Phylogeny of not-yet-cultured spirochetes from termite guts

• Published in: Applied and Environmental Microbiology 1996-02, Vol.62 (2), p.347-352
• Main Authors: Pastier, B.J. (Forsyth Dental Center, Boston, MA.), Dewhirst, F.E, Cooke, S.M, Fussing, V, Poulsen, L.K, Breznak, J.A
• Format: Article
• Language: English
• Subjects: ADN; Animals; APPAREIL DIGESTIF; ARN RIBOSOMAL; ARN RIBOSOMIAL; Base Sequence; Biological and medical sciences; Biological evolution; Cellular biology; CHIMIOTAXONOMIE; Deoxyribonucleic acid; DNA; DNA Probes - genetics; DNA, Bacterial - genetics; DNA, Ribosomal - genetics; FILOGENIA; FLORA MICROBIANA; FLORE MICROBIENNE; Fundamental and applied biological sciences. Psychology; GENE; GENES; Genetics of eukaryotes. Biological and molecular evolution; HIBRIDACION DE ADN; HYBRIDATION D'ADN; Insecta - microbiology; Insects; INTESTIN; INTESTINOS; Mastotermes darwiniensis; Microscopy, Electron; Molecular Sequence Data; NASUTITERMES; PHYLOGENIE; Phylogeny; QUIMIOTAXONOMIA; RIBOSOMAS; RIBOSOME; RNA, Bacterial - genetics; RNA, Ribosomal, 16S - genetics; SECUENCIA NUCLEICA; SEQUENCE NUCLEIQUE; SISTEMA DIGESTIVO; Spirochaeta - classification; Spirochaeta - genetics; Spirochaeta - isolation & purification; Spirochaeta caldaria; Spirochaeta stenostrepta; Spirochaetaceae - classification; Spirochaetaceae - genetics; Spirochaetaceae - isolation & purification; SPIROCHAETALES; Treponema - classification; Treponema - genetics; Treponema - isolation & purification; ULTRAESTRUCTURA; ULTRASTRUCTURE
• ISSN: 0099-2240; 1098-5336
• DOI: 10.1128/aem.62.2.347-352.1996

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 micrometers in size, with a wavelength and amplitude of about 10 micrometers and 0.8 to 1.6 micrometers, 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