基于EEM与PCR-DGGE技术分析温度对蚯蚓堆肥处理城镇污泥的影响

doi:10.16258/j.cnki.1674-5906.2021.05.019

生态环境学报 ›› 2021, Vol. 30 ›› Issue (5): 1060-1068.DOI: 10.16258/j.cnki.1674-5906.2021.05.019

基于EEM与PCR-DGGE技术分析温度对蚯蚓堆肥处理城镇污泥的影响

侯素霞¹(), 雷旭阳¹^,^*(), 张辉¹, 丁淑杰¹, 崔广宇²

1.邢台职业技术学院资源与环境工程系,河北邢台 054000
2.同济大学污染控制与资源化研究国家重点实验室,上海 200092

收稿日期:2021-01-26 出版日期:2021-05-18 发布日期:2021-08-06
通讯作者: * 雷旭阳（1989年生）,男,讲师,硕士,主要从事废水处理与污泥资源化方面的研究。E-mail:67227237@qq.com
作者简介:侯素霞（1981年生）,女,副教授,硕士,主要从事废水与污泥处理方面的研究。E-mail:housxhb@foxmail.com
基金资助:
河北省科技支撑计划项目(18273624);邢台市重点研发计划项目(2020ZC126)

Analysis of the Effect of Temperature on Vermicomposting of Municipal Sludge Based on EEM and PCR-DGGE

HOU Suxia¹(), LEI Xuyang¹^,^*(), ZHANG Hui¹, DING Shujie¹, CUI Guangyu²

1. Department of Resource and Environmental Engineering, Xingtai Polytechnic College, Xingtai 054000 China
2. State Key Laboratory of Pollution Control and Resource Reuse, Tongji University, Shanghai 200092 China

Received:2021-01-26 Online:2021-05-18 Published:2021-08-06

摘要/Abstract

摘要：

将城镇脱水污泥制成5 mm粒径颗粒分别在15、20、25 ℃条件下进行蚯蚓堆肥,利用三维荧光（EEM）考察了温度对蚯蚓堆肥处理城镇污泥过程腐熟程度的影响,并采用PCR-DGGE技术探究了不同温度条件下蚯蚓堆肥微生物种群结构的变化,旨在分析蚯蚓堆肥中的微生物种群结构的变化,进而验证污泥中腐殖酸类有机物熟化降解机理,并为蚯蚓堆肥处理城镇污泥实现工业化生产提供理论依据。EEM分析结果表明：在堆肥30 d时,堆肥系统中腐殖酸类物质已基本降解完全,且随着堆肥时间的延长,芳香类蛋白质一直在被蚯蚓和微生物所利用;同一时期随温度升高,蚯蚓吞食和系统内的微生物的协同增效作用能加快系统中DOM的降解效率、提高系统的矿化程度。依据PCR-DGGE技术测序结果可知：蚯蚓堆肥系统中微生物种群对于温度变化存在缓冲区间,当温度在15—20 ℃范围内变化时,细菌和真核微生物的种群多样性差异均较小;温度为25 ℃时,系统中细菌种群多样性降低,真核微生物种群多样性升高;堆肥结束时,3个温度组系统中共有细菌种群类别占系统中总细菌种群类别79%以上,共有真核微生物种群类别占系统总真核微生物种群类别75%以上,表明随着温度升高系统中主要微生物种群类别并未发生变化,仅共有微生物种群的相对数量发生了改变,且3个温度组系统中均以拟杆菌门（Bacteroidetes）和子囊菌门（Ascomycota）为优势种群,温度越高,相对数量越多,系统中易利用有机质所占比重小,系统更加稳定。

关键词: 蚯蚓堆肥, 温度, 城镇污泥, 三维荧光, PCR-DGGE, 水溶性有机质, 微生物种群

Abstract:

This study aimed to elucidate the effect of temperature on the process of vermicomposting with dewatered municipal sludge which made into 5mm particles, and research the changes at different temperatures (15 ℃, 20 ℃, 25 ℃) by EEM spectra. In addition, the microbial communities in different temperatures were analyzed by molecular biology PCR-DGGE assessment. The aim of this study was to verify the change of microbial population structure in earthworm composting by fingerprint sequencing results, and further verify the ripening and degradation mechanism of humic acid organic matter in sludge, and provide a theoretical basis for earthworm composting treatment of urban sludge to realize industrial production. The results of the EEM spectra showed that the humic acids in the composting system had been degraded completely after 30 days of composting, and with the extension of the composting time, the aromatic proteins were still used by earthworms and microorganisms until the end of composting. During the same period, with the increase of temperature, the synergistic effect of earthworm ingestion and microorganisms can accelerate the degradation efficiency of DOM in the system, improve the degree of mineralization of the system, and promote the stabilization efficiency. According to molecular biology PCR-DGGE assessment, the change of microbial population with temperature existed in the buffer zone, in the vermicomposting system. When the temperature changed in the range of 15?20 ℃, the difference of bacterial and eukaryotic microbial population diversity was small. When the temperature was 25 ℃, the bacterial population diversity decreased and the eukaryotic microbial population diversity increased. At the end of composting, The common bacterial population category accounted for more than 79% of the total bacterial population category, and the common eukaryotic microbial population category accounted for more than 75% of the total eukaryotic microbial population category in the three systems, indicating that the main microbial population category did not change with the increase of temperature, only the relative number of the common microbial population changed. At the same time, Bacteroidetes and Ascomycota predominated in three systems, and the relative quantities of the dominant microbial populations and the proportion of easy to use organic matter in the system reduced with the temperature increasing. Meanwhile, the system was more stable.

Key words: vermicomposting, temperature, municipal sludge, three-dimensional fluorescence, PCR-DGGE, dissolved organic matter, microbial population

中图分类号:

X703

侯素霞, 雷旭阳, 张辉, 丁淑杰, 崔广宇. 基于EEM与PCR-DGGE技术分析温度对蚯蚓堆肥处理城镇污泥的影响[J]. 生态环境学报, 2021, 30(5): 1060-1068.

HOU Suxia, LEI Xuyang, ZHANG Hui, DING Shujie, CUI Guangyu. Analysis of the Effect of Temperature on Vermicomposting of Municipal Sludge Based on EEM and PCR-DGGE[J]. Ecology and Environment, 2021, 30(5): 1060-1068.

图/表 7

表1 实验用泥开始和结束时理化、生化指标

Table 1 Physicochemical parameters of the initial substrate and the composting products

指标 Physicochemical Parameters	初始污泥 Initial Sludge	The Composting Products (60 d)
指标 Physicochemical Parameters	初始污泥 Initial Sludge	15 ℃	20 ℃	25 ℃
ω(OM)/%	68.0±7.7	55.0±0.01	51.7±0.02	49.8±0.01
ω(DOC)/(g∙kg^-1)	16.69±2.32	12.88±1.66	13.98±2.31	7.60±1.29
γ/(µS∙cm^-1)	573±8.49	1035.5±2.04	1275.0±6.94	2100.0±16.32
ω(NH₄⁺-N)/(mg·kg^-1)	7.36±0.08	64.99±0.76	107.32±0.23	296.20±2.90
ω(NO₃^--N)/(mg∙kg^-1)	10.26±2.0	219.91±45.15	285.10±9.10	1389.49±47.06
ω(MBC)/(g∙kg^-1)	105.61±10.60	15.63±0.98	8.50±0.18	6.71±1.08
DHA/[mg·g^-1∙h^-1]	28.06±1.34	0.49±0.08	0.49±0.02	0.27±0.01

图1 堆体不同阶段DOM的EEM谱图

Fig. 1 EEM spectra of DOM at different stages

图2 16S rDNA（A）和18S rDNA（B）的DGGE指纹图谱

Fig. 2 DGGE fingerprint and its schematic representation of 16S rDNA(A) and 18S rDNA(B) at three different temperature conditions

图3 不同温度16S rDNA和18S rDNA的DGGE指纹图谱Shannon-Wiener指数

Fig. 3 Shannon-Wiener index of biodiversity on 16S rDNA and 18S rDNA at three different temperature conditions

图4 16S rDNA（a）和18S rDNA（b）的DGGE指纹图谱PCA分析图“△”：15 ℃组独有。“□”：25 ℃组独有。“*”：15 ℃组和20 ℃组共有。“+”：20 ℃组和25 ℃组共有。“●”：15 ℃组、20 ℃组和25 ℃组共有

Fig. 4 Principal components analysis of the microbial community on 16S rDNA (a) and 18S rDNA (b)“△”: 15 ℃ group particular. “□”: 25 ℃ group particular. “*”: 15 ℃ group and 20 ℃ group common. “+”: 20 ℃ group and 25 ℃ group common. “●”: 15 ℃ group, 20 ℃ group and 25 ℃ group common

图5 16S rDNA的DGGE图谱系统发育树

Fig. 5 Neighbour-joining tree of partial 16S rDNA gene sequences from DGGE image

图6 18S rDNA的DGGE图谱系统发育树

Fig. 6 Neighbour-joining tree of partial 18S rDNA gene sequences from DGGE image

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基于EEM与PCR-DGGE技术分析温度对蚯蚓堆肥处理城镇污泥的影响

Analysis of the Effect of Temperature on Vermicomposting of Municipal Sludge Based on EEM and PCR-DGGE

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