土壤环境科学实验室知识库

      土壤动物的肠道微生物是土壤中隐藏的微生物组，在土壤生态过程中发挥着重要作用。 同时，其肠道微生物可能是抗生素抗性基因的潜在储存库，并在其迁
移 或者 被捕食过程造成抗生素抗性基因的扩散。目前土壤动物肠道微生物的研究
较为薄弱，特别是其携带的抗生素抗性基因情况尚不明确。
       本研究选择可能带来潜在环境污染的有机肥（禽畜粪便和污泥）作为研究典
型 土壤动物的肠道微生物及其抗性基因的切入点，通过有机肥施用野外长期定位
实验，采用高通量测序和高通量荧光定量 PCR，揭示有机肥施用下土壤性质和污
染物对典型土壤动物肠道微生物及其抗生素抗性基因的影响机制。同时利用室内
模拟实验验证野外实验得到的敏感因子对典型土壤动物肠道微生物组和抗性组的
作用机制。并 验证 有机肥资源化（生物炭）方式 是否可以减少有机肥对肠道微生物携带抗性基因的作用效果。 本研究的结果不仅对了解土壤动物的肠道微生物组成、阐明其所携带抗生素 抗性基因等基础 科学问题具有科学意义，而且为控制有机肥施用 造成的环境问题提供理论支持。主要研究结果包括：
（1）跳虫 微生物的群落结构不同于土壤，并且其多样性低于土壤。跳虫存在核心微生物 组，并且核心微生物组群落组成是一个非随机过程。施肥特别是有机肥 施 用可以改变跳虫的微生物组的群落组成。施肥造成 土壤微生物的变化在上述过程中发挥重要作用。
（2）蚯蚓肠道是抗生素抗性基因的重要储存库，有机肥施用造成了蚯蚓肠道中抗生素抗性基因的富集。肠道微生物的群落组成 、环境因子（土壤和肠道内容物性质）以及可移动遗传元件三个 因子 共 同驱动了蚯蚓肠道中抗性基因的变。
（3）重金属可以导致土壤动物肠道中抗性基因的共选择作用 ，并且抗生素和重 金属 对肠道抗性基因富集产生协同作用。 肠道微生物的 群落组成以及可移动遗传元件与抗生素抗性基因的组成密切相关。
（4 )六 种生物炭的施用可 以 改变土壤和肠道抗性基因的组成， 但并未显著影响土壤和肠道抗性基因的多样性和丰度，仅猪粪生物炭的施用造成肠道抗性基因的富集。猪粪生物炭的高浓度金属含量可能是造成抗性基因富集的主要因素，生物炭性质决定了其对肠道抗性组的影响方式。

      The soil fauna gut microbiota was an unexplored microbiome in soil and played crucial roles in soil ecological process. Meanwhile, the gut microbiota of soil fauna might be a reservoir of antibiotic resistance genes (ARGs), which could rapidly spread in the course of the migration and predation process of soil fauna. However, current research on soil fauna gut microbiota was still limited, especially the antibiotic resistance genes they carried were not unequivocal. In this study, organic fertilizer (animal manure and sewage sludge), which may bring potential environmental problem due to their application, was selected as the pointcut to study microbiota and ARGs in the gut of typical soil fauna. High throughput sequencing and high throughput fluorescence quantitative PCR were used to reveal the effect of soil property and contaminants on the soil fauna gut microbiota and ARGs in the long term field experiments of soil organic fertilizer application, and to verify the effects of these sensitive factors which were obtained in the field experiment on the gut associated ARGs in the indoor simulation experiment. Additionally, whether the reclamation of organic fertilizer (biochar) can reduce the enrichment of organic fertilizer on intestinal ARGs were also tested in this study. The results of these will not only be scientifically significant in understanding the composition of soil fauna gut microbiota and illuminating the ARGs that soil fauna gut microbiota carried, but also provide theoretical support to control the environmental problems induced by the soil organic fertilizer application. The main results were as follows:
(1) The collembolan bacterial communities differed significantly from their surrounding soil and their diversity was lower than that of the soil  icrobial community. A stable core microbiome existed in the collembolan ssociated--microbiota, and it was assembled non--randomly. Soil fertilization and in particularly application of inorganic fertilizer altered the bacterial community of the collembolan associated microbiota. Changes in the soil microbial community played an important role in the shift of in bacterial of collembolan associated microbiota.
(2) The gut microbiota of earthworms was a big reservoir of ARGs in the environment. Organic fertilization significantly increased the diversity and abundance of the gut associated ARGs. The gut microbial community, combined with environment factors (soil and gut content properties) shaped the ARG compositions in the earthworm gut.
(3) Heavy metals could lead to the co--selection of ARGs in collembolan gut, and heavy metal and antibiotic could induce a strong synergistic effect on the gut associated ARGs. The gut microbiota and mobile genetic elements (MGEs) were significantly correlated with ARG composition.
(4) The application of six types of biochar altered the profile of ARGs in collembolan gut, but did not significantly affect the diversity and abundance of soil and gut associated ARGs. However, only manure--derived biochar increased the relative abundance of gut--associated ARGs, and the high concentration of heavy metals in manure biochar might be the main driver of these enrichments. Biochar induced changes of the gut associated ARG compositions in collembolans were dependent on biochar quality.