Prevalence of intestinal nematodes of red foxes (Vulpes vulpes) in north-west Poland

Background: The red fox ( Vulpes vulpes ) is a widely distributed animal in the world. This wild carnivore is also a common host of several dangerous zoonotic parasites, primarily nematodes. Nematodes of red foxes, such as Toxocara canis and Uncinaria stenocephala , can cause numerous health problems in humans and domesticated animals. The aim of the study was to determine the parameters of occurrence of nematodes in red fox ( Vulpes vulpes ) in north-western Poland. Methods: The study was carried out in north-western Poland. The research material consisted of 620 red foxes ( Vulpes vulpes ). Parasitological sections of the foxes were taken using the sedimentation and counting technique. Results: The prevalence of infestations with nematodes was 77.3%, while the mean infection intensity was 20.1 per animal. The presence of Toxocara canis , Toxascaris leonina , Uncinaria stenocephala and Trichuris vulpis was noted. The greatest prevalence was presented by Uncinaria stenocephala (34.0%). Male and female foxes displayed a similar prevalence of nematodes. Their presence was recorded in the duodenum, jejunum, ileum and caecum of the foxes, and they were significantly more common in the jejunum than in other parts. The most commonly-observed coinfection was between Uncinaria stenocephala and Toxocara canis . Conclusions: It can be seen that nematodes are present in high numbers among foxes in north-western Poland. This phenomenon certainly contributes to an increased risk of transmission of parasites to humans and domestic animals, and this can represent a threat to health and even life.

infected foxes in urbanized areas increases the risk of contact with domestic dogs, and thus a greater risk of infection to humans (5,6,7). Infected foxes living in urban and suburban areas can pollute sandpits, parks and squares with their faeces, exposing people to contact with Toxocara canis eggs, together with other zoonotic nematode species such as Toxascaris leonina, Trichuris vulpis and nematodes from the Ancylostomatidae family (8,9,10,11,12).
Toxocariasis is considered to be the most widespread helminth zoonosis in developed countries.
Toxocariasis is responsible for the visceral and ocular larva migrans syndrome (3,13,14). In humans, as paratenic hosts, larvae of Toxocara canis do not develop into the adult stage, but migrate throughout the tissues and remain there as L3 arrested larvae for an extended period of time (15).
The number of seropositive cases in children is closely related to the economic status of countries and regions. The prevalence of anti-Toxocara canis serum antibody has been estimated at 1-6% in Japan, 2-4% in Denmark, 6% in Austria, 7% in Sweden, 14% in the USA, 22% in Iran, 81% in Nepal (16).
Serological tests have revealed the presence of antibodies in a significant percentage of the population in Poland, with the percentage of seropositive people ranging from 18.6-43% depending on the region (17,18).
The presence of Toxascaris leonina and Trichuris vulpis nematode larvae in the human body may cause the same symptoms as the presence of Toxocara canis larvae (2,19). Another potential threat to humans is posed by nematodes of the Ancylostomatidae family, such as Ancylostoma caninum and Uncinaria stenocephala (20). In humans, the invasive larvae of hookworms penetrate the skin and induce symptoms of larva migrans cutanea syndrome (21).
Previous findings, the growth in the fox population and the increasing number of diagnoses of parasitic diseases for whom foxes act as a reservoir, emphasise the need for further research into the helminth fauna of these predators. Therefore, the aim of this present study was to determine the parameters of occurrence of nematodes in red fox (Vulpes vulpes) in north-western Poland.

Methods
The study was carried out in north-western Poland. The research material consisted of 620 red foxes (Vulpes vulpes) (236 females, 384 males) obtained as part of a reduction shooting programme.
The research on foxes was carried out using security measures recommended by the WHO (1984).
The corpses were frozen at -70 °C to eliminate the invasiveness of Echinococcus multilocularis eggs.
After four days, parasitological sections of the foxes were taken using sectional methods described by Eckert et al. (22,23). During sectioning, the small intestine of foxes was isolated and divided into three sections (duodenum, jejunum, ileum) and the sex of the animals was identified based on the gonads. Nematodes were identified according to Stefański and Żarnowski (24) and preserved in 70% alcohol.
The Jaccard coexistence index (J) (25) was used to compare the coexistence of particular species of nematodes, where the feature is the presence of the species in the sample (environment). The values of the indicator will be 100% in cases when two analysed species of nematodes are always found to co-occur, regardless of their number, and 0% when these two species are never observed together.
where: Jp1p2 -co-existence index between species p1 and p2; a -the number of occurrences of species p1; b -number of occurrences of species p2; c -number of common occurrences of species p1 and p2 in the habitat.
Relative density was calculated as the arithmetic mean of the number of individuals of a particular parasite species per host examined.
The obtained results were analysed using Statistica 10.0. The significance of any differences in the occurrence of individual nematode species with regard to the sex of the host was determined using the Mann-Whitney U-test. The significance of any differences in the occurrence of nematodes in individual sections of the gastrointestinal tract was determined using the Kruskal-Wallis test.

Results
The prevalence of nematodes was 77.3%, while the mean infection intensity was 20  The most common nematode observed in males was Toxocara canis (32.0%), while Uncinaria stenocephala (37.7%) was most common in females; this difference in prevalence was not statistically significant. In both males and females, the highest mean intensity of infection was by Uncinaria stenocephala, which displayed higher intensity in females (21.7 individuals) than males (19.9 individuals). This difference, however, was not statistically significant ( Table 2). The presence of nematodes was noted in the duodenum, jejunum, ileum and caecum of the foxes.
They were not observed in the other parts of the digestive tract. In general, significantly the most prevalent nematodes were found in the jejunum than in the duodenum or the ileum (Kruskal-Wallis; H = 96.7; p = 0.000) ( Table 3).   (Table. 3).
In the duodenum, the most prevalent nematodes were Uncinaria stenocephala (20.7%) and Toxocara canis (20.7%); however, the highest mean intensity of infection was displayed by Uncinaria stenocephala (4.3 individuals per host) ( Table 3). The most prevalent and the highest mean intensity of infection in the jejunum and the ileum were associated with the occurrence of Uncinaria stenocephala (31.6%, 16.3 individuals and 14.2%, 7.0 respectively) ( Table 3).
Trichuris vulpis (11.9%) was found only in the caecum (11,9%) ( Table 3).   1 -the number of foxes in which the species of nematode was found; 2 -the number of co-occurring nematode species (prevalence); 3 -the Jaccard coexistence index between the species of nematode (%).

Discussion
In Europe, the most commonly found gastrointestinal nematode in foxes is Uncinaria stenocephala (3,26,27,28,29). Indeed, Uncinaria stenocephala was also found to be the most prevalent species in  (50). Interestingly, this parasite was not found in central Poland (51).
In other European countries, the prevalence of Trichuris vulpis among foxes varied considerably.
12.3% of tested foxes were found to be infected in Italy (27), 7.0% in Sweden (39), 5,5% in Switzerland (6), 4.8% in Ukraine (36) and 1.6% in Norway (38). However, Trichuris vulpis, which is localized in the large intestine, was very often not taken into consideration during post-mortem parasitological investigations because surveillances in foxes were often limited to the small intestine (1).
In the present study, similar levels of occurrence of nematodes were observed between males and females. Similarly, previous studies have also failed to note any difference in occurrence based on sex (2,30,52).
Unfortunately, it is difficult to find suitable comparisons for the results of the present study as This may indicate that these two species of parasites share an antagonistic relationship or are in competition with each other. The high prevalence of Toxocara canis among foxes and pets may lead to a low prevalence of Toxascaris leonina and vice versa (39). However, Jankovska et al. (2) report the co-occurrence of these two nematode species 70.6% of tested foxes in Czech Republic.
High prevalence of nematode has been observed in foxes inhabiting the region of north-west Poland.
A number of studies suggest that the occurrence of geohelminths is associated with climate conditions (temperature, air and soil humidity, mean annual rainfall) that extend the lifespan of their free-living developmental stages (1,2,53,54,55). The region of north-west Poland is situated close to the sea, has substantial forest cover and a temperate climate, as well as a total annual rainfall of 500-800 mm and a mean annual temperature of 7-9 °C, as well as significant air humidity, indicating that this area supports the development of parasites. These conditions certainly increase the threat to health, and even the life, of humans and their pets by increasing the risk of transmission of parasites; this risk is further increased by the growing numbers of foxes living nearby, which search for new places to live in areas inhabited by humans.

Conclusions
This study has shown that nematodes are present in high numbers among foxes in north-western Poland. This phenomenon certainly contributes to an increased risk of transmission of parasites to humans and domestic animals, and this can represent a threat to health and even life.
Abbreviations J: the Jaccard coexistence index Declarations Ethics approval and consent to participate

Consent for publication
Not applicable

Availability of data and materials
The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.

Competing interests
The authors declare that they have no competing interests

Authors' contributions
Study conception and design was carried out by AT and BP. Manuscript preparation was carried out by AT, BP, ATM and RP. Laboratory work was performed by AT. All authors read and approved the final manuscript.