title: Occurrence and molecular genotyping of Giardia duodenalis and Cryptosporidium spp. in wild mesocarnivores in Spain.
abstract: There is a surprisingly scarce amount of epidemiological and molecular data on the prevalence, frequency, and diversity of the intestinal protozoan parasites Giardia duodenalis and Cryptosporidium spp. in wildlife in general and mesocarnivore species in particular. Consequently, the extent of the cyst/oocyst environmental contamination attributable to these wild host species and their potential implications for public veterinary health remain largely unknown. In this molecular epidemiological survey a total of 193 individual faecal samples from badgers (Meles meles, n=70), ferrets (Mustela putorius furo, n=2), genets (Genetta genetta, n=6), Iberian lynxes (Lynx pardinus, n=6), beech martens (Martes foina, n=8), mongooses (Herpestes ichneumon, n=2), otters (Lutra lutra, n=2), polecats (Mustela putorius, n=2), red foxes (Vulpes vulpes, n=87), wildcats (Felis silvestris, n=2), and wolves (Canis lupus, n=6) were obtained from road-killed, hunted, and accidentally found carcasses, and from camera-trap surveys or animals entering rescue shelters, during the period December 2003-April 2016. Investigated specimens were collected in five Spanish autonomous regions including Andalusia (n=1), Asturias (n=69), Basque Country (n=49), Castile-La Mancha (n=38), and Extremadura (n=36). The presence of cysts/oocysts was confirmed by PCR-based methods targeting the small subunit (ssu) ribosomal RNA gene of these parasite species. Genotyping of the obtained isolates were attempted at appropriate markers including the glutamate dehydrogenase (G. duodenalis) and the 60-kDa glycoprotein (C. parvum and C. ubiquitum) loci. Overall, G. duodenalis was detected in 8% (7/87) of red foxes, a single beech marten, and a single wolf, respectively. Cryptosporidium was identified in 3% (2/70) of badgers, 8% (7/87) of red foxes, a single genet, and a single mongoose, respectively. None of the nine G. duodenalis isolates generated could be genotyped at the assemblage/sub-assemblage level. Out of the nine Cryptosporidium isolates successfully characterized, three were identified as C. canis (one in a mongoose and two in red foxes), and three as C. parvum (one in a badger and three in red foxes). The remaining three isolates were assigned to C. felis (in a red fox), C. hominis (in a badger), and C. ubiquitum (in a red fox), respectively. Two additional Cryptosporidium isolates infecting a badger and a genet, respectively, were untypable. The red fox was confirmed as a suitable host of potentially zoonotic Cryptosporidium species, mainly C. parvum and C. ubiquitum. The high mobility and wide home range of red foxes, together with their increasing presence in urban and peri-urban settings, may led to the overlapping of sylvatic and domestic cycles of the parasite, and consequently, to an increased risk of cryptosporidiosis in production animals and humans. The detection of C. hominis oocysts in a badger raises the question of whether this finding represents a true infection or a sporadic event of mechanical passage of C. hominis oocyst of anthroponotic origin.
authors.affiliation: Veterinary Faculty, Alfonso X El Sabio University, Avenida Universidad 1, 28691 Villanueva de la Cañada, Madrid, Spain.
authors.forename: Marta
authors.initials: M
authors.lastname: Mateo
authors.affiliation: Parasitology Service, National Centre for Microbiology, Health Institute Carlos III, Ctra. Majadahonda-Pozuelo Km 2, 28220 Majadahonda, Madrid, Spain.
authors.forename: Marta Hernández
authors.initials: MH
authors.lastname: de Mingo
authors.affiliation: Parasitology Service, National Centre for Microbiology, Health Institute Carlos III, Ctra. Majadahonda-Pozuelo Km 2, 28220 Majadahonda, Madrid, Spain.
authors.forename: Aida
authors.initials: A
authors.lastname: de Lucio
authors.affiliation: Parasitology Service, National Centre for Microbiology, Health Institute Carlos III, Ctra. Majadahonda-Pozuelo Km 2, 28220 Majadahonda, Madrid, Spain.
authors.forename: Lucía
authors.initials: L
authors.lastname: Morales
authors.affiliation: Centre for Animal Biotechnology (SERIDA), Camino de Rioseco 1225, La Olla, Deva, 33394 Gijón, Spain.
authors.forename: Ana
authors.initials: A
authors.lastname: Balseiro
authors.affiliation: Centre for Animal Biotechnology (SERIDA), Camino de Rioseco 1225, La Olla, Deva, 33394 Gijón, Spain.
authors.forename: Alberto
authors.initials: A
authors.lastname: Espí
authors.affiliation: Department of Animal Health, Basque Institute of Agricultural Research and Development (NEIKER), Berreaga 1, 48160 Derio, Bizkaia, Spain.
authors.forename: Marta
authors.initials: M
authors.lastname: Barral
authors.affiliation: SaBio Working Group, Institute for Game and Wildlife Research IREC (CSIC-UCLM-JCCM), Ronda de Toledo 12, 13005 Ciudad Real, Spain; SABIOTec Spin Off, Camino de Moledores s/n, 13005 Ciudad Real, Spain.
authors.forename: José Francisco
authors.initials: JF
authors.lastname: Lima Barbero
authors.affiliation: Department of Animal Health, Veterinary Sciences Faculty, Extremadura University, Avda. de la Universidad s/n, 10003, Cáceres, Spain.
authors.forename: Miguel Ángel
authors.initials: MÁ
authors.lastname: Habela
authors.affiliation: Area of Genetics and Animal Breeding, Department of Animal Production and Food Science, Veterinary Sciences Faculty, Extremadura University, Avda. de la Universidad s/n, 10003, Cáceres, Spain.
authors.forename: José L
authors.initials: JL
authors.lastname: Fernández-García
authors.affiliation: United States Department of Agriculture, Agricultural Research Service, Beltsville Agricultural Research Center, Animal Parasitic Diseases Laboratory, Building 1001, Beltsville, MD 20705-2350, USA.
authors.forename: Rafael Calero
authors.initials: RC
authors.lastname: Bernal
authors.affiliation: Fuendeverde Centre for Environmental Education and Natural History, Pilón Alto, s/n, 50142 Fuendetodos, Zaragoza, Spain.
authors.forename: Pamela C
authors.initials: PC
authors.lastname: Köster
authors.affiliation: Livestock Laboratory, Regional Government of Álava, Ctra. de Azua 4, 01520 Vitoria-Gasteiz, Spain.
authors.forename: Guillermo A
authors.initials: GA
authors.lastname: Cardona
authors.affiliation: Parasitology Service, National Centre for Microbiology, Health Institute Carlos III, Ctra. Majadahonda-Pozuelo Km 2, 28220 Majadahonda, Madrid, Spain. Electronic address: dacarmena@isciii.es.
authors.forename: David
authors.initials: D
authors.lastname: Carmena
mesh_terms: D000818:Animals; D002335:Carnivora / Q000469:parasitology*; D003457:Cryptosporidiosis / Q000453:epidemiology / Q000469:parasitology*; D003458:Cryptosporidium / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D016054:DNA, Protozoan / Q000737:chemistry / Q000302:isolation & purification; D004197:Disease Reservoirs; D005243:Feces / Q000469:parasitology; D005260:Female; D005589:Foxes / Q000469:parasitology; D005838:Genotype; D016829:Giardia lamblia / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D005873:Giardiasis / Q000453:epidemiology / Q000469:parasitology / Q000662:veterinary*; D058507:Host Specificity; D006801:Humans; D008297:Male; D017720:Molecular Epidemiology; D045969:Mustelidae / Q000469:parasitology; D033661:Oocysts; D010802:Phylogeny; D017422:Sequence Analysis, DNA / Q000662:veterinary; D013030:Spain / Q000453:epidemiology; D015047:Zoonoses
chemical_list: D016054:DNA, Protozoan
keywords: Cryptosporidium; Epidemiology; Genotyping; Giardia duodenalis; Mesocarnivores; Spain; Wildlife; Zoonotic transmission