Chemical energy in persistent non-buoyant plumes above vent fields
This study reports the detection and analysis of plumes associated with active venting from two mounds rising several hundred meters above the rift valley of the Gakkel Ridge. The Aurora mound is located at the westernmost end of the Western Volcanic Zone 14,20,33,34, the newly described mound “Polaris”, at the central part of the Eastern Volcanic Zone 35 (Fig. 1). In both areas, year-round full sea-ice coverage had prevented thorough studies of the source of the plumes prior to the RV Polarstern expeditions PS86 and PS101. Our main survey instruments were ocean floor observing systems (PS86:OFOS; PS101:OFOBS) for seafloor morphology and geobiology 36, and a CTD (conductivity, temperature, depth) rosette that was equipped with highly sensitive redox and turbidity sensors, 24 Niskin bottle samplers and in situ pumps. To survey the vent fields and sample their plumes, the icebreaker Polarstern had to drift with the ice floes across specific locations according to predicted paths 33,35. Numerous CTD casts were performed in tow-yo mode to assess the size and extent of the plumes. Wherever plumes were encountered, the water column was sampled with Niskin bottles and microorganisms and particles were sampled on filters with high volume in situ pumps. Additional dives with the OFOS/ OFOBS and were performed to inspect the seafloor directly under the plumes, especially in locations where buoyant hydrothermal plume signals were intercepted. Seafloor images collected with both system are available from the PANGAEA database, examples of images in which microbial life were visible are given in Supplementary Fig. 1 and the coordinates of all such observations across the Aurora and Polaris mounds from the PS86 and PS101 expeditions are given in Supplementary Table 1.
At the Aurora mound a Black Smoker vent field was first discovered during the PS86 expedition in 2014 (82°53.83’N; 6°15.32’W, in 3900 m water depth) at the southwestern flank of the mound (Fig. 1d-f) 33. Since then Aurora was revisited by other expeditions 26,34,37. The giant, highly active chimneys are black to bright yellow-orange, sulfidic and release substantial amounts of metal sulfides, visible as black particles. The surroundings of the Aurora vent field showed basalt rocks, populated by glass sponges, amphipods, limpets and snails of much higher density than in the surroundings, or at other mounds of this part of Gakkel Ridge (Boetius 2015). Near the vent field, a number of cracks and fissures with diffuse vents and dead chimneys were discovered, the latter overgrown with sponges (Fig 2e-h).
In 2016, during PS101, we visited the Eastern Volcanic Zone to investigate the source of a large hydrothermal plume hovering above the NW flank of a previously unnamed mound discovered 15 years earlier 14,20. We named this structure Polaris mound (86°87.5’N, 55°42.4’E; 3170 m water depth) (Fig. 1a-c). For further details, see Albers et al., (submitted). The NW flank was characterized by steep terraces with pillow basalts covered by minor gravel and discolored sediments, associated with deep fissures, yellow and orange bacterial mats, numerous fish skeletons, as well as abundant orange sulfur precipitates associated with small black chimneys emitting clear shimmering fluids (Fig 2 a-d). In the vicinity of the small vents, we observed an anomalous density of ophiuroids and polychaetes as well as sulfidic bacterial mats. The peak of the Polaris mound consisted of barely-sedimented pillow basalts, with abundant sea anemones. This are hosted only a few glass sponges, in contrast to the Aurora mound, where sponges were the dominant fauna in the vicinity of the vent field.
At Polaris, in situ sensors mounted on the OFOBS instrument intercepted an upwelling plume, which also spread horizontally between 400 to 800 m above the seafloor. Having surveyed approximately a third of the mound, the plume could not be identified as originating from a distinct large fluid source at the seafloor, with only a wide field of small vents, fissures and hydrothermal precipitates imaged during the expedition. The non-buoyant component of the hydrothermal plume assessed by tow-yo CTD was characterized by sharp anomalies in turbidity, redox and temperature extending from a depth of >2800 meters below sea level (mbsl) to ca. 2400 mbsl, with a horizontal stretch of over 1200 m according to the turbidity signal (Fig. 1 b). The CTD cast PS101-226 (86° 57.41 N, 55° 44.42 E), intercepted the buoyant part of plume a short distance above the seafloor (~2900m) with a temperature anomalies up to 160 mK above ambient seawater (−0.8 °C), suggesting a vigorous vent field near this position at the NW flank of the mound. Based on the plume rise-height and the measured stratification of the water column, we calculate a power of 130 MW, which at temperatures of 270° C translates into a water flux of approx. 11600 m3 per day (for calculation see Supplementary Discussion 1). The observed maximal redox anomaly was relatively low with −25 mV, and this redox signal disappeared faster than the turbidity signal, which was substantially higher at Polaris than at Aurora (Fig. 1 b, 3a). The low redox signal can be explained by the absence of reduced iron and manganese in the plume (Table 1). The Polaris plume was strongly enriched in 3He with δ3He values of up to 75% compared to atmospheric helium standards, indicating a substantial concentration of mantle-derived gas in the rising fluids (Fig 3b). The seawater samples from the buoyant part of the plume contained up to 300 nM CH4 and 360 nM H2 (Fig. 3c,d). With a temperature anomaly of >60 mK during the closure of the Niskin bottle, the fluids were diluted by at least a factor of 1:4500, assuming that based on the CH4/H2 ratio the fluid endmember should not be hotter than 270°C (see Table 1). This suggests that the original vent fluids contained about 1.35 mM CH4 and 1.6 mM H2. Based on the calculated fluid flux rates, the Polaris vents would emit about 25,600 mol hydrogen and 21,600 mol methane per day to feed the observed plume. We detected neither dissolved nor precipitated iron (detection limit between 10 and 50 µM), and also sulfide was analytically undetectable in the plume whilst using the Cline assay (detection limit ~10 µM). However, several plume samples had a noticeable sulfidic smell, translating to several 100 nM sulfide, which suggests that total sulfide emissions from the Polaris vents could be in a similar range as hydrogen and methane emissions 38.
At the Aurora mound, the plume also had a strong turbidity signal that could be followed over a distance of about 2 km, and for more than 800 m vertically (Fig. 1 d-f). Using OFOS, we were able to link the origin of this plume to a field of black smokers with brightly colored, yellow-orange chimneys at the southwestern flank of the mound, (Fig. 2) 34. Based on the observed plume characteristics we estimate that the vents have power of 24 MW (for calculation see Supplementary Discussion 1). Endmember fluids of volcanically-hosted black smokers at ultraslow spreading ridges typically have temperatures of 370°C 1,2,39,40), which would result in fluid fluxes of 1400 m3 per day to feed the observed plume. In all retrieved samples the temperature anomalies were low (< 8 mK), translating to dilution factors of ≥45×103 for all samples. This strong dilution would be entirely consistent with the Aurora plume 3He anomaly, which was rather small compared to Polaris, with δ3He values ≤10% (Fig. 1 g). By contrast, the redox anomaly at Aurora was much larger than that measured in the Polaris plume. A high particle load in the plume water matched the strong optical backscatter signals. The pronounced redox anomaly is consistent with the high concentrations of dissolved iron and manganese in the plume, derived from the large black smoker field at Aurora (Fig. 3f) 34. In situ filtration of the water yielded a yellowish residue, suggesting partial precipitation of elemental sulfur and metal sulfides. The calculated concentrations of metals in the vent fluid endmember (6.8 mM Fe, 0.7 mM Mn,) would thus be underestimated, because a fraction may have precipitated from the plume as polymetallic sulfides. Dissolved sulfide was not detectable, and the plume waters did not smell. The plume contained up to 30 nM methane 34, which translates 1.4 mM methane in the fluid endmember. Hydrogen concentrations were not measured on board. Because of underlying thermodynamics, the molar H2-CH4 ratio of the fluid endmember would be ≥10. Based on the calculated fluid volume flux rates (see earlier), we can calculate corresponding geochemical fluxes from the Aurora vents (Table 1): ≥ 17,000 mol H2 and >1,700 mol CH4 per day; >8000 mol Fe per day (168 tons per year) and ≥850 mol of Mn per day (17 tons per year).
Pronounced hydrogen metabolism in the plumes
To quantify the influence of the energy sources on the carbon fixation by the plume microbiota we collected water samples from the Polaris and the Aurora plumes, from their local background waters (above and below each plume) and from deep-sea reference water (>5 nautical miles from the vents). We incubated these waters with 14C-bicarbonate at in situ temperatures (−1 °C), and measured the transfer of the radioisotope into the particulate carbon fraction41. Both, reference waters distant from the vent, and local background water samples from above and below plume, showed carbon fixation rates between 0.5 to 1 µmol C m−3 d−1 (Fig. 4). These are typical values for dark carbon fixation in the oligotrophic deep ocean, as fueled by trace amounts of organic matter and ammonia oxidation 41. In contrast, plume water samples from both sites with notable in situ redox anomalies exhibited much higher carbon fixation rates between 5 to 45 µmol m−3 d−1. Assuming a mean carbon fixation of 15 µmol m−3, and a carbon content of 10 fmol per cell 42, an average of 1.5×109 autotrophic microorganisms could be formed per cubic meter of plume water each day. This translates into a maximal increase of cell mass of 60 % per day. This value matches well with the moderately increased cell numbers (Kruskal-Wallis chi-squared = 0.18182, df = 1, p-value = 0.6698). Only one of the plume samples at Polaris (PS101/139) had 3 times elevated cell numbers compared to the average in the background (Fig. 3e).
To evaluate the microbial metabolism associated with persistent plume waters, we tracked the development of hydrogen and methane concentrations in incubated waters from the hydrothermal plume, measured rates of dark carbon fixation (DCF) and analyzed the microbial community composition from both plumes and reference samples. (Fig. 5-7). Within the different plume water samples, the concentrations of methane correlated with the helium isotope ratios, which suggests pure dilution and no consumption for methane during plume dispersion 43. In contrast the hydrogen concentrations dropped faster than the helium isotope ratios (Fig. 3; Supplementary Fig. 2), indicating the microbial consumption of hydrogen. To further compare the microbial consumption of methane and hydrogen, replicate bottles of buoyant plume water, (3500 m water depth) and non-buoyant plume waters (3000 m water depth) from the Polaris vent and surrounding background waters were incubated with trace amounts of hydrogen and methane at in situ temperatures of −0.8°C (Fig. 5). In the background water (station PS101-175-5), concentrations of hydrogen (165 nM +/-8) and methane (31 nM; +/-3) remained stable throughout incubation times of up to 5 days, i.e. neither energy substrates were consumed by microorganisms. Similarly, in the buoyant plume waters, the natural concentrations of hydrogen (323 nM) and methane (272 nM) remained stable (Fig. 5a,b). In contrast, in samples of the non-buoyant plume, hydrogen concentrations of 150 nM decreased to 22 nM within 2 days, whereas methane remained stable for the entire observation period of five days. (Fig 5c). We explain the absence of hydrogenotrophic activity in the fluid-enriched buoyant plume sample with the quick rising period of the plume of about one hour 44. Within this short time frame, the ambient microbial communities were unlikely to respond to the presence of hydrogen or other reduced compounds. Furthermore, if transported with the fluids, microorganisms living in or at the vent chimney may not be adapted to temperatures of −0.8°C.. Aerobic hydrogen oxidizers have growth efficiencies of 25% to 30% of the reducing equivalents released during hydrogen oxidation 45. Hence, the hydrogen oxidation rate observed in our experiment may result in a carbon fixation of 15 to 20 µmol m−3 which may translate into the formation of 2×106 cells per liter. This increase is confirmed by our 16S rRNA analysis, which shows a bloom of hydrogenotrophic communities in the non-buoyant plume water. These hydrogen oxidizers must be recruited from the background ocean water column. This water mass contained bacteria such as SUP05 and Sulfurimonas, which can grow rapidly whenever they are exposed to their energy substrates methane or hydrogen 30,31. In contrast, methane-oxidizing bacteria that are also found in background seawater, do not seem to benefit from the supply of their substrate at the ice-cold temperatures of the Arctic Ocean, and consequently do not become enriched in the plume. We confirmed the absence of methanotrophy in the plumes using 14C-methane tracer assays, which also indicated that there was no methane oxidation in either the Aurora or the Polaris waters (see Supplementary Table 2). To our knowledge, similar experimental evidence for the fate of hydrogen and methane does not exist for any other vent plume. The influence of reduced metals and sulfide cannot be examined in similar experiments, because these oxidize abiotically in oxygen-saturated Arctic Ocean. Instead, we compared community-based genetic information to address the role of reduced sulfur as an energy source (see below)30,31,46,47.
We further analyzed the diversity of 16S rRNA genes and gene transcripts extracted from plume, local background and reference waters (Fig. 6, Fig.7, Supplementary Fig. 3). In the Polaris non-buoyant plume characterized by the highest carbon fixing activity, the most abundant taxon was SUP05 with 9 % (PS101/139) to 47% (PS101/159) of total 16S rRNA gene sequences (average of 25 % across all plume samples). In the background water and at reference stations this taxon had a much lower relative abundance of 1−9% and 3−4%, respectively. Sulfurimonas at Polaris contributed 4 to 19% in the non-buoyant plume compared to <1% in reference samples (Fig. 6). Most of the other active fraction of microbial communities was similar at both vents (Supplementary Fig. 3). In contrast, in the non-buoyant Aurora plume, Sulfurimonas dominated the microbial communities with 16% to 66% (PS86/055; Fig. 6) of bacterial 16S rRNA gene sequences and 69% to 79% of all SSU rRNA reads (PS86/055; Fig.6, Supplementary Fig. 3). SUP05 reached on average 10 % and 8% in the plume, of total and active community members, respectively (Fig.6 and Supplementary Fig. 3). The significant enrichments of SUP05 and Sulfurimonas (i.e. US. pluma) cells in Arctic plume waters were confirmed by microscopy applying specific oligonucleotide-probes, reaching values of 6−12% and 2−20%, respectively (Supplementary Table 3; Kruskal-Wallis chi-squared = 6.6, df = 1, p-value = 0.01). In the background waters Sulfurimonas still accounted for up to 2−10 % of all SSU rRNA reads waters (PS86/074 at 2500 m), and SUP05 was 2%. These numbers for Sulfurimonas appear very high and suggest either that Arctic deep-sea waters may be thoroughly mixed with hydrothermal waters from Gakkel Ridge 14, or that the mortality pressure is very low.
In both plumes, the increase of specific taxa resulted in a significant decrease of evenness as described by exponential Shannon entropy and inverse Simpson index, while the overall species richness of the water types was not affected (Supplementary Fig. 4,5, Supplementary Table 4). Aurora and Polaris plumes shared 36±6% of bacterial 16S rRNA gene sequences with reference seawater, with lowest values at station PS101/177 (25±2%) and PS86/055 (29±1%; Supplementary Table 5). On overage the two plumes shared 32±4% of the 16S rRNA genes, which is slightly lower than the value of 16S rRNA genes shared among the different non-plume waters (40±1%; Supplementary Table 5). As previously reported by Molari et al. (2023), phylogenetic comparison of the Sulfurimonas SSU rRNA gene sequences from both hydrothermal plumes revealed two main and distinct populations that share >99% SSU rRNA nucleotide sequence identity. Similarly, the analysis of V3−V4 region of the 16S rRNA gene of SUP05 revealed two ribotypes (similarity >98%). One ribotype dominates at Polaris, and the other is more dominant at Aurora (Supplementary Table 6). These results confirm the presence of two species of SUP05, as depicted by 16S rRNA comparison and three metagenomic assembled genomes obtained from Aurora and Polaris water samples (Scillipoti and Molari, submitted). Based on phylogenetic analysis of their 16S rRNA, one of the two SUP05 strains clusters with SUP05 sequences from Pacific Ocean hydrothermal plumes, and the other species was more similar to SUP05 sequences from the Atlantic and the Pacific oxygen minimum zone (Supplementary Fig. 5). In addition, some less abundant taxa show differential abundance. For instance, the Polaris plume was enriched in Oleispira that include alkane oxidizers from the Arctic Ocean 48) and methane oxidizers of the Methylococcales cluster Milano-WD1B-03 (Supplementary Fig. 3).
Transcripts for carbon fixation of Sulfurimonas via reverse tricarboxylic acid cycle (i.e. aclA, aclB) are highly enriched at Aurora, whereas at Polaris the transcripts of SUP05 for carbon fixation via the ribulose monophosphate pathway (rbcL) are enriched. This is in line with the dominance of Sulfurimonas and SUP05 in Aurora and Polaris, respectively. Genes for hydrogen and sulfur metabolism were highly expressed in the Polaris and the Aurora plume, but the transcription levels differed between the two plumes and the abundant taxa (Fig. 7). In both the Aurora and Polaris plume, the hydrogenase transcripts assigned to Sulfurimonas were strongly enriched. This is the prototypical oxygen-sensitive hydrogenase of hydrogen oxidizers isolated from hydrothermal vents and others redoxcline environments 31,49,50 (Supplementary Fig. 7), which supports the dominant role of hydrogen oxidation at these Arctic vents (Fig. 7). At Polaris the transcripts of SUP05 encoding the sulfur oxidation pathway, including sulfur oxidation to sulfate (dsrA, aprA) and thiosulfate oxidation to sulfate (soxA, SoxB), were also highly enriched.
The dataset also contains transcripts of the groups 1d and 1l [NiFe]-hydrogenase that belong to SUP05. The group 1d was highly enriched in Polaris plume, but largely depleted in Aurora plume. In contrast, the [NiFe]-hydrogenases group 1l was similarly expressed in both plumes (Fig. 7). These two hydrogenases most likely belong to two different SUP05 strains (Supplementary Fig. 6,7). The group 1d is the canonical enzyme for aerobic hydrogen oxidation and it catalyzes hydrogen oxidation in the SUP05 symbionts of Bathymodiolus mussels from hydrothermal vents 7. Recently, Lappan and coworkers reported that 1l [NiFe]-hydrogenases are widespread in marine microorganisms, however their function in many of these organisms remain unclear 49. SUP05 has been shown to grow on reduced sulfur compounds 51, and it was speculated that it also utilizes hydrogen 52. Yet, several SUP05 MAGs from hydrothermal plumes do not code for [NiFe]-hydrogenases 30,47 and thus it is unlikely that hydrogen is a pronounced energy substrate of SUP05 bacteria 53.
Puzzled by the lack of evidence for microbial oxidation of methane, we also looked for the transcription of methane monooxygenases (pmoA). Surprisingly, in both plumes pmoA transcripts were somewhat enriched, but this did not coincide with any measurable oxidation of methane. Also, we did not identify an enrichment of transcript related to iron oxidation within the pool of highly transcribed genes at Polaris and Aurora, suggesting a minor importance of microbial metal oxidation. 16,30. Only some metal related metabolic genes assigned to Sulfurimonas (cft, arsC) were highly transcribed in the plumes, indicating the importance of particulate metals for the physiology of Sulfurimonas 31. Overall, transcripts indicating hydrogen metabolism were more enriched in Aurora, supporting a role for hydrogen as the main energy source for the microorganisms in that plume. In the Polaris plume the transcripts coding for sulfur oxidation were six times more enriched than at Aurora. Thus, sulfur oxidation likely provides a substantial fraction of the energy for microbial growth at Polaris.