Chemosynthetic organisms flourish around deep-sea hydrothermal vents where energy-rich fluids are emitted from metal sulfide chimneys. However, microbial life hosted in mineral assemblages in extinct chimneys lacking fluid venting remains largely unknown. The interior of extinct chimneys remains anoxic where the percolation of oxygenated seawater is limited within tightly packed metal sulfide grains. Given the scarcity of photosynthetic organics in deep seawater, anaerobic microbes might inhabit the grain boundaries energetically depending on substrates derived from rock-water interactions. In this study, we reported ultra-small cells directly visualized in grain boundaries of CuFeS2 inside an extinct metal sulfide chimney from the southern Mariana Trough. Nanoscale solid analyses reveal that ultra-small cells are coated with Cu2O nanocrystals in grain boundaries enriched with C, N, and P. In situ spectroscopic and spectrometric characterizations demonstrate the distribution of organics with amide groups and a large molecular organic compound in the grain boundaries. We inferred that the ultra-small cells are anaerobes because of the fast dissolution of Cu2O nanocrystals in oxygenated solution. This Cu2O property also excludes the possibility of microbial contamination from ambient seawater during sampling. It is shown by 16S rRNA gene sequence analysis that the chimney interior is dominated by Pacearchaeota known to have anaerobic metabolisms and ultra-small cells. Our results support the potential existence of photosynthesis-independent microbial ecosystems in grain boundaries in submarine metal sulfides deposits on the early Earth.

Magnetosome-producing microorganisms can sense and move toward the redox gradient and have been extensively studied in terrestrial and shallow marine sediment environments. However, given the difficulty of sampling, magnetotactic bacteria (MTB) are poorly explored in deep-sea hydrothermal fields. In this study, a deep-sea hydrothermal vent chimney from the Southern Mariana Trough was collected using a remotely operated submersible. The mineralogical and geochemical characterization of the vent chimney sample showed an internal iron redox gradient. Additionally, the electron microscopy of particles collected by magnetic separation from the chimney sample revealed MTB cells with bullet-shaped magnetosomes, and there were minor occurrences of cuboctahedral and hexagonal prismatic magnetosomes. Genome-resolved metagenomic analysis was performed to identify microorganisms that formed magnetosomes. A metagenome-assembled genome (MAG) affiliated with Nitrospinae had magnetosome genes such as mamA, mamI, mamM, mamP, and mamQ. Furthermore, a diagnostic feature of MTB genomes, such as magnetosome gene clusters (MGCs), including mamA, mamP, and mamQ, was also confirmed in the Nitrospinae-affiliated MAG. Two lines of evidence support the occurrence of MTB in a deep-sea, inactive hydrothermal vent environment.

Chemosynthetic organisms flourish around deep-sea hydrothermal vents where energy-rich fluids are emitted from metal sulfide chimneys. In contrast to actively venting chimneys, the nature of microbial life in extinct chimneys without fluid venting remains largely unknown. Here, the occurrence of ultra-small cells in silica-filled grain boundaries inside an extinct chimney is demonstrated by high-resolution bio-signature mapping. The ultra-small cells are associated with extracellularly precipitated Cu2O nanocrystals. Single-gene analysis shows that the chimney interior is dominated by a member of Pacearchaeota known as a major phylum of DPANN. Genome-resolved metagenomic analysis reveals that the chimney Pacearchaeota member is equipped with a nearly full set of genes for fermentation-based energy generation from nucleic acids, in contrast to previously characterized Pacearchaeota members lacking many genes for nucleic acid fermentation. We infer that the ultra-small cells associated with silica and extracellular Cu2O nanocrystals in the grain boundaries are Pacearchaeota, on the basis of the experimentally demonstrated capability of silica to concentrate nucleic acids from seawater and the presence of Cu-exporting genes in a reconstructed Pacearchaeota genome. Given the existence of ~3-billion-year-old submarine hydrothermally deposited silica, proliferation of microbial life using silica-bound nucleic acids might be relevant to the primitive vent biosphere.