Desulfobacter latus Widdel
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2. Perform all steps under anaerobic conditions. (see below)
3. Let the frozen vial thaw at room temperature under a stream of oxygen free gas. Aseptically transfer the contents of the vial to a anaerobic tube of 1648 broth. Inoculate a plate of non-selective medium with 0.1 of the culture. Inoculate a non-selective tube of broth.
4. Seal the test tube with a rubber stopper and incubate anaerobically at 30oC. Incubate the plate(s) and aerobic broth at 30oC as a purity check.
5. After two weeks, growth should be evident by turbidity through out the broth. Once growth has been established, the culture should be transferred to fresh broth every week.
6. This culture is very sensitive to oxygen; therefore steps should be taken to avoid exposure to oxygen. When the culture exhibits good growth it will remain viable for up to 1 week if stored at 4oC under anaerobic conditions.
· Tubes of media are placed under a gassing cannula system hooked to a source of oxygen free gas.
· All transfers are performed while the test tubes are on the cannula system with a gentle stream of oxygen-free gas flowing through the system.
· As the test tubes are removed from the cannula system each is sealed with butyl rubber stopper thus maintaining the anaerobic headspace.
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Widdel F. New types of acetate-oxidizing sulfate-reducing Desulfobacter species Desulfobacter hydrogenophilus new species Desulfobacter latus new species and Desulfobacter curvatus new species. Arch. Microbiol. 148: 286-291, 1987.
Validation list no. 26. Int. J. Syst. Bacteriol. 38: 328-329, 1988.