再生固体牛粪垫料中细菌多样性分析及评价

doi:10.6048/j.issn.1001-4330.2021.12.021

新疆农业科学 ›› 2021, Vol. 58 ›› Issue (12): 2334-2341.DOI: 10.6048/j.issn.1001-4330.2021.12.021

再生固体牛粪垫料中细菌多样性分析及评价

刘建成¹(), 曾军², 丁峰¹, 许先查³, 窦晶晶¹, 陈开旭¹, 李凤鸣¹(), 高雁²()

1.新疆农业大学动物科学学院,乌鲁木齐 830052
2.新疆农业科学院微生物应用研究所/新疆特殊环境微生物实验室,乌鲁木齐 830091
3.新疆机场(集团)天缘绿色产业有限责任公司,新疆昌吉 831100

收稿日期:2021-03-29 出版日期:2021-12-20 发布日期:2021-12-31
通信作者: 李凤鸣,高雁
作者简介:刘建成(1982-),男,研究方向为养殖粪污处理及其资源化利用,(E-mail) 70964390@qq.com
基金资助:
新疆维吾尔自治区重大科技专项(2020A01001-2);新疆农业科学院科技创新重点培育专项(xjkcpy-2020005);新疆农业大学大学生创新项目(dxscx2020163)

Analysis of Bacterial Diversity and Safety Evaluation of the Recycled Manure Solids

LIU Jiancheng¹(), ZENG Jun², DING Feng¹, XU Xiancha³, DOU Jinjin¹, CHEN Kaixu¹, LI Fengmin¹(), GAO Yan²()

1. College of Animal Sciences, Xinjiang Agricultural University,Urumqi 830052,China
2. Institute of Applied Microbiology,Xinjiang Academy of Agricultural Sciences/Xinjiang Special Environmental Microbiology Laboratory,Urumqi 830091, China
3. Tianyuan Green Industry Co. Ltd., Xinjiang Airport(Group),Changji Xinjiang 831100,China

Received:2021-03-29 Online:2021-12-20 Published:2021-12-31
Correspondence author: LI Fengmin, GAO Yan
Supported by:
Ear-marked Fund of Major S & T Project of Xinjiang Uygur Autonomous Region(2020A01001-2);Key Scientific and Technological Innovation Project of Xinjiang Academy of Agriculture Sciences(xjkcpy-2020005);Xinjiang Agricultural University Student Innovation Project(dxscx2020163)

摘要/Abstract

摘要： 目的评估牛粪垫料再生系统(Bedding Recovery Unit,BRU)处理的再生固体牛粪垫料(Recycled Manure Solids,RMS)的安全性,为养殖场安全合理应用RMS提供理论依据和参考。方法采用可培养法对BRU系统处理前、处理后及堆存30 d的RMS样品中细菌总数、大肠杆菌、金黄色葡萄球菌、无乳链球菌和沙门氏菌的数量进行检测,并利用高通量测序方法分析样品中致病细菌群落组成和多样性。结果牛粪经BRU系统好氧发酵16~18 h后,细菌总数和大肠杆菌数量均显著下降,且都下降了一个数量级;金黄色葡萄球菌数量未发生显著变化(P>0.05);RMS在堆放30 d后,物料中的金黄色葡萄球菌数量显著下降;而细菌总数和大肠杆菌数量又呈现出了显著增加的趋势(P<0.05),在3种样品中均未检测到无乳链球菌和沙门氏菌。利用分子生物学方法仅在未经BRU系统处理的样本中检测到条件致病菌4个属:气球菌属(丰度为1.40%±0.8%)、费克蓝姆氏菌属(20.73%±3.97%)、厌氧梭菌属(7.20%±2.35%)和肉杆菌属(0.27%±0.15%)。经BRU系统处理后样品中细菌Alpha多样性指数呈现出极显著的下降趋势(P<0.01),表现在细菌OTU数量上减少了57.73%,细菌丰富度指数(ACE指数)降低了57.47%,香农-威纳多样性指数降低了81.63%。而RMS经过堆存30 d后,香农-威纳多样性指数显著增加了21.76%。经BRU系统处理显著的改变了细菌群落组成,并且其群落组成在堆存30 d后出现显著增加(P<0.01)。结论牛粪经BRU系统处理16~18 h后能够显著降低RMS中细菌多样性,改变群落组成,但其对除大肠杆菌外其他致病菌的杀灭作用有限,可通过延长堆放时间来降低其使用风险。

关键词: 牛粪垫料再生系统; 再生固体牛粪垫料; 高通量测序; 微生物多样性; 安全性评价

Abstract:

【Objective】 To evaluate the safety of recycled manure solids(RMS)treated by bedding recovery unit(BRU)in the hope of providing theoretical basis and reference for safe and reasonable application RMS on breeding farms. 【Methods】 Bacterial cultivation methods were used to detect the counts of the total bacteria, Escherichia coli, Salmonella, Staphylococcus aureus and parasitic eggs in RMS before and after the treatments by BRU. Besides, those detecting items in samples of the BRU treated by RMS that were stacked for a month were also determined and high-throughput sequencing method was used to detect the composition and diversity of pathogenic bacteria. 【Results】 After 16-18 h of aerobically fermentation, the counts of total bacterial and Escherichia coli in manure samples were decreased tenfold compared with fresh manure, but the counts of Staphylococcus aureus and parasitic eggs were not significantly changed (P>0.05). The counts of total bacterial number and Escherichia coli were significantly increased (P<0.05), but the counts of parasitic eggs were on the contrary (P<0.01). The pathogenic bacteria were only detected in manure samples untreated by BRU and they were divided into 4 genera: Aerococcus (1.40%±0.8%), Facklamia (20.73%±3.97%), Paeniclostridium (7.20%±2.35%) and Carnobacterium (0.27%±0.15%). Bacterial alpha-diversity indexes presented an significant decreasing trend after BRU treatment (P<0.01), mainly in terms of the bacterial OTU numbers, the bacterial richness (ACE index), and the Shannon-Weiner diversity index decreased 57.73%, 57.47% and 81.63%, respectively. While, the Shannon-Weiner diversity index of samples in RMS stacked for a month was significantly increased (21.76%) compared with the index in RMS treated by BRU. Furthermore, the bacterial beta-diversity analysis showed that the bacterial community structures were significantly changed after BRU treatment, and its communities were significantly increased in RMS stacked for a month (P<0.01). After 16-18 h of aerobically fermentation, the counts of total bacterial number, bacterial diversity and community structures were significantly changed. However, its killing effect on most pathogenic bacteria (apart from Escherichia coli) and parasitic eggs were still weak. 【Conclusion】 When RMS is treated with BRU system for 16-18 h, bovine manure can significantly reduce bacterial diversity and change community composition, but its killing effect on pathogenic bacteria and parasite eggs other than Escherichia coli is limited. Therefore, the risk can be reduced by prolonging stacking time.

Key words: bovine manure bedding recovery system; recycled manure solids; high-throughput sequencing; bacterial diversity; safety evaluation

中图分类号:

X713
S188

刘建成, 曾军, 丁峰, 许先查, 窦晶晶, 陈开旭, 李凤鸣, 高雁. 再生固体牛粪垫料中细菌多样性分析及评价[J]. 新疆农业科学, 2021, 58(12): 2334-2341.

LIU Jiancheng, ZENG Jun, DING Feng, XU Xiancha, DOU Jinjin, CHEN Kaixu, LI Fengmin, GAO Yan. Analysis of Bacterial Diversity and Safety Evaluation of the Recycled Manure Solids[J]. Xinjiang Agricultural Sciences, 2021, 58(12): 2334-2341.

图/表 4

参考文献 21

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检测指标 Detecting item	S₁ Sample 1	S₂ Sample 2	S₃ Sample 3
温度 Temperature(℃)	48.93±1.75^B	62.41±1.35^A	51.96±2.44^B
pH	7.29±0.32^a	7.16±0.14^a	7.45±0.03^a
水分 Moisture content(%)	51.72±2.18^a	48.91±1.46^a	43.01±1.92^b
干物质 Dry matter content(%)	48.62±2.60^a	51.19±1.94^a	56.43±0.71^b

检测指标 Detecting item	S₁ Sample 1	S₂ Sample 2	S₃ Sample 3
温度 Temperature(℃)	48.93±1.75^B	62.41±1.35^A	51.96±2.44^B
pH	7.29±0.32^a	7.16±0.14^a	7.45±0.03^a
水分 Moisture content(%)	51.72±2.18^a	48.91±1.46^a	43.01±1.92^b
干物质 Dry matter content(%)	48.62±2.60^a	51.19±1.94^a	56.43±0.71^b

检测指标 Testing index	S₁ Sample 1	S₂ Sample 2	S₃ Sample 3
细菌总数 Total number of bacteria (10⁷)	14.82±5.19^A	1.36±0.21^c	2.02±0.57^b
大肠杆菌 Escherichia coli Count (10⁵)	13.78±3.15^A	1.26±0.35^C	4.01±2.02^B
金黄色葡萄球菌 Staphylococcus aureus Count (10²)	5.14±2.83^a	3.98±2.20^a	2.24±0.94^b
无乳链球菌 Streptococcus agalactiae Count	ND	ND	ND
沙门氏菌 Salmonella Count	ND	ND	ND

检测指标 Testing index	S₁ Sample 1	S₂ Sample 2	S₃ Sample 3
细菌总数 Total number of bacteria (10⁷)	14.82±5.19^A	1.36±0.21^c	2.02±0.57^b
大肠杆菌 Escherichia coli Count (10⁵)	13.78±3.15^A	1.26±0.35^C	4.01±2.02^B
金黄色葡萄球菌 Staphylococcus aureus Count (10²)	5.14±2.83^a	3.98±2.20^a	2.24±0.94^b
无乳链球菌 Streptococcus agalactiae Count	ND	ND	ND
沙门氏菌 Salmonella Count	ND	ND	ND

样品 Samples	OTUs Operational Taxonomic Units	覆盖率 Coverage (%)	ACE丰度指数 Richness index of ACE	香农-威纳多样性指数 Shannon-Weiner diversity index	基于系统发育树多样性指数 Diversity index based on phylogenetic tree
S₁ Sample 1	149.00±8.00^A	91.50±0.70^a	302.44±34.72^A	5.06±0.28^A	14.78±1.76^A
S₂ Sample 2	60.00±9.00^B	96.60±0.80^a	128.60±34.62^B	1.93±0.11^C	6.88±0.83^B
S₃ Sample 3	40.00±9.00^C	98.4±0.3^a	60.01±9.51^C	2.35±0.34^B	3.65±0.51^C

再生固体牛粪垫料中细菌多样性分析及评价

Analysis of Bacterial Diversity and Safety Evaluation of the Recycled Manure Solids

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