New Freshwater molluscs from Crete, Greece (Gastropoda: Hydrobiidae, Bythinellidae, Valvatidae)

The syntype of Pseudamnicola brachia (Westerlund, 1886), the only known Pseudamnicola sp. from Crete, is compared with a recently collected Pseudamnicola sp. which could be identified as a species new to science. Comparisons with data in the literature show us that P. brachia (Westerlund, 1886) has nothing in common with P. brachia sensu Schütt, 1980 as well as P. brachia sensu Szarowska et al ., 2015, but both are possibly conspecific with the new species which is described here. In addition we describe two new Bythinella spp. from Crete which could be found in addition to B. cretensis Schütt, 1980, and a new Valvata ( Tropidina ) sp. from Lake Kournas.


Introduction
Many Pseudamnicola species have been found in the Aegean Islands as reported by Szarowska et al. (2015), Radea et al. (2015) and Glöer et al. (2018). In all cases these species are regional endemics. One part of our study was to identify Pseudamnicola brachia (Westerlund, 1886), the only Pseudamnicola sp. known so far from Crete. We found several populations of Pseudamnicola sp. and compared them with specimens presented in literature.
Formerly only one species of the genus Bythinella, B. cretensis Schütt, 1980, was known from Crete. Benke et al. (2011) sequenced the only sample of a Bythinella sp. from Crete they had, as B. cretensis, but unfortunately they did not collect the species from type locality. Szarowska et al. (2016) could show that there are more than one Bythinella spp. inhabiting Crete, as they found three distinct clades, but they incorrectly used the sampling site of Benke et al. (2011) as the supposed type locality of B. cretensis.
From the holarctic Valvatidae Gray, 1840, five species have been mentioned from Greece: Valvata  Schütt, 1962 from Lake Trichonis (Bank, 2004, Bank & Neubert, 2017. Valvata (Tropidina) macrostoma has been mentioned by Bank (2004: 57) and Bank & Neubert (2017) only from mainland Greece. Recently H-JH collected Valvata (Tropidina) sp. from Lake Ecologica Montenegrina 20: 10-23 (2019) This journal is available online at: www.biotaxa.org/em Kournas, which looks at a first glance like V. macrostoma. As the width of the umbilicus is suggested to be an important feature to distinguish between Valvata spp. (Vinarski et al., 2013), we compared the umbilicus of our material with V. macrostoma from Central Europe.

Material and Methods
During field trips in 2014, 2015 and 2016 we checked more than twenty springs for the occurrence of freshwater molluscs. If present, living snails were collected by hand by Andrea and Hans-Jürgen Hirschfelder, in part also by Theresia and Klaus Kittel, and fixed in 75% ethanol. Also the biggest freshwater lake of the island, Lake Kournas, was investigated, mainly the outflow at the northern end. Many empty shells of several species were deposited there.
To evaluate the current material we needed to review the historical material. To this end we were able to borrow the syntype of Pseudamnicola brachia from Westerlund's collection (NHMG, Göteborg) and obtain photos of the holotype and paratypes of Bythinella cretensis (SMF, Frankfurt a. Main).
The dissections and measurements of the genital organs and the shells were carried out using a stereo microscope (Zeiss). The type material is stored in the Zoological Museum of Hamburg (ZMH), Germany, whilst some paratypes are deposited in the private collections of the authors. The anatomical and shell photos were made with a digital camera system (Leica R8).

Results
We collected specimens of the genus Pseudamnicola Paulucci, 1878 in six springs and specimens of the genus Bythinella Moquin-Tandon, 1856 in ten springs. Recently one species of the genus Planorbis O.F. Müller, 1773 was detected and described as a new species (Glöer & Hirschfelder, 2015). At Lake Kournas, we collected several species of molluscs but no living specimens were found. After examination of the material collected we suspected a higher diversity of the freshwater mollusc fauna of Crete than previously thought.
Comparisons between our Pseudamnicola material and the syntype showed considerable differentiation. Specimens presented in literature which are very similar to ours were obviously misidentified. Thus we found an additional Pseudamnicola sp. and describe this species here as new to science. The holotype and one paratype are depicted in addition to the penis, which is characteristic for the genus.
The collected material of the genus Bythinella was compared with the original description of Bythinella cretensis Schütt 1980. We found three different species. We decribe herein two Bythinella spp. new to science. The comparison with Szarowska et al. (2016) showed that two of these species correspond very well with two of their proposed clades, the third species is totally new, and from the third clade in Szarowska et al. (2016) we had no material.
The comparison with Valvata macrostoma from Central Europe revealed that the Valvata sp. from Lake Kournas is a species new to science which is described herein.
In summary, at least two species of Pseudamnicola, four species of Bythinella and one species of Valvata live on the island of Crete.

Remarks:
The original description ( Fig. 1) corresponds with the syntype of Westerlund's collection (Fig. 3). Unfortunately Westerlund did not mention the exact type locality on Crete in his description. On the label only the town of Candia is noted which is the older name of the capital Iráklio. The specimens collected by H-JH do not correspond with the original description nor with the picture of the syntype of Pseudamnicola brachia of Westerlund's collection: "Gew. dick u. kurz, eifg., stumpf …". This translates to: "spire broad, short ovate, obtuse …". And the difference becomes particularly visible if we compare the type with a subadult specimen of Pseudamnicola occulta n. sp. of the same size (Figs. 3, 4).  (Westerlund, 1886) in coll. Westerlund; 1: Topólia, type locality of Pseudamnicola occulta n. sp.; green dot: Spíli, paratype locality of P. occulta n. sp.; red dots: additional material of P. occulta n. sp., 2 = Myrthios, 3 = Amári Valley between Méronas and Amári, 4 = Zou, 5 = Zákros; blue dot: Almyros spring in Georgioúpoli, sampling site of Pseudamnicola sp. in Szarowska et al. (2015), see Figs. 5, 6; yellow dot: Almira spring near Chaniá, sampling site of Pseudamnicola sp. in Schütt (1980, Fig. 29).  Schütt (1980: 133) listed Pseudamnicola brachia from many sampling sites on Crete, also from Spíli but not from Topólia. If we compare his photos with P. occulta n. sp. we see that Schütt had possibly the new species in hand, but not P. brachia. Also the photos presented in Szarowska et al. (2015)

Etymology:
The new species is named P. occulta which means "hidden". It has already been found by several authors but erroneously recognized as P. brachia. So the new species was "hidden" behind a wrong name for a long time.

Description
Shell: The brownish shell is glossy, conical to elongated conical with 4.5 convex whorls and a deep suture. The aperture is ovate, slightly oblique, with a sharp peristome which is thickened at the columella. The apex is pointed. The umbilicus is open. The shell is 2.7-3.4 mm high and 2.0-2.3 mm broad.
Animal: The animal is dark brown pigmented, with greyish tentacles. The penis is cylindrical with a broad basis and an obtuse penis tip.

Differentiating characters:
The spire in Pseudamnicola occulta n. sp. is higher than in P. brachia, the whorls are more convex and the suture is deeper. In addition P. occulta n. sp. is much larger than P. brachia. Additional material: In the following springs P. occulta n. sp. probably lives as well. The collected specimens correspond with the type series but are not treated as paratypes because only dry material could be examined and no anatomical investigation was made: Sampling site  Distribution: P. occulta n. sp. is known from several springs in western, central and eastern Crete and is probably widely distributed in the island.
Remarks: During our visit on Crete we found no Pseudamnicola in springs in the vicinity of Iráklio, i.e. in the big source of Almyros river 8 km west of the town center of Iráklio. Therefore the exact type locality and the distribution of the "true" P. brachia (Westerlund, 1886)

Description:
Shell: The shell is cylindrical with a small apex. The 4-4.5 whorls are slightly convex with a deep suture. The body whorl is prominent, especially in height. The aperture is ovate, narrowed at the top, with a peristome thickened at the columella. The umbilicus is slit-like to closed. The shell is up to 3.3 mm high and 1.5-2.0 mm broad.
Animal: The penis is shorter than the penial appendix. The tubular gland has the same width over the full length.
Sampling sites (numbers refer to the map in Fig. 19 A very small Bythinella population was also found 5 km W of Meso Potámi in a seeping spring at the monastery Kardiotisa, but the sample was too small for anatomical investigation. So the status of this population remains unknown.
Distribution: Lasíthi highlands and east central Crete.

Remarks:
For discussion see under Bythinella sitiensis n. sp.

Description:
Shell: The shell is cylindrical with a small apex. The 4-4.5 whorls are slightly convex with a deep suture. The aperture is ovate, narrowed at the top, with a sharp peristome. The umbilicus is slit-like. The shell is 2.7-2.9 mm high, 1.7-1.8 mm broad.
Animal: The penis is as long as the penial appendix. The tubular gland has the same width over the full length.
Additional material: B. magdalenae n. sp. probably also lives in the spring of Spíli (see above under Pseudamnicola occulta n. sp.). The collected specimens correspond with the type series but are not treated as paratypes because only dry material could be collected and no anatomical investigation was made. Etymology: Bythinella sitiensis n. sp. is named after Sitía in eastern Crete, in whose municipality (Dimos Sitías) the new species was found in two springs.

Description:
Shell: The shell is cylindrical with a small apex. The 4-4.5 whorls are slightly convex with a deep suture. The aperture is ovate, narrowed at the top, with a thickened peristome at the columella. The umbilicus is closed. The shell is 2.5 mm high, 1.5-1.7 mm broad.
Animal: The penis is as long as the penial appendix. The tubular gland becomes thicker at the distal end.
Distribution: Far eastern part of Crete.

Remarks:
The distribution map of our three Bythinella species (Fig. 19), we could determine by anatomical features, fits very well with two proposed clades in Szarowska et al. (2016). Clade "CR1" sensu Szarowska et al. (2016) is the true B. cretensis (their sampling site K07 is identical with our site 5), Clade "CR2" sensu Szarowska et al. (2016) should be B. magdalenae n. sp. The sampling site "Epis" (collected by Benke et al. 2011 as "B. cretensis") is 1100 m away from the type locality of B. magdalenae n. sp. In the region of clade "CR3" sensu Szarowska et al. (2016) we did not collect, and the region of East Crete, where we found B. sitiensis n. sp., was not included in the work of Szarowska et al. (2016). Anatomical and shell differences between the studied species are given in the key below.
Identification key for the Bythinella ssp. from Crete

Description:
Shell: The horn-colored shell has 3.5 circular whorls with a low spire and a blunt apex. The surface is densely covered by regular-silky ribs. The umbilicus is wide, shell diameter to diameter of umbilicus about 4 : 1. The aperture is circular and slightly angulated at the top. Shell height up to 2.6 mm, diameter 5.0 mm.
Differentiating characters: From the most similar Valvata macrostoma the new species differs in the narrower umbilicus and the slight angle at the top of the aperture which is missing in V. macrostoma.
Distribution: V. kournasi n. sp. is probably endemic to Lake Kournas.

Discussion
Seven Pseudamnicola spp. have been described from the Aegean islands (Bank, 2004, Delicado et al., 2014, Bank & Neubert, 2017, Glöer et al., 2018, most of them endemic to just one island. Only on Rhodes two distinct Pseudamnicola spp. could be found (P. exilis (Frauenfeld, 1863) and P. ianthe Radea & Parmakelis, 2015), and now also on Crete (P. brachia and P. occulta n. sp.). Thus eight Pseudamnicola spp. are currently known from the Aegean islands. Szarowska et al. (2016) stated that the differences between the Bythinella-clades form Crete are small and thus the morphology of the shells and the penis alone are not suitable to distinguish between Bythinella spp. We can confirm their results and in case of doubt the Bythinella spp. from Crete can be identified geographically and/or in combination with molecular data.
Valvata kournasi n. sp. is probably endemic to Lake Kournas because this is Crete's only freshwater lake of natural origin. In contrast to V. macrostoma which inhabits marshland, alluvial river flats, small ponds or temporary waters, the new species lives in open water of a large lake.
Springs are very sensitive ecosystems. All habitats where we collected Pseudamnicola and Bythinella are captured or partly captured and have been changed and modified for water supply. In several springs the snail populations were very small and therefore potentially endangered. This is especially true for the type locality in Topólia because it is a captured source in the center of the village. Many leaves of the surrounding trees where the molluscs feed on are deposited in the gutter. Someone might try to "clean" the water basins and the snails can be eradicated easily. This problem is evident in most of the springs. It is very important to inform the local people about their endemic snails and the need of protection.