Nitrogen Transformation Processes And The Diversity Of Functional Microbial Communities In Polar Environment

Nitrogen is one of the essential elements for all living organisms and the main nutrient limiting life on the earth.In nature,microorganisms can drive the nitrogen biogeochemical cycles,forming a complex nitrogen cycle network.In the past decade,many new metabolic capabilities and novel microorganisms related to nitrogen cycle were identified,such as anaerobic ammonium oxidation(Anammox),complete ammonia oxidation to nitrate(comammox),and ammonia-oxidizing archaea(AOA).These new discoveries have challenged the previous understanding of the biogeochemical cycle of nitrogen.Therefore,the biogeochemical cycle of nitrogen and its influence mechanism has been paid more attentions and become an international frontier topic recently.At present,extensive studies on the nitrogen transformation processes and the diversity for their function microorganisms have been carried out globally in terrestrial and aquatic ecosystems.However,little is known about nitrogen conversions and the diversity of functional microorganisms in polar ecosystems,especially in Antarctic tundra and lake environments.Whether the anaerobic ammonium oxidation(Anammox)exists in Antarctic extreme environments and the role that Anammox plays has not been reported in the literatures.Therefore,it is important to conduct the studies on nitrogen conversion and the function microorganisms in the context of global climate change.In this paper,the Fildes Peninsula in western Antarctica and Arctic Ny-Alesund were selected as study areas.The slurry experiments with 15N stable isotope tracer method and molecular biology methods,including clone libraries,metagenomics and real-time quantitative PCR,were used to(?)systematically study the rates of nitrogen conversion processes,including Anammox,denitrification,nitrification and DNRA in Antarctic and Arctic tundra soils,and lake/marine sediments,and their affecting factors;(?)to analysize the distribution and the diversity for ANAMMOX,AOB,AOA and denitrifying microorganisms and their affecting factors;(?)to comprehensively explain the effects of their effects on nitrogen cycles in polar tundra and lake environments;and(?)to discuss the effects of environmental factors and sea animal activities on nitrogen conversion and the diversity for their function microorganisms in polar environment.The main finfings are as following:1.Ammonia oxidation and diversity of functional microbial communities in Antarctic tundra soils.We chose seal colony tundra soils(SS),penguin colony soils(PS)and adjacent penguin-lacking tundra soils(PL),tundra marsh soils(MS),and background tundra soils(BS),to investigate the effects of sea animal colonization on the abundance,activity,and diversity of AOA and AOB in maritime Antarctica.Results indicated that AOB dominated over AOA in PS,SS,and PL;whereas AOB and AOA abundances were similar in MS and BS.Penguin or seal activities increased the abundance of soil AOB amoA genes,but reduced the abundance of AOA amoA genes,leading to very large ratios(1.5 × 102 to 3.2×104)of AOB to AOA amoA copy numbers.Potential ammonia oxidation rates(PAOR)were 0.64?9.91 nmol N g-1 h-1,significantly higher(P<0.05)in SS and PS than in PL,MS,and BS.PAOR was significantly positively correlated(P<0.01)with AOB amoA gene abundance,suggested that AOB play a more important role in the nitrification in animal colony soils.Sequence analysis for gene clones showed that AOA and AOB in tundra soils were from the Nitrososphaera and Nitrosospira lineages,respectively.The differences in AOB and AOA community structures were closely related to soil biogeochemical processes under the disturbance of penguin or seal activities:soil C:N alteration and sufficient input of NH4+-N and phosphorus from animal excrements.2.The Anammox,denitrification,DNRA and their contribution to nitrogen cycles in Antarctic tundra soil.It was found that the presence of Anammox bacteria(AnAOB)existed in Antarctic tundra soil by metagenomic sequencing.AnAOB in the penguin colony soil accounted for 0.18%of the total sequenced bacteria.Candidatus Kuenenia stuttgartiensis showed the highest abundance,accounting for 61.9%of the total AnAOB.The abundance of Anammox 16Sr RNA gene in Antarctic tundra soil was(9.79±0.11)x106?(1.61±0.08)× 108 copies g-1 soil,and was positively correlated with soil pH(P<0.01).The potential Anammox rate is 0.02?2.08 nmol N g-1 h-1,which is proportional to the NH4+-N(P<0.01),phosphorus(P<0.05)and sulfur(P<0.01)content in the soil,and inversely proportional to the soil C:N ratio(P<0.01).The contribution of Anammox to soil N2 production is 3.8%?62.0%.It is affected by soil C:N ratio(P<0.05)and NH4+-N/NO3--N ratio(P<0.01)that related to animal activities.In the non-animal activity area with high C:N ratio,the contribution of Anammox is higher;in the animal activity area,soil N2 production is dominated by denitrification.In addition,the potential DNRA rate in Antarctic tundra soil is 0.11?10.55 nmol N g-1 h-1,which is proportional to soil phosphorus(P<0.01)and sulfur(P<0.01)content,and inversely proportional to soil C:N ratio(P<0.05),contributing 3.5%?74.1%of soil total nitrate reduction.In the soil of non-animal activity areas,there is a coupling effect with Anammox and DNRA process.3.Nitrogen transformation process and functional microbial community in the sediments of Antarctic lakes.It was found that Anammox bacteria existed in the sediments of G Lake,Y3 Lake,Y4 Lake,Changhu Lake and Yanou Lake in the Fildes Peninsula of Antarctica by using the method of 15N-isotope tracer and molecular biology.The sequencing results indicated that the Anammox bacteria found in lake sediments were mostly related to the genus 'Brocadia'.The abundance of Anammox 16SrRNA gene ranged from(1.09±0.06)×106to(1.33±0.05)× 108 copies g-1 sediments,whose community structure was affected by environmental factors such as pH(P<0.05).AOB belonging to Nitrosospira was detected in sediments of Lake G,Y3 and Y4;while AOA belonging to group 1.1b Nitrososphaera cluster was detected in the sediments of Y2 and Yanou Lake.The study indicated that the C:N ratio was the key factor affecting the spatial distribution of AOA and AOB(P<0.05).DNRA is the most active reaction among the four nitrogen transformation processes in Antarctic lake sediments.The DNRA rates were 0.11?0.73 nmol Ng-1 h-1,which might be related to the high organic carbon content caused by low organic decomposition rate.The Anammox rates were 0.05?0.49 nmol Ng-1 h-1,which was coupled with DNRA to provide the contribution of N2 production up to 38.1%-91.4%in the Antarctic coastal lake sediments.4.Nitrogen transformation process and functional microbial community in Arctic Kongsfjorden sediments.The average gene abundance of AnAOB detected in the Kongsfjorden sediment was(1.30±0.82)x 107 copies g-1 sediment,belonging to the genus Candidatus Scalindua.Most of the AOA detected in the sediments belong to the group 1.1a Nitrososphaera cluster,while the AOB belong to the Nitrosolobus genus of the ?-proteobacteria.The abundance of AOB amoA gene in sediments was(1.61±1.03)x 108 copies g-1,higher than the abundance of AOA amoA[(6.42±5.52)x 105copies g-1 sediments].From inner basin to outer basin,the abundance of the Anammox 16S rRNA gene(P<0.01),nirS gene(P<0.05)and AOB amoA gene(P<0.01)increased with the increase of the TOC:N ratio in the sediment.The potential Anammox rate in Kongsfjorden sediments is(0.46±0.11)nmol N g-1 h-1,and the contribution to N2 production is 48.5%?62.7%.The DNRA rate in sediments was(0.90±0.62)nmol N g-1 h-1.DNRA accounted for 27.3%?67.3%of total nitrate reduction,and was negatively correlated with the content of NH4+-N(P<0.05).The potential nitrification rate is(0.69±0.36)nmol N g-1 h-1,and was inversely proportional to TOC in the sediments(P<0.05).5.Nitrogen transformation process,functional microbial community and N2O emission flux in Arctic tundra.Although the Anammox 16S rRNA gene abundance in Arctic Ny-Alesund tundra soil was as high as(8.48± 8.16)×107 copies g-1 soil,no Anammox activity was detected in Ny-Alesund tundra soil samples.The abundance of AOA amo A gene was(9.09±0.91)×106 copies g-1 soils,higher than the abundance of AOB amo A gene(P<0.05).Under the condition of 4? culture,the potential denitrification rate was(3.03± 1.79)nmol N g-1 h-1,higher than the potential nitrification rate which was(1.16±0.05)nmol N g-1 h-1.The soil moisture(P<0.05),TC(P<0.01),TOC(P<0.05),TN(P<0.05)and NH4+-N(P<0.05)levels were the key factors to determine the denitrification activity in Ny-Alesund tundra soils.The potential DNRA rate was(0.33±0.17)nmol N g-1 h-1,which was related to soil sulfur content(P<0.05).The N2O flux was determined to be 7.6?14.6?g N2O m-2 h-1,which was proportional to the potential nitrification rate(P<0.01).It is speculated that N2O was produced by the so-called nitrifier denitrification by AOA and AOB in Ny-Alesund tundra soils.