Screening and Identification of Efficient Degradation Fungi for Garden Waste Lignocellulose

doi:10.15933/j.cnki.1004-3268.2024.05.011

Abstract

Abstract: In order to screen fungal strains for efficient degradation of lignocellulose in garden waste,PDA medium was used to separate fungi from humus,cow manure,horse manure,dead branches and leaves,compost,straw and other environments and aniline blue medium and filter paper inorganic salt medium were used to qualitatively screen fungal strains with lignocellulosic degrading ability.The activity of lignocellulosic degrading enzymes and their degradation rate on garden waste were quantitatively measured，and excellent fungal strains for efficient degradation of lignocellulose in garden waste were screened.The results showed that a total of 109 strains of fungi were isolated. Among them，there were 32 fungi with lignin degradation ability，accounting for 29.36% of the tested strains;There were 40 fungi with cellulose degradation ability，accounting for 36.70% of the tested strains.Among the above strains，BF31‑32,BF32‑93,T2‑55,T1‑16，and T1‑24 could efficiently degrade lignocellulose from garden waste,with degradation rates of 50.30%,45.94%,44.49%,44.68%,and 46.81%,respectively.Based on the lignocellulosic degrading enzyme activity，morphological characteristics,and molecular identification results of fungi,strain BF31‑32 was identified as Penicillium brevicompactum,strain BF32‑93 as Aspergillus fumigatus,strains T1‑16 and T1‑24 as Fusarium equiseti,and strain T2‑55 as Paecilomyces lilacinus.

Key words: Garden waste, Lignin, Cellulose, Enzyme activity, Biodegradable fungi

摘要： 为筛选园林废弃物木质纤维素高效降解真菌菌株，利用PDA培养基分离腐殖土、牛粪、马粪、枯枝落叶、堆肥、秸秆等环境中的真菌，利用苯胺蓝筛选培养基、滤纸无机盐培养基定性筛选具有木质纤维素降解能力的真菌菌株，并定量测定其木质纤维素降解酶活性及其对园林废弃物的降解率，进而筛选出高效降解园林废弃物木质纤维素的优良真菌菌株。结果表明，共分离出109株真菌。其中，具有木质素降解能力的真菌有32株，占供试菌株的29.36%；具有纤维素降解能力的真菌有40株，占供试菌株的36.70%。上述菌株中，BF31-32、BF32-93、T2-55、T1-16、T1-24均可高效降解园林废弃物木质纤维素，其园林废弃物降解率分别为50.30%、45.94%、44.49%、44.68%、46.81%。结合真菌的木质纤维素降解酶活性、形态学特征及分子鉴定结果，将菌株BF31-32 鉴定为短密青霉菌（Penicillium brevicompactum）、菌株BF32-93鉴定为烟曲霉（Aspergillus fumigatus）、菌株T1-16和菌株T1-24鉴定为木贼镰孢菌（Fusarium equiseti）、菌株T2-55鉴定为淡紫拟青霉（Paecilomyces lilacinus）。

关键词: 园林废弃物, 木质素, 纤维素, 酶活性, 生物降解真菌

CLC Number:

S718.81

HUANG Qiuyue, HE Jianqing, LIU Haixin, NIE Zifang, XU Dong, PAN Changmang. Screening and Identification of Efficient Degradation Fungi for Garden Waste Lignocellulose[J]. Journal of Henan Agricultural Sciences, 2024, 53(5): 101-111.

黄秋月, 何建清, 刘海鑫, 聂子芳, 徐东, 潘长漭. 园林废弃物木质纤维素高效降解真菌的筛选与鉴定[J]. 河南农业科学, 2024, 53(5): 101-111.

References

［1］李婧男，汪群慧，梁宝瑞，等.园林废弃物对滨海盐渍土固碳减排效应与微生物群落结构的影响［J］.环境工程，2024，42（1）：95‑101.
LI J N，JIANG Q H，LIANG B R，et al.Effects of garden waste on emission reduction and microbial community in coastal saline soil［J］.Environmental Engineering，2024，42（1）：95‑101.
［2］林鹏.园林废弃物生态处理及资源化利用［J］.现代园艺，2023，46（6）：195‑197.
LIN P.Ecological treatment and resource utilization of garden waste［J］.Contemporary Horticulture，2023，46（6）：195‑197.

［3］李蕾.纤维素酶高产木霉菌株的筛选及其对秸秆的降解效果［D］.兰州：甘肃农业大学，2014.

LI L.Screening of cellulase highly producing Trichoderma strains and their degradation efficiency of crop straw［D］.Lanzhou：Gansu Agricultural University，2014.

［4］刘宽庆，张以恒.木质素的生物降解和生物利用［J/OL］.合成生物学：1‑14［2024‑04‑19］.http：//kns.cnki.net/kcms/detail/10.1687.Q.20231103.0847.002.html.

LIU K Q，ZHANG Y H.Biological degradation and utilization of lignin［J/OL］.Synthetic Biology Journal：1‑14［2024‑04‑19］.http：//kns.cnki.net/kcms/detail/10.1687.Q.20231103.0847.002.html.

［5］王国栋，陈晓亚.漆酶的性质、功能、催化机理和应用［J］.植物学通报，2003，38（4）：469‑475.

WANG G D，CHEN X Y.The properties，functions，catalytic mechanism and applicability of laccase［J］.Chinese Bulletin of Botany，2003，38（4）：469‑475.

［6］CHAKRAVARTY D， TEWARI H， BALLAL A.Harnessing salt stress to drive the efficient and inexpensive bioproduction of a valuable thermotolerant biocatalyst（Mn‑catalase） from cyanobacterial biomass［J］.Bioresource Technology Reports，2023，23：101535.

［7］HOSSEIN B，TAGHI G，ABDOLHAKIM T，et al.Growth performance，ruminal fermentation characteristics and microbial protein synthesis of lambs fed palmitic and stearic acid［J］.Animal Feed Science and Technology，2023，302：5674.

［8］ABDEL‑SALAM M S，HAFEZ S S，FADEL M，et al.Bio ethanol production from rice straw saccharification via avicelase gene in E. coli recombinant strain［J］.Clean Technologies，2023，5（2）：451‑465.

［9］高凤菊，李春香. 真菌与细菌纤维素酶研究进展［J］.唐山师范学院学报，2005，27（2）：7‑10.

GAO F J，LI C X. Studies on cellulase from Eubacterium and bacterium ［J］.Journal of Tangshang Teachers College，2005，27（2）：7‑10.

［10］张斯童，兰雪，李哲，等.微生物降解玉米秸秆的研究进展［J］.吉林农业大学学报，2016，38（5）：517‑522.

ZHANG S T，LAN X，LI Z，et al.Research progress of microbial degradation of corn straw［J］.Journal of Jilin

Agricultural University，2016，38（5）：517‑522.

［11］SHRIVASTAVA B，THAKUR S，KHASA Y P，et al.White‑rot fungal conversion of wheat straw to energy rich cattle feed［J］.Biodegradation，2011，22（4）：823‑831.

［12］黄亚丽，黄媛媛，马慧媛，等.低温秸秆降解真菌的筛选及在秸秆还田中的应用［J］.中国农学通报，2020，36（21）：53‑60.

HUANG Y L，HUANG Y Y，MA H Y，et al.Fungi with corn straw decomposing characterization at low‑temperature：Screening and application in straw returning field ［J］.Chinese Agricultural Science Bulletin，2020，36（21）：53‑60.

［13］刘起丽，张建新，葛文娇，等.一株产纤维素酶真菌的筛选及其对秸秆的降解效果研究［J］.现代食品科技，2014，30（6）：82‑86.

LIU Q L，ZHANG J X，GE W J，et al.The screening and identification of a cellulose degrading fungus and its straw degradating capability［J］.Modern Food Science and Technology，2014，30（6）：82‑86.

［14］宁振兴，陈皓睿，刘鸿，等.一株能高效降解梗丝果胶和纤维素的曲霉的筛选和鉴定［J］.基因组学与应用生物学，2016，35（8）：2077‑2082.

NING Z X，CHEN H R，LIU H，et al.Isolation and identification of an Aspergillus strain which is effective in the degration of pectin and cellulose for tobacco stem［J］.Genomics and Applied Biology，2016，35（8）：2077‑2082.

［15］BANSAL N，TEWARI R，SONI R，et al.Production of cellulases from Aspergillus niger NS‑2 in solid state fermentation on agricultural and kitchen waste residues［J］.Waste Management，2012，32（7）：1341‑1346.

［16］孙思琦，瓮岳太，邸雪颖，等.木质素降解真菌的筛选及其对森林地表可燃物的降解效果［J］.中南林业科技大学学报，2021，41（1）：29‑36.

SUN S Q，WENG Y T，DI X Y，et al.Screening of lignin‑degrading fungi and their degradation effect of forest surface fuel［J］.Journal of Central South University of Forestry & Technology，2021，41（1）：29‑36.

［17］戚业强.木质素降解菌株的筛选及其降解秸秆木质素的研究［D］.哈尔滨：东北农业大学，2021.

QI Y Q.Screening of lignin degrading strains and the degradation of lignin in straw［D］.Harbin：Northeast Agricultural University，2021.

［18］李雅琳，李素艳，孙向阳，等.1株木质素降解菌的筛选、鉴定及液态发酵条件优化［J］.浙江农林大学学报，2021，38（6）：1297‑1304.

LI Y L，LI S Y，SUN X Y，et al.Screening and identification of a lignin degrading strain and its optimized liquid fermentation conditions［J］.Journal of Zhejiang A & F University，2021，38（6）：1297‑1304.

［19］赵文萱，鞠志刚，郑亚强，等.一株高效降解麦秸木质素菌株的筛选及效能评价［J］.山东农业大学学报（自然科学版），2022，53（3）：386‑392.

ZHAO W X，JU Z G，ZHENG Y Q，et al.Screening and efficacy evaluation on a strain efficiently degrading wheat straw lignin［J］.Journal of Shandong Agricultural University（Natural Science Edition），2022，53（3）：386‑392.

［20］张漾泓，张少冰，张家顺，等.一株新分离的云芝栓孔菌Z-1应用于木质素降解及染料脱色［J］.菌物学报，2021，40（7）：1854‑1868.

ZHANG Y H，ZHANG S B，ZHANG J S，et al.A newly isolated Trametes versicolor Z‑1 and its application to lignin degradation and dye decolorization［J］.Mycosystema，2021，40（7）：1854‑1868.

［21］张芳芳，张桐，戴丹，等.高效木质素降解菌的筛选及其对玉米秸秆的降解效果［J］.菌物学报，2021，40 （7）：1869‑1880.

ZHANG F F，ZHANG T，DAI D，et al.Screening of efficient lignin‑degrading fungal strains and their degradation on cornstalk［J］.Mycosystema，2021，40 （7）：1869‑1880.

［22］王敬红，李灵灵，申贵男，等.木质素降解真菌菌群LDFC产酶条件优化［J］.黑龙江八一农垦大学学报，2022，34（2）：75‑80.

WANG J H，LI L L，SHEN G N，et al.Optimization of enzyme‑producing conditions of lignin‑degrading fungal flora LDFC［J］.Journal of Heilongjiang Bayi Agricultural University，2022，34（2）：75‑80.

［23］李灵灵，王敬红，赵铎，等.木质素降解菌BYL-7的筛选及降解条件优化［J］.微生物学通报，2020，47（12）：4059‑4071.

LI L L，WANG J H，ZHAO D，et al.Screening of lignin degrading strain BYL‑7 and optimization of degradation conditions［J］.Microbiology China，2020，47（12）：4059‑4071.

［24］张家齐.园林废弃物堆腐微生物过程及纤维素降解菌筛选研究［D］.北京：中国林业科学研究院，2013.

ZHANG J Q.The microbial process of garden waste decomposition and screening of cellulose‑degrading microbe［D］.Beijing：Chinese Academy of Forestry，2013.

［25］吴桐.园林生物质降解菌的筛选及其堆肥应用［D］.西安：西安科技大学，2021.

WU T.Screening of garden biomass degrading bacteria and the compost application［D］.Xi’an：Xi’an University of Science and Technology，2021.

［26］孙江慧.几株食用菌对秸秆木质纤维素降解能力的研究［D］.南京：南京农业大学，2012.

SUN J H.Study on potential ability of several edible fungi biodegradation of lignocelluloses of straw［D］.Nanjing：Nanjing Agricultural University，2012.

［27］冯茜，国巍，燕红.木质素降解菌的分离筛选及菌丝成球条件优化［J］.哈尔滨理工大学学报，2019，24（1）：138‑144.

FENG Q，GUO W，YAN H.Isolation and screening of the lignin‑degrading fungusand optimization of the Mycelium pellet cultivation conditions［J］.Journal of Harbin University of Science and Technology，2019，24 （1）：138‑144.

［28］王茂成，李勇，李世忠，等.木质素降解真菌的分离与相关酶活性测定及产酶条件优化［J］.西南师范大学学报（自然科学版），2013，38（11）：122‑126.

WANG M C，LI Y，LI S Z，et al.On separation of lignin degrading fungi，related enzyme activity assay and optimized conditions for enzyme production［J］.Journal of Southwest China Normal University（Natural Science Edition），2013，38（11）：122‑126.

［29］池玉杰，闫洪波.红平菇木质素降解酶系统漆酶、锰过氧化物酶及木质素过氧化物酶的检测［J］.林业科学，2009，45（12）：154‑158.

CHI Y J，YAN H B. Detection on laccase，manganese peroxidase and lignin peroxidase in ligninolytic enzymes of Pleurotus djamor［J］.Scientia Silvae Sinicae，2009，45（12）：154‑158.

［30］李美群，孟勇，杨明，等.不同处理方式对毛竹篼纤维素及木质素酶活的影响［J］.西南林业大学学报，2015，35（6）：31‑36.
LI M Q，MENG Y，YANG M，et al.Effects of different treatments on cellulase and lignin enzyme activity in bamboo stump［J］.Journal of Southwest Forestry University，2015，35（6）：31‑36.

［31］邹琦.植物生理学实验指导［M］.北京：中国农业出版社，2000：110‑130.
ZOU Q.Experimental supervision of plant physiology［M］.Beijing：China Agriculture Press，2000：110‑130.

［32］ZHANG Z，LIU J L，LAN J Y，et al.Predominance of Trichoderma and Penicillium in cellulolytic aerobic filamentous fungi from subtropical and tropical forests in China，and their use in finding highly efficient β‑glucosidase［J］.Biotechnology for Biofuels，2014，7 （1）：1‑14.

［33］赵昕，曲音波，高培基.抗黑液毒性的纤维素酶产生菌选育的探讨［J］.纤维素科学与技术，1993，1（2）：28‑32.
ZHAO X，QU Y B，GAO P J.The discussion on the selection of a cellulose：Production strain in black liquor［J］.Journal of Cellulose Science and Technology，1993，1（2）：28‑32.

［34］王欣宇.特异性产纤维素酶东北红豆杉内生真菌的筛选及初步应用［D］.哈尔滨：东北林业大学，2020.
WANG X Y.Screening of endophytic fungi of Taxus cuspidata S.et Z.for specific production of cellulase and its preliminary application［D］.Harbin：Northeast Forestry University，2020.

［35］刘孝文.木质纤维素降解菌及其组合对油棕棕榈丝降解的影响［D］.南京：南京农业大学，2017.
LIU X W.Influence of different combination of lignocellulose decomposing strains on oil palm fiber degradation［D］.Nanjing： Nanjing Agricultural University，2017.

［36］金伟，陈文静，缪礼鸿，等.1株耐热纤维素酶产生菌的筛选及其酶学特性［J］.江苏农业科学，2017，45（20）：272‑274.
JIN W，CHEN W J，MIAO L H，et al.Screening of a thermostable cellulase‑producing strain and its enzymatic characteristics［J］.Jiangsu Agricultural Sciences，2017，45（20）：272‑274.

［37］刘可，王远宏，常若葵，等.促秸秆腐熟真菌筛选实验初报［J］.浙江农业科学，2018，59（11）：2129‑2131.
LIU K，WANG Y H，CHANG R K，et al.Screening of soil fungus for straw maturing［J］.Journal of Zhejiang Agricultural Sciences，2018，59（11）：2129‑2131.

［38］LIU D Y，ZHANG R F，YANG X M，et al.Thermostable cellulase production of Aspergillus fumigatus Z5 under solid‑state fermentation and its application in degradation of agricultural wastes［J］.International Biodeterioration & Biodegradation，2011，65（5）：717‑725.

［39］孙曼钰，李栋梁，舒月力，等.尖孢镰刀菌诱导产酶及同步糖化发酵产纤维素乙醇［J］.江苏农业科学，2019，47（2）：277‑281.
SUN M Y，LI D L，SHU Y L，et al.Production of cellulase induced by Fusarium oxysporum and production of cellulosic ethanol by simultaneous saccharification fermentation［J］.Jiangsu Agricultural Sciences，2019，47（2）：277‑281.

［40］骆静怡.木腐真菌对金属硼处理材的腐朽能力研究［D］.杭州：浙江农林大学，2014.
LUO J Y.Decay of metaborates‑treated wood by wood‑rotting fungi［D］.Hangzhou：Zhejiang A & F University，2014.

［41］王和玉，刘延峰，张巧玲，等.1株来源于茅台酒酿造过程宛氏拟青霉MTDF-01的全基因组测序及分析［J］.食品科学，2019，40（24）：185‑192.
WANG H Y，LIU Y F，ZHANG Q L，et al.Whole‑genome sequencing and analysis of Paecilomyces variotii MTDF‑01，isolated during Maotai‑flavor liquor brewing［J］.Food Science，2019，40（24）：185‑192.

［42］蒋荣清，袁兴中，曾光明，等.一组高效木质素降解复合菌的筛选［J］.应用与环境生物学报，2010，16（2）：247‑251.
JIANG R Q，YUAN X Z，ZENG G M，et al.Screening of an efficient lignin‑degrading composite microorganisms［J］.Chinese Journal of Applied & Environmental Biology，2010，16（2）：247‑251.

［43］李春黎，李文豪，余君，等.烟秆高效复合降解菌群的筛选、鉴定及组合应用［J］.中国烟草科学，2019，40 （6）：26‑32.
LI C L，LI W H，YU J，et al.Screening，identification and combination of high efficiency degrading bacteria for tobacco stem［J］.Chinese Tobacco Science，2019，40（6）：26‑32.

[1]	LIU Lina, LI Shunfeng, XU Fangfang, CUI Guomei, GAO Shuaiping, WANG Anjian, TIAN Guangrui, WEI Shuxin. Characteristics of Lignocellulose Variation in Lentinus edodes Culture Material during Different Growth Periods [J]. Journal of Henan Agricultural Sciences, 2024, 53(3): 110-117.
[2]	KONG Fandan, ZHOU Lijun, ZHENG Meiyu, ZHANG Zuohe, YANG Zeyi, WU Juan. Effects of Straw Mulching on Soil Microbial Biomass，Enzyme Activity，and Soybean Yield in Black Soil Areas [J]. Journal of Henan Agricultural Sciences, 2024, 53(1): 87-95.
[3]	YU Gao, CHEN Fen, TIAN Xia, LU Xin, TENG Minghuan, XIE Wanying. Effects of Winter Mulching on Soil Chemical Properties and Enzyme Activities in Young Citrus Orchards [J]. Journal of Henan Agricultural Sciences, 2023, 52(9): 91-101.
[4]	YU Gao, WANG Hai, MO Zhongmei, CHEN Fen. Rhizome Quality Characteristics of Polygonatum rhizoma in Different Habitats and Driving Factors in Rhizosphere Soil Environment [J]. Journal of Henan Agricultural Sciences, 2023, 52(12): 57-68.
[5]	LIU Wei, ZHAO Yuanyuan, CHEN Xiaolong, WEI Jianyu, LI Juan, XIE Tianqi, SHI Hongzhi. Effects of Different Organic Fertilizers on Soil Nitrogen Mineralization and Enzyme Activity in the Tobacco Growing Areas of Central Henan and Western Yunnan [J]. Journal of Henan Agricultural Sciences, 2023, 52(11): 85-93.
[6]	WANG Yongsheng, ZHU Shuifen, CHEN Song, CHEN Yunming, JIANG Ya, FAN Yonghui, YANG Qihui, LI Qiongyan, YANG Hai. Effects of Artificial Diet and Mulberry Leaves on Activities of Three Metabolic Enzymes in Bombyx mori [J]. Journal of Henan Agricultural Sciences, 2023, 52(1): 154-160.
[7]	LOU Yusui, SHANG Hongquan, FAN Hongjie, LI Zheng, ZHANG Ke, LÜ Zhongwei, ZHANG Xiaofeng, WU Wenying. Changes in Berry Peel Structure and Related Substances of Shine Muscat Grape with Different Berry Russet Levels [J]. Journal of Henan Agricultural Sciences, 2022, 51(9): 105-113.
[8]	ZHANG Zhichao, GAO Ming, ZHAO Hao, WANG Xiaoya, CAO Zijing, WU Kun, XU Shuxia. Response of Soil Microecology to Quinclorac Contamination and Remediation by Enterobacter ludwigii EM1 [J]. Journal of Henan Agricultural Sciences, 2022, 51(4): 68-76.
[9]	WU Fengxia, SUN Yue, XIAO Qiang, ZHANG Shubin, LI Yufei, LIU Jianbin. Degradation Ability and Influencing Factors of Cellulose‑degradation Microbial Community PN‑8 on Wheat Straw [J]. Journal of Henan Agricultural Sciences, 2022, 51(4): 77-86.
[10]	WANG Yuanyuan, HAN Xiuli, YAN Zhenli, DU Chaojun, CHEN Yujie, CHANG Chun. Research Progress on Pretreatment Technology of Straw Biomass [J]. Journal of Henan Agricultural Sciences, 2022, 51(11): 1-10.
[11]	WU Yaoling, XI Xiaoli, ZENG Xianglian, LÜ Xibin, WANG Heyu, WANG Li. Isolation and Compost Application of High‐Activity Cellulose‐degrading Strains from Distiller’s Grains [J]. Journal of Henan Agricultural Sciences, 2022, 51(1): 79-88.
[12]	CHEN Fen, YU Gao, ZHANG Hongli, WU Hanqian, CHEN Rong, HOU Jianwei. Effects of Bio⁃organic Fertilizer Made from Chinese Traditional Herb Residues on the Content of Heavy Metals,Microbial Biomass Carbon and Nitrogen and Enzyme Activity in Hg and Cd Compound Contaminated Soil [J]. Journal of Henan Agricultural Sciences, 2021, 50(7): 66-75.
[13]	WU Jicheng, PAN Xiaoying, YANG Yonghui, GAO Cuimin, WANG Yue, . Effects of Long⁃term Application of Biogas Slurry on Soil Nutrient Content and Enzyme Activity [J]. Journal of Henan Agricultural Sciences, 2021, 50(7): 76-86.
[14]	WU Changwei, GONG Xinguo, ZHENG Lin, ZHANG Bo, JIANG Meihong, LI Xueshan, HUANG Zhiqiang, PU Xingwei, WU Mingmei, LI Xian. Degrading Tobacco Stem Cellulose by HF-09 Strain and Producing Aroma by Resting Cell Fermentation [J]. Journal of Henan Agricultural Sciences, 2021, 50(6): 171-180.
[15]	ZHENG Jianqiang, LIU Bin, WANG Xiaofei, JIA Xuewei, XU Chunping. Preparation and Characterization of Nanocellulose from Tobacco Straw [J]. Journal of Henan Agricultural Sciences, 2021, 50(3): 165-173.