Akabane, Aino and Chuzan virus are arthropod-borne (arbo)viruses mainly associated with reproductive failures in cattle. We investigated apoptosis in Vero cells (C-1586) infected with Akabane, Aino and Chuzan virus. The fragmentation of chromosomal DNA was simultaneously detected with the progress of cytopathic effect from 48 hr to 72 hr post infection, depending on viruses. Although the treatment of cycloheximide blocked apoptosis in Vero cells infected with three viruses, actinomycin D did not prevent DNA oligomerization, thus indicating that de novo viral protein synthesis is critical for viral apoptosis. In addition, the activation of caspase-3 was also detected in Vero cells by indirect fluorescent assay. From the present results, it is of future interest whether apoptotic characteristics of these viruses are related to pathogenecity in vivo.
Animals ; Apoptosis/*physiology ; Bunyaviridae/*physiology ; Caspase 3 ; Caspases/metabolism ; Cercopithecus aethiops ; Cytopathogenic Effect, Viral/*physiology ; DNA Fragmentation/physiology ; Dactinomycin ; Enzyme Activation ; Orbivirus/*physiology ; Vero Cells

Vector-borne arboviruses produce mild to severe symptoms in domestic animals. Bovine ephemeral fever (BEF), Akabane, Aino, and Chuzan virus have been primarily attributed to reproductive disorders or febrile diseases in cattle, and Japanese encephalitis virus (JEV) is mainly associated with reproductive failures in swine. We investigated antibody titers from domestic swine against four bovine arboviruses (BEF, Akabane, Aino, and Chuzan virus) and from cattle against JEV in Korea. While the positive rates for Akabane and BEF were 37.4% and 15.7%, the positive incidence of Chuzan and Aino were relatively low, with positive rates of 3.04% and 0.4%, respectively, based on a virus neutralization assay. Antibody titers against more than one virus were also frequently detected in domestic swine. The incidence of JEV was 51.3% among domestic cattle. In addition, one positive case was detected in the thoracic fluids from 35 aborted calves, based on the hemagglutination inhibition test. Our results indicate that swine are susceptible hosts of bovine arboviruses without showing clinical symptoms in a natural environment. Moreover, we confirmed that JEV could be associated with reproductive failure in pregnant cattle, as were other vector-borne bovine arboviruses assessed in this study.
Animals ; Antibodies, Viral/blood ; Cattle ; Cattle Diseases/*epidemiology/*virology ; Encephalitis Virus, Japanese/*immunology ; Encephalitis, Japanese/blood/epidemiology/*veterinary/virology ; Ephemeral Fever/blood/*epidemiology/virology ; Ephemeral Fever Virus, Bovine/*immunology ; Hemagglutination Tests ; Incidence ; Korea/epidemiology ; Neutralization Tests ; Swine ; Swine Diseases/*epidemiology/*virology

Bovine leukemia virus (BLV) envelope glycoprotein (gp51/gp30T-), consisting of BLV gp51 and BLV gp30 that lacked its C-terminal transmembrane domain, was expressed in insect cells under the control of the baculovirus polyhedron promoter. Recombinant BLV gp51/gp30T- secreted from insect cells was determined by immunofluorescence, enzyme-linked immunosorbent and western blot assays using a BLV-specific monoclonal antibody and BLV-positive bovine antibodies. An agar gel immunodiffusion (AGID) test using gp51/gp30T- as the antigen for the detection of BLV antibodies in serum was developed and compared to traditional AGID, which uses wild type BLV antigen derived from fetal lamb kidney cells. AGID with the recombinant BLV gp51/gp30T- was relatively more sensitive than traditional AGID. When the two methods were tested with bovine sera from the field, the recombinant BLV gp51/gp30T- and traditional antigen had a relative sensitivity of 69.8% and 67.4%, respectively, and a relative specificity of 93.3% and 92.3%. These results indicated that the recombinant BLV gp51/gp30T- is an effective alternative antigen for the diagnosis of BLV infection in cattle.
Agar ; Animals ; Antibodies, Viral/blood ; Antigens, Viral/immunology ; Baculoviridae/*metabolism ; Cattle ; Cell Line ; Enzootic Bovine Leukosis/blood/immunology ; Gene Expression Regulation, Viral/*physiology ; Immunodiffusion/methods/*veterinary ; Kidney/cytology ; Leukemia Virus, Bovine/genetics/*metabolism ; Molecular Biology ; Sheep ; Viral Envelope Proteins/genetics/*metabolism

To select a less pathogenic bovine viral diarrhea virus (BVDV) strain for the construction of chimeric pestivirus harboring classical swine fever virus (CSFV) E2 gene, five Korean BVDV isolates (four type 1 isolates and a type 2 isolate) were evaluated for their pathological and biological properties with respect to porcine infection. Each of five groups of 100-day-old pigs was inoculated intranasally with one of the five BVDV isolates. No clinical sign or leukopenia was observed in any pig throughout the duration of the experiment, but viruses were detected in blood, nasal discharges and postmortem samples using RT-PCR. These results indicated that although the five BVD viruses could infect pigs, they did not cause clinically apparent symptoms. Because of its proper infection dynamics shown in this preliminary animal study and its fast growth rate and quick CPE in cell culture, one isolate (KD26-1) was chosen among the five isolates to test its virulence and immunogenic properties in 40-day-old piglets. Neither clinical sign nor pathological lesion was observed in 40-day-old piglets during the course of infection of isolate KD26-1. The first neutralizing antibodies were detectable 14 days post-inoculation (PI) and increased to 1:128~1:256 28 days PI. A BVDV specific gene was detectable by RT-PCR in tonsil, spleen, inguinal lymph node and brain until 14 days PI. According to this study, it can be concluded that isolate KD26-1 has little pathological effect in pigs and is a candidate for construction of chimeric pestivirus harboring CSFV E2 gene.
Animals ; Antibodies, Neutralizing ; Brain ; Cell Culture Techniques ; Classical swine fever virus ; Diarrhea ; Leukopenia ; Lymph Nodes ; Palatine Tonsil ; Pestivirus ; Spleen ; Sprains and Strains ; Swine ; Viruses

To select a less pathogenic bovine viral diarrhea virus (BVDV) strain for the construction of chimeric pestivirus harboring classical swine fever virus (CSFV) E2 gene, five Korean BVDV isolates (four type 1 isolates and a type 2 isolate) were evaluated for their pathological and biological properties with respect to porcine infection. Each of five groups of 100-day-old pigs was inoculated intranasally with one of the five BVDV isolates. No clinical sign or leukopenia was observed in any pig throughout the duration of the experiment, but viruses were detected in blood, nasal discharges and postmortem samples using RT-PCR. These results indicated that although the five BVD viruses could infect pigs, they did not cause clinically apparent symptoms. Because of its proper infection dynamics shown in this preliminary animal study and its fast growth rate and quick CPE in cell culture, one isolate (KD26-1) was chosen among the five isolates to test its virulence and immunogenic properties in 40-day-old piglets. Neither clinical sign nor pathological lesion was observed in 40-day-old piglets during the course of infection of isolate KD26-1. The first neutralizing antibodies were detectable 14 days post-inoculation (PI) and increased to 1:128~1:256 28 days PI. A BVDV specific gene was detectable by RT-PCR in tonsil, spleen, inguinal lymph node and brain until 14 days PI. According to this study, it can be concluded that isolate KD26-1 has little pathological effect in pigs and is a candidate for construction of chimeric pestivirus harboring CSFV E2 gene.
Animals ; Antibodies, Neutralizing ; Brain ; Cell Culture Techniques ; Classical swine fever virus ; Diarrhea ; Leukopenia ; Lymph Nodes ; Palatine Tonsil ; Pestivirus ; Spleen ; Sprains and Strains ; Swine ; Viruses

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging zoonotic disease, which causes high fever, thrombocytopenia, and death in humans and animals in East Asian countries. The pathogenicity of SFTS virus (SFTSV) remains unclear. We intraperitoneally infected three groups of mice: wild-type (WT), mice treated with blocking anti-type I interferon (IFN)-α receptor antibody (IFNAR Ab), and IFNAR knockout (IFNAR−/−) mice, with four doses of SFTSV (KH1, 5 × 105 to 5 × 102 FAID50). The WT mice survived all SFTSV infective doses. The IFNAR Ab mice died within 7 days post-infection (dpi) with all doses of SFTSV except that the mice were infected with 5 × 102 FAID50 SFTSV. The IFNAR−/− mice died after infection with all doses of SFTSV within four dpi. No SFTSV infection caused hyperthermia in any mice, whereas all the dead mice showed hypothermia and weight loss. In the WT mice, SFTSV RNA was detected in the eyes, oral swabs, urine, and feces at 5 dpi. Similar patterns were observed in the IFNAR Ab and IFNAR−/− mice after 3 dpi, but not in feces. The IFNAR Ab mice showed viral shedding until 7 dpi. The SFTSV RNA loads were higher in organs of the IFNAR−/− mice compared to the other groups. Histopathologically,coagulation necrosis and mononuclear inflammatory cell infiltration in the liver and white pulp atrophy in the spleen were seen as the main lesions in the IFN signaling lacking mice. Immunohistochemically, SFTSV antigens were mainly detected in the marginal zone of the white pulp of the spleen in all groups of mice, but more viral antigens were observed in the spleen of the IFNAR−/− mice. Collectively, the IFN signaling-deficient mice were highly susceptible to SFTSV and more viral burden could be demonstrated in various excreta and organs of the mice when IFN signaling was inhibited.

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging zoonotic disease, which causes high fever, thrombocytopenia, and death in humans and animals in East Asian countries. The pathogenicity of SFTS virus (SFTSV) remains unclear. We intraperitoneally infected three groups of mice: wild-type (WT), mice treated with blocking anti-type I interferon (IFN)-α receptor antibody (IFNAR Ab), and IFNAR knockout (IFNAR−/−) mice, with four doses of SFTSV (KH1, 5 × 105 to 5 × 102 FAID50). The WT mice survived all SFTSV infective doses. The IFNAR Ab mice died within 7 days post-infection (dpi) with all doses of SFTSV except that the mice were infected with 5 × 102 FAID50 SFTSV. The IFNAR−/− mice died after infection with all doses of SFTSV within four dpi. No SFTSV infection caused hyperthermia in any mice, whereas all the dead mice showed hypothermia and weight loss. In the WT mice, SFTSV RNA was detected in the eyes, oral swabs, urine, and feces at 5 dpi. Similar patterns were observed in the IFNAR Ab and IFNAR−/− mice after 3 dpi, but not in feces. The IFNAR Ab mice showed viral shedding until 7 dpi. The SFTSV RNA loads were higher in organs of the IFNAR−/− mice compared to the other groups. Histopathologically,coagulation necrosis and mononuclear inflammatory cell infiltration in the liver and white pulp atrophy in the spleen were seen as the main lesions in the IFN signaling lacking mice. Immunohistochemically, SFTSV antigens were mainly detected in the marginal zone of the white pulp of the spleen in all groups of mice, but more viral antigens were observed in the spleen of the IFNAR−/− mice. Collectively, the IFN signaling-deficient mice were highly susceptible to SFTSV and more viral burden could be demonstrated in various excreta and organs of the mice when IFN signaling was inhibited.