The genus Ligidium Brandt, 1833, with 47 described species, is distributed in the northern hemisphere from West Europe to Japan and North America (Schmalfuss, 2003). Eight are endemic to Turkey, with ﬁve species restricted to the northwest of the country, six species are endemic to Greece, one is endemic to Romania, another to Bosnia, and two to the rest of Europe. The taxonomy within the genus has been based on the form of the uropods (relative length of exo- and endopodites), the shape and the number of setae on the ﬁrst and the second male pleopod exopodites, and the structure of the apex of the second male pleopod-endopodite. Sfenthourakis (1993) argued that at least for the species distributed in Greece, most of these characters exhibit high intraspeciﬁc variation, and may therefore be of limited taxonomic value. The apex of the second male pleopod endopodite, an important diagnostic character, is variable. In addition, its fragile structure is easily destructed, a fact that may cause inaccurate description of its character states due to the effects of malformations, improper handling etc. As a consequence, the actual taxonomic status of Ligidium populations, especially those distributed in southern Greece, remains unclear. Furthermore, it is likely that several of the species described from the Balkans and Asia Minor are synonyms, and probably identical with some broadly distributed European species. In order to shed light on the phylogenetic relationships of populations and resolve taxonomic uncertainties of the genus, we used a molecular approach that provides additional characters and a better insight to the processes of differentiation between populations. Molecular data have been sparsely used in terrestrial isopod phylogenetics (Marcadé et al., 1999; Wetzer, 2001, 2002; Charﬁ-Cheikhrouha, 2003; Mattern, 2003). Molecular data (mtDNA) from selected Ligidium species have been used in analyses with a broader taxonomic context (Michel-Salzat and Bouchon, 2000; Mattern and Schlegel, 2001; Mattern, 2003), or serving as outgroup taxa for analysis of a related genus (Taiti et al., 2003). One of these species (L. germanicum) has been reported from Greece (Schmalfuss, 1979), but the validity of the record was questioned by Sfenthourakis (1993).
All species of the genus are hygrophilic, living in wet habitats such as the dense leaf-litter layer on riverbanks. In Greece, Ligidium is found in riparian habitats, in the wet leaf-litter layer within a narrow zone (< 3 m) along permanent-ﬂow streams, and in the mosses around springs and waterfalls. Thus, their populations are very sensitive to ﬂuctuations in water ﬂow caused by exploitation of freshwater resources for irrigation and other uses by humans. This restriction to semi-isolated habitats may have led, at least in the drier areas of Ligidium’s distribution, to a pronounced metapopulational structure with limited gene ﬂow among local populations, especially between those belonging to different river systems.
The species known from Greece are L. beieri Strouhal, 1928, L. cycladicum Matsakis, 1979, L. euboicum Matsakis, 1975, L. germanicum Verhoeff, 1901, L. ghigii Arcangeli, 1928, L. mylonasi Sfenthourakis, 1992, and L. werneri Strouhal, 1937. All
Fig. 1. Map of central Greece indicating collecting sites (dots). Evvoia Island and the semi-isolated region of Peloponnisos are indicated in bold characters.
are endemic, except L. germanicum , which is widely distributed in central- eastern Europe.
As a ﬁrst step in our ongoing research project, we used nine populations from Greece and two from central Europe, we analyzed two mitochondrial gene segments of the 12S - and 16S rRNA subunits, and studied the intra- and interpopulation genetic variation using RLFP analysis. Herein, we focus mainly on the populations from the almost insular region of Peloponnisos (today it is separated from the Greek mainland by the isthmus of Korinthos), since these are the most difﬁcult to identify and they live in a piece of land that has gone through several events of isolation and reconnection to the Greek mainland (Creutzburg, 1963; Meulenkamp, 1985; Dermitzakis, 1990). According to Sfenthourakis (1993), the populations of Peloponnisos belong to L. euboicum , since male specimens seem to have the same genitalia morphology with the latter.