两种生物除磷系统活性污泥中除磷菌的甄别——淀粉-缺氧/好氧交替与乙酸盐-厌氧/好氧交替系统

为了直接识别出污泥中的聚磷细菌和其种属,本研究采用4',6-二脒基-2-苯基吲哚(DAPI)染色和流式细胞荧光分选技术(FACS)对以淀粉为唯一碳源的缺氧/好氧序批式活性污泥(SBR)系统(R1)的缺氧末期和好氧末期以及以乙酸盐为唯一碳源的厌氧/好氧SBR系统(R2)的好氧末期污泥的聚磷细菌进行了原位分选,并通过16S rRNA高通量测序技术鉴定了分选后细菌的种属.结果表明,在R1中,缺氧期和好氧期均进行生物除磷,且缺氧期吸磷量大于好氧期. R2中发生着厌氧期释磷、好氧期大量吸磷的传统生物除磷.利用FACS在R1和R2污泥中均分选得到10⁶个相对纯度为85%的具有聚磷颗粒的细菌.测序结果表明,在R1系统中,缺氧段优势的聚磷菌属为Halomonas(37.75%)、unclassified Brucellaceae(14.15%)、Pseudomonas(6.49%)、unclassified Chlamydiales(0.027%)和Sphingopyxis(0.007%);好氧段优势聚磷菌属为Halomonas(19.72%)、unclassified Brucellaceae(14.62%)、Pseudomonas(14.28%)、unclassified Comamonadaceae(0.046%)、unclassified Acidobacteria Gp3(0.036%)和Ferruginibacter(0.026%).R1系统中unclassified Chlamydiales和Sphingopyxis仅仅在缺氧条件下具有聚磷功能,而unclassified Comamonadaceae、unclassified Acidobacteria Gp3和Ferruginibacter仅在好氧条件下才具有聚磷功能.在R2系统中,优势聚磷菌群为Dechloromonas(11.06%)、unclassified Anaerolineaceae(9.29%)、unclassified Bacteroidetes(7.44%)、unclassified Gammaproteobacteria(7.34%)以及Acinetobacter(0.31%).这意味着在新型的除磷系统(R1)中,参与除磷过程的细菌包括好氧,缺氧和兼性缺氧聚磷细菌,而在传统的除磷系统(R2)中,参与除磷过程的细菌仅为好氧聚磷细菌.

Screening of phosphorus removing bacteria from activated sludge for biological phosphorus removal: starch-anoxic/aerobic alternation and acetate-anaerobic/aerobic alternation system

In order to identify phosphorus accumulating organisms(PAOs) in situ from activated sludge. 4', 6-diamidino-2- phenylindole (DAPI) staining and flow cytometry fluorescence sorting (FACS) were used to sceen the PAOs from the two sludge at the anoxic/aerobic SBR system (R1) with starch as the only carbon source and the anaerobic/aerobic system (R2) with acetate as the only carbon source. The species of the sorted bacteria were identified by 16S rRNA high-throughput sequencing. The results showed that in the R1, biological phosphorus removal was carried out in both the anoxic periods and aerobic periods. The phosphorus uptake in the anoxic period was greater than that in the aerobic period. In the R2, the phosphorus release in the anaerobic period and a large amount of phosphorus absorption in the aerobic period occurs. Results of the in situ fluorescence staining showed that 10⁶ bacteria with a relative purity of 85% with phosphorus-accumlating cell were sorted from the R1 and the R2. The sequencing results showed that in the R1, the dominant genera of PAOs in the anoxic periods were Halomonas (37.75%), unclassified Brucellaceae (14.15%), Pseudomonas (6.49%), unclassified Chlamydiales (0.027%) and Sphingopyxis (0.007%). The dominant PAOs in the aerobic periods were Halomonas (19.72%), unclassified Brucellaceae (14.62%), Pseudomonas (14.28%), unclassified Comamonadaceae (0.046%), unclassified Acidobacteria Gp3 (0.036%) and Ferruginibacter (0.026%). In the R1, unclassified Chlamydiales and Sphingopyxis only had phosphorus accumulation function under anoxic periods, while unclassified Comamonadaceae, unclassified Acidobacteria Gp3 and Ferruginibacter only had phosphorus accumulation function under aerobic periods. In the R2, the dominant PAOs were Dechloromonas (11.06%), unclassified Anaerolineaceae (9.29%), unclassified Bacteroidetes (7.44%), unclassified Gammaproteobacteria (7.34%) and Acinetobacter (0.31%). This means that in the R1, the bacteria involved in the phosphorus removal process include aerobic, anoxic and both aerobic and anoxic phosphorate accumulating bacteria three types. In the R2, the bacteria involved in the phosphorus removal process are only aerobic phosphorus accumulating bacteria.