Halanaerobium acetethylicum (Rengpipat et al.) Rainey et al.
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2. Perform all steps under anaerobic conditions.
3. Aseptically transfer 0.5 ml of #1476 broth to the vial and rehydrate the pellet. Transfer the suspension back into the broth tube. Inoculate a plate of a non-selective medium such as Tryptic Soy, Nutrient, or blood agar with 0.1 ml of the cell suspension.
4. Seal the tube with a rubber stopper and incubate anaerobically at 30oC. Incubate the plate(s) aerobically as a purity check.
5. After two or three days, growth should be evident as indicated by turbidity. Initial growth can take 7 days
6. When the culture exhibits good growth it will remain viable for up to 1 week if stored at 4oC under anaerobic condition.
· 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.
Always use freshly prepared anaerobic medium. If there is any question about the anaerobic condition of the medium, the medium can be reduced with the addition of 1.5% cysteine
(2.0 ml per 100 ml of medium).
Other commonly used reducing agents are sodium sulfide, cysteine, dithiothreitol, and titanium citrate. Cysteine is the reducing agent of choice since it does not cause the ferrous ammonium sulfate to precipitate.
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Validation list no. 29. Int. J. Syst. Bacteriol. 39: 205-206, 1989.
Rengpipat S, et al. Halobacteroides acetoethylicus sp.nov., a new obligately anaerobic halophile isolated from deep subsurface hypersaline environments. Syst. Appl. Microbiol. 11: 28-35, 1988.
Validation list no. 55. Int. J. Syst. Bacteriol. 45: 879-880, 1995.
Rainey FA, et al. The taxonomic study of the fermentative halophilic bacteria: description of Haloanaerobiales ord. nov., Halobacteroidaceae fam. nov., Orenia gen. nov. and further taxonomic rearrangements at the genus and species level. Anaerobe 1: 185-199, 1995.