Establishment Of Nitrogen-phosphorus Assimilation Marine Bacterial-algal Mutualistic System And Its Application In Saline Sewage Treatment

With the acceleration of urbanization and environmental pollution,the shortage of water resources has become an important problem in the world.To alleviate the shortage of freshwater resources and improve clean drinking water quality,more and more researchers have begun to seek alternative water sources.As the most economical alternative water source,seawater has been widely used in industry and agriculture.Especially,seawater for toilet flushing has covered 80%of the population in Hong Kong.However,the use of seawater substitutes still faces some problems and challenges.The seawater toilet flushing sewage(seawater blackwater)would enter to the municipal wastewater treatment system,treated together with other wastewaters and then be discharged into sea.The high salinity,high ammonium and low C/N in seawater blackwater have brought a great challenge on biological treatment system.In the current biological nitrogen removal methods,including nitrification/denitrification,ANAMMOX,SHARON,CANON,and HNAD,ammonium,nitrate,nitrite and other inorganic nitrogen all were converted to N2 and NxO(greenhouse gas)during the process.It will lead to nitrogen loss from aquatic systems and harm ecological environment.On the other hand,phosphate rock as a non-renewable resource will be depleted in 50-100 years.Phosphorus reserves have become the main factors of agriculture sustainable development.In fact,the main purpose of current wastewater treatment methods is ensure effluent meet the standards,the maximum nutrients recovery of nitrogen and phosphorus in the traditional tertiary treatment are only 15-20%and 20-30%,respectively.Thus nutrients recovery in sewage sludge for biofertilizer or livestock feed are a resource-oriented wastewater treatment scheme.S.platensis has some characteristics that high salt-tolerance,high growth rate,facultative heterotrophic with producing more oxygen,and superior performance in nitrogen and phosphorus removal by assimilation with harvesting valuable biomass.Although microalgae can achieve integration wastewater treatment and resource recovery,microalgae would release excessive alkali and organic carbon via overgrowth result in the deterioration of effluent water qualities.To address these challenges,a marine bacterium was used to achieve efficient removal of carbon and nutrients from synthetic seawater toilet flushing sewage in high organic load under single aerobic condition.Based on study the characteristics and mechanisms of nitrogen and phosphorus removal by the bacterium,a marine bacteria-S.platensis system was established and a photosynthetic membrane bioreactor was designed.The marine bacterial-algal system could achieve the treatment of seawater toilet flushing efficiently and energy-savingly,as well as harvest the valuable biomass with nutrients recovery.This paper mainly includes the following parts:(1)The mechanism of nitrogen removal and ecological toxicity of marine bacteria Vibrio.sp Y1-5 were systematically studied.The bacterium was assimilation nitrate reduction bacterium which can gradually reduce nitrate to nitrite,and then convert nitrite to ammonium under facultative aerobic conditions.Nitrate reductase is located in the cytoplasm and increasing oxygen concentration only accelerating nitrate conversion rather than affecting removal efficiency.Under single aerobic conditions,instead of nitrification,the strain assimilated ammonium directly,and it could tolerate as high as 1600 mg/L ammonium concentration while removing 844.6 mg/L.Ammonium assimilation occurred preferentially in the medium containing nitrate and ammonium.The ammonium assimilation has a certain inhibitory effect on nitrate reduction.The results of nitrogen balance illustrated that the removed nitrogen was all transformed to protein or stored as organic nitrogen substances in cells and no N was lost in the process.Toxicological studies with the brine shrimp species Artesia naupliia indicated that Vibrio sp.Y1-5 can be applied in aquatic ecosystems safely.(2)The characteristics of phosphorus removal and bioavailability of marine bacteria Vibrio.sp Y1-5 were systematically studied.The bacterium can grow well in salinity 1%-7%,pH 5-9,C/N 10-20,and initial phosphorus concentration 1-40 mg/L while 30 mg/L phosphorus were removed.Phosphorus removal by the bacterium needs pure aerobic conditions.The bacterium neither grows nor release of phosphorus under anaerobic condition.The phosphorus content in bacterial biomass accounts for 16.7%which is twice to PAOs.The proportion of OP+NAIP to TP was as high as 97.12%,implying high potentially mobile and bio-available P was stored in the bacterium.31P NMR showed that the bacterium can abort orthophosphate and gradually assimilate into phospholipids,pyrophosphate,polyphosphorus,etc.Among them,monoester-P and diester-P were accounted for 86.35%-76.68%of total phosphorus;phosphorus is the mainly IP which accounting for 13.65%-13.98%of TP;The proportion of pyrophosphorus and polyphosphate is very small.(3)Marine bacteria Vibrio.sp Y1-5 was used to treat different ratios(1:0,7:3,5:5,3:7)of seawater black water(seawater toilet flushing sewage)and grey water(municipal wastewater).The higher the ratio of black water,the better the bacterial growth status.The ammonium removal efficiency increased from 68.7%to 93.2%with the blackwater ratio decreased.The phosphorus removal efficiency was up to 97.39%and 98.12%in the ratio 5:5 and 3:7.The COD removal efficiency all were above 90%.The nutrients removal was directly related to biomass affected by dissolved oxygen concentration.During the nutrients removal process,626 mg/g-DW of extracellular polymers(EPSs)were produced,which could remove 63.9%of phosphorus and 71.7%of nitrogen,respectively.The removed nitrogen by EPSs was only stored as aromatic-protein substances without intermediate production in EPSs.Assimilated phosphorus was mainly orthophosphate and phospholipid which accounting for 70.28%of total phosphorus.Under the regulation of pH,metal and orthophosphate in the system were retarded by EPSs to form Mg3(PO4)2 and Ca(PO3)2 and other phosphates,which promoted the removal of phosphorus.(4)Based on the study of marine Vibrio.sp Y1-5 and characteristics of S.platensis,a marine bacterium-S.pirulina system was establishment for seawater flushing sewage treatment.The response surface experimental design study showed that the maximum removal efficiency obtained at the algal inoculum concentration of OD560 in mixture system 0.2 and the bacterial initial inoculum of mixture system 10%(v/v).Vibrio sp.Y1-5 in the algal-bacterial system has ability to regulate the pH of medium to lower than 9 which is of great significance to ensure the effluent standard and biomass yield.Both nutrients removal and biomass production exhibited the performance of algal-bacterial system was better than the sum of single systems suggesting bacteria and algae in the system was mutualistic.The TN removal rate was 85.4%(initial 200 mg/L),the total phosphorus removal rate was 87.8%(initial 40 mg/L),and the COD removal rate was 98.6%(initial 1600 mg/L)..Approximately 85.4%of TN(from 200 mg/L),87.8%of TP(from 40 mg/L)and 98.6%of COD(from 1600 mg/L)were removed from synthetic seawater toilet flushing sewage with 4.28 g/L biomass production containing 16.3%lipid and 62.6%protein.Nutrients removal was mainly assimilation/accumulation and chemical precipitation reinforced phosphorus removal.Mass balance analysis showed that 81 ± 14%of nitrogen and 86± 17%of phosphorus were recovered.Facultative heterotrophicsm of Spirulina ensured organic removal and avoiding excessive organic matter produced.PAC analysis confirmed that oxygen-producing by photosynthetic affect bacteria/algae biomass ratio;algae biomass,nutrient removal,and pH had a significant correlation;nitrogen removal mainly relied on algal assimilation;and the removal of organic matter and phosphorus relied on bacterium.(5)The marine bacterial-algal mutualistic system was operated and compared to single algal system for synthetic seawater toilet flushing sewage treatment and valuable biomass yield in DMPBR configuration at different carbon level.Compared with the single algae system,the bacteria-algal mutualistic system has shorter start-up time and faster dynamic membrane formation;the removal efficiency of ammonium,phosphorus and COD and biomass production by the mutualistic system is significantly higher than single algal system.With influent COD reduce from 1600 mg/L to 400 mg/L,nitrogen removal efficiency was decreased from 94.2%to 68.1%;while phosphorus removal increased firstly and then decreased.The removal efficiency of TP was up to 94.2%when influent COD was 800 mg/L.Removal efficiency of COD was higher than 90%during the whole operation period.When influent COD was 1200-400 mg/L,the effluent COD were below 50 mg/L.With,respect to biomass production and nutrient composition,the higher influent COD concentration,the greater biomass production and biomass productivity.When the influent COD was decreased from 1600 mg/L to 400 mg/L,the intracellular polysaccharide accumulation first increased and then decreased,the content was higher reported cultured microalgae.The protein content gradually increased,and the bacteria-algae system increased faster than the algal system,eventually reaching 61.9%-DW but still lower than cultured spirulina.The lipid production under salt stress was higher(>22%),and decreased slightly with carbon source decrease.The chlorophyll a content of the bacteria-algae system was similar to algal system suggesting that the mutualism of the bacteria and algae enhanced the quality of Spirulina.The analysis of extracellular polymer(SMP+eEPS)and SEM showed that the membrane interceptive was poor and effluent turbidity was too high in the early stage.After the membrane was successfully formed,no membrane fouling was observed.The effluent turbidity of mutualistic system is lower than that of single algal system.