Keita Noguchi

Emergence of pathogenic coronaviruses in cats by homologous recombination between feline and canine coronaviruses.

Type II feline coronavirus (FCoV) emerged via double recombination between type I FCoV and type II canine coronavirus (CCoV). In this study, two type I FCoVs, three type II FCoVs and ten type II CCoVs were genetically compared. The results showed that three Japanese type II FCoVs, M91-267, KUK-H/L and Tokyo/cat/130627, also emerged by homologous recombination between type I FCoV and type II CCoV and their parent viruses were genetically different from one another. In addition, the 3′-terminal recombination sites of M91-267, KUK-H/L and Tokyo/cat/130627 were different from one another within the genes encoding membrane and spike proteins, and the 5′-terminal recombination sites were also located at different regions of ORF1. These results indicate that at least three Japanese type II FCoVs emerged independently. Sera from a cat experimentally infected with type I FCoV was unable to neutralize type II CCoV infection, indicating that cats persistently infected with type I FCoV may be superinfected with type II CCoV. Our previous study reported that few Japanese cats have antibody against type II FCoV. All of these observations suggest that type II FCoV emerged inside the cat body and is unable to readily spread among cats, indicating that these recombination events for emergence of pathogenic coronaviruses occur frequently.

High Prevalence of Hepatitis E Virus in Wild Boar (Sus scrofa) in Yamaguchi Prefecture, Japan

Abstract Hepatitis E virus (HEV) causes a food- and water-borne disease in humans, and Japanese wild boar (Sus scrofa leucomystax) meat is one of the most important sources of infection in Japan. We tested 113 serum samples from wild boar captured in Shimonoseki City, Yamaguchi Prefecture, Japan from 2010 to 2012. Serum samples were tested by enzyme-linked immunosorbent assay (ELISA) using virus-like particles as antigen and nested reverse-transcription PCR (RT-PCR). Anti-HEV IgG antibodies were detected in 47 of the 113 wild boar serum samples (42%), and HEV RNA was detected in five samples (4%). Sequence analysis showed that the five HEV isolates belonged to genotype 4, forming a cluster with a previous isolate from a human hepatitis E case in this region in 2011. These results indicate that wild boar in this region are infected with potentially pathogenic HEV at a high prevalence.

Genetic characterization of coronaviruses from domestic ferrets, Japan.

We detected ferret coronaviruses in 44 (55.7%) of 79 pet ferrets tested in Japan and classified the viruses into 2 genotypes on the basis of genotype-specific PCR. Our results show that 2 ferret coronaviruses that cause feline infectious peritonitis–like disease and epizootic catarrhal enteritis are enzootic among ferrets in Japan.

Identification of a major immunogenic region of equine herpesvirus-1 glycoprotein E and its application to enzyme-linked immunosorbent assay.

A major immunogenic region of equine herpesvirus (EHV)-1 glycoprotein E (gE) was identified. Firstly, the various fragments of EHV-1 gE were expressed as fusion proteins with glutathione S-transferase (GST) in Escherichia coli and their antigenicities were compared by immunoblot analysis using sera from horses experimentally infected with EHV-1. Thirty-three amino acids of gE (a.a. 169-201) specifically and sensitively reacted with the antibodies induced by EHV-1 but not EHV-4 infection. The corresponding region of EHV-4 gE (a.a. 169-199) did not react with antibodies to EHV-1, indicating that this region is specific for each virus. In addition, when the antigenicities of three 20-mer synthetic peptides of EHV-1 gE, gE1(169-188), gE1(176-195) and gE1(182-201) were compared by enzyme-linked immunosorbent assay (ELISA), gE1(169-188) was found to contain a major B-cell epitope. ELISA using two synthetic peptides, gE1(169-188) and gG4(319-330), previously identified as the major EHV-4-specific epitope in gG, was developed and could specifically detect antibodies to EHV-1 and EHV-4, respectively. In Japan, the EHV-1 deleted in gE gene (EHV-1 ΔgE) virus is expected to be introduced in the field as a new modified live vaccine. This ELISA did not react with antibodies induced by inoculation with EHV-1 ΔgE, indicating that it is a useful method to differentiate between EHV-1 infection and EHV-1 ΔgE inoculation. In conclusion, the ELISA described herein, using synthetic peptides, is a simple method to distinguish between EHV-1 and EHV-4 infections and will be suitable as a vaccine marker after introduction of EHV-1 ΔgE into field horses.

Development and application of an indirect enzyme-linked immunosorbent assay for serological survey of Japanese encephalitis virus infection in dogs.

Japanese encephalitis virus (JEV) causes serious acute encephalitis in humans and horses. Although dogs are good sentinels for assessing the risk of JEV infection to humans, a virus neutralization test has been the only method available for measuring the levels of JEV antibody in dogs. In this study, an indirect enzyme-linked immunosorbent assay (ELISA) using purified viral particles as an antigen, was developed for serological survey of JEV infection in dogs. In dogs inoculated experimentally with JEV, the ELISA detected anti-JEV IgM 3 days after infection, with IgM levels peaking 7 days after infection. Anti-JEV IgG was detected 14 days after infection and peaked on 21-28 days after infection. Virus neutralization titers correlated with anti-JEV immunoglobulins measured by the ELISA. To test the utility of the new assay, the seroprevalence of JEV infection among 102 dogs in Kyushu, Japan, was examined by IgG ELISA and by virus neutralization. The correlation coefficient between the IgG ELISA and virus neutralization was 0.813 (p<0.001); comparison of the IgG ELISA and virus neutralization showed a sensitivity and specificity of 82% and 98%, respectively. The IgG ELISA was used to survey dogs in Bangkok, Thailand and 51% of these dogs were found seropositive for JEV. These data suggest that in the capital city of Thailand, the risk of infection with JEV remains high.

Characterization of Glycoproteins in Equine Herpesvirus-1

ABSTRACT In this study, we attempted to express twelve glycoproteins of equine herpesvirus-1 (EHV-1) in 293T cells and to characterize these using monoclonal antibodies (MAbs) and horse sera against EHV-1. Expression of glycoprotein B (gB), gC, gD, gG, gI and gp2 was recognized by immunoblot analysis using horse sera, but that of gE, gH, gK, gL, gM and gN was not. Four MAbs recognized gB, four recognized gC and one recognized gp2. Two MAbs against gB cross-reacted with EHV-4. Interestingly, coexpression of gE and gI and gM and gN enhanced their antigenicity. Furthermore, immunoblot analysis of gp2 showed that different molecular masses of gp2 were recognized by the MAb against gp2 and horse sera against EHV-1. In this study, it was demonstrated that at least six glycoproteins were immunogenic to horses, and coexpression of gE and gI and gM and gN was important for enhancement of antigenicity.

Fatal Tickborne Phlebovirus Infection in Captive Cheetahs, Japan

Two captive cheetahs from a zoo in Japan died of a severe fever with thrombocytopenia syndrome–like illness. Severe fever with thrombocytopenia syndrome virus, an endemic tickborne phlebovirus, was detected systemically with secretion of infectious viruses into the saliva. These cases highlight the risk for exposure of captive animals to endemic arthropodborne pathogens.

The complete genomic sequence of Rhinolophus gammaherpesvirus 1 isolated from a greater horseshoe bat

In a comprehensive research project on bat viruses, we successfully isolated a novel herpesvirus from the spleen of a greater horseshoe bat (Rhinolophus ferrumequinum) in Japan using a cell line established from the kidney of the same bat. This herpesvirus was a novel gammaherpesvirus (Rhinolophus gammaherpesvirus 1; RGHV-1), which belonged to the genus Percavirus. The whole RGHV-1 genome (147,790 bp) showed that 12 of the 84 genes predicted to contain open reading frames did not show any homology to those of other herpesviruses.

Getah virus epizootic among wild boars in Japan around 2012

In 2014, an outbreak of Getah virus (GETV) infection occurred in Japan in a horse population that was inoculated with a vaccine against GETV. In this study, we investigated the seroprevalence of GETV infection among wild boars in Japan. Interestingly, the highest rate of anti-GETV-positive wild boars was observed in 2013, which gradually decreased during 2014–2016. The results suggested that GETV spread among wild boars around 2012, resulting in the 2014 outbreak.