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Journal of Clinical Microbiology, May 2009, p. 1393-1401, Vol. 47, No. 5
0095-1137/09/$08.00+0     doi:10.1128/JCM.02469-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Rapid and Accurate Diagnosis of Human Intestinal Spirochetosis by Fluorescence In Situ Hybridization{triangledown}

Dinah Schmiedel,1 Hans-Jörg Epple,2 Christoph Loddenkemper,3 Ralf Ignatius,1 Jutta Wagner,1 Bettina Hammer,4 Annett Petrich,1 Harald Stein,3 Ulf B. Göbel,1 Thomas Schneider,2 and Annette Moter1*

Institut für Mikrobiologie und Hygiene, Charité—Universitätsmedizin Berlin, Charité Campus Mitte, Dorotheenstr. 96, D-10117 Berlin,1 Medizinische Klinik mit Schwerpunkt Gastroenterologie, Infektiologie, und Rheumatologie,2 Institut für Pathologie/Research Center ImmunoSciences (RCIS), Charité—Universitätsmedizin Berlin, Charité Campus Benjamin Franklin, Hindenburgdamm 30, 12200 Berlin,3 MVZ Labor Dr. Switkowski, Wagner, Dr. Bauermann, Karlsruher Str. 7A, 10711 Berlin, Germany4

Received 23 December 2008/ Returned for modification 2 February 2009/ Accepted 28 February 2009

Human intestinal spirochetosis (HIS) is associated with overgrowth of the large intestine by spirochetes of the genus Brachyspira. The microbiological diagnosis of HIS is hampered by the fastidious nature and slow growth of Brachyspira spp. In clinical practice, HIS is diagnosed histopathologically, and a significant portion of cases may be missed. Fluorescence in situ hybridization (FISH) is a molecular method that allows the visualization and identification of single bacteria within tissue sections. In this study, we analyzed intestinal biopsy samples from five patients with possible HIS. All specimens yielded positive results by histopathological techniques. PCR amplification and sequencing of the 16S rRNA gene were performed. Sequences of two isolates clustered in the group of Brachyspira aalborgi, whereas in three cases, the sequences were highly similar to that of Brachyspira pilosicoli. Three phylotypes showed mismatches at distinct nucleotide positions with Brachyspira sp. sequences published previously. In addition, culture for Brachyspira was successful in three cases. On the basis of these data, we designed and evaluated a Brachyspira genus-specific 16S rRNA-directed FISH probe that detects all of the Brachyspira spp. published to date. FISH of biopsy samples resulted in strong, unequivocal signals of brush-like formations at the crypt surfaces. This technique allowed simultaneous visualization of single spirochetes and their identification as Brachyspira spp. In conclusion, FISH provides a fast and accurate technique for the visualization and identification of intestinal spirochetes in tissue sections. It therefore represents a valuable tool for routine diagnosis of HIS.

* Corresponding author. Mailing address: Charité—Universitätsmedizin Berlin, Institut für Mikrobiologie und Hygiene, Dorotheenstr. 96, D-10117 Berlin, Germany. Phone: 49 30 450524226. Fax: 49 30 450524902. E-mail: annette.moter{at}charite.de

{triangledown} Published ahead of print on 11 March 2009.

Journal of Clinical Microbiology, May 2009, p. 1393-1401, Vol. 47, No. 5
0095-1137/09/$08.00+0     doi:10.1128/JCM.02469-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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