Skip to main content
PMC home page
Applied and Environmental Microbiology logoLink to Applied and Environmental Microbiology
. 1996 Dec;62(12):4433–4440. doi: 10.1128/aem.62.12.4433-4440.1996

Identifying numerically abundant culturable bacteria from complex communities: an example from a lignin enrichment culture.

J M González 1, W B Whitman 1, R E Hodson 1, M A Moran 1
PMCID: PMC168269  PMID: 8953714

Abstract

Culturable bacteria that were numerically important members of a marine enrichment community were identified and characterized phylogenetically. Selective and nonselective isolation methods were used to obtain 133 culturable bacterial isolates from model marine communities enriched with the high-molecular-weight (lignin-rich) fraction of pulp mill effluent. The culture collection was screened against community DNA from the lignin enrichments by whole-genome hybridization methods, and three marine bacterial isolates were identified as being numerically important in the communities. One isolate was in the alpha-subclass of Proteobacteria, and the other two were in the gamma-subclass of Proteobacteria. Isolate-specific 16S rRNA oligonucleotide probes designed to precisely quantify the isolates in the lignin enrichment communities indicated contributions ranging from 2 to 32% of enrichment DNA, values nearly identical to those originally obtained by the simpler whole-genome hybridization method. Two 16S rRNA sequences closely related to that of one of the isolates, although not identical, were amplified via PCR from the seawater sample originally used to inoculate the enrichment medium. Partial sequences of 14 other isolates revealed significant phylogenetic diversity and unusual sequences among the culturable lignin enrichment bacteria, with the Proteobacteria, Cytophaga-Flavobacterium, and gram-positive groups represented.

Full Text

The Full Text of this article is available as a PDF (275.9 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  2. Amann R. I., Ludwig W., Schleifer K. H. Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol Rev. 1995 Mar;59(1):143–169. doi: 10.1128/mr.59.1.143-169.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Barns S. M., Fundyga R. E., Jeffries M. W., Pace N. R. Remarkable archaeal diversity detected in a Yellowstone National Park hot spring environment. Proc Natl Acad Sci U S A. 1994 Mar 1;91(5):1609–1613. doi: 10.1073/pnas.91.5.1609. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Braun-Howland E. B., Vescio P. A., Nierzwicki-Bauer S. A. Use of a simplified cell blot technique and 16S rRNA-directed probes for identification of common environmental isolates. Appl Environ Microbiol. 1993 Oct;59(10):3219–3224. doi: 10.1128/aem.59.10.3219-3224.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Britschgi T. B., Giovannoni S. J. Phylogenetic analysis of a natural marine bacterioplankton population by rRNA gene cloning and sequencing. Appl Environ Microbiol. 1991 Jun;57(6):1707–1713. doi: 10.1128/aem.57.6.1707-1713.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Button D. K., Schut F., Quang P., Martin R., Robertson B. R. Viability and isolation of marine bacteria by dilution culture: theory, procedures, and initial results. Appl Environ Microbiol. 1993 Mar;59(3):881–891. doi: 10.1128/aem.59.3.881-891.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. DeLong E. F. Archaea in coastal marine environments. Proc Natl Acad Sci U S A. 1992 Jun 15;89(12):5685–5689. doi: 10.1073/pnas.89.12.5685. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. DeLong E. F., Wu K. Y., Prézelin B. B., Jovine R. V. High abundance of Archaea in Antarctic marine picoplankton. Nature. 1994 Oct 20;371(6499):695–697. doi: 10.1038/371695a0. [DOI] [PubMed] [Google Scholar]
  9. Del Sal G., Manfioletti G., Schneider C. The CTAB-DNA precipitation method: a common mini-scale preparation of template DNA from phagemids, phages or plasmids suitable for sequencing. Biotechniques. 1989 May;7(5):514–520. [PubMed] [Google Scholar]
  10. Fuhrman J. A., Comeau D. E., Hagström A., Chan A. M. Extraction from natural planktonic microorganisms of DNA suitable for molecular biological studies. Appl Environ Microbiol. 1988 Jun;54(6):1426–1429. doi: 10.1128/aem.54.6.1426-1429.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Fuhrman J. A., McCallum K., Davis A. A. Novel major archaebacterial group from marine plankton. Nature. 1992 Mar 12;356(6365):148–149. doi: 10.1038/356148a0. [DOI] [PubMed] [Google Scholar]
  12. Fuhrman J. A., McCallum K., Davis A. A. Phylogenetic diversity of subsurface marine microbial communities from the Atlantic and Pacific Oceans. Appl Environ Microbiol. 1993 May;59(5):1294–1302. doi: 10.1128/aem.59.5.1294-1302.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Giovannoni S. J., Britschgi T. B., Moyer C. L., Field K. G. Genetic diversity in Sargasso Sea bacterioplankton. Nature. 1990 May 3;345(6270):60–63. doi: 10.1038/345060a0. [DOI] [PubMed] [Google Scholar]
  14. Grimont P. A., Grimont F., Desplaces N., Tchen P. DNA probe specific for Legionella pneumophila. J Clin Microbiol. 1985 Mar;21(3):431–437. doi: 10.1128/jcm.21.3.431-437.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hobbie J. E., Daley R. J., Jasper S. Use of nuclepore filters for counting bacteria by fluorescence microscopy. Appl Environ Microbiol. 1977 May;33(5):1225–1228. doi: 10.1128/aem.33.5.1225-1228.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Huber R., Burggraf S., Mayer T., Barns S. M., Rossnagel P., Stetter K. O. Isolation of a hyperthermophilic archaeum predicted by in situ RNA analysis. Nature. 1995 Jul 6;376(6535):57–58. doi: 10.1038/376057a0. [DOI] [PubMed] [Google Scholar]
  17. Hyypiä T., Jalava A., Larsen S. H., Terho P., Hukkanen V. Detection of Chlamydia trachomatis in clinical specimens by nucleic acid spot hybridization. J Gen Microbiol. 1985 Apr;131(4):975–978. doi: 10.1099/00221287-131-4-975. [DOI] [PubMed] [Google Scholar]
  18. Kafatos F. C., Jones C. W., Efstratiadis A. Determination of nucleic acid sequence homologies and relative concentrations by a dot hybridization procedure. Nucleic Acids Res. 1979 Nov 24;7(6):1541–1552. doi: 10.1093/nar/7.6.1541. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Lane D. J., Pace B., Olsen G. J., Stahl D. A., Sogin M. L., Pace N. R. Rapid determination of 16S ribosomal RNA sequences for phylogenetic analyses. Proc Natl Acad Sci U S A. 1985 Oct;82(20):6955–6959. doi: 10.1073/pnas.82.20.6955. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Maidak B. L., Larsen N., McCaughey M. J., Overbeek R., Olsen G. J., Fogel K., Blandy J., Woese C. R. The Ribosomal Database Project. Nucleic Acids Res. 1994 Sep;22(17):3485–3487. doi: 10.1093/nar/22.17.3485. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Schmidt T. M., DeLong E. F., Pace N. R. Analysis of a marine picoplankton community by 16S rRNA gene cloning and sequencing. J Bacteriol. 1991 Jul;173(14):4371–4378. doi: 10.1128/jb.173.14.4371-4378.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Schut F., de Vries E. J., Gottschal J. C., Robertson B. R., Harder W., Prins R. A., Button D. K. Isolation of Typical Marine Bacteria by Dilution Culture: Growth, Maintenance, and Characteristics of Isolates under Laboratory Conditions. Appl Environ Microbiol. 1993 Jul;59(7):2150–2160. doi: 10.1128/aem.59.7.2150-2160.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Stahl D. A., Lane D. J., Olsen G. J., Pace N. R. Characterization of a Yellowstone hot spring microbial community by 5S rRNA sequences. Appl Environ Microbiol. 1985 Jun;49(6):1379–1384. doi: 10.1128/aem.49.6.1379-1384.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Tsai Y. L., Olson B. H. Rapid method for direct extraction of DNA from soil and sediments. Appl Environ Microbiol. 1991 Apr;57(4):1070–1074. doi: 10.1128/aem.57.4.1070-1074.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Voordouw G., Shen Y., Harrington C. S., Telang A. J., Jack T. R., Westlake D. W. Quantitative reverse sample genome probing of microbial communities and its application to oil field production waters. Appl Environ Microbiol. 1993 Dec;59(12):4101–4114. doi: 10.1128/aem.59.12.4101-4114.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Voordouw G., Strang J. D., Wilson F. R. Organization of the genes encoding [Fe] hydrogenase in Desulfovibrio vulgaris subsp. oxamicus Monticello. J Bacteriol. 1989 Jul;171(7):3881–3889. doi: 10.1128/jb.171.7.3881-3889.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Voordouw G., Voordouw J. K., Jack T. R., Foght J., Fedorak P. M., Westlake D. W. Identification of distinct communities of sulfate-reducing bacteria in oil fields by reverse sample genome probing. Appl Environ Microbiol. 1992 Nov;58(11):3542–3552. doi: 10.1128/aem.58.11.3542-3552.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Voordouw G., Voordouw J. K., Karkhoff-Schweizer R. R., Fedorak P. M., Westlake D. W. Reverse sample genome probing, a new technique for identification of bacteria in environmental samples by DNA hybridization, and its application to the identification of sulfate-reducing bacteria in oil field samples. Appl Environ Microbiol. 1991 Nov;57(11):3070–3078. doi: 10.1128/aem.57.11.3070-3078.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Ward D. M., Weller R., Bateson M. M. 16S rRNA sequences reveal numerous uncultured microorganisms in a natural community. Nature. 1990 May 3;345(6270):63–65. doi: 10.1038/345063a0. [DOI] [PubMed] [Google Scholar]
  30. Waterhouse R. N., Glover L. A. Differences in the hybridization pattern of Bacillus subtilis genes coding for rDNA depend on the method of DNA preparation. Appl Environ Microbiol. 1993 Mar;59(3):919–921. doi: 10.1128/aem.59.3.919-921.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Applied and Environmental Microbiology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES