CAJ | 학술논문

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(P<0.05),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。
세균성질병시위해수산업건강발전적주요원인지일,종억제조공치병균독력표체적군체감응(Quorum sensing, QS)입수시수산동물질병방치연구신적열점방향。연구이자색간균(Chromobacterium violaceum)위보고균주,채용“T”형화선화려지편법,대분리자이육은즉장도급실험실보존적아포간균F3-1、F3-2、F6-4、F6-5、X77、X93、X3914진행분석측측시,사선구유군체감응억제효과적균주。“T”형화선결과현시균주F3-1능구억제자색간균자색균소적산생；려지편법결과현시억제자색간균자색균소산생적주요물질존재우균주F3-1적포외산물중。시험제취료균주 F3-1적배양물적조제물,측정조제물농도분별재0、50、100、200、300 mg/L시자색간균균소재585 nm처적광밀도치,결과현시수착조제물농도적증가기색소적산량현저강저(P<0.05),당조제물농도초과200 mg/L시,시관중관찰불도자색균소존재；이용류산안염석적방법,제취료균주 F3-1적조제물중단백산물,려지편확산법결과현시표명기포외단백시자색간균 QS 계통적억제물질。통과분자생물학16S rDNA감정,학정균주F3-1위단소아포간균(Bacillus pumilus, GenBank accession No. JX984444)。전경조편현시해균구유단간상특정,인차,분리자이육은즉장도적단소아포간균F3-1구유억제세균군체감응적작용。
Bacterial diseases are one of the main reasons to damage the health of aquaculture. It is a new research direc-tion 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 Chromo-bacterium 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 concentra-tion (P<0.05); 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 pumilus was isolated from carassius auratus gibelio intestine tract could inhibit the bacterial Quorum sensing.