Study On The Anammox Bacteria Enrichment And Organotrophic Characteristics Of Predominant Bacteria Jettenia Asiatica

Anaerobic ammonia-oxidizing bacteria(AAOB) is a important player in the global nitrogen cycle. Under anaerobic condition, ammonium is oxidized into nitrogen gas by the AAOB along with the nitrite reduction. ANAMMOX process has been successfully applied in treating ammonium-rich wastewater for a low demand of energy and cost. AAOB are autotrophic organisms using bicarbonate as sole carbon source for synthesis of cell biomass. However, recent studies bring insights into the possibility of organotrophic process by a few AAOB species, such as Candidatus Kuenenia stuttgartiensi 、 Candidatus Brocadia fulgidaa and Candidatus Anammoxoglobus propionicus. In this study, anammox consortia dominated by Candidatus Jettenia asiatica was enrichment by EGSB reactor and the effects of C2/C3 organic matters stress on anammox activity and AAOB growth were investigated. Stable isotopic tracing technology and nanometre-scale secondary ion mass spectrometry(Nano SIMS) were employed to identify, quantify and visualize the incorporation of 13C-labeled carbon sources and 15N-labeled nitrogen sources. Then, the possible metabolic pathway was speculated.An EGSB reactor was set-up by seeding the stored ANAMMOX sludge at HRT 5.5~3 h, 33±2 ℃ for enrichment AAOB with bicarbonate as carbon source. The average ammonium and nitrite removal rates were 92.13 %, 92.12 % respectively. The maximum nitrogen efficiency was 6.59 kg-N·m-3·d-1 and the maximum nitrogen removal efficiency was 5.84 kg N/(m3·d). The ANAMMOX enrichment were red and granular, and the granules were irregular and formed by small granules. The predominant bacteria in ANAMMOX enrichment were cocci, and the cell morphology were consistent with 0.8~1.2 μm diameter. Fluorescence in situ hybridization(FISH) analysis showed the relative abundance of AAOB was increased from 57.69 % to 83.32 %. The results of q PCR further testified the gene copies of AAOB was increased from 0.49 hzo copies/ 16 S total bacteria copies to 0.83 hzo copies/16 S total bacteria copies. 16 S r RNA gene-cloning analysis indicated the predominant bacteria in the ANAMMOX enrichment culture was Jettenia asiatica.Based on the anammox consortia dominated by Jettenia asiatica, the effects of low-molecular-weight organic matters on anammox activity and AAOB growth were investigated. Low acetate(≤ 30mg/L) and propionate(≤ 50mg/L) concentration did not significantly influence ammonium oxidation. Molecular results clearly indicated that ANAMMOX bacteria Jettenia asiatica cells are capable of growing at the present of low-concentration acetate or propionate with a low growth rate. Acetate at 240 mg /L and propionate 400 mg /L could cause the decrease of ammonium consumption rate(P < 0.001).Isotopic tracing technology and secondary ion mass spectrometry technology were employed to identify, quantify and visualize the incorporation of 13 C and 15 N in Jettenia asiatica cells. The results clearly showed that Jettenia asiatica cells were capable of metabolizing [2-13C]acetate and [1-13C]propionate after 15 d cultivation. The addition of acetate or propionate promoted HCO3- uptake of Jettenia asiatica cells. The 15 N uptake was much faster than the 13 C, with the peak of 15N-incorporation at 20 min of incubation. The addition of acetateor and propionate improved the incorporation of 15NH4+-N by Jettenia asiatica. The acetate or propionate was primarily metabolized as electron donor.