Pilot-scale Columns Equipped with Aqueous and Solid-phase Sampling Ports Enable Geochemical and Molecular Microbial Investigations of Anoxic Biological Processes   

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A brief version of this protocol appeared in:
Environmental Science & Technology
Jan 2016


Column studies can be employed to query systems that mimic environmentally relevant flow-through processes in natural and built environments. Sampling these systems spatially throughout operation, while maintaining the integrity of aqueous and solid-phase samples for geochemical and microbial analyses, can be challenging particularly when redox conditions within the column differ from ambient conditions. Here we present a pilot-scale column design and sampling protocol that is optimized for long-term spatial and temporal sampling. We utilized this experimental set-up over approximately 2 years to study a biologically active system designed to precipitate zinc-sulfides during sulfate reducing conditions; however, it can be adapted for the study of many flow-through systems where geochemical and/or molecular microbial analyses are desired. Importantly, these columns utilize retrievable solid-phase bags in conjunction with anoxic microbial techniques to harvest substrate samples while minimally disrupting column operation.

Keywords: Environmental engineering, Geochemistry, Microbiology, Pilot-scale, Anaerobic respiration, Redox, Bioremediation, Bioreactor


The following describes an experimental design and sampling protocol that circumvents the obstacle of vertical coring for temporal and spatial resolution of column systems. The system has the further advantage of minimal disruption to physical, chemical and biological processes. The pilot-scale design incorporates vertically spaced sampling ports for collection of liquid and solid-phase substrate at discrete time points (Figure 1). Spatial sampling of solid-phase substrates that reside within these columns enables researchers to observe biologically relevant processes such as discrete zones of metal immobilization and shifts in microbial biofilm communities. The evolution of these vertical biogeochemical profiles can be tracked and related to performance over time. While optimized for anoxic systems as described below, this experimental design, which surmounts obstacles of more conventional flow-through column systems, could be applied for spatial inquiry into other systems that rely on aqueous and solid-phase interactions.

Of particular interest to our research, Sulfate Reducing Bioreactors (SRBRs) have been employed to mitigate the release of Mining Influenced Water (MIW) for approximately two decades (Wildeman et al., 1994). Due to the anoxic nature of these systems and their spatial heterogeneity, sampling SRBRs without operational disruption presents many challenges. Previously these systems were sampled with limited spatial resolution potentially excluding seminal processes occurring in regions within (Neculita et al., 2008). While sacrificial sampling can surmount this obstacle, it does so at the expense of temporal resolution. Furthermore, spatial inquiry into pilot-scale and larger SRBRs is challenging due to difficulty associated with coring saturated, heterogeneous organic materials (woodchips, sawdust, hay) and disruptions that can result from this form of sample collection. Our design and sampling procedure enabled us to examine the performance of sulfate reducing bioreactors that treat mining influenced water as a function of organic substrate, microbial community structure, water quality, and metal-sulfide precipitation yielding novel insights into the operation of these systems (Drennan et al., 2016).

Figure 1. Schematic of vertical down flow biochemical reactor columns. The three ports, indicated by circles on the column, were designed for solid substrate retrieval in conjunction with flow along the z-axis of the columns. The five liquid ports are depicted in blue along the side of the column. For discrete retrieval of solid substrates, columns were temporarily tilted to a horizontal plane using a custom built rack to mitigate water pressure complications and resultant loss (Figures 5 and 6). As labelled, ‘MIW Inf.’ indicates where the mining influenced water is introduced. The effluent from these columns was collected at the bottom liquid port as visualized in Figure 4A.

Copyright: © 2017 The Authors; exclusive licensee Bio-protocol LLC.
How to cite: Drennan, D. M., Almstrand, R., Lee, I., Landkamer, L., Figueroa, L. and Sharp, J. O. (2017). Pilot-scale Columns Equipped with Aqueous and Solid-phase Sampling Ports Enable Geochemical and Molecular Microbial Investigations of Anoxic Biological Processes. Bio-protocol 7(1): e2083. DOI: 10.21769/BioProtoc.2083.

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