Skip directly to site content Skip directly to page options Skip directly to A-Z link Skip directly to A-Z link Skip directly to A-Z link
Volume 16, Number 12—December 2010
Letter

Molecular Detection of Bartonella alsatica in Rabbit Fleas, France

On This Page
Tables
Article Metrics
6
citations of this article
EID Journal Metrics on Scopus

Cite This Article

To the Editor: Bartonella alsatica was first isolated from the blood of wild rabbits from the Alsace region in France (1). This bacterium is now considered an emerging infectious disease zoonotic agent in persons in close contact with rabbits; at least 2 human cases of endocarditis and 1 human case of lymphadenitis have been reported (24). In this study, we report the molecular detection of B. alsatica in fleas (Spilopsyllus cuniculi) collected from rabbits in southern France.

During January and February 2008, a total of 60 fleas were collected from wild rabbits (Oryctolagus cuniculus) from 3 regions in southern France: Canohes (42°38′N, 2°51′E), Pollestres (42°38′N, 2°52′E), and Toreilles (42°45′N, 2°58′E). The fleas were collected and identified phenotypically, kept in ethanol, and sent to Unité de Recherche sur les Maladies Infectieuses et Tropicales Emergentes in September 2009.

DNA from these fleas, as well as negative controls from uninfected lice maintained as colonies in our laboratory, were extracted by using a QIAmp Tissue Kit (QIAGEN, Hilden, Germany), as described (5). Identification of flea species at the molecular level was achieved by PCR amplification and sequencing of partial siphonapteran 18S rDNA gene (1.95 kbp) as described (5). Sequences were assembled in Sequencher 4.2 (GeneCodes Corporation, Ann Arbor, MI, USA). DNA was used as templates in a real-time quantitative PCR specific for a portion of the Bartonella genus 16S–23S intergenic spacer (ITS) performed in a Smart cycler instrument (Cepheid, Sunnyvale, CA, USA), as described (2). Positive samples at the genus level were confirmed by PCR amplification and sequencing of the Bartonella ITS region, as described (2). Finally, B. alsatica amplification and specific identification was confirmed by using 2 new specific PCRs with primers and TaqMan probes (Applied Biosystems, Courtaboeuf, France) specific for a portion of the heat shock protein 60 (hsp60) and the DNA gyrase subunit B (gyrB) genes of B. alsatica (Table). Specificity of these 2 PCRs was verified in silico (computer simulation) and by using a panel of 14 Bartonella species available in our laboratory (data not shown).

All fleas were morphologically identified as S. cuniculi by using current taxonomic criteria (6). Moreover, the 18S rRNA gene amplified and sequenced as described (6) from fleas gave a sequence with 100% similarity with the sequence of S. cuniculi fleas deposited in GenBank (accession no. EU336097). B. alsatica was detected by ITS reverse transcription–PCR in 8 (13.3%) of 60 fleas: 6 from Toreilles (17.6%, 6/34) region, 2 from Canohes (10.5%; 2/19), and none from Pollestres (0/7). Sequences obtained after PCR amplification and sequencing of partial ITS showed 96.6% identity with B. alsatica (GenBank accession no. HM060955). Using our 2 new PCRs specific for partial hsp60 and gyrB genes from B. alsatica, we identified all Bartonella spp.–positive fleas, which had cycle threshold values ranging from 12.15 to <32.35 and 13.21 to <36.99 for hsp60 and gyrB genes, respectively.

We report the specific detection of B. alsatica in S. cuniculi rabbit fleas from southern France using 4 different PCRs and sequencing, including 2 new reverse transcription PCRs described in this study. There is 1 report of molecular detection of B. alsatica from S. cuniculi fleas from a European wildcat (Felis silvestris silvestris) in Andalusia, Spain (7). Although S. cuniculi fleas are rare on cats, this study demonstrates that cats in contact with rabbits may be infected by these fleas and consequently become a potential source for B. alsatica transmission to humans. Márquez has also recently reported the molecular detection of B. alsatica in blood from 48/279 (17.2%) of wild rabbits (O. cuniculus) in Andalusia, Spain (8).

In conclusion, further research is needed to better understand the mode of transmission of B. alsatica in humans and mammals and the role of rabbit fleas for potential transmission for these bacteria. The recent description of B. alsatica as a human pathogen and the discovery of rabbit fleas as a potential vector reemphasize the emergence potential of this bacterium in humans who have close contact with rabbits.

Top

Acknowledgment

We are grateful to Jean-Claude Beaucournu for help in flea identification. We also thank Juline Collin and Oleg Mediannikov for their technical assistance.

Top

Tahar Kernif, Philippe Parola, Jean-Claude Ricci, Didier Raoult, and Jean-Marc RolainComments to Author 
Author affiliations: Author affiliations: Université de la Méditerranée, Marseille, France (T. Kernif, P. Parola, D. Raoult, J.-M. Rolain); Institut Méditerranéen du Patrimoine Cynégétique et Faunistique, Vergèze, France (J.-C. Ricci)

Top

References

  1. Heller  R, Kubina  M, Mariet  P, Riegel  P, Delacour  G, Dehio  C, Bartonella alsatica sp. nov., a new Bartonella species isolated from the blood of wild rabbit. Int J Syst Bacteriol. 1999;49:2838. DOIPubMedGoogle Scholar
  2. Raoult  D, Roblot  F, Rolain  J-M, Besnier  J-M, Loulergue  J, Bastides  F, First isolation of Bartonella alsatica from a valve of a patient with endocarditis. J Clin Microbiol. 2006;44:2789. DOIPubMedGoogle Scholar
  3. Jeanclaude  D, Godmer  P, Leveiller  D, Pouedras  P, Fournier  PE, Raoult  D, Bartonella alsatica endocarditis in a French patient in close contact with rabbits. Clin Microbiol Infect. 2009;15:1101. DOIPubMedGoogle Scholar
  4. Angelakis  E, Lepidi  H, Canel  A, Rispal  P, Perradeau  F, Barre  I, Human case of Bartonella alsatica lymphadenitis. Emerg Infect Dis. 2008;14:19513. DOIPubMedGoogle Scholar
  5. Whiting  MF. Mecoptera is paraphyletic: multiple genes and phylogeny of Mecoptera and Siphonaptera. Zool Scr. 2002;31:93104. DOIGoogle Scholar
  6. Bitam  I, Parola  P, Dittmar  DL, Matsumoto  K, Belkacem  B, Rolain  J-M, First molecular detection of Rickettsia felis in fleas from Algeria. Am J Trop Med Hyg. 2006;74:5325.PubMedGoogle Scholar
  7. Márquez  FJ, Millán  J, Rodríguez-Liébana  JJ, García-Egea  I, Muniain  MA. Detection and identification of Bartonella sp. in fleas from carnivorous mammals in Andalusia, Spain. Med Vet Entomol. 2009;23:3938. DOIPubMedGoogle Scholar
  8. Márquez  FJ. Molecular detection of Bartonella alsatica in European wild rabbits (Oryctolagus cuniculus) in Andalusia (Spain). Vector Borne Zoonotic Dis. 2010. In press. DOIPubMedGoogle Scholar

Top

Table

Top

Cite This Article

DOI: 10.3201/eid1612.100696

Related Links

Top

Table of Contents – Volume 16, Number 12—December 2010

EID Search Options
presentation_01 Advanced Article Search – Search articles by author and/or keyword.
presentation_01 Articles by Country Search – Search articles by the topic country.
presentation_01 Article Type Search – Search articles by article type and issue.

Top

Comments

Please use the form below to submit correspondence to the authors or contact them at the following address:

Jean-Marc Rolain, Université de la Méditerranée, URMITE UMR 6236 CNRS-IRD, Faculté de Médecine et de Pharmacie, Marseille, France

Send To

10000 character(s) remaining.

Top

Page created: August 29, 2011
Page updated: August 29, 2011
Page reviewed: August 29, 2011
The conclusions, findings, and opinions expressed by authors contributing to this journal do not necessarily reflect the official position of the U.S. Department of Health and Human Services, the Public Health Service, the Centers for Disease Control and Prevention, or the authors' affiliated institutions. Use of trade names is for identification only and does not imply endorsement by any of the groups named above.
file_external