BACKGROUND: Anaplasma phagocytophilum is characterized by a worldwide distribution and distinguished from other Anaplasmataceae by the broadest range of mammalian hosts and high genetic diversity. The role carnivores play in the life cycle of A. phagocytophilum in Europe is uncertain. Currently, only the red fox is considered a suitable reservoir host. In this study, we focused on native and invasive medium-sized carnivore species that live in sympatry and represent the most abundant species of wild carnivores in Poland. METHODS: A total of 275 individual spleen samples from six carnivore species (Vulpes vulpes, Meles meles, Procyon lotor, Nyctereutes procyonoides and Martes spp.) were screened combining nested PCR and sequencing for A. phagocytophilum targeting a partial groEL gene with subsequent phylogenetic analysis inferred by the maximum likelihood method. RESULTS: The DNA of A. phagocytophilum was detected in 16 of 275 individuals (5.8%). Eight unique genetic variants of A. phagocytophilum were obtained. All detected haplotypes clustered in the clade representing European ecotype I. Three variants belonged to the subclade with European human cases together with strains from dogs, foxes, cats, and wild boars. CONCLUSIONS: While carnivores might have a restricted role in the dissemination of A. phagocytophilum due to their relatively low to moderate infection rates, they hold significance as hosts for ticks. Consequently, they could contribute to the transmission of tick-borne infections to humans indirectly, primarily through tick infection. This underscores the potential risk of urbanization for the A. phagocytophilum life cycle, further emphasizing the need for comprehensive understanding of its ecological dynamics.

Biology Centre Institute of Parasitology Czech Academy of Sciences České Budějovice Czech Republic

Department of Botany and Zoology Faculty of Science Masaryk University Brno Czech Republic

Department of Chemistry and Biochemistry Mendel University Brno Czech Republic

Department of Parasitology Faculty of Biological Sciences University of Wrocław Wrocław Poland

Department of Veterinary Sciences Faculty of Agrobiology Food and Natural Resources Czech University of Life Sciences Prague Prague Czech Republic

Faculty of Medicine in Pilsen Biomedical Center Pilsen Czech Republic

Zobrazit více v PubMed

Ben Said M, Belkahia H, Messadi L. Anaplasma spp. in North Africa: a review on molecular epidemiology, associated risk factors and genetic characteristics. Ticks Tick Borne Dis. 2018;9:543–555. PubMed

Woldehiwet Z. The natural history of Anaplasma phagocytophilum. Vet Parasitol. 2010;167:108–122. PubMed

Rar V, Tkachev S, Tikunova N. Genetic diversity of Anaplasma bacteria: twenty years later. Infect Genet Evol. 2021;91:104833. PubMed

Jahfari S, Coipan EC, Fonville M, Van Leeuwen AD, Hengeveld P, Heylen D, Heyman P, Van Maanen C, Butler CM, Földvári G, Szekeres S. Circulation of four Anaplasma phagocytophilum ecotypes in Europe. Parasit Vectors. 2014;7:365. PubMed PMC

Jaarsma RI, Sprong H, Takumi K, Kazimirova M, Silaghi C, Mysterud A, Rudolf I, Beck R, Földvári G, Tomassone L, Groenevelt M. Anaplasma phagocytophilum evolves in geographical and biotic niches of vertebrates and ticks. Parasit Vectors. 2019;12:328. PubMed PMC

Stigum VM, Jaarsma RI, Sprong H, Rolandsen CM, Mysterud A. Infection prevalence and ecotypes of Anaplasma phagocytophilum in moose Alces alces, red deer Cervus elaphus, roe deer Capreolus capreolus and Ixodes ricinus ticks from Norway. Parasit Vectors. 2019;12:1–8. PubMed PMC

Hamšíková Z, Silaghi C, Takumi K, Rudolf I, Gunár K, Sprong H, Kazimírová M. Presence of roe deer affects the occurrence of Anaplasma phagocytophilum ecotypes in questing Ixodes ricinus in different habitat types of central Europe. Int J Environ Res Public Health. 2019;16:4725. PubMed PMC

Praskova I, Bezdekova B, Zeman P, Jahn P. Seroprevalence of Anaplasma phagocytophilum in horses in the Czech Republic. Ticks Tick Borne Dis. 2011;2:111–115. PubMed

Franzén P, Aspan A, Egenvall A, Gunnarsson A, Karlstam E, Pringle J. Molecular evidence for persistence of Anaplasma phagocytophilum in the absence of clinical abnormalities in horses after recovery from acute experimental infection. J Vet Intern Med. 2009;23:636–642. PubMed

Kapiainen S. The prevalence of Anaplasma phagocytophilum in Finnish dogs. 2016. https://core.ac.uk/works/37117259.

Karbowiak G, Vichova B, Majlathova V, Hapunik J, Petko B. Anaplasma phagocytophilum infection of red foxes (Vulpes vulpes) Ann Agric Environ Med. 2009;16:299–300. PubMed

Tolnai Z, Sréter-Lancz Z, Sréter T. Spatial distribution of Anaplasma phagocytophilum and Hepatozoon canis in red foxes (Vulpes vulpes) in Hungary. Ticks Tick Borne Dis. 2015;6:645–648. PubMed

Hodžić A, Alić A, Fuehrer HP, Harl J, Wille-Piazzai W, Duscher GG. A molecular survey of vector-borne pathogens in red foxes (Vulpes vulpes) from Bosnia and Herzegovina. Parasit Vectors. 2015;8:88. PubMed PMC

Földvári G, Jahfari S, Rigó K, Jablonszky M, Szekeres S, Majoros G, Tóth M, Molnár V, Coipan EC, Sprong H. Candidatus Neoehrlichia mikurensis and Anaplasma phagocytophilum in urban hedgehogs. Emerg Infect Dis. 2014;20:496–497. PubMed PMC

Ruszkowski JJ, Hetman M, Turlewicz-Podbielska H, Pomorska-Mól M. Hedgehogs as a potential source of zoonotic pathogens—a review and an update of knowledge. Animals. 2021;11:1754. PubMed PMC

Silaghi C, Skuballa J, Thiel C, Pfister K, Petney T, Pfäffle M, Taraschewski H, Passos LM. The European hedgehog (Erinaceus europaeus)—a suitable reservoir for variants of Anaplasma phagocytophilum? Ticks Tick Borne Dis. 2012;3:49–54. PubMed

Lesiczka PM, Hrazdilová K, Majerová K, Fonville M, Sprong H, Hönig V, Hofmannová L, Papežík P, Růžek D, Zurek L, Votýpka J. The role of peridomestic animals in the eco-epidemiology of Anaplasma phagocytophilum. Microb Ecol. 2021;82:602–612. PubMed

Hrazdilová K, Lesiczka PM, Bardoň J, Vyroubalová Š, Šimek B, Zurek L, Modrý D. Wild boar as a potential reservoir of zoonotic tick-borne pathogens. Ticks Tick Borne Dis. 2021;12:101558. PubMed

de la Fuente J, Gortazar C. Wild boars as hosts of human-pathogenic Anaplasma phagocytophilum variants. Emerg Infect Dis. 2012;18:2094–2095. PubMed PMC

Silaghi C, Pfister K, Overzier E. Molecular investigation for bacterial and protozoan tick-borne pathogens in wild boars (Sus scrofa) from southern Germany. Vector-Borne Zoonot Dis. 2014;14:371–373. PubMed PMC

Humair PF, Douet V, Cadenas FM, Schouls LM, Van De PI, Gern L. Molecular identification of bloodmeal source in Ixodes ricinus ticks using 12S rDNA as a genetic marker. J Med Entomol. 2007;44:869–880. PubMed

Hvidsten D, Frafjord K, Gray JS, Henningsson AJ, Jenkins A, Kristiansen BE, Lager M, Rognerud B, Slåtsve AM, Stordal F, Stuen S. The distribution limit of the common tick, Ixodes ricinus, and some associated pathogens in north-western Europe. Ticks Tick Borne Dis. 2020;11:101388. PubMed

Jahfari S, Ruyts SC, Frazer-Mendelewska E, Jaarsma R, Verheyen K, Sprong H. Melting pot of tick-borne zoonoses: the European hedgehog contributes to the maintenance of various tick-borne diseases in natural cycles urban and suburban areas. Parasit Vectors. 2017;10:1–9. PubMed PMC

Krawczyk AI, Van Leeuwen AD, Jacobs-Reitsma W, Wijnands LM, Bouw E, Jahfari S, van Hoek AH, van der Giessen JW, Roelfsema JH, Kroes M, Kleve J. Presence of zoonotic agents in engorged ticks and hedgehog faeces from Erinaceus europaeus in (sub) urban areas. Parasit Vectors. 2015;8:210. PubMed PMC

Baldridge GD, Scoles GA, Burkhardt NY, Schloeder B, Kurtti TJ, Munderloh UG. Transovarial transmission of Francisella-like endosymbionts and Anaplasma phagocytophilum variants in Dermacentor albipictus (Acari: Ixodidae) J Med Entomol. 2009;46:625–632. PubMed PMC

Bonnet SI, Binetruy F, Hernández-Jarguín AM, Duron O. The tick microbiome: why non-pathogenic microorganisms matter in tick biology and pathogen transmission. Front Cell Infect Microbiol. 2017;7:236. PubMed PMC

Szekeres S, Claudia Coipan E, Rigó K, Majoros G, Jahfari S, Sprong H, Földvári G. Candidatus Neoehrlichia mikurensis and Anaplasma phagocytophilum in natural rodent and tick communities in Southern Hungary. Ticks Tick Borne Dis. 2015;6:111–116. PubMed

Wirtgen M, Nahayo A, Linden A, Garigliany M, Desmecht D. Tickborne diseases: detection of Anaplasma phagocytophilum in Dermacentor reticulatus ticks. Vet Rec. 2011;168:195. PubMed

André MR. Diversity of Anaplasma and Ehrlichia/Neoehrlichia agents in terrestrial wild carnivores worldwide: implications for human and domestic animal health and wildlife conservation. Front Vet Sci. 2018;5:293. PubMed PMC

Matei IA, Ivan T, Ionică AM, D’amico G, Deak G, Nadas GC, Novac CS, Gherman CM, Mihalca AD. Anaplasma phagocytophilum in multiple tissue samples of wild carnivores in Romania. J Wildl Dis. 2021;57:949–953. PubMed

Szewczyk T, Werszko J, Myczka AW, Laskowski Z, Karbowiak G. Molecular detection of Anaplasma phagocytophilum in wild carnivores in north-eastern Poland. Parasit Vectors. 2019;12:1–5. PubMed PMC

Vercillo F, Lucentini L, Mucci N, Ragni B, Randi E, Panara F. A simple and rapid PCR-RFLP method to distinguishing Martes martes and Martes foina. Conserv Genet. 2004;5:869–871.

Kearse M, Moir R, Wilson A, Stones-Havas S, Cheung M, Sturrock S, Buxton S, Cooper A, Markowitz S, Duran C, Thierer T. Geneious basic: an integrated and extendable desktop software platform for the organization and analysis of sequence data. Bioinformatics. 2012;28:1647–1649. PubMed PMC

Nguyen LT, Schmidt HA, von Haeseler A, Minh BQ. IQ-TREE: a fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies. Mol Biol Evol. 2015;32:268–274. PubMed PMC

Kalyaanamoorthy S, Minh BQ, Wong TKF, von Haeseler A, Jermiin LS. ModelFinder: fast model selection for accurate phylogenetic estimates. Nat Methods. 2017;14:587–589. PubMed PMC

Minh BQ, Nguyen MAT, von Haeseler A. Ultrafast approximation for phylogenetic bootstrap. Mol Biol Evol. 2013;30:1188–1195. PubMed PMC

Guindon S, Dufayard JF, Lefort V, Anisimova M, Hordijk W, Gascuel O. New algorithms and methods to estimate maximum-likelihood phylogenies: assessing the performance of PhyML 3.0. Syst Biol. 2010;59:307–321. PubMed

Dugat T, Lagrée AC, Maillard R, Boulouis HJ, Haddad N. Opening the black box of Anaplasma phagocytophilum diversity: current situation and future perspectives. Front Cell Infect Microbiol. 2015;5:1–18. PubMed PMC

Hofmeester TR, Krawczyk AI, Van Leeuwen AD, Fonville M, Montizaan MGE, Van Den Berge K, Gouwy J, Ruyts SC, Verheyen K, Sprong H. Role of mustelids in the life-cycle of ixodid ticks and transmission cycles of four tick-borne pathogens. Parasit Vectors. 2018;11:1–3. PubMed PMC

Härtwig V, von Loewenich FD, Schulze C, Straubinger RK, Daugschies A, Dyachenko V. Detection of Anaplasma phagocytophilum in red foxes (Vulpes vulpes) and raccoon dogs (Nyctereutes procyonoides) from Brandenburg, Germany. Ticks Tick Borne Dis. 2014;5:277–280. PubMed

Sgroi G, Iatta R, Veneziano V, Bezerra-Santos MA, Lesiczka P, Hrazdilová K, Annoscia G, D’Alessio N, Golovchenko M, Rudenko N, Modrý D. Molecular survey on tick-borne pathogens and Leishmania infantum in red foxes ( Vulpes vulpes ) from southern Italy. Ticks Tick Borne Dis. 2021;12:101669. PubMed

Lesiczka PM, Rudenko N, Golovchenko M, Juránková J, Daněk O, Modrý D, Hrazdilová K. Red fox (Vulpes vulpes) play an important role in the propagation of tick-borne pathogens. Ticks Tick Borne Dis. 2023;14:102076. PubMed

Myśliwy I, Perec-Matysiak A, Hildebrand J. Invasive raccoon (Procyon lotor) and raccoon dog (Nyctereutes procyonoides) as potential reservoirs of tick-borne pathogens: data review from native and introduced areas. Parasit Vectors. 2022;15:1–11. PubMed PMC

Lindsø LK, Dupont P, Rød-Eriksen L, Andersskog IPØ, Ulvund KR, Flagstad Ø, Bischof R, Eide NE. Estimating red fox density using non-invasive genetic sampling and spatial capture–recapture modelling. Oecologia. 2022;198:139–151. PubMed PMC

Medkour H, Laidoudi Y, Marié JL, Fenollar F, Davoust B, Mediannikov O. Molecular investigation of vector-borne pathogens in red foxes (Vulpes vulpes) from southern France. J Wildl Dis. 2020;56:837–850. PubMed

Ebani VV, Verin R, Fratini F, Poli A, Cerri D. Molecular survey of Anaplasma phagocytophilum and Ehrlichia canis in red foxes (Vulpes vulpes) from central Italy. J Wildl Dis. 2011;47:699–703. PubMed

García-Pérez AL, Oporto B, Espí A, del Cerro A, Barral M, Povedano I, Barandika JF, Hurtado A. Anaplasmataceae in wild ungulates and carnivores in northern Spain. Ticks Tick Borne Dis. 2016;7:264–269. PubMed

Hildebrand J, Perec-Matysiak A, Popiołek M, Merta D, Myśliwy I, Buńkowska-Gawlik K. A molecular survey of spotted fever group rickettsiae in introduced raccoons (Procyon lotor) Parasit Vectors. 2022;15(1):162. doi: 10.1186/s13071-022-05280-0. PubMed DOI PMC

Kjær LJ, Jensen LM, Chriél M, Bødker R, Petersen HH. The raccoon dog (Nyctereutes procyonoides) as a reservoir of zoonotic diseases in Denmark. Int J Parasitol Parasites Wildl. 2021;16:175–182. PubMed PMC

Duscher T, Hodžić A, Glawischnig W, Duscher GG. The raccoon dog (Nyctereutes procyonoides) and the raccoon (Procyon lotor)—their role and impact of maintaining and transmitting zoonotic diseases in Austria. Central Europe Parasitol Res. 2017;116:1411–1416. PubMed PMC

Hildebrand J, Buńkowska-Gawlik K, Adamczyk M, Gajda E, Merta D, Popiołek M, Perec-Matysiak A. The occurrence of Anaplasmataceae in European populations of invasive carnivores. Ticks Tick Borne Dis. 2018;9:934–937. PubMed

Otranto D, Cantacessi C, Pfeffer M, Dantas-Torres F, Brianti E, Deplazes P, Genchi C, Guberti V, Capelli G. The role of wild canids and felids in spreading parasites to dogs and cats in Europe Part I: protozoa and tick-borne agents. Vet Parasitol. 2015;213:12–23. PubMed

Sutor A, Schwarz S, Conraths FJ. The biological potential of the raccoon dog (Nyctereutes procyonoides, Gray 1834) as an invasive species in Europe-new risks for disease spread? Acta Theriol (Warsz) 2014;59:49–59. PubMed PMC

Hofmeester TR, Jansen PA, Wijnen HJ, Coipan EC, Fonville M, Prins HHTT, Sprong H, van Wieren SE. Cascading effects of predator activity on tick-borne disease risk. Proc R Soc B Biol Sci. 1859;2017:20170453. PubMed PMC

Márton M, Markolt F, Szabó L, Heltai M. Niche segregation between two medium-sized carnivores in a hilly area of Hungary. Ann Zool Fenn. 2014;51:423–432.

Kauhala K, Holmala K. Landscape features, home-range size and density of northern badgers (Meles meles) Ann Zool Fenn. 2011;48:221–232.

Holmala K, Kauhala K. Habitat use of medium-sized carnivores in southeast Finland- key habitats for rabies spread? Ann Zool Fenn. 2009;46:233–246.