The Dynamics of Hepatic Fibrosis Related to Schistosomiasis and Its Risk Factors in a Cohort of China.

China has had a long history against schistosomiasis japonica. The most serious prognosis of chronic schistosome infection is hepatic fibrosis, which develops into advanced schistosomiasis if the process is not effectively controlled. After a more than seven decades endeavor, China has gained remarkable achievements in schistosomiasis control and achieved transmission control nationwide (infection rate of schistosomes in residents and domestic animals both less than 1%) by 2015. However, new advanced schistosomiasis cases emerge annually in China, even in areas where the transmission of schistosomiasis had been interrupted. In the present study, the residents (>5 years old) in a schistosomiasis endemic village were examined for schistosomiasis every year during 1995-2019 by the modified Kato-Katz thick smear method and/or miracidium hatching technique. Residents who were identified to have an active infection method were treated with praziquantel at a dose of 40 mg/kg body weight. Ultrasonography was carried out to assess the liver morbidity related to schistosomiasis in 1995 and 2019, respectively. The prevalence of schistosomiasis among residents presented a downward trend annually, from 17.89% (175/978) in 1995 to 0 (0/475) in 2019. Among 292 residents who received ultrasound scan both in 1995 and 2019, 141 (48.29%) presented stable liver damage, while liver fibrosis was developed severely in 86 (29.45%) and reversed in 65 (22.26%) residents. Univariate and multivariate analysis showed that anti-fibrosis treatment was the protective factor against schistosomiasis hepatic fibrosis. Males, residents aged 38 and above, fishermen, and people who did not receive anti-fibrosis treatment were groups with higher risk of liver fibrosis development. Our results revealed that although the infection rate of schistosome dropped significantly in endemic areas, liver fibrosis was still developing among some residents, even though they had received deworming treatment. Liver protection/anti-fibrosis treatment should be administered in endemic regions and regions with historically uncontrolled transmission to slow down the deterioration of hepatic fibrosis among patients in schistosomiasis endemic areas.

Effect of integrated control intervention on soil-transmitted helminth infections in Jiangxi province in southeast China.

Soil transmitted helminths (STHs) burden was enormous in China several decades ago, however, rigorous control efforts have been successful with appreciable reduction in diseases burden. Here, we assessed provincial-level data derived from cross sectional surveys, executed in 1989, 2002 and 2014, on the prevalence of STHs among populations in Jiangxi province, China. This study, also, reported STHs integrated control intervention aimed at reducing STHs transmission and worm burden among population at county-level. The intervention strategies included mass drug administration (MDA), health education, improved water supply for drinking, improved sanitary facilities and environmental modification in Guixi municipality. The overall infection rate of STHs in Jiangxi province decreased from 77.7% (1989) to 6.3% (2014), while Ascaris lumbricoides, hookworm and Trichuris trichiura decreased from 71.1%, 17.6% and 17.0% (1989) to 0.9%, 4.7% and 1.0% (2014), respectively. STHs infection rates in female population were higher than male in the three surveys. Reduction in STHs prevalence was observed in all age groups, but the decline was less in higher age group. STHs prevalence in Guixi intervention region indicated remarkable reduction from 31.8% (2006) to 6.1% (2009) (χ2＝255.22, P＜0.01). A. lumbricoides, hookworm and T. trichiura infection rates decreased from 10.4%, 17.0% and 7.1% (2006) to 0.1%, 4.1% and 2.2%, respectively (2009) (X2A.l = 110.23, P<0.01; X2hk = 103.57, P < 0.01; X2T.t = 32.0, P < 0.01). A. lumbricoides infection rate declined the most of all STHs. Following control efforts with integrated control intervention strategies, STHs prevalence in Jiangxi province experienced remarkable trend in decline between 1989 and 2014. Consolidating control efforts with sustained integrated control strategies is, therefore, important to achieving STHs elimination in China.

[Study on immune status of patients with schistosomiasis japonica in Poyang Lake region III Humoral and cellular immune characteristics between Schistosoma japonicum high and low antibody responders].

OBJECTIVE: To explore the immune mechanism of negative results of immune tests of schistosomiasis japonica patients. METHODS: Totally 142 schistosomiasis patients (positive stool examinations) of Poyang Lake region were tested by ELISA method, and the ROC curve was applied to determine the high and low response of the patients. The levels of cellular immunity and cytokines of high and low responders were compared. RESULTS: Totally eight schistosomiasis patients were found as low responders. Besides SWAP-IgA (t = -1.588, P > 0.1), the levels of isotype antibodies were significantly lower in the low responders compared with those in the high responders (t = -14.517 to -2.866, all P < 0.05). In the low responders, the propor- tion of CD3⁺T was increased; and the proportions of CD4⁺T, CD8⁺T, CD4⁺CD25⁺Treg, and the ratio of CD4⁺/CD8⁺ were all de- creased, but all of them were not significant (t = -1.72 to 0.974, all P > 0.05) compared with those in the high responders. The differences of IFN-γ and IL-10 between the high and low responders were both not significant (t = -2.426 to 0.216, all P > 0.05). CONCLUSIONS: There is a significant difference between the high and low responders only in the levels of isotype antibodies. One of the reasons of low response in the immune tests is the much lower antibody level after the antigen-antibody compound is completely formulated.

[Preservation of live eggs of Schistosoma japonicum].

OBJECTIVE: To observe the preservation time and activity of miracidium hatching from schistosome eggs preserved in different solutions, so as to obtain the optimal preservation conditions and then provide quality control products for field application. METHODS: The rectum stool of rabbits infected with Schistosoma japonicum was collected and the coarse fecal residue was removed with a series of sample sieves of 80, 100, 160 and 200 meshes respectively, and then the schistosome eggs were concentrated with the sample sieve of 260 meshes. The concentrated eggs were preserved in 0.9% sodium chloride solution, 1.2% sodium chloride solution, phosphate buffered saline solution (PBS, PH 7.2), 1.0% sucrose solution, and Mili-Q water, respectively, and then were conserved in a 4 °C refrigerator and 15 °C constant temperature incubator, respectively. The preserved eggs were hatched in different time (7-day interval) , the vitality and quantity of the miracidia were observed, and the hatching rates were calculated. RESULTS: Under the condition of 4 °C, the hatching rates of eggs dropped to 0 in 1.0% sucrose solution and 1.2% sodium chloride solution at the 49th and 126th day, respectively, and the hatching rates of eggs in the 0.9% sodium chloride solution and PBS solution dropped to 10% at the 112th day, and the activity of miracidium was weakened since 140th. In the Mili-Q water, the hatching rate dropped less than 10% at the 196th day and the activity of miracidia was weakened since the 280th day. Under the condition of 15 °C, the hatching rate of eggs in different solutions gradually dropped to 0 from the 49th day to 105th day. CONCLUSION: The eggs preserved in Mili-Q water at the temperature of 4 °C can be used as the positive reference for hatching tests within 196 days.

[Survey of epidemic status of principal human parasites in Jiangxi Province in 2014].

OBJECTIVE: To understand and analyze the epidemic status of principal human parasites in Jiangxi Province, so as to provide the evidence for formulating The 13rd Five-Year Project of Parasites Control. METHODS: A survey was performed according to the scheme of The 3rd National Survey of Principal Human Parasites as well as the specific situation of Jiangxi Province. The survey of the soil-transmitted nematodes and intestinal protozoa were performed based on the ecological regions and stratified by economic and geographic situation. The survey of Clonorchis sinensis was performed along with the soil-transmitted nematodes in rural, and was performed by the sample method of random cluster in cities and towns. RESULTS: Totally 23 606 residents of 92 survey sites from 32 counties were surveyed, all of them were tested for the infection of soil-transmitted nematodes, in which 21,569 residents were tested for intestinal protozoa infection, and 1 486 children were tested for eggs of Enterobius vermicularis. Twenty kinds of intestinal parasites were found, with the total infection rate of 9.64%, and 4.296 millions of patients were reckoned in the whole Jiangxi Province. The infection rate of children of E. vermicularis was 13.73%. The infection rates of C. sinensis and intestinal protozoa were 0.58% and 1.42% respectively. The infection rate of Toxoplasma gondii was 7.50%. CONCLUSION: In Jiangxi Province, the infection rates of soil-transmitted nematodes are decreasing but the infection rate of C. sinensis is increasing in local areas, and the control work of parasites still should be strengthened.

[Studies on changes of vegetation and Oncomelania hupensis snails in Poyang Lake after impoundment of Three Gorges Project].

OBJECTIVE: To explore the impact and change regularity of Oncomelania hupensis snails and vegetation in the marshlands of Poyang Lake caused by the impoundment of the Three Gorges Project. METHODS: By using the methods of stratified systematic sampling and pinpoint navigation, the succession of vegetation was observed and the distribution of O. hupensis snails was investigated in a period of 8 years on 4 typical marshlands in Poyang Lake. The regularity of changes in vegetation and 0. hupensis snails was analyzed on the marshlands at high, medium and low elevations, respectively. RESULTS: After the impoundment of Three Gorges Project, the average flooding days of these 4 marshlands in 2004 and in 2008 were reduced of 74.8 and 65.8 days compared with those in 2002, the year just before the impoundment of the project, respectively. The change of spatial distribution pattern and the succession of dominant species of vegetation on the marshlands of Poyang Lake have taken place since 2005, and several vegetations showed a trend of moving toward the center of the lake. The hygrophilous vegetation has degenerated and gradually been replaced by mesophily meadow in the marshland area above 14 m elevation. Carex, Phalaris arundinacea and Polygonum appeared in large area and expanded slowly towards the center of the lake on the marshland below 14 m elevation. O. hupensis snails were mostly found under the Carex association. The tendency of decreasing in density of snails and of migration of snail habitats towards low terrain was observed. CONCLUSIONS: The impoundment of Three Gorges Project has had a significant impact on hydrological regimes, and a new succession of vegetation has taken place in the marshlands of Poyang Lake. O. hupensis snails and the mesophily vegetation show a trend of migrating slowly to the areas of low elevation in the marshlands.

Immune responses result in misdiagnosis of Schistosoma japonicum by immunodiagnosis kits in egg-positive patients living in a low schistosomiasis transmission area of China.

BACKGROUND: In recent field surveys, we failed to detect the presence of specific antibody against Schistosoma japonicum in some egg-positive patients by commonly used immunodiagnostic kits. To find out whether low levels of specific antibody truly exist among egg-positive individuals and elucidate the underlying immune mechanisms, we carried out a cross-sectional epidemiologic study in a S. japonicum low transmission endemic area of Poyang Lake region, China and compared the humoral and cellular immune characteristics between S. japonicum high and low antibody responders. METHODS: Kato-Katz thick smear assay was used to determine the schistosomiasis status of 3,384 participants residing in two Poyang Lake region villages, Jiangxi, China. Among the 142 stool egg-positive participants, we identified low and high S. japonicum antibody responders with soluble egg antigen (SEA) and adult worm antigen (AWA) specific IgG levels by adopting ROC curve analysis. To compare the humoral and cellular immune responses between high and low S. japonicum antibody responders, serum specific antibody levels as well as the percentage of T lymphocyte subpopulation in PMBC, and cell stimulated cytokines (IFN- gamma and interlukin-10) were detected. RESULTS: Eight S. japonicum egg-positive participants were defined as low antibody responders. Although the percentage of CD3⁺T cells in low responders was slightly higher and the percentage of CD4⁺ T cells, CD8⁺ T cells, the ratio of CD4⁺/CD8⁺ and CD4⁺ CD25⁺ Treg cells were lower than those in high responders, the differences between the two groups were not significant (P > 0.05). AWA -stimulated interlukin-10 level was significantly higher in high responders, while other cytokines did not show differences between two groups. For antibody profiles, except AWA specific IgA, significant differences of each antibody isotype between low and high responders were detected (P < 0.05). CONCLUSIONS: Our study confirmed that there are S. japonicum antibody low responders among schistosome egg-positive residents in S. japonicum low-transmission areas in China. Thus, mis-diagnosis using immune-diagnosis kits do exist. Significant differences of responding antibody levels between low and high responders were detected, while no major cellular response changes were observed.

[Development and assessment of Schistosoma japonicum antibody test kit (IHA)].

OBJECTIVE: To develop a Schistosoma japonicum antibody test kit (IHA). METHODS: According to the requirement of Good Manufacturing Practice (GMP) and Quality Control (QC) of drugs, the components of kit as well as its preparation method and procedure were studied, and the test kit was assembled and its diagnostic effect was assessed. RESULTS: The sensitivity and specificity were up to 94.49% and 97.14% when testing the serum samples of chronic schistosomiasis patients and normal serum samples by the kit, respectively. The Youden Index of the Kit was 0.92. The cross reaction rates with paragonimus and trichina were 15.00% and 10.00%, respectively. Compared with ELISA and another IHA kit produced by An'Ji company which were used to test schistosome antibody in residents of endemic areas at the same time, the coincident rates of the developed kit were 93.06% and 92.25%, respectively. CONCLUSION: The IHA kit for Schistosomajaponicum antibody has a high sensitivity and specificity, and has the value of application and popularization in the field.

A novel procedure for precise quantification of Schistosoma japonicum eggs in bovine feces.

Schistosomiasis japonica is a zoonosis with a number of mammalian species acting as reservoir hosts, including water buffaloes which can contribute up to 75% to human transmission in the People's Republic of China. Determining prevalence and intensity of Schistosoma japonicum in mammalian hosts is important for calculating transmission rates and determining environmental contamination. A new procedure, the formalin-ethyl acetate sedimentation-digestion (FEA-SD) technique, for increased visualization of S. japonicum eggs in bovine feces, is described that is an effective technique for identifying and quantifying S. japonicum eggs in fecal samples from naturally infected Chinese water buffaloes and from carabao (water buffalo) in the Philippines. The procedure involves filtration, sedimentation, potassium hydroxide digestion and centrifugation steps prior to microscopy. Bulk debris, including the dense cellulosic material present in bovine feces, often obscures schistosome eggs with the result that prevalence and infection intensity based on direct visualization cannot be made accurately. This technique removes nearly 70% of debris from the fecal samples and renders the remaining debris translucent. It allows improved microscopic visualization of S. japonicum eggs and provides an accurate quantitative method for the estimation of infection in bovines and other ruminant reservoir hosts. We show that the FEA-SD technique could be of considerable value if applied as a surveillance tool for animal reservoirs of S. japonicum, particularly in areas with low to high infection intensity, or where, following control efforts, there is suspected elimination of schistosomiasis japonica.

[Antibody against schistosome and schistosome infection and reinfection].

Antibody against schistosome is related closely to the infection and reinfection of schistosome. This paper reviews the dynamic changes of schistosome-specific antibody under different infection situations, the effect and regulation of antibody to the reinfection, and the applicable value of antibody in diagnosis and effect evaluation of chemotherapy.

[Study on immune status of patients with schistosomiasis japonica in Poyang Lake region II characterization of cellular immune in schistosomiasis patients].

OBJECTIVE: To observe the characterization of cellular immune of schistosomiasis japonica patients in low endemic area of Poyang Lake region. METHODS: The schistosomiasis patients who were positive by stool detections were selected as study subjects, and the individuals who were negative by stool detections in three consecutive years were selected as negative controls. The CD4+T cell, CD8+T cell and CD4+CD25+Treg in venous blood of the study subjects were detected by using flow cytometry. In vitro, cytokine (IFN-gamma and IL-10) production of PBMC in response to soluble egg antigen (SEA) and soluble worm antigen preparation (SWA) were measured by ELISA. RESULTS: The levels of CD4+CD25+Treg in individuals who were positive by stool detections (5.99 +/- 1.60)% were higher than those of the negative (5.04 +/- 1.64)%, and the difference was significant (t = 3.261, P < 0.01). One year after the treatment, the levels of IFN-gamma were significantly higher (Stimulated by SEA before and after the treatment, the mean levels were 121.97 and 2 564.03 pg/ml, respectively; stimulated by SWA before and after treatment, the mean levels were 102.89 and 646.26 pg/ml, respectively) and IL-10 were significant lower in patients who were positive by stool detections (Stimulated by SEA before and after treatment, the mean level were 294.75 and 122.43 pg/ml, respectively; stimulated by SWAP before and after treatment, the mean cytokine levels were 110.10 and 31.85 pg/ml, respectively). CONCLUSION: The increase of induced CD4+CD25+Treg is one of causes that down-regulate cellular immune of human body after infection with Schistosoma japonicum.

[Field application of antibody detection in a low transmission area of Schistosoma japonicum].

OBJECTIVE: To evaluate the field application of IHA and ELISA for schistosomiasis japonica detection at low transmission status. METHODS: 728 and 799 persons were examined by Kato-Katz's method, IHA and ELISA for schistosomiasis in an endemic village in the year of 2008 and 2010, respectively. The results of IHA and ELISA was evaluated in comparison to that of Kato-Katz (27 slides with 3 stool specimens) used as gold standard. RESULTS: The positive rate of Kato-Katz's method, IHA and ELISA were 10.3% (75/728), 40.0% (291/728) and 40.1% (292/728) in 2008, and 3.8% (30/799), 31.5% (252/799) and 40.1% (320/799) in 2010 respectively, in which significant difference was observed for the result between Kato-katz's method (chi2 = 26.92, P < 0.05) and IHA (chi2 = 11.82, P < 0.05). The consistency between the result of antibody detection and that of Kato-Katz's method was poor, lower than 0.2 (P < 0.01). If routine screening diagnosis mode was adopted, namely, population screened with IHA or ELISA first and confirmed with Kato-Katz's method, correlation analysis showed that the positive rate of Kato-Katz's method increased with the number of stool specimens and slides (rIHA2008 = 0.922, rELISA2008 = 0.908, rIHA2010 = 0.749, rELISA = 0.798; P < 0.05) . Those with egg positive but missed by IHA or ELISA mainly were cases with low infection intensity. When EPG < or = 40, the rate of detection ranged from 66.1% (39/59) to 87.0% (20/23) with IHA, and 62.7% (37/59) to 100% (23/23) with ELISA. When EPG > 40, however, all cases could be detected with ELISA, but some missed with IHA. CONCLUSION: In low transmission areas, the determination of target population for chemotherapy should be based on the examination of nine slides per stool specimen by Kato-Katz's method after serological screening.

[Study on immune status of patients with schistosomiasis japonica in Poyang Lake region I characterization of antigen-specific antibody isotype responses to Schistosoma japonicum].

OBJECTIVE: To understand the characterization and levels of antibody isotype responses to soluble egg antigen (SEA) and adult worm antigen (AWA) of Schistosoma japonicum in schistosomiasis patients in Poyang Lake region. METHODS: Antibody isotype IgG, IgG1, IgG2, IgG3, IgG4, IgE, IgM and IgA responses to SEA and AWA in the sera of individuals from endemic and non-endemic areas were determined by indirect ELISA. RESULTS: The levels of antigen-specific antibody isotypes in the sera of patients with egg positive were significantly higher than those of people with egg negative and normal control population from non-endemic area (P < 0.001), and except for anti-SEA IgG2 and anti-AWA IgG4 and IgE antibodies, the levels of antibody isotypes in the sera of people with egg negative in endemic area were significantly higher than those of normal control population from non-endemic area (P < 0.05). In the population with egg positive, children had higher levels of anti-SEA IgM and IgG1 and anti-AWA IgM antibody, and lower AWA specific IgG than adults. There was no significant difference between the male and female except for the male had higher levels of IgG (F = 4.677, P = 0.032) and IgG1 (F = 5.55, P = 0.020)against AWA. Three months post-treatment, the levels of IgG1, IgG3, IgG4 antibody responses to SEA deceased significantly (P < 0.001, P = 0.029, P = 0.044), and almost all the antibody levels decreased significantly (P < 0.05) twelve months post-treatment, except for SEA specific IgE and IgM and AWA specific IgG4 and IgM antibodies. CONCLUSIONS: The levels of antigen-specific antibody isotypes in the sera of patients with egg positive are significantly higher than those of people with egg negative and normal control population from non-endemic area. Almost all of antibody levels decrease significantly twelve months post-treatment. The levels of antibody isotypes in the sera of people with egg negative in endemic area are significantly higher than those of population from non-endemic area, giving a clue that the selection of negative control of schistosomiasis diagnostic kit should be noticed.

[Current endemic situation and control strategy of schistosomiasis in lake and marshland regions in China].

This paper reviewed the current endemic features and situation of schistosomiasis in lake and marshland regions in China, and discussed the comprehensive control strategy for schistosomiasis with emphasis on infectious sources control, including its theoretical basis, the reality conditions of implementation and technical composition, and also introduced the effectiveness of the pilot trails and their large-scale application as well as several issues in the implementation. The aim of the review is to accelerate the process of schistosomiasis control in lake and marshland regions and promote the achievement of mid- and long-term goals in national schistosomiasis control program.

[Preparation and characteristics of Schistosoma japonicum signaling protein 14-3-3].

OBJECTIVE: To prepare soluble recombinant signal transduction protein 14-3-3 of Schistosoma japonicum (rSj14-3-3) and investigate its immunologic features. METHODS: The cDNA fragment of signal transduction protein 14-3-3 gene was prepared from Schistosoma japonicum adult worm mRNA, and was subcloned to the downstream of glutathione S transferase gene of expression vector pGEX-4T-3 to construct a recombinant expression plasmid 14-3-3/pGEX-4T-3. The recombinant plasmids were transferred into E. coli BL21, the transforments containing recombinant plasmid were induced by IPTG, and the expression situation of fusion protein GST-14-3-3 was observed by SDS-PAGE. The pure recombinant 14-3-3 protein was prepared by digesting the fusion protein GST-rSj14-3-3 with thrombin and affinity chromatography. The specific antibody against rSj14-3-3 protein was prepared by an immunized rabbit with rSj14-3-3 protein. The immunogenicity and immune reactivity of rSj14-3-3 protein were analyzed by Western blotting. RESULTS: The gene encoding signal transduction protein 14-3-3 of Schistosoma japonicum Jiangsu strain was cloned successfully, the homology of open read frame sequence to the registrated sequence of Sj14-3-3 protein in genbank was 99.08%. A 55 kilo Dalton fusion protein GST-rSj14-3-3 was expressed by transferring the recombinant plasmid 14-3-3/pGEX-4T-3 into E. coli BL21 and induced with IPTG. The high titer antibody against rSj14-3-3 was produced by the immunized rabbit with rSj14-3-3 ,and could recognize the nature and recombinant rSj14-3-3 proteins. CONCLUSIONS: The soluble rSj14-3-3 protein has been prepared successfully in this study, and it has good immunogenicity and reactivity.

[Diagnostic efficiency of Indirect Hemagglutination Assay Kit for antibody detection of schistosomiasis japonica].

OBJECTIVE: To evaluate the clinical diagnostic efficiency of Indirect Hemagglutination Assay Kit (IHA kit) for antibody detection of schistosomiasis japonica. METHODS: Schistosomiasis examinations by parasitological method were conducted in the schistosomiasis endemic areas in Jiangxi, Anhui and Hubei provinces, respectively. A total of 371 serum samples of schistosomiasis patients and 761 serum samples of non-patients without history of schistosome infection and infested water contact were collected. All the sera were detected by using IHA Kit. The diagnostic efficiency indicators of the kit, including sensitivity, specificity, positive predicted value, negative predicted value etc. were calculated. RESULTS: The sensitivity and specificity of the kit were 95.1% (95% confidence interval was between 92.9% and 97.3%) and 97.8% (95% confidence interval was between 96.8% and 98.8%), respectively; the positive and negative predicted values were 95.4% and 97.6%, respectively. No statistically significant difference was found in the prevalence of IHA positive detecting by the kit among the groups with different EPG levels (< or = 40, 41-100, > 100). Also, no significant difference was observed in sensitivity and specificity among the different age groups. CONCLUSIONS: The IHA kit shows a relative high efficiency for diagnosis and can be used for mass screening of schistosomiasis in field sites.

[Expression and characterization of thioredoxin glutathione reductase of Schistosoma japonicum].

OBJECTIVE: To prepare the recombinant thioredoxin glutathione reductase of Schistosoma japonicum (SjTGR) with biological activity. METHODS: The open reading frame DNA sequence of SjTGR was fused with a bacterial-type selenosysteine insertion sequence (SECIS) element by PCR to form a chimeric gene. The chimeric gene was subcloned into expression plasmid pET-41a to construct a recombinant plasmid SjTGR-pET-41a. Then the recombinant plasmid SjTGR-pET-41a was co-transformed into E. coli BL21 with plasmid pSU ABC. The SjTGR protein was expressed by inducing with IPTG. The recombinant SjTGR was purified from expression products by affinity chromatography with an adenosine 2', 5'- diphosphate agarose column. The polyclonal anti-serum against recombinant SjTGR was obtained by immunizing mice with purified SjTGR. The native TGR in S. japonicum was evidenced by using Western blotting. Thiorendoxin reductase (TrxR) activity, glutathione reductase (GR) activity and gluaredoxin (Grx) activity of recombinant TGR were analyzed according to the biochemical method. RESULTS: The chimeric gene of SjTGR with a bacterial-type selenosysteine insertion sequence (SECIS) element was constructed successfully. The bacteria containing the recombinant plasmid SjTGR-pET-41a could express the soluble SjTGR by inducing with IPTG at static growth stage for 24 h at 24 degrees C. The expressed products of plasmid pSU ABC could promote the integration of selenocysteine and increase the yield of selenoprotein. The result of Western blotting showed that the polyclonal antiserum against recombinant SjTGR could recognize the native TGR in S. japonicum adult worms. The enzymatic assay indicated that SjTGR was a multifunctional enzyme with activities of TrxR, GR and Grx. CONCLUSION: The recombinant SjTGR with biological activity is expressed successfully, which lays the foundation for further study on the function and applied values of SjTGR.

[Evaluation on application of common diagnosis methods for schistosomiasis japonica in endemic areas of China. III. Analysis and evaluation of underestimation of prevalence of Schistosoma japonicum infection by routine Kato-Katz technique].

OBJECTIVE: To understand the rates of underestimation of Schistosoma japonicum infection by Kato-Katz technique with different stool samples and different smears in field site. METHODS: With 27 Kato-Katz thick smears (3 stool samples with 9 thick smears each) for 3 consecutive years as a "gold standard", the rates of underestimation of schistosome infection by Kato-Katz technique with different stool samples and different smears were evaluated in a schistosomiasis endemic village. RESULTS: The examined rates of schistosome infection increased and the rates of underestimation decreased gradually with the increase of the number of Kato-Katz thick smears examined. The rates of underestimation of 3 smears were 40.98%-50.80% and the rates of underestimation of 6 smears were 25.48%-32.39%. The rates of schistosome infection based on the reading of 27 Kato-Katz thick smears were 10.96% in 2008, 8.54% in 2009 and 3.73% in 2010 respectively, but the rates of underestimation of various Kato-Katz thick smears were similar. There were no significant differences among the readings of 3, 6, and 9 smears of 1 stool sample, 3, 6 and 9 smears of 3 stool samples, 6 smears of 2 stool samples, 6 smears of 1 stool sample, and 6 smears of 3 stool samples. CONCLUSIONS: In a community with low-intensity of schistosome infection, the rate of underestimation is big by Kato-Katz technique with the reading of 3 smears of 1 stool sample. It suggested that it is better to increase smears rather than the number of stool samples examined.

[Gene synthesis, expression and characterization of egg protein IPSE of Schistosoma mansoni].

OBJECTIVE: To synthesize and express the gene of egg protein IPSE (IL-4-inducing principle of Schistosoma mansoni eggs) and evaluate its immunologic characteristics. METHODS: The IPSE gene of S. mansoni was synthesized by overlapping PCR, and confirmed by DNA sequencing, The recombinant plasmid IPSE-pET32a(+) was constructed by inserting the gene of IPSE into expression vector pET32a(+) at the downstream of thioredoxin (Trx) coding sequence. The recombinant plasmid IPSE-pET32a(+) was transformed into E. coli BL21(DE3) and followed by expression of the protein induced by IPTG. Large-scale fusion protein was prepared and purified with Ni affinity chromatography gel under denaturing conditions. A small amount of soluble Trx-IPSE was obtained by dialyzing the fusion protein in a large volume of PBS. Western blotting was used to detect if the recombinant IPSE was recognized by the IgG antibody in the pooled patient sera of schistosomiasis japonica and its binding capacity to the non-specific IgE antibody in the sera of healthy persons. RESULTS: DNA sequencing confirmed that the nucleotide sequence of synthesized IPSE gene was completely identical to the native one. SDS-PAGE showed that the recombinant plasmid IPSE/pET32a(+) expressed a fusion protein with an Mr 35700 after being induced by IPTG. The pure fusion protein Trx-IPSE reacted positively with the pooled sera of schistosomiasis patients under either denaturing or renaturing conditions. The protein Trx-IPSE also reacted with the nonspecific IgE in the sera of healthy persons. CONCLUSION: The IPSE gene of Schistosoma mansoni has been synthesized, and the recombinant can react with natural antibody IgG against S. japonicum and non-specifically bind to IgE antibody.

[Comparative evaluation of five test kits for antibody detection in Schistosoma japonicum endemic areas of Poyang Lake region].

OBJECTIVE: To evaluate the efficacy of five diagnostic test kits for the detection of antibodies to Schistosoma japonicum in endemic areas of Poyang Lake. METHODS: Residents over 5 years old in three schistosomiasis-endemic administrative villages in the lake region were simultaneously examined by modified Kato-Katz thick smears and nylon bag sedimentation/hatching method. At the same time, five sero-diagnostic test kits for detection of IgG antibody against soluble egg antigen of S. japonicum were carried out. RESULTS: The distribution of seropositive rates of S. japonicum IgG antibodies determined by sero-diagnostic tests showed similar trends with that of egg positive rates in different age groups, but the seropositive rate (33.1%-57.1%) was significantly higher than that of egg positive rate (5.0%-8.2%), both reached a peak in 40-49-year-old group. The antibody positive rates determined by each test increased with the infection intensity. The sensitivity of the evaluated test kits ranged from 81.2%-94.5%. 26.9% to 46.1% among those egg negatives were antibody positive by the kits. Furthermore, 5.5%-18.8% of the egg positives were antibody negative by the kits. CONCLUSION: The five sero-diagnostic kits can be used for epidemiological survey in schistosomiasis endemic areas at community level. Among them, enzyme-linked immunosorbent assay shows higher sensitivity.