Original Paper


, Volume 11, Issue 4, pp 563-576

First online:

Diversity of microbial eukaryotes in sediment at a deep-sea methane cold seep: surveys of ribosomal DNA libraries from raw sediment samples and two enrichment cultures

  • Kiyotaka TakishitaAffiliated withJapan Agency for Marine-Earth Science and Technology (JAMSTEC) Email author 
  • , Naoji YubukiAffiliated withGraduate School of Life and Environmental Sciences, University of Tsukuba
  • , Natsuki KakizoeAffiliated withJapan Agency for Marine-Earth Science and Technology (JAMSTEC)College of Bioresource Sciences, Nihon University
  • , Yuji InagakiAffiliated withGraduate School of Life and Environmental Sciences, University of TsukubaCenter for Computational Sciences and Institute of Biological Sciences, University of Tsukuba
  • , Tadashi MaruyamaAffiliated withJapan Agency for Marine-Earth Science and Technology (JAMSTEC)

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Recent culture-independent surveys of eukaryotic small-subunit ribosomal DNA (SSU rDNA) from many environments have unveiled unexpectedly high diversity of microbial eukaryotes (microeukaryotes) at various taxonomic levels. However, such surveys were most probably biased by various technical difficulties, resulting in underestimation of microeukaryotic diversity. In the present study on oxygen-depleted sediment from a deep-sea methane cold seep of Sagami Bay, Japan, we surveyed the diversity of eukaryotic rDNA in raw sediment samples and in two enrichment cultures. More than half of all clones recovered from the raw sediment samples were of the basidiomycetous fungus Cryptococcus curvatus. Among other clones, phylotypes of eukaryotic parasites, such as Apicomplexa, Ichthyosporea, and Phytomyxea, were identified. On the other hand, we observed a marked difference in phylotype composition in the enrichment samples. Several phylotypes belonging to heterotrophic stramenopiles were frequently found in one enrichment culture, while a phylotype of Excavata previously detected at a deep-sea hydrothermal vent dominated the other. We successfully established a clonal culture of this excavate flagellate. Since these phylotypes were not identified in the raw sediment samples, the approach incorporating a cultivation step successfully found at least a fraction of the “hidden” microeukaryotic diversity in the environment examined.


Cultivation Diversity Methane seep Microbial eukaryotes SSU rDNA