Microbiologically Influenced Corrosion Behavior And Mechanism Of X80 Pipeline Steel By Nitrate-reducing Bacterium,Bacillus Licheniformis

In this work,the microbiologically influenced corrosion(MIC)behavior and mechanism of X80 pipeline steel by nitrate-reducing bacterium Bacillus licheniformis were investigated.First,the impact of B.licheniformis on X80 steel corrosion was monitored for bacterial whole growth cycle by means of electrochemical analysises,weight loss test,etc.Second,a novel method was developed for detecting extracellular electron transfer(EET)from steel substrate to the cells and B.licheniformis was evidenced to be able to take up electrons from the X80 steel substrate for respiration.Third,the EET pathway from X80 steel to B.licheniformis was studied using electrochemical tests,proteomic analysis,etc.Finally,the B.licheniformis cell/X80 steel interface was revealed using focused-ion-beam(FIB)milling technique coupled with scanning/transmission electron microscope(SEM/TEM).The main results are as follows:Bioenergetic analysis implies that microorganisms can harvest the released energy from metal corrosion for survival via EET under carbon-starvation conditions.In this part,we combined carbon starvation with CTC redox fluorescent probe and developed a novel method for detecting EET from steel substrate to the cells.B.licheniformis was evidenced to be able to take up electrons from the X80 steel substrate for respiration,which accounts for a more critical role in substrate corrosion than the chemical effects of bacterially-secreted organic acids This study provides reliable support to the bioenergetic theory for MIC of carbon steel by nitrate-reducing bacterium.B.licheniformis can take up electrons from steel surface via three routes:direct electron transfer by cytochromes,direct electron transfer by flavin-bound cytochromes and mediated electron transfer by diffusible flavin.There are two types of cytochromes who have the capacity of electron transfer on the bacterial(near-)surface:P450 and c553.B.licheniformis can only secrete traces of riboflavin and it acquires extracellular electrons directly via the surface cytochromes without exogenous mediators.These riboflavin is mainly bound to the P450 cytochrome assisting electron transfer.Adding flavins can increase the bound amount and with the binding sites of P450 cytochrome becoming saturated,the rest of flavins will mediate electron transfer by diffusion.The B.licheniformis cell/X80 steel interface analysis provides structural and chemical insights to MIC mechanisms and permits to a better understanding of bacteria/surface interactions.B.licheniformis cells were found to be not in direct contact with steel surface.They were encrusted by amorphous Fe-phosphates like a capsule and attached on an amorphous aggregate layer of extracellular polymeric substances,Fe-(oxyhydr)oxides,sulphides etc.The amorphous Fe-phosphates capsule and aggregate layer can act as the channel for direct electron transfer and the encrusted cells are still able to assimilate nutrients(electron mediators for mediated electron transfer,for instance).B.licheniformis can accelerate corrosion of X80 steel in the early immersing by two ways:biocatalytic cathodic nitrate reduction and acidification by bacterially-secreted organic acids.The acidification can retard the passivation of steel surface and induce local corrosion and biocatalytic cathodic nitrate reduction reaction extracts the electrons from anodic dissolution,which accelerate the corrosion of steel substrate jointly.After immersing for ca.1 week in B.licheniformis culture,the corrosion rate of X80 steel decreased due to iron biomineralization.Although B.licheniformis is incapable of oxidizing Fe(II)enzymatically,it can generate nitrite under the starvation condition.Nitrate will abiotically oxidize Fe(II)and lead to the precipitation of amorphous Fe-phosphates.The Fe-phosphate coated steel substrate and inhibitted corrosion consequently.