基于电子分配的化能自养细菌培养策略  

Culture strategy of chemoautotrophic bacteria based on electron distribution

作　　者：何晓敏 林炜铁[1] 罗剑飞[1] HE Xiaomin;LIN Weitie;LUO Jianfei(School of Biology and Biological Engineering,South China University of Technology,Guangzhou,Guangdong,China)

机构地区：[1]华南理工大学生物科学与工程学院,广东广州

出　　处：《微生物学报》2025年第2期698-714,共17页Acta Microbiologica Sinica

基　　金：国家自然科学基金(91951118,41977034)。

摘　　要：【目的】针对化能自养细菌培养困难的问题,本研究基于电子分配理论分析它们难培养的原因,并基于纯培养探究电子分配策略对提高化能自养细菌生物量的可行性。【方法】从维持细胞内p H平衡和最适能量代谢的角度,计算得出硫氧化细菌盐硫小杆菌(Halothiobacillus)DCM-3、亚硝酸盐氧化细菌硝化杆菌(Nitrobacter)N1及氨氧化细菌亚硝化单胞菌(Nitrosomonas)SCUT-1氧化相应底物产生的电子最终分配给O_(2)与CO_(2)的最优比例,并以相应菌株为对象,分别设置不同的O_(2)与HCO_(3)-(CO_(2))摩尔量比例以形成不同的电子分配比例进行纯培养验证,用离子色谱仪和紫外分光光度计检测底物和产物浓度,用稀释涂布法测定细胞密度。【结果】DCM-3、N1和SCUT-1菌株的最优分配比例分别为0.733:0.267、0.867:0.133和0.6:0.4。若基于最优电子分配比例的条件,DCM-3、N1和SCUT-1菌株分别可合成3.967 ATP/S2O_(3)^(2-)、0.433 ATP/NO_(2)^(-)和1.35 ATP/NH_(3)。根据计算结果,其难培养的原因主要为单位底物提供的能量合成的ATP数量少,且要求适当控制低氧气浓度及补充适量的无机碳。纯培养验证中,DCM-3菌株在最优比例下的生物量为6.5×10^(7) CFU/mL,是不控制比例的对照组的2.2倍。N1菌株在最优比例下的生物量为7×10^(6) CFU/mL,与对照组无显著差异,但最优比例的HCO_(3)^(-)浓度(0.4 mmol/L)明显低于对照组(2.5 mmol/L),该菌株表现出倾向于较高氧气但对CO_(2)需求低的生长特性,与计算得到的最优比例相符。SCUT-1菌株在控制O_(2)和CO_(2)量的实验组中的单位NH_(4)^(+)浓度生物量积累达1.3×10^(6) CFU/(mL·(mmol/L))以上,比充足O_(2)和CO_(2)量条件的高25%-40%。【结论】基于电子分配的化能自养细菌培养策略,通过限制O_(2)和CO_(2)摩尔量并形成一定比例从而限制电子分配的培养条件,有助于提高同一底物条件下的生物量积累,为化能自养细菌的培养提�[Objective]In view of the difficulty in the culture of chemoautotrophic bacteria,this study analyzed the reasons for the difficulty based on the theory of electron distribution and explored the feasibility of using the electron distribution strategy for increasing the biomass of chemoautotrophic bacteria based on pure culture.[Methods]From the perspective of maintaining intracellular pH balance and optimal energy metabolism,we calculated the optimal distribution ratios of electrons produced by the sulfur-oxidizing bacterial strain Halothiobacillus sp.DCM-3,nitrite-oxidizing bacterial strain Nitrobacter sp.N1,and ammonia-oxidizing bacterial strain Nitrosomonas sp.SCUT-1 to O_(2)and CO_(2)by oxidizing corresponding substrates.Furthermore,different molar ratios of O_(2)to HCO_(3)-(CO_(2))were set respectively to form different electron distribution ratios for pure culture verification of the strains.Substrate and product concentrations were measured by ion chromatography and ultraviolet spectrophotometry,and cell density was measured by the dilution coating method.[Results]The optimal electron distribution ratios of strains DCM-3,N1,and SCUT-1 were 0.733:0.267,0.867:0.133,and 0.6:0.4,respectively.Based on the optimal electron distribution ratios,strains DCM-3,N1,and SCUT-1 could synthesize 3.967ATP/S_(2)O_(3)^(2-),0.433 ATP/NO_(2)^(-),and 1.35 ATP/NH_(3),respectively.According to the calculation results,the main reasons for the difficulty in culture were the small amount of ATP synthesized with the energy provided by per unit substrate and the need to control a low oxygen concentration and supplement an appropriate amount of inorganic carbon.The results of pure culture verification showed that the biomass of DCM-3 under the optimal ratio was 6.5×10^(7) CFU/mL,which was 2.2 times that of the control group.The biomass of N1 under the optimal ratio was 7×10^(6) CFU/mL,which was not significantly different from that of the control group.However,the HCO_(3)^(-)concentration(0.4 mmol/L)of the optimal ratio of strain N1

关 键 词：化能自养细菌 电子分配 能量代谢 纯培养 

分 类 号：Q93-335[生物学—微生物学]