Technical guidelines for genetic conservation and use Holm oak Quercus ilex Quercus ilex Quercus ilex Querc Bartolomeo Schirone1, Federico Vessella1 and Maria Carolina Varela2 1 University of Tuscia, Viterbo, Italy 2 National Institute for Biologic Resources, Forest Unit, Oeiras, Portugal

These Technical Guidelines are intended to assist those who cherish the valuable holm oak genepool and its inheritance, through conserving valuable seed sources or use in practical forestry. The focus is on conserving the genetic diversity of the species at the European scale. The recommendations provided in this module should be regarded as a commonly agreed basis to be complemented and further developed in local, national or regional conditions. The Guidelines are based on the available knowledge of the species and on widely accepted methods for the conservation of forest genetic resources.

Biology and ecology

Holm oak (Quercus ilex L.) is a monoecious, evergreen, wind- pollinated oak tree. It is a medi- um-sized tree (20–27 m tall), with finely square-fissured, blackish bark. The leaves are dark green above, and pale whitish-grey with dense short hairs under- neath. The leaf shape is variable (4–10 cm long and 1–3 cm wide). The acorns are dark brown, pe- dunculated, and held by cupulae with short, tight scales. The mat- uration is annual, but in some areas more flowerings in a year are possible. Q. ilex is polymorphous, but it exhib- its two main formae ranked at the subspecies level: Q. ilex subsp. ilex, and Q. ilex subsp. rotundifolia. There are several known natural hy- brids: Q. x morisii Borzì (with Q. suber), Q. x turn- eri Willd. (with Q. robur), Q. audleyensis Henry (with Q. pe- traea), Q. albescens Rouy (with Q. pubescens), Q. auzendi Green and Godr. (with Q. coccifera). HolmQuercus oakQuercus ilex Holm oakQuercus ilex Holm oakilexQuercus ilex Holm Quercus oakQuercus ilex Holm oakQuercus ilex Holm oakilexQuercus ilex Holm QuercusoakQuercus ilex Holm oakQuercus ilex Holm oak Quercusilex ilex Holm oakQuercusQuercus ilex Holm oakQuercus ilex Holm oak Quercusilex ilex Holm

Q. ilex is a very drought tol- Distribution Importance and use erant species among the ever- green Mediterranean oaks. Its ecological plasticity al- Holm oak shows a circum- Holm oak forests play a very lows it to thrive in almost all bio- Mediterranean range, absent important socio-economic role, climates, except extremely arid only in Egypt. It is more abun- owing to the diversification of ones, where annual precipitation dant in the western regions, products: wood, environmental is under 450 mm. It is highly where it forms large pure stands protection, tourism and forage termophilous, but it can resist (Morocco, the Iberian peninsula, resources. winter cold (as low as -20°C). the Atlantic and Mediterranean The ability to survive in re- Holm oak is indifferent to soils, coasts of France, the Italian pen- gions of extreme climates with but the best stations are found insula, the main Mediterranean cold winters and very hot and on calcareous brown or well- islands), than in the east, where dry summers, together with a drained soils. it can more usually be found in remarkable resistance to fire, It can be found from sea mixed stands (Balkan coasts, makes holm oak a key eco- level to 100–140 m above sea Greece, Crete, northern Anatolia, logical species virtually without level in the Black Sea area, up Black Sea and northern Leba- alternative in the harsh inland re- to 400-600 m in the Mediter- non). In Africa there are scat- gions of Portugal. Substitutions ranean, while on the Moroccan tered putative indigenous occur- tend to fail both on physiologi- Reef, it grows up to altitudes of rences along the Libyan coast cal and on economic aspects. 2000–2600 m. up to Cyrenaica. The northern- Where pines are used to replace When climatic conditions are most limit of the range is at the holm oak, fire risk increases in favourable, Q. ilex is strongly Gironde estuary; in the east it these areas. competitive and forms long-last- reaches the Turkish shores of the Holm oak forests cover ing, pure stands. These forests Aegean Sea and the Black Sea about 4,000,000 ha in Spain, are now becoming rare, mostly provinces of Zonguldak, Sinop and are mainly exploited in a sil- as a result of human and Samsun. Nowhere does it vo-pastoral regime for pig graz- activities over cen- penetrate inland. There ing. From the free-range black turies. are scattered occur- pigs that feed on acorns, the ex- rences in Cyprus and quisite delicacy jamon de bellota Lebanon. In Italy, Q. is produced, playing a major role ilex is absent in the rural economy in various in the Padan regions. The silvo-pastoral re- Plain, except gime for animal grazing benefit- for some iso- ing from the nutritious holm oak lated stands acorns is practised also in other around the countries, though on a smaller big pre-Al- scale than in Spain. pine lakes. The wood is hard and tough and has been used since ancient times for general construction purposes, such as pillars, tools, wagons, vessels and wine casks. It is also used as firewood, or for charcoal production. HolmQuercus oakQuercus ilex Holm oakQuercus ilex Holm oakilexQuercus ilex Holm Quercus oakQuercus ilex Holm oakQuercus ilex Holm oakilexQuercus ilex Holm QuercusoakQuercus ilex Holm oakQuercus ilex Holm oak Quercusilex ilex Holm oakQuercusQuercus ilex Holm oakQuercus ilex Holm oak Quercusilex ilex Holm

Holm oak is one of the top Genetic knowledge typic diversity. The analysis of three tree species used in the nuclear markers (isozymes) over establishment of truffle orchards, the whole species distribution or trufferies. Truffles grow in an The karyotype of Q. ilex con- revealed that some Sicilian pop- ectomycorrhizal association with sists of 18 metacentric, 4 sub- ulations are characterized by the the tree's roots. metacentric and 2 subtelocentric occurrence of private alleles. The acorns, like those of the chromosomes. Pair 1 shows a The reproductive profile is cork oak, are edible (toasted, as secondary constriction on the rather regular and balanced and a flour or boiled in water). When long arm; pair 2 bears a satel- the juvenile stage is much short- boiled they can also be used as a lite on the short arm. Holm oak er than in other oaks. This means medicinal treatment for disinfec- shows a different karyomorphol- that already at young age trees tion of wounds. The tree can be ogy from other oak species, but in a stand contribute to a high clipped to form a tall hedge, and is similar to Q. coccifera, thus effective population size of these it is suitable for coastal wind- confirming its belonging to the stands. This means good per- breaks, in any well drained soil. same subgenus Sclerophyllo- spectives for gene conservation It forms a picturesque rounded drys (D’Emerico et al. 2000). whenever natural regeneration is head, with pendulous low-hang- Lumaret et al. (2002) ana- encouraged (Varela, et al. 2008). ing branches. Its size and solid lyzed the variation of chloroplast evergreen character give it an DNA in Q. ilex over its imposing architectural presence whole distribution that makes it valuable in many range. urban and garden settings. Five macro-regions . of diversity were identified: (1) the Aegean peninsula and Crete, (2) the Italian peninsula, (3) North Africa, (4) eastern Iberia and France, (5) western Iberia and France. These results indicate that post-glacial recolonization probably started from the three Mediterranean peninsu- las. Italy and south-east- ern France appear to have been colonized by one major haplotype, most probably of Ital- ian origin. Additional haplotypes are rarely detected in continental Italy and on the islands of Cor- sica and Sicily, and few Balkan haplotypes are detected along the Italian Adriatic coast. Fin- eschi et al. (2005) concluded that Sicily represents the Italian reservoir for holm oak haplo- HolmQuercus oakQuercus ilex Holm oakQuercus ilex Holm oakilexQuercus ilex Holm Quercus oakQuercus ilex Holm oakQuercus ilex Holm oakilexQuercus ilex Holm QuercusoakQuercus ilex Holm oakQuercus ilex Holm oak Quercusilex ilex Holm oakQuercusQuercus ilex Holm oakQuercus ilex Holm oak Quercusilex ilex Holm

Threats to ing practices, in some zones Guidelines for genetic natural regeneration is poor, genetic diversity conservation and use thus creating a highly skewed Holm oak is affected by sev- age distribution towards old Seed propagation for high eral anthropogenic pressures trees. The combined effect of forests is the most suitable that could lead to a loss of old trees and cloned trees may management method to main- genetic diversity. Loss and/or be driving threats to the genetic tain and increase genetic diver- fragmentation of habitats by in- resources of the species, es- sity and, therefore, the conser- tense human impacts on forest pecially in particular ecological vation of the genetic resourc- and agricultural landscapes has conditions where local adap- es of the species. It is the been recorded for centuries in tation may have occurred most adequate regime for areas like Sicily. Replacement during evolution. landscape protection, or- with fast-growing species or Guidelines for spe- namental and recreational clear cuttings for tourist facili- cific management un- functionalities. High forest ties such as golf courses is also der coppice should merits are also on the use threatening the holm oak forests be part of the forest of water, which is a special in some parts of Portugal and management of the spe- feature for Mediterranean Spain. Indiscriminate cuttings cies. regions where increasing hu- and recurrent fires have mainly Holm oak stands man demands for water are affected all the Mediterranean are today often still expected together with cli- coastal stands, while overgraz- managed as coppice, mate change induced drought ing (especially during the re- sometimes with (Gracia, 2009). generation period) is particularly short cycles of Coppices for firewood pro- present in the Iberian peninsula. less than 20 duction (coppicing every 30– In addition, natural enemies, years. This is 40 years) usually host a large such as fungi and insects, have because the amount of the biodiversity of affected holm oak stands, in holm oak wood Mediterranean ecosystems, but particular in urban areas (central technological reduce the genetic diversity of and southern Italy). Nowadays, properties are not the coppiced species, in par- in the central-eastern Medi- as good as other ticular where up growing single terranean Basin (e.g. Italy and deciduous oaks stems per stump are not left. Dalmatia), holm oak, like other (e.g. Q. robur, Q. Therefore, the chance to con- oaks, is naturally prone to veg- petraea, Q. frainetto), vert coppices into high forests, etative sprouting from stumps which produce high through a gradual increase of or roots of dead or fallen trees. quality logs. Therefore, the number of single stems per Forest owners, especially when a considerable loss of intraspecific stump, should be considered. attempting to propagate inter- genetic diversity is observed in For instance, a coppice stand esting phenotypes, frequently holm oak, because adequate with a single stem per stump use this methodology, which high-forest management models system, with long cycles (not results in the cloning of these in which much longer cutting less than 40 to 50 years) and a trees. This leads to a decreased cycles take place, have not been large number of seed-bearing genetic diversity. developed yet. These models trees (even >200), contributes Adding to the potential re- could theoretically permit better to the regeneration of the popu- duction of genetic variability and conservation of the species lation and the maintenance of inbreeding from empirical clon- diversity. the understory and favours the QuercusHolm oakQuercus ilex Holm oakQuercus ilex Holm ilex oakQuercus ilex HolmQuercus oakQuercus ilex Holm oakQuercus ilex Holm ilexoakQuercus ilex HolmQuerc oakQuercus ilex Holm oakQue

Distribution range of holm oak

natural evolution of Q. ilex to- for conservation of genetic with the following objectives: wards its climax situation (which resources are established in conservation of endangered, is pure high forest). mixed oak- stands, hybridiza- marginal populations and habi- Holm oak hybridizes with tion monitoring is recommend- tats of Q. ilex; sampling the other oak species and hybridi- ed, whenever possible through genetic diversity; establishment zation may result in a loss or genomics, transcriptomics and of Dynamic Conservation Units an increase of genetic diver- progeny testing. based on long term autochtho- sity according to the concrete Since limited genetic infor- ny, high biodiversity value and situation of the population or mation about Q. ilex is available, location in ecologically diverse species evolution (Soltis & Solt- it is recommended that genetic regions of large populations is, 2009). Where populations conservation programmes start (> 1000 individuals). QuercusHolm oakQuercus ilex Holm oakQuercus ilex Holm ilex oakQuercus ilex HolmQuercus oakQuercus ilex Holm oakQuercus ilex Holm ilexoakQuercus ilex HolmQuerc oakQuercus ilex Holm oakQue

The series of these Technical Selected bibliography Guidelines and the distribution maps were produced by mem- bers of the EUFORGEN Net- D’Emerico S., Paciolla C. & Tommasi F., 2000. Contribution to the karyo- morphology of some species of genus Quercus. Silvae Genetica, 49, 6: works. The objective is to iden- 243–245. tify minimum requirements for Fineschi, S., Cozzolino, S., Migliaccio, M., Musacchio, A., Innocenti, M. & long-term genetic conservation Vendramin, G.G., 2005. Sicily represents the Italian reservoir of chloroplast in Europe, in order to reduce the DNA diversity of Quercus ilex L. (Fagaceae). Annals of Forest Science, 62: overall conservation cost and to 79-84. improve the quality of standards Gracia, C. 2009. Climate change, observed and expected trends and impact on Mediterranean forest ecosystems functioning. Communication at in each country. Regional Workshop Mediterranean Forest Genetic Resources and Climate Change, 24 – 26 November 2009, Chania, Crete, Greece. Lumaret, R., Mir, C., Michaud H. & Raynal, V., 2002. Phylogeographical varia- Citation: B. Schirone, F. Vessella tion of chloroplast DNA in holm oak (Quercus ilex L.). Molecular Ecology, and M.C. Varela. 2019. EUFOR­ 11: 2327-2336 GEN Technical Guidelines for Soltis P.S. & Soltis D.E. 2009. The Role of Hybridization in Plant Speciation. Annual Review of Plant Biology 60: 561-588. DOI: 10.1146/annurev. genetic conservation and use for arplant.043008.092039 Holm oak (Quercus ilex). Euro- Varela M.C., Brás R., Isabel R. Barros, Oliveira P. & Meierrose C., 2008. pean Forest Genetic Resources Opportunity for hybridization between two oak species in mixed stands as Programme (EUFOR­GEN), Euro- monitored by the timing and intensity of pollen production. Forest Ecology pean Forest Institute. 6 pages. and Management, 256 (8): 1546-1551. The distribution map is based on: Caudullo, G., Welk, E., San-Miguel-Ayanz, J., 2017. Chorological maps for the main European woody species. Data in Brief 12, 662-666. https://doi.org/10.1016/j.dib.2017.05.007 Drawings: Quercus ilex, Giovan- na Bernetti.

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