弓形虫主要表面抗原P30基因克隆与表达

将液氮保存的弓形虫 Ｎ Ｔ 株经小鼠复壮后,取腹腔液提取弓形虫基因组 Ｄ Ｎ Ａ,采用 Ｐ Ｃ Ｒ 方法从弓形虫 Ｎ Ｔ 株中扩增出约 ８００ ｂｐ 的片段。产物经 Ｅｃｏ Ｒ Ｉ和 Ｘｂａ Ｉ酶切后,克隆到 ｐ Ｕ Ｃ１９ 载体中,构建了 ｐ Ｂ Ｖ Ｐ３０ 非融合表达质粒和 ｐ Ｅ Ｔ Ｐ３０ 融合表达质粒。ｐ Ｂ Ｖ Ｐ３０ 转化到宿主菌 Ｄ Ｈ５α、ｐ Ｅ Ｔ Ｐ３０ 转化到宿主菌 Ｂ Ｌ２１ （ Ｄ Ｅ３）后,分别经温控诱导和 Ｉ Ｐ Ｔ Ｇ 诱导,产物经 Ｓ Ｄ Ｓ Ｐ Ａ Ｇ Ｅ 分析,ｐ Ｂ Ｖ Ｐ３０ 未发现表达产物,ｐ Ｅ Ｔ Ｐ３０ 出现约 ３０ ０００ 的产物。 Ｗ ｅｓｔｅｒｎ ｂｌｏｔｔｉｎｇ 显示,该蛋白与兔抗弓形虫血清发生特异性反应;薄层扫描显示,该蛋白占菌体总蛋白的 ２０％ 以上。     

弓形虫病的研究进展

弓形虫病给畜牧业的健康发展和公共卫生安全带来巨大威胁。本文结合国内外对弓形虫病的研究结果,对其研究进展进行了综合论述,并对进一步的研究作了展望,为有效防控该病提供参考。     

弓形虫病疫苗研究进展

综述了弓形虫病减毒和灭活疫苗、基因工程亚单位疫苗、核酸疫苗及活载体疫苗的研究进展。弓形虫是专性细胞内寄生原虫,全球分布,能引起人兽共患的弓形虫病。弓形虫病疫苗的研制对于弓形虫病的防治工作有着极其重要的意义。     

鸟弓形虫病的研究进展

刚地弓形虫(Toxoplasma gondii)是一种可寄生于包括鸟类在内的多种宿主、引起人兽共患弓形虫病的原虫.本文就鸟弓形虫病的发现、临床症状、血清学诊断、鸟弓形虫病人工感染模型的建立进行了概述,特别是对近年来基于PCR技术而发展起来的分子生物学技术在其基因型鉴定上的应用作了介绍.表明鸟类自然感染弓形虫病在世界范围内呈流行趋势,值得我们重视.     

弓形虫疫苗研究进展

弓形虫病是一种分布范围广、危害严重的人兽共患寄生虫病,不仅影响畜牧业发展以及肉食品卫生安全,而且也对人类公共卫生安全造成了极大的危害,因此疫苗的研制对于弓形虫病的防治工作意义重大。文章就弓形虫疫苗的研究现状进行综述,旨在为弓形虫疫苗的研制提供参考。     

猪弓形虫病研究进展

猪对许多国家的经济很重要,是人类食物的来源。在许多国家,受感染的猪肉是人和动物弓形虫感染的来源。这种寄生虫也会导致猪,特别是新生猪死亡。大多数猪在出生后由于摄入受污染环境的卵囊或摄入动物感染组织而被弓形虫感染。本文综述了猪的临床和无症状感染报告,比较了检测猪弓形虫的方法,对最近关于流行病学中关于猪流行的弓形虫菌株遗传分型的研究进行了讨论[1]。     

野生动物弓形虫病研究进展

弓形虫是一种重要的胞内寄生原虫,可以感染几乎所有的温血动物。食肉性动物中猫科动物对弓形虫病的传播起着重要的作用,它是惟一的可以排泄含有弓形虫的卵囊到环境中的动物,进而使全世界的人和动物都有机会感染弓形虫病,而草食性动物和杂食性动物则是重要的中间宿主。然而野生动物感染弓形虫病之后往往缺少明显的临床症状,很难在第一时间判断出是否感染弓形虫。野生动物关系到整个生态系统的稳定,对整个生物界起着至关重要的作用,应该引起我们极大的关注和重视。论文就弓形虫在野生动物中的感染情况及临床症状加以概括,为其在野生动物中的流行趋势、诊断和防控的深入研究提供资料。     

弓形虫病研究进展

弓形虫为人兽共患性寄生原虫,是一类重要的水源性和食源性专性细胞内寄生的寄生虫,能够感染几乎所有的温血动物,是引起人和动物流产的一个重要致病因素。弓形虫病世界五大洲各地均有报道,严重威胁着人类和动物的健康。论文对弓形虫病的病原体、流行病学、危害、诊断和防控等方面的研究进展进行了全面综述,为弓形虫病的相关研究和有效防控提供参考。     

弓形虫病的治疗研究进展

弓形虫病是一种世界性的人兽共患寄生虫病,不仅给畜牧业生产带来了严重危害,也严重影响人类健康。弓形虫病分为先天性弓形虫病和获得性弓形虫病,其中眼弓形虫病和脑弓形虫病是最常见的临床表现。乙胺嘧啶和磺胺嘧啶的联合用药是目前治疗弓形虫病的黄金标准。免疫力强的弓形虫病患者一般无需治疗,但对免疫功能不全或低下的患者可能会造成致命后果,需要长期监测和治疗。本文从先天性与获得性弓形虫病、弓形虫病临床用药、临床常见治疗方法以及新型抗弓形虫药物的研究进行综述。     

弓形虫病流行病学及防疫措施研究进展

弓形虫病流行病学及防疫措施研究进展李秉鸿广西兽医研究所５３０００１刚地弓形虫（Ｔｏｘｏｐｌａｓｍａｇｏｎｄｉ）是寄生于人类和许多动物组织细胞内的原虫，可侵犯脊椎动物的多种细胞，并在细胞内繁殖，最后破坏宿主细胞，释放出虫体，导致一系列病理变化。弓形虫病...     

弓形虫实验室诊断技术研究进展

弓形虫病（toxoplasmosis）是由刚地弓形虫（Toxoplasma gondii）感染引起的一种人兽共患寄生虫病,在世界各地都有分布,宿主种类十分广泛,人间和动物的感染率都比较高,对人类健康和公共卫生构成了威胁。目前,弓形虫病没有理想的药物用于治疗,早期诊断是防控该病的重要手段之一。病原学诊断是最早建立的弓形虫诊断方法,操作简单,结果可靠,但耗时较长,无法满足快速、高效、批量检测的需求,因此在实验室诊断方面应用逐步减少。免疫学检测方法以ELISA和IHA方法为主,已有较为成熟的商品化试剂盒或诊断试剂,在大规模的检测和流行病学调查中起到了很大作用。分子生物学方法与传统的病原学检测方法相比,其速度、通量、特异性、敏感性各方面都有提升,相较于免疫学诊断,避免了获得性免疫带来的假阳性结果,是未来弓形虫病实验室诊断的主要发展趋势。本文从病原学、分子生物学和免疫学方面对弓形虫的实验室诊断技术进行论述,以期为弓形虫的实验室诊断和防控措施的制定提供参考。     

弓形虫微线体蛋白研究新进展

微线体蛋白(microneme protein,MIC)是由位于弓形虫前端的微线体(microneme)分泌产生的,具有识别、黏附与侵染宿主细胞的性质。近年来研究结果证明,弓形虫的多种微线体蛋白在侵染宿主的过程中发挥重要作用,并且可以作为抗弓形虫病的疫苗候选分子。作者对目前研究较多的弓形虫微线体蛋白进行综述,为弓形虫疫苗的研究提供理论依据。     

Production of antibodies in murine mucosal immunization with Toxoplasma gondii excreted/secreted antigens

Toxoplasmagondii RH strain excreted/secreted antigens (ESA) were administrated weekly by the oral route, to two groups of 40 OF1 mice for 4 weeks. One group received ESA associated with cholera toxin (CT+) and the other, ESA only (CT-). Five animals from each group were sacrificed from day 4 (D4) to D49 following the first immunization and their feces and sera were collected and tested by ELISA for IgA, IgG and IgM antibody detection. In feces, IgA antibodies were detected on D4 and on D12 in the CT+ and CT- groups, respectively, and they persisted up to D49. IgG antibodies were detected from D12 to D41 in the CT+ group and on D12 only in the CT- group. No IgM antibodies were detected. In sera, IgA antibodies were detected on D27, D41 and D49 only in the CT+ group. IgG and IgM antibodies were found on D12 and D4, respectively, in the CT+ group and starting from D27 in the CT- group. To our knowledge, this is the first demonstration that ESA, with or without CT, are immunogenic when administrated by the oral route.     

山羊免疫弓形虫代谢分泌抗原后的免疫应答

将山羊随机分为6组,即E／SA纽、E／SA＋cpG组（未乳化）、E／SA＋CpG组、E／SA＋IL-2组、E／SA＋IL-2＋CpG组、对照组,每组3只,分别于免疫前、免疫后第2周、免疫后第4周、感染后第1周及感染后第2周采集山羊外周全血及涂血片,IHA法检测抗体滴度的动态变化;ELISA法检测IFN-γ、TNF—a、IL-2、IL-4的表达水平;ANAE染色法检测T淋巴细胞的动态变化。结果显示,免疫后各免疫组的IFN-γ、TNF-a、IL-2、IL-4及T淋巴细胞水平显著高于对照组（P〈0．05）,各免疫组的抗体水平均高于对照组。结果表明,E／SA可引起IFN—γ、TNF—a、IL-2、IL-4、T淋巴细胞及抗体水平的升高,说明E／SA免疫后可引起山羊较强的细胞免疫和体液免疫应答。     

Detection of Toxoplasma gondii in free-range chickens in China based on circulating antigens and antibodies

Toxoplasma gondii is widely distributed in humans and other animals including domestic poultry throughout the world, but the data on prevalence of T. gondii in free-ranged (FR) chickens in People's Republic of China (PRC) are limited. In the present study, the seroprevalence of T. gondii infection in FR chickens was investigated in 13 provinces/municipalities of China during the period from January to June 2010. A total of 1173 serum samples were collected and assayed for T. gondii circulating antigens (TCA) and antibodies (TCAb) using enzyme linked immunosorbent assay (ELISA) technique. Out of this number, 199 samples were TCA positive (16.97%), 226 samples were TCAb positive (19.27%), 69 samples were positive for both TCA and TCAb (5.88%), and the total seropositive rate was found in 356 of 1173 (30.36%). The results of the present survey indicated that infection with T. gondii in FR chickens is widely spread in China.     

细胞渗透肽TAT与3种弓形虫抗原的融合重组表达

为了探索新型弓形虫疫苗的传递系统,本试验分别构建了细胞渗透肽反式转录激活因子(TAT)与3种弓形虫抗原和增强型绿色荧光蛋白(EGFP)融合表达的重组质粒,即pET28a-TAT-EGFP,pET28a-TAT-SAG1,pET28a-TAT-GRA4和pET28a-TAT-AMA1质粒。重组质粒被转化到大肠杆菌中并通过IPTG诱导,成功进行了融合表达,表达产物采用Ni-NTA树脂进行纯化,然后进行SDS-PAGE分析。结果得到31 ku的TAT-EGFP融合蛋白、34 ku的TAT-SAG1融合蛋白、38 ku的TAT-GRA4融合蛋白和62 ku的TAT-AMA1融合蛋白,经过Western blotting分析,感染弓形虫的小鼠血清可特异性地识别融合TAT的SAG1、GRA4蛋白和微弱地识别TAT-AMA1蛋白。     

Immune responses against excreted/secreted antigens of Toxoplasma gondii tachyzoites in the murine model

In the present study, excretory secretory antigens (ESA) of Toxoplasma gondii were evaluated in immunization of 8-10 week inbred female Balb/c mice. Tachyzoites of the parasite were cultured in cell-free incubation medium (RPMI-1640), and then supernatant of the medium was loaded on an ion-exchange chromatography column. Two fractions (ESA-F(1) and ESA-F(2)) were collected from the column. For immunization of the mice, 50 were allocated into 5 groups of 10. The first, second, third, and fourth groups were immunized, twice with total-ESA, ESA-F(1), ESA-F(2) or toxoplasma lysate antigen (TLA), respectively. The fifth group was selected as a negative control group (non-immunized). The virulent RH strain of Toxoplasma gondii was used to challenge. Delayed-type hypersensitivity responses (DTHs) were measured by intra-footpad injection measuring induration at timed intervals. Lymphocyte transformation tests (LTTs) were done on lymph node cells using [3H] thymidine incorporation as an indication of reactivity. Peritoneal macrophages from sensitized mice were stimulated and nitric oxide was measured by Griess method. The ESA-F(1) and ESA-F(2) fractions were separated on poly acrylamide gel electrophoresis (PAGE) and SDS-PAGE. ESA-F(1) had 4 bands on PAGE and 14 bands on SDS-PAGE. ESA-F(2) had one band on PAGE and two bands on SDS-PGE. Sensitized mice showed DTH and lymphocyte transformation responses to total-ESA, ESA-F(1), and ESA-F(2) and peritoneal macrophages produce nitric oxide following stimulation. In challenge experiments, all non-immunized mice died within 10 days, whereas immunized mice survived for longer time periods (P<0.05). The highest survival rate was observed in mice that immunized with ESA-F(2). We suggest that these antigens especially ESA-F(2) should be of value for the development of new strategies for immunization against toxoplasmosis.     

Antibody responses to Toxoplasma gondii antigens in aqueous and cerebrospinal fluids of cats infected with T. gondii and FIV.

Antibodies to antigens of Toxoplasma gondii were measured in the aqueous and cerebrospinal fluid (CSF) of 16 specific-pathogen free kittens experimentally infected with feline immunodeficiency virus (FIV), T. gondii, or both pathogens. The results indicated that all cats infected with T. gondii had antibody responses to antigens of T. gondii in both aqueous fluids and CSF. Co-infection with FIV did not affect antibody levels. Aqueous fluids from eyes of cats with toxoplasmic retinochoroiditis did not necessarily have higher antibody levels than those from eyes without lesions. Antibodies to T. gondii were also detected in the CSF of two cats from whose brains no parasites were isolated by in vivo mouse inoculation. Total IgG did not increase significantly in the aqueous fluids and CSF of cats infected with T. gondii whether or not they were also infected with FIV.