Cyanobacterial calcification in modern microbialites at the submicrometer scale

International audienceThe search for microfossils in the geological record has been a long-term challenge. Part of the problem comes from the difficulty of identifying such microfossils unambiguously, since they can be morphologically confused with abiotic biomorphs. One route to improve our ability to correctly identify microfossils involves studying fossilization processes affecting bacteria in modern settings. We studied the initial stages of fossilization of cyanobacterial cells in modern microbialites from Lake Alchichica (Mexico), a Mg-rich hyperalkaline crater lake (pH 8.9) hosting currently growing stromatolites composed of aragonite [CaCO3] and hydromagnesite [Mg5(CO3)4(OH)2 * 4(H2O)]. Most of the biomass associated with the microbialites is composed of cyanobacteria. Scanning electron microscopy analyses coupled with confocal laser scanning microscopy observations were conducted to co-localize cyanobacterial cells and associated minerals. These observations showed that cyanobacterial cells affiliated with the order Pleurocapsales become specifically encrusted within aragonite with an apparent preservation of cell morphology. Encrustation gradients from non-encrusted to totally encrusted cells spanning distances of a few hundred micrometers were observed. Cells exhibiting increased levels of encrustation along this gradient were studied down to the nm scale using a combination of focused ion beam (FIB) milling, transmission electron microscopy (TEM) and scanning transmission x-ray microscopy (STXM) at the C, O and N K-edges. Two different types of aragonite crystals were observed: one type was composed of needle-shaped nano-crystals growing outward from the cell body with a crystallographic orientation perpendicular to the cell wall, and another type was composed of larger crystals that progressively filled the cell interior. Exopolymeric substances (EPS), initially co-localized with the cells, decreased in concentration and dispersed away from the cells while crystal growth occurred. As encrustation developed, EPS progressively disappeared, but remaining EPS showed the same spectroscopic signature. In the most advanced stages of fossilization, only the textural organization of the two types of aragonite recorded the initial cell morphology and spatial distribution

Biological Soil Crusts as Modern Analogues for the Archean Continental Biosphere: Insights from Carbon and Nitrogen Isotopes

Stable isotope signatures of elements related to life such as carbon and nitrogen can be powerful biomarkers that provide key information on the biological origin of organic remains and their paleoenvironments. Marked advances have been achieved in the last decade in our understanding of the coupled evolution of biological carbon and nitrogen cycling and the chemical evolution of the early Earth thanks, in part, to isotopic signatures preserved in fossilized microbial mats and organic matter of marine origin. However, the geologic record of the early continental biosphere, as well as its evolution and biosignatures, is still poorly constrained. Following a recent report of direct fossil evidence of life on land at 3.22 Ga, we compare here the carbon and nitrogen isotopic signals of this continental Archean biosphere with biosignatures of cyanobacteria biological soil crusts (cyanoBSCs) colonizing modern arid environments. We report the first extended δ13C and δ15N data set from modern cyanoBSCs and show that these modern communities harbor specific isotopic biosignatures that compare well with continental Archean organic remains. We therefore suggest that cyanoBSCs are likely relevant analogues for the earliest continental ecosystems. As such, they can provide key information on the timing, extent, and possibly mechanism of colonization of the early Earth's emergent landmasses

Prokaryotic and Eukaryotic Community Structure in Field and Cultured Microbialites from the Alkaline Lake Alchichica (Mexico)

The geomicrobiology of crater lake microbialites remains largely unknown despite their evolutionary interest due to their resemblance to some Archaean analogs in the dominance of in situ carbonate precipitation over accretion. Here, we studied the diversity of archaea, bacteria and protists in microbialites of the alkaline Lake Alchichica from both field samples collected along a depth gradient (0–14 m depth) and long-term-maintained laboratory aquaria. Using small subunit (SSU) rRNA gene libraries and fingerprinting methods, we detected a wide diversity of bacteria and protists contrasting with a minor fraction of archaea. Oxygenic photosynthesizers were dominated by cyanobacteria, green algae and diatoms. Cyanobacterial diversity varied with depth, Oscillatoriales dominating shallow and intermediate microbialites and Pleurocapsales the deepest samples. The early-branching Gloeobacterales represented significant proportions in aquaria microbialites. Anoxygenic photosynthesizers were also diverse, comprising members of Alphaproteobacteria and Chloroflexi. Although photosynthetic microorganisms dominated in biomass, heterotrophic lineages were more diverse. We detected members of up to 21 bacterial phyla or candidate divisions, including lineages possibly involved in microbialite formation, such as sulfate-reducing Deltaproteobacteria but also Firmicutes and very diverse taxa likely able to degrade complex polymeric substances, such as Planctomycetales, Bacteroidetes and Verrucomicrobia. Heterotrophic eukaryotes were dominated by Fungi (including members of the basal Rozellida or Cryptomycota), Choanoflagellida, Nucleariida, Amoebozoa, Alveolata and Stramenopiles. The diversity and relative abundance of many eukaryotic lineages suggest an unforeseen role for protists in microbialite ecology. Many lineages from lake microbialites were successfully maintained in aquaria. Interestingly, the diversity detected in aquarium microbialites was higher than in field samples, possibly due to more stable and favorable laboratory conditions. The maintenance of highly diverse natural microbialites in laboratory aquaria holds promise to study the role of different metabolisms in the formation of these structures under controlled conditions

Calcite-accumulating large sulfur bacteria of the genus Achromatium in Sippewissett Salt Marsh

Large sulfur bacteria of the genus Achromatium are exceptional among Bacteria and Archaea as they can accumulate high amounts of internal calcite. Although known for more than 100 years, they remain uncultured, and only freshwater populations have been studied so far. Here we investigate a marine population of calcite-accumulating bacteria that is primarily found at the sediment surface of tide pools in a salt marsh, where high sulfide concentrations meet oversaturated oxygen concentrations during the day. Dynamic sulfur cycling by phototrophic sulfide-oxidizing and heterotrophic sulfate-reducing bacteria co-occurring in these sediments creates a highly sulfidic environment that we propose induces behavioral differences in the Achromatium population compared with reported migration patterns in a low-sulfide environment. Fluctuating intracellular calcium/sulfur ratios at different depths and times of day indicate a biochemical reaction of the salt marsh Achromatium to diurnal changes in sedimentary redox conditions. We correlate this calcite dynamic with new evidence regarding its formation/mobilization and suggest general implications as well as a possible biological function of calcite accumulation in large bacteria in the sediment environment that is governed by gradients. Finally, we propose a new taxonomic classification of the salt marsh Achromatium based on their adaptation to a significantly different habitat than their freshwater relatives, as indicated by their differential behavior as well as phylogenetic distance on 16S ribosomal RNA gene level. In future studies, whole-genome characterization and additional ecophysiological factors could further support the distinctive position of salt marsh Achromatium

Diversity and mineral substrate preference in endolithic microbial communities from marine intertidal outcrops (Isla de Mona, Puerto Rico)

Endolithic microbial communities are prominent features of intertidal marine habitats, where they colonize a variety of substrates, contributing to their erosion. Almost 2 centuries worth of naturalistic studies focused on a few true-boring (euendolithic) phototrophs, but substrate preference has received little attention. The Isla de Mona (Puerto Rico) intertidal zone offers a unique setting to investigate substrate specificity of endolithic communities since various phosphate rock, limestone and dolostone outcrops occur there. High-throughput 16S rDNA genetic sampling, enhanced by targeted cultivation, revealed that, while euendolithic cyanobacteria were dominant operational taxonomic units (OTUs), the communities were invariably of high diversity, well beyond that reported in traditional studies and implying an unexpected metabolic complexity potentially contributed by secondary colonizers. While the overall community composition did not show differences traceable to the nature of the mineral substrate, we detected specialization among particular euendolithic cyanobacterial clades towards the type of substrate they excavate but only at the OTU phylogenetic level, implying that close relatives have specialized recurrently into particular substrates. The cationic mineral component was determinant in this preference, suggesting the existence in nature of alternatives to the boring mechanism described in culture that is based exclusively on transcellular calcium transport

Geomicrobiological study of modern microbialites from Mexico: towards a better understanding of the ancient fossil record

Microbialites are sedimentary formations that are found throughout the geological record and are usually considered as one of the oldest traces of life on Earth. Although they have been known for more than a century and hold as an emblematic object in Earth Sciences, we yet do not understand in details how they form and how microbial processes impact their chemistry, structure and macroscopic morphology. Here, we show recent advances achieved owing to funding provided by the EPOV program on the formation of modern microbialites in the crater Lake Alchichica (Mexico). We first show that very diverse microbial communities populate these microbialites, including diverse microbial groups able to induce carbonate precipitation. We demonstrate that this microbial diversity can be preserved for several years in laboratory aquaria offering a nice opportunity for future studies to assess the relationships between biodiversity and microbialite formation. We then detail the textural modifications affecting cyanobacterial cells during the first steps of fossilization as captured in Alchichica microbialites. Finally, we report the discovery of a new deepbranching cyanobacterium species, Candidatus Gloeomargarita lithophora, able to form intracellular Ca-, Mg-, Sr- and Ba-rich carbonates and discuss the implications for the interpretation of the fossil record

Extracellular polymeric matrix in biocrusts: its role as a repository for essential exometabolites

Matrice polisaccaridi come sito di accumulo di esometabolit

A novel method to evaluate nutrient retention by biological soil crust exopolymeric matrix

Aims: Biological soil crusts (biocrusts) are microbial communities commonly found in the upper layer of arid soils. These microorganisms release exopolysaccharides (EPS), which form the exopolymeric matrix (EPM), allowing them to bond soil particles together and survive long periods of dryness. The aim of this work is to develop methods for measuring metabolite retention by biocrust EPM and EPS. Methods: We report new methods for the investigation of metabolite sorption on biocrusts compared to the underlying unconsolidated subcrust fraction. A 13C–labeled bacterial lysate metabolite mixture was incubated with biocrust, subcrust and biocrust-extracted EPS. Non-sorbed metabolites were extracted and analyzed by liquid chromatography/mass spectrometry. Results: This simple and rapid approach enabled the comparison of metabolite sorption on the biocrust EPM or EPS versus mineral sorption on the underlying soils. Our results suggest that the biocrust (and its extracted EPS) sorb more metabolites, especially amino acids and organic acids, than the underlying subcrust. Conclusions: This study demonstrates a useful method to highlight the essential role of biocrust (especially the EPM), which acts as a passive nutrient filter, sequestering metabolites released by microbes during wetting events. This may facilitate recovery of the community upon wetting and further enhance biocrust survival and nutrient retention

Geomicrobiological study of modern microbialites from Mexico: towards a better understanding of the ancient fossil record

Microbialites are sedimentary formations that are found throughout the geological record and are usually considered as one of the oldest traces of life on Earth. Although they have been known for more than a century and hold as an emblematic object in Earth Sciences, we yet do not understand in details how they form and how microbial processes impact their chemistry, structure and macroscopic morphology. Here, we show recent advances achieved owing to funding provided by the EPOV program on the formation of modern microbialites in the crater Lake Alchichica (Mexico). We first show that very diverse microbial communities populate these microbialites, including diverse microbial groups able to induce carbonate precipitation. We demonstrate that this microbial diversity can be preserved for several years in laboratory aquaria offering a nice opportunity for future studies to assess the relationships between biodiversity and microbialite formation. We then detail the textural modifications affecting cyanobacterial cells during the first steps of fossilization as captured in Alchichica microbialites. Finally, we report the discovery of a new deepbranching cyanobacterium species, Candidatus Gloeomargarita lithophora, able to form intracellular Ca-, Mg-, Sr- and Ba-rich carbonates and discuss the implications for the interpretation of the fossil record