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<front>
<journal-meta>
<journal-id journal-id-type="publisher-id">AJBM</journal-id>
<journal-title-group>
<journal-title>Anales del Jard&#x00ED;n Bot&#x00E1;nico de Madrid</journal-title>
</journal-title-group>
<issn pub-type="ppub">0211-1322</issn>
<issn pub-type="epub">1988-3196</issn>
<publisher>
<publisher-name>Consejo Superior de Investigaciones Cientificas</publisher-name>
</publisher>
</journal-meta>
<article-meta>
<article-id pub-id-type="publisher-id">AJBM201710-2467</article-id>
<article-id pub-id-type="doi">10.3989/ajbm.2467</article-id>
<article-categories>
<subj-group subj-group-type="heading">
<subject>Articles</subject>
</subj-group>
</article-categories>
<title-group>
<article-title><italic>Delphinium fissum</italic> subsp. <italic>sordidum</italic> (Ranunculaceae) in Portugal: distribution and conservation status</article-title>
<trans-title-group xml:lang="es">
<trans-title><italic>Delphinium fissum</italic> subsp. <italic>sordidum</italic> (Ranunculaceae) en Portugal: distribuci&#x00F3;n y estatus de conservaci&#x00F3;n</trans-title>
</trans-title-group>
<alt-title alt-title-type="running-head"><italic>Delphinium fissum</italic> subsp. <italic>sordidum</italic> in Portugal</alt-title>
</title-group>
<contrib-group>
<contrib contrib-type="author" corresp="yes">
<name>
<surname>Ram&#x00ED;rez-Rodr&#x00ED;guez</surname>
<given-names>Rub&#x00E9;n</given-names>
</name>
<xref ref-type="aff" rid="aff0001">1</xref>
<xref ref-type="corresp" rid="cor1">&#x002A;</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Medina</surname>
<given-names>Leopoldo</given-names>
</name>
<xref ref-type="aff" rid="aff0002">2</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>de Sequeira</surname>
<given-names>Miguel Menezes</given-names>
</name>
<xref ref-type="aff" rid="aff0003">3</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Aguiar</surname>
<given-names>Carlos</given-names>
</name>
<xref ref-type="aff" rid="aff0004">4</xref>
</contrib>
<contrib contrib-type="author">
<name>
<surname>Amich</surname>
<given-names>Francisco</given-names>
</name>
<xref ref-type="aff" rid="aff0001">1</xref>
</contrib>
</contrib-group>
<aff id="aff0001">
<label>1</label>Evolution, Taxonomy, and Conservation Group (ECOMED), Department of Botany and Plant Physiology, University of Salamanca, E-37008 Salamanca, Spain; <email xlink:href="rubenrr@usal.es">rubenrr@usal.es</email>
</aff>
<aff id="aff0002">
<label>2</label>Real Jard&#x00ED;n Bot&#x00E1;nico CSIC, Plaza de Murillo 2, ES-28014 Madrid, Spain</aff>
<aff id="aff0003">
<label>3</label>GBM, Universidade da Madeira, Centro de Ci&#x00EA;ncias da vida, Grupo de Bot&#x00E2;nica da Madeira, Campus da Penteada, 9000-390 Funchal, Portugal</aff>
<aff id="aff0004">
<label>4</label>Centro de Investiga&#x00E7;&#x00E3;o de Montanha, Instituto Polit&#x00E9;cnico de Bragan&#x00E7;a, Apartado 1172, Bragan&#x00E7;a 5301-855, Portugal</aff>
<author-notes>
<corresp id="cor1">
<label>&#x002A;</label> Corresponding author</corresp>
<fn>
<p><bold>ORCID ID:</bold> R. Ram&#x00ED;rez-Rodr&#x00ED;guez (<ext-link ext-link-type="uri" xlink:href="http://orcid.org/0000-0003-4849-4233">http://orcid.org/0000-0003-4849-4233</ext-link>); L. Medina (<ext-link ext-link-type="uri" xlink:href="http://orcid.org/0000-0002-1675-4363">http://orcid.org/0000-0002-1675-4363</ext-link>); M. Menezes de Sequeira (<ext-link ext-link-type="uri" xlink:href="http://orcid.org/0000-0001-9728-465X">http://orcid.org/0000-0001-9728-465X</ext-link>); C. Aguiar (<ext-link ext-link-type="uri" xlink:href="http://orcid.org/0000-0001-8643-7112">http://orcid.org/0000-0001-8643-7112</ext-link>); F. Amich (<ext-link ext-link-type="uri" xlink:href="http://orcid.org/0000-0003-3546-4836">http://orcid.org/0000-0003-3546-4836</ext-link>).</p>
<p><bold>Associate Editor:</bold> Carlos Aedo.</p>
</fn>
</author-notes>
<pub-date pub-type="epub">
<day>31</day>
<month>05</month>
<year>2017</year>
</pub-date>
<pub-date pub-type="collection">
<year>2017</year>
</pub-date>
<volume>74</volume>
<issue>1</issue>
<elocation-id content-type="doi">10.3989/ajbm.2467</elocation-id>
<history>
<date date-type="received">
<day>22</day>
<month>12</month>
<year>2016</year>
</date>
<date date-type="accepted">
<day>15</day>
<month>01</month>
<year>2017</year>
</date>
<date date-type="published online">
<day>30</day>
<month>05</month>
<year>2017</year>
</date>
</history>
<permissions>
<copyright-statement>&#x00A9; 2017 CSIC</copyright-statement>
<copyright-year>2017</copyright-year>
<license license-type="open-access" xlink:href="http://creativecommons.org/licenses/by-nc/3.0/">
<license-p>This is an open-access article distributed under the terms of the Creative Commons Attribution-Non Commercial (by-nc) Spain 3.0 License.</license-p>
</license>
</permissions>
<abstract>
<p><italic>Delphinium fissum</italic> subsp. <italic>sordidum</italic> has been found in calcareous and schistaceous slopes on the Mines of Santo Adri&#x00E3;o, Tr&#x00E1;s-os-Montes region &#x2014;close to Bragan&#x00E7;a, northeastern Portugal&#x2014;. It is the first record of this taxon in Portugal and the westernmost locality of its general distribution area. Its conservation status, according to the most recent IUCN categories and criteria, should be &#x201C;Critically Endangered&#x201D; in Portugal.</p>
</abstract>
<trans-abstract xml:lang="es">
<title>Resumen</title>
<p>Se ha encontrado <italic>Delphinium fissum</italic> subsp. <italic>sordidum</italic> en laderas calc&#x00E1;reas y esquistosas en las Minas de Santo Adri&#x00E3;o, en la regi&#x00F3;n de Tr&#x00E1;s-os-Montes &#x2014;cerca de Bragan&#x00E7;a, en el noreste de Portugal&#x2014;. Es la primera cita de este taxon para Portugal y la localidad m&#x00E1;s occidental de su &#x00E1;rea general de distribuci&#x00F3;n. Su estado de conservaci&#x00F3;n, seg&#x00FA;n las categor&#x00ED;as y criterios de la UICN m&#x00E1;s recientes, deber&#x00ED;a ser de &#x201C;En Peligro Cr&#x00ED;tico&#x201D; en Portugal.</p>
</trans-abstract>
<kwd-group xml:lang="en">
<title>Keywords</title>
<kwd>Critically Endangered</kwd>
<kwd><italic>Delphinium</italic></kwd>
<kwd>IUCN</kwd>
<kwd>Mines of Santo Adri&#x00E3;o</kwd>
<kwd>Ranunculaceae</kwd>
<kwd>Tr&#x00E1;s-os-Montes</kwd>
</kwd-group>
<kwd-group xml:lang="es">
<title>Palabras clave</title>
<kwd><italic>Delphinium</italic></kwd>
<kwd>En Peligro Cr&#x00ED;tico</kwd>
<kwd>Minas de Santo Adri&#x00E3;o</kwd>
<kwd>Ranunculaceae</kwd>
<kwd>Tr&#x00E1;s-os-Montes</kwd>
<kwd>UICN</kwd>
</kwd-group>
</article-meta>
</front>
<body>
<sec id="sec1" sec-type="intro">
<title>INTRODUCTION</title>
<p>The genus <italic>Delphinium</italic> L. &#x2014;Ranunculaceae&#x2014; is distributed throughout the temperate and cold zones of the northern hemisphere. Its primary centre of differentiation ranges from the eastern Himalaya to the southwestern China, which includes the highest number of species within &#x2014;about 150 species&#x2014; (Blanch&#x00E9;, <xref ref-type="bibr" rid="cit0005">1990</xref>; Jabbour &#x0026; Renner, <xref ref-type="bibr" rid="cit0013">2012</xref>), as well as the most primitive perennial species as pointed out by Wang (<xref ref-type="bibr" rid="cit0025">1962</xref>) and Malyutin (<xref ref-type="bibr" rid="cit0016">1973</xref>). The larger sets of species, all perennial, are found in Asia and North America. Only a few annual, biennial or perennial species, with Mediterranean affinities, occur in the Iberian Peninsula. According to Blanch&#x00E9; &#x0026; Molero (<xref ref-type="bibr" rid="cit0007">1986</xref>), 11 taxa &#x2014;species and subspecies&#x2014; of <italic>Delphinium</italic> occur in Spain. More recently, the following 2 taxa have been reported for Spain: <italic>D. mansanetianum</italic> Pitarch, Peris &#x0026; Santis (Pitarch Garc&#x00ED;a, <xref ref-type="bibr" rid="cit0020">2002</xref>) and <italic>D. pentagynum</italic> subsp. <italic>formenteranum</italic> N. Torres, L. S&#x00E1;ez, Rossell&#x00F3; &#x0026; C. Blanch&#x00E9; (Torres &#x0026; al., <xref ref-type="bibr" rid="cit0024">2000</xref>). Likewise, <italic>D. fissum</italic> subsp. <italic>fontqueri</italic> Ascaso &#x0026; Pedrol was described by Ascaso &#x0026; Pedrol (<xref ref-type="bibr" rid="cit0002">1991</xref>), but later cytogenetic data (Simon &#x0026; al., <xref ref-type="bibr" rid="cit0023">1995</xref>) revealed that it is <italic>D. bolosii</italic> C. Blanch&#x00E9; &#x0026; Molero. In Portugal, 6 taxa &#x2014;species and subspecies&#x2014; occur, and only <italic>D. pentagynum</italic> Lam. is perennial (cf. Sequeira &#x0026; al., <xref ref-type="bibr" rid="cit0022">2011</xref>).</p>
<p><italic>Delphinium fissum</italic> subsp. <italic>sordidum</italic> (Cuatrec.) Amich, E. Rico &#x0026; J. S&#x00E1;nchez <italic>(D. sordidum</italic> Cuatrec., basion.) belongs to a group of species and subspecies with oriental origin &#x2014;<italic>D.</italic> (subgen. <italic>Oligophyllon</italic> Dimitrova) ser. <italic>Fissa</italic> B. Pawl.&#x2014;, which ranges throughout the Mediterranean region, being the Iberian Peninsula one of the extremes of its area of distribution (Blanch&#x00E9;, <xref ref-type="bibr" rid="cit0006">1991</xref>). This taxon is close to <italic>D. fissum</italic> Waldst. &#x0026; Kit. subsp. <italic>fissum</italic> &#x2014;southeastern France&#x2014; and <italic>D. bolosii</italic> &#x2014;northeastern Spain, with only 6 known populations up to now (cf. L&#x00F3;pez-Pujol &#x0026; al., <xref ref-type="bibr" rid="cit0015">2014</xref>)&#x2014;. The genetic diversity studies carried out by Orellana &#x0026; al. (<xref ref-type="bibr" rid="cit0018">2007</xref>) reinforce the hypothesis of Blanch&#x00E9; (<xref ref-type="bibr" rid="cit0006">1991</xref>) about an expansion of <italic>D. fissum</italic> subsp. <italic>fissum</italic> &#x2014;or an ancestor&#x2014; from the Alps to the Iberian Peninsula.</p>
<p><italic>Delphinium fissum</italic> subsp. <italic>sordidum</italic> was until now considered as an endemic taxon of Spain (Blanch&#x00E9; &#x0026; Molero, <xref ref-type="bibr" rid="cit0007">1986</xref>), where it is catalogued under the category of &#x201C;Endangered&#x201D; &#x2014;EN B2ab(v)c(iv);C2b&#x2014; in the <italic>Lista roja 2008 de la flora vascular espa&#x00F1;ola</italic> and its <italic>Addendum</italic> (Ba&#x00F1;ares &#x0026; al., <xref ref-type="bibr" rid="cit0003">2008</xref>, <xref ref-type="bibr" rid="cit0004">2010</xref>), respectively. Its occurrence in Portugal increases its distribution range to the western Iberian Peninsula; its implications, from a phytogeographical and conservational point of view, will be discussed below.</p>
</sec>
<sec id="sec2" sec-type="material|methods">
<title>MATERIAL AND METHODS</title>
<p>The specimens of <italic>D. fissum</italic> subsp. <italic>sordidum</italic> were collected during field trips in the municipality of Ca&#x00E7;arelhos &#x2014;Vi- mioso, Bragan&#x00E7;a, northeastern Portugal&#x2014; on 14 June 2010 and 14 June 2016. For the identification of the species, morphological characteristics were checked and compared to those given by Blanch&#x00E9; &#x0026; Molero (<xref ref-type="bibr" rid="cit0007">1986</xref>) &#x2014;perennial species, with linear-lanceolate leaf lobes, dense inflorescences, and densely pubescent flowers usually closed&#x2014;; all specimens were <italic>D. fissum</italic> subsp. <italic>sordidum.</italic> The reference material was deposited in the herbaria of the University of Salamanca &#x2014;SALA&#x2014; and the Royal Botanic Garden of Madrid &#x2014;MA.</p>
<p>We carried out a direct counting of reproductive individuals in the population. Nonetheless, we established a counting error of 15% because there are small forested areas at higher elevations where it is difficult to distinguish individual plants since they grow up in clumps and inaccessible places.</p>
<p>The International Union for Conservation of Nature assessment follows the latest guidelines of the IUCN (<xref ref-type="bibr" rid="cit0012">2017</xref>) and, specifically the IUCN regional guidelines (IUCN, <xref ref-type="bibr" rid="cit0011">2012</xref>). Species conservation status was evaluated using B, C, and D criteria. The area of occupancy &#x2014;AOO&#x2014; was calculated with ArcGIS 10.3.1 &#x2014;ESRI, Redlands, California, USA&#x2014;, creating a polygon which included the population area.</p>
<sec id="sec2.1">
<title>Herbarium specimens</title>
<p>PORTUGAL. <bold>Tr&#x00E1;s-os-Montes:</bold> Bragan&#x00E7;a, Vimioso, Ca&#x00E7;arelhos, Minas de Santo Adri&#x00E3;o, 29TQG1000, contact between schist and limestones, holm-oak forest, shrubs and grassland, 538 m a.s.l. <italic>R. Morales</italic> &#x0026; al. <italic>RM 2345,</italic> 14 June 2010, (MA 864924); <italic>ibidem,</italic> 29TQG1000, calcareous and schist slopes, 590 m a.s.l., <italic>F. Amich</italic> &#x0026; <italic>R. Ram&#x00ED;rez-Rodr&#x00ED;guez s.n.,</italic> 14 June 2016 (SALA 158950).</p>
</sec>
</sec>
<sec id="sec3" sec-type="results">
<title>RESULTS</title>
<p><italic>Delphinium fissum</italic> subsp. <italic>sordidum</italic> has been found on the Mines of Santo Adri&#x00E3;o, in the region of Tr&#x00E1;s-os-Montes (<xref ref-type="fig" rid="f0001">Fig. 1</xref>). The specimens collected on 14 June 2010 and 14 June 2016 were in bloom &#x2014;early stage&#x2014; (<xref ref-type="fig" rid="f0002">Fig. 2a</xref>). On 22 July 2016, the population was visited again, and fructifying specimens were seen and collected (<xref ref-type="fig" rid="f0002">Fig. 2b</xref>). The blooming period seems to be quite short, since it lasts about 4 weeks &#x2014;from mid-June to mid-July.</p>
<fig id="f0001">
<label>Fig. 1</label>
<caption>
<p>Distribution of <italic>Delphinium fissum</italic> subsp. <italic>sordidum</italic> in the Iberian Peninsula and location of the first citation in Portugal &#x2014;star symbol.</p>
</caption>
<graphic xlink:href="AJBM201710-2467-g001.tif" xmlns:xlink="http://www.w3.org/1999/xlink"/>
</fig>
<fig id="f0002">
<label>Fig. 2</label>
<caption>
<p><italic>Delphinium fissum</italic> subsp. <italic>sordidum</italic>: <bold>a,</bold> inflorescence; <bold>b,</bold> infrutescence (Mines of Santo Adri&#x00E3;o; photos 14 Jun 2016).</p>
</caption>
<graphic xlink:href="AJBM201710-2467-g002.tif" xmlns:xlink="http://www.w3.org/1999/xlink"/>
</fig>
<p>The taxon is found on a very small area close to the Mines of Santo Adri&#x00E3;o, which corresponds to the largest calcareous area in the northern Portugal. It lives in a holm-oak forest <italic>&#x2014;Quercus ilex</italic> subsp. <italic>ballota</italic> (Desf.) Samp.&#x2014; in a good state of conservation, and is mainly found in bottoms and south-facing slopes. On the slate and granites zones that surround the limestone area, an extensive cork oak forest &#x2014;<italic>Quercus suber</italic> L.&#x2014; grows. <italic>Delphinium fissum</italic> subsp. <italic>sordidum</italic> is located on slopes orientated to the southwest, within cleared holm oak forests, at an altitude of 590 m a.s.l., occupying an approximate area of 735 m<sup>2</sup> &#x2014;0.07 ha; 0.0007 km<sup>2</sup>&#x2014; where 353 reproductive individuals were counted, with an rough error of the 15% in the direct counting. With <italic>D. fissum</italic> subsp. <italic>sordidum</italic> the following species could also be seen: <italic>Quercus ilex</italic> subsp. <italic>ballota, Phillyrea angustifolia</italic> L., <italic>Cytisus scoparius</italic> (L.) Link, <italic>Ruscus aculeatus</italic> L., <italic>Thymus mastichina</italic> (L.) L., <italic>Asparagus acutifolius</italic> L., <italic>Jasminum fruticans</italic> L., <italic>Vincetoxicum nigrum</italic> (L.) Moench, <italic>Ruta montana</italic> (L.) L., <italic>Melica ciliata</italic> L., <italic>Tamus communis</italic> L., <italic>Galium glaucum</italic> L., <italic>Allium sphaerocephalon</italic> L., <italic>Phagnalon saxatile</italic> (L.) Cass., <italic>Sedum sediforme</italic> (Jacq.) Pau, <italic>Euphorbia falcata</italic> L., <italic>Reseda barrelieri</italic> Bertol. ex M&#x00FC;ll. Arg., <italic>Ornithogalum pyrenaicum</italic> L., <italic>Briza maxima</italic> L., <italic>Hypericum perforatum</italic> L., and <italic>Helichrysum stoechas</italic> (L.) Moench.</p>
<p>Although the Mines of Santo Adri&#x00E3;o are integrated into Natura 2000 network as a Site of Community Importance &#x2014;SCI&#x2014; and Special Protection Areas for birds &#x2014;SPA&#x2014;, the area is nonetheless subject to different threats, v.gr., fire and, possibly, collecting due to its showy flowers. Due to its restricted area of occupancy and extent of occurrence, the small number of mature individuals, which might have inter-annual variation due to environmental fluctuations and its unique biology as occurs in other studied populations and the identified threats that it faces, <italic>D. fissum</italic> subsp. <italic>sordidum</italic> is to be considered &#x201C;Critically Endangered&#x201D; &#x2014;CR B1ab(v)c(iv) + 2ab(v)c(iv);C2a(i,ii)b;D2&#x2014; in Portugal.</p>
<p>Applying the regional criteria, the population is found approximately 30 km away from the closest population, and this does not make possible the genetic flow, pollination or seed dispersal events. Thus, the category was not moved up or down. Likewise, the current status of <italic>D. fissum</italic> subsp. <italic>sordidum</italic> in the Iberian Peninsula does not vary in relation to the previously given for Spain &#x2014;EN B2ab(v)c(iv);C2b.</p>
</sec>
<sec id="sec4" sec-type="discussion">
<title>DISCUSSION</title>
<p><italic>Delphinium</italic> ser. <italic>Fissa</italic> is a group with oriental affinities. Several migration patterns can be recognized in the Iberian Peninsula according to Bocquet &#x0026; al. (<xref ref-type="bibr" rid="cit0008">1978</xref>) during the Messinian period. <italic>Delphinium bolosii</italic> may have appeared by some biogeographical barrier &#x2014;v.gr., Ebro River, allopatric speciation&#x2014; or by variation in chromosomes &#x2014;sympatric speciation&#x2014; (Orellana &#x0026; al., <xref ref-type="bibr" rid="cit0018">2007</xref>). Some populations of <italic>D. fissum</italic> subsp. <italic>fissum</italic> could have continued expanding to the western Peninsula and become differentiated as <italic>D. fissum</italic> subsp. <italic>sordidum.</italic> The Portuguese population increases its current distribution range to the western Iberian Peninsula (<xref ref-type="fig" rid="f0001">Fig. 1</xref>).</p>
<p>With <italic>D. fissum</italic> subsp. <italic>sordidum,</italic> the number of recognized taxa &#x2014;species and subspecies&#x2014; of the genus in Portugal increases to 7, with only 2 perennial taxa, <italic>D. pentagynum</italic> and <italic>D. fissum</italic> subsp. <italic>sordidum.</italic> In Spain, the main habitat of the taxon is constituted by formations of <italic>Castanea sativa</italic> Mill. &#x2014;Herv&#x00E1;s, C&#x00E1;ceres province&#x2014; and <italic>Quercus pyrenaica</italic> Willd. &#x2014;Linares de Riofr&#x00ED;o, Salamanca province&#x2014;, mainly on acidic soils. In the isolated population of Sierra M&#x00E1;gina &#x2014;Ja&#x00E9;n province&#x2014;, it lives in communities dominated by thorny and spiky deciduous shrubs. Likewise, in Portugal, the population appears on the calcareous soils of a holm-oak forest.</p>
<p>The new population size, 353 reproductive individuals, implies that demographic stochasticity may not be relevant &#x2014;n &#x003E; 50&#x2014; (Menges, <xref ref-type="bibr" rid="cit0017">1991</xref>; Lande, <xref ref-type="bibr" rid="cit0014">1998</xref>). Environmental stochasticity should be taken into account, especially in studies related to pollination biology and reproductive success, because the closest population is approximately 30 km away, implying that the new population is, in fact, isolated. According to Ellstrand (<xref ref-type="bibr" rid="cit0009">1992</xref>), &#x201C;most of the xenogamous threatened species that are separated by more than 10 km are not linked genetically through pollen flow&#x201D;. On the other hand, the area has a high risk of being affected by wild fires, which might reduce the population size and deteriorate the habitat quality, creating functional barriers that could limit the availability of the pollinators that ensure outcrossing (Pisanu &#x0026; al., <xref ref-type="bibr" rid="cit0019">2009</xref>).</p>
<p>The migration pattern of <italic>D.</italic> series <italic>Fissa,</italic> together with the small size of the populations, explain the low genetic diversity of the taxon, which can be considered depauperated (Orellana &#x0026; al., <xref ref-type="bibr" rid="cit0018">2007</xref>). It would be advisable to clarify the taxonomy of the Iberian populations of this series <italic>&#x2014;D. bolosii, D. mansanetianum,</italic> and <italic>D. fissum</italic> subsp. <italic>sordidum&#x2014;</italic> using ETS/ITS markers (L&#x00F3;pez-Pujol &#x0026; al., <xref ref-type="bibr" rid="cit0015">2014</xref>).</p>
<p>Plant species survival can also be affected by anthropogenic threats. The collection of plants, especially of the remarkable or rare species, is a potential threat for all them. However, botanists have generally been careful with collecting activities, and these botanical tasks have had very positive results for species conservation, providing timely and accurate information (Aedo &#x0026; al., <xref ref-type="bibr" rid="cit0001">2015</xref>). Moreover, we have not found evidences of herbivores, but we have observed it in other populations &#x2014;v.gr., Linares de Riofr&#x00ED;o and Sierra M&#x00E1;gina&#x2014;. Fire is the main threat to consider, because it occurs frequently in Mediterranean ecosystems. For instance, in Villarino de los Aires &#x2014;Salamanca province, Spain&#x2014;, we have evidences that fire affected negatively the population during the summer of 2006 (Ram&#x00ED;rez-Rodr&#x00ED;guez &#x0026; al., unpubl.). We have not observed that an increase of the weed had a negative effect on recruitment. On the contrary, the low recruitment of this species can be explained by its low survival rate of seedlings and samplings (Ram&#x00ED;rez-Rodr&#x00ED;guez &#x0026; al., unpubl.).</p>
<p>Habitat fragmentation has been shown as a critical factor for the loss of plant diversity, since it influences movement and dispersal of organisms, gene flow rates, and invasion by exotic plants, among many other factors (Heywood &#x0026; Iriondo, <xref ref-type="bibr" rid="cit0010">2003</xref>). Considering that it represents the only population currently known in Portugal, and its threat category is high, we propose to carry out intensive searches in the limestone environment of the Mines of Santo Adri&#x00E3;o and the annual monitoring of the population, to make germination studies and apply conservation measures to avoid its extinction. We believe that a detailed study of the biology of <italic>D. fissum</italic> subsp. <italic>sordidum</italic> is necessary in order to implement a recovery plan in Portugal, as well as for the populations of Castile and Leon (Ram&#x00ED;rez-Rodr&#x00ED;guez &#x0026; Amich, <xref ref-type="bibr" rid="cit0021">2014</xref>). Likewise, it is interesting to continue the fieldwork directed to find new populations in the Iberian Peninsula.</p>
</sec>
</body>
<back>
<ack>
<title>ACKNOWLEDGEMENTS</title>
<p>The authors thank the collaboration of the <italic>Flora iberica</italic> project (CGL2014-52787-C3-1-P) as well as the editor and two anonymous reviewers for their valuable comments and suggestions which helped us to improve the manuscript. Moreover, we would like to express our gratitude to Sergio Fuentes and Javier Garc&#x00ED;a for their assistance in the field work.</p>
</ack>
<ref-list>
<title>REFERENCES</title>
<ref id="cit0001">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Aedo</surname>
<given-names>C.</given-names>
</name>
<name>
<surname>Medina</surname>
<given-names>L.</given-names>
</name>
<name>
<surname>Barber&#x00E1;</surname>
<given-names>P.</given-names>
</name>
<name>
<surname>Fern&#x00E1;ndez-Albert</surname>
<given-names>M.</given-names>
</name>
</person-group>
<article-title>Extinctions of vascular plants in Spain</article-title>
<source>Nordic Journal of Botany</source>
<year>2015</year>
<volume>33</volume>
<fpage>83</fpage>
<lpage>100</lpage>
<comment><ext-link ext-link-type="uri" xlink:href="https://doi.org/10.1111/njb.00575">https://doi.org/10.1111/njb.00575</ext-link></comment>
</nlm-citation>
</ref>
<ref id="cit0002">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Ascaso</surname>
<given-names>J.</given-names>
</name>
<name>
<surname>Pedrol</surname>
<given-names>J.</given-names>
</name>
</person-group>
<article-title>De plantis vascularibus praesertim ibericis I</article-title>
<source>Fontqueria</source>
<year>1991</year>
<volume>31</volume>
<fpage>135</fpage>
<lpage>140</lpage>
</nlm-citation>
</ref>
<ref id="cit0003">
<mixed-citation publication-type="book">
<person-group person-group-type="editor">
<name>
<surname>Ba&#x00F1;ares</surname>
<given-names>&#x00C1;.</given-names>
</name>
<name>
<surname>Blanca</surname>
<given-names>G.</given-names>
</name>
<name>
<surname>G&#x00FC;emes</surname>
<given-names>J.</given-names>
</name>
<name>
<surname>Moreno</surname>
<given-names>J.C.</given-names>
</name>
<name>
<surname>Ortiz</surname>
<given-names>S.</given-names>
</name>
</person-group>
<year>2008</year>
<source>Lista roja 2008 de la flora vascular espa&#x00F1;ola</source>
<publisher-name>Direcci&#x00F3;n General de Medio Natural y Pol&#x00ED;tica Forestal</publisher-name>
<publisher-loc>Madrid</publisher-loc>
</mixed-citation>
</ref>
<ref id="cit0004">
<mixed-citation publication-type="book">
<person-group person-group-type="editor">
<name>
<surname>Ba&#x00F1;ares</surname>
<given-names>&#x00C1;.</given-names>
</name>
<name>
<surname>Blanca</surname>
<given-names>G.</given-names>
</name>
<name>
<surname>G&#x00FC;emes</surname>
<given-names>J.</given-names>
</name>
<name>
<surname>Moreno</surname>
<given-names>J.C.</given-names>
</name>
<name>
<surname>Ortiz</surname>
<given-names>S.</given-names>
</name>
</person-group>
<year>2010</year>
<source>Atlas y Libro Rojo de la Flora Vascular Amenazada de Espa&#x00F1;a. Adenda 2010</source>
<publisher-name>Direcci&#x00F3;n General de Medio Natural y Pol&#x00ED;tica Forestal</publisher-name>
<publisher-loc>Madrid</publisher-loc>
</mixed-citation>
</ref>
<ref id="cit0005">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Blanch&#x00E9;</surname>
<given-names>C.</given-names>
</name>
</person-group>
<article-title>
<italic>Delphinium</italic> L. subgen. <italic>Delphinium:</italic> origin and evolutionary trends</article-title>
<source>Collectanea Botanica</source>
<year>1990</year>
<volume>19</volume>
<fpage>75</fpage>
<lpage>95</lpage>
<comment><ext-link ext-link-type="uri" xlink:href="https://doi.org/10.3989/collectbot.1990.v19.118">https://doi.org/10.3989/collectbot.1990.v19.118</ext-link></comment>
</nlm-citation>
</ref>
<ref id="cit0006">
<mixed-citation publication-type="book">
<person-group person-group-type="author">
<name>
<surname>Blanch&#x00E9;</surname>
<given-names>C.</given-names>
</name>
</person-group>
<year>1991</year>
<source>Revisi&#x00F3; biosistem&#x00E1;tica del g&#x00E8;nere Delphinium L., a la Pen&#x00ED;nsula Ib&#x00E8;rica i a les Illes Balears</source>
<publisher-name>Institut d&#x00B4;Estudis Catalans. Arxius de la Secci&#x00F3; de Ci&#x00E8;ncies, n.&#x00BA; 98</publisher-name>
<publisher-loc>Barcelona</publisher-loc>
</mixed-citation>
</ref>
<ref id="cit0007">
<mixed-citation publication-type="book">
<person-group person-group-type="author">
<name>
<surname>Blanch&#x00E9;</surname>
<given-names>C.</given-names>
</name>
<name>
<surname>Molero</surname>
<given-names>J.</given-names>
</name>
</person-group>
<person-group person-group-type="editor">
<name>
<surname>Castroviejo</surname>
<given-names>S.</given-names>
</name>
<etal>al</etal>
</person-group>
<year>1986</year>
<chapter-title><italic>Delphinium</italic> L</chapter-title>
<source>Flora iberica</source>
<volume>vol. 1</volume>
<publisher-name>Real Jard&#x00ED;n Bot&#x00E1;nico de Madrid CSIC</publisher-name>
<publisher-loc>Madrid</publisher-loc>
</mixed-citation>
</ref>
<ref id="cit0008">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Bocquet</surname>
<given-names>G.</given-names>
</name>
<name>
<surname>Wilder</surname>
<given-names>B.</given-names>
</name>
<name>
<surname>Kiefer</surname>
<given-names>H.</given-names>
</name>
</person-group>
<article-title>The messinian model -a new outlook for the floristics and systematics of the Mediterranean area</article-title>
<source>Candollea</source>
<year>1978</year>
<volume>38</volume>
<fpage>269</fpage>
<lpage>287</lpage>
</nlm-citation>
</ref>
<ref id="cit0009">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Ellstrand</surname>
<given-names>N.C.</given-names>
</name>
</person-group>
<article-title>Gene flow by pollen: implications for plant conservation genetics</article-title>
<source>Oikos</source>
<year>1992</year>
<volume>63</volume>
<fpage>77</fpage>
<lpage>86</lpage>
<comment>
<ext-link ext-link-type="uri" xlink:href="https://doi.org/10.2307/3545517">https://doi.org/10.2307/3545517</ext-link>
</comment>
</nlm-citation>
</ref>
<ref id="cit0010">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Heywood</surname>
<given-names>V.H.</given-names>
</name>
<name>
<surname>Iriondo</surname>
<given-names>J.M.</given-names>
</name>
</person-group>
<article-title>Plant conservation: old problems, new perspectives</article-title>
<source>Biological Conservation</source>
<year>2003</year>
<volume>113</volume>
<fpage>321</fpage>
<lpage>335</lpage>
<comment><ext-link ext-link-type="uri" xlink:href="https://doi.org/10.1016/S0006-3207(03)00121-6">https://doi.org/10.1016/S0006-3207(03)00121-6</ext-link></comment>
</nlm-citation>
</ref>
<ref id="cit0011">
<mixed-citation publication-type="book">
<person-group person-group-type="author">
<collab>IUCN</collab>
</person-group>
<year>2012</year>
<source>Guidelines for application of IUCN Red List criteria at regional and national levels: version 4.0</source>
<publisher-name>IUCN Species Survival Commission</publisher-name>
<publisher-loc>Gland and Camnbrige</publisher-loc>
</mixed-citation>
</ref>
<ref id="cit0012">
<nlm-citation publication-type="webpage">
<person-group person-group-type="author">
<collab>IUCN</collab>
</person-group>
<source>Guidelines for using the IUCN Red List Categories and criteria: version 12</source>
<year>2017</year>
<comment>[<ext-link ext-link-type="uri" xlink:href="http://www.iucnredlist.org/documents/RedListGuidelines.pdf">http://www.iucnredlist.org/documents/RedListGuidelines.pdf</ext-link>]</comment>
</nlm-citation>
</ref>
<ref id="cit0013">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Jabbour</surname>
<given-names>F.</given-names>
</name>
<name>
<surname>Renner</surname>
<given-names>S.</given-names>
</name>
</person-group>
<article-title>A phylogeny of <italic>Delphinieae</italic> (Ranunculaceae) shows that <italic>Aconitum</italic> is nested within <italic>Delphinium</italic> and that Late Miocene transitions to long life cycles in the Himalayas and Southwest China coincide with bursts in diversification</article-title>
<source>Molecular Phylogenetics and Evolution</source>
<year>2012</year>
<volume>62</volume>
<fpage>928</fpage>
<lpage>942</lpage>
<comment><ext-link ext-link-type="uri" xlink:href="https://doi.org/10.1016/j.ympev.2011.12.005">https://doi.org/10.1016/j.ympev.2011.12.005</ext-link></comment>
</nlm-citation>
</ref>
<ref id="cit0014">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Lande</surname>
<given-names>R.</given-names>
</name>
</person-group>
<article-title>Anthropogenic, ecological and genetic factors in extinction and conservation</article-title>
<source>Researches on Population Ecology</source>
<year>1998</year>
<volume>40</volume>
<fpage>259</fpage>
<lpage>269</lpage>
<comment><ext-link ext-link-type="uri" xlink:href="https://doi.org/10.1007/BF02763457">https://doi.org/10.1007/BF02763457</ext-link></comment>
</nlm-citation>
</ref>
<ref id="cit0015">
<mixed-citation publication-type="book">
<person-group person-group-type="author">
<name>
<surname>L&#x00F3;pez-Pujol</surname>
<given-names>J.</given-names>
</name>
<name>
<surname>Bosch</surname>
<given-names>M.</given-names>
</name>
<name>
<surname>Mass&#x00F3;</surname>
<given-names>S.</given-names>
</name>
<name>
<surname>Molero</surname>
<given-names>J.</given-names>
</name>
<name>
<surname>Simon</surname>
<given-names>J.</given-names>
</name>
<name>
<surname>Rovira</surname>
<given-names>A.M.</given-names>
</name>
<name>
<surname>Blanch&#x00E9;</surname>
<given-names>C.</given-names>
</name>
</person-group>
<year>2014</year>
<source>Estudi preliminar de variabilitat isoenzim&#x00E0;tica de Delphinium bolosii al Parc Natural de Sant Lloren&#x00E7; del Munt i l&#x2019;Obac</source>
<publisher-name>Diputaci&#x00F3; de Barcelona</publisher-name>
<publisher-loc>Barcelona</publisher-loc>
</mixed-citation>
</ref>
<ref id="cit0016">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Malyutin</surname>
<given-names>N.I.</given-names>
</name>
</person-group>
<article-title>The phylogeny and the taxonomy of the <italic>Delphinium</italic> L. genus</article-title>
<source>Botanischeskii Zhurnal</source>
<year>1973</year>
<volume>58</volume>
<fpage>1710</fpage>
<lpage>1722</lpage>
</nlm-citation>
</ref>
<ref id="cit0017">
<mixed-citation publication-type="book">
<person-group person-group-type="author">
<name>
<surname>Menges</surname>
<given-names>E. S.</given-names>
</name>
</person-group>
<person-group person-group-type="editor">
<name>
<surname>Falk</surname>
<given-names>D.A.</given-names>
</name>
<name>
<surname>Holsinger</surname>
<given-names>K.E.</given-names>
</name>
</person-group>
<year>1991</year>
<chapter-title>The application of minimum viable population theory to plants</chapter-title>
<source>Genetics and Conservation of Rare Plants</source>
<publisher-name>Oxford University Press</publisher-name>
<publisher-loc>New York</publisher-loc>
</mixed-citation>
</ref>
<ref id="cit0018">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Orellana</surname>
<given-names>M.R.</given-names>
</name>
<name>
<surname>L&#x00F3;pez-Pujol</surname>
<given-names>J.</given-names>
</name>
<name>
<surname>Blanch&#x00E9;</surname>
<given-names>C.</given-names>
</name>
<name>
<surname>Bosch</surname>
<given-names>M.</given-names>
</name>
</person-group>
<article-title>Genetic diversity in the endangered dysploid larkspur <italic>Delphinium bolosii</italic> and its close diploid relatives in the series <italic>Fissa</italic> of the Western Mediterranean area</article-title>
<source>Biological Journal of the Linnean Society</source>
<year>2007</year>
<volume>92</volume>
<fpage>773</fpage>
<lpage>784</lpage>
<comment><ext-link ext-link-type="uri" xlink:href="https://doi.org/10.1111/j.1095-8312.2007.00910.x">https://doi.org/10.1111/j.1095-8312.2007.00910.x</ext-link></comment>
</nlm-citation>
</ref>
<ref id="cit0019">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Pisanu</surname>
<given-names>S.</given-names>
</name>
<name>
<surname>Filigheddu</surname>
<given-names>R.</given-names>
</name>
<name>
<surname>Farris</surname>
<given-names>E.</given-names>
</name>
</person-group>
<article-title>The conservation status of an endemic species of northern Sardinia: <italic>Centaurea horrida</italic> Badar&#x00F2; (Asteraceae)</article-title>
<source>Plant Biosystems</source>
<year>2009</year>
<volume>143</volume>
<fpage>275</fpage>
<lpage>282</lpage>
<comment><ext-link ext-link-type="uri" xlink:href="https://doi.org/10.1080/11263500902722493">https://doi.org/10.1080/11263500902722493</ext-link></comment>
</nlm-citation>
</ref>
<ref id="cit0020">
<mixed-citation publication-type="book">
<person-group person-group-type="author">
<name>
<surname>Pitarch Garc&#x00ED;a</surname>
<given-names>R.</given-names>
</name>
</person-group>
<year>2002</year>
<source>Estudio de la flora y vegetaci&#x00F3;n de las sierras orientales del Sistema Ib&#x00E9;rico: La Palomita, Las Dehesas, El Rayo y Mayabona (Teruel)</source>
<series>Serie Investigaci&#x00F3;n n.&#x00BA; 38</series>
<publisher-name>Consejo de Protecci&#x00F3;n de la Naturaleza de Arag&#x00F3;n</publisher-name>
<publisher-loc>Zaragoza</publisher-loc>
</mixed-citation>
</ref>
<ref id="cit0021">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Ram&#x00ED;rez-Rodr&#x00ED;guez</surname>
<given-names>R.</given-names>
</name>
<name>
<surname>Amich</surname>
<given-names>F.</given-names>
</name>
</person-group>
<article-title>Notes on rare and threatened flora in western-central Iberia</article-title>
<source>Lazaroa</source>
<year>2014</year>
<volume>35</volume>
<fpage>221</fpage>
<lpage>226</lpage>
<comment><ext-link ext-link-type="uri" xlink:href="https://doi.org/10.5209/rev_LAZA.2014.v35.46571">https://doi.org/10.5209/rev_LAZA.2014.v35.46571</ext-link></comment>
</nlm-citation>
</ref>
<ref id="cit0022">
<mixed-citation publication-type="book">
<person-group person-group-type="editor">
<name>
<surname>Sequeira</surname>
<given-names>M.</given-names>
</name>
<name>
<surname>Espirito-Santo</surname>
<given-names>M.D.</given-names>
</name>
<name>
<surname>Capelo</surname>
<given-names>J.</given-names>
</name>
<name>
<surname>Honrado</surname>
<given-names>J.</given-names>
</name>
</person-group>
<year>2011</year>
<source>Checklist da Flora de Portugal (Continental, A&#x00E7;ores e Madeira)</source>
<publisher-name>Associa&#x00E7;&#x00E3;o Lusitana de Fitossociologia</publisher-name>
<publisher-loc>Lisboa</publisher-loc>
</mixed-citation>
</ref>
<ref id="cit0023">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Simon</surname>
<given-names>J.</given-names>
</name>
<name>
<surname>Bosch</surname>
<given-names>M.</given-names>
</name>
<name>
<surname>Blanch&#x00E9;</surname>
<given-names>C.</given-names>
</name>
<name>
<surname>Molero</surname>
<given-names>J.</given-names>
</name>
</person-group>
<article-title>Delphinium</article-title>
<source>Flora Mediterranea</source>
<year>1995</year>
<volume>5</volume>
<fpage>323</fpage>
<lpage>331</lpage>
</nlm-citation>
</ref>
<ref id="cit0024">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Torres</surname>
<given-names>N.</given-names>
</name>
<name>
<surname>S&#x00E1;ez</surname>
<given-names>L.</given-names>
</name>
<name>
<surname>Rossell&#x00F3;</surname>
<given-names>J.A.</given-names>
</name>
<name>
<surname>Blanch&#x00E9;</surname>
<given-names>C.</given-names>
</name>
</person-group>
<article-title>A new <italic>Delphinium</italic> subsp. from Formentera (Balearic Islands)</article-title>
<source>Botanical Journal of the Linnean Society</source>
<year>2000</year>
<volume>133</volume>
<fpage>371</fpage>
<lpage>377</lpage>
<comment><ext-link ext-link-type="uri" xlink:href="https://doi.org/10.1111/j.1095-8339.2000.tb01552.x">https://doi.org/10.1111/j.1095-8339.2000.tb01552.x</ext-link></comment>
</nlm-citation>
</ref>
<ref id="cit0025">
<nlm-citation publication-type="journal">
<person-group person-group-type="author">
<name>
<surname>Wang</surname>
<given-names>W. T.</given-names>
</name>
</person-group>
<article-title>Critical review of the genus <italic>Delphinium</italic> from the Ranunculaceaous Flora of China</article-title>
<source>Acta Botanica Sinica</source>
<year>1962</year>
<volume>10</volume>
<fpage>87</fpage>
<lpage>282</lpage>
</nlm-citation>
</ref>
</ref-list>
</back>
</article>
