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This study presents a new method for energy-efficient wastewater treatment that synergizes the partial-denitrification, anammox, and in-situ fermentation (SPDAF) processes in an up-flow reactor. Nitrate-containing wastewater and actual domestic sewage were fed into this SPDAF system, which was operated for 180 days without the addition of external carbon sources and aeration. The total inorganic nitrogen (TIN) removal efficiency reached 93.1% with a low C/N ratio of 1.6, a NO3 ––N/NH4 +–N ratio of 1.13 and a TIN concentration of 92.5 mg N/L. The contribution of anammox to nitrogen removal accounted for 95.6%. Batch tests demonstrated that the partial-denitrification process was able to use organics from either the influent or those produced by fermentation, thus providing nitrite for anammox. Significantly, fermentation played a key role in using the slowly biodegradable organics and provided adequate electron donor for partial-denitrification. Metagenomic sequencing analysis showed that the genera related to partial-denitrification, anammox, and fermentation bacteria were coexisted in this SPDAF system. The key functional genes of anammox bacteria (Hzs, 3986 hits; Hdh, 2804 hits) were highly detected in this study. The abundances of cytoplasmic nitrate reductase (58 706 hits) and periplasmic nitrate reductase (70 540 hits) were much higher than copper nitrite reductase (16 436 hits) and cytochrome cd1 nitrite reductase (14 264 hits), potentially contributing to the occurrence of partial-denitrification. Moreover, different abundances of genes involved in fermentation metabolism suggested that fermentation likely generated easily biodegradable organics for partial-denitrification.

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