Taxonomic revision and dissolution of the genus Monachoides (Gastropoda, Stylommatophora)

Monachoides is a genus of medium‐sized European land snails in the family Hygromiidae. Systematics of this family was until recently based on traits of the genital system that were shown to be uninformative, resulting in revisions of classifications at the genus and family levels. Up to six species were recognized in Monachoides lately, but the inclusion of three of these species was questionable. Our phylogenetic analysis and morphological examinations show that the genus includes species previously classified in two different Hygromiidae subfamilies. After transferring three Balkan species to the unrelated genus Xerocampylaea, Monachoides emerged as a group that diversified relatively northerly, at the boundary between the Balkans and Central Europe. One species, Monachoides bacescui, is synonymized here and a previously overlooked one is redescribed, leaving only three valid species in Monachoides as currently understood: M. incarnatus, M. vicinus and M. welebitanus. These can be distinguished from other hygromiid species by their distinctive microsculpture of the shell surface, which also allows for identification of juveniles or fragmented fossil material. However, the clade comprising the three above species was found to also include the two species currently classified in Perforatella. As a result, the two genera are synonymized, with Perforatella having priority.

studied the systematics of the genus Monachoides Gude & Woodward, 1921, whose type species is not as well known to the public as H. pomatia, but is equally widely distributed and highly abundant in Central Europe.
Monachoides belongs to Hygromiidae Tryon, 1866, a diverse and genus-rich family of land snails, widely distributed across the Palaearctic region, and even reaching sub-Saharan Africa (Neiber et al., 2017).The systematics of Hygromiidae from the division into subfamilies down to the assignment of species to genera relied for a long time on the structure of the reproductive system (Forcart, 1946;Hesse, 1921Hesse, , 1931;;Nordsieck, 1987Nordsieck, , 1993;;Schileyko, 1970Schileyko, , 1972aSchileyko, , 1972bSchileyko, , 1978Schileyko, , 1991Schileyko, , 2006;;von Ihering, 1892;Wagner, 1915;Watson, 1919).The number and arrangement of dart sacs were among the most important characters; a single sac with a developed dart was one of the defining characters for the group into which Monachoides incarnatus (Müller, 1774) was classified.However, the morphology-based classifications were always controversial, and in many cases were eventually overturned by molecular phylogenetic analyses (Neiber et al., 2017;Razkin et al., 2015).Several characters traditionally used for systematics, including the number of dart sacs, were found to have evolved several times independently (Neiber et al., 2018;Neiber & Hausdorf, 2017).
At present, six species are considered members of Monachoides (MolluscaBase, 2023a;Welter-Schultes, 2012).Monachoides incarnatus (Central Europe, Balkans; Figure 1f-h) and M. vicinus (Eastern and Central Europe, Carpathians; Figure 1b,c) form the undoubted core of the genus.Clearly close to M. incarnatus is also Monachoides bacescui Grossu, 1979, described from western Romania.Besides these, there are three mutually similar species from the western Balkans (former Yugoslavia) whose genus-level placement is more problematic.Monachoides fallax is the most widely distributed of them (reported from Bosnia to North Macedonia, but the range is uncertain; Dhora & Welter-Schultes, 1996;Fehér & Erőss, 2009;Welter-Schultes, 2012), while the others have small ranges in the surroundings of their respective type localities: Monachoides taraensis de Winter & Maassen, 1992 is known from a part of the Tara river valley in Montenegro (Figure 1i), Monachoides kosovoensis de Winter & Maassen, 1992 was described from the eastern side of the Čakor Pass on the border of Montenegro and Kosovo.As already mentioned, Nordsieck (1993) proposed to transfer M. fallax to Urticicola, and de Winter & Maassen (1992) admitted that the placement of their two new species in Monachoides is provisional and is actually questioned by the presence of a small accessory sac in M. kosovoensis.De Winter and Maassen (1992) also noted a great conchological similarity of M. taraensis and M. kosovoensis to taxa that were recently shown to belong to the genus Xerocampylaea Kobelt, 1871 in the tribe Urticicolini Neiber, Razkin & Hausdorf, 2017 (Semifruticicola serbica Wagner, 1914 andHelix erjaveci Brusina, 1870;Neiber et al., 2017;Proćków et al., 2019) as well as to the type species of that genus, Xerocampylaea zelebori (Pfeiffer, 1853).
On the intraspecific level, two subspecies of M. incarnatus, aside from the nominotypical subspecies, were considered valid in recent decades (Falkner, 1990;MolluscaBase, 2023b;Štamol, 2010).One is currently called Monachoides incarnatus armatus (Stabile, 1859) and was described from near Lugano on the Swiss-Italian frontier and is characterized by a tooth on the lower apertural lip.The second subspecies is Monachoides incarnatus welebitanus (Pfeiffer, 1847) for which no type locality was indicated, and the original description mentions only a more solid [than typical M. incarnatus] and opaque shell with a straight lower apertural margin.Falkner (1990) newly characterized the taxon, stating among others that the surface microstructure of the shell is about twice as fine and dense as in M. incarnatus samples from north of the Alps.This subspecies should be distributed in the northwestern part of former Yugoslavia (welebitanus refers to the Velebit Mountains in western Croatia).Its status needs reassessment, also because of a passing note of Nordsieck (2017: inside of the back cover) on the syntopic presence of M. incarnatus and an unidentified, possibly undescribed Monachoides species (Figure 1d,e) in the Velebit Mountains.
We revise here the delimitation of the genus Monachoides and the species-level taxonomy of the taxa belonging to the genus, that is, M. incarnatus and its closest relatives as revealed by molecular phylogenetic analyses.While the current delimitation of Monachoides based on the presence of a single dart sac is rejected, we show that morphological characters, which were previously given less weight, support the new genus-level taxonomy or are informative for species identification.

| Note on sampling
We include here samples of five of the six species presently classified in Monachoides.We failed to obtain a sample of M. kosovoensis, although we included an unidentified juvenile collected near its type locality.Monachoides fallax is represented here only by samples from the type locality (Mt.Trebević near Sarajevo).On the intraspecific level, we did not sample marginal, postglacially colonized areas; we focused on the areas potentially relevant for the diversification of the genus.However, samples from the Romanian parts of the Carpathian Arc are largely missing due to logistic reasons.
The original data were supplemented by sequences from Neiber et al. (2017), who laid the foundations of the modern, phylogeny-based classification of Hygromiidae, and from Proćków et al. (2019), who focused on Xerocampylaea.We included all representatives of the tribes Perforatellini and Urticicolini with available sequences, which allowed for testing the hypothesis that M. fallax, M. taraensis (and by extension also M. kosovoensis) actually belong to the latter clade, as would hold if they are related to Xerocampylaea.

| Laboratory procedures
Total DNA was extracted using a spin column-based extraction kit for animal tissues (Tissue Genomic DNA Mini Kit, Geneaid) from ethanol-preserved pieces of the foot.Three loci were targeted: two partial mitochondrial genes (cytochrome c oxidase subunit I and 16S rRNA; cox1 and rrnL hereafter) and the nuclear internal transcribed spacer 2 (ITS2) with the flanking parts of the genes for 28S and 5.8S rRNA (hereafter 28S and 5.8S respectively).The nuclear marker was only sequenced for selected samples representing the major mitochondrial lineages.
The primer pairs LCO1490 + HCO2198 (producing 655 bp long amplicon; Folmer et al., 1994) and 16Scs1 + 16Scs2 (~850-860 bp; Chiba, 1999) were used for the amplification of cox1 and rrnL respectively.In a few samples where amplification of the rrnL fragment failed, we assembled it from two overlapping fragments using the primer pairs 16Sar + 16Scs2 (Palumbi et al., 1991) and 16Scs1 + 16S_MN3R (Neiber et al., 2017).For ITS2, we used the primer pair LSU1 + LSU3 (Wade & Mordan, 2000) for both PCR and sequencing.ITS2 is present in the genome in multiple copies that may differ in length (Nekola et al., 2023), preventing simple sequencing of the PCR product.We initially attempted to use internal primers (Neiber et al., 2017) to assemble the sequences from multiple reads positioned between the indels, but this turned out impractical in most cases due to the presence of more than one variant.We, therefore, performed the PCR and sequencing for multiple specimens per clade and kept those, for which the product could be sequenced in whole using the LSU1 and LSU3 primers or with the 1-2 internal primers in addition (MN2-F, MN3-F, MN2-R, MN4-R; Neiber et al., 2017).
PCR was mostly done with Platinum II Taq Hot-Start DNA Polymerase (Thermo Scientific) following the manufacturer's recommendations except for annealing temperature (50°C) and addition of 1.5 mM MgCl 2 .PCR products were purified with ExoSAP-IT™ (Thermo Scientific) and Sanger sequencing was performed in both directions by capillary electrophoresis (at Faculty of Science, Charles University, Prague or Macrogen Europe, Amsterdam).
All sequences were deposited in GenBank, and accession numbers are listed in Appendix S1.Most of the mitochondrial data from M. incarnatus were already reported in Korábek et al. (2023).
For model and partitioning scheme selection, the cox1 was split into three partitions by codon position; rrnL and ITS2 were treated as a single partition each.The best scheme selected was GTR + F + I + G4 (rrnL; in IQ-TREE notation), TIM2 + F + I + G4 (cox1 1st), K3Pu + F + I + G4 (cox1 2nd) and K3Pu + F + G4 (cox1 3rd) for the mtDNA alignment and TIM3 + F + G4 for ITS2.GTR model was specified for all partitions for analyses in MrBayes.Branch support was assessed with 500 bootstrap replicates with IQ-TREE.Analyses in MrBayes were performed in two replicates, each of six heated chains for 25.5 million generations (i.e., until the average standard deviation of split frequencies dropped below 0.01), sampling each 10,000th; the first 25% of the runs were discarded as burn-in.
Average p-distances between species and major intraspecific lineages, and maximum p-distances within them, were calculated for cox1 with MEGA 11 (Tamura et al., 2021).

| General conchological characteristics
We examined the general shape of the shells, the umbilicus, colouration and the surface sculpture.Whorl count follows the method favoured by Ehrmann (1933).Shell diameter and height in selected individuals were measured, as shown in Kerney et al. (1983), to assess size ranges of studied taxa.

| Surface microsculpture
We photographed the surface microsculpture of 118 individuals of Monachoides, Xerocampylaea and several other genera of the family Hygromiidae (Appendix S1) with a light microscope and a scanning electron microscope (SEM).Monachoides individuals were selected to represent all major intraspecific mitochondrial lineages.
The shell surface was studied primarily on the upper shell surface of adult shells, on the last three-quarters of the body whorl, but a few juveniles were also used.We also examined the structure of the shell in cross-section by studying the fracture surfaces of the mechanically broken wall of the last whorl and measured the thickness of the periostracum, the height of the periostracal scales and the scale density.This also allowed studying the periostracal (organic) versus ostracal (mineral) origin of the structure.The shell fragments were attached using double-sided adhesive carbon or copper tape on metal supports, dried in a desiccator and gold-plated.Scanning electron micrographs were taken with a SEM JEOL 6380VL device.We also photographed the surface structures under a stereomicroscope, as that is how they are typically examined during species identification.
The density of the tubercles/scales was counted from photos within a 200 × 200 μm square.A feature was counted if more than half of it was within the delimited area.

| Genital system morphology
Dissections of ethanol-preserved bodies were performed in search of characters that would allow for distinguishing M. incarnatus from the Croatian clade of Monachoides revealed here (see Results).A total of 21 M. incarnatus individuals representing five major mitochondrial clades (from Czechia, Croatia, Italy, Bosnia and Herzegovina and France; Appendix S1), 10 individuals belonging to the Croatian clade and four M. vicinus individuals were dissected.
Love darts of 22 specimens of M. incarnatus, 11 specimens belonging to the Croatian clade and six M. vicinus specimens were examined.Dart sacs were carefully cut out of the reproductive system and, following Koene and Schulenburg (2005), placed overnight in 1-M NaOH, which dissolved all the tissue and mucus but left the love dart intact.We would like to highlight, however, that the darts were very fragile after overnight incubation and it seems better to extract the dart from the tissue remnants immediately when it becomes visible in the dissolving dart sac.The love darts were prepared for scanning electron microscopy in the same way as the shell fragments.

| Phylogeny
The mitochondrial phylogeny (Figure 2) divides the presently accepted Monachoides species into two groups.
Monachoides incarnatus, M. vicinus and M. bacescui are embedded within the Perforatellini ("Perforatellini Monachoides" hereafter).These are split into three clades, of which only two correspond to accepted species (M.incarnatus and M. vicinus).Monachoides bacescui is found to be nested within M. incarnatus, specifically within its shallowly differentiated, widely distributed clade ranging from the southern Balkans to Poland in the north and France in the west.Besides these two species, there is a third clade comprising some of the samples from north-western Croatia ("Croatian clade" hereafter).Specimens of this clade occur sympatrically and even syntopically with M. incarnatus and represent a morphologically distinct form.The division into these three clades is supported also by the ITS2 phylogeny (Figure 3), which shows three Monachoides lineages matching the three mitochondrial clades.
The sample from the type locality of M. fallax, collected on the top of the Trebević Mountain in Bosnia, groups alongside M. taraensis with Xerocampylaea and Semifruticicola in the Urticicolini clade ("Urticicolini Monachoides" hereafter).That is the case also for the juvenile hygromiid collected near the type locality of M. kosovoensis.The sequence of the latter sample is close to that of a specimen collected almost precisely at the type locality of M. kosovoensis, identified as Xerocampylaea erjaveci floerickei in Neiber et al. (2017).In addition to the samples reported here, we analysed a number of samples of Trochulus-or Monachoideslike samples from within the possible range of M. fallax (Welter-Schultes, 2012), but these all also turned out to be close to Xerocampylaea (Proćków et al., unpublished data) and will be treated in detail elsewhere.We did not detect any lineage related to M. incarnatus in Bosnia, Serbia, Montenegro and Kosovo.
Our results show Monachoides, in the restricted sense comprising only the species related to M. incarnatus and placed in Perforatellini, as paraphyletic.Both species of Perforatella (P.bidentata and P. dibothrion) form a clade with M. vicinus in the mitochondrial phylogeny (Figure 2) and P. bidentata also in the ITS2 phylogeny (P.dibothrion ITS2 sequences could not be obtained; Figure 3).

| Overall appearance
The three clades of the "Perforatellini Monachoides" group correspond to three morphotypes that can in most cases be distinguished by outer appearance (Figure 1).Two of them are M. incarnatus (Figure 1f-h) and M. vicinus (Figure 1b,c); the differences between these species were described in great detail by Pawłowska-Banasiak (2008).They often differ in the colouration: M. vicinus tends to be paler and yellowish, M. incarnatus is often somewhat darker and with a reddish shell.The colour of the foot of M. incarnatus varies from pink to dark grey.The mantle is usually densely covered by a fine network of dark grey, rarely red spots.The foot of M. vicinus is yellow to grey and the mantle usually has large, widely spaced black spots.The shell of M. vicinus typically has no umbilicus, unlike M. incarnatus, in which the umbilicus is developed.The shells of Monachoides have a thickened rim inside the aperture, which is visible from the outside where it forms a yellowish to ochre band; from the inside, it is typically white in M. vicinus, in M. incarnatus also reddish.There is often an inconspicuous elongate tooth-like protuberance on the lower apertural margin in M. vicinus, but this may also be present in M. incarnatus.None of these differences is, however, entirely reliable by itself due to inter-individual and geographic variation.Monachoides vicinus may have brown shells, slightly open umbilicus, and dark bodies as is more typical for M. incarnatus, black spots on the mantle may be reduced in both species, M. incarnatus may be yellowish, with nearly (but never completely) missing umbilicus and wellseparated, compact spots on the mantle.Despite these overlaps, the two species can usually be distinguished in the area where they occur sympatrically.The Croatian clade is also distinguishable by morphological traits.We examined the phenotypic differences particularly at three localities where specimens belonging to this clade occur syntopically with M. incarnatus in northwestern Croatia: Švica, Vrbovsko and Frketić Selo (Appendix S1).At these localities, the unidentified species was distinctly smaller than M. incarnatus, which largely holds for the species as a whole.Shells are flatter than those of M. incarnatus.The shell surface of the unidentified species is dull, the shell itself is yellowish or brown.The last whorl appears relatively broader than that of M. incarnatus, the lower part of the apertural margin is usually straighter and the apertural margins and the rim inside the aperture are white.The umbilicus is narrower than in M. incarnatus but present.The colouration of the body and mantle is similar to M. vicinus, but the spots on the mantle are smaller and denser; there is typically a larger, elongated, transverse dark spot close to the aperture, which allows also for the identification of juveniles at the localities where this taxon occurs syntopically with M. incarnatus.We found no intermediate phenotypes at the syntopic sites.
The species grouping with Xerocampylaea ("Urticicolini Monachoides") have an appearance different from Monachoides (Perforatellini group).Their shells are more opaque, they have a much wider umbilicus, and in M. taraensis there is a conspicuous white band on the periphery (Figure 1i).There is also no pattern formed by dark spots on the mantle visible through the shell wall in M. taraensis and M. fallax.

| Shell surface microsculpture
The three species of the "Perforatellini Monachoides" group are well distinguished from each other by the microsculpture of the shell surface.Their shells are covered with diamond-shaped to oval ostracal tubercles, whose density differs among species (least dense in M. vicinus, the most dense in the Croatian clade).On top of these, there are arched, slightly curved periostracal scales (Figure 4a-e); between the scales of some individuals, there are fine but distinctive transverse ridges (Appendix S4: Figure 5c,g,k).In Monachoides vicinus, the scales are typically completely lost on adult shells but can be found well developed on juvenile shells.While in the other two species, the scales are typically lost when the whole periostracum peels off, in M. vicinus the periostracum appears to consist of two layers: a thin surface layer with the scales that peels off and a more persistent basal layer that remains on the shell even if there are no scales present anymore.While the density of the surface scales and tubercles usually allows for easy species identification, two things need to be noted.First, some of the western-most M. incarnatus samples from Italy had a relatively high density of sculptural elements, reaching that of some individuals of the unidentified species from Croatia.Second, the density seems to reflect the growth rate of the shell (scales are arranged in rows parallel to the direction of growth) and is thus slightly uneven.
Examination of the thickness of the periostracum, height of the periostracal scales and the surface of the protoconch did not reveal any interspecific differences.
The shell surface sculpture is different in the three species of the "Urticicolini Monachoides" group, which lack the periostracal scales altogether.It is quite smooth in M. taraensis, with only weakly developed tubercles parallel to the growth lines (Figure 4g).In M. fallax, the shell is covered by rounded tubercles (Figure 4i).According to F I G U R E 5 Approximate distributions of Perforatella incarnata (top), Perforatella vicina and P. welebitana (bottom), location of the sampling sites and the distribution of mitochondrial lineages.Revised nomenclature is used.Size of the dots reflects the number of individuals analysed; colours correspond to those in Figure 2. Eastern range limits of P. incarnata and Perforatella vicina are poorly documented.We did not check the reported occurrence of P. vicina in the south of Hungary.The range of P. welebitana likely extends into Slovenia.de Winter & Maassen (1992), the surface of M. kosovoensis consists of irregularly placed and shaped nodules and ridges (as in Figure 4h,i).

| Genital system morphology
The genital system of M. vicinus has been repeatedly described in the literature (Grossu, 1983;Pawłowska-Banasiak, 2008;Schileyko, 1978;Schmidt, 1854) and the last author concluded that no character can reliably discriminate M. vicinus from M. incarnatus, although the flagellum is usually slightly shorter than the epiphallus in M. incarnatus and longer in M. vicinus.
We focused on finding differences in the anatomy of the genital system useful for discriminating between M. incarnatus and the Croatian clade.Most characters are variable (Appendix S2: Figure S1), but individuals of the Croatian clade tend to have much shorter penis than M. incarnatus (relative to female parts as well as epiphallus).The flagellum is shorter than in both M. vicinus and M. incarnatus (relative to penis+epiphallus; Appendix S4: Figure S5).The bursa copulatrix has a bulge on the sac (missing in some individuals), which appears to be positioned more or less apically in both M. incarnatus and M. vicinus (where it can be very prominent) and more laterally in the Croatian clade.More individuals with undamaged bursa should be examined, however, to confirm this.In contrast to the soft parts of the reproductive system, the shape of the love dart is very different between the two species.M. incarnatus has a spirally twisted, curved love dart (Appendix S3: Figure S2a-c; Appendix S4: Figure S6a).The love dart of the Croatian clade is straighter and not twisted (Appendix S3: Figure S2d-f; Appendix S4: Figure S6b).Monachoides vicinus has a love dart similar to the Croatian clade (Appendix S3: Figure S2g-i; Appendix S4: Figure S6c).
The tip of the dart bears two main and several less developed blades in M. incarnatus, M. vicinus and the Croatian clade, while in M. kosovoensis (drawing by de Winter & Maassen, 1992) and M. fallax, the dart is a simple spike that is circular in cross-section and has no blades, similar to the dart of Xerocampylaea erjaveci (Wagner, 1915).We failed to retrieve the dart of M. taraensis.

| Distribution and intraspecific diversity
Monachoides incarnatus is the most broadly distributed species assigned to this genus (Figure 5), occurring from Bulgaria and Greece in the south across the Balkans and Central Europe to southern Scandinavia (Kerney et al., 1983;Welter-Schultes, 2012;Wiktor, 2004) in the north.The western limits of its range are reached in France (Kerney et al., 1983;Welter-Schultes et al., 2011) and northern Italy (Cossignani & Cossignani, 1995); the eastern limits are relatively poorly known.It occurs up to western Ukraine (Balashov, 2016) and Moldova (Balashov et al., 2013), but in Romania it lives only in the south and southwest of the country (Gheoca et al., 2024;Grossu, 1983;Poliński, 1924;Soós, 1943).There is a substantial intraspecific diversity of relatively deeply divergent mitochondrial lineages in M. incarnatus (Figure 2; intraspecific p-distance in the cox1 fragment reaching 16%).The distribution of these lineages (Figure 5) has been detailed by Korábek et al. (2023), who inferred that the glacial refugia from which the present distributions derived were located mainly in northern Croatia and Slovenia.However, there was another refugium inferred in northwestern Italy ("M.incarnatus armatus").
Monachoides vicinus is a species with a primarily Carpathian distribution (Figure 5).Its range extends in a discontiguous manner to the Białowieża Forest and Masuria in Poland in the north (Cameron et al., 2010;Riedel, 1988), and to western Ukraine (Balashov & Gural-Sverlova, 2012) and Romania (Grossu, 1983) in the east.To the west, the range extends from the Carpathians to the Sudeten Mountains and along their northern edge (Wiktor, 2004).The species does not extend south of the Carpathians.Besides this more or less contiguous range, there are isolated populations in Bavaria, Germany (Häßlein, 1960); additional isolated occurrences were reported from the southwest of Hungary (Pintér & Suara, 2004).Several earlier authors claim that this species also occurs in Croatia and Slovenia (Brusina, 1870;Clessin, 1879;Ehrmann, 1933;Erjavec, 1877;Hirc, 1881;Kormos, 1907;Poje & Rađa, 1975) but these reports were likely due to misidentifications (Štamol, 2010).Among the sites listed by Brusina (1870), one is very close to our sampling site of the Croatian clade in Vrbovsko (Stubica) and at another one we sampled untypically coloured M. incarnatus with a nearly absent umbilicus (Podsused, Figure 1h).
Like in M. incarnatus, there are divergent intraspecific mitochondrial lineages within M. vicinus (p-distances in cox1 exceeding 15%), but the sampling is insufficient to describe their distribution.Most crucially, we lacked samples from the Romanian Carpathians and, therefore, additional major lineages could have been missed.However, we were able to analyse samples from an isolated Bavarian population and found it to be very closely related to samples from Poland and Slovakia.
The Croatian clade was found only in Croatia (Figure 5).To the west, the occurrence of this species extends to Vela Učka National Park, in the northeast we found it up to Karlovac, while the eastern-most record is close to Bihać.In the south, it was found in the Velebit Mts.southwest of Gospić.Its range lies completely within the range of M. incarnatus, but it is distributed southwest of the glacial refugia of M. incarnatus inferred by Korábek et al. (2023).Based on the material analysed so far, the Croatian clade has a very shallow intraspecific variation in the mitochondrial markers (pdistance in cox1 under 2%).
Complete overview of the intraspecific (as well as interspecific) sequence variation of the studied cox1 fragment is provided in Appendix S5.Grossu (1979) distinguished M. bacescui largely based on characteristics of the genital system, but the only character that should differentiate it from M. incarnatus and is actually depicted in his comparative drawings are long mucous glands.Indeed, we collected two adult snails at its type locality and both had long mucous glands (Appendix S2: Figure S1e).The shell characters (size, colour, surface) were not unusual for M. incarnatus and the length of the mucous glands is variable in M. incarnatus (Appendix S2: Figure S1).Both the mitochondrial and ITS2 phylogenies place M. bacescui in M. incarnatus.We, thus, consider M. bacescui a synonym of the latter.

| Validity of problematic taxa
The probable syntype of M. incarnatus armatus (deposited in Museo cantonale di Storia naturale, Lugano) has the usual appearance of M. incarnatus and lacks the tooth on the lower apertural margin that should characterize the taxon.We did not analyse samples from the type locality of M. incarnatus armatus near Lugano, only from even more westerly locations in northern Italy; all eight fully developed adult shells had a tooth on the lip.The samples yielded two divergent, endemic mitochondrial lineages embedded within M. incarnatus.Shell and body colour are not unusual, but the surface microsculpture of the shell tends to be more dense than typical for M. incarnatus (Figure 4c).We do not recognize any subspecies here formally, but we acknowledge that the sampled populations from northwestern Italy exhibit some regionally specific characteristics.They have been probably geographically isolated at least since the last interglacial (Korábek et al., 2023).
The type material of M. incarnatus welebitanus was most probably destroyed with Pfeiffer's whole collection (Zilch, 1967) and the description (Pfeiffer, 1848) is useless for identification.Wohlberedt (1909) used the name, with doubt, for large and reddish individuals from Montenegro; generally, it was presumed to be a taxon from Velebit Mts.
or more broadly the western Balkans (von Möllendorff, 1873;Jaeckel et al., 1957).Our M. incarnatus samples collected close to the Velebit were large but otherwise ordinary, and the molecular data show that these are closely related to central European M. incarnatus (Korábek et al., 2023).Falkner (1990) provided under the name M. incarnatus welebitanus a description that, apart from the larger size, fits the Croatian clade studied by us.We follow this interpretation and attribute the name to the Croatian clade by selecting a neotype for Helix welebitana Pfeiffer, 1847.
Detailed descriptions of the species of the "Perforatellini Monachoides" and information on types and synonymies are provided in Appendix S4.
We designate here a neotype for Helix welebitana Pfeiffer, 1847 to clarify its taxonomic status because the original description does not allow to identify the taxon (and in particular to distinguish it from Perforatella incarnata, which may occur syntopically), and also to fix the type locality of the taxon, which was originally not stated (Art. 75.3.1;ICZN, 1999).Helix welebitana is re-described (Art.75.3.2) and the neotype figured (Art. 75.3.3) in Appendix S4 (Figure S8a).No type material of Helix welebitana Pfeiffer, 1847 could be found in major old collections (Appendix S4) and the author's collection is believed to have been destroyed (Zilch, 1967) (Art. 75.3.4).There are two species around the Velebit Mts. that the original description may refer to; by the neotype selection we have chosen the one that presumably better fits to the characters given by the author of the description (see Appendix S4 for details) (Art.75.3.5).The new type locality is close to the Velebit Mts., from where the Pfeiffer's material likely originated given the taxon name, the biological species to which the neotype belongs does occur in the Velebit, the neotype is not genetically different from the populations in Velebit, and the chosen type locality yielded a representative series of individuals (Art.75.3.6).The selected individual was deposited in National Museum, Prague, Czechia under the number P6M 42979 (Art.75.3.7).

Monachoides
Our phylogenetic analyses show that the genus Monachoides as recently recognized contained two groups of very distantly related species.On the one hand, there is the type species M. incarnatus and two related species, which together with the two species of Perforatella belong to the subfamily Hygromiinae, tribe Perforatellini.On the other hand, there are M. fallax and M. taraensis, which group with Xerocampylaea in Trochilinae, tribe Urticicolini.The placement of the latter group in Monachoides was uncertain already before our study.Nordsieck (1993) suggested transferring M. fallax to Urticicola (thus close to its actual position revealed by molecular phylogenetic analyses), and de Winter & Maassen (1992) were themselves not confident about the generic placement of the two species they described, M. taraensis and M. kosovoensis, and noted similarities to Xerocampylaea.Although we could not analyse samples unambiguously identified as M. kosovoensis, Xerocampylaea samples were found in the area of its type locality (which lies on the border between Kosovo and Montenegro on the east side of Čakor Pass) by us as well as by Neiber et al. (2017), and the similarities to M. taraensis and M. fallax leave little doubt that this species also belongs to Xerocampylaea.Its exclusion from Monachoides is supported also by the presence of a small accessory sac in the genital system (de Winter & Maassen, 1992).
The genus Xerocampylaea was, until the work of Neiber et al. (2017), considered a monotypic genus with a restricted range.Since then, Xerocampylaea emerged as a more diversified group widely distributed in the western Balkans and the southwest of the Pannonian Basin from Kosovo to Slovenia and Hungary (Proćków et al., 2019).It now includes taxa previously classified in Trochulus Chemnitz, 1786(Trochulinae: Trochulini Lindholm, 1927), and we here show that also taxa classified as Monachoides belong to this group (X.fallax, X. taraensis and X. kosovoensis).A more detailed study of Xerocampylaea, based on a broad sample of sequenced individuals, is in preparation.
We found three species in what remained of Monachoides after excluding the misclassified Balkan taxa.Of the three originally recognized, Monachoides bacescui is indistinguishable from M. incarnatus both morphologically and genetically, so we consider it a junior synonym of the latter.However, the number remains at three, as we confirmed the distinctness of another species from western Croatia, hitherto considered a subspecies or synonym of M. incarnatus.It is clearly differentiated in both mitochondrial and nuclear markers and can be reliably distinguished by its finer, more dense surface sculpture.Also, the overall appearance is different from the other two species.All three species appear to be reproductively isolated, as the range of M. incarnatus overlaps with the ranges of the other two, and frequently occurs with either of them syntopically.The Croatian clade is matched here with M. welebitanus (Pfeiffer, 1847) by designating a neotype for that taxon; this interpretation largely follows the description given by Falkner (1990).
However, as M. vicinus is shown here to be more closely related to both species of Perforatella than to the other two Monachoides species while sharing conchological characteristics with the latter, the two genera can no longer be treated as separate.Perforatella was characterized by shells with regular riblets but no periostracal structures and two large teeth on the lower apertural margin, traits which appear to be derived in light of our phylogenetic analyses.But apart from these differences, the two genera share similarities in the genital system, although these are not diagnostic: single dart sac without traces of an accessory sac (unlike Lozekia, Kovacsia, Chilanodon, Lindholmomneme; Schileyko, 1978;Fehér et al., 2009), and short, but not highly reduced flagellum.Furthermore, the shell morphology of P. bidentata and P. dibothrion is probably an ecological adaptation (a very similar morphology is seen in Chilanodon bicallosa (L.Pfeiffer, 1853)) and apparently derived, and as such taxonomically uninformative.Finally, the periostracal scales typical for Monachoides are present on the shells of Perforatella hatchlings (Kuźnik-Kowalska & Roksela, 2009).We propose to reunite the two genera in Perforatella Schlüter, 1838 with five species: Perforatella bidentata (Gmelin, 1791), Perforatella dibothrion (Kimakowicz, 1884), Perforatella vicina (Rossmässler, 1842), Perforatella incarnata (Müller, 1774) and Perforatella welebitana (Pfeiffer, 1847).

| Biogeographic implications
We have previously shown that even though the range of P. incarnata includes also most of the non-Mediterranean Balkans, the diversification centre of the species is located more northerly, in Slovenia and Croatia (Korábek et al., 2023).Perforatella vicina is a primarily Carpathian species, not occurring south of the Carpathians.The newly defined P. welebitana lives north of 44.4° of latitude, and both P. bidentata and P. dibothrion are distributed exclusively north of the Balkans.The Balkan species previously included in Monachoides were transferred to the very distantly related genus Xerocampylaea (Trochulinae: Urticicolini) here.Perforatella, thus, belongs to a group of several Hygromiidae genera that apparently diversified and persisted over the Quaternary glacial cycles north of the south European peninsulas in the Alps, Carpathians, and Central Europe.Other examples are Petasina Beck, 1847, Plicuteria Schileyko, 1978, Prostenomphalia Baidashnikov, 1985and Trochulus Chemnitz, 1786, after many species were excluded from the last-mentioned genus following molecular phylogenetic analyses (Neiber et al., 2017).
Snails with this type of distribution are interesting from the palaeoecological point of view, because they likely survived the glacial cycles rather northerly, yet are mostly still more dependent on woody flora and less tolerant to harsh climatic conditions than the typical land snail fauna of the cold stadials of Central Europe (Juřičková et al., 2014).These taxa could, therefore, be particularly informative about the location of the northern-most glacial refugia of the temperate invertebrate fauna in Europe, considering that the shells of land snails frequently leave a well-preserved and recognizable fossil record.
Further studies are needed to confirm if the species of the "Perforatellini Monachoides" can be distinguished from all other hygromiids using the surface sculpture of the shell only.We found that the three "Perforatellini Monachoides" species differ from each other in the shell surface sculpture even after the periostracum is lost (Appendix S4: Figure S4d,h,l), which is important for species identification in the fossil record.The Quaternary fossil record of land snails is extensive, but medium-sized snails like Perforatella are typically found in the form of small shell fragments, which can often be identified only based on the surface structure (Ložek, 1964).It remains to be demonstrated that the surface sculpture of the "Perforatellini Monachoides" after periostracum loss is unique (e.g., with respect to Lozekia and Kovacsia).This is important because in the fossil record we cannot rely on present distributions when identifying the material.
The functional significance of periostracal surface sculptures in land snails remains virtually unexplored.There have been speculations on the function of periostracal hair in Trochulus (Trochulini) with regard to the water that can be held between the hairs (Pfenninger et al., 2005;Proćków et al., 2018;Shvydka et al., 2020).Such hairs occur in many unrelated lineages of land snails and was even reported from a Cretaceous caenogastropod land snail (Neubauer et al., 2019), suggesting some significant function, but there is no well-supported consensus on what advantage the hairs convey.Why species from different lineages have mutually similar sickle-like periostracal scales is then completely unclear.However, regardless of their possible functions, they can be a valuable character for taxonomy and for species identification from shell fragments.

| CONCLUSIONS
The land snail family Hygromiidae is an example where traditional morphology-based taxonomy was unable to uncover the true relationships among species, genera and family-group taxa.The earlier reliance on the genital system morphology led to incorrect family-and genus-level systematics.A crucial defining character of Monachoides was thought to be a single dart sac, but our results show that this trait is shared by distantly related taxa in the family.Three species classified so far in Monachoides were found to belong to Xerocampylaea, a monophyletic clade that now contains species with one or two dart sacs (with or without accessory sacs).Conchological characters are also unreliable at the genus level.The remaining species of Monachoides share a typical microsculpture on the shell surface.However, as a group, they were found to be paraphyletic with respect to Perforatella, a genus comprising so far two species lacking the microsculpture and differing from Monachoides in having prominent teeth on the lower apertural margin.Based on the results of phylogenetic analyses, Monachoides is here synonymized with Perforatella, which is an older name.
The change in the taxonomic perspective is accompanied also by a change in the biogeographic perspective.As long as Monachoides included narrow-range endemics from the western Balkans, it was natural to expect that its diversity, both species-level and intraspecific, follows the pattern usual for mid-to large-sized land snails, for which the centre of diversity is located in the south of the taxon's range (e.g., Hofman et al., 2022;Korábek et al., 2022;Pinceel et al., 2005;Ramos-Gonzales et al., 2022;Welter-Schultes, 2012).We have shown previously that this is not true for the intraspecific diversity of P. incarnata (Korábek et al., 2023), and here we demonstrate the same for the whole genus.We did uncover an overlooked species, but not in the south of the distribution range of Perforatella.After our revision, Perforatella is a genus from southeastern Central Europe with five species.Being related to genera from the Carpathians and southern Siberia (Neiber et al., 2017), it is part of a group that diversified in relatively high latitudes and apparently never had any close relationship to the hotspot of land snail diversity in the Balkans.

F
I G U R E 2 Mitochondrial phylogeny of Monachoides.Maximum likelihood tree based on concatenated partial rrnL and cox1 sequences.Support values represent SHaLRT, standard bootstrap (both from the ML analysis) and posterior probabilities from a Bayesian analysis respectively.The supports are shown for branches where SHaLRT >95 or bootstrap >70 or posterior probability >0.95.Species classified so far in Monachoides and samples collected at or near the type localities of Monachoides kosovoensis are highlighted in bold.Revised taxonomic names (see Systematics) are shown in bold on the right.The colours correspond to those in Figures 3 and 5. * data from Neiber et al. (2017), ** data from Proćków et al. (2019), *** data from Pfenninger et al. (2005).

F
Nuclear phylogeny of Monachoides sp.("Perforatellini Monachoides").Maximum likelihood tree based on the internal transcribed spacer 2 and the flanking parts of 5.8S rRNA and 28S rRNA genes.Support values represent SHaLRT, standard bootstrap (both from the ML analysis) and posterior probabilities from a Bayesian analysis respectively.The supports are shown for branches where SHaLRT >95 or bootstrap >70 or posterior probability >0.95.Coloured dots indicate intraspecific mitochondrial lineage of each analysed M. incarnatus individual.An asterisk indicates data from Neiber et al. (2017); M. incarnatus and M. vicinus sequences from that study were not used, as the former formed distinctively the longest branch within M. incarnatus and the latter branched off basally of a clade comprising our M. vicinus samples and Perforatella bidentata.Attempts to sequence Perforatella dibothrion failed.Revised taxonomic names (see Systematics) are shown in bold on the right.