Table of Content

28 February 2005, Volume 23 Issue 1

Previous Issue    Next Issue

论著

Enhancement of the Protective Effect of SjC23 DNA Vaccine against Schistosoma japonicum Infection by Immunostimulatory Sequence

ZHAOSong;ZHUYin-chang*;D.A.Harn;SIJin;RENJian-gong;YINXu-ren;HEWei;LIANGYou-sheng;XUMing;XUYong-liang

2005, 23(1):  1-5. 

Asbtract ( )   PDF (376KB) ( )  

Related Articles | Metrics

Objective To investigate the effect of immunostimulatory sequence on SjC23 DNA vaccine against Schistosoma japonicum infection. Methods SjC23 gene fragment was inserted into pcDNA3.1-CpG to construct pcDNA3.1-SjC23/CpG. BALB/c mice in 4 groups were immunized intramuscularly 3 times at 2 week intervals, with 100 μg plasmid DNA per injection. Four weeks after the 3rd immunization, all mice were challenged with 45±1 cercariae of S.japonicum by abdominal skin penetration. After 45 days post-challenge, mice were perfused and the number of recovered worms and of eggs in liver was counted. Blood samples were collected from the tail vein of all mice 2 days before the 1st immunization and before challenge respectively. IgG, IgG1 and IgG2a in sera were detected. Three weeks after the 3rd inoculation, the spleen cells of 2 mice from each group were cultured and stimulated with ConA and recombinant peptide. The supernatant was collected to detect IL-2, IL-4 and IFN-γ. Simultaneously, the cytotoxic activity was detected with 51 Cr release assay in vitro. Results The worm reduction rate in SjC23 group and SjC23/CpG group was 28.1% and 35.1%, the hepatic egg reduction rate was 21.6% and 26.5%, respectively, compared with the control group. The level of protection in SjC23/CpG group was higher than that in SjC23 group (P<0.05). ELISA results indicated that mice immunized with pcDNA3.1-SjC23 and SjC23/CpG produced specific IgG to rSjC23, while mice immunized with pcDNA3.1 and pcDNA3.1-CpG did not. Mice in SjC23 group and SjC23/CpG group also produced IgG1 and IgG2a antibody isotypes, with the ratio of IgG2a/IgG1 10.1 and 12.2, respectively. In comparison with the control, the level of IL-2 and IFN-γ in mice immunized with pcDNA3.1-SjC23 and pcDNA3.1-SjC23/CpG was augmented. The cytotoxic activity of spleen cells from mice in SjC23/CpG group was augmented from 9.7% to 40.0% compared with that in SjC23 group. Conclusion The study indicates that immunostimulatory sequence appears to increase the level of protection induced by immunization with pcDNA3.1-SjC23 vaccine.


Studies on the Association of Human Leukocyte Antigen Class Ⅱ Alleles with Advanced Hepatosplenic Schistosomiasis japonica

ZHANGJun-hua;LIUWen-qi;LIYong-long*;LONGXiao-chun

2005, 23(1):  2-9. 

Asbtract ( )   PDF (167KB) ( )  

Related Articles | Metrics

Objective To study the association of human leukocyte antigen classⅡ(HLA-Ⅱ) alleles with genetic susceptibility and resistance to advanced hepatosplenic schistosomiasis japonica. Methods The allelic types of HLA-DRB1, DPA1, DQA1 and DQB1 were detected by polymerase chain reaction with sequence-specific primers (PCR-SSP) technique in 46 patients with advanced hepatosplenic schistosomiasis, characterized with extensive liver fibrosis. Another 43 subjects with chronic schistosomiasis were used as control. The statistical significance of differences in allelic frequencies was determined by χ ² test. Results The frequencies of HLA-DRB1*04, DPA1*0103, DQA1*0601, DQB1*0201 in advanced patients were markedly higher than those in control group, while the frequencies of HLA-DQA1*0501 and DQB1*0601 in control group were higher than those in advanced patients. Conclusion The study indicated that HLA-DRB1*04, DPA1*0103, DQA1*0601 and DQB1*0201 showing a positive, statistically significant (P<0.05) association with advanced hepatosplenic schistosomiasis japonica may be the susceptible genes, whereas HLA-DQA1*0501 and DQB1*0601 may be more relevant to a resistance to the disease.

Clinical Analysis on Hepatic Hydatid Disease in Yili River Valley

GAOYong-sheng;ZHUMa-bai;GUOYong-zhong;DILMura-ti;Ar-xen;WANGYan;CHUYI-ming;WENHao;LIANGDong;LIShi-cai;LIChang-yu

2005, 23(1):  3-13. 

Asbtract ( )   PDF (110KB) ( )  

Related Articles | Metrics

Objective To analyze the clinical diagnosis and treatment of hepatic hydatid disease and its epidemiological characteristics in Yili river valley. Methods Retrospective investigation was carried out on 2 049 cases collected in 1993-2003. Clinical diagnosis was made by ways of intradermal test, serological test, ultrasound, X-ray, CT and/or MRI, majority of them received surgical operation. Results Among the 2 049 cases, cystic hydatidosis occupied 96%( 1 965 / 2 049 ), while 4%(84/ 2 049 ) were alveolar hydatidosis. 99% ( 2 034 / 2 049 ) accepted surgery including hepatolobectomy, endocystomy and hydatidostomy in 302 cases (14.7%) without relapses. 754 cases (36.7%) received chemotherapy (praziquantel, albendazole) after surgical operation. The disease distributed in agri-pastoral areas along the valley. Local residents from different minorities had a close contact with dogs, 54% of the cases were females and 48% of the cases were in the group of 25-49 years old. The incidence tends to decline in the years. Conclusion Hydatidosis is still an important health problem in the region. Further practices for improving treatment especially surgical intervention and for epidemiological investigation are needed.

Construction of Monovalent and Compound Nucleic Acid Vaccines Against Toxoplasma gondii with Gene Encoding p30

YANGTing-ting;HEShen-yi*;JIANGHua;GUQin-min;CONGHua;ZHOUHuai-yu;ZHANGJia-qin;LIYing;ZHAOQun-li

2005, 23(1):  4-17. 

Asbtract ( )   PDF (591KB) ( )  

Related Articles | Metrics

Objective To construct a monovalent gene vaccine pcDNA3.1-p30 and a compound gene vaccine pcDNA3.1-p30-ROP2 and assess the protective effect of the two vaccines against Toxoplasma gondii. Methods The sequences encoding p30 and ROP2 were amplified from the genomic DNA of T.gondii RH strain by polymerase chain reaction (PCR) and inserted into eukaryotic vector pcDNA3.1 to construct pcDNA3.1-p30 and pcDNA3.1-p30-ROP2. Mice were injected with the recombinant plasmid to observe the immunoprotectivity of the nucleic acid vaccine by using ELISA for detection of total IgG and observing the survival time after tachyzoites challenge. Results The recombinant plasmids pcDNA3.1-p30 and pcDNA3.1-p30-ROP2 were constructed. Mice in pcDNA3.1-p30-ROP2 group showed higher IgG(P<0.05) and survived longer than those in pcDNA3.1-p30 group(P<0.01)after challenged with T.gondii. Conclusion Compound vaccine of genes from different stages of T.gondii elicits stronger immunoprotectivity in mice than a single gene vaccine.

Cloning and Prokaryotic Expression of Transcriptional Co-activator Gene of Clonorchis sinensis and Functional Analysis of the Expressed Protein

ZHANGYong-li;YUXin-bing*;WUDe;WUZhong-dao;BIHui-xiang

2005, 23(1):  5-23. 

Asbtract ( )   PDF (478KB) ( )  

Related Articles | Metrics

Objective To construct prokaryotic recombinant plasmids of transcriptional co-activator (TC) gene of Clonorchis sinensis, express and purify the recombinant protein and analyze its biological function. Methods A pair of primers was designed according to the known sequence of TC gene. The TC gene fragment was amplified by PCR. After purification and digestion with BamHⅠ and SalⅠ , the TC gene was connected to the prokaryotic expression vectors, pGEX-4T-1 and pET30a(+). By cloning target gene into these vectors, pGEX-4T-1 and pET30a(+), prokaryotic recombinant plasmids of TC gene were constructed and transferred into E.coli BL21. The positive expressed recombinants were detected by SDS-PAGE and Western blotting. Immobilized metal (Ni2+ ) chelation affinity chromatography was used to purify His-TC produced by the expression of the recombinant protein pET30a(+)-TC. Results The recombinant plasmids, pGEX-4T-1-TC and pET30a(+)-TC, were constructed successfully. SDS-PAGE testified that the molecular weight of the recombinant protein was correct. Western blot analysis of GST-TC recombinant protein testified that the recombinant protein could be recognized by immunized rabbit serum, which means the protein is GST-immune active and the clone can express recombinant Clonorchis sinensis antigen. After affinity chromatography of the pET-TC protein, there was only one protein band with expected size on the SDS-PAGE gel. Conclusion The TC gene was screened from cDNA library of adult Clonorchis sinensis, cloned, expressed and purified. The purified protein of TC gene will be of importance for further research on the biological function of the gene.

Cloning and Sequence Analysis of Trichomonas vaginalis Ferredoxin Gene

XIEHui;WANGYa-jing;TIEChao-nan;BIShi-liang;LIUPei-na

2005, 23(1):  6-26. 

Asbtract ( )   PDF (282KB) ( )  

Related Articles | Metrics

Objective To construct a recombinant plasmid containing ferredoxin gene of Trichomonas vaginalis. Methods Total DNA was extracted from Trichomonas vaginslis with Chelex-100 method and used as templates for PCR. Primers were designed based on the published sequence of the ferredoxin gene and used to amplify the Trichomonas vaginalis gene using PCR method. The ferredoxin gene obtained by PCR technique was directionally cloned into plasmid pMD-18T simple vector. The constructed recombinant plasmid was transferred into E.coli JM109. The transformants were screened and identified by PCR and restriction analysis. The DNA sequence of the gene was determined by Sanger’s method. Results The size of amplified ferredoxin gene was 306bp. The correct recombinant plasmid was isolated and confirmed by PCR and restriction analysis. The DNA sequence of cloned gene was the same as the published sequence. Conclusion The ferredoxin gene was successfully amplified and cloned into plasmid pMD-18T simple vector. The cloned ferredoxin gene could be used to produce recombinant protein and for study of its function.

Fluctuation in the Resistance of Plasmodium falciparum to Chloroquine in China

LIUDe-quan;FENGXiao-ping;YANGHeng-lin;LINShi-gan;CHENWen-jiang;YANGPin-fang

2005, 23(1):  7-31. 

Asbtract ( )   PDF (192KB) ( )  

Related Articles | Metrics

Objective To investigate whether chloroquine-resistance of Plasmodium falciparum had changed after stopping or reducing the use of chloroquine as an antimalarial in Hainan and Yunnan provinces. Methods WHO standard in vitro microtest and 4-week in vivo test were used, assays were carried out in different time after stopping or reducing the use of chloroquine. Results In vitro test in Hainan indicated that the rate of chloroquine-resistant P.falciparum was 97.9% in 1981, and dropped to 26.7% in 1997(P<0.01). The mean concentration of chloroquine for complete inhibition of schizont formation was 10.46±7.14 pmol/μl blood in 1981, decreased to 1.63±1.47 pmol/μl blood in 1997(P<0.01). The proportion of samples taken from malaria cases that required high concentration (>6.4 pmol/μl blood ) of chloroquine for complete inhibition of schizont formation was 83.3% in 1981 and only 6.7% in 1997(P<0.01). In the 4-week in vivo test, the rate of chloroquine-resistant P.falciparum decreased from 84.2% in 1981 to 18.4% in 1997(P<0.01). RⅢ cases accounted for 53.1% of the total resistant cases in 1981, and for 14.3% in 1997(P<0.01). In vitro test in Yunnan revealed that the rate of chloroquine-resistant P.falciparum, the mean concentration of chloroquine for complete inhibition of schizont formation and the proportion of samples taken from malaria cases that required>6.4 pmol/μl blood of chloroquine for complete inhibition of schizont formation were 97.4%, 17.2±12.6 pmol/μl blood and 58.9% in 1981 respectively, and dropped to 70.4%(P<0.01), 4.0±3.3 pmol/μl blood(P<0.01) and 16.6%(P<0.01) in 2002 respectively. Conclusion The resistance of P.falciparum to chloroquine declined progressively after its use had been stopped or reduced in Hainan and Yunnan provinces.

Expression and Antigenicity Analysis of p46 000 Antigen from Newborn Larvae of Trichinella spiralis

YUANLi-hong;FUBao-quan;LIUMing-yuan*;GaoFei;ZHANGYa-lan;WUXiu-ping;LINBen-fu;LILian-rui;LUQiang;CHENQi-jun;P.Boireau

2005, 23(1):  8-35. 

Asbtract ( )   PDF (254KB) ( )  

Related Articles | Metrics

Objective To express and analyze p46 000 antigen from newborn larvae of Trichinella spiralis. Methods p46 000 antigen gene was subcloned to the pET28a expression system by PCR. The recombinant transformant was induced by IPTG and the antigenicity was analyzed with ELISA and Western blotting. Results The molecular weight of the expressed protein was about Mr 48 000 . ELISA and Western blotting showed that this recombinant protein could be recognized by T.spiralis infected swine serum and rabbit anti-recombinant protein serum, and the rabbit anti-recombinant protein serum could recognize a Mr 46 000 protein from newborn larvae of T.spiralis. Conclusion p46 000 recombinant antigen from newborn larvae of T.spiralis was expressed which shows a specific antigenicity.

Antigen Analysis of Angiostrongylus cantonensis in Different Developmental Stages

LIHua;CHENXiao-guang*;SHENHao-xian;PENGHong-juan;ZHAOXing-cun

2005, 23(1):  9-39. 

Asbtract ( )   PDF (376KB) ( )  

Related Articles | Metrics

Objective To analyze the difference among antigens of Angiostrongylus cantonensis in different developmental stages and identify dominant diagnostic antigen for angiostrongyliasis. Methods Antigens of A.cantonensis in different developmental stages were analyzed by SDS-PAGE and immunoblot. Results The protein bands of all developmental stages were similar on SDS-PAGE. The Mr 40 000 , 50 000 , 66 000 and 80 000 antigens reacted not only with the sera of rats infected by A.cantonensis but also with the sera of normal rats. The Mr 104 000 antigen could be discerned by sera of rats infected with A.cantonensis for 2 weeks. The Mr 32 000 antigen could be recognized by sera of rats 2 weeks after infection, and the reaction became stronger with the infection continued. Conclusion The Mr 40 000 , 50 000 , 66 000 and 80 000 antigens might result in the unspecific reaction in the immunodiagnosis of angiostrongyliasis using the crude antigen of A.cantonensis. The Mr 104 000 of larva, Mr 33 000 of adult females and Mr 32 000 of the worms might be used as candidate antigens in early diagnosis and epidemiological survey of angiostrongyliasis.


Study on the Biological Characteristics of Permethrin-Resistant andSusceptible Strains of Culex pipiens pallens

SONGFeng-lin;ZHAOTong-yan*;DONGYan-de;LUBao-lin

2005, 23(1):  10-42. 

Asbtract ( )   PDF (152KB) ( )  

Related Articles | Metrics

Objective To compare the biological characteristics of Culex pipiens pallens between permethrin-resistant strain and the susceptible strain. Methods In laboratory, the biological characteristics about bloodsucking, reproduction and development of permethrin-resistant and susceptible strains were observed and recorded. Life table of the experimental populations was constituted. Results The bloodsucking rate of resistant strain was lower than susceptible strain and the difference was significant. The life-span of adults of resistant strain and the developmental duration of eggs, larvae and pupae were longer than susceptible strain and the difference was significant. The death rate of eggs of resistant strain was higher than that of susceptible strain and the difference was significant. The permethrin-resistant strain showed a fitness value of 0.58 relative to the susceptible strain. Conclusion The permethrin resistance of Culex pipiens pallens may result in a disadvantage to its reproduction and development.

Construction of Prokaryotic Expression Vector of the Fusion Gene IFN-α1b/CSPⅡ and Expression in E.coli

CHENHui-hong;YUXin-bing;GAOXing-zheng

2005, 23(1):  11-47. 

Asbtract ( )   PDF (548KB) ( )  

Related Articles | Metrics

Objective To Construct the prokaryotic expression vector of the fusion gene IFN-α1b/CSPⅡ. MethodsIFN-α1b was amplified from the human genomic DNA by PCR and cloned into prokaryotic expression vector pGEX-4T-1. The recombinant plasmid pGEX-4T-1/IFN-α1b was constructed. Circumsporozoite proteinⅡ(CSPⅡ) was amplified from the Plasmodium falciparum genomic DNA by PCR and was cloned into the prokaryotic expression vector pGEX-4T-1. The recombinant plasmid pGEX-4T-1/CSPⅡ was constructed. IFN-α1b was cut from the recombinant plasmid pGEX-4T-1/IFN-α1b digested with BamHⅠ and EcoRⅠ and ligated with the recombinant plasmid pGEX-4T-1/CSPⅡ also digested with BamHⅠ and EcoRⅠ . The recombinant prokaryotic plasmid pGEX-4T-1/IFN-α1b/CSPⅡ was constructed. The fusion gene IFN-α1b/CSPⅡ was expressed in E.coli by IPTG. Results The prokaryotic expression vector pGEX-4T-1/IFN-α1b, pGEX-4T-1/CSPⅡ and pGEX-4T-1/IFN-α1b/CSPⅡ were identified by PCR, enzyme digestion and gene sequencing. The expressed fusion protein/IFN-α1b/CSPⅡ in E.coli was identified by SDS-PAGE and Western blot. Conclusion The prokaryotic expression vector of the fusion gene IFN-α1b/CSPⅡ was successfully constructed, which was then expressed in E.coli .

实验报道

Detection of Pneumocystis carinii DNA in Rats by PCR-ELISA

CHENSheng-xia;JIANGXu-gan;QIUJin-bo;XUHui-juan;SHUAILian-yun

2005, 23(1):  12-50. 

Asbtract ( )   PDF (184KB) ( )  

Related Articles | Metrics

　Objective To establish a PCR-ELISA and evaluate its use in detecting DNA of Pneumocystis carinii(P.c) in rat model. Methods SD rats and Wistar rats were used in the experiment. P.c DNA from rat lung tissue and BALF was amplified by PCR. The amplified products were visualized by ethidium bromide (EB) staining after agarose gel electrophoresis or detected by ELISA. The results were compared with that by Giemsa stain. Results The positive rate in the two species of rats by the two methods was 96.4% and 100% in lung tissue, 96.4% and 100% in BALF, respectively, with no significant difference (P>0.05). Giemsa positive samples were all positive by PCR-ELISA. The negative control group had one positive by ELISA in lung tissue and BALF respectively. Conclusion PCR-ELISA shows a high sensitivity and specificity in detecting the DNA of Pneumocystis carinii, which is a secure and easy use method.