Analysis Of Nitrogen Metabolism And Arsenic Tolerance Based On The Whole Genome Of Alsobacter Methylodruns SK200a-9

The heavy metal-tolerant strain Alsobacter methylodruns SK200a-9 is a type strain of the new family Alsobacteraceae.So far,Alsobacter methylodruns SK200a-9and Alsobacter soli SH9 are two species of the genus Alsobacter,and they are widely distributed in different habitats,implying that they have a potential ecological role in the global biogeochemical cycle.However,at present,apart from a general understanding of the heavy metal tolerance of SK200a-9,little is known about their functions and activities.Here,whole genome sequencing and biological information analysis of SK200a-9 was carried out.Based on whole genome analysis and comparative genome analysis,it was found that the arsenic tolerance genes of the two strains had good collinearity.Moreover,genes are quite different in nitrogen metabolism.Therefore,combined genomic and physiological activity experiments revealed the nitrogen metabolism and arsenic tolerance mechanism.The results are as follows:1.The results of whole gene sequencing show that the total length of the genome is 5101366 bp,and the average GC content is 67.59%.The genome predicted 4895genes,53 tRNAs,and 3 16S-23S-5S rRNA operons.2.Comparative genome analysis found that the genetic differences of the two strains of this family are manifested in environmental information processing,respiration,nitrogen metabolism,stress response,membrane transport and so on.In addition,SK200a-9 has more pathogenicity-related type VI secretion system gene clusters than SH9.In the classification of virulence and defense genes,it includes cobalt,zinc,cadmium,copper,mercury,zinc,magnesium,nickel,iron and other metal transport and resistance genes.3.The measurement results of nitrogen metabolism activity show that SK200a-9cannot utilize nitrate,while SH9 has dissimilation nitrate reduction activity.Both SK200a-9 and SH9 have NO₂^--N reduction activity,but their tolerance to nitrite is very different.The tolerance of SH9 to nitrite is more than fifty times that of SK200a-9.The nitrite-sensitive transcriptional repressor(nsr R)of SH9 may play a key role in improving nitrite tolerance.Both SK200a-9 and SH9 possess nor BC gene and activity.SH9 has clade?type nos Z gene and regulatory gene nos DFYLR.And confirmed that SH9 has a high N₂O reduction activity,SK200a-9 does not have N₂O reduction gene and activity.4.The results of arsenic tolerance test showed that the MIC₅₀ of As(?)of SK200a-9 was 11.7mM,and the MIC₅₀ of SH9 was 14.4mM.Scanning electron microscopy showed that cells without arsenic added were smooth and regular in shape,while cells under arsenic stress had strange shape.The results of adsorption accumulation or oxidation-reduction activity measurement show that SK200a-9 has a prominent adsorption and accumulation capacity,and SH9 has a prominent arsenic reduction capacity.Both SK200a-9 and SH9 have the cytoplasmic arsenic-reducing ars gene cluster,and SH9 also has the respiratory arsenic-reducing gene arr AB.The phylogenetic analysis of these 4 types arsenic-resistant proteins(Ars C,Arr AB,Acr3,Ars H)was carried out.5.The test results of the tolerance of the two strains to heavey metals showed that there was only a significant difference in thallium.The MIC₅₀ of SK200a-9 to Ti⁺was7.2 mM,while the MIC₅₀ of SH9 to Ti⁺was 0.6 mM,the thallium tolerance of SK200a-9 is 12 times that of SH9.In short,whole-genome analysis pointed out the direction for the study of its origin,evolution,and metabolic activity.On the basis of genome analysis,experiments confirmed that two strains of the Alsobacteraceae family had significant activity in arsenate tolerance and nitrogen metabolism activity.The whole genome data of SK200a-9 filled the blank of the bacterial gene database,and the experimental determination of the activity of the strains of this family provided a basic basis for reducing farmland N₂O and arsenic pollution.