Genetic Diversity, Genetic Drift and Mixed Mating System in Small Subpopulations of Dyckia Ibiramensis , a Rare Endemic Bromeliad from Southern Brazil

Conserv Genet DOI 10.1007/s10592-011-0183-3

RESEARCH ARTICLE

Genetic diversity, genetic drift and mixed mating system in small subpopulations of Dyckia ibiramensis , a rare endemic bromeliad from Southern Brazil

Karina Vanessa Hmeljevski • Ademir Reis • Tiago Montagna • Maurı´cio Sedrez dos Reis

Received: 5 July 2010 / Accepted: 6 January 2011 Ó Springer Science+Business Media B.V. 2011

Abstract Dyckia ibiramensis is a naturally rare, endemic adult subpopulations indicate pronounced genetic structure and threatened bromeliad which occurs naturally on 4 km ^0 (GST ¼ 0:674). D. ibiramensis has a mixed mating system ^ of rocky river outcroppings in Southern Brazil. For this and multilocus outcrossing rates tm were variable between study, subpopulations of the species were characterized subpopulations. This study demonstrates the importance of based on size and genetics, to compile information for in in situ preservation of all subpopulations for the mainte- situ and ex situ conservation strategies. A census of the nance of species diversity. For effective ex situ conserva- rosettes was undertaken for each subpopulation and seven tion, it would be necessary to collect seeds from 52 to 99 allozyme polymorphic loci were used to estimate genetic seed-rosettes, depending on the target population. diversity and structure of adults and offspring and assess the mating system. In general, the subpopulations were Keywords Allozyme Á Bromeliaceae Á Conservation Á small and most of the rosettes were aggregated into Founder effects Á Metapopulation clumps. The species showed a high genetic diversity ^ ^ (He ¼ 0:219) and signiﬁcant ﬁxation index ( f ¼ 0:642 ; P B 0.05). The estimate of differentiation among all Introduction

Small populations are more prone to demographic, environmental and genetic stochasticity (Lande 1988 ; Ooster- Electronic supplementary material The online version of this meijer et al. 2003 ) than large populations due to the loss of article (doi: 10.1007/s10592-011-0183-3 ) contains supplementary genetic variation through random genetic drift, increased material, which is available to authorized users. selfing, mating among related individuals, and inbreeding K. V. Hmeljevski ( &) (Ellstrand and Elam 1993 ; Honnay and Jacquemyn 2007 ). Instituto de Pesquisas Jardim Botaˆnico do Rio de Janeiro, Recent meta-analyses have found that population size has a Pacheco Leaõ 915, Jardim Botaˆnico, Rio de Janeiro, highly significant effect on population genetic diversity, RJ 22460-030, Brazil thus suggesting that small populations consistently contain e-mail: [email protected] significantly less genetic variation than large populations A. Reis (Aguilar et al. 2008 ; Honnay and Jacquemyn 2007 ; Leimu Laborato´rio de Restauraçaõ Ambiental Sisteˆmica, Departamento et al. 2006 ). According to Honnay and Jacquemyn ( 2007 ), de Botaˆnica, Centro de Cieˆncias Biolo´gicas, Universidade loss of alleles through population bottlenecks and random Federal de Santa Catarina, CP 476, Floriano´polis, SC 88040-900, Brazil genetic drift play an important role in the genetic impoverishment of plant populations. These recent studies also T. Montagna Á M. S. dos Reis suggest that there is no significant correlation between Nućleo de Pesquisas em Florestas Tropicais, Departamento de population size and inbreeding coefficients. Although Fitotecnia, Centro de Cieˆncias Agra´rias, Universidade Federal de Santa Catarina, Rodovia Admar Gonzaga 1346, Itacorubi, several explanations have been mentioned by the authors in Floriano´polis, SC 88034-001, Brazil an attempt to explain this, further research is necessary to

123 Conserv Genet examine the uncertain relationship between homozygote point out that the genetic differentiation between popula- excess and plant population size, because an excess in tions may be complicated considering metapopulation homozygotes affects short-term fitness and future adapt- dynamics, depending on whether new colonies are founded ability (Honnay and Jacquemyn 2007 ). If inbreeding and by individuals originating all from a single population (the drift are important constituents of current population fit- propagule-pool model) or randomly sampled from the ness, then heterozygosity and population size should be entire metapopulation (the migrant-pool model) (Pannell positively correlated with fitness among populations of a and Charlesworth 2000 ). species (Reed and Frankham 2003 ). Nevertheless, Tero et al . (2003 ) and Honnay et al. The strength and tendency of the relationships among (2010 ) emphasize that genetic diversity and genetic dif- parameters such as plant population size, fitness and ferentiation of plant populations along rivers can be genetic variation may depend on different plant charac- understood only by incorporating a metapopulation per- teristics, especially life span, mating system and rarity spective. From a practical conservation approach, knowl- (Leimu et al. 2006 ). Endemics and rare species are believed edge of the distribution of genetic variation is important to have reduced genetic variability (Cole 2003 ; Ellstrand because it can be valuable in determining how many and and Elam 1993 ; Hamrick and Godt 1989 ; Karron 1991 ; which populations to protect as well as guiding policies for Kruckeberg and Rabinowitz 1985 ); this is mainly attributed seed collection and the establishment of new populations to the influence of evolutionary forces like genetic drift and (Baskauf and Burke 2009 ). selection or events such as genetic bottlenecks and founder Dyckia ibiramensis is a rare endemic bromeliad from effect (Barrett and Kohn 1991 ; Karron 1991 ). The natural Southern Brazil. The species occurs in only nine patches rarity of the species can also determine susceptibility to distributed along 4 km of the rocky banks of a river. In genetic erosion, since once common and now rare species 1992, it was classified as Critically Endangered and since are expected to experience a greater impact on genetic then it has remained on the Official List of the Endangered diversity than naturally rare species (Aguilar et al. 2008 ). Species of the Brazilian Flora (MMA 2008 ). Nevertheless, Plant species considered to be rare can display a wide range in 2004 authorization was given to build a small hydro- of distribution patterns and maintain different levels and electric facility in the region where the species occurs. patterns of genetic diversity (Gitzendanner and Soltis 2000 ; Development in this area has become the biggest threat to in Holmes et al. 2009 ). situ conservation of D. ibiramensis in the short-term and has Many plant populations are naturally isolated, small, and generated demand for more knowledge about the ecology subdivided into local breeding units, a condition that can and genetics of the species in order to undertake suitable lead to the genetic differentiation of local demes (Tero preservation strategies. In this context, the aim of this study et al. 2003 ). Meanwhile, riparian plant populations can is to investigate the size of each subpopulation, the effective occur as discrete patches because of the erratic distribution size, as well as the population genetics of adults and off- of suitable, often ephemeral, habitats along rivers (Honnay spring of D. ibiramensis to compile information for in situ et al. 2010 ). Studying the riparian species Silene tatarica , and ex situ conservation strategies. We tested the hypothesis Tero et al. ( 2003 ) described five different possible models that the species will show low levels of genetic diversity, of population structure and migration patterns between high genetic drift, and structured subpopulations, because of linearly arranged patches or subpopulations: (a) genetically its restricted and clumped distribution. From this perspec- uniform population with free gene flow across the popu- tive, several questions were raised: (1) what is the total lation; (b) fragmentized population without recurrent gene number of rosettes and the number of fertile rosettes in each flow between the subpopulations; (c) stepping-stone pop- subpopulation? (2) What is the level of genetic diversity and ulation; (d) source-sink population; and (e) ‘classical’ how it is distributed among subpopulations? (3) Which metapopulation. The last two models are metapopulation subpopulations are priorities for in situ conservation and models in which the subpopulations are more or less how many seed-rosettes are necessary for seed collection ephemeral. In a recent study on riparian and aquatic plant for ex situ conservation programs? species, Honnay et al. ( 2010 and references there in) determine that seed flow between linearly arranged populations may occur in two ways: predominantly between Materials and methods adjacent populations, where genetic distances between populations are expected to increase monotonically with Study species and study area geographical distance; and secondly, by long-distance seed dispersal between non-adjacent populations, resulting in Dyckia ibiramensis Reitz (Bromeliaceae) is a naturally rare limited effects of isolation due to distance and little genetic and endemic species found in the city of Ibirama, State differentiation among populations. However, the authors of Santa Catarina, Brazil (Reitz 1983 ). It is classified as

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Fig. 2 The ﬁgure shows a clump and diameter classes: smaller than 15 cm (class 1), between 15 and 30 cm (class 2) and larger than 30 cm (class 3)

also registered to determine the effective size. The spatial distribution of the rosettes was identified as an isolated rosette or a clump. Fig. 1 Location of the subpopulations of D. ibiramensis along Itajaı´ do Norte River, Santa Catarina State, Brazil Sampling and laboratory methodology rupicolous, heliophyte and rheophyte (Klein 1990 ). The In early 2008, leaf samples of fertile rosettes were collected species occurs as discrete patches or subpopulations, dis- from P1, P3, P4, P6, P7, and P8, for a total of 271 samples. tributed along approximately 4 km of the rocky banks of Since the genetic similarity is unknown within each clump, the Itajaı´ do Norte River (Fig. 1). The plants grow on rock the sampling strategy was to collect from only one indi- fractures, next to the beds of swift-running streams. They vidual per clump in order to avoid sampling the same are subject to strong river currents and habitats are often genet. This strategy was possible in subpopulations where submerged in times of floods and experience drought and the number of fertile rosettes was greater than 30. In sub- sun exposure during the ebb (Klein 1979 ). D. ibiramensis populations with an effective size of less than 30, samples shows iteroparity, therefore the rosette does not die after a from all the fertile rosettes were collected. In January 2006, reproductive event, and the plant exhibits clonal growth, seeds were collected from randomly selected seed-rosettes resulting in the formation of several clumps. A clump of the subpopulations P4, P6, and P7. Seeds were germi- consists of a spatial aggregation of individuals (Fig. 2) with nated in a greenhouse under vermiculite and organic sub- a variable number of rosettes in close proximity, whose strate. After 1 year, leaf samples were collected from 10 genetic similarity is unknown. The carpenter bee Xylocopa seed-rosettes (23–26 seedlings/seed-rosette) originating (Neoxylocopa ) brasilianorum and hummingbird Thalura- from each point (P4, P6, and P7), totaling 30 seed-rosettes. nia glaucopis are the main species’ pollinators (Hmeljevski Genetic characterization was carried out using allozyme 2007 ). Seeds are dispersed by wind (Downs 1974 ), gravity, markers, according to the methodology of Kephart ( 1990 ) and possibly by water. The study was conducted in the city and Alfenas ( 1998 ). For the migration of the enzymes, gels of Ibirama, and all patches (P1–P9) of D. ibiramensis were prepared with 13% hydrolyzed starch (penetrose 30) were studied (Fig. 1), referred to here as ‘‘subpopulations’’. used. For the extraction of the enzymes, leaf tissue was ground with a mortar and mixed with extraction solution 1 Subpopulation sizes and effective size following Alfenas ( 1998 ). The buffers histidine (H—tri- sodium citrate 105.82 g/l, pH 8.0) and tris citrate (TC—tris In January 2008, all rosettes in all subpopulations were 27 g/l, citric acid 16.52 g/l, pH 7.5) were used in electro- counted, to determine subpopulation size, and classified phoresis (Alfenas 1998 ). Staining recipes followed the according to diameter: smaller than 15 cm (class 1), methodology described by Alfenas ( 1998 ). Seven enzyme between 15 and 30 cm (class 2) and larger than 30 cm systems were studied: 6-phophogluconate dehydrogenase (class 3) (Fig. 2). The reproductive state of the rosettes was (locus 6pgdh , E.C. 1.1.1.44), malic enzyme (locus Me1 ,

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^ ^ E.C. 1.1.1.40) and phosphoglucoisomerase (locus Pgi2 , effective number of pollen donors ( Nep ¼ 1rpðmÞ), E.C. 5.3.1.9) for H; and esterase (fluorescent) (loci Est2 described by Ritland ( 1989 ). Additionally, the coefficient and Est3 , E.C. 3.1.1.1), phosphoglucomutase (locus Pgm , of correlation among offspring within progenies was esti- E.C. 5.4.2.2), malate dehydrogenase (locus Mdh2 , E.C. mated according to Ritland ( 1989 ) as r^xy ¼ 0:25 ð1 þ 1.1.1.37) and peroxidase (loci Prx2 and Prx3 , E.C. F^ Þ½ 4s^þ ð ^t2 þ ^ts^r^ Þð 1 þ r^ Þ , where F^ is the coefficient 1.11.1.7) for TC. Locus 6pgdh was identified only in off- p s pðmÞ p ^ spring and used only in the mating system analysis. of inbreeding in the parental generation (here we used the f values of each correspondent fertile adults’ population) and ^ Genetic diversity and structure s^ is the selfing rate ( s^ ¼ 1 À tm). The coancestry coefficient ^ within progeny was estimated from r^xy as hxy ¼ r^xy =2 Genetic diversity was analyzed using the estimated allele (Sebbenn 2002 ). The variance effective size was estimated ^ ^ ^ frequencies, the average number of alleles per locus ( A), as NeðvÞ ¼ 0:5=hxy (Cockerham 1969 ). The necessary ^ effective number of alleles per locus ( Ae), observed het- number of seed-rosettes for seed collection with the aim of ^ ^ erozygosity ( Ho), expected heterozygosity in Hardy– retaining a reference effective population size ( Neðreference Þ) ^ ^ ^ ^ ^ Weinberg equilibrium ( He), and fixation index ( f ). The of 100 was estimated as m ¼ Neðreference Þ=NeðvÞ (Bittencourt ^ statistical significance of f values was evaluated using and Sebbenn 2007 ). permutations. The effective number of alleles per locus was ^ ^ calculated by Ae ¼ 1=ð1 À HeÞ. In order to investigate if ^ Results there were differences in the levels of genetic diversity ( Ho ^ ^ and He) and f between fertile adults and offspring, the one- sided test described by Goudet ( 2002 ) was used and the Subpopulation size and effective size significance was tested using permutations. All of these ^ The subpopulation sizes of D. ibiramensis were in general analyses, except Ae, were run using the program FSTAT ^ ^ ^ small, but variable. Approximately 15% of the total version 2.9.3.2. (Goudet 2002 ). The values of He; A; and f rosettes were fertile, of which 41% were classified as size were correlated with the subpopulation size and effective class 3. Although class 1 contained the majority of the total size for adults using a Spearman rank correlation. number of rosettes, it had only 0.13% of the fertile rosettes. The genetic differentiation was investigated separately Only subpopulations P4, P6, and P7 had a considerable among all subpopulations’ adults and offspring by the number of fertile rosettes ( [50), while no fertile rosettes 0 standardized G ST statistic (Hedrick 2005 ). In order to study were identified in P2, P5, and P9. the association between pair wise population differentia- The spatial distribution of the rosettes in the subpopu- ^0 ^0 tion [ GST =ð1 À GST Þ] and linear geographic distances lations showed an aggregated distribution with the number among subpopulations, the Mantel test was applied using of clumps being considerably larger than the number of the TFPGA program version 1.3 (Miller 1997 ). isolated rosettes. In all subpopulations few fertile isolated rosettes were registered. Mating system Genetic diversity and structure The mating system was analyzed on the basis of the mixed mating model and correlated mating model, implemented From the 9 allozyme loci tested, 7 were polymorphic and 2 in the software Multilocos MLTR version 3.4 (Ritland were monomorphic ( Pgi2 and Prx2 ). The polymorphic loci 2009 ). The estimated parameters were the multilocus out segregated from 1 to 3 alleles, with a total of 18 alleles in ^ crossing rate ( tm) using the Expectation–Maximization all subpopulations. In general, the allele frequencies were ^ method, single-locus out crossing rate ( ts), out crossing rate very similar between adults and offspring; however, allele ^ ^ among relatives ( tm À ts), allele frequencies of the ovules 2 of locus Mdh2 appeared only in the offspring of P4 and ^ and pollen, selfing correlation ( rs) and multilocus paternity P6, and alleles 1 and 3 of locus Est3 were exclusive to P4 ^ correlation ( rpðmÞ). The standard error ( SE ) of the estimates (and its offspring) and P3, respectively (see Appendix S1 in was obtained by 1,000 bootstraps among progenies. The Supplementary Material). It is important to mention the homogeneity of the allele frequencies of the ovules and presence of many alleles at an intermediate frequency ^ pollen was tested using FST (Nei 1977 ) and its significance (0.05 \ P B 0.25) in all subpopulations as well as very provided by the v2 test (Workman and Niswander 1970 ). rare alleles ( P B 0.01) in offspring, suggesting the occur- ^ From rpðmÞ parameter it was possible to estimate the rence of genetic drift. This is reflected in the effective

123 Conserv Genet number of alleles per locus, which was lower than the Mating system number of alleles per locus (Table 2). Observed and expected heterozygosities varied from low Heterogeneity of ovules and pollen allele frequencies was to high in adult subpopulations and offspring, but the overall detected for all loci, except for Pgm1 in P6 offspring (see ^ He for both was high (Table 2). The majority of the genetic Appendix S2 in Supplementary Material), indicating non- diversity was found in P4, a subpopulation located in the random sampling of the pollen pool by each seed-rosette. center of the species’ distribution. The genetic diversity Dyckia ibiramensis has a mixed mating system, showed ^ seems to decrease with distance from the centre of the spe- by subpopulations’ tm, which varied from 0.893 (P4 off- cies’ range, reaching zero at the limits of its distribution spring) to 0.563 (P7 offspring) and were significantly dif- (Fig. 1, Table 2). There was a significant excess of homo- ferent from unity (Table 3). Data suggest that there are few zygotes in each subpopulation, except for P3 which showed a or no matings among related individuals, as shown by the ^ ^ ^ negative value for f . Comparing the adults with offspring minimal differences between tm and ts (Table 3). The ^ (Table 2), there were no significant differences in the aver- values of rs were significant in all offspring, suggesting high variation in individual outcrossing rate. The high and age observed and expected heterozygosities ( Pone-tailed = significant values of r^ indicate that many of the off- 0.592 and Pone-tailed = 0.551, respectively) and fixation pðmÞ index ( Pone-tailed = 0.406). spring have the same paternal and maternal genitor, it Because of the significant pairwise association between means that part of the offspring were full-sibs. These ^ population size and effective size ( r = 1, P \ 0.001), the results are consistent with the estimates of Nep (Table 3), values of correlation calculated with these two variables which imply that, on average, 3 pollen donors contributed are presented together. A significant positive correlation to the production of offspring. In the offspring of P4, the ^ ^ was found between He and population size/effective size hxy was close to that expected for full-sibs ( hxy ¼ 0:250). In (r = 0.9429, P = 0.004) and between A^ and population the offspring of P6 and P7 the results were higher and, ^ size/effective size ( r = 0.9276, P = 0.008). There was no consequently, the estimates of NeðvÞ were lower (Table 3) ^ ^ significant association between f and population size/ than expected in panmitic populations ( NeðvÞ ¼ 4). effective size ( r = - 0.3000, P = 0.6238). Accordingly, the estimated m^ varied from 52 to 99 seed- The estimate of differentiation among all adult sub- rosettes (Table 3). ^0 populations shows a significantly high value ( GST ¼ 0:674), suggesting that approximately 68% of genetic diversity was distributed among subpopulations and 32% within subpopulations. When the analysis was run for Discussion ^0 offspring, the value was reduced ( GST ¼ 0:346). Mantel test revealed non-significant ( r = 0.4813, P = 0.1175; High diversity, founder effect and genetic drift Fig. 3) isolation due to distance between subpopulations of D. ibiramensis . Dyckia ibiramensis is a naturally rare bromeliad, distributed in small subpopulations, each unique in its composition. This variety is the result of the habitat, effective size (Table 1), high levels of genetic diversity variation (Table 2) and significant genetic divergence among the ^0 subpopulations ( GST ¼ 0:674). The overall genetic diversity ^ of the species ( He ¼ 0:219) was high compared to other species of Bromeliaceae (Soltis et al. 1987 ; Murawski and Hamrick 1990 ; Izquierdo and Pin˜ero 2000 ; Sarthou et al. 2001 ; Gonza´lez-Astorga et al. 2004 ), monocotyledons in general (Hamrick and Godt 1989 ), and even in comparison with rare (Cole 2003 ) and endemic species (Hamrick and Godt 1989 ; Godt and Hamrick 1996 ). According to Leimu et al . (2006 ), hybridization, recent speciation, multiple ori- gins or recent population bottlenecks may result in high levels of genetic variation in rare species. Similar to results found by recent meta-analyses ^0 ^0 Fig. 3 Plots of Mantel test for the correlation between GST =ð1 À GST Þ (Aguilar et al. 2008 ; Honnay and Jacquemyn 2007 ; Leimu and geographic distance et al. 2006 ), positive correlations between population size/

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Table 1 Area of occupancy, number of clumps, isolated rosettes, total rosettes by size classes, population size and effective size for each subpopulation of D. ibiramensis Subpopulation Area of occupancy Number of Number of isolated Total number of rosettes Population size/ (m 2) clumps rosettes effective size Class 1 Class 2 Class 3

P1 245 5/1 1/1 3/0 10/0 15/2 28/2 P2 16 9/0 2/0 26/0 24/0 12/0 62/0 P3 27 8/3 12/0 55/0 26/0 30/13 111/13 P4 8400 260/122 101/1 1174/3 1128/159 969/326 3271/488 P5 40 6/0 3/0 25/0 15/0 5/0 45/0 P6 2175 41/20 19/1 152/0 83/20 173/101 408/121 P7 936 313/132 116/2 1643/1 966/124 748/369 3357/494 P8 228 10/2 4/0 38/0 10/0 35/7 83/7 P9 – 0/0 1/0 1/0 0/0 0/0 1/0 Overall 12067 652/280 259/5 3117/4 2262/303 1987/818 7366/1125 Data are: number of rosettes/number of fertile rosettes

Table 2 Allozyme genetic diversity indices and sample size ( n) for impoverishment of plant subpopulations in the upper fertile adults and offspring of D. ibiramensis reaches of the river, and also no distance isolation. The ^ ^ ^ ^ ^ subpopulation variation in genetic diversity associated with Subpopulation n A Ae Ho He f the occurrence of genetic drift in offspring and high genetic Fertile adults differentiation may reflect a colonization and extinction P1 2 1.0 1.0 0.000 0.000 – dynamic, strongly influenced by the founder effect. When P3 13 1.4 1.1 0.115 0.098 -0.180 new populations are established by a small number of P4 95 1.8 1.3 0.152 0.240 0.369* original founders, heterozygosity may decrease due to P6 44 1.4 1.1 0.023 0.104 0.780* random drift. This is compounded by inbreeding which is P7 110 1.6 1.1 0.040 0.112 0.644* associated with increases in the levels of homozygosity P8 7 1.1 1.0 0.000 0.036 1.000 (Templeton 1980 ). Studies with others riparian species also Overall 271 2.0 1.3 0.064 0.219 0.642* found highly significant genetic differentiation among Offspring populations (Tero et al. 2003 ; Honnay et al. 2010 ). P4 255 2.0 1.3 0.182 0.248 0.269* In a metapopulation scenario, it is possible that the P6 250 1.9 1.1 0.024 0.094 0.741* colonization of new patches of D. ibiramensis occurs P7 244 1.8 1.2 0.068 0.146 0.530* according to the propagule-pool model (Pannell and Overall 749 2.0 1.2 0.092 0.199 0.539* Charlesworth 2000 ). Furthermore, the source-sink meta- ^ ^ population structure (Tero et al. 2003 ) seems to be plau- A number of alleles per locus; Ae effective number of alleles per ^ ^ sible in this species, with P4 being the actual source locus; Ho observed heterozygosity; He expected heterozygosity; f^ fixation index subpopulation. * P B 0.05 The mating system is another factor that can influence the spatial distribution of genetic variation within and ^ ^ effective size and He and A were significant. In contrast, a among populations (Loveless and Hamrick 1984 ). In sel- positive correlation was not found for f^. The loss of alleles, fing plants, like D. ibiramensis , most of the genetic varia- detected with allozyme markers, suggests a strong random tion is found among populations (Loveless and Hamrick genetic drift in D. ibiramensis subpopulations. In addition, 1984 ; Hamrick and Godt 1989 ). increased selfing and mating among closely related indi- Although the lower genetic differentiation found among ^0 viduals may result in inbreeding and a reduction of the the offspring ( GST ¼ 0:346) in comparison with the adult number of heterozygotes (Honnay and Jacquemyn 2007 ). subpopulations may indicate effective pollen flow among In self-compatible species, inbreeding could be high in subpopulations, further studies are needed to clarify these small populations irrespective of their size. results. As stated by Honnay et al . (2010 ), inferring gene- As evidenced by Honnay et al . (2010 ) for other riparian flow patterns from simple measures of genetic differenti- species, D. ibiramensis showed no indication of down- ation or isolation by distance patterns is inappropriate stream accumulation of genetic diversity or genetic in the riparian ecosystems because of genetic diversity

123 Conserv Genet distribution in this ecosystem simply cannot be understood the pollen is deposited in the first flower visited after col- without metapopulation theory. lection. The amount of pollen deposited in subsequent flowers decreases rapidly; less than 1% of the collected Mixed mating system pollen survives on pollinator after eight successive visits to flowers (Richards 1997 ). The analysis of the reproductive system of D. ibiramensis The respective values of coancestry were greater or showed that the species has a mixed mating system. The similar to that expected for full-sibs in all subpopulations estimates of selfing correlation were high (Table 3), indi- (Table 3). These high estimates are probably due to the cating that there is strong tendency of some seed-rosettes to association of selfing and mating among relatives with produce more descendants from selfing (or matings) than the correlated outcrossings, resulting in an increase of the others (Gusson et al. 2006 ). coancestry above the expected value for offspring of half- Paternity correlations revealed correlated mating in the sibs (Sebbenn 2003 ). According to Sebbenn ( 2003 ), this subpopulations, indicating that offspring are not composed reduction in the variance effective size (Table 3) results in exclusively by half-sibs, but rather of a mixture of half- the need to collect a larger number of seed-rosettes for ex sibs, full-sibs and selfing-sibs. This result is consistent with situ conservation than would be necessary if outcrossing the heterogeneity detected in the allele frequencies of were perfectly panmictic. pollen and ovules, which indicates that the pollen con- tributing to the formation of the seedlings did not originate Implications of the results for in situ and ex situ from a representative sample of the population. Rather, the conservation pollen originated systematically from the same plants. High estimates of paternity correlations are common in species Considering the results discussed in this study and taking with aggregated distribution that depend on pollinator into account that D. ibiramensis is a rare, endemic, and insects (Kageyama et al. 2003 ). Observations of the endangered bromeliad, the maintenance of all existent behavior of the pollinators of D. ibiramensis (Hmeljevski subpopulations in situ is necessary for its conservation in 2007 ) are consistent with this situation as they systemati- the mid- and long-term. However, subpopulation P4 pre- cally visit all the open flowers of an inflorescence and sents the best characteristics for promoting the success of subsequently move to the next closest inflorescence. In the perpetuation of the species in its natural environment spite of small populations favoring crosses between dif- and is therefore essential for conservation. ferent populations (Ellstrand and Elam 1993 ), plants with If construction of the small hydroelectric facility noted clonal growth increase floral display through the multipli- in the introduction is permitted to go forward, subpopula- cation of flowering ramets (Charpentier 2002 ). This can tions P2, P3, P4, P5, and part of P6, would be completely interfere in the behavior of the pollinators causing more submerged. This would result in the flooding of 50% of the dislocations between nearby inflorescences due to the species’ rosettes and the loss of the subpopulation with the availability of resources of grouped form, reducing so the most significant genetic diversity. Fortunately, because of middle distance of dispersal of pollen (Richards 1997 ). the results presented herein along with the incompatibility The average number of plants that contributed to the of construction plans with the conservation of D. ibiram- pollination in each population was relatively low (Table 3). ensis in its natural habitat, the environmental agency has Besides the behavior of pollinators, another factor that can not approved the construction of the hydroelectric facility. influence this low value is that, typically, more than 50% of As a consequence, the company responsible for the facility

Table 3 Estimates of the Parameters P4 P6 P7 mating system parameters of the D. ibiramensis ^ a Multilocus outcrossing rate ( tm) 0.893 (0.034) 0.774 (0.052) 0.563 (0.039) ^ Single-locus outcrossing rate ( ts) 0.896 (0.032) 0.760 (0.047) 0.561 (0.035) ^ ^ Outcrossing rate among relatives ( tm À ts) -0.003 (0.015) 0.014 (0.018) 0.002 (0.015)

Selﬁng correlation ( r^s) 0.091 (0.038) 0.425 (0.105) 0.747 (0.093) ^ Multilocus paternity correlation ( rpðmÞ) 0.326 (0.059) 0.470 (0.072) 0.316 (0.062) ^ Number of pollen donors ( Nep ) 3.1 2.1 3.2 ^ Coancestry coefﬁcient within progeny ( hxy ) 0.256 0.421 0.495 ^ Variance effective size ( NeðvÞ) 1.952 1.186 1.011 a Number of seed-rosettes to retain the 52 85 99 SE obtained by 1,000 effective size of 100 ( m^) bootstraps

123 Conserv Genet has suggested a relocation of the dam. This new proposal Downs RJ (1974) Anatomy and physiology. In: Smith LB, Downs RJ will result in the submersion of subpopulations P2 and P3, (eds) Pitcairnioideae ( Bromeliaceae ), flora neotropica mono- graph. Hafner Press, New York, pp 2–28 which correspond to 2.6% of the rosettes and will no longer Ellstrand NC, Elam DR (1993) Population genetic consequences of affect subpopulation P4. The company has also proposed small population size: implications for plant conservation. Annu the creation of a specific conservation area that will include Rev Ecol Syst 24:217–242 all remaining subpopulations of D. ibiramensis (Hmeljev- Gitzendanner MA, Soltis PS (2000) Patterns of genetic variation in rare and widespread plant congeners. Am J Bot 87:783–792 ski and Reis 2009 ). Godt MJW, Hamrick JL (1996) Conservation genetics os endemic For ex situ conservation of the species, our results plant species. 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