Novel Candidatus Rickettsia Species Detected in Nostril Tick from Human, Gabon, 2014

We report the identification of a nymphal nostril tick (Amblyomma sp.) from a national park visitor in Gabon and subsequent molecular detection and characterization of tickborne bacteria. Our findings provide evidence of a potentially new Rickettsia sp. circulating in Africa and indicate that tick bites may pose a risk to persons visiting parks in the region.

T icks are hematophagous arthropods that parasitize different species of vertebrates, and they serve as intermediate hosts for infectious pathogens that can have serious implications for humans. Because of climate change and socioeconomic factors, tickborne diseases have increased in the past 3 decades, and these arthropods are second only to mosquitoes as vectors of human infectious diseases (1,2). Many ixodid tick species are found in Africa, and tickborne diseases in travelers returning from that continent have been reported worldwide (3). Among the travelassociated cases of African tick-bite fever, most occur in persons returning from travel to southern Africa with fever and systemic illness (4).

The Study
A 21-year-old female field worker from Spain visited Lopé National Park in Gabon (Africa) for 13 days during January-February 2014 to observe chimpanzees and gorillas. Four days before returning to Spain, she noticed a foreign body (black spot) inside her left nostril but had no signs or symptoms of illness. After returning home, the woman sought care at the Tropical Medicine Centre at University Hospital Ramón y Cajal in Madrid, Spain, where a tick attached to the anteroinferior part of the left nasal septum (the Kiesselbach area) was extracted with forceps during rhinofibroscopy. The tick was sent to the Center of Rickettsiosis and Arthropod-Borne Diseases at Hospital San Pedro-Center of Biomedical Research of La Rioja in Logroño, Spain, for identification and molecular detection of tickborne bacteria. The tick was photographed (Figure) and identified, by morphologic features, as an Amblyomma sp. nymph, according to taxonomic keys (5).
Immature stages of Amblyomma ticks cannot be identified to the species level on the basis of morphologic features without allowing the nymph to molt. Thus, we conducted genetic analysis to identify the tick. To extract genomic DNA, we incubated the tick with ammonium hydroxide (1 mL of 25% ammonia and 19 mL of sterile water) for 20 min at 100°C and for another 20 min at 90°C. The DNA was used as template in PCR assays targeting the tick mitochondrial 16S rRNA (6), mitochondrial 12S rRNA (7), and nuclear 5.8S-28S rRNA intergenic transcribed spacer 2 (ITS2) (8). As a positive control, we used DNA extract from a tick of known identity (Haemaphysalis punctata) that was collected in La Rioja, Spain.
To screen for the presence of A. phagocytophilum and Borrelia spp., we performed PCR targeting the partial msp2 gene (334 bp) and Borrelia genus-specific 16S rRNA ( For the strain in this study, single bands of the expected sizes for the 2 ompB rickettsial fragment genes analyzed were detected. A BLAST search revealed that these 2 sequences were genetically most similar (97.2% and 98.3% identity) to the ompB gene of Rickettsia japonica and Rickettsia heilongjiangensis, respectively (Table). The nucleotide sequence of ompA was closest (99.8% identity) to that of the ompA of Rickettsia sp. strain Davousti, and showed maximum identity (97.2%) with R. heilogjiangensis as validated species. When compared with sequences of validly published Rickettsia spp. available in GenBank, the 16S rRNA and sca4 gene sequences showed the highest identity with R. japonica (99.4%-99.6% and 98.5%, respectively). The gltA sequence shared 99.1% identity with R. japonica and R. heilogjiangensis (Table). These results are in accordance with the genetic criteria for identifying the rickettsia as Candidatus Rickettsia sp. (online Technical Appendix, reference 16).
Although reports about the circulation of Rickettsia spp. in Gabon are scarce, Rickettsia sp. strain Davousti was detected in Amblyomma tholloni ticks from African elephants in that country (online Technical Appendix, reference 22). In addition, the gltA sequence obtained in our study was 99.9% identical to the gltA sequence of Rickettsia sp. strain Davousti (Table). These findings suggest that both strains could belong to the same Rickettsia sp. No other sequences, apart from those for ompA and gltA, of Rickettsia sp. strain Davousti have been deposited in GenBank; however, on the basis of findings in the previous report (online Technical Appendix, reference 22), we propose the name Candidatus R. davousti for the strain in this study.
In addition, we did not obtain amplicons for the msp2 gene of A. phagocytophilum or 16S rRNA gene specific for Borrelia genus. For each set of PCR primers, no bands were detected on agarose gels for negative control samples.

Conclusions
We report the detection of a potentially novel Rickettsia sp. from an Amblyomma sp. nymphal tick that was removed from the nostril of a field researcher when she returned to Spain after visiting Gabon's Lopé National Park; we propose the name Ca. R. davousti for this Rickettsia sp. strain.  Our findings provide further evidence of the presence of circulating Rickettsia sp. in Africa and indicate that tick bites may be a threat to persons visiting national parks in Africa. Further studies are needed to determine the prevalence of Ca. R. davousti and to establish whether this bacterium is pathogenic for humans.