常温和低温纤维素降解菌的筛选及其酶活特性

doi:10.12160/j.issn.1672-5190.2023.02.002

摘要/Abstract

摘要：

[目的]筛选适宜在我国北方地区应用的常温和低温纤维素高效降解菌株。[方法]利用羧甲基纤维素钠培养基，从内蒙古地区采集的腐殖土、稻草、羊粪、驴粪样本中分离并初步筛选能够降解纤维素的细菌及真菌；分别采用16S rDNA和ITS基因序列分析技术确定细菌和真菌菌株的种属；通过纤维素刚果红水解圈试验、滤纸条降解试验对菌株进行进一步筛选。采用3，5-二硝基水杨酸（3，5-dinitrosalicylic acid，DNS）比色法测定菌株的纤维素酶活性。[结果]共分离获得44株纤维素降解菌，包括43株细菌和1株真菌，其中，27株为30 ℃条件下分离得到的常温纤维素降解菌，17株为18 ℃条件下分离得到的低温纤维素降解菌。在属水平上，芽孢杆菌属（Bacillus）为常温纤维素降解菌中第1优势菌属，假单胞杆菌属（Pseudomonas）为第2优势菌属；假单胞杆菌属为低温纤维素降解菌中第1优势菌属，不动杆菌属（Acinetobacter）和芽孢杆菌属为第2优势菌属。经纤维素刚果红水解圈试验、滤纸条降解试验进一步筛选出纤维素降解效果更好的35株菌，在此基础上通过DNS比色法筛选出3株羧甲基纤维素酶（carboxymethyl cellulose，CMCase）活性较高的菌株，分别为T-8-2（芽孢杆菌属）、LF-7（芽孢杆菌属）、LF-5（曲霉属），其CMCase活性分别为（169.24±2.18）、（136.79±2.05）、（116.01±0.52） U/mL，该3株菌的β-Gase、FPA、C₁活性也较高。[结论]研究揭示了内蒙古地区可培养纤维素降解菌的类群组成及纤维素降解能力。筛选出的3株CMCase、β-Gase、FPA、C₁活性较高的常温纤维素降解菌株可以作为有效降解纤维素的潜力菌株应用于粪污堆肥发酵剂的研制。

关键词: 纤维素降解菌, 常温, 低温, 分离筛选, 羧甲基纤维素酶活性

Abstract:

[Objective] The aim of this study was to screen high-efficient cellulose-degrading bacterial and fungal strains suitable for use in northern China at room and low temperatures. [Method] Using sodium carboxymethyl cellulose medium, bacteria and fungi capable of degrading cellulose were isolated and preliminary screened from the samples of humus soil, rice straw, sheep feces and donkey feces collected from Inner Mongolia. The species of the obtained bacterial and fungal strains were molecularly identified by using 16S rDNA and ITS gene sequence analysis, respectively. The strains were further screened on cellulose Congo-red agar medium and by filter paper degradation test. 3,5-dinitrosalicylic acid （DNS） colorimetry was used to measure the cellulase activities of the strains. [Result] A total of 44 cellulose-degrading strains, comprising 43 bacterial strains and 1 fungal strain, were isolated. Of these, 27 strains were isolated at 30 ℃ and exhibited cellulose-degrading ability at room temperature, and 17 strains were isolated at 18 ℃ and exhibited cellulose-degrading ability at low temperature. At the genus level, Bacillus was the first dominant genus among room temperature cellulose-degrading bacteria, followed by Pseudomonas. Among low temperature cellulose-degrading bacteria, Pseudomonas was the first dominant genus, followed by Acinetobacter and Bacillus. A total of 35 strains with improved cellulose-degrading efficiency were subsequently identified using cellulose Congo-red agar medium and filter paper degradation test. Furtherly, 3 strains, T-8-2 （Bacillus spp.）, LF-7 （Bacillus spp.） and LF-5 （Aspergillus spp.）, with higher carboxymethyl cellulase （CMCase） activities were isolated by DNS colorimetry, with CMCase activities of （169.24 ± 2.18）, （136.79 ± 2.05） and （116.01 ± 0.52） U/mL, respectively. These 3 strains also had higher activities of β-Gase, FPA and C₁. [Conclusion] This study characterizes the community composition and cellulose-degrading ability of the culturable cellulose-degrading bacteria and fungi in Inner Mongolia. The screened 3 strains exhibiting cellulose-degrading ability at room temperature with high activities of CMCase, β-Gase, FPA and C₁ can be used as potential strains for effectively degrading cellulose in the development of fermentation agents for manure composting.

Key words: cellulose-degrading strain, room temperature, low temperature, isolation and screening, carboxymethyl cellulase activity

中图分类号:

岳林芳, 成立新, 李蕴华, 苏少锋, 于朝晖, 王志铭, 吴海青. 常温和低温纤维素降解菌的筛选及其酶活特性[J]. 畜牧与饲料科学, 2023, 44(2): 9-16.

YUE Linfang, CHENG Lixin, LI Yunhua, SU Shaofeng, YU Zhaohui, WANG Zhiming, WU Haiqing. Screening and Enzyme Activity Characteristics of Cellulose-degrading Bacteria and Fungi Isolated at Room and Low Temperatures[J]. Animal Husbandry and Feed Science, 2023, 44(2): 9-16.

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