Enolase is one kind of important glycolytic enzymes which widely exists in most organisms. A number of recent studies confirm that this enzyme has the functions of activating the plasminogen involving in the processes of infection and mi-gration of parasites reducing the immune function of the host as well as preventing parasites from the immune attack of the host. This paper reviews the current research advances in the parasite enolase and explores its potential for diagnosis drug develop-ment and vaccine target of parasitic diseases.

A kind of colloidal blue dye(D-l)used as a staining reagent for immunoassay was first selected from the dyes produced in China in this study. The optimun condition for labelling the dye onto sheep antihuman IgG antibody was explored. The dye-labelled antibody could react and stain with relative antigens. The minimal concentration of human IgG protein could be detected with the labelled dye by Dot Double Antibody Sandwich Assay was 20-10ng/ ml. 61 of 62 sera samples of schistosomiasis japonica were positive,the positive rate was 98.4%. Just 1 out of 30 healthy people's sera sambles was positive,the false positive rate was 3.3%. All of 40 Fasciolopsiasis sera samples were negative. The results were similar to that of Dot-ELISA. The activity of antibody labelled with colloidal dye could be maintained for 1 week in room temperature and be presered in lyophilized condition for a longer period. The assay was less expensive,simple to conduct and required no special equipment. It suggested that the Dot Colloidal Dye Immunoassay(DIA)might have potential value for use in schisto-somiasis endemic areas.

Objectives To express the miracidial antigen from eggs of Schistosoma japonicum (Chinese mainland strain) (SjMP10), and investigate the role of the miracidial antigen during the hepatic granuloma formation of schistosomiasis. Methods A pair of specific primers was designed and synthesized according to the nucleotide sequence of the open reading frame of the miracidial antigen gene. The open reading frame of the miracidial antigen gene was amplified, digested by restrictive enzyme(BamHI, SalI), and cloned directly into the expression plasmid pGEX-4T-3 to construct the recombinant plasmid. The recombinant plasmids were transformed into E. coli BL21(DE3), and induced by IPTG to express the fusion protein of GST-SjMP10. The expressed fusion protein was purified by electric elution method, and its antigenicity was examined by Western blotting and lymphocyte proliferation test. Results The gene of miracidial antigen was cloned into the expression plasmid pGEX-4T-3. After induced by IPTG, the recombinant expressed a fusion protein of GST-SjMP10, with a molecular weight of 39 000 approximately. The purified fusion protein showed proper antigenicity that could be recognized by the sera of rabbits heavily infected by Schistosoma japonicum and could stimulate the proliferation of splenic lymphocytes of infected BALB/c mice. Conclusion The miracidial antigen from eggs of Schistosoma japonicum was expressed successfully.

Objective To observe the effects of road transport on hematological and biochemical parameters in New Zealand rabbits.Methods A total of 12 healthy New Zealand rabbits were selected for 2 h road transport.Blood samples were collected at 0, 24, 48, 72 and 96 h after transport, respectively.White blood cells (WBC), red blood cells (RBC), hemoglobin (HGB), hematocrit (MCV), mean erythrocyte hemoglobin content (MCH), mean erythrocyte hemoglobin concentration (MCHC) and platelets (PLT) were measured using a blood analyzer.Blood alanine aminotransferase (ALT), aspartate aminotransferase (AST), albumin (ALB), total protein (TP), urea nitrogen (UREA), creatinine, uric acid (UA), triglycerides (TG), total cholesterol (COHL), glucose (GLU), hypersensitive C-reactive protein (CRP), α-amylase (AMYL), and creatine kinase (CK) were detected by an automatic biochemical analyzer.Results Compared the parameters before and after transport, The WBC count was increased first (P< 0.05 or P < 0.01) and then decreased after transport, the levels of RBC, HGB, HCT and PLT were decreased first (P< 0.05 or P < 0.01) and then increased after transport, and MCV was significantly high at 96 h after transport (P< 0.05).Among the clinical biochemical parameters, ALT, AST and BUN were firstly elevated (P< 0.05 or P < 0.01) and then decreased.TP, ALB as well as CREA and TG were firstly decreased (P< 0.05 or P < 0.01) and then increased.GLU was significantly low at 24 h after transport (P< 0.05).All parameters except MCV at 96 h after transport were not significantly different from those before transport.Conclusions Changes of blood routine, liver and kidney function indexes, lipid metabolism indexes, glucose metabolism index and creatine kinase index are observed in the New Zealand rabbits after 2-hour road transportation, and all the indicators except MCV return to pre-transport levels within 96 h.

Objective To investigate the value of recombinant fusion protein (GST-HD)of the large hydrophilic domain (HD) of 23 kDa membrane protein of Schistosoma japonicum with the Glu-tathione-S-transferase (GST) of S. japonicum for early diagnosis of schistosorniasis. Methods The rabbits were infected with cercariae of S. japonicum Chinese mainland strain (300 per one). The rabbits' sera before infection and after being infected at different time were collected. The antibodies IgG(s) against recombinant GST-HD and SEA were measured respectively by ELISA to observe the rabbits' immune reaction status to GST-HD and SEA at different time after being infected with the cercariae. At the same time, 90 serum samples of patients with acute schistosorniasis and 30 samples of healthy persons were checked with GST-HD to evaluate its value for early diagnosis of schistosorniasis. Results At the 17th, 21st and 24th day after infection, the positive rates of antibody IgG of rabbits sera against GST-HD were 42.85%, 92.80% and 100.00% respectively, but the positive rates of antibody IgG against SEA were 14. 28% , 50. 00% and 84.60% respectively. The sensitivity of GST-HD for detecting early schistosome infection was higher than that of SEA significantly. The predictive values of positive and negative of GST-HD for detecting acute schistosorniasis was 98. 89% and 96.67%,respectively, and the diagnostic efficacy was 98. 33%. Conclusion The recombinant GST-HD fusion protein has high early diagnostic value for schistosorniasis.

Objective To screen the bacteria and its components which are toxic to Oncomelania hupensis. Methods The samples of Oncomelania hupensis on the point of death and the soil around the snails were collected. The bacteria existing in the snails and soil were isolated and identified by using regular methods. After being fermented, the toxicity of the bacterium components including the ferment supernatant, split products and bacteria to the snail were tested by the toxicity test. Results Totally, 104 strains of bacteria were isolated from the snails and soil samples, which included Gram positive cocci or bacilli, Gram negative cocci or bacilli. The fermenting supernatant, splitting products and 10 strains of bacteria showed different level of toxicity against the snail respectively. All the fermenting supernatant of bacterium PY1-1, PY7-2, PY3-5, PY19-3, PY2-2-2, PY8-2-2, PY16-1 could kill more than 50 percent of snails. Conclusion The bacteria which are poisonous to snails have been isolated that make a good basis for identifying single toxic component against snails.

Objective To evaluate the molluscicidal effect of colistin E on Oncomelania hupensis. Methods The molluscicidal effect of colistin E on O. hupensis was checked by using the immersion method and spraying method. The toxicity of colistin E on fish was observed by using the toxic test. Results The snail mortality of each group was 100% when the snails were immersed in the colistin E solutions at a concentration of 5. 0, 1. 0 g/L for 24, 48 h and 72 h separately. When the snails were immersed in the colistin E solution at a concentration of 0. 5 g/L for 24, 48 h and 72 h, the death rates were 95% , 100% and 100% respectively; when the snails were immersed in the colistin E solution at a concentration of 0. 1 g/L for 24, 48 h and 72 h, the mortality was 90%, 95% and 100% respectively; when the snails were immersed in the colistin E solution at a concentration of 0. 01 g/L for 24, 48 h and 72 h, the mortality was 70%, 86% and 100% respectively. The snail mortality by the spraying method in a dose of 35, 70, 140 mg/m2 of colistin E was 60%, 100% and 100%. The result of toxic test showed that the toxicity of colistin E on fish was low. Conclusion Colistin E is an effective molluscicide, and is worthy of investigation further.

Objective To analyze the full length cDNA sequence of Schistosoma japonicum egg miracidia genes harboring signal sequence.Methods The gene specific primers were designed and synthesized according to S.japonicum egg miracidia cDNA fragment containing signal sequence identified by signal sequence trapping method previously. The 5′and 3′ end cDNA fragments of each egg miracidia cDNA fragment harboring signal sequence were amplified by nest PCR using the first strand cDNA of S.japonicum as the template. The specific PCR fragments were cloned by TA clone method and sequenced. The full length cDNA sequence of each gene with signal sequence was constructed by comparing the cDNA sequence identified with signal sequence trapping method and the 5′ end sequence, the 3′ end sequence and deleting the overlapping fragments. The splicing model between mRNA of signal sequence and one of mature portions of S.japonicum egg miracidia gene was checked by analyzing the genomic DNA sequence structure of some genes with signal sequence. Results The 5′ and 3′ end cDNA fragments of sixteen among thirty cDNA fragment with signal sequence were amplified successfully, and their DNA sequences were determined. The full length cDNA sequences of sixteen egg miracidia genes were obtained by sequence matching and splicing. The results of deduced amino acid analysis found that the signal peptide of gene SjP4001 was the same to the one of SjP1531 and the signal peptide of gene SjP1183 was similar to the one of gene SjP3742. It confirmed that different genes could share the same or similar signal peptide. The data of S.japonicum genomic DNA sequence analysis showed that the S.japonicum could obtain its signal sequence by alternative splicing model or trans-splicing model. Conclusions The full length cDNA sequences of sixteen S.japonicum egg miracidia genes with signal sequence have been defined, it indicated that the S.japonicum egg miracidia genes could get its signal sequence by alternative splicing model or trans-splicing model was found in this study.

Objective:To investigate the influencing factors for synchronous colorectal liver metastasis (synCRLM).Methods:The retrospective case-control study was conducted. The clinicopathological data of 3 172 patients with primary colorectal cancer (CRC) who were admitted to the Affiliated Hospital of Qingdao University from January 2010 to January 2016 were collected. There were 1 946 males and 1 226 females, aged (63±12)years, with a range from 21 to 97 years. Observation indicators: (1) general data analysis; (2) clinicopathological data analysis; (3) analysis of influencing factors for synCRLM. Measurement data with normal distribution were represented as Mean±SD. Count data were represented as absolute numbers. The influencing factors for synCRLM were analyzed after excluding missing data of tumor differentiation degree, tumor diameter, pathological T stage and N stage. Univariate analysis was conducted by chi-square test or Logistic regression model. Multivariate analysis was conducted by Logistic regression model. Results:(1) General data analysis: among the 3 172 patients, cases with age ≤29 years, from 30 to 39 years, from 40 to 49 years, from 50 to 59 years, from 60 to 69 years, from 70 to 79 years, and ≥80 years were 15, 82, 342, 774, 965, 759 and 235, respectively. There were 2 972 patients in Qingdao, 172 cases in Yantai and 28 cases in Weihai. Of the 2 972 patients in Qingdao, there were 422 cases in Shinan District, 658 cases in Shibei District, 457 cases in Huangdao District, 144 cases in Laoshan District, 188 cases in Licang District, 205 cases in Chengyang District, 252 cases in Jimo District, 221 cases in Jiaozhou City, 255 cases in Pingdu City, 170 cases in Laixi City. (2) Clinico-pathological data analysis: among the 3 172 patients, there were 1 639 cases of colon cancer including 972 cases with left colon cancer and 667 cases with right colon cancer, 1 533 cases of rectal cancer. There were 2 981 cases of adenocarcinoma, 165 cases of mucinous adenocarcinoma, 10 cases of signet ring cell carcinoma and 16 cases of other types including carcinoid tumor, squamous carcinoma, tubular adenocarcinoma, etc.There were 162 cases with highly differentiated adenocarcinoma, 5 cases with highly-moderately differentiated adenocarcinoma, 2 338 cases with moderately differentiated adenocarcinoma, 80 cases with moderately-poorly differentiated adeno-carcinoma, 396 cases with poorly differentiated adenocarcinoma and 191 cases missing tumor differentiation data. There were 708 cases with tumor diameter <4 cm, 1 957 cases with tumor diameter ≥4 cm and 507 cases missing tumor diameter data. There were 486 cases in T1 or T2 stage of pathological T stage, 2 169 cases in T3 or T4 stage of pathological T stage and 517 cases missing tumor pathological T staging data. There were 1 563 cases in N0 stage of pathological N staging, 1 062 cases in N1 or N2 stage of pathological N staging and 547 cases missing tumor pathological N staging data. There were 2 895 cases without synCRLM and 277 cases with synCRLM. There were 2 799 cases without diabetes and 373 cases with diabetes. There were 2 931 cases without fatty liver and 241 cases with fatty liver. There were 2 989 cases negative for hepatitis B surface antigen (HBsAg) and 183 cases positive for HBsAg. (3) Analysis of influencing factors for synCRLM. Results of univariate analysis showed that gender, tumor location, tumor differentiation degree, tumor diameter, pathological T stage, fatty liver, HBsAg were related factors for synCRLM in primary colorectal cancer ( χ2=7.400, 7.577, 7.111, 4.513, 12.125, 5.686, 5.919, P<0.05), and neutrophils counts, lymphocytes counts, platelet counts, alanine aminotransferase (ALT), aspartate aminotrans-ferase (AST), total bilirubin, γ-glutamyltransferase (GGT), triacylglycerol (TG), total cholesterol (TC), carcinoembryonic antigen (CEA), and CA19-9 were related factors for synCRLM in primary colorectal cancer ( odds ratio=1.101, 0.807, 1.002, 1.017, 1.023, 1.027, 1.012, 0.686, 1.169, 1.007, 1.004, 95% confidence interval as 1.048-1.156, 0.678-0.960, 1.001-1.004, 1.011-1.024, 1.016-1.031, 1.011-1.044, 1.009-1.015, 0.541-0.869, 1.047-1.306, 1.006-1.008, 1.003-1.004, P<0.05). Results of multivariate analysis showed that cases as male, case with positive HBsAg, AST, GGT, TC, CEA and CA19-9 were independent risk factors for synCRLM in primary colorectal cancer ( odds ratio=1.503, 2.492, 1.018, 1.007, 1.301, 1.005, 1.003, 95% confidence interval as 1.038-2.178, 1.443-4.304, 1.003-1.034, 1.003-1.011, 1.112-1.522, 1.003-1.006, 1.002-1.003, P<0.05), and lymphocytes, ALT and TG were independent protective factors for synCRLM in primary colorectal cancer ( odds ratio=0.777, 0.983, 0.602, 95% confidence interval as 0.608-0.993, 0.966-0.999, 0.421-0.862, P<0.05). Conclusion:Cases as male, case with posotive HBsAg, AST, GGT, TC, CEA and CA19-9 are independent risk factors for synCRLM in primary colorectal cancer, while lymphocytes, ALT and TG are independent protective factors for synCRLM in primary colorectal cancer.

Objective To study the effect of praziquantel on cells within sehistosomal ovum granuloma in the lung of mice.Methods Forty-eight mice were divided into 4 groups.Group A:first,the mice were injected with sehistosomal ova hypodermically in abdomen,and 10 days later,injected with schistosomal ova intravenously in the cauda;Group B:in addition to the injection of schistosomal ova as the same of Group A,the mice were administered with praziquantel[300 mg/(kg·d)]for 3 days,one day before the intravenous injection of the ova;Group C:in addition to the injection of schistosomal ova as the same of Group A,the mice were administered with praziquantel(75 mg/kg,B.i.d.) for 5 days weekly until the mice were sacrificed;Group D:the same as Group C but praziquantel was given to the mice from the 29th after the intravenous injection of the ova.Three mice of each group were sacrificed on the 7th,14th,28th,56th day after the intravenous injection of the ova and the lung tissues were fixed with formalin and the slices were HE stained.Twenty-five to thirty pieces of schistosomal ovum granuloma were examined and the neutrophilic granulocytes,eosinocytes,lymphocytes,fibroblasts and macrophages within the ovum granulomas were counted and the mean numbers of them of each group were calculated and compared.Results Compared with Group A,the mean numbers of neutrophilic granulocytes,eosinocytes and macrophages within the ovum granulomas were decreased significantly,and the extend of the increase of fibroblasts reduced significantly in the three groups administered with praziquantel,and especially in Group C.On the 56th day after the intravenous injection of the ova,the mean numbers of neutrophilic granulocytes,eosinocytes and macrophages decreased by 54.4%、87.0% and 23.1%,and the extend of the increase of fibroblasts reduce by 59.4%,respectively in Group C,compared with Group A.The numbers of lymphocytes did not change very much in 4 groups.Conclusion Praziquantel can restrain inflammatory cells and the hyperplasia of fibroblasts within schistosomal ovum granulomas.