[Seasonal changes of glycogen level in Oncomelania hupensis].

Oncomelania hupensis snails were collected from Qingyi River of Wuhu City from August 2012 to July 2013. Livers and pedal muscles of snails were dissected. Anthrone colorimetric method was used to evaluate the glycogen concentrations of whole-body, liver and muscle. The concentration of whole-body and liver glycogen decreased from September to next June. The whole body glycogen content in female (0.55 microg/mg) and male (0.88 microg/mg) snails was the lowest in June and April, respectively. The mean whole-body glycogen concentration in females and males was 2.99 and 3.39 microg/mg, respectively. Liver glycogen concentration was lowest in May (female = 0.29 microg/mg, male = 0.22 microg/mg), and reached peak level in August (female = 2.49 microg/mg, male = 2.78 microg/mg). The average liver glycogen concentration in female and male snails was 1.09 and 0.89 microg/mg, respectively. The muscle glycogen concentration gradually decreased from February to June, the lowest was found in June (female = 0.25 microg/mg, male = 0.41 microg/mg), and reached peak level in December (female =16.59 microg/mg, male = 10.06 microg/mg). The average muscle glycogen concentration in female and male snails was 799 and 605 microg/mg, respectively. There was a positive linear correlation between whole-body and liver glycogen concentrations (P < 0.05), and both of them had the similar trend in their monthly change. A positive linear correlation was found among whole-body, liver and muscle glycogen concentrations (P < 0.05).

[Extraction and morphological observation of Oncomelania hupensis haemocytes].

Haemocytes were collected from Oncomelania hupensis snails by using tissue disruption and filtration method, stained by Giemsa and methylene blue, respectively, and observed under microscope. Number of haemocytes from one snail was counted. Out of 54 haemocytes, 3 types of cells were found: big round cells with particles, small round cells with oval nuclei and spindle cells with oblong nuclei. The diameter of big round cells with particles and small round cells with oval nuclei was (21.59-31.97) and (13.24-20.77) microm, respectively. Spindle cells with oblong nuclei was about (17.60-25.47) microm x (27.19-30.25) microm. The nucleocytoplasmic ratio of the above three type cells was 0.38, 0.44 and 0.38, and occupied 35.95% (19/54), 12.42% (28/54) and 51.63% (7/54), respectively. About 194 600 haemocytes were filtered from one single snail. It means that this filtration method is an effective one to extract haemocytes from O. hupensis.

[Extraction of agglutinin from Oncomelania hupensis and its haemagglutination activity].

OBJECTIVE: To explore the extraction methods of agglutinin from Oncomelania hupensis snail and study its haemagglutination activity. METHODS: Protein obtained by ammonium sulfate fractionation precipitation with 20%-100% saturation of ammonium sulfate. Its haemagglutination activity was determined by rabbit erythrocytes. The precipitation which could agglutinate rabbit erythrocytes was diluted with 2.5 mg/ml D-galactose, D-fructose, D-glucose, saccharose, maltose and lactose, respectively, and then their haemagglutination activity was tested. Snail agglutinin were incubated at different temperatures (25-90 degrees C) and assayed for agglutinating activity. The effect of pH on the haemagglutination activity was determined by using the PBS buffer at different pH values (3.0-10.0). RESULTS: Oncomelania snail agglutinin exhibited high haemagglutination activity in 20%-40% saturated ammonium sulfate pellet. Lactose and galactose could inhibit the haemagglutination activity of snail agglutinin. The agglutinin showed maximum activity at pH 7.0. In temperature range of 30-70 degrees C, the haemagglutination activity decreased with increasing temperature, and all activity lost beyond 80 degrees C. CONCLUSION: Galactose/lactose specific agglutinin exists in Oncomelania snail, its haemagglutination activity is affected by pH and temperature.

[Isolation and purification of Oncomelania hupensis agglutinin and determination of its molecular weight].

OBJECTIVE: To isolate and purify agglutinin from Oncomelania hupensis snail and determine its molecular weight. METHODS: Agglutinin was preliminarily isolated from snail tissue homogenate by 0%-40% saturated ammonium sulfate, and then successively purified with Sephadex G-75 gel filtration and Sepharose 4B affinity chromatography. Bradford assay was used to determine the protein content. The agglutination activity was determined by rabbit erythrocytes. The purity of agglutinin preparations was assessed by SDS-PAGE. The molecular weight of agglutinin subunit was determined by Sephadex G-75 gel filtration. RESULTS: The specific activity of snail tissue homogenate was 21.74 titer/mg. After ammonium sulfate precipitation, Sephadex G-75 gel filtration and Sepharose 4B affinity chromatography, the specific activity of snail agglutinin from the homogenate solution increased to 61.93 titer/mg, 75.89 titer/mg and 963.86 titer/mg, respectively. SDS-PAGE analysis indicated that snail agglutinin (M, 53,000) was purified by Sephadex G-75 gel filtration and Sepharose 4B chromatography. The molecular weight of the snail agglutinin produced by Sephadex G-75 gel filtration was Mr 78,000. CONCLUSION: Combined use of salt fractionation, gel filtration and affinity chromatography can be efficient for extraction and purification of agglutinin from Oncomelania hupensis species. The snail agglutinin is characterized as mono subunit protein with a molecular weight of Mr 78,000.

[Composition and diversity of Acaroid mites' communities in indoor environments of kindergartens in Wuhu City].

OBJECTIVE: To understand the breeding situation of Acaroid mites in indoor environments of kindergartens in Wuhu City, so as to provide the evidence for its prevention and control. METHODS: From March to June and September to December in 2014, dust samples were collected from 15 kindergartens of 3 ranks every month. Acaroid mites in the samples were isolated, identified and counted. RESULTS: Totally 360 samples were selected, and 169 samples (46.94%) had mite infestation. Pyroglyphidae accounted for the most in the population, with the breeding rate of 45.00%. Totally 18 504 mites were found, and Dermatophagoides farina and Dermatophagoides pteronyssinus were the dominant species, with the composition ratios of 30.21% (5 590 mites) and 17.83% (3 300 mites), respectively. The third-ranked kindergartens were in the most serious situation concerning with 81.67% (98/120) of the breeding rate of Acaroid mites. The differences were statistically significant among the kindergartens with different ranks (F = 6.048, χ2 = 73.523, both P < 0.05). There were no significant differences about the Acaroid mite population among different grades in kindergartens (F = 0.132, χ2 = 2.377, both P > 0.05). CONCLUSION: Acaroid mites in the kindergartens in Wuhu City were in serious condition, and the population and species composition of Acaroid mite community is influenced by the human behavior and indoors environments.

[Experimental study on determination of viability of Schistosoma japonicum cercariae by staining].

OBJECTIVE: To determine the viability of Schistosoma japonicum cercariae by staining. METHODS: Schistosoma japonicum cercariae were stained by 0.4% trypan blue, 0.5% methylene blue-eosin-borax (M.E.B), 0.5% eosin, 0.5% methylene blue and 0.05% neutral red, respectively, for 5 min, then they were observed under a stereoscopic microscope. RESULTS: The dead cercariae were stained in the trypan blue, M.E.B, eosin and neutral red, but unstained in the methylene blue. The vital cercariae were unstained in all the five kinds of dyes. CONCLUSION: The staining methods by using 0.4% trypan blue, 0.5%M.E.B, 0.5% eosin and 0.05% neutral red can be used to determine the viability of S. japonicum cercariae.

Investigations on factors that influence the moving neutralization reaction boundary method for capillary electrophoresis and isoelectric focusing.

We investigated several factors, such as temperature, current intensity (i), time (t) and the product (mA min mm(-2), viz., C mm(-2)) of i and t, etc., that obviously affect the moving neutralization reaction boundary method (MNRBM). The results manifest that the temperature and the product ti have a strong influence on the movement rate of the boundary. The data prove that about 0.6 C mm(-2) (being equivalent to 10 mA min mm(-2)) is a critical point. If the product ti is lower than the critical point, a good quantitative agreement exists between the observed and theoretical values, but if it is higher than the critical point, the agreements are poor. The optimized experimental conditions are: (1) 18-20 degrees C room temperature, (2) 0.6-0.8 mA mm(-2), (3) less than 10 mA min mm(-2), (4) 1% agarose gel, (5) daily prepared solution and gel containing NaOH. The optimized MNRBM is of benefit for the studies on MNRB itself, isoelectric focusing and capillary zone electrophoresis as will be partially shown in this paper.

Improving separation efficiency of capillary zone electrophoresis of tryptophan and phenylalanine with the transient moving chemical reaction boundary method.

A simple and convenient mode--moving chemical reaction boundary method-capillary zone electrophoresis (MCRBM-CZE)--was designed for the enhancement of separating efficiency of CZE. In this mode, the transient MCRBM is used for the on-line pre-treatment of sample. By analyses of tryptophan (Trp) and phenylalanine (Phe) as an example, the experiments by MCRBM-CZE were carried out and further compared with those by normal CZE without the transient MCRBM. The results reveal that by carefully selected appropriate electrolytes, a strong condensation effect can be achieved by using MCRBM-CZE; this effect can greatly improve the separation efficiency, resolution and peak height of Trp and Phe in CZE as compared with those of normal CZE of Trp and Phe. Even if the sample comprises high concentrations of salt, such as 80 mM NaCl (concentration of sodium ion up to 145.6 mM), the same condensation effect can also been observed; this implies obvious significance for biological samples like urine and serum. However, if the electrolytes was chosen inappropriately only a poor compression effect of sample was observed in the MCRBM-CZE runs.

[Relationship between Yangtze River floodplain micro ecological environment and distribution of Oncomelania hupensis snails].

OBJECTIVE: To explore the relationship between the Yangtze River floodplain ecological environment (vegetation, soil, water and light intensity) and the distribution of Oncomelania hupensis snails, so as to provide the evidence for ecological snail control. METHODS: Three regions (the Lu-Gang Bridge, Dragon Nest Lake in the bund, and Dragon Nest lake beach) were selected to investigate the plant characteristics (species, height, coverage, frequency and strain of clusters), soil characteristics (temperature, humidity, light intensity) and pH value. All the results were analyzed statistically with SPSS 18 software. RESULTS: A total of 920 boxes were investigated. The vegetation coverage was 3.7%-63.5%, and the dominant population was Cyperusrotundus L. cluster on the marshland. The soil temperature was 19.0 degrees C-24.0 degrees C, pH 5.0-5.7, and humidity 53%-75%. There were statistical significants in average number of living snails and dead snails among 3 groups (P < 0.05). As the light intensity was strong in summer and weak in winter, the snails were in the back surface in summer and front surface in winter respectively. The average number of living snails was the most near the water. The difference was statistical significant (P < 0.01) among three ranges (0-1 m, 1-3 m and 3-5 m). CONCLUSION: The snail survival and distribution have close relations with micro ecological environment factors, such as vegetation, soil, water and light intensity.

[Seasonal changes of glucose levels in Oncomelania hupensis].

OBJECTIVE: To study the seasonal changes of glucose levels per unit soft tissue of Oncomelania hupensis. METHODS: O. hupensis snails were collected from the beach of the Qingyi River in Wuhu City, Anhui Province from August 2012 to July 2013. They were kept in minus 80 degrees C refrigerator immediately. The male snails were distinguished from female, and their soft tissues were collected separately after crushing their shells. The hexokinase method was used to measure the glucose concentrations, and the results were analyzed statistically. RESULTS: The contents of glucose decreased from March to July. The lowest glucose content in the female was 1.87 µg/mg in March, and that in male was 3.70 µg/mg in July. Both of them increased from August and reached peak levels in September (Female = 57.38 µg/mg, male = 44.39 µg/mg), and then gradually decreased from October to next January and increased in February. CONCLUSION: The contents of glucose have seasonal changes regularly in O. hupensis.

[Experimental study on quick determination of viability of Oncomelania hupensis by staining].

OBJECTIVE: To find out a quick, simple and convenient method of determining the viability of Oncomelania hupensis. METHODS: O. hupensis snails were stained for 30 minutes by 0.05% water soluble dye neutral red, 0.5% methylene blue, red ink, methylene blue-eosin-borax (MEB) and 0.4% trypan blue, respectively. The soft tissue samples of the snails were observed by a stereoscopic microscope after crushing their shells. RESULTS: The vital snails were stained and the dead were unstained in the neutral red. The vital and dead snails were unstained in methylene blue. However, the vital and dead snails were stained in red ink. The partial vital and dead snails were stained in MEB. The vital snails were stained and the partial dead were stained in trypan blue. CONCLUSION: The use of 0.05% water soluble dye neutral red is simple, rapid and accurate in determination of the viability of O. hupensis.

[Lysozyme activities of Oncomelania hupensis in different seasons and ages of snails].

OBJECTIVE: To explore the relationship between the lysozyme activities of Oncomelania hupensis snails among different seasons, and the difference of the lysozyme activities among the different ages of the snails. METHODS: The homogenate soft tissues of O. hupensis were dissolved with Tris-HCl-TritonX-114 buffer and concentrated, and the enzyme was extracted by centrifugation. RESULTS: There were significant differences of lysozyme activities among different seasons (Spring, Summer, Autumn, Winter) with the results of 71.33 +/- 4.16 U/mg, 93.67 +/- 7.10 U/mg, 150.33 +/- 10.01 U/mg, 358 +/- 35.38 U/mg,respectively (F = 144.455, P < 0.01). There were also significant differences of the activities between two age groups (adults and young snails) with the results of 93.67 +/- 7.10 U/mg and 69.33 +/- 5.03 U/mg,respectively (t = 4.845, P < 0.05). CONCLUSION: There are significant differences of the lysozyme activities among different seasons and ages of snails, and the highest lysozyme activity is in winter. The lysozyme activity of adult snails is higher than that of the youth.

[Killing effect of exogenous NO on cercariae of Schistosoma japonicum in vitro].

OBJECTIVE: To explore the killing effects of exogenous NO on the cercariae of Schistosoma japonicum in vitro and the blocking effects of NO inhibitors. METHODS: The cercariae of S. japonicum were collected from naturally infected snails, and then formulated into a 1000 cercariae/ml suspension with RPMI 1640 medium. The relationship between the killing effects and doses of exogenous NO on cercariae were investigated through the suspension with different concentrations of NO generating agents (SNP) and negative control (without SNP). On the other hand, the blocking effects were also investigated through the suspensions being added 2.00 mmol/L SNP and four different NO inhibitors, Hb, FeSO4, L-cyst, L-arg and their combination. Additionally, the appropriate positive(2.00 mmol/L SNP) and negative controls (without NO generating agent) were used. RESULTS: The mortality rates of cercariae were (8.3 +/- 1.1)%, (6.26 +/- 2.3)%, and (9.3 +/- 1.0)% in the SNP 0.06, 0.1 and 0.2 mmol/L groups, respectively, and there were no statistically significant differences between the blank control group and the above-mentioned three groups, respectively (P > 0.05). The mortality rates of cercariae were(23.5 +/- 3.9)%, (46.0 +/- 1.1)%, and (59.4 +/- 0.5)% in the SNP 0.50, 1.00 and 2 mmol/L groups, respectively, and there were statistically significant differences between the blank control group and the above-mentioned groups, respectively (P < 0.05). The mortality rates of cercariae were (30.1 +/- 1.2)%, (45.1 +/- 1.4)%, (31.1 +/- 1.3)%, (34.2 +/- 3.1)%, (47.8 +/- 2.0)%, (49.1 +/- 0.6)%, and (44.2 +/- 0.1)% in the 2.00 mmol/L SNP suspensions which were added of Hb, FeSO4, L-cyst, L-arg, FeSO4+L-cyst, FeSO4+L-arg, FeSO4+L-arg+L-cyst, respectively. Compared with the 2.00 mmol/L SNP suspension, the mortality rates of the above-mentioned groups declined, and there were statistically significant differences (P < 0.05). CONCLUSION: There are killing effects of exogenous NO on the cercariae of S. japonicum in vitro, and four different NO inhibitors, Hb, FeSO4, L-cyst, L-arg and their combinations have some different degrees of blocking effects.

Stacking ionizable analytes in a sample matrix with high salt by a transient moving chemical reaction boundary method in capillary zone electrophoresis.

The paper presents a novel on-line transient moving chemical reaction boundary method (tMCRBM) for simply but efficiently stacking ionizable analytes in high-salt matrix in capillary zone electrophoresis (CZE). The powerful function and stability of the tMCRBM are elucidated with the ionizable test analytes of L-phenylalanine (Phe) and L-tryptophan (Trp) in the matrix with 85.6-165.6 mM sodium ion and further compared with the normal CZE of Phe and Trp samples dissolved in running buffer. The results verify that (1) the on-line tMCRBM mode can evidently increase separation efficiency, peak height, and resolution, (2) with the mode, the analytes in a 28-cm high-salt matrix plug can be stacked successfully and further separated well, (3) the values of relative standard deviation of peak height, peak area, and migrating time range from 3.9% to 6.1%; the results indicate the high stability of the technique of tMCRBM-CZE. The techniques implies obvious potential significance for those ionizable analytes, e.g., protein, peptide, and weak alkaline or acidic compound, in such matrixes as serum, urine, seawater, and wastewater, with high salt, which has a deleterious effect on isotachophoresis (ITP) and especially on electrostacking and field-amplified sample injection (FASI). The mechanism of stacking of zwitterionic analytes in a high-salt matrix by the tMCRBM relies on non-steady-state isoelectric focusing (IEF) but not on transient ITP, electrostacking, and FASI.