Objective： To observe the discrepancies of responses induced by Schistosoma japonicum （S. japonicum） normal cercaria antigen （NCA） and ultraviolet （UV） -radiation-attenuated cercaria antigen （UVACA） in an in ...Objective： To observe the discrepancies of responses induced by Schistosoma japonicum （S. japonicum） normal cercaria antigen （NCA） and ultraviolet （UV） -radiation-attenuated cercaria antigen （UVACA） in an in vitro system. Methods： S. japonicum cercariae were collected and UVACA and NCA were prepared. Mouse macro- phage model cells （RAW 264.7） were treated with medium, NCA （40 μg/mL） or UVACA （40 μg/mL） in the presence or absence of recombinant mouse interferon gamma （rmIFN-γ; 4 ng/mL） for 48 h. Cell surface staining and flow cytometry were used to assess the major histocompatibility complex （MHC） Ⅱ expression, and data were expressed as mean fluorescence intensities （MFI）. Interleukin （IL） -10, IL-6 and prostaglandin E2 （PGE2） in cell culture supernatant were evaluated by commercial enzyme-linked immunosorbent assays. Results： NCA significantly suppressed IFN-7-induced MHC Ⅱ expression on RAW 264.7 cells. In the presence of 1FN-7, NCA significantly promoted IL-6, IL-10 and PGE2 secretion from RAW 264.7 cells. In the presence of IFN-γ, UVACA significantly promoted IL-10 but not IL-6 and PGE2 secretion from RAW 264.7 cells and showed no effect on IFN-γ-induced MHC Ⅱ expression. Compared with UVACA, NCA significantly suppressed IFN-γ-induced MHC Ⅱ expression and significantly promoted IL-6, PGE2 and IL-10 secretion from RAW 264.7 cells. Conclusion： RAW 264.7 cells respond differently to NCA and UVACA. NCA can significantly suppress IFN-γ-induced MHC Ⅱ expression and significantly promote IL-6, IL-10 and PGE2 secretion from RAW 264.7 cells compared with UVACA.展开更多
Aim To explore if Echinochasmus liliputanus cercariae can develop into metacercariae both invivo and in vitro and biological activity of the metacercariae,then to determine the effects of silver ni-trate on eereariae ...Aim To explore if Echinochasmus liliputanus cercariae can develop into metacercariae both invivo and in vitro and biological activity of the metacercariae,then to determine the effects of silver ni-trate on eereariae encystation in vitro.Methods Cereariae of Echinochasmus liliputanus from Bellamyaaeruginosa snails treated with or without silver nitrate were used to infected goldfish,the second inter-mediate host,or to incubated in many different solutions for 24 h to record the encystation rates.Themetacercariae formed both in vivo and in vitro were then used to infected New Zealand rabbits to testtheir infectivity to its definitive hosts or to exeyst in 0.1% sodium deoxycholate excystation medium at37℃ for 1 h.Results In vivo encystment of cercariae occurred in the gills of goldfish.However,thecercariae were also able to encyst in vitro in Locke’s solution,NaCl solution,artificial gastric juice orhuman gastric juice with eneystation rates of 74.28 %,44.94 %,8.37 % and 10.79 %.0.7×-1.2×Locke’s or 0.7%-1.2% NaCl solution was shown to be appropriate for in vitro encystment to occurwithin 24 hour,however,full- strength Locke’s solution was shown to be optimal.The one- day- oldencysted metacereariae formed in vivo showed 88.53 % exeystation when treated in 0.1% sodium de-oxycholate exeystation medium at 37℃ for 1h.The metacercariae formed in vitro,however,showed88.60% and 84.95% excystation for normal and abnormal ones respectively.While abnormal cysts atroom temperature usually die within 10 days,about 70% normal cyst,both in vivo and in vitro,canstill excyst after stored in 0.5×Locke’s at 4℃ for 3 mouths.Cysts formed in vivo and in vitro were e-qually infective to rabbits.1 uM silver nitrate had a dramatic effect on the cercariae encysting in vitro.When treated with silver nitrate,the cereariae encystation rates decreased to 16.25% in Locke’s solu-tion and 6.69% in NaCl solution,however,the encystment was largely restored wben the cercariaewere washed to remove Ag+.Conclusion The finding of E.liliputanus cercariae encysting in vitro,especially in human gastric juice,might be helpful in elucidating mechanisms of the definitive hosts in-fected by the cercariae directly.The encystment of the cercariae in vitro could be inhibited when thecercariae were treated with 1 uM silver nitrate.As silver nitrate binds to the papillae,especially to theciliated papillae,on the cercarial surface,it is suggested that papillar chemoreceptors may be involved inencystment of the cercariae.展开更多
Objective:To investigate the epidemiological situation of cercarial trematodes infection in freshwater snails from different water resources in Chiang Mai province,Thailand.Methods:The snail specimens were collected f...Objective:To investigate the epidemiological situation of cercarial trematodes infection in freshwater snails from different water resources in Chiang Mai province,Thailand.Methods:The snail specimens were collected from 13 districts of Chiang Mai province during April 2008 to February 2012.The prevalence of cercarial infection in snails was investigated using the crushing method.The drawing was done with the help of a camera lucida for the morphological study.Results:A total of 2479 snail individuals were collected and classified into 7 families,11 genera,and 14 species,Among them,8 snails species were found to be infected with an overall prevalence of 17.27%(428/2479),which infected with nine groups of cercariae;gymnocephalous cercaria,strigea cercaria,megalurous cercaria,monostome cercaria,parapleurolophocercous cercaria(Haplorchis cercaria),pleurolophocercous cercaria,furcocercous cercaria(Transversotrema cercaria),xiphidiocercaria,and virgulate cercaria.The parapleurolophocercous cercaria was found to be the dominant type among the cercarial infection in the snails(64.25%).Conclusions:The various species of snails found in the research location act as the intermediate hosts for the high prevalence of parasitic infection of many species of mammals.This work will provide new information on both the distribution and first intermediate host of trematodes.展开更多
The migratory pattern and attrition of<sup>75</sup>Se-labelled S.mansoni challenge cercariae in CF1mice immunized 0,1,3, or 5 times with 12kR-or 48kR-irradiated cercariae were investigatedby compressed org...The migratory pattern and attrition of<sup>75</sup>Se-labelled S.mansoni challenge cercariae in CF1mice immunized 0,1,3, or 5 times with 12kR-or 48kR-irradiated cercariae were investigatedby compressed organ autoradiography.Mice were percutaneously inoculated with 500-1000 cer-cariae for each immunization and with a known count of approximately 90 cercariae at adifferent site for challenge.The skin,lungs,liver,other organs,and catrcass were processedfor autoradiography.Hepatic and mesenteric perfusions for worm collection were alsosampled from day 19 through 34.In naive tnice,25% of challenge cercariae died in theskin.Skin attrition of challenge cercariae in mice immunized with 12kR-irradiated cercariaewas 43% with 1 immunization,54% with 3,and 58% with 5,whereas in mice immunized with48kR-irradiated cercariae,skin,attrition was48% with 1,63% with 3,and 76% with 5 immunni-zations.In all immunized mice,a high percentage of migrating sehistosomula died in the skinwithin 24 hours of a challenge infection.Utilizing autoradiography,it was found that thesilver foci created by single labelled worms were difficult to differentiate from those of pairedworms.This study documents the induction of acquired resistance against a challenge infectionin the skin by vaccination with highly irradiated cercariae,with the degree of attemtation ofimmunizing cercariae and the number of immunizations being closely related to the attritiotn.rate in the skin.Finally,we discuss the contradictory views concerning the site of attritiotn ofchallenge cercariae.展开更多
Objective:To explore larvicidal effects of anthelmintic drugs on Opisthorchis viverrini(O.viverrini) for alternative approach to interrupting its cycle for developing a field-based control program.Methods:The larvicid...Objective:To explore larvicidal effects of anthelmintic drugs on Opisthorchis viverrini(O.viverrini) for alternative approach to interrupting its cycle for developing a field-based control program.Methods:The larvicidal activities of albendazole(A1),artesunate(Ar),praziquantel(Pzq) and miltefosine(Mf) on O.viverrini cercariae and mature metacercariae were investigated.Lethal concentrations(LC_(50) and LC_(95)) of these drugs were determined.Mature metacercariae previously exposed to various concentrations of the drugs were administered to hamsters.Worms were harvested 30 d post infection and worm recovery rates calculated.Al,Ar,Pzq and Mf produced morphological degeneration and induced shedding tails of cercariae after 24 h exposure.Results:The LC_(50) and LC_(95) of Al,Ar,Pzq and Mf on cercariae were 0.720 and 1.139,0.350 and 0.861,0.017 and 0.693,and 0.530 and 1.134 ppm,respectively.LC_(50) and LC_(95) of Ar on mature metacercariae were 303.643 and 446.237 ppm and of Mf were 289.711 and 631.781 ppm,respectively but no lethal effect in Pzq-and Al-treated groups(up to 1 ppt).No worms were found in hamsters administered Pzq-treated metacercariae.The adult worms from Al-treated metacercariae were significantly bigger in size compared to the control group(P<0.05).Fecundity and body width were greater in adults from Mf-treated metacercariae compared to the control group(P<0.05).Conclusions:The larvicidal effects of these drugs were high efficacy to O.viverrini cercariae but lesser efficacy to metacercariae.It should be further studied with the eventual aim of developing a field-based control program.展开更多
Objective： To better understand the reason that Schistosoma japonicurn （S. japonicum） ultraviolet （UV）- radiated cercariae could not induce high level of protection in C57BL/6 mice. Methods： Microarray technolog...Objective： To better understand the reason that Schistosoma japonicurn （S. japonicum） ultraviolet （UV）- radiated cercariae could not induce high level of protection in C57BL/6 mice. Methods： Microarray technology was performed to investigate the gene transcription profile in skin draining lymph nodes （sdLNs） at 1 w after exposure to attenuated cercariae （AC） or normal cercariae （NC） of S. japonicum in C57BL/6 mice. The expressions of some representative genes were further confirmed by real-time PCR. Subsequently, the expressions of Th1/Th2 cytokine genes, cytotoxicity-related genes, as well as co-stimulator genes in spleens from AC-vaccinated and NC- infected mice were analyzed by real-time PCR at w 3 and 6 post-exposure. Results： The gene expressions of Th1 cytokines, including interferon-y （IFN-γ）, interleukin （IL）-12 and tumor necrosis factor-α （TNF-α） in the sdLNs were significantly lower in AC-vaccinated mice than in NC-infected mice. Furthermore, the gene expressions of Th1- and Th2- cytokines, including IFN-γ, IL-12, TNF-α, IL-4 and IL-10, in the spleens from AC-vaccinated mice showed little changes at w 3 and 6 post-vaccination. In addition, cytotoxicity-related molecules including granzyme A, granzyme B, granzyme K, perforin 1 and Fas L were up-regulated from the early stage of vaccination, and peaked at the 3rd w after vaccination with UV-AC. Conclusion： UV-AC of S. japonicum could not ef- fectively induce a Thl response in C57BL/6 mice, which may be an explanation for the low protection against parasite challenge, and the role played by up-regulated expression of cytotoxicity-related genes in mice needs to be further investigated.展开更多
Objective:To investigate the prevalence of cercarial infections in freshwater snails from several water sources in Nakhon Nayok,Nonthaburi,and Pathum Thani provinces of Central Thailand,and to reconstruct a phylogenet...Objective:To investigate the prevalence of cercarial infections in freshwater snails from several water sources in Nakhon Nayok,Nonthaburi,and Pathum Thani provinces of Central Thailand,and to reconstruct a phylogenetic tree for improved understanding of the relationships in the cercarial stage.Methods:The snail specimens were collected from 34 total sampling sites and investigated for cercarial infections using the crushing method.The cercarial specimens were classified and used for the phylogenetic tree analysis using the Internal Transcribed Spacer 2(ITS2).Results:A total of 1921 snail specimens were classified into five families and seven species.The results showed that four snail species were identified as intermediate hosts of the larval stages of trematodes,with an overall prevalence of infection of 2.45%(47/1921).The infected snail specimens included five groups of the cercarial type:cercariaeum cercariae,echinostome cercaria,megalurous cercaria,parapleurolophocercous cercaria,and xiphidiocercariae.This is particularly true of xiphidiocercariae,which was found to be the dominant type among cercarial infections in bithyniid snails by approximately 38.00%.With regard to molecular identification,the phylogenetic tree was reconstructed using the neighbor-joining method with 10000 bootstraps and separated the trematodes into three clades:Echinostomatoidea,Microphalloidea and Opisthorchioidea.Conclusions:The study reveals a high prevalence of cercarial infection for each cercarial type and maturation into a definite trematode genus and delineates morphological characteristics and evolutionary trends among each larval trematode in Nakhon Nayok,Nonthaburi and Pathum Thani provinces.In addition,the ITS2 sequence data of cercariae could be used to examine classification of these species at the family level.展开更多
Recently we applied randomly amplified polymorphic DNA (RAPD) fingerprinting to detect clonal variability among individual cercariae within daughter sporocysts and rediae of 10 digenean trematodes (Platyhelminthes: Tr...Recently we applied randomly amplified polymorphic DNA (RAPD) fingerprinting to detect clonal variability among individual cercariae within daughter sporocysts and rediae of 10 digenean trematodes (Platyhelminthes: Trematoda). The most variable RAPD patterns were obtained for Schistosomatidae representative-avian schistosome Trichobilharzia szidati. In this work, 50 polymorphic DNA fragments of approximately 300-1500 bp from RAPD patterns of individual T. szidati cercariae were cloned and sequenced. As a result genomic DNA sequences (total length of approximately 41,000 bp) revealing clonal variability in T. szidati cercariae were obtained and analyzed. The analysis indicated that these sequences contained tandem, inverted and dispersed repeats as well as regions homological to retroelements of two human parasites, Schistosoma mansoni and S. japonicum. Tandem and inverted repeats constituted 8.9% and 22.1% respectively, while the percentage of dispersed repeats was 21.0%. The average content of these components was 41.7% with the average AT content being 59.0%. About 40% of sequences included regions ranging in length from 96 to 1005 bp which displayed amino acid homology with open reading frame pol products of S. mansoni and S. japonicum retroelements: non-long terminal repeat retrotransposons (nLTRs, 76%), long terminal repeat retrotransposons (LTRs, 14%), and Penelope-like elements (PLEs, 10%). Most of these regions (86.4%) contained frameshifts, gaps, and stop-codons. The largest portion of them was homological to nLTRs of the RTE clade (67%). The number of sequences homologous to the members of CR1 lineage was 7 times smaller (9%). Homology with LTRs of Gypsy/Ty3 and BEL clades was revealed in 5% and 9% of cases respectively. We assume that the repetitive elements including retroelement-like sequences described in the current study may serve as the source of clonal variability detected previously in T. szidati and other digenean trematodes. Such genome regions rapidly accumulate mutations and thus may play an important functional role in the life history of the species.展开更多
目的探究采用不同数量微卫星位点标记对日本血吸虫种群遗传多样性分析的影响,为日本血吸虫种群遗传学研究提供参考。方法自湖北省公安县某地野外荒滩采集湖北钉螺,采用直管逸蚴法筛选出37只血吸虫感染性钉螺。分别收集每只感染性钉螺逸...目的探究采用不同数量微卫星位点标记对日本血吸虫种群遗传多样性分析的影响,为日本血吸虫种群遗传学研究提供参考。方法自湖北省公安县某地野外荒滩采集湖北钉螺,采用直管逸蚴法筛选出37只血吸虫感染性钉螺。分别收集每只感染性钉螺逸出的单条尾蚴,各随机挑选10条尾蚴提取DNA。以前期经大规模样本验证过的9个微卫星位点以及自参考文献和GenBank数据库中筛选且可稳定扩增的15个微卫星位点作为分子标记,利用Type-it微卫星PCR试剂盒对上述尾蚴DNA进行3组8重微卫星PCR扩增,通过毛细管电泳检测样本基因型。对上述日本血吸虫尾蚴DNA进行种群遗传多样性分析,评估等位基因数(observed number of alleles,Na)、有效等位基因数(effective number of alleles,Ae)、观察杂合度(observed heterozygosity,Ho)、期望杂合度(expected heterozygosity,He)和多态信息含量(polymorphism information content,PIC)等多态性指标,采用Hardy-Weinberg平衡检验和连锁不平衡评估进行尾蚴种群遗传结构分析。此外,为进一步探究微卫星位点数量对日本血吸虫种群遗传多样性的影响,依次将微卫星位点数量设定为1~24个,计算不同位点数量时日本血吸虫种群Na均数及其标准差,并计算等位基因数变异系数,观察Na随微卫星位点数量增加的变化。结果共选取345条日本血吸虫尾蚴DNA,用上述24个微卫星位点进行检测,结果显示全部位点均满足连锁平衡标准化[连锁不平衡系数(D′)<0.7,r^(2)<0.3],均偏离Hardy-Weinberg平衡(P<0.001)。日本血吸虫尾蚴种群在24个微卫星位点的Na、Ae、Ho和He均值分别为27.46±2.18、12.46±0.95、0.46±0.03和0.91±0.01,PIC值为0.85~0.96,提示24个位点在全基因组微卫星水平上均具有较好的代表性。采用经前期验证的9个微卫星位点进行分析时,日本血吸虫种群Na-Ae均值为19.88±8.43,高于使用全部24个位点分析时的结果(14.99±8.09)。随着微卫星位点数量的增加,Na均值虽无明显变化,但标准差逐渐变小;尤其是当位点数为18个及以上时,标准差变化幅度明显减小;当位点数为18个时,Na标准差小于位点数为24个时Na均值的5%,变异系数为4.6%。结论微卫星位点数量可显著影响日本血吸虫种群遗传多样性分析结果。在目前低感染率和血吸虫遗传分布不平衡的背景下,推荐选取≥18个微卫星位点进行日本血吸虫种群遗传多样性分析。展开更多
Objective To study the developing cercaria of Schistosoma Japonicum (S. japonicum) on ultrastructural level by transmission electron microscope (TEM) for analyzing the morphological dynamic changes of the tegume...Objective To study the developing cercaria of Schistosoma Japonicum (S. japonicum) on ultrastructural level by transmission electron microscope (TEM) for analyzing the morphological dynamic changes of the tegument, glands and musculature. Methods Artificial infected Oncomelania hupensis were dissected under dissecting microscope and the daughter sporocysts picked up for studying the germinal cells stage (S1). The other embryonic cercaria were selected according to the modified parameter of Chen and Bier (1972). The specimens were prepared by conventional procedure of the laboratory of TEM and were observed by Hitachi H600. Results Besides the germinal cell stage (S1), this is the first chronological study on the morphological development of S. japonicum cercaria from S2 S5 concerning the tegument and its elements (glycocalyx, sensory papilla, basal lamina and spine), head and acetabular gland and musculature of the body and tail. This article discusses the ultrastructural morphological differences from prior authors and emphasizes on the postacetalar gland and the pattern of tail musculature of the matured stage cercaria. Conclusions According to the cell division and differentiation, the process of development may be divided into 5 stages: S1, the single germinal cell stage; S2, the germinal ball stage; major multiplication and minor differentiation; S3, tail budding embryonic stage, both active in multiplying and differentiating; S4, the pre matured stage, major differentiation and minor multiplication; and S5, the fully matured stage (completing the differentiation).展开更多
Objective:To investigate the genetic diversity and population structure of the helminth intermediate host,Anentome(A.)helena,populations from northern Thailand and reveal the effect of parasitic infection on snail hos...Objective:To investigate the genetic diversity and population structure of the helminth intermediate host,Anentome(A.)helena,populations from northern Thailand and reveal the effect of parasitic infection on snail host populations.Methods:A.helena was collected throughout river drainages from northern Thailand and investigated for larval trematode infection.Seventy-three A.helena specimens were used to examine and re-construct the molecular phylogenetics(neighbour-joining,maximum likelihood,and Bayesian inference methods),genetic diversity,and population structure based on the concatenated datasets of COI and 16S rRNA(1178 bp).Results:The infection of zoogonid cercariae,heterophyid metacercariae,and echinostome metacercariae in the snail was found from 2 localities,including Mae Sa and Mae Pu Kha populations,with a total prevalence of 6.7%and 16.3%,respectively.The phylogenetic tree and haplotype network revealed three main clades and three haplogroups for each region(COI and 16S rRNA).High levels of genetic diversity in terms of haplotype diversity were found in parasitic infected populations.Conclusions:This is the first time that Echinostoma mekongi has been discovered from A.helena in northern Thailand.The results also showed that parasitic infection can affect the population genetic structure of the snail intermediate host,A.helena.Moreover,habitat and historical demographic changes could be other important factors that can affect the degree of population genetic structure in A.helena.Understanding the population structure of intermediate host species can inform parasite transmission dynamics and is useful for the creation of effective public health management strategies.展开更多
Objective:To detect the species of larval trematodes(cercariae) in Melanopsis praemorsa snails from 5 different fresh water bodies in Palestine.Methods:A total of 1 880 Melanopsis praemona snails were collected from d...Objective:To detect the species of larval trematodes(cercariae) in Melanopsis praemorsa snails from 5 different fresh water bodies in Palestine.Methods:A total of 1 880 Melanopsis praemona snails were collected from different fresh water bodies in Palestine from October,2008 to November,2010.Cercariae in Melanopsis praemona snails were obtained by lighting and crushing methods.The behavior of cercariae was observed using a dissecting microscope.Results:Three different species of larval trematodes were identified from Melanopsis praemona snails collected only from Al-Bathan fresh water body,while snails from other water bodies were not infected. These species were microcercous cercaria,xiphidiocercaria and brevifurcate lophocercous cercaria.These cercariae called Cercaria melanopsi palestinia 1,Cercaria melanopsi palestinia H and Cercaria melanopsi palestinia III have not been described before from this snail in Palestine. The infection rate of Melanopsis praemona collected from Al-Bathan fresh water body was 5.7%, while the overall infection rate of snails collected from all fresh water bodies was 4.3%.Details are presented on the morphology and behavior of the cercariae as well as their development within the snail.Conclusions:These results have been recorded for the first time and these cercariae may be of medical and veterinary importance.展开更多
基金supported by a grant from National Nature Science Found(No.30430600)
文摘Objective： To observe the discrepancies of responses induced by Schistosoma japonicum （S. japonicum） normal cercaria antigen （NCA） and ultraviolet （UV） -radiation-attenuated cercaria antigen （UVACA） in an in vitro system. Methods： S. japonicum cercariae were collected and UVACA and NCA were prepared. Mouse macro- phage model cells （RAW 264.7） were treated with medium, NCA （40 μg/mL） or UVACA （40 μg/mL） in the presence or absence of recombinant mouse interferon gamma （rmIFN-γ; 4 ng/mL） for 48 h. Cell surface staining and flow cytometry were used to assess the major histocompatibility complex （MHC） Ⅱ expression, and data were expressed as mean fluorescence intensities （MFI）. Interleukin （IL） -10, IL-6 and prostaglandin E2 （PGE2） in cell culture supernatant were evaluated by commercial enzyme-linked immunosorbent assays. Results： NCA significantly suppressed IFN-7-induced MHC Ⅱ expression on RAW 264.7 cells. In the presence of 1FN-7, NCA significantly promoted IL-6, IL-10 and PGE2 secretion from RAW 264.7 cells. In the presence of IFN-γ, UVACA significantly promoted IL-10 but not IL-6 and PGE2 secretion from RAW 264.7 cells and showed no effect on IFN-γ-induced MHC Ⅱ expression. Compared with UVACA, NCA significantly suppressed IFN-γ-induced MHC Ⅱ expression and significantly promoted IL-6, PGE2 and IL-10 secretion from RAW 264.7 cells. Conclusion： RAW 264.7 cells respond differently to NCA and UVACA. NCA can significantly suppress IFN-γ-induced MHC Ⅱ expression and significantly promote IL-6, IL-10 and PGE2 secretion from RAW 264.7 cells compared with UVACA.
文摘Aim To explore if Echinochasmus liliputanus cercariae can develop into metacercariae both invivo and in vitro and biological activity of the metacercariae,then to determine the effects of silver ni-trate on eereariae encystation in vitro.Methods Cereariae of Echinochasmus liliputanus from Bellamyaaeruginosa snails treated with or without silver nitrate were used to infected goldfish,the second inter-mediate host,or to incubated in many different solutions for 24 h to record the encystation rates.Themetacercariae formed both in vivo and in vitro were then used to infected New Zealand rabbits to testtheir infectivity to its definitive hosts or to exeyst in 0.1% sodium deoxycholate excystation medium at37℃ for 1 h.Results In vivo encystment of cercariae occurred in the gills of goldfish.However,thecercariae were also able to encyst in vitro in Locke’s solution,NaCl solution,artificial gastric juice orhuman gastric juice with eneystation rates of 74.28 %,44.94 %,8.37 % and 10.79 %.0.7×-1.2×Locke’s or 0.7%-1.2% NaCl solution was shown to be appropriate for in vitro encystment to occurwithin 24 hour,however,full- strength Locke’s solution was shown to be optimal.The one- day- oldencysted metacereariae formed in vivo showed 88.53 % exeystation when treated in 0.1% sodium de-oxycholate exeystation medium at 37℃ for 1h.The metacercariae formed in vitro,however,showed88.60% and 84.95% excystation for normal and abnormal ones respectively.While abnormal cysts atroom temperature usually die within 10 days,about 70% normal cyst,both in vivo and in vitro,canstill excyst after stored in 0.5×Locke’s at 4℃ for 3 mouths.Cysts formed in vivo and in vitro were e-qually infective to rabbits.1 uM silver nitrate had a dramatic effect on the cercariae encysting in vitro.When treated with silver nitrate,the cereariae encystation rates decreased to 16.25% in Locke’s solu-tion and 6.69% in NaCl solution,however,the encystment was largely restored wben the cercariaewere washed to remove Ag+.Conclusion The finding of E.liliputanus cercariae encysting in vitro,especially in human gastric juice,might be helpful in elucidating mechanisms of the definitive hosts in-fected by the cercariae directly.The encystment of the cercariae in vitro could be inhibited when thecercariae were treated with 1 uM silver nitrate.As silver nitrate binds to the papillae,especially to theciliated papillae,on the cercarial surface,it is suggested that papillar chemoreceptors may be involved inencystment of the cercariae.
基金Supported by National Research Council of Thailand(NRCT)(Grant No.2555A10402010)
文摘Objective:To investigate the epidemiological situation of cercarial trematodes infection in freshwater snails from different water resources in Chiang Mai province,Thailand.Methods:The snail specimens were collected from 13 districts of Chiang Mai province during April 2008 to February 2012.The prevalence of cercarial infection in snails was investigated using the crushing method.The drawing was done with the help of a camera lucida for the morphological study.Results:A total of 2479 snail individuals were collected and classified into 7 families,11 genera,and 14 species,Among them,8 snails species were found to be infected with an overall prevalence of 17.27%(428/2479),which infected with nine groups of cercariae;gymnocephalous cercaria,strigea cercaria,megalurous cercaria,monostome cercaria,parapleurolophocercous cercaria(Haplorchis cercaria),pleurolophocercous cercaria,furcocercous cercaria(Transversotrema cercaria),xiphidiocercaria,and virgulate cercaria.The parapleurolophocercous cercaria was found to be the dominant type among the cercarial infection in the snails(64.25%).Conclusions:The various species of snails found in the research location act as the intermediate hosts for the high prevalence of parasitic infection of many species of mammals.This work will provide new information on both the distribution and first intermediate host of trematodes.
文摘The migratory pattern and attrition of<sup>75</sup>Se-labelled S.mansoni challenge cercariae in CF1mice immunized 0,1,3, or 5 times with 12kR-or 48kR-irradiated cercariae were investigatedby compressed organ autoradiography.Mice were percutaneously inoculated with 500-1000 cer-cariae for each immunization and with a known count of approximately 90 cercariae at adifferent site for challenge.The skin,lungs,liver,other organs,and catrcass were processedfor autoradiography.Hepatic and mesenteric perfusions for worm collection were alsosampled from day 19 through 34.In naive tnice,25% of challenge cercariae died in theskin.Skin attrition of challenge cercariae in mice immunized with 12kR-irradiated cercariaewas 43% with 1 immunization,54% with 3,and 58% with 5,whereas in mice immunized with48kR-irradiated cercariae,skin,attrition was48% with 1,63% with 3,and 76% with 5 immunni-zations.In all immunized mice,a high percentage of migrating sehistosomula died in the skinwithin 24 hours of a challenge infection.Utilizing autoradiography,it was found that thesilver foci created by single labelled worms were difficult to differentiate from those of pairedworms.This study documents the induction of acquired resistance against a challenge infectionin the skin by vaccination with highly irradiated cercariae,with the degree of attemtation ofimmunizing cercariae and the number of immunizations being closely related to the attritiotn.rate in the skin.Finally,we discuss the contradictory views concerning the site of attritiotn ofchallenge cercariae.
基金The Higher Education Research Promotion and National Research University Project of Thailand,Office of the Higher Education Commission,though the Health Cluster(SHePGMS),Khon Kaen University to student,Miss Phornphitcha Pechdee and advisor,Assoc.Prof.Dr.Smarn Tesana for financial support of this work
文摘Objective:To explore larvicidal effects of anthelmintic drugs on Opisthorchis viverrini(O.viverrini) for alternative approach to interrupting its cycle for developing a field-based control program.Methods:The larvicidal activities of albendazole(A1),artesunate(Ar),praziquantel(Pzq) and miltefosine(Mf) on O.viverrini cercariae and mature metacercariae were investigated.Lethal concentrations(LC_(50) and LC_(95)) of these drugs were determined.Mature metacercariae previously exposed to various concentrations of the drugs were administered to hamsters.Worms were harvested 30 d post infection and worm recovery rates calculated.Al,Ar,Pzq and Mf produced morphological degeneration and induced shedding tails of cercariae after 24 h exposure.Results:The LC_(50) and LC_(95) of Al,Ar,Pzq and Mf on cercariae were 0.720 and 1.139,0.350 and 0.861,0.017 and 0.693,and 0.530 and 1.134 ppm,respectively.LC_(50) and LC_(95) of Ar on mature metacercariae were 303.643 and 446.237 ppm and of Mf were 289.711 and 631.781 ppm,respectively but no lethal effect in Pzq-and Al-treated groups(up to 1 ppt).No worms were found in hamsters administered Pzq-treated metacercariae.The adult worms from Al-treated metacercariae were significantly bigger in size compared to the control group(P<0.05).Fecundity and body width were greater in adults from Mf-treated metacercariae compared to the control group(P<0.05).Conclusions:The larvicidal effects of these drugs were high efficacy to O.viverrini cercariae but lesser efficacy to metacercariae.It should be further studied with the eventual aim of developing a field-based control program.
基金supported by the National Basic Research Program of China(973 Program,No.2007CB513106)the National Science Foundation of China(NSFC,No.30430600)
文摘Objective： To better understand the reason that Schistosoma japonicurn （S. japonicum） ultraviolet （UV）- radiated cercariae could not induce high level of protection in C57BL/6 mice. Methods： Microarray technology was performed to investigate the gene transcription profile in skin draining lymph nodes （sdLNs） at 1 w after exposure to attenuated cercariae （AC） or normal cercariae （NC） of S. japonicum in C57BL/6 mice. The expressions of some representative genes were further confirmed by real-time PCR. Subsequently, the expressions of Th1/Th2 cytokine genes, cytotoxicity-related genes, as well as co-stimulator genes in spleens from AC-vaccinated and NC- infected mice were analyzed by real-time PCR at w 3 and 6 post-exposure. Results： The gene expressions of Th1 cytokines, including interferon-y （IFN-γ）, interleukin （IL）-12 and tumor necrosis factor-α （TNF-α） in the sdLNs were significantly lower in AC-vaccinated mice than in NC-infected mice. Furthermore, the gene expressions of Th1- and Th2- cytokines, including IFN-γ, IL-12, TNF-α, IL-4 and IL-10, in the spleens from AC-vaccinated mice showed little changes at w 3 and 6 post-vaccination. In addition, cytotoxicity-related molecules including granzyme A, granzyme B, granzyme K, perforin 1 and Fas L were up-regulated from the early stage of vaccination, and peaked at the 3rd w after vaccination with UV-AC. Conclusion： UV-AC of S. japonicum could not ef- fectively induce a Thl response in C57BL/6 mice, which may be an explanation for the low protection against parasite challenge, and the role played by up-regulated expression of cytotoxicity-related genes in mice needs to be further investigated.
基金Srinakharinwirot University,Thailand for providing funding(Project Nos.071/2562,184/2563)
文摘Objective:To investigate the prevalence of cercarial infections in freshwater snails from several water sources in Nakhon Nayok,Nonthaburi,and Pathum Thani provinces of Central Thailand,and to reconstruct a phylogenetic tree for improved understanding of the relationships in the cercarial stage.Methods:The snail specimens were collected from 34 total sampling sites and investigated for cercarial infections using the crushing method.The cercarial specimens were classified and used for the phylogenetic tree analysis using the Internal Transcribed Spacer 2(ITS2).Results:A total of 1921 snail specimens were classified into five families and seven species.The results showed that four snail species were identified as intermediate hosts of the larval stages of trematodes,with an overall prevalence of infection of 2.45%(47/1921).The infected snail specimens included five groups of the cercarial type:cercariaeum cercariae,echinostome cercaria,megalurous cercaria,parapleurolophocercous cercaria,and xiphidiocercariae.This is particularly true of xiphidiocercariae,which was found to be the dominant type among cercarial infections in bithyniid snails by approximately 38.00%.With regard to molecular identification,the phylogenetic tree was reconstructed using the neighbor-joining method with 10000 bootstraps and separated the trematodes into three clades:Echinostomatoidea,Microphalloidea and Opisthorchioidea.Conclusions:The study reveals a high prevalence of cercarial infection for each cercarial type and maturation into a definite trematode genus and delineates morphological characteristics and evolutionary trends among each larval trematode in Nakhon Nayok,Nonthaburi and Pathum Thani provinces.In addition,the ITS2 sequence data of cercariae could be used to examine classification of these species at the family level.
文摘Recently we applied randomly amplified polymorphic DNA (RAPD) fingerprinting to detect clonal variability among individual cercariae within daughter sporocysts and rediae of 10 digenean trematodes (Platyhelminthes: Trematoda). The most variable RAPD patterns were obtained for Schistosomatidae representative-avian schistosome Trichobilharzia szidati. In this work, 50 polymorphic DNA fragments of approximately 300-1500 bp from RAPD patterns of individual T. szidati cercariae were cloned and sequenced. As a result genomic DNA sequences (total length of approximately 41,000 bp) revealing clonal variability in T. szidati cercariae were obtained and analyzed. The analysis indicated that these sequences contained tandem, inverted and dispersed repeats as well as regions homological to retroelements of two human parasites, Schistosoma mansoni and S. japonicum. Tandem and inverted repeats constituted 8.9% and 22.1% respectively, while the percentage of dispersed repeats was 21.0%. The average content of these components was 41.7% with the average AT content being 59.0%. About 40% of sequences included regions ranging in length from 96 to 1005 bp which displayed amino acid homology with open reading frame pol products of S. mansoni and S. japonicum retroelements: non-long terminal repeat retrotransposons (nLTRs, 76%), long terminal repeat retrotransposons (LTRs, 14%), and Penelope-like elements (PLEs, 10%). Most of these regions (86.4%) contained frameshifts, gaps, and stop-codons. The largest portion of them was homological to nLTRs of the RTE clade (67%). The number of sequences homologous to the members of CR1 lineage was 7 times smaller (9%). Homology with LTRs of Gypsy/Ty3 and BEL clades was revealed in 5% and 9% of cases respectively. We assume that the repetitive elements including retroelement-like sequences described in the current study may serve as the source of clonal variability detected previously in T. szidati and other digenean trematodes. Such genome regions rapidly accumulate mutations and thus may play an important functional role in the life history of the species.
文摘目的探究采用不同数量微卫星位点标记对日本血吸虫种群遗传多样性分析的影响,为日本血吸虫种群遗传学研究提供参考。方法自湖北省公安县某地野外荒滩采集湖北钉螺,采用直管逸蚴法筛选出37只血吸虫感染性钉螺。分别收集每只感染性钉螺逸出的单条尾蚴,各随机挑选10条尾蚴提取DNA。以前期经大规模样本验证过的9个微卫星位点以及自参考文献和GenBank数据库中筛选且可稳定扩增的15个微卫星位点作为分子标记,利用Type-it微卫星PCR试剂盒对上述尾蚴DNA进行3组8重微卫星PCR扩增,通过毛细管电泳检测样本基因型。对上述日本血吸虫尾蚴DNA进行种群遗传多样性分析,评估等位基因数(observed number of alleles,Na)、有效等位基因数(effective number of alleles,Ae)、观察杂合度(observed heterozygosity,Ho)、期望杂合度(expected heterozygosity,He)和多态信息含量(polymorphism information content,PIC)等多态性指标,采用Hardy-Weinberg平衡检验和连锁不平衡评估进行尾蚴种群遗传结构分析。此外,为进一步探究微卫星位点数量对日本血吸虫种群遗传多样性的影响,依次将微卫星位点数量设定为1~24个,计算不同位点数量时日本血吸虫种群Na均数及其标准差,并计算等位基因数变异系数,观察Na随微卫星位点数量增加的变化。结果共选取345条日本血吸虫尾蚴DNA,用上述24个微卫星位点进行检测,结果显示全部位点均满足连锁平衡标准化[连锁不平衡系数(D′)<0.7,r^(2)<0.3],均偏离Hardy-Weinberg平衡(P<0.001)。日本血吸虫尾蚴种群在24个微卫星位点的Na、Ae、Ho和He均值分别为27.46±2.18、12.46±0.95、0.46±0.03和0.91±0.01,PIC值为0.85~0.96,提示24个位点在全基因组微卫星水平上均具有较好的代表性。采用经前期验证的9个微卫星位点进行分析时,日本血吸虫种群Na-Ae均值为19.88±8.43,高于使用全部24个位点分析时的结果(14.99±8.09)。随着微卫星位点数量的增加,Na均值虽无明显变化,但标准差逐渐变小;尤其是当位点数为18个及以上时,标准差变化幅度明显减小;当位点数为18个时,Na标准差小于位点数为24个时Na均值的5%,变异系数为4.6%。结论微卫星位点数量可显著影响日本血吸虫种群遗传多样性分析结果。在目前低感染率和血吸虫遗传分布不平衡的背景下,推荐选取≥18个微卫星位点进行日本血吸虫种群遗传多样性分析。
文摘Objective To study the developing cercaria of Schistosoma Japonicum (S. japonicum) on ultrastructural level by transmission electron microscope (TEM) for analyzing the morphological dynamic changes of the tegument, glands and musculature. Methods Artificial infected Oncomelania hupensis were dissected under dissecting microscope and the daughter sporocysts picked up for studying the germinal cells stage (S1). The other embryonic cercaria were selected according to the modified parameter of Chen and Bier (1972). The specimens were prepared by conventional procedure of the laboratory of TEM and were observed by Hitachi H600. Results Besides the germinal cell stage (S1), this is the first chronological study on the morphological development of S. japonicum cercaria from S2 S5 concerning the tegument and its elements (glycocalyx, sensory papilla, basal lamina and spine), head and acetabular gland and musculature of the body and tail. This article discusses the ultrastructural morphological differences from prior authors and emphasizes on the postacetalar gland and the pattern of tail musculature of the matured stage cercaria. Conclusions According to the cell division and differentiation, the process of development may be divided into 5 stages: S1, the single germinal cell stage; S2, the germinal ball stage; major multiplication and minor differentiation; S3, tail budding embryonic stage, both active in multiplying and differentiating; S4, the pre matured stage, major differentiation and minor multiplication; and S5, the fully matured stage (completing the differentiation).
基金financially supported by the Human Resource Development in Science Project(Science Achievement Scholarship of Thailand,SAST)partially supported by Chiang Mai University.
文摘Objective:To investigate the genetic diversity and population structure of the helminth intermediate host,Anentome(A.)helena,populations from northern Thailand and reveal the effect of parasitic infection on snail host populations.Methods:A.helena was collected throughout river drainages from northern Thailand and investigated for larval trematode infection.Seventy-three A.helena specimens were used to examine and re-construct the molecular phylogenetics(neighbour-joining,maximum likelihood,and Bayesian inference methods),genetic diversity,and population structure based on the concatenated datasets of COI and 16S rRNA(1178 bp).Results:The infection of zoogonid cercariae,heterophyid metacercariae,and echinostome metacercariae in the snail was found from 2 localities,including Mae Sa and Mae Pu Kha populations,with a total prevalence of 6.7%and 16.3%,respectively.The phylogenetic tree and haplotype network revealed three main clades and three haplogroups for each region(COI and 16S rRNA).High levels of genetic diversity in terms of haplotype diversity were found in parasitic infected populations.Conclusions:This is the first time that Echinostoma mekongi has been discovered from A.helena in northern Thailand.The results also showed that parasitic infection can affect the population genetic structure of the snail intermediate host,A.helena.Moreover,habitat and historical demographic changes could be other important factors that can affect the degree of population genetic structure in A.helena.Understanding the population structure of intermediate host species can inform parasite transmission dynamics and is useful for the creation of effective public health management strategies.
文摘Objective:To detect the species of larval trematodes(cercariae) in Melanopsis praemorsa snails from 5 different fresh water bodies in Palestine.Methods:A total of 1 880 Melanopsis praemona snails were collected from different fresh water bodies in Palestine from October,2008 to November,2010.Cercariae in Melanopsis praemona snails were obtained by lighting and crushing methods.The behavior of cercariae was observed using a dissecting microscope.Results:Three different species of larval trematodes were identified from Melanopsis praemona snails collected only from Al-Bathan fresh water body,while snails from other water bodies were not infected. These species were microcercous cercaria,xiphidiocercaria and brevifurcate lophocercous cercaria.These cercariae called Cercaria melanopsi palestinia 1,Cercaria melanopsi palestinia H and Cercaria melanopsi palestinia III have not been described before from this snail in Palestine. The infection rate of Melanopsis praemona collected from Al-Bathan fresh water body was 5.7%, while the overall infection rate of snails collected from all fresh water bodies was 4.3%.Details are presented on the morphology and behavior of the cercariae as well as their development within the snail.Conclusions:These results have been recorded for the first time and these cercariae may be of medical and veterinary importance.
