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BMC Evolutionary Biology 2008, 8:56                            http://www.biomedcentral.com/1471-2148/8/56




            record [22]. So far, no phylogeographic signature of the  Bufo siculus n. sp
            Pliocene subdivision  of  Sicily into two islands was  Holotype
            detected. The effective population size of mtDNA in Bufo  An adult female, P215 (Museum of Terrasini, Palermo,
            siculus n.sp., as inferred from θ (Fluctuate) is small (Table  Figure  5a, c), collected November, 17, 2006, by M. Lo
            2).                                                 Valvo,  Monte  Pellegrino reserve (38.170 N, 13.351 E),
                                                                Palermo, Sicily, Italy; for nomenclature and morphologi-
            The distribution analysis indicated an observed distribu-  cal description: Additional files 2 and 4.
            tion in the B. siculus clade that was unimodal and visually
            congruent with the distribution of expected values for a  Paratypes
            sudden population expansion, although not quite statisti-  ZFMK 85896 (juv.,  topotypic, M. Lo  Valvo  leg.  2005),
            cally significant (Figure 3f, p = 0.077). The distribution for  ZFMK 85778 (ad. male, Sicily, Monte Carbonara region,
            this clade is extremely compressed to the left, indicating  W. Haas leg., ca. 1991, GenBank EU497506), ZMB 69556
            very low numbers of mismatches and low variance. This  (juv. female, EU497501) and MVZ 250741 to MVZ
            profile may  indicate an even more recent expansion in  250743 (two juv. and one subad. female, EU497502 to
            this group than in the mainland B. balearicus group. The  EU497504):  Italy, southeastern Sicily,  mouth of San
            Fluctuate analysis also did not statistically permit rejection  Leonardo River, M. Lo Valvo leg. 2005; MZPA A95 (adult
            of a no-growth scenario for this clade, and Tajima's D did  female, La Fossa, NW  Sicily, A. Sicilia leg. 2006,
            not fit the expectation for an expanded population.  EU497598); NME A 1490/08 (adult male, La Fossa, NW
                                                                Sicily, F. Marrone leg.). For morphometric details of two
            However, the high similarity among mtDNA haplotypes  adult paratypes see Table 5 in Additional file 4.
            across the entire island is consistent with the known high
            mobility of green toads,  exemplified  by fast post-Pleis-  Diagnosis
            tocene re-colonization of northern Europe by two haplo-  A medium- to large-sized green toad that differs from all
            type groups [28]. The current range of B. siculus appears to  other, especially all circum-Mediterranean green toad spe-
            follow a pattern known in another Sicilian endemic, the  cies, by its distinct mitochondrial haplotype group. Bufo
            lizard Podarcis wagleriana, that is widespread across Sicily  siculus exhibits strong variability in coloration with adult
            but absent in the very northeast [42,73].

            Our data reveal taxonomic identity (B. siculus) of toads on
            Favignana Island (loc. 25) and Ustica Island (loc. 24);
            green toads on the Aeolian Islands may be members of the
            geographically proximate B. balearicus (see Additional file
            3 for further comments on Circum-Sicilian islands).
            Detection of Calabrian B. balearicus haplotypes in north-
            eastern Sicily (loc. 20) suggests their relatively recent inva-
            sion; either during  Pleistocene sea level lows [72],  by
            rafting across the narrow Strait of Messina, or possibly by
            human introduction. It remains unknown whether both
            taxa (B. balearicus  and  B. siculus) occur in sympatry  in
            northern Sicily as range maps suggest [42], and whether
            they would hybridize, given the considerable mtDNA and
            breeding-phenology differences.  Bioacoustic data in  the
            potential contact zone will  also help to illuminate this
            question.

            Description of a new species
            Mitochondrial and nuclear, morphometric and other pre-
            liminary biological data show that green toads from most
                                                                speci
                                                                Figure 5
                                                                Holotype of Bufo siculus n.sp. and chromosomes of the new es
            of Sicily represent a separate lineage. Sicilian green toads  Holotype of Bufo siculus n.sp. and chromosomes of
            have been physically separated by the Strait of Sicily from
                                                                the new species. (a, c) Holotype of Bufo siculus n.sp. (P215,
            their closest African relatives for long evolutionary peri-  Museum of Terrasini, Palermo), adult female, in life (Photo:
            ods. In order to acknowledge these facts, and to raise the  M. Lo Valvo). (b) Somatic metaphase from a paratype (MVZ
            potential conservation status of this form, which repre-  250742), southeastern Sicily, San Leonardo River, with 2n =
            sents an island endemic, we hereby describe it as a new  22 chromosomes (Photo: M. Stöck).
            species.

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