Microbial ecology of the newly discovered serpentinite-hosted Old City hydrothermal field (southwest Indian ridge)

Abstract

Lost City (mid-Atlantic ridge) is a unique oceanic hydrothermal field where carbonate-brucite chimneys are colonized by a single phylotype of archaeal Methanosarcinales, as well as sulfur- and methane-metabolizing bacteria. So far, only one submarine analog of Lost City has been characterized, the Prony Bay hydrothermal field (New Caledonia), which nonetheless shows more microbiological similarities with ecosystems associated with continental ophiolites. This study presents the microbial ecology of the 'Lost City'-type Old City hydrothermal field, recently discovered along the southwest Indian ridge. Five carbonate-brucite chimneys were sampled and subjected to mineralogical and geochemical analyses, microimaging, as well as 16S rRNA-encoding gene and metagenomic sequencing. Dominant taxa and metabolisms vary between chimneys, in conjunction with the predicted redox state, while potential formate- and CO-metabolizing microorganisms as well as sulfur-metabolizing bacteria are always abundant. We hypothesize that the variable environmental conditions resulting from the slow and diffuse hydrothermal fluid discharge that currently characterizes Old City could lead to different microbial populations between chimneys that utilize CO and formate differently as carbon or electron sources. Old City discovery and this first description of its microbial ecology opens up attractive perspectives for understanding environmental factors shaping communities and metabolisms in oceanic serpentinite-hosted ecosystems.

Fig. 1. Microbathymetric map (in meters below sea level) of the Old City hydrothermal field (27° 50′ 6″ S to 64° 35′ 6″ E; southwest Indian ridge) and locations of vents (with dive photographs) sampled for this study using the remotely operated vehicle Victor 6000.

Samples were collected from the recently formed white mineral deposit of potentially active chimneys (BaC, Chan, and OCT) or from apparently less active chimneys with dark coating made of biological mats and manganese deposits (Tomo and Pimo). Details on samples can be found in Supplementary Table S1 and Supplementary methods. Videos of chimney sampling are also available at https://campagnes.flotteoceanographique.fr/campagnes/16002000/fr/. The red star in the context map reproduced from [22] indicates the region of the SWIR investigated during the ROVSMOOTH cruise. CIR central Indian ridge, RTJ Rodrigues triple junction, SEIR southeast Indian ridge.

Fig. 2. Relative abundance of taxa obtained from representative ASV sequences retrieved from Old City chimneys (at the genus and phylum level for Archaea and Bacteria, respectively and using the SILVA database release 132 [26]).

Semi-quantitative X-ray diffraction results presenting the mineralogy of sampled chimneys are also shown as pie charts. The predominance of brucite is observed in most samples while ToMo is dominated by calcium carbonates (calcite and aragonite) and PiMo shows a similarly high relative content in calcite. Cand. Nitrosopumilus Candidatus Nitrosopumilus, MBG-A Marine Benthic Group-A, BHI80–139 candidate phylum BHI80–139/NPL-UPA2. Nitrosopumilaceae and Methermicoccaceae are two archaeal families with unclassified genera (gen.). “Others” includes all taxa that do not show a relative abundance higher than 1% in at least one chimney sample. It includes Actinobacteria and Bipolaricaulota (formerly Acetothermia) phyla.

Fig. 3. Confocal laser scanning microscopy images showing different microbial morphotypes in chimney samples.

Rod-shaped cells (white arrows in a, f) and coccoidal cells (a–d) sometimes organized in filaments (d, f) are dominant in vent conduits where they attach to minerals here autofluorescing in blue (a–c). Diplococci, characteristic of methylotrophs, can be observed in Chan sample (e).

Fig. 4. Maximum likelihood phylogenetic tree of Nitrospirae (Thermodesulfovibrionia), Bacteroidetes (Cyclobacteriaceae), candidate phylum BHI80–139/NPL-UPA2, Chlamydiae, Acetothermia (Acetothermiia), Chloroflexi, Actinobacteria (Actinomarinales) and Patescibacteria (Gracilibacteria and Parcubacteria) based on near full-length 16S rRNA-encoding gene reconstructed from metagenomic reads.

To highlight phylotype diversity, only abundant sequences (>1.5%) are represented. Sequences from this study are colored with respect to chimney samples (Supplementary Table S1) with normalized relative abundance in brackets (%). Environmental clone accession numbers are indicated in brackets with serpentinization-related ecosystems in bold. They correspond to the closest hits to the near full-length 16S rRNA-encoding gene sequences reported in this study using the SILVA database release 132 [26]. Bootstrap values over 70% support based on 1,000 replicates are indicated by black dots on their respective nodes. LCHF Lost City hydrothermal field, PBHF Prony Bay hydrothermal field, EPR East Pacific rise, JdFR Juan de Fuca ridge, GoM, Gulf of Mexico.

Fig. 5. Normalized abundance of key genes in chimney metagenomes according to PROKKA [39] and KEGG orthology [41] annotations.

a Genes involved in C1 compounds’ metabolisms. b Genes involved in sulfur, nitrogen, and oxygen metabolisms. Each gene and its function is described in Supplementary Table S3.

Fig. 6. Overview of metabolic pathways’ completion and of the presence of key enzyme-encoding genes in dominant microbial lineages of chimney metagenomes.

Colored boxes highlight the presence and completion of metabolic pathways (a) or key genes coding for the enzyme indicated (b) as determined from KEGG module and orthology [41]. Metabolic pathway that are missing half of all required genes are considered undetected. Nitrogen fixation and methanogenesis are not indicated due to their absence or poor completion in all microbial lineages investigated.

Fig. 7. Normalized abundance of genes encoding hydrogenases.

The different groups or subgroups [42, 43] are identified in brackets. Each group of hydrogenases identified in our metagenomic data and its functions is described in Supplementary Table S3.

Fig. 8. Distribution of average carbon oxidation state (Z_C) estimated for the subsampled proteins predicted from metagenomes.

For each chimney sample, the vertical distribution was artificially computed to avoid overlap. The gray dashed vertical line highlights mean Z_C value (−0.168) obtained at OCHF for all metagenomes and dark dots indicate the mean Z_C value for each predicted proteome with distance from the mean Z_C value. Z_C values calculated by [44] for Lost City hydrothermal chimneys (red dashed line) and ambient seawater near the Axial seamount, north east Pacific Ocean (blue dashed line), are indicated for comparison. Cand. candidate phylum.