Morphological and ontogenetic characteristics of Miridex putorii (Acariformes: Demodecidae), a new genus and species of skin mite specific to the European polecat Mustelaputorius

Among carnivorans, mites of the family Demodecidae are mainly represented by the eight species of the genus Demodex known to cause demodecosis in domestic dog Canis lupus familiaris Linnaeus, 1758 and domestic cat Felis catus Linnaeus, 1758. However, nine other Demodex species from wild carnivorans are also known; in addition they are only known from few records. Previously unknown demodecid mites have been isolated from European polecats, Mustela putorius Linnaeus, 1758, originating from Germany. The specimens are characterized of by an aedeagus with a posterior end located between the opisthosoma and podosoma and an anterior end in the gnathosoma area, with a genital opening in the epistome area; aedeagus length corresponds to 53% (45–59%) of male body length. The mites were isolated from the head skin in 16 of 21 polecats examined (76.2%), mainly in the mystacial vibrissae area (84.8%) and less often in adjacent areas. However, the mite infestation did not appear to cause skin abnormality. Based on the morphological analysis of the adult mites and their morphological ontogenesis, including significant characteristics in demodecid taxonomy, the mite specimens have been classified as representatives of a new species and genus, described as Miridex putorii gen. nov., sp. nov.


Introduction
The Demodecidae (Acariformes: Prostigmata) of carnivoran mammals have been relatively poorly studied. Most information regarding mites of the genus Demodex concern domestic animals, dogs and cats, each harboring four species; however, some of these have been discovered only in recent years Rolbiecki, 2018, 2020;Morita et al., 2018). The majority of known information concerns the clinical signs of the parasitosis in the hosts, canine demodecosis and feline demodecosis, respectively.
The present study describes the discovery of Demodecidae mites from the European polecat M. putorius Linnaeus, 1758 originating from Germany. The new taxon exhibits characteristics that are distinct not only from all known demodecid mite species, but also from the eight genera known thus far; therefore, it has been classified in a separate, new genus as Miridex putorii gen nov., sp. nov.

Materials and methods
In the course of a survey of the parasite fauna of the European polecat in Germany (Kretschmar, 2016) The demodecid mites were recovered by digestion of host skin fragments (Izdebska, 2004). The method was modified to suit the examined host. Skin fragments of 1 cm 2 were taken from several head regions, including the area around the eyes, nose, vibrissae, lips, chin, cheeks, and vertex. The samples, which had been preserved in 70% ethanol, were subjected to digestion in 10% potassium hydroxide solution. The resulting solution was decanted and examined under phase-contrast microscopy (Nikon Eclipse 50i); a 1 cm 2 of skin sample yielded approximately 100 wet preparations. Any mites were placed in polyvinyl-lactophenol solution, measured (micrometers), photographed, and graphically documented. The following measurements were made: total body length = length of gnathosoma, podosoma and opisthosoma; gnathosomal width = width at base; podosomal and opisthosomal width = maximum width.
The specimen depository are cited using the abbreviation UGDIZP, University of Gdańsk, Department of Invertebrate Zoology and Parasitology, Gdańsk, Poland (Zhang, 2018).
The species description adopted the nomenclature commonly used for the Demodecidae (Nutting, 1976). It was completed with the nomenclature proposed by Bochkov (2008) for the superfamily Cheyletoidea (Acariformes: Prostigmata) and by . The scientific and common names of the hosts follow Wilson and Reeder (2005)
In development, adult stages are located in the middle region of deutonymph (gnathosoma of adults at level of I-II pair of deutonymph legs); furthermore, two deutonymph morphotypes were observed, one with a shorter (Fig. 3C) and one with a longer (Fig. 3D) opisthosoma: the former probably male and latter female.

Etymology
As is common practice regarding the creation of generic names in the Demodecidae, a name was set combining mirus (peculiar, unusual), referring to the peculiar feature (regarding location of the male genital opening and the length aedeagus) with the word dex -'a worm'. In turn, the specific epithet putorii was added to refer to the host species name.

Infestation and location in the host
Miridex putorii sp. nov. was found in 16 of 21 European polecats examined (76.2%); in total, 158 specimens (56 males, 51 females, three larvae, six protonymphs, 42 deutonymphs) and 11 eggs were found. Mites were found in the skin of the mystacial vibrissal area (134 specimens, 84.8%) and in adjacent areas (24 specimens, 15.2%); eggs were found only in the mystacial vibrissal area. The infestation did not cause skin lesions in the mite positive polecats.

Differential diagnosis
The feature distinguishing M. putorii sp. nov. from other known Demodecidae is the genital opening of the male, which is located on the dorsal side of the gnathosoma (on the epistome); in other genera, it is always located on the dorsal side of the podosoma. Another specific feature is the presence of the massive claws, strongly bifurcated at the end; in Demodex, they are clearly less bifurcated, with different shapes being observed in other genera. Miridex putorii sp. nov. most closely resembles some Demodex species in the habit, shape and arrangement of its epimeral plates; in these species, the epimeral plates come into contact in the midline of the ventral part of the body. However, the palps and legs of Demodex are strongly shortened, being composed of overlapping segments which are sometimes difficult to distinguish. While M. putorii sp. nov. resembles Glossicodex, insofar that its leg segments are clearly separated, in the case of the latter, the leg segments are elongated and end with hooked, non-bifurcated claws. In addition, a relatively large, conical subgnathosomal setae can be found on both sides of the pharyngeal bulb on M. putorii sp. nov. These are pronounced in the larvae in the form of long, vimineous setae. In other Demodecidae, the subgnathosomal setae are typically fine, and often difficult to observe.

Discussion
Our taxonomic analysis, including data from the morphological ontogenesis (including juvenile stages) supports that the specimens identified from the European polecat should be classified into a new (separate) genus. This genus appears to exhibit certain archaic and specific features which reflect unique adaptations and whose significance is difficult to interpret.
The representatives of individual Demodecidae genera can often be distinguished based on the structure of the legs (i.e. the shape of the claws and the shape and arrangement of the epimeral plates), as well as the form and location of the structures related to the gnathosoma (Fain, 1959;Lukoschus and Nutting, 1979;Bukva, 1996;). However, it should be added that these features are not homogeneous within the genus Demodex, and they should be subject to revision.
In hitherto described Demodecidae species, the female's vulva is located on the ventral side of the body; however, it is observed at different levels, ranging from the border between podosoma and opisthosoma (partially within IV. pair of epimeral plates) or within the opisthosoma, directly under the edge of the last pair of epimeral plates, or even lower. In contrast, the genital opening of the male is always located on the dorsal side of the podosoma. Its precise position depends on the location of the aedeagus (whose length corresponds to approx. 10-15% length of the male body); in M. putorii sp. nov. the aedeagus is very long, corresponding to 45-59% of male's body length, and it reaches from the margin of podosoma and opisthosoma to the midgnathosoma, with an opening within the epistome. Among the known demodecid mites, a relatively long aedeagus is characteristic of Glossicodex musculi , however, it covers a space corresponding to three epimeral plates (corresponding to 20-25% of male body length) with a genital opening located on the podosoma . Therefore, the aedeagus location observed for M. putorii sp. nov. is a unique feature within the Demodecidae.
On the other hand, the clear separation of palps and leg segments, claw-like spines on the palps, or larger, more distinct setae, particularly in juvenile stages, may be of a primary nature. Most importantly, the long, vimineous subgnathosomal setae on the gnathosoma of juvenile stages form a clear link to other Cheyletoidea, which are phylogenetically more distinct from Demodecidae (Bochkov, 2008 Izdebska, Rolbiecki et Fryderyk, 2022 was found solely in the region of the lips, nose and chin, whereas D. cati Megnin, 1877 prefers the nasal region, areas of the eyes and auricles (Izdebska and Rolbiecki, 2013;Izdebska et al., , 2022.
The lack of data regarding the Demodecidae in the large, biodiverse and widely distributed mustelid group can probably be attributed to the asymptomatic nature of the infestation. Demodecosis is usually known from humans and domestic animals, but constitutes a rare observation in wild animals, even at high infestation levels: no skin abnormalities were observed in the case of D. melesinus in the European badger or D. lutrae in the Eurasian otter, despite very high mite densities Rolbiecki and Izdebska, 2014;.
More surprising is, however, the scarcity of data regarding demodecid mites from mustelids farmed for fur or kept as pets, such as the species of the genus Mustela. While a report of "Demodex bonapartei" Nutting, 1950 from M. erminea cicognanii Bonaparte, 1838 has been given in international literature (Nutting, 1950(Nutting, , 1964, no species description has yet been published: the report is included in an unpublished doctoral thesis. A report also exists on demodecosis associated with local skin lesions (alopecia, pruritus) in ferrets M. putorius furo Linnaeus, 1758, caused by the adult and juvenile forms of unidentified Demodex (Noli et al., 1996). These mites were found in skin scrapings and exudate from the ear of two ferrets; they were very small, with short, blunt opisthosoma and were reminiscent of D. criceti Nutting et Rauch, 1958 from Mesocricetus auratus (Waterhouse, 1839) or to the unnamed cat mite (Noli et al., 1996). Furthermore, while information exists on the isolation of an unknown Demodex from the long-tailed weasel M. frenata Lichtenstein, 1831 and M. putorius putorius Linnaeus, 1758, no detailed data have been provided (Noli et al., 1996).
Many mammals have been found to demonstrate synhospital, cooccurring Demodecidae species, these being specific for the given host species, and occupying different locations . It is therefore likely that further representatives of this mite family will be found in the European polecat and other members of the Mustelidae.

Declaration of competing interest
Authors have no conflict of interest to declare.