Anaerobic Bio-dechlorination Of Chloroethenes Enhanced With Legacy Cobamides Offering By Rhodococcus Biphenylivorans TG9 During Its Dormancy

As common pollutants in soil and groundwater,chlorinated organics could be eliminated by microbial anaerobic dechlorination and aerobic degradation in the aerobic-anaerobic natural environment and several reaction systems.Dehalococcoides strains carrying out obligate organic dechlorinating respiration play important roles in anaerobicly dechlorinating chlorinated organics,while lack the ability to biosynthesize cobamides that function as cofactors of reductive dehalogenases and need to be provided exogenously.In this study,we identified the biosynthesis of the cobamide monomer-Vitamin B12(VB12)in aerobic bacterium Rhodococcus biphenylivorans TG9,added intracellular extracts of TG9 into an exogenously cobamide-dependent dechlorinating enrichment culture PSM to detect chloroethenes dechlorination,and constructed the PSM-TG9 coculture to investigate legacy VB12 and anaerobic dormancy of TG9.Another cobamide-insufficient dechlorinating enrichment culture TS was analyzed the promotion of chloroethenes dechlorination when added TG9.The main results are as follows:(1)Without the addition of VB12,the culture PSM could not perform the reductive dechlorination of chloroethenes.After adding VB12 standard for 21 days,the PSM dechlorinated 0.6-m M perchloroethylene(PCE)to 0.57-m M trichloroethylene(TCE)and 0.03-m M dichloroethylenes(DCEs).The dechlorinating genus was Dehalococcoides,and the functional gene was pcb A1.The biosynthesis of VB12 was detected in the intracellular of TG9 cultured aerobicly.After adding intracellular extracts of TG9 to the PSM for 21 days,0.6 m M PCE was converted into 0.58 m M TCE and 0.02 m M DCEs,Dehalococcoides 16 S r RNA coding gene and pcb A1 gene copy numbers increased.(2)In the PSM-TG9 coculture,0.6 m M PCE was converted into 0.59 m M TCE and 0.01 m M DCEs after 47 days,and the copy numbers of Dehalococcoides 16 S r RNA coding gene and pcb A1 gene increased.99.99% of TG9 was died after 49 days in anaerobic conditions,and released legacy VB12 to promot the reductive dechlorination of the culture PSM.TG9 could be induced to the “viable but nonculturable”(VBNC)state in anaerobic conditions after 14 days,0.01% of TG9 was still alive in VBNC state after 49 days,which could resuscitated after 24 hours by adding oxygen.(3)After 28 days,the dechlorinating enrichment culture TS only dechlorinated 0.07 m M PCE to TCE with its own insufficient cobamides,while converted 0.6 m M PCE to 0.44 m M TCE,0.11 m M trans-DCE and 0.05 m M cis-DCE with intracellular extracts of TG9 added.0.6 m M PCE was dechlorinated to 0.57 m M TCE and 0.03 m M DCEs in the TS-TG9 coculture.Dechlorinating bacteria detected were Dehalococcoides,and genes related were cbdb A1588 and cbdb A80.In this study,we analyzed the mechanism of cobamide-auxotroph bacteria Dehalococcoides performing chloroethenes dechlorination promoted by the legacy cobamides released from aerobic cobamide-producer bacterium TG9 in anaerobic conditions and the mechanism of dormancy and resuscitation under aerobic and anaerobic conditions,laying the foundation for clarifying the cofactor supply relationship between aerobic cobamide-producer bacteria and anaerobic dechlorinating bacteria,and providing new ideas for eliminating the cobamide limitation in the microorganism remediation of chlorinated organics.