Advertisement

Archives of Microbiology

, Volume 151, Issue 5, pp 399–406 | Cite as

Enrichment and isolation of Acetitomaculum ruminis, gen. nov., sp. nov.: acetogenic bacteria from the bovine rumen

  • R. C. Greening
  • J. A. Z. Leedle
Original Papers

Abstract

Five strains of acetogenic bacteria were isolated by selective enrichment from the rumen of a mature Hereford crossbred steer fed a typical high forage diet. Suspensions of rumen bacteria, prepared from contents collected 7 h postfeeding, blended and strained through cheesecloth, were incubated in a minimal medium containing 10% clarified rumen fluid under either H2:CO2 (80:20) or N2:CO2 (80:20) headspace atmosphere. The selection criterion was an increment of acetate in the enrichments incubated under H2:CO2. Periodically, the enrichment broths were plated onto agar media and presumed acetogenic bacteria subsequently were screened for acetate production. Selected acetogenic bacteria utilized a pressurized atmosphere of H2:CO2 to form acetate in quantities 2 to 8-fold higher than when grown under N2:CO2. All presumptive acetogenic isolates were derived from either the 10-7 or 10-8 dilutions of rumen contents. All 5 strains were Gram-positive rods, and all utilized formate, glucose and CO. One strain required, and all were stimulated by, rumen fluid. No spores were observed with phase-contast microscopy and two strains were motile. No methane was detected in the headspace of pure cultures grown under either gas phase. The isolation of these bacteria indicates that acetogenic bacteria are inhabitants of the rumen of the bovine fed a typical diet and suggests that they may be participants in the utilization of hydrogen in the rumen ecosystem. Strain 139B (= ATCC 43876) is named Acetitomaculum ruminis gen. nov., sp. nov. and is the type strain of this new species.

Key words

Acetitomaculum ruminis Hydrogen oxidation Acetate production Rumen 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Balch WE, Wolfe RS (1976) A new approach to the cultivation of methanogenic bacteria: 2-mercaptoethanesulfonic acid (HS-CoM)-dependent growth of Methanobacterium ruminantium in a pressurized atmosphere. Appl Environ Microbiol 32:781–791PubMedGoogle Scholar
  2. Balch WE, Wolfe RS (1979) Specificity and biological distribution of coenzyme M (2-mercaptoethanesulfonic acid). J Bacteriol 137:256–263PubMedGoogle Scholar
  3. Balch WE, Schoberth S, Tanner RS, Wolfe RS (1977) Acetobacterium, a new genus of hydrogen-oxidizing, carbon dioxide-reducing, anaerobic bacteria. Int J Syst Bacteriol 27:355–361Google Scholar
  4. Balch WE, Fox GE, Magrum LJ, Woese CR, Wolfe RS (1979) Methanogens: reevaluation of a unique biological group. Microbiol Rev 43:260–296PubMedGoogle Scholar
  5. Bouwer EJ, McCarty PL (1983) Effects of 2-bromoethanesulfonic acid and 2-chloroethanesulfonic acid on acetate utilization in a continuous-flow methanogenic fixed-film column. Appl Environ Microbiol 45:1408–1410PubMedGoogle Scholar
  6. Boyde A, Vesely P (1972) Comparison of fixation and drying procedure for preparation of some cultured cell lines for examination in the SEM. In: Johari O, Corvin I (eds) Proceedings of the Fifth Annual Scanning Electron Microscope Symposium. IIT Research Institute, Chicago, IL, pp 265–272Google Scholar
  7. Braun M, Gottschalk G (1982) Acetobacterium wieringae sp. nov., a new species producing acetic acid from molecular hydrogen and carbon dioxide. Zentralbl Bakteriol Parasitenkd Infektioskr Hyg Abt 1 Orig. Reihe C 3:368–376Google Scholar
  8. Braun M, Mayer F, Gottschalk G (1981) Clostridium aceticum (Wieringa), a microorganism producing acetic acid from molecular hydrogen and carbon dioxide. Arch Microbiol 128:288–293PubMedCrossRefGoogle Scholar
  9. Braun M, Schoberth S, Gottschalk G (1979) Enumeration of bacteria forming acetate from H2 and CO2 in anaerobic habitats. Arch Microbiol 120:201–204PubMedCrossRefGoogle Scholar
  10. Breznak JA, Switzer JM (1986) Acetate synthesis from H2 plus CO2 by termite gut microbes. Appl Environ Microbiol 52:623–630PubMedGoogle Scholar
  11. Bryant MP, Small N, Bouma C, Robinson I (1958) Studies on the composition of the ruminal flora and fauna of young calves. J Dairy Sci 41:1747–1767CrossRefGoogle Scholar
  12. Canby CA, Dogan U, Tomanck RJ (1985) Hexamethyldisilazane (HMDS) for mammalian tissue: an alternative method to critical point drying (CPD). J Electron Microsc Tech 2:653Google Scholar
  13. Cottyn BG, Boucque CV (1968) Rapid method for the gas-chromatographic determination of volatile fatty acids in rumen fluid. J Agric Fd Chem 16:105–107CrossRefGoogle Scholar
  14. Genthner BRS, Davis CL, Bryant MP (1981) Features of rumen and sewage strains of Eubacterium limosum, a methanol- and H2:CO2-utilizing species. Appl Environ Microbiol 42:12–19PubMedGoogle Scholar
  15. Gunsalus RP, Romesser JA, Wolfe RS (1978) Preparation of coenzyme M analogs and their activity in the methyl-coenzyme M reductase in Methanobacterium thermoautotrophicum. Biochemistry 17:2374–2377PubMedCrossRefGoogle Scholar
  16. Heinrikson RL, Meredith SC (1984) Amino acid analysis by reversephase high-performance liquid chromatography: precolumn derivatization with phenylisothiocyanate. Anal Biochem 136:65–74PubMedCrossRefGoogle Scholar
  17. Hermann M, Popoff MR, Sebald M (1987) Sporomusa paucivorans sp. nov., a methylotrophic bacterium that forms acetic acid from hydrogen and carbon dioxide. Int J Syst Bacteriol 37:93–101Google Scholar
  18. Holdeman LV, Cato EP, Moore WEC (1977) Anaerobic laboratory manual, 4th ed. Virginia Polytechnic Institute and State University, BlacksburgGoogle Scholar
  19. Hungate RE (1969) A roll tube method for cultivation of strict anaerobes. In: Norris R, Ribbons DW (eds) Methods in microbiology, vol 3 B. Academic Press, New York, pp 117–132Google Scholar
  20. Hungate RE (1976) The rumen fermentation. In: Schlegel HG, Gottschalk G, Pfennig N (eds) Microbial production and utilization of gases. Goltze, Göttingen, pp 119–124Google Scholar
  21. Huss V, Schleifer KH, Lindal E, Schwan O, Smyth CJ (1982) Peptidoglycan type, base composition of DNA, and DNA-DNA homology of Peptostreptococcus indolicus and Peptostreptococcus asaccharolyticus. FEMS Microbiol Lett 15:285–289CrossRefGoogle Scholar
  22. Johnson JL (1981) Genetic characterization. In: Gerhardt P, Murray RGE, Costilow RN, Nester EW, Wood WA, Krieg NR, Phillips GB (eds) Manual of methods for general bacteriology, 1st ed. American Society for Microbiology, Washington DC, pp 450–472Google Scholar
  23. Johnson JL, Francis BS (1975) Taxonomy of the clostridia: ribosomal ribonucleic acid homologies among the species. J Gen Microbiol 88:229–244PubMedGoogle Scholar
  24. Kandler O, Schoberth S (1979) Murein structure of Acetobacterium woodii. Arch Microbiol 120:181–183CrossRefGoogle Scholar
  25. Leedle JAZ, Butine TJ (1987) Enumeration of cellulolytic anaerobic bacteria from the bovine rumen: comparison of three methods. Curr Microbiol 15:77–79CrossRefGoogle Scholar
  26. Leedle JAZ, Greening RC (1988) Methanogenic and acidogenic bacteria in the bovine rumen: postprandial changes after feeding high- or low-forage diets once daily. Appl Environ Microbiol 54:502–506PubMedGoogle Scholar
  27. Leedle JAZ, Hespell RB (1980) Differential carbohydrate media and anaerobic replica techniques in delineating carbohydrate-utilizing subgroups in rumen bacterial populations. Appl Environ Microbiol 39:709–719PubMedGoogle Scholar
  28. Leigh JA, Mayer F, Wolfe RS (1981) Acetogenium kivui, a new thermophilic hydrogen-oxidizing acetogenic bacterium. Arch Microbiol 129:275–280CrossRefGoogle Scholar
  29. Mayhew JW, Gorbach SL (1977) Internal standards for the gas chromatographic analysis of metabolic end products from anaerobic bacteria. Appl Environ Microbiol 33:1002–1003PubMedGoogle Scholar
  30. Möller B, Oßmer R, Howard BH, Gottschalk G, Hippe H (1984) Sporomusa, a new genus of Gram-negative anaerobic bacteria including Sporomusa sphaeroides spec. nov. and Sporomusa ovata spec. nov. Arch Microbiol 139:388–396CrossRefGoogle Scholar
  31. Moore WEC, Holdeman LV (1986) Genus Eubacterium. In: Sneath PHA, Mair NS, Sharpe ME, Holt JG (eds) Bergey's manual systematic bacteriology, vol 2. Williams and Wilkins, Baltimore, pp 1353–1373Google Scholar
  32. Ohwaki D, Hungate RE (1977) Hydrogen utilization by clostridia in sewage sludge. Appl Environ Microbiol 33:1270–1274PubMedGoogle Scholar
  33. Prins RA, Lankhorst A (1977) Synthesis of acetate from CO2 in the cecum of some rodents. FEMS Microbiol Lett 1:255–258CrossRefGoogle Scholar
  34. Reynolds ES (1963) The use of lead citrate at high pH as an electron-dense stain in electron microscopy. J Cell Biol 17:208–213PubMedCrossRefGoogle Scholar
  35. Robinson JA, Strayer RF, Tiedje JM (1981) Method for measuring dissolved hydrogen in anaerobic ecosystems: application of the rumen. Appl Environ Microbiol 41:545–548PubMedGoogle Scholar
  36. Schleifer KH, Kandler O (1972) Peptidoglycan types of bacterial cell walls and their taxonomic implications. Bacteriol Rev 36:407–477PubMedGoogle Scholar
  37. Schleifer KH, Nimmermann E (1973) Peptidoglycan types of the strains of the genus Peptococcus. Arch Mikrobiol 93:245–258PubMedCrossRefGoogle Scholar
  38. Sleat R, Mah RA, Robinson R (1985) Acetoanaerobium noterae gen. nov., sp. nov.: an anaerobic bacterium that forms acetate from H2 and CO2. Int J Syst Bacteriol 35:10–15CrossRefGoogle Scholar
  39. Smith MR, Mah RA (1981) 2-Bromoethanesulfonate: a selective agent for isolating resistant Methanosarcina mutants. Curr Microbiol 6:321–326CrossRefGoogle Scholar
  40. Supelco, Inc. (1975) Separation of VFA C2−C5, Bulletin 749D. Bellefonte, PAGoogle Scholar
  41. Tanner RS, Stackebrandt E, Fox GE, Woese CR (1981) A phylogenetic analysis of Acetobacterium woodii, Clostridium barkeri, Clostridium butyricum, Clostridium lituseburense, Eubacterium limosum, and Eubacterium tenue. Curr Microbiol 5:35–38CrossRefGoogle Scholar
  42. Teather RM (1982) Maintenance of laboratory strains of obligately anaerobic rumen bacteria. Appl Environ Microbiol 44:499–501PubMedGoogle Scholar
  43. Weiss N, Schleifer KH, Kandler O (1981) The peptidoglycan types of Gram-positive anaerobic bacteria and their taxonomic implications. Rev Inst Pasteur (Lyon) 14:3–12Google Scholar
  44. Zehnder AJB, Huser BA, Brock TD, Wuhrmann K (1980) Characterization of an acetate-decarboxylating non-hydrogen oxidizing methane bacterium. Arch Microbiol 124:1–11PubMedCrossRefGoogle Scholar
  45. Zinder SH, Anguish T, Cardell SC (1984) Selective inhibition by 2-bromoethanesulfonate of methanogenesis from acetate in a thermophilic anaerobic digestor. Appl Environ Microbiol 47:1343–1345PubMedGoogle Scholar

Copyright information

© Springer-Verlag 1989

Authors and Affiliations

  • R. C. Greening
    • 1
  • J. A. Z. Leedle
    • 1
  1. 1.Microbiology and Nutrition ResearchThe Upjohn CompanyKalamazooUSA

Personalised recommendations