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Applied and Environmental Microbiology, November 1999, p. 5059-5065, Vol. 65, No. 11
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Reactor-Scale Cultivation of the Hyperthermophilic Methanarchaeon Methanococcus jannaschii to High Cell Densities

Biswarup Mukhopadhyay,* Eric F. Johnson, and Ralph S. Wolfe

Department of Microbiology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801

Received 16 July 1999/Accepted 25 August 1999

For the hyperthermophilic and barophilic methanarchaeon Methanococcus jannaschii, we have developed a medium and protocols for reactor-scale cultivation that improved the final cell yield per liter from ~0.5 to ~7.5 g of packed wet cells (~1.8 g dry cell mass) under autotrophic growth conditions and to ~8.5 g of packed wet cells (~2 g dry cell mass) with yeast extract (2 g liter-1) and tryptone (2 g liter-1) as medium supplements. For growth in a sealed bottle it was necessary to add Se to the medium, and a level of 2 µM for added Se gave the highest final cell yield. In a reactor M. jannaschii grew without added Se in the medium; it is plausible that the cells received Se as a contaminant from the reactor vessel and the H2S supply. But, for the optimal performance of a reactor culture, an addition of Se to a final concentration of 50 to 100 µM was needed. Also, cell growth in a reactor culture was inhibited at much higher Se concentrations. These observations and the data from previous work with methanogen cell extracts (B. C. McBride and R. S. Wolfe, Biochemistry 10:4312-4317, 1971) suggested that from a continuously sparged reactor culture Se was lost in the exhaust gas as volatile selenides, and this loss raised the apparent required level of and tolerance for Se. In spite of having a proteinaceous cell wall, M. jannaschii withstood an impeller tip speed of 235.5 cms-1, which was optimal for achieving high cell density and also was the higher limit for the tolerated shear rate. The organism secreted one or more acidic compounds, which lowered pH in cultures without pH control; this secretion continued even after cessation of growth.

* Corresponding author. Mailing address: University of Illinois at Urbana-Champaign, Department of Microbiology, B103 Chemical and Life Sciences Laboratory, 601 S. Goodwin Ave., Urbana, IL 61801. Phone: (217) 333-1397. Fax: (217) 244-8485. E-mail: biswarup{at}life.uiuc.edu.

Applied and Environmental Microbiology, November 1999, p. 5059-5065, Vol. 65, No. 11
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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