Study On Sulfur Oxidation And Nitrogen Fixation Of Chemoautotroph Halothiobacillus Sp.LS2

Microbial-driven element biogeochemical cycle is not only the basic science of human cognition of the earth and nature,but also provides the key technology for solving the problems of resource shortage,energy shortage,ecological crisis and so on.The continuous study of the biogeochemical cycle of various elements in nature is not only an in-depth exploration of the earth's ecology,but also a solution to the problems facing mankind.In this paper,the bacteria,which can be isolated from the water of the Pearl River,can be used to autotrophic thiobacillus thiooxidans.The whole genome of LS2 was sequenced,and it was found that its genome contains a number of coding genes and complete nitrogen-fixing enzyme gene cluster,and it is presumed that it has the ability to fix N2 in addition to other sulfur-oxidizing bacteria.The relationship between the physiological and biochemical characteristics of nitrogen fixation under aerobic condition and the function of sulfur oxidation was studied,and the expressio n level of the gene was analyzed by fluorescence quantitative PCR.The results showed that the Nif gene cluster of Halthiobacillus sp.LS2 and the Acidithiobacillus ferrooxidans ATCC 23270 is very similar,and it can be assumed that the strain LS2 has a similar nitrogen fixation function to the Acidithiobacillus ferrooxidans ATCC 23270.In addition,the strain LS2 is the only sulfur-oxidizing bacteria with the complete nif gene cluster in the genus Halothiobacillus and the genus which is relatively close to it,and is the only sulfur-oxidizing bacteria which contains the Fix and the Rnf complex at the same time.According to the evolution and development analysis of three related functional genes of nifHDK,fixB and rnfB,and the gene cluster structure analysis to obtain the sulfur oxidation-nitrogen fixation metabolic pathway based on the whole genome level of the strain LS2.In addition,the strain LS2 contains a plurality of antioxidant enzymes and enzymes related to oxygen consumption,and the strain LS2 of these enzymes is presumed to be protected by the nitrogen fixation under the aerobic condition.Based on the results of the analysis of the relevant gene,the physiological and biochemical research of the sulfur oxidation and the nitrogen fixation of the strain LS2 was carried out,and it was found that the strain LS2 can be grown by the fixed nitrogen without anmmonia nitrogen,and the growth of the strain LS2 was less than that of the experimental group with the addition of ammonia nitrogen.The growth of the strain LS2 under the condition of low concentration ammonia nitrogen?1 mM NH₄⁺?use the ammonia nitrogen provided in the culture medium,and fix nitrogen at the time.The nitrogen fixation of the strain LS2 under the condition of high concentration of ammonia nitrogen?50 mM NH₄⁺?was not completely inhibited.The nitrogen concentration has little influence on the growth of the strain LS2,and the addition of 5%nitrous oxide gas can inhibit the nitrogen fixation of the strain to a certain extent.The nitrogen fixation effect can promote the growth of the strain LS2,and the growth of the strain LS2 can be inhibited when the nitrogen fixation is inhibited.Acetylene replaces nitrogen as the electron acceptor,which is more beneficial to the nitrogen fixation of the strain LS2.The oxygen limitation?5%O2?has no effect on the oxidation of the strain LS2 sulfur.Based on the analysis of the expression of the gene,it was found that the expression of the fixB,rnfB and nifH genes were down-regulated under the condition of high concentration of ammonia nitrogen.It was further explained that the nitrogen fixation of the strain LS2 under high concentration conditions was inhibited.When the concentration of the sulfur substrate is low,the strain LS2 is grown by the initial nitrogen source,and the nitrogen fixation effect is inhibited;when the concentration of the sulfur substrate is high?>1mM?,the expression amount of the nifH gene is increased as the concentration of the substrate increases.When the concentration of oxygen is limited,the expression of nifH gene can increase obviously,and the oxidation of sulfur is not affected by the concentration of oxygen.The strain LS2 can still perform the nitrogen fixation under the condition of high oxygen concentration mainly rely on the regulation and protection of the antioxidant enzyme system and oxygen consumption related enzyme.