Table of Content

30 August 2001, Volume 19 Issue 4

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论著

Cloning and Identification of Deltamethrin-Resistance or Susceptibility Associated Genes of Culex pipiens pallens

TIANHai-sheng;ZHUChang-liang*;GAOXiao-hong;MALei;SHENBo;LIXiu-lan;WUGuan-ling

2001, 19(4):  1-197. 

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　Objective To obtain deltamethrin-resistance or susceptibility associated genes of Culex pipiens pallens. \ Methods\ The differentially expressed genes were obtained by suppression subtractive hybridization (SSH), and identified by cDNA microarray and reverse Northern blotting.\ Results\ 523 and 286 clones were selected respectively in the two directional SSH. 155 and 42 genes were respectively expressed 2-3 and >3 times higher in the insecticide-resistant strain than in the susceptible strain; 15 and 9 genes were respectively expressed 2-3 and >3 times higher in the susceptible strain than in the resistant strain. There were 2 genes only expressed in the insecticide-resistant strain. 51 three times differentially expressed clones and 2 specially expressed clones were sequenced. 44 sequences were obtained which belong to 13 new genes. There were 8 over-expressed genes in resistant strain, 7 of which were similar respectively to mitochondrion rRNA gene, 60S ribosomal protein gene, 40S ribosomal protein S4 gene, trypsin gene, chymotrypsin A gene, opsin gene, and 16S ribosomal RNA gene. There were 5 over-expressed genes in susceptible strain, 2 of them being similar with 40S ribosomal protein S29 gene and myosin regulatory light chain 2 gene. In addition, 2 genes specially expressed in resistant strain were similar respectively to glycogen branching enzyme gene and ribosomal protein 46 gene.\ Conclusion\ The differentially expressed genes may be associated with deltamethrin-resistance or susceptibility of Culex pipiens pallens.


Induced Expression of the Variable Region of AMA-1 from Plasmodium falciparum

NIEBen-yong;ZHANGLong-xing;PANWei-qing;QIANFeng

2001, 19(4):  2-200. 

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　Objective To express the variable region of AMA-1 gene from Plasmodium falciparum in Escherichia coli . \ Methods Genomic DNA of FCC1/HN was used as template and the variable region of AMA-1 gene was amplified by polymerase chain reaction(PCR). The PCR products were digested by endonuclease Bam HⅠ and Hin dⅢ, cloned into pBlu2KSP. The nucleotide sequences of the variable region of AMA-1 gene were determined by sequencing. The AMA-1 gene fragment was subcloned into plasmid pQE, expressed in E.coli and induced by IPTG. The fusion product as identified by SDS-PAGE gel electrophoresis and Western blotting were proceeded with anti-AMA-1 sera from rabbit.\ Results The size of the variable region of AMA-1 gene from FCC1/HN was 506 bp and encoded 168 amino acids. On SDS-PAGE gel dyed with Coomassie brilliant blue R250, no specific protein band can be discerned, but Western blotting proceeded with anti-AMA-1 sera from rabbit demonstrated that the specific protein band was about 23.0 kDa.\ Conclusion The variable region of AMA-1 gene from FCC1/HN was able to be expressed in E.coli and analysis of Western blotting demonstrated that the AMA-1 fussion protein contained specific antigenic epitopes.


Study on Molecular Phylogeny of Schistosoma sinensium Based on Nuclear Ribosomal DNA

ZHANGGuang-jun;QIUChi-ping;QIUDong-chuan;CHANGZheng-shan;QINZhi-hui;XIAMing-yi*

2001, 19(4):  3-204. 

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　Objective To determine the phylogenetic relationships between Schistosoma sinensium and other Schistosomatid species using DNA sequence data. Two segments of the nuclear rDNA repeat, the second internal spacer (ITS2) and large subunit (LSU/12S) were selected for sequencing. \ Methods Adult worms stored in 100% methanol were washed 3 times with 0.1×TE (pH8.0) and the genomic DNA was extracted by the GNT-K method. The target regions were amplified by PCR using specific primers. The PCR products were purified before ligation into the plasmid pT-adv (Clontech). Recombinant plasmids were amplified in E.coli (strain TOP10), extracted and purified using routine methods and then sequenced using M13 primers (F/R) on a Licor long-read auto-sequencer. Sequences of related schistosomes were retrieved from GenBank and aligned with our data in the sequence editor ESEE. Gene trees were constructed in PHYLIP (Version 3.6 alpha July, 2 000) and MEGA (version 2.0 beta build 3) using both Maximum Parsimony and Neighbor-Joining methods. For parsimony analysis, all characters were treated as unordered and with equal weights. At least 3 000 cycles of bootstrapping were carried out. For analysis in MEGA, all gap columns were deleted. Schistosomatium douthitti and Trichobilharzia were used as outgroups. \ Results The ITS2 and LSU sequences of Schistosoma sinensium were obtained. The ITS2 sequence of Trichobilharzia sp. was reported here for the first time. \ Conclusion The phylogenetic trees from these data of nuclear rDNA suggested that S.sinensium belongs to the Asian schistosome group. And this species might be an ancient member in the Asian clade.


Construction of Multivalent DNA Vaccine against Schistosoma japonicum

LILiu-zhe;SHIYou-en;JIAGNKan

2001, 19(4):  4-208. 

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　Objective To construct a multivalent DNA vaccine.\ Methods\ The multivalent DNA vaccine candidates pBK Sj26 Sj23,pBK Sj32 Sj23 were constructed based on the plasmids pBluescript Sj26,pBluescript Sj32 and pBluescript Sj23 with three pairs of specific primers using DNA recombinant technique. In the primers, a synthetic linker sequence encoding a peptide was designed,and the antigen genes Sj26 and Sj23,Sj32 and Sj23 were then ligated. After identification, the quadriceps muscle of mice were immunized with the multivalent antigen genes. Four weeks after immunization, the multivalent antigen genes were present in the muscular tissue of mice by PCR.\ Results\ The eukaryotic plasmids including multivalent antigens of S.japonicum were constructed successfully, and the plasmids including multivalent antigen gene could be stably existing in the muscle tissue of mice and the multivalent antigens could be expressed in the muscle tissue cells of mice.\ Conclusion\ A multivalent S.japonicum DNA vaccine has been established.


Aedes albopictus: Cloning and Identification of the Acetylcholinesterase Gene Fragment from the Mosquito

WUMing-wei;ZHANGLing-min;HUANGJiong-lie;ZHOUGuo-li;WUYu;ZHAOShuang-xing

2001, 19(4):  5-212. 

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　Objective To isolate, clone and identify the acetylcholinesterase (AChE) fragment from the mosquito, Aedes albopictus, in relation to exploring mechanism of insecticide resistance. \ Methods\ The genome DNA extracted from the mosquito was used for degenerate polymerase chain reaction (PCR) and the two pairs of oligonucleotides encoding the highly conserved protein sequences were used as primers. The reaction products were cloned to T-vector and transfected into E.coli JM 109. The replicative form DNA of recombinant vector extracted from E.coli JM 109 through alkalilysis was identified by the methods of digestion with EcoRⅠand SalⅠ and PCR. \ Results\ The products of degenerate primers polymerase chain reaction were obtained and the identified clone belongs to the AChE fragment of the mosquito.\ Conclusion\ The clone was identified as the AChE fragment of Aedes albopictus.


Antigen Analysis of Trichomonas vaginalis Trophozoite by SDS-PAGE and Two-Dimensional Gel Electrophoresis

GAOXing-zheng;JosephWingOnTam

2001, 19(4):  6-216. 

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　Objective To analyze soluble antigens of Trichomonas vaginalis. \ Methods\ Soluble antigens of the parasite from a patient suffering from trichomonad vaginitis were analyzed by SDS-PAGE, immunoblotting and two-dimensional gel electrophoresis. \ Results\ A total of 26 distinct protein bands were demonstrated by using 10% resolution gel. Nine of them were main bands, eight with MWs 15-62 kDa, one with MW 97 kDa. By immunoblotting the specific anti-T. vaginalis antibodies raised in rabbit recognized 24 protein bands with 8 main bands in them. Two-dimensional gel electrophoresis revealed up to 43 individual trichomonad polypeptide spots, among which, 9 were main ones. The pI and MWs of these spots were 3.65-5.84 and 27->100 kDa respectively. \ \{Conclusion\ Eight\} protein bands out of 26 soluble antigen bands of the parasite showed high immunogenicity. There were 9 main polypeptide spots in 43 polypeptide spots of the parasite.


Leishmania mexicana: The Circular DNA 1 (CD1) Element Contains Genes Encoding Nucleotide-Binding Protein

WANGJun-yun;YANGYue-tao;BAOYi-fang;QUJing-qi

2001, 19(4):  7-220. 

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　Objective To determine the nucleotide sequence of cloned CD1 fragments from Leishmania mexicana and find ORFs predicted to have protein coding function. \ Methods\ CD1 element was separated by CHEF and recovered by agarase, and the digested CD1 fragments were cloned into pZero vector. Nucleotide sequences were determined by the dideoxy chain termination method with the automatic sequencing system ALF using the M13 universal primers. Sequences were analyzed using GCG-PCGENE computer programs. \ Results\ The sequence with \{4 385\} nucleotides was determined and two ORFs were considered to have protein coding function (encoding nucleotide-binding protein). \ Conclusion\ Genes encoding nucleotide-binding protein were identified from the amplified CD1 element of Leishmania mexicana.


The Role of Anopheles anthropophagus in Malaria Transmission in in Xinyang City of Henan Province

GUZheng-cheng;SHANGLe-yuan;CHENJian-she;ZHENGXiang;SUYu-jie;LIAi-min;LIUHui;LUOMan-zhen;QIANHui-lin;TANGLin-hua

2001, 19(4):  8-224. 

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　Objective To study the role of Anopheles anthropophagus in malaria transmission and transmission threshold so as to provide basis for vector surveillance and malaria control strategy. Methods Parasitological and entomological methods were used in the investigation at 5 villages of Xinyang City, Henan Province. Results From July to August, 1999, 74 febrile cases (10.9% of the total population) were examined. Among them 50 were infected, the incidence in the population of surveyed spots was 7.4%. Active detection was made in another randomly selected two villages and found that the parasite rate in the inhabitants was 2.0%, and the positive rate of IFA was 8.4%. Only vivax malaria was detected. An.anthropophagus and An.sinensis were collected, with An.anthropophagus as the predominant one in human dwellings. The estimated man-biting rate and the human blood index were 4.9388 and 0.7858 respectively. The vectorial capacity of An. anthropophagus was 5.5296. The critical man-biting rate of An.anthropophagus was 0.2407 as calculated by the formula (ma=-rlnP/abPn) according to Macdonald′s model.The local man-biting rate was 20 times higher than that of the critical man-biting rate. Conclusion The results demonstrated that An.anthropophagus is the principal vector in malaria transmission in the area. The findings imply that the critical man-biting rate is of practicable importance in vector surveillance.


防治经验

Study on Schistosomiasis Control Strategy in Ertan Reservoir

GUYong-gang;XIALong-fa;LIZai-wen;ZHAOMing-fu;YANGHuan-yin;LUOQian-yin;XIAWen-hua;FENGQing-yuan

2001, 19(4):  9-228. 

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　Objective To explore means and strategies of preventing the spread of schistosomasis transmission due to the building of Ertan Dam.\ Methods\ To eliminate the infection sources and Oncomelania snails. To install concrete irrigation and piping system of water supply. To encourage the immigrants to build methane-generating tanks and improve sanitary facilities and conditions for families who live near the water-retaining line.\ Results\ \{2 360\} people and 152 cattle were treated for schistosome infection. Mollusciciding and environmental modification were made for eliminating snails at an area of \{3 634 580\} m 2 and \{67 105.5\} m 2 respectively. The length of concrete irrigation and piping system installed was 51.13 and 104.895 km respectively. Methane-generating tanks, water-heating instruments using solar energy and other sanitary facilities were established in \{1 781\} households. After three-year intervention, no infected snails were found and no infected human being, cattle and wild rats were detected.\ Conclusion\ Schistosomiasis control was financially supported since the very beginning of the Ertan Dam project, which provided a condition for sustainable development. Continued surveillance of snails and infection sources should be carried out, which will provide scientific basis for schistosomiasis control in the Three Gorges region as well as other new projects of hydropower and water conservancy in endemic area.


实验报道

Experimental Study on the Pathogenesis of Entamoeba gingivalis

LIUGuang-ying;CHENJin-fu;WENWang-rong;CHENWen-lie;LINLi-qun;HONGHang

2001, 19(4):  10-232. 

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　Objective To study the pathogenesis of Entamoeba gingivalis ( E.g .) and its relation to periodontal diseases.\ Methods Rats were treated with immuno-inhibitor for one week and the neck of incisor teeth of the rats was bound with steel wire. They were randomly divided into three groups: the first group was infected by E.g . in the periodontal tissue, the second group was infected by symbiotic bacteria (s.b.),and the third group was given physiological saline as control.Observation on the periodontal inflammation was made for each group of rats, and the purulent secretion from periodontal abscess was examined for living pathogens.\ Results The incidence of periodontal diseases in rats infected by E.g . was higher than that of symbiotic bacteria group and that of control ( P <0.05), the incidence of periodontal diseases in rats infected by s.b. was higher than that of control group ( P <0.05). Living pathogens were found in the abscess liquid.\ Conclusion E.g . is an opportunistic pathogen, which, together with synergistic symbiotic bacteria, can cause periodontal diseases in hosts with low immunity.


Effect of Experimental Infection with Schistosoma japonicum on the Pregnancy of Mice

WANGYan-nan;MAXi-mei;LIHong;ZHANGXiao-yan;HUANGWen-chang

2001, 19(4):  11-235. 

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　Objective To understand the effect of Schistosoma infection on the gestation in mice.\ Methods Female mice infected with Schistosoma japonicum cercariae, and mated with male mice (uninfected) at 40 d and 100 d post-infection, the changes during pregnant period and the growth of offspring were observed until birth. The serum level of estradiol and progesterone of the infected mice was measured by RIA at oestrus.\ Results The level of estradiol and progesterone, and the pregnant rate were much lower in schistosome infected group than that of the control. The rate of abortion, the mortality of pregnant mice and the death rate due to abortion of infected mice increased significantly. The mortality increased with the time of merging ♀ and ♂mice in one cage prolonged. The body weight and length of the offspring in both infected and control groups were found no significant difference.\ Conclusion The results revealed that schistosome infection may suppress estradiol and progesterone secretion, decrease the rate of pregnancy, and that it may also increase the complications and mortality during the gestation periods.


临床研究

Pathological Changes of Diffuse Pneumocystis carinii Infection in the Liver of an AIDS Patient

GUANXiao-qin;ZHOULi-chun;LIAOXiao-gang;LINXiao;LIYuan-yuan

2001, 19(4):  12-238. 

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　Objective To examine the pathological changes in the liver of an AIDS patient with complicated infection of Pneumocystis carinii(PC). \ Methods\ A liver biopsy was made. The tissue was stained with HE, PAS, Giemsa, GMS, and acid-fast staining, and examined under light microscope and transmission electron microscope. \ Results\ Granulomas (acid-fast negative) in the tissue and numerous pathogens (PAS positive) in hepatic sinusoids were detected. Giemsa and GMS staining and electron microscopy all confirmed that the pathogen was Pneumocystis carinii. \ Conclusion\ The pathological findings revealed a diffuse extrapulmonary infection of Pneumocystis carinii in the patient of AIDS.


调查报告

Investigation on the Prevalence of Human Demodex Among 2 248 Medical Students in Inner Mongolia

HUQun;WANGYan

2001, 19(4):  13-240. 

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　Objective To understand the difference of Demodex prevalence among medical students living and studying together with different classes, sexes and nationalities. \ Methods\ Demodex folliculorum and Demodex brevis were detected by using cellophane tape on the nasolabial grooves. \ Results\ The overall prevalence was 51.5% in 2 248 medical students. Prevalence in different classes and sexes was: freshman 42.6%, sophomore 49.6% and third year students 66.8%; 55.7% in males and 48.7% in females. Prevalence in students with different nationalities was: Mongolian 49.8%, Han 52.8% and other minorities 64.3%. \ Conclusion\ The Demodex prevalence in students of senior classes was higher than those from junior ones (P<0.01), higher in males than in females (P<0.01). Prevalence in Han students was slightly higher than in Mongolian with no statistical difference.