Molecular and histopathological detection of Hepatozoon canis in red foxes (Vulpes vulpes) from Portugal

Particle and Fibre Toxicology

Gad Baneth¹  Paula A Rodrigues³  Maria João Vila-Viçosa⁴  Ana Patrícia Lopes³  Antónia Reis⁴  Osnat Eyal¹  Helder CE Cortes⁴  Luís Cardoso² 

[1] Koret School of Veterinary Medicine, The Hebrew University of Jerusalem, Rehovot, Israel;Parasite Disease Group, Instituto de Biologia Molecular e Celular, Universidade do Porto, Oporto, Portugal;Animal and Veterinary Research Centre, UTAD, Vila Real, Portugal;Victor Caeiro Laboratory of Parasitology, Instituto de Ciências Agrárias e Ambientais Mediterrânicas, Universidade de Évora, Évora, Portugal

关键词: Vulpes vulpes;    Red fox;    Portugal;    Polymerase chain reaction;    Histopathology;    Hepatozoon canis;    Canine vector-borne diseases;   

Others  :  809738
DOI  :  10.1186/1756-3305-7-113

 received in 2013-12-08, accepted in 2014-01-11,  发布年份 2014

【 摘 要 】

Background

Hepatozoon canis is a protozoan tick-borne pathogen of dogs and wild canids. Hepatozoon spp. have been reported to infect foxes in different continents and recent studies have mostly used the polymerase chain reaction (PCR) for the detection and characterization of the infecting species. Surveying red foxes (Vulpes vulpes) may contribute to better understanding the epidemiology of canine vector-borne diseases, including hepatozoonosis caused by H. canis in domestic dogs. The present study investigated the prevalence of Hepatozoon spp. by means of histopathology and molecular analysis of different tissues in red foxes from different parts of Portugal.

Methods

Blood and tissues including bone marrow, heart, hind leg muscle, jejunum, kidney, liver, lung, popliteal or axillary lymph nodes, spleen and/or tongue were collected from 91 red foxes from eight districts in northern, central and southern Portugal. Tissues were formalin-fixed, paraffin-embedded, cut and stained with hematoxylin and eosin. Polymerase chain reaction (PCR) amplified a ~650 bp fragment of the 18S rRNA gene of Hepatozoon spp. and the DNA products were sequenced.

Results

Hepatozoon canis was detected in 68 out of 90 foxes (75.6%) from all the sampled areas by PCR and sequencing. Histopathology revealed H. canis meronts similar in shape to those found in dogs in the bone marrow of 11 (23.4%) and in the spleen of two (4.3%) out of 47 foxes (p = 0.007). All the 11 foxes found positive by histopathology were also positive by PCR of bone marrow and/or blood. Positivity by PCR (83.0%) was significantly higher (p < 0.001) than by histopathological examination (23.4%) in paired bone marrow samples from the same 47 foxes. Sequences of the 18S rRNA gene of H. canis were 98–99% identical to those in GenBank.

Conclusions

Hepatozoon canis was found to be highly prevalent in red fox populations from northern, central and southern Portugal. Detection of the parasite by histopathology was significantly less sensitive than by PCR. Red foxes are a presumptive reservoir of H. canis infection for domestic dogs.

【 授权许可】

   
2014 Cardoso et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

【 参考文献 】

• [1]Baneth G: Hepatozoonosis – Hepatozoon canis infection. In Infectious Diseases of the Dog and Cat. 4th edition. Edited by Greene CE. St. Louis: Elsevier Saunders; 2012:750-757.
• [2]Baneth G: Perspectives on canine and feline hepatozoonosis. Vet Parasitol 2011, 181:3-11.
• [3]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
• [4]Dantas-Torres F, Capelli G, Giannelli A, Ramos RA, Lia RP, Cantacessi C, de Caprariis D, De Tommasi AS, Latrofa MS, Lacasella V, Tarallo VD, Di Paola G, Qurollo B, Breitschwerdt E, Stanneck D, Otranto D: Efficacy of an imidacloprid/flumethrin collar against fleas, ticks and tick-borne pathogens in dogs. Parasit Vectors 2013, 6:245. BioMed Central Full Text
• [5]Christophers SR: The sexual life cycle of Leucocytozoon canis in the tick. Sci Mem Off Med Sanit Dep Gov India 1907, 28:1-11.
• [6]Baneth G, Samish M, Shkap V: Life cycle of Hepatozoon canis (Apicomplexa: Adeleorina: Hepatozoidae) in the tick Rhipicephalus sanguineus and domestic dog (Canis familiaris). J Parasitol 2007, 93:283-299.
• [7]Murata T, Inoue M, Tateyama S, Taura Y, Nakama S: Vertical transmission of Hepatozoon canis in dogs. J Vet Med Sci 1993, 55:867-868.
• [8]Baneth G, Samish M, Alekseev E, Aroch I, Shkap V: Transmission of Hepatozoon canis to dogs by naturally-fed or percutaneously-injected Rhipicephalus sanguineus ticks. J Parasitol 2001, 87:606-611.
• [9]Baneth G, Weigler B: Retrospective case–control study of hepatozoonosis in dogs in Israel. J Vet Intern Med 1997, 11:365-370.
• [10]Otranto D, Dantas-Torres F, Weigl S, Latrofa MS, Stanneck D, Decaprariis D, Capelli G, Baneth G: Diagnosis of Hepatozoon canis in young dogs by cytology and PCR. Parasit Vectors 2011, 4:55. BioMed Central Full Text
• [11]Baneth G, Barta JR, Shkap V, Martin DS, Macintire DK, Vincent-Johnson N: Genetic and antigenic evidence supports the separation of Hepatozoon canis and Hepatozoon americanum at the species level. J Clin Microbiol 2000, 38:1298-1301.
• [12]Baneth G, Sheiner A, Eyal O, Hahn S, Beaufils JP, Anug Y, Talmi-Frank D: Redescription of Hepatozoon felis (Apicomplexa: Hepatozoidae) based on phylogenetic analysis, tissue and blood form morphology, and possible transplacental transmission. Parasit Vectors 2013, 6:102. BioMed Central Full Text
• [13]Criado-Fornelio A, Buling A, Cunha-Filho NA, Ruas JL, Farias NA, Rey-Valeiron C, Pingre JL, Etievant M, Barba-Carretero JC: Development and evaluation of a quantitative PCR assay for detection of Hepatozoon sp. Vet Parasitol 2007, 150:352-356.
• [14]Oyamada M, Davoust B, Boni M, Dereure J, Bucheton B, Hammad A, Itamoto K, Okuda M, Inokuma H: Detection of Babesia canis rossi, B. canis vogeli, and Hepatozoon canis in dogs in a village of eastern Sudan by using a screening PCR and sequencing methodologies. Clin Diagn Lab Immunol 2005, 12:1343-1346.
• [15]Sasanelli M, Paradies P, Greco B, Eyal O, Zaza V, Baneth G: Failure of imidocarb dipropionate to eliminate Hepatozoon canis in naturally infected dogs based on parasitological and molecular evaluation methods. Vet Parasitol 2010, 171:194-199.
• [16]Conceição-Silva FM, Abranches P, Silva-Pereira MC, Janz JG: Hepatozoonosis in foxes from Portugal. J Wildl Dis 1988, 24:344-347.
• [17]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, 119:21-26.
• [18]Criado-Fornelio A, Ruas JL, Casado N, Farias NA, Soares MP, Müller G, Brumt JG, Berne ME, Buling-Saraña A, Barba-Carretero JC: New molecular data on mammalian Hepatozoon species (Apicomplexa: Adeleorina) from Brazil and Spain. J Parasitol 2006, 92:93-99.
• [19]Majláthová V, Hurníková Z, Majláth I, Petko B: Hepatozoon canis infection in Slovakia: imported or autochthonous? Vector Borne Zoonotic Dis 2007, 7:199-202.
• [20]Deždek D, Vojta L, Ćurković S, Lipej Z, Mihaljević Ž, Cvetnić Ž, Beck R: Molecular detection of Theileria annae and Hepatozoon canis in foxes (Vulpes vulpes) in Croatia. Vet Parasitol 2010, 172:333-336.
• [21]Gabrielli S, Kumlien S, Calderini P, Brozzi A, Iori A, Cancrini G: The first report of Hepatozoon canis identified in Vulpes vulpes and ticks from Italy. Vector Borne Zoonotic Dis 2010, 10:855-859.
• [22]Allen KE, Yabsley MJ, Johnson EM, Reichard MV, Panciera RJ, Ewing SA, Little SE: Novel Hepatozoon in vertebrates from the southern United States. J Parasitol 2011, 97:648-653.
• [23]Giannitti F, Diab SS, Uzal FA, Fresneda K, Rossi D, Talmi-Frank D, Baneth G: Infection with a Hepatozoon sp. closely related to Hepatozoon felis in a wild Pampas gray fox (Lycalopex – Pseudalopex – gymnocercus) co-infected with canine distemper virus. Vet Parasitol 2012, 186:497-502.
• [24]Almeida AP, Souza TD, Marcili A, Labruna MB: Novel Ehrlichia and Hepatozoon agents infecting the crab-eating fox (Cerdocyon thous) in southeastern Brazil. J Med Entomol 2013, 50:640-646.
• [25]Cardoso L, Cortes HC, Reis A, Rodrigues P, Simões M, Lopes AP, Vila-Viçosa MJ, Talmi-Frank D, Eyal O, Solano-Gallego L, Baneth G: Prevalence of Babesia microti-like infection in red foxes (Vulpes vulpes) from Portugal. Vet Parasitol 2013, 196:90-95.
• [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, 136:307-316.
• [28]Inokuma H, Okuda M, Ohno K, Shimoda K, Onishi T: Analysis of the 18S rRNA gene sequence of a Hepatozoon detected in two Japanese dogs. Vet Parasitol 2002, 106:265-271.
• [29]Petrie A, Watson P: Statistics for Veterinary and Animal Science. 2nd edition. Oxford: Blackwell Publishing; 2006.
• [30]Criado-Fornelio A, Martinez-Marcos A, Buling-Saraña A, Barba-Carretero JC: Molecular studies on Babesia, Theileria and Hepatozoon in southern Europe. Part I. Epizootiological aspects. Vet Parasitol 2003, 113:189-201.
• [31]Gimenez C, Casado N, Criado-Fornelio A, de Miguel FA, Dominguez-Peñafiel G: A molecular survey of Piroplasmida and Hepatozoon isolated from domestic and wild animals in Burgos (northern Spain). Vet Parasitol 2009, 26:147-150.
• [32]Criado-Fornelio A, Rey-Valeiron C, Buling A, Barba-Carretero JC, Jefferies R, Irwin P: New advances in molecular epizootiology of canine hematic protozoa from Venezuela, Thailand and Spain. Vet Parasitol 2007, 144:261-269.
• [33]Little SE, Allen KE, Johnson EM, Panciera RJ, Reichard MV, Ewing SA: New developments in canine hepatozoonosis in North America: a review. Parasit Vectors 2009, 2(Suppl 1):S5. BioMed Central Full Text
• [34]Caeiro V: General review of tick species present in Portugal. Parassitologia 1999, 41(Suppl 1):11-15.
• [35]Giannelli A, Ramos RA, Di Paola G, Mencke N, Dantas-Torres F, Baneth G, Otranto D: Transstadial transmission of Hepatozoon canis from larvae to nymphs of Rhipicephalus sanguineus. Vet Parasitol 2013, 196:1-5.
• [36]Johnson EM, Panciera RJ, Allen KE, Sheets ME, Beal JD, Ewing SA, Little SE: Alternate pathway of infection with Hepatozoon americanum and the epidemiologic importance of predation. J Vet Intern Med 2009, 23:1315-1318.
• [37]Cassini R, Zanutto S, Frangipane di Regalbono A, Gabrielli S, Calderini P, Moretti A, Tampieri MP, Pietrobelli M: Canine piroplasmosis in Italy: epidemiological aspects in vertebrate and invertebrate hosts. Vet Parasitol 2009, 165:30-35.
• [38]Menn B, Lorentz S, Naucke TJ: Imported and travelling dogs as carriers of canine vector-borne pathogens in Germany. Parasit Vectors 2010, 3:34. BioMed Central Full Text
• [39]Vilhena H, Martinez-Díaz VL, Cardoso L, Vieira L, Altet L, Francino O, Pastor J, Silvestre-Ferreira AC: Feline vector-borne pathogens in the north and centre of Portugal. Parasit Vectors 2013, 6:99. BioMed Central Full Text
• [40]Lindblad-Toh K, Wade CM, Mikkelsen TS, Karlsson EK, Jaffe DB, Kamal M, Clamp M, Chang JL, Kulbokas EJ 3rd, Zody MC, Mauceli E, Xie X, Breen M, Wayne RK, Ostrander EA, Ponting CP, Galibert F, Smith DR, DeJong PJ, Kirkness E, Alvarez P, Biagi T, Brockman W, Butler J, Chin CW, Cook A, Cuff J, Daly MJ, DeCaprio D, Gnerre S, et al.: Genome sequence, comparative analysis and haplotype structure of the domestic dog. Nature 2005, 438:803-819.