Skip to main content
PMC home page
Applied and Environmental Microbiology logoLink to Applied and Environmental Microbiology
. 1996 Feb;62(2):347–352. doi: 10.1128/aem.62.2.347-352.1996

Phylogeny of not-yet-cultured spirochetes from termite guts.

B J Paster 1, F E Dewhirst 1, S M Cooke 1, V Fussing 1, L K Poulsen 1, J A Breznak 1
PMCID: PMC167805  PMID: 8593040

Abstract

Comparisons of 16S rDNA sequences were used to determine the phylogeny of not-yet-cultured spirochetes from hindguts of the African higher termite, Nasutitermes lujae (Wasmann). The 16S rRNA genes were amplified directly from spirochete-rich hindguts by using universal primers, and the amplified products were cloned into Escherichia coli. Clones were screened with a spirochete-specific DNA probe. Analysis of 1,410 base positions of the 16S rDNA insert from one spirochete clone, designated NL1, supported its assignment to the genus Treponema, with average interspecies similarities of ca. 85%. The sequence of NL1 was most closely related (ca. 87 to 88% similarity) to sequences of Spirochaeta stenostrepta and Spirochaeta caldaria and to a previously published sequence (ca. 87% similarity) of spirochetal clone MDS1 from the Australian lower termite, Mastotermes darwiniensis (Froggatt). On the basis of 16S rRNA sequence comparisons and individual base signatures, clones NL1 and MDS1 clearly represent two novel species of Treponema, although specific epithets have not yet been proposed. The gross morphology of NL1 was determined from in situ hybridization experiments with an NL1-specific, fluorescently labeled oligonucleotide probe. Cells were approximately 0.3 to 0.4 by 30 microns in size, with a wavelength and amplitude of about 10 microns and 0.8 to 1.6 micron, respectively. Moreover, electron microscopy of various undulate cells present in gut contents confirmed that they possessed ultrastructural features typical of spirochetes, i.e., a wavy protoplasmic cylinder, periplasmic flagella, and an outer sheath. The sequence data suggest that termite gut spirochetes may represent a separate line of descent from other treponemes and that they constitute a significant reservoir of previously unrecognized spirochetal biodiversity.

Full Text

The Full Text of this article is available as a PDF (936.6 KB).

Selected References

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

  1. Amann R. I., Krumholz L., Stahl D. A. Fluorescent-oligonucleotide probing of whole cells for determinative, phylogenetic, and environmental studies in microbiology. J Bacteriol. 1990 Feb;172(2):762–770. doi: 10.1128/jb.172.2.762-770.1990. [DOI] [PMC free article] [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. Berchtold M., Ludwig W., König H. 16S rDNA sequence and phylogenetic position of an uncultivated spirochete from the hindgut of the termite Mastotermes darwiniensis Froggatt. FEMS Microbiol Lett. 1994 Nov 1;123(3):269–273. doi: 10.1111/j.1574-6968.1994.tb07235.x. [DOI] [PubMed] [Google Scholar]
  4. Bermudes D., Chase D., Margulis L. Morphology as a basis for taxonomy of large spirochetes symbiotic in wood-eating cockroaches and termites: Pillotina gen. nov., nom. rev.; Pillotina calotermitidis sp. nov., nom. rev.; Diplocalyx gen. nov., nom. rev.; Diplocalyx calotermitidis sp. nov., nom. rev.; Hollandina gen. nov., nom.[TRUNCATED]. Int J Syst Bacteriol. 1988 Jul;38(3):291–302. doi: 10.1099/00207713-38-3-291. [DOI] [PubMed] [Google Scholar]
  5. Breznak J. A., Pankratz H. S. In situ morphology of the gut microbiota of wood-eating termites [Reticulitermes flavipes (Kollar) and Coptotermes formosanus Shiraki]. Appl Environ Microbiol. 1977 Feb;33(2):406–426. doi: 10.1128/aem.33.2.406-426.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Brune A., Emerson D., Breznak J. A. The Termite Gut Microflora as an Oxygen Sink: Microelectrode Determination of Oxygen and pH Gradients in Guts of Lower and Higher Termites. Appl Environ Microbiol. 1995 Jul;61(7):2681–2687. doi: 10.1128/aem.61.7.2681-2687.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Choi B. K., Paster B. J., Dewhirst F. E., Göbel U. B. Diversity of cultivable and uncultivable oral spirochetes from a patient with severe destructive periodontitis. Infect Immun. 1994 May;62(5):1889–1895. doi: 10.1128/iai.62.5.1889-1895.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Defosse D. L., Johnson R. C., Paster B. J., Dewhirst F. E., Fraser G. J. Brevinema andersonii gen. nov., sp. nov., an infectious spirochete isolated from the short-tailed shrew (Blarina brevicauda) and the white-footed mouse (Peromyscus leucopus). Int J Syst Bacteriol. 1995 Jan;45(1):78–84. doi: 10.1099/00207713-45-1-78. [DOI] [PubMed] [Google Scholar]
  9. Fox J. G., Yan L. L., Dewhirst F. E., Paster B. J., Shames B., Murphy J. C., Hayward A., Belcher J. C., Mendes E. N. Helicobacter bilis sp. nov., a novel Helicobacter species isolated from bile, livers, and intestines of aged, inbred mice. J Clin Microbiol. 1995 Feb;33(2):445–454. doi: 10.1128/jcm.33.2.445-454.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Krogfelt K. A., Poulsen L. K., Molin S. Identification of coccoid Escherichia coli BJ4 cells in the large intestine of streptomycin-treated mice. Infect Immun. 1993 Dec;61(12):5029–5034. doi: 10.1128/iai.61.12.5029-5034.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. 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]
  12. Paster B. J., Dewhirst F. E., Weisburg W. G., Tordoff L. A., Fraser G. J., Hespell R. B., Stanton T. B., Zablen L., Mandelco L., Woese C. R. Phylogenetic analysis of the spirochetes. J Bacteriol. 1991 Oct;173(19):6101–6109. doi: 10.1128/jb.173.19.6101-6109.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Poulsen L. K., Lan F., Kristensen C. S., Hobolth P., Molin S., Krogfelt K. A. Spatial distribution of Escherichia coli in the mouse large intestine inferred from rRNA in situ hybridization. Infect Immun. 1994 Nov;62(11):5191–5194. doi: 10.1128/iai.62.11.5191-5194.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Saitou N., Nei M. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol. 1987 Jul;4(4):406–425. doi: 10.1093/oxfordjournals.molbev.a040454. [DOI] [PubMed] [Google Scholar]
  15. Stackebrandt E., Liesack W., Goebel B. M. Bacterial diversity in a soil sample from a subtropical Australian environment as determined by 16S rDNA analysis. FASEB J. 1993 Jan;7(1):232–236. doi: 10.1096/fasebj.7.1.8422969. [DOI] [PubMed] [Google Scholar]
  16. 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]
  17. Weisburg W. G., Barns S. M., Pelletier D. A., Lane D. J. 16S ribosomal DNA amplification for phylogenetic study. J Bacteriol. 1991 Jan;173(2):697–703. doi: 10.1128/jb.173.2.697-703.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Woese C. R., Kandler O., Wheelis M. L. Towards a natural system of organisms: proposal for the domains Archaea, Bacteria, and Eucarya. Proc Natl Acad Sci U S A. 1990 Jun;87(12):4576–4579. doi: 10.1073/pnas.87.12.4576. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Zhou C., Yang Y., Jong A. Y. Mini-prep in ten minutes. Biotechniques. 1990 Feb;8(2):172–173. [PubMed] [Google Scholar]

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

RESOURCES