反硝化菌群的萘代谢与反硝化偶联机制

Investigation of the Coupling Mechanism Between Naphthalene Degradation and Denitrification of a Naphthalene Degraded Bacterial Consortium Under Denitrification

为探寻反硝化与萘代谢过程的偶联机制,从潜在PAHs(polycyclic aromatic hydrocarbons)污染的土壤中富集获取了萘的反硝化降解菌群.通过Illumina Mi Seq测序对其细菌群落结构进行了解析,并研究其萘代谢过程中反硝化电子受体[硝酸根(NO_3^-)、亚硝酸根(NO_2^-)]浓度、气态还原产物[氧化亚氮(N_2O)、氮气(N_2)]产生速率及反硝化微生物相关的nar G(periplasmic nitrate reductase gene)和nir S(cd1-nitrite reductase gene)基因丰度的动态变化.Illumina Mi Seq测序结果表明,变形菌门中的Pseudomonas是该富集菌群中丰度最高的菌属.富集获取的萘反硝化降解菌群9 d内对萘的降解率为49.11%,培养初期(1~3 d)及末期(7~9 d)萘的降解速率无差异,但它们均显著高于培养中期(3~7 d)的降解速率(P<0.05).培养期间,培养液中NO_3^-浓度呈逐渐下降趋势,而NO_2^-积累出现在第1~3 d.培养的3~9 d,NO_2^-浓度迅速下降,但在培养中期(3~7 d)未检测到气体产生,只在培养末期(7~9 d)检测到明显的N2O[3.39μg·(L·h)-1]和N2[8.97μg·(L·h)-1]的产生.在培养期间nar G及nir S基因的丰度均随培养时间而上升,表明该富集菌群中反硝化微生物丰度的逐渐增加.综上,NO_3^-还原过程及随后的NO_2^-还原过程等产气过程均可能与萘的厌氧降解过程相偶联,该结果可为进一步深入探讨萘的反硝化代谢机制打下基础.

To investigate the coupling mechanism between naphthalene degradation and denitrification using a liquid media containing naphthalene（ sole carbon source） and nitrate,a naphthalene degradation bacterial consortium under denitrification was enriched from a soil with potential risk of PAH（ polycyclic aromatic hydrocarbon） contamination. The bacterial community composition of the enriched consortium was analyzed by Illumina Mi Seq Sequencing. Subsquently,the enriched consortium was cultured under anaerobic conditions for 9 days,and the concentrations of electron acceptors（ nitrate and nitrite） for denitrification,gaseous reduction products（ N2O and N2） involved in denitrification,and abundances of denitrification concerned genes（ nar G： periplasmic nitrate reeducates gene; nir S：cd1-nitrite reductase gene） were detected at days 1,3,7,and 9. The result of Illumina Mi Seq Sequencing showed that Pseudomonas（ Proteobacteria） was the most dominant genus in this enriched consortium. Under anaerobic conditions,the naphthalene removal rate of this enriched consortium was 49. 11% within 9 days. Relative higher naphthalene degradation rates were found both at the beginning（ day 1-3） and the end（ day 7-9） of incubation,and these were significantly higher than at the middle（ day 3-7） of the incubation stage（ P 0. 05）. The concentration of nitrate decreased during the whole culture period,while the concentration of nitrite increased during the initial incubation（ day 1-3） and rapidly decreased from day 3 to 9. Furthermore,obvious productions of N2O [3. 39μg·（ L·h）-1]and N2[8. 97 μg·（ L·h）-1]were only measured at the end of incubation（ day 7-9）. The abundances of both nar G and nir S increased during the incubation,indicating the continuous growth of denitrifiers in the enriched consortium during the incubation period. In summary,this study illustrated that both the nitrate reducing stage and gas producing steps of denitrification could be coupled with anaerobic naphthalene degradation,which might be helpful for a deeper investigation regarding the coupling mechanism between denitrification and anaerobic PAH degradation.