The Preliminary Study On Gene Cloning,gas Production And Application In The Process Of Denitrification Of Pseudomonas Aeruginosa YY24 Strain

Nitrogen is a kind of important pollutants in water pollution,great harm to human health and the environment.The biological denitrification method is considered to be the most practical method for wastewater denitrification with its non-polluting and cost-effective characteristics.In recent years,many of the newly discovered aerobic denitrifying bacteria and heterotrophic denitrifying bacteria can achieve simultaneous nitrification and denitrification in the same aerobic tank,providing a new way for biological denitrification.In this study,the gene cloning of nitrification and denitrification was studied from the perspective of molecular biological level based on the identified Pseudomonas aeruginosa YY24,which was based on the biological denitrification in aquaculture water environment.Heterotrophic nitrification-aerobic denitrification in the process of control genes;The effects of different nitrogen sources on the growth,nitrogen removal ability and gas products of strain YY24 were investigated.The effects of strain YY24 on the growth and nitrogen removal capacity of strain YY24 were studied.First,nine kinds of nitrifying and denitrifying genes of Pseudomonas aeruginosa YY24 were amplified,identified and sequenced.Advances in gene cloning of isoxic nitrification-good denitrification in strain YY24 from the genetic level.The results showed that there were nitric oxide synthase gene type amo A and nitrite reductase gene type nar G,nitrite reductase gene type nir S,nitric oxide reductase gene type qnor B and nitrous oxide reductase gene type nos Z,which control nitrification in Pseudomonas aeruginosa YY24.According to the above analysis,we proposed the hypothetical pathway of NH₄⁺-N→NO₃--N→NO₂--N→NO→N2O→N2,and the key enzymes of control reaction were amo A,nar G,Nir S,qnor B and nos Z.The effects of different nitrogen sources on the simultaneous nitrification and denitrification and gas production of strain YY24 were investigated by using the gas generating device and the gas detector to analyze the gas species and concentration produced during the denitrification.The results showed that strain YY24 could efficiently denote nitrification and aerobic denitrification with a single nitrogen source.303.69 mg/L ammonium was reduced to 6.74 mg/L at 120 h and the removal rate was 97.78%,101.89 mg/L nitrate was reduced to 0.04 mg/L and the removal rate was 99.96%,60.72 mg/L nitrite was reduced to 16.59 mg/L and the removal rate was 72.68%.The three nitrogen sources are suitable for the growth of nitrogen as a strain YY24,with ammonium nitrogen as the best nitrogen source,the final product was N2.NO,N2 O was not detected.Pseudomonas aeruginosa YY24 is a strain with strong heterotrophic nitrification and aerobic denitrification.According to the strain YY24,the denitrification intermediate product of single nitrogen source is analyzed.The accumulation of NO₂--N was in the process of heterotrophic nitration and de-ammoniumation was the accumulation of NO₃--N,and there was no accumulation of NO₂--N in the process of aerobic denitrification.The effects of different ways and different concentrations of Pseudomonas aeruginosa YY24 on the growth and water quality of Litopenaeus vannamei were studied by adding Pseudomonas aeruginosa YY24 to Litopenaeus vannamei cultured water.The results showed that Pseudomonas aeruginosa YY24 had strong competitive ability,nitrogen removal function and activity,and did not show pathogenicity in Litopenaeus vannamei aquaculture water.Compared with the control group,the survival rate and the weight gain rate of each test group were significantly higher than those of the control group（P<0.05）.The control group could obviously purify the water quality,and the shrimp growth was the best in the same concentration In a single delivery group,where G group is optimal;The concentration of NH₄⁺-N、NO₃--N and NO₂--N in the test group reached the lowest value at 96 h,the nitrogen removal efficiency of strain YY24 was high and the time required for the reaction was shorter.The initial delivery of 106 cfu/ml concentration of strain YY24,96 h to remove ammonia nitrogen effect is better,and every other day to vote 104 cfu/ml concentration strain YY24 sustained effect is better.