Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
doi: 10.1111/j.1420-9101.2007.01446.x Correlations between sex rate estimates and fitness across predominantly parthenogenetic flatworm populations T. G. D’SOUZA & N. K. MICHIELS Animal Evolutionary Ecology, Zoological Institute, Faculty of Biology, University of Tuebingen, Tuebingen, Germany Keywords: Abstract evolution of sex; fitness; genetic diversity; heritability; natural selection; occasional sex; parthenogenesis; Platyhelminthes; pseudogamy; reproduction. One explanation for the success of sexual reproduction is that sex increases the efficacy of natural selection. Recombination and segregation lead to fitness variance among offspring which then offers a wider target for natural selection. Consequently, adaptation to changing environments is accelerated and population mean fitness will increase.We investigated whether low levels of sex are associated with increased fitness variance and mean in parthenogenetic biotypes of the planarian flatworm Schmidtea polychroa. Parthenogenetic S. polychroa are triploid and reproduce clonally with occasional sexual reproduction. By-products and measures of occasional sex are the local presence of tetraploids and elevated levels of genotypic diversity. We correlated the proportion of tetraploids and genotypic diversity with fitness attributes of six genetically differentiated locations within one meta-population. Results indicate strong, positive correlations with variance and with mean offspring number produced during a 5-week period. The ecological and evolutionary implications for the maintenance of parthenogenetic S. polychroa are discussed. Introduction Sexual reproduction is believed to be beneficial because it creates genetic variation in fitness among offspring and therefore increases the efficacy of natural selection. This hypothesis originally dates back to Weismann (1886). Several theories explaining the maintenance of sex are based on Weismann’s principle, such as the Fisher– Muller hypothesis, Muller’s ratchet and the deterministic mutation hypothesis (Kondrashov, 1993; Burt, 2000). For instance, through recombination and segregation, independently arisen beneficial mutations can be united in the same genome and therefore offspring with above average fitness can be produced (Fisher–Muller hypothesis, Bell, 1982). Moreover, sex enhances the fixation of beneficial mutations by separating them from deleterious background (‘Ruby in the Rubbish’ hypothesis, Peck, 1994). As with beneficial mutations, sex also concenCorrespondence: Thomas D’Souza, Animal Evolutionary Ecology, Zoological Institute, Faculty of Biology, University of Tuebingen, Auf der Morgenstelle 28, D-72076 Tuebingen, Germany. Tel.: +49 (0)7071 2974842; fax: +49 (0)7071 295634; e-mail: [email protected] 276 trates deleterious mutations (Deterministic mutation hypothesis). Offspring carrying deleterious mutations of both parents will be negatively selected. Hence, sex allows for the more efficient elimination of deleterious mutations from the population (Kondrashov, 1984, 1988). In purely clonal populations variability among offspring can be created only by mutations. Moreover, in the absence of recombination and segregation the ability to concentrate beneficial or harmful mutations into single offspring is lacking. In addition, beneficial mutations can be trapped in a deleterious genetic background, which reduces the likelihood of fixation (Peck, 1994; Rice, 2002). Hence, adaptability of asexuals can only be achieved under restricted conditions such as high rates of beneficial mutations or large population sizes (Colegrave, 2002; De Visser & Rozen, 2005). Thus the response to natural selection should be low in asexuals relative to sexuals and, consequently, adaptation should be hampered (but see Doroszuk et al., 2006). In a strictly Weismannian sense, the benefit of sex is an increase in fitness variance, not mean, allowing a more effective response to selection in the long term (Burt, ª 2007 THE AUTHORS. J. EVOL. BIOL. 21 (2008) 276–286 JOURNAL COMPILATION ª 2007 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY Sex rate estimates and fitness in parthenogens 2000; Agrawal, 2006). However, in order for this to work negative fitness correlations across loci are required (Kondrashov, 1993; Barton & Charlesworth, 1998; Burt, 2000). In the presence of positive correlations, sex actually causes decreased variance (Agrawal, 2006). Moreover, destroying positive associations (e.g. favourable gene combinations) typically reduces mean fitness (Barton & Charlesworth, 1998). Hence, the advantage of sex is that it breaks up negative associations among loci, which results in increased fitness variation (Burt, 2000; Agrawal, 2006). Negative associations may be caused deterministically by epistatic interactions between loci or stochastically by genetic drift in finite populations (Kondrashov, 1993; Peters & Otto, 2003; De Visser & Elena, 2007). The effect of sex to increase genetic variance in fitness is beneficial under constant selection if epistasis is negative. Alternatively, fluctuating selection and ⁄ or random genetic drift in finite populations may also create those benefits of sex (Peters & Otto, 2003 and references therein). Especially negative associations arising stochastically slow down natural selection (Hill–Robertson effect), and therefore are likely to be longer preserved than positive associations (Peters & Otto, 2003). Theoretical models including genetic drift as a source of negative associations seem less restrictive and hence more broadly applicable in explaining the ubiquity of sex than models with negative epistasis (Otto & Gerstein, 2006). Assuming negative associations among loci, the benefit of sex in increasing the efficiency of natural selection applies both to individual and group selection (Barton & Charlesworth, 1998; Burt, 2000). However, in order to be strong enough to offset a twofold cost of sex (Williams, 1975; Maynard Smith, 1978), group selection is more important in Weismann’s argumentation: The selective advantage of a modifier gene that increases the rate of recombination is much smaller compared to effect of sexual clade competing with an asexual clade (Burt, 2000; but see Holmström & Jensen, 2004). Most evidence for Weismann’s principle focuses on the increased adaptation rate of recombining compared with nonrecombining populations. A faster response to selection is seen as an indicator for an increased genetic variance in fitness by sex. Indications for faster adaptation has been shown for recombining regions ⁄ chromosomes compared with nonrecombining ones in several Drosophila species (Rice & Chippindale, 2001; Bachtrog & Charlesworth, 2002; Betancourt & Presgraves, 2002; Bachtrog, 2003; Presgraves, 2005). Induction of sexual episodes increased adaptation to novel environments in yeast (Zeyl & Bell, 1997; Grimberg & Zeyl, 2005) and in the green alga Chlamydomonas reinhardtii (Colegrave, 2002; Colegrave et al., 2002; Kaltz & Bell, 2002). However, direct evidence of how sex affects the mean and variance of fitness in natural systems is scarce (Burt, 2000; Agrawal, 2006). Furthermore, it is difficult to draw 277 any general conclusions about the benefits of sex from these few studies (Agrawal, 2006). In the present study we ask whether rare sex is accompanied by increased fitness variation and mean fitness in sperm-dependent parthenogenetic biotypes of the freshwater planarian Schmidtea (formerly Dugesia) polychroa. We correlated the proportion of tetraploids and genotypic diversity, used as estimates for the rate of sex (see below), with mean fitness and fitness variance of six highly genetically differentiated locations of the same lake. In addition, further correlations and path analyses were performed to show whether the putative effect of rare sex on fitness is mediated by fertility (a relative measure of embryo survival) or fecundity (an absolute measure of female investment in cocoons). Materials and methods Model organism Schmidtea polychroa is a nonselfing, simultaneously hermaphroditic flatworm with diploid, obligate sexual and polyploid (mostly triploid or tetraploid) parthenogenetic biotypes. The investigated meta-population exclusively consists of parthenogenetic biotypes that produce their eggs meiotically after premeiotic doubling, which restores the maternal ploidy level. Crossing over does not lead to genetic recombination as identical, duplicated sister chromatids pair (Benazzi Lentati, 1970). Parthenogenetic S. polychroa are sperm-dependent (pseudogamous), i.e. allosperm is required to trigger zygote development, but paternal genetic contribution to offspring is generally suppressed (Benazzi Lentati, 1970; Fig. 1A,B [P]). Because parthenogens produce haploid, fertile sperm (Storhas et al., 2000), they do not rely on sexual spermdonors, which are absent in Central and Western Europe (Beukeboom et al., 1996; Pongratz et al., 2003). This is important as the production of fertile sperm allows for occasional sex in otherwise parthenogenetic populations (D’Souza et al., 2004, 2006). Sexual processes in parthenogenetic individuals result from a combination of at least three mechanisms. First, triploid eggs can fail to expel the triggering sperm leading to genome addition and increase of offspring ploidy (3x fi 4x, Fig. 1A [S1]). Second, tetraploid eggs can undergo meiosis without premeiotic doubling resulting in diploid oocytes which then fuse with haploid sperm resulting in triploid offspring (4x fi 3x, Fig. 1B [S2]). This ploidy cycle from triploidy to tetraploidy and back represents a two-step form of sexual reproduction (Fig. 1C) and leads to stable ploidy frequencies, despite low fitness of tetraploids (D’Souza et al., 2005). Triploid karyotypes represent the original parthenogenetic forms in S. polychroa as they arose from diploid sexuals most probably after fertilization of unreduced eggs (Benazzi & Benazzi Lentati, 1976). Thus, tetraploids are considered a by-product of this two-step sexual cycle and indicate the presence of ª 2007 THE AUTHORS. J. EVOL. BIOL. 21 (2008) 276–286 JOURNAL COMPILATION ª 2007 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY 278 T. G. D’SOUZA AND N. K. MICHIELS (a) (b) BU S1 S3 P S1 S3 3x (c) S2 BB 4x S3 P S2 S3 LS W Fig. 1 Mating system in sperm-dependent parthenogenetic, triploid (a) and tetraploid (b) Schmidtea polychroa. The number of bars indicates the ploidy level. Four different outcomes of sperm–egg interaction are shown: (P) Pseudogamy: Sperm is used to trigger zygote development and resulting offspring are clonally produced. This applies both to triploids and tetraploids. The other mechanisms include syngamy of polyploid eggs (filled bars) and haploid sperm (open bars). (S1) Chromosome addition: Sperm pronucleus fuses with triploid egg and fails to be expelled. Resulting offspring is tetraploid. (S2) Chromosome loss: Tetraploid individual produces diploid, reduced egg, which fuses with haploid sperm nucleus. The resulting zygote then develops into triploid offspring. (S3) Chromosome displacement: Instead of paternal chromosomes one maternal chromosome set is expelled from the zygote which results in offspring with a mixture of paternal and maternal alleles, but unaltered ploidy level. (c) The unique two-step cycle of occasional sex including ploidy alterations from triploidy to tetraploidy and back to triploidy. Alternatively, but more rarely sex may occur without ploidy changes. occasional sex in parthenogenetic populations (D’Souza & Michiels, 2006). A third mechanism involves egg and sperm fusion without ploidy change. In such cases, one maternal chromosome set instead of a paternal one is expelled from the zygote, thus restoring ploidy (D’Souza et al., 2004, 2006; Fig. 1A,B [S3]). SR N E S HE 5 km WW Fig. 2 Sample locations. All sample locations are situated on the East shore of the lake Ammersee, a 46 km2, eutrophic lake in Southern Germany. Location abbreviations: Buch (BU), Breitbrunn (BB), Schloss Ried (SR), Lochschwaben (LS), Herrsching (HE), Wartaweil (WW). dark cycle) and temperature (16 C) conditions. Schmidtea polychroa produces spherical cocoons from which up to 10 offspring hatch. Cocoons were collected daily for 35 days and kept isolated in compartments of ice-cube trays under constant 20 C. Cocoons were checked daily for offspring and were defined as sterile when no offspring hatched after 45 days (ntotal = 1004, nsterile = 141). Average hatching time (±SD) was 13.84 ± 1.79 days (noffspring = 3302). Sampling and collection sites A total of 254 flatworms were collected from six locations (19–59 individuals per location) of the eastern shore of the Ammersee, a eutrophic lake in Bavaria, Germany in July 2004 (Fig. 2). Animals were sampled from stones and transported to the laboratory. Sampling locations had been analysed previously and found to be genetically isolated as shown by significant differentiation and little overlap in genotype composition (see also this study). Sites differed in genotypic diversity and genotypic evenness, which is likely to be caused by varying degrees of occasional sex (D’Souza & Michiels, 2006). Culture conditions Individuals were randomized and kept isolated in 200 mL plastic vials under constant light (12 : 12 light ⁄ Fitness measurements Total number of offspring per individual (produced over 35 days) was used as the main fitness parameter. Fecundity was defined as the total number of (fertile and sterile) cocoons per individual and fertility as the mean number of offspring per cocoon. Thus, individual fitness is the product of fertility and fecundity per individual. These parameters have been used as standard measures of fitness before (Weinzierl et al., 1999; Storhas et al., 2000; D’Souza et al., 2005). Importantly offspring number and cocoon number show highly significant, interclonal variation indicating a genetic basis of fitness (D’Souza et al., 2005). For all further analyses fitness parameters were determined for triploids only to exclude any confounding effects resulting from ploidy differences among locations. ª 2007 THE AUTHORS. J. EVOL. BIOL. 21 (2008) 276–286 JOURNAL COMPILATION ª 2007 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY Sex rate estimates and fitness in parthenogens Body size measurements As body size is often correlated with fitness, we measured planar body area. Digital images were taken from slowly moving animals. Body area was then determined using image-analysis software (ImageJ v1.34s, National Institutes of Health, Bethesda, Maryland, USA). Karyology To determine ploidy level, a modified protocol of Redi et al. (1982) was used (Beukeboom et al., 1996). Briefly, partially regenerated individuals were treated with colchicine for 10 h to arrest cell cycle in metaphase. Chromosomes were then counted using phase contrast microscopy. As expected, only triploid (3x = 12) and tetraploid (4x = 16) parthenogens were found. Genotyping and genotypic diversity Two highly polymorphic trinucleotide microsatellite loci (SpATT12 and SpATT20) were used to determine twolocus genotypes. In the present study, the number of alleles for SpATT12 and SpATT20 was 15 and 19 respectively. For details on DNA isolation, PCR conditions and fragment size analysis see Pongratz et al. (2001) and D’Souza et al. (2004). In order to avoid that genetic differences may be due to de novo mutations, genotypes were pooled, when differing by fewer than three trinucleotide repeat units across all loci, leading to conservative diversity estimates (Bruvo et al., 2004; D’Souza & Michiels, 2006). Genotypic differentiation among locations was tested by applying log-likelihood G-statistics (Goudet et al., 1996) as described in D’Souza & Michiels (2006). For this, multilocus genotypes were coded as different alleles of a single locus. We used h, the Weir and Cockerham estimator, for overall FST (Weir & Cockerham, 1984). For further analysis of population differentiation we counted the number of private genotypes (genotypes that are restricted to one location) and unique genotypes (genotype that is represented by one individual) (Menken et al., 1995). Simpson’s index D for genotypic (or clonal) diversity was used to characterize the genetic diversity of the locations. D expresses the probability, that two randomly sampled individuals are genotypically different. To correct for sampling size differences the following formula was used: ! m X n 2 Dg ¼ pi 1 n1 i¼1 where n = sampling size, pi = relative frequency of ith genotype, m = number of multilocus genotypes. D ranges from 0 (i.e. all individuals show the same genotype) to 1 (i.e. all individuals have different genotypes). In contrast 279 to genotypic diversity, allelic diversity (also known as genetic diversity or heterozygosity) was also calculated but revealed only little variation across locations (mean of both loci ranged from 0.733 to 0.880). Estimates of sex rate Occasional sex in S. polychroa includes the creation of tetraploids from triploids and vice versa. These ploidy transitions lead to a cyclic gene flow from the more ancestral triploids to tetraploids and back (D’Souza et al., 2004; Fig. 1C). Without these ploidy transitions triploids will displace tetraploids within few generations as a consequence of increased fitness of triploids compared with tetraploids. Especially the recurrent production of tetraploids leads to stable proportions of both ploidy types (D’Souza et al., 2005). Thus tetraploids originate and are maintained as a result of occasional sex. For that reason tetraploids emerge as a by-product of occasional sex in S. polychroa. That tetraploids indicate the rate of sex is especially reflected by the strong, positive correlation between tetraploid proportion and triploid genotypic diversity (D’Souza & Michiels, 2006; this study). This suggests that the production of tetraploids leads to an increase of genotypic diversity among triploids. Hence, especially the proportion of tetraploids can be used as a reliable estimate of the rate of (rare) sex in S. polychroa. The association between genotypic diversity (in contrast to genetic or allelic diversity) and fitness is less often studied, but may provide valuable insights into the ecological and evolutionary importance of genotypic diversity (Reusch & Hughes, 2006). Genotypic diversity is typically increased by sexual reproduction (Balloux et al., 2003; Bengtsson, 2003), but is also affected by other factors. For example, besides rare sex, mutations and multiple origins from sexual ancestors are known to influence genotypic diversity (Parker, 1979). Moreover, migration and population size may also alter genotypic diversity (Leimu et al., 2006). For S. polychroa we can exclude mutations, multiple origins from sexuals and migration as central sources of diversity: in S. polychroa microsatellite mutation rate is low (D’Souza et al., 2006), sexual conspecifics presumably have never been present since the emergence of the investigated meta-population (Beukeboom et al., 1996; Pongratz et al., 2003), and migration is virtually absent (Pongratz et al., 2002; D’Souza & Michiels, 2006). However, as population size is difficult to determine in S. polychroa we cannot exclude that locations differ in population size. Therefore, we concentrate on the proportion of tetraploids as an estimate of rare sex and use triploid genotypic diversity to support the findings. Statistical analysis One major assumption of Weismann’s principle is that fitness must be strongly heritable (Burt, 2000). For this ª 2007 THE AUTHORS. J. EVOL. BIOL. 21 (2008) 276–286 JOURNAL COMPILATION ª 2007 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY 280 T. G. D’SOUZA AND N. K. MICHIELS we estimated genetic variation for offspring number using a mixed-model A N O V A with genotype as a random factor and location as a fixed factor, while genotype is nested within location. Only genotypes with at least four representatives were considered for the analysis. Splitting phenotypic variance into genetic and environmental variance was performed according to Becker (1992) and Vorburger (2005). Variance among genotypes served as an estimate for genetic variance and the residual variance for environmental variance. The ratio of genetic to phenotypic variance was used to estimate broad-sense heritability, H2. An approximate standard error for H2 was calculated using a formula taken from Becker (1992), which corrects for unequal numbers of individuals per genotype (Vorburger, 2005). The F-test for the effect of the factor genotype resulting from the mixed-model A N O V A was used to indicate whether heritability of offspring number differed significantly from zero. The main goal of the study was to determine whether a relationship exists between sex rate estimates (proportion of tetraploids and genotypic diversity of triploids) and fitness of triploids (expressed as offspring produced in 5 weeks). This effect was broken down into effects of sex rate on fertility, fecundity and body size. For this we used nonparametric Spearman rank correlations. For each correlation the mean and variance of fitness, fertility, fecundity and body size were taken per location to correct for sample size differences and to avoid pseudoreplication. Path analysis was applied to calculate the extent of the effect of sex rate on triploid fitness via fertility or fecundity. For this, path coefficients were estimated as standardized partial regression coefficients using maximum likelihood (Sokal & Rohlf, 2003). As there is no nonparametric alternative to regression analysis, coefficients were calculated although the formal assumptions were not fulfilled. Therefore corresponding P-values have to be taken with care. We used sex rate as an exogenous, independent variable and fitness as a dependent, endogenous variable. Fecundity, fertility and size were endogenous, intermediary variables. See path diagram for relationships (Fig. 3). Note, as fitness (offspring number) is the product of fertility (offspring number per cocoon) and fecundity (cocoon number), there is no direct path leading from genotypic diversity to fitness in the model. The unexplained variance (U) of the dependent variable fitness was estimated as one minus the squared total effect of genotypic diversity on fitness (Sokal & Rohlf, 2003). Path analysis was separately performed for mean and variance of the endogenous variables. The reliability of both models was tested with a goodness-of-fit v2-test. As the path model with variances differed significantly from the observed data (v23 = 17.997, P < 0.001), we only refer to the results from the path analysis of means (v23 = 3.079, P = 0.38). Fecundity 0.751* 0.384*** 0.296 –0.179 Proportion of tetraploids Size Fitness 0.342 0.572 0.678*** 0.784*** Fertility U = 0.328 Fig. 3 Path diagram. The effect of genotypic diversity on mean fitness of triploids mediated by mean fecundity, mean fertility and mean body size of six locations. Arrows indicate the extent and direction of a causal relationship between two variables. Path coefficients are estimated as partial regression coefficients using maximum likelihood. Significant relationships are indicated with ***P < 0.001 and *P < 0.05. See Methods for statistical analysis. U is the proportion of variance of mean fitness which is not explained by the model. Model and observed data do not significantly differ (v23 = 3.079, P = 0.38). Approximately 67% of the variance in mean fitness can be explained by genotypic diversity. Genotypic diversity was calculated with G E N O D I V E (Meirmans & Van Tienderen, 2004). F S T A T (v 2.9.3) was used to test population differentiation (Goudet, 1995, 2001). Path analysis was performed using A M O S 5 and all other analyses with J M P I N (v 5.1). Scatter plots were created with Sigma Plot (v 9.0) (Systat Software Inc., San Jose, California, USA). Means are given (± standard deviation) unless stated otherwise. Results Genotypic diversity We identified a total of 57 triploid and 19 tetraploid genotypes from 254 individuals. Overall genotypic diversity of the six locations ranged from 0.476 to 0.924 (Table 1). When excluding tetraploids, the diversity indices of triploids were slightly smaller. As predicted, genotypic diversity of triploids increased significantly with the proportion of tetraploids across sites (Spearmann, rs,6 = 0.943, P = 0.005). Population differentiation Highly significant genotypic differentiation indicates strong sub-structuring of the meta-population (G-test, P = 0.0001, h = 0.206). Approximately 84% of the 76 observed genotypes were private to a single location. Hence, only 16% of the genotypes occurred in more than one location. In addition, 70% of all observed genotypes are represented by a single individual (unique genotype). The high proportion of private and unique genotypes implies little overlap in clonal composition and highlights the extreme genotypic differentiation among locations, despite the fact that they were all along the shoreline of the same lake. ª 2007 THE AUTHORS. J. EVOL. BIOL. 21 (2008) 276–286 JOURNAL COMPILATION ª 2007 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY 281 Sex rate estimates and fitness in parthenogens Location Sample size Proportion tetraploids Number of genotypes Private genotypes Unique genotypes Total genotypic diversity Triploid genotypic diversity BU BB SR LS HE WW Total 46 19 39 51 40 59 254 0.130 0.158 0.154 0.098 0.075 0.102 0.110 16 12 20 15 9 20 76 12 8 16 9 5 14 64 8 8 14 8 3 12 53 0.894 0.924 0.872 0.671 0.474 0.749 0.907 0.878 0.892 0.820 0.600 0.387 0.700 0.883 Table 2 Results from mixed-model A N O V A on offspring number as response variable and factor genotype (random) nested within factor location (fixed). The significant effect of genotype shows strong heritability of offspring number (H2 = MSgenotype ⁄ (MSgenotype + MSresidual) = 0.755 ± 0.082. Source of variation df SS MS F-ratio P Genotype Location (genotype) Residual Total 10 4 147 161 5761.650 3505.570 27468.044 35989.809 576.165 876.393 186.857 3.083 4.690 0.001 0.001 HE Mean fitness of triploids ± sd Table 1 Sample sizes and genetic characteristics of each location. Position of sampling locations is depicted in Fig. 1. LS WW BU SR BB 40 30 20 10 0 0.08 0.10 0.12 0.14 0.16 Proportion of tetraploids Heritability of fitness Mixed-model A N O V A revealed substantial variation among locations and among genotypes for offspring number (Table 2). The factors location as well as genotype contribute significantly to the variation in fitness (both P = 0.001). The significant effect of genotype also indicates that broad-sense heritability of trait offspring number differs significantly from zero. Fitness exhibits a high heritability value of 0.755 ± 0.082 (H2 ± SE). Proportion of tetraploids and fitness attributes of triploids The variance and mean of offspring number of triploids increased with the proportion of tetraploids (Variance: Spearman, rs,6 = 0.886, P = 0.019; Mean: Spearman, rs,6 = 0.886, P = 0.019; Fig. 4). To verify that fertility or fecundity accounted for the relationship between proportion of tetraploids and fitness of triploids, we also correlated fertility and fecundity with proportion of tetraploids. Neither mean nor variance of fertility and fecundity was significantly correlated with proportion of tetraploids (Table 3a). Moreover, there was no significant relationship between proportion of tetraploids and (mean and variance of) body size (Table 3a). Considering triploids and tetraploids for the calculation of population’s fitness attributes does not change significances (results not shown). Fig. 4 Correlation between proportion of tetraploids and fitness of triploids. Proportion of tetraploids, an estimate for occasional sex, is significantly, positively correlated with mean fitness (Spearman, rs,6 = 0.886, P = 0.019) and fitness variance (Spearman, rs,6 = 0.886, P = 0.019). Names of the locations are given in the upper part of the diagram. Triploid genotypic diversity and fitness attributes of triploids Correlations between triploid genotypic diversity and mean ⁄ variance of fitness, fertility, fecundity and body size of triploids revealed similar results as with proportion of tetraploids and fitness attributes: only the mean and variance of fitness correlates significantly with triploid genotypic diversity (Table 3b). Path analysis We used path analysis to partition the effect of proportion of tetraploids on mean fitness of triploids through effects mediated by the mean of fertility, fecundity and body size. The path model (Fig. 3), explained approximately 67% of the variance in mean fitness across the six populations (R2 = 0.672, U = 0.328). Observed data and path model did not differ significantly (v23 = 3.079, P = 0.38) suggesting a good fit of the model. The total effect (direct + indirect via body size) of tetraploid ª 2007 THE AUTHORS. J. EVOL. BIOL. 21 (2008) 276–286 JOURNAL COMPILATION ª 2007 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY 282 T. G. D’SOUZA AND N. K. MICHIELS Table 3 Results of the Spearman rank correlations between sex rates estimates, (a) proportion of tetraploids, (b) triploid genotypic diversity and fitness parameters of triploids. Only correlations with mean fitness and fitness variance revealed significant results (n = 6). Correlation coefficient (rs) (a) Fitness Variance Mean Fertility Variance Mean Fecundity Variance Mean Body size Variance Mean (b) Fitness Variance Mean Fertility Variance Mean Fecundity Variance Mean Body size Variance Mean P-value 0.886 0.886 0.019 0.019 0.771 0.771 0.072 0.072 0.486 0.600 0.329 0.208 )0.086 0.257 0.872 0.623 0.943 0.886 0.005 0.019 0.771 0.771 0.072 0.072 0.543 0.486 0.266 0.329 )0.143 0.0857 0.787 0.872 proportion on fecundity is R = 0.751 and on fertility R = 0.784. Combined with the effect of fecundity (R = 0.384) and fertility (R = 0.678) on fitness, this indicates that the impact of tetraploid proportion on fitness via fertility (R = 0.784Æ0.678 = 0.531) is stronger than the effect via fecundity (R = 0.751Æ0.384 = 0.288). In other words, 28% (R2 = 0.282) of the variation in mean fitness can be explained by tetraploid proportion via fertility and 8.3% (R2 = 0.083) by tetraploid proportion via fecundity. Path analyses with triploid genotypic diversity as the exogenous, independent variable revealed similar results and trends (data not shown). Discussion Indications for increased efficacy of natural selection We showed that increased presence of tetraploids, a byproduct of occasional sex, is associated with increased fitness variance and mean fitness across six genetically isolated subpopulations of parthenogenetic S. polychroa. This pattern is in accordance with Weismann’s principle as it shows that (occasional) sex increases fitness variance and mean fitness, making natural selection more effective. To our knowledge the present study provides the first empirical support to validate Weismann’s principle in natural subpopulations with variable degrees of sex. However, describing correlative effects in ‘natural experiments’ like these, does not exclude other differences affecting fitness traits between locations. Local subpopulations may differ in age, food quality, resource availability and other ecological or genetic factors. One alternative explanation for the significant relationship between proportion of tetraploids and fitness could be that tetraploids may show a higher fitness mean and variance compared with triploids. However, we can exclude this possibility for the following reasons: First, contrary to other systems (for a review see Otto & Whitton, 2000), higher ploidy level is not associated with increased fitness in S. polychroa, as tetraploids exhibit reduced fitness compared with triploids (D’Souza et al., 2005). Secondly, we considered only triploids in the fitness calculation making the proportion of tetraploids as an estimate of sex doubtless independent from the populations’ fitness parameters. Hence, fitness effects caused by different ploidy levels do not confound the results. Locations may vary in mutation rates caused, e.g. by different levels of stress, mutagenic substances, or temperature. Increased mutation rates may lead to increased defects in gametogenesis causing the formation of more tetraploid individuals, and increased fitness variance. Failures during oogenesis in triploid S. polychroa should also lead to the production of aneuploid or hexaploid offspring after meiotic ⁄ mitotic dysfunctions or chromosome doubling respectively. However, as such ploidy types have been never observed in the field (Beukeboom et al., 1996; D’Souza & Michiels, 2006) nor in laboratory experiments (Benazzi Lentati, 1970, 1979), they are either not produced or not viable. In the case of an early mortality mean fitness of those populations with increased mutations rates (and therefore high tetraploid proportion) should be reduced, which contradicts our findings. Therefore, the most parsimonious explanation of the observed fitness pattern in S. polychroa is indeed the proportion of tetraploids and other sex-related features. Although we did not experimentally control the amount of sex, the strength of this study is that we used a system of naturally varying degrees of sex within a single meta-population. Contrary to other systems, we did not artificially induce sex, which may cause other confounding effects. For instance, in Chlamydomonas reinhardtii and yeast, sex induction by starvation leads to higher mutation rates (Marini et al., 1999; Goho & Bell, 2000). For C. reinhardtii it has been shown that higher mutation rates already lead to an increased variance in fitness, which is also expected to be the result of sexual reproduction (Goho & Bell, 2000). ª 2007 THE AUTHORS. J. EVOL. BIOL. 21 (2008) 276–286 JOURNAL COMPILATION ª 2007 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY Sex rate estimates and fitness in parthenogens Weismann’s principle and occasional sex Weismann’s principle and other theories of sex were originally developed to compare obligate sexuals and obligate asexuals. A theoretical model by Burt (2000) extends this limitation by showing that Weismann’s principle is also valid for species that only have sex occasionally. Low levels of sex and recombination are sufficient to increase mean fitness by enhancing the efficacy of natural selection in predominantly clonal populations (Burt, 2000). Moreover, even single episodes of recombination after long phases of asexuality may release most of the hidden genetic variation. Consequently, the rate of phenotypic evolution after sex may increase to an extent comparable with continuously sexuals (Lynch & Gabriel, 1983). This allows parthenogens to rapidly adjust to new selection pressures and lead to an increased probability of long-term survival (Lynch & Gabriel, 1983). Mutations or environmental factors explaining sex? Many models explaining the maintenance of sex rely on the diversifying character of recombination and segregation. They can be classified as either mutational models (Deterministic mutation hypothesis, Muller’s ratchet) or environmental models (Fisher–Muller hypothesis, Red Queen) (Hurst & Peck, 1996; West et al., 1999). Comparisons between sexual and parthenogenetic individuals revealed that major predictions of both models are found in S. polychroa. Parthenogens showed both indications for increased mutational load (Storhas et al., 2000) as well as parasite load (Michiels et al., 2001). Moreover, parthenogenetic lineages show significant correlation between mutation and parasite load giving evidence to the so-called pluralistic approach (West et al., 1999; WestEberhard, 2005), which combines both mutational and environmental models (Bruvo et al., 2007). Reduction in fertility most likely results from developmental problems and increased embryo mortality which in turn was used as an indicator for the accumulation of deleterious mutations in previous studies (Storhas et al., 2000; Bruvo et al., 2007). However, we could not identify any significant correlation between rate of sex and fertility. This indicates that either the sex rate is too low to yield a clear effect on mutational load or that deleterious mutations are not the only factor maintaining occasional sex. The interplay of occasional sex and fitness The effect of proportion of tetraploids on fitness traits is only significant for total number of offspring, which was used as the primary fitness estimate. Compared to published accounts, broad-sense heritability of total offspring number is extremely high in S. polychroa (e.g. Vorburger, 2005). This stresses the genetic basis of fitness 283 which is supported by our previous finding of significant interclonal differences in several fitness traits (D’Souza et al., 2005). The mixed-model A N O V A revealed not only significant fitness differences between clones, but also between locations. However, total number of offspring depends on the number of cocoons an individual produces (fecundity) and the number of offspring per cocoon (fertility). A previous study revealed that triploid and tetraploid clonal lineages differ in fecundity, but not in fertility, resulting in a higher total offspring production of triploids (D’Souza et al., 2005). Hence, one would expect that locations with large proportions of tetraploids show a reduced fecundity. However, there was no evidence for such a relationship in the present study (Table 3a), indicating that the benefits of occasional sex may compensate for the fecundity disadvantage of tetraploids. Fertility is largely determined by the reproductive mode of the sperm donor in S. polychroa: sperm from obligate sexuals increases fertility of parthenogenetic sperm receivers (Storhas et al., 2000). Additionally, sperm from occasionally sexual parthenogens increases fertility of more clonal receivers (D’Souza et al., 2007). Although no significant relationship between both sex estimates and fertility was found, the importance of fertility on the sex–fitness relationship is indicated by results of the path analysis. The relative contribution of fertility is high, while fecundity merely plays a minor role (Fig. 3). Ecological implications of genotypic diversity The relationship between genetic diversity (in the sense of allele diversity, population heterozygosity) and fitness is commonly investigated. Genetic diversity is used as an indicator of inbreeding and reduced fitness is expected when genetic diversity is low due to inbreeding depression, genetic drift, fixation of deleterious mutations and other processes (David, 1998; Booy et al., 2000; Hansson & Westerberg, 2002; Reed & Frankham, 2003; Field et al., 2007). However, the ecological and evolutionary significance of genotypic diversity is less often studied. Using manipulative field experiments, it was demonstrated that genotypic diversity of the seagrass Zostera marina enhances ecosystem stability and therefore buffers against biotic or abiotic disturbances (Hughes & Stachowicz, 2004; Reusch et al., 2005; Reusch & Hughes, 2006). Transects with high genotypic diversity recovered faster from grazing (Hughes & Stachowicz, 2004) and after a high temperature period compared with monoclonal transects (Reusch et al., 2005). Moreover, genotypic diversity increased resistance against parasites in leafcutting ants (Hughes & Boomsma, 2004). Similarly, a theoretical model showed that the presence of parasites selects for clonal diversity and that genetically diverse clones can outcompete sexual populations in the absence of deleterious mutations (Lively & Howard, 1994). ª 2007 THE AUTHORS. J. EVOL. BIOL. 21 (2008) 276–286 JOURNAL COMPILATION ª 2007 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY 284 T. G. D’SOUZA AND N. K. MICHIELS Genotypically diverse populations also bear a competitive advantage towards monoclonal populations in Daphnia (Tagg et al., 2005a,b). Genotypic diversity increases competitiveness in invading and resisting invasions of monoclonal populations (Tagg et al., 2005b). In summary, high genotypic diversity serves as a buffer against interfering biotic and abiotic disturbances and may also aid in the persistence of parthenogenetic populations of S. polychroa. Through increased competitive ability, genotypically diverse subpopulations may resist displacement by the immigrant planarian Girardia (Dugesia) tigrina, which has expanded its distribution across Britain during the last 50 years (Young & Reynoldson, 1999) and has also recently been found in lake Ammersee (T.G. D’Souza, personal observation). Conclusions In summary, we found indications that occasional sex in parthenogenetic S. polychroa leads to an increase in fitness variance at the local population level, which in turn increases mean fitness, which can be interpreted as a signature of a more effective natural selection. Consequently, occasional sex may prevent the extinction of parthenogenetic subpopulations and explain the success of this particular type of parthenogenesis in this species. Acknowledgments We thank Rebecca Schulte and Vera Bellenhaus for help in collecting flatworms and Gaby Niester and Barbara Hasert for technical assistance in the laboratory. We are grateful to Claudia Böhnisch, Beatriz Sanchez Navarro, Rebecca Schulte Nadine Timmermeyer and Hinrich Schulenburg for fruitful discussions. Thanks to Christoph Vorburger for providing advice in heritability analysis. Paul Sunnucks, Stu Field, Beatriz Sanchez Navarro and two anonymous referees gave valuable comments to improve earlier versions of the manuscript. This study was supported by a grant from the German Science Foundation (DFG) no. MI 482 ⁄ 11-1. References Agrawal, A.F. 2006. Evolution of sex: why do organisms shuffle their genotypes? Curr. Biol. 16: R696–R704. Bachtrog, D. 2003. Adaptation shapes patterns of genome evolution on sexual and asexual chromosomes in Drosophila. Nat. Genet. 34: 215–219. Bachtrog, D. & Charlesworth, B. 2002. Reduced adaptation of a non-recombining neo-Y chromosome. Nature 416: 323–326. Balloux, F., Lehmann, L. & de Meeus, T. 2003. The population genetics of clonal and partially clonal diploids. Genetics 164: 1635–1644. Barton, N.H. & Charlesworth, B. 1998. Why sex and recombination? Science 281: 1986–1990. Becker, W.A. 1992. Manual of Quantitative Genetics. Academic Enterprises, Pullman, WA. Bell, G. 1982. The Masterpiece of Nature. The Evolution and Genetics of Sexuality. University of California press, Berkely and Los Angeles, CA. Benazzi, M. & Benazzi Lentati, G. 1976. Platyhelminthes. Gebrueder Borntraeger, Berlin, Stuttgart. Benazzi Lentati, G. 1970. Gametogenesis and egg fertilization in planarians. Int. Rev. Cytol. 27: 101–179. Benazzi Lentati, G. 1979. Polypoidy and types of development in the Triclad Dugesia lugubris s.l.: interaction between the ploidy level and the origin of amphimixis or pseudogamy. Caryologia 32: 247–263. Bengtsson, B.O. 2003. Genetic variation in organisms with sexual and asexual reproduction. J. Evol. Biol. 16: 189–199. Betancourt, A.J. & Presgraves, D.C. 2002. Linkage limits the power of natural selection in Drosophila. Proc. Natl. Acad. Sci. USA 99: 13616–13620. Beukeboom, L.W., Weinzierl, R.P., Reed, K.M. & Michiels, N.K. 1996. Distribution and origin of chromosomal races in the freshwater planarian Dugesia polychroa (Turbellaria: Tricladida). Hereditas 124: 7–15. Booy, G., Smulders, M.J.M., Vosman, B., Hendriks, R.J.J. & Van Groenendael, J.M. 2000. Genetic diversity and the survival of populations. Plant. Biol. 2: 379–395. Bruvo, R., Michiels, N.K., D’Souza, T.G. & Schulenburg, H. 2004. A simple method for the calculation of microsatellite genotype distances irrespective of ploidy level. Mol. Ecol. 13: 2101–2106. Bruvo, R., Schulenburg, H., Storhas, M. & Michiels, N.K. 2007. Synergism between mutational meltdown and Red Queen in parthenogenetic biotypes of the freshwater planarian Schmidtea polychroa. Oikos 116: 313–323. Burt, A. 2000. Perspective: Sex, recombination, and the efficacy of selection - Was Weismann right? Evolution 54: 337–351. Colegrave, N. 2002. Sex releases the speed limit on evolution. Nature 420: 664. Colegrave, N., Kaltz, O. & Bell, G. 2002. The ecology and genetics of fitness in Chlamydomonas. VIII. The dynamics of adaptation to novel environments after a single episode of sex. Evolution 56: 14–21. D’Souza, T.G. & Michiels, N.K. 2006. Genetic signatures of occasional sex in parthenogenetic subpopulations of the freshwater planarian Schmidtea polychroa. Freshw. Biol. 51: 1890–1900. D’Souza, T.G., Storhas, M., Schulenburg, H., Beukeboom, L.W. & Michiels, N.K. 2004. Occasional sex in an ‘asexual’ polyploid hermaphrodite. Proc. R. Soc. Lond. B Biol. Sci. 271: 1001–1007. D’Souza, T.G., Storhas, M. & Michiels, N.K. 2005. The effect of ploidy level on fitness in parthenogenetic flatworms. Biol. J. Linn. Soc. 85: 191–198. D’Souza, T.G., Schulte, R.D., Schulenburg, H. & Michiels, N.K. 2006. Paternal inheritance in parthenogenetic forms of the planarian Schmidtea polychroa. Heredity 97: 97–101. D’Souza, T.G., Bellenhaus, V., Wesselmann, R. & Michiels, N.K. 2007. Sperm length and quality in sperm-dependent parthenogens. Biol. J. Linn. Soc. 92, in press. David, P. 1998. Heterozygosity-fitness correlations: new perspectives on old problems. Heredity 80: 531–537. De Visser, J.A.G.M. & Elena, S.F. 2007. The role of sex: empirical insights into the roles of epistasis and drift. Nat. Rev. Genet. 8: 139–149. De Visser, J.A.G.M. & Rozen, D.E. 2005. Limits to adaptation in asexual populations. J. Evol. Biol. 18: 779–788. ª 2007 THE AUTHORS. J. EVOL. BIOL. 21 (2008) 276–286 JOURNAL COMPILATION ª 2007 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY Sex rate estimates and fitness in parthenogens Doroszuk, A., Wojewodzic, M.W. & Kammenga, J.E. 2006. Rapid adaptive divergence of life-history traits in response to abiotic stress within a natural population of a parthenogenetic nematode. Proc. R. Soc. Lond. B Biol. Sci. 273: 2611–2618. Field, S.G., Lange, M., Schulenburg, H., Velavan, T.P. & Michiels, N.K. 2007. Genetic diversity and parasite defense in a fragmented urban metapopulation of earthworms. Anim. Conserv. 10: 162–175. Goho, S. & Bell, G. 2000. Mild environmental stress elicits mutations affecting fitness in Chlamydomonas. Proc. R. Soc. Lond. B Biol. Sci. 267: 123–129. Goudet, J. 1995. FSTAT (vers.1.2): a computer program to calculate F-statistics. J. Hered. 86: 485–486. Goudet, J. 2001. FSTAT, a program to estimate and test gene diversities and fixation indices (version 2.9.3). Available from http://www.unil.ch/izea/softwares/fstat.html. Goudet, J., Raymond, M., Rousset, F. & De Meeüs, T. 1996. Testing differentiation in diploid populations. Genetics 144: 1933–1940. Grimberg, B. & Zeyl, C. 2005. The effects of sex and mutation rate on adaptation in test tubes and to mouse hosts by Saccharomyces cerevisiae. Evolution 59: 431–438. Hansson, B. & Westerberg, L. 2002. On the correlation between heterozygosity and fitness in natural populations. Mol. Ecol. 11: 2467–2474. Holmström, K. & Jensen, H.J. 2004. Who runs fastest in an adaptive landscape: sexual versus asexual reproduction. Physica A Statist. Mech. Appl. 337: 185–195. Hughes, W.O.H. & Boomsma, J.J. 2004. Genetic diversity and disease resistance in leaf-cutting ant societies. Evolution 58: 1251–1260. Hughes, A.R. & Stachowicz, J.J. 2004. Genetic diversity enhances the resistance of a seagrass ecosystem to disturbance. Proc. Natl. Acad. Sci. USA 101: 8998–9002. Hurst, L.D. & Peck, J.R. 1996. Recent advances in understanding of the evolution and maintenance of sex. Trends Ecol. Evol. 11: 46–52. Kaltz, O. & Bell, G. 2002. The ecology and genetics of fitness in Chlamydomonas. XII. Repeated sexual episodes increase rates of adaptation to novel environments. Evolution 56: 1743–1753. Kondrashov, A.S. 1984. Deleterious mutations as an evolutionary factor. I. The advantage of recombination. Genet. Res. 44: 199–217. Kondrashov, A.S. 1988. Deleterious mutations and the evolution of sexual reproduction. Nature 336: 435–440. Kondrashov, A.S. 1993. Classification of hypothesis on the advantage of amphimixis. J. Hered. 84: 372–387. Leimu, R., Mutikainen, P., Koricheva, J. & Fischer, M. 2006. How general are positive relationships between plant population size, fitness and genetic variation? J. Ecol. 94: 942–952. Lively, C.M. & Howard, R.S. 1994. Selection by parasites for clonal diversity and mixed mating. Philos. Trans. R. Soc. Lond. B Biol. Sci. 346: 271–281. Lynch, M. & Gabriel, W. 1983. Phenotypic evolution and parthenogenesis. Am. Nat. 122: 745–764. Marini, A., Matmati, N. & Morpurgo, G. 1999. Starvation in yeast increases non-adaptive mutation. Curr. Genet. 35: 77–81. Maynard Smith, J. 1978. The Evolution of Sex. Cambridge University Press, Cambridge, London, New York, Melbourne. Meirmans, P.G. & Van Tienderen, P.H. 2004. GENOTYPE and GENODIVE: two programs for the analysis of genetic diversity of asexual organisms. Mol. Ecol. Notes 4: 792–794. 285 Menken, S.B.J., Smit, E. & Den Nijs, H.C.M. 1995. Genetical population structure in plants: gene flow between diploid sexual and triploid asexual dandelions (Taraxacum section Ruderalia). Evolution 49: 1108. Michiels, N.K., Beukeboom, L.W., Pongratz, N. & Zeitlinger, J. 2001. Parthenogenetic flatworms have more symbionts than their coexisting, sexual conspecific, but does this support the Red Queen? J. Evol. Biol. 14: 110–119. Otto, S.P. & Gerstein, A.C. 2006. Why have sex? The populations genetics of sex and recombination Biochem. Soc. Trans. 34: 519– 522. Otto, S.P. & Whitton, J. 2000. Polyploid incidence and evolution. Annu. Rev. Genet. 34: 401–437. Parker, E.D. 1979. Ecological Implications of clonal diversity in parthenogenetic morphospecies. Am. Zool. 19: 753–762. Peck, J.R. 1994. A ruby in the rubbish - Beneficial mutations, deleterious mutations and the evolution of sex. Genetics 137: 597–604. Peters, A.D. & Otto, S.P. 2003. Liberating genetic variance through sex. Bioessays 25: 533–537. Pongratz, N., Gerace, L., Alganza, A.M., Beukeboom, L.W. & Michiels, N.K. 2001. Microsatellite development and inheritance in the planarian flatworm Schmidtea polychroa. Belg. J. Zool. 131: 71–75. Pongratz, N., Gerace, L. & Michiels, N.K. 2002. Genetic differentiation within and between populations of a hermaphroditic freshwater planarian. Heredity 89: 64–69. Pongratz, N., Storhas, M., Carranza, S. & Michiels, N.K. 2003. Phylogeography of competing sexual and parthenogenetic forms of a freshwater flatworm: patterns and explanations. BMC Evol. Biol. 3. Presgraves, D.C. 2005. Recombination enhances protein adaptation in Drosophila melanogaster. Curr. Biol. 15: 1651–1656. Redi, C.A., Garagna, S. & Pellicciari, C. 1982. Chromosome preparation from planarian blastemas: a new procedure suitable for cytogenetic and cytochemical studies. Stain Technol. 57: 190–192. Reed, D.H. & Frankham, R. 2003. Correlation between fitness and genetic diversity. Conserv. Biol. 17: 230–237. Reusch, T.B.H. & Hughes, A.R. 2006. The emerging role of genetic diversity for ecosystem functioning: Estuarine macrophytes as models. Estuar. Coast 29: 170–175. Reusch, T.B.H., Hämmerli, A., Ehlers, A. & Worm, B. 2005. Ecosystem recovery after climatic extremes enhanced by genotypic diversity. Proc. Natl. Acad. Sci. USA 102: 2826– 2831. Rice, W.R. 2002. Experimental tests of the adaptive significance of sexual recombination. Nat. Rev. Genet. 3: 241–251. Rice, W.R. & Chippindale, A.K. 2001. Sexual recombination and the power of natural selection. Science 294: 555–559. Sokal, R.R. & Rohlf, F.J. 2003. Biometry, 3d edn. W.H. Freeman and Company, New York. Storhas, M., Weinzierl, R.P. & Michiels, N.K. 2000. Paternal sex in parthenogenetic planarians: a tool to investigate the accumulation of deleterious mutations. J. Evol. Biol. 13: 1–8. Tagg, N., Doncaster, C.P. & Innes, D.J. 2005a. Resource competition between genetically varied and genetically uniform populations of Daphnia pulex (Leydig): Does sexual reproduction confer a short-term ecological advantage? Biol. J. Linn. Soc. 85: 111–123. Tagg, N., Doncaster, C.P. & Innes, D.J. 2005b. Outcomes of reciprocal invasions between genetically diverse and ª 2007 THE AUTHORS. J. EVOL. BIOL. 21 (2008) 276–286 JOURNAL COMPILATION ª 2007 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY 286 T. G. D’SOUZA AND N. K. MICHIELS genetically uniform populations of Daphnia obtusa (Kurz). Oecologia 143: 527–536. Vorburger, C. 2005. Positive genetic correlations among major life-history traits related to ecological success in the aphid Myzus persicae. Evolution 59: 1006–1015. Weinzierl, R.P., Schmidt, P. & Michiels, N.K. 1999. High fecundity and low fertility in parthenogenetic planarians. Invertebr. Biol. 118: 87–94. Weir, B.S. & Cockerham, C.C. 1984. Estimating F-statistics for the analysis of population structure. Evolution 38: 1358–1370. Weismann, A. 1886. The significance of sexual reproduction in the theory of natural selection. In: Essays Upon Heredity and Kindred Biological Problems (E. B. Poulton, S. Schönland & A. E. Shipey, eds), pp. 251–332. Clarendon Press, Oxford. West, S.A., Lively, C.M. & Read, A.F. 1999. A pluralist approach to sex and recombination. J. Evol. Biol. 12: 1003–1012. West-Eberhard, M.J. 2005. The maintenance of sex as a developmental trap due to sexual selection. Q. Rev. Biol. 80: 47–53. Williams, G. 1975. Sex and Evolution. Princeton University Press, Princeton, NJ. Young, J.O. & Reynoldson, T.B. 1999. Continuing dispersal of freshwater triclads (Platyhelminthes; Turbellaria) in Britain with particular reference to lakes. Freshw. Biol. 42: 247–262. Zeyl, C. & Bell, G. 1997. The advantage of sex in evolving yeast populations. Nature 388: 465–468. Received 6 August 2007; revised 13 September 2007; accepted 14 September 2007 ª 2007 THE AUTHORS. J. EVOL. BIOL. 21 (2008) 276–286 JOURNAL COMPILATION ª 2007 EUROPEAN SOCIETY FOR EVOLUTIONARY BIOLOGY