RESUMEN
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.
RESUMEN
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.
Asunto(s)
Animales , Adenosina Difosfato/farmacología , Adenosina Trifosfato , Moluscocidas/farmacología , Gel de Sílice/farmacología , Caracoles , Streptomyces , AguaRESUMEN
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.
RESUMEN
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.
RESUMEN
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.
RESUMEN
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.
RESUMEN
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.
RESUMEN
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.