本文選題：B型流感嗜血桿菌 + 莢膜多糖��； 參考：《重慶醫(yī)科大學(xué)》2011年碩士論文

【摘要】：目的: 本研究旨在關(guān)注引發(fā)小兒嚴(yán)重侵襲性疾病病原體B型流感嗜血桿菌(Haemophilus influenzae type b,Hib),以動物為模型,評價(jià)Hib特異性莢膜多糖的免疫原性及其與抗體水平之間的劑量關(guān)系,評估PCR技術(shù)在檢測小兒呼吸道疾病中Hib的應(yīng)用價(jià)值,探討與莢膜劑量對應(yīng)的基因分型在反映菌株侵襲力方面的應(yīng)用潛力及可行性,指導(dǎo)兒童臨床用藥及接種疫苗,為研發(fā)新疫苗提供一定的實(shí)驗(yàn)基礎(chǔ)。 方法: 1.以SD大鼠為模型,分別給予不同劑量莢膜多糖的B型流感嗜血桿菌PRP-T結(jié)合疫苗免疫大鼠,設(shè)生理鹽水為對照,ELISA法測定血清中Hib多糖抗體濃度,用SAS 8.1統(tǒng)計(jì)軟件處理數(shù)據(jù),分析其Hib莢膜多糖的免疫原性及劑量效應(yīng)。 2.搜集2009-2010年重慶兒童醫(yī)院呼吸道感染患兒深部痰標(biāo)本分離的流感嗜血桿菌菌株159份。針對Hib莢膜基因合成型特異性引物及編碼轉(zhuǎn)運(yùn)莢膜多糖基因的引物,應(yīng)用PCR技術(shù)對臨床分離菌株進(jìn)行Hib檢測及亞型鑒定,對陽性標(biāo)本進(jìn)行PCR產(chǎn)物序列測定分析。 結(jié)果: 1.末次免疫后給予不同劑量莢膜多糖Hib結(jié)合疫苗的各實(shí)驗(yàn)組誘導(dǎo)產(chǎn)生PRP IgG抗體平均濃度均高于0.15μg/ml。免疫后1、2、4周各實(shí)驗(yàn)組內(nèi)PRP IgG抗體水平逐漸升高,其差異有顯著性統(tǒng)計(jì)學(xué)意義(P0.01)。 2.給予不同劑量莢膜多糖的Hib結(jié)合疫苗各實(shí)驗(yàn)組誘導(dǎo)大鼠產(chǎn)生抗PRP IgG抗體水平隨著莢膜多糖劑量的升高而升高。將免疫前及免疫后1、2、4周同一時(shí)間點(diǎn)的各實(shí)驗(yàn)組平均抗體水平進(jìn)行組間比較,免疫前各實(shí)驗(yàn)組組間無顯著性差異(p0.01)。免疫后1周0.8μg多糖組與2.5μg多糖組差異無統(tǒng)計(jì)學(xué)意義(P0.01),其余各實(shí)驗(yàn)組組間抗體水平差異有統(tǒng)計(jì)學(xué)意義(p0.01)。 3.159株流感嗜血桿菌中,肺炎患兒中分離Hi 90株,占56.6%;支氣管炎患兒39株,占24.5%。 4.應(yīng)用bex A基因特異性擴(kuò)增159株流感嗜血桿菌,4株陽性。其中69號臨床分離菌株經(jīng)Hi-b、HiHcsA-I特異性編碼基因擴(kuò)增陽性,PCR產(chǎn)物序列測定結(jié)果與GenBank數(shù)據(jù)庫內(nèi)基因序列進(jìn)行比對分析,一致性分別為98.5%、99%。 5.159株流感嗜血桿菌中有4株bex A基因擴(kuò)增陽性,為莢膜型流感嗜血桿菌,占流感嗜血桿菌2.52%,不可分型流感嗜血桿菌占97.48%;69號臨床菌株Hi-b及HiHcsA-I基因擴(kuò)增條帶陽性,經(jīng)基因序列比對分析鑒定為B型流感嗜血桿菌I型。 結(jié)論: 1.Hib結(jié)合疫苗中莢膜多糖能誘導(dǎo)大鼠產(chǎn)生較好的免疫應(yīng)答,其抗體水平隨時(shí)間及莢膜多糖含量的升高而升高,有良好的劑量效應(yīng)。 2. Hi是引起小兒呼吸道疾病的主要病原體之一,應(yīng)用PCR技術(shù)對臨床菌株進(jìn)行Hib檢測及分型具有較高靈敏度及特異性。針對HiHcsA基因的Hib分型可能反映臨床菌株的侵襲力。
[Abstract]:Objective: The aim of this study was to evaluate the immunogenicity of Hib specific capsule polysaccharides and their dose-dependent relationship with antibody levels in animals. To evaluate the value of PCR in the detection of Hib in children with respiratory diseases, to explore the potential and feasibility of genotyping corresponding to capsule dose in reflecting the invasiveness of strains, and to instruct children to use drugs and vaccinations in clinic. To provide a certain experimental basis for the development of new vaccines. Methods: 1. SD rats were immunized with different doses of Haemophilus influenzae type B (Haemophilus influenzae B) combined with PRP-T vaccine. The concentration of Hib polysaccharides antibody in serum was determined by using normal saline as control Elisa, and the data were processed by SAS 8.1 software. The immunogenicity and dose effect of Hib capsule polysaccharides were analyzed. 2. A total of 159 Haemophilus influenzae strains isolated from deep sputum samples of children with respiratory tract infection in children's hospitals in Chongqing were collected from 2009-2010. According to the specific primers of Hib capsule gene synthesis and the primers encoding the translocation of capsule polysaccharide gene, the Hib and subtype identification of clinical isolates were carried out by PCR technique. The positive samples were sequenced by PCR products. Results: 1. After the last immunization, the average concentration of PRP IgG antibody was higher than 0.15 渭 g / ml in each experimental group treated with different doses of capsule polysaccharide Hib conjugated vaccine. The level of PRP IgG antibody increased gradually in each experimental group at 4 weeks after immunization, and the difference was statistically significant (P 0.01). 2. The level of anti PRP IgG antibody induced by Hib conjugated vaccine with different doses of capsule polysaccharide increased with the increase of the dose of capsule polysaccharide. The average antibody levels of each experimental group at the same time point of 4 weeks before and after immunization were compared. There was no significant difference between the experimental groups before immunization and at the same time point (p0.01). At 1 week after immunization, there was no significant difference between 0.8 渭 g polysaccharide group and 2.5 渭 g polysaccharide group, but there was significant difference in antibody level among other experimental groups. Among the 3.159 Haemophilus influenzae strains, 90 strains (56.6%) were isolated from pneumonia and 39 strains (24.555%) from bronchitis. 4. Nine strains of Haemophilus influenzae were specifically amplified by bex A gene and 4 strains of Haemophilus influenzae were positive. The results of PCR products sequencing of 69 clinical isolates were compared with those in GenBank database by Hi-bSA-I specific coding gene amplification. The consistency was 98.5% and 99g, respectively. Of the 5.159 Haemophilus influenzae strains, 4 were positive for bex A gene amplification, accounting for Haemophilus influenzae, accounting for 2.52% of Haemophilus influenzae, 97.48% for Haemophilus influenzae, and 97.48% for Haemophilus influenzae. Haemophilus influenzae type B was identified by gene sequence alignment analysis. Conclusion: Capsule polysaccharides in 1.Hib conjugated vaccine could induce a good immune response in rats, and the antibody level increased with the increase of time and the content of capsule polysaccharides, which had a good dose effect. 2. Hi is one of the main pathogens causing respiratory tract diseases in children. PCR technique has high sensitivity and specificity in the detection and typing of clinical strains by Hib. The Hib typing of HiHcsA gene may reflect the invasiveness of clinical strains.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2011
【分類號】：R392.1

【參考文獻(xiàn)】

相關(guān)期刊論文 前10條

1 沈賽爾,倪贊明;應(yīng)用ELISA診斷b型流感嗜血桿菌性腦膜炎[J];上海醫(yī)科大學(xué)學(xué)報(bào);1993年02期

2 吳昕;黎明強(qiáng);甘志高;崔雪蓮;王曉偉;賈作民;馬晶巖;;國產(chǎn)b型流感嗜血桿菌結(jié)合疫苗免疫原性觀察[J];現(xiàn)代生物醫(yī)學(xué)進(jìn)展;2010年03期

3 王藝;王傳清;王曉紅;;細(xì)菌性腦膜炎266例病原學(xué)與耐藥性分析[J];實(shí)用兒科臨床雜志;2006年06期

4 劉梅影;王建華;陳蕓;許正莉;;DTaP-Hib聯(lián)合疫苗中Hib-TT免疫原性研究[J];微生物學(xué)免疫學(xué)進(jìn)展;2008年04期

5 李云;;老年人呼吸道流感嗜血桿菌的感染及耐藥分析[J];西北國防醫(yī)學(xué)雜志;2009年05期

6 彭焱;賀加;;重慶市渝中區(qū)兒童B型流感嗜血桿菌疫苗接種情況分析[J];現(xiàn)代預(yù)防醫(yī)學(xué);2010年03期

7 聶大平,李冬,劉敏,周力音;用套式PCR檢測b型流感嗜血桿菌[J];預(yù)防醫(yī)學(xué)文獻(xiàn)信息;1998年03期

8 王亞娟,楊永弘,沈敘莊,王亞亭,胡允文,姚德秀,佟月娟,Heikki Peltola,Didier Leboulleux;b型嗜血流行性感冒桿菌在急性下呼吸道感染病原學(xué)中的地位[J];中國計(jì)劃免疫;2004年03期

9 任紅宇;安淑一;李馬超;徐麗;朱兵清;邵祝軍;;細(xì)菌性腦膜炎多重聚合酶鏈反應(yīng)檢測方法的研究[J];中國疫苗和免疫;2010年01期

10 聶大平,李東,劉敏,周力音;聚合酶鏈反應(yīng)檢測細(xì)菌性腦膜炎患兒腦脊液中b型流感嗜血桿菌[J];中華傳染病雜志;2001年02期

，

本文編號：1937284