Rickettsiae in ticks from wild and domestic carnivores of Doñana National Park (Spain) and surrounding area

Abstract

The aim of this study was to determine the vector distribution and the prevalence of Rickettsia spe- cies that circulate among populations of several wild and domestic carnivores in Donana National Park, a nature reserve located in southern Spain. The presence of Rickettsia in hard ticks was evaluated using molecular techniques (PCR amplification and sequencing) over a sample of 430 specimens belonging to eight tick species: Ixodes (I.) ricinus (Linnaeus, 1789); I. (I.) ventalloi Gil Collado, 1936; Pholeoixodes hexagonus (Leach, 1815); Hyalomma (Euhyalomma) lusitanicum Koch, 1844; Rhipicephalus (Rhipicephalus) sanguineus (Latreille, 1806); Rh. (Rh.) turanicus Pomerantsev, 1940; Rh. (Rh.) pusillus Gil Collado, 1938, and Rh. (Digeneus) bursa Canestrini & Fanzago, 1878. These ticks were parasitising Iberian lynx (Lynx pardinus), common genet (Genetta genetta), Egyp- tian mongoose (Herpestes ichneumon), Eurasian badger (Meles meles) and red fox (Vulpes vulpes) studied in the programme for Iberian lynx con- servation in Donana National Park (1). In addi- tion, samples from free-roaming cats and dogs were included in this study (Table 1). After collection, the ticks were immediately placed in vials with 70% ethanol, properly labelled, and were later identified in the laboratory by species, gender and stage using existing taxonomic keys (2). DNA was extracted individually or from monospecific lots using the kit Nucleo Spin Tissue (Macherey-Nagel, Duren, Germany) and specific rickettsial sequences were detected by using PCR primers that amplify a portion of glta, ompA and ompB genes, respectively (3,4). Positive PCR products were sequenced using PCR primers and the GenomeLab DTCS- Quick Start kit (Beck- man Coulter Life, Brea, CA, USA) and a CEQ 2000XL capillary DNA sequencer (Beckman Coul- ter) according to the manufacturer's instructions. Sequences were manually aligned and analysed with Bioedit vers. 7.0.1. and identified using the BLAST feature of GenBank. Overall, specific rickettsial DNA was detected in 52 (31.7%) of 164 of the examined tick lots by PCR amplifying of at least two rickettsiae-specific fragments. Sequence analysis of amplicons of glta, ompA and ompB genes revealed the presence of three rickettsiae species. Rickettsia monacensis and R. helvetica were found in Ixodes ricinus and I. ventalloi (prevalence of 32.3 and 5.9%, respec- tively) parasitising five species of wild carnivores included in this study (lynx, genet, mongoose, badger and fox). Ticks of the Rhipicephalus san- guineus group (Rh. sanguineus, Rh. turanicus and Rh. pusillus) from lynx, mongoose, fox, cat and dog appeared infected with R. massiliae (preva- lence of 28.3%). None of the Ph. hexagonus, H. lusitanicum or Rh. bursa specimens harboured rickettsiae.