1997 Vol.12(1)
随着对虾养殖业的发展，其病害亦日趋突出。对虾病原的研究，特别是病毒性病原的研究，是当前我国发展对虾养殖业的重要课题，它对虾病的防治与检测提供重要的科学依据。为此，我们对近十多年来已发表的有关对虾病毒方面研究的论文和资料作一概述，希望能对同行的研究工作...
The genes of poliovirus type I isolated from environmental water were studied by PCR (polymerase chain reaction). It was found that 22 strains of the virus were all Sabin-related. The result showed that the wild polioviruses in the environmental water were substituted by the sabinstrain completely after carrying on the conduct of mass immunization campaignS and EPI in thisarea. The method was suitable for the gene analysis of poliovirus type I.
The replication of dengue-2 virus in cultured human endothelial cells derived from humanumbilical veins was demonstrated. The viruses were detected in the supernatant as early as 12 h after infection by microplaque forming assay on C6/36 monolayer cells. The highest viral titers were obtained at 48h, then rapidly declined to very low titers at 96h. Dengue-2 virus antigenpositive human endothelial cell were demonstrated by an indirect immunofluorescent assay. No obvious differences in morphology and structure were observed between the virus infected cells and uninfected cells under phase-contrast microscope and transmission electron microscope. Viral production of infeted cells was depended on the multiplicity of infection (MOI). As the viral dose was increased from MOI of 0.1 PFU/cell to 100 PFU/cell, significant increase of viral production was observed.
Reverse transcription polymerase chain reaction was used to detect hepatitis C virus (HCV) RNA in 95 serum specimens of blood donors with anti-HCV IgG negative. Six out of 8 detections, HCV RNA was detected in 17 of 95 specimens (17. 9% ). These 17 HCV RNA positive specimens were detected once more for anti HCV IgG to confirm their negative detection. Sequencing data of hypervariable region from 8 of 17 positive PCR products showed that the amplificated sequences came from different HCV strains, and were not the false amplification because of PCR contamination. In these 8 PCR products, two nucleotide sequences were determined at fulllength, and showed higher homology of 77%~79% with the corresponding sequences of representative strain of HCV genotype Ⅱ than that of 62%~69% with the sequences of HCV genotypes Ⅰ, Ⅲ, Ⅳ, and were verified as HCV genotype Ⅱ sequences which are most popular among Chinese HCV infected cases. Our data demonstrated that anti-HCV IgG detection is not sensitive enough for HCV scre
The primary manufactory technique for Vero cell rabies vaccine was established. The quality of the experimental vaccine meet "the requirment for Vero cell rabies vaccine" introduced byWHO in 1986.The Vero cell strain used for the vaccine production was from ATCC(U. S. A. ). RFD virusstrain used as seed virus for the vaccine was established by our laboratory in 1989.The Vero cell was cultured in X-celligen(5L) with microcarrier and then were infectedwith RFD fixed virus. The supernatant containing rabies virus was harvested at 2~3 days intervals. The virus titer of the supernatants was 6~8 log LD(50)/mL. The virus pool was concentratedby ultrafiltration and the virus components were isolated by chromatograph. The final productcontains human albumin as stabilizer and Al(OH)3 as adjuvant. So, it is an adsorbed purified Verocell rabies vaccine.Six lots of the vaccine were detected by QC group. The results showed that the vaccine is safeand potent.
A partial gene of HGV was cloned and sequenced from the serum of a blood donor in Hebei Province of China. The partial sequence from the nostructural region gene showed 90. 5% nucleotide identity and 95 % amino acid identity over the corresponding region of isolates in America. One hundred and fifteen samples collected in Jangshu and Hebei Provinces were detected for HGV RNA by RT-PCR. The HGV RNA positive rate is 3.47%.
The cloned coat protein (CP) gene from Ys strain of papaya rinspot virus (PRV) was sequenced by dideoxy-chain termination method. Results show that the full length of the CP genewas 858 nucleotides (nt). Comparison among CP genes of 16 PRV strains or isolates from Chinaand abroad indicated that Ys shared high CP gene homologous sequence (94.44%~97.68%)with the other strains or isolates from China, while it shared lower CP gene homologous sequence(88.88%-92.70% )with the strains or isolates abroad. It was noted that there were 6 nt missing after the 63rd nt in Ys CP gene, and there were 3 nt missing in CP genes of Sin. G. THAI.SRI. Ys CP cDNA was inserted into the cloning sites between CaMV 35S promoter and nos terminator of the middle plasmid pRok Ⅱ to produce the plant expression vector pRPCY. LBA44004of Agrtobacterium tumefasciens containing the binary vector system pRPCY/pAL4404 was obtained through triparental mating, and will be used to introduce CP gene into papaya genome.
Because of the close phylogenetic relationship, the simian acquired immunodeficiency syndrome (SAIDS) is considered as an important model in search of new drugs in prevention and treatment of the pandemic human AIDS. The simian AIDS type D retroviruses (SRV) and simian immunodeficiency viruses (SIV) are the causative agents of SAIDS. A SRV model to screen anti -AIDS drugs in vitro was established. AZT and trichosanthin (TCS) were shown to inhibit SRV1 syncytium formation obviously. The selectivity indexes (SI) of AZT and TCS were 13500 and 8812, respectively. Forty-six natural compounds isolated from the Chinese medical herbs were screened by using this model. There were eleven natural compounds with obvious antiviral activity in vitro. These data indicated that SRV model for screening anti-AIDS drugs was feasible.
In vivo transfection with a plasmid-linked tandem DHBV DNA dimer on 2-day old DHBV-free Furong ducklings was carried out. Most of the animals (86%) developed transient viremia. Serum DHBs/pre S Ag and DHBV DNA appeared on 9th day, reached to its peak on12th~14th day and disappeared on 28th after transfection. Whereas, there were a few ducks whose viremia persisted for more than 50 days. DHBV DNA replicative intermediates were alsofound in the liver tissue of transfected ducks. DHBV intact viral panicals were detected in viremic sera under electronic microscope. Intraperitoneal injection of the transfected ducks' viremic sera into 1-day old DHBV-free ducklings resulted in productive DHBV infection on 60% animals,demonstrating the production of biologically active virus after in via transfection. The establishmant of this method is valuable for the research of DHBV variant and the relation between DHBVgene structure and its function.
The viruses were isolated by cocultivation of sheep foetal lung cells and the peripheral blood mononuclear cells of goats inoculated with ovine progressive pneumonia virus (OPPV). The result indicates that OPPV can infect goats. The isolated viruses were characterized by observations of cytopathogenic effects, indirect fluorescent antibody assay, electron microscopy, and polymerase chain reaction (PCR), and were further identified as OPPV. The isolation results show that this is a sensitive method of isolating OPPV. Sheep foetal lung cells can be passaged for forty times, and each subculture can be used to isolate virus, so it is a very practical method of isolating OPPV.
By studying the flue structure of GCHV (Grass Carp Hemorrhage Virus), It was found that the capsid of purified GCHV can be degraded spontaneously into its morphological units by prologing in a buffer of low ionic strength, but not very completely. The virus preparation was obtained from GCHV infected cell, then treated with 1% NP40 reagent in low-salt solution and fieezed thaw ng, finally purified by differential centrifugation. Using negative staining method, out, core capsid and capsomeres of the virus were showed by electron micrograph. Afterwards, the preparation digested by RNase was equally mixed with Freund's adjuvant and immunized rabbits. The antibody was tested by ELISA and neutralization test, which demostrated the preparation can elicit good levels of antibody. The RNA-free preparation was detected by PAGE and DIG-labelled GCHV-cDNA hybridization. The sensitivity of the test is approximately to 10pg, the samples of subunit all showed negative.
This study showed that 300 W/cm2 UV irradiation for 5 min could inactivate 94.63% ofHAV. No obvious changes were found experimentally under this irradiation dose, other than the decrease of RNA transfection ability. After 20 min UV irradiation, all of HAV were inactivated. For the RNA sedimentation, the original single UV absorption peak (32.9S) became two (11 S and 15.2S) after the irradiation. The fragment of 5'-untranslated region (HAV RNA 222~ 819) could not be detected with RT-PCR. The RNA transfection became negative. But the fragment of P1 region (HAV RNA 2823~3049) could still be detected with RT-PCR, the hybridhation shadow of 3'-terminal poly(A) tail with oligo(dT) probe labelled with -(32)P also became slightly lighter, and no obvious changes were found in the antigenicity of HAV detectedwith ELISA and in 4 capsid polypeptides which was separated by 15% SDS-polyacrylamide gel containing 4 mol/L urea.
Peptides of coat proteins of cucumber mosaic virus isolates infecting banana from Guangdongprovince were first analysed. The coat proteins of different isolates were digested with Chymotrypsin and Pepsin to produce different peptide patterns. They were evaluated by Western blotusing the antiserum to CMV-T37. It was found that MM contained more quantitative peptidesof antigen determinants to antiserum CMV-T37 than BS or CS did.
Serological characterization of three strains (BS, CS and MM) from Guangdong and one strain (HI) from Hawaii of cucumber mosaic virus infecting banana were first studied. These fourstrains were confirmed to belong to CMV DTL serotype. The serological relationships between BSand CS were closed while theprelationshi s between BS and MM were less closed.
