Inbreeding depression in small populations of self-incompatible plants.

Self-incompatibility (SI) is a widespread mechanism that prevents inbreeding in flowering plants. In many species, SI is controlled by a single locus (the S locus) where numerous alleles are maintained by negative frequency-dependent selection. Inbreeding depression, the decline in fitness of selfed individuals compared to outcrossed ones, is an essential factor in the evolution of SI systems. Conversely, breeding systems influence levels of inbreeding depression. Little is known about the joint effect of SI and drift on inbreeding depression. Here we studied, using a two-locus model, the effect of SI (frequency-dependent selection) on a locus subject to recurrent deleterious mutations causing inbreeding depression. Simulations were performed to assess the effect of population size and linkage between the two loci on the level of inbreeding depression and genetic load. We show that the sheltering of deleterious alleles linked to the S locus strengthens inbreeding depression in small populations. We discuss the implications of our results for the evolution of SI systems.     

Breakdown of self-incompatibility in the perennial Arabidopsis lyrata (Brassicaceae) and its genetic consequences

Mating systems in plants are known to be highly labile traits, with frequent transitions from outcrossing to selfing. The genetic basis for breakdown in self-incompatibility (SI) systems has been studied, but data on variation in selfing rates in species for which the molecular basis of SI is known are rare. This study surveyed such variation in Arabidopsis lyrata (Brassicaceae), which is often considered an obligately outcrossing species, to examine the causes and genetic consequences of changes in its breeding system. Based on controlled self-pollinations in the greenhouse, three populations from the Great Lakes region of North America included a minority of self-compatible (SC) individuals, while two showed larger proportions of SC individuals and all populations contained some individuals capable of setting selfed seeds. Loss of SI was not associated with particular haplotypes at the S-locus (as estimated by alleles amplified at the SRK locus, the gene controlling female specificity) and all populations contained similar numbers of SRK alleles, suggesting that some other genetic factor is responsible for modifying the SI reaction. The loss of SI has resulted in an effective shift in the mating system, as the two populations with a high frequency of SC individuals showed significantly lower microsatellite-based multilocus outcrossing rates and higher inbreeding coefficients than the other populations. Based on microsatellites, observed heterozygosities and genetic diversity were also significantly depressed in these populations. These findings provide the unique opportunity to examine in detail the consequences of mating system changes within a species with a well-characterized SI system.     

THE ROLE OF GENES OF LARGE EFFECT ON INBREEDING DEPRESSION IN MIMULUS GUTTATUS

Severe inbreeding depression is routinely observed in outcrossing species. If inbreeding load is due largely to deleterious alleles of large effect, such as recessive lethals or steriles, then most of it is expected to be purged during brief periods of inbreeding. In contrast, if inbreeding depression is due to the cumulative effects of many deleterious alleles of small effect, then it will be maintained in the face of periodic inbreeding. Whether or not inbreeding depression can be purged with inbreeding in the short term has important implications for the evolution of mating systems and the probability that a small population will go extinct. In this paper I evaluate the extent to which the tremendous inbreeding load in a primarily outcrossing population of the wildflower, Mimulus guttatus, is due to alleles of large effect. To do this, I first constructed a large outbred “ancestral” population by randomly mating plants collected as seeds from a natural population. From this population I formed 1200 lines that were maintained by self-fertilization and single seedling descent: after five generations of selling, 335 lines had survived the inbreeding process. Selection during the line formation is expected to have largely purged alleles of large effect from the collection of highly inbred lines. Because alleles with minor effects on fitness should have been effectively neutral, the inbreeding depression due to this class of genes should have been unchanged. The inbred lines were intercrossed to form a large, outcrossed “purged” population. Finally, I estimated the fitness of outbred and selfed progeny from the ancestral and purged populations to determine the contribution of major deleterious alleles on inbreeding depression. I found that although the average fitness of the outcrossed progeny nearly doubled following purging, the limited decline in inbreeding depression and limited increase in inbred fitness indicates that alleles of large effect are not the principle cause of inbreeding depression in this population. In aggregate, the data suggest that lethals and steriles make a minority contribution to inbreeding depression and that the increased outbred fitness is due primarily to adaptation to greenhouse conditions.     

Digest: Exposing the role of rare alleles in inbreeding depression in monkeyflower*

Outcrossing is maintained in many hermaphroditic species despite theoretical work suggesting that alleles increasing selfing should invade outcrossing populations. Brown and Kelly (2019) identify reasons why this may not have occurred in an outcrossing population of monkeyflower, namely that inbreeding depression causes strong reductions in fitness, resulting in selection for the maintenance of outcrossing. They find that genetic load imposed by rare alleles is inversely correlated with fitness-associated traits, providing evidence that recessive, deleterious alleles contribute to inbreeding depression.     

Effects of inbreeding on male function and self-fertility in the partially self-incompatible herb Campanula rapunculoides (Campanulaceae)

We examined the effect of inbreeding on fitness (through both male and female functions) and changes in self-fertility in the partially self-incompatible species Campanula rapunculoides. Individuals in natural populations of C. rapunculoides varied extensively in their strength of self-incompatibility (SI). We crossed 11 individuals that differed in their strength of SI to generate families with four levels of inbreeding (f = 0.0, 0.25, 0.5, and 0.75). Progeny were scored for three traits related to male fitness and for outcrossed and selfed seed production. Analyses of variance revealed significant inbreeding depression for the three male traits and seed set. Families with strong or weak SI differed in their response to inbreeding. Families with weak SI had lower levels of inbreeding depression for most traits than families with strong SI, but strong SI families had a greater increase in selfed seed set, but not self-fertility, with inbreeding. Finally, we found evidence of a significant linear response to inbreeding for all three male reproductive traits and outcrossed seed, indicating that inbreeding depression was primarily caused by partially or fully recessive deleterious alleles. Variation in genetic load was associated with variation in self-fertility, a finding that suggests an evolutionary role for partial self-fertility in natural populations of C. rapunculoides.     

Inbreeding load, average dominance and the mutation rate for mildly deleterious alleles in Mimulus guttatus

The goal of this study is to provide information on the genetics of inbreeding depression in a primarily outcrossing population of Mimulus guttatus. Previous studies of this population indicate that there is tremendous inbreeding depression for nearly every fitness component and that almost all of this inbreeding depression is due to mildly deleterious alleles rather than recessive lethals or steriles. In this article I assayed the homozygous and heterozygous fitnesses of 184 highly inbred lines extracted from a natural population. Natural selection during the five generations of selfing involved in line formation essentially eliminated major deleterious alleles but was ineffective in purging alleles with minor fitness effects and did not appreciably diminish overall levels of inbreeding depression. Estimates of the average degree of dominance of these mildly deleterious alleles, obtained from the regression of heterozygous fitness on the sum of parental homozygous fitness, indicate that the detrimental alleles are partially recessive for most fitness traits, with h approximately 0.15 for cumulative measures of fitness. The inbreeding load, B, for total fitness is approximately 1.0 in this experiment. These results are consistent with the hypothesis that spontaneous mildly deleterious mutations occur at a rate >0.1 mutation per genome per generation.     

Mating system and ploidy influence levels of inbreeding depression in Clarkia (Onagraceae)

Inbreeding depression is the reduction in offspring fitness associated with inbreeding and is thought to be one of the primary forces selecting against the evolution of self-fertilization. Studies suggest that most inbreeding depression is caused by the expression of recessive deleterious alleles in homozygotes whose frequency increases as a result of self-fertilization or mating among relatives. This process leads to the selective elimination of deleterious alleles such that highly selfing species may show remarkably little inbreeding depression. Genome duplication (polyploidy) has also been hypothesized to influence levels of inbreeding depression, with polyploids expected to exhibit less inbreeding depression than diploids. We studied levels of inbreeding depression in allotetraploid and diploid species of Clarkia (Onagraceae) that vary in mating system (each cytotype was represented by an outcrossing and a selfing species). The outcrossing species exhibited more inbreeding depression than the selfing species for most fitness components and for two different measures of cumulative fitness. In contrast, though inbreeding depression was generally lower for the polyploid species than for the diploid species, the difference was statistically significant only for flower number and one of the two measures of cumulative fitness. Further, we detected no significant interaction between mating system and ploidy in determining inbreeding depression. In sum, our results suggest that a taxon's current mating system is more important than ploidy in influencing levels of inbreeding depression in natural populations of these annual plants.     

MUTATIONAL MELTDOWN IN SELFING ARABIDOPSIS LYRATA

The majority of plant species and many animals are hermaphrodites, with individuals expressing both female and male function. Although hermaphrodites can potentially reproduce by self‐fertilization, they have a high prevalence of outcrossing. The genetic advantages of outcrossing are described by two hypotheses: avoidance of inbreeding depression because selfing leads to immediate expression of recessive deleterious mutations, and release from drift load because self‐fertilization leads to long‐term accumulation of deleterious mutations due to genetic drift and, eventually, to extinction. I tested both hypotheses by experimentally crossing Arabidopsis lyrata plants (self‐pollinated, cross‐pollinated within the population, or cross‐pollinated between populations) and measuring offspring performance over 3 years. There were 18 source populations, each of which was either predominantly outcrossing, mixed mating, or predominantly selfing. Contrary to predictions, outcrossing populations had low inbreeding depression, which equaled that of selfing populations, challenging the central role of inbreeding depression in mating system shifts. However, plants from selfing populations showed the greatest increase in fitness when crossed with plants from other populations, reflecting higher drift load. The results support the hypothesis that extinction by mutational meltdown is why selfing hermaphroditic taxa are rare, despite their frequent appearance over evolutionary time.     

The inbreeding depression cost of selfing: Importance of flower size and population size in Collinsia parviflora (Veronicaceae)

Inbreeding depression should evolve with selfing rate when frequent inbreeding results in exposure of and selection against deleterious alleles. The selfing rate may be modified by plant traits such as flower size, or by population characteristics such as census size that can affect the probability of biparental inbreeding. Here we quantify inbreeding depression (δ) among different population sizes of Collinsia parviflora, a wildflower with interpopulation variation in flower size, by comparing fitness components and multiplicative fitness of experimentally produced selfed and outcrossed offspring. Selfed offspring had reduced multiplicative fitness compared to outcrossed offspring, but inbreeding depression was low in all combinations of population size and flower size (δ ≤ 0.05) except in large populations of large-flowered plants (δ = 0.45). The decrement to multiplicative fitness with inbreeding was not affected by population size nested within flower size, but differed between small- and large-flowered plants: small-flowered populations had lower overall inbreeding depression (δ = 0.04) compared to large-flowered populations (δ = 0.25). The difference in load with flower size suggests that either selection has removed deleterious recessive alleles or these alleles have become fixed in small-flowered, potentially more selfing populations, but that purging has not occurred to the same extent in presumably outcrossing large-flowered populations.     

Genetic causes and consequences of the breakdown of self-incompatibility: case studies in the Brassicaceae.

The genetic consequences of inbreeding is a subject that has received thorough theoretical attention and has been of interest to empirical biologists since the time of Darwin. Particularly for species with genetically controlled mechanisms to promote outcrossing (self-incompatibility or SI systems), it is expected that high levels of genetic load should accumulate through sheltering of deleterious recessive mutations. Nevertheless, transitions to selfing are common across angiosperms, which suggests that the potentially negative consequences of reduced heterozygosity and genetic diversity are balanced by other factors, such as reproductive assurance. This mini-review focuses on empirical research in the Brassicaceae to emphasize some of the genetic consequences of shifts to inbreeding in terms of mechanisms for loss of SI, changes in genetic diversity following loss of SI, and inbreeding depression in relation to outcrossing history. Despite the long history of theoretical attention, there are still some surprisingly large gaps in our understanding in each of these areas. Rather than providing a complete overview, examples are drawn predominantly from published and emerging data from Arabidopsis thaliana and its relatives to highlight recent progress and remaining questions. We are currently on the brink of major breakthroughs in understanding due both to advances in sequencing technology and a shift in focus from crop plants to natural populations, where critical factors such as population structure, phylogeography, demographic history, partial compatibility and individual variation can be taken into account when investigating the nature of the selective forces regulating mating system evolution.     

Modes and rates of selfing and associated inbreeding depression in the self-incompatible plant Senecio squalidus (Asteraceae): a successful colonizing species in the British Isles

The strength of the self-incompatibility (SI) response in Senecio squalidus was measured across its British range. Geographic variation in SI was investigated and the extent and inheritance of pseudo-self-compatibility (PSC) and inbreeding depression were determined. Mean self-fruit-set per capitulum was calculated for individuals and sample populations. The heritability of PSC and the magnitude of inbreeding depression were assessed by comparing selfing rates and fitness trait values between SI and PSC parent-progeny lines. SI was found to be strongly expressed in S. squalidus throughout its British range, with only 3.1% of the individuals sampled showing PSC. This PSC had relatively low heritability with stronger expression of SI in selfed progeny relative to PSC parents. Inbreeding depression was shown to be great in S. squalidus, with mean life history stage values ranging from 0.18 to 0.25. The strength of SI in S. squalidus appears not to have weakened in response to its rapid colonization of Britain. The avoidance of inbreeding depression is likely to be the primary factor maintaining strong SI in this successful colonizing species.     

Genetic diversity in Leavenworthia populations with different inbreeding levels.

Levels of neutral genetic diversity within and between populations were compared between outcrossing (self-incompatible) and inbreeding populations in the annual plant genus Leavenworthia. Two taxonomically independent comparisons are possible, since self-incompatibility has been lost twice in the group of species studied. Within inbred populations of L.uniflora and L.crassa, no DNA sequence variants were seen among the alleles sampled, but high diversity was seen in alleles from populations of the outcrosser L. stylosa, and in self-incompatible L. crassa populations. Diversity between populations was seen in all species. Although total diversity values were lower in the sets of inbreeding populations, between-population values were as high or higher, than those in the outcrossing taxa. Possible reasons for these diversity patterns are discussed. As the effect of inbreeding appears to be a greater than twofold reduction in diversity, we argue that some process such as selection for advantageous mutations, or against deleterious mutations, or bottlenecks occurring predominantly in the inbreeders, appears necessary to account for the findings. If selection for advantageous mutations is responsible, it appears that it must be some form of local adaptive selection, rather than substitution of alleles that are advantageous throughout the species. This is consistent with the finding of high between-population diversity in the inbreeding taxa.     

GENETIC ARCHITECTURE OF INBREEDING DEPRESSION AND THE MAINTENANCE OF GAMETOPHYTIC SELF‐INCOMPATIBILITY

Gametophytic self‐incompatibility (GSI) is a widespread genetic system, which enables hermaphroditic plants to avoid self‐fertilization and mating with close relatives. Inbreeding depression is thought to be the major force maintaining SI; however, inbreeding depression is a dynamical variable that depends in particular on the mating system. In this article we use multilocus, individual‐based simulations to examine the coevolution of SI and inbreeding depression within finite populations. We focus on the conditions for the maintenance of SI when self‐compatible (SC) mutants are introduced in the population by recurrent mutation, and compare simulation results with predictions from an analytical model treating inbreeding depression as a fixed parameter (thereby neglecting effects of purging within the SC subpopulation). In agreement with previous models, we observe that the maintenance of SI is associated with high inbreeding depression and is facilitated by high rates of self‐pollination. Purging of deleterious mutations by SC mutants has little effect on the spread of those mutants as long as most deleterious alleles have weak fitness effects: in this case, the genetic architecture of inbreeding depression has little effect on the maintenance of SI. By contrast, purging may greatly enhance the spread of SC mutants when deleterious alleles have strong fitness effects.     

The genetic basis of inbreeding depression

Data on the effects of inbreeding on fitness components are reviewed in the light of population genetic models of the possible genetic causes of inbreeding depression. Deleterious mutations probably play a major role in causing inbreeding depression. Putting together the different kinds of quantitative genetic data, it is difficult to account for the very large effects of inbreeding on fitness in Drosophila and outcrossing plants without a significant contribution from variability maintained by selection. Overdominant effects of alleles on fitness components seem not to be important in most cases. Recessive or partially recessive deleterious effects of alleles, some maintained by mutation pressure and some by balancing selection, thus seem to be the most important source of inbreeding depression. Possible experimental approaches to resolving outstanding questions are discussed.     

No inbreeding depression in an outcrossing alpine species: The breeding system of Campanula thyrsoides

Plants that live in fragmented landscapes, where populations are isolated from each other and in which long-distance dispersal is essential for colonization of empty sites, reproduction should be favoured by self-compatibility (Baker's law). Nevertheless, outcrossing mechanisms, such as self-incompatibility and dichogamy, are common in many species and are often maintained by inbreeding depression in the fitness of selfed progeny. Here, we studied the breeding system and the consequences of selfing and sister mating in Campanula thyrsoides, a short-lived perennial monocarp, which is found in the naturally fragmented landscape of the Alps. An experiment with controlled pollinations was set up in the common garden with plants grown from seeds originating from 14 seed families, collected in the siliceous Central Alps, where this plant is found on isolated carbonate bearing outcrops.Our results indicate that C. thyrsoides has a strong self-incompatibility system (SI) with no or low seed set in selfed flowers compared to outcrossed and sister-crossed flowers. Moreover, the SI system in C. thyrsoides did not break down with flower age as in some other Campanula species. Surprisingly, there was no significant difference in seed set, seed weight, germination percentage, seedling survival and size between outcrossed and sister-crossed offspring, which indicates no inbreeding depression.We suggest that the absence of inbreeding depression in this outcrossing species might be a result of frequent bottlenecks during colonization of the isolated habitats in the alpine landscape.     

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.     

Slow Recovery from Inbreeding Depression Generated by the Complex Genetic Architecture of Segregating Deleterious Mutations

The deleterious effects of inbreeding have been of extreme importance to evolutionary biology, but it has been difficult to characterize the complex interactions between genetic constraints and selection that lead to fitness loss and recovery after inbreeding. Haploid organisms and selfing organisms like the nematode Caenorhabditis elegans are capable of rapid recovery from the fixation of novel deleterious mutation; however, the potential for recovery and genomic consequences of inbreeding in diploid, outcrossing organisms are not well understood. We sought to answer two questions: 1) Can a diploid, outcrossing population recover from inbreeding via standing genetic variation and new mutation? and 2) How does allelic diversity change during recovery? We inbred C. remanei, an outcrossing relative of C. elegans, through brother-sister mating for 30 generations followed by recovery at large population size. Inbreeding reduced fitness but, surprisingly, recovery from inbreeding at large populations sizes generated only very moderate fitness recovery after 300 generations. We found that 65% of ancestral single nucleotide polymorphisms (SNPs) were fixed in the inbred population, far fewer than the theoretical expectation of ∼99%. Under recovery, 36 SNPs across 30 genes involved in alimentary, muscular, nervous, and reproductive systems changed reproducibly across replicates, indicating that strong selection for fitness recovery does exist. Our results indicate that recovery from inbreeding depression via standing genetic variation and mutation is likely to be constrained by the large number of segregating deleterious variants present in natural populations, limiting the capacity for recovery of small populations.     

Antagonism between local dispersal and self-incompatibility systems in a continuous plant population

Many self-incompatible plant species exist in continuous populations in which individuals disperse locally. Local dispersal of pollen and seeds facilitates inbreeding because pollen pools are likely to contain relatives. Self-incompatibility promotes outbreeding because relatives are likely to carry incompatible alleles. Therefore, populations can experience an antagonism between these forces. In this study, a novel computational model is used to explore the effects of this antagonism on gene flow, allelic diversity, neighbourhood sizes, and identity by descent. I confirm that this antagonism is sensitive to dispersal levels and linkage. However, the results suggest that there is little to no difference between the effects of gametophytic and sporophytic self-incompatibility systems (GSI and SSI) on unlinked loci. More importantly, both GSI and SSI affect unlinked loci in a manner similar to obligate outcrossing without mating types. This suggests that the primary evolutionary impact of self-incompatibility systems may be to prevent selfing, and prevention of biparental inbreeding might be a beneficial side-effect.     

Mitosis, stature and evolution of plant mating systems: low-Phi and high-Phi plants

There is a long-recognized association in plants between small stature and selfing, and large stature and outcrossing. Inbreeding depression is central to several hypotheses for this association, but differences in the evolutionary dynamics of inbreeding depression associated with differences in stature are rarely considered. Here, we propose and test the Phi model of plant mating system evolution, which assumes that the per-generation mutation rate of a plant is a function of the number of mitoses (Phi) that occur from zygote to gamete, and predicts fundamental differences between low-Phi (small-statured) and high-Phi (large-statured) plants in the outcomes of the joint evolution of outcrossing rate and inbreeding depression. Using a large dataset of published population genetic studies of angiosperms and conifers, we compute fitted values of inbreeding depression and deleterious mutation rates for small- and large-statured plants. Consistent with our Phi model, we find that populations of small-statured plants exhibit a range of mating systems, significantly lower mutation rates, and intermediate inbreeding depression, while large-statured plants exhibit very high mutation rates and the maximum inbreeding depression of unity. These results indicate that (i) inbred progeny typically observed in large-statured plant populations are completely lost prior to maturity in nearly all populations; (ii) evolutionary shifts from outcrossing to selfing are generally not possible in large-statured species, rather, large-statured species are more likely to evolve mating systems that avoid selfing such as self-incompatibility and dioecy; (iii) destabilization of the mating system-high selfing rate with high-inbreeding depression-might be a common occurrence in large-statured species; and (iv) large-statured species in fragmented populations might be at higher risk of extinction than previously thought. Our results help to unify and simplify a large and diverse field of research, and serve to emphasize the importance that developmental and genetic constraints play in the evolution of plant mating systems.     

The transition to social inbred mating systems in spiders: role of inbreeding tolerance in a subsocial predecessor

The social spiders are unusual among cooperatively breeding animals in being highly inbred. In contrast, most other social organisms are outbred owing to inbreeding avoidance mechanisms. The social spiders appear to originate from solitary subsocial ancestors, implying a transition from outbreeding to inbreeding mating systems. Such a transition may be constrained by inbreeding avoidance tactics or fitness loss due to inbreeding depression. We examined whether the mating system of a subsocial spider, in a genus with three social congeners, is likely to facilitate or hinder the transition to inbreeding social systems. Populations of subsocial Stegodyphus lineatus are substructured and spiders occur in patches, which may consist of kin groups. We investigated whether male mating dispersal prevents matings within kin groups in natural populations. Approximately half of the marked males that were recovered made short moves (< 5m) and mated within their natal patch. This potential for inbreeding was counterbalanced by a relatively high proportion of immigrant males. In mating experiments, we tested whether inbreeding actually results in lower offspring fitness. Two levels of inbreeding were tested: full sibling versus non-sib matings and matings of individuals within and between naturally occurring patches of spiders. Neither full siblings nor patch mates were discriminated against as mates. Sibling matings had no effect on direct fitness traits such as fecundity, hatching success, time to hatching and survival of the offspring, but negatively affected offspring growth rates and adult body size of both males and females. Neither direct nor indirect fitness measures differed significantly between within patch and between-patch pairs. We tested the relatedness between patch mates and nonpatch mates using DNA fingerprinting (TE-AFLP). Kinship explained 30% of the genetic variation among patches, confirming that patches are often composed of kin. Overall, we found limited male dispersal, lack of kin discrimination, and tolerance to low levels of inbreeding. These results suggest a history of inbreeding which may reduce the frequency of deleterious recessive alleles in the population and promote the evolution of inbreeding tolerance. It is likely that the lack of inbreeding avoidance in subsocial predecessors has facilitated the transition to regular inbreeding social systems.