Enrichment, Isolation And Ammonia Oxidation Capacity Analysis Of Ammonia Oxidizing Archaea In Urban Polluted Rivers

Water shortages caused by environmental pollution is a major problem today, and sewage recycling is an effective method to solve the problem. The most mature and widely used technology for sewage treatment is biological method both at home and abroad. Traditional concepts think that nitrification process was mainly performed by the ammonia oxidizing bacteria(AOB) in wastewater treatment, but the archaea discovered recent years also has the function of ammonia oxidation. Archaea was widely distributed and there are large numbers in the natural environment. Thus, ammonia oxidation mechanism of ammonia oxidizing archaea(AOA) was been studied, and using their numerical advantage to study the nitrogen removal efficiency of AOA.In this paper, water samples of Shenzhen Urban polluted river(Dasha river) were collected for AOA enrichment. A rapid enrichment method of AOA has been established through the medium selection and antibacterial programs selection. Changes of ammonia, nitrate and nitrite concentration were investigated and population of AOA was analyzed. Various separation methods were used in AOA preliminary separation from AOA enrichment broth. The ammonia oxidation performance of AOA was evaluated by single factor experiment, orthogonal design and the actual sewage application experiment.The best enrichment effect was obtained in medium A of the program which alternate antibiotic every month. Degradation rate of ammonia was 70%, AOA numbers reached 8.7×106 copies/ng DNA and remain stable after one month. Combination of five antibiotics has the best enrichment effect in medium B with the concentration of AOA reached 3.7×107 copies/ng DNA, and the most significant effect was obtained in samples B-1-b with a concentration of 7.3×107 copies/ng DNA. AOB was not detected at all enrichment samples after four months. The proportion of AOA accounted for 96.74% and 95.18% in samples A-1-a and B-2-b. Only one OTU was detected in Sample A-1-a and B-2-b, and AOA in A-1-a and B-2-b were identified as new spices.All colonies isolated by agar and gellan gum tablet were bacteria. The bacteria isolated from the agar plate were divided into two OTUs, and 14 OTUs from Gellan gum plate. Pure culture of AOA were not isolated from liquid medium, but symbiotic relationship was discovered between AOA and other bacteria.Results of single-factor experimental using AOA enrichment broth showed that the fastest ammonia degradation rate of 90% was obtained ender temperature 40 ℃ compared with 45 ℃, with AOA concentration reached 1.87×107 copies/ng DNA, ammonia degradation rate of 93.5% was obtained under p H 7.0 copared with p H 8.0, with AOA concentration reached 7.15×107 copies/ng DNA, in addition, concentration of AOA was relatively constant at 107-108 copies/ng DNA. Range analysis of orthogonal experiment results showed that R(temperature)> R(p H)> R(ammonia). 37 ℃, p H 7.0 and 1.5 m M ammonium was the optimal growth conditions of AOA and also the best ammonia oxidation conditions of AOA.While studied AOA ammonia oxidation performance in synthetic wastewater, city sewage and river sewage, similar nitrogen removal effect was found in control and experimental group, which indicated that AOB plays a dominant role in ammonia degrading rather than the AOA.