本文選題：剛地弓形蟲 + 酶聯(lián)免疫吸附試驗　； 參考：《吉林大學(xué)》2011年碩士論文

【摘要】：剛地弓形蟲(Toxoplasma gondii)是一種專性細胞內(nèi)的頂復(fù)門原蟲,能感染所有的恒溫脊椎動物,包括哺乳動物和鳥類。剛地弓形蟲在人群中廣泛分布,世界上1/3的人口呈慢性感染。根據(jù)不同的環(huán)境特點和飲食習(xí)慣,血清陽性率在不同地理區(qū)域中是不同的:在歐洲,育齡期女性血清陽性率在英國為10-18%,西班牙為19-29%,意大利為40%,法國為55%。通過食用未煮熟的或生的被感染的肉(例如豬肉,羊肉)或通過貓排泄的卵囊污染的食物,水,或未洗的蔬菜而感染弓形蟲。感染弓形蟲常見但通常無癥狀,但是卻對免疫受損的患者如癌癥病人、艾滋病、器官移植患者等產(chǎn)生威脅。因為孕期病原會通過胎盤傳播給胎兒,所以孕期初次和再次感染尤其對胎兒來說是高危因素。先天性弓形蟲病會導(dǎo)致許多各種不同的臨床表現(xiàn),從輕微癥狀,視覺缺陷或視網(wǎng)膜脈絡(luò)膜炎的形成到嚴重的腦積水,小頭畸形,智能缺陷,甚至死亡。 在南美洲和歐洲,剛地弓形蟲是高度克隆的,有三種主要的系譜組成(I型,II型和III型),然而,在南美洲,分布于人群中的一些蟲株呈高度多樣性,這些蟲株在生物學(xué)和遺傳方面不同于南美和歐洲的蟲株,這些顯示弓形蟲種群整體的多樣性很高。目前,關(guān)于亞洲剛地弓形蟲多樣性的數(shù)據(jù)很有限。 在這項研究中,多基因位點聚合酶鏈反應(yīng)-限制性片段長度多態(tài)性(PCR-RFLP)分析被用于確定與中國人弓形蟲病相關(guān)的剛地弓形蟲(I型、II型、III型)不同基因型的流行情況。 首先,通過間接酶聯(lián)免疫吸附試驗(ELISA)檢測血清樣本中抗剛地弓形蟲IgG抗體的水平。然后,從IgG抗體陽性孕婦外周血中提取DNA樣本,通過聚合酶鏈反應(yīng)(PCR)檢測剛地弓形蟲529bp重復(fù)序列。最后,用12個基因標記物包括SAG1, 5’SAG2, 3’SAG2, alt.SAG2,SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1和APICO通過多基因位點聚合酶鏈反應(yīng)-限制性片段長度多態(tài)性(PCR-RFLP)進行剛地弓形蟲分離株的基因鑒定。 中國東北地區(qū)840份孕婦血清樣本用間接ELISA檢測抗剛地弓形蟲IgG抗體水平, 80名孕婦抗剛地弓形蟲IgG抗體為陽性的,血清陽性率為9.5%。從IgG抗體陽性孕婦的外周血中提取DNA樣本通過聚合酶鏈反應(yīng)用于檢測529bp重復(fù)序列,33份樣本為聚合酶鏈反應(yīng)陽性。通過多基因位點聚合酶鏈反應(yīng)-限制性片段長度多態(tài)性分析對陽性DNA樣本進行基因分型,8個樣本基因分型為II型。
[Abstract]:Toxoplasma gondii (Toxoplasma gondii) is a specific intracellular protozoa that infects all isothermal vertebrates, including mammals and birds. Toxoplasma gondii is widespread among people, with a third of the world's population chronically infected. According to different environmental characteristics and dietary habits, the positive rate of serum is different in different geographical regions: in Europe, the positive rate of seropositive in women of childbearing age is 10-18 in Britain, 19-29in Spain, 40 in Italy and 5555 in France. Toxoplasma gondii is infected by eating uncooked or raw infected meat (such as pork or mutton) or by excreting food, water, or unwashed vegetables. Infection with Toxoplasma gondii is common but usually asymptomatic, but poses a threat to immunocompromised patients such as cancer patients, AIDS, and organ transplant patients. As pathogens transmit to the fetus through the placenta during pregnancy, first and second infections during pregnancy are particularly high risk factors for the fetus. Congenital toxoplasmosis can lead to a wide variety of clinical manifestations ranging from mild symptoms visual defects or retinal choroiditis to severe hydrocephalus microcephaly mental retardation and even death. In South America and Europe, Toxoplasma gondii is highly cloned, with three major pedigree types, type I and III. However, in South America, some of the species of Toxoplasma gondii are highly diverse. These strains are biologically and genetically different from those in South America and Europe, indicating a high diversity of Toxoplasma gondii populations as a whole. Currently, data on the diversity of Toxoplasma gondii in Asia are limited. In this study, polygenic locus polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis was used to determine the prevalence of different genotypes of Toxoplasma gondii type I and type II associated with Toxoplasma gondii in Chinese. First, the level of IgG antibody against Toxoplasma gondii in serum samples was detected by indirect enzyme-linked immunosorbent assay (Elisa). Then, DNA samples were extracted from the peripheral blood of pregnant women with positive IgG antibody, and the 529bp repeats of Toxoplasma gondii were detected by polymerase chain reaction (PCR). Finally, 12 gene markers, including SAG1, SAG2, SAG2, alt.SAG2, BTUB, GRA6, c22-8, c29-2, L358, PK1 and APICO, were used to identify Toxoplasma gondii isolates by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Indirect ELISA was used to detect the level of IgG antibody against Toxoplasma gondii in 840 pregnant women in northeast China. The positive rate of IgG antibody against Toxoplasma gondii in 80 pregnant women was 9.5%. DNA samples were extracted from peripheral blood of pregnant women with positive IgG antibody. 33 samples of 529bp repeats were detected by polymerase chain reaction (PCR). The positive DNA samples were genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2011
【分類號】：R382.5

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