KARYOLOGICAL COMPARISONS OF THE EUROPEAN HARE ( LEPUS EUROPAEUS PALLAS , 1778 ) FROM THE ASIAN PART OF TURKEY , WITH MORPHOLOGICAL CONTRIBUTIONS

We focused on a comparison of karyological data for the European brown hare Lepus europaeus, from the Asian part of Turkey. In Turkish L. europaeus, the diploid number (2n), the fundamental number of chromosomal arms (FN) and the number of autosomal arms (FNa) were determined to be 2n=48, 84 and 80, respectively. The autosomes are composed of three pairs of metacentric chromosomes, four pairs of submetacentric chromosomes, ten pairs of subtelocentric chromosomes and six pairs of acrocentric chromosomes. The X chromosome was a medium-large submetacentric and the Y chromosome was a very small acrocentric. This is the third report for L. europaeus from Turkey and confirmed the previous results with regard to 2n. However, when comparing our findings with those of other authors, there were karyotypic differences among the chromosomes. These differences were related to the number of chromosome arms. Additionally, we present the skull measurements of seven samples for the Turkish brown hare collected from three localities, and these measurements conformed to those of previous studies.

Lepus europaeus Pallas 1778 was firstly defined as a subspecies of L. capensis L. 1758 and both taxa are currently accepted as two different species (Flux and Angermann, 1990;Wilson andReeder, 1993, 2005).The European hare, L. europaeus, is widely distributed throughout Palaearctic and other regions, except for Antarctica (Chapman and Flux, 1990;Burkitt, 1995;Wilson andReeder, 1993, 2005).Although there are 30 known subspecies belonging to this species today, the status of these subspecies is considered to be very variable (Flux and Angermann, 1990;Wilson andReeder, 1993, 2005).
As in all Lepus, the karyotype of L. europaeus has 48 chromosomes over this taxon's range (Flux and Angermann, 1990) and was described by Gustavsson (1971) from Sweden, Schröder et al. (1978a,b) from Finland, and different researchers (see Zima and Kral, 1984) from Germany, (former) Yugoslavia, Spain and (former) Czechoslovakia.Recently, data on the karyology of the European hare was reported by Arslan (2010) and Demirbaş et al. (2010) from Turkey, In the present study, we aimed to report on the cranial measurements and to confirm the karyotype of L. europaeus from the Asian part of Turkey.

MATERIALS AND METHODS
In this study, we examined seven samples of L. europaeus from three different localities (Kayseri, 3; Çankırı-Ilgaz, 3; Nevşehir-Gülşehir, 1) in the Asian part of Turkey (Fig. 1).All samples were obtained from hunters during legal hunting seasons in 1998-1999.Of these samples, blood samples of two individuals were collected from Kayseri and Nevşehir and used to prepare karyotypes.The blood was taken from the marginal ear vein and by cardiac puncture, and stored in heparinized tubes at +4 o C until treatment.Chromosome analysis was carried out in accordance with the method of Rooney and Czepulkowski (1987).Nomenclature for the centromeric position on chromosomes follows Levan et al. (1964): metacentric (m); submetacentric (sm); subtelocentric (st); and acrocentric (a).

Skull characteristics:
The skull measurements of Turkish brown hares are given in Table 1.The skull of Turkish brown hare is shown in Fig. 2.

Status:
In "IUCN Red List of Threatened Species", L. europaeus was evaluated to be of "Least Concern (LC)" by Smith and Johnston (2008).The population density of this species has gradually been decreasing because of extreme hunting and habitat spoiling in Turkey.Therefore, in the near future, we predict that it will be listed as "Vulnerable" for Turkey.

DISCUSSION
Lagomorph mammals known as "hares, rabbits and pikas" arose in the Paleocene (Harrison, 1972;Burkitt, 1995).The genus Lepus is represented by 11 to 15 species in Eurasia (Flux and Angermann, 1990;Wilson andReeder, 1993, 2005).The taxonomic status of these species has been discussed in detail by various authors (Riga et al., 2001;Pierpaoli et al., 1999;Suchentrunk et al., 2000).L. europaeus was previously evaluated to be a subspecies of L. capensis, a cline in morphological features over most of its range (Wilson andReeder, 1993, 2005).A cline in morphological characteristics may reflect genetic differences originating from the divergence time between two species, or adaptation to different environments (see Riga et al., 2001).According to Wilson andReeder (1993, 2005), L. europaeus is larger in terms of its size than L. capensis.This size difference was also noted by Gramov and Erbajeva (1995) who mapped the distribution of the genus Lepus in Russia.According to the distributional map of Gramov and Erbajeva (1995), L. europaeus was also distributed in Asia Minor.In this study, the skull characteristics of the Turkish samples were similar to those of the Russian L. europaeus reported by Gramov and Erbajeva (1995).
Recent molecular genetic studies, including the mtDNA control region and cytochrome b, have also maintained that L. europaeus and L. capensis are two separate species (Halanych et al., 1999;Pierpaoli et al., 1999;Slimen et al., 2006Slimen et al., , 2008)).Even though all species of the genus Lepus have the same 2n=48, the fundamental number of chromosomal arms (FN) or number of autosomal arms (FNa) and G banded patterns, are very similar.This may imply that karyotypic conservatism in the genus Lepus has occured in the course of its chromosomal evolution (see Robinson 1981in Lorenzo et al., 2003).
Karyotypes of animals that are known as chromosome complements exhibit a great diversity in their number and morphology (Ferguson-Smith and Tri- fonov, 2007).Among mammals, the diploid numbers range from 6 to 102 (see Ferguson-Smith and Tri-fonov, 2007;O'Brien et al., 2006).In all karyotyped Lepus species, they have closely similar or identical karyotypes, showing 2n = 48 (Flux and Angermann, 1990).This appears to be conserved within the genus Lepus.It is indicated by some authors that the lack of chromosomal diversity within the genus Lepus points to the mechanisms of speciation that is not valid in the chromosomal models proposed for other mammals.These researchers have stated that isolation mechanisms may be the reason for speciation within the genus Lepus (see Halanych et al., 1999) because all species of the genus Lepus have very similar karyotypes with the same diploid number of chromosomes (2n=48) (Gustavsson, 1971;see Schröder et al., 1978b; see Zima and Kral, 1984;Flux and Angermann, 1990), although there are minor differences resulting from the amount of C-heterochromatin among the karyotypes of species belonging to the genus Lepus.Centric fusion and heterochromatin addition are regarded as the causes of these differences (see Zima and Kral, 1984).
The first record on the karyotypes of European brown hare was presented by Gustavsson (1971) from Sweden.In the Swedish L. europaeus, Gustavsson (1971) described five submetacentric pairs, three metacentric pairs, nine subtelocentric pairs and six telocentric pairs of chromosomes, as well as identical results between hybrids of L. europaeus and L. timidus.Schröder et al. (1978a) compared the karyotype of European brown hare (L.europaeus) with that of the jackrabbit (L.californicus deserticola) and observed that both taxa have identical karyotypes.However, these authors stated that there are differences in terms of heterochromatin staining on two chromosomes due to paracentric inversions.Furthermore, Schröder et al., (1978b) compared karyotypes obtained from the lymphocytes of rabbit (Oryctolagus cuniculus) and two hare species (L.europaeus and L. timidus), and reported that the karyotypes of both hare species were the same, (2n=48) (except for chromosomal rearrangements; especially deletions and duplications), but differed in the rabbit (2n=44).The lower 2n in the rabbit (O.cuniculus) had resulted from two centric fusions (see Schröder et al., 1978b).In spite of great differences regarding 2n between rabbit and hares species, Schröder et al. (1978b) stated that great similarities were found in the G-banding of the chromosomes of L. europaeus, L. timidus and O. cuniculus.
It was also reported by Zima and Kral (1984) that the karyotype of L. europaeus consists of 48 chromosomes, and 3 metacentric, 5 submetacentric, 9 subtelocentric and six acrocentric pairs.According to Zima and Kral (1984), the X chromosome is submetacentric or metacentric, but the Y is a very small acrocentric chromosome.In addition, the same karyotype was described differently from country to country in Europe (Germany, Spain, (the former) Yugoslavia and (the former) Czechoslovakia) (see Zima and Kral, 1984).
In this study, the conventional karyotype of L. europaeus is presented for the third time from Turkey.In two studies by Arslan (2010 andDemirbaş et al. (2010), as expected, the 2n of brown hares occurring in Turkey was the same.However, comparing the values of the FN and FNa for Turkish brown hares, the present study had a lower number of chromosomal arms and was therefore different from previous studies (Arslan, 2010;Demirbaş et al., 2010) (Table 2).Furthermore, Demirbaş et al. (2010) reported that the karyotype of the Kilis sample had a duplication in one of the long arms in a pair of 14.
In conclusion, the karyotypes reported here and by Arslan (2010) and Demirbaş et al. (2010) may be representatives of the karyological forms of L europaeus in Turkey (Table 2); however, the differences between karyotypes of Turkish brown hare could have resulted from the classification system used by each author; they could also be a reflection of interindividual variations in the procedure used for chromosomal measurements.REFERENCES Alves, P.C., Branco, M., Matias, O., and N. Ferrand (2000).New genetic variation in European hares Lepus granatensis and L. europaeus.Biochem.Gen. Vol.38(3-4), 87-96.

Table 1 .
The skull measurements (mm) of the Turkish Lepus europaeus (N -number of specimens;, SD -standard deviation; SE -standard error).