瓜类细菌性果斑病菌对硫酸铜的敏感性检测与分析

果斑病菌两个亚群的143个菌株继代培养10代后,通过平板画线法对各菌株的硫酸铜敏感性检测结果表明：在硫酸铜浓度为300 μg/mL(1.88 mmol/L)时只有一株菌株表现敏感,在浓度达到750 μg/mL(4.69 mmol/L)时所有菌株均表现敏感;浓度为450~650 μg/mL (2.81~4.06 mmol/L)时分离自甜瓜的敏感菌株所占比率显著低于西瓜菌株;继代培养后所有菌株的硫酸铜敏感性没有改变,证实果斑病菌具有稳定的抗铜性。     

西瓜嗜酸菌趋化性的初步研究

瓜类细菌性果斑病是世界范围的检疫性细菌病害,病原菌为西瓜嗜酸菌,带菌种子为主要侵染源。病原细菌在寄主表面的定殖能力与其致病能力关系密切,而趋化性是决定定殖能力的关键因素之一,研究不同物质对西瓜嗜酸菌趋化性的影响对防治瓜类细菌性果斑病具有重要意义。本文采用毛细管法,研究了碳源、氨基酸、有机酸及其他物质对西瓜嗜酸菌趋化性的影响。结果表明,所测碳源中,麦芽糖、葡萄糖、乳糖、蔗糖和半乳糖均显著促进西瓜嗜酸菌的趋化性;所测氨基酸中,L-精氨酸、L-天冬氨酸、L-组氨酸、L-谷氨酸、L-亮氨酸、L-缬氨酸、L-谷氨酰胺和L-丙氨酸显著促进西瓜嗜酸菌的趋化性;所测有机酸中,琥珀酸、半乳糖醛酸和酒石酸显著促进西瓜嗜酸菌的趋化性;氯化钠、硫酸镁等对西瓜嗜酸菌的趋化性无显著影响。     

基于环介导等温扩增技术检测瓜类细菌性果斑病菌

本文利用环介导等温扩增技术(loop-mediated isothermal amplification,LAMP),针对西瓜嗜酸菌(Acidovorax citrulli)基因组中Aave_4063和Aave_4064序列设计了2对特异引物Ac-F3/Ac-B3和Ac-FIP/Ac-BIP,建立了西瓜嗜酸菌的LAMP检测体系。利用该体系在65℃保温1h,通过荧光显色即可完成检测。设计的引物特异性强,其检测灵敏度为2.0×101 cfu/mL。该方法为西瓜嗜酸菌的检疫及其所致病害的快速诊断提供了新的技术。     

Characterization of Acidovorax citrulli strains isolated from solanaceous plants

Acidovorax citrulli is the causal agent of bacterial fruit blotch disease of cucurbits. Strains of this pathogen are distributed into two major groups: Group I strains have been mainly isolated from melon and other non-watermelon cucurbits, while Group II strains have been mainly recovered from watermelon. Here we report the characterization of strains T1 and EP isolated from diseased tomato and eggplant plants, respectively, and further confirmed to belong to A. citrulli species. Based on PCR, PFGE, and rep-PCR, these strains showed high similarity to the Group II strain 7a1. Sequencing and comparative analyses revealed that the genomes of T1 and EP aligned with that of the Group II model strain AAC00-1, over 97.88% and 99.22%, respectively. The virulence of T1, EP, and 7a1 determined on tomato, eggplant, and watermelon was similar and significantly higher than that of Group I strain M6. In contrast, M6 was more virulent on melon. Expression levels of seven virulence genes measured 24 hr after inoculation of tomato, eggplant, watermelon, and melon showed that the expression pattern was generally similar in strains 7a1, T1, and EP, whereas for M6 the expression was high only on melon. Overall, our results indicate that the solanaceous strains belong to Group II. To the best of our knowledge, this is the first study that reports characterization of A. citrulli strains isolated from solanaceous species. The fact that A. citrulli is able to naturally colonize and cause disease in non-cucurbit crops poses additional challenges for management of this important pathogen.     

哈蜜瓜种子带细菌性果斑病菌检测技术的研究

哈蜜瓜细菌性果斑病是世界性的检疫对象之一,主要以种子带菌进行远距离传播,本文通过温室育苗、室内分离和PCR(聚合酶链式反应)技术对哈蜜瓜种子带菌情况和主要带菌部位进行了检验,结果表明:3种方法都可以对种子带菌情况进行检测,分离检测和PCR检测还可以确定种子的带菌部位,PCR技术具有特异性强、灵敏度高、检测时间短等特点.另外检测结果也表明种子带菌以种皮为主,种仁的带菌量较少.     

西瓜细菌性果斑病菌快速免疫PCR检测

对种子携带的西瓜细菌性果斑病菌(Acidovorax avenae subsp. citrulli)进行简单抽提和纯化,不经过DNA提取而以病原菌为模板进行PCR检测.结果显示,对带菌种子提取液采用直接PCR法最低检出限为3600个细菌/mL,而免疫PCR最低检出限可达到600个细菌/mL.免疫PCR法可以有效富集病原后再扩增,方便快速,成本低,灵敏度高,适用于种子携带微量的西瓜细菌性果斑病菌的快速鉴定.     

瓜类种子细菌性果斑病菌防治研究进展

细菌性果斑病是典型的由种子带菌引起的传染性病害,解决种子带菌问题是防治该病的关键。通过综述近年国内外瓜类种子的发酵处理、热处理、药剂处理以及快速干燥对BFB的防治,对种子处理剂提出新的研究方向,为从事相关研究的工作人员提供参考和借鉴。     

哈密瓜细菌性果斑病菌群体感应系统的检测

本文利用群体感应信号报告菌Agrobacterium tumefaciens NTL4(pZLR4),在LB报告平板上对燕麦食酸菌西瓜亚种的19个菌株进行了初步检测,发现18个菌株有群体感应信号产生.用琼脂条法对菌株Pslb-94进一步检测,证实菌株Pslb-94存在群体感应系统.提取了该菌株信号物质,反相薄层层析(TLC)检测证明该菌株能产生群体感应信号分子.     

瓜类果斑病菌致病基因及靶标基因的预测

哈密瓜果斑病是严重危害瓜类的种传细菌性病害,其病原菌为嗜酸菌属西瓜种(Acidovorax citrulli)。本文首先用Bioedit软件建立瓜类果斑病菌全氨基酸序列本地资源库,运用比较基因组学的方法,通过大量查找嗜酸菌属、欧文氏菌属、假单胞菌属、黄单胞菌属、劳尔氏菌属、土壤杆菌属、木质部小菌属中的致病基因和参考文献中已报道的致病基因,下载相关致病基因的氨基酸序列,与瓜类果斑病菌的全氨基酸序列进行localblast,取E<10-5的为候选基因,得到果斑病菌中与致病性相关的基因,并对未知蛋白进行产物预测。同时,根据已经公布的靶标基因,用同样的比对方法,得到果斑病菌中潜在的靶标基因。本研究通过保守估计,得到77个致病性相关蛋白,预测了其中7个未知蛋白的产物,得到46个靶标蛋白。致病基因和靶标基因的分析预测,对以后研究此类基因提供了指导作用,为更好地防治该病害提供了理论基础。     

利用GICA-PCR快速检测瓜类果斑病菌

瓜类果斑病菌(Acidovorax avenae subsp.citrulli,Aac)是瓜类作物上重要的病原细菌,为我国进境植物检疫性有害生物。胶体金免疫层析试纸条方便快捷,应用广泛。该方法使用不当会出现假阳性问题,仅适用于病原菌的初筛检测。本研究将Aac胶体金免疫层析方法(GICA)与PCR技术相结合,建立了GICA-PCR检测方法。检测结果表明,该方法在蛋白与核酸2个层面上从发病西瓜叶片上检测到瓜类果斑病菌,有效解决了试纸条检测的假阳性问题,提高了瓜类果斑病菌检测的准确性,值得推广应用。     

西瓜细菌性果斑病研究进展

西瓜细菌性果斑病是由类产碱假单胞菌西瓜亚种〔Pseudomonas pseudoalcaligenes subsp. citrulli (Schaad et al. 1978)〕所引起，1992年该病菌改名为燕麦食酸菌西瓜亚种〔Acidovorax avenae subsp. citrulli (Willems et al. 1992) 〕。革兰氏阴性菌，属rRNA组I。不产生荧光和其他色素，单根极生鞭毛，严格好氧。不产生精氨酸水解酶，能在41℃下生长，但不能在4℃生长，明胶液化力弱，氧化酶和2－酮葡糖酸试验阳性。利用葡萄糖和蔗糖作碳源结果不一致；但利用β－丙氨酸、柠檬酸盐、乙醇、乙醇胺、果糖、L－亮氨酸和D－丝氨酸结果一致…     

High-throughput screening reveals small molecule modulators inhibitory to Acidovorax citrulli

Acidovorax citrulli is a seed-transmitted gram-negative bacterium that can cause substantial economic yield loss in watermelon and melon production worldwide. Four small-molecule libraries containing 4,952 compounds were selected for high-throughput screening against A. citrulli wild-type strain Xu3-14 by evaluation of growth inhibition. One hundred and twenty-seven molecules (2.5% hit rate) were identified as bactericidal or bacteriostatic against A. citrulli at 100 μM. Secondary screens indicated that 27 candidate compounds were more effective against A. citrulli Group II strains than Group I strains (classified using repetitive element PCR). Several compounds were inhibitory to other pathogenic bacteria, including Clavibacter michiganensis subsp. michiganensis and Xanthomonas campestris pv. campestris, but did not affect the growth of plant beneficial bacteria Pseudomonas fluorescens and Bacillus subtilis. More than half of the compounds did not inhibit germination of Arabidopsis or watermelon seeds. The effect of small molecules on A. citrulli seed-to-seedling transmission was evaluated by applying each compound to inoculated watermelon seeds and assessing seedling infection. Nine compounds were chosen for further investigation based on their reduction of percentage seedling infection and compiling scores on their specificity, sensitivity, and phytotoxicity obtained in the secondary screens. The five best compounds were selected (thiamphenicol, nadifloxacin, pipemidic acid, ciclopirox, and zinc pyrithione) for greenhouse tests and were found to effectively reduce the seed-to-seedling transmission of A. citrulli in both artificially and naturally infested seeds. These top five compounds provide a basis for future development of an A. citrulli-specific bactericide.     

进境西瓜种子中西瓜细菌性果斑病菌的检测鉴定

由嗜酸菌属西瓜种(Acidovorax citrulli)引起的西瓜细菌性果斑病是西瓜等葫芦科作物上的一种极具毁灭性的病害,该病原细菌可以由种子携带传播。本研究通过直接研磨种子,用双抗体夹心酶联免疫吸附测定法(DAS-ELISA)和免疫捕捉聚合酶链式反应法(IC-PCR)对进境的西瓜种子进行检测,并且对PCR产物进一步克隆测序。结果表明,在DAS-ELISA产生阳性结果的样品中,IC-PCR结果也产生阳性。序列分析表明,该序列与已知的该病菌16S rRNA基因的相应序列具有100%同源性。因此,2006年这批来自台湾的西瓜种子携带有嗜酸菌属西瓜种(Acidovorax citrulli)。     

瓜类果斑病菌胶体金试纸条的检测应用

瓜类果斑病是一种严重为害西甜瓜作物的细菌病害,胶体金试纸条是对其进行现场检测的主要手段.将中国检验检疫科学研究院研制的瓜类果斑病菌胶体金检测试纸条与市场上商品化的美国Agdia公司同类试纸条进行比较,发现两种试纸条的灵敏度和特异性相当,适用于检测出现疑似果斑病症状的叶片和果实;也可用于种子带菌检测,但易出现假阳性结果....     

我国瓜类细菌性果斑病发生现状和综合防控策略

瓜类细菌性果斑病是西甜瓜等葫芦科作物上的一种检疫性病害,本文从病害的发生历史、传播及流行特点等方面综述了瓜类细菌性果斑病在我国的发生现状,提出了政府部门、种子种苗生产单位和种植户应共同采取措施对病害进行综合防控,以防止病害进一步传播扩散的策略。     

Pathways of bacterial invasion and watermelon seed infection by Acidovorax citrulli

This study explored the pathways of ingress of Acidovorax citrulli, the causal agent of bacterial fruit blotch of cucurbits, into watermelon seeds. Up until 7 days post‐inoculation (DPI), a significantly higher percentage of watermelon seeds was infected with A. citrulli when the bacteria were applied (c. 1 × 10⁶ colony‐forming units) to stigmas versus ovary pericarps of female flowers. Immunofluorescence microscopy showed that, with stigma inoculation, A. citrulli colonized style and ovary tissues by 1 DPI, and the bacteria co‐localized with pollen germ tubes in these tissues. With ovary pericarp inoculation, A. citrulli cells penetrated the epicarp and mesocarp tissues by 1 DPI but did not reach endocarp until 4 DPI. Finally, manual pollination followed by stigma inoculation led to >53% A. citrulli‐infected seed lots, while A. citrulli was not detected in seeds/ovules generated by stigma inoculation without pollination (chemically induced parthenocarpy). These results show that stigma inoculation results in faster colonization of watermelon ovules by A. citrulli than pericarp inoculation, even though there is no difference in the levels of infection in mature seeds. The data also indicate that pollen germ tubes play an important role in A. citrulli ingress into watermelon seeds via stigmas.     

瓜类细菌性果斑病研究进展

瓜类细菌性果斑病是发生在甜瓜、西瓜等葫芦科植物上的一种严重的世界性病害,此病是典型的种传细菌性病害,病原为嗜酸菌属西瓜种(Acidovorax citrulli)。本文围绕瓜类细菌性果斑病菌的分离检测、致病机理、遗传多样性及防治等方面的研究进展作一概述,阐明了瓜类细菌性果斑病的研究现状。     

应用蛋白宏阵列快速检测西瓜细菌性果斑病菌

蛋白阵列(Protein array)也叫蛋白芯片,是生物芯片(Biochip)的一种,主要利用抗原/抗体可特异性结合原理制备而成。按阵列样点大小,蛋白阵列又可分为微阵列(Microarray)和宏阵列(Mac-roarray)两类。微阵列集成度极高,可进行高通量