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DETECTION OF PLASMODIUM VIVAX BY NESTED PCR AMPLIFICATION OF DRIED BLOOD SPOTS ON FILTER PAPERS

Chun NIU ; Xinping ZHU ; Lei ZHOU ; Qiang LIU

Acta Parasitologica et Medica Entomologica Sinica.2000;Ⅶ(1):7-10. doi:10.3969/j.issn.1005-0507.2000.01.002

Malaria parasites DNA was eluted from dried blood spot on filter paper by Chelex treatment and amplified by PCR to detect low parasitemia of Plasmodium vivax. Sensitivity and specificity of nested PCR assay were confirmed by amplifying a 121 bp DNA fragment of SSUrRNA gene of Plvivax.Density of about 25 parasites per ul of blood can be detected successfully.None of other three human malaria species(Plasmodium falciparum,Plasmodium malariae and Plasmodium ovale)samples was positive.The ease of collection and transport of filter paper specimens combined with the sensitive and specific detection of P.vivax by nested PCR suggest that this method might be a valuable tool for moleculr epidemiological study of P.vivax.

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PRELIMINARY STUDIES ON MOUSE PLASMODIUM YOELII MODEL FOR THE EVALUATION OF A MULTI-EPITOPE VACCINE CANDIDATE OF PLASMODIUM FALCIPARUM

Wenqi DONG ; Huixang BI ; Ming LI ; Lizhi QU

Acta Parasitologica et Medica Entomologica Sinica.2001;8(1):1-6. doi:10.3969/j.issn.1005-0507.2001.01.001

To find a suitable animal model for the vaccine assessment of Plasmodium falciparum,Plasmodium yoelii and Plasmodium berghei of mouse were used to evaluate the protection for a multi-epitope vaccine candidate of Plasmodium falciparum. Results showed that mice of the group firstly immunized with the protein expressed in E.coli,then boosted with the recombinant vaccinia virus from the same epitopes survived longer than that of groups immunized with the wild vaccinia virus or the blank control (P<0.01).This indicates that Plasmodium yoelii of mouse could be used as a protective model for the evaluation of multi-epitope vaccine candidate of Plasmodium falciparum.

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OBSTRUCTION AND SEQUENCE DETERMINATION OF E.COLI-MYCOBACTERIA RECOMBINANT SHUTTLE PLASMID OF CSP GENE FRAGMENT OF PLASMODIUM FALCIPARUM

Chunfu ZHENG ; Shaoting WU ; Yatang CHEN ; Shitong GAO ; Min LIN

Acta Parasitologica et Medica Entomologica Sinica.2001;8(1):7-12. doi:10.3969/j.issn.1005-0507.2001.01.002

The circumsporozoite protein(CSP) gene fragment of Plasmodium falciparum was cloned and sequenced.A pair of primer was designed according to the CSP encoding sequence of 837 isolate(Thailand isolate),and the CSP gene fragment was amplified by polymerase chain reaction(PCR) from Plasmodium falciparum FCC-1/HN isolate,it spanned the conserved region I.the central immunodominant repeat region\,the variable region behind the repeats and the conserved II region.After purification,the CSP gene fragment was digested with restriction enzyme BamH I and Kpn I,and ligated with the pBCG5.6 digested with the same enzyme.The recombinant pBCG5.6/CSP was transformed into E.coli DH5α.Positive clones were screened and identified by PCR technique and digestion with restriction enzyme.Nucleotide sequence was determined by dideotide chain-termination method.The results indicated that the CSP gene was successfully amplified and cloned,its nucleotide sequence was about 1 171 bp and was in accordance with the expected one.Sequence determination results showed that the cloned gene fragment was the same with the CSP encoding gene.

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ENZYME-LINKED IMMUNOELECTROTRANSFER BLOT ASSAY (EITB)FOR DETECTING IGG AND IGG4 ANTIBODY IN SERUM OF HUMAN NEUROCYSTICERCOSIS

Jing WU ; Yajie LI ; Ping LIU ; Hui WANG

Acta Parasitologica et Medica Entomologica Sinica.2001;8(1):13-18. doi:10.3969/j.issn.1005-0507.2001.01.003

Total IgG and IgG4 subclass antibody were investigated in serum of neurocysticercosis patients who were divided into four groups.The first group consisted of parasitological and clinically confirmed patients before treatment.The second,third and fourth groups comprised treated patients after 1～3,4～6 and 7～9 therapeutic courses respectively(Each therapeutic course includes 10～14 days)*.241 serum samples from these four groups were detected with EITB using lentil-lectin affinity purified antigens for total IgG and IgG4.36 sera from healthy individuals were used as negative control.We also detected 27 sera from patients of echinococcosis and clonorchiosis by the same method.Compared to negative control the total IgG and IgG4 subclass antibody levels in four different groups were respectively 96.3% and 97.5% for the lst group;93.3% and 78.6% for the 2ed;88.0% and 38.0% for the 3rd;86.1% and 13.9% for the 4th.There was no significant difference for the total IgG among these four groups(P>0.01).In contrast,the levels of IgG4 antibodies in the post-treatment patients were lower than that of the before-treatment patients.The positive rate of IgG4 antibody in symptomatic post-treatment patients is 77.0% but in asymptomatic post-treatment patients is only 21.3%(P<0.001).None of the antigens recognized by IgG was unique to the four groups.GP42 and GP24 were the most common bands recognized by many patients for IgG,but in the latter two groups,IgG4 distinctively recognized low molecular weight antigen of 18kD and 13kD.36 sera from healthy individuals were all negative.The positive rate of total IgG antibody in 27 sera from heterologous infections is 7.5%,in contrast,IgG4 antibody of these sera were all negative.These observation confirmed that IgG4 is an important diagnostic parameter for cure of human neurocysticercosis.

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CAPTURING EFFECT OF ARTHROBOTRYS OLIGOSPORA ON THE 3RD LARVAE OF PARASITIC NEMATODES OF SHEEP IN VITRO

Zerong QIN ; Zuoqing MIAO ; Ming KANG ; Gang CHEN ; Meizi LI

Acta Parasitologica et Medica Entomologica Sinica.2001;8(1):19-22. doi:10.3969/j.issn.1005-0507.2001.01.004

The capturing effect of 5 strains of Arthrobotrys oligospora on the 3rd larvae of parasitic nematodes of sheep was described in the present study. The presence of nematode larvae induced fungi to produce trapping loops. Five strains of A.oligospora reduced the nematode larvae,respectively,by 72.6%\,89.0%、80.0%、84.4%、90.2%.

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CLONE AND ANALYSIS ON GENE FRAGMENTS OF NEW MEMBERS OF CYP6 FAMILY FROM GENOME OF AEDES ALBOPICTUS

Yu WU ; Jionglie HUANG ; Guoli ZHOU ; Xiaoxin LIU ; Zuoliang ZHONG

Acta Parasitologica et Medica Entomologica Sinica.2001;8(1):23-30. doi:10.3969/j.issn.1005-0507.2001.01.005

Aim:To obtain new members'gene fragments of CYP6 family from Aedes albopictus. Method: A pair of degenerate primers according to the homologous amino acid sequences of CYP6A1,CYP6D1 in Musca domestica and CYP6E1 in Culex quinquefasciatus was designed and used to amplify CYP6 gene from genomic DNA of sensitive and deltamethrin-resistant strains of Ae.albopictus by PCR technique. Five DNA fragments sized as designed were obtained. These fragments were recombined by T-A ligation, and cloned into JM109.Five positive clones were selected to sequence. Homologous analysis on these deduced amino acid sequences were executed. Results: Five nucleotide sequences were obtained, of which the length is from 236bp to 240bp.The identities of these cloned sequences to other known members of CYP6 are from 23.1% to 53.8%,to CYP3 are from 25.6% to 43.9%,and to CYP4 are from 13.9% to 24.4%. Conclusion:These cloned sequences are gene fragments from new members of CYP6 family.

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DYNAMIC STUDIES OF PATHOGEN VARIETY AND EPIDEMIOLOGY OF HYPODERMIASIS IN THE XING'AN LEAGUE,INNER MONGOLIA

Qingfeng LI

Acta Parasitologica et Medica Entomologica Sinica.2001;8(1):34-40. doi:10.3969/j.issn.1005-0507.2001.01.007

From July in 1997 to March in 2000,the writer undertook epidemiological investigation of hypodermiasis to 483 cattle at 15 villages and examined the seasonal fluctuation of infection of each stadium larvae of hypodermiasis to 307 cattle at 21 villages of five counties in Xing'an League,Inner Mongolia.The results of the investigation are as follows:It concluds that the main pathogenes of hypodermiasis in Xing'an League are H.lineatum maggot and H.bovis maggot,and　H.lineatum maggot is the local superior species.The clinical examination has showed that the infection rate of hypodermiasis in Xing'an League was 80.35%,and the average infection intensity was 2～73.Surveillance of each stadium of hypoderma larvae,infection rate was 50%～100%,infection intensity was 1～193.The unique geological environment,the natural condition and the characteristec of cattle husbandry in Xing'an League are the reasons which make hypoderma prevalence in this area.

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TEM OBSERVATUION ON THE LARVAL CUTICLE LAYER OF ALDRICHINA GRAHAMI (ALDRICH)

Yukun MA ; Cui HU ; Jian HONG ; Jianxiong MIN

Acta Parasitologica et Medica Entomologica Sinica.2001;8(1):41-45. doi:10.3969/j.issn.1005-0507.2001.01.008

The larval cuticle layer of Aldrichina grahami was studied under constant temperatures 15℃,20℃,25℃ and 30℃. The results showed that (1) The largest number of cuticle layers for the first instar larva was eight, the second eleven, but the third variable depending on temperature, for 15℃, 20℃,25℃ and 30℃ the numbers were sixty-nine, seventy-seven, ninety-two and eighty-one.(2) New secreted layer was thicker than the old one, the average thickness of the layer increased as the developing time, after getting the largest thickness the layer got thinner as the time elapsed. (3) Depositive rates of the layer at different constant temperatures were variable. There was not a linear relationship between the depositive rate of the layer and the developing time. It is a useful way to estimate insect daily age by the larval cuticle chitin layers changing, and this can be used in forensic entomology through establishing suitable mathematic model by further study.

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DESCRIPTION OF SIMULIUM (EUSIMULIUM) TAISHANENSE SP.NOV.FROM CHINA (DIPTERA:SIMULIIDAE)

Jian XUE ; Jiyao AN

Acta Parasitologica et Medica Entomologica Sinica.2001;8(1):46-49. doi:10.3969/j.issn.1005-0507.2001.01.009

Description and illustrations of a new species of the genus Simulium Latreille collected from Mount Tai,Shandong Province,China on adult,larval and pupal stages.All the specimens are kept in the Medical Entomology Collection Gallery,Academy of Military Medical Sciences,Beijing.The new species is assigned to the subgenus Eusimulium and closely related to Simulium (Eusimulium) satsumense Takaoka,1976,S.(E.)kuandianensis Chen et Cao,1983,S.(E.)weiningense Chen et Zhang,1997,but it is easily distinguished from three species mentioned above by the shape of genital fork of female,ventral plate,median sclerite of male,larval head spots,cleft and pupal setae and spines of terga and sterna.Holotype ♀,paratype 1,2pupae,7 mature larvae,from Changshou Bridge (500m) (36.3N,117.1E),Mount Tai,Shandong Province,China 23 June,1999.

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CALORIE RESTRICTION RELATED GENE SELECTION IN TRICHOMONAS VAGINALIS BY SUPPRESSION SUBTRACTIVE HYBRIDIZATION

Hong LIU ; Yu-Cai FU ; Ke-Hao ZHANG ; Ming-Yan XU ; Li-Li LUO

Acta Parasitologica et Medica Entomologica Sinica.2005;12(3):129-133. doi:10.3969/j.issn.1005-0507.2005.03.001

That calorie restriction (CR) extends lifespan has been observed in most laboratory species, and the only way to slow down aging in mammals. We established a model of CR in the protist Trichomonas vaginalis. In this system, the cell cycle of organism could be extended by reducing the concentration of glucose. Differences in gene expression are likely to explain the cell cycle extension in CRT. vaginalis. To screen differentially expressed genes in the normal and CRT.vaginalis, suppression subtractive hybridization (SSH) was utilized and 15 differentially expressed clones were isolated.Sequence analysis revealed that 9 were unknown genes which did not match to any previously reported genes, the other 6were homologues of known genes.

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