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Although classical swine fever occurred in September 2018 for the first time in 26 years, its virulence is thought to be moderate based on field observations by veterinary authorities and our previous experimental infections. We quantified viremia and viral shedding in pigs infected with recent Japanese classical swine fever virus isolates, as well as a highly virulent strain. The results show that pigs infected with the Japanese strains exhibited lower viremia and viral shedding than those infected with the highly virulent strain. However, horizontal transmission occurred in pigs infected with the Japanese strains, similar to those infected with the highly virulent strain. Additionally, viremia and neuralization antibodies coexisted in pigs infected with the Japanese strains, presenting challenges for control measures. [ABSTRACT FROM AUTHOR]

After 26 years, another classical swine fever virus (CSFV) outbreak in domestic pigs and wild boars occurred in Japan 2018. Herein, we investigated the entry and the spatial dynamics of the CSFV outbreak in Japan using the nearly complete genomes of strains isolated from both wild boars and domestic pigs during this epidemic. Phylogenetic analysis showed that the most recent common ancestor (MRCA) of the Japanese lineage emerged 146 days (95% highest posterior density (HPD): 85-216 days) before the index case was detected. Based on epidemiological analysis, the period for the 95% HPD was 1 month earlier than the time of virus introduction into the index farm. The disease mainly spreads to the adjoining regions during the epidemic, with no spread to the nonadjacent regions. This result indicates that human activities, such as the movement of vehicles, contributed to the infection spread. As cases occurred in nonadjacent regions, the MRCA for the epidemic in the Saitama prefecture was estimated to have emerged 93 days before the date of detection in the initial farm in this region. Similarly, the MRCA for the epidemic in Okinawa prefecture, more than 1,300 km away from the other infected regions, was estimated to have emerged 34 days before the date of detection in the region's primary farm. Therefore, our results indicate that if exotic diseases emerge after a long period of absence or in a disease-free country, a longer period of time will elapse before detection, resulting in further spread. Additionally, subsequent infections occurring in regions distant from the original infected region will require less time for detection than in the original region. This study provides valuable insights into a CSFV outbreak that occurred in a previously CSFV-free country and thus beneficial in enhancing producers' awareness and allow for better preparation for infections. [ABSTRACT FROM AUTHOR]

Classical swine fever (CSF) is a worldwide devastating disease of the pig industry caused by classical swine fever virus (CSFV). In September 2018, an outbreak of CSF occurred in Japan where the disease had been eradicated and was officially designated a CSF-free country since 2015. Following the detection of the first 2018 case on a farm in Gifu Prefecture, the disease spread among both farm pigs and wild boars and still continues. Epigenome analysis using whole-genome information is helpful in identifying the infection route, but the current approaches provide an insufficient resolution. In this study, a novel method of using single-nucleotide variants (SNVs) was employed to identify the associations among 158 isolates (65 from farms and 93 from wild boars). The identified groups of CSFV strains were plotted in different colours on a map, identifying the location where each strain was collected. The lack of an SNV set shared between the index case and the other strains suggested the first infection in Japan during the outbreak occurred in wild boars, not at the index farm. For the Atsumi Peninsula outbreaks, where nine farms were found infected within a 10-km radius area, the farm strains were assembled into three groups, suggesting these outbreaks resulted from at least three different infection events in this area. For the infections in the area around Saitama Prefecture, an area remote from the epicentre, strains from both the farms and wild boars were identified as being in the same group, suggesting they resulted from one viral introduction. Likewise, seven infected farms in Okinawa Prefecture, almost 1,500 km from Gifu Prefecture, were identified as being in a common, but separate group. By demonstrating the variety of transmission routes and possibility of long-distance infection, these results will help improve disease control measures. [ABSTRACT FROM AUTHOR]

Classical swine fever (CSF) re-emerged in Japan in 2018 for the first time in 26 years. The disease has been known to be caused by a moderately pathogenic virus, rather than the highly pathogenic virus that had occurred in the past. However, the underlying pathophysiology remains unknown. This study conducted an experimental challenge on specific pathogen-free (SPF) pigs in a naïve state for 2, 4, and 6 weeks and confirmed the disease state during each period by clinical observation, virus detection, and pathological necropsy. We revealed the pathological changes and distribution of pathogens and virus-specific antibodies at each period after virus challenge. These results were comprehensively analyzed and approximately 70% of the pigs recovered, especially at 4- and 6-week post-virus challenge. This study provides useful information for future countermeasures against CSF by clarifying the pathogenicity outcomes in unvaccinated pigs with moderately pathogenic genotype 2.1 virus., (© 2024 The Author(s). Microbiology and Immunology published by The Societies and John Wiley & Sons Australia, Ltd.)

Classical swine fever virus (CSFV) is a member of the genus Pestivirus, which causes serious economic losses. The re-emergence of the disease in Japan in 2018 has increased awareness of CSFV. In this study, Balb/c mice were immunized with plant-derived E2 protein, and four monoclonal antibodies (mAbs) 4B11, 7B3, 11A5 and 6F3 were generated. Two of these mAbs, 4B11 and 7B3, effectively blocked CSFV infection of PK-15 cells. Both mAbs recognized a novel linear epitope, 256CLIGNTTVKVHASDER271. The neutralizing ability of anti-CSFV serum decreased 63%, when pre-incubated with the linear peptide at 200 μg/mL. Structural analysis showed that this linear epitope is present at the border of Domain C and Domain D on the surface of the E2 protein. Alignment of amino acid sequences showed that the epitope was conserved in different subgroups of CSFV but not in other members of the Pestivirus genus. Consistently with the analysis above, this epitope distinguished antibodies against CSFV from those against bovine viral diarrhea virus (BVDV). Our study provides an ideal candidate peptide for new vaccine design and differential diagnosis of CSFV. These findings will contribute to the control and eradication of classical swine fever. • Four mAbs against CSFV E2 were generated. Two of them presented neutralizing reactivity against CSFV. • A novel linear neutralizing epitope P33 located at the border of Domain C and Domain D of the E2 protein was identified. • Epitope P33 distinguished antibodies to CSFV from those to BVDV. [ABSTRACT FROM AUTHOR]

We evaluated the role of classical swine fever virus (CSFV) in the formation of button ulcers in the mucosa of the gastrointestinal tract. Histopathological and immunohistochemical analyses of pigs experimentally infected with a subgenotype 2.1 isolate of CSFV, which was isolated in Japan in 2019, revealed follicular necrosis in the submucosal mucosa-associated lymphoid tissue and herniation of crypts as factors that contribute to the development of button ulcers during CSFV infection. These findings indicate that CSFV induces follicular necrosis and is one of the causative agents of button ulcers in pigs. [ABSTRACT FROM AUTHOR]

There is a possibility that classical swine fever (CSF) virus outbreak has negative impacts on wild boar. To adequately manage native wild boar populations, wildlife managers need to gather the field data on wild boar and implement population management practices. We aimed to report the relative abundance index of wild boar before and after this outbreak. Our results showed that relative abundance index declined from 2017 (8.88 wild boars/100 trap days) to 2019 (2.03 wild boars/100 trap days), because of the negative impact of this virus and continuous culling programs. Although the eradication risk from the synergistic effect is low, wildlife managers need to consider the relationship between the trade-off between the risk of CSF and the conservation ecology risk of native species eradication. [ABSTRACT FROM AUTHOR]

Following an outbreak of classical swine fever (CSF) in Japan, 2018, CSFV JPN/1/2018 was isolated from an infected pig sample. In this study, we carried out a comparative experimental infection in pigs using this strain and the highly virulent ALD strain and compared outcomes, including clinical manifestation, virus shedding patterns and antibody responses. Although pigs inoculated orally or intramuscularly with JPN/1/2018 developed hyperthermia and had decreased leucocyte numbers, they survived for the whole experimental period and showed less severe clinical signs than those infected with the ALD strain. We confirmed the presence of characteristic multifocal infarction of the margin of the spleen that arises following infection with JPN/1/2018, albeit that this finding was not observed in all infected pigs. Both viruses efficiently spread to contact pigs in a similar manner, suggesting in transmissibility between the two strains. Viral RNAs were detected in all clinical samples, especially whole blood samples, before the pigs developed hyperthermia until at least approximately 2 weeks after inoculation. Our findings will be valuable for the investigations into epidemic events occurring in Japan and for establishing diagnostic strategies and control measures against CSF. [ABSTRACT FROM AUTHOR]

In September 2018, classical swine fever reemerged in Japan after 26 years, affecting domestic pigs and wild boars. The causative virus belongs to the 2.1 subgenotype, which caused repeated outbreaks in eastern and Southeast Asia. Intensive surveillance of swine and vaccination of wild boars will help control and eradicate this disease in Japan. [ABSTRACT FROM AUTHOR]

Classical swine fever virus (CSFV) causes a highly contagious disease among swine that has an important economic impact worldwide. CSFV strain LOM is an attenuated virus of low virulent strain of Miyagi isolated from Japan in 1956. Eight DNA fragments representing the genome of the CSFV strain LOM were obtained by RT-PCR. These were used to determine the complete nucleotide sequence and construct a full-length cDNA clone which was called Flc-LOM. Sequence analysis of the recombinant clone (Flc-LOM) revealed the presence of eight mutations, resulting in two amino acid substitutions, when compared to the parental sequence. RNA transcripts of both LOM and Flc-LOM were directly infectious in PK-15 cells. The rescued Flc-LOM virus grew more slowly than the parental virus, LOM, in the cells. Intramuscular immunization with Flc-LOM was safe and highly immunogenic in pigs; no clinical signs or virus transmission to sentinel animals were observed after 35 days. CSFV-specific neutralizing antibodies were detected 14 days post-infection. After challenge with the virulent CSFV strain SW03, pigs immunized with Flc-LOM were shown to be fully protected. Thus, our newly established infectious clone of CSFV, Flc-LOM, could serve as a vaccine candidate. [ABSTRACT FROM AUTHOR]

Classical swine fever (CSF) is a worldwide devastating disease of the pig industry caused by classical swine fever virus (CSFV). In September 2018, an outbreak of CSF occurred in Japan where the disease had been eradicated and was officially designated a CSF-free country since 2015. Following the detection of the first 2018 case on a farm in Gifu Prefecture, the disease spread among both farm pigs and wild boars and still continues. Epigenome analysis using whole-genome information is helpful in identifying the infection route, but the current approaches provide an insufficient resolution. In this study, a novel method of using single-nucleotide variants (SNVs) was employed to identify the associations among 158 isolates (65 from farms and 93 from wild boars). The identified groups of CSFV strains were plotted in different colours on a map, identifying the location where each strain was collected. The lack of an SNV set shared between the index case and the other strains suggested the first infection in Japan during the outbreak occurred in wild boars, not at the index farm. For the Atsumi Peninsula outbreaks, where nine farms were found infected within a 10-km radius area, the farm strains were assembled into three groups, suggesting these outbreaks resulted from at least three different infection events in this area. For the infections in the area around Saitama Prefecture, an area remote from the epicentre, strains from both the farms and wild boars were identified as being in the same group, suggesting they resulted from one viral introduction. Likewise, seven infected farms in Okinawa Prefecture, almost 1,500 km from Gifu Prefecture, were identified as being in a common, but separate group. By demonstrating the variety of transmission routes and possibility of long-distance infection, these results will help improve disease control measures., (© 2021 Wiley-VCH GmbH.)

The GPE− strain is a live attenuated vaccine for classical swine fever (CSF) developed in Japan. In the context of increasing attention for the differentiating infected from vaccinated animals (DIVA) concept, the achievement of CSF eradication with the GPE− proposes it as a preferable backbone for a recombinant CSF marker vaccine. While its infectious cDNA clone, vGPE−, is well characterized, 10 amino acid substitutions were recognized in the genome, compared to the original GPE− vaccine seed. To clarify the GPE− seed availability, this study aimed to generate and characterize a clone possessing the identical amino acid sequence to the GPE− seed. The attempt resulted in the loss of the infectious GPE− seed clone production due to the impaired replication by an amino acid substitution in the viral polymerase NS5B. Accordingly, replication-competent GPE− seed variant clones were produced. Although they were mostly restricted to propagate in the tonsils of pigs, similarly to vGPE−, their type I interferon-inducing capacity was significantly lower than that of vGPE−. Taken together, vGPE− mainly retains ideal properties for the CSF vaccine, compared with the seed variants, and is probably useful in the development of a CSF marker vaccine. [ABSTRACT FROM AUTHOR]

• The diversification of CSFV genome is limited to ensure viral protein functions. • SAVs inherited by at least two viruses were separately evaluated as shared SAVs. • Shared SAVs were not present in active or receptor domains on viral glycoproteins. • SNVs or SAVs in NS4A were not inherited by two or more viruses. • Stable structure of the NS3-4A complex was suggested to be indispensable for CSFV. Although RNA viruses exhibit extensive sequence diversity, the mutation rate must be limited to ensure protein functions that maintain the viral life cycle. Here, we compared the whole genome sequences of 150 isolates of classical swine fever virus (CSFV), obtained from a single epidemic that occurred in Japan during 2018–2020. After the detection of the first case, the disease spread among both farm pigs and wild boars and caused severe impact on the pig industry. To evaluate the diversification of the CSFV genome that eliminated mutations negatively affecting viral transmission, the substitution sets inherited by at least two isolates were separately evaluated as shared single nucleotide variants (SNVs) or shared single amino acid variants (SAVs). Comparisons of 12 protein-coding regions indicated that the percentages of SNVs and SAVs in the multifunctional nonstructural protein NS3 were the lowest, and shared SAVs were not detected in another nonstructural protein, NS4A. This demonstrated purifying negative selection suppressing changes in the protein sequences of NS3 and NS4A during virus transmission in the field. In contrast, a high possibility of nonsynonymous substitution among shared SNVs was detected only in genes encoding the secreted protein Erns and the nonstructural protein NS2, suggesting positive selection during the epidemic. Mapping of shared SAVs to the three-dimensional structure of Erns revealed that shared SAVs were not present in the substrate-binding sites but were instead localized to the peripheral region of the protein. These data will support efforts toward the development of diagnostic methods, recombinant vaccines, and antiviral agents targeting conserved and indispensable viral genes. [ABSTRACT FROM AUTHOR]

Classical swine fever virus (CSFV) in the wild boar population has been spreading in Japan, alongside outbreaks on pigs, since classical swine fever (CSF) reemerged in September 2018. The vaccination using oral bait vaccine was initially implemented in Gifu prefecture in March 2019. In the present study, antibodies against CSFV in wild boar were assessed in 1443 captured and dead wild boars in Gifu prefecture. After the implementation of oral vaccination, the increase of the proportion of seropositive animals and their titer in wild boars were confirmed. Quantitative analysis of antigen and antibodies against CSFV in wild boar implies potential disease diversity in the wild boar population. Animals with status in high virus replication (Ct < 30) and non- or low-immune response were confirmed and were sustained at a certain level after initial oral vaccination. Through continuous vaccination periods, the increase of seroprevalence among wild boar and the decrease of CSFV-positive animals were observed. The epidemiological analysis based on the quantitative virological outcomes could provide more information on the efficacy of oral vaccination and dynamics of CSF in the wild boar population, which will help to improve the implementation of control measures for CSF in countries such as Japan and neighboring countries. [ABSTRACT FROM AUTHOR]

We evaluated the role of classical swine fever virus (CSFV) in the formation of button ulcers in the mucosa of the gastrointestinal tract. Histopathological and immunohistochemical analyses of pigs experimentally infected with a subgenotype 2.1 isolate of CSFV, which was isolated in Japan in 2019, revealed follicular necrosis in the submucosal mucosa-associated lymphoid tissue and herniation of crypts as factors that contribute to the development of button ulcers during CSFV infection. These findings indicate that CSFV induces follicular necrosis and is one of the causative agents of button ulcers in pigs.

There is a possibility that classical swine fever (CSF) virus outbreak has negative impacts on wild boar. To adequately manage native wild boar populations, wildlife managers need to gather the field data on wild boar and implement population management practices. We aimed to report the relative abundance index of wild boar before and after this outbreak. Our results showed that relative abundance index declined from 2017 (8.88 wild boars/100 trap days) to 2019 (2.03 wild boars/100 trap days), because of the negative impact of this virus and continuous culling programs. Although the eradication risk from the synergistic effect is low, wildlife managers need to consider the relationship between the trade-off between the risk of CSF and the conservation ecology risk of native species eradication.

We analyzed the pathogenicity of a recent Japanese classical swine fever virus (CSFV) to wild boars via an experimental infection using boar-pig hybrids as an alternative to wild boars. We also investigated the effectiveness of a bait vaccine against the CSFV. Naïve boar-pig hybrids and pigs showed clinical signs such as fever, leucopenia, anorexia and conjunctivitis following the experimental infection. In contrast, the boar-pig hybrids administered the bait vaccine did not show any clinical signs. Our data indicated that boar-pig hybrids and domestic pigs have similar susceptibility to the recent Japanese CSFV. Additionally, the bait vaccine is effective against the CSFV.

Following an outbreak of classical swine fever (CSF) in Japan, 2018, CSFV JPN/1/2018 was isolated from an infected pig sample. In this study, we carried out a comparative experimental infection in pigs using this strain and the highly virulent ALD strain and compared outcomes, including clinical manifestation, virus shedding patterns and antibody responses. Although pigs inoculated orally or intramuscularly with JPN/1/2018 developed hyperthermia and had decreased leucocyte numbers, they survived for the whole experimental period and showed less severe clinical signs than those infected with the ALD strain. We confirmed the presence of characteristic multifocal infarction of the margin of the spleen that arises following infection with JPN/1/2018, albeit that this finding was not observed in all infected pigs. Both viruses efficiently spread to contact pigs in a similar manner, suggesting in transmissibility between the two strains. Viral RNAs were detected in all clinical samples, especially whole blood samples, before the pigs developed hyperthermia until at least approximately 2 weeks after inoculation. Our findings will be valuable for the investigations into epidemic events occurring in Japan and for establishing diagnostic strategies and control measures against CSF.