H2S events in the Peruvian oxygen minimum zone facilitate enhanced dissolved Fe concentrations

Dissolved iron (DFe) concentrations in oxygen minimum zones (OMZs) of Eastern Boundary Upwelling Systems are enhanced as a result of high supply rates from anoxic sediments. However, pronounced variations in DFe concentrations in anoxic coastal waters of the Peruvian OMZ indicate that there are factors in addition to dissolved oxygen concentrations (O2) that control Fe cycling. Our study demonstrates that sediment-derived reduced Fe (Fe(II)) forms the main DFe fraction in the anoxic/euxinic water column off Peru, which is responsible for DFe accumulations of up to 200 nmol L−1. Lowest DFe values were observed in anoxic shelf waters in the presence of nitrate and nitrite. This reflects oxidation of sediment-sourced Fe(II) associated with nitrate/nitrite reduction and subsequent removal as particulate Fe(III) oxyhydroxides. Unexpectedly, the highest DFe levels were observed in waters with elevated concentrations of hydrogen sulfide (up to 4 µmol L−1) and correspondingly depleted nitrate/nitrite concentrations (<0.18 µmol L−1). Under these conditions, Fe removal was reduced through stabilization of Fe(II) as aqueous iron sulfide (FeSaqu) which comprises complexes (e.g., FeSH+) and clusters (e.g., Fe2S2|4H2O). Sulfidic events on the Peruvian shelf consequently enhance Fe availability, and may increase in frequency in future due to projected expansion and intensification of OMZs.

Iron (Fe) forms an important micronutrient, controlling marine phytoplankton growth and nitrogen fixation in vast regions of the global ocean [1][2][3] . Shelf sediments are recognized as an important source of Fe to coastal waters and the open ocean [4][5][6] . In particular, Fe supplied from shelf sediments in oxygen minimum zones (OMZs) results in highly elevated dissolved Fe (DFe) concentrations in coastal waters depleted in oxygen (O 2 ) to concentrations below 2 µmol L −1 7,8 , thereby providing a potential Fe source for offshore transport 9 . The strength of the Fe flux from shelf sediments is significantly enhanced in case sediments become euxinic (hydrogen sulfide (H 2 S) present) [10][11][12] .
Upwelling of nutrient-rich deep waters across the Peruvian shelf region results in extremely high primary productivity. The sediments and water column in this region are severely depleted in dissolved oxygen (O 2 ) as a result of bacterial respiration of sinking organic matter 13 , augmented by a sluggish ventilation accounting for a reduced lateral and vertical flux of oxygen into the Peruvian shelf region 14,15 . The Peruvian sediment pore water concentrations of DFe, predominantly in the reduced form Fe(II), are in the 1-30 micromolar range and hence several orders of magnitude higher than in the overlying bottom waters 16 . The steep DFe concentration gradients and anoxic conditions of the bottom waters facilitate enhanced Fe fluxes out of the sediments reaching 10 to 866 µmol DFe m −2 d −1 8 , which result in bottom water DFe concentrations of 20-300 nmol L −1 7,17 , with most of the DFe being present in the reduced form Fe(II) [18][19][20][21] .
Iron(II) is formed in anoxic sediments by the dissimilatory reduction of Fe(III) oxides as part of a sequence of microbially mediated redox reactions 22,23 . The layer of Fe(II) enrichment in the pore waters of the sediments is located above that of elevated H 2 S levels. Below the Fe(II)-rich zone, H 2 S is formed by the dissimilatory reduction of sulfate (SO 4 2− ) and reacts with dissolved Fe(II) to form a wide variety of aqueous Fe complexes ([FeSH] + , [Fe(SH) 2 ]°, [Fe(SH) 3 ] − , etc.) and clusters (e.g., Fe n S n 0 | 4H 2 O (n = 2 or 4)) (summarized as FeS aqu ) 24 followed by amorphous particulate Fe(II) monosulfides (Fe(II)S) (e.g. mackinawite (FeS m )) [25][26][27] . Aqueous Fe sulfide clusters are ligated directly to H 2 O and the structure of the cluster is similar to the basic structure of mackinawite 24  to similar structural homologies found for aqueous zinc and copper sulfide clusters, it has been suggested that the structure of the cluster in solution determines the form of the initial particulate phase 28,29 . However, mackinawite formation is thermodynamically favored when the ion activity product of H 2 S and Fe(II) exceeds the thermodynamic stability product of FeS m (log K sp = −3.6) 26 . This condition is typically met in euxinic sediments (e.g. Fe(II) >30 µmol L −1 ; H 2 S >10 µmol L −1 30 ), and represents the initial step of Fe pyrite (FeS 2 ) formation 25,27,31 .
The formation of FeS m is also reported to occur in anoxic coastal waters and has been observed in permanently euxinic bottom waters of the Framvaren fjord (Norway), where Fe(II) and H 2 S levels are in the millimolar range 32 . Because H 2 S concentrations in these waters are extremely high, Fe sulfide precipitates in the water column as framboidal pyrite 33 . It is, however, unclear whether favorable conditions for Fe sulfide precipitation also occur in the coastal OMZ regions off Peru. The Peruvian OMZ features extremely low O 2 concentrations (<50 nmol kg −1 ) 34-37 , with anoxic waters prevailing at depths between 10 to 500 m, which in near shore regions are in direct contact with shelf sediments 28 . The positioning of the OMZ over euxinic shelf sediments facilitates benthic supply of H 2 S and Fe(II), which accumulates periodically in the overlying waters 7,38 . The transient presence of H 2 S in Peruvian coastal waters have mainly been observed during the austral summer season and their occurrences have recently been associated with stagnant flow on the shelf 39 . H 2 S and DFe in the Peruvian OMZ waters have been reported to reach concentrations as high as 13 µmol L −1 and 300 nmol L −17 , respectively, during the upwelling season in austral summer and the formation of Fe(II)S minerals resulting in DFe removal has been suggested 40 .
Here we present new data from three locations with different environmental settings on the Peruvian shelf regarding DFe, Fe(II), H 2 S, nitrate (NO 3 − ), nitrite (NO 2 − ) and ammonium (NH 4 + ) (with the sum of all N species = DIN), O 2 , and dissolved inorganic phosphorous (DIP) expressed as P* (P* = DIP − DIN/16) 41 . We examine the mechanisms that control DFe in the Peruvian OMZ, under contrasting conditions of presence and absence of enhanced water column H 2 S concentrations.

Results and Discussion
Water column anoxia occurred at all three study sites during the upwelling season in December/January 2008/09 ( Fig. 1A-C). The surface mixed layer extended down to ca. 10 m depth at site 1, 20 m at site 2 and 60 m depth at site 3, and was assessed using a fixed density difference criterion (Δσ = 0.125) 42 . At all sites, O 2 concentrations decreased below the surface mixed layer to levels below the LOD of the CTD mounted O 2 sensors. At site 2, O 2 The water column at site 2 featured enhanced H 2 S concentrations, coinciding with high DFe levels ( Fig. 1B,E). However, an increase in H 2 S concentrations at site 2 was observed with depth including a mid-depth maximum at 50 m (~4 µmol L −1 ; Fig. 1E) and ~3 µmol L −1 near the seafloor. Elevated DFe concentrations of up to 300 nmol L −1 during a H 2 S event reaching ~ 10 µmol H 2 S L −1 on the Peruvian shelf in 2012 have also been reported by Scholz, et al. 7 , with DFe and H 2 S being released by anoxic sediments.
Removal of DFe through the formation of Fe(II) sulfide minerals has been reported for euxinic sediments 27 , deep-sea hydrothermal vent systems 43 , euxinic fjord waters 32 , and anoxic waters in the bottom boundary layer of the Peruvian OMZ 40 . Using Visual MINTEQ. 3.1 44 , we calculated the species distribution of dissolved Fe(II), aqueous sulfide (FeS aqu : log K = 5.62 24 ), and Fe(II)S in the crystal structure of mackinawite (log K = −3.6 24 ), an ubiquitous mineral in low temperature aqueous environments, to determine if Fe(II)S formation in the water column at site 2 was feasible ( Fig. 2) (concentrations of model parameters used are listed in supplementary material S1). At 13 °C and pH 7.65, which is typical for subsurface waters on the Peruvian shelf 45,46 , and concentrations of Fe(II) of 200 nmol L −1 and H 2 S levels of 3 µmol L −1 , which reflect the bottom water composition at site 2, the ion activity product of the educts (log IAP = − 5.466) is below the typical apparent solubility product of mackinawite of log K' sp = − 3.6 44 (undersaturation with respect to mackinawite: log IAP -log K' sp <0). To validate the robustness of the model a sensitivity test was executed by increasing pH and temperature which would increase log IAP and enhance the likelihood of mackinawite formation. At 40 °C and pH 8.5, obviously very unrealistic conditions for the Peruvian OMZ, the log IAP increased slightly to −4.61. This suggests that under the transient conditions typically encountered, the formation of mackinawite is thermodynamically unfavorable in the anoxic bottom waters off Peru.
The mackinawite formation will only commence at Fe(II) and H 2 S concentrations in the higher micromolar range. Our modelling indicates that at pH 7.65, 13 °C and 200 nmol Fe(II) L −1 more than 2 mmol L −1 H 2 S are required to facilitate the formation of mackinawite (Fig. 2). Further, noticeably less H 2 S is required (~10 µmol L −1 ) at higher Fe(II) levels of >3 µmol L −1 to attain mackinawite saturation. These required enhanced H 2  In H 2 S free seawater more than 75% of Fe(II) occurs as a truly dissolved free cation, while the remainder will form complexes with hydroxide (OH − ), carbonate (CO 3 2− ), and chloride (Cl − ) ions 52 . In the presence of an excess of H 2 S over Fe(II) and above pH 7.5, FeS aqu becomes the dominant Fe(II) species 24 (Fig. 2). Formation of thiols in sediments 53 may result in Fe(II)-thiol complexes, with thiols also facilitating reduction of Fe(III) 54 . Organic ligands complexing with Fe(III) serve a similar purpose in oxic waters to stabilize Fe 55 . In the sulfide containing Peruvian waters at site 2, the Fe distribution in the water column was therefore controlled by the total Fe flux from the sediments rather than the equilibrium concentrations of their solid phases. Prolonged sulfidic periods will therefore result in an increase in DFe concentrations in the anoxic water column, as formation of Fe(II)S precipitates like mackinawite is unfavorable at the Fe(II) and H 2 S concentrations so far observed in this region.
The formation of FeS aqu stabilizes DFe via an increased soluble pool (as opposed to oxygenated waters in which DFe is dominated by Fe(III)-ligands and colloids 56 ), and hence facilitates vertical diffusive DFe fluxes in the Peruvian OMZ. Vertical diffusive DFe fluxes were determined for site 2 by combining eddy diffusivities determined from microstructure measurements sampled in the same region during January 2012 (FS Meteor cruise M92 39 ) and DFe concentration from this study. Altogether, 102 microstructure profiles were collected at shelf stations with bottom depth between 80 m and 100 m. The microstructure shear and temperature measurements were performed using a MSS90-D profiler (S/N 32, Sea & Sun Technology). Standard processing procedures were used to determine the rate of kinetic energy dissipation of turbulence in the water column (please see Schafstall et al. 57 for a detailed description). Subsequently, eddy diffusivities were determined from ε = Γ ρ − K N 2 where N is stratification and Γ is mixing efficiency for which a value of 0.2 was used (Fig. S1) 58 . Between mid-depth waters (40 m) and the surface (10 m), an average eddy diffusivity of K ρ = 3.1 × 10 −4 m 2 s −1 was obtained (Fig. S1). We employed the method of de Jong et al. 59 8 . It indicates an important role of FeS aqu in the anoxic waters of the Peruvian OMZ, which through stabilization facilitates Fe supply to surface ocean phytoplankton communities on the shelf and possibly further afield by filaments and mesoscale eddies that move off shore, away from the coast zone 60 .
In the absence of H 2 S in the anoxic water column at sites 1 and 3, DFe concentrations were 72 to 94% lower in bottom waters than at site 2. Vertical diffusive water column fluxes calculated using a K ρ of 3.1 × 10 −4 m 2 s −1 at sites 1 and 3 were concurrently reduced by 88 to 97% (7.60 ± 4.13 µmol m −2 d −1 ), which indicates that supply and removal processes in anoxic water columns may differ and may depend on controls other than O 2 .
Organic matter remineralization processes in anoxic environments are mainly controlled by anaerobic microbial processes involving nitrogen, sulfur and Fe for electron-transfer reactions 61 . Of relevance for the Peruvian OMZ is the reduction of nitrate, coupled with the oxidation of H 2 S by filamentous bacteria of the family Desulfobulbaceae 62,63 . A similar coupling of anaerobic microbial denitrification with Fe(II) oxidation has been documented for chemolithotrophic organisms in freshwater sediments 64,65 , suboxic aquifers 66 , marine coastal sediments 67 , and for the anoxic water column off the coast of Peru, where Fe(II) oxidizers such as Marinobacter aquaeolei are active 7 .
Transient accumulations of H 2 S have been reported for anoxic Peruvian, Namibian and Indian shelf waters that were depleted in nitrate/nitrite [68][69][70][71][72] . These euxinic waters form the extreme end point of ocean conditions, with fully oxygenated waters being the opposite end point 73 . We suggest that the coupled process of anaerobic microbial denitrification and H 2 S oxidation depletes nitrate and allows DFe to accumulate in anoxic bottom waters in the Peruvian OMZ. The extent of anaerobic microbial denitrification in the Peruvian anoxic environments is mainly controlled by H 2 S levels and less by Fe(II) given that H 2 S concentrations are an order of magnitude higher than those of Fe(II).
Depending on H 2 S and nitrate concentrations, two distinct and one advanced scenarios can be envisaged (Fig. 3). Scenario 1: Under conditions of an anoxic water column (no measurable H 2 S) above the sediments and in the presence of nitrate and nitrite (i.e. denitrification and anammox has not fully removed nitrate and nitrite) bacteria at the sediment-water interface will reduce these nitrogen species and thereby utilize both H 2 S and Fe(II). During that process all the sulfide and a significant fraction of the Fe(II) will be oxidized. Insoluble Fe(III)oxyhydroxide particles are consequently formed near the sediment-bottom water interface, leading to an accumulation of reactive solid Fe phases near the sediment surface. Any Fe(II) that diffuses across the sediment-bottom water interface into the water column is then oxidized by O 2 /H 2 O 2 or, in the absence of O 2 , via nitrate-dependent Fe(II) oxidizing microbes with the Fe remaining either as organically complexed Fe(III) (FeL), or is lost by scavenging processes and insoluble particle formation, as was documented by Heller et al. 18 . This situation was observed at sites 1 and 3 (Fig. 1A-C and F) and is illustrated in Fig. 3 (left schematic).
Scenario 2: Reductive removal of the nitrate and nitrite pool in the sediment, in association with H 2 S oxidation, allows enhanced sediment-water H 2 S and Fe(II) fluxes resulting in an accumulation of these species in the water column. The oxycline in the upper water serves as a removal filter, with e.g. oxidation of Fe(II) to Fe(III) in the presence of oxygen and hydrogen peroxide 74 . This scenario is proposed for site 2 ( Fig. 1B and E) and resulted in the sulfidic event with H 2 S concentrations of up to 4 µM and Fe(II) concentrations of up to 200 nM (Fig. 3, middle schematic). We assume that such a scenario is temporarily confined. Advanced scenario 2: Long lasting release of sedimentary H 2 S and Fe(II) raises concentrations of both compounds in the water column, with highest concentrations expected at the sediment-bottom water interface. We hypothesize that under such conditions, log IAP > log K' sp , resulting in the formation of mackinawite at the sediment-bottom water interface (Fig. 3, right  schematic). Due to a lack of observational data, we do not know if such an advanced scenario can occur in the more turbulent Peruvian OMZ, or if it just arises in water bodies as stagnant as the Black Sea 49  The coupling of Fe(II) and nitrite in OMZs is under debate in the scientific community. Hong and Kester 19 found a linear relationship between nitrite and Fe(II) for samples from the OMZ along the Peruvian shelf and suggested this represented a common sediment source. More recently several trace metal studies conducted in open ocean OMZs observed a deep Fe(II) maximum that coincided with elevated nitrite and DFe (Fe(II) + Fe(III)) levels at the same depths. In the Peruvian OMZ a filament with elevated Fe(II) and nitrite concentrations in the center of the otherwise DFe rich OMZ has been located in the anoxic core zone between 300 m and 400 m water depth 9,20,75 . The filament extended from the coast to ca. 1,000 km off-shore and was related to off-shore Fe transport. A similar pattern has also been described for the OMZ in the Arabian Sea 76,77 and both were interpreted as a coupling between nitrate reduction and Fe(II) accumulation, with Fe(III) being microbially reduced 78 . In contrast, Rickard and Luther III 24 ,Scholz,et al. 7 , and Heller, et al. 18 suggested a role for nitrite in the oxidation of Fe(II) and an accumulation of particulate Fe oxyhydroxides in the anoxic water column off Peru.
Periodic sulfidic events occur in the Peruvian OMZ. Our observations indicate that the present conditions are not favorable for the formation of mackinawite in the anoxic Peruvian waters given that Fe(II) and H 2 S concentrations remain consistently too low to exceed the solubility product of Fe(II) sulfide minerals. At micromolar H 2 S concentrations, aqueous Fe sulfide complexes and clusters become the dominant Fe(II) species and buffer DFe through prevention of scavenging, thereby enhancing Fe solubility in the euxinic water column.
Oxygen minimum zones are projected to expand and intensify as a consequence of reduced oxygen solubility related to ocean warming, increased stratification of the water column 15,79 , changes in oxygen consumption via biotic respiration, and changes in the large-scale overturning circulation 80 . This will likely result in more frequent H 2 S events and associated enhanced Fe(II) concentrations in coastal OMZs 72 . For the eastern tropical South Pacific off Peru it has been shown that eddies frequently forming at the coast can transport coastal waters far offshore within days and weeks 81 . This scenario may enhance the supply of DFe to local surface waters and potentially to the Fe depleted South Pacific gyre system 3,82,83 , with positive feedbacks for primary productivity and nitrogen fixation.  (Fig. 1). The water depth was 112 m at site 1, 98 m at site 2 and 255 m at site 3. Samples for trace metal analysis were collected using trace metal clean Go-Flo bottles (General Oceanics) attached to a Kevlar wire. Go-Flo bottles were deployed once at sites 2 and 3, while at site 1 the Go-Flo bottles were deployed five times within 24 h. After recovery, bottles were transferred into a clean lab container. The seawater samples were filtered using cartridge filters (0.2 µm, Sartobran 300, Sartorius) and dispensed into acid cleaned 1 L low density polyethylene (LDPE, Nalgene) bottles. The samples were then acidified with quartz distilled hydrochloric acid to pH 1.8 (17.8 µmol H + L −1 ). Unfiltered seawater samples for on-board Fe(II) analysis were dispensed into opaque acid cleaned 60 mL LDPE bottles under normal filtered air. The Fe(II) analysis was carried out immediately after sample collection. All sample handling was performed in a laminar flow hood. Sample analysis. Dissolved Fe concentrations were determined half a year later using graphite furnace atomic absorption spectrometry (Perkin Elmer, 4100 ZL) following Grasshoff et al. 84 . The blank and limit of detection (LOD) (three times the standard deviation of the blank measurement) for Fe concentrations were 0.104 nmol L −1 and 0.079 nmol L −1 , respectively. The accuracy of the analytical procedure was evaluated by the analysis of certified seawater standard NASS-5 (National Research Council of Canada) and SAFe. Our Fe values agree well with the certified values for NASS 5 and the SAFe data (NASS 5: 26.3 ± 1.1 nmol kg −1 (certified: 25.7 ± 2.0 nmol kg −1 );SAFe S: 0.112 ± 0.013 nmol kg −1 (census: 0.093 ± 0.008 nmol kg −1 ); SAFe D2: 0.83 ± 0.13 nM Fe (census: 0.933 ± 0.023 nmol kg −1 )). The precision of the method is 3-5%.
Fe(II) concentrations were measured on samples collected at site 2 immediately upon collection by a chemiluminescence flow injection analysis following the method of Croot and Laan 85 , which has a LOD of 0.1 nmol L −1 Fe(II).
Nutrient and O 2 samples were obtained using Niskin bottles (General Oceanics) on a stainless steel CTD rosette deployed at the same locations. Nutrient samples were analyzed for NO 3 − and DIP using an autoanalyzer (TRAACS800, Bran&Lubbe) following Grasshoff et al. 86 . NO 2 − was determined spectrophotometrically 86 and NH 4 was analyzed fluorometrically 87 on board. Oxygen concentrations in the water column were measured by a Seabird O 2 sensor that was calibrated with oxygen concentrations determined from CTD water samples (Winkler method 84 ). The latter caused a LOD of 2 µmol kg −1 . Additionally, at site 2 a switchable trace O 2 sensor (STOX) with a LOD of 50 nmol kg −1 was used 36,37 . For H 2 S measurements seawater samples were collected with a pump-CTD 38 and were analyzed spectrophotometrically 88 . At the pH of seawater HS − represents the dominant hydrogen sulfide species, which is why the term H 2 S for our study refers to the sum of H 2 S, HS − , and S 2+ . Data availability. The dataset generated during this study is available in the GEOMAR-OSIS repository (https://portal.geomar.de/de/osis).