Skip directly to local search Skip directly to A to Z list Skip directly to navigation Skip directly to site content Skip directly to page options
CDC Home

Volume 19, Number 7—July 2013

Letter

Spotted Fever Group Rickettsiae in Questing Ticks, Central Spain

Suggested citation for this article

To the Editor: The number of spotted fever group (SFG) rickettsiae that cause diseases in humans is rapidly increasing (1,2); infections have been described in ticks and humans in Spain (3,4). However, in Castilla-La Mancha, central Spain, where recreational parks and hunting estates are abundant and humans may be exposed to infected ticks, information on such infections is not available. Therefore, it is worthwhile to characterize Rickettsia spp. found in this area for epidemiologic studies and proper diagnosis of possible rickettsial diseases.

Figure

Thumbnail of Rickettsia species in questing ticks collected in central Spain. A) Study area with 20 collection sites where ticks were found (black dots) of the 39 sites surveyed (white and black dots). B) Multilocus sequence analysis of Rickettsia spp. The evolutionary history was inferred by using the neighbor-joining method of ompA-ompB concatenated sequences (total length = 1,189 nt). The optimal tree with the sum of branch length = 0.15227017 is shown. The percentage of replicate trees in wh

Figure. . Rickettsia species in questing ticks collected in central Spain. A) Study area with 20 collection sites where ticks were found (black dots) of the 39 sites surveyed (white and black...

In this study, we obtained 148 questing adult ticks, representing the most abundant species in the area: 12 Dermacentor marginatus, 26 Rhipicephalus bursa, 41 Rh. sanguineus, 15 Rh. turanicus, 8 Rh. pusillus, 2 Haemaphysalis punctata, 11 Hyalomma lusitanicum, and 33 Hyalomma marginatum (5). The ticks were collected from the vegetation at natural sites surveyed in Castilla-La Mancha by blanket dragging with a cotton flannelette during fall 2009 and spring–summer 2010 (Figure, panel A) and classified (5).

Total DNA was extracted from dissected tick internal organs by using the DNeasy Blood & Tissue Kit (QIAGEN, Düsseldorf, Germany) and used to analyze Rickettsia spp. DNA by PCR, cloning, and sequence analysis of the amplicons. At least 3 clones were sequenced for each amplicon. Genes targeted by PCR included fragments of adenosine triphosphate synthase α subunit (atpA), heat-shock protein 70 (dnaK), outer membrane protein A (ompA), outer membrane protein B (ompB), citrate synthase (gltA), 16S rRNA, recA, and initiator protein of DNA replication (dnaA) (6,7). To characterize Rickettsia spp., we compared nucleotide sequence identity to reference strains and carried out multilocus analysis using ompA-ompB sequences and in silico PstI and RsaI restriction analysis of ompA sequences (7).

Ticks were first screened by 16S rRNA PCR, and positive samples were analyzed for all targeted genes. The results showed that 27 (18.2%) of the 148 ticks analyzed were positive for Rickettsia spp. Of these, 11 were confirmed as R. massiliae in Rh. sanguineus, Rh. turanicus, and Rh. pusillus, 3 as R. raoultii in D. marginatus, 2 as R. slovaca in D. marginatus, and 2 as R. sibirica subsp. mongolitimonae in H. marginatum and Rh. pusillus (Figure, panel B). These species had >99% pairwise nucleotide sequence identity to reference strains R. massiliae MTU5 (GenBank accession no. NC_009900), R. slovaca 13-B (accession no. NC_016639), and R. sibirica subsp. mongolitimonae HA-91 (accession no. AHZB00000000) genome sequences for all genes analyzed, and the only R. raoultii reported sequences (accession nos. JQ792107, JQ792166, JQ792134, and NR_043755 for ompB, ompA, gltA, and 16S rRNA, respectively). The sequences obtained in this study were deposited in the GenBank under accession nos. KC427998–KC428040.

Multilocus sequence analysis of ompA-ompB sequences (Figure, panel B) and in silico PstI and RsaI restriction analysis of ompA sequences also confirmed the identity of the Rickettsia spp. identified in this study. As previously shown (7,8), multilocus analysis with ompA-ompB sequences was highly informative about the phylogenetic relationship between Rickettsia spp. (Figure, panel B), with similar results for maximum likelihood, maximum parsimony, and neighbor-joining methods (data not shown). Furthermore, the results suggested the tick vectors for these Rickettsia spp. in the study area (Figure, panel B) match those reported or suspected previously for these Rickettsia spp. (14), but for the first time, R. sibirica subsp. mongolitimonae was identified in Hyalomma and Rhipicephalus spp. ticks in Spain (4).

These tick species are frequently found in the same area feeding on Eurasian wild boar (Sus scrofa) and red deer (Cervus elaphus), which may act as hosts for these pathogens (5,9). To test this hypothesis, we determined the seroprevalence for SFG rickettsiae in these host species in Castilla-La Mancha. Serum samples from 235 red deer and 206 wild boar were analyzed for the presence of anti-SFG Rickettsia antibodies by ELISA (Spotted Fever Rickettsia IgG EIA Antibody Kit, Fuller Laboratories, Fullerton, CA, USA). The ELISA was adapted to test ungulate serum specimens by substituting antihuman IgG-horseradish by protein G-horseradish peroxidase (Sigma-Aldrich, Madrid, Spain). Specific SFG-Rickettsia antibodies were detected in 146 (70.9%) of 206 wild boar and 174 (74.0%) of 235 red deer, indicating a high seroprevalence in these species and thus the possibility that they can serve as hosts for these pathogens.

These tick species also infest humans, thus posing a risk for transmission of rickettsiae that are pathogenic in humans (1). In fact, Castilla-La Mancha is one of the regions in Spain where a high number of SFG rickettsioses are reported ([10]; http://pagina.jccm.es/sanidad/salud/epidemiologia/3507.pdf).

In conclusion, these results demonstrate that SFG rickettsiae with public health relevance are found in ticks in central Spain as in other regions in Spain. In central Spain, the widespread distribution of tick vectors and possible wildlife hosts, the presence of persons in tick-infested recreational and hunting areas, and the transstadial and transovarial transmission of the pathogen in ticks may favor transmission to humans.

Isabel G. Fernández de Mera, Francisco Ruiz-Fons, Gabriela de la Fuente, Atilio J. Mangold, Christian Gortázar, and José de la FuenteComments to Author 
Author affiliations: Instituto de Investigación en Recursos Cinegéticos (IREC)–CSIC-UCLM-JCCM, Ciudad Real, Spain (I.G. Fernández de Mera, F. Ruiz-Fons, G. de la Fuente, C. Gortázar, J. de la Fuente); Universidad Complutense de Madrid, Madrid, Spain (I.G. Fernández de Mera); Instituto Nacional de Tecnología Agropecuaria, Santa Fe, Argentina (A.J. Mangold); Oklahoma State University, Stillwater, Oklahoma, USA (J. de la Fuente).

Acknowledgments

We thank M. Durán-Martínez and R. Sobrino for help with tick surveys.

F. R.-F. and I.G.F.M. are supported by a Juan de la Cierva contract from the Spanish Ministry for Economy and Competitiveness. Research supported by POII09-0141-8176 and European Union FP7 ANTIGONE (Anticipating the Global Onset of Novel Epidemics) project number 278976.

References

  1. Raoult D, Roux V. Rickettsioses as paradigms of new or emerging infectious diseases. Clin Microbiol Rev. 1997;10:694719 .PubMed
  2. Uchiyama T. Tropism and pathogenicity of rickettsiae. Front. Microbiol. 2012;3:230.
  3. Márquez FJ. Spotted fever group Rickettsia in ticks from southeastern Spain natural parks. Exp Appl Acarol. 2008;45:18594 and. DOIPubMed
  4. Aguirrebengoa K, Portillo A, Santibáñez S, Marín JJ, Montejo M, Oteo JA. Human Rickettsia sibirica mongolitimonae infection, Spain. Emerg Infect Dis. 2008;14:5289 and. DOIPubMed
  5. Ruiz-Fons F, Fernández de Mera IG, Acevedo P, Höfle U, Vicente J, de la Fuente J, Ticks (Acari: Ixodidae) parasitizing Iberian red deer (Cervus elaphus hispanicus) and European wild boar (Sus scrofa) from Spain: geographical and temporal distribution. Vet Parasitol. 2006;140:13342 and. DOIPubMed
  6. Fernández de Mera IG, Zivkovic Z, Bolaños M, Carranza C, Pérez-Arellano JL, Gutiérrez C, Rickettsia massiliae in the Canary Islands. Emerg Infect Dis. 2009;15:186970 and. DOIPubMed
  7. Torina A, Fernández de Mera IG, Alongi A, Mangold AJ, Blanda V, Scarlata F, Rickettsia conorii Indian tick typhus strain and R. slovaca in humans. Sicily. Emerg Infect Dis. 2012;18:100810 . DOIPubMed
  8. Zhu Y, Fournier PE, Eremeeva M, Raoult D. Proposal to create subspecies of Rickettsia conorii based on multi-locus sequence typing and an emended description of Rickettsia conorii. BMC Microbiol. 2005;5:11 and. DOIPubMed
  9. de la Fuente J, Naranjo V, Ruiz-Fons F, Vicente J, Estrada-Peña A, Almazán C, Prevalence of tick-borne pathogens in ixodid ticks (Acari: Ixodidae) collected from European wild boar (Sus scrofa) and Iberian red deer (Cervus elaphus hispanicus) in central Spain. Eur J Wildl Res. 2004;50:18796. DOI
  10. Bartolomé J, Lorente S, Hernández-Pérez N, Martínez-Alfaro E, Marín-Orsa A, Crespo MD. Estudio clínico-epidemiológico de las rickettsiosis del grupo de las fiebres exantemáticas en Albacete. Enferm Infecc Microbiol Clin. 2005;23:1946 and. DOIPubMed

Figure

Suggested citation for this article: Fernández de Mera IG, Ruiz-Fons F, de la Fuente G, Mangold AJ, Gortázar C, de la Fuente J. Spotted fever group rickettsiae in questing ticks, central Spain [letter]. Emerg Infect Dis [Internet]. 2013 Jul [date cited]. http://dx.doi.org/10.3201/eid1907.130005

DOI: 10.3201/eid1907.130005

Related Links

Top of Page

Table of Contents – Volume 19, Number 7—July 2013

Comments to the Authors

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

José de la Fuente, Instituto de Investigación en Recursos Cinegéticos (IREC-CSIC-UCLM-JCCM), Ronda de Toledo s/n, 13005 Ciudad Real, Spain





characters(s) remaining.

Comment submitted successfully, thank you for your feedback.

Comments to the EID Editors

Please contact the EID Editors via our Contact Form.

 

Past Issues

Select a Past Issue:

Art in Science - Selections from Emerging Infectious Diseases
Now available for order



CDC 24/7 – Saving Lives, Protecting People, Saving Money. Learn More About How CDC Works For You…

USA.gov: The U.S. Government's Official Web PortalDepartment of Health and Human Services
Centers for Disease Control and Prevention   1600 Clifton Rd. Atlanta, GA 30333, USA
800-CDC-INFO (800-232-4636) TTY: (888) 232-6348 - Contact CDC–INFO