华支睾吸虫代谢抗原用于免疫诊断的试验研究

有人报道用华支睾吸虫成虫代谢抗原检测兔体内抗体取得了较满意的结果。本文应用了无血清1640培养液内活虫培养的代谢产物作为抗原与华支睾吸虫全虫抗原进行比较,评价代谢抗原在检测华支睾吸虫抗体中的价值。一、材料与方法(一)抗原制备1.代谢抗原:自人工感染华支睾吸虫囊蚴的猫肝脏中无菌取出华支睾吸虫成虫,用含双抗的无菌生理盐水漂洗后,分别装于100ml 含有1640培养液的三角烧     

同一鱼塘中3种鱼华支睾吸虫囊蚴感染度的调查

珠江三角洲是华支睾吸虫 (Clonorchis sinensis)的主要分布区。虽然随着人们防病意识的增强 ,使许多地区塘鱼的华支睾吸虫的感染有所降低 ,但由于当地人群独特的饮食习惯使华支睾吸虫的感染仍然普遍存在 [1 ]。本实验对广东顺德市杏坛镇同一鱼塘中的 3种鱼进行了华支睾吸虫囊蚴感染度的调查 ,以了解不同鱼类对华支睾吸虫囊蚴的感染情况。1　材料和方法1.1　鱼的来源　 3种鱼均来源于广东省顺德市杏坛镇某同一鱼塘。选择其中的草鱼 (Ctenopharygodon idellus) 5 0条 (平均体重 2 .8g/条 )、鳙鱼 (Aristichthys nobilis) 5 0条 (平均体重 9.…     

7种抗蠕虫药物的体外抗华支睾吸虫作用

目的观察吡喹酮、三苯双脒、左旋咪唑、蒿甲醚、青蒿琥酯、阿苯达唑和甲苯达唑体外对华支睾吸虫成虫的作用。方法 70只大鼠于感染华支睾吸虫囊蚴(50～100个/只)5～7周后解剖,从胆总管内采集华支睾吸虫成虫,亨氏盐平衡溶液培养。取24孔培养板,每孔放置华支睾吸虫3～4条,加入不同浓度的上述药物,于药物处理后1、4、24、48和72 h,在倒置显微镜下观察成虫的活动和形态变化。结果吡喹酮可迅速减弱华支睾吸虫的活动,使口吸盘丧失吸附皿壁能力,虫体蜷缩、皮层出现空泡。吡喹酮对华支睾吸虫的最低致死浓度为0.1μg/ml。三苯双脒0.5、1和10μg/ml组虫体接触药物后迅速麻痹伸直呈松弛状。三苯双脒对华支睾吸虫的最低致死浓度为0.05μg/ml。左旋咪唑10和20μg/ml组华支睾吸虫的活动经药物作用后逐渐减弱,虫体松弛,但48h后,大部分虫体和口吸盘明显恢复活动。左旋咪唑的浓度高达50μg/ml时,虫体立即伸直麻痹,其表现与三苯双脒组相仿。蒿甲醚和青蒿琥酯10μg/ml和50μg/ml组虫体和口吸盘的活动减弱,虫体收缩,继则松弛,体表出现空泡,培养72 h后两组均有半数以上虫体死亡。阿苯达唑和甲苯达唑10μg/ml和5...     

三苯双脒、青蒿琥酯和蒿甲醚抗华支睾吸虫及其他吸虫的实验研究进展

吡喹酮是目前用于治疗血吸虫病和其他吸虫病的主要药物。近年的实验研究发现,治疗肠道线虫感染的新药三苯双脒具有抗华支睾吸虫、麝猫后睾吸虫和卡氏棘口吸虫(Echinostoma caproni)的作用,特别是对感染华支睾吸虫的大鼠,其顿服接近治愈的剂量为300 mg/kg,低于吡喹酮的治愈剂量375～500 mg/kg。抗疟药青蒿琥酯和蒿甲醚不仅已被发展为预防血吸虫病的药物,而且发现它们对多种吸虫有效,特别是对华支睾吸虫,其顿服对大鼠华支睾吸虫感染的治愈或高效剂量为75 mg/kg。本文系综述这些药物单用或联合用药治疗大鼠、小鼠、仓鼠或犬的华支睾吸虫及其他吸虫感染的实验研究进展。     

华支睾吸虫病荧光抗体试验的初步观察

1984年以来,我们应用华支睾吸虫成虫冷冻切片为抗原,对华支睾吸虫病患者和正常人及动物进行了初步观察。 材料与方法 抗原制备 从感染华支睾吸虫囊蚴后30d的家兔胆管中取出成虫,用生理盐水清洗。为方便于切片,另取一正常小鼠肝,洗净后用刀片切开一道裂缝,然后将成虫插入间隙内,在-10℃下切成5μm     

广州珠三角地区家猫自然感染华支睾吸虫的调查及豚鼠模型的构建北大核心CSCD

目的了解广州珠三角地区家猫自然感染华支睾吸虫状况,观察豚鼠感染华支睾吸虫后肝组织病理变化。方法解剖当地家猫,取肝脏,检查华支睾吸虫感染情况,阳性肝脏收集成虫,并取肝组织制作切片;采集阳性鱼,分离华支睾吸虫囊蚴,经口感染豚鼠,60个囊蚴/只,60d后解剖豚鼠,检查肝脏,收集成虫,取病变肝组织制作切片,作病理检查。实验设未感染对照组。结果当地猫华支睾吸虫自然感染率为41.47%(214/516);感染豚鼠华支睾吸虫成虫回收率50.83%(305/600)。与对照组比较,实验组豚鼠肝脏肿大,边缘呈球状隆起,部分肝叶表面可见水泡状凸起病变,水泡内液清亮;病变组织横切面可见胆管管壁增厚,管腔内有华支睾吸虫成虫寄生;镜下可见华支睾吸虫虫卵切面,周围大量炎症细胞浸润,胆管管壁增厚,管周纤维组织增生伴胆管扩张,肝小叶结构破坏。结论广州珠三角地区家猫华支睾吸虫感染率较高。豚鼠是华支睾吸虫合适的终末宿主,且肝脏病变明显。     

吡喹酮治疗后华支睾吸虫的组织化学变化

吡喹酮是目前治疗华支睾吸虫病的最有效药物。关于该药对华支睾吸虫的杀虫作用,迄今仅见虫体组织学方面的研究,为了进一步探讨该药对华支睾吸虫的作用机理。本文进行了组织化学方面的研究。     

亲和层析法纯化华支睾吸虫抗原的研究

目的探索华支睾吸虫特异性抗原的纯化方法。方法从华支睾吸虫病流行区的淡水鱼中分离华支睾吸虫囊蚴,接种动物,收集成虫,制备匀浆液(粗抗原),借助偶联华支睾吸虫病人血清抗体的Sepharose4B亲和层析柱,提取特异性抗原,用浓度梯度聚丙烯酰胺凝胶电泳(CG-PAGE)和免疫印迹方法鉴定纯化抗原的纯度和特异性。结果亲和层析法纯化抗原经CG-PAGE显示5条蛋白带,免疫印迹试验鉴定均有抗原性,强阳性带蛋白分子质量单位为31ku,各区带蛋白与并殖吸虫、血吸虫血清抗体无交叉反应。粗抗原有16条蛋白带。结论经亲和层析法纯化的华支睾吸虫抗原特异性高。亲合层析是获取华支睾吸虫诊断用抗原的较佳方法。     

吡喹酮对华支睾吸虫的皮层、表皮细胞和肌肉影响的超微结构观察

我们在进行吡喹酮治疗华支睾吸虫病的实验研究中,发现感染大鼠一次口服吡喹酮300mg/kg后12小时,虫体即发生明显的挛缩、卷曲,表皮有轻重不同的皱褶,这提示本药对虫体体壁有损伤作用。关于本药对华支睾吸虫超微结构的影响,迄今只见Rim等的报道,本文对吡喹酮损害的华支睾吸虫的皮层、表皮细胞和肌纤维进行了动态观察。     

吡喹酮对体外培养华支睾吸虫的作用

体外培养寄生虫是进行实验研究的有效方法之一.目前已被广泛应用于寄生虫学的研究。为寻求简易的华支睾吸虫体外培养方法,我们进行了本实验,同时观察了吡喹酮的杀虫作用. 材料与方法 一、从江苏邳县购得的麦穗鱼体内分离华支睾吸虫囊蚴,感染家兔,两个月后从肝胆管中取成虫,     

日本血吸虫副肌球蛋白全基因核酸疫苗在小鼠诱生的保护性免疫力

目的： 研究日本血吸虫大陆株副肌球蛋白全基因核酸疫苗免疫Ｃ５７ ＢＬ／６ 及ＢＡＬＢ／ｃ 小鼠诱生的保护性免疫力。方法： 将构建的日本血吸虫大陆株副肌球蛋白全基因核酸疫苗 （ｐＣＭＶ－ＳｊＣ９７） 经后腿胫前肌免疫Ｃ５７ＢＬ／６及ＢＡＬＢ／ｃ小鼠, 共免疫３ 次, 每次间隔３ ｗ ｋ。末次免疫后３ ｗ ｋ 以血吸虫尾蚴攻击感染, ６ ｗｋ 后计数成虫负荷及肝、脾、肠组织虫卵数。设不含ＳｊＣ９７ 编码基因的空载体质粒免疫组为对照组。结果： ｐＣＭＶ－ＳｊＣ９７ 免疫Ｃ５７ ＢＬ／６ 小鼠主要诱生ＩｇＧ２ａ 和ＩｇＧ２ｂ 亚类, 而免疫ＢＡＬＢ／ｃ小鼠除诱生ＩｇＧ２ａ 和ＩｇＧ２ｂ 亚类外, 还诱生ＩｇＧ１。ｐＣＭＶ－ＳｊＣ９７ 免疫Ｃ５７ＢＬ／６ 小鼠诱生较明显的减虫率 （３５．５％ ～４１．１％ ） 和减卵率 （肝、脾及肠组织减卵率分别为４４．５％ ～５９．６％ 、５６．７％ ～８２．４％ 及５７．９％ ）, 而对ＢＡＬＢ／ｃ小鼠未诱生保护性。结论： ｐＣＭＶ－ＳｊＣ９７ 核酸疫苗能对Ｃ５７ＢＬ／６ 小鼠诱生较明显保护性免疫力, 而对ＢＡＬＢ／ｃ 小鼠未诱生免疫保护力     

SjAWMAg免疫血清和吡喹酮联用体内杀不同发育期血吸虫的实验研究

目的　观察日本血吸虫成虫表膜抗原(SjAWMAg)免疫血清与吡喹酮合用对不同发育期血吸虫的杀虫效果,探讨体液免疫在吡喹酮杀虫中的作用。　方法　提取 SjAWMAg,并用 SDS PAGE 对其进行鉴定,制备高滴度(1∶25 600)SjAWMAg抗血清,采用Western blot分析该血清与肺期童虫、肝期童虫及成虫抗原的反应性;将 SjAWMAg抗血清与吡喹酮合用治疗血吸虫感染后第2 d、第14 d和第35 d的小鼠,在感染后第49 d剖杀小鼠收获虫体,计算减虫率和减卵率。结果　Western blot分析表明,SjAWMAg抗血清与肺期及肝期童虫抗原存在交叉反应; 联合用药的 3 个治疗组(感染2、14、35 d)与单用吡喹酮组比较杀虫作用均有显著提高,减虫率分别提高了 38.79%、29.80%和 46.82%,其中以感染35 d治疗组提高最显著;减卵率分别提高了 62.04%,42.78%和 29.81%,以感染 2 d治疗组更为显著。结论抗SjAWMAg多克隆抗体与吡喹酮联合使用,均可提高吡喹酮对不同发育期血吸虫的杀灭效果,揭示体液免疫在吡喹酮杀血吸虫过程中具有重要作用,同时提示这种联合用药可能解决单用吡喹酮治疗血吸虫早期感染效果不佳的难题。     

噬菌体表达短肽模拟旋毛虫抗原表位及其抗血吸虫保护性免疫研究

目的　从噬菌体随机肽库中筛选出模拟旋毛虫特异性抗原表位的短肽分子 ,探讨其抗血吸虫的交叉免疫保护效果。　方法　以纯化的旋毛虫感染鼠血清IgG为配基 ,亲合筛选法富集特异性噬菌体 ,随机挑取噬菌体克隆用ELISA检测其特异性 ;混合噬菌体克隆经皮下免疫小鼠 3次 ,攻击感染后第 4 5天剖杀小鼠 ,观察减虫和减卵效果。　结果　经 3轮筛选 ,特异性噬菌体得到了有效的富集 ,第三轮洗脱噬菌体的产量约为第一轮的 15 0倍。随机挑取 2 4个噬菌体克隆经ELISA测定 ,有 2 1个克隆能与旋毛虫感染鼠血清IgG特异性反应。与对照组相比 ,混合噬菌体克隆免疫小鼠的减虫率与减卵率分别为 4 2 8%与 6 6 3% (P <0 0 0 1)。　结论　利用噬菌体随机肽库技术可获得模拟旋毛虫特异性抗原表位的短肽分子 ,这些短肽分子能诱导明显的抗血吸虫的保护性免疫。     

Artemether, artesunate, praziquantel and tribendimidine administered singly at different dosages against Clonorchis sinensis: a comparative in vivo study

We comparatively assessed the in vivo efficacy of artemether, artesunate, praziquantel and tribendimidine against different stages of Clonorchis sinensis. Rats were infected with 40-50 C. sinensis metacercariae, and drugs were administered singly by the oral route at different dosages. Rats were dissected 2-4 weeks post-treatment and C. sinensis trematodes were removed from the liver and bile ducts and counted. We used a negative binomial regression model to test the effect of drug and dosage in terms of worm burden reduction. Single 150 mg/kg oral doses of artesunate, artemether, tribendimidine and praziquantel, administered to rats infected with adult C. sinensis, resulted in mean worm burden reductions of 100, 100, 89.5 and 80.7%, respectively (all P<0.001). Halving the dose to 75 mg/kg still resulted in highly significant worm burden reductions for artesunate, artemether and tribendimidine (71.4-100%), but not for praziquantel (20.7%). In the juvenile infection model, a single 150 mg/kg oral dose of tribendimidine and praziquantel resulted in mean worm burden reductions of 99.1 and 90.0%, respectively, whereas considerably lower reductions were observed for artemether (59.2%) and artesunate (57.6%) when used at the same single dose. The in vivo results presented here with the artemisinins and tribendimidine provide further data for clinical investigations to assess the safety and efficacy of these drugs in clonorchiasis patients.     

最大耐受剂量卡维地洛对充血性心力衰竭的干预作用及其抗氧化效应

目的 :观察第三代 β 受体阻滞剂卡维地洛治疗国人充血性心力衰竭 (CHF)患者的最大耐受剂量 ,临床疗效及其抗氧化作用。方法 :CHF患者 4 0例 (扩张型心肌病 2 8例 ,缺血性心肌病 12例 ) ,随机分为常规治疗组 (C组 ,18例 ,予以洋地黄、利尿剂和ACEI等扩血管药物 )与卡维地洛治疗组 (CA组 ,2 2例 ,在常规治疗基础上加用卡维地洛 )。治疗前和治疗 3个月后对患者的超声心动图、6min步行试验、心功能以及氧化应激等指标进行检测 ,用药期间记录所有的药物副作用。结果 :CA组卡维地洛的最大耐受剂量为 ( 2 8 75± 7 2 3)mg d( 10～ 4 0mg d)。 2组患者心功能分级均有显著改善 ,但CA组的临床疗效好于C组。CA组SOD、GSH均有明显升高 (P <0 0 5 ,P <0 0 0 1) ,GSH含量也显著高于常规治疗组 (P<0 0 5 ) ;MDA则明显下降 (P <0 0 5 )。常规治疗组下降不明显。结论 :本组HCF患者卡维地洛治疗的平均最大耐受剂量为 ( 2 8 75± 7 2 3)mg d。卡维地洛治疗有利于改善CHF患者的心功能和心室重构。国人CHF患者长期服用卡维地洛有良好的耐受性。卡维地洛对CHF的治疗作用可能部分地与抗氧化效应有关     

Induction of immunity in mice to Fasciola hepatica with a Fasciola/Schistosoma cross-reactive defined immunity antigen

The paradox of schistosomiasis is that infection confers immunity to its host, yet immunization with subcellular antigens of the parasite does not, in general, induce protective immunity. Infection or immunization with subcellular antigens of Fasciola hepatica confers high levels of immunity to a challenge infection with another trematode, Schistosoma mansoni. We have isolated by antibody affinity chromatography a Fasciola hepatica/Schistoma mansoni cross-reactive antigen, designated FhSmIII(M), and also have shown that this antigen confers immunity in mice to a challenge infection with S. mansoni. This antigen was compared with a crude F. hepatica worm extract (FhWWE) as to its ability to induce an IgG antibody response in mice, and to determine whether it had a protective effect in mice to a challenge infection with F. hepatica metacercariae. Mice immunized with FhSmIII(M) or FhWWE, and subsequently infected with F. hepatica, developed higher IgG antibody levels to FhSmIII(M), as measured by ELISA, than F. hepatica-infected controls. Mice immunized with FhWWE did not develop significant levels of resistance to challenge with F. hepatica metacercariae. Mice immunized with FhSmIII(M) and infected with F. hepatica metacercariae developed 69%-78% less worms than controls. An F. hepatica/S. mansoni cross-reactive, cross-protective defined immunity antigen confers in mice significant levels of protection to a challenge infection with F. hepatica.     

Active immunization of mice with Schistosoma mansoni worm membrane antigens enhances efficacy of praziquantel

The efficacy of praziquantel treatment was significantly enhanced (P < 0.01) in CBA/Ca mice that had been immunized prior to Schistosoma mansoni infection with a crude extract of worm membrane antigens. In Western immunoblots sera from the worm antigen-immunized animals had a poly specific antibody response, with a 25–27 kDa antigen being reacted against with particular intensity. A molecule of similar size was also recognized by rabbit antisera raised against an antigen with esterase activity that has been previously identified as a sensitive target for drug-antibody synergy. The increase in efficacy of subcurative doses of praziquantel in immunized animals is attributed to drug-induced tegumental damage causing antigens to become exposed on the worm surface. Thus, specific antigens, including the 25–27kDa antigen, become accessible to circulating schistosomicidal antibodies. The role of antibodies that can synergize with praziquantel to kill schistosome worms is discussed.,     

Trichinella spiralis: Low vaccine potential of glutathione S-transferase against infections in mice

We have previously reported that Trichinella spiralis glutathione-S-transferase (TsGST) gene is an up-regulated gene in intestinal infective larvae (IIL) compared to muscle larvae (ML). In this study, the TsGST gene was cloned, and recombinant TsGST (rTsGST) was produced. Anti-rTsGST serum recognized the native TsGST by Western blotting in crude antigens of ML, adult worm (AW) and newborn larvae (NBL) of T. spiralis, but not in ML excretory–secretory (ES) antigens. Expression of TsGST was observed in all different developmental stages (IIL, AW, NBL and ML). An immunolocalization analysis identified TsGST in the cuticle, stichosome and genital primordium of the parasite. The rTsGST had GST enzymatic activity. After a challenge infection with T. spiralis larvae, mice immunized with rTsGST displayed a 35.71% reduction in adult worms and a 38.55% reduction in muscle larvae. The vaccination of mice with rTsGST induced the Th1/Th2-mixed type of immune response with Th2 predominant (high levels of IgG1) and partial protective immunity against T. spiralis infection.     

Synergistic action of praziquantel and host specific immune response against Schistosoma mansoni at different phases of infection

The interaction between specific immune response to Schistosoma mansoni and praziquantel (PZQ) was studied in mice. In mice harboring concomitant immunity, 6-day-old parasites treated with PZQ were more effectively removed than 24 h treated parasites despite both had a significant worm burden reduction when compared with respective treated controls. These results show that PZQ can be effective at the skin and lung stages of parasite's development mainly acting with a established specific immune response, and particularly at the lung phase.     

Therapeutic effect of subcurative dose praziquantel on Schistosoma mansoni infected mice and resistance to challenge infection after treatment

The therapeutic effect of a subcurative dosage of praziquantel (PZQ) on Schistosoma mansoni infected mice and resistance to challenged worm infection after treatment were assessed and compared with conventional treatment using a curative dosage of PZQ. S. mansoni infected mice were treated with PZQ at a curative dosage (600 mg kg(-1)) or a subcurative dosage (300 mg kg(-1)) at 9 weeks after infection. Untreated mice and non-infected mice were added as controls. The therapeutic effect of the drug was evaluated in terms of the mortality of mice after treatment, and the parasitological and pathological findings in mice sacrificed at 1 week, 1 month, or 3 months after treatment. Another sample of mice was not killed but challenged with S. mansoni cercariae at 1 week, 1 month, or 3 months after treatment. Resistance to re-infection was evaluated by the extent of challenged worm reduction. In conclusion, there was no significant difference in mortality, or parasitological and pathological findings between mice treated with PZQ at the two dosages. However, resistance to challenged worm infection was more sustained in the group treated with subcurative dose PZQ, especially at 3 months after treatment.