Effects of praziquantel isomers on the proliferation and activation of the LX-2 human hepatic stellate cell line / 中国血吸虫病防治杂志

Objective To compare the effects of levo-praziquantel (L-PZQ) and dextro-praziquantel (D-PZQ) on the proliferation and activation of the human hepatic stellate cell line LX-2 in vitro. Methods LX-2 cells were stimulated with transforming growth factor-β (TGF-β). LX-2 cell proliferation was measured using the CCK-8 assay after 24 h stimulation with 0 to 50 μg/mL concentrations of praziquantel, and the gene and protein expression of type Ⅰ collagen (collagen Ⅰ), type Ⅲ collagen (collagen Ⅲ) and α-smooth muscle actin (α-SMA) was quantified in LX-2 cells using quantitative real-time PCR (qPCR) and Western blotting assays 24 h and 48 h following stimulation with 15 μg/mL praziquantel to detect LX-2 cell activation. Results There were significant differences in the survival rate of LX-2 cells between L-PZQ and D-PZQ treatments at all concentrations (F = 6.119 and 79.180, both P values < 0.05). Either L-PZQ or D-PZQ at a concentration of < 30 μg/mL showed no remarkableeffectsonthe LX-2 cell proliferation (both P values > 0.05), and L-PZQ at a concentration of > 50 μg/mL and D-PZQ at a concentration of > 40 μg/mL inhibited the LX-2 cell proliferation (both P values < 0.05), while D-PZQ at concentrations of 40 μg/mL and 50 μg/mL showed greater inhibition on LX-2 cell proliferation than L-PZQ (t = 3.419 and 8.776, both P values < 0.05). There were significant differences in the collagen Ⅰ, collagen Ⅲ and α-SMA expression in LX-2 cells at both transcriptional (F = 21.55, 79.99 and 46.70, all P values < 0.05) and translational levels (F = 20.12, 30.29 and 32.93, all P values < 0.05) among the blank control group, TGF-β stimulation group, L-PZQ treatment group and D-PZQ treatment group. L-PZQ treatment resulted in remarkable inhibition on collagen Ⅲ and α-SMA gene expression in LX-2 cells (both P values < 0.05); however, the treatment showed no remarkable inhibition collagen Ⅰ gene expression or collagen Ⅰ, collagen Ⅲ or α-SMA protein expression in LX-2 cells (all P values > 0.05). In addition, D-PZQ treatment resulted in significant inhibition on collagen Ⅰ, collagen Ⅲ and α-SMA expression in LX-2 cells at both translational and transcriptional levels (all P values < 0.05), and D-PZQ showed higher inhibition on collagen Ⅰ, collagen Ⅲ and α-SMA gene expression in LX-2 cells than L-PZQ (all P values < 0.05). Conclusions Both L-PZQ and D-PZQ inhibit the proliferation and activation of LX-2 cells, and D-PZQ shows a higher inhibitory activity than L-PZQ.

Molluscicidal activity of the secondary metabolites from Streptomyces nigrogriseolus XD 2-7 against Oncomelania hupensis and its preliminary mechanisms of molluscicidal actions / 中国血吸虫病防治杂志

OBJECTIVE@#To evaluate the storage stability of metabolites from actinomycetes Streptomyces nigrogriseolus XD 2-7 and the mollcuscicidal activity against Oncomelania hupensis in the laboratory, and to preliminarily explore the mechanisms of the molluscicidal activity.@*METHODS@#The fermentation supernatant of S. nigrogriseolus XD 2-7 was prepared and stored at -20, 4 °C and 28 °C without light for 10 d; then, the molluscicidal effect was tested against O. hupensis following immersion for 72 h. The fermentation supernatant was boiled in a 100 °C water bath for 30 min and recovered to room temperature, and then the molluscicidal effect was tested against O. hupensis following immersion for 72 h. The pH values of the fermentation supernatant were adjusted to 4.0, 6.0 and 9.0 with concentrated hydrochloric acid and sodium hydroxide, and the fermentation supernatant was stilled at room temperature for 12 h, with its pH adjusted to 7.0; then, the molluscicidal effect was tested against O. hupensis following immersion for 72 h. The fermentation product of S. nigrogriseolus XD 2-7was isolated and purified four times with macroporous resin, silica gel and octadecylsilane bonded silica gel. The final products were prepared into solutions at concentrations of 10.00, 5.00, 2.50, 1.25 mg/L and 0.63 mg/L, and the molluscicidal effect of the final productswas tested against O. hupensis following immersion for 72 h, while dechlorination water served as blank controls, and 0.10 mg/L niclosamide served as positive control. The adenosine triphosphate (ATP) and adenosine diphosphate (ADP) levels were measured in in O. hupensis soft tissues using high performance liquid chromatography (HPLC) following exposure to the final purified fermentation products of S. nigrogriseolus XD 2-7.@*RESULTS@#After the fermentation supernatant of S. nigrogriseolus XD 2-7 was placed at -20, 4 °C and 28 °C without light for 10 d, immersion in the stock solution and solutions at 10- and 50-fold dilutions for 72 h resulted in a 100% (30/30) O. hupensis mortality. Following boiling at 100 °C for 30 min, immersion in the stock solution and solutions at 10- and 50-fold dilutions for 72 h resulted in a 100.00% (30/30) O. hupensis mortality. Following storage at pH values of 4.0 and 6.0 for 12 h, immersion in the fermentation supernatant of S. nigrogriseolus XD 2-7 for 72 h resulted in a 100.00% (30/30) O. hupensis mortality, and following storage at a pH value of 9.0 for 12 h, immersion in the fermentation supernatant of S. nigrogriseolus XD 2-7 for 72 h resulted in a 33.33% (10/30) O. hupensis mortality (χ2 = 30.000, P < 0.05). The minimum concentration of the final purified fermentation products of S. nigrogriseolus XD 2-7 was 1.25 mg/L for achieving a 100% (30/30) O. hupensis mortality. The ATP level was significantly lower in O. hupensis soft tissues exposed to 0.10 mg/L and 1.00 mg/L of the final purified fermentation products of S. nigrogriseolus XD 2-7 than in controls (F = 7.274, P < 0.05), while no significant difference was detected in the ADP level between the treatment group and controls (F = 2.485, P > 0.05).@*CONCLUSIONS@#The active mollcuscicidal ingredients of the S. nigrogriseolus XD 2-7 metabolites are maintained stably at -20, 4 °C and 28 °C for 10 d, and are heat and acid resistant but not alkali resistant. The metabolites from S. nigrogriseolus XD 2-7 may cause energy metabolism disorders in O. hupensis, leading to O. hupensis death.

Activity of aromatic pyrrole-based compounds against of Schistosoma japonicum cercariae and acute toxicity to fish / 中国血吸虫病防治杂志

Objective To test the activity of aromatic pyrrole-based compounds against cercariae of Schistosoma japonicum and test their acute toxicity to fish. Methods A series of aromatic pyrrole-based compounds were synthesized using 4-benzyl-5-(trifluoromethyl)-1H-pyrrole-3-nitrile as the lead compound. The synthesized compounds were prepared into solutions at concentrations of 10.00, 1.00, 0.10, 0.01 mg/L, and the activity of these solutions against S. japonicum cercariae was tested in 30 min, while 0.10 mg/L and 0.01 mg/L niclosamide solutions served as a positive control and dechlorinated water with 1% dimethyl sulfoxide (DMSO) was used as a negative control, with 10 to 30 cercariae of S. japonicum in each group. In addition, the compounds were prepared into solutions at concentrations of 0.50, 0.25, 0.12, 0.06, 0.03 mg/L, and their toxicity to zebrafish was tested in 72 h, while 0.15 mg/L and 0.30 mg/L niclosamide solutions served as a positive control and dechlorinated water with 1% DMSO was used as a negative control, with 10 zebrafishes in each group. Results A total of 7 aromatic pyrrole-based compounds were successfully synthesized. Treatment with compounds 102, 104 and 106 at a concentration of 0.01 mg/L for 30 min killed all S. japonicum cercariae, and compounds 105 and 107 showed no activity against cercariae. No death of cercariae was found in the blank control group, while treatment with 0.10 mg/L niclosamide for 10 min caused a 100% mortality rate of S. japonicum cercariae and 0.01 mg/L niclosamide failed to kill S. japonicum cercariae. No zebrafish death was found 72 h post-treatment with compounds 101, 104 and 105 at a concentration of 0.03 mg/L, and exposure to compounds 102, 103 and 106 at a concentration of 0.03 mg/L for 12 h resulted in a 100% mortality rate of zebrafish. No zebrafish death occurred 72 h post-treatment with 0.50 mg/L Compound 104, and no zebrafish death was found in the blank control group, while treatment with 0.30 mg/L niclosamide for 24 h resulted in a 100% mortality rate of zebrafish. Conclusions Compound 104 achieves a 100% mortality rate against S. japonicum cercariae at a concentration of 0.01 mg/L for 30 min, and causes no death of zebrafish at a concentration of 0.50 mg/L for 72 h, which may serve as a cercaricide candidate.

Studies on resistance of Schistosoma to praziquantel XVIII Sensitivity to praziquantel in filial generations of praziquantel-resistant and -sensitive Schistosoma japonicum mixed infections / 中国血吸虫病防治杂志

Objective To investigate the sensitivity of adult worms of filial generations from praziquantel-resistant and -sensitive Schistosoma japonicum mixed infections to praziquantel. Methods Mice were infected with the cercariae of an experimentally generated praziquantel-resistant S. japonicum isolate [median effective dose (ED50) = 277.4 mg/kg] and a laboratory-maintained praziquantel-sensitive S. japonicum isolate (ED50 = 99.6 mg/kg) at a mixture ratio of 1:1 and 2:1, which was maintained in the laboratory via the mouse-snail cycle for 8 generations. Then, mice were infected with the cercariae of the 8th filial-generation parasite, and grouped 35 days post-infection. Mice in the 5 treatment groups were given praziquantel treatment by gavage at a single oral dose of 37.5, 75, 150, 300 mg/kg and 600 mg/kg, while animals in the control group was administered orally with 2.5% cremophor EL. All mice were sacrificed 14 days post-treatment and adult worms were collected by perfusion of the portal vein. The worm burden reductions and praziquantel ED50 values were calculated. The praziquantel-resistant S. japonicum isolate generated from experimental induction with 12 rounds of praziquantel treatment with sub-curative doses was maintained in the laboratory via the mouse-snail cycle, and mice were infected with the cercariae of the 8th filial-generation parasite. The praziquantel ED50 value against the 8th filial-generation adults was measured. Results After mice were infected with the mixture of cercariae of PZQ-resistant and -sensitive S. japonicum isolates at a ratio of 1:1, the praziquantel ED50 was 135.2 mg/kg against the adults of the 8th filial-generation parasite. After mice were infected with the mixture of cercariae of PZQ-resistant and -sensitive S. japonicum isolates at a ratio of 2:1, the praziquantel ED50 was 129.2 mg/kg against the adults of the 8th filial-generation parasite. In addition, the praziquantel ED50 was 208.4 mg/kg against the adults of the 8th filial-generation S. japonicum without the selection pressure of praziquantel. Conclusions Compared with the experimentally induced praziquantel-resistant S. japonicum isolate, the adult worms of the filial-generation S. japonicum show a reduced sensitivity to praziquantel in the same host following infection with the mixture of cercariae of praziquantel-resistant and -sensitive S. japonicum isolates. The adult worms of the filial generation of the praziquantel-resistant S. japonicum isolate without the selection pressure of praziquantel may still maintain the resistance to praziquantel.

Study on the factors affecting the degradation of niclosamide in the soil / 中国血吸虫病防治杂志

ObjectiveTo investigate the factors affecting the degradation of niclosamide in the soil, so as to provide the evidence for the assessment of the environmental safety in the field snail control with niclosamide. MethodsA high performance liquid chromatography was established for the determination of niclosamide in the field. Then, the degradation of niclosamide was investigated in soils with different moistures (10%, 30%, 50%, 70% and 90%), temperatures [(15 ± 1), (25 ± 1), (35 ± 1) °C], initial concentrations (1, 5, 10 mg/kg) and in sterilized and non-sterilized soils. In addition, the degradation of niclosamide was fitted with the first-order kinetics equation, and the degradation half-life was calculated. Results The niclosamide residues gradually decreased over time in soils with different moistures, and a higher rate of degradation was seen in soils with a higher moisture. The degradation half-life of niclosamide reduced from 4.258 d in the soil with a 10% moisture to 2.412 d in the soil with a 90% moisture. The niclosamide residues gradually decreased over time in soils with different temperatures, and a higher rate of degradation was seen in soils with a higher temperature. The degradation half-life of niclosamide reduced from 4.398 d in the soil with a temperature of (15 ± 1) °C to 2.828 d in the soil with a temperature of (35 ± 1) °C. The degradation half-lives of niclosamide were 3.212, 3.333 d and 3.448 d in soils containing niclosamide at initial concentrations of 1, 5 mg/kg and 10 mg/kg, and > 30 d and 3.273 d in sterilized and non-sterilized soils. Multiple linear regression analysis revealed that soil microorganisms (P = 0.010), moisture (P = 0.000) and temperature (P = 0.002) affected the half-life of niclosamide degradation. Conclusions The degradation of niclosamide in soils fits the first-order kinetics equation, and presence of microorganisms, a high temperature and high moisture may accelerate the degradation of niclosamide in the soil.

[Study on biological characteristics: reproduction and viability of Biomphalaria glabrata as an intermediate host of Schistosoma mansoni].

OBJECTIVE: To compare the difference of fertility of Biomphalaria glabrata snails between self-fertilization and cross-fertilization and to observe the circadian rhythm of laying eggs, the effect of light on laying eggs and the tolerance of the snail to water and food deficiency, so as to provide the evidence for control and elimination of B. glabrata snails in the field. METHODS: Under laboratory conditions, a single B. glabrata egg for self-fertilization was separated and hatched individually, and young snails were raised in different plastic boxes individually. The eggs for cross-fertilization were hatched and the young snails were fed in the same plastic box. The ability of spawn, the development of the eggs, and the number of snails growing from young to adult snails were compared between the self-fertilization and cross-fertilization. The snails were in the water under four environments, all day illumination, all day without illumination, daytime lighting and night without illumination, and daytime without illumination but night lighting. The eggs were collected and counted daily. The circadian rhythm of spawn and the effect of illumination on spawn were observed. The adult snails were divided into 6 groups and exposed to the environments with relative humidity of 0, 65%, 87% and 100%, respectively. The survival rates of the adult snails exposed to the different environments after different time were observed. The adult snails were placed at 25 °C in the oven to remove water content from the soft body of snails. When the dehydration rates of the soft bodies achieved 10%, 20%, 30%, 40%, 50%, 52%, 55%, 57%, 60%, and 70% respectively, the survival rates of the adult snails exposed to the oven were observed. RESULTS: In the 25 °C water, the average laying egg number for 15 days per snail was (8.77 ± 16.92) eggs/snail in the self-fertilization snail. The average laying egg number for 15 days per snail was (149.71 ± 142.28) eggs/snail in the cross-fertilization snails. There was a significant difference between the self-fertilization snail and cross-fertilization snail (t = 0.999 999, P < 0.01). The hatching rate and reproductive maturation rate of the self-fertilization snails and cross-fertilization snails were 50.1% and 78.9%, and 19.3% and 3.8%, respectively, There was a significant difference (the hatching rate: χ2 = 18.18, P < 0.01, the reproductive maturation rate: χ2 = 11.83, P < 0.01) . In the natural environment of daytime with illumination and nighttime with darkness, the amount of laying 20 eggs of B. glabrata snail was (944.07 ± 392.53) eggs/day during a whole day, among them the amount of laying eggs during daytime account for 10.1% and the amount of laying eggs during nighttime account for 89.9%, and the laying egg was given priority to with the night. The above results suggested that the dark environment was conducive to B. glabrata snails to lay eggs. The above results suggested that light can promote the increase of spawning of B. glabrata. When B. glabrata was exposed to the environments with the relative humidity of 0, 65%, 87% and 100% at 25 °C, respectively, and the longest survival times of snails were 7, 70, 150 d and 100 d, respectively. In the 25 °C water, the snails could survive for 50 days without food. The adult snails were placed at 25 °C in the oven to remove water content from the soft body of snails. When the dehydration rates of the soft bodies achieved 10%, 20%, 30%, 40%, 50%, 52%, 55%, 57%, 60%, and 70% respectively, the survival rates of the adult snails exposed to the oven were 100%, 100%, 100%, 100%, 70%, 30%, 0, 0, 0 and 0, respectively. CONCLUSIONS: B. glabrata can achieve the reproductive process by cross-fertilization or self-fertilization. There is a significant difference in reproductive ability between the cross-fertilization snail and self-fertilization snail, cross-fertilization is stronger than self-fertilization, but the rate of reproduction in the self-fertilization is higher than that in the cross-fertilization. It is indicated that B. glabrata that survive after the dry season plays an important role in the maintenance of local snail populations and transmission of schistosomiasis mansoni.

Study on biological characteristics: reproduction and viability of Biomphalaria glabrata as an intermediate host of Schistosoma mansoni / 中国血吸虫病防治杂志

Objective To compare the difference of fertility of Biomphalaria glabrata snails between self-fertilization and cross-fertilization and to observe the circadian rhythm of laying eggs, the effect of light on laying eggs and the tolerance of the snail to water and food deficiency, so as to provide the evidence for control and elimination of B. glabrata snails in the field. Methods Under laboratory conditions, a single B. glabrata egg for self-fertilization was separated and hatched individually, and young snails were raised in different plastic boxes individually. The eggs for cross-fertilization were hatched and the young snails were fed in the same plastic box. The ability of spawn, the development of the eggs, and the number of snails growing from young to adult snails were compared between the self-fertilization and cross-fertilization. The snails were in the water under four environments, all day illumination, all day without illumination, daytime lighting and night without illumination, and daytime without illumination but night lighting. The eggs were collected and counted daily. The circadian rhythm of spawn and the effect of illumination on spawn were observed. The adult snails were divided into 6 groups and exposed to the environments with relative humidity of 0, 65%, 87% and 100%, respectively. The survival rates of the adult snails exposed to the different environments after different time were observed. The adult snails were placed at 25 °C in the oven to remove water content from the soft body of snails. When the dehydration rates of the soft bodies achieved 10%, 20%, 30%, 40%, 50%, 52%, 55%, 57%, 60%, and 70% respectively, the survival rates of the adult snails exposed to the oven were observed. Results In the 25 °C water, the average laying egg number for 15 days per snail was (8.77 ± 16.92) eggs/snail in the self-fertilization snail. The average laying egg number for 15 days per snail was (149.71 ± 142.28) eggs/snail in the cross-fertilization snails. There was a significant difference between the self-fertilization snail and cross-fertilization snail (t = 0.999 999, P < 0.01). The hatching rate and reproductive maturation rate of the self-fertilization snails and cross-fertilization snails were 50.1% and 78.9%, and 19.3% and 3.8%, respectively, There was a significant difference (the hatching rate: χ2 = 18.18, P < 0.01, the reproductive maturation rate: χ2 = 11.83, P < 0.01) . In the natural environment of daytime with illumination and nighttime with darkness, the amount of laying 20 eggs of B. glabrata snail was (944.07 ± 392.53) eggs/day during a whole day, among them the amount of laying eggs during daytime account for 10.1% and the amount of laying eggs during nighttime account for 89.9%, and the laying egg was given priority to with the night. The above results suggested that the dark environment was conducive to B. glabrata snails to lay eggs. The above results suggested that light can promote the increase of spawning of B. glabrata. When B. glabrata was exposed to the environments with the relative humidity of 0, 65%, 87% and 100% at 25 °C, respectively, and the longest survival times of snails were 7, 70, 150 d and 100 d, respectively. In the 25 °C water, the snails could survive for 50 days without food. The adult snails were placed at 25 °C in the oven to remove water content from the soft body of snails. When the dehydration rates of the soft bodies achieved 10%, 20%, 30%, 40%, 50%, 52%, 55%, 57%, 60%, and 70% respectively, the survival rates of the adult snails exposed to the oven were 100%, 100%, 100%, 100%, 70%, 30%, 0, 0, 0 and 0, respectively. Conclusions B. glabrata can achieve the reproductive process by cross-fertilization or self-fertilization. There is a significant difference in reproductive ability between the cross-fertilization snail and self-fertilization snail, cross-fertilization is stronger than self-fertilization, but the rate of reproduction in the self-fertilization is higher than that in the cross-fertilization. It is indicated that B. glabrata that survive after the dry season plays an important role in the maintenance of local snail populations and transmission of schistosomiasis mansoni.

Study on biological characteristics: reproduction and viability of Biomphalaria glabrata as an intermediate host of Schistosoma mansoni / 中国血吸虫病防治杂志

Objective To compare the difference of fertility of Biomphalaria glabrata snails between self-fertilization and cross-fertilization and to observe the circadian rhythm of laying eggs, the effect of light on laying eggs and the tolerance of the snail to water and food deficiency, so as to provide the evidence for control and elimination of B. glabrata snails in the field. Methods Under laboratory conditions, a single B. glabrata egg for self-fertilization was separated and hatched individually, and young snails were raised in different plastic boxes individually. The eggs for cross-fertilization were hatched and the young snails were fed in the same plastic box. The ability of spawn, the development of the eggs, and the number of snails growing from young to adult snails were compared between the self-fertilization and cross-fertilization. The snails were in the water under four environments, all day illumination, all day without illumination, daytime lighting and night without illumination, and daytime without illumination but night lighting. The eggs were collected and counted daily. The circadian rhythm of spawn and the effect of illumination on spawn were observed. The adult snails were divided into 6 groups and exposed to the environments with relative humidity of 0, 65%, 87% and 100%, respectively. The survival rates of the adult snails exposed to the different environments after different time were observed. The adult snails were placed at 25 °C in the oven to remove water content from the soft body of snails. When the dehydration rates of the soft bodies achieved 10%, 20%, 30%, 40%, 50%, 52%, 55%, 57%, 60%, and 70% respectively, the survival rates of the adult snails exposed to the oven were observed. Results In the 25 °C water, the average laying egg number for 15 days per snail was (8.77 ± 16.92) eggs/snail in the self-fertilization snail. The average laying egg number for 15 days per snail was (149.71 ± 142.28) eggs/snail in the cross-fertilization snails. There was a significant difference between the self-fertilization snail and cross-fertilization snail (t = 0.999 999, P < 0.01). The hatching rate and reproductive maturation rate of the self-fertilization snails and cross-fertilization snails were 50.1% and 78.9%, and 19.3% and 3.8%, respectively, There was a significant difference (the hatching rate: χ2 = 18.18, P < 0.01, the reproductive maturation rate: χ2 = 11.83, P < 0.01) . In the natural environment of daytime with illumination and nighttime with darkness, the amount of laying 20 eggs of B. glabrata snail was (944.07 ± 392.53) eggs/day during a whole day, among them the amount of laying eggs during daytime account for 10.1% and the amount of laying eggs during nighttime account for 89.9%, and the laying egg was given priority to with the night. The above results suggested that the dark environment was conducive to B. glabrata snails to lay eggs. The above results suggested that light can promote the increase of spawning of B. glabrata. When B. glabrata was exposed to the environments with the relative humidity of 0, 65%, 87% and 100% at 25 °C, respectively, and the longest survival times of snails were 7, 70, 150 d and 100 d, respectively. In the 25 °C water, the snails could survive for 50 days without food. The adult snails were placed at 25 °C in the oven to remove water content from the soft body of snails. When the dehydration rates of the soft bodies achieved 10%, 20%, 30%, 40%, 50%, 52%, 55%, 57%, 60%, and 70% respectively, the survival rates of the adult snails exposed to the oven were 100%, 100%, 100%, 100%, 70%, 30%, 0, 0, 0 and 0, respectively. Conclusions B. glabrata can achieve the reproductive process by cross-fertilization or self-fertilization. There is a significant difference in reproductive ability between the cross-fertilization snail and self-fertilization snail, cross-fertilization is stronger than self-fertilization, but the rate of reproduction in the self-fertilization is higher than that in the cross-fertilization. It is indicated that B. glabrata that survive after the dry season plays an important role in the maintenance of local snail populations and transmission of schistosomiasis mansoni.

[Studies on colonization risk and potential geographical distribution of Biomphalaria glabrata as an intermediate host of Schistosoma mansoni in Mainland China].

OBJECTIVE: To predict the colonization risk and potential geographical distribution of Biomphalaria glabrata in the Mainland China based on the past period temperature data. METHODS: The survival extreme high temperatures and low temperatures of B. glabrata eggs, young and adult B. glabrata snails and the average effective accumulated temperature of generation development were determined in laboratory conditions. The temperature data in January and July from 1955 to 2010 were collected from the national meteorological monitoring sites in the southern part of China, including Chongqing, Zhejiang, Yunnan, Sichuan, Jiangxi, Hunan, Hainan, Guizhou, Guangdong, Guangxi and Fujian provinces (11 provinces). A database of ambient temperature related to B. glabrata was established based on the Geographic Information System (GIS). The colonization risk and potential geographical distribution of B. glabrata in the southern part of China were analyzed and predicted by ArcGIS 10.1 software. RESULTS: The half lethal low temperatures of B. glabrata eggs, young and adult B. glabrata snails were 6.80, 6.34 ℃ and 6.60 ℃ respectively; the half lethal high temperatures of B. glabrata eggs, young and adult B. glabrata snails were 35.99, 33.59 ℃ and 32.20 ℃, respectively. The developmental threshold temperature was 7.16 ℃; the average effective accumulated temperature of generation development was (1 970.07 ± 455.10) days-degree. The GIS overlay analysis of the half lethal low and high temperatures of B. glabrata showed that the local temperature conditions in all Hainan and part regions in Yunnan, Guangxi, Guangdong and Fujian were conformed to the survival temperature of B. glabrata snails. The regions, where the average effective accumulated temperature was more than the average effective accumulated temperature of generation development of B. glabrata, were Guangdong and Hainan, and part regions of other 9 provinces. The overlay analysis of GIS maps of the survival extreme high temperatures and low temperatures of B. glabrata with the GIS map of the average effective accumulated temperature of generation development in 2010 showed that the whole region of Hainan and part regions of Guangdong, Guangxi, Yunnan and Fujian were potential geographical distribution regions of colonization risk of B. glabrata. The overlay analysis of GIS maps of the survival extreme high temperatures and low temperatures of B. glabrata with the GIS map of the average effective accumulated temperature of generation development from 1955 to 2010 showed that the potential geographical distribution regions of B. glabrata was expanding from the whole region of Hainan and part regions of Guangdong in 1955 to the whole region of Hainan and part regions of Guangdong, Guangxi, Yunnan and Fujian in 2010. CONCLUSIONS: If B. glabrata snails were introduced into the Mainland China, the potential geographical distribution regions would be the whole region of Hainan and part regions of Guangdong, Guangxi and Yunnan. The changes of risk range and risk intensity present the trends of expanding and increasing from the south to the north gradually.

Studies on colonization risk and potential geographical distribution of Biom-phalaria glabrata as an intermediate host of Schistosoma mansoni in Main-land China / 中国血吸虫病防治杂志

Objective To predict the colonization risk and potential geographical distribution of Biomphalaria glabrata in the Mainland China based on the past period temperature data.Methods The survival extreme high temperatures and low tem-peratures of B.glabrata eggs,young and adult B.glabrata snails and the average effective accumulated temperature of genera-tion development were determined in laboratory conditions.The temperature data in January and July from 1955 to 2010 were collected from the national meteorological monitoring sites in the southern part of China,including Chongqing,Zhejiang,Yun-nan,Sichuan,Jiangxi,Hunan,Hainan,Guizhou,Guangdong,Guangxi and Fujian provinces(11 provinces).A database of ambient temperature related to B.glabrata was established based on the Geographic Information System(GIS).The colonization risk and potential geographical distribution of B.glabrata in the southern part of China were analyzed and predicted by ArcGIS 10.1 software.Results The half lethal low temperatures of B.glabrata eggs,young and adult B.glabrata snails were 6.80,6.34℃ and 6.60℃ respectively;the half lethal high temperatures of B.glabrata eggs,young and adult B.glabrata snails were 35.99,33.59℃ and 32.20℃,respectively.The developmental threshold temperature was 7.16℃;the average effective accumu-lated temperature of generation development was(1 970.07 ± 455.10)days-degree.The GIS overlay analysis of the half lethal low and high temperatures of B.glabrata showed that the local temperature conditions in all Hainan and part regions in Yunnan,Guangxi,Guangdong and Fujian were conformed to the survival temperature of B.glabrata snails.The regions,where the aver-age effective accumulated temperature was more than the average effective accumulated temperature of generation development of B.glabrata,were Guangdong and Hainan,and part regions of other 9 provinces.The overlay analysis of GIS maps of the sur-vival extreme high temperatures and low temperatures of B.glabrata with the GIS map of the average effective accumulated tem-perature of generation development in 2010 showed that the whole region of Hainan and part regions of Guangdong,Guangxi,Yunnan and Fujian were potential geographical distribution regions of colonization risk of B.glabrata.The overlay analysis of GIS maps of the survival extreme high temperatures and low temperatures of B.glabrata with the GIS map of the average effective accumulated temperature of generation development from 1955 to 2010 showed that the potential geographical distribution re-gions of B.glabrata was expanding from the whole region of Hainan and part regions of Guangdong in 1955 to the whole region of Hainan and part regions of Guangdong,Guangxi,Yunnan and Fujian in 2010.Conclusions If B.glabrata snails were intro-duced into the Mainland China,the potential geographical distribution regions would be the whole region of Hainan and part re-gions of Guangdong,Guangxi and Yunnan.The changes of risk range and risk intensity present the trends of expanding and in-creasing from the south to the north gradually.

[Advances in researches of biogenic molluscicides].

Biogenic molluscicides refer to the use of plants, animals and micro-organisms or their metabolites, and synthesis biomimetic molluscicides to kill Oncomelania hupensis snails. With the rapid development of science and technology, new biogenic molluscicides are continuously emerging and the category also continues to expand. According to the molluscicidal active ingredient and sources, at present, the biogenic molluscicides with in-depth studies include plant-derived molluscicides, micro-organism molluscicides, microbial metabolite molluscicides and animal molluscicides. This paper reviews the advances in the researches of biogenic molluscicides in recent years.

[Role of goat in transmission of schistosomiasis japonica III Effect of temperature and humidity on survival of eggs in goat feces and egg survival in natural environments].

OBJECTIVE: To investigate the survival of Schistosoma japonicum eggs in goat feces in natural marshlands and the factors affecting its survival, so as to provide evidences for understanding of the role of eggs in goat feces in the transmission of schistosomiasis and the development of the interventions pertaining to disease control and elimination. METHODS: The goat animals of schistosomiasis japonica were modeled in laboratory, and the feces of infected goat were collected. In laboratory, the effects of environmental temperature and water content in goat feces on egg hatching were evaluated, and in the field, the effect of duration of goat feces on marshland on egg hatching and the effect of direct sunshine on egg survival were evaluated. RESULTS: At 25°C in laboratory, the hatching rate of eggs in goat feces washigh-positively correlated with the water content in goat feces (r = 0.87). If the water content reduced to 7.6% in goat feces, the eggs in goat feces lost the ability to hatch. Under the same water content in goat feces, the hatching rate of eggs gradually decreased with the extension of the duration of exposure of goat feces to -5 °C, which reduced to 0 following 5 h exposure. At 5, 15 and 25 °C, the hatching rates of eggs gradually decreased with the extension of the duration of exposure of goat feces, and themiracidium hatching ratesof eggs were 2.3%, 5% and 0.9% respectively following the exposure for 52 d. At 35°C, the hatching rate of eggs gradually decreased with the extension of the duration of exposure, which reduced to 0 following 13 d exposure. In winter (-2-10 °C), the hatching rate of eggs gradually decreased with the extension of the duration of exposure of goat feces on marshlands, which reduced to 0 after 21 d of exposure, and in spring (16-19 °C), the hatching rate of eggs gradually decreased with the extension of the duration of exposure of goat feces on marshlands, which reduced to 0.9% after 5 d of exposure. At the same time point on the same marshland, the hatching rate of eggs in goat feces exposed to marshlands with direct sunshine was lower than that without direct sunshine. CONCLUSIONS: The survival of S. japonicum eggs in goat feces is associated with environmental temperature and water content (humidity) in goat feces, and the temperature and humidity are major natural factors affecting egg hatching.

[Role of goat in transmission of schistosomiasis japonica III Environmental contamination by goat feces and prediction of environments at high risk of S. japonicum infection].

OBJECTIVE: To quantitatively estimate the range and area of environmental contamination by the feces of Schistosoma japonicum-infected that were freely grazed, so as to provide the theoretical evidence for the scientific assessment of the role of the freely grazed goat in the transmission of schistosomiasis japonica and development of control strategy. METHODS: All the fecal samples excreted by the infected goat at daytime (12 h) were collected by using a self-made goat fecal collector, weighed and counted. The quantity and dispersal of the feces excreted by the freely grazed goat at daytime under a natural condition were investigated, and the walking route and speed of the freely grazed goat at daytime were recorded with a multifunction GPS data logger. The maximum range and area of the environment contaminated by the feces of the freely grazed goat at daytime were estimated, and the maximum range and area of the Oncomelania hupensis snails that may be infected by the schistosome miracidium released from the eggs in the fecal samples of the freely graze goat at daytime were calculated. RESULTS: During the walking along the marshland at daytime (12 h), the quantity of the feces execrated by the freely grazed infected goat was (232.8 ± 39.8) g per goat, and the fecal samples were composed of (819.2 ± 152.1) pellets. The goat had a mean walking speed of (0.522 7 ± 0.099 7) km/h, and the longest distance, largest radius and largest range of walking activity were (6.272 4 ± 1.195 8) km, 3.136 2 km and (3 191.113 0 ± 1 189.709 4) hm2 at daytime, respectively. The area of the snails that may be infected by the miracidium released from the eggs in the fecal samples of the freely graze goat (range of key regions for infected snails detection and control) at daytime was estimated to be (3 210.717 5 ± 1 190.907 3) hm2. CONCLUSIONS: The intensity of environmental contamination by the eggs in the fecal samples of the freely grazed goat is linked to the number of infected goat. The contamination range caused by the feces of the freely grazed goat with fixed fences is relatively stably kept within the walking range at day-time, and the range and area of goat fecal contamination is associated with the number of households that breed goat and the distribution of goat fence. The area of the snails that may be infected by the miracidium released from the eggs in the fecal samples of the freely graze goat is larger than the area of setting contaminated by the eggs in the goat feces, indicating that the range of infected snail examination and control is larger than the range of goat feces detected.

[Role of goat in transmission of schistosomiasis japonica III Development of goat feces collector and optimization of fecal hatching technique].

OBJECTIVE: To develop a simple, feasible goat feces collector to improve the collection accuracy and integrity of goat fecal samples without pollution, and to modify the miracidium hatching test with a plastic tube to achieve simple, standard and comparative procedures, so as to provide technical support for pathogenic diagnosis and scientific research of goat schistosomiasis japonica. METHODS: According to the body features of goat in marshland regions, a goat fecal collector, which was made of coarse fabric cottons, was devised, which was able to be fixed onto the goat buttocks and avoid urine pollution. Prior to miracidium hatching test, the goat fecal samples were pieced by using a mechanical method instead of the conventional artificial piecing method, and the effect of mechanical piecing treatment on miracidium hatching was evaluated. A filter membrane was added between the tube and rubbery ring to block the floater in fecal residues into the tube. The effects on miracidium hatching by using thin fat-free cotton, thick fat-free cotton, nylon gauze at 100 pores/25.4 mm2 and 150 pores/25.4 mm2 were compared. RESULTS: The goat feces collector was composed of foreleg fixing garment, hindleg fixing garment and stool bag. The functions of the fixing garment were as a fixed collector to allow non-shift and tolerance of weight during goat activity, while the major function of stool bag was in storage of stool. The goat activity did not affect by the use of collector, and all fecal samples were excreted to the bag. This collector was easy to perform and could avoid urine pollution, which was reusable after cleaning. Prior to miracidium hatching, the goat fecal samples, together with water, were pieced at 18000 to 23000 r/min for successive three times in a cooking machine, of 10 s each time at an interval of 5 s. Mechanical piecing had no clear-cut effect on miracidium hatching of eggs in fecal samples. A total of 541, 620, 344 and 211 miracidia were detected by using the miracidium hatching test with nylon gauze at 100 pores/25.4 mm2 and 150 pores/25.4 mm2, thin fat-free cotton and thick fat-free cotton respectively, indicating a better detection efficacy by using nylon gauze at 100 pores/25.4 mm2 and 150 pores/25.4 mm2. CONCLUSIONS: The goat fecal collector is a easy-to-perform, accurate, unpolluted and reusable device to collect goat feces, which is suitable for pathogenic diagnosis of goat schistosomiasis. Mechanical piecing and use of nylon gauze at 150 pores/25.4 mm2 allow a simple, accurate and stable technique for parasitological diagnosis of schistosomiasis japonica, which provides a reliable tool for schistosomiasis control and research.

[Role of goat in transmission of schistosomiasis japonica â £ Schistosome development in goat and egg count and distribution in goat feces].

OBJECTIVE: To describe the growth and development of Schistosoma japonicum in goat and the intensity and temporal distribution of eggs excreted by goat feces, so as to provide baseline data for the control and elimination of the role of goat in the transmission of schistosomiasis. METHODS: The goat animal models of schistosomiasis were established, and stool samples were collected for parasitological examinations. The number of adult worms recovered, variation of schistosomes in goat at different time points post-infection, number of eggs in schistosomes, variation in number and temporal profiles of eggs excreted from goat feces were observed. RESULTS: Of the 6 schistosome-infected goat, 415 adult worms were recovered, with a mean adult worm recovery of 34.58% (range, 23.00% to 45.50%). Among the 5 goat infected with 200 cercariae each, 47, 93, 77, 74 and 73 adult worms were recovered 2, 5, 8, 11 and 14 months post-infection, respectively. There were (200.00±42.33), (226.20±45.88), (168.20±25.85), (183.80±55.13) and (190.80±53.53) eggs detected in female schistosomes. The mean prepatent period of eggs excreted by 10 infected goat was (37.7±3.02) d. From 2 to 14 months post-infection, 7 batches of goat feces were hatched, and there were 30, 23, 14, 1 and 2 times for miracidium intensity of "++++", "+++", "++", "+" and "-", respectively, with 42.86%, 32.86%, 20.00%, 1.43% and 2.86% constituent ratios of miracidium intensity. CONCLUSIONS: Approximately 1/3 S. japonicum cercariae may develop to adults in goats post-infection, and the prepatent period of eggs is (37.7±3.02) d. There is no remarkable decrease seen in the number of adult worms, eggs in female schistosomes and eggs in goat feces within 14 months post-infection. Our findings suggest a long duration for infected goat in the transmission of schistosomiasis, and there is no evidence to prove the "self-cure" phenomenon in goat, indicating that goat is an important source of infection for schistosomiasis japonica.

[Studies on spray of niclosamide ethanolamine salt. I. development and effect assessment of spray of niclosamide ethanolamine salt].

OBJECTIVE: To develop a spray of niclosamide ethanolamine salt for prevention of bovine from Schistosoma japonicum infection, and explore its characteristics and effect. METHODS: The solubilizers, penetrating agents, emulsifiers were screened, and the spray of niclosamide ethanolamine salt was formulated according to the screening results. The niclosamide ethanolamine salt was determined by using a HPLC technique, and the stability was observed. The preventive effect of the spray was assessed by in-vitro trials against cercariae and protection trials in mice. RESULTS: The screened formulation was presented as following: 1% niclosamide ethanolamine salt was dissolved in 18% dimethyl sulfoxide, and then added with 1% azone and 2% span, together with 78% ethanol, to yield a 1% spray of niclosamide ethanolamine salt. The spray appeared golden flowing liquid, with 1% niclosamide ethanolamine salt in content (W/W), pH 7.4-7.8, and good thermal and cold stability. All cercariae died (100%) while exposed to the spray at a concentration of 1.00 mg/L for 2 min, and the similar effect was achieved while exposed to 0.50 mg/L of the spray for 5 min or 0.10 mg/L for 30 min. The spray at concentrations less than 0.05 mg/L had no evident toxicity to cercariae. A volume of 0.5 ml of the 1% spray was sprayed on the abdomen of mice, 1-3 d later, the mice were infected with S. japonicum cercariae on the spraying sites, no mice were infected, with a protection rate of 100%. Five days post-spraying, the protection rate was 40%, and the worm burden reduction rate was 65.87%. Ten days later, all the mice were infected, however, the worm burden reduction rate was 51.98%. The worm burdens on days 5 or 10 post-spraying were significantly lower than those of the control (P < 0.01). The spray exhibited a good preventive efficacy to mice from S. japonicum infection in lab. CONCLUSIONS: The spray of niclosamide ethanolamine salt has stable physical and chemical property, and is a novel liquid preventive agent against bovine schistosomiasis. In addition, it has a rapid activity against S. japonicum cercariae, so can prevent bovine from S. japonicum infection.