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中文题名:

 有机肥矿化及其提高辣椒产量的微生物生态学机制    

姓名:

 刘超    

学号:

 2017203020    

保密级别:

 公开    

论文语种:

 chi    

学科代码:

 090302    

学科名称:

 农学 - 农业资源利用 - 植物营养学    

学生类型:

 博士    

学位:

 农学博士    

学校:

 南京农业大学    

院系:

 资源与环境科学学院    

专业:

 农业资源与环境    

研究方向:

 农业废弃物资源化的生物学过程    

第一导师姓名:

 沈标    

第一导师单位:

 南京农业大学    

完成日期:

 2021-06-30    

答辩日期:

 2021-09-04    

外文题名:

 The Mineralization of Organic Fertilizer and Its Microbiological Ecological Mechanism to Increase The Yield of Pepper    

中文关键词:

 有机肥料 ; 分解 ; 土壤微生物群落 ; 群落演替 ; 促生作用    

外文关键词:

 Organic fertilizer ; Decomposition ; Soil microbial community ; Community succession ; Growth promotion    

中文摘要:

我国是农业大国,每年产生的农业有机固体废弃物高达40亿吨,随着国家对环境保护和农业发展的重视,这些农业有机废弃物的资源化利用逐渐形成了“肥料化”、“饲料化”、“燃料化”、“食用菌基料化”和“工业原料化”的“农用为主的五料化”格局。其中,有机物料的肥料化是实现包括作物秸秆、畜禽粪便和菇渣在内的农业废弃物资源化利用的有效途径,既有利于保护环境,又可以有效改善土壤理化性状,优化土壤结构,提高土壤微生物活性,提高农作物产量。土壤微生物是农业生态系统有机质分解、养分循环过程中的核心要素,其生长繁殖得益于有机肥料的施用,微生物活性的增高反过来进一步推动了有机肥料的分解。

本论文运用原位分解试验、盆栽试验、田间试验方法,结合高通量测序技术等研究了1)不同纬度地域影响有机肥分解的生物和非生物因素;2)有机肥分解过程中微生物群落的演替规律及其优势微生物;3)有机肥分解对土壤微生物群落组成、功能、构建及其促生能力的影响;4)田间连续施用有机肥对土壤微生物群落组成和辣椒产量的影响。本文试图揭示有机肥分解的微生物学过程及其对土壤微生物群落的影响,阐明有机肥提高辣椒产量的微生物生态学机制。主要研究结果如下:

1.通过土壤原位分解试验,分别研究了有机肥在中国吉林、江苏和云南三个不同纬度地区的矿化特征及影响因素。结果表明,有机肥在三个不同地点的分解速率显著不同,云南的分解速率最高,江苏次之,吉林的最低;未灭菌有机肥的分解速率明显高于灭菌有机肥;三个试验点的土壤微生物多样性和群落结构存在显着差异,江苏试验点的土壤中细菌丰富度和多样性最高,云南试验点的土壤次之,吉林试验点的土壤最小,但云南试验点的土壤真菌丰富度最高,明显高于江苏和吉林试验点土壤,但三个试验点的土壤真菌多样性没有显著差异;梯度决策树(ABT)分析结果表明,土壤真菌丰富度和年平均温度(MAT)分别是影响有机肥分解的最重要的生物和非生物因素;年平均温度和年平均降水量(MAP)是影响真菌丰富度的重要环境因素;年平均温度与真菌丰富度显著正相关,年平均温度、年平均降水量和土壤真菌丰富度分别与有机肥的分解速率常数显著正相关;LEfSe差异分析结果表明,有机肥在吉林试验点土壤中显著富集了未定义腐生真菌和欧石南类菌根真菌,在云南试验点土壤中显著富集了外生菌根真菌、丛枝菌根真菌、地衣类真菌和附生植物的真菌;且外生菌根真菌、丛枝菌根真菌、地衣类真菌的相对丰度与有机肥分解速率常数显著正相关。

2.在有机肥的土壤原位分解试验中,研究了有机肥分解过程中微生物群落的演替特征和优势微生物种群。结果表明,地域因素、时间因素和处理因素(未灭菌和灭菌)及其交互效应均显著影响有机肥分解过程中的细菌丰富度和多样性;时间因素和地域因素显著改变了未灭菌和灭菌有机肥中的细菌群落结构,地域因素、时间因素及其交互效应对真菌群落丰富度也有显著影响,处理因素、时间因素及其交互效应对真菌群落多样性有显著影响,地域因素和时间因素的交互效应、地域因素和处理因素的交互效应以及三种因素的交互效应对真菌群落多样性有显著影响。

对不同处理和每个试验点的群落结构的进一步分析发现,处理因素(灭菌与否)显著改变了吉林和江苏试验点有机肥分解过程中的细菌群落结构,而时间因素和处理因素显著改变了云南试验点有机肥分解过程中的细菌群落结构。对真菌而言,时间因素和地域因素显著改变了未灭菌和灭菌有机肥分解过程中的真菌群落结构,处理因素(灭菌与否)显著改变了吉林试验点有机肥分解过程中的真菌群落结构,而时间因素和处理因素显著改变了云南试验点有机肥分解过程中的真菌群落结构;随机森林模型和相关性分析结果表明,TrueperaLewinellaThermoactinomycesWinogradskyellaErythromicrobiumSubsaxibacterHalieaMuricaudaChondromyces是三个试验点均与未灭菌肥料分解速率显著正相关的共有细菌属,TrueperaAnaerolinea是三个试验点均与灭菌肥料分解速率显著正相关的共有细菌属,同时也是吉林试验点与灭菌和未灭菌肥料分解速率显著正相关的共有细菌属。而江苏试验点和云南试验点只有Truepera是与灭菌和未灭菌肥料分解速率显著正相关的共有细菌属。对真菌而言,Mortierella是三个试验点中与未灭菌有机肥料分解速率显著正相关的共有真菌。

3. 在有机肥的土壤原位分解试验中,取不同时期的未灭菌有机肥网袋外围的土壤,研究了有机肥矿化对土壤微生物群落的影响,并通过室内辣椒盆栽实验,评估了有机肥不同矿化时期的外围土壤微生物群落的促生能力。结果表明,有机肥分解过程中,三个试验点有机肥网袋外围的土壤微生物群落的促生能力随着分解时间而逐渐减弱,均与分解时间显著负相关;方差分解结果表明,分解时间是影响辣椒鲜重的主要因子;土壤微生物群落结构PCoA分析结果显示,地域因素和时间因素显著影响了土壤细菌和真菌群落结构的变化;土壤pH和土壤有机质与土壤细菌和真菌群落组成显著相关;吉林试验点肥料网袋外围土壤中Gp1Thielavia的相对丰度与辣椒生物量显著正相关;江苏试验点肥料网袋外围土壤中AzohydromonasThermomonasMethyloversatilisPontibacterAscobolusMortierella的相对丰度与辣椒生物量显著正相关;云南试验点肥料网袋外围土壤中Gemmatimonas的相对丰度与辣椒生物量显著正相关;吉林试验点未灭菌肥料网袋外围土壤微生物群落主要通过影响细菌群落的丰富度、多样性和关键微生物类群来影响辣椒生长,江苏试验点未灭菌肥料网袋外围土壤微生物群落主要通过影响细菌群落结构和关键微生物类群来影响辣椒生长,而云南试验点未灭菌肥料网袋外围土壤微生物群落主要通过影响关键微生物类群来影响辣椒生长。

4. 利用田间试验研究了连续2年施用不同有机肥料对设施辣椒产量、土壤理化、土壤微生物群落组成的影响。结果表明,相比于施用化肥,施用有机肥改善了辣椒植株的农艺性状,显著提高了辣椒的株高、茎粗和产量;显著提高了土壤pH和有机质含量;显著改变了土壤微生物群落结构,增加了细菌群落的多样性;显著改变细菌群落门水平的组成,降低了变形菌门的相对丰度,提高了酸杆菌门的相对丰度,提高了土壤中GemmatimonasNitrolanccaConexibacter属的相对丰度,降低了GP6GaiellaMassiliaLysobacterGeodermatophilus属的相对丰度;Mantel检验和相关性分析分别表明土壤细菌群落、土壤细菌丰富度和多样性均与辣椒产量显著性正相关;相关性网络分析结果表明,施用有机肥富集的与辣椒产量显著正相关的微生物类群通过分解有机肥释放养分和提高根际有益菌丰度来促进辣椒增产。

综上所述,区域尺度下年平均气温和土壤真菌群落分别是影响有机肥分解的主要的环境和生物因素,虽然不同纬度地区的土壤微生物结构不同,但参与有机肥矿化的优势细菌和真菌类群有许多是共有的;有机肥的分解显著影响了土壤微生物群落结构,微生物群落对辣椒的促生能力随有机肥分解时间而逐渐减弱。连续施用有机肥料显著提高了土壤有机质,调控了土壤微生物群落,促进了辣椒增产。

外文摘要:

China, as a large agricultural country, produced about 4 billion tons of agricultural organic solid wastes annually. Since the government pays more attention to environmental protection and agricultural development, these organic waste has gradually formed the five major modes of resource utilization as "fertilization", "feed", "Fuelization", "Edible fungus substrateization" and "Industrial raw materialization". Among them, the fertilization of organic wastes is an effective way to realize the resource utilization of agricultural wastes including crop straws, livestock and poultry manures and mushroom residues. It is not only conducive to environmental protection but also can effectively improve soil physical and chemical properties and optimize soil structure. In addition, it can improve soil microbial activity and increase crop yields. Soil microorganisms are the core elements during the decomposition and nutrient cycling of organic fertilizer in agricultural ecosystems. Soil microbial growth and reproduction benefit from the application of organic fertilizer. The increase in microbial activity in turn further promotes the decomposition of organic fertilizer.

In this paper, in-situ decomposition experiments, pot experiments, field experiments and high-throughput sequencing were used to study the following four scientific issues. 1) Biological and non-biological factors that affected the decomposition of organic fertilizer in different regions. 2) The succession of the microbial community during the decomposition of organic fertilizer and the microbial driving mechanism of decomposition. 3) The effect of the organic fertilizer decomposition on the composition, function, construction and growth-promoting ability of the surrounding soil microbial community. 4) The effect of continuous application of organic fertilizer in the field on the composition of soil microbial community and pepper yield. This article attempts to reveal the biological process of organic fertilizer decomposition and its influence on the surrounding soil microbial community, and then clarify the microbial process of organic fertilizer decomposing in the soil and the microbiological ecological mechanism of increasing the yield of pepper. This will provide a theoretical basis for the rational and wide application of organic fertilizer and the development of bio-organic fertilizer. The main results are as follows:

1. In-situ decomposition experiments aimed at exploring the decomposition characteristics and influencing factors of organic fertilizers were carried out in Jilin, Jiangsu and Yunnan at three different latitudes. The results showed that organic composts had different decomposition rates in the three different sites, with the highest in Yunnan, followed in Jiangsu and Jilin. The decomposition rates of unsterilized organic composts were significantly greater than these of sterilized organic compost. The soil microbial diversity and community structure among the three sites are significantly different. The soil bacterial richness and diversity in Jiangsu was the highest, followed by Yunnan and Jilin. The soil fungal richness in Yunnan was the highest, which is significantly higher than that of Jiangsu and Jilin.  There was no significant difference in soil fungal diversity in three different sites. The results of Gradient Decision Tree (ABT) show that soil fungal richness and annual average temperature (MAT) were the most important biological and non-biological control factors affecting the decomposition of organic fertilizer while MAT and annual average precipitation (MAP) were the important environmental factors affecting soil fungal richness. The results of the correlation analysis showed that the MAT was significantly positively correlated with the soil fungal richness and the decomposition rate constant of organic fertilizer while MAP and soil fungal richness were significantly positively correlated with the decomposition rate constant of organic fertilizer. The results of the LEfSe difference analysis showed that unidentified saprophytes and heather fungi were significantly enriched in Jilin soil. Animal pathogenic fungi were significantly enriched in Jiangsu soil. Ectophytes, arbuscular mycorrhizas, lichen fungi and epiphytes plants were significantly enriched in Yunnan soil. The relative abundance of ectomycorrhizal, arbuscular mycorrhizal and lichen fungi was significantly positively correlated with the decomposition rate constant of organic fertilizer.

2. Through an in-situ decomposition experiment, we evaluated the succession characteristics of microbial communities during the decomposition of organic fertilizer. The results showed that location, decomposition time, treatment and their interaction effects had significant effects on the changes in bacterial richness and diversity. Location, decomposition time and their interaction effects had significant effects on the changes in fungal richness. Sterilization or not, decomposition time and their interaction effects had a significant impact on the change of fungal diversity. The interaction effect of location and decomposition time, the interaction effect of location and treatment, and the interaction effect of the three factors also had a significant impact on the change of fungal diversity. Time and location significantly changed the bacterial community structure in unsterilized and sterilized organic fertilizer. We further studied the community structure of different treatments and different experimental sites and found that sterilization or not significantly changed the bacterial community structure of organic fertilizer decomposed in Jilin and Jiangsu, while time and treatment significantly changed the bacterial community structure of organic fertilizer decomposed in Yunnan. Time and location significantly changed the fungal community structure in unsterilized and sterilized organic fertilizer. We further studied the community structure of different treatments and different experimental sites and found that sterilization or not significantly changed the fungal community structure of organic fertilizer decomposed in Jilin, while time and treatment significantly changed the fungal community structure of organic fertilizer decomposed in Jiangsu and Yunnan. The results of the random forest model and correlation analysis showed that among the OTUs that were significantly positively correlated with the decomposition rate of organic fertilizer, Truepera, Lewinella, Thermoactinomyces, Winogradskyella, Erythromicrobium, Haliea, Subsaxibacter, Muricauda and Chondromyces were the shared bacterial genera that were significantly positively correlated with the decomposition rate of unsterilized fertilizers in the three sites, Truepera and Anaeolinea were shared bacterial genera that were significantly positively correlated with the decomposition rate of sterilized fertilizers in the three sites. They were also bacterial genera shared by sterilized and unsterilized fertilizers in Jilin. Truepera was a shared bacterial genus that had a significant positive correlation between the decomposition rate of sterilized and unsterilized fertilizers in Jiangsu. It was also a shared bacterial genus shared by sterilized and non-sterilized fertilizers in the Yunnan experimental site. For fungi, Mortierella was a shared fungal genus that was significantly positively correlated with the decomposition rate of unsterilized organic fertilizers among the three sites.

3. Through indoor pot experiments, we took the microbial community in the surrounding soil of unsterilized organic fertilizer as the research object, and evaluated the influence of the decomposition of organic fertilizer on the succession and growth-promoting ability of the surrounding soil microbial community. The results showed that during the decomposition process of organic fertilizer, the growth-promoting ability of the surrounding soil microbial communities gradually weakened with the decomposition time, which were significantly negatively correlated with the decomposition time. The result of VPA showed that the decomposition time was the main factor affecting the fresh weight of pepper. The decomposition of organic fertilizer changed the soil microbial diversity. In addition to the significant increase in bacterial richness and diversity in Jiangsu over time, the bacterial richness and diversity in Jilin and Yunnan showed a trend of increasing firstly and then decreasing. The soil fungal richness of Yunnan declined firstly and then increased while the soil fungal richness in the other sites gradually declined. The soil fungal diversity among three sites gradually declined. The PCoA analysis results of the microbial community structure showed that the location and time significantly affected the changes of soil bacterial and fungal community structure. Soil pH and organic matter were significantly related to the composition of soil bacterial and fungal community. The relative abundance of Gp1 and Thielavia in the soil outside the unsterilized fertilizer in Jilin was significantly positively correlated with the pepper biomass. The relative abundances of Azohydromonas, Thermomonas, Methyloversatilis, Pontibacter, Ascobolus, and Mortierella in the surrounding soil of the unsterilized fertilizer in Jiangsu was significantly positively correlated with the pepper biomass. The relative abundance of Gemmatimonas in the soil surrounding the unsterilized fertilizer in Yunnan was significantly positively correlated with pepper biomass. The soil microbial community outsided the unsterilized organic fertilizer in Jilin mainly affected the growth of peppers by affecting bacterial richness, diversity and key microbial groups. The soil microbial community outsided the unsterilized organic fertilizer in Jiangsu mainly affected the growth of peppers by affecting the bacterial community structure and key microbial groups. The soil microbial communities outsided the unsterilized organic fertilizer in Yunnan mainly affected the growth of peppers by affecting key microbial groups.

4. Field experiments were used to study the effects of applying organic fertilizers for two consecutive years on pepper biomass, yield, soil physicoshemical properties, and microbial community composition. The results showed that compared with the application of chemical fertilizers, the application of organic fertilizer significantly increased plant height, stem diameter and yield of pepper, and soil pH and organic matter content. The application of organic fertilizer significantly changed the soil microbial community and increased the diversity of the bacterial community. The application of organic fertilizer significantly changed the composition of the microbial community, reduced the relative abundance of the Proteobacteria, while increased the relative abundance of Acidobacteria. The application of organic fertilizer increased the relative abundance of Gemmatimonas, Nitrolancca and Conexibacter while decreased the relative abundance of GP6, Gaiella, Massilia, Lysobacter, and Geodermatophilus. The result of Mantel test and correlation analysis showed that soil bacterial communities composition, bacterial richness and bacterial diversity were significantly positively correlated with pepper yield. The results of correlation network analysis showed that the microbial groups, which enriched in organic fertilizer treatments and significantly positively correlated with pepper yield, decomposed organic fertilizer to release nutrients and increased the relative abundance of beneficial bacteria in the rhizosphere of plant to increase the production of pepper.

In summary, the application of organic fertilizers at a regional scale should consider climatic factors and soil fungal communities. The dominant bacteria and fungal groups participated in the decomposition of organic fertilizer. The growth-promoting ability of the peripheral soil microbial community gradually weakens with the decomposition time of organic fertilizer. The application of organic fertilizers can significantly increase soil organic matter, regulate soil microbial communities, and promote pepper production.

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中图分类号:

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 2021-10-08    

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