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At particular stages during their life cycles, fungi use multiple strategies to form specialized structures to survive unfavorable environmental conditions. These strategies encompass sporulation, as well as cell-wall melanization, multicellular tissue formation or even dimorphism. The resulting structures are not only used to disperse to other environments, but also to survive long periods of time awaiting favorable growth conditions. As a result, these specialized fungal structures are part of the microbial seed bank, which is known to influence the microbial community composition and contribute to the maintenance of diversity. Despite the importance of the microbial seed bank in the environment, methods to study the diversity of fungal structures with improved resistance only target spores dispersing in the air, omitting the high diversity of these structures in terms of morphology and environmental distribution. In this study, we applied a separation method based on cell lysis to enrich lysis-resistant fungal structures (for instance, spores, sclerotia, melanized yeast) to obtain a proxy of the composition of the fungal seed bank. This approach was first evaluated in-vitro in selected species. The results obtained showed that DNA from fungal spores and from yeast was only obtained after the application of the enrichment method, while mycelium was always lysed. After validation, we compared the diversity of the total and lysis-resistant fractions in the polyextreme environment of the Salar de Huasco, a high-altitude athalassohaline wetland in the Chilean Altiplano. Environmental samples were collected from the salt flat and from microbial mats in small surrounding ponds. Both the lake sediments and microbial mats were dominated by Ascomycota and Basidiomycota, however, the diversity and composition of each environment differed at lower taxonomic ranks. Members of the phylum Chytridiomycota were enriched in the lysis-resistant fraction, while members of the phylum Rozellomycota were never detected in this fraction. Moreover, we show that the community composition of the lysis-resistant fraction reflects the diversity of life cycles and survival strategies developed by fungi in the environment. To the best of our knowledge this is the first time that the fungal diversity is explored in the Salar de Huasco. In addition, the method presented here provides a simple and culture independent approach to assess the diversity of fungal lysis-resistant cells in the environment.

International audience; Soil organic matter (SOM) is a complex heterogeneous mixture formed through decomposition and organo-mineral interactions, and characterization of its composition and biogeochemical stability is challenging. From this perspective, Rock-Eval® is a rapid and efficient thermal analytical method that combines the quantitative and qualitative information of SOM, including several parameters related to thermal stability. This approach has already been used to monitor changes in organic matter (OM) properties at the landscape, cropland, and soil profile scales. This study was aimed to assess the stability of SOM pools by characterizing the grain size fractions from forest litters and topsoils using Rock-Eval® thermal analysis. Litter (organic) and topsoil samples were collected from a beech forest in Normandy (France), whose management in the last 200 years has been documented. Fractionation by wet sieving was used to separate large debris (> 2 000 μm) and coarse (200–2 000 μm) and fine particulate OM (POM) (50–200 μm) in the organic samples as well as coarse (200–2 000 μm), medium (50–200 μm), and fine (< 50 μm) fractions of the topsoil samples. Rock-Eval® was able to provide thermal parameters sensitive enough to study fine-scale soil processes. In the organic layers, quantitative and qualitative changes were explained by the progressive decomposition of labile organic compounds from plant debris to the finest organic particles. Meanwhile, the grain size fractions of topsoils presented different characteristics. The coarse organo-mineral fractions showed higher C contents, albeit with a different composition, higher thermal stability, and greater decomposition degree than the plant debris forming the organic layer. These results are consistent with those of previous studies that microbial activity is more effective in this fraction. The finest fractions of topsoils showed low C contents, the highest thermal stability, and low decomposition degree, which can be explained by the stronger interactions with the mineral matrix. Therefore, it is suggested that the dynamics of OM in the different size fractions be interpreted in the light of a plant-microbe-soil continuum. Finally, three distinct thermostable C pools were highlighted through the grain size heterogeneity of SOM: free coarse OM (large debris and coarse and fine particles), weakly protected OM in (bio)aggregates (coarse fraction of topsoil), and stabilized OM in the fine fractions of topsoil, which resulted from the interactions within organo-mineral complexes. Therefore, Rock-Eval® thermal parameters can be used to empirically illustrate the conceptual models emphasizing the roles of drivers played by the gradual decomposition and protection of the most thermally labile organic constituent

Bacteria–fungi interactions (BFIs) are essential in ecosystem functioning. These interactions are modulated not only by local nutritional conditions but also by the physicochemical constraints and 3D structure of the environmental niche. In soils, the unsaturated and complex nature of the substrate restricts the dispersal and activity of bacteria. Under unsaturated conditions, some bacteria engage with filamentous fungi in an interaction (fungal highways) in which they use fungal hyphae to disperse. Based on a previous experimental device to enrich pairs of organisms engaging in this interaction in soils, we present here the design and validation of a modified version of this sampling system constructed using additive printing. The 3D printed devices were tested using a novel application in which a target fungus, the common coprophilous fungus Coprinopsis cinerea, was used as bait to recruit and identify bacterial partners using its mycelium for dispersal. Bacteria of the genera Pseudomonas, Sphingobacterium and Stenotrophomonas were highly enriched in association with C. cinerea. Developing and producing these new easy-to-use tools to investigate how bacteria overcome dispersal limitations in cooperation with fungi is important to unravel the mechanisms by which BFIs affect processes at an ecosystem scale in soils and other unsaturated environments., Fungal highway columns produced by additive printing.

Summary This study presents the evolution of organic matter content using Rock-Eval® pyrolysis in Beech forest soils after their particle Size-fractionation. The amount of C in soil depends in microbial and physicochemical interactions between various biomass affecting its accumulation and its stability. As far as thermal stability of soil OM is supposed to mimic its biogeochemical stability, Rock-Eval® pyrolysis method is an efficient tool to quantify the organic and mineral carbon. To this end, litter and topsoil samples from a beech forest located in Normandy (France) were selected and fractionated by sieving from large debris to A-fine fractions. The TOC decreases from large debris to A-medium and drops in A-fine. Thermal stability inputs are highlighted through RE parameters when comparing litters and particle size fractions separated from topsoil samples (increase of TpS2 and OI, decrease of HI with decreasing fraction size in A horizons). Three distinct thermal stability C pools are identified

Non-anthropogenic earth mounds, defined as mima-like mounds in this study, have recently been observed in non-carbonate watersheds along the Sudano-Sahelian belt in the Chad Basin. In the Diamare piedmont (northern Cameroon) they are particularly well developed within stream networks. In less eroded areas, they occur as whaleback, flattened morphologies, or even as buried features. All these shapes are composed of clay-rich sediment associated with high proportions of secondary carbonate nodules and Fe Mn micro-nodules. Their soil structure is prismatic to massive and vertical cracks are observed locally. Grain-size distributions emphasize the clay-rich nature of the sediment, with average clay contents of 32% ± 12.8% (n = 186), which is significantly higher than the clay content in the adjacent sediments in the landscape (mean = 10% ± 4%, n = 21). Moreover, high proportions of smectite characterize the soil, with average contents of 34 ± 7% (n = 25). At the micro-scale, the groundmass has a cross-striated b-fabric, with embedded smooth subangular quartz and feldspar grains of the silt-size fraction. All the characteristics point to altered vertic properties in the clay-rich sediment composing the mima-like mounds. Mima-like mounds are thus interpreted as degraded Vertisols. Compared to present-day Vertisols occurring in the piedmont, mima-like mounds are located upstream. It is thus proposed that the Vertisol areas were more extensive during a former and wetter period than the present-day. Subsequent changing climatic conditions increased erosion, revealing the gilgai micro-relief by preferential erosion in micro-lows rather than in micro-highs. Mima-like mounds of the Chad Basin might thus result from pedogenesis combined with later erosion. These local processes can be inherited from regional climatic variations during the Late Pleistocene-Holocene and likely be related to the African Humid Period.

In the field of microbial biomineralization, much of the scientific attention is focused on processes carried out by prokaryotes, in particular bacteria, even though fungi are also known to be involved in biogeochemical cycles in numerous ways. They are traditionally recognized as key players in organic matter recycling, as nutrient suppliers via mineral weathering, as well as large producers of organic acids such as oxalic acid for instance, an activity leading to the genesis of various metal complexes such as metal-oxalate. Their implications in the transformation of various mineral and metallic compounds has been widely acknowledged during the last decade, however, currently, their contribution to the genesis of a common biomineral, calcite, needs to be more thoroughly documented. Calcite is observed in many ecosystems and plays an essential role in the biogeochemical cycles of both carbon (C) and calcium (Ca). It may be physicochemical or biogenic in origin and numerous organisms have been recognized to control or induce its biomineralization. While fungi have often been suspected of being involved in this process in terrestrial environments, only scarce information supports this hypothesis in natural settings. As a result, calcite biomineralization by microbes is still largely attributed to bacteria at present. However, in some terrestrial environments there are particular calcitic habits that have been described as being fungal in origin. In addition to this, several studies dealing with axenic cultures of fungi have demonstrated the ability of fungi to produce calcite. Examples of fungal biomineralization range from induced to organomineralization processes. More examples of calcite biomineralization related to direct fungal activity, or at least to their presence, have been described within the last decade. However, the peculiar mechanisms leading to calcite biomineralization by fungi remain incompletely understood and more research is necessary, posing new exciting questions linked to microbial biomineralization processes.

Background and Aims The oxalate-carbonate pathway (OCP) has been observed in acidic tropical soils with low alkaline cation content where compartments are transient and fed by the rapid turnover of organic matter. By acting on edaphic parameters, the OCP may influence soil nutrient distribution. This study aims at assessing the influence of the OCP on soil nutrients within an agroforestry system associated to oxalogenic iroko trees. Methods Soil nutrient distribution was studied in a 30 m long and 1 m deep transect starting at the iroko tree towards the vegetation surrounding it. Results Processes controlling nutrient distributions varied with both distance and depth. The tree drastically impacted edaphic variables, in the first instance pH. Changes in pH generated gradients of calcium and magnesium, both of the exchangeable and the total fraction. In contrast, total phosphorus and potassium distribution were mostly influenced by depth. Conclusions This downward gradient fits the "plant cycling model" that explains the effect of vegetation on the recharge of soil nutrients. This is the first example of the effect of the OCP on soil nutrient distribution. Considering that the OCP is not restricted to the iroko tree, this study highlights a more general pattern of forest dynamics developed on highly weathered tropical soils.

A continuum of carbon, from atmospheric CO2 to secondary calcium carbonate, has been studied in a soil associated with scree slope deposits in the Jura Mountains of Switzerland. This approach is based on former studies conducted in other environments. This C continuum includes atmospheric CO2, soil organic matter (SOM), soil CO2, dissolved inorganic carbon (DIC) in soil solutions, and secondary pedogenic carbonate. Soil parameters (pCO2, temperature, pH, Cmin and Corg contents), soil solution chemistry, and isotopic compositions of soil CO2, DIC, carbonate and soil organic matter (δ13CCO2, δ13CDIC, δ13Ccar and δ13CSOM values) have been monitored at different depths (from 20 to 140 cm) over one year. Results demonstrated that the carbon source in secondary carbonate (mainly needle fiber calcite) is related to the dissolved inorganic carbon, which is strongly dependent on soil respiration. The heterotrophic respiration, rather than the limestone parent material, seems to control the pedogenic carbon cycle. The correlation of δ13Corg values with Rock-Eval HI and OI indices demonstrates that, in a soil associated to scree slope deposits, the main process responsible for 13C-enrichment in SOM is related to bacterial oxidative decarboxylation. Finally, precipitation of secondary calcium carbonate is enhanced by changes in soil pCO2 associated to the convective movement of air masses induced by temperature gradients (heat pump effect) in the highly porous scree slope deposits. The exportation of soil C-leachates from systems such as the one studied in this paper could partially explain the “gap in the European carbon budget” reported by recent studies.

Calcitic nanofibres are ubiquitous habits of secondary calcium carbonate (CaCO3) accumulations observed in calcareous vadose environments. Despite their widespread occurrence, the origin of these nanofeatures remains enigmatic. Three possible mechanisms fuel the debate: (i) purely physicochemical processes, (ii) mineralization of rod-shaped bacteria, and (iii) crystal precipitation on organic templates. Nanofibres can be either mineral (calcitic) or organic in nature. They are very often observed in association with needle fibre calcite (NFC), another typical secondary CaCO3 habit in terrestrial environments. This association has contributed to some confusion between both habits, however they are truly two distinct calcitic features and their recurrent association is likely to be an important fact to help understanding the origin of nanofibres. In this paper the different hypotheses that currently exist to explain the origin of calcitic nanofibres are critically reviewed. In addition to this, a new hypothesis for the origin of nanofibres is proposed based on the fact that current knowledge attributes a fungal origin to NFC. As this feature and nanofibres are recurrently observed together, a possible fungal origin for nanofibres which are associated with NFC is investigated. Sequential enzymatic digestion of the fungal cell wall of selected fungal species demonstrates that the fungal cell wall can be a source of organic nanofibres. The obtained organic nanofibres show a striking morphological resemblance when compared to their natural counterparts, emphasizing a fungal origin for part of the organic nanofibres observed in association with NFC. It is further hypothesized that these organic nanofibres may act as templates for calcite nucleation in a biologically influenced mineralization process, generating calcitic nanofibres. This highlights the possible involvement of fungi in CaCO3 biomineralization processes, a role still poorly documented. Moreover, on a global scale, the organomineralization of organic nanofibres into calcitic nanofibres might be an overlooked process deserving more attention to specify its impact on the biogeochemical cycles of both Ca and C.

The oxalate–carbonate pathway (OCP) is a biogeochemical process, which has been described in Milicia excelsa tree ecosystems of Africa. This pathway involves biological and geological parameters at different scales: oxalate, as a by-product of photosynthesis, is oxidized by oxalotrophic bacteria leading to a local pH increase, and eventually to carbonate accumulation through time in previously acidic and carbonate-free tropical soils. Former studies have shown that this pedogenic process can potentially lead to the formation of an atmospheric carbon sink. Considering that 80% of plant species are known to produce oxalate, it is reasonable to assume that M. excelsa is not the only tree that can support OCP ecosystems. The search for similar conditions on another continent led us to South America, in an Amazon forest ecosystem (Alto Beni, Bolivia). This area was chosen because of the absence of local inherited carbonate in the bedrock, as well as its expected acidic soil conditions. Eleven tree species and associated soils were tested positive for the presence of carbonate with a more alkaline soil pH close to the tree than at a distance from it. A detailed study of Pentaplaris davidsmithii and Ceiba speciosa trees showed that oxalotrophy impacted soil pH in a similar way to at African sites (at least with 1 pH unit increasing). African and South American sites display similar characteristics regarding the mineralogical assemblage associated with the OCP, except for the absence of weddellite. The amount of carbonate accumulated is 3 to 4 times lower than the values measured in African sites related to M. excelsa ecosystems. Still, these secondary carbonates remain critical for the continental carbon cycle, as they are unexpected in the acidic context of Amazonian soils. Therefore, the present study demonstrates the existence of an active OCP in South America. The three critical components of an operating OCP are the presence of: i) local alkalinization, ii) carbonate accumulations, and iii) oxalotrophic bacteria, which were identified associated to the oxalogenic tree C. speciosa. If the question of a potential carbon sink related to oxalotrophic–oxalogenic ecosystems in the Amazon Basin is still pending, this study highlights the implication of OCP ecosystems on carbon and calcium biogeochemical coupled cycles. As previously mentioned for M. excelsa tree ecosystems in Africa, carbonate accumulations observed in the Bolivian tropical forest could be extrapolated to part or the whole Amazon Basin and might constitute an important reservoir that must be taken into account in the global carbon balance of the Tropics.

A technique based on an inverted Petri dish system was developed for the growth and isolation of soil oxalotrophic bacteria able to disperse on fungal mycelia. The method is related to the 'fungal highways' dispersion theory in which mycelial fungal networks allow active movement of bacteria in soil. Quantification of this phenomenon showed that bacterial dispersal occurs preferentially in upper soil horizons. Eight bacteria and one fungal strain were isolated by this method. The oxalotrophic activity of the isolated bacteria was confirmed through calcium oxalate dissolution in solid selective medium. After separation of the bacteria-fungus couple, partial sequencing of the 16S and the ITS1 and ITS2 sequences of the ribosomal RNA genes were used for the identification of bacteria and the associated fungus. The isolated oxalotrophic bacteria included strains related to Stenotrophomonas, Achromobacter, Lysobacter, Pseudomonas, Agrobacterium, Cohnella, and Variovorax. The recovered fungus corresponded to Trichoderma sp. A test carried out to verify bacterial transport in an unsaturated medium showed that all the isolated bacteria were able to migrate on Trichoderma hyphae or glass fibers to re-colonize an oxalate-rich medium. The results highlight the importance of fungus-driven bacterial dispersal to understand the functional role of oxalotrophic bacteria and fungi in soils.

Oxalate oxidation and carbonate precipitation associated with the oxalogenic tree Terminalia bellirica were investigated. Calcium oxalate crystals, oxalotrophic bacteria (dominated by genera Methylobacterium and Burkholderia), and carbonate accumulation (82% dry weight), were detected in the bark. In contrast, only a slight accumulation of carbonate was observed in soil (1.5%). The combined geochemical and microbiological analyses suggest that bark of an Indian living tree represents a novel habitat for the accumulation of carbonate associated with bacterial oxalate oxidation. The importance of these types of habitats needs to be quantified when considering the biogeochemical cycling of carbon and calcium in tropical ecosystems.

[Departement_IRSTEA]Territoires [TR1_IRSTEA]SEDYVIN; International audience; This paper aims to develop a new approach to interpret soil organic matter (SOM) dynamics from Rock-Eval pyrolysis. Rock-Eval standard parameters (TpS2, HI, OI) are limited when applied to SOM, as they were defined for tracking the origin of sedimentary organic matter (i.e. terrestrial vs aquatic and lacustrine vs marine). This study proposes new Rock Eval based indices, projected on a new diagram plotting > 1300 samples, including litter and soil horizons, pure compounds and organic material. These new parameters reflect the thermal stability of SOM rather than its bulk chemistry. Their calculations are based on the contribution of four different areas (A1 to A4) integrated below the S2 pyrogram (amounts of released hydrocarbon compounds during the pyrolysis step). Results demonstrate that the relative values of A1 to A4 parameters can be used to survey the evolution of SOM during pedogenesis. These Rock-Eval parameters revealed a consistent thermal differentiation of SOM with depth, from thermally labile biogenic SOM in soil organic horizons to thermally stable mineral-associated SOM compounds in organo-mineral and mineral soil horizons, indicating a pedogenetic stabilization of SOM. Finally, newly defined I- and R-indices integrate the respective parts of A1 to A4 parameters into SOM dynamics, the I-index emphasizing the degree of transformation of the immature organic fraction (related to SOM stabilization), the R-index highlighting the contribution of the most refractory fraction or persistent SOM (related to pedogenic and inherited contributions). Analyses of a wide range of soils and reference samples (cellulose, lignin, humic substances, lignite, charcoal, coal, etc.) allow end-members as well as particular trends (humic, spodic, inherited) to be drawn. Specific examples are provided in order to illustrate some applications and uses for this new Rock-Eval based I/R diagram, from the study of grain size fractions to the evolution of SOM in soil chronosequences.

Summary The oxalate-carbonate pathway involves the oxida- tion of calcium oxalate to low-magnesium calcite and represents a potential long-term terrestrial sink for atmospheric CO2. In this pathway, bacterial oxalate degradation is associated with a strong local alkalini- zation and subsequent carbonate precipitation. In order to test whether this process occurs in soil, the role of bacteria, fungi and calcium oxalate amend- ments was studied using microcosms. In a model system with sterile soil amended with laboratory cul- tures of oxalotrophic bacteria and fungi, the addition of calcium oxalate induced a distinct pH shift and led to the final precipitation of calcite. However, the simultaneous presence of bacteria and fungi was essential to drive this pH shift. Growth of both oxalo- trophic bacteria and fungi was confirmed by qPCR on the frc (oxalotrophic bacteria) and 16S rRNA genes, and the quantification of ergosterol (active fungal biomass) respectively. The experiment was replicated in microcosms with non-sterilized soil. In this case, the bacterial and fungal contribution to oxalate deg- radation was evaluated by treatments with specific biocides (cycloheximide and bronopol). Results showed that the autochthonous microflora oxidized calcium oxalate and induced a significant soil alkali- nization. Moreover, data confirmed the results from the model soil showing that bacteria are essentially responsible for the pH shift, but require the presence of fungi for their oxalotrophic activity. The combined results highlight that the interaction between bacteria and fungi is essential to drive metabolic processes in complex environments such as soil.

Needle fiber calcite (NFC) is an ubiquitous terrestrial sec ondary calcium carbonate mineral often associated with calcitic nanofibers. NFC’s origin has been debated for a long time and a fungal origin is often proposed. Fungi are known to be involved in mineral weathering and production of metal oxalate but little information exists regarding the genesis of other minerals such as calcite. In this study a comparison of similar ultrastructural characteristics of fungal hyphae and NFC has been performed to highlight analogies between both features. These analogies clearly demonstrate the probable close relationship between fungal fil aments (hyphae and rhizomorphs) and NFC and its associated nanofibers.

Studies of the influence of microbial communities on calcium carbonate deposits mostly rely on classical or molecular microbiology, isotopic analyses, and microscopy. Using these techniques, it is difficult to infer microbial activities in such deposits. In this context, we used isothermal microcalorimetry, a sensitive and non- destructive tool, to measure microbial activities associated with moonmilk ex-situ. Upon the addition of diluted LB medium and other carbon sources to fresh moonmilk samples, we estimated the number of colony forming units per gram of moonmilk to be 4.8 3 10 5 6 0.2 3 10 5 . This number was close to the classical plate counts, but one cannot assume that all active cells producing metabolic heat were culturable. Using a similar approach, we estimated the overall growth rate and generation time of the microbial community associated with the moonmilk upon addition of various carbon sources. The range of apparent growth rates of the chemoheterotrophic microbial community observed was between 0.025 and 0.067 h 21 and generation times were between 10 and 27 hours. The highest growth rates were observed for citrate and diluted LB medium, while the highest carbon-source consumption rates were observed for low molecular weight organic acids (oxalate and acetate) and glycerol. Considering the rapid degradation of organic acids, glucose, and other carbon sources observed in the moonmilk, it is obvious that upon addition of nutrients during snow melting or rainfall these communities can have high overall activities comparable to those observed in some soils. Such communities can influence the physico-chemical conditions and participate directly or indirectly to the formation of moonmilk.

Stable carbon and oxygen isotope analyses were conducted on pedogenic needle fibre calcite (NFC) from seven sites in areas with roughly similar temperate climates in Western Europe, including the Swiss Jura Mountains, eastern and southern France, northern Wales, and north-eastern Spain. The δ(13)C values (-12.5 to-6.8 ‰ Vienna Pee Dee Belemnite (VPDB)) record the predominant C(3) vegetation cover at the sites. A good correlation was found between mean monthly climatic parameters (air temperature, number of frost days, humidity, and precipitation) and δ(18)O values (-7.8 to-3.4‰ VPDB) of all the NFC. Similar seasonal variations of δ(18)O values for monthly NFC samples from the Swiss sites and those of mean monthly δ(18)O values of local precipitation and meteorological data point out precipitation and preferential growth/or recrystallisation of the pedogenic needle calcite during dry seasons. These covariations indicate the potential of stable isotope compositions of preserved NFC in fossil soil horizons as a promising tool for palaeoenvironmental reconstructions.

An African oxalogenic tree, the iroko tree (Milicia excelsa), has the property to enhance carbonate precipitation in tropical oxisols, where such accumulations are not expected due to the acidic conditions in these types of soils. This uncommon process is linked to the oxalate-carbonate pathway, which increases soil pH through oxalate oxidation. In order to investigate the oxalate-carbonate pathway in the iroko system, fluxes of matter have been identified, described, and evaluated from field to microscopic scales. In the first centimeters of the soil profile, decaying of the organic matter allows the release of whewellite crystals, mainly due to the action of termites and saprophytic fungi. In addition, a concomitant flux of carbonate formed in wood tissues contributes to the carbonate flux and is identified as a direct consequence of wood feeding by termites. Nevertheless, calcite biomineralization of the tree is not a consequence of in situ oxalate consumption, but rather related to the oxalate oxidation inside the upper part of the soil. The consequence of this oxidation is the presence of carbonate ions in the soil solution pumped through the roots, leading to preferential mineralization of the roots and the trunk base. An ideal scenario for the iroko biomineralization and soil carbonate accumulation starts with oxalatization: as the iroko tree grows, the organic matter flux to the soil constitutes the litter, and an oxalate pool is formed on the forest ground. Then, wood rotting agents (mainly termites, saprophytic fungi, and bacteria) release significant amounts of oxalate crystals from decaying plant tissues. In addition, some of these agents are themselves producers of oxalate (e.g. fungi). Both processes contribute to a soil pool of "available" oxalate crystals. Oxalate consumption by oxalotrophic bacteria can then start. Carbonate and calcium ions present in the soil solution represent the end products of the oxalate-carbonate pathway. The solution is pumped through the roots, leading to carbonate precipitation. The main pools of carbon are clearly identified as the organic matter (the tree and its organic products), the oxalate crystals, and the various carbonate features. A functional model based on field observations and diagenetic investigations with δ13C signatures of the various compartments involved in the local carbon cycle is proposed. It suggests that the iroko ecosystem can act as a long-term carbon sink, as long as the calcium source is related to non-carbonate rocks. Consequently, this carbon sink, driven by the oxalate carbonate pathway around an iroko tree, constitutes a true carbon trapping ecosystem as defined by ecological theory.

Carbon and oxygen isotope compositions of the needle fibre calcite (NFC) from two soils observed in quarries (Villiers and Savagnier) in the Swiss Jura Mountains are used to investigate the processes involved in the formation of these calcite crystals in pedogenic environments. NFC occurs in soils with different macroscopic and microscopic morphologies. Three groups of microscopic morphologies are distinguished: simple needles (SN), simple needles with overgrowths (SNO), and simple needles with nanofibres (SNN), nanofibres being either organic or mineral in nature. These groups correspond to different stages in the formation and evolution of NFC. Evaluation of the δ13C and δ18O values of NFC and late calcite cements (LCC) in relationship to the composition of the carbonate host rock (CHR), and δ13CDIC from the soil waters, indicate that both NFC and LCC are precipitated in isotopic equilibrium with the soil waters. Similar δ13C and δ18O values of the elongated NFC and the rhombohedral calcite crystals of the LCC suggest that the precipitation of these different calcite habits is not due to changes in physicochemical conditions. The growth of NFC crystals inside an organic mould formed by the fungal hyphae could explain the formation of calcite crystals in the shape of a needle in isotopic equilibrium with the local environment. The comparison of the δ13C and δ18O signatures of the various NFC morphological groups suggests that they likely evolve in the soil solution from calcite crystals liberated during the degradation of fungal hyphae walls. The NFC may be a proxy of the soil conditions (temperature and vegetation) during its precipitation, and also an indicator of biogenic (fungal) activity.

The origin of soil mineralized nanofibres remains controversial. It is attributed to either biogenic factors or physicochemical processes. Scanning electron microscope and transmission electron microscope observations show that nanofibres could originate from the breakdown of fungal hyphae, especially its cell wall. It is hypothesized that during the decay of organic matter, cell wall microfibrils are released in the soil where they are exposed to mineralizing pore fluids, leading to their calcitic pseudomorphosis and/or are used as a template for calcite pre-cipitation. When associated with needle fibre calcite bundles, nanofibres could indicate the relict of an organic sheath in which calcite has precipitated. This paper emphasizes the important roles of both organic matter and fungi in carbonatogenesis, and consequently in the soil carbon cycle.

Calcium carbonate nanofibres are found in numerous terrestrial environments, often associated with needle fibre calcite. This study attempts to mimic the natural system and generate comparable crystalline structures. A comparison of natural and synthesized nanofibre structures, using HRTEM as well as electron energy loss spectroscopy (EELS) and electron spectroscopic imaging (ESI), has demonstrated that this type of nanocrystal can result from precipitation on organic templates, most likely cellulose nanofibres. This study emphasizes the fundamental role of organic templates in the precipitation of calcium carbonate in vadose environments, even at the nanoscale.

Biologically induced accumulations of calcium carbonate have been found inside orthox soils, under and around the native iroko tree Milicia excelsa (Moraceae) in Biga (Ivory Coast). The nature of these accumulations and their origin were studied in two soil profiles, directly under the tree and at a distance of 30 cm from the trunk. Microscale forms of CaCO3 include: (1) wood pseudomorphic structures such as parenchyma cells, cellulose fibers, and calcitic vessel infillings; (2) three types of rhombohedra; and (3) needle fiber calcite (NFC). In addition, large scale blocks exhibit three types of textures: (1) micritic calcite, which seems to be the original material; (2) light-colored sparite in moldic voids; and (3) asymmetrical radiaxial laminated fibrous cement. Some micritic aggregates and hemi-spherulites (vaterite) were found in the sap on the trunk as well as in soils on silica grains and the wood itself. The mineralogy of all these carbonate forms is mainly a stoichiometric calcite or a moderately enriched Mg calcite. However, some samples contain monohydrocalcite, as well as two polymorphs of calcium oxalate (weddellite and whewellite). Calcite precipitation is facilitated by the oxidation of oxalate by soil bacteria that contributes to the increase in pH in Biga soils. This is in contrast to conventional orthox soils. Therefore, three conditions are necessary for biologically induced precipitation of calcium carbonate in orthox soils associated with iroko trees: the presence of a large amount of oxalate (originating from the tree and fungi), the existence of an oxalotrophic flora for oxalate oxidation into carbonate, and a dry season.

Iroko trees ( Milicia excelsa ) in Ivory Coast and Cameroon are unusual because of their highly biomineralized tissues, which can virtually transform the trunk into stone. Oxalic acid (C 2 O 4 H 2 ) and metal-oxalate play important roles in their ecosystems. In this study, the various forms of oxalate and carbonate mineralization reactions are investigated by using scanning electron microscopy and X-ray diffraction. Calcium oxalate monohydrate is associated with stem, bark and root tissues, whereas calcium oxalate dihydrate is found with wood rot fungi in soils, as well as in decaying wood. Laboratory cultures show that many soil bacteria are able to oxidize calcium oxalate rapidly, resulting in an increase in solution pH. In terms of M . excelsa , these transformations lead to the precipitation of calcium carbonate, not only within the wood tissue, but also within the litter and soil. We calculate that c . 500 kg of inorganic carbon is accumulated inside an 80-year-old tree, and c . 1000 kg is associated with its surrounding soil. Crucially, the fixation of atmospheric CO 2 during tree photosynthesis, and its ultimate transformation into calcite, potentially represents a long-term carbon sink, because inorganic carbon has a longer residence time than organic carbon. Considering that calcium oxalate biosynthesis is widespread in the plant and fungal kingdoms, the biomineralization displayed by M. excelsa may be an extremely common phenomena.

This study stresses the role of specific bacterial outer structures (such as glycocalix and parietal polymers) on calcium carbonate crystallization in terrestrial environments. The aim is to compare calcium carbonate crystals obtained in bacterial cultures with those obtained during abiotically mediated synthesis to show implications of exopolysaccharides and amino acids in the mineralogy and morphology of calcium carbonate crystals produced by living bacteria. This is done using various amounts of purified exopolysaccharide (xanthan EPS) and L-amino acids with a range of acidities. Amino acids and increasing xanthan content enhance sphere formation in calcite and vaterite. Regarding calcite, the morphology of crystals evolves from rhombohedral to needle shape. This evolution is characterized by stretching along the c axis as the amino acid changes from glutamine to aspartic acid and as the medium is progressively enriched in EPS. Regarding vaterite, the spherulitic habit is preserved throughout the morphological sequence and starts with spheres formed by the agglomeration of short needles, which are produced in a xanthan-free medium with glutamine. Monocrystals forming spheres increase in size as xanthan is added and the acidity of amino acids (glutamic and aspartic acids) is increased. At high xanthan concentrations, amino acids, and mainly aspartic and glutamic acids, induce vaterite precipitation. The role of the carboxyl group is also probably critical because bacterial outer structures associated with peptidoglycan commonly contain carboxyl groups. This role, combined with the results presented here, clearly demonstrate the influence of bacterial outer structure composition on the morphology and mineralogy of bacterially induced calcium carbonate. This point should not be neglected in the interpretation of calcite cements and carbonate accumulations in terrestrial environments.

International audience; In mine soil, quantification of soil organic carbon (OC) derived recently from biomass decomposition is complicated by the presence of fossil (geogenic) C derived from coal, oil shale, or similar material in the overburden. The only reliable method for such measurement is 14C analysis (i.e. radiocarbon dating) using instrumentation such as accelerator mass spectrometry, which is too expensive for routine laboratory analysis. We tested two previously used and two new methods for recent C quantification and compared them with 14C AMS radiocarbon dating as a reference using a set of soil samples (n=14) from Sokolov, Czech Republic: (i) 13C isotope ratio composition, (ii) cross polarization magic angle spinning 13C nuclear magnetic resonance (CPMAS 13C NMR) spectroscopy, (iii) near infrared spectroscopy (NIRS) coupled with partial least squares regression and (iv) Rock\textendashEval pyrolysis. Conventional methods for OC determination (dry combustion, wet dichromate oxidation, loss-on-ignition) were also compared to quantify any bias connected with their use. All the methods provided acceptable recent carbon estimates in the presence of mostly aliphatic fossil C from kerogen. However, the most accurate predictions were obtained with two approaches using Rock\textendashEval pyrolysis parameters as predictors, namely (i) S2 curve components and (ii) oxygen index (OI). The S2 curve approach is based on the lower thermal stability of recent vs. fossil organic matter. The OI approach corresponded well with 13C NMR spectra, which showed that samples rich in recent C were richer in carboxyl C and O-alkyl C. These two methods showed the greatest potential as routine methods for recent C quantification.

The oxalate–carbonate pathway (OCP) is a biogeochemical set of reactions that involves the conversion of atmospheric CO2 fixed by plants into biomass and, after the biological recycling of calcium oxalate by fungi and bacteria, into calcium carbonate in terrestrial environments. Oxalotrophic bacteria are a key element of this process because of their ability to oxidize calcium oxalate. However, the diversity and alternative carbon sources of oxalotrophs participating to this pathway are unknown. Therefore, the aim of this study was to characterize oxalotrophic bacteria in tropical OCP systems from Bolivia, India, and Cameroon. Ninety-five oxalotrophic strains were isolated and identified by sequencing of the 16S rRNA gene. Four genera corresponded to newly reported oxalotrophs (Afipia, Polaromonas, Humihabitans, and Psychrobacillus). Ten strains were selected to perform a more detailed characterization. Kinetic curves and microcalorimetry analyses showed that Variovorax soli C18 has the highest oxalate consumption rate with 0.240 μM h-1. Moreover, Streptomyces achromogenes A9 displays the highest metabolic plasticity. This study highlights the phylogenetic and physiological diversity of oxalotrophic bacteria in tropical soils under the influence of the oxalate–carbonate pathway.

The oxalate-carbonate pathway (OCP) leads to a potential carbon sink in terrestrial environments. This process is linked to the activity of oxalotrophic bacteria. Although isolation and molecular characterizations are used to study oxalotrophic bacteria, these approaches do not give information on the active oxalotrophs present in soil undergoing the OCP. The aim of this study was to assess the diversity of active oxalotrophic bacteria in soil microcosms using the Bromodeoxyuridine (BrdU) DNA labeling technique. Soil was collected near an oxalogenic tree (Milicia excelsa). Different concentrations of calcium oxalate (0.5%, 1%, and 4% w/w) were added to the soil microcosms and compared with an untreated control. After 12 days of incubation, a maximal pH of 7.7 was measured for microcosms with oxalate (initial pH 6.4). At this time point, a DGGE profile of the frc gene was performed from BrdU-labeled soil DNA and unlabeled soil DNA. Actinobacteria (Streptomyces- and Kribbella-like sequences), Gammaproteobacteria and Betaproteobacteria were found as the main active oxalotrophic bacterial groups. This study highlights the relevance of Actinobacteria as members of the active bacterial community and the identification of novel uncultured oxalotrophic groups (i.e. Kribbella) active in soils.

The oxalate-carbonate pathway involves the oxidation of calcium oxalate to low-magnesium calcite and represents a potential long-term terrestrial sink for atmospheric CO(2). In this pathway, bacterial oxalate degradation is associated with a strong local alkalinization and subsequent carbonate precipitation. In order to test whether this process occurs in soil, the role of bacteria, fungi and calcium oxalate amendments was studied using microcosms. In a model system with sterile soil amended with laboratory cultures of oxalotrophic bacteria and fungi, the addition of calcium oxalate induced a distinct pH shift and led to the final precipitation of calcite. However, the simultaneous presence of bacteria and fungi was essential to drive this pH shift. Growth of both oxalotrophic bacteria and fungi was confirmed by qPCR on the frc (oxalotrophic bacteria) and 16S rRNA genes, and the quantification of ergosterol (active fungal biomass) respectively. The experiment was replicated in microcosms with non-sterilized soil. In this case, the bacterial and fungal contribution to oxalate degradation was evaluated by treatments with specific biocides (cycloheximide and bronopol). Results showed that the autochthonous microflora oxidized calcium oxalate and induced a significant soil alkalinization. Moreover, data confirmed the results from the model soil showing that bacteria are essentially responsible for the pH shift, but require the presence of fungi for their oxalotrophic activity. The combined results highlight that the interaction between bacteria and fungi is essential to drive metabolic processes in complex environments such as soil.

Needle fibre calcite is one of the most ubiquitous habits of calcite in vadose environments (caves deposits, soil pores, etc.). Its origin, either through inorganic, indirect or direct biological processes, has long been debated. In this study, investigations at 11 sites in Europe, Africa and Central America support arguments for its biogenic origin. The wide range of needle morphologies is the result of a gradual evolution of the simplest type, a rod. This rod is the elementary brick which, by aggregation and welding, builds more complex needles. The absence of cross-welded needles implies that they are welded in a mould, or under a longitudinal and unidirectional constraint, before being released inside the soil pores. The difference between the lengthening of the needles and the c axis can be explained by the existence of needles observed under a scanning electron microscope in organic sleeves, which can act as a mould during rod growth. Complex morphologies with epitaxial outgrowths on straight rods cannot have grown entirely inside organic microtubes; they must result from soil diagenesis after the release of straight rods in a soil-free medium. Whisker crystals are interpreted as the result of growth and coalescence of euhedral crystals on a rod. Rhomb chains are considered to be the consequence of successive epitaxial growth steps on a needle during variations in growth conditions. Isotopic signatures for needle fibre calcite vary from −16·63‰ to +1·10‰ and from −8·63‰ to −2·25‰ for δ13C and δ18O, respectively. The absence of high δ18O values for needle fibre calcite precludes a purely physicochemical origin (evaporative) for this particular habit of calcite. As epitaxial growth cannot precipitate in the same conditions as initial needles, needle fibre calcite stable isotopic signatures should be used with caution as a proxy for palaeoenvironmental reconstructions. In addition, it is suggested that the term needle fibre calcite should be kept for the original biogenic form. The other habit should be referred to as epitaxial forms of needle fibre calcite.

Introduction Although fungi are generally disregarded in the biogeochemical literature, they undoubtedly constitute crucial biogeochemical factors in many elemental cycles. This fact, combined with their abundance in the soil, warrants greater detailed study into their geoecological impact. The network formed by fungal filaments can represent 10 000 km of thread-like mycelia in 1 m 2 of fertile soil. Their mass is evaluated at 3500 kg ha − 1 at a depth of 20 cm in an average continental soil, i.e. taking into account all the different terrestrial environments on Earth (Gobat et al ., 2004). In comparison, bacteria and algae would represent 1500 and 10–1000 kg ha − 1 respectively, in the same virtual average soil. Fungi are not only biologically important as saprophytes in the recycling of organic matter, but also play a geological role by excreting notable amounts of organic acids, among which oxalic acid is particularly important (Gadd, 1999), contributing to continental weathering as well as to mineral neogenesis (Verrecchia & Dumont, 1996; Verrecchia, 2000; Burford et al ., 2003 a , b ). The first fossil fungi have been identified in rocks dated from the Ordovician, i.e. 460 to 455 Ma ago (Redecker et al ., 2000). However, molecular clock estimates for the evolution of fungi have suggested a Late Precambrian (600 Ma) colonization on land (Berbee & Taylor 2000). Recent molecular studies, based on protein sequence analysis, indicate that fungi were present on continents 1 billion years ago and possibly affected (together with plants) the evolution of Earth's atmosphere and climate since 700 Ma (Heckman et al ., 2001).

Carbon sequestration in the global carbon cycle is almost always attributed to organic carbon storage alone, while soil mineral carbon is generally neglected. However, due to the longer residence time of mineral carbon in soils (10(2)-10(6) years), if stored in large quantities it represents a potentially more efficient sink. The aim of this study is to estimate the mineral carbon accumulation due to the tropical iroko tree ( Milicia excelsa) in Ivory Coast. The iroko tree has the ability to accumulate mineral carbon as calcium carbonate (CaCO(3)) in ferralitic soils, where CaCO(3) is not expected to precipitate. An estimate of this accumulation was made by titrating carbonate from two characteristic soil profiles in the iroko environment and by identifying calcium (Ca) sources. The system is considered as a net carbon sink because carbonate accumulation involves only atmospheric CO(2) and Ca from Ca-carbonate-free sources. Around one ton of mineral carbon was found in and around an 80-year-old iroko stump, proving the existence of a mineral carbon sink related to the iroko ecosystem. Conservation of iroko trees and the many other biomineralizing plant species is crucial to the maintenance of this mineral carbon sink.

Aims The Oxalate-Carbonate Pathway (OCP) is a biogeochemical process that transfers atmospheric CO2 into the geologic reservoir as CaCO3; however, until now all investigations on this process have focused on species with limited food benefits. This study evaluates a potential OCP associated with Brosimum alicastrum, a Neotropical species with agroforestry potential (ca. 70–200 kg-nuts yr−1), in the calcareous soils of Haiti and Mexico. Methods / results Enzymatic analysis demonstrated significant concentrations of calcium oxalate (5.97 % D.W.) were associated with B. alicastrum tissue in all sample sites. The presence of oxalotrophism was also confirmed with microbiological analyses in both countries. High concentrations of total calcium (>7 g kg−1) and lithogenic carbonate obscured the localised alkalinisation and identification of secondary carbonate associated with the OCP at most sample sites, except Ma Rouge, Haiti. Soils adjacent to subjects in Ma Rouge demonstrated an increase in pH (0.63) and CaCO3 concentration (5.9 %) that, when coupled with root-like secondary carbonate deposits in Mexico, implies that the OCP does also occur in calcareous soils. Conclusions Therefore this study confirms that the OCP also occurs in calcareous soils, adjacent to B. alicastrum, and could play a fundamental and un-accounted role in the global calcium-carbon coupled cycle.