Unravelling the genetic diversity and relatedness of Echinococcus multilocularis isolates in Eurasia using the EmsB microsatellite nuclear marker

The cestode Echinococcus multilocularis is the causative agent of alveolar echinococcosis, a severe helminthic zoonotic disease distributed in the Northern Hemisphere. The lifecycle of the parasite is mainly sylvatic, involving canid and rodent hosts. The absence of genetic data from most eastern European countries is a major knowledge gap, affecting the study of associations with parasite populations in Western Europe. In this study, EmsB microsatellite genotyping of E. multilocularis was performed to describe the genetic diversity and relatedness of 785 E. multilocularis isolates from four western and nine eastern European countries, as well as from Armenia and the Asian parts of Russia and Turkey. The presence of the same E. multilocularis populations in the Benelux resulting from expansion from the historical Alpine focus can be deduced from the main profiles shared between these countries. All 33 EmsB profiles obtained from 528 samples from the nine eastern European countries belonged to the European clade, except one Asian profile form Ryazan Oblast, Russia. The expansion of E. multilocularis seems to have progressed from the historical Alpine focus through Hungary, Slovakia, the Czech Republic and southern Poland towards Latvia and Estonia. Most of the samples from Asia belong to the Asian clade, with one EmsB profile shared between Armenia and Turkey, and two between Turkey and Russia. However, two European profiles were described from two foxes in Turkey, including one harboring worms from both European and Asian clades. Three EmsB profiles from three Russian samples were associated with the Arctic clade. Two E. multilocularis profiles from rodents from Lake Baikal belonged to the Mongolian clade, described for the first time here using EmsB. Further worldwide studies on the genetic diversity of E. multilocularis using both mitochondrial sequencing and EmsB genotyping are needed to understand the distribution and expansion of the various clades.     

Using the genetics of Echinococcus multilocularis to trace the history of expansion from an endemic area

Alveolar echinococcosis, caused by the cestode Echinococcus multilocularis, is the most serious parasitic disease for humans in Europe, with a sylvatic life cycle generally between small rodents and red foxes. General expansion of the range of E. multilocularis has been observed across Europe over the last 15 years. In France, a westward spread of the known endemic areas of the parasite was described recently. For genotyping, the microsatellite EmsB was used to trace expansion in five French areas. A total of 22 EmsB profiles were identified, with five similar to those previously described in other parts of Europe. An imbalance of genetic diversity was observed between the five areas which also revealed their interconnection with the presence of common profiles, notably the two main profiles both present in all regions except one in the North. These two findings are similar to those described at the European level, highlighting transmission of the parasite by a mainland–island system. A spatio-temporal scenario of the expansion of E. multilocularis can be proposed with spread from the French historical focus in eastern France to the Lorraine, the Champagne-Ardenne and finally the North, while simultaneously another expansion has occurred from the historical focus into the West. The colonization by the parasite into the West and North areas from the historical focus was probably due to the migration of foxes several decades ago. Recent detection of the parasite in new endemic “départements” may be due to more active research rather than a recent spread of the parasite. Regarding the numerous data obtained by the different EmsB analyses, principally across Europe, centralization of all the profiles described in a public databank appears necessary in order to obtain a precise understanding of transmission of the parasite from one country to another.     

Genetic diversity of Echinococcus multilocularis on a local scale

Echinococcusmultilocularis is the causative agent of human Alveolar Echinococcosis (AE), and it is one of the most lethal zoonotic infections in the Northern Hemisphere. In France, the eastern and central regions are endemic areas; Franche-Comté, Lorraine and Auvergne are particularly contaminated. Recently, several human cases were recorded in the French Ardennes area, a region adjacent to the western border of the E. multilocularis range in France. A previous study in this focus described a prevalence of over 50% of the parasite in red foxes. The present study investigated the genetic diversity of adult worms collected from foxes in a 900km(2) area in the Ardennes. Instead of a conventional mitochondrial target (ATP6), two microsatellite targets (EmsB and NAK1) were used. A total of 140 adult worms isolated from 25 red foxes were genotyped. After hierarchical clustering analyses, the EmsB target enabled us to distinguish two main assemblages, each divided into sub-groups, yielding the differentiation of six clusters or assemblage profiles. Thirteen foxes (52% of the foxes) each harbored worms from at least two different assemblage profiles, suggesting they had become infected by several sources. Using the NAK1 target, we identified 3 alleles, two found in association with the two EmsB assemblages. With the NAK1 target, we investigated the parasite breeding system and the possible causes of genetic diversification. Only one fox harbored hybrid worms, indicative of a possible (and rare) occurrence of recombination, although multiple infections have been observed in foxes. These results confirm the usefulness of microsatellite targets for assessing genetic polymorphism in a geographically restricted local range.     

Genomic characterization of EmsB microsatellite loci in Echinococcus multilocularis

EmsB is a molecular marker applied to Echinococcus multilocularis genotyping studies. This marker has largely been used to investigate the epidemiology of the parasite in different endemic foci. The present study has lifted the veil on the genetic structure of this microsatellite. By in silico analysis on the E. multilocularis genome the microsatellite was described in about 40 copies on the chromosome 5 of the parasite. Similar structure was found in the relative parasite Echinococcus granulosus, where the microsatellite was firstly described. The present study completes the first investigations made on the EmsB microsatellite origins and confirms the reliability of this highly discriminant molecular marker.     

Echinococcus multilocularis in Svalbard, Norway: microsatellite genotyping to investigate the origin of a highly focal contamination

Echinococcus multilocularis is a threatening cestode involved in the human alveolar echinococcosis. The parasite, mainly described in temperate regions of the Northern hemisphere was described for the first time in 1999 in the High Arctic Svalbard archipelago, Norway. The origin of this contamination could be due to an anthropogenic introduction from mainland Europe by domestic dogs or with the introduction of the sibling vole, perhaps from mainland Russia (St. Petersburg area), or with roaming Arctic foxes, known as the main definitive host of the parasite in Arctic regions. The genetic diversity of E. multilocularis in Svalbard was investigated here for the first time by genotyping using EmsB microsatellite and compared to other genotyped populations in the main worldwide endemic areas. We found low polymorphism amongst the 27 metacestode isolates from sibling voles trapped in the core of the distribution area of the vole on Svalbard. E. mutilocularis Arctic populations, using the Arctic fox as the definitive host, were genetically separated from European temperate populations that use the red fox, but closely related to St. Lawrence Island samples from Alaska. The result is inconsistent with the hypothesis of an anthropogenic introduction from mainland Europe, but can be seen as consistent with the hypothesis that Arctic foxes introduced E. multilocularis to Svalbard.     

Genetic diversity of the cardiopulmonary canid nematode Angiostrongylus vasorum within and between rural and urban fox populations

Angiostrongylus vasorum is an emerging parasitic cardiopulmonary nematode of dogs, foxes, and other canids. In dogs, the infection causes respiratory and bleeding disorders along with other clinical signs collectively known as canine angiostrongylosis, while foxes represent an important wildlife reservoir. Despite the spread of A. vasorum across various countries in Europe and the Americas, little is known about the genetic diversity of A. vasorum populations at a local level in a highly endemic area. Thus, in the present study, we investigated the genetic diversity of 323 adult A. vasorum nematodes from 64 foxes living in the canton of Zurich, Switzerland. Among those, 279 worms isolated from 20 foxes were analyzed separately to investigate the genetic diversity of multiple worms within individual foxes. Part of the mitochondrial cytochrome c oxidase subunit I (mtCOI) gene was amplified and sequenced. Overall, 16 mitochondrial haplotypes were identified. The analysis of multiple worms per host revealed 12 haplotypes, with up to 5 different haplotypes in single individuals. Higher haplotype diversity (n = 10) of nematodes from foxes of urban areas than in rural areas (n = 7) was observed, with 5 shared haplotypes. Comparing our data with published GenBank sequences, five haplotypes were found to be unique within the Zurich nematode population. Interestingly, A. vasorum nematodes obtained from foxes in London and Zurich shared the same dominating haplotype. Further studies are needed to clarify if this haplotype has a different pathogenicity that may contribute to its dominance. Our findings show the importance of foxes as a reservoir for genetic parasite recombination and indicate that high fox population densities in urban areas with small and overlapping home ranges allow multiple infection events that lead to high genetic variability of A. vasorum.     

Anthelmintic Baiting of Foxes against Echinococcus multilocularis in Small Public Area,Japan

We distributed anthelmintic baits on a university campus in Japan inhabited by foxes infected with Echinococcus multilocularis to design an effective baiting protocol for small public areas. High-density baiting can reduce the risk for human exposure to the parasite to near zero. However, monthly baiting is recommended to maintain this effect.     

Vaccine-induced rabies case in a cow (Bos taurus): Molecular characterisation of vaccine strain in brain tissue

Rabies is a fatal neuropathogenic zoonosis caused by the rabies virus of the Lyssavirus genus, Rhabdoviridae family. The oral vaccination of foxes — the main reservoir of rabies in Europe — using a live attenuated rabies virus vaccine was successfully conducted in many Western European countries. In July 2015, a rabies vaccine strain was isolated from the brain tissues of a clinically suspect cow (Bos taurus) in Romania. The nucleotide analysis of both N and G gene sequences showed 100% identity between the rabid animal, the GenBank reference SAD B19 strain and five rabies vaccine batches used for the national oral vaccination campaign targeting foxes.     

Anthelmintic baiting of foxes against urban contamination with Echinococcus multilocularis

In recent years, increases in the urban fox population have been observed in many countries of the Northern Hemisphere. As a result, Echinococcus multilocularis has entered the urban environment. Because of a possible increased risk for alveolar echinococcosis, intervention strategies need to be evaluated. In Zürich, Switzerland, 50 praziquantel-containing baits per km² were distributed monthly in six 1-km² bait areas and one 6-km² bait area from April 2000 through October 2001. The proportion of E. multilocularis coproantigen–positive fox fecal samples collected remained unchanged in six control areas but decreased significantly in the 1-km² bait areas (from 38.6% to 5.5%) and in the 6-km² bait area (from 66.7% to 1.8%). E. multilocularis prevalence in the intermediate host Arvicola terrestris also decreased significantly in baited areas. This controlled baiting study shows that a pronounced reduction of E. multilocularis egg contamination is feasible in urban areas where the organism is highly endemic.The zoonotic tapeworm Echinococcus multilocularis is typically perpetuated in a wild life cycle, which includes foxes (genera Vulpes and Alopex) as definitive hosts and various rodent species as intermediate hosts (1). In addition, eggs are accidentally ingested by humans; the metacestodes enter mainly the liver and cause alveolar echinococcosis, a severe, sometimes fatal disease if left untreated (2,3).Few studies have been performed on the epidemiology of alveolar echinococcosis. Risk factors for alveolar echinococcosis may include occupational and behavioral activities. However, hunters, trappers, and persons working with fur were not at increased risk for alveolar echinococcosis in South Dakota (4). Data from Europe have indicated that farming activities increase the risk for infection (5,6). Contamination of the rural environment with E. multilocularis connected with farming activities was indirectly demonstrated by high prevalences of alveolar echinococcosis in sows kept indoors but fed with grass (7). Areas with high water vole (Arvicola terrestris) densities yielded a 10-fold higher risk for human alveolar echinococcosis compared with areas with low densities of this important intermediate host (8). In an area where the organism is highly endemic, up to 39% of A. terrestris and up to 7% of dogs with free access to rodents were infected with E. multilocularis (9), and persons who have kept dogs around dwellings were at higher risk for alveolar echinococcosis on St. Lawrence Island, Alaska (10).Red foxes (Vulpes vulpes) are likely to be the most important final host in many regions (11). In the past 2 decades, foxes have started to colonize in cities around the world (12–14), and evidence of the parasite cycle in urban areas is increasing (13,15,16). In Zürich, Switzerland, one study found that 47% of the urban fox population was infected with E. multilocularis (17).The high number of infected foxes in cities and villages, in close contact with domestic pets and humans, could increase the risk of alveolar echinococcosis (16). The disease has an incubation period of 5 to 15 years; therefore, whether the actual incidence rate of alveolar echinococcosis reflects a continuing stable and low infection risk or whether the increased infection pressure in highly populated areas will lead to a delayed increase in the incidence of alveolar echinococcosis cases in the future is unclear (3). However, ecologic changes resulted in a very high alveolar echinococcosis prevalence of 4.0% in China, which is highly endemic for the organism (18). The high prevalence of E. multilocularis in densely populated areas and the increase of foxes living in close vicinity to humans strongly suggest that evaluating possible intervention strategies is prudent.Few field studies focus on anthelmintic treatment of definitive hosts. Rausch et al. (10) demonstrated in a village that was hyperendemic for the organism (St. Lawrence Island, Alaska) that continual treatment of dogs with praziquantel reduces infection pressure of E. multilocularis, resulting in lower prevalence in locally trapped voles. In extended rural areas of Germany and Japan, praziquantel baits lowered the prevalence of E. multilocularis in foxes (19–21). These results cannot be transferred to the condition of agglomerations and urban areas, where until now no attempt has been made to evaluate an intervention strategy for foxes.The urban cycle of E. multilocularis was studied intensively in Zürich, Switzerland (16,17,22). Analyses of fox stomachs indicated that A. terrestris was the most frequently consumed intermediate host (23), and E. multilocularis is highly prevalent (mean 9.1%, maximum 20.9%) in this vole species, which lives predominantly along the city border (22). Accordingly, the prevalence of E. multilocularis in foxes was significantly higher in the urban periphery than in more central areas (17), and the infection risk for alveolar echinococcosis might therefore be concentrated mainly in delimited areas in the urban periphery (16). Since urban inhabitants frequently use the zones of highest contamination for recreational activities and their domestic cats and dogs have access to infected voles, the urban periphery may represent a risk for alveolar echinococcosis.In this controlled experimental field study, we investigated the effect of anthelmintic baiting in defined urban areas where the organism is highly endemic and tested whether E. multilocularis egg contamination was significantly reduced. We also examined whether, as an expected consequence, its prevalence in urban intermediate hosts diminished.     

A field study to control Echinococcus multilocularis-infections of the red fox (Vulpes vulpes) in an endemic focus.

Foxes harbouring E. multilocularis represent an important source for human infection with this parasite which causes alveolar echinococcosis. To minimize the risk of human infection, a control study was conducted to reduce the prevalence of E. multilocularis-infection in foxes in an focal endemic area of 5000 km2. Foxes were given access to baits containing 50 mg praziquantel. Twenty baits per km2 were distributed by airplane during 14 campaigns. The effects of control measures were monitored by parasitological examination of 9387 foxes shot before and during the control trial. A distinct reduction of the prevalence of E. multilocularis was observed for both, the initially endemic area and the low-endemic periphery. The effect was more pronounced in adult than in juvenile foxes. Under control conditions, the risk area decreased in size. However, an eradication of the parasite was not reached with the chosen strategy.     

Red fox (Vulpes vulpes) play an important role in the propagation of tick-borne pathogens

The red fox (Vulpes vulpes) is the most widespread free-living carnivore in the world. Over the years, foxes have been recognized as hosts for a number of tick-borne pathogens. However, their role as reservoirs for zoonotic tick-borne diseases is poorly understood. The aim of our study was to investigate tick-borne pathogens in the red fox population in the Czech Republic. Out of 117 red foxes, 110 (94.02%) individuals tested positive for the presence of at least one pathogen by the combined PCR and sequencing approach. Hepatozoon canis was the most frequently detected pathogen (n = 95; 81.2%), followed by Babesia vulpes (n = 75; 64.1%). Babesia canis was not detected in our study. Four (3.42%) red foxes were positive for Candidatus Neoehrlichia sp., 3 (2.56%) for Anaplasma phagocytophilum, and one red fox (0.85%) tested positive for the presence of Ehrlichia sp. DNA. Overall, DNA of spirochetes from the Borrelia burgdorferi s.l. complex was detected in 8.6% of the foxes and B. miyamotoi in 5.12% of the samples. As a carnivore found in all ecosystems of Central Europe, foxes obviously contribute to transmission of tick-borne pathogens such as A. phagocytophilum, B. burgdorferi s.l., and B. myiamotoi. In addition, foxes apparently harbour a community of pathogens, associated with this host in local ecological context, dominated by H. canis and B. vulpes (possibly also Candidatus Neoehrlichia sp.). These species have the potential to spread to the domestic dog population and should be included in the differential diagnosis of febrile diseases with hematologic abnormalities in dogs.     

Echinococcus multilocularis in Urban Coyotes, Alberta, Canada

Echinococcus multilocularis is a zoonotic parasite in wild canids. We determined its frequency in urban coyotes (Canis latrans) in Alberta, Canada. We detected E. multilocularis in 23 of 91 coyotes in this region. This parasite is a public health concern throughout the Northern Hemisphere, partly because of increased urbanization of wild canids.     

Genetic evidence of interspecies introgression of mitochondrial genomes between Trichinella spiralis and Trichinella britovi under natural conditions

Trichinellosis is a zoonotic disease caused by Trichinella muscle larvae (ML) through ingestion of raw or undercooked meat. To date, 12 taxa are recognized in this genus, of which four are circulating in Europe (Trichinella spiralis, Trichinella nativa, Trichinella britovi and Trichinella pseudospiralis). T. spiralis and T. britovi circulate in European wildlife and occur simultaneously in the same host species. The possibility of hybrid formation between T. britovi and T. spiralis has hardly been addressed and so far, results of experimental hybridisation attempts between T. britovi and T. spiralis are inconclusive. The aim of the present study was to analyse molecular polymorphisms of single T. spiralis and T. britovi ML from natural infections based on nuclear 5S rDNA intergenic spacer region (5S rDNA-ISR) and mitochondrial cytochrome c oxidase 1 (CO1) gene sequences. Six haplotypes of the 5S rDNA intergenic spacer region (5S rDNA-ISR) and 14 of the cytochrome c oxidase 1 (CO1) gene were demonstrated in 89 individual T. britovi ML from Latvia and Poland. In contrast, only two haplotypes were observed at both 5S rDNA-ISR and CO1 of 57 individual T. spiralis ML from Polish wild boar and red foxes. Moreover, this study demonstrates hybridisation in eight individual ML between T. britovi and T. spiralis under natural conditions in four Polish wild boar and two red foxes, revealed by combining 5S rDNA-ISR and CO1 sequence information of individual Trichinella ML. To our knowledge, this is the first report of interspecies hybridisation between T. spiralis and T. britovi under field conditions.     

The fox as a definitive host of Echinococcus and its role in the spread of hydatid disease

In the first part of this discussion of the significance of the fox in the epidemiology of hydatid disease in man, the author reviews the literature on vulpine echinococciasis. He points out that the evidence available from field surveys and laboratory observations suggests that the fox is not a definitive host of Echinococcus granulosus—the parasite responsible for the most common form of hydatid disease in man—but is a host of E. multilocularis, which is found in parts of Europe and Asia, in the islands of the Bering Sea and in Alaska, and is known to be the causal agent of alveolar hydatid disease.     

Characteristics of immune-competent amoebocytes non-invasively retrieved from populations of the sentinel earthworm Lumbricus rubellus (Annelida; Oligochaeta; Lumbricidae) inhabiting metal polluted field soils

Lumbricus rubellus is a cosmopolitan earthworm devoid of riboflavin-storing eleocytes; its immune competent coelomocytes are predominantly amoebocytes. Our aim was to determine whether amoebocyte cytometrics in L. rubellus are robust biomarkers for innate immunological responses to environmental pollutants. Investigations were conducted on populations inhabiting three unpolluted and five metalliferous (mainly Pb+Zn+Cd) habitats in the UK and Poland. Inter-population differences in worm mass and amoebocyte numbers did not consistently reflect soil or tissue metal concentrations. Flow cytometry indicated that autofluorescence of the amoebocytes differs between cells from the unpolluted and metal-polluted worms, and pinocytosis of neutral red by amoebocytes was lower (especially at 15 versus 60 min incubation) in worms from the polluted Poland site compared with the reference population. To conclude, amoebocyte cytometrics and functionality are potentially useful for environmental diagnostics; deployment is contingent on better understanding potential confounders.     

A foreign invader or a reclusive native? DNA bar coding reveals a distinct European lineage of the zoonotic parasite Schistosoma turkestanicum (syn. Orientobilharzia turkestanicum ())

Natural foci of Schistosoma turkestanicum (syn. Orientobilharzia turkestanicum) has been identified in the Gemenc Forest regions of Hungary utilising red deer as the definitive host. In order to identify the origins of this parasite in Europe standard DNA bar coding techniques were employed to sequence fragments of the cytochrome oxidase 1 (cox1) and the nuclear ribosomal internal transcribed region (ITS) from 10 individual adult male worms. Phylogenetic reconstruction using maximum likelihood phylogenetic reconstruction and haplotype networks of the cox1 showed all the worms to be of a distinct unique Hungarian lineage although some ITS haplotypes were shared with worms from populations in China and Iran. Molecular clock analysis suggests an early divergence event around 270,000 years before present (YBP) between all S. turkestanicum populations giving rise to the Chinese, Iranian and Hungarian lineages. However, divergence of the sequences within the Hungarian population appears to have occurred approximately 63,000 YBP suggesting a long established population of S. turkestanicum in Europe. This suggests that the Hungarian population of S. turkestanicum has been native since the Ice Age and probably established itself during the last interglacial period as red deer moved into Europe from North Africa and the Middle East. This may also indicate that the parasite may have unknown populations established in several other countries in Eastern, Central and Southern Europe.     

Identical 18S rRNA haplotypes of Hepatozoon canis in dogs and foxes in Brandenburg,Germany

Hepatozoon canis is a blood parasite of the suborder Adeleorina infecting wild and domestic canids. Transmission occurs by oral uptake of Rhipicephalus sanguineus sensu lato vector ticks infected with H. canis, but vertical transmission is also assumed to be possible. In German foxes, a high prevalence of H. canis has previously been reported despite the fact that R. sanguineus s.l. is not endemic. In the absence of knowledge about local transmission pathways, foxes should be considered to be possible reservoirs of H. canis and contribute to infection of domestic dogs. The present study aimed to determine how often foxes and dogs are infected in Brandenburg (Germany) and if identical or different H. canis 18S rRNA haplotypes are found in these host species. Hepatozoon spp. were detected by PCR in 46/1050 (4.4 %) of dog blood and 176/201 (77.6 %) of fox spleen samples from Brandenburg. Sequencing of 19 dog and 56 fox samples identified all as H. canis. For nine positive dogs, owners stated that they had never left Germany suggesting that autochthonous transmission occurs not only in foxes but also in dogs. Sequences for seven of these possible autochthonous cases were obtained and six were identical to the predominant haplotype found in the foxes. Haplotype network analysis confirmed that many dogs, including some without travel history, carried the same or very similar 18S rRNA haplotypes as the foxes suggesting that both hosts participate in the same epidemiological cycle.     

Risk of alveococcosis for humans in Poland

The paper presents data on occurrence, diagnosis and treatment of alveolar echinococcosis in humans as well as possible methods of its prevention. Studies done in 2001-2003 in Poland have shown the high prevalence of the tapeworm in red foxes in the north-east (34.5%) and south-east (39.3%) of Poland with foci of infection in some counties (up to 70% foxes infected). It makes the high potential risk for human infection in these areas.     

European echinococcosis registry: human alveolar echinococcosis,Europe, 1982-2000

Surveillance for alveolar echinococcosis in central Europe was initiated in 1998. On a voluntary basis, 559 patients were reported to the registry. Most cases originated from rural communities in regions from eastern France to western Austria; single cases were reported far away from the disease-"endemic" zone throughout central Europe. Of 210 patients, 61.4% were involved in vocational or part-time farming, gardening, forestry, or hunting. Patients were diagnosed at a mean age of 52.5 years; 78% had symptoms. Alveolar echinococcosis primarily manifested as a liver disease. Of the 559 patients, 190 (34%) were already affected by spread of the parasitic larval tissue. Of 408 (73%) patients alive in 2000, 4.9% were cured. The increasing prevalence of Echinococcus multilocularis in foxes in rural and urban areas of central Europe and the occurrence of cases outside the alveolar echinococcosis-endemic regions suggest that this disease deserves increased attention.     

Elucidating the spread of the emerging canid nematode Angiostrongylus vasorum between Palaearctic and Nearctic ecozones

Angiostrongylus vasorum is an emerging parasite that is currently distributed through Western Europe and parts of South America. An isolated population is also present in Newfoundland, Canada. This presents a risk of onward spread into North America, but its origin is unknown. To ascertain the phylogeographic relationships and genetic diversity of A. vasorum within the western Palaearctic and eastern Nearctic ecozones, a total of 143 adult and larval nematode specimens were collected from foxes (Vulpes vulpes) and dogs (Canis lupus familiaris) in Canada, Denmark, France, Germany, Ireland, the Netherlands, Portugal and the United Kingdom, and a coyote (Canis latrans) in Canada. DNA was extracted and the second internal transcribed spacer and two mitochondrial loci were amplified and sequenced. Multiple haplotypes (n = 35) based on combined mitochondrial sequences (1078 bp) of the partial cytochrome oxidase subunit I (COI), large subunit ribosomal RNA (rrnL) and the complete nicotinamide adenine dinucleotide dehydrogenase 3 (NADH3) sequences, were observed throughout the Palaearctic countries sampled; however, only a single haplotype was observed for the Canadian A. vasorum population. The likely origin of A. vasorum in Newfoundland is therefore inferred to be within the western Palaearctic. There was no evidence of genetic segregation of parasites in dogs, foxes and coyotes, supporting the hypothesis that transmission occurs between wild and domestic canids. The transmission dynamics and population structure of this nematode are further discussed.