Nematoda from the terrestrial deep subsurface of South Africa

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Since its discovery over two decades ago, the deep subsurface biosphere has been considered to be the realm of single-cell organisms, extending over three kilometres into the Earth’s crust and comprising a significant fraction of the global biosphere1, 2, 3, 4. The constraints of temperature, energy, dioxygen and space seemed to preclude the possibility of more-complex, multicellular organisms from surviving at these depths. Here we report species of the phylum Nematoda that have been detected in or recovered from 0.9–3.6-kilometre-deep fracture water in the deep mines of South Africa but have not been detected in the mining water. These subsurface nematodes, including a new species, Halicephalobus mephisto, tolerate high temperature, reproduce asexually and preferentially feed upon subsurface bacteria. Carbon-14 data indicate that the fracture water in which the nematodes reside is 3,000–12,000-year-old palaeometeoric water. Our data suggest that nematodes should be found in other deep hypoxic settings where temperature permits, and that they may control the microbial population density by grazing on fracture surface biofilm patches. Our results expand the known metazoan biosphere and demonstrate that deep ecosystems are more complex than previously accepted. The discovery of multicellular life in the deep subsurface of the Earth also has important implications for the search for subsurface life on other planets in our Solar System.

At a glance


  1. General morphology of H. mephisto.
    Figure 1: General morphology of H. mephisto.

    Light microscopy drawings of female holotype and scanning electron microscopy photograph of head. a, Entire body; b, neck region; c, anterior region; d, scanning electron microscope face view (scale bar, 1μm; black arrowheads indicate the positions of two cephalic papillae; black arrow indicates amphid opening); e, reproductive system; f, tail. SE, secretory–excretory.

  2. Bayesian-interference 50%-majority-rule consensus phylogenies based
on small-subunit rDNA data.
    Figure 2: Bayesian-interference 50%-majority-rule consensus phylogenies based on small-subunit rDNA data.

    H. mephisto with GenBank sequences of closely related taxa. Branch support is indicated with posterior probability values. Scale bar, expected substitutions per site. SSU, small subunit.

Accession codes

Primary accessions



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Author information


  1. Department of Biology, Nematology Section, Ghent University, Ledeganckstraat 35, B9000 Ghent, Belgium

    • G. Borgonie &
    • W. Bert
  2. Metagenomics Platform, Department of Microbial, Biochemical and Food Biotechnology, University of the Free State, PO Box 339, Bloemfontein 9300, South Africa

    • A. García-Moyano,
    • D. Litthauer,
    • A. Bester,
    • E. van Heerden,
    • C. Möller &
    • M. Erasmus
  3. Laboratory of Nematology, Department of Plant Sciences, Wageningen University, 6708 Wageningen, The Netherlands

    • W. Bert
  4. Department of Geosciences, Princeton University, Princeton, New Jersey 08544, USA

    • T. C. Onstott
  5. Present address: Department of Biology, University of Bergen, Postbox 7803, N-5020 Bergen, Norway.

    • A. García-Moyano


A.G.-M., D.L. and W.B. all contributed equally to this study. G.B., A.G.-M., D.L., A.B. and M.E. collected the filtered samples and the control samples and performed field analyses. G.B. carried out the enrichments. A.G.-M. performed microbial DNA extraction and 16S rRNA amplification, sequencing and tree construction. C.M. performed DNA analyses on filters of mining water. W.B. provided the nematode identification, their morphological description and their molecular analyses. T.C.O. modelled the geochemical, 3H and 14C data. G.B. wrote the paper with input from W.B., A.G.-M., T.C.O. and E.v.H.

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The authors declare no competing financial interests.

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Sequence information for H. mephisto has been deposited at GenBank under accession number GQ918144.

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Supplementary information

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  1. Supplementary Figures (1.8M)

    This file contains Supplementary Figures 1-9 with legends.

  2. Supplementary Information (895K)

    This file contains Supplementary Tables 1-5, Supplementary Methods, a Supplementary Discussion and additional references.

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