糙皮侧耳栽培料中纤维素降解菌的筛选及其复合菌系降解效果评价

doi:10.13560/j.cnki.biotech.bull.1985.2024-1167

摘要/Abstract

摘要：

目的从糙皮侧耳栽培料中筛选纤维素降解菌并构建高效复合菌系用于降解农业废弃物。方法以羧甲基纤维素钠为唯一碳源从糙皮侧耳栽培料中分离纯化菌株，刚果红染色进行初筛得到具有纤维素降解性能的细菌，利用分子生物学对其鉴定并构建系统发育进化树。选取相互无拮抗作用的菌株构建11个复合菌系，测定不同发酵时间下各复合菌系的纤维素相关酶活力。将最优纤维素降解复合菌系进行水稻秸秆和棉籽壳降解实验。结果筛选出4株纤维素降解能力较强的共生细菌，分别为链霉菌（Streptomyces sp.）、灿烂类芽胞杆菌（Paenibacillus lautus）、树状微杆菌（Microbacterium arborescens）和微杆菌（Microbacterium aoyamense）。在11个复合菌系中，由Streptomyces sp. Q5和P. lautus Q6组成的复合菌系H为最优纤维素降解复合菌系，培养第4天时其滤纸酶（filter paper cellulase, FPase）酶活相比单个菌株分别提高了229.97%和134.29%，羧甲基纤维素酶（carboxymethyl cellulase, CMCase）酶活相比单个菌株分别提高了92.81%和21.94%。复合菌系H处理15 d后水稻秸秆降解率为38.72%，棉籽壳降解率为35.76%，复合菌系H对水稻秸秆的木质素、纤维素和半纤维素的降解率分别为33.94%、31.17%和22.43%，对棉籽壳中木质素、纤维素和半纤维素的降解率分别为27.95%、25.56%和53.86%。扫描电镜结果表明水稻秸秆和棉籽壳经复合菌系处理后被充分降解。结论构建的复合菌系能够高效降解秸秆和棉籽壳，研究结果为提高农业废弃物资源的有效利用提供理论基础。

关键词: 复合菌系, 糙皮侧耳栽培料, 纤维素降解, 农业废弃物降解

Abstract:

Objective To screen cellulose-degrading bacteria from the cultivation substrate of Pleurotus ostreatus and construct an efficient microbial consortium for the degradation of agricultural waste. Method Bacterial strains were isolated and purified using sodium carboxymethyl cellulose (CMC-Na) as the sole carbon source from the cultivation substrate of P. ostreatus, and those with cellulose-degrading capabilities were preliminarily identified through Congo red staining. Molecular biology techniques were employed for their identification and to construct a phylogenetic tree. Eleven microbial consortiums were established from strains without mutual antagonism and the cellulase activities of these communities were measured at different fermentation times. The optimal cellulose-degrading microbial consortium was then used in degradation experiments with rice straw and cottonseed hulls to determine its cellulose-degrading capability. Result Four symbiotic bacteria with efficient cellulose-degrading capabilities were identified: Streptomyces sp., Paenibacillus lautus, Microbacterium arborescens and Microbacterium aoyamense. The results indicated that composite microbial system H, composed of Streptomyces sp. Q5 and P. lautus Q6, showed as the most effective cellulose-degrading strain. On the fourth day of culture, its filter paper cellulase (FPase) enzyme activity increased by 229.97% and 134.29%, respectively, compared to the single strains. Additionally, the carboxymethyl cellulase (CMCase) enzyme activity increased by 92.81% and 21.94%, respectively, when compared to the individual strains. After 15 d of treatment with composite microbial system H, the degradation rate was 38.72% for rice straw and 35.76% for cottonseed hulls. Specifically, the degradation rates of lignin, cellulose, and hemicellulose in the rice straw were 33.94%, 31.17%, and 22.43%, respectively, while the corresponding rates for cottonseed hulls were 27.95%, 25.56%, and 53.86%. Further analysis of the surface morphological changes of both rice straw and cottonseed hulls before and after degradation confirmed that composite microbial system H possessed a superior cellulose-degrading capability. Conclusion The microbial consortium constructed in this study can efficiently degrade straw and cottonseed hulls, providing a theoretical basis for improving the effective utilization of agricultural waste resources.

Key words: microbial consortium, Pleurotus ostreatus cultivation substrate, cellulose degradation, degradation of agricultural waste

刘姣姣, 穆德梅, 夏丽明, 方勇, 刘祚军. 糙皮侧耳栽培料中纤维素降解菌的筛选及其复合菌系降解效果评价[J]. 生物技术通报, 2025, 41(7): 326-335.

LIU Jiao-jiao, MU De-mei, XIA Li-ming, FANG Yong, LIU Zuo-jun. Screening of Cellulose-degrading Bacteria in Pleurotus ostreatus Cultivation Substrate and Evaluation of Degradation Effect of Microbial Consortium[J]. Biotechnology Bulletin, 2025, 41(7): 326-335.

图/表 9

表1 试验分组设计

Table 1 Experimental group design

复合菌系 Microbial consortium	菌株名称 Name of strain
A	Q3
B	Q5
C	Q6
D	Q7
E	Q3、Q5
F	Q3、Q6
G	Q3、Q7
H	Q5、Q6
I	Q5、Q7
J	Q6、Q7
K	Q3、Q5、Q6
L	Q3、Q5、Q7
M	Q3、Q6、Q7
N	Q5、Q6、Q7、
O	Q3、Q5、Q6、Q7

表2 纤维素降解菌株透明圈直径和菌落直径比

Table 2 Ratio of hyaline circle diameter to colony diameter of cellulose-degrading strains

菌株编号 Strain No.	透明圈直径 Transparent circle diameter （D）（cm）	菌落直径 Colony diameter （d）（cm）	D/d
Q1	0.44±0.06 d	0.19±0.01 f	2.40±0.36 e
Q2	0.72±0.21 d	0.26±0.01 d	2.83±0.93 de
Q3	1.47±0.13 c	0.21±0.01 ef	7.18±0.61 b
Q4	/	0.07±0.01 g	/
Q5	2.72±0.21 b	0.65±0.15 a	4.19±0.35 c
Q6	2.97±0.37 b	0.38±0.01 c	7.81±1.19 b
Q7	0.75±0.09 d	0.22±0.01 e	3.42±0.50 cd
Q8	/	0.06±0.01 g	/
Q9	0.57±0.19 d	0.39±0.08 c	1.47±0.55 f
糙皮侧耳	4.39±0.11 a	0.44±0.03 b	10.08±0.31 a

图1 纤维素降解菌株的刚果红透明圈CK：对照组；Q1-Q9：纤维素降解菌；P：糙皮侧耳

Fig. 1 Congo red clearance zones of cellulose-degrading bacterial strainsCK: Control group; Q1-Q9: cellulose-degrading bacteria; P: Pleurotus ostreatus

图2 基于16S rRNA序列构建的细菌系统发育树标注T的为模式菌株

Fig. 2 Phylogenetic tree based on the 16S rRNA sequences of the bacteriaThe strain marked with T is the model strain

表3 菌株之间的拮抗关系

Table 3 Stain relations between the antagonism

菌株编号 Strain No.	Q3	Q5	Q6	Q7
Q3	/	-	-	-
Q5	-	/	-	-
Q6	-	-	/	-
Q7	-	-	-	/

图3 单菌和复合菌系的酶活力A：单菌和复合菌系的滤纸酶活；B：单菌和复合菌系的CMCase酶活

Fig. 3 Enzyme activities of single strains and microbial consortiumA: Filter paper enzyme activity of single and compound bacterial systems; B: CMCase activity of single and compound bacterial systems

图4 菌株生长曲线图

Fig. 4 Growth curve of bacterial strain

图5 复合菌系对水稻秸秆和棉籽壳的降解率

Fig. 5 Degradation rates of rice straw and cottonseed hulls by microbial consortium

图6 水稻秸秆（A）和棉籽壳（B）经复合菌系降解前后的扫描电镜图RS-1：降解前水稻秸秆；RS-2：降解后水稻秸秆；CH-1：降解前棉籽壳；CH-2：降解后棉籽壳

Fig. 6 Graph of rice straw (A) and cottonseed hulls (B) before and after degradation by microbial consortiumsRS-1: Rice straw before degradation; RS-2: rice straw after degradation; CH-1: cottonseed hulls before degradation; CH-1: cottonseed hulls after degradation

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