First report of Anaplasma platys infection in red foxes (Vulpes vulpes) and molecular detection of Ehrlichia canis and Leishmania infantum in foxes from Portugal

Particle and Fibre Toxicology

Gad Baneth²  Paula A Rodrigues⁴  Margarida Simões¹  Maria João Vila-Viçosa¹  Ana Patrícia Lopes³  Yaarit Nachum-Biala²  Helder CE Cortes¹  Matan Gilad²  Luís Cardoso⁴ 

[1] Victor Caeiro Laboratory of Parasitology, Instituto de Ciências Agrárias e Ambientais Mediterrânicas (ICAAM), University of Évora, Évora, Portugal;Koret School of Veterinary Medicine, The Hebrew University of Jerusalem, Rehovot, Israel;Animal and Veterinary Research Centre, UTAD, Vila Real, Portugal;Department of Veterinary Sciences, School of Agrarian and Veterinary Sciences, Universidade de Trás-os-Montes e Alto Douro (UTAD), Vila Real, Portugal

关键词: Vulpes vulpes;    Canine vector-borne diseases;    Red foxes;    Portugal;    Polymerase chain reaction;    Leishmania infantum;    Ehrlichia canis;    Anaplasma platys;   

Others  :  1146434
DOI  :  10.1186/s13071-015-0756-y

 received in 2015-01-29, accepted in 2015-02-20,  发布年份 2015

【 摘 要 】

Background

The bacteria Anaplasma platys and Ehrlichia canis and the protozoan Leishmania infantum are vector-borne agents that cause canine vector-borne diseases, some of which are zoonotic. The present survey investigated the prevalence of Anaplasma, Ehrlichia and Leishmania in red foxes (Vulpes vulpes) from Portugal by molecular analysis, in order to evaluate the epidemiological role of these canids as reservoirs of infection.

Methods

Blood and/or bone marrow samples were collected from 78 red foxes obtained in eight districts of northern, central and southern Portugal. Real-time polymerase chain reactions (PCR) amplified a 123 bp fragment of the 16S rRNA gene of Anaplasma spp. and Ehrlichia spp. and a 265 bp fragment of the L. infantum internal transcribed spacer one (ITS1) region of the rRNA operon evaluated by PCR-high resolution melt analysis (PCR-HRM), with sequencing of the DNA products. A phylogenetic analysis was carried out to compare these to other sequences from Anaplasma spp. and Ehrlichia spp. deposited in GenBank®.

Results

A. platys was detected in 10 (14.5%) and E. canis in two (2.9%) out of 69 foxes; and L. infantum was detected in one (1.3%) of the 78 foxes. The prevalence of A. platys was significantly different from the prevalence of E. canis (p=0.016) and from that of L. infantum (p=0.002). No co-infections were found in any one of the 78 foxes. No statistically significant differences were found between the type of sample (blood and bone marrow), geographic regions (north/centre and south), age (<2 years and ≥2 years) and gender for any one of the agents.

Conclusions

This is the first known report of A. platys in red foxes worldwide, as well as the first molecular evidence of E. canis in foxes from Portugal. The moderate prevalence of A. platys suggests that red foxes may play a role in the epidemiology of infection with this bacterium and serve as a reservoir for domestic dogs.

【 授权许可】

   
2015 Cardoso et al.; licensee BioMed Central.

【 图 表 】

Figure 1.

【 参考文献 】

• [1]Otranto D, Dantas-Torres F, Breitschwerdt EB. Managing canine vector-borne diseases of zoonotic concern: part one. Trends Parasitol. 2009; 25:157-63.
• [2]Dantas-Torres F, Latrofa MS, Annoscia G, Giannelli A, Parisi A, Otranto D. Morphological and genetic diversity of Rhipicephalus sanguineus sensu lato from the New and Old Worlds. Parasit Vectors. 2013; 6:213. BioMed Central Full Text
• [3]Shaw SE, Day MJ, Birtles RJ, Breitschwerdt EB. Tick-borne infectious diseases of dogs. Trends Parasitol. 2001; 17:74-80.
• [4]Alleman AR, Wamsley HL. An update on anaplasmosis in dogs. Vet Med. 2008; 103:212-20.
• [5]Harrus S, Aroch I, Lavy E, Bark H. Clinical manifestations of infectious canine cyclic thrombocytopenia. Vet Rec. 1997; 141:247-50.
• [6]De Tommasi AS, Otranto D, Furlanello T, Tasca S, Cantacessi C, Breitschwerdt EB et al.. Evaluation of blood and bone marrow in selected canine vector-borne diseases. Parasit Vectors. 2014; 7:534. BioMed Central Full Text
• [7]Maggi RG, Mascarelli PE, Havenga LN, Naidoo V, Breitschwerdt EB. Co-infection with Anaplasma platys, Bartonella henselae and Candidatus Mycoplasma haematoparvum in a veterinarian. Parasit Vectors. 2013; 6:103. BioMed Central Full Text
• [8]Breitschwerdt EB, Hegarty BC, Qurollo BA, Saito TB, Maggi RG, Blanton LS et al.. Intravascular persistence of Anaplasma platys, Ehrlichia chaffeensis, and Ehrlichia ewingii DNA in the blood of a dog and two family members. Parasit Vectors. 2014; 7:298. BioMed Central Full Text
• [9]Arraga-Alvarado CM, Qurollo BA, Parra OC, Berrueta MA, Hegarty BC, Breitschwerdt EB et al.. Molecular evidence of Anaplasma platys infection in two women from Venezuela. Am J Trop Med Hyg. 2014; 91:1161-5.
• [10]Neer TM, Breitschwerdt EB, Greene RT, Lappin MR. Consensus statement on ehrlichial disease of small animals from the infectious disease study group of the ACVIM. J Vet Intern Med. 2002; 16:309-15.
• [11]Harrus S, Bark H, Waner T. Canine monocytic ehrlichiosis: an update. Comp Cont Educ Pract Vet. 1997; 19:431-44.
• [12]Fishman Z, Gonen L, Harrus S, Strauss-Ayali D, King R, Baneth G. A serosurvey of Hepatozoon canis and Ehrlichia canis antibodies in wild red foxes (Vulpes vulpes) from Israel. Vet Parasitol. 2004; 119:21-6.
• [13]Maia C, Ramos C, Coimbra M, Bastos F, Martins A, Pinto P et al.. Bacterial and protozoal agents of feline vector-borne diseases in domestic and stray cats from southern Portugal. Parasit Vectors. 2014; 7:115. BioMed Central Full Text
• [14]Perez M, Bodor M, Zhang C, Xiong Q, Rikihisa Y. Human infection with Ehrlichia canis accompanied by clinical signs in Venezuela. Ann N Y Acad Sci. 2006; 1078:110-7.
• [15]Schallig HD, Cardoso L, Semião-Santos SJ. Seroepidemiology of canine leishmaniosis in Évora (southern Portugal): 20-year trends. Parasit Vectors. 2013; 6:100. BioMed Central Full Text
• [16]Bourdeau P, Saridomichelakis MN, Oliveira A, Oliva G, Kotnik T, Gálvez R et al.. Management of canine leishmaniosis in endemic SW European regions: a questionnaire-based multinational survey. Parasit Vectors. 2014; 7:110. BioMed Central Full Text
• [17]Desjeux P. Worldwide increasing risk factors for leishmaniasis. Med Microbiol Immunol. 2001; 190:77-9.
• [18]Koutinas AF, Polizopoulou ZS, Saridomichelakis MN, Argyriadis D, Fytianou A, Plevraki KG. Clinical considerations on canine visceral leishmaniasis in Greece: a retrospective study of 158 cases (1989–1996). J Am Anim Hosp Assoc. 1999; 35:376-83.
• [19]Solano-Gallego L, Miró G, Koutinas A, Cardoso L, Pennisi MG, Ferrer L et al.. LeishVet guidelines for the practical management of canine leishmaniosis. Parasit Vectors. 2011; 4:86. BioMed Central Full Text
• [20]Baneth G, Dank G, Keren-Kornblatt E, Sekeles E, Adini I, Eisenberger CL et al.. Emergence of visceral leishmaniasis in central Israel. Am J Trop Med Hyg. 1998; 59:722-5.
• [21]Duscher G, Fuehrer HP, Kübber-Heiss A. Fox on the run–molecular surveillance of fox blood and tissue for the occurrence of tick-borne pathogens in Austria. Parasit Vectors. 2014; 7:521.
• [22]Cardoso L, Tuna J, Vieira L, Yisaschar-Mekuzas Y, Baneth G. Molecular detection of Anaplasma platys and Ehrlichia canis in dogs from the North of Portugal. Vet J. 2010; 183:232-3.
• [23]Cardoso L, Yisaschar-Mekuzas Y, Rodrigues FT, Costa A, Machado J, Diz-Lopes D et al.. Canine babesiosis in northern Portugal and molecular characterization of vector-borne co-infections. Parasit Vectors. 2010; 3:27. BioMed Central Full Text
• [24]René-Martellet M, Lebert I, Chêne J, Massot R, Leon M, Leal A et al.. Diagnosis and incidence risk of clinical canine monocytic ehrlichiosis under field conditions in Southern Europe. Parasit Vectors. 2015; 8:3. BioMed Central Full Text
• [25]Cardoso L, Cortes HC, Eyal O, Reis A, Lopes AP, Vila-Viçosa MJ et al.. Molecular and histopathological detection of Hepatozoon canis in red foxes (Vulpes vulpes) from Portugal. Parasit Vectors. 2014; 7:113. BioMed Central Full Text
• [26]Saénz-de-Buruaga M, Lucio AJ, Purroy J. Reconocimiento de Sexo y Edad en Especies Cinegéticas. Gobierno Provincial de Álava, Vitoria-Gasteiz; 1991.
• [27]Davidson RK, Gjerde B, Vikøren T, Lillehaug A, Handeland K. Prevalence of Trichinella larvae and extra-intestinal nematodes in Norwegian red foxes (Vulpes vulpes). Vet Parasitol. 2006; 136:307-16.
• [28]Peleg O, Baneth G, Eyal O, Inbar J, Harrus S. Multiplex real-time qPCR for the detection of Ehrlichia canis and Babesia canis vogeli. Vet Parasitol. 2010; 173:292-9.
• [29]Waner T, Nachum-Biala Y, Harrus S. Evaluation of a commercial in-clinic point-of-care polymerase chain reaction test for Ehrlichia canis DNA in artificially infected dogs. Vet J. 2014; 202:618-21.
• [30]Parola P, Roux V, Camicas JL, Baradji I, Brouqui P, Raoult D. Detection of ehrlichiae in African ticks by polymerase chain reaction. Trans R Soc Trop Med Hyg. 2000; 94:707-8.
• [31]Talmi-Frank D, Nasereddin A, Schnur LF, Schönian G, Töz SO, Jaffe CL et al.. Detection and identification of old world Leishmania by high resolution melt analysis. PLoS Negl Trop Dis. 2010; 4:e581.
• [32]Tamura K, Stecher G, Peterson D, Filipski A, Kumar S. MEGA6: Molecular Evolutionary Genetics Analysis version 6.0. Mol Biol Evol. 2013; 30:2725-9.
• [33]Petrie A, Watson P. Statistics for Veterinary and Animal Science. 3rd ed. Wiley-Blackwell, Oxford; 2013.
• [34]Cardoso L, Cortes HC, Reis A, Rodrigues P, Simões M, Lopes AP et al.. Prevalence of Babesia microti-like infection in red foxes (Vulpes vulpes) from Portugal. Vet Parasitol. 2013; 196:90-5.
• [35]Maia C, Ferreira A, Nunes M, Vieira ML, Campino L, Cardoso L. Molecular detection of bacterial and parasitic pathogens in hard ticks from Portugal. Ticks Tick Borne Dis. 2014; 5:409-14.
• [36]Caeiro V. General review of tick species present in Portugal. Parassitologia. 1999; 41 Suppl 1:11-5.
• [37]Santos-Silva MM, Beati L, Santos AS, De Sousa R, Núncio MS, Melo P et al.. The hard-tick fauna of mainland Portugal (Acari: Ixodidae): an update on geographical distribution and known associations with hosts and pathogens. Exp Appl Acarol. 2011; 55:85-121.
• [38]Santos AS, Alexandre N, Sousa R, Núncio MS, Bacellar F, Dumler JS. Serological and molecular survey of Anaplasma species infection in dogs with suspected tickborne disease in Portugal. Vet Rec. 2009; 164:168-71.
• [39]Maia C, Almeida B, Coimbra M, Fernandes MC, Cristóvão JM, Ramos C, et al. Bacterial and protozoal agents of canine vector-borne diseases in the blood of domestic and stray dogs from southern Portugal. Parasit Vectors. in press.
• [40]Alexandre N, Santos AS, Núncio MS, De Sousa R, Boinas F, Bacellar F. Detection of Ehrlichia canis by polymerase chain reaction in dogs from Portugal. Vet J. 2009; 181:343-4.
• [41]Millán J, Candela MG, Palomares F, Cubero MJ, Rodríguez A, Barral M et al.. Disease threats to the endangered Iberian lynx (Lynx pardinus). Vet J. 2009; 182:114-24.
• [42]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.
• [43]Torina A, Blanda V, Antoci F, Scimeca S, D’Agostino R, Scariano E et al.. A molecular survey of Anaplasma spp., Rickettsia spp., Ehrlichia canis and Babesia microti in foxes and fleas from Sicily. Transbound Emerg Dis. 2013; 60 Suppl 2:125-30.
• [44]Cortes S, Vaz Y, Neves R, Maia C, Cardoso L, Campino L. Risk factors for canine leishmaniasis in an endemic Mediterranean region. Vet Parasitol. 2012; 189:189-96.
• [45]Maia C, Gomes J, Cristóvão J, Nunes M, Martins A, Rebêlo E et al.. Feline Leishmania infection in a canine leishmaniasis endemic region, Portugal. Vet Parasitol. 2010; 174:336-40.
• [46]Vilhena H, Martinez-Díaz VL, Cardoso L, Vieira L, Altet L, Francino O et al.. Feline vector-borne pathogens in the north and centre of Portugal. Parasit Vectors. 2013; 6:99. BioMed Central Full Text
• [47]Abranches P, Conceição-Silva FM, Silva-Pereira MC. Kala-azar in Portugal. V. The sylvatic cycle in the enzootic endemic focus of Arrabida. J Trop Med Hyg. 1984; 87:197-200.
• [48]Rioux JA, Albaret JL, Houin R, Dedet JP, Lanotte G. Écologie des leishmanioses dans le sud de la France: 2. Les réservoirs selvatiques. Infestation spontanée de renard (Vulpes vulpes L.). Ann Parasitol Hum Comp. 1968; 43:421-8.
• [49]Bettini S, Pozio E, Gradoni L. Leishmaniasis in Tuscany (Italy): (II) Leishmania from wild Rodentia and Carnivora in a human and canine leishmaniasis focus. Trans R Soc Trop Med Hyg. 1980; 74:77-83.
• [50]Martín-Iniesta F, Martín-Iniesta E, Martin-Luengo F. Papel de perros y zorros como reservorio de leishmaniasis en la región murciana. Resultados preliminares. Rev Iber Parasitol. 1982; 42:307-13.
• [51]Sobrino R, Ferroglio E, Oleaga A, Romano A, Millan J, Revilla M et al.. Characterization of widespread canine leishmaniasis among wild carnivores from Spain. Vet Parasitol. 2008; 155:198-203.
• [52]Davoust B, Mary C, Marié JL. Detection of Leishmania in red foxes (Vulpes vulpes) from southeastern France using real-time quantitative PCR. J Wildl Dis. 2014; 50:130-2.
• [53]Dipineto L, Manna L, Baiano A, Gala M, Fioretti A, Gravino AE et al.. Presence of Leishmania infantum in red foxes (Vulpes vulpes) in southern Italy. J Wildl Dis. 2007; 43:518-20.
• [54]Verin R, Poli A, Ariti G, Nardoni S, Fanucchi Bertuccelli M, Mancianti F. Detection of Leishmania infantum DNA in tissues of free-ranging red foxes (Vulpes vulpes) in Central Italy. Eur J Wildl Res. 2010; 56:689-92.
• [55]Rioux JA, Lanotte G, Destombes P, Vollhardt Y, Croset H. Leishmaniose expérimental du renard. Rec Med Vet. 1971; 147:489-98.
• [56]Millán J, Ferroglio E, Solano-Gallego L. Role of wildlife in the epidemiology of Leishmania infantum infection in Europe. Parasitol Res. 2014; 113:2005-14.
• [57]Cardoso L, Mendão C, Madeira De Carvalho L. Prevalence of Dirofilaria immitis, Ehrlichia canis, Borrelia burgdorferi sensu lato, Anaplasma spp. and Leishmania infantum in apparently healthy and CVBD-suspect dogs in Portugal–a national serological study. Parasit Vectors. 2012; 5:62. BioMed Central Full Text
• [58]Campino L, Maia C. Epidemiologia das leishmanioses em Portugal. Acta Med Port. 2010; 23:859-64.