NEW BYTHINELLA SPECIES FROM NORTHERN ROMANIA (GASTROPODA: RISSOOIDEA)

Recent samples of freshwater molluscs from Romania contained six new Bythinella species: B. blidariensis n. sp., B. georgievi n. sp., B. falniowskii n. sp., B. feheri n. sp., B. sirbui n. sp., and B. szarowskae n. sp. Since B. molcsanyi H. Wagner, 1941 sometimes co-occurred with B. grossui Falniowski, Szarowska et Sîrbu, 2009 or with B. feheri n. sp., differences between these species were studied, to identify diagnostic features. Apart from shell morphology, also the tubular gland proved to be important for identification of Bythinella spp.


INTRODUCTION
Species of the genus Bythinella Moquin-Tandon, 1856 are distributed from Europe to Turkey and North Africa.They inhabit springs and caves in the mountains (BOETERS 1998).The highest species richness is known from France and the Balkan Peninsula (GEORGIEV & STOYCHEVA 2011).Passive dispersal of photophobic species is difficult due to their habitat preferences, thus most of the species are locally endemic (GLÖER & GEORGIEV 2011).

MATERIAL AND METHODS
Dr. ZOLTÁN FEHÉR (Hungarian Natural History Museum, Budapest) collected 19 samples of Bythinella spp. in Romania in May 2009 (Fig. 1, Table 1).The snails were collected by hand and with a sieve from freshwater habitats.The samples were put into 75% ethanol.The dissections and measurements of genital  organs and shells were carried out using a stereo microscope (Zeiss); photographs of the shells and the genital anatomy were taken with a digital camera system (Leica R8).
To identify Bythinella spp., the following features have been applied: (i) shell morphology: height, ratio of shell height to the width of the last whorl (H:W), aperture height to shell height ratio (A:H), umbilicus type, aperture shape, and (ii) the male copulatory organ (Fig. 2): length ratio of the penis to the penial appendix, number of half loops of the tubular gland, and width of the proximal and distal parts of the tubular gland.The tubular gland is not considered by many authors, but it is an important feature to distinguish Bythinella spp.To select constant features of the male copulatory organ, it was studied in at least three specimens from every sample.Shell variability was assessed on the basis of measurements.
The female reproductive system is not considered here because the differences did not provide any useful features to distinguish between the Bythinella spp.under discussion.
For species delimitation, I had to work with the morphological species concept (QUEIROZ 2005, SINGH 2012), based on features that are intraspecifically constant but interspecifically different.A good test was that there were three samples in which the Bythinella specimens could be separated into two distinct clusters each, identified as B. molcsanyi and B. grossui (038RO, 053RO: Fig. 3, Table 1), or B. molcsanyi and B. feheri n. sp.(054RO: Table 1), respectively.Sympatric populations confirm indirectly the biological species concept, which could support the morphological species concept in this case.In addition, morphological distinctness of B. molcsanyi and B. grossui All materials are stored in the Hungarian Natural History Museum (HNHM) and some paratypes in my own collection.

RESULTS
Identification of Bythinella spp. is not easy because these species have only few distinguishing features.Therefore, only the differentiating features have been used for species description.The following diagnostic features were recognized in this study: (i) shell size, (ii) morphometry of the tubular gland, and (iii) proportions of the penis and penial appendix.The main feature is the tubular gland, which can be (i) evenly thick over the whole length; (ii) thinner or thicker in the proximal part, and (iii) evenly thick or bulbed in the distal part.In addition, the number of half loops of the tubular gland is a good distinguishing feature (Fig. 2).
Differentiating features: This species differs from the other Romanian species in its shell height and the short tubular gland with 3 half loops, which is similar to those of B. feheri and B. szarowskae but in the latter species the distal end of the tubular gland is bulbed.IDENTIFICATION KEY OF BYTHINELLA SPP.OF NORTHERN ROMANIA For identification, mainly the features presented in Table 2 are used.For nomenclature of the male copulatory organ, see Fig. 2. Abbreviations: tgl -tubular gland half loops, H:W -shell height to width ratio, A:H -aperture height to shell height ratio.Description: Shell horn-coloured, cylindrical and slim (Fig. 14), its 4.5 whorls regularly rounded, with a deep suture.Surface silky and finely striated.Apex broad and obtuse, umbilicus slit-like.Aperture oval.Shell height 2.7-3.1 mm, mean 3.0 mm (s = ±0.17,n = 11), width 1.5-1.7 mm, mean 1.6 mm (s = ±0.17),aperture height to shell height ratio 0.36 (s = ±0.11).Penis as long as penial appendix (Fig. 15), tubular gland short, with 3 half loops, thin proximally and bulbed distally.In one specimen, the penial appendix had 2 tubular glands (Fig. 16).

Differentiating features:
The ratio of aperture height to shell height is 0.36 and the tubular gland is short and club-shaped at the distal end.The aperture is oval and not angled, as it is in B. falniowskii and B. feheri.

Differentiating features:
The tubular gland is long and evenly thick, with 5 half loops.

DISCUSSION
The syntopic species Bythinella molcsanyi and B. grossui, as well as B. feheri n. sp. and B. molcsanyi, differ in shell proportions as well as in thickness of the proximal part of the tubular gland or length of the tubular gland.These are the main features used for species delimitation in Bythinella spp., which has already been pointed out by GLÖER & GEORGIEV (2011).Because all specimens were collected in the same month (1-3 May 2009), the differences in the tubular gland cannot be a result of different sexual activities.A multivariate cluster analysis (Fig. 28) revealed that it is possible to distinguish the N Romanian Bythinella spp.from each other by the features listed in Table 2.
All Bythinella spp.from N Romania seem to be locally endemic (Fig. 1), as is the case in other mountainous countries where Bythinella spp.occur.Recent investigations of Bythinella spp. in the neighbouring Bulgaria revealed 18 distinct species (GEORGIEV 2009, GLÖER & GEORGIEV 2011, GEORGIEV & STOYCHEVA 2011), so it is possible that more Bythinella spp.might be found in Romania in the future.

Table 2 .
Distinguishing features of Bythinella spp. in N Romania