东方蝼蛄肠道菌分离鉴定及产纤维素酶菌株的筛选与酶活力测定  

Screening and enzyme activity determination of cellulase-producing strains from the gut bacteria of Gryllotalpa orientalis

作　　者：陈广青 陈芳敏 王琪 付晶晶 夏琬婷 王金星 CHEN Guangqing;CHEN Fangmin;WANG Qi;FU Jingjing;XIA Wanting;WANG Jinxing(Liaoning Key Laboratory of Urban Integrated Pest Management and Ecological Security,College of Life Science and Bioengineering,Shenyang University,Shenyang 110044,Liaoning,China;Key Laboratory of Eco-restoration of Regional Contaminated Environment of Ministry of Education,Shenyang University,Shenyang 110044,Liaoning,China)

机构地区：[1]沈阳大学生命科学与工程学院,辽宁省城市有害生物治理与生态安全重点实验室,辽宁沈阳110044 [2]沈阳大学区域污染环境生态修复教育部重点实验室,辽宁沈阳110044

出　　处：《微生物学通报》2024年第10期4257-4269,共13页Microbiology China

基　　金：国家自然科学基金(42177406);辽宁省自然科学基金(2023-MS-319)。

摘　　要：【背景】蝼蛄(Gryllotalpa spp.)是我国分布广泛的农业地下害虫,其肠道菌的组成及功能菌资源的挖掘都有待研究。【目的】获得蝼蛄肠道可培养菌株,对其进行鉴定和产纤维素酶功能菌株的筛选,并检测其酶活性,对活性较高菌株的产酶条件进行探究。【方法】采用传统的微生物分离培养方法,结合分子生物学技术对东方蝼蛄肠道菌进行分离和鉴定,采用羧甲基纤维素钠(carboxymethylcellulose sodium,CMC-Na)水解圈法筛选产纤维素酶菌株,并用3,5-二硝基水杨酸(3,5-dinitrosalicylic acid,DNS)法测定纤维素酶活力。【结果】从东方蝼蛄肠道共分离出23个菌株,可归为7属10种:其中芽孢杆菌属(Bacillus)4种,肠球菌属(Enterococcus)、葡萄球菌属(Staphylococcus)、不动杆菌属(Acinetobacter)、变形杆菌属(Proteus)、肠杆菌属(Enterobacter)和乳球菌属(Lactococcus)各1种。结果显示,除菌株N18外的其余22个菌株均具有产纤维素酶能力,其中菌株N3和N14产酶能力最强,其次是N15和N21。培养基的pH、培养时间、菌液接种量对菌株的产酶能力影响各异。在实验设定的梯度范围内,所测各单因素影响下,均为菌株N3和N14产酶能力最强。【结论】蝼蛄肠道可培养菌资源丰富,获得菌株中高达95%具有产纤维素酶的功能,其中枯草芽孢杆菌(Bacillus subtilis)N3和N14产酶能力最强,最具开发潜力。[Background]Mole crickets(Gryllotalpa spp.)are widely distributed agricultural underground pests in China,and the composition of their gut bacteria and the functional bacterial resources remain to be studied.[Objective]To obtain the culturable bacteria from the gut of G.orientalis,screen out the strains capable of producing cellulase,measure the cellulase activities of the strains,and explore the culture conditions of the strains with high cellulase activities.[Methods]The gut bacteria of G.orientalis were isolated by the culture method and identified by methods of molecular biology.The cellulose-producing strains were screened by the carboxymethylcellulose sodium(CMC-Na)hydrolysis assay,and the cellulase activity was determined by the 3,5-dinitrosalicylic acid(DNS)method.[Results]A total of 23 strains were isolated from the gut of G.orientalis,belonging to 10 species of 7 genera(4 species of Bacillus and 1 species of Enterococcus,Staphylococcus,Acinetobacter,Proteus,Enterobacter,and Lactococcus,respectively).The results showed that all the 22 strains except strain N18 had cellulase-producing capacity,among which strains N3 and N14 had the strongest cellulase-producing capacity,followed by N15 and N21.The medium pH,culture time,and inoculum amount had different effects on the cellulase production.Within the ranges set in this study,strains N3 and N14 always had the highest cellulase production under the influences of different single factors.[Conclusion]The gut of G.orientalis harbors rich culturable bacterial resources,and 95%of the isolates of gut bacteria have the function of producing cellulase.Among the isolates,Bacillus subtilis N3 and N14 with the highest cellulase production demonstrate the most potential for development.

关 键 词：蝼蛄 肠道菌 产纤维素酶功能菌株 纤维素酶活性 

分 类 号：S433.84[农业科学—农业昆虫与害虫防治]