1887

Abstract

The polymerase chain reaction (PCR) was used to amplify small-subunit ribosomal DNA from the anaerobic ciliated protozoon , and from its uncultured endosymbiotic bacteria. This was accomplished directly from total DNA extracted from protozoa without prior isolation or enrichment for symbiont cells. The double-stranded amplification products were precipitated and directly sequenced using the linear PCR reaction. Fluorescent oligonucleotide probes were designed and used in whole-cell hybridizations to provide direct visual evidence that the sequences originated from the host ciliate and from the endosymbiont. Phylogenetic analysis of the sequence consistently placed it as a deep-branching lineage near the root of the ciliate tree. However, the present data were insufficient to resolve the detailed relationship between and and thus to determine if the heterotrichs are mono- or paraphyletic. Phylogenetic analysis of the symbiont partial sequence clearly demonstrated that it is an archaeobacterium and that it is closely related to, but distinct from, .

Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-138-7-1479
1992-07-01
2024-10-11
Loading full text...

Full text loading...

/deliver/fulltext/micro/138/7/mic-138-7-1479.html?itemId=/content/journal/micro/10.1099/00221287-138-7-1479&mimeType=html&fmt=ahah

References

  1. Amman R. I., Krumholz L., Stahl D. A. 1990; Fluorescent oligonucleotide probing of whole cells for determinative, phylogenetic, and environmental studies in microbiology. Journal of Bacteriology 172 762 770
    [Google Scholar]
  2. Amman R. I., Springer N., Ludwig W., Gortz H. D., Schleifer K. H. 1991; Identification and phylogeny of uncultured bacterial symbionts. Nature, London 351 161 164
    [Google Scholar]
  3. Boone D. R., Mah R. A. 1989; Methanobacteriaceae. In Bergey’s Manual of Systematic Bacteriology vol. 3 pp. 2175 2178 Edited by Staley J. T., Bryant M. P., Pfennig N., Holt J. G. Baltimore: Williams & Wilkins;
    [Google Scholar]
  4. Corliss J. O. 1974; Remarks on the composition of the large ciliate class Kinetofragminophora de Puytorac et al., 1974, and recognition of several new taxa therein, with emphasis on the primitive order Protociliatida n. ord. Journal of Protozoology 21 207 220
    [Google Scholar]
  5. Delong E. F., Wickham G. S., Pace N. R. 1989; Phylogenetic stains: ribosomal RNA based probes for the identification of single cells. Science 243 1360 1363
    [Google Scholar]
  6. Edman J. C., Kovacs J. A., Masur H., Santi D. V., Elwood H. J., Sogin M. L. 1988; Ribosomal RNA sequences shows Pneumocystis carinii to be a member of the fungi. Nature, London 334 519 522
    [Google Scholar]
  7. Elwood H. J., Olsen G. J., Sogin M. L. 1985; The small subunit ribosomal RNA gene sequences from the hypotrichous ciliates Oxytricha nova and Stylonichia pustulata. Molecular Biology and Evolution 2 399 410
    [Google Scholar]
  8. Embley T. M. 1991; The linear PCR reaction: a simple and robust method for sequencing amplified rRNA genes. Letters in Applied Microbiology 13 171 174
    [Google Scholar]
  9. Fenchel T., Finlay B. J. 1990; Anaerobic free-living protozoa: growth efficiencies and the structure of anaerobic communities. FEMS Microbiology Ecology 74 269 276
    [Google Scholar]
  10. Fenchel T., Finlay B. J. 1991a; The biology of free living anaerobic ciliates. European Journal of Protistology 26 201 215
    [Google Scholar]
  11. Fenchel T., Finlay B. J. 1991b; Endosymbiotic methanogenic bacteria in anaerobic ciliates : significance for the growth efficiency of the host. Journal of Protozoology 38 18 22
    [Google Scholar]
  12. Fenchel T., Perry T., Thane A. 1977; Anaerobiosis and symbiosis with bacteria in free-living ciliates. Journal of Protozoology 24 154 163
    [Google Scholar]
  13. Finlay B. J. 1990; Physiological ecology of free living protozoa. Advances in Microbial Ecology 11 1 35
    [Google Scholar]
  14. Finlay B. J., Fenchel T. 1989; Hydrogenosomes in some anaerobic bacteria resemble mitochondria. FEMS Microbiology Letters 65 311 314
    [Google Scholar]
  15. Finlay B. J., Fenchel T. 1991; An anaerobic protozoon, with symbiotic methanogens, living in municipal landfill material. FEMS Microbiology Ecology 85 169 180
    [Google Scholar]
  16. Fitch W. M., Margoliash E. 1967; Construction of phylogenetic trees. A method based on mutation distances as estimated from cytochrome c sequences is of general applicability. Science 155 279 284
    [Google Scholar]
  17. Fry N. K., Rowthbottom T. J., Saunders N. A., Embley T. M. 1991; Direct amplification and sequencing of the 16S ribosomal DNA of an intracellular Legionella species recovered by amoebal enrichment from the sputum of a patient with pneumonia. FEMS Microbiology Letters 83 165 168
    [Google Scholar]
  18. Gillespie J. H. 1986; Rates of molecular evolution. Annual Review of Ecology and Systematics 17 637 665
    [Google Scholar]
  19. Giovannoni S. J., DeLong E. F., Olsen G. J., Pace N. R. 1988; Phylogenetic group-specific oligonucleotide probes for the identification of single microbial cells. Journal of Bacteriology 170 720 726
    [Google Scholar]
  20. Giovannoni S. J., Britschgi T. B., Moyer C. L., , & Field K. G. 1990; Genetic diversity in the Sargasso Sea bacterioplankton. Nature, London 345 60 63
    [Google Scholar]
  21. Greenwood S. J., Schlegel M., Sogin M. L., Lynn D. H. 1991a; Phylogenetic relationships of Blepharisma americanum and Colpoda inflata within the phylum Ciliophora inferred from the complete small subunit rRNA gene sequences. Journal of Protozoology 38 1 6
    [Google Scholar]
  22. Greenwood S. J., Sogin M. L., Lynn D. H. 1991b; Phylogenetic relationships within the class Oligohymenophora, phylum Ciliophora, inferred from the complete small subunit rRNA gene sequences of Colpidium campylum, Glaucoma chattoni and Opisthon-ecta henneguyi. Journal of Molecular Evolution 33 163 174
    [Google Scholar]
  23. Jin L., Nei M. 1991; Limitations of the evolutionary parsimony method of phylogenetic analysis. Molecular Biology and Evolution 7 82 102
    [Google Scholar]
  24. Jones W. J., Nagle D. P., Whitman W. B. 1987; Methanogens and the diversity of archaebacteria. Microbiological Reviews 51 135 177
    [Google Scholar]
  25. Jukes T. H., Cantor C. R. 1969; Evolution of protein molecules. In Mammalian Protein Metabolism pp. 21 132 Edited by Munro H. N. New York: Academic Press;
    [Google Scholar]
  26. Lake J. 1987; A rate-independent technique for analysis of nucleic acid sequences: evolutionary parsimony. Molecular Biology and Evolution 4 167 191
    [Google Scholar]
  27. Lechner K., Wich G., Bock A. 1985; The nucleotide sequence of the 16S rRNA gene and flanking regions from Methanobacterium formicium: on the phylogenetic relationship between methanogenic and halophilic archaebacteria. Systematic and Applied Microbiology 6 157 163
    [Google Scholar]
  28. Lynn D., Corliss J. 1991; Ciliophora. In Microscopic Anatomy of Invertebrates, vol. 1, Protozoa pp. 333 467 Edited by Harrison F., Corliss J. O. New York: Wiley-Liss;
    [Google Scholar]
  29. Margulis L., Corliss J. O., Melkonian M., Chapman D. J. 1989 Handbook of Protoctista Boston: Jones & Bartlett;
    [Google Scholar]
  30. Medlin L., Elwood H. J., Stickel S., Sogin M. L. 1988; The characterisation of enzymatically amplified eukaryotic 16S-like rRNA coding regions. Gene 71 491 499
    [Google Scholar]
  31. Muller M. 1988; Energy metabolism of protozoa without mitochondria. Annual Review of Microbiology 42 465 488
    [Google Scholar]
  32. Neefs J.-M., De Wachter R. 1991; A proposal for the secondary structure of a variable area of eukaryotic small ribosomal subunit RNA involving the existence of a pseudoknot. Nucleic Acids Research 18 5695 5704
    [Google Scholar]
  33. Neefs J.-M., Van de Peer Y., De Rijk P., Goris A., De Wachter R. 1991; Compilation of small ribosomal subunit RNA sequences. Nucleic Acids Research 19 Supplement 1987 2015
    [Google Scholar]
  34. Nickrent D. L., Sargent M. L. 1991; An overview of the secondary structure of the V4 region of eukaryotic small-subunit ribosomal RNA. Nucleic Acids Research 19 227 235
    [Google Scholar]
  35. Olsen G. J., Lane D. J., Giovannoni S. J., Pace N. R., Stahl A. 1986; Microbial ecology and evolution: a ribosomal RNA approach. Annual Review of Microbiology 40 337 365
    [Google Scholar]
  36. Pitcher D., Saunders N. A., Owen R. J. 1989; Rapid extraction of bacterial genomic DNA with guanidium thiocyanate. Letters in Applied Microbiology 8 151 156
    [Google Scholar]
  37. Saiki R. K., Gelfand D. H., Stoffel S., Scharf S. J., Higuchi R., Horn G. T., Mullis K. B., Erlich H. A. 1988; Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 239 487 491
    [Google Scholar]
  38. Saitou N., Nei M. 1987; The neighbour joining method: a new method for constructing phylogenetic trees. Molecular Biology and Evolution 4 406 425
    [Google Scholar]
  39. Schlegel M. 1991; Protist evolution and phylogeny as discerned from small subunit ribosomal RNA sequence comparisons. European Journal of Protistology 27 207 219
    [Google Scholar]
  40. Schlegel M., Elwood H. J., Sogin M. L. 1991; Molecular evolution in hypotrichous ciliates: sequence of the small subunit ribosomal RNA genes from Onychodromus quadricornutus and Oxytricha granulifera (Oxytrichidae, Hypotrichida, Ciliophora). Journal of Molecular Evolution 32 64 69
    [Google Scholar]
  41. Sidow A., Wilson A. C. 1990; Compositional statistics: an improvement of evolutionary parsimony and its application to deep branches in the tree of life. Journal of Molecular Evolution 31 51 68
    [Google Scholar]
  42. Sogin M. L., Swanton M. T., Gunderson J. H., Elwood H. J. 1986; Sequence of the small subunit ribosomal RNA gene from the hypotrichous ciliate Euplotes aediculatus. Journal of Protozoology 33 26 29
    [Google Scholar]
  43. Stahl D. A., Amman R. I. 1991; Development and application of nucleic acid probes in bacterial systematies. In Nucleic Acid Techniques in Bacterial Systematies pp. 205 248 Edited by Stackebrandt E., Goodfellow M. Chichester: John Wiley;
    [Google Scholar]
  44. Stahl D. A., Flesher B., Mansfield H. R., Montgomery L. 1988; Use of phylogenetically based hybridization probes for studies of ruminal microbial ecology. Applied and Environmental Microbiology 54 1079 1084
    [Google Scholar]
  45. Swoffford D. 1991; paup: phylogenetic analysis using parsimony, version 3.0. Computer program distributed by the Illinois Natural History Survey. Champaign, Illinois, USA:
    [Google Scholar]
  46. Van Bruggen J. J. A., Zwart K. B., Van Assema R. M., Stumm C. K., Vogels G. D. 1984; Methanobacterium formicicum, an endosymbiont of the anaerobic ciliate Metopus striatus McMurrich. Archives of Microbiology 139 1 7
    [Google Scholar]
  47. Van Bruggen J. J. A., Zwart K. B., Hermans J. G. F., Van Hove E. M., Stumm C. K., Vogels G. D. 1986; Isolation and characterisation of Methanoplanus endosymbiosus sp. nov., an endosymbiont of the marine sapropelic ciliate Metopus contortus Quennerstedt. Archives of Microbiology 144 367 374
    [Google Scholar]
  48. Wagener S., Pfennig N. 1987; Monoxenic culture of the anaerobic ciliate Trimyema compressum Lackey. Archives of Microbiology 149 4 11
    [Google Scholar]
  49. Wagener S., Stumm C. K., Vogels G. D. 1986; Electromigration, a tool for studies on anaerobic ciliates. FEMS Microbiology Ecology 38 197 203
    [Google Scholar]
  50. Woese C. R. 1987; Bacterial classification. Microbiological Reviews 51 221 271
    [Google Scholar]
  51. Woese C. R., Kandler O., Wheelis M. L. 1990; Towards a natural system for organisms: proposal for the Domains Archaea, Bacteria and Eucarya. Proceedings of the National Academy of Sciences of the United States of America 87 4576 4579
    [Google Scholar]
/content/journal/micro/10.1099/00221287-138-7-1479
Loading
/content/journal/micro/10.1099/00221287-138-7-1479
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error