Diversity of microorganisms in Hyalomma aegyptium collected from spur-thighed tortoise (Testudo graeca) in North Africa and Anatolia

Ticks carry a diverse community of microorganisms including non-pathogenic symbionts, commensals, and pathogens, such as viruses, bacteria, protozoans, and fungi. The assessment of tick-borne microorganisms (TBM) in tortoises and their ticks is essential to understand their eco-epidemiology, and to map and monitor potential pathogens to humans and other animals. The aim of this study was to characterize the diversity of microorganisms found in ticks collected from the spur-thighed tortoise (Testudo graeca) in North Africa and Anatolia. Ticks feeding on wild T. graeca were collected, and pathogens were screened by polymerase chain reaction using group-specific primers. In total, 131 adult Hyalomma aegyptium ticks were collected from 92 T. graeca in Morocco (n = 48), Tunisia (n = 2), Algeria (n = 70), and Turkey (n = 11). Bacteria and protozoa detected included Hemolivia mauritanica (22.9%), Midichloria mitochondrii (11.4%), relapsing-fever borreliae (8.4%), Ehrlichia spp. (7.6%), Rickettsia spp. (3.4%), Borrelia burgdorferi s.l. (0.9%), Francisella spp. (0.9%), and Wolbachia spp. (0.8%). The characterization of Rickettsia included R. sibirica mongolitimonae (Algeria), R. aeschlimannii (Turkey), and R.africae (Morocco). Hemolivia mauritanica and Ehrlichia spp. prevalence varied significantly with the sampling region/country. We did not detect significant associations in microorganism presence within ticks, nor between microorganism presence and tick mitochondrial DNA haplogroups. This is the first report of Francisella persica-like, relapsing fever borreliae, M. mitochondrii, and Wolbachia spp. in H. aegyptium ticks collected from wild hosts from the South and Eastern Mediterranean region, and of R. sibirica mongolitimonae and R. africae in H. aegyptium from Algeria and Morocco, respectively. Given that T. graeca is a common species in commercial and non-commercial pet trade, the evaluation of the role of this species and its ticks as hosts for TBM is particularly relevant for public health.This study received financial support from Fundação para a Ciência e a Tecnologia by the strategic program of MARE (MARE - UID/MAR/04292/2020) and the transitory norm contract DL57/2016/CP1370/CT89 to ACN and by the FCT IF contract (IF/01257/2012) and project (IF01257/2012/CP0159/CT0005) to AP. The work of EG and AG was funded by the Spanish Ministry of Science through projects CGL2015-64144 and PID2019-105682RA-I00/AEI/10.13039/5011 00011033 (the first with the support of the European Regional Development Fund, MINECO/FEDER).