The stages of formation and development of activated sludge biofilm carrying out the anammox process in a laboratory sequencing batch reactor (SBR) with complete biomass retention on the carrier were investigated using light, epifluorescence, and electron microscopy. Light microscopy revealed biofilm formation on the carrier to occur within one week. Rod-shaped and filamentous microorganisms were the first to attach on the carrier and acted as a skeleton for biofilm formation, thus playing an important role in colonization of the carrier. Epifluorescence microscopy revealed physiologically active anammox bacteria of the genera Candidatus “Brocadia” and Ca. “Jettenia” in the biofilms. Selective autofluorescence of the colonies of anammox bacteria Ca. “Jettenia” was observed. This autofluorescence was probably caused by specific proteins in the extracellular polymeric substances (EPS), their synthesis and/or amount depending on the colony age and the physiological state of the cells. Structural organization of the colonies of anammox bacteria was investigated by electron microscopy. High-throughput sequencing of the 16S rRNA gene fragments revealed the presence of sequences affiliated with members of the phyla Chloroflexi, Bacteroidetes, Planctomycetes, and Proteobacteria in the biofilms. Apart from anammox bacteria, anaerobic and facultatively anaerobic organotrophs, stage I nitrifiers, denitrifiers, and sulfate reducers were detected. Members of the genus Ca. “Brocadia” were predominant among the anammox bacteria, probably due to better adhesion of their cells to the carrier or to their competitive advantage over Ca. “Jettenia” in the presence of organic acids (acetate and formate) in the medium.

中文翻译：

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通过光、落射荧光和电子显微镜研究厌氧氨氧化生物膜的形成和发展

使用光、落射荧光和电子显微镜研究了在实验室序批式反应器 (SBR) 中进行厌氧氨氧化过程的活性污泥生物膜的形成和发展阶段，其中生物量完全保留在载体上。光学显微镜显示载体上的生物膜形成在一周内发生。杆状和丝状微生物最先附着在载体上，作为生物膜形成的骨架，在载体的定殖中发挥着重要作用。落射荧光显微镜显示具有生理活性的厌氧氨氧化菌属 Candidatus “Brocadia” 和 Ca。生物膜中的“Jettenia”。厌氧氨氧化细菌 Ca 菌落的选择性自发荧光。观察到“Jettenia”。这种自发荧光可能是由细胞外聚合物 (EPS) 中的特定蛋白质引起的，它们的合成和/或数量取决于集落年龄和细胞的生理状态。通过电子显微镜研究厌氧氨氧化细菌菌落的结构组织。16S rRNA 基因片段的高通量测序揭示了生物膜中存在与 Chloroflexi、Bacteroidetes、Planctomycetes 和 Proteobacteria 成员相关的序列。除了厌氧菌外，还检测到厌氧和兼性厌氧有机营养菌、I 阶段硝化菌、反硝化菌和硫酸盐还原菌。Ca 属的成员。“Brocadia”在厌氧氨氧化细菌中占主导地位，可能是因为它们的细胞更好地粘附在载体上，或者它们比 Ca 具有竞争优势。