Research Article |
Corresponding author: Ondřej Korábek ( ondrej.korabek@gmail.com ) Academic editor: Frank Köhler
© 2023 Ondřej Korábek, Igor Balashov, Marco T. Neiber, Frank Walther, Bernhard Hausdorf.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Korábek O, Balashov I, Neiber MT, Walther F, Hausdorf B (2023) The Caucasus is neither a cradle nor a museum of diversity of the land snail genus Helix (Gastropoda, Stylommatophora, Helicidae), while Crimea is home to an ancient lineage. Zoosystematics and Evolution 99(2): 535-543. https://doi.org/10.3897/zse.99.110610
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The Caucasus and the adjacent Pontic Mountains in north-eastern Anatolia are home to numerous endemic land snail genera and species. The diversity of the region is the result of both intra-regional speciation and the persistence of relict lineages. The same seemed to be true for the genus Helix, which has been present in the Greater Caucasus since the Miocene. In the Caucasus region, there are three Helix species. Helix buchii (Pontic Mountains and Georgia) and Helix albescens (southern Ukraine to northern Lesser Caucasus) are both separated by deep splits from the major Helix clades in the mitochondrial phylogeny. In contrast, Helix lucorum belongs to the Anatolian radiation of Helix. At least part of its intraspecific diversification may have occurred in north-eastern Anatolia and the adjacent parts of the Caucasus. Here, we report new evidence suggesting that the Caucasus and the Pontus regions were less important as a refugium of ancient Helix lineages or as a diversification centre than previously hypothesised. Helix lucorum probably diversified more westwards, while H. buchii is a less ancient lineage than previously thought. Helix albescens had its long-term refugium on the Crimean Peninsula in southern Ukraine, not in the Caucasus. The Caucasus is close to the eastern limit of the distribution range of the genus and, although the fossil record shows that Helix was present there as early as the Miocene, the current diversity of the genus there is the result of much later colonisation.
Anatolia, fossil, land snail, phylogeography, refugia, Ukraine
The Caucasus ecoregion (
Phylogenetic relationships between some Pontic land snail taxa on the one hand and those from southern Anatolia on the other hand suggest that the Pontic region contains relicts of lineages that were eliminated in central Anatolia by the uplift and aridification of the Anatolian Plateau and its surrounding mountain ranges (
In addition to the two possibly relictual species, there is a third Helix species in the Caucasus, Helix lucorum Linnaeus, 1758. It is distributed all over the Caucasus region, Anatolia and the southern Balkan Peninsula. The occurrence of most of the main intraspecific mitochondrial clades in the Caucasus region or adjacent north-eastern Anatolia, with endemic clades in the south-eastern Caucasus (
Based on our expanded sampling, we tested the hypothesis that the Caucasus region and the Pontic Mountains were the area where the intraspecific diversity of H. lucorum originated and that the other two species are old, phylogenetically isolated lineages that persisted in the Caucasus (H. albescens) or the Pontic Mountains (H. buchii).
Partial sequences of the mitochondrial genes for cytochrome c oxidase subunit I (cox1), 16S rRNA (rrnL) and 12S rRNA (rrnS) were analysed. Laboratory methods were as described in
The new data were combined with previously published sequences (
Intraspecific mitochondrial lineages of the Caucasian Helix species and their distribution. A–C. Distribution of major clades (i.e. the branches labelled with correspondingly coloured circles in the trees). Non-native or potentially non-native occurrences are not distinguished. A. H. albescens; B. H. buchii; C. H. lucorum; D. Classification of the discussed regions: southern Ukraine (mainland, orange), Crimea (yellow), the Caucasus region (red), eastern and western part of Anatolia (dark and light blue) and the Balkans (green). The east-west division of Anatolia is for plotting purposes only and coincides with the western distribution limit of H. buchii, which also marks the limit of the wettest area of the Pontic Mountains; E–G. Bayesian mitochondrial trees based on concatenated alignments of partial rrnL, rrnS (only in H. buchii) and cox1 sequences. Supported nodes are indicated by small circles at nodes (omitted from the shallowest nodes), large circles on branches denote the clades used for plotting the maps along with the corresponding numbers of the clades as given in
Alignments were done with MAFFT 7.487 (E-INS-i algorithm;
PCR and sequencing with the primer pair LCO1490 and HCO219 targeting the “barcoding” fragment of cox1 produced for two H. buchii individuals an incomplete pseudogene sequence containing an in-frame stop codon and frame-shift indels. Two phylogenetically distant mitochondrial lineages are known from H. buchii, one of which was probably acquired from an unknown, possibly extinct species (
The sequence data underlying this article are available from the NCBI Nucleotide database (available at: www.ncbi.nlm.nih.gov/Genbank). Accession numbers are provided in the online supplementary material along with sampling localities and voucher information.
We found two major clades, each of them with only shallow internal divergences (Fig.
Two divergent lineages have been found in H. buchii. All but one of the analysed individuals yielded haplotypes belonging to the ‘majority lineage’, which is deeply separated from the major Helix radiations (Figs
Maximum Likelihood tree based on rrnL and cox1 sequences showing the relationships of H. buchii. Both mitochondrial lineages and the cox1 pseudogene (presumed NUMT) sequence obtained from two of its individuals with the majority lineage mitochondrion are included. The samples listed in Suppl. material
Partial, but overlapping cox1 pseudogene sequences, were obtained from two individuals (GenBank accessions: isolate SP2 OQ148366, isolate 6916 OQ148367). These samples otherwise yielded haplotypes of the clades 44 and 45 of the majority lineage. Phylogenetic analysis placed the pseudogene sequences into the Anatolian clade of Helix (
In Armenia and Azerbaijan, all but one sample (from Tatev Monastery, Armenia) yielded haplotypes from two co-occurring clades (Fig.
The haplotypes of the clade 89 form four well-supported groups. One of them was found only in a non-native sample from Spain, but one was recorded so far only in the eastern Balkans and none of the remaining two is unique to the Caucasus and Pontus. Furthermore, there are nearly identical haplotypes (1 bp difference over 1469 bp of the rrnL+cox1 alignment) in the Caucasus on the one hand and the eastern Balkans on the other hand. The sister lineage of the whole clade (red in Fig.
Populations with globular shells, which share mitochondrial clade 93 (green in Fig.
Both major mitochondrial clades within H. albescens occur in the Crimean Mountains and the lowlands of southern Ukraine, while only one of them occurs in the Caucasus (Fig.
The extent of the native distribution of H. albescens on the East European Plain is unclear. It is relatively continuously distributed across the steppe ecoregion of southern Ukraine and in adjacent regions of Russia south to the Caucasus, both in natural and transformed habitats. In Ukraine, there are no Quaternary fossil records of this species outside Crimea (
Based on the distribution of individual clades from the majority lineage, Helix buchii did not originate in the Caucasus hotspot or the centre of land snail diversity in the mountains around Artvin in Turkey. The data show generally low intraspecific diversity in Georgia, whereas multiple lineages (including the minority lineage) were found in north-eastern Turkey. Our results obtained with improved sampling thus confirm earlier observations (
The phylogenetic placement of the cox1 pseudogene (presumed NUMT) sequenced here in the Anatolian clade indicates that the minority mitochondrial lineage is the original mitochondrial lineage of H. buchii. This means that H. buchii is younger and less isolated than suggested by the sequence data representing the majority mitochondrial lineage. The origin of the majority lineage remains unclear. Perhaps the inferred early branching is an artefact caused by its long branch. In any case, the closest relatives of H. buchii appear to be species from the northwest of Anatolia.
With two sympatric lineages restricted to the south-eastern Caucasus region (Azerbaijan, Armenia, adjacent Iran), the species is not a complete newcomer to the area, although it is still only a very young group in the context of the Helix phylogeny (
The geographic origin of the widespread clade 89 (see Fig.
The current distribution of different H. lucorum lineages within the species’ range is influenced by anthropogenic translocations (
It is likely that Helix buchii is not an ancient and isolated lineage of Helix in the Caucasus region, but part of a group of species that diversified in Anatolia (the ‘Anatolian clade’ of
The hypothesis of the Caucasus and the adjacent Pontus region of Turkey as a place where ancient lineages of Helix (not belonging to the European, Mediterranean or Anatolian clades of Helix) persisted for a long period of time (
We thank colleagues who provided us samples for analysis or helped with collecting, especially S. Kramarenko (Mykolayiv National Agrarian University, Ukraine), M. Son (Institute of Marine Biology NAS Ukraine, Odesa, Ukraine), E. Rybalchenko (Poltava, Ukraine) and P. Romanov (Kryvyi Rih, Ukraine). The work was funded by the Alexander von Humboldt Foundation. The publication of this article was funded by the Open Access Fund of the Leibniz Association.
List of sequences
Data type: xlsx
Explanation note: table S1. List of sequences used for intraspecific phylogenetic analyses presented in Fig.