Putative novel hydrogen- and iron-oxidizing sheath-producing Zetaproteobacteria thrive at the Fåvne deep-sea hydrothermal vent field

Abstract

Knowledge on microbial iron oxidation is important for understanding the cycling of iron, carbon, nitrogen, nutrients, and metals. The current study yields important insights into the niche sharing, diversification, and Fe(III) oxyhydroxide morphology of Ghiorsea, an iron- and hydrogen-oxidizing Zetaproteobacteria representative belonging to Zetaproteobacteria operational taxonomic unit 9. The study proposes that Ghiorsea exhibits a more extensive morphology of Fe(III) oxyhydroxide than previously observed. Overall, the results increase our knowledge on potential drivers of Zetaproteobacteria diversity in iron microbial mats and can eventually be used to develop strategies for the cultivation of sheath-forming Zetaproteobacteria.

Keywords: Zetaproteobacteria; genome-resolved metagenomics; hydrogen oxidation; hydrothermal vents; iron oxidation; microbial mats.

Fig 1

Fe mats at Fåvne are dominated by Fe(III) oxyhydroxide sheaths produced by Zetaproteobacteria. (a) Fe mats on black smoker chimneys at Fåvne vent field with hydraulic suction device (biosyringe) used for sampling on the right. (b) Measuring temperatures within Fe mats using an isobaric gas-tight sampler (55). (c) Fe mats dominated by Fe(III) oxyhydroxide sheaths of two different widths produced by Zetaproteobacteria, scanning electron microscopy. Two Fe(III) oxyhydroxide sheath morphotypes, either 2 µm or 1 µm wide. (d) Zetaproteobacteria cells inside Fe(III) oxyhydroxide sheaths (stained with Zeta674 fluorescence in situ hybridization probe). Overlay of phase-contrast and florescence images.

Fig 2

Phylogeny of Zetaproteobacteria in black smoker Fe mats at Fåvne. The tree is based on a concatenated alignment of a manually curated set of 12 single-copy gene markers (see Table S5 at https://doi.org/10.5281/zenodo.8297777) using MAGs from this study and references. Blue genomes have been reconstructed from the Fåvne vent field. Genomes highlighted in orange are present in Fe mat associated to a focused flow black smoker (Ghiorsea, ZetaOTU9). MAGs above 0.5 coverage were selected. Black node circles mark branches with support values higher than 75% with standard bootstrapping and 1,000 iterations. The maximum likelihood tree with substitution model Qpfam + F + I + I + R7. The Ghiorsea genus is based on GTDB taxonomy r214 and AAI values within the proposed 65% AAI cutoff for genus (69).

Fig 3

Phylogeny of the large subunit of uptake Ni,Fe hydrogenase (hya; 1d). Phylogenetic tree of the large subunit of uptake Ni,Fe hydrogenase (hya; 1d) present in MAGs in the black smoker Fe mat and in all publicly available Zetaproteobacteria genomes, with closest relative reference using BLAST. Blue MAGs have been reconstructed from the Fåvne vent field. Black node circles mark branches with support values higher than 75% with standard bootstrapping and 1,000 iterations. Maximum likelihood tree with substitution model LG + I + I + R7.

Fig 4

Phylogeny of outer membrane cytochrome Cyc2. Phylogenetic tree of Fe oxidation cytochrome Cyc2 present in MAGs in the black smoker Fe mat including amino acid sequences from all publicly available Zetaproteobacteria genomes with closest relative references using BLAST. Blue labels are sequences from MAGs reconstructed from the Fåvne vent field in the current study. Support values for branches are calculated with standard bootstrapping and 1,000 iterations. The maximum likelihood tree was constructed using the substitution model Qpfam + F + I + I + R5.

Fig 5

Functional characterization of the top 25 abundant MAGs in the black smoker Fe mat. Distribution of genes involved in the utilization of a range of electron donors and electron acceptors. The number of genomes in each taxonomic class cluster is indicated in parenthesis, and the color gradient refers to the percentage of genomes per class that encode the genes. Average completeness and contamination values for each taxonomic class cluster are based on CheckM2 predictions. Top 25 most abundant MAGs account for 87% of MAG coverages.

Fig 6

Membrane complexes in Ghiorsea. Electrons coming from the oxidation of Fe²⁺ are passed all the way to the high oxygen affinity terminal oxidase, leading to the generation of a proton motive force. Reverse electron transport is necessary to regenerate NADH needed for CO₂ fixation. NADH could also get replenished with the help of Ni,Fe uptake hydrogenase instead of the energy-intensive reverse electron transport. Hydrogenase can also donate electrons to the electron transport chain. ATP is generated by ATP synthase. A schematic representation of the metabolic potential of Ghiorsea, based on Ghiorsea MAGs from Fåvne and previous studies (40, 53). Created with BioRender.com.

Fig 7

Conceptual model of Fe mats on black smoker chimneys at Fåvne vent field. Fe mats are found on black smoker chimneys with focused flow high-temperature venting of fluids containing iron and hydrogen. The temperature in the Fe mat close to the chimney exterior was ~50°C. The model is based on metabolic reconstruction of MAGs of the most abundant microbial groups.