Prevalence of hepatitis E virus (HEV) infection in wild boars (Sus scrofa leucomystax) and pigs in Gunma Prefecture, Japan.

The prevalence of hepatitis E virus (HEV) infection in wild boars and pigs in Gunma Prefecture, Japan, was serologically and genetically examined. The positive detection rates of anti-HEV IgG and HEV RNA in the wild boars were 4.5% (4/89) and 1.1% (1/89), whereas those in the pigs were 74.6% (126/169) and 1.8% (3/169), respectively. The positive rates of anti-HEV IgG and HEV RNA on the 17 pig farms in the present study ranged from 20% to 100%, respectively. One male wild boar approximately 5 years of age was positive for HEV RNA but was negative for anti-HEV IgG. Three pigs from 2 farms were positive for HEV RNA; 2 of these pigs were negative for HEV IgG, and the other was positive. A phylogenetic analysis revealed that all of the HEV ORF1 genes detected in the present study belonged to genotype III. In Gunma Prefecture, HEV is highly prevalent and widespread, and uncooked wild boar and pig meat may have the potential to transmit HEV to humans.

Role of GPI-anchored NCAM-120 in rabies virus infection.

Although the neural cell adhesion molecule (NCAM) -140 and -180 have been shown to serve as a receptor for rabies virus (RV), it was not known whether the other major isoform of NCAM, GPI-anchored NCAM-120 functions as RV receptor. In this study, we have established HEp-2 cells stably expressing NCAM-120 or NCAM-140, and their susceptibilities to RV infection were compared. The results demonstrated that NCAM-120 served as virus attachment protein; however, the cells expressing NCAM-120 did not support efficient RV replication. Furthermore, the level of IFN-ss mRNA was apparently elevated in NCAM-120 expressing cells but not in NCAM-140 expressing cells, suggesting that GPI-anchored NCAM-120 suppressed RV replication via induction of IFN-ss even though NCAM-120 was able to promote virus penetration into the cells.

Detection of rabies-specific antigens by egg yolk antibody (IgY) to the recombinant rabies virus proteins produced in Escherichia coli.

We obtained rabies-specific egg yolk antibodies (IgY) by immunizing hens with recombinant His-tagged nucleoprotein and phosphoprotein (rN, rP) of the rabies virus (CVS-11 strain) expressed in Escherichia coli. The anti-rN and rP IgY were shown to bind specifically to the respective proteins of the CVS-11 strain of rabies virus by Western blotting, immune fluorescent assay and immunohistochemistry, indicating that IgY to rabies recombinant proteins could serve as a reagent for diagnosis of rabies virus infection.

The absence of anti-rabies antibody in the sera of feral raccoons (Procyon lotor) captured in Hokkaido, Japan.

Feral raccoons captured in Hokkaido, Japan were examined for the presence of rabies virus neutralizing antibody (VNA). Between 2000 to 2002, 741 serum samples were collected and then subjected to VNA titer determination by rapid fluorescent focus inhibition test (RFFIT). Sera showing RFFIT titers of > or =1:25 have been considered as positive according to previous reports. VNA was detected in none of serum samples from the feral raccoons. The present study provides valuable background information for rabies prevention in Japan. The potential risk of raccoon-derived rabies in the wilderness has been of concern because of the increasing number of feral raccoons originally introduced from rabies-endemic countries. The continuous monitoring of sick and/or dead wild raccoons would help to prevent an epidemic spread of raccoon rabies.

Safe and easy monitoring of anti-rabies antibody in dogs using his-tagged recombinant N-protein.

The virus neutralization (VN) test is a reliable indicator of adequate vaccination in animals. However, the VN test is tedious and complicated to perform. Enzyme-linked immunosorbent assay (ELISA), though rapid and simple compared to the VN test, is complicated and hazardous during preparation of the viral antigen. In an effort to overcome the disadvantage of ELISA, the recombinant His-tagged nucleoprotein (His-rNP) expressed in Escherichia coli was used as a safe antigen for ELISA (i.e., live virus was not used). Anti-rabies antibody levels were determined by fluorescent ELISA (FELISA) using His-rNP as an antigen. The presence of anti-rabies VN antibody was determined by the rapid fluorescent focus inhibition test (RFFIT). The VN titers by RFFIT were found to correlate well with the FITC-signal determined by the FELISA (r = 0.616). The sensitivity and specificity of the FELISA were 91.7 and 100%, respectively. This study showed that the His-rNP could be useful as an antigen of ELISA to test for anti-rabies antibody in vaccinated dogs. Several studies in Japan have investigated the antibody level in the sera of vaccinated dogs. A safe and convenient test using His-rNP would contribute to our understanding of the status of herd immunity among not only domestic dogs but also stray dogs in Japan.

Production of rabies neutralizing antibody in hen's eggs using a part of the G protein expressed in Escherichia coli.

In an attempt to produce anti-rabies immunoglobulin affordable for people living in developing countries, we have immunized layer chickens with a part of the G protein of rabies virus expressed in Escherichia coli. Immunoglobulin (IgY) was purified from the yolks of eggs layed by immunized hens. It was revealed in vitro that the antibody specifically bound to virions as well as cells infected with rabies virus. Moreover, the antibody apparently neutralized rabies virus infectivity. Inoculation of the antibody into mice infected with rabies virus reduced the mortality caused by the virus, suggesting that IgY directed to the part of the G protein expressed in E. coli could serve as a possible alternative to currently available anti-rabies human or equine immunoglobulins.

Characterization of spotted fever group rickettsiae detected in dogs and ticks in Okinawa, Japan.

Spotted fever group (SFG) rickettsial DNAs were detected in 2.4% of 340 canine blood samples and a pool of 84 tick pool samples (229 ticks) collected in Okinawa, Japan by PCR using a citrate synthase and an SFG rickettsial 190-kDa surface antigen gene primer pair. The sequences of both genes from canine blood and tick samples showed high levels of similarity with those of Rickettsiajaponica and several SFG rickettsiae (R. aeschlimannii, R. massiliae, R. rhipicephali and Bar-29 strain). Phylogenesis of canine blood and tick samples was closely related to that of reference SFG rickettsiae. Serological evidence of SFG rickettsial infection in dogs and humans in Okinawa, where no clinical human cases have been reported, has been obtained. In this study, genetical characterization of SFG rickettsia in Okinawa was investigated phylogenetically.