Screening Of Denitrifying Bacterias And Their Effects For The Remediation Of The Environment Of Aquaculture Ponds

In recent years,aquaculture industry in China has developed rapidly.With the increase of intensive and high-density aquaculture,nitrogen pollution is gradually increasing.Nitrogen is one of the main pollutants causing the deterioration of aquaculture water.It not only causes serious harm to aquatic organisms?fish,shrimp and crab?,but also causes serious environmental problems?red tide,et al.?,and even affects the health of human through bioconcentration.Biology method is one of the most economic and effective ways for the remediation of aquaculture waste at present.Nitrifying bacteria play an important role in the process of biological nitrogen removal.Three highly efficient heterotrophic nitrifying bacteria were isolated from the water of tilapia?Oreochromis spp?recirculation aquaculture systems.They were classified and identified,and their nitrification properties were studied then.Those three strains were used to remediate the water and sediment of tilapia ponds.Their effects on the pond environment quality were studied.The main findings are as follows:1.Isolation and denitrification performance of a heterophic nitrifying bacterium P5-2A highly efficient heterotrophic nitrifying bacterium P5-2 was isolated from the water of recirculation aquaculture systems.The strain was identified as Bacillus megaterium by morphological observation,physiological and biochemical reaction identification and 16S rRNA gene sequence analysis.The results of Gram staining showed that the strain was Gram-positive.The result of determination of extracellular enzyme showed that starch and casein could be utilized by the strain.When temperature was 30°C,salinity was 10‰,and pH was 7,the strain grew fastest,and first reached the logarithmic growth period and the stable period.The results of nitrification experiments showed that environmental factors such as carbon source,carbon-nitrogen ratio and dissolved oxygen concentration could influence its rate of ammonia nitrogen removal.When sodium citrate was used as carbon source,carbon-nitrogen ratio was 15,pH was 7,and rotational speed was 180 rpm,the highest ammonia nitrogen removal rate of 99.07%was achieved after 24 hours.2.Isolation and denitrification performance of a heterophic nitrifying bacterium P3-1A highly efficient heterotrophic nitrifying bacterium P3-1 was isolated from the water of recirculation aquaculture systems.The strain was identified as Alcaligenes faecalis by morphological observation,physiological and biochemical reaction identification and 16S rRNA gene sequence analysis.The results of Gram staining showed that the strain was Gram-negative.The result of determination of extracellular enzyme showed that casein could be utilized by the strain.When temperature was 30°C,salinity was 10‰,and pH was 7 or 9,the strain grew fastest,and first reached the logarithmic growth period and the stable period.The results of nitrification experiments showed that environmental factors such as carbon source,carbon-nitrogen ratio and dissolved oxygen concentration could influence its rate of ammonia nitrogen removal.When sodium citrate was used as carbon source,carbon-nitrogen ratio was 10,pH was7,and rotational speed was 180 rpm,the highest ammonia nitrogen removal rate of 96%was achieved after 24 hours.3.Isolation and denitrification performance of a heterophic nitrifying bacterium P1-2A highly efficient heterotrophic nitrifying bacterium P1-2 was isolated from the water of recirculation aquaculture systems.The strain was identified as Rhodococcus pyridinivorans by morphological observation,physiological and biochemical reaction identification and 16S rRNA gene sequence analysis.The results of Gram staining showed that the strain was Gram-positive.The result of determination of extracellular enzyme showed that casein could be utilized by the strain.When temperature was 40°C,salinity was 10‰,and pH was 7,the strain grew fastest,and first reached the logarithmic growth period and the stable period.The results of nitrification experiments showed that environmental factors such as carbon source,carbon-nitrogen ratio and dissolved oxygen concentration could influence its rate of ammonia nitrogen removal.When sodium citrate was used as carbon source,carbon-nitrogen ratio was 15,pH was7,and rotational speed was 180 rpm,the highest ammonia nitrogen removal rate of99.63%was achieved after 24 hours.4.Effects of the three heterotrophic nitrifying bacteria for the remediation of aquaculture ponds of tilapiaThe strains P1-2,P3-1,and P5-2 were added into pond water of tilapia every 7 days?The final concentration of the bacteria of experimental groups is 5.0×10⁵ cfu/mL.The same volume of normal saline was added into the blank control group and bottom aeration control group?.Water quality parameters were monitored.?1?The result showed that for the water samples,the TP,NH₄⁺-N,TN and NO₂^--N removal rate of the strain P5-2 was significantly higher than that of blank control group?P<0.05?.For the sediment sampes,the NH₄⁺-N and TN removal rate of the strain P5-2 were 65.63%and55.35%,which were higher than the blank control?P<0.05?.?2?For the water samples,the TP,NH₄⁺-N,TN and NO₂^--N removal rate of the strain P3-1 was significantly higher than that of blank control group?P<0.05?.For the sediment samples,the NH₄⁺-N and TN removal rate of the strain P3-1 were 40.8%and 59.23%,which were higher than the blank control?P<0.05?.?3?For the water samples,the NH₄⁺-N and TN removal rate of the strain P1-2 was significantly higher than that of blank control group?P<0.05?.For the sediment samples,the NH₄⁺-N and TN removal rate of the strain P1-2 were 60%and 51.9%,which were higher than the blank control?P<0.05?.The results showed that three strains could significantly improve the quality of the aquaculture environment.All the three strains reduced the NH4⁺-N and TN concentrations of the water and sediment of tilapia ponds through nitrification.