<rahmann et al.
(19°C) using Shimadzu® 1800 UV—VIS double—-beam
spectrophotometer within 230-750 nm wavelength range against
MilliQ water at 1 nm intervals. More details on the spectroscopic
analyses may be found in Loginova et al. (2015, 2016, 2020).
Microbial Oxygen Consumption, Nitrogen
Transformation, and Primary Productivity Rates
Jinitrogen (N2) and carbon (C) fixation rates were measured
an nine cruises (Löscher and Mehrtens, 2021b; see Table 2
and Supplementary Table 17) using shipboard incubation
experiments, complemented with nutrient and oxygen
manipulations. During cruises M77/3, M77/4, and M80/2,
N2 fixation was measured using the bubble addition method
following Montoya et al. (1996). During M80/2 a novel method
based on !°N; gas pre-dissolution, which was developed by
Mobr et al. (2010), was tested in parallel to the method of
Montoya et al. (1996). An underestimation of N fixation rates
by the classic method has been observed (Großkopf et al., 2012)
and therefore the novel ‘pre-dissolution method’ was applied
during the following cruises (M83/1, M90, M91, M93, M97,
M104, and M107). Single cell N, fixation rates to differentiate
the contribution of different clades of N, fixers were measured
using a NanoSIMS (Martinez-Perez et al., 2016). C fixation was
determined using !*C-bicarbonate additions (e.g., Großkopf
at al., 2012; Löscher et al., 2014) and heterotrophic C turnover
was determined using !*C-glucose additions (Löscher et al.,
2014, 2016).
Potential rates for microaerobic respiration and aerobic
organic matter degradation as a source of ammonia (NH4") in
the Peruvian OMZ was assessed using an !®O>; labeling approach
suitable for microaerobic respiration (Holtappels et al., 2014).
Further, the effects of O» depletion associated with marine snow
particles on microbial respiration was explored by combining
802 labeling experiments with in situ particle size analysis
and modeling of aggregate-size dependent respiration (Kalvelage
2t al., 2015). Anammox, denitrification, and nitrification, as
well as NO production rates were measured on several cruises
(Kalvelage et al., 2011; Löscher et al., 2012; Bourbonnais et al.,
2017; Callbeck et al., 2017; Frey et al., 2020; Löscher and
Mehrtens, 2021a; see Table 2 and Supplementary Table 18)
using isotope fractionation studies, N tracer additions, and
inhibitor studies.
Marine Microbial Diversity Function
[n order to identify key groups of microbes for C, N,
and O>» turnover, microbial metabolic rate measurements
were complemented with analyses of metagenomes and
metatranscriptomes from the Eastern Tropical South Pacific
(ETSP) and Eastern Tropical North Atlantic (ETNA). In addition,
key gene and transcript characterization and quantification for
aerobic respiration (Kalvelage et al., 2015), N2 fixation (Großkopf
zt al., 2012; Löscher et al., 2014, 2015, 2016, 2020), anammox,
denitrification, and nitrification (Löscher et al., 2012, 2015, 2016;
Kalvelage et al., 2013) were carried out using Sanger sequencing
and quantitative real time polymerase chain reactions (PCRs) as
described in Löscher et al. (2012, 2014). To assure high quality
sampling of nucleic acids, sample filtration times did not exceed
-rontiers in Marine Science | www.frontiersin.orc
SFB754 Data Legacy
20 min and samples were shock-frozen in liquid N, and stored
at —80°C (e.g., Löscher et al., 2014). Early metagenomic and
-transcriptomic analyses targeted an understanding of microbial
communities in the surface waters above the OMZ, the oxyclines,
OMZ core waters, and sulfidic anoxic waters, as summarized
in Löscher et al. (2016) and were based on Pyrosequencing
technology (e.g., Desai et al., 2013; Schunck et al., 2013). Due to
‘he rapid advance in sequencing technologies, it was possible to
generate more conclusive metagenomes for targeted studies on
sulfur, N, and O, cycling during M90-M93. Nine metagenomes
were sequenced using Illumina HiSeq technology (Callbeck et al.,
2018) from those cruises. On those data sets, genome assemblies
and phylogenetic classifications were carried out to explore the
role of a key microbial cluster, SUP05, and its role in OMZ
sulfur and nitrogen turnover. Metagenomes from the ETNA
cruise M107 were sequenced in the context of the development
of anoxic water masses in collaboration with the DFG-funded
Cluster of Excellence “The Future Ocean’ (Löscher et al., 2015). In
addition to full metagenomes, targeted community studies were
carried out using 16S rDNA amplicon sequencing sequenced
on Illumina MiSeq sequencers from the same anoxic eddy in
the ETNA and from the Peruvian OMZ (Löscher et al., 2015;
Scholz et al., 2016). All published sequences were submitted to
ihe National Center for Biotechnology Information’s archives
(NCBI'; see Table 2 and Supplementary Table 19). Physical
DNA libraries are archived at GEOMAR and the University of
Southern Denmark, and subsamples are available on request
from C. Löscher.
In addition to this mainly pelagic work, transcriptomes, and
genomes of the denitrifying benthic foraminifera Globobulimina
turgida and G. auriculata from the seasonally hypoxic Swedish
Gullmar Fjord were analyzed (Woehle et al., 2018). The
obtained information was used to describe the foraminifera
unique eukaryotic ability to denitrify and colonize low-oxygen
environments. Sequences were submitted to the NCBI's
Sequence Read Archive (accession numbers SRR6202052 -
S$RR6202078) and to the transcriptome sequencing archive
(accession numbers GGCE00000000 and GGCD00000000).
The genome assembly was submitted to NCBI (draft
genomes PIVH00000000- PIWH00000000; unassigned contigs:
PJEL00000000). Furthermore, individually amplified 18S$ rRNA
gene sequences of the two analyzed foraminiferal species were
submitted to GenBank (MG800664 to MG800667).
Zooplankton and Particle Distribution
A Hydrobios Multinet Midi with an aperture of 0.25 m?
and five nets (mesh size 200 wm) was deployed for vertically
stratified tows on several cruises (Hauss et al., 2021la; see
Table 2 and Supplementary Table 20), mostly in paired day-
night tows to quantify diel vertical migration. Standard depths
used for these deployments were 1000-600-300-200-100-0 m.
On cruise M93, a Multinet Maxi (nine nets, 333 wm mesh)
was used instead. Samples were fixated in 4% formaldehyde
in seawater solution, scanned at GEOMAR or at the Ocean
Science Center Mindelo, Mindelo/Cape Verde, and analyzed
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