Molecular Phylogenetics and Evolution
Vol. 22, No. 2, February, pp. 174-183, 2002

Phylogenetic Relationships among the Species of the Genus
Testudo (Testudines: Testudinidae) Inferred from
Mitochondrial 12S rRNA Gene Sequences

Antoinette C. van der Kuyl,*,1 Donato L. Ph. Ballasina, John T. Dekker,* Jolanda Maas,* Ronald E. Willemsen, and Jaap Goudsmit*

*Department of Human Retrovirology, Academic Medical Center, University of Amsterdam, Meibergdreef 15, 1105 AZ Amsterdam, The Netherlands; and Centro CARAPAX, CP 34, 58024 Massa Marittima (GR), Italy

Received January 4, 2001, revised August 6, 2001

To test phylogenetic relationships within the genus Testudo (Testudines: Testudinidae), we have sequenced a fragment of the mitochondrial (mt) 12S rRNA gene of 98 tortoise specimens belonging to the genera Testudo, Indotestudo, and Geochelone. Maximum likelihood and neighbor-joining methods identify two main clades of Mediterranean tortoises, one composed of the species Testudo graeca, Testudo marginata, and Testudo kleinmanni and a second of Testudo hermanni, Testudo horsfieldii, and Indotestudo elongata. The first clade, but not the second, was also supported by maximum parsimony analysis. Together with the genus Geochelone, a star-like radiation of these clades was suggested, as a sister-group relationship between the two Testudo clades could not be confirmed. The intraspecies genetic variation was examined by sequencing the mt 12S rRNA fragment from 28 specimens of T. graeca and 49 specimens of T. hermanni from various geographic locations. Haplotype diversity was found to be significantly larger in T. graeca compared with T. hermanni, suggestive of reduced genetic diversity in the latter species, perhaps due to Pleistocene glaciations affecting northern and middle Europe or other sources of lineage reduction. No ancient mt 12S rRNA gene haplotypes were identified in T. graeca and/or T. hermanni originating from islands in the Mediterranean Sea, suggesting that these islands harbor tortoise populations introduced from the European and African mainland.

Key Words: Testudinidae; Mediterranean tortoises; 12S rRNA gene; phylogeny.