The Tinto River system - Wiley Online Library

JOURNAL OF GEOPHYSICAL RESEARCH, VOL. 108, NO. E7, 5080, doi:10.1029/2002JE001918, 2003

Geological record of an acidic environment driven by iron hydrochemistry: The Tinto River system David C. Fernańdez-Remolar, Nuria Rodriguez, and Felipe Go´mez Centro de Astrobiologıá, INTA-CSIC, Torrejoń de Ardoz, Spain

Ricardo Amils Centro de Astrobiologıá, INTA-CSIC, Torrejoń de Ardoz, Spain Centro de Biologıá Molecular, Universidad Autońoma de Madrid, Cantoblanco, Madrid, Spain Received 16 April 2002; revised 21 March 2003; accepted 15 May 2003; published 30 July 2003.

[1] The existence of possible hematitic strata on the surface of Mars demands a search for

terrestrial analogues formed in unusual environments. This will help us to recognize and interpret environmental and, perhaps, biological signatures preserved in Mars’ hematites. Such an analogue would allow us to establish valid reference systems based on geomicrobial and biogeochemical signatures. Two different aspects place the Tinto River inside the boundaries of a natural extreme system: its high level of biological diversity and the presence of fluvial rocks formed in the same acidic conditions as in the modern system, which could predate the Tertiary. Study of both the modern system and the ancient system is necessary to understand the formation of biosignatures. A chemolithotrophic community that biooxidizes the Iberian Pyritic Belt, acidifying water (pH between 0.9 and 3.0) and favoring high concentrations of ferric iron in solution (up to 20 gL1), maintains this iron-driven system. In spite of these extreme conditions, high microbial diversity was found. Its acidic bacteria, archaea, and eukarya constitute a complex community supported by algal biomass in highly stable hydrochemical conditions, which are achieved through iron buffering. The pH is maintained at constant low levels even at very high water dilution. In these conditions, iron minerals as oxyhydroxides, hydroxides, and sulfates are formed. The modern and recent parageneses contrast with the ancient Tinto River terrace mineral associations, which show dehydrated and desulfated iron oxides. If this dehydration process is considered, these Tinto River ironstones may be a key for knowing some aquatic habitats, which may have hosted a part INDEX TERMS: 3665 Mineralogy, Petrology, and Mineral Physics: of the early Mars biosphere. Mineral occurrences and deposits; 3675 Mineralogy, Petrology, and Mineral Physics: Sedimentary petrology; 6225 Planetology: Solar System Objects: Mars; KEYWORDS: Extreme acidic environment, iron biogeochemistry, chemolithotrophs, Mars analog Citation: Fernańdez-Remolar, D. C., N. Rodriguez, F. Go´mez, and R. Amils, Geological record of an acidic environment driven by iron hydrochemistry: The Tinto River system, J. Geophys. Res., 108(E7), 5080, doi:10.1029/2002JE001918, 2003.

1. Introduction [2 ] The mining of metallic ores produces dramatic changes in the hydrochemistry of aquatic ecosystems. The contamination of natural freshwaters by acid mine drainage is a classic example described in different treatises on environmental geology. Chemolithotrophic microorganisms obtain energy by oxidation of the metalliferous substrate, mainly reduced iron and sulfur, producing low pH, which facilitates the solution of metal cations, especially ferric iron, and their transfer among the different reservoirs of the system. Once the mining leaching waters reach a habitat, a community with low diversity of extremophilic chemolitotrophs replaces the original one. These communities with Copyright 2003 by the American Geophysical Union. 0148-0227/03/2002JE001918

low diversity are considered depending on the characteristics of water which are sustained by metal ore mining activities. Moreover, when sulfide metallic ores are subjected to warm or temperate climates, acidic and ferruginous aquifers are formed as a consequence of metal mineral bioleaching [Munchmeyer, 1996]. The Tinto River is a plausible example of these environments. [3] The Tinto River area has been mined for centuries. Tartessians (1,500 B.C.), Phoenicians and Romans carried out intensive mining activity in different localities. The mines were almost inactive from the 4th until the 18th century. Mineral extraction rates from the early period can be considered negligible in comparison to the 19th and 20th centuries [Davis et al., 2000; Leblanc et al., 2000], when British and, to a lesser extent, Spanish companies mined the metalliferous substrate for sulfide, copper, silver and gold extraction. Considered a highly polluted area, the microbial

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´ NDEZ-REMOLAR ET AL.: TINTO RIVER GEOMICROBIAL SYSTEM FERNA

communities were perceived as a colonization of a contaminated habitat by chemolitotrophic microorganisms, which use iron and sulfur as electron donors and acceptors in the respiration chain to obtain energy. However, the discovery of the presence of a diverse microbial community formed not only by chemolitotrophic prokaryotes but heterotrophic bacteria, cyanobacteria, algae, fungi and heterotrophic protists, comparable to non-contaminated freshwater communities, strongly suggested that the Tinto River was an unusual natural habitat [Go´mez et al., 1993; Lo´pez-Archilla et al., 1993, 2001; Gonza´lez-Toril et al., 2001]. [4] Although we do not as yet have direct access to environment within the actively leaching ore body, we believe for the following reasons that the pyrite oxidation is under biological control [Gonza´lez-Toril et al., 2001]. The presence of the chemolithotrophic prokaryotes detected in the water table in a microcosmos experiment showed that they accelerate by several orders of magnitude the oxidation of sulfide and iron from complex sulfides. Negative control experiments showed that the oxidation rates of complex sulfides in the absence of bacteria with only water or even sulfuric acid is negligible. Major players detected by in situ hybridization and amplification with PCR were Leptospirillum ferrooxidans (oxidize only iron), Acidithiobacillus ferrooxidans (aerobically oxidize iron and sulfide) and Acidithiobacillus spp. (reduces iron in both aerobic and anaerobic conditions). In addition there are other minor players (less than 1% of the biodiversity) as Ferroplasma, Ferromicrobium spp. (iron oxidizer in aerobic conditions and reducer in anaerobic conditions) Acidimicrobium spp. (similar metabolism and phylogenically very close to Ferromicrobium), Acidithiobacillus thiooxidans (oxidize sulfides and sulfur in aerobic conditions) and Acidithiobacillus caldus (oxidize reduced sulfur compounds in aerobic conditions). Basically the iron oxidizers produce ferric iron and protons which then can be used use to oxidize other sulfides. Also some sulfur oxidizers can oxidize sulfides producing sulfuric acid. Therefore iron and sulfur oxidation by these microorganisms is the main process that drives the bioleaching of the metallic ore. [5] Moreover, these iron and sulfur cycles are fully operative. As it is well known, Acidithiobacillus ferrooxidans and other bacteria can reduce iron in anaerobic conditions. This bacterium appears in important numbers in the Tinto River system. In the lab cycles of iron oxidation can be ran followed by iron reduction only introducing or removing oxygen from the flask. The excess of ferric iron and sulfate allows the geomicrobial system to be regulated by pH, formation of oxides (ferrihydrite, goethite, hematite, magnetite), oxyhydroxides and iron sulfates. All the critical cations and anions are products of the chemolithotrophic metabolism and they can be produced by the isolated microbes in controlled experiments. Because the abiotic reactions are much slower, it is reasonable to say that the extreme conditions of the Tinto River seems to be produced by chemolithotrophic microorganisms. One of the most important conclusions for Planetary research is that the system can operate underground because light is not needed. [6] An integrated bio-geological study of the Tinto system is changing the conception of such an environment, understanding it as a community adapted to an extreme environment, that resulted in a complex system which emerged from

the interaction of, at least, three groups of variables and is driven by a diverse acidophilic community [Lo´pez-Archilla et al., 2001]. Evidence will be provided to support this hypothesis, such as the interaction between the hydrochemistry and the iron chemolithotrophic core, and the ancient hematitic sedimentary rocks deposited in a fluvial acidic environment [Phillips, 1881]. Moreover, different iron laminated structures that grow seasonally forming discontinuous beds on the riverbanks are very similar to the ancient laminated ironstones (>15% Fe). The probable presence on Mars (Sinus Meridianis) of hematitic strata [Christensen et al., 2000] increases the interest in understanding the biogeological variables that characterize iron-driven systems. In fact, the Tinto River environment is maintained by a subterranean microbial community that acidifies water and supplies ferric iron to the system by oxidation of the complex sulfides of the Iberian Pyritic Belt. The by-products of this type of community are easily recognized and can be considered biosignatures thus facilitating the design of methodologies and instruments for future planet exploration missions. [7] It is not known if analogues of the Tinto River could have colonized and developed on Mars. Moreover, by analyzing Mars Global Surveyor Thermal Spectrometer data, Christensen et al. [2000] have proposed five different environments that may produce large hematitic deposits by abiogenic pathways. It can be then argued that the Tinto River Basin and its ferric by-products are inadequate to be applied for understanding Mars early environments. Thus the study of this readily accessible environment may help to confirm or discard first syn- and diagenetic analogies and second the existence of similar communities at some period of the red planet’s history. [8] In this work, the different variables and the geomicrobial record that characterize the acidic Tinto River environment are described. The hydrochemistry and physical variables, such as iron balance and climate, are the main variables, which modulated by the chemolithotrophic community, result in the environmental stability of the system by iron buffering. Interestingly, in high acidic conditions a buffer reaction mediated by ferric iron (Fe3+ + H2O () Fe[OH]2+ + H+) regulates the sedimentary processes. Orange colored waters have been observed in winter, which indicate colloidal ferric oxihydroxide aggregation. Conversely, water acquires dark red color in summer, which is produced by a high concentration in iron. Thus the seasonal alternation will have significant consequences in the mineral paragenesis of the Tinto River sediments. The buffer reaction has been reproduced in the laboratory adding ferric clorure to an acid sulfidric solution, producing analogous alternation in water coloration. Finally, given the importance of having a reference for terrestrial and hypothetical Martian analogues of this iron geomicrobial system, the iron bioinduced sediments are also described.

2. Geographical and Geological Settings [9] Huelva province, located in southwestern Spain is crossed obliquely by the Tinto River Basin. The basin can be divided into three main zones on the basis of topological, geological and geochemical characteristics: the north, the transitional and the estuary, which have been differentiated by a Pliocene half-graben activity with a NE-SW direction


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Figure 1. Geographical cartography of the Tinto River Basin. (a) Situation of the basin in the South Atlantic area of the Iberian Peninsula (Huelva Region). (b) Tinto River Basin domains: the north area, the transitional area and the estuary area, from the source (Penã de Hierro) to the mouth (Huelva City). (c) Detailed map of the north area including modern and ancient sampling sites: Penã de Hierro 1, Alto de la Mesa 2, las Zarandas 3, Berrocal 4 and La Palma del Condado 5. [Flores, 1996] (Figure 1). Whereas the north zone occupies an uplift, with a changing landscape due to recent tectonic movements, the subsidence in the estuary zone favors a high sedimentation rate in a littoral regimen, which was started by the Flandrian transgression [Rodrı´guez et al., 1996; Clemente et al., 1998]. [10] The north zone comprises Riotinto, Nerva, Berrocal, La Palma del Condado and Niebla localities (Figure 1). This area is characterized by highlands (100 to 660 m) and a high

stability within the hydrochemical parameters such as pH, which remains between 0.9 and 3 (mean value of 2.3), and a high concentration of iron in solution (between 1.5 and 20 gL1). The geological substrate is Paleozoic and presents graywackes, shales and volcanic materials with hydrothermal mineralizations of metallic composition that make up the iron and sulfur source that sustain the Tinto River system. In these environmental conditions the iron oxyhydroxides and sulfates are the main mineral parageneses to be formed.

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[11] The transitional area is located between Huelva and Palos de la Frontera (Figure 1), although its position varies depending on the tidal regimen and hydric seasonal changes of the river. Thus fine to medium-size supratidal and intertidal alternating with sandy wedges and ferruginous deposits are the characteristic facies of the area. Therefore the hydrochemical parameters experience spatial and periodic changes manifested downstream as a pH increase (higher than 3) by seawater neutralization, and a diminution in the ferric iron concentration of fluvial water (lower than 0.2 gL1). In this area the relief is smooth and has a lower altitude (less than 100 m), with the substrate varying from Paleozoic shales to Tertiary sandstones and carbonates that are accompanied by an extensive Quaternary terrace system. [12] Near San Juan del Puerto (Figure 1) tidal activity influences the geochemistry of the water by forming reducing environments that favor ferric iron precipitation and reduction in the form of sulfides. Therefore, in these hydrochemical conditions, limolitic and pyritic sediments enriched in organic matter alternating with fine sandstone are the main facial by-products, with biogenic pyrite being the mineral paragenesis. In the estuary area, the marine influence and to a lesser extend the Odiel River produce a notable pH increase (higher than 5), as well as a further reduction in ferric iron concentration. The diminution of iron concentration in the estuary water can be correlated with the high pyrite content of the cores sampled from the sediments, indicating massive sequestering of iron by sulfate-reducing bacteria. [13] The Iberian Pyritic Belt consists of a 250-km long geological entity that is embedded in the South-Portuguese geotectonical zone of the Iberian Peninsula. These metallic ores were formed in the Hercynian Orogenesis by Vissean hydrothermalism, which intruded the acid materials of an upper Devonian to a Carboniferous volcano-sedimentary complex (Figures 2b and 2c) generated by a previous rifting event [Boulter, 1996]. The generation of the Iberian Pyrite Belt was favored by the presence of large sills that acted as caps and host rock for the ascendant fluids [Boulter, 1996]. Massive bodies of iron and copper sulfides, as well as minor quantities of lead and zinc, represent the metallic ores. This special geochemistry is also partially recorded and fractionated in the water budget and in the current bioinduced ironstones and sulfate parageneses, which can be used as tracers to follow the geobiological processes through space and time throughout the whole system. [14] Even though chemolitotrophic activity has been detected in the transitional and estuary zones, the absence of seawater influx marks the north zone as an ideal area to study in detail the geomicrobiology of the system. Moreover, the north area includes the Iberian Pyritic Belt, the mineral substrate that originates and maintains the Tinto River system by iron and sulfur bioleaching. Thus substrate, aqueous matrix and iron metabolic by-products, which result from the community interaction, can be studied in a small area of a few hundreds of square kilometers.

3. System Parameters [15] The interaction between external and internal parameters grouped into three different sets of variables drives the stability, structure and dynamic of the Tinto River geo-

microbial system. Understanding such interplay is fundamental to determine the biogeochemical pathways of matter and energy transference, which may be detected by in situ measurement (survey sensors as pH-, Eh-, O2- or conductivity-meters) and by analyzing the biomineral record in the laboratory (FTIR, XRD, TXR, ICP and MS). These qualitative and quantitative data will establish a biogeochemical model of such an iron-driven system that may be useful for predicting analogue systems elsewhere. 3.1. Climate [ 16 ] Water and temperature are external variables addressed by climate. Whereas water is an essential matrix for matter and energy exchange between different reservoirs of a given system, temperature influences and even increases the metabolic reactions in the bioreactor that sustains the whole system. Thus water and temperature are the initial parameters that must be studied in order to understand the extraordinary conditions prevalent in the Tinto River system. In this context, the existence of at least three ancient iron levels of lacustrial and fluvial origin (see below) could indicate that the paleoclimatic parameters were similar to the present climate. [17] When the temperature and water input (rainfall) are analyzed along the Tinto River Basin, a climatic gradient based on water availability and average temperature is obtained. In this sense, the climate variation is arranged approximately accordingly to the distribution of the north, transitional and estuary zones. In the north area, the thermal and pluviosity parameters correspond to a subhumid lower mesomediterranean to upper thermomediterranean climatic stage (Figure 2a), whereas the transitional and estuary areas are included in a subhumid to dry thermomediterranean climate with semiarid conditions. The highlands (north zone) present a thermal index of nearly 340, an average minimum temperature of 6°C and an Im humidity index of 21.8 [Thornthwaite, 1948], indicating temperate and semihumid conditions. Moreover, the pluviosity is greater than the potential evapotranspiration, reaching in some areas very wet conditions (higher than 100 mm each month). In these conditions the hydric balance favors a plentiful underground reservoir of water (Figure 2b) as well as the presence of water in the whole river basin even in the driest years. [18] Under these hydric and thermal circumstances, the climograms of the north, transitional and estuary zones denote bimodality in the annual water availability (Figure 2a), consisting of a humid and temperate season alternating with a dry and warm season. This seasonal bimodality influences greatly the community activity by changing hydrophysical and chemical parameters: water flow, oxygenation, salinity, element concentration, stagnation and density of water. As the chemolithotrophic community plays an important role in the sedimentation processes (see below), changes in these hydrophysical parameters will also have consequences in the sulfate and iron oxide biosedimentation. 3.2. Hydrochemistry [19] As stated before, the water chemistry of the Tinto River is strongly dependent on the composition of the metallic massif at its source. Therefore iron (1.5 to 20 gL1) and sulfur (2 to 16 gL1) are the dominant


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Figure 2. Climograms for different Tinto River Basin localities and views of the hydrothermal metallic massif in the north area of the Tinto River Basin. (a) A climatic gradient can be observed between the localities corresponding to highlands (Zufre and El Guijo) with hyperhumid conditions and excess of water (P-EVT > 0) to lowlands (Bonares and Huelva) with pluviosity below 100 mm per month and P-EVT < 0. (b) Massive sulfides (green to gray rocks of the scarp in the detail) and dark red water (emerging from underground) in Penã de Hierro’s acidic lake, two characteristic features of the Tinto system, can be observed. This location is considered the source of the Tinto River. (c) Corta Atalaya locality, showing the net boundary between a hydrothermal metallic ore and Carboniferous shales. elements in the water column, supported by chemolithotrophic pyrite oxidation (Figures 2b and 2c). The biooxidation of metallic sulfide is accompanied by proton release, lowering the environmental pH to a range between 0.9 and 3, and inducing ferric iron solution. The Tinto River water

analysis shows a high concentration of silicon in the water (0.5 to 2 gL1) as well. This element is released by the decomposition of the metallic massif during the biooxidation processes, which is essential in the biology of certain microorganisms such as diatoms. Moreover, the biooxida-

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tion of the metallic complex is accompanied by the release of many other cations such as copper (3.7 mgL1), potassium (9.2 mgL1), magnesium (445 mgL1), sodium (12.6 mgL1), calcium (70 mgL1), cobalt (3.7 mgL1), chromium (0.6 mgL1), zinc (116.2 mgL1) and lead (0.1 mgL1). [20] The iron hydrochemistry in the Tinto River is the basis of the microbial community’s regulation of the environment, since ferric iron acts as an excellent buffer keeping the pH at a constant value of 2.3. Thus, at low pH conditions, ferric iron from ferric hidroxide precipitates dissolves consuming protons to compensate the acidity, whereas at high pH values, it precipitates as ferric hydroxide supplying protons to the hydric reservoir (Figure 3a). The pH buffering maintains this particular hydrochemical environment even during high-water episodes, when torrential rains discharge huge quantities of water in minutes. In these cases, sands cemented with iron oxyhydroxides are deposited on the riverbanks as result of the iron buffering system (Figure 3a). [21] Both redox potential and oxygen content are two other important hydrochemical parameters, which result from the interaction between the metallic ore bioleaching and the climatic conditions. The redox potential ranges between 280 and 610 mV, and the oxygen content varies from atmospheric (around 8 mgL1) to anoxic concentrations (close to 0 mgL1). As stated above, the redox values and oxygen concentration depend on bioleaching. Also water density is dependent on bioleaching, but the climatic parameters control water agitation favoring oxygenation or, conversely, water stratification and stagnation. Thus, in the higher water content phase, agitation favors the oxygenation of the water column also increasing the redox potential, whereas in the drier season anoxygenic conditions and Eh is lowered by water stagnation, as a consequence of the decrease in oxygen diffusion along the water column. However, some Tinto River pools or acidic dams present long-term anoxygenic conditions, since stagnation stability is highly dependent on the water column depth. 3.3. Microbiota [22] The Tinto River ecosystem is unusual in that the biological communities are exclusively microbial and the extreme conditions of the habitat are the consequence of very active chemolithotrophy performed by iron- and sulfuroxidizing prokaryotes growing in the rich complex metallic sulfide deposits of the Iberian Pyritic Belt. In spite of the acidic pH and heavy metal concentrations of the Tinto River, a high level of biodiversity can be found in its waters: members of representative groups of Bacteria, Archaea and Eukarya (Figures 3b– 3e) [Lo´pez-Archilla et al., 2001]. [23] Using conventional and molecular ecology techniques different iron oxidizing prokaryotes were isolated and identified as Acidithiobacillus ferrooxidans (formerly Thiobacillus ferrooxidans), Leptospirillum ferrooxidans and Ferroplasma spp. In addition to the members of the Acidithiobacillus genus (At. thiooxidans and At. cuprithermicus) other sulfur oxidizing bacteria unable to oxidize iron have been also isolated [Lo´pez-Archilla et al., 2001; Gonza´lez-Toril et al., 2001]. Most of these chemolithotrophic prokaryotes are autothotrophic, thus in addition to

promoting the extreme conditions of the habitat they are also primary producers. At. ferrooxidans is able to reduce ferric iron through anaerobic respiration of reduced sulfur compounds. Probably this is an important role of this type for bacteria in the Tinto system. Using fluorescent hybridization probes members of the Ferrimicrobium and Acidimicrobium genera were also detected at rather low concentrations and at specific locations on the river. These Gram-positive bacteria were originally described as iron oxidizers and more recently as ferric iron respirers using reduced carbon compounds as electron donors in anaerobic conditions [Johnson, 1998, 1999]. Their role in the iron cycle of the Tinto system requires further investigation. [24] Acidophilic photosynthetic protists (algae) accounted for the greatest proportion of the river’s biomass (over 65%). Together with the chemolithotrophic microorganisms, algae constitute the main primary producers of the habitat. Members of the Bacillariophyta (Diatoms), Chlorophyta (Chlamydomonas, Klebsormidium and Zignema), Euglenophyta (Euglena) and Rhodophyta (Galdieria) phyla have been identified and some of them isolated [Lo´pez-Archilla et al., 2001]. There is strong evidence (rDNAs) of the existence of acidophilic cyanobacteria in the Tinto River, although we have been unable, so far, to purify them. Further characterization will be required to ascertain the role of oxygenic photosynthetic bacteria in the primary production of the Tinto ecosystem [Gonza´lez-Toril et al., 2001]. [25] A large number of heterotrophic bacteria were isolated from enrichment cultures. Many of them did not grow after the second or third transfer, probably as a result of the dilution by serial transfer of a limiting growth factor. Some of the isolates were identified as members of the Acidiphilium genus [Lo´pez-Archilla et al., 2001; Gonza´lez-Toril et al., 2001]. All known species of the heterotrophic genus Acidiphilium are also facultative anaerobic respirers, capable to couple the oxidation of organic substrates to the reduction of ferric iron. In contrast with At. ferrooxidans, ferric reduction by some Acidiphilium spp. can occur in the presence of oxygen [Bridge and Johnson, 2000]. These bacteria are important elements in the Tinto system iron cycle, as they accelerate the reductive dissolution of many ferric iron-containing minerals, like ferrihydrite, jarosites and goethite. Other bacterial isolates were Gram-positive bacilli, aerobic spore formers of the genus Bacillus. These bacteria must exist in the river as active members (vegetative state) and not as passive resistant forms (spore) because they are sensitive to heat denaturation. rDNA amplification allowed sequences with a high level of homology with sulfate reducing bacteria to be identified [Gonza´lez-Toril et al., 2001]. So far, these important microorganisms related with the sulfur cycle, have not been isolated in the most acidic part of the river, although they have been isolated in the estuary zone. They are responsible for the massive pyrite precipitation in the anoxic sediments of this area. Recent reports described the isolation of acidophilic sulfate reducing bacteria in acid mine drainage ecosystems, which means that it is reasonable to assume that they may exist in the acidic zones of the Tinto River [Johnson, 1999]. [26] Within the decomposers, fungi, including both yeast and filamentous forms, showed a high abundance and diversity. A high percentage of the isolated hyphomycetes were able to grow in the Tinto River conditions. Some of


Figure 3. Schematic representation of the geomicrobiological process operating in the Tinto River and some representatives of the Tinto River microbiota. (a) The biooxidation of the sulfide massif acidifies the environment and releases ferric and ferrous iron that reach the basin surface originating the extreme conditions found on the river. In these hydrochemical conditions, ferric iron in solution forms hydroxide complexes releasing protons, which maintain the pH. In some cases the ferric iron is reduced to ferrous iron, which can be then reoxidized by chemolithotrophic prokaryotes. Matter and energy transfer from highlands to lowlands is continuously produced. (b) Electron microscopy of Acidithiobacillus cuprithermicus, a chemolithotrophic bacteria isolated from the Tinto River growing on chalcopyrite (white arrows). (c) Fluorescent microscopy of a sample from the origin stained with a specific probe for Leptospirillum ferrooxidans. (d) Acidophilic diatoms. (e) Acidophilic dematiaceous fungi.

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the isolated yeast species can be found in other less extreme aquatic environments. But the isolated dematiaceous fungi seem to be specific for the extreme conditions of the river, since they are rarely found in neutral fresh waters [Lo´pez-Archilla et al., 2001]. [27] Among the eukaryotes, heterotrophic protists constitute the major consumer group in the Tinto ecosystem. Different flagellates (phylum Zoomastigina), amoeba of the class Lobosea (phylum Rhizopoda), some representatives of the class Heliozoa (phylum Actinopoda) and ciliates ( phylum Ciliophora) have been observed mainly associated to biofilms [Lo´pez-Archilla et al., 2001]. Studies of the microbial ecology of the acidophilic community of the Tinto River show that the river ecosystem can be described as function of low pH, high iron and metal concentration, and high biological productivity [Lo´pez-Archilla et al., 2001].

4. Geomicrobial System By-Products: Sulfateand Iron-Rich Sediments [28] The interplay between climate, hydrochemistry and microbiota produces iron-rich deposits, which are useful to understand and validate the Tinto model system. By considering in this model the buffer reaction stated before, which is fed by the pyritic massif bioleaching, different iron oxihydroxide facies should be generated. As stated, the study of the bed architecture and mineralogy of iron sediments is vital to the future recognition of similar systems in other terrestrial and extraterrestrial environments. The characterization of biogeochemical signatures of acidic irondependent communities based on minor and trace element geochemistry, as well as on isotope analysis, will be the next step to be taken in the geomicrobiological study of the Tinto system. In order to achieve this goal we have started with the structural, mineralogical, sedimentological, paleontological, isotopic and trace element geochemistry characterization of the metabolic products of metallic sulfidic chemolithotrophy. [29] Another significant aspect of the Tinto River system is the existence of ancient iron deposits formed in different fluvial regimens, but under similar hydrochemical and microbial conditions. First, they are primordial to demonstrate that the present Tinto River geomicrobial system is a natural entity and not a highly contaminated habitat. Second, they are the starting point in the elaboration of a reference for detecting similar traces of chemosynthetic independent paleosystems on other planets. 4.1. Iron Formations [30] In the north area of the Tinto River Basin, three ironstone formations of hematitic and goethitic mineralogy have been detected. They originated in different sedimen-

tary environments, but their hydrochemistry has been inferred to be similar to that of modern iron rich deposits from analogous sedimentological characteristics. These three units are located near the Rıótinto and Nerva (Figure 1), occupying different heights above the modern river channel (Figures 4a – 4d). [31] The first and most extensive formation appears at the top of the ‘‘Alto de la Mesa’’ (more than 35 m above the river) near Riotinto town. It was first discovered by Phillips [1881], who described fossiliferous bog iron-ore at the top of the ‘‘Mesa de los Pinos’’ (Alto de la Mesa), underlying its sedimentary origin (Figure 4a). This lithostratigraphical unit seems to be formed by three different subunits, which show a coarsening upward tendency. The lower subunit is composed of approximately five meters of well-rounded conglomerates with thin laminar and massive levels. The second unit consists of ten meters of massive, laminar and conglomeratic strata (around 45% Fe), which can be divided into two parts. The lower is mainly made up of paraconglomerate and massive facies, whereas the upper shows a greater diversity of laminar and massive facies, with conglomeratic facies as well. The third unit consists of one-meter thick paraconglomeratic facies that are covered by thinner beds with laminar structure. In some laminar levels plant fossils have been found which correspond to the Quercus and Equisetum genera [Phillips, 1881] along with fungal hyphae. Moreover, bacteria-like morphologies in the laminar and fossiliferous facies are frequently observed by SEM (Figure 4b). [32] The facies analysis and the stratigraphical architecture study, along with the morphology and continuity of beds, indicate that an extensive fluvial environment produced this iron rich formation. In fact, the coarsening upward sequences in the middle subunit could be interpreted as fluvial episodes of channel infilling. The massive facies may belong to deeper pool environments, while the conglomeratic levels (more abundant at the top of the subunit) might represent more energetic areas of active channels. The presence of similar conglomeratic, massive and laminar beds inside the Odiel River Basin (more than 20 km from the ‘‘Alto de la Mesa’’ outcrop) would support the existence of an extensive fluvial system, although geochronological dating is needed to confirm this hypothesis. [33] The intermediate unit (at 20– 30 m above the river) crops out near the town of Nerva. It consists of terrace sediments of goethitic composition that occupy the valley of the modern river. Their characteristic facies are five-meter thick dark greenish and reddish massive ironstones (around 40% Fe) with laminar beds, although massive paraconglomerate beds with boulders have been also recognized (Figure 4c). Laminar and dome-shaped structures with several centimeters of diameter and fibrous frameworks are

Figure 4. (opposite) Images of ancient ironstones, different structures corresponding to modern laminated sediments from the Tinto River Basin and SEM from modern sediments and minerals. (a) Image composition of the oldest fluvial ironstone level at El Alto de la Mesa. (b) Two to three micron spherical structures of a sample from the oldest ironstone level that may represent record of bacterial microfossils. (c) Second fluvial ironstone level close to Nerva. (d) Holocene fluvial bars of the youngest ironstone level at Las Zarandas, which are being entrenched by the modern river. (e) Filamentous structures in modern laminated sediments of organic matter containing biogenic pyritic grains. (f and g) Details of the alternation between thin dark and thick orange lamina. (h) General aspect of a fluvial plain at Berrocal during the summer showing laminated sediments (white arrows in f ) and sulfates. (i) Iron oxide cover on streamer biofilms forming terraces during late winter.


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entities of possible biogenic origin. In some places the strata are organized in sinuous wedges of metric scale dipping toward the modern channel. If these stratigraphical characteristics are considered as a whole, the origin of the ferruginous sediments may be related to an acidic meandering river. [34] The youngest unit (1 – 2 m above the river) corresponds to abandoned river bars which are being entrenched by the modern fluvial system. Their facies are mainly represented by one-meter thick and laterally thinned strata of conglomerates and paraconglomerates with a massive matrix of iron oxide and badly sorted boulders (Figure 4d). Moreover, secondary massive laminar iron facies can be also observed locally. Although the stratigraphy and sedimentology of this unit have not been studied in detail, these sediments seem to have been form in an acidic high-energy water stream. The geometry and texture of these materials are congruent with gravel sedimentation in a braided river (conglomerate and ferric cement generation) alternating with drier anoxygenic-water episodes (non-grain-supported paraconglomerates with boulders). [35] The age of the three ferruginous units is unknown; no geochronological analysis or palaeontological record is available to provide consistent results with which to determine absolute or relative ages, respectively. However, if the climatic parameters that drive the modern sediments are considered and the paleoclimatical record is taken into account, a preliminary estimation of its stratigraphical position can then be established. In this sense, Rodrı´guez-Vidal et al. [1985] identify one ‘‘alteration profile’’ near the Bonares locality that is composed of laminar crusts and ferruginous pisolites. These materials have been interpreted as having been formed by laterization processes in a wet (1000 – 1200 mm) and warm tropical climate after a marine regression from latest lower Pliocene infralittoral to upper Pliocene environments. However, this new ironstone level rests on fluvial sand [Rodrı´guez-Vidal et al., 1985] not on iron-silicated rocks, which would have acted as a source of real ferruginous laterites after silicon leaching of the rock source. Moreover, the ‘‘laterite’’ level is covered by fluvial sands of the lowest Quaternary ‘‘Alto Nivel Aluvial’’ [Pedoń and Rodrı´guez-Vidal, 1986], which is the first marker of the fluvial imposition after the upper Neogene marine regression. Taking into account the chronostratigraphical position of the Bonares ‘‘laterite’’, an upper Pliocene to lower Pleistocene age may be inferred either for the first ironstone fluvial unit of the Tinto River Basin or for the second fluvial terrace unit. [36] By considering the stratigraphical situation of the third ironstone unit, which appears as laminated ferruginous sediments and iron-cemented bars eroded by the modern channel system, a late Pleistocene to early Holocene origin may be inferred. This estimation is consistent with the development of the Flandrian transgression that started 10,000 B.P., and reached maximum flooding 6,500 B.P. [Clemente et al., 1985; Lario et al., 2002], probably as a sedimentary infilling in the highest marine level. Afterward, the post-Flandrian regression would have driven the channel incision of the ferruginous fluvial deposits that were previously sedimented. Moreover, the ironstone genesis of these facies coincides approximately with the installation of the Mediterranean climate in the last 10,000 years [Jalut et al., 2000]. In these warm climatic conditions the iron leaching

would have produced similar iron-enriched sediments as formed in the modern environment. [37] The study of the geobiological processes that form the modern river’s iron laminar sediment allowed us to understand the origin of this facies and their environmental significance. Apparently, they are formed by cyclic succession between seasonal periods of low water table stagnation with calm water, and high water table with diluted solutions and turbulent conditions. The repetition of the cycle resulted in laminated structures comprised of bioinduced (biological uptake) and chemoinduced (buffer activation) laminas. However, massive beds were not recognized in recent sediments; perhaps they are being formed underwater and so are never exposed even in the driest period, making them impervious to seasonal influence, where the biological recycling of iron has a greater continuity. [38] The study of old and recent iron sediments, water biogeochemistry and the present community of the Tinto River give us a vision of the temporal and spatial evolution of the geomicrobiological system. For example, the determination of sulfur and iron transfer throughout the different reservoirs can be used as a tool for detecting ancient or recent biological activity; its association to bioinduced structures such as non carbonate stromatolites can be useful for establishing organic and non-organic biomarkers in these extreme conditions, as well as for predicting similar systems on the basis of chemosynthetic primary producers. Once a thorough investigation has been carried out, the Tinto River community may prove to be a valuable reference system. 4.2. Iron Mineral Parageneses [39] Iron oxides and oxyhydroxides associated to sulfates are the characteristic minerals that are formed in the modern sediments (Figure 4h). The Tinto River mineral parageneses result from the alternation between oxide and saline facies due to the annual climatic cycle. However, sulfates and oxides can also precipitate together depending on the sulfate concentration of water, which would explain the ubiquitous presence of sulfates in the oxide-dominated sediments. Moreover, the presence of some transitional oxide-sulfate minerals cannot be discounted [Schwertmann and Fitzpatrick, 1992]. The oxide facies are composed of a combination of ferrihydrite (Fe5O7(OH)4H2O), goethite (Fe2O3H2O and FeOOH), hematite (Fe2O3) and magnetite (Fe3O4), their content depending on the degree of the dehydration of the mineral. Thus hematite has been detected in both recent and old sediments at different dehydration degrees, with ferrihydrite existing only in hydrated and fresh sediments, which are or have recently been precipitated onto the riverbanks or the river channel. This would explain the prevalence of hematite in the ancient iron formations in contrast to the modern sediments, which present a higher diversity in iron oxides. [40] As stated earlier, both the bioleaching process and the high evaporation rate induce the formation of peculiar acidic brines [Eugster and Hardie, 1978; Long et al., 1992], which are mainly composed of SO4 complexes [Elbaz-Poulichet and Dupuy, 1999] and metals. Two different parageneses, gypsum (CaSO42H2O) and jarosite (K2Fe6(SO4)4(OH)12), are precipitated from these brines, which are associated to different Fe-bearing sulfates [Hudson-Edwards et al., 1999]. Copiapite, coquimbite and schwertmannite have been


detected associated to jarosite-dominated sediments, with the gypsum parageneses being poorer in a number of salt species. The genetic relationship between both parageneses is not still understood, although a differential distribution has been observed along the length of the basin. Thus gypsum is predominant in areas located close to the river source, whereas the jarosite content increases downstream, where the gypsum remains a secondary phase or even diminishes to a negligible concentration. The decrease in gypsum content could be related to massive Ca sequestering in the source area because this mineral is probably less soluble than the Fe-bearing sulfates. [41] Iron sulfates, which are ubiquitous in modern sediments of the fluvial basin, decrease to negligible content in the ancient iron levels. This lowering of sulfate is also related to sulfur loss in the ancient iron levels. For example, the sulfur content in the ironstone terraces at Nerva locality reaches as much as 1.3%, with a concentration of 8.2% in some modern sediments of the source area. Therefore the lack of sulfate in the older levels cannot be explained by the reduction of sulfur, from sulfate to sulfide, which is recorded as pyrite. Probably, most of the sulfur has been solved directly by higher-pH meteoric waters during the early diagenesis [Fernańdez-Remolar et al., 2002]. Probably, the sulfate dissolution is recorded by goethite precipitation that fill the porosity produced during the sulfate removing. To a minor extent, H2S emission may be another pathway for sulfur removing. However, these hypotheses require a well-detailed mass balance that must be based first on in situ measurement of the H2S emission in different localities of the Tinto River Basin modern system.

5. Biosedimentary Model [42] As has been shown, the interplay between these three main variables (climate, hydrochemistry and microbiota) results, first, in acidification and iron bioleaching, and produces, second, the generation of a peculiar chemo- and biomineral reservoir composed of iron oxides and sulfates. Moreover, biogeochemical pathways coupled with the seasonal cycles induce the formation of laminated structures, which represent a large part of the mineral reservoir as byproducts of the transference of matter through the system. As this transfer of matter and energy maintains the geomicrobial system at a regional scale, a global mass balance of the river basin should be considered. In this sense, understanding of the Tinto River geomicrobial system requires detailed monitoring of the transfer of the main compounds through the different reservoirs, taking into account the seasonal cycles and spatial exchange of matter. Unfortunately, the cationic complexes and organics that are involved in such interactions have not been identified yet. The elemental analysis to quantify the degree of temporal and spatial exchange between the hydric and biomineral reservoirs is the first step in this process (D. C. Fernańdez-Remolar et al., work in preparation, 2003). [43] However, field observations, obtained in laminated complexes of the Berrocal locality, have shown how the seasonal matter exchange that originates the iron sedimentation occurs. These laminated sediments represent the biomineral reservoir and are essential for iron sequestering in the geological record. The record of iron sequestering by

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chemical and biological processes is formed by the repetition of couplets of red and orange laminas (Figures 4f –4c), which represent the seasonal variations of the system: orange lamina corresponding to iron hydroxides sedimentation in humid seasons, and red lamina to the sedimentation of iron oxyhydroxide and sulfate mixtures in dry seasons. [44] The model herein established is qualitative and is based on the chemical control by pH buffering that seasonal variations cause (humid versus dry season) in their parameters (Figure 5). The existence of such a hydrochemical stabilizer and its interaction with the climate would explain alternation in sedimentary processes and therefore would produce this peculiar laminated sedimentological record. Therefore, to maintain the pH stable in high water conditions the buffer reaction provides H+ by generation of colloidal iron hydroxide (Fe(OH)2+) (Figures 5a and 5b). In these humid conditions, the aqueous environment is turbulent, the oxygen content is increased up to atmospheric levels and light transmission is lowered due to the high concentration of water in colloidal hydroxides and clays. The turbulence prevents stratification in the water column and the microbial colonization of the channel substrate. Once the turbulent conditions cease, a transitional stage is reached, and the colloids precipitate as amorphous iron hydroxides and, to a minor extend, as ferric hydroxysulfates [Clarke et al., 1997]. These sediments appear as orange laminas of amorphous matter (Figure 5b). An important question is whether the orange lamina precipitation is mediated by microbial activity (Figure 5). In this sense, Clarke et al. [1997] describe passive formation of epicellular iron hydroxides by ‘‘oxidation and hydrolysis of cell-bound ferrous iron’’, as well as ‘‘binding of cationic colloidal species’’ (Figure 4i). [45] By considering the buffer reaction that drives the model, it may be expected that once the humid seasonal activity decreases, temperature and evaporation rates be increased. In these conditions colloidal hydroxides are then removed from the water solution and protons begin to concentrate in the river solution. The rise in evaporation rates in the dry season subsequently increases the proton and sulfate concentration, lowering the pH solution (Figure 5c). In this case, pH stability is also maintained by the reverse buffer reaction, i.e., ferric iron released from the dissolving colloidal ferric hydroxides consuming protons. This process can be monitored by direct observation of the river water coloration, which turns from orange to darkish red. Moreover, the flow diminution induces the changes from turbulent to laminar conditions in the water mass, favoring the microbial colonization of the channel substrate by microbial filaments. [46] The higher evaporation rate produces peculiar brines dominated by sulfate and ferric iron. Precipitation of iron oxyhydroxide and sulfate mixtures at lower pH values may be expected by the evaporation of these acidic brines, as shown by Schwertmann and Fitzpatrick [1992]. A higher concentration of iron in the sediments would produce the dark red lamina alternating with the previous orange lamina. This is consistent with the pH buffer model because of the iron releasing from hydroxides. However, prokaryotic, algal and fungal biofilms may locally form massive filamentous frameworks that induce the cationic sequestering. Thus microbial activity may partially induce the dark red lamina of oxyhydroxide and sulfate mixtures that may be accreted

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Figure 5. Scheme of the successive stages generating the modern Tinto River laminated records. (a) From early fall to early winter, the water supply from rain produces turbulences disrupting the water stratification. At this stage the ferric iron reservoir forms hydroxide complexes maintaining the pH. (b) Once the turbulent conditions are lowered, from late winter to middle spring, iron hydroxides complexes are originated either as amorphous precipitated materials or as iron hydroxides trapped by local biofilms; during this wet and temperate season orange thick lamina are formed. (c) Subsequently, in the middle to late spring, and due to a decrease of the water column, water stratification starts and ferric iron is transferred from hydroxide complexes to the water matrix consuming protons to maintain stable the pH. The sulfate increasing in water by a higher evaporation rate favors the precipitation of hydroxisulfate and oxyhydroxide mixtures as well. Most of the ferric iron is sedimented as oxyhydroxides but some may be trapped in biofilms, in oxygenated areas, whereas in anoxygenic pools is reduced to ferrous iron, being again available for bacterial oxidation. (d and e) Two final stages that produce the thin dark red lamina during the hot summers and which are dependent on the water column thickness are shown. (d) In the channel area water acquire a thick anoxygenic hypolimnion by low diffusion of oxygen and its consumption in the epilimnion by microbiota, which favors ferric iron and sulfate reduction. (e) Sulfate, ferryhidrite and pyrite are formed in the fluvial flat, depending in the dehydration degree and oxygen content.


to the geomicrobial complexes, reinforced by fungal and prokaryotic biomineralization inside the ferric sediments. In some cases, when neutral streams meet acidic waters, the origin of the red lamina can be associated to ferrihydrite or goethite precipitation by pH increasing [Clarke et al., 1997]. However, the acidic conditions of the Tinto River water prevents inorganic formation of pure oxyhydroxide lamina, remaining the formation of iron- and sulfateenriched sediments and biological mineralization as the most probable processes for the red lamina formation. Further research on the role of the microbial mineralization versus chemical precipitation is required to recognize the biogeochemical pathways that are involved in the formation of the ferruginous materials of the Tinto River Basin. [47] In the dry season, the water column experiences dramatic changes in its structure. The higher density of water, which has been increased by evaporation, and the microbial activity induce stratification and even stagnation (Figures 5c and 5d). The balance between oxygen release and uptake must determine the kind of microbial communities that colonize the substrate. Moreover, the light available in the water column is another main variable that drives the presence of algal communities in the water. Red lamina records the succession of communities of the laminated complexes. In these anoxygenic conditions, the ferric iron is reduced to ferrous iron by oxyhydroxide and sulfate mixtures dissolution using direct microbial anaerobic respiration of oxidized iron or reaction with the H2S produced by sulfate reducing bacteria, thus precipitating biogenic pyrite [Bridge and Johnson, 2000; Neal et al., 2001].

6. Analogies Between the Tinto River Basin and the Mars Hematite Sites [48] Some previous considerations concerning water content and environment temperature are required for considering the Tinto River Basin as an analog for Mars’ hematitic sites. As previously indicated, liquid water is not scarce but abundant in the Tinto River Basin. Water conversely appears as solid or vapor phases on the present-day Mars’ surface due to its harsh and dry climatic conditions. Although some theoretical and experimental studies suggest that liquid water may be present on Mars’ surface [cf. Haberle et al., 2001; Kuznetz and Gan, 2002], it rapidly ablates in the low-pressure conditions present at the surface. These environmental conditions contrast with the Tinto River, which is an extreme Terrestrial habitat sustained by high water availability. However, regional and high-resolution images of Mars, as well as spectral data provided by planetary probes provide evidential support for distinctive episodes of water release on Mars’ surface during the past [Carr, 1995, 1996; Baker, 2001]. The morphological characteristics of Mars’ northern plains were previously used to support the hypothesis of an early Mars ocean basin [Baker et al., 1991], but have lately been reconsidered as much younger. Moreover, it is well known that some craters have acted as traps for sediments deposited since the Noachian period in lake-related systems [Cabrol et al., 1999]. [49] Liquid water on ancient Mars implies warmer and wetter climatic conditions than observed on the present-day Mars. Climatic studies of the early atmospheric evolution of Mars, which used the same geomorphological and isotopic

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fractionation data [Ghosh and Mahajan, 1998] as primary evidences, indicate that during the Noachian epoch the atmospheric volatile pressure was high enough to sustain a warm and wet surface environment [Carr, 1999; Jakosky and Phillips, 2001]. Some climate models on atmospheric modeling [Forget and Pierrehumbert, 1997; Carr, 1999] suggest that the Noachian Mars, under a 0.5 to 1 bar CO2 atmosphere, hosted stable water masses that could have sustained microbial communities with different metabolic strategies [Nealson, 1997]. Although many of the physical and chemical properties of the early Mars climate [Haberle, 1998] are still not well understood, new interpretations of the Martian landscape suggest that periods of rainfall may have produced surface runoff during the early history of Mars [Craddock and Howard, 2002]. [50] A wetter scenario for Mars makes it comparable to some Terrestrial environments, in which the Tinto River and other may be included. However, the discovery of some Noachian iron lithological units on Mars, i.e., the Meridiani Hematite Formation, suggests the Tinto River Basin as a possible analog for understanding those environmental processes that drove the generation of iron oxides on the Noachian Mars. The spectral analyses of data obtained by TES indicate that Mars hematite corresponds to a gray coarse-grained phase that is similar to that found in aqueous environments [Christensen et al., 2000]. Moreover, comparisons between Sinus Meridiani TES data and Terrestrial midinfrared spectra support the claim that the Meridiani Formation is composed of platy hematite, implying burial and recrystallization as secondary processes, which was primary precipitated from Fe-rich aqueous and/or hydrothermal solutions on Early Mars [Lane et al., 2002]. Therefore, if the Sinus Meridiani oxides represent relicts of an early basin that hosted Fe-rich waters under milder climatic conditions than observed on present-day Mars, the Tinto River Basin may turn into a valuable living system for understanding the geological processes that drove the hematite genesis. Moreover, as stated earlier, the existence of mineral relicts in the Tinto River Basin will help unravel those rock-forming processes involved in the transformation of primary iron-rich sediments. Understanding these process are essential for de-encrypting the primary geological information produced during the early diagenesis. Whether the hematitic formations on Mars were formed by microbial-induced oxidation is unclear, although the high metabolic diversity of the prokaryotic life in the Tinto River suggests that microbes may have thrived in the water masses of early Mars that generated them.

7. Conclusions [51] High microbial diversity and the existence of ancient iron levels indicate that the origin of the Tinto River system is unrelated to industrial activity. Mining activities may have influenced the leaching and weathering rates in recent times, but in the absence of this peculiar chemolithotrophic community such a low-pH environment with a high iron concentration would not have occurred. Water and temperature are the main physical conditions that the microbiota needs to maintain the system. Thus the high availability of superficial and underground water, as well as an average temperature of over 15°C, accelerates the enzymatic reac-

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tions involved in the biogeochemical cycling of iron and sulfur throughout the system. This means that the three different ancient ironstone levels were formed in similar climatic conditions, since this environment is unlikely to develop in cold or drier climates. [52] Both the high microbial diversity and the existence of the three ironstone levels demonstrate that the Tinto River geomicrobial system must be considered an example of the diversity of the natural world. The existence of such a system shows that microbial activity may be able to modulate the conditions of the medium by iron buffering. This can be understood as a homeostatic process. Moreover, while the chemolithotrophic core of the Tinto River community controls environmental stability through chemical control, the photosynthetic core produces the biomass. This new microbial interplay produces a highly adaptive symbiotic relationship that may allow occupying a planetary surface in two stages. The first stage chemically and physically stabilizes the environment, and the second stage carries out the increase on biodiversity. [53] One essential parameter to be regulated in Mars’ thin atmosphere is the flux of electromagnetic spectra that reaches the water mass. However, the high concentration of ferric iron in these acidic conditions could act as an effective shield against the ultraviolet radiation. UV regulation would be the first stage of the microbial occupation of an inhospitable area. Hydrochemical stability of the environment and UV blocking effect are two important features of the Tinto River system that must be considered when searching for extant or fossil life on Mars. Whether the Martian hematitic record was the result of the activity of such an acidophillic community is unknown, but it is a good starting point for understanding how signatures of biological origin are formed in Fe-driven systems and how they can be detected in these strange environments. [54] Acknowledgments. The authors are grateful to Dr. Andrew H. Knoll and Dr. Juan Pe´rez Mercader, their comments, suggestions and continuous support. We thank to Dr. Carol Cleeland’s suggestions that have improved the manuscript. The work has been supported by grants BIO99-0184 from the Ministerio de Educacioń y Cultura and BXX20001385 supported by the DGICYT.

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R. Amils, Centro de Biologıá Molecular, Universidad Autońoma de Madrid, Cantoblanco, Madrid, E-28049, Spain. D. C. Fernańdez-Remolar, F. Go´mez, and N. Rodriguez, Centro de Astrobiologıá, INTA-CSIC, Carretera de Ajalvir km. 4, Torrejoń de Ardoz, E-28850, Spain. ([email protected])