Isolation And Characterization Of A Novel Heterotrophic Nitrification-aerobic Denitrification Bacterium From Pufferfish Takifugu Rubripes RAS

In the recirculating aquaculture systems (RAS), where for the effective treatmentand safe recycle of wastewater is the key, the discharge water that after thepurification treatment will cycle to the culture pond. Biological treatment system asthe core unit of the wastewater treatment system plays a vital role in the RAS. Thenitrogen accumulating in the culture wastewater are the pollutants that the hazard isserious. Thus, its degradation and removal has become the emphasis and difficulty inthe biological treatment system. The aim of the thesis was to screen efficient nitrogendegrading bacteria that was expected to apply to the RAS treatment system. For thispurpose, with the Pufferfish Takifugu rubripes RAS as the research respect, this paperscreen a strain that have the ability of heterotrophic nitrification. At the same time, thepreliminary investigation of the microbe to the actual wastewater treatment effect wasconducted. The main conculusions were as follows:1. Through the primary and secondary screening, strain AD7was isolated fromthe sample of biofilm samplinged from the Pufferfish Takifugu rubripes RAS storedin lab. Strain AD7was identified as Kocuria polaris according to its morphologicalproperties, physiological and biochemical characteristics and the physiological andbiochemical characteristics and the phylogenetic analysis based on16S rRNA genesequence. So far, it was for the first report about Kocuria genus’s ability ofdenitrification.2. Kocuria polaris AD7is a bacteria with a certain heterotrophic nitrification. Bythe single factor experiment we determined the optimal conditions of strain AD7forits heterotrophic nitrification were as follows: glucose as carbon source, C/N5, initialpH8, culturing temperature30℃, initial ammonia nitrogen concentration60-90mg/L,shaking speed130r/min, initial inoculation concentration0.9%, salinity26.7%. Withthe aforementioned conditions, the ammonia nitrogen removal rate was57.33%(4d)and the maximum removal speed of ammonia nitrogen was1.26mg·N/（L·h）.3. Meanwhile, Kocuria polaris AD7also behaved certain aerobic denitrification capacity. The experimental results showed that strain AD7expressed splendid aerobicdeitrification ability when KNO3as the substrate, for the nitrate nitrogen removal ratereached80.70%, but accompanied by a large number of the accumulation of nitritenitrogen and relatively low actual denitrification rate.4. By means of the analysis of the treatment effect of the wastewater of Atalanticsalmon RAS by strain AD7, we didn’t monitored the bacteria growth after inoculation48h and the concentration of the water quality indators including ammonia nitrogen,nitrite nitrogen and nitrate nitrogen changed unconspicuous. The conclusion abovedmay be due to the type of carbon source and lower nitrogen concentration in theactual culture wastewater. The final results of experiment which was added additionalcarbon and nitrogen source showed that the cell growth increased with the increase ofammonia nitrogen concentration after48h incolution. Meanwhile, the removal rate ofammonia nitrogen went up along with very low accumulation of nitrite nitrogen(around0.1mg/L) and the nitrate nitrogen removal rate also has a rising trend. Thecell polulation OD600of strain AD7reached0.02after48h incolution when theconcentration of ammonia nitrogen increased to5mg/L and the ammonia nitrogenefficiency was close to40%. This results indicated that the feasibility that applyingstrain AD7to the biological treatment facilities which has higher pollutantconcentration is good.5. The conclusion of the experiment which inoculating strain AD7to artificialdomestic sewage showed that the cell population OD600of strain AD7reached0.27after48h and ammonia removal efficiency over50%. Visibly, applying strain AD7tosewage to remove ammonia nitrogen is feasible and effective.The study of this paper about strain AD7at the aspect of the heterotrophicnitrification–aerobic denitrification performance and the efficiency of the actualwastewater treatment provides the support of data and theory to put it into practicaluse in future.