Distribution And Correlation Of Novel Ammonia Oxidizing Microorganisms In Riparian Zone

The riparian zone ecosystem has the good characteristics of the land-freshwater interfaces and has long been regarded as biogeochemical hot zones of nitrogen cycling,which has a good interception effect on ammonia nitrogen.Comammox,anammox,AOA,and AOB play a major role in ammonia nitrogen removal.The relative contribution of four ammonia-oxidizing microorganisms to ammonia nitrogen removal has not been clarified to date.So,we determined the abundance and community compositions of comammox,anammox,AOA and AOB to study the environmental factors affecting the distribution and occurrence of comammox,anammox,AOA and AOB in riparian zone,which is important for the evaluation of ammonia oxidation process and ammonia nitrogen removal in riparian zone.The objective aim of the study was to investigate the occurrence,distribution and contribution of comammox in riparian zone and to establish a novel understanding of the role of comammox in ammonia oxidation in riparian zone,as compared to anammox and traditional ammonia oxidation.Rhizosphere and non-rhizosphere samples were collected under different seasons and spatial distributions,using molecular biology methods,¹⁵N isotope tracing techniques,high-throughput sequencing,clone library and mathematical statistics analysis.The main research results are as follows:?1?The abundance of the comammox bacteria based on the amoA functional gene ranged from 1.04±0.22×10⁸ to 3.19±0.39×10⁸ copies g^-1 at the soils/sediment samples in riparian zone.The abundance of the AOA archaea based on the amo A functional gene ranged from 7.34±0.15×10⁸ to 2.39±0.31×10⁹ copies g^-1,while the abundance of the AOB bacteria based on the amoA functional gene ranged from6.33±0.40×10⁶ to 5.13±0.27×10⁷ copies g^-1.Moreover,the abundance of Nitrospira bacteria based on the nxrB functional gene ranged from 2.43±0.88×10⁸ to1.10±0.52×10⁹ copies g^-1.The abundance of the anammox bacterial based on the hzsB ranged from 9.93±3.14×10³ to 1.49±0.24×10⁷ copies g^-1.Furthermore,the potential activity of anammox bacteria ranged from 0.00±0.00 to 6.55±0.61 nmol N g^-1 h^-1.The abundance and potential activity of anammox bacteria reached the maximum at the riparian interface sediment?Site A?irrespective of the season and the soil type?rhizosphere/non-rhizosphere?,as well as the abundance and potential activity of anammox bacteria gradually decreased from the riparian interface sediment to the central riparian soil?Site D?.In summary,the abundance of AOA was the highest,followed by Nitrospira,comammox,AOB and anammox bacteria irrespective of season and soil type?rhizosphere/non-rhizosphere?,indicating that they coexist and have a certain competitive relationship in riparian zone.Based on above,suggesting that the removal of ammonia nitrogen may be dominated by AOA,and the comammox bacteria played a key role in the process of ammonia oxidation and even the whole process of nitrogen cycle as well as the anammox bacteria played a leading role in sediments in riparian zone.?2?Pearson's correlation analysis showed that comammox and Nitrospira abundance were not closely associated with NO₃^-,NO₂^-and NH₄⁺concentrations,suggesting that the abundance of comammox bacteria may not be regulated by the substrate after the substrate concentration is below a certain threshold.The abundance and potential activity of anammox bacterial were significantly positive correlated with water content?WC?,but negatively correlated with TC,while the abundance and potential activity of anammox bacterial were not associated with NO₂^-and NH₄⁺,indicating that water content is an important parameter for the distribution and contribution of anammox in riparian zone.Moreover,the abundance of AOA was significantly positive correlated with TC and TOM,while the abundance of AOB was significantly positive correlated with TN and TC,which suggests that substrates are unlikely to be the only factor determining the contribution and distribution of comammox,Nitrospira,anammox,AOA and AOB bacteria in different ecological niches,whereas are driven by other key environmental factors?WC,TN,TC and TOM?as well as microorganisms.?3?The abundance of comammox bacteria was positively correlated with the relative abundance of Candidatus Nitrospira nitrificans,and it was also positively correlated with the sum of the relative abundances of the species Candidatus Nitrospira nitrificans and Candidatus Nitrospira nitrosa,suggesting that the abundance of comammox bacteria contributed a higher Nitrospira abundance.The abundance of Nitrospira bacterial was positively correlated with the relative abundance of Candidatus Nitrospira nitrificans and the sum of the relative abundances of the species Candidatus Nitrospira nitrificans and Candidatus Nitrospira nitrosa.Over all,indicating that both Candidatus Nitrospira nitrificans and Candidatus Nitrospira nitrosa play an important role in comammox and Nitrospira.Furthermore,the potential activity of anammox bacterial was significantly positive correlated with the anammox abundance.The abundance and potential activity of anammox bacterial were significantly negative associated with comammox abundance,indicating that anammox and comammox bacteria coexist in riparian zone and may be competitive each other.In summary,we demonstrate the existence of complete nitrification by a single organism?comammox?and a significant and previously unrecognized contribution of comammox to the mediation of autotrophic nitrificationin riparian zone ecosystems.These results will greatly promote the understanding of comammox processes in widely distributed riparian zone ecosystems.This study is the first observation of the occurrence,distribution and contribution of comammox,as well as studying the interactions among comammox,anammox and traditional ammonia oxidation microorganisms.Filling this gap in current knowledge is important,as ammonia oxidation is a crucial component of the biogeochemical nitrogen cycle.