Polyethersulfone filter

An acetogenic microbiome previously enriched on graphite granule cathodes at potentials from -590 to -800 mV vs. SHE was used to inoculate the bioreactors (LaBelle et al., 2014 (link)). The cells were drawn from the cathode compartment of an active reactor and were concentrated using tangential flow filtration using a 0.2 μm polyethersulfone filter (Sartorius), spun at 5000 (RCF) for 10 min and re-suspended in fresh medium before transfer to the RVC cathode reactors. The culture, inoculum and reactor, were treated as an open, non-aseptic system.
The reactor was operated at 25°C with constant current supplied from a VMP3 Potentiostat (BioLogic) set in galvanostatic mode and the voltage monitored with EC Lab software. A galvanostatic operation of 8 A/L_catholyte was chosen to overcome electron limitation to a biofilm immobilized on the cathode and promote fast colonization. Humidified 100% CO₂ was passed through the catholyte headspace using Norprene tubing at an initially set rate of 25 mL/min. Filter sterilized medium was flowed into the base of the cathode compartment and exited just above the top of the RVC cathode at a rate of 250 mL/day (a dilution rate of 5 day^-1) using a peristaltic pump and PharMed BPT tubing. Deionized water was added to ports made from syringe housing on the anolyte compartment daily to compensate for water oxidation and evaporation.

Sampling took place in the summer of 2019, part of the PACES project, in the Lanaudières and Laurentides regions in Quebec, Canada. The groundwater samples that we used in this study to compare with aquatic surface samples are part of a bigger groundwater sample collection that was initially published in Groult et al. [12 (link)]. Lakes, rivers, and stream water samples were collected the same day as the surrounding groundwater samples and were chosen as likely discharge zones of groundwater flowing back to the surface (S. Gagné personal communication, Figure 1). Six surface samples did not seem connected to any groundwater systems but were still used as representatives of surface aquatic samples (Supplemental Table S1). Lake, river, and stream water samples were collected in sterilized polypropylene bottles (Nalgene, Rochester, NY, USA), transported on ice, and stored at 4 °C until filtration in the lab, which was done the same day as sampling. Filtration was carried out using a 0.2 µm polyethersulfone filter (Sartorius, Göttingen, Germany) with 1 L of water. Filters were subsequently stored at −20 °C.

Growth rates were determined based on biomass dry weight samples collected during the exponential growth phase. First a known amount of cell culture (typically around 3 mL) was filtered through pre-dried and weighted 0.45 µm polyether sulfone filters (Sartorius Stedim Biotech) which was then washed with demineralized water. The filter was folded to lock the biomass inside and dried in a microwave at 150 W for 20 min. The dry weight was measured after a cooling period of approximately 30 min in a desiccator.

DGT samplers (DGT Research Ltd., Lancaster, UK) were used for both solution and soil tests. Polyethersulfone filters (0.45 μm pore size, 0.13 mm thick, Sartorius Stedim, Goettingen, DE) were used as a protective membrane. The membranes were washed in 5 % HNO₃ (w/w) and stored in an aqueous 10 mmol L⁻¹ NaNO₃ solution (Reagent Plus, Sigma Aldrich, Buchs, CH). Agarose cross-linked polyacrylamide (APA) diffusive hydrogels of 0.8 mm thickness were prepared according to [11 (link)] and cut to discs. Anion exchange resin (Amberlite IRA-400; chloride form, Sigma Aldrich, Buchs, CH) hydrogels for S sampling (0.4 mm thickness) were prepared according to [10 ]. Therefore, 3 g of the resin was ground with a ball mill for 10 min, passed through a 200-μm sieve, and washed in 10 % HCl (p.a., Merck). The resin was mixed with 10 mL gel solution [11 (link)], 60 μL of riboflavin solution (0.01 g riboflavin ((-)-Riboflavin, Sigma Aldrich) in 10 mL H₂O), and 20 μL of tetramethylethylendiamine (TEMED; VWR Int., Randor, USA). The solution was shaken well and cast between two acid-washed glass plates (6 × 20 cm) separated by a U-shaped acid-washed plastic spacer (0.4 mm thickness). The glass plate with the freshly coated gel solution was left under fluorescent light overnight for photopolymerization. The gels were hydrated and cut to discs. A 10 mmol L⁻¹ NaNO₃ solution was used for storage of all gels.

In Sep 2014 and June 2017 groundwater samples (40 ml) were collected on two dates, filtered in the field through 0.2 μm polyethersulfone filters (Sartorius Stedim Biotech GmbH, Germany) and stored at -20°C in 50 ml polyethylene screw cap tubes. Samples were analysed (Dept. of Catchment Hydrology, UFZ, Germany) for the isotopic composition of NO₃^- (^15/14N and ^18/16O), NH₄⁺ (^15/14N), and H₂O (^2/1H and ^18/16O). Gas exetainers of 12 ml were additionally used for dissolved-N₂O (^15/14N and ^18/16O). Isotope values are reported in δ‰ relative to international standards (AIR for N and VSMOW (Vienna Standard Mean Ocean Water) for O and H). Water δ¹⁸O and δD (δ²H) signatures for H₂O were analysed in accordance with [5 ,6 ]. The results are interpreted according to a modified Rayleigh equation: f = 1—e^{(δs–δs0)/ε}, where f is the fraction of substrate remaining, ε is the isotope enrichment factor, and δs and δs0 are the isotopic composition of the residual substrate and initial substrate, respectively. The δ¹⁵N-NH₄⁺ was measured in subsurface locations with detectable NH₄⁺-N concentration. The δ¹⁵N signature for NH₄⁺ was obtained as in Zhang et al. [25 (link)]. The δ¹⁵N-NO₃^- and δ¹⁸O-NO₃^- were obtained as in [26 (link)]. The δ¹⁵N and δ¹⁸O composition of the produced N₂O (plus that of the dissolved N₂O samples) were measured as in [5 ].