Microsatellites and the Genetics of Highly Selfing Populations in the Freshwater Snail Bulinus Truncatus

Hermaphrodite tropical freshwater snails provide a good opportunity to study the effects of mating system and genetic drift on population genetic structure because they are self-fertile and they occupy transient patchily distributed habitats (ponds). Up to now the lack of detectable allozyme polymorphism prevented any intrapopulation studies. In this paper, we examine the consequences of selfing and bottlenecks on genetic polymorphism using microsatellite markers in 14 natural populations (under a hierarchical sampling design) of the hermaphrodite freshwater snail Bulinus truncatus. These population genetics data allowed us to discuss the currently available mutation models for microsatellite sequences. Microsatellite markers revealed an unexpectedly high levels of genetic variation with <=41 alleles for one locus and gene diversity of 0.20-0.75 among populations. The values of any estimator of F(is) indicate high selfing rates in all populations. Linkage disequilibria observed at all loci for some populations may also indicate high levels of inbreeding. The large extent of genetic differentiation measured by F(st), R(st) or by a test for homogeneity between genic distributions is explained by both selfing and bottlenecks. Despite a limited gene flow, migration events could be detected when comparing different populations within ponds.     

Population genetic structure and levels of inbreeding depression in the Mediterranean island endemic Cyclamen creticum (Primulaceae)

The variation and evolution of reproductive traits in island plants have much attention from conservation and evolutionary biologists. However, plants on islands in the Mediterranean region have very little attention. In the present study, we examine the floral biology and mating system of Cyclamen creticum , a diploid perennial herb endemic to Crete and Karpathos. Our purpose is to quantify (1) variation and covariation of floral traits related to the mating system, (2) the ability of the species to self in the absence of pollinators and its relative performance on selfing and outcrossing and (3) generic diversity within and among populations. Pollen/ovule ratios were indicative of a xenogamous species. A controlled pollination experiment showed that the species is self-compatible but is unable to set seed, in the absence of pollinators, probably due to stigma-anther separation. A multiplicative estimate of inbreeding depression based on fruit maturation, seed number and percentage seed germination gave δ= 0.38 Population genetic diversity was high, 54.76% polymorphic loci, a mean of 1.78 alleles per locus and a mean observed heterozygosity of 0.053. F -statistics nevertheless indicated high inbreeding rates (mean F ^is= 0.748) in natural populations, and low levels of population differentiation (mean F^is= 0.168). C. creticum thus appears to have a mixed-mating system with high levels of (pollinator) mediated inbreeding (either by facilitated selfing, geitonogamy or biparental inbreeding) in natural populations.     

Inbreeding depression is high in a self‐incompatible perennial herb population but absent in a self‐compatible population showing mixed mating

High inbreeding depression is thought to be one of the major factors preventing evolutionary transitions in hermaphroditic plants from self‐incompatibility (SI) and outcrossing toward self‐compatibility (SC) and selfing. However, when selfing does evolve, inbreeding depression can be quickly purged, allowing the evolution of complete self‐fertilization. In contrast, populations that show intermediate selfing rates (a mixed‐mating system) typically show levels of inbreeding depression similar to those in outcrossing species, suggesting that selection against inbreeding might be responsible for preventing the transition toward complete self‐fertilization. By implication, crosses among populations should reveal patterns of heterosis for mixed‐mating populations that are similar to those expected for outcrossing populations. Using hand‐pollination crosses, we compared levels of inbreeding depression and heterosis between populations of Linaria cavanillesii (Plantaginaceae), a perennial herb showing contrasting mating systems. The SI population showed high inbreeding depression, whereas the SC population displaying mixed mating showed no inbreeding depression. In contrast, we found that heterosis based on between‐population crosses was similar for SI and SC populations. Our results are consistent with the rapid purging of inbreeding depression in the derived SC population, despite the persistence of mixed mating. However, the maintenance of outcrossing after a transition to SC is inconsistent with the prediction that populations that have purged their inbreeding depression should evolve toward complete selfing, suggesting that the transition to SC in L. cavanillesii has been recent. SC in L. cavanillesii thus exemplifies a situation in which the mating system is likely not at an equilibrium with inbreeding depression.     

Quantifying inbreeding in natural populations of hermaphroditic organisms

We review molecular methods for estimating selfing rates and inbreeding in populations. Two main approaches are available: the population structure approach (PSA) and progeny-array approach (PAA). The PSA approach relies on single-generation samples and produces estimates that integrate the inbreeding history over several generations, but is based on strong assumptions (for example, inbreeding equilibrium). The PSA has classically relied on single-locus inbreeding coefficients averaged over loci. Unfortunately PSA estimates are very sensitive to technical problems such as the occurrence of null alleles at one or more of the loci. Consequently inbreeding might be substantially overestimated, especially in outbred populations. However, the robustness of the PSA has recently been greatly improved by the development of multilocus methods free of such bias. The PAA, on the other hand, is based on the comparison between offspring and mother genotypes. As a consequence, PAA estimates do not reflect long-term inbreeding history but only recent mating events of the maternal individuals studied ('here and now' selfing). In addition to selfing rates, the PAA allows estimating other mating system parameters, including biparental inbreeding and the correlation of selfing among sibs. Although PAA estimates could also be biased by technical problems, incompatibilities between the mother's genotype and her offspring allow the identification and correction of such bias. For all methods, we provide guidelines on the required number of loci and sample sizes. We conclude that the PSA and PAA are equally robust, provided multilocus information is used. Although experimental constraints may make the PAA more demanding, especially in animals, the two methods provide complementary information, and can fruitfully be conducted together.     

Consequences of multiple mating‐system shifts for population and range‐wide genetic structure in a coastal dune plant

Evolutionary transitions from outcrossing to selfing can strongly affect the genetic diversity and structure of species at multiple spatial scales. We investigated the genetic consequences of mating‐system shifts in the North American, Pacific coast dune endemic plant Camissoniopsis cheiranthifolia (Onagraceae) by assaying variation at 13 nuclear (n) and six chloroplast (cp) microsatellite (SSR) loci for 38 populations across the species range. As predicted from the expected reduction in effective population size (N_e) caused by selfing, small‐flowered, predominantly selfing (SF) populations had much lower nSSR diversity (but not cpSSR) than large‐flowered, predominantly outcrossing (LF) populations. The reduction in nSSR diversity was greater than expected from the effects of selfing on N_e alone, but could not be accounted for by indirect effects of selfing on population density. Although selfing should reduce gene flow, SF populations were not more genetically differentiated than LF populations. We detected five clusters of nSSR genotypes and three groups of cpSSR haplotypes across the species range consisting of parapatric groups of populations that usually (but not always) differed in mating system, suggesting that selfing may often initiate ecogeographic isolation. However, lineage‐wide genetic variation was not lower for selfing clusters, failing to support the hypothesis that selection for reproductive assurance spurred the evolution of selfing in this species. Within three populations where LF and SF plants coexist, we detected genetic differentiation among diverged floral phenotypes suggesting that reproductive isolation (probably postzygotic) may help maintain the striking mating‐system differentiation observed across the range of this species.     

No evolutionary shift in the mating system of north American Ambrosia artemisiifolia (Asteraceae) following its introduction to China

The mating system plays a key role during the process of plant invasion. Contemporary evolution of uniparental reproduction (selfing or asexuality) can relieve the challenges of mate limitation in colonizing populations by providing reproductive assurance. Here we examined aspects of the genetics of colonization in Ambrosia artemisiifolia, a North American native that is invasive in China. This species has been found to possess a strong self-incompatibility system and have high outcrossing rates in North America and we examined whether there has been an evolutionary shift towards the dependence on selfing in the introduced range. Specifically, we estimated outcrossing rates in one native and five invasive populations and compared levels of genetic diversity between North America and China. Based on six microsatellite loci we found that, like the native North American population, all five Chinese populations possessed a completely outcrossing mating system. The estimates of paternity correlations were low, ranging from 0.028-0.122, which suggests that populations possessed ~8-36 pollen donor parents contributing to each maternal plant in the invasive populations. High levels of genetic diversity for both native and invasive populations were found with the unbiased estimate of gene diversity ranging from 0.262-0.289 for both geographic ranges based on AFLP markers. Our results demonstrate that there has been no evolutionary shift from outcrossing to selfing during A. artemisiifolia's invasion of China. Furthermore, high levels of genetic variation in North America and China indicate that there has been no erosion of genetic variance due to a bottleneck during the introduction process. We suggest that the successful invasion of A. artemisiifolia into Asia was facilitated by repeated introductions from multiple source populations in the native range creating a diverse gene pool within Chinese populations.     

COMPARATIVE FITNESS AND REPRODUCTIVE ISOLATION BETWEEN TWO BULINUS GLOBOSUS(GASTROPODA: PLANORBIDAE) POPULATIONS

Bulinus globosus, an intermediate host of schisto-somes, isa self-fertile hermaphrodite species whose regular mating systemis cross-fertilization. Fitness of selfing individuals originatingfrom two geographically remote populations (Elevi, Ivory Coast;Daikaina, Niger) was scored. Individuals were then paired eitherwith individuals from their own population, or with individualsfrom the other population. Electrophoretic analysis showed thatpairing ‘female’ Elevi and ‘male’ Daikainaindividuals results in only a low level of outcrossing, whereasthe reverse pairings produced almost only outcrossed offspring.This could reflect unilateral partial isolation between thetwo populations. However, analysis of fitness values withinand among groups of self-fertilizing and cross-fertilizing individuals,as well as electrophoretic data, suggest that selfing (or partialselfing) is the regular mating system in Elevi. Such experimentsare relevant to the biological control of schistosiomasis byintroducing resistant snails from allopatric populations, becausethey allow an assessment of the fate of introduced resistantgenes. (Received 30 December 1991; accepted 13 March 1992)     

Low selfing in a mass-flowering, endangered perennial, Eryngium alpinum L. (Apiaceae)

We investigated the reproductive ecology of an endangered alpine species, Eryngium alpinum L., to determine its selfing rate and to propose possible mechanisms that may shape its breeding system. Whereas pollinators' foraging behavior suggested a high potential for geitonogamy (70% of the flights occur within plants), microsatellite analyses of seed progenies demonstrated that plants are primarily outcrossing (outcrossing rate [tm] = 0.65, 0.96, and 1 in three populations). Given the relatively long pollen viability (at least 4-5 d) and the high number of simultaneously opened flowers on each plant, protandry is not sufficient to eliminate selfing. Second, controlled crosses demonstrated not only auto-fertility, but also partial self-incompatibility. Partial self-incompatibility is probably due to the competitive advantage of cross vs. self-pollen, and, together with protandry, could lead the species to selfing as a reproductive assurance. These results are encouraging for the maintenance of large populations. However, higher selfing was observed in a small population that could suffer inbreeding depression, as observed on experimentally selfed seeds. Thus, these populations should be carefully monitored. Finally, this study shows how molecular markers and field experiments may complement each other in our reaching a global understanding of mating patterns.     

A novel mating system analysis for modes of self-oriented mating applied to diploid and polyploid arctic Easter daisies (Townsendia hookeri)

We have developed a new model for mating system analysis, which attempts to distinguish among alternative modes of self-oriented mating within populations. This model jointly estimates the rates of outcrossing, selfing, automixis and apomixis, through the use of information in the family structure given by dominant genetic marker data. The method is presented, its statistical properties evaluated, and is applied to three arctic Easter daisy populations, one consisting of diploids, the other two of tetraploids. The tetraploids are predominantly male sterile and reported to be apomictic while the diploids are male fertile. In each Easter daisy population, 10 maternal arrays of six progeny were assayed for amplified fragment length polymorphism markers. Estimates, confirmed with likelihood ratio tests of mating hypotheses, showed apomixis to be predominant in all populations (ca. 70%), but selfing or automixis was moderate (ca. 25%) in tetraploids. It was difficult to distinguish selfing from automixis, and simulations confirm that with even very large sample sizes, the estimates have a very strong negative statistical correlation, for example, they are not independent. No selfing or automixis was apparent in the diploid population, instead, moderate levels of outcrossing were detected (23%). Low but significant levels of outcrossing (2-4%) seemed to occur in the male-sterile tetraploid populations; this may be due to genotyping error of this level. Overall, this study shows apomixis can be partial, and provides evidence for higher levels of inbreeding in polyploids compared to diploids and for significant levels of apomixis in a diploid plant population.     

Genetic Diversity and Population Structure in Polygonum cespitosum: Insights to an Ongoing Plant Invasion

Molecular markers can help elucidate how neutral evolutionary forces and introduction history contribute to genetic variation in invaders. We examined genetic diversity, population structure and colonization patterns in the invasive Polygonum cespitosum, a highly selfing, tetraploid Asian annual introduced to North America. We used nine diploidized polymorphic microsatellite markers to study 16 populations in the introduced range (northeastern North America), via the analyses of 516 individuals, and asked the following questions: 1) Do populations have differing levels of within-population genetic diversity? 2) Do populations form distinct genetic clusters? 3) Does population structure reflect either geographic distances or habitat similarities? We found low heterozygosity in all populations, consistent with the selfing mating system of P. cespitosum. Despite the high selfing levels, we found substantial genetic variation within and among P. cespitosum populations, based on the percentage of polymorphic loci, allelic richness, and expected heterozygosity. Inferences from individual assignment tests (Bayesian clustering) and pairwise F _ST values indicated high among-population differentiation, which indicates that the effects of gene flow are limited relative to those of genetic drift, probably due to the high selfing rates and the limited seed dispersal ability of P. cespitosum. Population structure did not reflect a pattern of isolation by distance nor was it related to habitat similarities. Rather, population structure appears to be the result of the random movement of propagules across the introduced range, possibly associated with human dispersal. Furthermore, the high population differentiation, genetic diversity, and fine-scale genetic structure (populations founded by individuals from different genetic sources) in the introduced range suggest that multiple introductions to this region may have occurred. High genetic diversity may further contribute to the invasive success of P. cespitosum in its introduced range.     

The interactive effects of herbivory and mixed mating for the population dynamics of Impatiens capensis

In this study, we examine the demographic consequences of mixed mating and explore the interactive effects of vegetative herbivory and mating system for population dynamics of Impatiens capensis, a species with an obligate mixed mating system (i.e., individuals produce both obligately selfing cleistogamous and facultatively outcrossing chasmogamous flowers). In two natural populations, we followed seeds derived from cleistogamous and chasmogamous flowers subject to different herbivory levels throughout their life cycle. Using a mating system-explicit projection matrix model, we found that mating system types differed in important vital rates. Cleistogamous individuals had higher rates of germination than did chasmogamous individuals, whereas chasmogamous individuals expressed a fecundity advantage over cleistogamous individuals. In addition, population growth was most sensitive to changes in vital rates of cleistogamous individuals, indicating the demographic importance of selfing for these populations. Herbivory also had demographic consequences; a 33%-49% reduction in herbivory caused the population growth rates to increase by 104%-132%, primarily because of effects on vital rates of selfed individuals. Our results not only uncover a novel consequence of mating system expression, that is, mating system influences population dynamics, but also shed light on the role of herbivores in maintaining mixed mating.     

Increased selfing and decreased effective pollen donor number in peripheral relative to central populations in Picea sitchensis (Pinaceae)

Because mating system can be influenced by effective neighborhood size, density, and isolation, populations at range peripheries may differ from those in the center. The importance of peripheral populations to conservation and evolution is controversial, and additional information about their genetic structure and evolutionary dynamics will inform conservation strategies. In wind-pollinated species, selfing rate is generally negatively correlated with population size and density, and inbreeding may therefore increase toward range peripheries. Picea sitchensis has a long and narrow range along the Pacific Coast of North America that tapers toward the northern and southern peripheries. We investigated whether central and peripheral populations differ in mating system parameters. The results suggest that population position within the range has a strong effect on mating system, and geographic isolation appears to be associated with higher selfing. The estimated effective number of pollen donors was much higher in the center of the range (mean = 18.5) than at the periphery (mean = 3.6), while selfing rate increased from 7.3% in central populations to as high as 35.2% in the northern, isolated population. These strong geographical patterns suggest mating system is influenced by both population size and isolation at range peripheries.     

Partial selfing, ecological disturbance and reproductive assurance in an invasive freshwater snail

Although reproductive assurance (RA) might play a central role in the evolution of the selfing rate, this hypothesis has never been seriously investigated in an hermaphroditic animal. We studied the mating system of the freshwater snail Physa acuta in which the availability of mating partners might be highly variable, because this species is an efficient colonizer occupying unstable habitats. A total of 11 populations differing in ecological disturbance regime (water level, openness) and snail densities were monitored over 2 years. The outcrossing rate was estimated in ca 10 families per population using microsatellite markers and the progeny-array approach. Components of fecundity and survival were recorded for each progeny. Predominant outcrossing (t(m)=0.94) was detected, with a few individuals (4%) purely selfing. The outcrossing rate did not explain among-family variation in fitness components. None of the predictions formulated under the RA hypothesis were verified: (i) selfing was related neither to disturbed habitats, nor to temporal density fluctuations, (ii) it was positively related to population density, (iii) it co-occurred with multiple paternity, and (iv) it did not induce delayed reproduction. Explanations for these negative results are discussed in light of other arguments supporting the RA hypothesis in P. acuta, as well as alternative theories explaining the occurrence of partial selfing, as either a genetically fixed or plastic trait.     

RECONSTRUCTING ORIGINS OF LOSS OF SELF‐INCOMPATIBILITY AND SELFING IN NORTH AMERICAN ARABIDOPSIS LYRATA: A POPULATION GENETIC CONTEXT

Theoretical and empirical comparisons of molecular diversity in selfing and outcrossing plants have primarily focused on long‐term consequences of differences in mating system (between species). However, improving our understanding of the causes of mating system evolution requires ecological and genetic studies of the early stages of mating system transition. Here, we examine nuclear and chloroplast DNA sequences and microsatellite variation in a large sample of populations of Arabidopsis lyrata from the Great Lakes region of Eastern North American that show intra‐ and interpopulation variation in the degree of self‐incompatibility and realized outcrossing rates. Populations show strong geographic clustering irrespective of mating system, suggesting that selfing either evolved multiple times or has spread to multiple genetic backgrounds. Diversity is reduced in selfing populations, but not to the extent of the severe loss of variation expected if selfing evolved due to selection for reproductive assurance in connection with strong founder events. The spread of self‐compatibility in this region may have been favored as colonization bottlenecks following glaciation or migration from Europe reduced standing levels of inbreeding depression. However, our results do not suggest a single transition to selfing in this system, as has been suggested for some other species in the Brassicaceae.     

Mating system parameters at hierarchical levels of fruits, individuals and populations in the Brazilian insect-pollinated tropical tree, Tabebuia roseo-alba (Bignoniaceae)

For many tree species, mating system analyses have indicated potential variations in the selfing rate and paternity correlation among fruits within individuals, among individuals within populations, among populations, and from one flowering event to another. In this study, we used eight microsatellite markers to investigate mating systems at two hierarchical levels (fruits within individuals and individuals within populations) for the insect pollinated Neotropical tree Tabebuia roseo-alba. We found that T. roseo-alba has a mixed mating system with predominantly outcrossed mating. The outcrossing rates at the population level were similar across two T. roseo-alba populations; however, the rates varied considerably among individuals within populations. The correlated paternity results at different hierarchical levels showed that there is a high probability of shared paternal parentage when comparing seeds within fruits and among fruits within plants and full-sibs occur in much higher proportion within fruits than among fruits. Significant levels of fixation index were found in both populations and biparental inbreeding is believed to be the main cause of the observed inbreeding. The number of pollen donors contributing to mating was low. Furthermore, open-pollinated seeds varied according to relatedness, including half-sibs, full-sibs, self-sibs and self-half-sibs. In both populations, the effective population size within a family (seed-tree and its offspring) was lower than expected for panmictic populations. Thus, seeds for ex situ conservation genetics, progeny tests and reforestation must be collected from a large number of seed-trees to guarantee an adequate effective population in the sample.     

Reliable selfing rate estimates from imperfect population genetic data

Genotypic frequencies at codominant marker loci in population samples convey information on mating systems. A classical way to extract this information is to measure heterozygote deficiencies (FIS) and obtain the selfing rate s from FIS = s/(2 - s), assuming inbreeding equilibrium. A major drawback is that heterozygote deficiencies are often present without selfing, owing largely to technical artefacts such as null alleles or partial dominance. We show here that, in the absence of gametic disequilibrium, the multilocus structure can be used to derive estimates of s independent of FIS and free of technical biases. Their statistical power and precision are comparable to those of FIS, although they are sensitive to certain types of gametic disequilibria, a bias shared with progeny-array methods but not FIS. We analyse four real data sets spanning a range of mating systems. In two examples, we obtain s = 0 despite positive FIS, strongly suggesting that the latter are artefactual. In the remaining examples, all estimates are consistent. All the computations have been implemented in a open-access and user-friendly software called rmes (robust multilocus estimate of selfing) available at http://ftp.cefe.cnrs.fr, and can be used on any multilocus data. Being able to extract the reliable information from imperfect data, our method opens the way to make use of the ever-growing number of published population genetic studies, in addition to the more demanding progeny-array approaches, to investigate selfing rates.     

Postcopulatory sexual selection reduces genetic diversity in experimental populations of Caenorhabditis elegans

Postcopulatory sexual selection affects the evolution of numerous features ranging from mating behavior to seminal fluid toxicity to the size of gametes. In an earlier study of the effect of sperm competition risk on sperm size evolution, experimental populations of the nematode Caenorhabditis elegans were maintained either by outcrossing (sperm competition present) or by selfing (no sperm competition), and after 60 generations, significantly larger sperm had evolved in the outcrossing populations. To determine the effects of this selection on population genetic variation, we assessed genetic diversity in a large number of loci using random amplification of polymorphic DNA-PCR. Nearly 80% of the alleles present in parental strain populations persisted in the 6 experimental populations after the 60 generations and, despite a 2.2-fold difference in expected heterozygosity, the resulting levels of genetic variation were equivalent between the outcrossing and selfing experimental populations. By inference, we conclude that genetic hitchhiking due to sexual selection in the experimental populations dramatically reduced genetic diversity. We use the levels of variation in the selfing populations as a control for the effects of drift, and estimate the strength of sexual selection to be strong in obligatorily outcrossing populations. Although sequential hermaphrodites like C. elegans probably experience little sexual selection in nature, these data suggest that sexual selection can profoundly affect diversity in outcrossing taxa.     

An explicit model for the inbreeding load in the evolutionary analysis of selfing

I present analytical predictions for the equilibrium inbreeding load expected in a population under mutation, selection, and a regular mating system for any population size and for any magnitude and recessivity of the deleterious effects. Using this prediction, I deduce the relative fitness of mutant alleles with small effect on selfing to explore the situations where selfing or outcrossing are expected to evolve. The results obtained are in agreement with previous literature, showing that natural selection is expected to lead to stable equilibria where populations show either complete outcrossing or complete selfing, and that selfing is promoted by large deleterious mutation rates. I find that the evolution of selfing is favored by a large recessivity of deleterious effects, while the magnitude of homozygous deleterious effects only becomes relevant in relatively small populations. This result contradicts the standard assumption that purging in large populations will only promote selfing when homozygous deleterious effects are large, and implies that previously published results obtained assuming lethal mutations in large populations can be extrapolated to nonlethal alleles of similar recessivity. This conclusion and the general approach used in this analysis can be useful in the study of the evolution of mating systems.     

Transposable element number in mixed mating populations

Theoretical population genetic studies of transposable elements focus almost exclusively on random mating species, whereas many plants reproduce through partial or substantial self-fertilization. Here I develop computer simulation and analytic approximations of simplified element dynamics (transposition balanced by selective elimination) in partially self-fertilizing populations, using Ty1-copia elements for biological inspiration. Under the most plausible models and parameter values, element numbers decrease with self-fertilization when element insertions are deleterious, but may increase when ectopic exchange regulates element number. Conclusions for models of ectopic exchange depend in part on parameters for which little firm empirical evidence is available. Small changes in selfing rate can lead to abrupt changes in element number when homozygous and heterozygous elements have markedly different fitness effects. Equilibrium element numbers can be sensitive to population size, especially at high selfing rates. Elements are frequently lost in small highly selfing populations under the deleterious insertion model. In contrast, small highly selfing populations can accumulate very large numbers of elements under ectopic exchange. Empirical data on element number and localization in plants with different mating systems suggests that deleterious insertion, rather than ectopic exchange, may regulate element number. Limitations to available empirical data, especially the lack of comparison between closely related species differing in mating system, mean that this conclusion is tentative.     

Mating system variation in the hermaphroditic brooding coral, Seriatopora hystrix

Self-compatible, hermaphroditic marine invertebrates have the potential to self-fertilize in the absence of mates or under sperm-limited conditions, and outcross when sperm is available from a variety of males. Hence, many hermaphroditic marine invertebrates may have evolved mixed-mating systems that involve facultative self-fertilization. Such mixed-mating strategies are well documented for plants but have rarely been investigated in animals. Here, I use allozyme markers to make estimates of selfing from population surveys of reef slope and reef flat sites, and contrast this with direct estimates of selfing from progeny-array analysis, for the brooding coral Seriatopora hystrix. Consistent heterozygote deficits previously reported for S. hystrix suggests that inbreeding (including the extreme of selfing) may be common in this species. I detected significant levels of inbreeding within populations (F(IS)=0.48) and small but significant differentiation among all sites (F(ST)=0.04). I detected no significant differentiation among habitats (F(HT)=0.009) though among site differentiation did occur within the reef slope habitat (F(SH)=0.06), but not within the reef flat habitat (F(SH)=0.015). My direct estimates of outcrossing for six colonies and their progeny from a single reef flat site revealed an intermediate value (t(m) (+/-s.d.)=0.53+/-0.20). Inbreeding coefficients calculated from progeny arrays (F(e)=0.31) were similar to indirect estimates based on adult genotype frequencies for that site (F(IS)=0.38). This study confirms that the mating system of this brooding coral is potentially variable, with both outcrossing and selfing.