Eucoleus contortus (Nematoda: Capillariidae), a parasite of Cairina moschata domestica (Anseriformes: Anatidae) on Marajó Island, Pará State, in Brazilian Amazon

The family Capillariidae is one of the most important in the superfamily Trichinelloidea, with 27 genera and more than 300 species parasitizing vertebrates. This study considers the morphology, morphometry and prevalence of Eucoleus contortus parasitizing the Muscovy duck Cairina moschata domestica esophagus from Marajó Island, in State of Pará, Brazil. Morphologically the nematodes had a filiform body, with transversely striated cuticle, long esophagus, divided into two parts, one muscular and another consisting of stichocytes, spicule weakly sclerotized, spiny sheath and pseudobursa present in males. Females had a pre-equatorial vulva, barrel-shaped eggs and were bioperculated. In the northern region of Brazil, the Muscovy duck is an abundant bird, and one of the items in the food supply for human communities. The occurrence of E. contortus adds data to the biodiversity of parasites described in Anseriform birds of the Brazilian Amazon, especially those used as source of protein by human communities of Marajó.


Introduction
The family Capillariidae Neveu-Lemaire, 1936 is one of the most important in the superfamily Trichinelloidea Ward, 1907Ward, (1879, with more than 300 known species parasitizing all vertebrate classes around the world (ANDERSON, 2000;GIBSON et al., 2014). The classification of capillariids is one of the most complex and unsatisfactory among nematodes, due to the scarcity of good morphological characteristics (SPRATT, 2006). Moreover, there are still problems with the correct identification of these parasites, and at present the classification system is based mainly on the morphology of males (MORAVEC & JUSTINE, 2010). This fact often results in descriptions of species lacking elucidative morphological data (FREITAS & ALMEIDA, 1935;MORAVEC, 1982;STAPF et al., 2013).
In the Baylis (1928) conception, the morphological classification of capillariid members included all of them in the genus Capillaria sensus Zeder, 1800, and is not currently accepted by most helminthologists, but remains in use in the medical literature (MORAVEC, 2001). Because of this classification today, the genus Capillaria has a large number of synonymies resulting from different attempts to reorganize the genus taxonomically (BUTTERWORTH & BEVERLEY-BURTON, 1980).
Cairina moschata domestica Linnaeus, 1758 (Muscovy ducks) is an important source of food for several human populations, since they provide meat and eggs (MATTOS et al., 2008). In the northern region of Brazil, the Muscovy duck is an abundant bird, composing one of the items in the food supply for human communities. As part of an ongoing study of the helminths of birds on Marajó Island, Brazil, samples of ducks were collected and necropsied. The objective of this study is to characterize the morphology, morphometry and prevalence of nematode Capillariidae parasites in Muscovy duck collected on Marajó Island, Brazilian Amazon.

Materials and Methods
The study used Nematoda Capillariidae obtained from 19 males and 11 females of C. moschata domestica aged 4-8 months and acquired from rural properties of the municipality of Soure (00° 43' 00" S; 48° 31' 24" W), in Marajó Island, Brazil. The Muscovy ducks were components of small extensively reared herds with free access to the environment, for the purpose of providing meat and eggs for families or for sale at local markets. These birds were slaughtered stunning with a club, cutting the blood vessels of the neck, exsanguination on the farm and only the organs of the digestive tract were transported to the Laboratório de Histologia e Embriologia Animal, Universidade Federal Rural da Amazônia, Campus Belém (Brazil). In the laboratory, the organs were separated and placed in Petri dishes with saline solution NaCl 0.9% and examined using a stereomicroscope. The recovered nematodes were fixed in a solution of AFA (93 parts 70% ethyl alcohol, 5 parts formaldehyde, and 2 parts glacial acetic acid) and processed using light microscopy and scanning electron microscopy according to method described by Pinheiro et al. (2018). A total of 10 male specimens, 10 female specimens and 50 eggs were used for the morphometric analysis of the nematodes. Measurements are given in micrometers unless otherwise noted and are presented as the range (minimum and maximum values) followed by the mean in parentheses. Taxonomic classification of nematodes was in accordance with Vicente et al. (1995) and Gibbons (2010).  Creplin, 1839(Gagarin, 1951 (Based on light microscopy and scanning electron microscopy examination: Figures 1-3) Medium-sized nematodes in relation to their congeners, filiform, with finely transversely striated cuticle. Cephalic region in button format (Figures 1a, 2a). Oral aperture circular. Muscular esophagus short, narrow. Nerve ring circulating the muscular esophagus in its initial portion. Stichosome consisting of single row of about 33 elongate stichocytes with distinct transverse annulae (mean of the three initial, middle and final stichocytes); nuclei of stichocytes large and fragmented (Figure 1b). Two wing-like pseudocoelomatic glandular cells present at esophagus-intestinal junction. Two bacillary lateral bands along the body, more numerous in females.

Discussion
The nematodes found inserted in the mucosa of the esophagus of Muscovy ducks raised extensively in the municipality of Soure, Marajó Island, State of Pará (Brazil), have characteristics similar to those of the Capillariidae family. Gibbons (2010) (MORAVEC, 1982;MORAVEC et al., 1987;STAPF et al., 2013).
The genus Eucoleus is composed of species that parasitize the respiratory tract, mucosa of the esophagus, buccal cavity and stomach of birds and mammals (MORAVEC, 1982;VICENTE et al., 1995;GIBBONS, 2010). Although Anderson (2000) and Anderson et al. (2009), employ Eucoleus as a synonym of Capillaria, Moravec (1982), Vicente et al. (1995) and Gibbons (2010) accept Eucoleus as a valid genus, with morphologically presenting males with thin spicule, moderately sclerotized with a long spinal sheath covered with cuticular spines, caudal lateral alae absent, two small lobes rounded laterally towards the posterior supporting a pseudobursa, and females with vulvae without an appendix.
The nematodes of the present study presented morphological and morphometric characteristics compatible with E. contortus. Creplin (1839) proposed E. contortus as a parasite of the oral cavity and esophagus of different birds (Anseriformes, Charadriiformes, Galliformes Falconiformes and Passeriformes) in Germany, but later, different authors re-described or added a new geographic distribution for the species, including Diesing (1851) in Falconiformes, Passeriformes and Charadriiformes from Austria; Eberth (1863) in Passeriformes from Germany; Linstow (1877) in Passeriformes, Charadriiformes and Anseriformes from Berlin; Railliet & Lucet (1889) in Anseriformes from Paris; Cram (1936) in Galliformes, Charadriiformes and Anseriformes from United States; Chabaud (1952) in Charadriiformes from Paris and Mettrick (1959) in Galliformes and Anseriformes from England.
Muscovy ducks are extensively reared in most part of Brazil, and on the Marajó Island they are of great importance to the human populations as a source of food, although there are few data on their parasitic fauna. In Brazil, the first record of bird parasitic nematodes was made by Travassos (1915) in Anseriformes, Charadriiformes, Passeriformes, and Falconiformes, followed by Freitas & Almeida (1935) in Passeriformes, Anseriformes, Charadriiformes, Falconiformes, and Galliformes; Freitas et al. (1959) in Galliformes, Piciformes and Psittaciformes, Mattos et al. (2008) in Anseriformes and Stapf et al. (2013) in Anseriformes. Although Brazil is one of the main refuges for resident and visiting birds, in discussions of work related to the parasites of these animals, only Vicente et al. (1995) has reviewed nematodes in birds in Brazil.
The use of scanning electron microscopy, although very common for nematodes, has rarely been applied for members of the Capillariidae family (MORAVEC & BARTON, 2018), especially because of the difficulty in processing the samples. In this study using SEM, we observed details of the cephalic region in button shape, in female vulva with unraised lip, in males tail with pseudobursa, supported by two lobes, each lobe having a papilla, spinal sheath armed with spines, besides the difference in quantity of bacillary bands between males and females. Morphometric comparisons between Eucoleus contortus and its other re-descriptions, in addition to the comparison with other species found in Brazil, are presented in Table 1.
Eggs of E. contortus morphologically have a barrel shape, with polar plugs and a rough surface; morphometrically the eggs presented similarity in size with that already described in the literature for the species (see Table 1). Campbell & Little (1991) states in his study that eggs of E. boehmi are characterized by having a barrel shape and polar plugs with small morphological differences. Macchioni et al. (2013), when analyzing dog feces in Italy with a prevalence of 7.4% of capillary parasites (E. aerophilus and E. boehmi) affirm that the use of molecular biology should be fundamental for the specific identification of family Capillariidae.
In this study, 76.6% of the analyzed ducks were parasitized by E. contortus in the esophageal mucosa. Different authors report parasitism and a high prevalence of E. contortus parasitizing the esophagus in different birds: Betlejewska et al. (2002) reported by 52.3% of parasitism in Anas platyrhynchos in the Northwest of Poland. Mattos et al. (2008) described the occurrence of E. carinae with prevalence of 6.6% in Muscovy ducks raised extensively in the State of Rio de Janeiro. Stapf et al. (2013) reported 24% of prevalence in A. platyrhynchos and 40% of prevalence in
In the Marajó Island, the local human population commonly uses Muscovy ducks for food and commerce. Most of these birds live in an open environment, using this space as shelter and to obtain food, besides being raised together with other birds, and domestic and wild animals. Endoparasite infections are almost inevitable in an extensive system due to the prolonged survival of eggs in the environment, especially when there is high humidity, which allows the greater survival of immature forms of helminths and increases the number of infectious stages in the soil, when capillary eggs can survive up to 11 months viable in the environment (YADAV & TANDON, 1991;PERMIN et al., 2002;CARDOZO & YAMAMURA, 2004;SOBRAL et al., 2010). Ruff (1999) and Vita et al. (2014) observed that for birds that are raised free, having access to other birds and domestic or wild animals in places with poor hygiene, in addition to direct contact with the ground, this is the ideal environment for the proliferation of parasitic diseases.
The high prevalence of nematodes in Muscovy duck from the Marajó Island may be related to the interaction of these birds with the soil, which is essential for the maintenance of the life cycle of many parasites, such as E. contortus, where the birds ingest the intermediate host, possibly earthworms, besides the viable eggs in the environment (CARDOZO & YAMAMURA, 2004).

Conclusions
The Muscovy duck is an important source of animal protein for the human population of Marajó and the knowledge of its nematofauna is important information to understand the pathogens that can affect poultry and that reduce poultry production in the country, so that measures can be taken for the purpose of avoiding possible zoonoses.