Novel experimental methods for the identification of anoxic micro-niches in porous media.

Porous media found in the shallow subsurface host an extremely heterogeneous environment arising from the complex coupling of abiotic (e.g. chemical reactions and anomalous solute transport) and biotic (e.g. metabolism and growth) processes. This heterogeneity is expected to characterize oxygen concentration distribution which is one of the major drivers for both abiotic and biotic redox reactions. Anoxic micro-niches, i.e. small portions of medium characterized by disproportionately different physical-chemical properties and microbial community composition compared to those characterizing the medium bulk, are expected to occur and persist even in averagely well-oxygenated porous media explaining macroscopic observed phenomena. However, the current lack of non-invasive technologies to observe the oxygen concentration field in porous media at spatial scales of interest for bacteria (i.e., 10 - 100 μm) structures still limits our ability to attain a quantitative description of anoxic micro-niches formation phenomenology in terms of their spatial distribution, average inter-niche distances, and proportion between oxygenated and anoxic pore-volume. This work presents the development, the implementation and preliminary as- sessment of a novel experimental methodology to observe oxygen concentration gradients and their evolution in space and time. This methodology combines the use of: a) PDMS microfluidics devices, which mimicking natural porous media geometries; b) planar transparent optodes which are fluorescent chemical sensors whose fluorescence intensity is quenched as a function of the oxygen concentration; and c) fully-automated microscope which allows to collect large images. The dynamics of oxy- gen concentration fields generated by pure physical processes are compared to those generated by the coupled effect of solute transport and the metabolism of aerobic bacteria. Our results allow to a) demonstrate the compatibility of microfluidics devices and optodes, b) highlight the strengths and challenges of the proposed novel methodology and c) reveal the ability of the planar optodes to capture fast evolving and sharp gradients associated with oxygen within porous media environment.