1996 Vol.11(4)



Advances of research on chicken anaemia virus

LU Chun, CHEN Bu-Yan, CA Bao-Xiang

1996, 11(4): 289

Advances in transmission mechanism of nonpersistent viruses by aphids

WU Yun-Feng, ZHOU Guang-He

1996, 11(4): 296

Research progeress on human papilloma viruses

YU Xiu-Beng, BIAN Ji-Feng

1996, 11(4): 301

Detection of hepatitis B virus in perinatal transmission by nested polymerase chain reaction

HUANG Xiao-Jun, HUANG Ya-Rong, ZHENG Hui-Tong, FU Yu-Liang, CHEN Huo-Sheng

1996, 11(4): 313

high sensitive method of nested-polymerase chain reaction(Nested-PCR)was estab-lished to detect HBV-DNA in peripheral serum from 169 HBsAg or HBsAg/HBeAg positivepregnant women and their neonates。Positive of HBV-DNA was 72.8% in 103 positive of HB-sAg mothers and 33.0% in their neOnates.In the other 66 both HBsAg and HBeAg positivemothers and their neonates,the HBV-DNA positive was 86.4% and 43.9%,Fifty five samplesof the colostrum from 169 HBsAg or HBsAg/HBeAg positive mothers were detected,the HBV-DNA was found in 20 mothers.These results suggested the e antigen was important in HBV peri-natal transmission.Additional detection was made in 105 cases of 6 months infants from atoveneonates after injection of NBV vaccine and hepatitis B immunoglobulin,23 positive samples werefound in serum。

Genomic Typing and Sequence Analysis of Hantavirus

SUN Yong-Tao, BAI Xue-Fan, YANG Wei-Song, HUANG Chang-Xing, ZHOU Yong-Xin

1996, 11(4): 319

The present study reports the use of reverse transcriptase polymerase chain reaction(RT-PCR)for typing 18 independently isolated Hantaviruses from Asia-Pacific area by 2 sets of type-specific primers and the sequence analysis.The results of PCR showed that Hantaan specificprimers could amplify all 10 isolates in the Hantaan serotype,while Seoul specific primers werereactive to all 7 isolates in Seoul serotype and Leakey.No cross-reaction was obserred in the ex-periment.The PCR products were analysed by nested PCR and restriction endonuclease digestionfor further confirmation。The sequence analysis of PCR products revealed that the sequence simi-larities of R36 between 76-118 and R22 were 78.4% and 68.1%,respectively. In addition,apairwise comparison of sequence homology revealed that R36 shares a higher degree of sequencehomology with isolates in Hantaan serotype than those in Seoul serotype.Lekey could be ampli-fied by Seoul specific primers

Expression of HCV Core Protein in Insect Cells and Evaluation 0f Its Im munological properties

XIANG Jin-Zhong, GUO Liang, ZHANG Peng-Fei

1996, 11(4): 325

Full-length cDNA coding HCV core protein was isolated from human HCV positiveserum by RT-PCR and inserted into baculovirus transfer vector.Sf9 insect cells were co-transfect-ed with the recombinant transfer vector plasmid and wild type baculovirus DNA.The recombi-nant baculoviruses containing gene of HCV core protein were obtained by plaque screening.Therecombinant HCV core proteins(20 kD)were expressed by Sf9 cells infected with recombinantviruses.Western blot and EIA revealed that the recombinant proteirls can be recognized by humanHCV positive sera,and the recombinant protein ean elicited specific anti-HCV antibody in ani-mals.

In Vitro Transcription of HCV 5 Non-Coding Region cDNA

LI Gang, YAO Ji-Lu, PANG Wen-Wei, WANG Bin, CHEN Qing

1996, 11(4): 332

epatitis C virus(HCV)5'non-coding region(5'NCR)cDNA with 302 bp obtained byreverse transcription polymerase chain reaction(RT-PCR)from the serum of a patient withchronic hepatitis C in Guangdong.Province was subsequently filled in recessed ends,purified andinserted into pUC19 plasmid vector.The recombinant plasmid pUN was sequenced.The targetgene in pUN was subcloned into EcoR I/Pst I sites of pSPORT I transcription vector.Alter therecombinant pSN was linearized,the transcription reaction in vitro was performed using SP6RNA polymerase or T_7 RNA polymerase。The sense RNA with 429 bp and anti-sense RNA with362 bp synthesized by SP6 RNA polymerase and T7 RNA polymerase respectively were identifiedby electrophoresis on agarose gel and by RT-PCR using HCV-specific primers.It was also ver-ified that the RNAs were transcripted from HCV5'NCR cDNA.The 5'NCR cDNA clone con-structed and RNA synthesized can be used as effective positive control templat for PCR and RTrespectively,which will be helpful in eliminating false

The Pr~luetion ofTNF-~ I~dueed by HI-IV ——6 in Vitro

FAN Ping, TAO Kun, JI Xiao-Hui, ZHOU Yao-Xi

1996, 11(4): 338

here are high levels of TNF-α in the supernatants of lymphocyte culture infected with HHV-6 GS strain and the Nanjing loeal strains.TNF-α was detected by two methods:biological ac-tivity assay using L929 cells and ELISA.HHV-6 can induce TNF-α with in 24 h,maximal re-lease of TNF-α occurred during 48 h to 72 h post infection and then gradually decreased.The levelof TNF-α in infected culture suprenatants is far higher than that of uninfecter cells(P0.1).Three lo-cal strains have same result.Specific monoclonal antibody against TNF-α can completely neutralizethe activity of TNF in the supernatants of these cells infected with HHV-6.The levels of TNF-αinduced by HHV-6 is much higher than that induced by LPS as a positive control.

Morphology and morphogenesis of the isolated strain of rubella virus in BHK21 cell

WANG Zhi-Yu, SONG Yan-Yan, TAO Ping, WANG Gui-Ting, HU Hong-Zhi

1996, 11(4): 343

ltrathin secction electron microscopy was used to investigate the morphology and morpho-genesis of rubella virus(RV)JR23 and Gos-10 strains in BHK21 cells and its effects on cellularultrastructures.The results showed that virions were observed in the cytoplasm at 6 h postinfec-tion and a large number of enveloped virions were seen at 96 h postinfection.The virion wasspherical,45 to 75 nm in diameter,with a 25 ̄35 nm electron-dense core covered by a double-layered lcose envelope.A numbers of roughly spherieal electron-dense particles,20 ̄30 nm indiameter,could be seen in the cytoplasm.The virus was enveloped by budding through cytoplas-mic or vacuole membrane.There was no difference between the two strains in the morphologyand morphogenesis.

Pharm acodynam ics Studies of Alternanthera philoxeroides Griseb in Oral for EHFV Infection in Suckling Mice

XIAO Hong, YANG Zhan-Qiu, WEN Li, LIU Yan-Wen

1996, 11(4): 348

Antiviral effect of Alternanthera philoxeroides Griseb(APG)was studied in oral for epi-demic hemorrhagic fever virus(EHFV)infection in suckling mice.The APG dosages of A,BandC groups were 5mg/(g·d),7.5mg/(g·d)and 10mg/(g·d),respectively. The results showedthat the survival rate of A,B and C group were 80.0%,72.2% and 40.0%,the means time day(MTD)were 56.5±0.9, 53.5±1.1 and 41.5±2.7 day,respectively.The survival rate and MTD were 0.0% and 26.3±0.8 day in untreated viral control group.The antiviral effect of theAPG was similar with the ribavirin.This indicated that the APG was effective in treatment of epidemic hemorrhagic fever.

Isolation and Identification of an Avian Reovirus Variant Strain

DIAO Zhen-Fen, XU Wei-Yan, DU Nian-Xin, XUE Heng-Beng

1996, 11(4): 352

virus strain was isolated from the liver of a sick chicken and identified to be an avian re-ovirus(RV)by observation of electron-microscopy,analysis of physical-chemieal characteris-tics,SDS-PAGE of nucleic acid and neutralization test.The virus strain produced evident cyto-pathic effect(CPE)only on chicken embryo liver cell(CELi)but not on CEF and Vero cell,andwas inactive at 50℃ for 1 hour,even in Mg ̄(2+)solution.Additionally,the migration rate of foursmall segments of it's genome was slightly different from ARV FDO and S_(1133) strain.The experi-ment suggested that the virus is a variant strain of ARV.

Amplification and cloning of protein kinase gene of pseudorabies virus by PCR

LUO Man-Lin, DING Jian-Hua, WANG Jia-Fu, ZHANG Chu-Yu

1996, 11(4): 360

Pseudorabies virus(PRV)was mumplicated on BHK-21 monolayer cells.After concentra-tion with centrifuge,viral DNA was isolated by SD-proteinase K digestion method.According to the sequences of protein kinase genes of PRV Ka and NIA-3 strains,the primers with 26 bpand 32 bp were designed.With pured PRV geneome DNA as template,the PK gene of PRV wasamplified successfully by PCR,and cloned into pUC19 vector.Restriction enzyme analysis showedthat the cIoned PK gene at Smal I,Xho I and Sal I site was the same as that of PRV NIA-3strain。This work laid foundation for constructing the PRV vaccine with deletion of PK gene.

Immunological detection of two kinds of viruses from Penaeus chinensis

SHI Zheng-Li, XIAO Lian-Chun, GAO Wei, CHEN Dai-Hua

1996, 11(4): 365

Using two kinds of spherical viruses from diseased Penaeus chinensis as antigens,the relativeantibodies were prepared.Then diseased shrimp samples from Jiangsu,Hebei,Qingdao,Fujianand Dalian were detected by counter immunoelectrophoresis,ELISA and immunological EM tech-nique.The results showed that samples from Jiangsu,Qingdao and Hebei have relative viralpathogens.

Study on ELISA method for detecting specific circulating immune complexes in sera of patients with HFRS

ZHANG Dong-Hai, SUN Hui, GAO Feng

1996, 11(4): 369

new ELISA method was established for detecting specific IgG,IgM,IgA type circulatingimmune complexes(CIC)in sera of patients with Hemorrhagic Fever with Renal Sydrome(HFRS).The rabbit anti-HFRS virus antibody was used as coated antibody and anti-humanIgG,IgM,IgA monoclonal antibodies were used as HRP conjugates.The results showed that thedetected rates of specific IgG,IgM,IgA types CIC in 73 acute phases(1 ̄8days after illness)were 69.49%,69.86% and 56.16% respectively,in 54 convalescence phases(10 ̄28 days afterillness)were 90.74%,85.19% and 75.93% respectively,28 sera in 1 ̄3 years after illness wereall 0.00%,36 serum specimens from normal and hepatitis B patients were all negative.The developed method showed highl specificity and sensitirity as well as reproducibility(CV:IGG12.07%,IgM6.29% and IgA12.89%)for the detection of HFRS specific CIC.

The effect of satellite RNAs on replication of cucunber mosaic virus genomic RNAs in vitro

YANG Hai-Hua, KANG Liang-Yi, ZHAO Da-Jian, TIAN Bo

1996, 11(4): 373

RNA - dependent RNA polymerase extracted from the cucumber cotyledons four days afterinoculation by CMV-B-1 containing attenuation satellite RNA was used to initiate synthesis of(-)sense RNA from(+)sense RNA,using both CMV genomic or satellite RNAs as tem-plates.In presence of satellite RNA,the synthesis of CMV genomic RNA1,RNA2 and RNA4was inhibited.However,different satellite RNAs showed different effects on the synthesis of viralRNAs.Adisease-attenuated satellite RNA-1 decreased the synthe.is of CMV RNA4,but thetomato necrosis-induced satellite RNA-D increased the synthesis of CMV RNA4.The resultindicated that the satellite RNAs could interfere with synthesis of CMV coat protein in the CMV-infected tissues and modulate plant symptoms by regulating the synthesis of CMV RNA1,RNA2 and RNA4.

Sequence Homology and Base Pairing between Cucumber M osaic Virus Satellite RNA an d Potato Spindle Tube r Viroid


1996, 11(4): 378

The results of computer analysis showed that,(1)47.8~66.7% seq homolo~ between CARNA5D (t-)RNA and PS『rVld(+)RNA;(2)48.1~63.4% sequeEtce homology be.tween CARNA5D(4-)RNA and PSTVd (一)RNA;(3)six dotplot between CARN^5D(+)RNA and PSTV_d(+)RNA;and(4)six base pairing regions between CARNASD(+)RNA and P明、Vd(4-)RNA containing high G :C ratio and high△ G values:KJ/tool= 一872.4~一975.9.

Isolation of a new psittacine virus in China

FENG Feng, JIA Wei, ZHAO Lin, CHEN Sheng-Liang, GONG Shao-Yu

1996, 11(4): 384

strain of new virus was isolated from fledgling budgetigars.The viruses were purified byusing Sepharose-2B chromatography.The size fo the virions was about 50 nm by electronic mi-croscopy.Pure virions were inoculated to healthy young fledgling budgerigars(not older than 5days).All of the birds were clinically abnormal,11 days later they died one by one.So we con-cluded that the virns is an important pathogen for the young fledgling budgerigars in China.