ALBERT

Description: Upwelling systems are significant sources of atmospheric nitrous oxide (N₂O). The Benguela Upwelling System is one of the most productive regions worldwide and a temporally variable source of N₂O. Strong O₂ depletions above the shelf are favoring periodically OMZ formations. We aimed to assess underlying N₂O production and consumption processes on different temporal and spatial scales during austral winter in the Benguela Upwelling System, when O₂-deficiency in the water column is relatively low. The fieldwork took place during the cruise M157 (August 4th – September 16th 2019) onboard the R/V METEOR. This expedition included four close-coastal regions around Walvis Bay at 23°S, which presented the lowest O₂ concentrations near the seafloor and thus may provide hotspots of N₂O production. Seawater was collected in 10 L free-flow bottles by using a rosette system equipped with conductivity-temperature-depth (CTD) sensors (SBE 911plus, Seabird-electronics, USA). Incubation experiments were performed using stable isotope ¹⁵N-tracers. Seawater samples for ¹⁵N-tracer incubations and natural abundance N₂O analysis were collected from 10 L free-flow bottles and filled bubble-free into 125 mL serum bottles. The samples for natural abundance N₂O analysis were immediately fixed with saturated HgCl₂ and stored in the dark. To perform the incubation, we added ¹⁵N-labeled NO₂-, NO₃⁻ and NH₄⁺ to estimate the in-situ N₂O production rates and associated reactions. To determine a single rate, the bottles were sacrificed after tracer addition, and within the time interval of 12 h, 24 h and 48 h by adding HgCl₂. Rates were calculated based on a linear regression over time. Total N₂O and natural abundance isotopologues of N₂O were analyzed by using an isotope ratio mass spectrometer (IRMS, Delta V Plus, Thermo Scientific). NO₂- production was additionally analyzed by transforming ¹⁵NO₂- to ¹⁵N₂O following the azide method after McIlvin & Altabet (2005) and the nitrogen isotope ratio of N₂O was measured by an IRMS. N₂ production was determined via an IRMS (Flash-EA-ConfloIV-DELTA V Advanced, Thermo Scientific) by injecting headspace from exetainers. The N₂O yield per nitrite produced and the N₂O yield during denitrification was calculated. Samples for natural abundance N₂O was sampled and measured in triplicates and is shown as an average with standard deviation (SD). In order to estimate the contribution of different N₂O producing pathways by major biological processes and the extent of N₂O reduction to N₂, the dual-isotope mapping approach was applied to natural abundance isotopologues of N₂O, which uses the relative position of background-subtracted N₂O samples in a δ¹⁵Nˢᴾ-N₂O vs. δ¹⁸O-N₂O diagram (Yu et al., 2020; Lewicka-Szczebak et al., 2020).

Description: © The Author(s), 2019. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Torres-Beltran, M., Mueller, A., Scofield, M., Pachiadaki, M. G., Taylor, C., Tyshchenko, K., Michiels, C., Lam, P., Ulloa, O., Jurgens, K., Hyun, J., Edgcomb, V. P., Crowe, S. A., & Hallam, S. J. Sampling and processing methods impact microbial community structure and potential activity in a seasonally anoxic fjord: Saanich Inlet, British Columbia. Frontiers in Marine Science, 6,(2019):132, doi:10.3389/fmars.2019.00132.

Description: The Scientific Committee on Oceanographic Research (SCOR) Working Group 144 Microbial Community Responses to Ocean Deoxygenation workshop held in Vancouver, B.C on July 2014 had the primary objective of initiating a process to standardize operating procedures for compatible process rate and multi-omic (DNA, RNA, protein, and metabolite) data collection in marine oxygen minimum zones and other oxygen depleted waters. Workshop attendees participated in practical sampling and experimental activities in Saanich Inlet, British Columbia, a seasonally anoxic fjord. Experiments were designed to compare and cross-calibrate in situ versus bottle sampling methods to determine effects on microbial community structure and potential activity when using different filter combinations, filtration methods, and sample volumes. Resulting biomass was preserved for small subunit ribosomal RNA (SSU or 16S rRNA) and SSU rRNA gene (rDNA) amplicon sequencing followed by downstream statistical and visual analyses. Results from these analyses showed that significant community shifts occurred between in situ versus on ship processed samples. For example, Bacteroidetes, Alphaproteobacteria, and Opisthokonta associated with on-ship filtration onto 0.4 μm filters increased fivefold compared to on-ship in-line 0.22 μm filters or 0.4 μm filters processed and preserved in situ. In contrast, Planctomycetes associated with 0.4 μm in situ filters increased fivefold compared to on-ship filtration onto 0.4 μm filters and on-ship in-line 0.22 μm filters. In addition, candidate divisions and Chloroflexi were primarily recovered when filtered onto 0.4 μm filters in situ. Results based on rRNA:rDNA ratios for microbial indicator groups revealed previously unrecognized roles of candidate divisions, Desulfarculales, and Desulfuromandales in sulfur cycling, carbon fixation and fermentation within anoxic basin waters. Taken together, filter size and in situ versus on-ship filtration had the largest impact on recovery of microbial groups with the potential to influence downstream metabolic reconstruction and process rate measurements. These observations highlight the need for establishing standardized and reproducible techniques that facilitate cross-scale comparisons and more accurately assess in situ activities of microbial communities.

Description: This work was performed under the auspices of the Scientific Committee on Oceanographic Research (SCOR), the United States Department of Energy (DOE) Joint Genome Institute, an Office of Science User Facility, supported by the Office of Science of the United States Department of Energy under Contract DE-AC02- 05CH11231, the G. Unger Vetlesen and Ambrose Monell Foundations, the Tula Foundation-funded Centre for Microbial Diversity and Evolution, the Natural Sciences and Engineering Research Council of Canada, Genome British Columbia, the Canada Foundation for Innovation, and the Canadian Institute for Advanced Research through grants awarded to SH. McLane Research Laboratories and Connie Lovejoy contributed access to instrumentation for field work. Ship time support was provided by NSERC between 2007 and 2014 through grants awarded to SC, SH and Philippe Tortell MT-B was funded by Consejo Nacional de Ciencia y Tecnología (CONACyT) and the Tula Foundation.