaccine strain GPE^-, N^pro acquired a mutation that abolished its capacity to bind and degrade IRF3, rendering it unable to prevent IFN-α/β induction. In a previous study, we showed that the GPE^- vaccine virus became pathogenic after forced serial passages in pigs, which was attributed to the amino acid substitutions T830A in the viral proteins E2 and V2475A and A2563V in NS4B. Interestingly, during the re-adaptation of the GPE^- vaccine virus in pigs, the IRF3-degrading function of N^pro was not recovered. Therefore, we examined whether restoring the ability of N^pro to block IFN-α/β induction of both the avirulent and moderately virulent GPE^--derived virus would enhance pathogenicity in pigs. Viruses carrying the N136D substitution in N^pro regained the ability to degrade IRF3 and suppress IFN-α/β induction in vitro. In pigs, functional N^pro significantly reduced the local IFN-α mRNA expression in lymphoid organs while it increased quantities of IFN-α/β in the circulation, and enhanced pathogenicity of the moderately virulent virus. In conclusion, the present study demonstrates that functional N^pro influences the innate immune response at local sites of virus replication in pigs and contributes to pathogenicity of CSFV in synergy with viral replication.

Since September 2018, nearly 900 notifications of classical swine fever (CSF) have been reported in Gifu Prefecture (Japan) affecting domestic pig and wild boar by the end of August 2019. To determine the epidemiological characteristics of its spread, a spatio-temporal analysis was performed using actual field data on the current epidemic. The spatial study, based on standard deviational ellipses of official CSF notifications, showed that the disease likely spread to the northeast part of the prefecture. A maximum significant spatial association estimated between CSF notifications was 23 km by the multi-distance spatial cluster analysis. A space-time permutation analysis identified two significant clusters with an approximate radius of 12 and 20 km and 124 and 98 days of duration, respectively. When the area of the identified clusters was overlaid on a map of habitat quality, approximately 82% and 75% of CSF notifications, respectively, were found in areas with potential contact between pigs and wild boar. The obtained results provide information on the current CSF epidemic, which is mainly driven by wild boar cases with sporadic outbreaks on domestic pig farms. These findings will help implement control measures in Gifu Prefecture.

Highly pathogenic avian influenza viruses (HPAIVs) A(H5N6) were concurrently introduced into several distant regions of Japan in November 2016. These viruses were classified into the genetic clade 2.3.4.4c and were genetically closely related to H5N6 HPAIVs recently isolated in South Korea and China. In addition, these HPAIVs showed further antigenic drift.