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宋增福, 范斌, 郭婧, 李娟英, 潘连德, 陈彪, 张庆华. 具有群体感应抑制效果芽孢杆菌的筛选与鉴定[J]. 水生生物学报, 2013, 37(6): 1059-1065. DOI: 10.7541/2013.142
引用本文: 宋增福, 范斌, 郭婧, 李娟英, 潘连德, 陈彪, 张庆华. 具有群体感应抑制效果芽孢杆菌的筛选与鉴定[J]. 水生生物学报, 2013, 37(6): 1059-1065. DOI: 10.7541/2013.142
Song zeng-fu, Fan bin, Guo jing, Li juan-ying, Pan Lian-de, CHEN Biao, Zhang qing-hua. SCREENING AND IDENTIFICATION OF BACILLUS TO INHIBIT QUORUM SENSING[J]. ACTA HYDROBIOLOGICA SINICA, 2013, 37(6): 1059-1065. DOI: 10.7541/2013.142
Citation: Song zeng-fu, Fan bin, Guo jing, Li juan-ying, Pan Lian-de, CHEN Biao, Zhang qing-hua. SCREENING AND IDENTIFICATION OF BACILLUS TO INHIBIT QUORUM SENSING[J]. ACTA HYDROBIOLOGICA SINICA, 2013, 37(6): 1059-1065. DOI: 10.7541/2013.142

具有群体感应抑制效果芽孢杆菌的筛选与鉴定

SCREENING AND IDENTIFICATION OF BACILLUS TO INHIBIT QUORUM SENSING

  • 摘要: 细菌性疾病是危害水产业健康发展的主要原因之一,从抑制调控致病菌毒力表达的群体感应(Quorum sensing,QS)入手是水产动物疾病防治研究新的热点方向。研究以紫色杆菌 (Chromobacterium violaceum)为报告菌株,采用T型划线和滤纸片法,对分离自异育银鲫肠道及实验室保存的芽孢杆菌F3-1、F3-2、F6-4、F6-5、X77、X93、X3914进行分析测测试,筛选具有群体感应抑制效果的菌株。T型划线结果显示菌株F3-1能够抑制紫色杆菌紫色菌素的产生;滤纸片法结果显示抑制紫色杆菌紫色菌素产生的主要物质存在于菌株F3-1的胞外产物中。试验提取了菌株F3-1的培养物的粗提物,测定粗提物浓度分别在0、50、100、200、300 mg/L时紫色杆菌菌素在585 nm处的光密度值,结果显示随着粗提物浓度的增加其色素的产量显著降低(P0.05),当粗提物浓度超过200 mg/L时,试管中观察不到紫色菌素存在;利用硫酸铵盐析的方法,提取了菌株F3-1的粗提物中蛋白产物,滤纸片扩散法结果显示表明其胞外蛋白是紫色杆菌QS系统的抑制物质。通过分子生物学16S rDNA鉴定,确定菌株F3-1为短小芽孢杆菌(Bacillus pumilus,GenBank accession No. JX984444)。电镜照片显示该菌具有短杆状特征,因此,分离自异育银鲫肠道的短小芽孢杆菌F3-1具有抑制细菌群体感应的作用。

     

    Abstract: Bacterial diseases are one of the main reasons to damage the health of aquaculture. It is a new research direction to prevent and control aquatic animal diseases by inhibiting the Quorum sensing (QS) that could regulate the pathogen virulence. The purpose of the present experiment was to screen the Bacillus strain to inhibit the QS, the Chromobacterium violaceum was used as the reporting strain to analyze the Bacillus strain F3-1, F3-2, F6-4, F6-5, X77, X93 and X3914 that the isolations from carassius auratus gibelio intestine tract or preserved in the lab by using T type streak cultivation and filter-paper-diffusion. The results of T type streak cultivation showed that strain F3-1 could inhibit the production of violacein of Chromobacterium violaceum; the results of filter-paper-diffusion indicated that the materials to inhibit the production of violacein of Chromobacterium violaceum was in the extracellular products of strain F3-1.Then, the crude extracts was taken from the culture of strain F3-1, and the OD585 of violacein of Chromobacterium violaceum was determined at the concentration of crude extracts of 0, 50, 100, 200 and 300 mg/L, and the results indicated that the production of violacien significantly decreased with the increasing of crude extracts concentration (P0.05); No violacien was observed in the test tube when the concentration of crude extracts reached 200mg/L. The protein of crude extracts was taken by the method of ammonium sulfate precipitation. The results showed that the extracellular proteins were probably the material to inhibit the QS of Chromobacterium violaceum. The strain F3-1 was identified as Bacillus pumilus (GenBank accession No. JX984444) by 16S rDNA sequence analysis. So, Bacillus pumiluswas isolated from carassius auratus gibelio intestine tract could inhibit the bacterial Quorum sensing.

     

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