Abstract

The deep sedimentary biosphere, extending 100s of meters below the seafloor harbors unexpected diversity of Bacteria, Archaea, and microbial eukaryotes. Far less is known about microbial eukaryotes in subsurface habitats, albeit several studies have indicated that fungi dominate microbial eukaryotic communities and fungal molecular signatures (of both yeasts and filamentous forms) have been detected in samples as deep as 1740 mbsf. Here, we compare and contrast fungal ribosomal RNA gene signatures and whole community metatranscriptomes present in sediment core samples from 6 and 95 mbsf from Peru Margin site 1229A and from samples from 12 and 345 mbsf from Canterbury Basin site U1352. The metatranscriptome analyses reveal higher relative expression of amino acid and peptide transporters in the less nutrient rich Canterbury Basin sediments compared to the nutrient rich Peru Margin, and higher expression of motility genes in the Peru Margin samples. Higher expression of genes associated with metals transporters and antibiotic resistance and production was detected in Canterbury Basin sediments. A poly-A focused metatranscriptome produced for the Canterbury Basin sample from 345 mbsf provides further evidence for active fungal communities in the subsurface in the form of fungal-associated transcripts for metabolic and cellular processes, cell and membrane functions, and catalytic activities. Fungal communities at comparable depths at the two geographically separated locations appear dominated by distinct taxa. Differences in taxonomic composition and expression of genes associated with particular metabolic activities may be a function of sediment organic content as well as oceanic province. Microscopic analysis of Canterbury Basin sediment samples from 4 and 403 mbsf produced visualizations of septate fungal filaments, branching fungi, conidiogenesis, and spores. These images provide another important line of evidence supporting the occurrence and activity of fungi in the deep subseafloor biosphere.

Highlights

  • The marine deep subsurface biosphere includes both sedimentary and oceanic crustal habitats hydrated by subsurface fluid flows

  • We analyzed sediments from the same Peru Margin site as well as aerobic sediments from Canterbury Basin (Site U1352) in order to expand on those first data on subsurface gene expression for the whole community, and to evaluate whether transcript expression is distinct at comparable depths in these two very different habitats

  • In our eukaryote-focused metatranscriptome from 345 mbsf at Canterbury Basin, celldivision transcripts were retrieved indicating actively dividing fungal cells at 345 mbsf., Fungal transcripts involved in mycelium development, filamentous growth, fungal cell membrane, and hyphal growth were detected, providing concrete evidence for active fungal growth in deeply buried sediment

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Summary

Introduction

The marine deep subsurface biosphere includes both sedimentary and oceanic crustal habitats hydrated by subsurface fluid flows. Total prokaryotic cell abundance in marine sediments was recently estimated to be 5.39 × 1029 cells, which is comparable to global estimates of cell numbers in seawater and soil (Parkes et al, 2014) This accounts for only 0.18– 3.6% of total global biomass (Kallmeyer et al, 2012), given the physical extent of potential sedimentary and rock subsurface habitats, contributions to global elemental cycling may be nontrivial in locations where sufficient carbon, electron donors and acceptors, and other nutrient sources exist (reviewed by Orcutt et al, 2013). Microorganisms in subsurface habitats confront increased challenges with increasing depth below seafloor These include increased pressure, elevated temperatures, more refractory organic carbon pools, pH gradients resulting from fluid flow through sediments and rocks with different mineralogy, and in some habitats, decreased pore spaces. The marine subsurface is overall, a very heterogeneous amalgam of habitats more or less conducive to microbial life, with gradients that can vary on the sub millimeter scale (Schrenk et al, 2010)

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