A 240-nucleotide sequence of the capsid/premembrane gene region of 23 Japanese encephalitis virus (JEV) strains isolated in Tokyo and Oita, Japan was determined and phylogenetic analyses were performed. All the strains clustered into two distinct genotypes (III and I). All strains isolated before 1991 belonged to genotype III, while those isolated after 1994 belonged to genotype I. In addition, the strains of the genotype I isolated in Japan showed a close genetic relationship with those from Korea and Malaysia. * CSF ‫ס‬ cerebrospinal fluid. † MVE virus was used as an outgroup in all phylogenetic analyses.

Hepatitis delta virus (HDV) infection is relatively common in the Miyako Islands, Okinawa, Japan, where the infection has been reported to be associated with low pathogenicity. HDV RNA extracted from each of 6 patients with HDV-related chronic liver disease living in the islands was amplified by reverse transcription-polymerase chain reaction and examined genetically to determine the HDV genotype. All isolates from the 6 patients were classified as genotype II by the neighbor-joining method. However, these isolates had relatively low homology (75-81%) to the HDV genotype II isolate reported from Japan, and showed relatively high identity (83-95%) to the novel genotype II isolate (HDV genotype IIb) recently reported from Taiwan. Phylogenetic analysis showed that the 6 isolates form a novel group within HDV genotype II. Furthermore, there was notable variation in sequence among the 6 isolates compared with the relatively close clustering of HDV isolates within limited areas (e.g., United States, Archangelos, Turkey, Albania, Peru). HDV genotype II in the Miyako Islands is therefore unique, and HDV infection may have been introduced at a relatively early time in this area.

SUMMARYThe dengue 4 virus (DEN-4) core gene and part of the PreM genes were inserted into the baculovirus polyhedrin gene region. The recombinant baculovirus directed the synthesis of the DEN-4 core protein fused to a part of the polyhedrin protein (Mr 25K), as determined by Western blot analysis using DEN-4 core monoclonal antibody. A mouse polyclonal antibody prepared against the DEN-4 core fusion protein showed antigenic reactivity with the authentic DEN-4 core protein (Mr 15.5K) present in the nucleus as well as in the cytoplasm of DEN-4-infected Veto cells as demonstrated by the peroxidase-antiperoxidase staining method. This antibody did not react with cells infected with DEN-l, -2, -3 or Japanese encephalitis virus, or mock-infected cells.

The Miyako Islands, located in the southernmost part of Japan, have been reported to be endemic for hepatitis delta virus (HDV). The majority of HDV patients in this area exhibit a relatively mild course of infection that evolves into a quiescent cirrhotic condition. The entire nucleotide sequence of the Miyako isolate (L215) of HDV obtained from a cirrhotic patient infected with HDV was determined. This isolate, L215, comprises 1682 nt and encodes 213 aa of the hepatitis delta antigen. Phylogenetic analysis showed that L215 is closely related to the Taiwanese genotype IIb HDV isolate. In addition, the predicted folding structure of the antigenomic RNA substrate was different from those of the published genotype II sequences.Hepatitis delta virus (HDV) is a unique, viroid-like human pathogen that needs to be associated with hepatitis B virus infection (Monjardino, 1996). It was first detected in the liver cell nuclei of patients with hepatitis B surface antigen (HBsAg)-positive chronic liver disease (Rizzetto et al., 1977). HDV has a single-stranded, negative-sense, circular RNA genome of 1?7 kb in length (Wang et al., 1986(Wang et al., , 1987Makino et al., 1987;Saldanha et al., 1990;Imazeki et al., 1991;Casey et al., 1993;Lee et al., 1996;Wu et al., 1998). The RNA genome encodes two proteins, 24 and 27 kDa, referred to as the small hepatitis delta antigen (S-HDAg) and the large hepatitis delta antigen (L-HDAg), respectively (Bonino et al., 1986;Bergmann & Gerin, 1986). S-and L-HDAg share the same initiation codon but use different termination codons (Wang et al., 1986;Weiner et al., 1988). Each HDAg is translated from a distinct mRNA species, arising from a unique RNA-editing event. The two forms of HDAg have distinct and opposing functions: S-HDAg is required for HDV replication, whereas L-HDAg inhibits HDV replication and is required for virion formation (Gerin et al., 2001).Based on geographical origin and genomic divergence, HDV strains are divided into at least three genotypes: I, II and III (Casey et al., 1993;Lai, 1995). Recently, Wu et al. (1998) reported a novel strain of genotype II in Taiwan and proposed genotypes IIa and IIb. The Miyako Islands, which are located in the southernmost part of Japan, about 390 km east of Taiwan, are one of the areas in Japan that is endemic for HDV (Sakugawa et al., 1995(Sakugawa et al., , 1997a(Sakugawa et al., , b, 1999Nakasone et al., 1998;Arakawa et al., 2000). The majority of patients with chronic HDV-related liver disease in the Miyako Islands exhibit a relatively mild course of infection that evolves to a quiescent cirrhotic condition (Sakugawa et al., 1997a). Sakugawa et al. (1999) sequenced the 645 nt of HDAg (position 940-1584; numbering according to Wang et al., 1987) of six strains of genotype II, collected from patients with HDV-related chronic liver disease in the Miyako Islands, and reported the presence of genotype IIb. However, the entire genome sequence of the Miyako isolate has not been reported. Hence, we describe the entire nucleotide sequence o...

In order to elucidate the molecular characteristics of Japanese encephalitis (JE) virus in Okinawa, 23 strains of JE virus isolated in a 25-year span were sequenced for the 240 nucleotides of the C-preM junction region and 111 nucleotides of the E gene region and compared with those of reference strains isolated in mainland Japan. The results of phylogenic analysis showed that although all the Okinawan isolates showed more than 96% homology in the nucleotide sequence in each region, they were chronologically divided into two groups: the old group (nine strains) and a new group (14 strains). On the other hand, in a comparison with reference strains in mainland Japan, the Okinawan isolates showed more than 94% nucleotide sequence homology in both regions, indicating that the Okinawan strains belong to the same genotype as that of JE strains in mainland Japan. The nucleotide homology of the old group was relatively higher than that of the new group. Among the 14 strains in the new group, 13 strains were isolated from mosquitoes collected from a pig farm from 1986 through 1992. These strains showed higher nucleotide divergence than the old group strains, isolated from mosquitoes and swine sera collected at several sites, in both regions. A nucleotide substitution at the position 1920 in the E gene was identified in three isolates. This substitution generated an asparagine-proline-threonine sequence capable of serving as an attachment site of carbohydrate.

In May 1994, an epidemic of dengue fever (DF) / dengue hemorrhagic fever (DHF) was reported in Ban Horn commune in Vientiane municipality. This was after a six-year period of non-epidemic DF/ DHF in the municipality. In early June, we collected 81 serum specimens from children with fever of unknown origin (FUO) in the commune. Of 81 specimens, 48 were taken from children within 5 days after the onset of FUO and subjected to dengue (DEN) virus isolation. Four strains of DEN-1 and 12 strains of DEN-2 viruses were isolated from 15 serum specimens (31.3%) . A case of dual infection with DEN-1 and DEN-2 viruses was detected by enzyme antibody method using type-specific monoclonal antibodies and reverse transcription polymerase chain reaction (RT-PCR). The percentage incidences of the positive virus isolation from the serum specimens taken on the second and the third day of fever were 41.8% and 21.1%, respectively, and no virus was isolated from the specimens other than those from these two days. All the specimens were examined by the IgM-capture ELISA. Thirty-two specimens (39.5%) were positive for DEN IgM antibody.

Dengue fever (DF) is a major public health concern in the Dominican Republic. In recent years, several epidemics of DF have been reported to the Pan American Health Office (PAHO), but the extent of the epidemics has not been clearly understood yet. Therefore, we conducted a nationwide seroepidemiology of dengue (DEN) infection. At the same time, we conducted an interview survey to assess public awareness regarding the disease. The serum samples were collected at seven main cities in the Dominican Republic and screened for DEN antibody with a commercial ELISA kit. A total of 2007 serum specimens were examined. The prevalence of DEN antibody in the seven cities varied between 43.1 and 89.7%. Neutralization (N) test carried out on the ELISA-positive serum from Samana, one of the high antibody-prevalent cities, revealed that all the sera showed positive to at least two DEN serotypes. Geometric mean N titers against DEN-1, 2, 3 and 4 were 40.5, 463.7, 59.9 and 454.4 respectively. No difference in antibody prevalence was observed between males and females. It appeared that a high level of awareness regarding DF did little affect DEN prevalence. Strong, concrete public health strategies that motivate the local community to combat DF are required.