Summary auto-generated
Researchers sequenced 16S rRNA genes from 47 oral Treponema strains collected by Dr. Robert Smibert over 20 years to determine their phylogenetic relationships. Five distinct species were identified based on 16S rRNA sequence analysis, with a species designation cutoff at approximately 98% sequence similarity. Two species were newly named: Treponema maltophilum (strain D4B-1) and Treponema amylovorum (strain D62CR-I2). Three additional species—Treponema Smibert-2, Smibert-3, and Smibert-5—have not yet been formally characterized. Smibert-2 contained nine isolates with identical sequences but differing cellular fatty acid profiles, indicating poor correlation between phylogenetic and biochemical classification methods. The study also characterized previously unanalyzed species including T. vincentii and three T. socranskii subspecies (socranskii, buccale, and paredis), which formed separate phylogenetic branches at 98% sequence similarity. Several reference strains were identified as T. denticola, and some cultures were contaminated with mycoplasmas, highlighting the need for mycoplasma screening in spirochaete cultures.
Key findings
- Five distinct cultivable oral Treponema species were identified from the Smibert collection, including two newly named species (T. maltophilum and T. amylovorum) and three uncharacterized species
- Cellular fatty acid analysis showed poor correlation with phylogenetic classification, as nine Treponema Smibert-2 strains with identical 16S rRNA sequences belonged to six different fatty acid groups
- T. vincentii's closest relative was T. medium at 98.7% sequence similarity, and T. socranskii subspecies formed three separate phylogenetic branches with approximately 98% intragenic sequence similarity
- Multiple historical reference strains previously thought to be different species were identified as T. denticola through 16S rRNA sequencing
- Some spirochaete cultures were contaminated with mycoplasmas, indicating the need for screening protocols
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Abstract
Dr Robert Smibert from the Virginia Polytechnic Institute, USA, isolated and collected over 200 strains of oral treponemes over a 20-year period. Dr Smibert, Dr W. E. C. Moore and Dr L. V. Moore separated these isolates and reference strains into different groups on the basis of cellular fatty acid analysis. In this study, the 16S rRNA genes were sequenced for 47 strains that were representative of these groups. Five distinct species were identified on the basis of 16S rRNA sequence comparisons; two of these species are newly named and three have not yet been characterized. The first species, designated Treponema Smibert-1, was represented by the single strain D4B-1 and was later identified as the newly described Treponema maltophilum. However, strain D4B-1 possessed a different flagellar arrangement to that of T. maltophilum. The second species, Treponema Smibert-2, was represented by nine isolates that possessed identical 16S rRNA gene sequences. The closest relatives of this species were Treponema Smibert-3 and Treponema Smibert-4 at approximately 90% sequence similarity. Within Treponema Smibert-2, there was no correlation between phylogenetic analysis and cellular fatty acid analysis since six different cellular fatty acid groups represented the nine strains. Treponema Smibert-3 (strain D36ER-1) and Treponema Smibert-4 (D62CR-12) were each represented by only a single strain and were closely related to each other at 98% sequence similarity. Strain D36ER-1 of Treponema Smibert-3 was identified as belonging to the not-yet-cultivated phylotype 20 [Choi, B. K., Paster, B. J., Dewhirst, F. E. & Göbel, U. B. (1994). Infect Immun 62,1889–1895]. Strain D62CR-12 of Treponema Smibert-4 was nearly identical in sequence to the newly described Treponema amylovorum. The fifth species, Treponema Smibert-5, was represented by a single strain, D120CR-1, and was closely related at about 98% sequence similarity to the three subspecies of Treponema socranskii. The phylogenetic analyses of strains of Treponema vincentii and of subspecies of T. socranskii are also reported. The closest oral relatives of T. vincentii were Treponema medium at 98·7% sequence similarity and Treponema denticola at 91·5% sequence similarity. T. socranskii subspp. socranskii, buccale and paredis formed three separate phylogenetic branches with sequence similarities of about 98% to each other. The closest relative of the subspecies of T. socranskii and of Smibert-5 was Smibert-2 at about 86% sequence similarity. Historic reference strains Fuji, Treponema ambigua, Fm, lchelson-2, N-39, TD2, TRRD, MRB, IPP, Jethro and T32A, as well as an unknown strain designated only as Treponema oralis, were identified as strains of T. denticola. Reference strains Fuji, Jethro, T32A and IPP plus three isolates of the Smibert collection were also contaminated with a mycoplasma as determined by 16S rRNA comparative analysis. Consequently, spirochaetal cultures should be screened for mycoplasmas. There are presently at least ten species of cultivable oral species of Treponema with the cut-off for separate species