Iron Tolerant Cyanobacteria as an Effective Tool to Study Early Evolution of Life and the Development of Biosignatures

We are currently conducting preliminary studies on the diversity of iron-tolerant cyanobacteria (CB) isolated from iron-depositing hot springs in and around Yellowstone National Park (WY, USA). In conclusion, there is no consensus on the divergence of cyanobacteria from a common ancestor for either anoxygenic or oxygenic phototrophs. Anoxygenic photosynthesis may have provided energy for the common ancestor, but it is unclear what environmental pressure induced the evolving of oxygenic phototrophs. It is supposed, however, that predecessors of contemporary CB were capable of oxidizing various substrates other than water , and it is likely that Fe2+ could be one of those substrates . If that were the case, the work of entire photosystems in Precambrian cyanobacteria and/or in their predecessors could follow three scenarios (at least): 1) ferrous iron may have been oxidized in PS II but without significant effects on oxygen evolution, and environmental iron could have been oxidized either enzymatically or chemically; 2) ferrous iron may have been oxidized only enzymatically by PS II, accompanied by the repression of O2 evolution; or 3) ferrous iron may have been oxidized by PS I upon the prevalence of anoxygenic photosynthesis or without any effect on PS II. All of these scenarios will be the subject of our future studies with the aim to understand which line-ages of CB could be typical for Precambrian time

Diversity and Physiology of Siderophilic Cyanobacteria: Implication for the Bioenergetics

Prior to 2.4 Ga, global oceans were likely significantly enriched in soluble iron (Rouxel, Bekker, Edwards, 2005), a condition that is not conducive to the growth of most contemporary mesophilic cyanobacteria (CB). Recent studies of the mechanisms of iron-deficiency stress in CB suggest that contemporary mesophilic freshwater and marine B underwent long-term adaptation to a permanent decrease in soluble iron in the ocean environment (Boyer, et al., 1987; Braun, Hantke, and Koster, 1998). Of all extant environments, iron-depositing hot springs may constitute the most appropriate natural models for analysis of the transition of ancestral cyanobacteria (CB) or protocyanobacteria (PCB) (Olson, 2001) from anoxygenic photosynthesis to oxygenic one and biogeochemical processes in the late Archean and early Paleoproterozoic eras. In particular, Olson (2001) proposed the definition for PCB and postulated that the common ancestor of PCB and CB might well have used Fe(OH)+ as the principal electron donor for CO2 fixation (Widdel, et al., 1993; Ehrenreich and Widdel, 1994; Pierson and Olson, 1989; Olson, 2006). Olson (2001) proposed that the driving force for the evolution of RC2, in addition to RC1, was the necessity to use Fe(OH)+ effectively for CO2 fixation in the absence of reduced sulfur compounds. The global decrease of dissolved environmental reduced iron could have been the driving force for the transition from anoxygenic to oxygenic photosynthesis (Brown et al., 2007). Despite the insights into the ecology, evolutionary biology, paleogeobiochemistry, and astrobiology the examination of iron depositing hot springs (IDHS) could potentially provide, very few studies dedicated to the diversity and physiology of cyanobacteria inhabiting IDHS have been conducted. Here we describe the phylogeny, physiology and ultrastructure and biogeochemical activity of several recent CB isolates from two different greater Yellowstone area IDHS, e.g. LaDuke and Chocolate Pots. Phylogenetic analysis of 16S rRNA genes indicated that 6 of 12 new isolates examined could not be placed within established CB genera. Some of the isolates exhibited pronounced requirements for elevated iron concentrations, with maximum growth rates observed when 0.4-1 mM Fe(3+) was present in the media. However, the pronounced effect of iron limitation on the proliferation of siderophilic CB can be observed only after several passages through iron "free" media. TEM studies of several species of siderophilic CB revealed that the cultures JSC-3 and -11 are probably capable of some sort of pinocytosis of precipitated iron. This phenomenon may explain high tolerance of siderophilic CB to iron deficit. We also found that the stimulation of the growth of siderophilic CB by oxidized iron is accompanied by the decrease of O2 evolution by some species after addition Fe(2+) in iron "free" medium

Iron-Tolerant Cyanobacteria for Human Habitation beyond Earth

In light of the President's Moon/Mars initiative, lunar exploration has once again become a priority for NASA. In order to establish permanent bases on the Moon and proceed with human exploration of Mars, two key problems will be addressed: first, the production of O2 and second, the production of methane (CH4). While O2 is required for life support systems (LSS), both liquid O2 and CH4 are needed as an oxidizer and a propellant, respectively for the Lunar Surface Access Module (LSAM) and the Crew Exploration Vehicle (CEV). Unlike previous propulsion systems, the new CEV will use liquid oxygen (LO2) as an oxidizer and liquid methane (LCH4) as a propellant. Existing technology (e.g. hydrogen reduction) for the production of liquid oxygen from lunar regolith is very energy intensive and requires high temperature reactors. We propose an alternative approach using iron-tolerant cyanobacteria. We have found that iron-tolerant cyanobacteria (IT CB) are capable of etching iron-bearing minerals, which may lead to bonds breaking between Fe and O of common lunar mare basalt Feoxides including ilmenite, pseudobrookite, ferropseudobrookite, and armalcolite with the subsequent release of both Fe, Ti and oxygen as by-products. We also propose to use CB biomass for CH4 production as carbon stock and a propellant. Both processes can be accomplished in an energy and cost effective manner because sunlight will be used as an energy source and allows the reactions at ambient temperatures between 10-60 C. Current evaluations include assessing the thermodynamics of such biogenic reactions using a variety of nutrients and atmospheric parameters, as well as assessing the rates and species variation effects of the driving reactions

Cyanobacteria for Human Habitation beyond Earth

In light of the President s Moon/Mars initiative, lunar exploration has once again become a priority for NASA. In order to establish permanent bases on the Moon and proceed with human exploration of Mars, two key problems will be addressed: first, the production of O2 and second, the production of methane (CH4). While O2 is required for life support systems (LSS), both liquid O2 and CH4 are needed as an oxidizer and a propellant, respectively for the Lunar Surface Access Module (LSAM) and the Crew Exploration Vehicle (CEV). Unlike previous propulsion systems, the new CEV will use liquid oxygen (LO2) as an oxidizer and liquid methane (LCH4) as a propellant. Existing technology (e.g. hydrogen reduction) for the production of liquid oxygen from lunar regolith is very energy intensive and requires high temperature reactors. We propose an alternative approach using iron-tolerant cyanobacteria. We have found that iron-tolerant cyanobacteria (IT CB) are capable of etching iron-bearing minerals, which may lead to bonds breaking between Fe and O of common lunar mare basalt Fe-oxides including ilmenite, pseudobrookite, ferropseudobrookite, and armalcolite with the subsequent release of both Fe, Ti and oxygen as byproducts. We also propose to use CB biomass for CH4 production as carbon stock and a propellant. Both processes can be accomplished in an energy and cost effective manner because sunlight will be used as an energy source and allows the reactions at ambient temperatures between 10-60 C. Current evaluations include assessing the thermodynamics of such biogenic reactions using a variety of nutrients and atmospheric parameters, as well as assessing the rates and species variation effects of the driving reactions

High-Quality Draft Genome Sequence of Fischerella thermalis JSC-11, a Siderophilic Cyanobacterium with Bioremediation Potential.

Here, we report the draft genome sequence of the siderophilic cyanobacterium Fischerella thermalis JSC-11, which was isolated from an iron-depositing hot spring. JSC-11 has bioremediation potential because it is capable of both extracellular absorption and intracellular mineralization of colloidal iron. This genomic information will facilitate the exploration of JSC-11 for bioremediation

Biogeochemical Activity of Siderophilic Cyanobacteria: Implications for Paleobiogeochemistry

Understanding the patterns of iron oxidation by cyanobacteria (CB) has tremendous importance for paleobiogeochemistry, since cyanobacteria are presumed to have been involved in the global oxidation of ferrous iron during the Precambrian (Cloud, 1973). B.K. Pierson (1999, 2000) first proposed to study iron deposition in iron-depositing hot springs (ID HS) as a model for Precambrian Fe(2+) oxidation. However, neither the iron-dependent physiology of individual species of CB inhabiting iron-depositing hot springs nor their interactions with minerals enriched with iron have been examined thoroughly. Such study could shed light on ancient iron turnover. Cyanobacterial species isolated from ID HS demonstrate elevated tolerance to colloidal Fe(3+) (= 1 mM), while a concentration of 0.4 mM proved toxic for mesophilic Synechocystis PCC 6803. Isolates from ID HS require 0.4-0.6 mM Fe3+ for maximal growth while the iron requirement for Synechocystis is approximately one order of magnitude lower. We have also demonstrated that thick polysaccharide sheaths around cells of CB isolated from ID HS serve as repositories for precipitated iron. The growth of the mesophilic cyanobacteria Phromidium aa in iron-saturated (0.6 mM) DH medium did not lead to iron precipitation on its filament surfaces. However, a 14.3 fil.2 culture, isolated from an ID HS and incubated under the same conditions, was covered with dense layer of precipitated iron. Our results, taken together with Pierson s data concerning the ability of Fe2+ to stimulate photosynthesis in natural CB mats in ID HS, suggest that CB inhabiting ID HS may constitute a new group of the extremophiles - siderophilic CB. Our recent experiments have revealed for the first time that CB isolates from ID HS are also capable of biodeterioration - the etching of minerals, in particular glasses enriched with Fe, Al, Ti, O, and Si. Thus, Precambrian siderophilic cyanobacteria and their predecessors could have been involved not only in iron deposition but also in the global release of elements. The ability of siderophilic CB to participate in iron turnover make them appropriate candidates for biotechnological processes

High-Quality Draft Genome Sequence of the Siderophilic and Thermophilic Leptolyngbyaceae Cyanobacterium JSC-12.

The siderophilic, thermophilic Leptolyngbyaceae cyanobacterium JSC-12 was isolated from a microbial mat in an iron-depositing hot spring. Here, we report the high-quality draft genome sequence of JSC-12, which may help elucidate the mechanisms of resistance to extreme iron concentrations in siderophilic cyanobacteria and lead to new remediation biotechnologies

A <i>Pseudomonas aeruginosa</i> EF-Hand Protein, EfhP (PA4107), Modulates Stress Responses and Virulence at High Calcium Concentration

<div><p><i>Pseudomonas aeruginosa</i> is a facultative human pathogen, and a major cause of nosocomial infections and severe chronic infections in endocarditis and in cystic fibrosis (CF) patients. Calcium (Ca²⁺) accumulates in pulmonary fluids of CF patients, and plays a role in the hyperinflamatory response to bacterial infection. Earlier we showed that <i>P. aeruginosa</i> responds to increased Ca²⁺ levels, primarily through the increased production of secreted virulence factors. Here we describe the role of putative Ca²⁺-binding protein, with an EF-hand domain, PA4107 (EfhP), in this response. Deletion mutations of <i>efhP</i> were generated in <i>P. aeruginosa</i> strain PAO1 and CF pulmonary isolate, strain FRD1. The lack of EfhP abolished the ability of <i>P. aeruginosa</i> PAO1 to maintain intracellular Ca²⁺ homeostasis. Quantitative high-resolution 2D-PAGE showed that the <i>efhP</i> deletion also affected the proteomes of both strains during growth with added Ca²⁺. The greatest proteome effects occurred when the pulmonary isolate was cultured in biofilms. Among the proteins that were significantly less abundant or absent in the mutant strains were proteins involved in iron acquisition, biosynthesis of pyocyanin, proteases, and stress response proteins. In support, the phenotypic responses of FRD1 Δ<i>efhP</i> showed that the mutant strain lost its ability to produce pyocyanin, developed less biofilm, and had decreased resistance to oxidative stress (H₂O₂) when cultured at high [Ca²⁺]. Furthermore, the mutant strain was unable to produce alginate when grown at high [Ca²⁺] and no iron. The effect of the Δ<i>efhP</i> mutations on virulence was determined in a lettuce model of infection. Growth of wild-type <i>P. aeruginosa</i> strains at high [Ca²⁺] causes an increased area of disease. In contrast, the lack of <i>efhP</i> prevented this Ca²⁺-induced increase in the diseased zone. The results indicate that EfhP is important for Ca²⁺ homeostasis and virulence of <i>P. aeruginosa</i> when it encounters host environments with high [Ca²⁺].</p></div

Sequence analyses of EfhP.

<p><b>a.</b> Sequence alignment of the EF-hand domains in EfhP (PA4107) and CasA (YP_472788) from <i>Rhizobium etli </i><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0098985#pone.0098985-Xi1" target="_blank">[24]</a>. The predicted Ca²⁺-binding loops are underlined. <b>b.</b> The predicted transmembrane (TM) region and two Ca²⁺-binding loops (1 and 2) are shown. EF-hand domains were predicted by PROSITE. Transmembrane region and cellular localization of the protein were predicted by TMHMM.</p

Free [Ca²⁺]_in profiles of PAO1 WT (black line) and <i>efhP</i> mutant strain PAO1043 (grey line).

<p>Cultures were grown in 0²⁺. After the basal level of [Ca²⁺]_in was monitored for 1 min, 1 mM Ca²⁺ was added (indicated by the arrow) followed by further [Ca²⁺]_in measurement for 20 min.</p