“A meta-enzyme approach” is proposed as an ecological enzymatic method to explore the potential functions of microbial communities in extreme environments such as the deep marine subsurface. We evaluated a variety of extra-cellular enzyme activities of sediment slurries and isolates from a deep subseafloor sediment core. Using the new deep-sea drilling vessel “Chikyu”, we obtained 365 m of core sediments that contained ∼2% organic matter and considerable amounts of methane from offshore the Shimokita Peninsula in Japan at a water depth of 1,180 m. In the extra-sediment fraction of the slurry samples, phosphatase, esterase, and catalase activities were detected consistently throughout the core sediments down to the deepest slurry sample from 342.5 m below seafloor (mbsf). Detectable enzyme activities predicted the existence of a sizable population of viable aerobic microorganisms even in deep subseafloor habitats. The subsequent quantitative cultivation using solid media represented remarkably high numbers of aerobic, heterotrophic microbial populations (e.g., maximally 4.4 × 107 cells cm−3 at 342.5 mbsf). Analysis of 16S rRNA gene sequences revealed that the predominant cultivated microbial components were affiliated with the genera Bacillus, Shewanella, Pseudoalteromonas, Halomonas, Pseudomonas, Paracoccus, Rhodococcus, Microbacterium, and Flexibacteracea. Many of the predominant and scarce isolates produced a variety of extra-cellular enzymes such as proteases, amylases, lipases, chitinases, phosphatases, and deoxyribonucleases. Our results indicate that microbes in the deep subseafloor environment off Shimokita are metabolically active and that the cultivable populations may have a great potential in biotechnology.
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Bale SJ, Goodman K, Rochelle PA, Marchesi JR, Fry JC, Weightman AJ, Parkes RJ (1997) Desulfovibrio profundus sp. nov., a novel barophilic sulfate-reducing bacterium from deep sediment layers in the Japan Sea. Int J Syst Bacteriol 47:515–521
Batzke A, Engelen B, Sass H, Cypionka H (2007) Phylogenetic and physiological diversity of cultured deep-biosphere bacteria from equatorial Pacific Ocean and Peru margin sediments. Geomicrobiol J 24:261–273
Biddle JF, House CH, Brenchley JE (2005) Microbial stratification in deeply buried marine sediment reflects changes in sulfate/methane profiles. Geobiology 3:287–295
Boschker HTS, Cappenberg TE (1998) Patterns of extracellular enzyme activities in littoral sediments of Lake Gooimeer, The Netherland. FEMS Microbiol Ecol 25:79–86
D’Hondt S, Rutherfold S, Spivack AJ (2002) Metabolic activity of subsurface life in deep-sea sediments. Science 295:2067–2070
D’Hondt S, Jørgensen BB, Miller DJ, Batzke A, Blake R, Cragg BA, Cypionka H, Dickens GR, Ferdelman T, Hinrichs K-U, Holm NG, Mitterer R, Spivack A, Wang G, Bekins B, Engelen B, Ford K, Gettemy G, Rutherford SD, Sass H, Skilbeck CG, Ailleo IW, Guerin G, House CH, Inagaki F, Meister P, Naehr T, Niitsuma S, Parkes RJ, Schippers A, Smith DC, Teske A, Wiegel J, Padilla CN, Acosta JLS (2004) Distribution of microbial activities in deep subseafloor sediments. Science 306:2216–2221
Hoppe H-G (2003) Phosphatase activity in the sea. Hydrobiologia 493:187–200
Inagaki F, Suzuki M, Takai K, Oida H, Sakamoto T, Aoki K, Nealson K, Horikoshi K (2003) Microbial communities associated with geological horizons in coastal subseafloor sediments from the Sea of Okhotsk. Appl Environ Microbiol 69:7224–7235
Inagaki F, Nunoura T, Nakagawa S, Teske A, Lever A, Lauer A, Suzuki M, Takai K, Delwiche M, Colwell FS, Nealson KH, Horikoshi K, D’Hondt S, Jørgensen BB (2006) Biogeographical distribution and diversity of microbes in methane hydrate-bearing deep marine sediments on the Pacific Ocean margin. Proc Natl Acad Sci USA 103:2815–2820
Kendall MM, Liu Y, Sieprawska-Lupa M, Stetter KO, Whitman WB, Boone DR (2006) Methanococcus aeolicus sp. nov., a mesophilic, methanogenic archaeon from shallow and deep marine sediments. Int J Syst Evol Microbiol 56:1525–1529
Kormas KA, Smith DC, Edgcomb V, Teske A (2003) Molecular analysis of deep subsurface microbial communities in Nankai Trough sediments (ODP Leg 190, Site 1176). FEMS Microbiol Ecol 45:115–125
Lane DJ (1991) 16S/23S sequencing. In: Stackbrandt E, Goodfellow M (eds) Nucleic acid techniques in bacterial systematics. John Wiley & Sons, New York, pp 115–176
Ludwig W, Strunk O, Westram R, Richter L, Yadhukumar HM, Buchner A, Lai T, Steppi S, Jobb G, Förster W, Brettske I, Gerber S, Ginhart AW, Gross O, Grumann S, Hermann S, Jost R, König A, Liss T, Lüßmann R, May M, Nonhoff B, Reichel B, Strehlow R, Stamatakis A, Stuckmann N, Vilbig A, Lenke M, Ludwig T, Bode A, Schleifer KH (2004) ARB: a software environment for sequence data. Nucleic Acids Res 32:1363–1371
Mikuchi JA, Liu Y, Delwiche M, Colwell FS, Boone DR (2003) Isolation of a methanogen from deep marine sediments that contain methane hydrates, and description of Methanoculleus submarines sp. nov. Appl Environ Microbiol 69:3311–3316
Newberry CJ, Webster G, Cragg BA, Parkes RJ, Weightman AJ, Fry JC (2004) Diversity of prokaryotes and methanogenesis in deep subsurface sediments from the Nankai Trough, Ocean Drilling Program Leg 190. Environ Microbiol 6:274–287
Parkes RJ, Cragg BA, Bale SJ, Getliff JM, Goodman K, Rochelle PA, Fry JC, Weightman AJ, Harvey SM (1994) Deep bacterial biosphere in Pacific Ocean sediments. Nature 371:410–413
Parkes RJ, Cragg BA, Wellsbury P (2000) Recent studies on bacterial populations and progresses in subseafloor sediments: a review. Hydrogeol J 8:11–28
Parkes RJ, Webster G, Cragg BA, Weightman AJ, Newberry CJ, Ferdelman TG, Kallmeyer J, Jørgensen BB, Aiello IW, Fry JC (2005) Deep sub-seafloor prokaryotes stimulated at interfaces over geological time. Nature 436:390–394
Poremba K, Hoppe H-G (1995) Spatial variation of benthic microbial production and hydrolytic enzymatic activity down the continental slope of the Celtic Sea. Mar Ecol Prog Ser 118:237–245
Reed DW, Fujita Y, Delwiche ME, Blackwelder DB, Sheridan PP, Uchida T, Colwell FS (2002) Microbial communities from methane hydrate-bearing deep marine sediments in a forearc basin. Appl Environ Microbiol 68:3759–3770
Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425
Schippers A, Neretin LN, Kallmeyer J, Ferdelman TG, Cragg BA, Parkes RJ, Jørgensen BB (2005) Prokaryotic cells of the deep sub-seafloor biosphere identified as living bacteria. Nature 433:861–864
Sørensen KB, Teske A (2006) Stratified communities of active archaea in deep marine subsurface sediments. Appl Environ Microbiol 72:4596–4603
Süß J, Engelen B, Cypionka H, Sass H (2004) Quantitative analysis of bacterial communities from Mediterranean sapropels based on cultivation-dependent methods. FEMS Microbiol Ecol 51:109–121
Taira A (2005) Shimokita area site survey: Northern Japan Trench seismic survey, northern Honshu, Japan. CDEX Tech Rep Vol. 2, pp 155. Available from: http://sio7.jamstec.go.jp/publication/pdf/TechRep/CDEXTRV02.pdf
Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG (1997) The clustal X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res 24:4876–4882
Wellsbury P, Goodman K, Cragg BA, Parkes RJ (2000) The geomicrobiology of deep marine sediments from Black Ridge containing methane hydrate (sites 994, 995, and 997) In: Paul CK, Matsumoto R, Wallace PJ, Dillon WP (eds) Proc ODP Sci Results 164, vol 164. Ocean Drilling Program, College Station, TX, pp 379–391
Whitman WB, Coleman DC, Wiebe WJ (1998) Prokaryotes: the unseen majority. Proc Natl Acad Sci USA 95:6578–6583
Wobus A, Bleul C, Maassen S, Scheerer C, Schuppler M, Jacobs E, Röske I (2003) Microbial diversity and functional characterization of sediments from reservoirs of different trophic states. FEMS Microb Ecol 46:331–347
Communicated by G. Antranikian.
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Kobayashi, T., Koide, O., Mori, K. et al. Phylogenetic and enzymatic diversity of deep subseafloor aerobic microorganisms in organics- and methane-rich sediments off Shimokita Peninsula. Extremophiles 12, 519–527 (2008). https://doi.org/10.1007/s00792-008-0157-7
- Deep subseafloor biosphere
- Subseafloor sediment
- Aerobic bacteria
- Methane hydrate