Investigating The Sulfide Oxidation Pathway In Ruegeria Pomeroyi DSS-3 And Its Role In S-N Coupling

Sulfur cycling in nature is tightly related to the cycling of carbon and nitrogen,and can be coupled through various biological processes.The bottom of the sea is the main site for the biogeographic sulfur cycle.Marine cycle of inorganic sulfur mainly includes H2S oxidation and sulfate reduction.In the seabed sediments,sulfate can be used as electron acceptor for organic mineralization to be reduced to H2S in the anaerobic zone,and a large amount of H2S generated can be diffused to the oxygen-containing zone in the upper layer to generate sulfate again through multiple reactions.At present,there is no clear conclusion as to how H2S is oxidized in marine sediments,despite a large number of relevant studies.Current studies suggest that sulfur oxidation in sediments is mainly through chemical oxidation and some autotrophic sulfur-oxidizing bacteria through their unique mechanism.Since the chemical oxidation rate is slow and sulfur oxidation capacity the above two bacteria of is limited,we speculate that there may be other sulfur oxidation pathways in the seabed sediments.A study in our laboratory found that heterotrophic bacteria have a wide range of ability to rapidly oxidize H2S complete sulfur oxidation pathway of heterotrophic bacteria.Sulfide:quinone oxidoreductases(SQR)and flavocytochrome c-sulfide dehydrog-enases(FCSDs)are two main enzymes for H2S removal.SQR is more active than FCSDs for sulfide oxidation.In heterotrophic bacteria,SQR can cooperate with persulfide dioxygenase(PDO)to oxidize sulfide.SQR oxidizes H2S to zero valence sulfur with intracellular GSH as sulfur acceptors to produce different sulfane sulfur species.Then PDO oxidizes GSSH into sulfite.The latter reacts with zero valeance sulfur to form thiosulfate spontaneously.Sulfur transferase speeds up this process.Finally,thiosulfate will be oxidized to sulfate by Sox system.In this paper,we measured sulfides in offshore sediments and found that the content of S0 was significantly higher in the deeper sediments.Thus,we hypothesized that with the decrease of oxygen content in the sediments,the activity of PDO was restricted,leading to the accumulation of polysulfide.These results suggests that PDO pathway may play an important role in the sulfur oxidation pathway of sediments.Marine microorganisms play a very important role in driving carbon,nitrogen and sulfur cycles and coupling due to their wide distribution,diverse metabolic types and large quantities.Roseobacter is the main flora in seawater and offshore sediments.They are heterotrophic bacteria which widely contain SQR-PDO and Sox systems in the genome of the known Roseobacter.These findings suggest that heterotrophic bacteria may be involved in H2S oxidation in marine sediments.In order to verify the sulfur oxidation function of Roseobacter,we use the model strain R.pomeroyi DSS-3 as the research object,and it is found through experiments that DSS-3 can metabolize H2S rapidly,and further generate thiosulfate,which is finally oxidized to sulfate.Furthermore,we optimized the genetic manipulation of the bacteria and knocked out the key sulfur oxidation genes.Finally we verified the function of sulfur oxidation genes in its genome through metabolic experiments.It shows that the pdo pathway may play an important role in sediment sulfur oxidation pathway.Studies have found that in the subaerobic layer of offshore sediments,the coupling of H2S oxidation and denitrification is crucial.It is generally believed that the chemical autotrophic microorganisms are functional flora that couples H2S oxidation and denitrification.DSS-3 also contains a relatively complete denitrifying enzyme system,so it may couple H2S oxidation with denitrification to adapt to the micro-aerobic and anaerobic environment in marine sediments.In this paper,we tested whether DSS-3 can use nitrate to oxidize H2S through physiological experiments to explore the coupling effect of R pomeroyi DSS-3 in sulfur and nitrogen metabolism.In summary,through a series of experiments,we verified the function of R.pomeroyi DSS-3 sulfur oxidation gene and explained its sulfur oxidation pathway.At the same time,we explored whether it can couple H2S oxidation with nitrate reduction,revealing the important role of heterotrophic microorganisms in sulfur oxidation in offshore sediments.