Phytoplanktonic species in the haloalkaline Lake Dziani Dzaha select their archaeal microbiome.

Microorganisms are key contributors of aquatic biogeochemical cycles but their microscale ecology remains largely unexplored, especially interactions occurring between phytoplankton and microorganisms in the phycosphere, that is the region immediately surrounding phytoplankton cells. The current study aimed to provide evidence of the phycosphere taking advantage of a unique hypersaline, hyperalkaline ecosystem, Lake Dziani Dzaha (Mayotte), where two phytoplanktonic species permanently co-dominate: a cyanobacterium, Arthrospira fusiformis, and a green microalga, Picocystis salinarum. To assay phycospheric microbial diversity from in situ sampling, we set up a flow cytometry cell-sorting methodology for both phytoplanktonic populations, coupled with metabarcoding and comparative microbiome diversity. We focused on archaeal communities as they represent a non-negligible part of the phycospheric diversity, however their role is poorly understood. This work is the first which successfully explores in situ archaeal diversity distribution showing contrasted phycospheric compositions, with P. salinarum phycosphere notably enriched in Woesearchaeales OTUs while A. fusiformis phycosphere was enriched in methanogenic lineages affiliated OTUs such as Methanomicrobiales or Methanofastidiosales. Most archaeal OTUs, including Woesearchaeales considered in literature as symbionts, were either ubiquitous or specific of the free-living microbiome (i.e. present in the 3-0.2 µm fraction). Seminally, several archaeal OTUs were enriched from the free-living microbiome to the phytoplankton phycospheres, suggesting (i) either the inhibition or decrease of other OTUs, or (ii) the selection of specific OTUs resulting from the physical influence of phytoplanktonic species on surrounding Archaea.

The Dziani Dzaha Lake: A long-awaited modern analogue for superheavy pyrites.

Sedimentary records of superheavy pyrites in Phanerozoic and Proterozoic successions (i.e., extremely positive δ34 Spyrite values together with higher δ34 Spyrite than coeval δ34 SCAS ) are mostly interpreted as resulting either from secondary postdepositional processes or from multiple redox reactions between sulfate and sulfide in stratified sulfate-poor environments. We report here the first observation of strongly positive δ34 S values for both dissolved sulfate and sulfide (average δ34 Sdiss.sulfate value of 34.6‰ and δ34 Sdiss.sulfide values of 36.7‰) compared to the present-day seawater δ34 Sdiss .sulfate (~21‰), with a negative apparent fractionation between sulfate and sulfide (∆34 Sdiss.sulfate-diss.sulfide ~ -2.1 ± 1.4‰), in the sulfate-poor (<3 mm) modern thalassohaline lacustrine system Dziani Dzaha (Mayotte, Indian Ocean). Overall, surface sediments faithfully record the water column isotopic signatures including a mainly negative ∆34 Ssed.sulfate-sed.sulfide (-4.98 ± 4.5‰), corresponding to the definition of superheavy pyrite documented in the rock record. We propose that in the Dziani Dzaha this superheavy pyrite signature results from a two-stage evolution of the sulfur biogeochemical cycle. In a first stage, the sulfur cycle would have been dominated by sulfate from initially sulfate-rich marine waters. Overtime, Raleigh distillation by microbial sulfate reduction coupled with sulfide burial in the sediment would have progressively enriched in 34 S the water column residual sulfate. In a second still active stage, quantitative sulfate reduction not only occurs below the halocline during stratified periods but also in the whole water column during fully anoxic episodes. Sulfates are then regenerated by partial oxidation of sulfides as the oxic-anoxic interface moves downward. These results demonstrate that the atypical superheavy pyrite isotope signature does not necessarily require postdepositional or secondary oxidative processes and can result from primary processes in restricted sulfate-poor and highly productive environments analogous to the Dziani Dzaha.

Influence of aphotic haloclines and euxinia on organic biomarkers and microbial communities in a thalassohaline and alkaline volcanic crater lake.

Studies on microbial communities, and their associated organic biomarkers, that are found thriving in the aphotic euxinic waters in modern stratified ecosystems are scarce compared to those undertaken in euxinic photic zones. The Dziani Dzaha (Mayotte, Indian Ocean) is a tropical, saline, alkaline crater lake that has recently been presented as a modern analog of Proterozoic Oceans due to its thalassohaline classification (having water of marine origin) and specific biogeochemical characteristics. Continuous intense photosynthetic production and microbial mineralization keep most of the water column permanently aphotic and anoxic preventing the development of a euxinic (sulfidic and anoxic) photic zone despite a high sulfide/sulfate ratio and the presence of permanent or seasonal haloclines. In this study, the molecular composition of the organic matter in Lake Dziani Dzaha was investigated and compared to the microbial diversity evaluated through 16S rRNA gene amplicon sequencing, over two contrasting seasons (rainy vs. dry) that influence water column stratification. Depth profiles of organic biomarker concentrations (chlorophyll-a and lipid biomarkers) and bacterial and archaeal OTU abundances appeared to be strongly dependent on the presence of aphotic haloclines and euxinia. OTU abundances revealed the importance of specific haloalkaliphilic bacterial and archaeal assemblages in phytoplanktonic biomass recycling and the biogeochemical functioning of the lake, suggesting new haloalkaline non-phototrophic anaerobic microbial precursors for some of the lipid biomarkers. Uncultured Firmicutes from the family Syntrophomonadaceae (Clostridiales), and Bacteroidetes from the ML635J-40 aquatic group, emerged as abundant chemotrophic bacterial members in the anoxic or euxinic waters and were probably responsible for the production of short-chain n-alkenes, wax esters, diplopterol, and tetrahymanol. Halocline-dependent euxinia also had a strong impact on the archaeal community which was dominated by Woesearchaeota in the sulfide-free waters. In the euxinic waters, methanogenic Euryarchaeota from the Methanomicrobia, Thermoplasmata, and WSA2 classes dominated and were likely at the origin of common hydrocarbon biomarkers of methanogens (phytane, pentamethyl-eicosenes, and partially hydrogenated squalene).

Carbon isotope evidence for large methane emissions to the Proterozoic atmosphere.

The Proterozoic Era records two periods of abundant positive carbon isotope excursions (CIEs), conventionally interpreted as resulting from increased organic carbon burial and leading to Earth's surface oxygenation. As strong spatial variations in the amplitude and duration of these excursions are uncovered, this interpretation is challenged. Here, by studying the carbon cycle in the Dziani Dzaha Lake, we propose that they could be due to regionally variable methane emissions to the atmosphere. This lake presents carbon isotope signatures deviated by ~ + 12‰ compared to the modern ocean and shares a unique combination of analogies with putative Proterozoic lakes, interior seas or restricted epireic seas. A simple box model of its Carbon cycle demonstrates that its current isotopic signatures are due to high primary productivity, efficiently mineralized by methanogenesis, and to subsequent methane emissions to the atmosphere. By analogy, these results might allow the reinterpretation of some positive CIEs as at least partly due to regionally large methane emissions. This supports the view that methane may have been a major greenhouse gas during the Proterozoic Era, keeping the Earth from major glaciations, especially during periods of positive CIEs, when increased organic carbon burial would have drowned down atmospheric CO2.

Spatiotemporal variations in microbial diversity across the three domains of life in a tropical thalassohaline lake (Dziani Dzaha, Mayotte Island).

Thalassohaline ecosystems are hypersaline environments originating from seawater in which sodium chloride is the most abundant salt and the pH is alkaline. Studies focusing on microbial diversity in thalassohaline lakes are still scarce compared with those on athalassohaline lakes such as soda lakes that have no marine origin. In this work, we investigated multiple facets of bacterial, archaeal and eukaryotic diversity in the thalassohaline Lake Dziani Dzaha using a metabarcoding approach. We showed that bacterial and archaeal diversity were mainly affected by contrasting physicochemical conditions retrieved at different depths. While photosynthetic microorganisms were dominant in surface layers, chemotrophic phyla (Firmicutes or Bacteroidetes) and archaeal methanogens dominated deeper layers. In contrast, eukaryotic diversity was constant regardless of depth and was affected by seasonality. A detailed focus on eukaryotic communities showed that this constant diversity profile was the consequence of the high predominance of Picocystis salinarum, while nondominant eukaryotic groups displayed seasonal diversity turnover. Altogether, our results provided an extensive description of the diversity of the three domains of life in an unexplored extreme environment and showed clear differences in the responses of prokaryotic and eukaryotic communities to environmental conditions.

Key Role of Alphaproteobacteria and Cyanobacteria in the Formation of Stromatolites of Lake Dziani Dzaha (Mayotte, Western Indian Ocean).

Lake Dziani Dzaha is a thalassohaline tropical crater lake located on the "Petite Terre" Island of Mayotte (Comoros archipelago, Western Indian Ocean). Stromatolites are actively growing in the shallow waters of the lake shores. These stromatolites are mainly composed of aragonite with lesser proportions of hydromagnesite, calcite, dolomite, and phyllosilicates. They are morphologically and texturally diverse ranging from tabular covered by a cauliflower-like crust to columnar ones with a smooth surface. High-throughput sequencing of bacterial and archaeal 16S rRNA genes combined with confocal laser scanning microscopy (CLSM) analysis revealed that the microbial composition of the mats associated with the stromatolites was clearly distinct from that of the Arthrospira-dominated lake water. Unicellular-colonial Cyanobacteria belonging to the Xenococcus genus of the Pleurocapsales order were detected in the cauliflower crust mats, whereas filamentous Cyanobacteria belonging to the Leptolyngbya genus were found in the smooth surface mats. Observations using CLSM, scanning electron microscopy (SEM) and Raman spectroscopy indicated that the cauliflower texture consists of laminations of aragonite, magnesium-silicate phase and hydromagnesite. The associated microbial mat, as confirmed by laser microdissection and whole-genome amplification (WGA), is composed of Pleurocapsales coated by abundant filamentous and coccoid Alphaproteobacteria. These phototrophic Alphaproteobacteria promote the precipitation of aragonite in which they become incrusted. In contrast, the Pleurocapsales are not calcifying but instead accumulate silicon and magnesium in their sheaths, which may be responsible for the formation of the Mg-silicate phase found in the cauliflower crust. We therefore propose that Pleurocapsales and Alphaproteobacteria are involved in the formation of two distinct mineral phases present in the cauliflower texture: Mg-silicate and aragonite, respectively. These results point out the role of phototrophic Alphaproteobacteria in the formation of stromatolites, which may open new perspective for the analysis of the fossil record.

Microbial Diversity and Cyanobacterial Production in Dziani Dzaha Crater Lake, a Unique Tropical Thalassohaline Environment.

This study describes, for the first time, the water chemistry and microbial diversity in Dziani Dzaha, a tropical crater lake located on Mayotte Island (Comoros archipelago, Western Indian Ocean). The lake water had a high level of dissolved matter and high alkalinity (10.6-14.5 g L-1 eq. CO32-, i.e. 160-220 mM compare to around 2-2.5 in seawater), with salinity up to 52 psu, 1.5 higher than seawater. Hierarchical clustering discriminated Dziani Dzaha water from other alkaline, saline lakes, highlighting its thalassohaline nature. The phytoplankton biomass was very high, with a total chlorophyll a concentration of 524 to 875 µg chl a L-1 depending on the survey, homogeneously distributed from surface to bottom (4 m). Throughout the whole water column the photosynthetic biomass was dominated (>97% of total biovolume) by the filamentous cyanobacteria Arthrospira sp. with a straight morphotype. In situ daily photosynthetic oxygen production ranged from 17.3 to 22.2 g O2 m-2 d-1, consistent with experimental production / irradiance measurements and modeling. Heterotrophic bacterioplankton was extremely abundant, with cell densities up to 1.5 108 cells mL-1 in the whole water column. Isolation and culture of 59 Eubacteria strains revealed the prevalence of alkaliphilic and halophilic organisms together with taxa unknown to date, based on 16S rRNA gene analysis. A single cloning-sequencing approach using archaeal 16S rDNA gene primers unveiled the presence of diverse extremophilic Euryarchaeota. The water chemistry of Dziani Dzaha Lake supports the hypothesis that it was derived from seawater and strongly modified by geological conditions and microbial activities that increased the alkalinity. Dziani Dzaha has a unique consortium of cyanobacteria, phytoplankton, heterotrophic Eubacteria and Archaea, with very few unicellular protozoa, that will deserve further deep analysis to unravel its uncommon diversity. A single taxon, belonging to the genus Arthrospira, was found responsible for almost all photosynthetic primary production.