本文關(guān)鍵詞： 布魯氏菌 基因分型 DFR MLST MLVA　出處：《吉林大學(xué)》2012年博士論文　論文類型：學(xué)位論文

【摘要】：布魯氏菌病是由布魯氏菌侵入機(jī)體引起的一類傳染－變態(tài)反應(yīng)性的全球性分布的人畜共患病，在許多發(fā)展中國家流行并造成很大的經(jīng)濟(jì)損失。依據(jù)對(duì)宿主偏好性以及培養(yǎng)和生物學(xué)特性的不同，布魯氏菌分為6個(gè)經(jīng)典的種，其中羊種、牛種、豬種、犬種4個(gè)種能夠使人致病。布魯氏菌基因的高度保守性使得建立種型之間的遺傳關(guān)系極具挑戰(zhàn)性。了解布魯氏菌基因水平的差異，有助于了解布魯氏菌的進(jìn)化與毒力特征。目前布魯氏菌基因分型方法中以多位點(diǎn)序列分型(MLST)和多位點(diǎn)VNTR (MLVA)法重復(fù)性好、分辨率高，但是由于布魯氏菌的變異，在實(shí)際工作中已遇到問題。尋找一種更好的分型方法或者是多種方法的聯(lián)合運(yùn)用，可能會(huì)更好地解決布魯氏菌的分種分型問題。 通過十株已測(cè)序的布魯氏菌全基因組的比較分析和布魯氏菌全基因組芯片雜交的結(jié)果，共鑒定出53個(gè)差異基因區(qū)段（DFR），其中位于I號(hào)染色體上的DFR有26個(gè)，，II號(hào)染色體上的DFR為27個(gè)，II號(hào)染色體的DFR密度遠(yuǎn)遠(yuǎn)高于I號(hào)染色體。探討這些差異區(qū)段在19株標(biāo)準(zhǔn)菌株和129株我國不同種型布魯氏菌分離株的分布特征表明，差異區(qū)段可能是進(jìn)化過程中獲得或缺失的，與菌株的遺傳關(guān)系沒有必然的聯(lián)系，為研究不同疫源地或不同年代的布魯氏菌菌株基因水平差異提供了依據(jù)。 為了提高傳統(tǒng)的MLST方法（CMLST）的分型分辨率，我們?cè)诒狙芯恐刑接懲ㄟ^加長基因的PCR擴(kuò)增片段來用于MLST分型研究，即為改良的MLST方法（EMLST）。對(duì)129株我國布魯氏菌分離株進(jìn)行了CMST和EMLST的分析，用CMLST方法定義了23個(gè)ST型，而用EMLST方法則定義了35個(gè)ST型，多種方法分析表明，EMLST較CMLST具有更高的分型分辨率。和已報(bào)道的國外布魯氏菌菌株的CMLST結(jié)果比較分析發(fā)現(xiàn)，我國布魯氏菌分離株具有自己的基因型構(gòu)成特點(diǎn)�；诳醇一虻母牧嫉腗LST方法適合于親緣關(guān)系較遠(yuǎn)的布魯氏菌菌株的分型。 傳統(tǒng)的MLVA方法采用PCR擴(kuò)增并電泳區(qū)分的方法，但是存在不穩(wěn)定及費(fèi)時(shí)費(fèi)力的缺點(diǎn)，因此本研究采用測(cè)序的方法對(duì)120株分離株的15個(gè)多態(tài)性較高的MLVA位點(diǎn)進(jìn)行分析，共分為87個(gè)型�；赩NTR位點(diǎn)的改良的MLVA方法應(yīng)用于我國布魯氏菌分離株的結(jié)果表明，該方法適合于親緣關(guān)系較近的布魯氏菌菌株的分型。對(duì)120株CMLST、EMLST以及MLVA數(shù)據(jù)綜合比較分析表明，EMLST較CMLST具有更多的分支，有著更高的分辨率，而MLVA則是三種分型方法中分辨率最高的方法。
[Abstract]:Brucellosis is a class of infectious-allergic global zoonosis caused by brucellosis. Brucellosis is prevalent in many developing countries and causes great economic losses. According to the difference of host preference, culture and biological characteristics, Brucella is divided into six classical species, sheep, cattle and pig. The highly conserved nature of Brucella genes makes it challenging to establish genetic relationships among species. Understand the differences in Brucella gene levels. It is helpful to understand the evolutional and virulence characteristics of Brucella. At present, the methods of multilocus sequence typing (MLSTs) and multilocus VNTR (MLVA) methods have good reproducibility. High resolution, but because of the variation of brucella, has encountered problems in practical work. To find a better method of typing or combination of multiple methods. It may be better to solve the classification problem of Brucella. Based on the comparative analysis of the whole genome of ten strains of Brucella and the results of whole genome chip hybridization of Brucella, a total of 53 differentially expressed gene regions (DFR) were identified. Among them, there are 26 DFR on chromosome I and 27 DFR on chromosome II. The DFR density of chromosome II was much higher than that of chromosome I. the distribution characteristics of these differential regions in 19 standard strains and 129 strains of Brucella strains of different species in China were studied. The differential region may be acquired or missing in the course of evolution, which is not necessarily related to the genetic relationship of the strains, which provides a basis for the study of gene level differences of Brucella strains in different foci or in different years. In order to improve the typing resolution of the traditional MLST method, we studied the use of the extended PCR fragment in the study of MLST typing. The CMST and EMLST of 129 strains of Brucella in China were analyzed by CMST and EMLST for the improved MLST method. 23 St types were defined by CMLST method. In contrast, 35 St types were defined by EMLST method. EMLST has higher typing resolution than CMLST. The results of CMLST analysis of Brucella strains have been compared and analyzed. Brucella isolates in China have their own genotypic characteristics. The improved MLST method based on housekeeping gene is suitable for the genotyping of Brucella strains with distant genetic relationship. The traditional MLVA method is based on PCR amplification and electrophoresis, but it is unstable and time-consuming. Therefore, 15 polymorphic MLVA loci of 120 isolates were analyzed by sequencing. The modified MLVA method based on VNTR locus was applied to Brucella isolates in China. This method is suitable for the genotyping of brucella strains with close genetic relationship. The comprehensive analysis of 120 strains of CML-ST-EMLST and MLVA data shows that this method is suitable for the typing of Brucella strains. EMLST has more branches and higher resolution than CMLST, and MLVA is the highest resolution of the three typing methods.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2012
【分類號(hào)】：R378

【參考文獻(xiàn)】

相關(guān)期刊論文 前7條

1 吳從雅;布魯氏菌屬細(xì)菌分種分型簡史和研究進(jìn)展(綜述)[J];地方病通報(bào);1990年01期

2 尚德秋;布氏菌病研究進(jìn)展[J];中國地方病防治雜志;2004年04期

3 崔步云,尹繼明,李蘭玉,樸冬日,趙鴻雁,尚德秋;布魯氏菌的Rep-PCR分型研究[J];疾病監(jiān)測(cè);2005年08期

4 姜海;崔步云;李蘭玉;趙鴻雁;樸東日;;用MLVA技術(shù)和多重PCR對(duì)犬種布氏菌基因分型[J];生物技術(shù)通訊;2009年03期

5 張秀梅,郭秀蘭,馬志東;用紫外可見分光光度計(jì)分析法測(cè)定布魯氏菌DNA[J];中國人獸共患病雜志;1993年06期

6 李元?jiǎng)P，邱海燕，馬燁，尚德秋;布魯氏菌非典型菌株及R型菌株生物學(xué)特性和產(chǎn)生原因的探討[J];中國人獸共患病雜志;1996年05期

7 齊景文;布魯氏菌病概述[J];中國獸醫(yī)雜志;2004年09期

本文編號(hào)：1489251