Bovine growth hormone (bGH) gene was expressed in an insect Spodoptera frugiperda cell line using a Baculovirus, Hyphantria cunea nuclear polyhedrosis virus (HcNPV). The bGH gene in pbGH plasmid was sequenced and amplified by PCR technique with two primers containing NcoI sites. The bGH gene consisted of 654 bp (217 amino acid residues), the 5'-untranslated region of the cloned bGH cDNA contains 56 bp, and the 3'-untranslated region contains 145 bp and two pallindromic regions. The amplified bGH gene DNA fragment (654 bp) was inserted into the NcoI site of the pHcEVII vector, which was named pHcbGH. The pHcbGH transfer vector DNA and the wild type HcNPV DNA were cotransfected into S. frugiperda cells to construct a recombinant virus. Eight recombinant viruses were selected and named HcbGH. One clone, HcbGH-4-1 showed largest plaque size, therefore the recombinant virus was further studied. The multiplication pattern of the recombinant HcbGH-4-1 was similar to that of the wild type HcNPV. The bGH gene DNA in the HcbGH-4-1 recombinant was confirmed by Southern blot hybridization. The amount of the bGH (217 amino acid residues, 21 kDa) produced in S. frugiperda cells infected with the HcbGH-4-1 recombinant was approximately 5.5 ng per ml ($10^6$ cells) by radioimmunoassay.

Recombinant polioviruses have been developed by many research groups for use as vaccine vector because poliovirus induces mucosal immunity as well as humoral immunity through oral uptake. We assessed the potential use of poliovirus as a T-cell epitope carrier. Recombinant poliovirus V129 5L was constructed to have a substituted T-helper epitope from the core protein of Hepatitis B virus at neutralization antigenic site 1 on its VP1 capsid protein. The recombinant virus replicated less efficiently than type 1 poliovirus Mahoney strain. The V129 5L formed a little smaller plaques than the Mahoney strain and showed some 1.25 log unit lower titer at the peak in the one-step growth kinetics though it had similar growth profile to that of the Mahoney strain. Since V129 5L recombinant virus was genetically stable even after 24 successive passages in HeLa cells, the antigenic site 1 on VP1 capsid protein was confirmed for its ability of carrying T cell epitope. The genetic stability of V129 5L also indicated that recombinant poliovirus can be successfully utilized for the development of the multivalent vaccines.

Hantavirus is a genus of the Bunyaviridae family consisting following serotype groups: Hantaan, Seoul, Puumala, Prospect Hill, Thailand, Belgrade, Thotta palayam, Sin Nombre. Most of Hantavirus group have been associated with many clinically similar disease known collectively as hemorrhagic fever with renal syndrome (HFRS). Hantaan virus is the prototype of the genus hantavirus, originally isolated from Apodemus agrarius. Bat was found as a natural host for Hantaan virus in Lee's lab for the first time. Then, Hantaan-like virus was isolated Hantaan-like virus from bat. To identify hantaviruses that are present in Korea among bats, bats were collected from Jeong-Sun, Won-Joo, Chung-Ju and Hwa-Cheon area, RNA was isolated from lung and serum. RT-PCR was performed with a universal primer from M segment. Nested RT-PCR was carried out to differentiate Hantaan, Seoul and Puumala virus using serotype specific primers. As we expected, Hantaan viruses were detected in bats and Seoul virus was not detected. Interestingly, Puumala viruses were also detected in bats from Won-Ju, but not in other areas. Puumala virus is originally isolated from Clethrinomys glareolus, and cause light HFRS. Recently, Paradoxomis webbiana, a wild bird turn out to be a reservoir for Puumala virus in Korea. These data indicate that bat is a new natural reservoir of Puumala virus.

Various hantaviruses were isolated from HFRS patients and various rodent species, in many parts of the world. Bandicotas were captured at Yogyakarta, east region of Sumatura island, Indonesia; and 4 rodents species including Bandicotas were captured at Chiang Rai in Thailand during 1995. Sera were collected from captured bandicotas and other rodent spicies were screened for antibody test against Hantaan (HTN), Seoul (SEO), Puumala (PUU) and Sin Nombre (SN) viruses by immunofluoresence antibody assay (IFA). Hantavirus antigen in lung tissues were tested by IFA. Among 55 captured Bandicota indica in Indonesia, 14 (25.5%) were antibody positive against HTN, SEO, PUU and SN virus. Hantavirus antigen were detected from 5 (9.0%) out of 55 lungs tested. Among 34 captured Bandicota indica in Thailand, 9 (26.5%) were antibody positive against HTN, SEO, PUU and SN virus. Among 34 lungs tissues of Bandicota indica examined, 3 (8.8%) were antigen positive. In other rodent species, antibody positive against Hantaviruses of Rattus rattus, Rattus losea and Mus cervicolor were 4/62 (6.5%), 5/25 (20%), 1/1 (100%), respectively. But no one has antigen in their lung tissues. Antigen positive lungs suspension were inoculated into vero E6 cells for virus isolation and 4 viruses were isolated from Indonesian Badicota and 3 viruses from Thailand.

Characterizations of the VP4 (P type) and VP7 (G type) genes of Korean isolates of bovine rotavirus were performed using RT-PCR/RFLP and nucleotide sequencing analysis. After RT-PCR amplification of partial length (1094bp) of the VP4 and full length (1062bp) of the VP7 genes, amplified PCR products were digested with restriction endonucleases and digestion patterns were compared with those of reference rotaviruses. With the VP4 genes, four RFLP (A-D) profiles were observed; three (A, Band C) were the same as those of bovine rotavirus NCDV (P[1]), IND (P[5]) and B223 (P[11]), respectively. Profile D was the same as that of porcine rotavirus OSU (P[7]). With the VP7 genes, five RFLP profiles (I-V) were observed; three of them (I, II and III) were the same as those of bovine rotavirus NCDV (G6), Cody 1-801 (G8), and B223 (G10), respectively. Profile IV and V were atypical to those of reference bovine rotaviruses used in this study. These two profiles were identified as G6 and G5, respectively, after analyzing and comparing the nucleotide sequences. The G typing analysis revealed that 61.9% (26/42) were G6, which included G6 subtype; 28.6% (12/42) were G5; 7.1% (3/42) were G10; 2.4% (1/42) were G8. The P typing analysis revealed that 54.8% (23/42) were P[5]; 28.6% (12/42) were P[7]; 11.8% (5/42) were [11]; 4.8% (2/42) were P[1]. Our results showed that G6/P[5] were the most prevalent rotaviruses in diarrheic calves in Korea. Also, this is the first report that G5/P[7] rotaviruses were identified from cattle with diarrhea.

We investigated the rate of hepatitis G virus infection among 50 patients who were not infected with the hepatitis C virus but showed symptoms of hepatitis. Viral RNA was extracted from the patients' sera and cDNA was synthesized and amplified by RT-PCR (reverse transcription-polymerase chain reaction) using random hexamer and 5 primers (470-20-1-77F, 470-20-1-211R, 470-20-1-211R-biotin, GV57-4512MF, GV57-4657MR). The amplified PCR products were confirmed by electrochemiluminescence (ECL), liquid hybridization (LH) and Southern blotting (SB). Among the 50 PCR products, by means of ECL, we found 4 samples to be positive and 5 samples to be indeterminate. The GV45-89M probe (5'-CYCGCTGRTITGGGGTGTACfGGAAGGC-3') was end-labelled with gamma-$^{32}P$ ATP and used for liquid hybridization with the PCR products. By using liquid hybridization, we detected specific bands from 4 positive sera and also from one indeterminate serum as determined by ECL. An 1.5% agarose gel electrophoresis of the 9 PCR products which were HGV positive or indeterminate as determined by ECL showed a 160bp band from 4 positive and one indeterminate serum. The 5 PCR products proved to be positive when SB was applied with the GV45-89M probe as well as when LH was applied. LH and SB were shown to have higher sensitivity and specificity than ECL. Two cases among 5 positive cases had relatively high SGOT, SGPT, ALP values when compared with other 48 cases. In summary, we confirmed hepatitis G virus infection in 5 cases among 50 Korean patients showing symptoms of viral hepatitis.

DNA methylation degree in the several murine brain and liver genes of different ages and after scrapie infection have been examined by using methylation-sensitive restriction endonuclease digestion. We found that the methylation of c-fos and c-myc in the brain and liver was increased during the late fetal to one month postnatal developmental periods. However, those of the SGP-2, $S100{\beta}$, APP950, PrP, and APLP1 genes were decreased at the same periods. The comparison of the DNA methylation patterns between scrapie infected brains and controls demonstrated there is no significant difference in methylation degree of scrapie-infected brains. These observations indicate that DNA methylation might be importantly related to the aging process. The scrapie-infected murine brain was not significantly developed more senescent than the same age-controls did.

In Sprague Dawley (SD) rats, antibodies against strains of Orientia tsutsugamushi, Kato, Karp and Gilliam, were produced in order to investigate their longevity and cross-reactivities to their corresponding homologous and heterologous antigens. By immunofluorescence assay (IFA) of IgG and IgM, it was shown that the immunity to the homologous strains persisted at a higher level (longevity of at least 34 weeks with higher IFA titers). On the other hand, the immunity to the heterologous strains persisted at a lower level (longevity of 10 to 34 weeks with lower IFA titers). Since infection with one strain of O. tsutsugamushi does not preclude reinfection with other strains, understanding of the antigenic diversity of O. tsutsugamushi and duration of the immunity to both homologous and heterologous strain is very important in diagnosis of scrub typhus.