Diversity of sexual systems within different lineages of the genus Silene

Research Article Diversity of sexual systems within different lineages of the genus Silene Ine ´s Casimiro-Soriguer1,2*, Maria L. Buide1and Eduardo Narbona1 1 A ´rea de Bota ´nica, Departamento de Biologı ´a Molecular e Ingenierı ´a Bioquı ´mica, Universidad Pablo de Olavide, Ctra. de Utrera, km 1, 41013 Sevilla, Spain 2 A ´rea de Bota ´nica, Departamento de Biologı ´a Vegetal y Ecologı ´a, Universidad de Sevilla, Avenida Reina Mercedes s/n, 41012 Sevilla, Spain Received: 16 December 2014; Accepted: 26 March 2015; Published: 10 April 2015 Associate Editor: Diana Wolf Citation: Casimiro-Soriguer I, Buide ML, Narbona E. 2015. Diversity of sexual systems within different lineages of the genus Silene. AoB PLANTS 7: plv037; doi:10.1093/aobpla/plv037 Abstract. Species and populations can be categorized by their sexual systems, depending on the spatial distribution of female and male reproductive structures within and among plants. Although a high diversity of sexual systems exists in Silene, their relative frequency at the genus and infrageneric level is unknown. Here, we carried out an extensive literature search for direct or indirect descriptions of sexual systems in Silene species. We found descriptions of sexual systems for 98 Silene species, where 63 and 35 correspond to the phylogenetically supported subgenera Silene and Behenantha, respectively. Hermaphroditism was the commonest sexual system (58.2 %), followed by dioecy (14.3 %), gynodioecy (13.3 %) and gynodioecy–gynomonoecy (i.e. hermaphroditic, female and gynomonoecious plants coexisting in the same population; 12.2 %). The presence of these sexual systems in both subgenera suggests their multiple origins. In 17 species, the description of sexual systems varied, and in most cases these differences corresponded to variations within or among populations. Interestingly, the poorly studied gynodioecy–gynomonoecy sexual system showed similar frequency to dioecy and gynodioecy in both subgenera. In addition, the incidence of gynodioecy–gynomonoecy was analysed in the species of section Psammophilae (Silene littorea,S. psammitis, S. adscendens and S. cambessedesii), in a survey of 26 populations across the distribution area of the species. The four species showed gynomonoecy–gynodioecy in most populations. Hermaphrodites were the most frequent morph, with a low number of females and gynomonoecious plants in all populations. The frequency of sexual morphs varied significantly among the studied populations but not among species. Female plants generally produced smaller numbers of flowers than hermaphroditic or gynomonoecious plants, and the percentages of female flowers per population were low. All these findings suggest that the gynodioecious–gynomonoecious sexual system in section Psammophilae is closer to hermaphroditism or gynomonoecy than gynodioecy. Keywords: Behenantha; Caryophyllaceae; dioecy; gynodioecy; gynodioecy–gynomonoecy; hermaphroditism; Psammophilae; sexual polymorphism; sexual system; Silene. Introduction The study of the diversity and evolution of sexual systems in plants has been the focus of many scientists since early days. Species or populations may be categorized by sexual system, depending on the spatial distribution of male and female reproductive structures within and *Corresponding author’s e-mail address: [email protected] Published by Oxford University Press on behalf of the Annals of Botany Company. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/ licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited. AoB PLANTS www.aobplants.oxfordjournals.org &The Authors 2015 1 among plants (Bawa and Beach 1981). Although many different sexual systems may exist (Sakai and Weller 1999), most of the angiosperm species belong to one of the five main types: hermaphroditism (72 %), gynodioecy (7 %, female and hermaphroditic individuals), monoecy (5 %, individuals with female and male flowers), dioecy (5–6 %, female and male individuals) and gynomonoecy (3 %, individuals with female and hermaphroditic flowers) (Richards 1997;Renner 2014). Although only 5–6 % of total angiosperms are dioecious, dioecious species are present in 43 % of families, and from 871 to 5000 independent origins of dioecy have been proposed (Renner 2014). Therefore, the evolutionary pathways to dioecy have been the focus of interesting debate, specially the transition from hermaphroditism to dioecy, with gynodioecy or monoecy as intermediate steps (Charlesworth 1999). The association of gynodioecy or monoecy with dioecy at the family or genera level suggests that both are possible pathways to dioecy (Renner and Ricklefs 1995;Dufay et al. 2014;Renner 2014). Gynomonoecyoccurs frequently in families such as Compositae or Chenopodiaceae (Yampolsky and Yampolsky 1922;Torices et al. 2011), and has been considered the main route to monoecy from hermaphroditism and vice versa (Torices et al. 2011). The genus Silene (Caryophyllaceae) has been widely used to study the evolution of sexual systems and gender variation (Meagher 2007;Bernasconi et al. 2009;Charlesworth 2013;Weingartner and Delph 2014), and is one of the groups used for the phylogenetic approach (Desfeux et al. 1996;Rautenberg et al. 2010;Marais et al. 2011). Nonetheless, the complete phylogenetic relationship within this genus is not yet resolved (Rautenberg et al. 2010;Petri et al. 2013). What seems clear is the subdivision of Silene (sensu Oxelman and Lide ´n 1995)intotwoclades:subgenera Silene and Behenantha (Popp and Oxelman 2004,2007; Rautenberg et al. 2010). The first of these phylogenetic studies found that dioecy appeared independently at least twice (in subsection Otites and section Melandrium; according to Oxelman et al. 2013), and that gynodioecy was the most probable ancestral condition for the genus (Desfeux et al. 1996). More recently, Marais et al. (2011) found that either gynodioecy or hermaphroditism could be the ancestral condition of Silene.InOtites and Melandrium, different types of sex-determining systems with a different date of origin are implicated (Ka ¨fer et al. 2013; Slancarova et al. 2013). In addition to dioecy, hermaphroditism and gynodioecy are common in Silene (Desfeux et al. 1996;Ju ¨rgens et al. 2002). However, monoecy is not present, suggesting the evolution of dioecy through the gynodioecy pathway. Gynomonoecy, andromonoecy (individuals with male and hermaphroditic flowers) and trioecy (populations with hermaphroditic, male and female individuals) have also been reported for Silene, but are very rare (Desfeux et al. 1996;Ju ¨rgens et al. 2002; present study). However, in a non-negligible number of gynodioecious species, the existence of gynomonoecious individuals (i.e. plants with female and hermaphroditic flowers) in the populations is reported (e.g. Shykoff 1988;Talavera et al. 1996; Lafuma and Maurice 2006;Dufay et al. 2010). Species or populations containing hermaphroditic, female and gynomonoecious individuals must be considered as gynodioecious–gynomonoecious (Gd–Gm hereafter) (Desfeux et al. 1996). The frequency of gynomonoecious plants may be highly variable among populations and species; in some cases, this sexual morph is rare and in others it may be the most frequent (Charlesworth and Laporte 1998;Maurice 1999;Dufay et al. 2010;Casimiro-Soriguer et al. 2013). The genetic mechanism for sex determination of gynodioecy may be based on the interaction of cytoplasmic male sterility genes with nuclear restorers of male fertility (Bailey and Delph 2007), as found in S. vulgaris (Charlesworth and Laporte 1998). In some cases, the incomplete restoration of the cytoplasmic male sterility factors or heteroplasmy (the occurrence of different cytotypes within an individual) can cause partially male-sterile plants that are able to produce females and hermaphroditic flowers (i.e. gynomonoecious plants) (Koelewijn and Van Damme 1996;McCauley et al. 2005). Thus, although the genetic basis for gynomonoecious and female individuals in Gd–Gm species has been hypothesized in Silene species (Glaettli and Goudet 2006; Garraud et al. 2011), their incidence remains unclear. Silene littorea is one of the most studied species with a Gd–Gm sexual system (Guitia ´n and Medrano 2000;Vilas and Garcı ´a 2006;Vilas et al. 2006;Casimiro-Soriguer et al. 2013). In several populations from two contrasting sites in their distribution area, the frequency of hermaphroditesorgynomonoeciousplantsvariedhighlyamong populations, but female plants were always rare (Guitia ´n and Medrano 2000;Casimiro-Soriguer et al. 2013). Analysis of functional gender showed that nearly all plants in the population transmit their genes via both pollen and ovules; thus, the Gd–Gm sexual system of S. littorea seems to be closer to hermaphroditism or gynomonoecy than gynodioecy (Casimiro-Soriguer et al. 2013). Interestingly, S. stockenii also shows a Gd–Gm sexual system with a very low frequency of female plants (Talavera et al. 1996). Both species belong to the section Psammophilae, composed of three other annual species (S. adscendens, S. cambessedesii and S. psammitis). Therefore, the question which arises from these findings is whether the Gd–Gm sexual system is widespread in the whole Psammophilae section. In addition, the reproductive output of the different morphs may vary in the Gd–Gm sexual system, which may be important to the stable maintenance 2AoB PLANTS www.aobplants.oxfordjournals.org &The Authors 2015 Casimiro-Soriguer et al. — Sexual systems in Silene and section Psammophilae of these morphs in the populations (Dufay et al. 2010). For instance, Shykoff et al. (2003) found that overall females produce more but smaller flowers, set more fruits and produce more and heavier seeds than hermaphrodites. In this study, two different approaches were used to evaluate the occurrence of sexual systems, particularly gynodioecy–gynomonoecy, in Silene.Forthegeneral approach, we searched the literature extensively to locate any direct or indirect description of the sexual system of the species of Silene.Thissearchallowsustoknowthe frequency of sexual systems at the genus and infrageneric level as well as their variability within species. Accurate estimates of the frequency of the Gd–Gm sexual system may shed light on their possible evolution and stability in Silene, and also in other groups of angiosperms. For the specific approach, we have studied the sexual systems of a total 26 populations of the species of the section Psammophilae. Specifically, we seek to answer the following questions. (i) Is the Gd–Gm sexual system widespread throughout the distribution area of S. littorea and the other species of section Psammophilae? (ii) What is the frequency of the different sexual morphs and types of flowers in the populations? (iii) Are there differences in the number of flowers produced by each morph? Methods Study system Silene littorea,S. cambessedesii,S. psammitis and S. stockenii are endemic to the Iberian Peninsula and Balearic Islands (Talavera 1979). Talavera (1979) included these taxa together with S. almolae,S. germana and S. pendula within the section Erectorefractae. We follow Greuter (1995) who proposed the section Psammophilae, previously considered a subsection of Erectorefractae (Talavera 1979). Oxelman et al. (2013) consider the species status of S. adscendens (previously considered a subspecies of S. littorea). All the species are spring-flowering annuals and grow in different types of soil: sandy substrates from the coast (S. cambessedesii,S. littorea), dolomites or slates (S. psammitis), calcareous sandstones (S. stockenii)orschists (S. adscendens)(Talavera 1979). Analysis of the sexual system of section Psammophilae During the peak of the flowering period from 2010 to 2012, we visited 5 populations of S. adscendens,8of S. cambessedesii,11ofS. littorea and 4 of S. psammitis (Fig. 1)[see Supporting Information]. We did not include S. stockenii because: (i) it is a critically endangered species Figure 1. Populations sampled from the different species of section Psammophilae: nine populations of S. littorea (grey dots), five populations of S. adscendens (black triangles), eight populations of S. cambessedesii (black dots) and four populations of S. psammitis (grey squares). AoB PLANTS www.aobplants.oxfordjournals.org &The Authors 2015 3 Casimiro-Soriguer et al. — Sexual systems in Silene and section Psammophilae with only a few populations (Ban ˜ares et al.2004), (ii) Talavera et al. (1996) have already studied the sexual system of S. stockenii in the most important population and (iii) we visited some of the remnant populations, detecting high florivory levels and a small number of individuals. We performed single-day linear transects of 100 plants, with the exception of some very small populations [see Supporting Information]. We chose plants separated by at least 1 m to avoid microhabitat or clustering effects in sex expression (Klaas and Olson 2006). For each plant, we counted all the flowers in anthesis, and noted their sex (female or hermaphroditic). Withered flowers were also analysed when sex differentiation was possible. Plants bearing only female or hermaphroditic flowers were considered female or hermaphroditic individuals, respectively, whereas individuals with female and hermaphroditic flowers were considered gynomonoecious. In a previous study in S. littorea,Casimiro-Soriguer et al. (2013) found that the probability of recording female flowers in gynomonoecious plants was higher when the whole flowering period of a plant was studied than when estimates were based on a single census. Thus, our sampling methodology would underestimate the frequency of gynomonoecious plants in the population. In addition, plants with a large number of flowers would have a higher probability to be classified as gynomonoecious. Literature search on Silene sexual systems We performed a literature search in the SCOPUS and JSTOR databases including the terms: Silene, breeding system, sexual system, hermaphroditic, hermaphrodite, hermaphroditism, dioecious, dioecy, gynodioecious, gynodioecy, gynomonoecious, gynomonoecy, androdioecious, androdioecy, andromonoecious, andromonoecy, monoecious and monoecy. We also revised the description of the Silene species in main floras and revision studies or books previous to 1938 that contain information about plant sexuality, see Table 1and [Supporting Information]. In addition, for those species with numerous sexual system descriptions or present in the Euro +Med database, specific individual searches were performed. We annotated the information about the sexual systems of the species in one of the following categories: hermaphrodite (H), dioecious (D), gynodioecious (Gd), gynomonoecious (Gm), androdioecious (Ad), andromonoecious (Am) or trioecious (T). In some cases, mixed sexual systems were found within a single population, for instance Am–Ad (male, hermaphroditic and andromonoecious plants), H–Gm (hermaphrodites and gynomonoecious plants) and gynodioecious–gynomonoecious (Gd–Gm, female, hermaphroditic and gynodioecious plants). When various studies described a species with different sexual systems, all of them were annotated with the respective reference; however, the principal or most frequent sexual system was used for calculating the frequency at the genus or subgenus level (see Ju ¨rgens et al. 2002 for similar criteria). The H–Gm category was assigned as H because in most cases gynomonoecious individuals are extremely rare and bear only a few female flowers (e.g. A. Ju ¨rgens, pers. comm.; Gime ´nez-Benavides et al. 2007). We will follow the classification criteria of Oxelman et al. (2013) for Silene and the infrageneric level. The subspecies level was not considered. Statistical analysis To test for differences in the proportion of the different sexual morphs among species and populations, a generalized linear model (GLM) with a multinomial distribution and a probit link function was carried out. We considered the sexual morph of each individual (female, hermaphrodite or gynomonoecious) as the multinomial response variable; and species and population (nested within species) as fixed factors. Population was treated as a fixed factor rather than a random factor because we are interested in examining the differences in morph frequencies among our specific populations, and the same populations would be analysed in future studies (Bennington and Thayne 1994;Potvin 2001). Comparisons of the number of flowers between female plants and hermaphrodite or gynomonoecious plants were performed using GLMs with a log link function and a Poisson error distribution. The dependent variable was the number of flowers produced by each individual; and sexual morph, population (nested within species) and species were included as fixed factors. On the other hand, the frequency of each sexual system between the subgenus Silene and Behenantha was compared using x 2 tests for contingency tables (Quinn and Keough 2002). All the analysis were carried out in IBM w SPSS w Statistics v.22. Results Sexual system of the section Psammophilae A total of 2478 individuals belonging to 26 populations were surveyed. In general, each studied taxa of section Psammophilae showed Gd and Gd–Gm populations, although Gd–Gm populations were the most frequent (Fig. 2). Silene littorea,S. adscendens and S. psammitis showed one Gd population each, whereas S. cambessedesii showed two Gd populations. The remaining populations were all Gd–Gm (Fig. 2). Overall, the most frequent morph of section Psammophilae was the hermaphrodite, with an 86.8 % of individuals included in this category, followed by the female 4AoB PLANTS www.aobplants.oxfordjournals.org &The Authors 2015 Casimiro-Soriguer et al. — Sexual systems in Silene and section Psammophilae .......................................................................... ............................................................................................................................................................................ Table 1. Sexual systems in Silene. Sexual system description recognizes all the sexual systems described for the species in the literature. The sexual system assigned here is the principal or most frequent sexual system for the species according to our review. Species classification follows Oxelman et al. (2013). H, hermaphrodite; D, dioecious; Gd, gynodioecious; Gm, gynomonoecious; Ad, androdioecious; Am, andromonoecious and T, trioecious. Mixed sexual systems are denoted by a dash. Subgenus, section, species Sexual system Described in literature Assigned Subgenus Behenantha (Otth) Endl. Section Behenantha Otth S. pendula L. Gd 1,2,51 Gd S. uniflora Roth Gd 3,4,5 Gd S. vulgaris (Moench) Garcke Am–Ad 6 ;Gd 2 ;Gd–Gm 6,7,8,9,10 Gd–Gm Section Conoimorpha Otth S. conica L. H 2,7 ;H–Gm 2 H S. conoidea L. H 1,2 H S. subconica Friv. Gd 2 Gd Section Dichotomae (Rohrb.) Chowdhuri S. dichotoma Ehrh. Gd 2,6,7 Gd Section Elisanthe (Fenzl) Fenzl S. noctiflora L. H 11 ;H–Gm 2,7 ;Gm 12,13 ;Gd–Gm 1 Gm Section Erectorefractae Chowdhuri S. germana Gay H 51 H Section Melandrium (Ro ¨hl.) Rabeler S. astrachanicum (Pacz.) Takht. D 14 D S. diclinis (Lag.) M.Laı ´nz D 1,8,14,15,16,17 D S. dioica (L.) Clairv. Am 6 ;D 2,6,7,8,14,15 D S. integripetala Bory and Chaub. H–Gm 30 H Section Viscosae (Boiss.) C.L.Tang S. viscosa (L.) Pers. H 2,7 H Others S. acutifolia Link ex Rohrb. H 8,31 H S. elisabethae Jan H 6,7 H Subgenus Silene Section Auriculatae (Boiss.) Schischkin S. disticha Willd. H 2,8 H S. echinata Otth H 8 ;H–Gm 2 H S. linicola C.C.Gmel. H 2,6,8 H S. schafta J.G.Gmel. ex Hohen. Gd 2 Gd S. spergulifolia (Willd.) M.Bieb. H 2 H S. vallesia L. Gd 2,7 Gd Section Silene S. apetala Willd. H 1,2 H S. ciliata Pourr. H–Gm 32 H Continued AoB PLANTS www.aobplants.oxfordjournals.org &The Authors 2015 5 Casimiro-Soriguer et al. — Sexual systems in Silene and section Psammophilae .......................................................................... ............................................................................................................................................................................ Table 1. Continued Subgenus, section, species Sexual system Described in literature Assigned S. colorata Poir. H 8,33 ;H–Gm 2 H S. gallica L. H 8 ;H–Gm 2 H S. gracilis DC. H 51 H S. micropetala Lag. H 8 ;H–Gm 2 H S. nicaeensis All. H 2,8,51 H S. nocturna L. H 1,2,8 H S. pseudoatocion Desf. Gd–Gm 2 Gd–Gm S. ramosissima Desf. H 8,51 H S. scabriflora Brot. H 51 H S. roemeri Friv. Gd–Am 15 ;H–Gd 2 Gd S. saxifraga L. Am–Gm–T 6 ;Gm 1 ;Gd–Gm 2,7 Gd–Gm S. sendtneri Boiss. D 15 D S. senneni Pau H 45 H S. thessalonica Boiss. and Heldr. H–Gm 2 H S. viridiflora L. H 7 ;Gd–Gm 2 Gd–Gm S. waldsteinii Griseb. H–Gm 2 H S. wolgensis (Hornem.) Otth D 36 D Section Spergulifoliae (Boiss.) Schischkin S. brahuica Boiss. Gd 21 Gd Others S. bupleuroides L. H 2,7 H S. heuffelii Soo ´D 14 D S. latifolia Poir. Am 6 ;D 2,6,7,8,14,15 D S. marizii Samp. D 8 D Section Physolychnis (Bentham) Bocquet S. caroliniana Walter H 18 H S. douglasii Hook. H 11,19 H S. gangotriana Pusalkar, D.K.Singh and Lakshmin H 20 H S. laxantha Majumdar Gd 20,21 Gd S. regia Sims H 22,23 H S. rotundifolia Nutt. H 23 H S. scouleri Hook. H 11 H S. stellata (L.) W.T. Aiton H 18,24 H S. tibetica Lide ´n and Oxelman H–Am 25 Am S. virginica L. H 18,26 H S. zawadzkii Herbich H 2 H Continued 6AoB PLANTS www.aobplants.oxfordjournals.org &The Authors 2015 Casimiro-Soriguer et al. — Sexual systems in Silene and section Psammophilae .......................................................................... ............................................................................................................................................................................ Table 1. Continued Subgenus, section, species Sexual system Described in literature Assigned Section Psammophilae (Talavera) Greuter S. adscendens Lag. Gd–Gm 50 Gd–Gm S. cambessedesii Boiss. and Reut. Gd–Gm 50 Gd–Gm S. littorea Brot. Gd–Gm 2,27,28,50 Gd–Gm S. psammitis Link ex Spreng Gd–Gm 50 Gd–Gm S. stockenii Chater Gd–Gm 29,51 Gd–Gm Section Sedoideae Oxelman and Greuter S. secundiflora Otth H 2,8 H S. sericea All. Gd 2 Gd S. succulenta Forssk. H 2 H Section Siphonomorpha Otth S. acaulis (L.) Jacq. D 2,6,7,14 ;T 6,7,15,34 ;Gd–Gm 35 D S. andryalifolia Pomel Gd 2 Gd S. borysthenica (Gruner) Walters D 15,36 D S. colpophylla Wrigley D 37 D S. cyri Schischkin D 14 D S. fernandezii Jeanm. H–Gm 8 H S. flavescens Waldst. and Kit. H–Gm 2 H S. fruticosa L. H 2 H S. gazulensis Gala ´n, Corte ´s, Orell. and Morales Alonso H 38 H S. gigantea L. H–Gm 39 H S. hayekiana Hand.-Mazz. and Janch. Gd 2 Gd S. hellmannii Claus D 14,15 D S. hifacensis Rouy Gd 23 ;Gd–Gm 40 Gd–Gm S. italica (L.) Pers. H 7 ;Gd 2 ;Gd–Gm 1,41,42 Gd–Gm S. multicaulis Guss. H 2 H S. multiflora (Ehrh) Pers. H 7 H S. nocteolens Webb and Berthel. H 38 H S. nutans L. Am–Ad 6 ;Gd–Gm 1,2,6,7,43 Gd–Gm S. otites (L.) Wibel Ad 6 ;D 1,2,6,7,15,36 D S. paradoxa L. H 2,23 H S. parnassica Boiss. and Spruner H 2 H S. patula Desf. H 44 H S. capitellata Boiss. H 46 H S. chlorantha (Willd.) Ehrh H 2,7,47 H S. cretica L. H 2,7,8 H S. friwaldskyana Hampe. H 2 H S. hawaiiensis Sherff H 23,48 H Continued AoB PLANTS www.aobplants.oxfordjournals.org &The Authors 2015 7 Casimiro-Soriguer et al. — Sexual systems in Silene and section Psammophilae and the gynomonoecious morphs (7.9 and 5.3 %, respectively). The proportion of hermaphroditic plants within populations ranged from 64.0 % (in S. littorea)to 99 % (in S. adscendens), whereas the proportion of female plants varied from 1.0 to 18.0 % (in S. littorea), and that of gynomonoecious plants ranged from zero (at least one population in each species) to 20 % (in S. psammitis)(Fig.2). The frequency of sexual morphs per population varied significantly among the studied populations (Wald x 2 ¼62.62, df ¼22, P,0.0001) but not among species (Wald x 2 ¼4.59, df ¼3, P¼0.21). On the whole, 92.1 % of the 7001 flowers analysed were hermaphrodites, and 7.9 % were females. At the species level, S. psammitis showed the highest proportion of female flowers (16.1 %) in the population and S. adscendens the lowest (4.8 %) [see Supporting Information]. The predominance of hermaphrodite flowers was also found at the population level; the mean percentage of female flowers per population ranged from 1.3 to 18.7 % in S. littorea, from 0.8 to 10.2 % in S. adscendens, from 1.1 to 14.1 % in S. cambessedesii and from 7.1 to 29.4 % in S. psammitis [see Supporting Information].On the other hand, the percentage of female flowers in gynomonoecious individuals was 34.8+2.0 % (mean+1 SE) in S. littorea, 34.5+5.2 % in S. adscendens,31.4+3.0 % in S. cambessedesii and 43.4 +3.1 % in S. psammitis [see Supporting Information]. The average number of flowers in female plants was generally smaller than in hermaphroditic or gynomonoecious plants [see Supporting Information]. The numberof flowers per individual showed significant differences among sex morphs (Wald x 2 ¼328.85, df ¼2, P,0.0001), populations (Wald x 2 ¼2126.73, df ¼22, P,0.0001) and species (Wald x 2 ¼571.88, df ¼3, P,0.0001). Diversity and frequency of sexual systems in Silene We found that 98 Silene species have been specifically studied or described in terms of the sexual system (Table 1). We have collected the data from 46 different species in addition to those formerly found by Desfeux et al. (1996) and Ju ¨rgens et al. (2002). The numberof species described in the subgenus Silene (63 species) is nearly double that in subgenus Behenantha (35 species) (Table 1). The most frequent sexual system at the genus level is hermaphroditism (58.2 %), followed by dioecy (14.3 %), gynodioecy (13.3 %) and gynodioecy–gynomonoecy (12.2 %). Interestingly, all four sexual systems are present in both subgenera, with a statistically similar frequency (P.0.43 for all the sexual systems, except for hermaphroditism, that showed marginally significant differences P¼0.09) (Fig. 3). In addition, one Gm and one Am species were found, and both belong to the subgenus Behenantha. The fact that 13 of our assigned H species (21.1 %) are described as H–Gm in the literature is worthy of mention. There are some sections whose species present mainly the same sexual system. For instance, section Melandrium are all dioecious, section Psammophilae are all Gd–Gm and section Physolychnis are all hermaphroditic except one species (S. laxantha, Table 1). Other sections seem more variable. Thus, section Silene includes H, Gd and Gd–Gm species, and section Siphonomorpha includes all the sexual systems present in the subgenus; however, it can be said that both sections have the highest number of species with known sexual systems. .......................................................................... ............................................................................................................................................................................ Table 1. Continued Subgenus, section, species Sexual system Described in literature Assigned S. inaperta L. H 1,2 H S. isaurica Contandr. and Que ´zel Gd 46 Gd S. kemoniana C. Brullo, Brullo, Giusso, Ilardi and Sciandr. H 49 H S. muscipula L. H 2 H S. portensis L. H 2,51 H S. struthioloides A.Gray H 23,48 H 1 Desfeux et al. (1996), 2 Ju ¨rgens et al. (2002), 3 Baker and Dalby (1980), 4 Pettersson (1997), 5 Warren and James (2008), 6 Knuth (1908), 7 Meusel and Mu ¨hlberg (1979), 8 Talavera (1990), 9 Glaettli and Goudet (2006), 10 Miyake and Olson (2009), 11 Touzet and Delph (2009), 12 Folke and Delph (1997), 13 Davis and Delph (2005), 14 Schischkin (1970), 15 Chater and Walters (1964), 16 Prentice (1976), 17 Montesinos et al. (2006), 18 Reynolds et al. (2009), 19 Kephart et al. (1999), 20 Pusalkar et al. (2004), 21 Tropicos.org (2014), 22 Dolan (1994), 23 Moyle (2006), 24 Castillo et al. (2013), 25 Oxelman et al. (2001), 26 Dudash and Fenster (2001), 27 Guitia ´n and Medrano (2000), 28 Casimiro-Soriguer et al. (2013), 29 Talavera et al. (1996), 30 Oxelman (1995), 31 Buide and Guitia ´n (2002), 32 Gime ´nez-Benavides et al. (2007), 33 Terrab et al. (2007), 34 Alatalo and Molau (2001), 35 Shykoff (1992), 36 Lihua et al. (2001), 37 Mrackova et al. (2008), 38 Ban ˜ares et al. (2004), 39 Ghazanfar (1989), 40 Prentice et al. (2003), 41 Maurice (1999), 42 Lafuma and Maurice (2006), 43 Dufay et al. (2010), 44 Naciri et al. (2010), 45 Martinell et al. (2010), 46 Yildiz and Çirpici (2013), 47 Lauterbach et al. (2011), 48 Westerbergh and Saura (1994), 49 Brullo et al. (2012), 50 Casimiro-Soriguer et al. present study and 51 E. Narbona, M. L. Buide and I. Casimiro-Soriguer, pers. observations. 8AoB PLANTS www.aobplants.oxfordjournals.org &The Authors 2015 Casimiro-Soriguer et al. — Sexual systems in Silene and section Psammophilae Figure 2. Frequency of hermaphroditic (grey), gynomonoecious (white) and female (black) individuals of species from section Psammophilae in each population. The number of individuals per sexual morph sampled in each population is shown elsewhere [see Supporting Information]. Figure 3. Proportion of sexual systems in subgenus Behenantha (black bars) and subgenus Silene (white bars). H, Hermaphroditism; D, dioecy; Gd–Gm, gynodioecy–gynomonoecy; Gd, gynodioecy; Gm, gynomonoecy; Am, andromonoecy. AoB PLANTS www.aobplants.oxfordjournals.org &The Authors 2015 9 Casimiro-Soriguer et al. — Sexual systems in Silene and section Psammophilae