ppk1作为遗传标记解析污水脱氮除磷系统中Candidatus Accumulibacter的菌群结构和动态变化

Phosphorus (P) from wastewater is considered as a key factor causing surface water eutrophication, and thus enhanced biological phosphorus removal (EBPR) has been widely applied in many wastewater treatment plants. Candidatus Accumulibacter has been identified as a dominant polyphosphate accumulating organism (PAO) in EBPR activated sludge. Community structure and metabolic activity of Candidatus Accumulibacter has an important effect on the performance of EBPR. However, the relevance of different Accumulibacter clades to the performance of biological nutrients removal (BNR) from real wastewater is unknown. .In this study, the poly-P kinase 1 (ppk1) gene is used as phylogenetic marker to characterize Candidatus Accumulibacter populations in different BNR systems for real wastewater treatment. Four typical BNR systems are selected, that is, wastewater treatment plants, lab-scale conventional BNR systems, BNR via nitrite pathway and denitrifying phosphorus removal system. Clade-level population structure and transcriptional level gene expression of Candidatus Accumulibacter are analyzed by using molecular biological methods e.g. reverse transcription quantifying PCR (RT-qPCR), fluorescence in situ hybridization (FISH) + Flow-Cytometric Sorting and terminal-restriction fragment lengths polymorphism (T-RFLP) based on ppk1 gene, combined with traditional kinetic and stoichiometric studies. Dynamics of gene expression of ppk1, nitrate reductase gene (nar) and nitrite reductase genes (nirK and nirS) and clade-level population structure of Candidatus Accumulibacter corresponding to BNR performance will be established to reveal the major factors leading to unstability of EBPR performance..Furthermore, the effect of nitrite accumulation from nitritation on the clade-level population structure of Candidatus Accumulibacter and EBPR performance is also be investigated. An effective strategy will be studied to alleviate or eliminate the inhibition of nitrite buildup resulted from nitritation to metabolic activities of Candidatus Accumulibacter. Whether or not denitrifying phosphorus removal can be achieved under certain Candidatus Accumulibacter population structure and the different nitrifying mode will also be further studied. Based on the investigation above, important microbiological information is provided for BNR stable performance.

Candidatus Accumulibacter是污水生物除磷的优势聚磷菌，其菌群结构与代谢活性直接影响除磷效果，但Accumulibacter各进化枝与实际污水脱氮除磷系统运行的相关性仍然未知。.本研究以编码聚合磷酸盐激酶的功能基因ppk1作为遗传标记，对城市污水处理厂、实验室规模传统脱氮除磷、短程脱氮除磷和反硝化除磷系统中Accumulibacter进化枝水平的菌群结构、转录水平的关键功能基因表达进行分析。建立Accumulibacter 菌群ppk1，nirK，nirS和nar等基因表达及菌群结构对系统运行状态变化的动态响应机制。特别关注短程硝化引起的亚硝酸盐积累对Accumulibacter菌群结构和代谢活性的的影响，以及不同硝化模式下Candidatus Accumulibacter各进化枝的菌群构成和反硝化性能，为实际污水脱氮除磷系统的稳定运行奠定微生物学理论基础。

Candidatus Accumulibacter是污水生物除磷的优势聚磷菌，其菌群结构与代谢活性直接影响除磷效果。本课题以编码聚合磷酸盐激酶的功能基因ppk1作为遗传标记，对城市污水处理厂传统脱氮除磷、短程脱氮除磷和反硝化除磷系统中Accumulibacter进化枝水平的菌群结构、转录水平的关键功能基因表达进行分析。建立Accumulibacter 菌群ppk1，nirK，nirS和nar等基因表达及菌群结构对系统运行状态变化的动态响应机制。. 北京10座城市污水处理厂中都含有Accumulibacter的6个进化枝 （IA、IIA、IIB、IIC、IID、IIF）。其中A2O工艺中总Acc丰度最高，达到22.77%。IIC和IID是城市污水处理厂中的优势聚磷菌。游离亚硝酸（free nitrous acid, FNA）抑制使系统中的生物多样性显著降低。IIF分支对FNA抑制更为敏感，而IID分支具有更强的FNA耐受能力。首次报道了在处理实际生活污水的系统中以亚硝酸盐作为电子受体进行反硝化除磷的Acc-IID始终是优势聚磷菌，达到总Accumulibacter的92%以上，甚至接近100%，亚硝酸盐浓度是影响其丰度变化的重要因素。Acc-IID作为优势聚磷菌保证了亚硝酸型反硝化除磷的稳定运行。在全程硝化模式下的反硝化除磷系统中，Acc-IIC和IIF是优势聚磷菌，在以硝酸盐为电子受体的反硝化除磷过程中发挥重要作用。(nirS+nirK)/ppk1比值能够用来评价具有反硝化除磷功能的Acc在总Acc中所占的比例。. 关于短程硝化引起的亚硝酸盐积累对Accumulibacter菌群结构和代谢活性的的影响，以及不同硝化模式下Candidatus Accumulibacter各进化枝的菌群构成和反硝化性能，为实际污水脱氮除磷系统的稳定运行奠定微生物学理论基础。