Sampling and Processing Methods Impact Microbial Community Structure and Potential Activity in a Seasonally Anoxic Fjord: Saanich Inlet, British Columbia

2019
The Scientific Committee on Oceanographic Research (SCOR) Working Group 144 Microbial Community Responses to Ocean Deoxygenationworkshop held in Vancouver, B.C on July 2014 had the primary objective of initiating a process to standardize operating proceduresfor compatible process rate and multi-omic (DNA, RNA, protein and metabolite) data collection in marine oxygen minimum zones(OMZs) 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 situversus 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 rRNAgene (rDNA) amplicon sequencing followed by downstream statistical and visual analyses. Results from these analyses showed that significant community shifts occurred between in situversus on ship processed samples. For example, Bacteroidetes, Alphaproteobacteriaand Opisthokonta associated with on-ship filtration onto 0.4 µm filters increased 5-fold compared to on-ship in-line 0.22 µm filters or 0.4 µm filters processed and preserved in situ. In contrast, Planctomycetesassociated with 0.4 µm in situfilters increased 5-fold compared to on-ship filtration onto 0.4 µm filters and on-ship in-line 0.22 µm filters. In addition, candidate divisionsand 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 fixationand fermentation within anoxic basin waters. Taken together, filter size and in situversus 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 situactivities of microbial communities.
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