Table of Content

30 November 1989, Volume 7 Issue 4

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A PRELIMINARY STUDY ON DIAGNOSIS OF SCHISTOSOMIASIS JAPONICA BY INDIRECT IMMUNOPEROXIDASE ASSAY USING DAUGHTER SPOROCYST-CONTAINING ONCOMELANIA TISSUE SECTIONS

1989, 7(4):  245-248. 

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Schistosoma japonicum daughter sporocysts in Oncomelania tissue sections were, for the first time, used as antigen in the diagnosis of schistosomiasis japonica by indirect immunoperoxidase assay (IIPA-DS). The most strong peroxidase reaction was localized on the tegument of the daughter sporocysts and cercariae, and at some parts of cercariaf parenchyma, when IIPA-DS was carried out. Of 112 sera from proven cases of schisto-Bomiasis japonica, 106 (94.6%) were positive, the range of titre dilution was from 1:1 to 1:160, the geometrie mean of titres was 1:20. IIPA-DS highly coincided with IHA, COP and ELJSA in both sensitivity and strength of reaction. Sera from 101 blood donors and 24 normal rabbits were all negative. No cross reaction with sera from 24 patients with paragonimiasis was observed.

CELL LINE OF HYBRIDOMA SECRETING MONOCLONAL ANTIBODIES AGAINST ECHINOCOCCUS GRANULOSUS ANTIGENS

1989, 7(4):  249-251. 

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In order to produce specific monoclonal antibodies for immunodiagnosis and immu noprophylaxis in hydatid disease, a monoclonal antibody cell line NIA2 secreting specific antibodies was established, through fusion of myeloma cells SP2/0 and spleen cells of BALB/c mice immunized with hydatid fluid from a local (Ningxia) patient and repeated screenings and clonings. The specificity was assayed via ELISA, immunoprecipitating and ultramicroscopy. Antibody titer of 1:2560 was obtamed in the supernatant of tne culture medium by ELISA and a high titcr of 3:163840 in seeded ascitic fluid of mice of the same isolate. Immunoglobulin subclass of the NIA2 secreting antibody was deter-mined to be IgG1 which showed no cross reaction to Cysticercus cellulosae. Cloned cell lines from the hybridomas were found consistent in secreting anti-Echinococcus gronulosus antibody through serial passage in tissue culture for ten months.

OBSERVATION ON THE DEVELOPMENT OF PLASMODIUM FALCIPARUM IN ANOPHELES DIRUS

1989, 7(4):  252-255. 

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HThis paper is a record of our observation on the stages of development of Plasmodium Jalciparum in Anopheles dirus. The malarial parasites were derived from 5 infected pa-tients living in Guizhou Province and used to infect 8 batches of An,dirus. The morpho-logy of various developmental stages studied under light microscope and their average size were as follows: Microgametes were filament-shaped, 13.31 ±2.Z2μm; macrogametes and zygotes oval or round, 4.36 ± 0.59μm and 3.39±0.39μm respectively; ookinetes banana shaped, 13.56 ± 0.80μm× 2.90± 0.48μm; oocysts ovoid or spherical in shape, the smallest one being 7.086μm in equivalate diameter (2 days old) and the largest one-72.60μm (11 day old); slender sporozoites measured 10.625 ±0.82μ× 1.179± 1.3μm. The late sporogonic stage of P. fulciparum was observed with scanning electron mi-croscope. The sporozoite buds developed on the surface of the sporoblast bodies, being round or elliptic or irregular-shaped. The anterior end of sporozoites was truncate and sometimes a micropyle could be seen at a distance 1/3 from the anterior end.A description was given of the different characteristics of the macrogametes and zygotes, together with the arrangement of the pigment granules of oocysts.

COMPARATIVE STUDIES ON SEVERAL BIOCHEMICAL INDICES OF ANOPHELES ANTHROPOPHAGUS AND ANOPHELES SIN ENSIS

1989, 7(4):  256-258. 

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Proteins, sugars and esterase isoenzymes of An. anthropophagus and An. sinensis were compared by IEF and two dimensional gel electrophoresis.The results show that there are some differences in the electrophoretic patterns between An. anthropophagus and An. sinensis. The glycoprotein, lipidprotein, glycolipi-dprotein, protein, polysaccharide and esterase isosnzymES showed 10, O, 6, 14, 2 and 13 bands in An. anthropophagus; 10, l, 5, 16, 3 and 15 in An. sinensis. There exist 234 and 240 polypeptide spots in An. anthropophagus and in An. sinensis, respectively, alto-gether 27.8% of polypeptide spots being different.

CELLULAR RESPONSES ELICITED BY CHALLENGED SCHISTOSOMULA OF SCHISTOSOMA JAPONICUM IN THE SKIN OF NAIVE AND CHRONICALLY INFECTED MICE

1989, 7(4):  259-262. 

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Naive and chronically infected C57BL/6 mice were challenged percutaneously over the ear pinna with Schistosoma japonicum cercariae. After 15 hours, the number of EOS increased significantly in the skin of chronically infected mice. Inflammatory cells aggre-gated in the vicinity of schisto.somula or entrapped intact and disintegrated schistosomula, forming granulocytic micro-abscesses in both groups. Ultrastructure studies revealed that flattened EOS tightly attached to the schistosomulum surface and degranulated. Local tegument damage occurred in the area of attacbment. NEU adherence did not seem to be as intimate as EOS, and degranulation was not seen. The tegument of the attached schis-tosoniulum remained normal. The result suggested that EOS appeared to be the efficient killer cell against skin phase schistosomula of S. japonicum (Figs. 1-6).

HUMORAL AND CELLULAR IMMUNITY IN MICE IMMUNIZED WITH MUTAGENIC NTG-ATTENUATED CERCARIAE OF SCHISTOSOMA JAPONICUM

1989, 7(4):  267-270. 

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Observation was made on humoral and cellular immunity in mice immunized with NTG-attenuated cercariae. Results showed that the specific antibody in serum of immunized mice increased gradually. The titer by ELISA and IFA were more than 1:1280 respectively at week 7 post-immunization with NTG-attenuated cercariae (15 min. pc or 60 min. ip). SDS-PAGE electrophoresis showed that there was an extra-band of IgG with MW. between 50 000 and 40 000 in the serum of immunized mice and there was no obvious difference in IgM bands. It suggested that there were differences in not only the increase of antibody titre, but also the composition of antibody post-immunization. The results also indicated that there were differences in levels of lymphocyte proliferative response to schistosome antigens at different weeks post-immunization. The proliferative response to SEA was greater in immunized mice than in infected mice, especially in lymph node lymphocytes. It suggested that NTG-attenuated cercariae did stimulate mice to produce immunity. The dynamic characteristics of humoral and cellular immunity may play roles in the induction of protective immunity against S. japonicum.

STUDIES ON ANTIBODY-DEPENDENT CELL-MEDIATED CYTOTOXICITY TO SCHISTOSOMA JAPONICUM SCHISTOSOMULA IN MICE

1989, 7(4):  271-275. 

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In the presence of antischistosomular serum(Ab), the adherence of macrophages(Mφ). eosinophils(Eos) and neutrophils(Neu) to and the killing effect on schistosomula of Schistosoma japonicum were studied with a mouse model. In vitro experiment showed that all the three kinds of effectcr cells could adhere to the surface of the schistosomula when opsonized with specific Ab resulting in the significant increase in the percentage of dead schistosomula. When SPA was added to the system, the percentage of schistosomula with adherent cells decreased markedly. It was revealed that the adherence of cells was at least partially through the binding of Fc fragment of IgG to Fc receptor on the cell surfaces. After having been incubated with Ab and/or cells for 18 h in vitro, the schistosomula were inoculated into the peritoneal cavity of mice. The adult recovery rate 6 weeks after inoculation in groups Eos+Ab and Mφ + Ab were significantly lower than that of the control group (P0.01).

FURTHER STUDY ON IN VITRO CULTURE OF THIRD-STAGE LARVAE OF WUCHERERIA BANCROFTI AND BRUGIA MALAYI

1989, 7(4):  276-279. 

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This paper reported on improved in vitro cultute system of human lymphatic filarial larvae. Four culture systems were used. Third-stage larvae of Brugia malayi were best maintained, developing and molted twice in the medium containing modified RPMI-1640 medium, supplemented with 20% newborn calf serum and fauman embrsyonic kidney cell line as feeder layer. This culture system kept larvae alive up to 54 days. Brugia malayl third-stage larvae began to moult on the 8-10th day and again on the 32-36th day; Wuchereria bancrofti third stage larvae grew and developed to the fourth-stage and juvenile and survived to 57 days. They began to moult on the 12-18th days and again on the 32-44th day. This culture system was thought to be useful for studies on morphology and sensitivity to drugs.We also studied several cell-free culture systems. Among them, Best survival, growth and devclopment were obtained in 1:1 mixture of modified RPMI-1640 and TC199 medium supplemented with 20% newborn calf serum. Both Wuchereria bancrofti. and Brugia malayi third-stage larvae grew and developed to the fourth stage larvae and juveniles and survived to 36 days and 42 days respectively. The availability of such culture systems for human filariasis would facilitate studies of biochemistry, immunology, duction of moncclonal antibodies and vaccine.

ASSESSMENT OF INTRACUTANEOUS TEST IN LONGITUDINAL SURVEILLANCE FOR LYMPHATIC FILARIASIS

1989, 7(4):  280-283. 

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In order to evaluate the usefulness of intracutaneous test (IT) in longitudinal sur-veillance of filariasis, two administrative villages selected from Queshan County, Henan Province of China, endemic for Wuchereria bancrofti, were surveyed in 1982, 1983 and 1987 respectively. by IT with antigen FPT derived from Dirofilaria immitis.The result showed that the original level of IT to antigen FPT in the population was consistent with the data of either etiological or entomological investigation before chemotherapy. When the microfilaraemia rate and naturall filarial infection rate of mos-quitoes in a village were high, the positive rate and frequency of strong positivity (skin wheel diamater≥1.3mm) for immediate hypersensitivity reaction would be high too; and the reverse was true. It is suggested that both criteria of IT mentioned above may be useful in assessing endemicity of lymphatic filariasis before mass chemotherapy.The speed of negative conversion of IT in both groups, the previously microfilarae-mic patients and the amicrofilaraemic inhabitants positive to immediate hypersensitivity reaction before chemotherapy, were different, the former being significantly slower than that of the latter after mass and selective chemotherapy with diethylcarbamazine.AU of the three criteria for immediate hypersensitivity, positive rate, frequency of strong positivity and positive conversion rate, decreased gradually after a mass and selective DEC treatment. Until 1987, the 5th year after the chemotherapy, the average positive rate in the two villages dropped to 20.0% from 55.4% in 1982, the frequency of strong positivity to 2.8% from 23.8% in 1982, and the positive conversion rate to 9.7% (1984-1987) form 19.2% (1982-1983). This result indicated that the three criteria of IT reflect not only the status but also the trend of filariasis transmission after chemotherapy in an endemic area, and may be also useful in longitudinal seroepidemiological surveillance of lymphatic filariasis.

OPTIMUM NUMBER OF MIXED PERIPHERAL BLOOD SAMPLES BY MEMBRANE FILTRATION TECHNIQUE FOR MASS BLOOD SURVEY OF FILARIASIS

1989, 7(4):  284-287. 

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The membrane filtration technique has been used widely in the evaluation of effect of control and survey of filariasis. The presem study was made to explore an optimum number of mixed peripheral blood samples and a mathematical model of work load for this method in surveying filariasis. By analysing the correlation between the microfilare-mia rate and the optimum number of mixed peripheral blood samples and applying the heory of Binomial Distribution or Poisson Distribution, the authors reckoned a table for estimating the number of filariasis cases in villages with different microfilarial rates and different population as well as the optimum number of mixed peripheral blood samples.