The Biodiversity of Demodecid Mites (Acariformes: Prostigmata), Specific Parasites of Mammals with a Global Checklist and a New Finding for Demodex sciurinus

: Demodecidae are the most specialized parasitic mites of mammals; they typically inhabit the skin, but they have been found in other tissues and organs. They can cause demodecosis (a disease which is hazardous and difficult to cure ) in humans, domestic animals and livestock. They are para sites with high host and topical specificity. They have been found for most orders of mammals, and they are common in the populations of numerous host species. Therefore, they not only constitute an important subject of veterinary and medical study, but also comprise an excellent model for faunistic and parasitological analyses concerning different aspects of functioning and evolution of the host– parasite relationship. The current level or knowledge of demodecid mites is irregular and fragmentary, and numerous questions require elaboration and ordering, from the taxonomic diversity to geographic distribution and relations with hosts. Such data may be of use i.a. for the development of more efficient a nd reliable diagnostic methods, as well as understanding the etiology and pathogenesis mechanisms of demodecosis, currently a contentious issue. The present paper lists all formally -described valid species of demodecid mites, together with other functioning specific names, verified and with comments on their status. This is significant for correct species identification and demodecosis diagnostics. The list has been drawn up on the basis of data acquired in the period 1842 − 2020. It contains 122 valid species of parasite, including their hosts and geographic distribution, data on parasitism, as well as only the second record of Demodex sciurinus in Eurasian red squirrel Sciurus vulgaris in over 100 years since its initial discovery.


Introduction
The members of the Demodecidae are specialized, typically monoxenic mammal parasites, and are likely abundant within host populations. They are stationary parasites, with their whole life cycle spent on the host; however, depending on the species, its topography, seasonal dynamics and transmission mechanism, they may exhibit a variable level of infestation prevalence, which may reach up to 100% [1,2]. Typically, the presence of demodecid mites does not produce disease symptoms, even at high infestation intensity and high density on the skin [3][4][5]. However, under favorable host circumstances, the high density of these mites may be linked to the development of demodecosis (formerly demodicosis, demodicidosis). Demodecosis often has a complicated course, depending on various factors including the species constituting the etiologic factor. Its symptoms typically include the presence of various skin lesions with different topography, hair loss, eyelid margin inflammation and conjunctivitis, and changes within gum second finding of Demodex sciurinus globally, confirming the existence of this species close to one hundred years from its original discovery. Another significant objective of the revision is to organize currentlyavailable Demodecidae records, not only for faunistic purposes, but also for parasitological, veterinary and medical research. A key value of such a summary of the current state of research is that it also highlights the absence of information from numerous countries where demodecid mites, and its relationship with demodecosis, are a significant area of study: In some of these areas, no information on the distribution of demodecid mite species has been published. Our global data also constitute a significant starting point for future, more comprehensive regional analyses, as well as the development of diversity models in the context of host-parasite relationships. These more specific findings would be of great value in the development of more efficient and reliable diagnostic methods, and in improving our understanding of the etiological mechanisms and pathogenesis of demodecosis, which is currently a contentious issue.
Demodecid mites were isolated using skin digestion methods [19]. Skin fragments of 1 cm 2 were collected from several body regions, including the head (around eyes, ear pinnae, nose, lips, chin, cheeks and vertex), neck, abdomen, back, limbs, tail and genital-anal area. Skin samples were preserved in 70% ethanol and digested in 10% potassium hydroxine solution. The obtained samples were decanted and analyzed using phase-contrast microscopy; an examination of 1 cm 2 of skin was equal to the analysis of approximately 100 wet preparations. The mites were mounted in polyvinyl-lactophenol solution and photographed. The following measurements (µm) were taken as follows: Total body length equals length of gnathosoma, podosoma and opisthosoma; gnathosomal width equals width at base; and podosomal and opisthosomal width equals maximum width. The specimens were deposited in scientific collections within the framework of the Collection of Extant Invertebrates in Department of Invertebrate Zoology and Parasitology, University of Gdańsk, Poland (UGDIZP).

Literature Review-The Checklist Structure, Biogeographic and Parasitological Data Analysis
The checklist has been drawn up based on manuscripts published during the period 1842-2020 (278 items). It also contains a new record, marked in the Table 1 as the present study. Demodecidae species have been listed in systematic order, and in alphabetical order within the genera. The list includes all formally described species and other functioning specific names; all of which are verified and provided with comments on their status. Information on dates of host species, as well as the occurrence have been also included. Wherein, for cosmopolitan demodecid mite species, selected records from various range regions were given. Host records related to unidentified Demodex spp. have not been included.
The scientific names, common names, and systematics of the hosts follow Wilson and Reeder [20] and the Taxonomic Information System [21].

A New Record of Demodex sciurinus
The examined squirrel specimens were found to have D. sciurinus (Table 2, Figure 1). A total number of 13 females and 8 males were identified, as well as several specimens at nymphal stages; male and immature stages were demonstrated for the first time. All mites were found in the skin of the penis. The presence of demodecid mites was not associated with demodecosis symptoms.

Biodiversity and Geographic Distribution of Demodecidae Mites
A total of 122 demodecid mite species with verified systematic status are presently known, of which one represents Apodemodex, 106 Demodex, one Glossicodex, seven Ophthalmodex, one Pterodex, one Rhinodex, one Soricidex and four Stomatodex ( Table 1). Representatives of the Demodecidae have been recorded on all continents outside of the polar regions (Figure 2), and their presence is typically dictated by the presence of a typical host; however, no studies of this group have been conducted in numerous areas of its range, even for common mammal species. Many species of hosts have wide distribution ranges and are also considered cosmopolitan.

Demodecidae Parasitism and Relationships with Hosts
The greatest diversity of Demodecidae has, thus far, been described for bats (five genera), followed by rodents (three genera) and soricomorphs (two genera) as hosts; only Demodex representatives have been recorded in the remaining mammal orders. In turn, the species diversity in the individual host groups typically corresponds to the species diversity of the host (Figures 3 and 4). Hence the highest number of Demodecidae species have been described from the most abundant groups of mammals, e.g., 43 from rodents, 27 from bats, 17 from ungulates and 15 from carnivorans. In contrast, the Demodecidae from primates (six species) or marsupials (three species) have been especially poorly studied, which may be associated with limited access to the material. More than one species of Demodecidae has been recorded in 27 species of mammals, and the greatest number of synhospital species have been described among rodents and bats (Table 3).

Discussion
Although the Demodecidae are associated with almost all of the modern mammalian orders (Table 1), their distribution and occurrence in host populations have been poorly and unevenly studied. This paucity of information has been attributed to the difficulty in detecting asymptomatic infestation, the low number of faunistic studies and issues associated with correct species identification. This is confirmed by the high number of records for demodecid mites without species identification, listed as Demodex sp. e.g., [22][23][24][25][26][27][28][29], including valuable new records of hosts.

Demodecidae Biodiversity Analysis in the Light of Taxonomic Identification Problems
The number of demodecid mite species described thus far (122) does not seem to be very high when considering that over 5000 mammal species could act as potential hosts. This primarily stems from technical issues linked to their detection (e.g., miniature size, secretive life history with rare manifesting of their presence in the form of demodecosis), and the strict species description criteria used for the group: Representative series of specimens of both sexes are used, often with juvenile stages [9,11].
Demodecidae species associated with human, domestic and livestock mammals are an important issue in the comprehensive elaboration of this group. Although demodecid mites have been listed in handbooks and other overview papers, in lists with parasites with data on their pathogenic importance for hosts in different countries and continents, comparatively few published records exist. Therefore, verification of whether these purported Demodecidae indeed occur in these sources, or can be potentially detected in them, has proven difficult. A series of demodecosis descriptions, case studies or clinical studies exist indicating occurrence of individual demodecid mites in a given area; however, these descriptions typically lack the information needed to determine the geographic locality of the parasite or its frequency of occurrence, or whether earlier records exist. Often the identification of the demodecid mite is limited only to the genus e.g., [30][31][32][33][34][35].
In addition, a number of species with unverified status or nomen nudum exist in the parasitological or veterinary literature. As the Demodecidae are monoxenic parasites, hosts are often assigned species solely on the basis of an alleged host specificity. Recording the presence of a Demodex species in a new host species suggests a high probability that this species is new to Science; however, this requires an appropriate taxonomic analysis to be conducted and a description in accordance with the International Commission on Zoological Nomenclature (ICZN) requirements to be published [2,10]. Unfortunately, such newlydiscovered taxa have been assigned unsupported names with suitable descriptions; these have also been copied in other publications, thus becoming established in the specialist literature. For instance, the lists of the parasitofauna of the gerbil, a commonly-used laboratory animal, frequently contain a reference to Demodex merioni [36,37]: An alleged species not supported with a description and not assigned to any concrete host species, since the term gerbil is used to refer to a multitude of taxa. A similar situation concerned D. cornei from the domestic dog. The name had functioned for many years to describe an alleged species referred to as "short form" from the dog epidermis. Although morphological and morphometric research [8,38] has confirmed the existence of such species, the lack of a formal, unambiguous description meant that it was impossible to verify records from different parts of the world, as these were mostly based on the criterion of length, possibly confirmed with topical distinctiveness. Although, eventually, the specific status has been explained and confirmed with an appropriate species description [2], this does not provide any possibility to verify earlier records not supported with morphological characteristics. The groundlessness of using size as a criterion to identify species within the same host has been further confirmed by the discovery of another "short" canine demodecid species, D. cyonis. Furthermore, literature data including the nomina nuda D. araneae and D. bonaparti has been published without an appropriate description [39], despite being correctly distinguished by the author: A specialist on Demodecidae research. Likewise, the description of D. myotidis, D. sciurei, D. sylvilagi or D. transitionalis were included in an unpublished dissertation, a procedure that does not meet the ICZN criterion on the publication of species descriptions ( Table 1). At present, numerous directions in Demodecidae research have employed molecular methods; however, their outcomes are not reliable given the lack of correlation between morphological taxonomy and molecular divergence. An example here would be D. "felis", which was recorded for the domestic cat in a study solely based on molecular analyses [40]. The authors, who assigned it a temporary name, giving the impression of a species name, stipulate that it is only a working name proposal, intended for the purpose of distinguishing the alleged species, which, according to those authors, differs from other feline demodecid mites e.g., [41][42][43][44]. Such a study where cladistics is based solely on molecular data without confirming that the inferred genetic distance is a reliable evidence at the infraspecific level, cannot provide a sufficient basis to assign a species name for an identified demodicid mite; this is especially true as the results, in this case, have not been supported with any other evidence, including morphology, and it cannot be said which taxa they concern. However, despite this irregularity, the species already functions in the veterinary literature at the specific name level.
Another issue in the study of Demodecidae distribution concerns the existence of uncertainties with regards to correct identification. Individual species can differ with regard to sets of characters and small morphological elements which may be only several micrometers in size, sometimes less than 1 µm; such minute variation requires the use of suitable preparation methods, phase contrast techniques and immersion microscopy, as well as experience in such taxonomic analyses. At the same time, some studies use alleged host specificity or, sometimes, size as the basis for identification; however, the study methods described in the works do not leave any doubt that a correct identification had not been possible. As a host may be associated with different specific Demodecidae species with similar sizes and proportions, any application of the host specificity criterion in species identification is not only insufficient, but also groundless. An accidental transfer onto atypical hosts cannot be excluded, which may happen under favorable conditions, even in the case of highly-specialized parasites.

State and Perspectives for the Study on Geographic Distribution
In view of presented data, the highest number of species (confirmed records) have been recorded in Poland (51 species), the USA (23), Czech Republic (18) and Great Britain (18). Naturally, this does not stem from any special preferences of the Demodecidae for the hosts occurring in those countries, but it is consequence of sampling bias. A clear contrast can be seen between Demodecidae records obtained from wild, domestic and livestock mammals. Detections in wild animals are rare, because demodecid mites rarely manifest their presence in the form of demodecosis.
It is also possible that the occurrence of the known Demodecidae species is considerably wider than that indicated by the published data, and likely coincides with the ranges of their hosts. This has been confirmed by the latest records of D. chiropteralis, D. melesinus and, the present record of, D. sciurinus: Species formerly known only from individual records from England, and a single observation recorded a hundred years previously from distant Poland. The currently identified individuals of D. sciurinus exhibited traits complying with the description and figures published by Hirst [45], despite the description deviating from the modern standards assumed for the Demodecidae taxonomy. Therefore, a redescription will definitely be necessary in the future; this would include an initial description of the juvenile stages, which will be possible after collecting a wider range of material from a greater number of hosts. Current intensive research conducted within the area of Poland has further confirmed the presence of almost all species formerly described from the Czech Republic, the Netherlands or the USA, provided that typical hosts are to be found. It should be added that many species exhibit very high infestation prevalence in the host populations, reaching up to 100% e.g., [1,4,12,18,19].
It is currently important to organize the diverse body of data concerning the occurrence of species of medical and veterinary significance, i.e., to verify and correlate data on the occurrence of demodecosis in various host species with information on the occurrence of the agent species, as confirmed by taxonomic identification.

Host-Parasite Relationships
The diversity of Demodecidae in individual host groups is typically convergent with the species richness of the host group (Figures 2 and 3). Moreover, the study of this group of species is also related to the incidence rate of demodecosis and the practical importance of hosts, thus the number of species described from carnivores is relatively higher (with regards to the biodiversity of this group) than for other mammals. Among the 17 demodecid mite species described for carnivorous mammals, six originate from domestic dogs and cats. In addition, the availability of material for study, and the technical issues related to detection, are other significant factors. The detection of asymptomatic infestation is labor-intensive, with an efficiency that is inversely proportionate to the host size. It is therefore unsurprising that the highest number of species have been described from small mammals. In contrast, the Demodecidae from primates (six species), or marsupials (three species) have been especially poorly studied, which may be associated with the limited access to the material, with only individual specimens of selected species obtained from zoological gardens being tested, and the fact that certain areas of the world, such as Asia and Australia, are absent from the body of data (Table 1). Therefore, it is possible that the number of existing species is considerably higher than presented herein.
The Demodecidae exhibit high specificity towards hosts (monoxeny). Only five species are considered oligoxenic, i.e., recorded from more than one host; however, all host species were closely related. Of this group, D. apodemi and S. corneti have been described according to insufficient criteria, based on the current state of knowledge, and hence require redescription. In turn, D. sabani and D. kutzeri also require taxonomic revision, as they may constitute aggregate species, with their taxa being difficult to distinguish according to morphological criteria; such revision should include additional criteria such as ontogeny, molecular characteristics and parasitological testing. Furthermore, the host status is not always clear, e.g., D. kutzeri has been recorded from various deer species, with ambiguous species status (Cervus canadensis or C. elaphus canadensis).

Conclusions
The Demodecidae have high veterinary and medical importance, and these aspects have directed the majority of research into the family. Despite the fact that such research stretches back to the 19th Century, appropriate zoological studies (taxonomy, fauna) are scarce and limited to species descriptions, typically based on singular records from one locality and a single host. More detailed biodiversity studies, based on the analysis of the occurrence of the parasite in different populations of individual host species, or the co-occurrence of different Demodecidae in one host species, have been published only in Poland. Even those studies have been limited by the availability of the host or the need to conduct comprehensive studies of its entire parasitofauna.
Unfortunately, no comparable data on Demodecidae are available from other regions; in addition, the group has never been included in holistic parasitofauna studies, or even studies of parasitic arthropods. The greatest obstacles to such studies are associated with parasite detection, particularly since most infestations are asymptomatic, and problems with species identification. This stands in contrast with the multitude of global medical and veterinary reports on demodecosis, or routine testing for this disease conducted at diagnostic laboratories.
There is a clear need to integrate zoological and parasitological research with medical and veterinary studies, or to perform further interdisciplinary studies. Only, such, broader approaches will provide a greater understanding of the key issues concerning Demodecidae parasitism, allow the development of efficient diagnostic methods and deepen our understanding of the causes and mechanisms of demodecosis.

Conflicts of Interest:
The authors declare no conflict of interest.