Inbreeding depression and low between-population heterosis in recently diverged experimental populations of a selfing species

In fragmented populations, genetic drift and selection reduce genetic diversity, which in turn results in a loss of fitness or in a loss of evolvability. Genetic rescue, that is, controlled input of diversity from distant populations, may restore evolutionary potential, whereas outbreeding depression might counteract the positive effect of this strategy. We carried out self-pollination and crosses within and between populations in an experimental subdivided population of a selfing species, Triticum aestivum L., to estimate the magnitude of these two phenomena. Surprisingly, for a self-fertilizing species, we found significant inbreeding depression within each population for four of the six traits studied, indicating that mildly deleterious mutations were still segregating in these populations. The progeny of within- and between-population crosses was very similar, indicating low between-population heterosis and little outbreeding depression. We conclude that relatively large population effective sizes prevented fixation of a high genetic load and that local adaptation was limited in these recently diverged populations. The kinship coefficient estimated between the parents using 20 neutral markers was a poor predictor of the progeny phenotypic values, indicating that there was a weak link between neutral diversity and genes controlling fitness-related traits. These results show that when assessing the viability of natural populations and the need for genetic rescue, the use of neutral markers should be complemented with information about the presence of local adaptation in the subdivided population.     

Estimating inbreeding depression in natural plant populations using quantitative and molecular data

Inbreeding and inbreeding depression are important issues in the biology and conservation of natural plant and animal populations, primarily when subpopulation sizes are reduced due to habitat fragmentation. In this study, we propose a method for estimating inbreeding depression in progenies of natural plant populations, combining the estimation of the fixation index by codominant markers with the experimental evaluation of quantitative traits. Our technique estimates apparent inbreeding depression in structured natural populations using the linear regression of phenotypic means on the inbreeding coefficients estimated with codominant markers. This method was applied to data from 112 maternal progenies of 10 natural subpopulations of Eugenia dysenterica DC, a fruiting tree species from the Brazilian savanna (Cerrado). The results show that the proposed method was efficient at detecting the presence of inbreeding depression for seedling emergence and initial growth traits in the species. This corroborates the importance of maintaining high levels of heterozygosity for in situ conservation or genetic restoration of natural populations.     

Inbreeding and outbreeding depression in Stylidium hispidum: implications for mixing seed sources for ecological restoration

The benefits of composite rather than local seed provenances for ecological restoration have recently been argued, largely on the basis of maximizing evolutionary potential. However, these arguments have downplayed the potentially negative consequences of outbreeding depression once mixed provenances interbreed. In this study, we compared intraspecific F1 hybrid performance and molecular marker differentiation among four populations of Stylidium hispidum, a species endemic to Southwestern Australia. Multivariate ordination of 134 AFLP markers analyzed genetic structure and detected two clusters of paired sites that diverged significantly for marker variation along a latitudinal boundary. To test for outbreeding depression and to determine the consequences of molecular population divergence for hybrid fitness, we conducted controlled pollinations and studied germination and survival for three cross categories (within‐population crosses, short‐ and long‐distance F1 hybrids) for paired sites distributed within and between the two genetically differentiated regions. We found evidence of outbreeding depression in long‐distance hybrids (111–124 km), and inbreeding depression among progeny of within‐population crosses, relative to short‐distance (3–10 km) hybrids, suggesting an intermediate optimal outcrossing distance in this species. These results are discussed in light of the evolutionary consequences of mixing seed sources for biodiversity restoration.     

Effects of competition on lifetime estimates of inbreeding depression in the outcrossing plant Crepis sancta (Asteraceae)

Inbreeding depression was studied in two populations of a Mediterranean allogamous colonizing species Crepis sancta. In order to test the hypothesis that the magnitude of inbreeding depression can be modified by successional processes, the growth and survival of individuals resulting from two generations of inbred crosses including selfing were analysed with interspecific competition (in natural vegetation) and without interspecific competition (by removing natural vegetation). Inbreeding depression was weak for seed production. Germination was little affected by inbreeding but mortality and the number of capitula showed inbreeding depression, especially in the presence of competition. This suggests that inbreeding depression is very sensitive to variations in environmental conditions such as interspecific competition. As a consequence, inbreeding depression cannot be considered as constant in natural conditions.     

Effects of outcrossing in fragmented populations of the primarily selfing forest herb <Emphasis Type="Italic">Geum urbanum</Emphasis>

In fragmented landscapes, small populations may be subjected to inbreeding or genetic drift. Gene flow is expected to alleviate the burden of deleterious mutations in such populations. The beneficial effects of outcrossing may, however, depend on life history characteristics such as the species’ breeding system. Frequent selfing is expected to purge (sub)lethal alleles and mitigate inbreeding depression, at least if the load of mildly deleterious mutations has not accumulated through genetic drift in populations with a small effective size. Gene-inflow from distant source populations can cause outbreeding depression due to genomic incompatibilities. We tested these predictions using highly fragmented populations of the self-compatible forest herb Geum urbanum. Assessment of mating system parameters using microsatellite markers inferred high selfing rates (92.5%), confirming the predominantly self-fertilizing character of the study species. We conducted experimental pollinations with self and outcross pollen collected from populations at different distances from the target populations. There were no significant signs of inbreeding depression, even in very small target populations. Except for a minor negative effect on the germination rate for the long-distance crosses, we found no effects of outbreeding on fitness estimates.     

How relatedness between mates influences reproductive success: An experimental analysis of self‐fertilization and biparental inbreeding in a marine bryozoan

Kin associations increase the potential for inbreeding. The potential for inbreeding does not, however, make inbreeding inevitable. Numerous factors influence whether inbreeding preference, avoidance, or tolerance evolves, and, in hermaphrodites where both self‐fertilization and biparental inbreeding are possible, it remains particularly difficult to predict how selection acts on the overall inbreeding strategy, and to distinguish the type of inbreeding when making inferences from genetic markers. Therefore, we undertook an empirical analysis on an understudied type of mating system (spermcast mating in the marine bryozoan, Bugula neritina) that provides numerous opportunities for inbreeding preference, avoidance, and tolerance. We created experimental crosses, containing three generations from two populations to estimate how parental reproductive success varies across parental relatedness, ranging from self, siblings, and nonsiblings from within the same population. We found that the production of viable selfed offspring was extremely rare (only one colony produced three selfed offspring) and biparental inbreeding more common. Paternity analysis using 16 microsatellite markers confirmed outcrossing. The production of juveniles was lower for sib mating compared with nonsib mating. We found little evidence for consistent inbreeding, in terms of nonrandom mating, in adult samples collected from three populations, using multiple population genetic inferences. Our results suggest several testable hypotheses that potentially explain the overall mating and dispersal strategy in this species, including early inbreeding depression, inbreeding avoidance through cryptic mate choice, and differential dispersal distances of sperm and larvae.     

Mixing source populations increases genetic diversity of restored rare plant populations

The genetic diversity of germplasm used in reintroduction and restoration efforts can influence how resulting populations establish, reproduce, and evolve over time, particularly in disturbed and changing conditions. Regional admixture provenancing, mixing seeds derived from multiple populations within the same region as the target site, has been suggested to produce genetically diverse germplasm. Yet little empirical evidence shows how genetic diversity in germplasm resulting from this approach compares to source populations, or how it varies in restored populations. Here, we use neutral molecular markers to follow genetic diversity through production and use of germplasm when mixing multiple source populations in nursery production beds. Castilleja levisecta is a rare species experiencing inbreeding depression in remaining populations, with a federal recovery plan requiring the re‐establishment of populations in areas where it has been extirpated. Specifically, we track diversity from wild‐collected source populations through different production approaches and reintroductions using two propagule types. We show that measures of genetic diversity, inbreeding, and relatedness change during the production and use of material produced with a regional admixture provenancing approach, with the step at which source populations are mixed and germplasm type used influencing whether all source populations are equally represented. While genetic diversity increased throughout the process, inbreeding and relatedness increased in nursery production beds but decreased in reintroductions, with the lowest inbreeding and relatedness in populations restored using seeds rather than plugs. The results highlight the importance of taking an integrated approach informed by research when planning and implementing reintroductions with mixed‐source germplasm.     

Molecular and pedigree measures of relatedness provide similar estimates of inbreeding depression in a bottlenecked population

Individual‐based estimates of the degree of inbreeding or parental relatedness from pedigrees provide a critical starting point for studies of inbreeding depression, but in practice wild pedigrees are difficult to obtain. Because inbreeding increases the proportion of genomewide loci that are identical by descent, inbreeding variation within populations has the potential to generate observable correlations between heterozygosity measured using molecular markers and a variety of fitness related traits. Termed heterozygosity‐fitness correlations (HFCs), these correlations have been observed in a wide variety of taxa. The difficulty of obtaining wild pedigree data, however, means that empirical investigations of how pedigree inbreeding influences HFCs are rare. Here, we assess evidence for inbreeding depression in three life‐history traits (hatching and fledging success and juvenile survival) in an isolated population of Stewart Island robins using both pedigree‐ and molecular‐derived measures of relatedness. We found results from the two measures were highly correlated and supported evidence for significant but weak inbreeding depression. However, standardized effect sizes for inbreeding depression based on the pedigree‐based kin coefficients (k) were greater and had smaller standard errors than those based on molecular genetic measures of relatedness (RI), particularly for hatching and fledging success. Nevertheless, the results presented here support the use of molecular‐based measures of relatedness in bottlenecked populations when information regarding inbreeding depression is desired but pedigree data on relatedness are unavailable.     

Recent approaches into the genetic basis of inbreeding depression in plants

Predictions for the evolution of mating systems and genetic load vary, depending on the genetic basis of inbreeding depression (dominance versus overdominance, epistasis and the relative frequencies of genes of large and small effect). A distinction between the dominance and overdominance hypotheses is that deleterious recessive mutations should be purged in inbreeding populations. Comparative studies of populations differing in their level of inbreeding and experimental approaches that allow selection among inbred lines support this prediction. More direct biometric approaches provide strong support for the importance of partly recessive deleterious alleles. Investigators using molecular markers to study quantitative trait loci (QTL) often find support for overdominance, though pseudo-overdominance (deleterious alleles linked in repulsion) may bias this perception. QTL and biometric studies of inbred lines often find evidence for epistasis, which may also contribute to the perception of overdominance, though this may be because of the divergent lines initially crossed in QTL studies. Studies of marker segregation distortion commonly uncover genes of major effect on viability, but these have only minor contributions to inbreeding depression. Although considerable progress has been made in understanding the genetic basis of inbreeding depression, we feel that all three aspects merit more study in natural plant populations.     

Partial self-incompatibility and inbreeding depression in a native tree species of La Réunion (Indian Ocean)

We investigated the reproductive system of the threatened taxon Dombeya acutangula ssp. acutangula Cav. (Sterculiaceae), an endemic tree of the Mascarene archipelago (Indian Ocean). A controlled crossing experiment was performed in two natural populations located in the remnants of the low-elevation dry forest on the island of La Réunion. Active pollination, probably mainly by insects, was necessary for reproduction in this species. Individuals varied in their degree of self-sterility from 0 to 100%. Outcrossing between nearby individuals produced lower seed set than did crosses between more distant individuals within one of the two tested populations. The variation in reproductive success on selfing and in the different types of crosses could result from inbreeding depression causing embryo death, and we provide evidence that progenies from selfing have lower seed size and quality. However, for inbreeding depression to account for the dramatic variation in seed set found in our crossing experiment, the distribution of genetic load and number of lethal factors required appear unrealistic. We favour an alternative interpretation, that D. acutangula possesses an incompatibility system similar to that found in other Sterculiaceae species such as Theobroma cacao L. Such an incompatibility system allows a certain amount of selfing, and different individuals vary in their degree of self-incompatibility. The low success of crosses among close neighbours in one population suggests that there was spatial structure for incompatibility alleles in that population. This could partly explain the decline of the species in fragmented and disturbed habitats, since relatedness at incompatibility loci may increase in small or isolated population and thus reduce mate availability. Received: 2 March 1998 / Accepted: 3 August 1998     

Estimating levels of inbreeding using AFLP markers

In the absence of detailed pedigree records, researchers have attempted to estimate individuals' levels of inbreeding using molecular markers, generally making use of heterozygosity measures based on microsatellite markers. Here we report and validate a method for estimating an individual's inbreeding coefficient, f, using amplified fragment length polymorphism (AFLP) markers. We use simulations to confirm that our measure scales appropriately with f when allele frequencies can be estimated from a subset of outbred individuals. We also present an approach for obtaining satisfactory estimates even in the absence of an independent set of known outbred individuals from which to estimate allele frequencies. We then test our method against empirical data from 179 wild and captive-bred old-field mice, Peromyscus polionotus subgriseus, comprising pedigree-based estimates of f, along with genetic data from 94 AFLP markers and 12 microsatellites. Inbreeding estimates based on both AFLP and microsatellite markers were found to correlate strongly with pedigree-based inbreeding coefficients. Owing to their ease of amplification in any species, AFLP markers may prove to be a valuable new tool for estimating f in natural populations and for examining correlations between heterozygosity and fitness.     

Comparing molecular measures for detecting inbreeding depression

Correlations between heterozygosity and components of fitness have been investigated in natural populations for over 20 years. Positive correlations between a trait of interest and heterozygosity (usually measured at allozyme loci) are generally recognized as evidence of inbreeding depression. More recently, molecular markers such as microsatellites have been employed for the same purpose. A typical study might use around five to ten markers. In this paper we use a panel of 71 microsatellite loci to: (1) Compare the efficacy of heterozygosity and a related microsatellite‐specific variable, mean d², in detecting inbreeding depression; (2) Examine the statistical power of heterozygosity to detect such associations. We performed our analyses in a wild population of red deer (Cervus elaphus) in which inbreeding depression in juvenile traits had previously been detected using a panel of nine markers. We conclude that heterozygosity‐based measures outperform mean d²‐based measures, but that power to detect heterozygosity‐fitness associations is nonetheless low when ten or fewer markers are typed.     

Release from natural enemies mitigates inbreeding depression in native and invasive Silene latifolia populations

Inbreeding and enemy infestation are common in plants and can synergistically reduce their performance. This inbreeding ×environment (I × E) interaction may be of particular importance for the success of plant invasions if introduced populations experience a release from attack by natural enemies relative to their native conspecifics. Here, we investigate whether inbreeding affects plant infestation damage, whether inbreeding depression in growth and reproduction is mitigated by enemy release, and whether this effect is more pronounced in invasive than native plant populations. We used the invader Silene latifolia and its natural enemies as a study system. We performed two generations of experimental out‐ and inbreeding within eight native (European) and eight invasive (North American) populations under controlled conditions using field‐collected seeds. Subsequently, we exposed the offspring to an enemy exclusion and inclusion treatment in a common garden in the species’ native range to assess the interactive effects of population origin (range), breeding treatment, and enemy treatment on infestation damage, growth, and reproduction. Inbreeding increased flower and leaf infestation damage in plants from both ranges, but had opposing effects on fruit damage in native versus invasive plants. Inbreeding significantly reduced plant fitness; whereby, inbreeding depression in fruit number was higher in enemy inclusions than exclusions. This effect was equally pronounced in populations from both distribution ranges. Moreover, the magnitude of inbreeding depression in fruit number was lower in invasive than native populations. These results support that inbreeding has the potential to reduce plant defenses in S. latifolia, which magnifies inbreeding depression in the presence of enemies. However, future studies are necessary to further explore whether enemy release in the invaded habitat has actually decreased inbreeding depression and thus facilitated the persistence of inbred founder populations and invasion success.     

Inbreeding depression and heterosis in populations of Schiedea viscosa, a highly selfing species

Progeny from self-pollinations and intrapopulation crosses were examined in Schiedea viscosa to determine the extent of inbreeding depression in this highly selfing species. Progeny of interpopulation crosses were also investigated to determine whether deleterious alleles have been fixed in populations of S. viscosa. There was no inbreeding depression at early life history stages, including seeds per capsule, seed mass, or germination. Inbreeding depression was detected for the later life history stage of fruit biomass, although not for survival or life span. Heterosis for vegetative biomass and fruit biomass was detected in progeny from crosses between populations. Levels of inbreeding depression in S. viscosa are low relative to out-crossing species of Schiedea, especially when early life history stages are compared.     

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.     

The genetic rescue of two bottlenecked South Island robin populations using translocations of inbred donors

Populations forced through bottlenecks typically lose genetic variation and exhibit inbreeding depression. ‘Genetic rescue’ techniques that introduce individuals from outbred populations can be highly effective in reversing the deleterious effects of inbreeding, but have limited application for the majority of endangered species, which survive only in a few bottlenecked populations. We tested the effectiveness of using highly inbred populations as donors to rescue two isolated and bottlenecked populations of the South Island robin (Petroica australis). Reciprocal translocations significantly increased heterozygosity and allelic diversity. Increased genetic diversity was accompanied by increased juvenile survival and recruitment, sperm quality, and immunocompetence of hybrid individuals (crosses between the two populations) compared with inbred control individuals (crosses within each population). Our results confirm that the implementation of ‘genetic rescue’ using bottlenecked populations as donors provides a way of preserving endangered species and restoring their viability when outbred donor populations no longer exist.     

Insight into post-mating interactions between the sexes: relatedness suppresses productivity of singly mated female Drosophila melanogaster

Post-mating, prefertilization inbreeding avoidance (PPIA) is well established in plants but not in animals. Support for animal PPIA comes from sperm competition studies showing success of a male's gametes declining with his relatedness to the multiply mated female; however, such studies confound female-male and male-male interaction. To avoid this problem, we investigated offspring productivity of singly mated Drosophila melanogaster females using flies from four different genetic backgrounds. Our experiments established that intrapopulation crosses using highly related parents (within-strain) were significantly less productive than intrapopulation crosses using unrelated individuals from the same population (between-strain). Furthermore, we showed that these effects were not due to inbreeding depression. The average decrease in offspring productivity of within-strain crosses relative to between-strain crosses was 18.3% [nonlaboratory populations: Zimbabwe 20.3%, Riverside 11.4%, neither of which showed inbreeding depression; and temperature-adapted laboratory populations, uncorrected (corrected) for nonsignificant inbreeding depression: 18 degrees C, 26.5% (24.2%) and 29 degrees C, 20.1% (9.5%)]. The significant reduction of within-cross productivity demonstrates PPIA in the absence of multiple mating.     

Understanding the relationship between the inbreeding coefficient and multilocus heterozygosity: theoretical expectations and empirical data

Geneticists have been interested in inbreeding and inbreeding depression since the time of Darwin. Two alternative approaches that can be used to measure how inbred an individual is involve the use of pedigree records to estimate inbreeding coefficients or molecular markers to measure multilocus heterozygosity. However, the relationship between inbreeding coefficient and heterozygosity has only rarely been investigated. In this paper, a framework to predict the relationship between the two variables is presented. In addition, microsatellite genotypes at 138 loci spanning all 26 autosomes of the sheep genome were used to investigate the relationship between inbreeding coefficient and multilocus heterozygosity. Multilocus heterozygosity was only weakly correlated with inbreeding coefficient, and heterozygosity was not positively correlated between markers more often than expected by chance. Inbreeding coefficient, but not multilocus heterozygosity, detected evidence of inbreeding depression for morphological traits. The relevance of these findings to the causes of heterozygosity--fitness correlations is discussed and predictions for other wild and captive populations are presented.     

Close‐kin mating,but not inbred parents,reduces hatching rates and offspring quality in a threatened tortoise

Inbreeding depression, the reduction in fitness due to mating of related individuals, is of particular conservation concern in species with small, isolated populations. Although inbreeding depression is widespread in natural populations, long‐lived species may be buffered from its effects during population declines due to long generation times and thus are less likely to have evolved mechanisms of inbreeding avoidance than species with shorter generation times. However, empirical evidence of the consequences of inbreeding in threatened, long‐lived species is limited. In this study, we leverage a well‐studied population of gopher tortoises, Gopherus polyphemus, to examine the role of inbreeding depression and the potential for behavioural inbreeding avoidance in a natural population of a long‐lived species. We tested the hypothesis that increased parental inbreeding leads to reduced hatching rates and offspring quality. Additionally, we tested for evidence of inbreeding avoidance. We found that high parental relatedness results in offspring with lower quality and that high parental relatedness is correlated with reduced hatching success. However, we found that hatching success and offspring quality increase with maternal inbreeding, likely due to highly inbred females mating with more distantly related males. We did not find evidence for inbreeding avoidance in males and outbred females, suggesting sex‐specific evolutionary trade‐offs may have driven the evolution of mating behaviour. Our results demonstrate inbreeding depression in a long‐lived species and that the evolution of inbreeding avoidance is shaped by multiple selective forces.     

Inbreeding depression in self-incompatible and self-compatible populations of Leavenworthia alabamica

Inbreeding depression is one of the leading factors preventing the evolution of self-fertilization in plants. In populations where self-fertilization evolves, theory suggests that natural selection against partially recessive deleterious alleles will reduce inbreeding depression. The purpose of this study was to evaluate this hypothesis by comparing the magnitude of inbreeding depression in self-incompatible and self-compatible populations of Leavenworthia alabamica. Within-population crosses were conducted to compare the quantity and quality of offspring produced by outcrossing and self-fertilization. These progeny were grown in a common greenhouse and inbreeding depression was measured in germination, survival, biomass, transition rate to flowering, flower number, petal length, pollen grains/anther, pollen viability, and ovule number. In comparison to outcrossing, self-fertilization led to the production of fewer and smaller seeds within self-incompatible populations. Moreover, inbreeding depression was observed in eight of 11 offspring traits within self-incompatible populations of L. alabamica. In contrast, there was significant inbreeding depression only in flower number within self-compatible populations. The results of this study are consistent with the idea that self-fertilization selectively removes partially recessive deleterious alleles causing inbreeding depression in natural plant populations. However, in plant species such as L. alabamica where self-compatibility may evolve in small populations following long-distance dispersal, declines in inbreeding depression may also be facilitated by genetic drift.