Constructing simplified microbial consortia that couple lactic acid and ethanol utilization to highly produce caproic acid from liquor-making wastewater

IF 11.4 1区环境科学与生态学 Q1 ENGINEERING, ENVIRONMENTAL

Water Research Pub Date : 2025-06-07 DOI:10.1016/j.watres.2025.123973

Jinfeng Xiang , Zihan Zhou , Zhihao Liu , Cong Ren , Yan Xu

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Abstract

Converting biodegradable carbon in wastewater into medium-chain fatty acids (MCFAs) through stable microbiota is highly attractive. In this study, we utilized a top-down approach for constructing MCFA-producing microbial consortia. Specifically, an enrichment and plating-screening strategy employing lactic acid and ethanol as selective carbon sources was applied to isolate simplified caproic acid-producing microbial consortia from liquor-making pit mud. The representative microbial consortium SimpCom3 demonstrated high level of caproic acid production (14.62 ± 0.48 g/L) in a semi-synthetic medium, significantly outperforming consortium SimpCom1 (5.96 ± 0.11 g/L) and consortium SimpCom2 (9.63 ± 0.16 g/L). This performance of microbial consortium SimpCom3 was attributed to its ability to co-utilize lactic acid and ethanol, produce fewer odd-chain fatty acids byproducts, and maintain pH self-regulation between 6.45 and 8.29. Metagenomic analyses revealed the dominance of Clostridium kluyveri (30.69 %–50.46 %), C. butyricum (6.71 %–13.98 %) and C. tyrobutyricum (37.11 %–58.07 %) in consortium SimpCom3, which synergistically converted lactic acid and ethanol to caproic acid via reverse β-oxidation. Stable performance over 56 days of cyclic-batch fermentation processes confirmed the robustness of consortium SimpCom3. When applying consortium SimpCom3 to unsterilized liquor-making wastewater in a fermenter with a fed-batch approach, 22.13 g/L caproic acid was produced with 66.38 % selectivity, and microbial dynamics analysis demonstrated the consortium's high adaptability to real wastewater. Metabolic analysis based on high-quality assembly metagenomes (HQ-MAGs) revealed a novel cooperative metabolism: cross-feeding between Clostridium kluyveri (which utilizes ethanol and produces caproic acid) and lactate-utilizing butyrate producers maintained consortium stability and enhanced caproic acid production. Crucially, the decarboxylation of lactic acid counteracted acidification caused by ethanol-driven caproic acid synthesis, enabling self-regulated pH stability within the simplified microbiome system. Together, this study presents a simplified microbial consortium construction method for caproic acid production from liquor-making wastewater, overcoming the limitations of synthetic co-cultures and enhancing the viability of chain-elongation biorefineries in wastewater treatment.

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构建乳酸和乙醇耦合利用的简化微生物群落，从酿酒废水中高效生产己酸

通过稳定的微生物群将废水中的可生物降解碳转化为中链脂肪酸（MCFAs）具有很高的吸引力。在这项研究中，我们采用了自上而下的方法来构建产生mcfa的微生物群落。具体而言，采用乳酸和乙醇作为选择性碳源的富集和电镀筛选策略，从酿酒窖泥中分离出简化的己酸产菌群。具有代表性的微生物联合体SimpCom3在半合成培养基中表现出较高的己酸产量（14.62±0.48 g/L），显著优于联合体SimpCom1（5.96±0.11 g/L）和联合体SimpCom2（9.63±0.16 g/L）。微生物联合体SimpCom3的这种表现归因于其能够共同利用乳酸和乙醇，产生较少的奇链脂肪酸副产物，并保持pH在6.45至8.29之间的自我调节。宏基因组分析显示，菌株SimpCom3中kluyveri梭状芽胞杆菌（30.69% ~ 50.46%）、C. butyricum（6.71% ~ 13.98%）和C. tyrobutyricum（37.11% ~ 58.07%）占主导地位，通过β-反氧化作用将乳酸和乙醇协同转化为己酸。56天循环分批发酵过程的稳定表现证实了财团SimpCom3的稳健性。将联合体SimpCom3应用于发酵罐中未灭菌的酿酒废水，采用间歇进料法，自己酸产率为22.13 g/L，选择性为66.38%，微生物动力学分析表明联合体对实际废水具有较高的适应性。基于高质量组装宏基因组（hg - mags）的代谢分析揭示了一种新的合作代谢：克卢维梭菌（利用乙醇生产己酸）和利用乳酸的丁酸生产者之间的交叉饲养维持了联盟的稳定性，并提高了己酸的产量。至关重要的是，乳酸的脱羧抵消了乙醇驱动的己酸合成引起的酸化，使简化的微生物群系统能够自我调节pH稳定性。总之，本研究提出了一种简化的微生物联合体构建方法，用于酿酒废水生产己酸，克服了合成共培养的局限性，提高了延伸链生物精炼厂在废水处理中的可行性。

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来源期刊

Water Research 环境科学-工程：环境

CiteScore

20.80

自引率

9.40%

发文量

1307

审稿时长

38 days

期刊介绍： Water Research, along with its open access companion journal Water Research X, serves as a platform for publishing original research papers covering various aspects of the science and technology related to the anthropogenic water cycle, water quality, and its management worldwide. The audience targeted by the journal comprises biologists, chemical engineers, chemists, civil engineers, environmental engineers, limnologists, and microbiologists. The scope of the journal include: •Treatment processes for water and wastewaters (municipal, agricultural, industrial, and on-site treatment), including resource recovery and residuals management; •Urban hydrology including sewer systems, stormwater management, and green infrastructure; •Drinking water treatment and distribution; •Potable and non-potable water reuse; •Sanitation, public health, and risk assessment; •Anaerobic digestion, solid and hazardous waste management, including source characterization and the effects and control of leachates and gaseous emissions; •Contaminants (chemical, microbial, anthropogenic particles such as nanoparticles or microplastics) and related water quality sensing, monitoring, fate, and assessment; •Anthropogenic impacts on inland, tidal, coastal and urban waters, focusing on surface and ground waters, and point and non-point sources of pollution; •Environmental restoration, linked to surface water, groundwater and groundwater remediation; •Analysis of the interfaces between sediments and water, and between water and atmosphere, focusing specifically on anthropogenic impacts; •Mathematical modelling, systems analysis, machine learning, and beneficial use of big data related to the anthropogenic water cycle; •Socio-economic, policy, and regulations studies.