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Journal of Clinical Microbiology, October 1999, p. 3159-3166, Vol. 37, No. 10
0095-1137/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Molecular Analysis of Riboflavin Synthesis Genes in Bartonella henselae and Use of the ribC Gene for Differentiation of Bartonella Species by PCR

Stefan Bereswill,* Silke Hinkelmann, Manfred Kist, and Anna Sander

Department of Microbiology and Hygiene, Institute of Medical Microbiology and Hygiene, University of Freiburg, D-79104 Freiburg, Germany

Received 29 March 1999/Returned for modification 19 May 1999/Accepted 24 June 1999

The biosynthesis pathway for riboflavin (vitamin B2), the precursor of the essential cofactors flavin mononucleotide and flavin adenine dinucleotide, is present in bacteria and plants but is absent in vertebrates. Due to their conservation in bacterial species and their absence in humans, the riboflavin synthesis genes should be well suited either for detection of bacterial DNA in human specimens or for the differentiation of pathogenic bacteria by molecular techniques. A DNA fragment carrying the genes ribD, ribC, and ribE, which encode homologues of riboflavin deaminase (RibD) and subunits of riboflavin synthetase (RibC and RibE), respectively, was isolated from a plasmid-based DNA library of the human pathogen Bartonella henselae by complementation of a ribC mutation in Escherichia coli. Sequence analysis of the ribC gene region in strains of B. henselae, which were previously shown to be genetically different, revealed that the ribC gene is highly conserved at the species level. PCR amplification with primers derived from the ribC locus of B. henselae was used to isolate the corresponding DNA regions in B. bacilliformis, B. clarridgeiae, and B. quintana. Sequence analysis indicated that the riboflavin synthesis genes are conserved and show the same operon-like genetic organization in all four Bartonella species. Primer oligonucleotides designed on the basis of localized differences within the ribC DNA region were successfully used to develop species-specific PCR assays for the differentiation of B. henselae, B. clarridgeiae, B. quintana, and B. bacilliformis. The results obtained indicate that the riboflavin synthesis genes are excellent targets for PCR-directed differentiation of these emerging pathogens. The PCR assays developed should increase our diagnostic potential to differentiate Bartonella species, especially B. henselae and the newly recognized species B. clarridgeiae.

* Corresponding author. Mailing address: Institute of Medical Microbiology and Hygiene, Department of Microbiology and Hygiene, University of Freiburg, Hermann-Herderstr. 11, D-79104 Freiburg, Germany. Phone: 49-761-203-6539. Fax: 49-761-203-6562. E-mail: bereswil{at}sun1.uk1.uni-freiburg.de.

Journal of Clinical Microbiology, October 1999, p. 3159-3166, Vol. 37, No. 10
0095-1137/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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