本文選題：幽門螺桿菌 + 多位點(diǎn)序列分型　； 參考：《天津醫(yī)科大學(xué)》2011年碩士論文

【摘要】：目的：研究幽門螺桿菌(H. pylori)中國分離株的基因分型特征。 方法：對(duì)分離自中國7個(gè)不同地區(qū)(北京、黑龍江、煙臺(tái)、浙江、湖北、西安、云南)、不同胃、十二指腸相關(guān)疾病(慢性胃炎、消化性潰瘍、胃癌、功能性消化不良)患者的119株H.pylori的7個(gè)看家基因(atpA, efp, mutY,ppa, trpC, urel和yphC)和毒力相關(guān)基因vacA,應(yīng)用多位點(diǎn)序列分型(multilocus sequence typing, MLST)的方法獲得菌株的等位基因譜(allelic profile)和序列型(sequnce type,ST),與HP MLST數(shù)據(jù)庫中世界范圍內(nèi)的菌株信息進(jìn)行比較。應(yīng)用軟件BURST (Based Upon Related Sequence types)運(yùn)算法則將菌株歸為譜系(clonal complex),應(yīng)用軟件START2 (Sequence Type Analysis and Recombinational Tests, version 2)計(jì)算相關(guān)系數(shù)(Index of Association, IA),應(yīng)用軟件MEGA4.0(Molecular Evolutionary Genetics Analysis, version 4.0)構(gòu)建進(jìn)化樹進(jìn)行分子進(jìn)化遺傳分析。對(duì)vacA等位基因(sla, slb, s2, mla, mlb, m2, il和i2)進(jìn)行特異引物聚合酶鏈?zhǔn)椒磻?yīng)(polymerase chain reaction, PCR),根據(jù)擴(kuò)增產(chǎn)物片段大小確定vacA基因型,對(duì)基因型分布應(yīng)用SPSS13.0軟件數(shù)據(jù)包進(jìn)行統(tǒng)計(jì)學(xué)分析、并應(yīng)用層級(jí)聚類分析軟件Mev_4_0 (Multiple Experiment Viewer, TIGR)進(jìn)行聚類分析。 結(jié)果：MLST共確定了107個(gè)序列型,都是HP MLST數(shù)據(jù)庫中未曾報(bào)道過的。119株H.pylori菌株被分為3個(gè)譜系,其他均為獨(dú)特型。START2重組分析顯示,107個(gè)STs的IA為2.1398。MEGA4.0軟件將119株中國菌株分布在3個(gè)祖先群。111株分布在hpEastAsia,6株分布在hpEurope,2株分布在hpAsia2。119株H.pylori vacA基因以sla,m2和i1型為主,分別為97.5%(116/119),68.9%(82/119)和91.6%(109/119)。26.1%(31／119)為mlb。slb和mla未檢出。vacA組合基因型以sla/m2/il為主(62.2%,74/119),sla/mlb/il次之(25.2%,30/119)。不同地域、不同疾病來源菌株sla分布差異無統(tǒng)計(jì)學(xué)意義(P0.05)。而m區(qū)基因多態(tài)性在疾病類型及分離地區(qū)間差異均有統(tǒng)計(jì)學(xué)意義(P0.01)。不同疾病來源菌株間i區(qū)基因分型差異無統(tǒng)計(jì)學(xué)意義(P0.05),但不同分離地區(qū)菌株間差異有統(tǒng)計(jì)學(xué)意義(P0.01)。119株H.pylori vacA基因序列聚類為三個(gè)不同組群。一株獨(dú)在一個(gè)組群,8株在另一個(gè)組群,其余的菌株聚為一群。 結(jié)論：本研究中H.pylori中國分離株絕大多數(shù)起源于hpEastAsia,很少部分屬于hpAsia2(?)和hpEurope,尚未發(fā)現(xiàn)有起源于非洲的菌株。H.pylori中國分離株vacA基因型以sla m2 il組合型為主,sla分型結(jié)果與菌株分離地域及患者疾病類型無關(guān)。不同地域、不同疾病來源菌株m區(qū)基因分型不同。不同地域菌株vacA基因i區(qū)分型不同,但i區(qū)基因分型與菌株的疾病來源沒有顯著相關(guān)性。中國H.pylori菌株有一定的地域相關(guān)性,但疾病相關(guān)性不明顯。
[Abstract]:Objective: to study the genotyping characteristics of Helicobacter pylori H. pylori isolates from China. Methods: isolated from 7 different regions of China (Beijing, Heilongjiang, Yantai, Zhejiang, Hubei, Xi'an, Yunnan, different stomach and duodenal diseases (chronic gastritis, peptic ulcer, gastric cancer). Seven housekeeping genes of 119 H.pylori strains from patients with functional dyspepsia (H.pylori) were identified by multilocus sequence typing (MLSTs) and virulence related genes (Vaca) and virulence related genes (Vaca). The allelic and serotype sequnce type STs were obtained by multilocus sequence typing (MLSTs). The results were compared with the worldwide strain information in HP MLST database. The phylogenetic analysis was carried out by using the algorithm of BURST based Upon Related Sequence types, and START2 sequence Type Analysis and Recombinational tests, version 2, MEGA4.0(Molecular Evolutionary Genetics Analysis, version 4.0) was used to calculate the correlation coefficient of association, IAA, MEGA4.0(Molecular Evolutionary Genetics Analysis, version 4.0) to construct the phylogenetic analysis. The phylogenetic analysis was carried out by using the software START2 sequence Type Analysis and Recombinational tests (version 2) to calculate the correlation index of association, IAA, MEGA4.0(Molecular Evolutionary Genetics analysis, version 4.0). Specific primer polymerase chain reaction (PCR) was performed on the alleles of vacA, such as SLAA, SLB, S2, mla, MLB, m2il and i2). The genotypes of vacA were determined according to the fragment size of the amplified products, and the distribution of vacA genotypes was analyzed statistically by SPSS13.0 software. The hierarchical cluster analysis software Mev_4_0 multiple Experiment Viewer (TIGR) was applied to cluster analysis. Results A total of 107 sequence types were identified, all of which were not reported in HP MLST database. 119 H.pylori strains were divided into 3 lineages. The recombination analysis of Idiotype. START2 showed that 107 STs IA software distributed 119 Chinese strains in 3 progenitor groups. 111 strains distributed in hpEastAsiaS6 strains distributed in hpAsia2.119 strain H.pylori vacA gene mainly distributed in hpAsia2.119 strain SLAM 2 and I 1 type. 97.5% 11.6 / 119 / 68.9 / 82 / 119 and 91.6 / 109 / 119 / 26.1 / 31 / 119 respectively. For mlb.slb and mla, the main genotype of Vaca was 62.2% / 119slar / 119slar / ml / ml / 119g, and 25.2nb / 119b / r 25.2nb / 119a, respectively, for mlb.slb and mla with no Vaca genotype 62.2% / 119slar / ml / r / il. There was no significant difference in the distribution of sla between different regions and different disease sources (P 0.05). But the gene polymorphism in m region was significantly different in disease type and isolation region (P 0.01). There was no significant difference in I region genotyping among strains from different disease sources, but there were significant differences in H.pylori vacA gene sequences among different isolates from different regions. There were three different groups of H.pylori vacA gene sequences. One strain was isolated in one group, 8 strains in another group, and the other strains were clustered into a group. Conclusion: most of the Chinese isolates of H.pylori in this study originated from hpEastAsia, and few of them belong to hpAsia2. No vacA genotypes of H.pylori strains originated in Africa were found in China, and sla m2il combinations were the dominant genotypes. The results were not related to the region of isolates and the disease types of patients. The genotyping of m region was different in different regions and different disease sources. The genotype of vacA gene I was different in different regions, but there was no significant correlation between the genotype of I region and the source of disease. H.pylori strains in China have a certain geographical correlation, but the disease correlation is not obvious.
【學(xué)位授予單位】：天津醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2011
【分類號(hào)】：R378

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本文編號(hào)：1788514