Contribution to the vascular flora of Chalki Island (East Aegean, Greece) and biomonitoring of a local endemic taxon

Abstract: 
 The island of Chalki, located W of Rhodos, belongs to the East Aegean Islands and is situated at the E part of the South Aegean Island Arc. The flora of Chalki consists of 519 vascular plant taxa, 29 of which are under statutory protection, 22 are Greek endemics and 109 are reported here for the first time. We show that Chalki has the second highest percentage of Greek endemics in the phytogeographical region of the East Aegean Islands. The known distribution of Limonium ocymifolium, L. sitiacum and Phoenix theophrasti is expanded, being reported for the first time for the phytogeographical region of the East Aegean Islands. Finally, we assess for the first time the conservation status of Allium chalkii, a single-island endemic, by biomonitoring its populations. Citation: Tsakiri M., Kougioumoutzis K. & Iatrou G. 2016: Contribution to the vascular flora of Chalki Island (East Aegean, Greece) and biomonitoring of a local endemic taxon. – Willdenowia 46: 175–190. doi: http://dx.doi.org/10.3372/wi.46.46114 Version of record first published online on 6 April 2016 ahead of inclusion in April 2016 issue.


Introduction
With more than 7000 islands and islets (Triantis & Mylonas 2009), the Aegean archipelago is one of the largest archipelagos in the world, after the Caribbean, Indonesia and the South Pacific (Blondel & al. 2010). The Aegean has long served as a meeting ground for species of varying origin, since it lies at the crossroads of three continents, namely europe, Asia and Africa (Poulakakis & al. 2014). Its high environmental and topographical heterogeneity (Blondel & al. 2010), complex geological and palaeogeographical history, as well as its high diversity and endemism (Strid 1996) render it as an ideal stage for biodiversity studies. Intensive field work has taken place in the Aegean phytogeographical regions of greece, and numerous biodiversity and biogeographical studies have been carried out in recent years (e.g. Livaniou-Tiniakou & al. 2003;Kallimanis & al. 2010;Kagiampaki & al. 2011;Raus 2012;Trigas & al. 2013;Kougioumoutzis & al. 2014a) and more specifically in the phytogeographical region of the east Aegean Islands (e.g. Panitsa & al. 2006Constantinidis 2013), which has been characterized as one of the most floristically explored phytogeographical regions of greece (Dimopoulos & al. 2013;Bergmeier & Strid 2014). However, certain islands have not yet been the main focal areas of floristic research, leaving our knowledge of the flora of the east Aegean islands still incomplete. mid bioclimatic zone with a warm winter and also to the Thermo-mediterranean zone. Rechinger (1944) was the first botanist to report on plants from Chalki. Most records, however, were reported thereafter in the frame of the project Flora of Turkey and the East Aegean Islands (Davis 1965(Davis -1985Davis & al. 1988;güner & al. 2001), as well as by Carlström (1987). Information on some endemic and/or rare taxa occurring in the area is given by Strid & Tan (1997, 2002. From a phytogeographical point of view, however, the interesting flora of the island of Chalki has not yet received the attention it deserves in spite of these earlier records.
Finally, Allium chalkii Tzanoud. & Kollmann is confined to Chalki, and thus Chalki is the smallest island in the phytogeographical region of the east Aegean Islands that hosts at least one single-island endemic. even though A. chalkii is a very rare taxon, it has never before been monitored or evaluated under IuCn criteria.
Therefore, the scope of the present study aims at thoroughly investigating (1) the flora of Chalki; (2) whether its Fig. 1. Location of Chalki Island in relation to nearby islands and in the Se Aegean (inset). Willdenowia 46 -2016 flora is impoverished or not, compared to that of the other east Aegean islands; and (3) the conservation status of Allium chalkii with a preliminary assessment of its population size, area of occupancy and extent of occurrence.

Material and methods
Several collection and field observation trips to the study area were carried out from April 2012 to May 2014 in order to acquire an integrated knowledge of the flora of Chalki. Herbarium specimens are deposited at the Botanical Museum of the university of Patras (uPA; herbarium code according to Thiers [continuously updated]). Species identification and nomenclature are according to Davis (1965Davis ( -1985, Tutin & al. (1968Tutin & al. ( -1980Tutin & al. ( , 1993Tutin & al. ( ), greuter & al. (1984Tutin & al. ( -1989, Da vis & al. (1988), Strid & Tan (1997, 2003), güner & al. (2001, Tan & al. (2001), greuter & Raab-Straube (2008 and Dimopoulos & al. (2013). For family delimitations we follow Dimopoulos & al. (2013). The status of the alien taxa occurring in the study area is according to Arianoutsou & al. (2010) and Dimopoulos & al. (2013). The nomenclature and status of the endemic taxa recorded from Chalki are based on Dimopoulos & al. (2013). The life-form and chorological categories follow Dimopoulos & al. (2013 -see Appendix for abbreviations used). Data regarding the endemism levels in the phytogeographical region of the east Aegean Islands are based on  and Dimopoulos & al. (2013).
In order to compare the floristic diversity between islands of different size, we investigated the island speciesarea relationships (ISARs) for the native and endemic species occurring in the east Aegean islands (as in , by fitting the logarithmic transformation of the Arrhenius (1921) power model: Log(S) = c + z*Log(A), where S is the value of each of the richness metrics used, A is the area of the respective island in km 2 and c, z the fitted parameters. A value of 1 was added to certain cases before log 10 transformation, because zero values were reported for some islands. Then, we used the values of the residuals from ISAR log 10 -transfomed models of the native and endemic species occurring in the east Aegean islands, as these values are interpreted as a measure of island species diversity and do not reflect the influence of island area (Hobohm's α-index;Hobohm 2000Hobohm , 2003. Positive and negative values of Hobohm's α-index refer to areas with species diversity above and below average, respectively (Hobohm 2000(Hobohm , 2003. All the diversity analyses were carried out in the R computing environment (R Core Team 2015) using core functions and functions from the ggplot2 package (Wickham 2009).
Furthermore, extensive field research has taken place in order to locate the exact locus classicus of Allium chalkii, and to discover possible new localities and subpopulations of this taxon in the study area. We used the Technical guide for Mapping (Dafis & al. 2001) for the determination of the habitat types where A. chalkii occurs. The terms population size, subpopulation, location, area of occupancy (Aoo) and extent of occurrence (eoo) are used according to the definitions established by IuCn (IuCn Standards and Petitions Subcommittee 2010). The RAMAS Red List 3.0 software package was used for the conservation status assessment of A. chalkii according to the IuCn criteria (Akcakaya & Ferson 2007). All known localities of A. chalkii were surveyed for two consecutive years (2013 -2014). Detailed mapping was performed using a gPS device and occasionally a tape measure. Polygons of the local extent of occurrence of each subpopulation were constructed. The QgIS (QgIS Development Team 2015) software package was used for data digitization and area estimations. Population size was measured by counting every individual that was at anthesis. Finally, the direct threats in the environment where A. chalkii occurs and the stresses they cause to its individuals were recorded and classified according to the IuCn categories (IuCn & CMP 2006).

Flora
The vascular flora of Chalki comprises 519 taxa, belonging to 296 genera and 79 families (Table 1). nine alien taxa are included in the plant list, but have not been considered in the floristic analysis.
The literature survey revealed 408 bibliographical reports for the study area (Rechinger 1944;Davis 1965Davis -1986Carlström 1987;Tzanoudakis & Kollmann 1991;Strid & Tan 1997, 2002Cattaneo 2015). We report 109 taxa as new to Chalki (see Appendix). Twenty-two taxa are greek endemics, four of which are new records for the study area. eleven of the new records and 29 taxa overall are under statutory protection.
According to their general distribution, the vascular plants of Chalki can be classified into 16 main chorologi- cal categories ( Table 2). The endemic group represents 4.31 % of the total flora with 22 taxa. Phytogeographically, the endemic element is the most important group and is discussed separately. The Mediterranean chorological group predominates, highlighting the geographical position and climatic characteristics of Chalki. Within this group, the circum-Mediterranean elements are dominant, followed by the east Mediterranean taxa. The other elements are represented in lower percentages, with a relatively high portion of SW Asian elements due to the phytogeographical position of the study area. The alien flora of Chalki comprises nine taxa (1.73 %), belonging to nine genera and nine families, all reported for the first time from Chalki. eight of them are neophytes, and the most prominent among the invasive species are Nicotiana glauca graham, Opuntia ficus-indica (L.) Mill. and Oxalis pes-caprae L., which occupy large areas.

Endemism
The level of endemism in Chalki (4.31 %) is rather low compared to that in the whole of greece (22.2 % -Dimopoulos & al. 2013) but, taking into consideration the small size of the study area, the short distance from the mainland, as well as the intense human pressure present on the island (i.e. high-intensity grazing, track and road construction, discarded waste materials), this percentage is rather significant. Furthermore, compared to endemism levels of those east Aegean Islands with a larger size than that of the study area (Table 3), that of Chalki is rather high, even appearing to be the second highest in the phytogeographical region of the east Aegean Islands.
The endemic species belong to 15 families and 20 genera. Families rich in endemic species in absolute numbers are Asteraceae and Plumbaginaceae, their degree of endemism (5.48 % and 75.00 %, respectively) being higher than that of the general flora (4.31 %). These results agree with the trend observed in the whole greek endemic flora (georghiou & Delipetrou 2010).
Among the 22 greek endemic taxa (Table 4), there are   A very interesting record is Phoenix theophrasti greuter, a taxon known to occur in the Aegean region, mainly in Kriti, as well as in three areas of SW Turkey (Boydak & Barrow 1995). We believe its occurrence on Chalki to be of native origin, as its individuals were found to occupy similar ecological niches as in the Cretan locus classicus (stream banks near sea level), in two different localities. Thus, P. theophrasti is recorded as an indigenous member of the flora of the east Aegean Islands, whereas previous reports from Kalymnos, Kos, nisyros and Symi have not yet been confirmed (Turland & al. 1993: 194;Thymakis 2009;Dimopoulos & al. 2013). Phoenix theophrasti has also been recorded from a single locality in Peloponnisos Willdenowia 46 -2016 (Thymakis 2009) and in the phytogeographical region of the Kiklades on Milos (Kalheber in Raus 2012), Anafi (Kougioumoutzis & al. 2012) and Amorgos (Biel & Tan 2015). The presence of P. theophrasti on Chalki and probably other east Aegean Islands bridges the range gap between Kriti and SW Turkey.

Plant diversity of Chalki Island
using Hobohm's α-index (Table 5) for the greek native and endemic species occurring in 20 east Aegean islands, Chalki presents values well above average for both of these categories (0.74 for native, 1.61 for endemic). Lesvos could be regarded as floristically impoverished, as it presents very low α-values for both the native and endemic species. Chalki, Ikaria and Samos have high α-values for both native and endemic species (Fig. 2), while more than half of the east Aegean islands display low α-values for both species-richness metrics. our results indicate that Chalki, Ikaria and Samos in fact constitute plant diversity hotspots, whereas islands with a large total number of native and greek endemic species, such as Chios and Lesvos, are not characterized as such. Rhodos seems to be an endemic plant diversity hotspot in the phytogeographical region of the east Aegean Islands.

Preliminary assessment and conservation status of Allium chalkii
Allium chalkii is an erect, bulbous geophyte up to 8 cm tall, with membranous outer bulb tunics, bearing 2 or 3 glabrous leaves, a persistent, erect, sometimes divaricate spathe and a fastigiate, few-flowered inflorescence. The ovary is ellipsoid to oblong-ellipsoid and the capsule is globose, 3-valved, c. 3.5 mm in diam., containing black, 2 -2.5 mm-long seeds. Allium chalkii grows in calcareous, rocky and stony crevices at 100 -350 m a.s.l. and flowers from April to June (Fig. 3). Historical knowledge about this single-island endemic is scarce: A. chalkii was described in 1991 (Tzanoudakis & Kollmann 1991) from specimens collected in the frame of the project Flora Hellenica from several localities between Chorio and Kastro settlements (locus classicus) in 1986 by g. Iatrou and Th. georgiadis and in rocky, calcareous escarpments on Chalki in 1988 by e. Karavokyrou.
Allium chalkii has not been evaluated before and its eoo, Aoo and population size are reported here for the first time.
The current and past distribution of Allium chalkii is shown in Fig. 4 and Table 6. The entire population of the species consists of three subpopulations (AC1, AC2

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Tsakiri & al.: Vascular flora of Chalki Island and biomonitoring a local endemic taxon and AC3), corresponding to the localities Lendaki, Mesovounia and Petrolakki for AC1, Agia Varvara and Kakoskali for AC2, and finally Ais giorgis tou Riakiou for AC3. The area of the locus classicus was searched repeatedly (2012 -2014) and exhaustively, but no plants were found. The fact that A. chalkii has not been reported from its locus classicus since 1988, suggests that this subpopulation may be extinct.
All subpopulations are scattered in seven different locations (Fig. 4, Table 6). During the two years of monitoring (2013 -2014), both the eoo and the Aoo -based on a 1 × 1 km grid -were estimated at 5 km 2 . The whole population local extent was 225 m 2 in 2013 and 250 m 2 in 2014.
Population size, expressed as the total number of mature individuals in all subpopulations, as well as the size of each subpopulation, exhibited a slight annual increase in 2014 compared to 2013 (Table 6). Total population size, local extent and the number of plants per m 2 (a rough estimation of plant density) were highest in 2014 in subpopulation AC1.
Allium chalkii occurs in stony or rocky meadows with typical Mediterranean phry gana, dominated by Sarcopoterium spinosum (L.) Spach and other spiny and aromatic shrubs, as well as in steep, calcareous cliffs near the sea, at 100 -270 m a.s.l. Allium chalkii does not seem to face any imminent and direct threats, as its occurrence inside spiny phrygana and on precipitous cliffs protects it from grazing and trampling. However, habitat degradation and/or loss due to excessive grazing (stress codes 1.3 & 2.2) and road construction works (stress code 4.1), may constitute a potential threat for this species in the near future.
Allium chalkii is suggested to be classified as endangered (en), according to the IuCn (2012) criterion D, since its total population consists of fewer than 250 mature individuals.
According to Arianoutsou & al. (2010), the total number of alien taxa accounts for c. 5 % of the native flora of greece, and this percentage is significantly higher than that of Chalki (1.54 %). nevertheless, on Chalki, where grazing grounds and abandoned farm lands occupy large areas, Nicotiana glauca, Opuntia ficus -indica and Oxalis pes-caprae have heavily contaminated and altered these habitats, which would otherwise be colonized by native pioneer herbs and shrubs. This phenomenon is also observed in other Aegean islands (Arianoutsou & al. 2010;Kougioumoutzis & al. 2012Kougioumoutzis & al. , 2014bKougioumoutzis & al. , 2015. The high percentages of chamaephytes and hemicryptophytes depend on the frequency of limestone cliffs, which very often harbour endemic taxa (Kypriotakis 1998;Kypriotakis & Tzanoudakis 2001;Tzanoudakis & al. 2006). In Chalki, the majority (63.6 %) of the en demic plants are chamaephytes or hemicryptophytes, which are scattered in the numerous steep calcareous cliffs and crevices present on the island. Chalki is floristically more diverse than the other parts of the phytogeographical region of the east Aegean Islands, probably because of the increased habitat diversity due to its great local topographic and geological heterogeneity (intense hilly relief with many different inclinations and exposures, and numerous geological substrates of different age), factors known to promote species richness (Whittaker & Fernández-Palacios 2007;Sfenthourakis & Triantis 2009;. Its rather high level of endemism is due to its palaeogeographical history, as Chalki, contrary to the majority of the east Aegean islands (Parmakelis & al. 2006;Triantis & al. 2008), has been isolated from the adjacent mainland (Asia Minor) for the last 0.8 My (Perissoratis & Conispoliatis 2003). our results are in accordance with Sfenthourakis & Panitsa (2011), who stated that in small Aegean islands diversity at the whole-island scale is shaped mainly by heterogeneity among local communities. According to Hobohm's index, Chalki seems to be a plantdiversity hotspot in the phytogeographical region of the east Aegean Islands, since it has a higher Ho bohm's α-index value than Ikaria and Rhodos, two islands considered highly important in terms of phytogeography and endemism (Carlström 1986;Christodoulakis 1996b).
The palaeogeographical events that shaped the presentday South Aegean region are reflected in the occurrence of several taxonomically isolated and relict species in the flora of Chalki. Among these, Allium chalkii is the most prominent representative, since it constitutes a very rare, ancient, geographically isolated, single-island endemic confined to refuge habitats and considered as a member of the Tertiary Aegean flora because it bears some primitive morphological features (i.e. erect-divaricate-spathe, fewflowered inflorescence; Tzanoudakis & Kollmann 1991;Brullo & al. 2001). Arenaria aegaea Rech. f., Asyneuma giganteum (Boiss.) Bornm., Centaurea lactucifolia Boiss. and Umbilicus albido-opacus Carlström are probably Pliocene relicts (Carlström 1986), while the occurrence of Lomelosia variifolia (Boiss.) greuter & Burdet on Karpathos, Chalki and Rhodos seems to reflect the South Aegean palaeogeography during the early Pliocene, since Karpathos was joined with Chalki, Rhodos and Anatolia at that time (Daams & van der Weerd 1980). The distributions of Colchicum macrophyllum B. L. Burtt and Valeriana asarifolia Dufr. reflect the land-bridge connection that existed in the South Aegean during the Messinian salinity crisis (Carlström 1986;Schüle 1993;Cattaneo 2015): the former can be found on Kriti, Karpathos, Chalki and Rhodos, extending to SW Anatolia, while the latter is a South Aegean Island Arc endemic, occurring on Antikythira, Kriti, Karpathos and Chalki. on the other hand, Campanula rhodensis A. DC., Dianthus fruticosus subsp. rhodius (Rech. f.) Runemark, Nigella arvensis subsp. brevifolia Strid and Symphytum circinale may have resulted from random genetic drift during the climatic fluctuations and sea-level oscillations of the Pleistocene (Carlström 1986;Comes & al. 2008).
nearly half (nine) of the endemic taxa found on Chalki correspond to one or two phytogeographical areas (Table 4), thus providing valuable information regarding the phytogeographical position of the study area, as the existence of biregional endemics is a good indication of phytogeographical connections between regions (georghiou & Delipetrou 2010). It would be expected that Chalki shows higher affinities with the phytogeographical area of the Kiklades (Kik) since, according to georghiou & Delipetrou (2010), the phytogeographical area of the east Aegean Islands (eAe) is chorologically closer connected to Kik than to that of Kriti and Karpathos (KK). While this may be true for the majority of the east Aegean Islands, our results demonstrate that Chalki is phytogeographically closer to KK, as we recorded three endemic taxa (Asyneuma giganteum, Limonium sitiacum and Lomelosia variifolia) occurring exclusively in KK and eAe and only one taxon (Limonium ocymifolium) occurring exclusively in Kik and eAe. More specifically, A. giganteum and L. variifolia are found only on Karpathos, Chalki and Rhodos, while L. sitiacum occurs only  (Carlström 1986). This phenomenon could possibly be attributed to (1) the close palaeogeographical distance between these islands and Kriti and Karpathos during the Messinian salinity crisis (Hsü 1972), as well as during the Pliocene (Creutzburg 1963;greuter 1970;Daams & van der Weerd 1980;Wiedenbein 1991c;Lymberakis & Poulakakis 2010) and the Pleistocene (Dermitzakis & de Vos 1987); and (2) the South Aegean Island Arc (of which Chalki is a part) being isolated from the rest of the Aegean region by extensive saline deserts or even saline/hypersaline lakes during the Messinian salinity crisis, even though dispersal through land was feasible at that time (Poulakakis & al. 2014). Five very narrowly distributed taxa are present on Chalki (Allium chalkii, Campanula rhodensis, Centaurea lactucifolia, Dianthus fruticosus subsp. rhodius and Umbilicus albido-opacus), with C. lactucifolia being the most variable taxon, especially in terms of leaf and involucral appendage shape; it was once divided by Wagenitz (1975) into two varieties, var. lactucifolia and var. halkensis (Fors.-Major & Barbey) Wagenitz. Chalki is floristically more closely connected with Rhodos, since all of these taxa -except for A. chalkii -also occur on Rhodos. This phenomenon can be attributed to the two islands having been connected until the offset of the last glacial maximum (Lykousis 2009).
Chalki also seems to have close phytogeographical affinities with the Muğla province of Turkey, and more specifically, with the Marmaris (Datça) peninsula, since virtually all (apart from Ophrys speculum subsp. regisferdinandii) of its range-restricted east Mediterranean taxa occur also there. This phenomenon can be attributed to the cliff flora of the outer Marmaris (Datça) peninsula mainly resembling a slightly impoverished Aegean flora, since it hosts several typical Aegean species, such as Campanula hagielia, Symphytum circinale and Verbascum propontideum (Carlström 1986).
Finally, regarding the conservation status of Allium chalkii, its known population size and density and its area of occupancy were increased during the two years of monitoring, although the population at the type locality was apparently lost. Habitat loss due to human-induced activities constitutes the main threat faced by this taxon. ex situ conservation seems to be an appropriate measure for A. chalkii, while an effort should be made to minimize the direct threats it is facing by informing the local residents about its rarity and importance.