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Journal of Clinical Microbiology, July 2001, p. 2425-2430, Vol. 39, No. 7
Unité des Rickettsies, CNRS UPRES-A
6020, Faculté de Médecine, Université de la
Méditerranée, Marseille, France
Received 8 January 2001/Returned for modification 11 March
2001/Accepted 8 April 2001
Until recently no isolate of Tropheryma whippelii
was available, and therefore genetic studies were limited to those
based on PCR amplification of conserved genes. In this study we
determined the nucleotide sequence of rpoB (encoding the
0095-1137/01/$04.00+0 DOI: 10.1128/JCM.39.7.2425-2430.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
rpoB Sequence Analysis of Cultured
Tropheryma whippelii
-subunit of RNA polymerase) from a cultured strain of T.
whippelii using degenerate consensus PCR and genome walking.
The T. whippelii rpoB consists of 3,657 bp with a 50.4%
GC content and encodes 1,218 amino acids with a calculated molecular
mass of 138 kDa. Comparison of T. whippelii RpoB with
other eubacterial RpoB proteins indicated sequence similarity
ranging from 57.19 (Mycoplasma pneumoniae) to 74.63%
(Mycobacterium tuberculosis). Phylogenetic analysis of
T. whippelii based on comparison of its RpoB sequence
with sequences available for other bacteria was consistent with that previously derived from the 16S ribosomal DNA (rDNA) sequence, indicating that it belongs to the actinomyces clade. The sequence comparison allowed the design of a primer pair, TwrpoB.F and TwrpoB.R, specific for T. whippelii rpoB. When incorporated into a
PCR, this primer pair allowed the detection of T. whippelii
rpoB in three of three 16S rDNA PCR-positive biopsy specimens
and zero of seven negative controls. rpoB could
therefore be targeted in PCR-mediated detection and identification of
this emerging bacterial species. This approach has previously been
shown useful for the identification of related mycobacteria. This study
underscores that a method involving isolation and then propagation of
emerging bacteria is a useful way to quickly achieve extensive
molecular knowledge of these pathogens.
*
Corresponding author. Mailing address: Unité des
Rickettsies, Faculté de Médecine, 27 Boulevard Jean Moulin,
13385 Marseille cedex 5, France. Phone: 33 (0)4 91 32 43 75. Fax: 33 (0)4 91 38 77 72. E-mail:
Didier.Raoult{at}medecine.univ-mrs.fr.
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