Study On The Oxidation Of Ammonium Coupling With Sulfur Transformation And The Function Of EPS Of Anammox Sludge

Anaerobic ammonia oxidation(Anammox)has been widely concerned at home and abroad in recent decades.Although a large number of studies have been carried out around Anammox as well as some wastewater treatments applied,our understanding of anaerobic ammonia oxidation bacteria(An AOB)is still limited.It has been shown that the Anammox sludge existed mostly as granule or biofilm in reactors and exhibited red color(commonly known as“red bacteria”).In addition,its extracellular polymeric substances(EPS)content was relatively higher than other kinds of nitrogen removal sludges.The EPS of Anammox sludge showed typical red characteristics as well as the bacteria.Previous studies on the EPS of sludge mainly focused on flocculation,adsorption,hydrophilic/hydrophobic,sedimentation,etc.,but there were few on the response mechanism of the EPS when An AOB was subjected to the substrate inhibition or other adverse conditions.Moreover,“excessive”removal of ammonium was often observed in Anammox reactors with long-term supply of sulfate.Whether this phenomenon was related to sulfur metabolism still lacked of research.Aimed to above questions,this study used two sequencing batch reactors(SBR).On the basis of rapid startup of Anammox,the research firstly focused on the roles of EPS in Anammox granulation,substrate inhibition as well as the reconstruction of Anammox activity.Then,the red characteristics and biochemical catalytic activity of the EPS of Anammox granules were studied.Finally,the operation characteristics of Anammox reactor under low Rs _IN(influent NO₂^--N/NH₄⁺-N)were investigated,and the capacities of An AOB were discussed,which exhibited sulfate reducing anaerobic ammonium oxidation(SRAAO)and the sulfur-based autotrophic denitrification with long-term sulfate supply,meanwhile,the microbial community of the reactors were analyzed.The main results obtained from this study are as follows:1.During the granulation of Anammox sludge,there was no significant correlation between EPS content and the particle size.But the ratio of protein content to polysaccharide content(PN/PS)increased firstly,then started to drop when more than half of the granules in the reactor in size was turned from small size(<300?m)to medium size(300 to 500?m).Protein was the main component of the EPS of the Anammox granules.As the granules size increased,the values of?-helix/(?-sheet+random coil)in loosely bound EPS(LB-EPS)and tightly bound EPS(TB-EPS)gradually decreased from0.58 and 0.67 to 0.36 and 0.29,respectively.This change was beneficial to hydrophobicity on the surface of the granules and promoted the granulation of the sludge.2.When concentration of NO₂^--N in the reactor was 209 mg/L under pH=7.74,concentration of FNA exceeded 0.0072 mg N/L,substrate inhibition appeared in the SBR-Anammox system,leading to a 45%reduction of the specific Anammox activity.During the inhibition,the Anammox granules tended to secrete a large amount of EPS,and the protein content increased with high PN/PS ratio.In the recovery period,the EPS increment was positively correlated with the increment of the nitrogen removal rate(NRR).After the recovery of Anammox activity,the EPS content remained stable,and no longer increased in spit of the loading rised.It was believed that the An AOB could resist and remove the FNA inhibition by secreting more EPS,which also maintained the stability of the grannule structure.This was a kind of self-protection mechanism of the bacteria.3.The EPS of Anammox granules contained two multiheme enzymes,hydroxylamine oxidoreductasec(HAO)and hydrazine-oxidizing enzyme(HZO),which made the EPS exhibit significantly biology catalytic activity,and endowed the two parts of the EPS in various shades of blood red.The catalytic activities of LB-EPS with hydroxylamine and hydrazine were both lower than that of TB-EPS.The maximum hydroxylamine catalytic rates of HAO in the two kinds of EPS were 0.010 mmol·(L·h)^-1(LB-EPS)and 0.171 mmol·(L·h)^-1(TB-EPS),as well as the maximum hydrazine catalytic rates of HZO were 0.054 mmol·(L·h)^-1(LB-EPS)and 0.382 mmol·(L·h)^-1(TB-EPS),respectively.5.A significant co-metabolism of nitrogen and sulfur was observed in the Anammox system supplying sulfate for a long-term operation.With the declination of Rs _IN,the over-consumption of ammonium became more distinct in the reactor,meanwhile,the potential of nitrogen removal ability dominated by Anammox decreased gradually.The An AOB enriched by this Anammox system could not only remove NH₄⁺and SO₄^2-synchronically,but also had a typical sulfur autotrophic denitrification capacity.The N/S ratio in substrate had an influence on the products composition of sulfide in the liquid phase and the yields of elemental sulfur.With the high N/S(=2.5)ratio,S₂O₃^2-was the main product in the liquid and less elemental sulfur in the solid.The main product with low N/S(?1)ratio was SO₄^2-.The Sulfate Reducing Anaerobic Ammonium Oxidation(SRAAO)and Dissimilar Nitrate Reduction to Ammonium(DNRA)were observed during the tests.High-throughput sequencing results showed that the dominant An AOB in the long-term sulfate supply Anammox system were Anammoxoglobus.The short rod and filamentous bacteria formed as inner skeleton-structure of the granules,dividing it into several small compartments,in which a large number of spherical Anammoxoglobus embedded tightly.The Thioprofundum lithotrophica was spiral rod,which was closely related to sulfur metabolism,distributed in the outer edge of the granular,or formed dense colony adhering to the surface of it.In the anaerobic sulphate-ammonia oxidation system,the coupling transformation of nitrogen and sulfur was a complex biochemical process in which a variety of bacteria participated,competed and cooperated with each other.