Quercus suber Distribution Revisited

Abstract

The complex evolutionary history of Quercus suber is still under debate. Also, data and evidence at the eastern end of the species range are largely incomplete. In this study, historic floras, fossil data, and local toponyms were surveyed and genetic analyses and linguistic research used in order to point towards a previously neglected occurrence of Q. suber east of Italy. Such a multidisciplinary approach depicts a scenario in which cork oak survived in the Balkan Peninsula until recently and suggests how Q. suber might have been evicted and relegated westward, due to climate changes, ecologic competition and human impact. Our findings also suggest that the differentiation core of Q. suber was in a yet unidentified area corresponding to present-day central Europe to southwestern Asia. Radiation occurred during the Middle Miocene, with a later extension into southern Europe, Iberia and North Africa, as documented by samples collected from the late Miocene-early Pliocene. The causes of extinction in the east, however, should be investigated further and may lead to investigations about other species that might have experienced range shifts similar to that of cork oak.

Appendix 1

1.1 Transliteration from Greek to Latin

Greek	Latin	Meaning
άκυλος	akylos	acorn
αλιφλοίος	aliphloios	Q. cerris
Aντίφελλος	Antiphellos	The name of an ancient town
αρία	aria	Q. ilex
βαλάνος	balanos	Acorn
δρῦς	drys	Oak
φελλεύς	phelleus	Rocky
Φελλία	Phellia	The name of an ancient settlement
Φελλίνη	Phelline	The name of an ancient settlement
φελλόδρυς	phellodrys	Q. suber (or Q. pseudosuber)
Φελλόη	Phelloe	The name of an ancient settlement
φελλóς	phellos	Cork (cork oak)
Φελλούσα	Phellousa	The name of an ancient Island
Φελλών	Phellon	The name of an ancient settlement
Φλέω	phleo	To overflow
Φλοιός	phloios	Bark
Φολόη	Pholoe	The name of an ancient settlement
Πρίνος	prinos	Q. coccifera (Q. ilex)
Σούβουρ	Soubour	The name of an ancient settlement
Σύβαρις	Sybaris	The name of an ancient settlement
σῦφαρ	syphar	Wrinkled skin
ῦφεαρ	yphear	Viscum album

1.2 Appendix 2: Materials and Methods for the Genetic Investigations

Plant material – Plants were all sampled in the wild. Leaves were dried in silica gel, preserved as herbarium specimens and liophilised. Vouchers with institutional accession numbers were prepared for each sample and are stored at the Forest DNAbank of the University of Tuscia (Italy). DNA extractions were performed with the Dneasy Plant Minikit (QIAGEN), following the maufacturer’s instructions.

Chloroplast DNA PCR-RFLP – Chloroplast DNA was amplified with universal primers (fragments TF, CD, DT, AS, SR; Taberlet et al. 1991; Demesure et al. 1995). Amplification, digestion, and electrophoretic procedures followed Jimenez et al. (2004). Statistical parsimony was used to reconstruct phylogenetic relationships between haplotypes (TCS, version 1.21; Clement et al. 2000).

Nuclear and Chloroplast DNA sequencing – Plastid primers for matK and trnH-psbA fragments, and PCR conditions were the same as in Piredda et al. (2011); oligos for the nuclear ribosomal ITS were designed as follows: 5′-ACGACTCTCGGCAACGGATA-3′ (5,8S_Fw), 5′-CAGCGGGTAGTCCCGCCTGA-3′ (25S_Rev); thermocycling conditions were 98°C for 3 min, followed by 35 cycles of 98°C for 30 s (s), 60°C for 30 s and 72°C for 30 s, with a final extension step of 5 min at 72°C. DNAs (ca. 40 ng) were amplified with RTG PCR beads (GE Healthcare). PCR products were cleaned with the Illustra DNA Purification Kit (GE Healthcare), and standardized aliquots were then submitted to Eurofins MWG Operon (http://www.eurofinsdna.com) for sequencing. Electropherograms were edited with CHROMAS 2.3 (http://www.technelysium.com.au) and checked visually. Potential ITS pseudo-genes were filtered according to Denk and Grimm (2010). ITS boundaries were determined by comparison with Bellarosa et al. (2005). Optimal multiple alignments were obtained with CLUSTALW 1.81 (Thompson et al. 1994) and checked visually. The phylogenetic investigation was performed on both the cpDNA (joined matK + trnH-psbA sequences) and cpDNA + ITS datasets. Split networks for the ITS, CpDNA, and ITS + CpDNA data sets were generated with the Neighbor-Net method implemented in SplitsTree4 (Bryant and Moulton 2004).

1.3 References for Appendix 2

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