Identification And Characterization Of The Alkalophilic Pseudomonas Stutzeri HK13 With High Denitrification Capacity

The biological denitrification of nitrifiers and denitrifiers has many advantages.Therefore,it is widely used in wastewater treatment systems.In recent years,the discharge standard of total nitrogen(TN)becomes increasingly strict.Ammonia nitrogen levels below the discharge standard but total nitrogen levels exceeding that due to high-concentration nitrate in effluent in many in wastewater treatment plants.Therefore,denitrifiers with high denitrification capacity are urgently required to strengthen the Biological Denitrification Process.In this study,an efficient alkaliophilic facultative anaerobic denitrifier HK13 was isolated from a coal chemical wastewater treatment plant.Based on morphological observations and 16S rRNA gene sequence analysis,strain HK13 was identified as Pseudomonas stutzeri,named as Pseudomonas stutzeri HK13 with the GenBank accession number of MN294682.Single factor experiments were operated using nitrate wastewater.The conclusions were as follows:(1)The optimum denitrification conditions of strain HK13 were:sodium citrate as the carbon source,C/N=8,initial pH at 7?10,rotation speed at 100 rpm and temperature at 30??40?.(2)The denitrification efficiency and rate of strain HK13 was notable.Under the optimum conditions with an initial TN concentration of 107.00 mg/L,the denitrification rate reached 20.03 mg/(L·h)between 9 h and 12 h,and the TN removal efficiency was above 98%at 15 h without nitrite accumulation in the system.(3)The suitable temperature and pH range for strain HK13 were 20?-40? and 7-10,respectively,suggesting that strain HK13 had a wide range of temperature and pH to adapt to the environment.The nitrogen removal abilities of strain HK13 in different wastewater were investigated.The results showed that:(1)When N02--N of 105.26 mg/L was used as sole nitrogen source,the NO2--N and TN removal efficiencies were 100%and 98.58%at 15 h with the denitrification rate of 21.77 mg/(L·h)between 9 h and 12 h.This result indicated that strain HK13 could degrade nitrite rapidly and efficiently.(2)When the concentration of initial NO3--N was 455.04 mg/L,the average NOx--N removal amount was more than 200 mg/L/day and the TN in the broth decreased to 9.5 mg/L at 56 h.These results suggested that strain HK13 could be used to treat wastewater with high concentrations of nitrate and nitrite.(3)Strain HK13 exhibited efficient denitrification ability with the NOx--N removal efficiency of 99.64%in real wastewater.The denitrification functional genes of strain HK13 were analyzed.The results showed that the strain had a complete enzyme genes system of denitrification(narG,nirS,norB and nosZ),and it was deduced that strain HK13 presented the metabolic pathway of NO3-?NO2-?NO?N2O?N2.