Okamoto Akihiro, Rowe Annette, Deng Xiao, Nealson Kenneth H
International Center for Materials Nanoarchitectonics, National Institute for Materials Science;
Department of Earth Sciences, University of Southern California.
J Vis Exp. 2018 Jul 24(137):57632. doi: 10.3791/57632.
Anaerobic respiration coupled with electron transport to insoluble minerals (referred to as extracellular electron transport [EET]) is thought to be critical for microbial energy production and persistence in many subsurface environments, especially those lacking soluble terminal electron acceptors. While EET-capable microbes have been successfully isolated from various environments, the diversity of bacteria capable of EET is still poorly understood, especially in difficult-to-sample, low energy or extreme environments, such as many subsurface ecosystems. Here, we describe an on-site electrochemical system to enrich EET-capable bacteria using an anode as a respiratory terminal electron acceptor. This anode is connected to a cathode capable of catalyzing abiotic oxygen reduction. Comparing this approach with electrocultivation methods that use a potentiostat for poising the electrode potential, the two-electrode system does not require an external power source. We present an example of our on-site enrichment utilized in an alkaline pond at the Cedars, a terrestrial serpentinization site in Northern California. Prior attempts to cultivate mineral reducing bacteria were unsuccessful, which is likely due to the low-biomass nature of this site and/or the low relative abundance of metal reducing microbes. Prior to implementing our two-electrode enrichment, we measured the vertical profile of dissolved oxygen concentration. This allowed us to place the carbon felt anode and platinum-electroplated carbon felt cathode at depths that would support anaerobic and aerobic processes, respectively. Following on-site incubation, we further enriched the anodic electrode in the laboratory and confirmed a distinct microbial community compared to the surface-attached or biofilm communities normally observed at the Cedars. This enrichment subsequently led to the isolation of the first electrogenic microbe from the Cedars. This method of on-site microbial enrichment has the potential to greatly enhance the isolation of EET-capable bacteria from low biomass or difficult to sample habitats.
厌氧呼吸与电子传递至不溶性矿物质（称为细胞外电子传递[EET]）被认为对许多地下环境中微生物的能量产生和生存至关重要，尤其是那些缺乏可溶性终端电子受体的环境。虽然已成功从各种环境中分离出具有EET能力的微生物，但对具有EET能力的细菌多样性仍知之甚少，特别是在难以采样、低能量或极端环境中，如许多地下生态系统。在此，我们描述了一种现场电化学系统，该系统使用阳极作为呼吸终端电子受体来富集具有EET能力的细菌。该阳极连接到能够催化非生物氧还原的阴极。将这种方法与使用恒电位仪来保持电极电位的电培养方法进行比较，两电极系统不需要外部电源。我们展示了在加利福尼亚北部陆地蛇纹石化地点锡达尔斯的一个碱性池塘中利用我们的现场富集方法的一个例子。先前培养矿物还原细菌的尝试均未成功，这可能是由于该地点生物量低的性质和/或金属还原微生物的相对丰度低。在实施我们的两电极富集之前，我们测量了溶解氧浓度的垂直剖面。这使我们能够将碳毡阳极和镀铂碳毡阴极分别放置在支持厌氧和好氧过程的深度。现场培养后，我们在实验室进一步富集阳极电极，并确认与通常在锡达尔斯观察到的表面附着或生物膜群落相比，有一个独特的微生物群落。这种富集随后导致从锡达尔斯分离出第一种产电微生物。这种现场微生物富集方法有可能极大地增强从低生物量或难以采样的栖息地中分离具有EET能力的细菌。
