Porewater and microbial community properties of permeable riverbed sediments across the south of England, 2018

Two field campaigns were conducted in 2018 in winter [February] and summer [2018] seasons) to measure in situ rates of nitrogen transformations, porewater chemistry and microbial gene abundance and transcripts in 12 rivers with permeable beds and varying concentrations of phosphorous in the Hampshire Avon catchment, Kent, and Essex.

Nitrogen transformations were measured and porewater samples collected with a "push-pull" technique. Briefly, 15 bespoke stainless steel probes were installed in the riverbed at two depths (7 at 5cm and 8 at 10 cm) to cover three unvegetated sediment patches. Porewater samples were obtained by a syringe connected to the porewater probe and applying a slight vacuum (by hand). From each probe, the O2 and pH content of the porewater was first determined and a newly collected porewater sample was preserved for gas analysis. Porewater Temperature and pH were measured with a Hach multi-meter and O2 concentrations were measured with a fast response microelectrode. Finally, another freshly collected porewater sample was passed through a 0.45 micron polypropylene filter and frozen for later nutrient (ammonium, nitrite, nitrate phosphate) analysis on a Skalar SAN++ auto analyser. To measure rates of nitrogen transformations, 15N-labelled ammonium or nitrate was injected into riverbed and porewater samples for gas and nutrient/chloride analysis were recovered from the same probe over time. The tracer was 300 microM, 98 atom % 15N-ammonium or nitrate in a synthetic river water matrix with additional KCl (approx. 150 mg per L). Each experiment used 25 mL of tracer, consisted of 4-time points (immediately following the injection plus three others) and lasted for no more than 60 minutes in total. For each sediment patch, a single 2cm sediment core was taken (before push-pull analyses) and subsampled at 5 and 10 cm depths for molecular analyses. The core was positioned no less than 20cm from a push-pull probe to avoid surface water intrusion into the porespace. Subsamples were then cryopreserved in liquid N2.

16S rRNA and N-cycle functional genes and transcripts were quantified by qPCR and RT-qPCR on a on a BioRad CFX384 following extraction of RNA and DNA with PowerSoil DNA or RNA Isolation kits (Qiagen). Genes were also amplified via PCR, indexed using Nextera XT indices (Illumina) and sequenced using MiSeq Reagent Kits v3 (600-cycle)on an Illumina MiSeq sequencer in foursequencing runs.