Study On Phenotype And Genotype Of Halophilic Archaea And The Dominated Nitrogen Cycling In Saline Soil

Halophilic archaea are halophilic,widely distributed in high-salinity ecosystems such as salt lakes,salt mines,and saline soils.With researchers'exploring of extreme environmental biological resources and the development of genome sequencing technology,more and more halophilic archaea have been identified,described and sequenced.In order to study the growth,metabolic characteristics and potential ecological function of halophilic archaea,this paper integrated and analyzed the physiological and biochemical characteristics and genome data of halophilic archaea.High throughput sequence and genomic data were combined to analyze the dominated nitrogen cycle through denitrification and nitrate dissimilation reduction to ammonium(DNRA).Through the analysis of the halophilic archaeal genomes,the phylogenetic tree of 207 species of halophilic archaea based on the tandem sequence of 122 marker proteins could distinguish the three orders of halophilic archaea more clearly.Through the analysis of the basic genome information,habitat and growth conditions of halophilic archaea,it was found that the haloarchaea of the order Halobacteriales were widely distributed in ecosystems such as terrestrial,marine and high-salt food,the genome size ranged from 2.09 Mb to 5.61 Mb,the number of protein coding sequences ranged from 2140 to 5647,the G+C mol%content ranged from 47.7 to 68.7,and the growth range of p H,Na Cl and temperature were 5.0 to 10.0,0.8 to 6.0 M,15 to 60?,respectively;the haloarchaea of the order Haloferacales were mainly distributed in terrestrial and marine ecosystems.These halophilic archaea could be clearly distinguished in the genomic phylogenetic trees,with a genome size ranging from 2.47 Mb to 5.19 Mb,the number of protein coding sequences being between 2570 and 5299,the G+C mol%content ranging from 58.9 to 69.2,and the growth range of p H,Na Cl and temperature were 5.0 to 11.0,0.9 to 6.0 M,10 to 60?,respectively;the haloarchaea of the order Natrialbales were halophilic and alkalophilic,which were mainly distributed in terrestrial alkaline ecosystems.Their genome size ranged from 3.22Mb to 5.44 Mb,the number of protein coding sequences from 3124 to 5209,the G+C mol%content from 60.2 to 66.1,and their growth range of p H,Na Cl,and temperature were 6.0 to 11.0,1.3 to 6.0 M,and 17 to 60?,respectively.Through the analysis of utilizable carbon sources and energy substances of halophilic archaea,it was found that most halophilic archaea had a preference utilization of glucose,mannose,sucrose and glycerol.The utilizable alcohols,amino acids and organic acid in halophilic archaea of the order Natrialbales are less than those of Halobacteriales and Haloferacales,which indicated that the Natrialbales have higher requirements for carbon source or energy sources.The wide distribution and energy sources utilization of halophilic archaea enable them to perform ecological functions in a variety of ecosystems without being restricted by environment.The analysis of the genome annotation results of 207 species of halophilic archaea found that,in addition to the 6 strains of inorganic heterotrophic or oligotrophic halophilic archaea isolated from hypoxic and sulfide-rich environments,such as Halodesulfurarchaeum formicicum,Halococcoides cellulosivorans,et al.,the other halophilic archaea contained complete genes related to the TCA cycle.We speculated that halophilic archaea participate in the carbon cycle through processes such as the tricarboxylic acid cycle,glycolysis,and pentose phosphate pathways.In addition,we found some halophilic archaea also possess encoding genes of key enzymes in the chitin deacetylation and phenolic acid degradation pathways(Gentisate 1,2-dioxygenase,?-ketoadipate enolactone hydrolase),which is related to the conversion of chitin and phenolic acids in the environment.The three orders of halophilic archaea could perform nitrate assimilation,but the process of nitrate dissimilation is mainly dominated by Halobacteriales and Haloferacales.Among them,Halobacteriales and Haloferacales can reduce nitrate to nitrite,mainly through denitrification to dissimilate nitrate with product of N₂O and N₂;Natrialbales cannot dissimilate nitrate,but they can use nitrite as a substrate to participate in the nitrogen cycle in the ecosystem through denitrification and DNRA.Among all the denitrifying halophilic archaea,Haloferacales and Natrialbales could employ N₂ as the end product,while Halobacteriales can only employ greenhouse gas N₂O as the end product.Besides,some halophilic archaea possessed genes encoding thiosulfate reductase(phs ABC)and arsenic oxidase,played a role in the process of sulfur reduction and arsenic oxidation.As a result,halophilic archaea play an irreplaceable role in the biogeochemical cycle of C,N,S and other elements in the high-salt ecosystem.Through physical and chemical properties determination of 3 inland saline soil samples and5 coastal saline soil samples,it was found that the saline soil could provide the suitable salinity for the growth of halophilic archaea(19.34?29.07 g·kg^-1).The micorcosm experiment showed that the archaeal community had a higher contribution to the microbial denitrification than the bacterial and fungal communities of saline soil.High-throughput sequencing technology was employed to analyze the archaeal communities of saline soils,and we found that in both inland and coastal saline soil samples,halophilic archaea were the dominant groups.Halobacteriales,Haloferacales and Natrialbales were distributed in the inland saline soil samples,including the unique Natrialbales.The halophilic archaea in the coastal saline soil samples were mainly Halobacteriales and Haloferacales.By combining high-throughput sequencing results with genome annotations results,aimed to analyze the nitrate and nitrite cycle dominated by haloarchaea,all the saline soils samples contained high relative abundance of denitrifying haloarchaea and low relative abundance of DNRA haloarchaea,and the relative abundance of halophilic archaea with nor was higher than that with nos Z,so we speculate that the denitrification products of halophilic archaea in saline soil are mainly greenhouse gas N₂O.By predicting the metabolic pathway of the nitrogen cycle in halophilic archaea,we think that when denitrification is performed by haloarchaea,the electrons needed are produced by the organic matter in the soil to participate in glycolysis and TCA cycle.In addition,haloarchaea could also participate in the cycle of S and As elements in the saline soil ecosystem through the sulfur reduction and arsenic oxidation.Therefore,we believe that halophilic archaea dominate the regulation of nitrate and nitrite in high-salinity ecosystems through denitrification,and have non-negligible contributions to nitrogen cycle in high-salt ecosystems.In this paper,the distribution,growth conditions,phenotype and genotype characteristics of halophilic archaea were analyzed.And the role and potential function of halophilic archaea in the biogeochemical cycle of high salinity ecosystem were revealed at the genomic level.The nitrogen cycle in saline soil dominated by haloarchaea through denitrification and DNRA was also studied.The purpose is to provide a theoretical basis for the excavation of halophilic archaea with excellent traits,and to provide guidance for the cultivation and screening of halophilic archaea from the perspective of phenotype and genotype,and improve the efficiency of screening targeted halophilic archaea.