一株肠膜明串珠菌的分离鉴定及其抑菌特性

Isolation,identification,and antibacterial characterization of Leuconostoc mesenteroides

【背景】水产病原细菌严重威胁水产动物健康且制约水产养殖业发展,细菌性鱼病的有效防治成为水产养殖领域亟待解决的问题。【目的】筛选对水产病原细菌有抑制效果的菌株,并研究其抑菌特性及其在水产细菌病害防治中的实际效果。【方法】通过16S rRNA基因测序、构建系统发育树和生理生化鉴定确定筛选菌株的进化地位,通过乙酸乙酯萃取获得抑菌物质粗提物,通过偶氮酪蛋白法检测菌株胞外蛋白酶活力,采用结晶紫染色法对菌株的生物膜形成能力进行测定,通过浸浴攻毒模型确定所筛菌株对维氏气单胞菌的防治作用。【结果】从泡菜发酵物中筛选出一株乳酸菌DH,经16SrRNA基因测序、发育树分析和生理生化鉴定确定其为肠膜明串珠菌,该菌分泌的胞外抑菌物质对鼠伤寒沙门氏菌、大肠埃希氏菌、铜绿假单胞菌、杀鲑气单胞菌、希瓦氏菌和维氏气单胞菌表现出抑菌效果,其抑菌物质能被乙酸乙酯萃取并且具有热稳定性。菌株DH能够显著抑制待测菌株的蛋白酶产量和生物膜形成能力,并且对维氏气单胞菌浸浴攻毒有防治作用。【结论】肠膜明串珠菌DH通过分泌抑菌物质抑制水产病原细菌的生长,能够为细菌性鱼病的防治提供一定的理论和应用潜力。

[Background]Aquatic pathogenic bacteria threaten the health of aquatic organisms and restrict the development of aquaculture.The effective prevention and control of bacterial diseases in fish has become an urgent problem in aquaculture.[Objective]To screen out a bacterial strain with inhibitory effect on aquatic pathogenic bacteria,and to study its antibacterial properties and performance in controlling aquatic bacterial diseases.[Methods]The strain was identified after full consideration of the phylogenetic tree constructed based on the 16 S rRNA gene and the physiological and biochemical characteristics.The antibacterial substances were extracted with ethyl acetate.The effects of the strain on the extracellular protease production and biofilm-forming ability of pathogenic bacteria were determined by azo casein method and crystal violet staining,respectively.Furthermore,the inhibitory effect of the strain on Aeromonas veronii Z12 was evaluated by animal experiment.[Results]The lactic acid bacterial strain DH was screened out from the fermented product of pickle and identified as Leuconostoc mesenteroides.The strain secreted extracellular antimicrobial substances,and showed antibacterial effect on Salmonella typhimurium,Escherichia coli,Pseudomonas aeruginosa,Aeromonas salmonicida,Shewanella putrefaciens,and A.veronii.The antibacterial substances could be extracted with ethyl acetate and had thermal stability.Strain DH inhibited the extracellular protease production and biofilm-forming ability of the tested pathogenic bacteria,and exhibited protection against the challenge of Aeromonas veronii Z12.[Conclusion]L.mesenteroides DH can inhibit the growth of aquatic pathogenic bacteria by secreting antibacterial substances,which has the potential for the prevention and control of bacterial diseases in fish.