The Architecture of Iron Microbial Mats Reflects the Adaptation of Chemolithotrophic Iron Oxidation in Freshwater and Marine Environments.

Microbes form mats with architectures that promote efficient metabolism within a particular physicochemical environment, thus studying mat structure helps us understand ecophysiology. Despite much research on chemolithotrophic Fe-oxidizing bacteria, Fe mat architecture has not been visualized because these delicate structures are easily disrupted. There are striking similarities between the biominerals that comprise freshwater and marine Fe mats, made by Beta- and Zetaproteobacteria, respectively. If these biominerals are assembled into mat structures with similar functional morphology, this would suggest that mat architecture is adapted to serve roles specific to Fe oxidation. To evaluate this, we combined light, confocal, and scanning electron microscopy of intact Fe microbial mats with experiments on sheath formation in culture, in order to understand mat developmental history and subsequently evaluate the connection between Fe oxidation and mat morphology. We sampled a freshwater sheath mat from Maine and marine stalk and sheath mats from Loihi Seamount hydrothermal vents, Hawaii. Mat morphology correlated to niche: stalks formed in steeper O2 gradients while sheaths were associated with low to undetectable O2 gradients. Fe-biomineralized filaments, twisted stalks or hollow sheaths, formed the highly porous framework of each mat. The mat-formers are keystone species, with nascent marine stalk-rich mats comprised of novel and uncommon Zetaproteobacteria. For all mats, filaments were locally highly parallel with similar morphologies, indicating that cells were synchronously tracking a chemical or physical cue. In the freshwater mat, cells inhabited sheath ends at the growing edge of the mat. Correspondingly, time lapse culture imaging showed that sheaths are made like stalks, with cells rapidly leaving behind an Fe oxide filament. The distinctive architecture common to all observed Fe mats appears to serve specific functions related to chemolithotrophic Fe oxidation, including (1) removing Fe oxyhydroxide waste without entombing cells or clogging flow paths through the mat and (2) colonizing niches where Fe(II) and O2 overlap. This work improves our understanding of Fe mat developmental history and how mat morphology links to metabolism. We can use these results to interpret biogenicity, metabolism, and paleoenvironmental conditions of Fe microfossil mats, which would give us insight into Earth's Fe and O2 history.

Comparative Genomic Insights into Ecophysiology of Neutrophilic, Microaerophilic Iron Oxidizing Bacteria.

Neutrophilic microaerophilic iron-oxidizing bacteria (FeOB) are thought to play a significant role in cycling of carbon, iron and associated elements in both freshwater and marine iron-rich environments. However, the roles of the neutrophilic microaerophilic FeOB are still poorly understood due largely to the difficulty of cultivation and lack of functional gene markers. Here, we analyze the genomes of two freshwater neutrophilic microaerophilic stalk-forming FeOB, Ferriphaselus amnicola OYT1 and Ferriphaselus strain R-1. Phylogenetic analyses confirm that these are distinct species within Betaproteobacteria; we describe strain R-1 and propose the name F. globulitus. We compare the genomes to those of two freshwater Betaproteobacterial and three marine Zetaproteobacterial FeOB isolates in order to look for mechanisms common to all FeOB, or just stalk-forming FeOB. The OYT1 and R-1 genomes both contain homologs to cyc2, which encodes a protein that has been shown to oxidize Fe in the acidophilic FeOB, Acidithiobacillus ferrooxidans. This c-type cytochrome common to all seven microaerophilic FeOB isolates, strengthening the case for its common utility in the Fe oxidation pathway. In contrast, the OYT1 and R-1 genomes lack mto genes found in other freshwater FeOB. OYT1 and R-1 both have genes that suggest they can oxidize sulfur species. Both have the genes necessary to fix carbon by the Calvin-Benson-Basshom pathway, while only OYT1 has the genes necessary to fix nitrogen. The stalk-forming FeOB share xag genes that may help form the polysaccharide structure of stalks. Both OYT1 and R-1 make a novel biomineralization structure, short rod-shaped Fe oxyhydroxides much smaller than their stalks; these oxides are constantly shed, and may be a vector for C, P, and metal transport to downstream environments. Our results show that while different FeOB are adapted to particular niches, freshwater and marine FeOB likely share common mechanisms for Fe oxidation electron transport and biomineralization pathways.

Using in situ voltammetry as a tool to identify and characterize habitats of iron-oxidizing bacteria: from fresh water wetlands to hydrothermal vent sites.

Iron-oxidizing bacteria (FeOB) likely play a large role in the biogeochemistry of iron, making the detection and understanding of the biogeochemical processes FeOB are involved in of critical importance. By deploying our in situ voltammetry system, we are able to measure a variety of redox species, specifically Fe(ii) and O2, simultaneously. This technique provides significant advantages in both characterizing the environments in which microaerophilic FeOB are found, and finding diverse conditions in which FeOB could potentially thrive. Described here are four environments with different salinities [one fresh groundwater seep site, one beach-groundwater mixing site, one hydrothermal vent site (Mid-Atlantic Ridge), and one estuary (Chesapeake Bay)] where in situ voltammetry was deployed, and where the presence of FeOB were confirmed by either culturing methods or molecular data. The sites varied in both O2 and Fe(ii) content with O2 ranging from below the 3 µM detection limit of the electrodes at the Chesapeake Bay suboxic zone, to as high 150 µM O2 at the vent site. In addition, a range of Fe(ii) concentrations supported FeOB communities, from 3 µM Fe(ii) in the Chesapeake Bay to 300 µM in the beach aquifer. In situ electrochemistry provides the means to quickly measure these redox gradients at appropriate resolution, making it possible in real time to detect niches likely inhabited by microaerophilic FeOB, then accurately sample for proof of FeOB presence and activity. This study demonstrates the utility of this approach while also greatly expanding our knowledge of FeOB habitats.

Ferriphaselus amnicola gen. nov., sp. nov., a neutrophilic, stalk-forming, iron-oxidizing bacterium isolated from an iron-rich groundwater seep.

A neutrophilic, stalk-forming, iron-oxidizing bacterium, strain OYT1(T), which was isolated from a groundwater seep in Ohyato Park, Tokyo, Japan, was subjected to taxonomic analysis. OYT1(T) was a motile, bean-shaped, Gram-negative bacterium that was able to grow at 8-30 °C (optimally at 25-30 °C) and at pH 5.6-7.3 (optimally at pH 6.1-6.5). The strain grew microaerobically and autotrophically. Major cellular fatty acids detected were C16 : 1ω7c/C16 : 1ω6c and C16 : 0. The total DNA G+C content was 57.6 mol%. 16S rRNA gene sequence analysis revealed that strain OYT1(T) was affiliated with the class Betaproteobacteria and clustered with iron-oxidizing bacteria isolated from groundwater seeps and wetlands and with uncultured clones detected in freshwater iron-rich environments. Based on the phenotypic and phylogenetic characteristics of strain OYT1(T), we propose that the strain represents a novel species in a new genus, for which the name Ferriphaselus amnicola gen. nov., sp. nov. is proposed; the type strain of Ferriphaselus amnicola is OYT1(T) ( = JCM 18545(T) = DSM 26810(T)).

Isolation and characterization of a novel biomineral stalk-forming iron-oxidizing bacterium from a circumneutral groundwater seep.

The Fe-depositing microorganism Gallionella ferruginea was first described in 1836 based on its association with Fe-rich environments and its distinctive morphology. Since then, this morphology has been widely used to identify G. ferruginea. Researchers have isolated several Fe-oxidizing bacteria (FeOB) related to Gallionella; however, few isolates have produced organized extracellular biomineral structures, and of these, only one stalk former has a sequenced 16S rRNA gene, listed as G. ferruginea in the GenBank database. Here we report the isolation and characterization of a novel stalk-forming Fe-oxidizing bacterium, strain R-1, from a freshwater Fe seep. Despite a strong morphological similarity to G. ferruginea, this isolate has only 93.55% 16S rRNA gene sequence similarity with the previously determined sequence. R-1 only grows on Fe(II) substrates, at pH 5.6 to 7.0 and from 10°C to 35°C, with a doubling time of ∼15 h at pH 6.3 and 22°C. It is a Betaproteobacterium, most closely related to uncultured bacteria from microaerobic Fe(II)-rich groundwater springs. The most closely related isolates are Sideroxydans spp. (94.05-94.42% sequence similarity), FeOB that are not known to produce morphologically distinct minerals. To our knowledge, this is the first reported stalk-forming freshwater FeOB isolate distinct from Gallionella.