【作者】 毛小芳；

【导师】 胡锋；

【作者基本信息】 南京农业大学 ， 生态学， 2006， 博士

【摘要】 食细菌线虫是土壤中重要的食细菌微型动物，由于其在土壤中数量多，代谢活性高，与微生物关系密切，关于食细菌线虫与微生物的相互作用及其生态效应的研究引起了生态学家普遍的关注。尽管研究结果不尽相同，但总体趋势都揭示食细菌线虫对细菌的取食所产生的直接或间接作用促进了土壤氮素矿化及氮素养分供应，促进了多数植物的生长和氮素营养状况改善。但是食细菌线虫与细菌之间的相互作用必然也会受到线虫种类、密度，细菌种类及环境因素等的影响。并且基于原生动物对植物生长促进作用的激素效应的研究，由于食细菌线虫在生理及生态上与原生动物极相似，并且在土壤中其数量和原生动物相当甚至超过原生动物，因此有理由认为食细菌线虫对植物生长的促进作用机理也可能存在线虫作用下的激素效应。本文在实验室已有工作基础上，首先采用悉生培养系统，并在设置了不同线虫牧食强度和不同湿度条件下，研究模式食细菌线虫（C.elegans）与细菌（B.subtilis）的相互作用及其对土壤氮素矿化的影响；其次，在自然土培条件下，研究供试土壤土著食细菌线虫及模式线虫（C.elegans）对番茄（Lycopersicon esculentum Mill）和小麦（宁麦9号）根系生长的影响，并初步探讨其中的激素作用及同时对土壤微生物群落结构的影响。旨在进一步阐明土壤食细菌线虫与细菌的相互作用及其对植物生长的促进作用机理。研究结果表明：1．土壤食细菌线虫与细菌的相互作用对细菌数量的影响有别于一般的牧食关系，食细菌线虫的取食促进了细菌的繁殖，并且能够促进细菌的活性。土壤食细菌线虫与细菌的相互作用进一步促进了土壤氮素的矿化，尤其是铵态氮和矿质氮含量的提高，但是，在悉生培养条件下，土壤硝态氮表现出强烈的降低趋势。2．食细菌线虫对细菌的促进作用存在“适度牧食”密度，即过度牧食的情况下，线虫对细菌的促进作用下降，同时线虫的增长幅度也下降。在试验设置的三个线虫取食密度下，线虫对细菌的促进作用表现为接种20条线虫·g^-1土处理＞10条线虫·g^-1土＞40条线虫·g^-1土。并且，线虫对细菌过度牧食在相对降低细菌数量的同时，对土壤氮素的矿化作用也相应下降。不同取食密度处理间，线虫对土壤氮素矿化的促进作用程度与对细菌的增殖促进作用程度趋势一致。3．食细菌线虫与细菌之间的相互作用受土壤水分含量的影响，在试验设置的含水量条件下，线虫对细菌的增殖促进作用总体表现为23％含水量处理＞17％含水量处理＞28％含水量处理，并且对土壤氮素矿化的促进作用也表现为23％含水量处理高于17％和28％两个含水量处理，而后两个含水量处理之间没有明显差异。因为线虫是水生动物，在土壤中需要水膜来生存和运动。但是，线虫在薄的水膜中的迁移活动比在自由水中要快，而线虫在不同含水量条件下迁移、分布和活动的改变也就影响了对细菌生物量周转以及土壤养分的作用。4．通过添加基质（猪粪和稻草）培养获得食细菌线虫的大量繁殖，并根据线虫虫体大小的特点采用两种孔径的网袋将内层添加基质土壤和外层未加基质土壤区分，从而内层大量繁殖的线虫通过1mm网袋迁移至外层未加基质的土壤，而采用5μm网袋则限制了线虫向外层土壤的迁移，从而获得土著食细菌线虫大量富集的外层土壤及对照土壤。并且1mm网袋外层主要是Protorhabditis.sp线虫的增加。添加两种基质的5μm网袋处理，外层土壤线虫数和空白处理差异不大。添加猪粪处理对食细菌线虫的繁殖作用优于添加稻草处理，这主要可能是由于猪粪的C：N比比较小，分解较快，线虫的繁殖，尤其是食细菌线虫的繁殖，和有机物料的分解速度直接相关。5．无论是直接原位富化获得土著食细菌线虫的大量繁殖，还是接种外源食细菌线虫，都促进了植物（番茄和小麦）的根系生长，主要是促进了根系形态的改变，即根系发育成了更长、更细、有更多分支的根系系统。6．食细菌线虫富集处理与对照处理相比，在种植番茄和小麦的土壤中IAA和GA₃的含量都显著提高。将食细菌线虫各处理作总平均值的分析表明，在种植番茄的土壤中，食细菌线虫使IAA和GA₃含量分别提高了72.7％和71.4％；种植小麦的土壤中，食细菌线虫分别使IAA和GA₃含量提高了90.7％和41％。通过Biolog的测定考察食细菌线虫对种植植物土壤微生物（细菌）群落的影响表明食细菌线虫的存在促进了土壤微生物群落的总体活性（AWCD值表示），并显著地改变了土壤微生物群落结构：接种食细菌线虫处理，土壤微生物对碳源类群的利用发生了显著的改变。而土壤中激素含量的提高主要应该是由于线虫的选择性取食对土壤微生物群落结构的改变。将这些结果与食细菌线虫对植物根系生长的促进作用相结合，在一定程度上说明了食细菌线虫同原生动物一样，存在对植物根系生长促进作用的激素效应。7．食细菌线虫对微生物群落结构的影响以及对植物根系的促进作用大小受线虫种类的影响，试验土壤富化的土著食细菌线虫对植物根系生长的促进作用优于实验室培养基上培养获得的外源食细菌线虫C.elegans，对土壤微生物群落的总体活性（AWCD值表示）的促进作用也表现为前者大于后者，并且两个处理微生物群落对碳源类群的利用也不同。而食细菌线虫与微生物和根系之间的相互作用同样受植物种类的影响，因为不同植物其根系分泌物不同，对微生物也会产生不同的影响。反映了植物根际土壤动物-微生物-根系之间复杂的相互作用。本研究进一步明确了土壤食细菌线虫与细菌之间相互作用及其对土壤氮素矿化的影响因素，并初步探讨了食细菌线虫对植物根系生长的促进作用及其基于改变微生物群落结构促进有益微生物生长的激素机理。本研究可为今后有效开发利用土壤有益线虫资源改善土壤养分管理、提高作物生产提供理论依据和潜在的技术储备，有较重要的理论意义和一定的应用前景。更多还原

【Abstract】 Bacterial-feeding nematodes is one of the most important bacterial grazer in soil.Interactions between bacterial-feeding nematodes and microbe and their ecological effectsin terrestrial ecosystems have caused great attention of ecologists due to their high biomassand turnover and the close interactions with microbe. Despite the results were notconsistent up to now, mostly researches found that the direct or indirect effects due tobacterial-feeding nematodes grazing on bacteria increased the nitrogen mineralization andimproved the status of the supply of inorganic nitrogen, so subsequently the plant growthwas stimulated. But this effect must be affected by the species and density of nematode andbacteria and the surrounding environmental condition, such as the humidity. Researchesabout protozoa found that there were hormonal effects on plant growth by protozoangrazing changing the rhizosphere bacterial community to stimulate plant growth-promotingrhizobacteria. Considering the similarly of the physiology and zoology betweenbacterial-feeding nematodes and protozoa, and that the bacterial-feeding nematodes occurat equal or greater biomass in the rhizosphere than protozoa, it is believed that the activityof bacterial-feeding nematodes in the rhizosphere will also stimulate root proliferation byhormonal effects as seen for protozoan grazing.On the base of previous experiments in our lab, the gnotobiotic microcosmexperiments were conducted to study the interactions between bacterial-feeding nematode（Caenorhabditis elegans） and bacteria （Bacillus subtilis） and their effects on soil nitrogenmineralization at different nematode density or different soil moisture content. Then,experiments with natural soil （soil didn’t sterile but directly incubated for enriched in soilnative nematodes or inoculated with nematodes） were manipulated to determine the effectsof bacterial-feeding nematodes on the root development of tomato and wheat seedlings. Thehormonal effect by nematode grazing changing the microbiall community was also discussed.The main results were as follows:1. Interactions between bacterial-feeding nematodes and bacteria were different fromthose between other hunters and preys. The grazing of nematodes significantly stimulatedthe growth and activity of the bacteria. Furthermore, the activity of nematodes mainlyenhanced soil nitrogen mineralization, especially remarkably increased the soil NH₄⁺-N andmineral N. But the NO₃^--N was significantly decreased with the inoculation ofbacterial-feeding nematodes in the experiments.2. The effect of bacterial-feeding nematodes on bacteria growth was exhibited a"moderate density" of nematodes, which means when the number of nematodes was morethan this density, the increment of the nematodes on bacteria was decreased. With thetreatments designed in the experiment, the effect of nematodes on bacteria was in the orderof 20 nematodes·g^-1 dry soil＞10 nematodes·g^-1 dry soil＞40 nematodes·g^-1 dry soil. At thesame time, the enhancement of nematodes on soil nitrogen mineralization was alsodecreased in the excessive grazing of nematodes on bacteria, and the increment ofnematodes on nitrogen mineralization was also in the same order of nematode densities asaffected on bacterial population in the experiment.3. Soil moisture content affected the interactions between bacterial-feeding nematodesand bacteria. The stimulated effect of nematodes on bacteria growth was different with thesoil moisture content in the order of 23% water content （w/w） treatment＞17% watercontent （w/w） treatment＞28% water content （w/w） treatment as designed in the experiment.The increment of nematodes on soil nitrogen mineralization was also greater in the 23%water content （w/w） treatment than the other two treatments, while there were nosignificantly difference between those two. These results indicated that at the lower soilmoisture content, effects of nematodes on bacteria growth and soil nitrogen mineralizationwere enhanced with the increasing of soil moisture content. But these effects weredecreased in the soil with too higher moisture content. Nematodes are aquatic and theirsurvival and movement need water films, and they can move quickly in thin water film than in free water. The difference of movement, distribution and activity of nematodes atdifferent soil moisture contents will affect their effects on bacteria growth and soil nutrientmineralization.4. Soil mixed with either pig manure or rice straw was placed in a mesh bag （1mm or5μm）, and then surrounded by an outer layer of unamended soil. The greater populations ofbacterial-feeding nematodes that generated in the soil mixed with pig manure or rice strawmigrated through the 1 mm diameter mesh bag into the outer soil, thus giving greaterpopulations than in soil surrounding the control treatment of 5μm diameter mesh bag,through which nematodes cannot migrate. The increased bacterial-feeding nematodes wereprimarily Protorhabditis. sp. There were more nematodes in the pig manure treatment thanin the rice straw treatment. This is probably related to the more rapid bacterialdecomposition of the pig manure, as reflected in its lower C: N ratio. The increase innematode numbers, especially bacterial-feeders, is directly related to the rate ofdecomposition of different organic amendments.5. Tomato and wheat seedlings grown in the soils containing more bacterial-feedingnematodes （no matter by stimulating the native population or by adding additionalnematodes） developed a highly branched root system with longer and thinner roots.6. The contents of IAA and GA₃ were significantly enhanced in the soils containing morebacterial-feeding nematodes compared with the control soils, no matter planted with tomatoor wheat. The mean values of all nematode treatments showed that in the presence of morebacterial-feeding nematodes, IAA and GA₃ increased by 72.7% and 71.4% respectively inthe soils planted with tomato, while in the soils planted with wheat, increased by 90.7% and41% respectively. Bacterial-feeding nematodes also stimulated the total microbial activity（indicated by AWCD） and significantly changed the soil microbial community structure thatthe soil microbe’s functions shifted evidently based on the sole carbon use pattern derivedform Biolog method. The increment of IAA and GA₃ in the soils should be caused by thechanges of soil microbial community structure due to the selective grazing of nematodes.Combined these results with the effects of bacterial-feeding nematodes on plant root proliferation, in some extent illustrated that bacterial-feeding nematodes affect plant rootgrowth likely through the hormonal effects by a grazing-induced change of soil microbialcommunity as found in the presence of protozoa.7. Bacterial-feeding nematode’s effects on soil microbial community and plant rootgrowth were affected by nematode species. Tomato and wheat seedlings grown in the soilscontaining more native bacterial-feeding nematodes developed a more highly branched rootsystem than those grown in the soils inoculated with C.elegans. The microbial activity wasalso higher in the soil with more native nematodes than with C.elegans and the soilmicrobe’s use pattern of the sole carbon was different in the presence of these twonematodes species. While the plant species also affected the interactions amongbacterial-feeding nematodes, microbe and plant root, for different plant will affect the soilmicrobe by releasing different root exudates. That the interactions among soil microfauna,microbe and root were complex.To summarize, the research farther discussed the factors affected the interactionsbetween bacterial-feeding nematodes and bacteria and their effects on soil nitrogenmineralization, and probed into bacterial-feeding nematode’s effects on plant root growththrough the hormonal effects by grazing-induced changes of soil microbial community.These findings supported theoretic knowledge and technic to host soil nematodes as animportant bio-resource in soil nutrient management and plant growth. The present studymay be valid not only for theory, but also for future application in agriculture.更多还原