【作者】 陈涛；

【导师】 胡白石；

【作者基本信息】 南京农业大学 ， 植物病理学， 2008， 硕士

【摘要】 哈密瓜细菌性果斑病菌（Acidovorax avenae subsp.citrulli）是西瓜、甜瓜、哈密瓜等葫芦科植物上的一种重要的病原细菌。近年来果斑病的发生趋势有所上升,对我国哈密瓜及其他瓜类生长造成严重威胁,常造成严重的经济危害。目前,对果斑病菌的研究主要集中于传播机理,防治措施等方面,对于其致病机理的研究少有报道。本研究利用AHL超敏感生物检测菌株JZA1（pJZ372）（pJZ384）（pJZ410）对哈密瓜细菌性果斑病菌（A.avenae subsp.citrulli）产生群体感应信号分子的能力进行了检测,定量的β-半乳糖苷酶活性检测与定性的反向薄层层析检测表明该菌株可以产生一定活性的、1种类型（C₈-AHL）的群体感应信号分子,说明哈密瓜果斑病菌中存在QS系统。同时,在我们检测的57株哈密瓜果斑病菌菌株中,39株菌株能产生群体感应信号分子,18株未检测到AHL类信号分子,说明QS系统在哈密瓜果斑病菌中存在差异。利用分子生物信息学,通过同源性比对,在哈密瓜果斑病菌的全基因中发现了信号分子的合成基因acluxⅠ。通过PCR技术,从AHL产生菌株xjL12中扩增了acluxⅠ基因。将克隆的acluxⅠ基因在autoinducer-Ⅰ（AI-1）AI-1缺陷的菌株Escherichia coliDH5α中原核表达,发现DH5α可恢复产生AI-1的能力。利用同源重组技术,构建了acluxⅠ基因的缺失突变株,信号分子检测和致病性测定试验表明,acluxⅠ缺失突变株完全丧失了合成AI-Ⅰ的能力,同时其在西瓜果实和幼苗上的致病性也显著降低,但不影响其生长能力。我们研究表明,群体感应调节系统在哈密瓜果斑病菌的致病性中扮演着十分重要的角色。更多还原

【Abstract】 Bacterial fruit blotch（BFB）,caused by Acidovorax avenae subsp,citrulli,is one of the most devastating and difficult-to-control diseases.Recently,this disease is becoming more and more serious in China,and has caused great damage on growth of Hamirmelons and economy.Till now,many researches focused on improving the BFB management, including chemical seed treatment and biological control.Little information on the virulence determinants of this pathogen was availableIn this study,we detected the autoinduer（AI） activity in the supernant of A.avenae subsp,citrulli by a ultrasensitive AHL bioassay strain JZA1（pJZ372）（pJZ384）（pJZ410）. The results of quantitative LacZ activity and qualitative TLC detection showed that A. avenae subsp,citrulli xjL12 could produce high active and one type AHLs（C₈-AHL）.In addition,other 56 strains were also selected for AHL bioasaay.Among them,39 strains had the ability to produce AI-1（AHLs）,and 18 strains were showed no AHL-activity.Together, these results indicated that quorum sensing system was existed in A.avenase subsp,citrulli, but different in the different strains.A search of the genome of A.avenae subsp,citrulli revealed that a acluxI homologue to that of Pesudomonas syringe pv.tomato DC3000 was found.By PCR method,the acluxI homologue was amplified from AHL-producing strain xjL12,and was expressed in Escherichia coli DH5α.Interestingly,the AI-1-deficient E.coli DH5αhad the ability to produce AHLs by expression of luxI homologue from strain xjL12.Furthermore,the acluxI homologue was knocked out by recombinant homologue technique The results of AHL detection and pathogenicity test showed that the acluxI mutant had no AHL activity and showed significant reduction of virulence on fruit and seedlings of Hamimelon.These results revealed that Quorum sensing system might play a key role in the virulence of A. avenae subsp,citrulli.更多还原