Table of Content

30 October 2000, Volume 18 Issue 5

Previous Issue    Next Issue

论著

CLONING AND SEQUENCE ANALYSIS OF THE LIGHT CHAIN VARIABLE REGION GENE OF MONOCLONAL ANTI-IDIOTYPIC ANTIBODY NP30 OF SCHISTOSOMA JAPONICUM

SONGXiaotong;FENGZhenqing;QIUZhenning;LIYunqian;YUXiaocong;XIONGYing;YINChangcheng;HUANGHualiang;GUANXiaohong

2000, 18(5):  1-259. 

Asbtract ( )   PDF (278KB) ( )  

Related Articles | Metrics

　Objective] To amplify and sequence the light chain of anti idiotypic monoclonal antibody NP30 of Schistosoma japonicum. [Methods] By comparing the conserved regions at each end of the nucleotide sequences of murine germ line genes enco ding FR1 and FR4 regions of immunoglobulin light chain variable regions, we designed a set of primers for amplification of V L gene. The hybridoma cells secreting anti idiotypic monoclonal antibody NP30 of Schistosoma japonicum were cultured and their genome DNAs were extracted and used as templates for PCR. The PCR product was then cloned into pUC19 vector. The recombinants were sequenced by Sanger′s method. The V L gene was compared with GenBank and published mouse V L genes. [Results] The full length of V L gene was 318 bp. The V L gene was a member of mouse Ig κ light chain subgroup IV and generated from rearrangement of germ line V and Jκ 4 genes. The V L gene sequence has been registered by GenBank(accession No. AF206720). [Conclusion] The obtained V L gene was a potentially functional gene of anti idiotypic monoclonal antibody NP30 of Schistosoma japonicum .


CLONING AND SEQUENCE ANALYSIS OF SSU rRNA GENE OF CUTANEOUS LEISHMANIASIS PATHOGEN FROM XINJIANG OF CHINA

ZHENGXueli**;HUXiaosu;YANGWentian;ZHANGTao;JINGBaoqian

2000, 18(5):  2-262. 

Asbtract ( )   PDF (276KB) ( )  

Related Articles | Metrics

　Objective] By sequencing of SSU rRNA gene cloning from Xinjiang cutaneous leishmaniasis pathogen (XJCLP) to provide evidence for identification of the pathogen. [Methods] By PCR assay with primers R222 and R333, the specific fragment had been produced from SSU rRNA gene of XJCLP , L infantum, L tropica and cloned into pGEM ○[KG-6/7]R T Easy vector .The clones were sequenced by the Sanger dideoxy mediated chain termination method, analysis of SSU rRNA gene sequences from XJCLP, L tropica, L infantum with DNASIS. [Results] Sequence analysis showed that the specific fragment of SSU rRNAgene from XJCLP, L infantum,L tropica , were all 394 bp in length. There were 391 bases identical and three point mutations between the sequences of XJCLP and L tropica , the similarity being 99 2%; 390 bases identical and three point mutations and one insertion /deletion between the sequences of XJCLP and L infantum , the similarity being 99 0%. One insertion/deletion between the sequences of L tropica and L infantum , the similarity being 99 7%. The primary and secondary structures of SSU rRNA gene from XJCLP differed from those of L infantum and L tropica .A retrieval from GenBank confirmed that these 394 bp sequence are new gene sequences. [Conclusion]The primary and secondary structures of SSU rRNA gene from XJCLP, L infantum , L tropica were different. 394 bp sequence from SSU rRNA gene of XJCLP is a new gene sequence.


CLONING AND IDENTIFICATION OF CYTOCHROME P450 RESISTANCE RELATED GENES IN THE MOSQUITO, CULEX PIPIENS PALLENS

ZHUChangliang;LIJianmin;TIANHaisheng;MALei;LIXiulan;WUGuanling

2000, 18(5):  3-268. 

Asbtract ( )   PDF (302KB) ( )  

Related Articles | Metrics

　Objective] To inquire into the relationship between cytochrome P450 and deltamethrin resistance. [Methods] 24 new cDNA sequences encoding cytochrome P450 were amplified respectively from deltamethrin susceptible and resistant strains of Culex pipiens pallens with a pair of degenerate primers according to the conservative amino acid sequences of CYP4 in insects by RT PCR and the Direct Cloning Method, and then were identified by cDNA chip and reverse Northern. [Results] 112 positive clones were obtained, of which 24 were shown to be new sequences encoded for cytochrome P450. They have been lodged in GenBank and were appraised by the Nomenclature Committee of Cytochrome P450, belonging to the subfamily CYP4C,CYP4D,CYP4H and CYP4J in CYP4 family. The hybrid signal values of 6 P450 sequences (NYDS3, NYDS5, NYDR6, NYDR9, NYDR15 and NYDR17) were 3.1～9.7 times higher in the resistance probe than in the susceptible probe, and NYDR17 only reacted with the resistance probe. The result of reverse Northern in NYDR15 was similar to that of cDNA chip. [Conclusion] CYP4 is related to deltamethrin resistance and the specific expression caused by point mutation of cytochrome P450 gene may exist in deltamethrin resistant Cx.pipiens pallens .


STUDIES ON APOPTOSIS AND ITS INDUCTION IN SCHISTOSOMA JAPONICUM

WANGWenshi;LIYonglong

2000, 18(5):  4-271. 

Asbtract ( )   PDF (245KB) ( )  

Related Articles | Metrics

　Objective] To explore the apoptosis and its induction in S.japonicum. [Methods] The eggs and adult cells of S. japonicum were isolated. The adult cells were incubated with H 2O 2, dexamethasone and cyclosporin A, cyclosporin A vehicle solution, respectively in order to initiate the apoptosis. The eggs were treated at 4℃, 37℃ and 42℃, respectively for induction of apoptosis.The occurrence of apoptosis was supervised by agarose gel electrophoresis and in situ end labelling. [Results] DNA ladder band characteristic of apoptosis appeared in the cells of adult worms after incubated with H 2O 2, dexamethasone or cyclosporin A and the eggs treated at 42℃, whereas no DNA ladder band was found in other control groups. In situ end labeling revealed that the positive fluorescence could be seen in the cells incubated with H 2O 2, dexamethasone or cyclosporin A and few nuclei in the control group and cyclosporin A vehicle solution group appeared positive fluorescence. [Conclusion] Apoptosis in the cells of S.japonicum adults and eggs induced by chemical or physical agents was observed.


STUDIES ON IDENTIFICATION OF CIRCUMSPOROZOITE PROTEIN GENOTYPING OF PLASMODIUM VIVAX

HUANGTianyi;WANGXiaoli;LIXueming;HUANGYaming;ZENGFengxiu;CHEYing;ZHANGSimiao;FUWeizhong;ZHANGZaixing;ZHANGGuosengCAIXianzheng;WANGShanqing;WANGGuangze

2000, 18(5):  5-276. 

Asbtract ( )   PDF (506KB) ( )  

Related Articles | Metrics

　Objective] To develop a new method of genotyping circumsporozoite protein (CSP) gene for identification of field isolates of Plasmodium vivax . [Methods] Improved Chelex 100 ion exchange method was used to extract DNA from blood filter paper samples, nested PCR and allele specific PCR techniques, agarose gel electrophoresis analysis and dot/southern blotting probe hybridization were employed for amplification, resolution and identification of the diagnostic fragments. [Results] Using the nest allele specific PCR assay reported here, small amounts of DNA extracted from a piece of blood filter paper sample were amplified which produced three different size ranges of diagnostic bands: 650～770 bp PV species specific band, 170～230 bp diagnostic band for temperate zone family and 588 bp band for PV type 2. The sizes and patterns of the bands produced by the reference strains were consistent with those of designed target sequences. Of 59 examined isolates from 6 provinces of China, 42 temperate zone family strains, 15 tropical zone family strains and two PV type 2 strains were identified. [Conclusion] 1, Three genotype strains of P.vivax mentioned above could be identified by this method with only two rounds of PCR and without probe hybridization. 2, The preliminary results showed that PV type 1 including temperate zone family strains and tropical zone family strains as well as PV type 2 strains are present in China. In addition, another CSP genotype with both sequence characteristics of temperate zone and tropical zone family might also be present in China.


ANALYSIS OF PAGUMOGONIMUS SKRJABINI ANTIGEN AND ITS APPLICATION IN SERODIAGNOSIS

ZHANGXilin;DUANJianhua;WANGYing;KUANGMingshu;HUANGFusheng

2000, 18(5):  6-281. 

Asbtract ( )   PDF (584KB) ( )  

Related Articles | Metrics

　Objective] To analyse the soluble antigens of different developmental stages of Pagumogonimus skrjabini and deve lop a specific and sensitive serodiagnostic method for pagumogonimiasis. [Methods] The soluble antigens of P.skrjabini of various stages were separated by SDS PAGE. The specific antigen of the adult fluke was recognized immunologically by immunoblot assay. The protein bands between 10～30 kDa purified by SDS PAGE and electrophoretic elution were used in dot ELISA. [Results] Using dot ELISA, the soluble antigens of adult were recognized by sera infected with P skrjabini . More reactive bands appeared at 10～30 kDa, but major protein bands were at 22、24 and 26 kDa. However, using sera from patients infected with other trematodes including schistosome and Clonorchis , cross reaction bands appeared within 60 to 90 kDa. When compared with ELISA of crude adult antigens for detecting 28 suspected patients, there was no significant difference between the two methods. The sera of 38 patients with other diseases were also detected by the two tests. No cross reaction occurred with the purified adult antigen dot ELISA while 13 2%(5/38) of the sera cross reacted in ELISA of crude adult antigens. [Conclusion] Dot ELISA using 10～30 kDa antigen might be a specific and sensitive serodiagnostic method for diagnoing pagumogonimiasis.


STUDIES ON PHARMACOKINETICS OF CHLOROQUINE IN MICE INFECTED WITH CHLOROQUINE-RESISTANT STRAIN OF PLASMODIUM BERGHEI

SHIJiong;SONGGuanhong;NIYichang;WANGMingjie

2000, 18(5):  7-285. 

Asbtract ( )   PDF (376KB) ( )  

Related Articles | Metrics

　Objective] To compare the pharmacokinetic differences of chloroquine in normal mice and the mice infected with the N and the RC strains of Plasmodium berghei . [Methods] The concentrations of chloroquine in the plasma of normal mice and the mice infected with the N or the RC strains of P. berghei were analyzed by reverse phase HPLC. The pharmacokinetic parameters were measured with software 3P87. [Results] The t 1/2 β value was significantly lower in the mice infected with the RC strain than in normal mice and the mice infected with the N strain( P <0.05), however, there were no significant differences between the mice infected with the N strain and normal mice. [Conclusion] Elimination of chloroquine in the mice infected with the RC strain of P. berghei speed up significantly comparing with the mice infected with the N strain.


EFFECT OF NIPPOSTRONGYLUS BRASILIENSIS INDUCED ALTERATIONS IN T HELPER CELL SUBSETS ON PLASMODIUM BERGHEI INFECTION IN MICE

XIAONing;TakahishaFURUTA;TakaneKIGUCHI;SomeiKOJIMA

2000, 18(5):  8-290. 

Asbtract ( )   PDF (488KB) ( )  

Related Articles | Metrics

　Objective] To observe the anti P.berghei ability of C57BL/6 mice after infected with Nippostrongylus brasiliensis alterations in T helper cell subsets in the course of Plasmodium infection, and the effect of the alteration on host′s prognoses. [Methods] C57BL/6 mice were infected with Nippostrongylus brasiliensis subcutaneously, 3 wk later, the mice were injected with Plasmodium berghei intraperitoneally. The parasitemia was monitored daily. On days 0,3 and 9, RNA from the spleens of infected mice was prepared for PT PCR to analyse the changes of IFN γ and IL 4 mRNA during the infection course.[Results] Compared with control group, the peak reaching time of parasitemia in the experiment group was delayed, and the bearing capacity of mice to malaria infection and the surviving time increased obviously. The IL 4 level in the experiment group was higher than that in the control group on day 0 of P.berghei infection, but all raised abnormally in both groups at the early stage of the infection; while IFN γ level in the experiment group was higher than that in the control group on day 3 after infection, and then began to reduce to some extent on day 9 of the infection in the experiment group. [Conclusion] T helper cell subsets play an important role in antimalarial immunoregulation in mice.


OBSERVATION ON SPECIFIC IgG4 ANTIBODY OF SCHISTOSOMIASIS PATIENTS BEFORE AND AFTER TREATMENT

FENGZheng;QIULishu;ZHANGYonghong;LIHao

2000, 18(5):  9-292. 

Asbtract ( )   PDF (181KB) ( )  

Related Articles | Metrics

　Objective] To observe the alteration of specific IgG4 antibody of schistosomiasis patients before and after treatment. [Methods] ELISA. [Results] The SEA IgG4 and AWA IgG4 positive rates of 27 schistosomiasis cases were 96 3% and 100%, respectively,their average OD values were 1 62 and 0 72. 6 months post treatment 18 cases were followed up, the positive rates were 94 4% and 100%, respectively, their average OD values were 1 06 and 0 56, respectively. 12 months post treatment all cases were followed up, the positive rates of SEA IgG4 and AWA IgG4 were 96 3% and 92 6%, resepectively ,their average OD values were 0 99 and 0 58, respectively. [Conclusion] No obvious changes were found in the SEA IgG4 and AWA IgG4 positive rates of 27 schistosomiasis cases before and after treatment, whereas the antibody level of specific IgG4 was decreased.


ULTRA-PATHOLOGICAL STUDY ON THE SYNCYTIUM OF SCHISTOSOMA MANSONI EXPOSED TO CYCLOSPORIN A IN VITRO

LIUJianfa;L.H.Chappell

2000, 18(5):  10-295. 

Asbtract ( )   PDF (360KB) ( )  

Related Articles | Metrics

　Objective] To study the ultra pathological changes of syncytium of Schistosoma mansoni after cyclosporin A (CsA) treatment. [Methods] MF1 mice were infected with Schistosoma mansoni cercariae. Six weeks later, the adult worms were recovered by portal vein perfusion. After the worms were exposed to CsA of 20 μg/ml for 24 h, the drug induced damage of the worm surface was observed by SEM and TEM. [Results] Incubation of male and female schistosomes with 20 μg/ml of CsA for 24 h resulted in disruption of the tegument and rupture of the spines. Progressive surface damage and swelling and vacuolization of the tegument led to eventual disruption of the syncytium. [Conclusion]The antischistosomal action of CsA is direct, the syncytium is the main site for CsA attack.


OCCURRENCE OF PAGUMOGONIMUS VEOCULARIS IN FUJIAN PROVINCE

LIYousong;CHENYouzhu;LINChenxing;LUChengjiangYEXiaoping;WUJinyou;LINJinxiang

2000, 18(5):  11-300. 

Asbtract ( )   PDF (451KB) ( )  

Related Articles | Metrics

　Objective]To prove that Fujian Province is also a natural focus of Pagumogonimus veocularis(Pv).[Methods]The adult worms were obtained from a cat fed with Pv metacercariae.[Results]Pv were found in Jianou,Fujian Province.All 1 873 Semisulcospira libertina showed negative.The positive rate of Tricula fujianensis and Erhaia jianouenensis were 0.10%(1/695) and 0.25%(5/2 038), respectively. The main crab host was S.fujianensis.Ps alone and mixed infection with Pv were found in the Sinopotamon ,the infection rates were 36.8%(43/117)and 20.5%(24/117), respectively. The numbers of the metacercariae were 806 and 40, respectively. A cat was infected with 12 metacercariae of Pv , eggs were found in the stool 56 days after infection,and 6 worms were found in the lungs 68 days after infection. [Conclusion]Fujian is one of the natural focus of Pv, cat is the adequate host. The fluke was identified as Pv according to the characteristics of the metacercariae.


实验研究

LABORATORY DIAGNOSIS OF ACANTHAMOEBA KERATITIS

DENGXinguo;LIJiachen;ZHULei

2000, 18(5):  12-304. 

Asbtract ( )   PDF (390KB) ( )  

Related Articles | Metrics

Objective] To find a rapid method for diagnosing Acanthamoeba keratitis and identifing Acanthamoeba . [Methods] 10% potassium hydroxide(KOH) wet mount preparations, Acanthamoeba culture, inverted phase contrast microscopy,and pathological examination using H.E. staining and PAS staining. [Results]Using corneal scrapings and corneal materials obtained from surgery,7 cases and 5 cases of Acanthamoeba keratitis were diagnosed by 10% KOH wet mount preparations. 6 strains of Acanthamoeba were isolated in corneal materials of 6 cases by protozoa culture method. The cysts, trophozoites and pseudopods on the trophozoites of Acanthamoeba were directly observed under the inverted phase contrast microscope. The cysts and trophozoites of Acanthamoeba were seen by H.E. staining and PAS staining with 20 h. [Conclusion] Acanthamoeba keratitis could be rapidly diagnosed by 10% KOH wet mount preparations and inverted phase contrast microscopy. Acanthamoeba organisms could be directly observed and identified under inverted phase contrast microscope.

实验报道

EFFECTS OF LAGENIDIUM GIGANTEUM ON THE ACTIVITIES OF THREE ENZYMES OF CULEX QUINQUEFASCIATUS LARVAE

WUJiahong;BAOHuaien

2000, 18(5):  14-311. 

Asbtract ( )   PDF (440KB) ( )  

Related Articles | Metrics

　Objective] To investigate the possible mechanism underlying the killing of Culex quinquefasciatus larvae by Lagenidium giganteum from the biochemical point of view. [Methods] The activities of alkaline phosphatase(AKP), acid phosphatase(ACP) and esterase(EST) were observed by using calcium cobalt method, lead nitrate method and naphthol method. Results were photomicrographed and analysed quantitatively by image analysis using computer. [Results] The activities of AKP and ACP were decreased while the activities of EST were increased among the infected groups with different infection intensities. [Conclusions] The changes of these enzyme activities might be one of the important mechanisms of action of Lagenidium giganteum against Culex quinquefasciafus larvae.