Stromatolites are laminated, presumably microbial, structures, consisting largely of an authigenic precipitate, and manifest the appearance of microbial life in the geological rock record at least 3.4 Ga ago. Thus, stromatolites provide unique geochemical archives of aqueous environments on Earth and their habitability. It is, however, still incompletely understood under which physico-chemical conditions stromatolites formed and how these environments changed with the co-evolution of the atmosphere-hydrosphere-lithosphere systems through deep time.

This contribution targets the potential and pitfalls of emerging and established isotope applications to stromatolites based on improved and newly developed analytical and technical facilities in the last decades. I will provide an overview of present data and the interpretation of novel applications of stable and radiogenic isotope systems in stromatolites. Although the behaviour and fractionation processes of different isotope systems in stromatolites and microbial mats are sometimes incompletely understood, the different isotope proxies have the unique potential to better understand and reconstruct microbial habitats through deep time. Primarily, radiogenic isotopes are used to directly date stromatolites and determine the source of elements in ancient stromatolite environments; stable isotopes are used to understand redox conditions, metal availability, and (biogenic) metal cycling processes in microbial habitats. I provide insights into different isotope applications and their future perspectives to bridge the gap between geochemistry and microbiology and better understand the evolution of microbial life in stromatolite-forming environments on Earth and beyond.

The global record of early life is only poorly preserved, but has an ark in the 3.2 Ga Moodies Group of the Barberton Greenstone Belt, South Africa and Eswatini. It preserves silicified photosynthetic and sulfate-reducing metabolic signatures in sandstone-dominated, terrestrial to shallow-marine strata. Large fluid-escape structures are common in thick-bedded kerogen-laminated sandstones of (sub-)tidal facies. We document and interpret silicified, massive and laminated carbonate aggregates and beds, both of likely microbial origin, within and on top of these syndepositional and early diagenetic features, not previously described from Archean shallow-water fluid-escape structures. We distinguish three morphotypes: (1) cm-scale, silicified, bulbous aggregates aligned within fluid-escape conduits; (2) up to dm-scale, dolomitized, finely-laminated conical and tabular mounds on top of the conduits; (3) cm-scale, isolated, silicified, finely-laminated, stromatolitic aggregates. In-situ SIMS isotope analyses from traverses across the best-preserved laminae of a mound yielded δ¹³C_(PDB) values relative to a dolomite standard of -2.5 to 0.5‰, and -3.5 to 4.0‰ for δ³⁴S_(VCDT) from diagenetic rims of nearby detrital pyrite grains, respectively. Values and ranges are consistent with a near-complete hydrothermal alteration. Facies context, location within and on top of the fluid-escape structures, stromatolitic morphology, and carbonate composition suggest that robust microbial communities utilized one or several carbon-based redox pathways in this siliciclastic tidal setting. Methanogens, methanotrophs, sulfate reducers and photosynthesizers may have colonized these tidal-zone sand volcanoes at 3.2 Ga, collectively forming a diverse microbial community.

The Earth’s atmosphere was without free oxygen until the Great Oxygenation Event, thought to have been driven by oxygenic photosynthesis. The expansion of early Cyanobacteria was proposed to be restricted by the lack of bioavailable nitrogen. The effects of an anoxic Archean atmosphere on the growth of a the nitrogen fixing Cyanobacterium Nostoc sp. PCC7524 was compared to control cultures grown under present day atmospheric levels (PAL) of O₂ and CO₂. Additionally, we assessed how the early Archean atmosphere affected the gas diffusion barrier, consisting of heterocyte glycolipids, of the heterocyte and the ability of early Cyanobacteria to fix N₂.

While no significant changes were observed for growth rates under N-depleted conditions in the experimental and control atmospheres, upregulation of the C- and N₂-fixation associated genes, were observed under Archean conditions relative to PAL. This correlated with increased levels of the C-fixing Rubisco protein and O₂ production. The glycogen and protein content of the Archean endpoint culture material showed raised levels of these long-term storage compounds compared to those grown under PAL conditions. No significant changes in the heterocyte glycolipid content or composition was observed.

This data suggests that diazotrophic Cyanobacteria were able to fix nitrogen and carbon more efficiently under the anoxic conditions of the Archean, thereby releasing more biologically available carbon and nitrogen into the immediate environment than under PAL conditions. The fact that no significant changes in the heterocyte glycolipid content occurred suggests they are suitable biomarkers for cyanobacterial N₂-fixation in geological records.

Archean Cyanobacteria oxygenated Earth’s atmosphere during the Great Oxygenation Event (GOE) through the action of oxygenic photosynthesis. The photosynthetic apparatus relies on metalloproteins, many of which contain iron. Cyanobacteria use several specific transporters to meet their high iron requirements. In the ferruginous anoxic Archean ocean, the FeoABC transporter was thought to be the primary means of Fe(II) uptake.

Our goal is to investigate the distribution of inorganic iron uptake mechanisms among Cyanobacteria and to determine the emergence of core iron receptors in the Cyanobacterial lineage.

Essential iron uptake transporters and regulators were identified in 125 Cyanobacteria using in silico analysis. We reconstructed the Baysean phylogeny of the Fe(II) receptor FeoB, the high affinity Fe(III) permease, FutB, and cyanobacterial FTR1. Additionally, the arrival of these iron receptors in the Cyanobacterial lineage was timed using a molecular clock. The expression of cftr1 (Pse7367_Rs12485), furA (Pse7367_Rs06445) and cyoC (Pse7367_Rs00935) was determined by quantitative RT-PCR against the reference gene, rpoC1 (Pse7367_Rs07505), in the basal clade cyanobacterium Pseudanabaena PCC7367, grown under simulated Archean conditions.

Genome analysis shows an absence of the high affinity Fe(II) transporter, FeoB, in most basal Cyanobacteria. Moreover, evolutionary dating timed the arrival of FeoB, cFTR1 and FutB in the cyanobacterial lineage during the Proterozoic. Furthermore, cftr1 is constitutively expressed in Pseudanabaena PCC7367, even after the addition of Fe(II).

This study highlights the need for a reappraisal of iron uptake systems in early Cyanobacteria, as Fe(II) does not appear to have been their primary source of iron in the ferruginous Archean oceans.

Cyanobacteria are able to conduct oxygenic photosynthesis and are thought to have been responsible for the Great Oxygenation Event (GOE). The effect of increasing levels of atmospheric O₂ on the physiology of Cyanobacteria is unknown. Cyanobacteria produce toxic superoxide ions during photosynthesis through the hydrolysis of water. In this project, we investigate the expression of the Superoxide Dismutase (SOD) enzyme, which is responsible for eliminating the superoxide ion, in an ancestral marine species, Pseudanabaena sp. PCC7367.

Growth curves based on Chlorophyll a and protein content were conducted under an anoxic atmosphere representing the ‘Archean’, and one representing Present Atmospheric Levels (PAL) of CO₂ and O₂. Expression of SOD genes was monitored over a day: night cycle, in conjunction with measuring oxygen release. The activity of the enzymes was assessed using native gel assays.

The growth rate for Pseudanabaena sp. PCC7367 was highest for cultures grown under the anoxic atmosphere suggesting that modern levels of atmospheric O₂ impair the growth of Cyanobacteria compared to the ‘Archean’ atmosphere. SOD gene expression was highest during the day when O₂ levels were at their highest. Relative gene expression under both atmospheres was not significantly different, suggesting that the expression of SOD depends on cellular O₂ production rather than atmospheric O_2. Enzyme activity assays confirmed the synthesis of the SODs.

In conclusion, this study suggests that increased atmospheric O₂ levels would not have restricted the spread of Cyanobacteria as they would have required SODs once they acquired the ability to conduct oxygenic photosynthesis.

The so-called Permian – Triassic mass extinction was followed by a prolonged period of ecological recovery that lasted until the Middle Triassic. Triassic stromatolites from the Germanic Basin seem to be an important part of the puzzle, but have barely been investigated so far. Here we analyzed late Anisian (upper Middle Muschelkalk) stromatolites from across the Germanic Basin by combining petrographic approaches (optical microscopy, micro X-ray fluorescence, Raman imaging) and geochemical analyses (sedimentary hydrocarbons, stable carbon and oxygen isotopes). Paleontological and sedimentological evidence, such as Placunopsis bivalves, intraclasts and disrupted laminated fabrics, indicate that the stromatolites formed in subtidal, shallow marine settings. This interpretation is consistent with δ¹³C_carb of about -2.1 ‰ to -0.4 ‰. Remarkably, the stromatolites are composed of microbes (perhaps cyanobacteria and sulfate reducing bacteria) and metazoans (non-spicular demosponges, Placunopsis bivalves, and/or Spirobis-like worm tubes). Therefore, they should more correctly be referred to as microbe-metazoan build-ups. They are characterized by diverse lamination types, including planar, wavy, domal and conical ones. Microbial mats likely played an important role in forming the planar and wavy laminations. Domal and conical laminations commonly show clotted to peloidal features and mesh-like fabrics, attributed to fossilized non-spicular demosponges. In the light of our findings, it appears plausible that the involved organisms benefited from elevated salinities. Another possibility is that the mutualistic relationship between microbes and non-spicular demosponges enabled these organisms to fill ecological niches cleared by the Permian – Triassic Crisis and maintain their advantage until the Middle Triassic.