The coevolution of ova defensiveness with sperm competitiveness in house mice.

Theoretical models have suggested that sperm competition can lead to increased ova resistance to fertilization. While there is some comparative evidence that this might be true, there is no experimental evidence to show that ova defensiveness evolves in response to sperm competition. We performed a series of in vitro fertilization assays to gauge the fertilizability of ova produced by female house mice from experimental populations that evolved either with or without sperm competition. Our analysis revealed that after 24 generations of experimental evolution, females that evolved under a polygamous regime produced more defensive ova than females that evolved under a monogamous regime. We therefore provide the first direct line of evidence that sperm competition can generate sexual conflict at the gametic level and lead to asymmetries in fertilization rates among populations. Our results show that females respond to sperm competition via fertilization barriers that have the potential to mediate sperm entry.

Postcopulatory sexual selection increases ATP content in rodent spermatozoa.

Sperm competition often leads to increase in sperm numbers and sperm quality, and its effects on sperm function are now beginning to emerge. Rapid swimming speeds are crucial for mammalian spermatozoa, because they need to overcome physical barriers in the female tract, reach the ovum, and generate force to penetrate its vestments. Faster velocities associate with high sperm competition levels in many taxa and may be due to increases in sperm dimensions, but they may also relate to higher adenosine triphosphate (ATP) content. We examined if variation in sperm ATP levels relates to both sperm competition and sperm swimming speed in rodents. We found that sperm competition associates with variations in sperm ATP content and sperm-size adjusted ATP concentrations, which suggests proportionally higher ATP content in response to sperm competition. Moreover, both measures were associated with sperm swimming velocities. Our findings thus support the idea that sperm competition may select for higher ATP content leading to faster sperm swimming velocity.

Heterozygosity-fitness correlations and inbreeding depression in two critically endangered mammals.

The relation among inbreeding, heterozygosity, and fitness has been studied primarily among outbred populations, and little is known about these phenomena in endangered populations. Most researchers conclude that the relation between coefficient of inbreeding estimated from pedigrees and fitness traits (inbreeding-fitness correlations) better reflects inbreeding depression than the relation between marker heterozygosity and fitness traits (heterozygosity-fitness correlations). However, it has been suggested recently that heterozygosity-fitness correlations should only be expected when inbreeding generates extensive identity disequilibrium (correlations in heterozygosity and homozygosity across loci throughout the genome). We tested this hypothesis in Mohor gazelle (Gazella dama mhorr) and Iberian lynx (Lynx pardinus). For Mohor gazelle, we calculated the inbreeding coefficient and measured heterozygosity at 17 microsatellite loci. For Iberian lynx, we measured heterozygosity at 36 microsatellite loci. In both species we estimated semen quality, a phenotypic trait directly related to fitness that is controlled by many loci and is affected by inbreeding depression. Both species showed evidence of extensive identity disequilibrium, and in both species heterozygosity was associated with semen quality. In the Iberian lynx the low proportion of normal sperm associated with low levels of heterozygosity was so extreme that it is likely to limit the fertility of males. In Mohor gazelle, although heterozygosity was associated with semen quality, inbreeding coefficient was not. This result suggests that when coefficient of inbreeding is calculated on the basis of a genealogy that begins after a long history of inbreeding, the coefficient of inbreeding fails to capture previous demographic information because it is a poor estimator of accumulated individual inbreeding. We conclude that among highly endangered species with extensive identity disequilibrium, examination of heterozygosity-fitness correlations may be an effective way to detect inbreeding depression, whereas inbreeding-fitness correlations may be poor indicators of inbreeding depression if the pedigree does not accurately reflect the history of inbreeding.

Postnatal testicular development in mouse species with different levels of sperm competition.

Postcopulatory sexual selection leads to an increase in sperm numbers which is partly the result of an increase in relative testes mass and could also be the consequence of changes in testis architecture or function. Very little is known regarding developmental changes during the first spermatogenic wave that may lead to enhanced spermatogenic efficiency and increased sperm production. We examined testicular development after birth in four mouse species with different sperm competition levels to assess changes in testicular architecture and function. Differences in relative testes mass between species appeared soon after birth and were exacerbated thereafter. The volume of testes occupied by seminiferous tubules differed between species postnatally and were associated with sperm competition levels. Finally, changes over time in the proportions of tubules with different germ cell types were also associated with sperm competition levels, with the time taken for the transition between various cell stages being negatively associated with levels of sperm competition. We conclude that postnatal testis development differs between closely related species with different sperm competition levels influencing testis architecture and the rate of progression of spermatogenesis, leading to differences in testis function at reproductive maturity.

Sperm competition differentially affects swimming velocity and size of spermatozoa from closely related muroid rodents: head first.

Sperm competition favours an increase in sperm swimming velocity that maximises the chances that sperm will reach the ova before rival sperm and fertilise. Comparative studies have shown that the increase in sperm swimming speed is associated with an increase in total sperm size. However, it is not known which are the first evolutionary steps that lead to increases in sperm swimming velocity. Using a group of closely related muroid rodents that differ in levels of sperm competition, we here test the hypothesis that subtle changes in sperm design may represent early evolutionary changes that could make sperm swim faster. Our findings show that as sperm competition increases so does sperm swimming speed. Sperm swimming velocity is associated with the size of all sperm components. However, levels of sperm competition are only related to an increase in sperm head area. Such increase is a consequence of an increase in the length of the sperm head, and also of the presence of an apical hook in some of the species studied. These findings suggest that the presence of a hook may modify the sperm head in such a way that would help sperm swim faster and may also be advantageous if sperm with larger heads are better able to attach to the epithelial cells lining the lower isthmus of the oviduct where sperm remain quiescent before the final race to reach the site of fertilisation.

Mass-specific metabolic rate and sperm competition determine sperm size in marsupial mammals.

Two complementary hypotheses have been proposed to explain variation in sperm size. The first proposes that post-copulatory sexual selection favors an increase in sperm size because it enhances sperm swimming speed, which is an important determinant of fertilization success in competitive contexts. The second hypothesis proposes that mass-specific metabolic rate acts as a constraint, because large animals with low mass-specific metabolic rates will not be able to process resources at the rates needed to produce large sperm. This constraint is expected to be particularly pronounced among mammals, given that this group contains some of the largest species on Earth. We tested these hypotheses among marsupials, a group in which mass-specific metabolic rates are roughly 30% lower than those of eutherian mammals of similar size, leading to the expectation that metabolic rate should be a major constraint. Our findings support both hypotheses because levels of sperm competition are associated with increases in sperm size, but low mass-specific metabolic rate constrains sperm size among large species. We also found that the relationship between sperm size and mass-specific metabolic rate is steeper among marsupials and shallower among eutherian mammals. This finding has two implications: marsupials respond to changes in mass-specific metabolic rate by modifying sperm length to a greater extent, suggesting that they are more constrained by metabolic rate. In addition, for any given mass-specific metabolic rate, marsupials produce longer sperm. We suggest that this is the consequence of marsupials diverting resources away from sperm numbers and into sperm size, due to their efficient sperm transport along the female tract and the existence of mechanisms to protect sperm.

Sperm competition, sperm numbers and sperm quality in muroid rodents.

Sperm competition favors increases in relative testes mass and production efficiency, and changes in sperm phenotype that result in faster swimming speeds. However, little is known about its effects on traits that contribute to determine the quality of a whole ejaculate (i.e., proportion of motile, viable, morphologically normal and acrosome intact sperm) and that are key determinants of fertilization success. Two competing hypotheses lead to alternative predictions: (a) sperm quantity and quality traits co-evolve under sperm competition because they play complementary roles in determining ejaculate's competitive ability, or (b) energetic constraints force trade-offs between traits depending on their relevance in providing a competitive advantage. We examined relationships between sperm competition levels, sperm quantity, and traits that determine ejaculate quality, in a comparative study of 18 rodent species using phylogenetically controlled analyses. Total sperm numbers were positively correlated to proportions of normal sperm, acrosome integrity and motile sperm; the latter three were also significantly related among themselves, suggesting no trade-offs between traits. In addition, testes mass corrected for body mass (i.e., relative testes mass), showed a strong association with sperm numbers, and positive significant associations with all sperm traits that determine ejaculate quality with the exception of live sperm. An "overall sperm quality" parameter obtained by principal component analysis (which explained 85% of the variance) was more strongly associated with relative testes mass than any individual quality trait. Overall sperm quality was as strongly associated with relative testes mass as sperm numbers. Thus, sperm quality traits improve under sperm competition in an integrated manner suggesting that a combination of all traits is what makes ejaculates more competitive. In evolutionary terms this implies that a complex network of genetic and developmental pathways underlying processes of sperm formation, maturation, transport in the female reproductive tract, and preparation for fertilization must all evolve in concert.

Why mammalian lineages respond differently to sexual selection: metabolic rate constrains the evolution of sperm size.

The hypothesis that sperm competition should favour increases in sperm size, because it results in faster swimming speeds, has received support from studies on many taxa, but remains contentious for mammals. We suggest that this may be because mammalian lineages respond differently to sexual selection, owing to major differences in body size, which are associated with differences in mass-specific metabolic rate. Recent evidence suggests that cellular metabolic rate also scales with body size, so that small mammals have cells that process energy and resources from the environment at a faster rate. We develop the 'metabolic rate constraint hypothesis' which proposes that low mass-specific metabolic rate among large mammals may limit their ability to respond to sexual selection by increasing sperm size, while this constraint does not exist among small mammals. Here we show that among rodents, which have high mass-specific metabolic rates, sperm size increases under sperm competition, reaching the longest sperm sizes found in eutherian mammals. By contrast, mammalian lineages with large body sizes have small sperm, and while metabolic rate (corrected for body size) influences sperm size, sperm competition levels do not. When all eutherian mammals are analysed jointly, our results suggest that as mass-specific metabolic rate increases, so does maximum sperm size. In addition, species with low mass-specific metabolic rates produce uniformly small sperm, while species with high mass-specific metabolic rates produce a wide range of sperm sizes. These findings support the hypothesis that mass-specific metabolic rates determine the budget available for sperm production: at high levels, sperm size increases in response to sexual selection, while low levels constrain the ability to respond to sexual selection by increasing sperm size. Thus, adaptive and costly traits, such as sperm size, may only evolve under sexual selection when metabolic rate does not constrain cellular budgets.

Sperm competition and the evolution of sperm design in mammals.

BACKGROUND: The influence of sperm competition upon sperm size has been a controversial issue during the last 20 years which remains unresolved for mammals. The hypothesis that, when ejaculates compete with rival males, an increase in sperm size would make sperm more competitive because it would increase sperm swimming speed, has generated contradictory results from both theoretical and empirical studies. In addition, the debate has extended to which sperm components should increase in size: the midpiece to accommodate more mitochondria and produce more energy to fuel motility, or the principal piece to generate greater propulsion forces. RESULTS: In this study we examined the influence of sperm competition upon sperm design in mammals using a much larger data set (226 species) than in previous analyses, and we corrected for phylogenetic effects by using a more complete and resolved phylogeny, and more robust phylogenetic control methods. Our results show that, as sperm competition increases, all sperm components increase in an integrated manner and sperm heads become more elongated. The increase in sperm length was found to be associated with enhanced swimming velocity, an adaptive trait under sperm competition. CONCLUSIONS: We conclude that sperm competition has played an important role in the evolution of sperm design in mammals, and discuss why previous studies have failed to detect it.

Protamine 3 shows evidence of weak, positive selection in mouse species (genus Mus)--but it is not a protamine.

Protamines are short and highly basic sperm-specific nuclear proteins that replace somatic histones during spermiogenesis in a process that is crucial for sperm formation and function. Many mammals have two protamine genes (PRM1 and PRM2) located in a gene cluster, which appears to evolve fast. Another gene in this cluster (designated protamine 3 [PRM3]) encodes a protein that is conserved among mammals but that does not seem to be involved in chromatin condensation. We have compared protein sequences and amino acid compositions of protamines in this gene cluster, searched for evidence of positive selection of PRM3, and examined whether sexual selection (sperm competition) may drive the evolution of the PRM3 gene. Nucleotide and amino acid analyses of mouse sequences revealed that PRM3 was very different from PRM1 and from both the precursor and the mature sequences of PRM2. Among 10 mouse species, PRM3 showed weak evidence of positive selection in two species, but there was no clear association with levels of sperm competition. In analyses from among mammalian species, no evidence of positive selection was found in PRM3. We conclude that PRM3 exhibits several clear differences from other protamines and, furthermore, that it cannot be regarded as a true protamine.

Sexual selection halts the relaxation of protamine 2 among rodents.

Sexual selection has been proposed as the driving force promoting the rapid evolutionary changes observed in some reproductive genes including protamines. We test this hypothesis in a group of rodents which show marked differences in the intensity of sexual selection. Levels of sperm competition were not associated with the evolutionary rates of protamine 1 but, contrary to expectations, were negatively related to the evolutionary rate of cleaved- and mature-protamine 2. Since both domains were found to be under relaxation, our findings reveal an unforeseen role of sexual selection: to halt the degree of degeneration that proteins within families may experience due to functional redundancy. The degree of relaxation of protamine 2 in this group of rodents is such that in some species it has become dysfunctional and it is not expressed in mature spermatozoa. In contrast, protamine 1 is functionally conserved but shows directed positive selection on specific sites which are functionally relevant such as DNA-anchoring domains and phosphorylation sites. We conclude that in rodents protamine 2 is under relaxation and that sexual selection removes deleterious mutations among species with high levels of sperm competition to maintain the protein functional and the spermatozoa competitive.

High levels of DNA fragmentation in spermatozoa are associated with inbreeding and poor sperm quality in endangered ungulates.

Inbreeding is known to cause deleterious effects upon reproduction and survival, but its effects upon sperm DNA integrity have not been examined. In the present study, we analyzed this relationship among three endangered ungulates: Gazella cuvieri, Gazella dama mhorr, and Gazella dorcas neglecta. In addition, we examined whether levels of sperm DNA fragmentation are associated with semen quality. The magnitude of sperm DNA damage in the two species with high levels of inbreeding (G. cuvieri and G. dama mhorr) was extremely high when compared to the species with low levels of inbreeding (G. dorcas neglecta) and to values previously reported for outbred populations. Levels of sperm DNA fragmentation significantly increased with inbreeding and age. Increased DNA damage in sperm was associated with increased sperm head abnormalities, lower percentage of sperm with an intact acrosome, and poor motility. Our findings suggest that the link between inbreeding and semen quality is mediated by the effects of inbreeding upon sperm DNA damage. The deleterious effects of inbreeding upon the paternal genome likely decrease male fertility and may cause genetic damage to future generations. Because inbreeding is common among endangered species, high levels of sperm DNA damage may have considerable impact upon the viability of their populations.

Paternal levels of DNA damage in spermatozoa and maternal parity influence offspring mortality in an endangered ungulate.

Understanding which factors influence offspring mortality rates is a major challenge since it influences population dynamics and may constrain the chances of recovery among endangered species. Most studies have focused on the effects of maternal and environmental factors, but little is known about paternal factors. Among most polygynous mammals, males only contribute the haploid genome to their offspring, but the possibility that sperm DNA integrity may influence offspring survival has not been explored. We examined several maternal, paternal and individual factors that may influence offspring survival in an endangered species (Gazella cuvieri). Levels of sperm DNA damage had the largest impact upon offspring mortality rates, followed by maternal parity. In addition, there was a significant interaction between these two variables, so that offspring born to primiparous mothers were more likely to die if their father had high levels of sperm DNA damage, but this was not the case among multiparous mothers. Thus, multiparous mothers seem to protect their offspring from the deleterious effects of sperm DNA damage. Since levels of sperm DNA damage seem to be higher among endangered species, more attention should be paid to the impact of this largely ignored factor among the viability of endangered species.

Reproductive traits in captive and free-ranging males of the critically endangered Iberian lynx (Lynx pardinus).

The Iberian lynx (Lynx pardinus) is the most endangered felid in the world. Adequate genetic management of in situ and ex situ populations, and linkage between both, require knowledge on male reproductive biology and factors influencing it. We examined the influence of age, free-ranging versus captive conditions and seasonality on phenotypic, endocrine and semen traits, and links between reproductive traits and male fertility. Males had relatively small testes, produced low sperm numbers, a low proportion of normal sperm, and a high proportion of motile sperm. Young (2-year-old) males had lower testosterone levels, fewer sperm, and a lower proportion of motile and normal sperm than > or =4-year-old males. No major differences were found in semen traits before and after the mating season or between free-ranging and captive males, although the latter had better sperm motility. Males with larger relative testes weight and more sperm copulated more frequently, whereas males that produced more sperm with higher motility produced more cubs per female. In conclusion, small relative testes size and low sperm quality could indicate either low levels of sperm competition or high levels of inbreeding. Young males are probably subfertile; there is a slight trend for males in the captive breeding programme to have better semen quality than wild males, and males with higher sperm production are sexually more active and more fertile. These findings have major implications for decisions regarding which males should breed, provide samples for the genetic resource bank, or participate in programmes involving the use of assisted reproductive techniques.

The effect of glucocorticoids on mouse oocyte in vitro maturation and subsequent fertilization and embryo development.

Increased glucocorticoid levels, due to medical therapy or stress-related, may affect reproduction via the hypothalamus-pituitary-axis or directly at the oocyte level. We examined the effects of natural (corticosterone) or synthetic (dexamethasone) glucocorticoids on mouse oocyte maturation and underlying changes in extracellular signal-regulated kinase (ERK) phosphorylation patterns. Fertilization and progression up to the blastocyst stage were also evaluated. Oocytes were exposed to corticosterone or dexamethasone (0, 0.25, 2.5, 25 or 250microM) for 17h during in vitro maturation. After maturation, ERK-1/2 activation in oocytes was assessed by SDS-PAGE and immunoblotting, and fertilization and developmental capacity were examined in vitro. Corticosterone exposure during oocyte maturation significantly decreased progression to metaphase II, fertilization and embryo development at the highest concentration. Corticosterone caused a concentration-dependent inhibition of ERK-1/2 activation, with the highest concentration resulting in considerable inhibition of oocyte ERK-1/2 phosphorylation and no blastocyst development. In contrast, dexamethasone had no effect on maturation, fertilization and cleavage, and no effect was seen on ERK-1/2 phosphorylation. Based on these in vitro findings, high glucocorticoid levels may have consequences for subsequent development, although a short exposure to physiologic or stress-related glucocorticoid levels may not represent a hazard to meiosis progression of the oocyte.

The effect of glucocorticoids on ERK-1/2 phosphorylation during maturation of lamb oocytes and their subsequent fertilization and cleavage ability in vitro.

High levels of glucocorticoids may alter reproduction, but little is known about their direct actions on oocyte maturation, fertilization and subsequent development. Earlier work suggested negative effects of cortisol or dexamethasone on oocyte maturation but differences were noted between animal models. Both glucocorticoids reduce the p34(cdc2)-cyclin B1 complex but it is unknown if other signaling pathways important for meiosis progression are affected. In this study, using sheep oocytes as a model system, we assessed in vitro the effects of increasing concentration of glucocorticoids (0-250 microM) on oocyte maturation and underlying changes in the MAP kinase pathway, and the ability of oocytes to undergo fertilization and embryo development. Cortisol decreased oocyte maturation but only at the highest concentration, whereas dexamethasone had no effect. Fertilization and cleavage were not affected. On the other hand, both cortisol and dexamethasone inhibited ERK-1/2 activation in a concentration-dependent manner. It thus seems that oocytes can overcome deleterious effects of glucocorticoids during maturation despite the decrease in ERK-1/2 activity, but repercussions in vivo should be further explored.

Assessment of semen quality, sperm cryopreservation and heterologous IVF in the critically endangered Iberian lynx (Lynx pardinus).

Semen traits and factors affecting sperm cryopreservation were assessed in the Iberian lynx (Lynx pardinus), a species regarded as the most endangered felid in the world. For cryopreservation, semen was washed, resuspended in a Tes-Tris-based diluent (TEST) or a Tris-based diluent (Biladyl), both with 20% egg yolk and 4% glycerol, loaded into straws, cooled to 5 degrees C using an automated programmable system and frozen on nitrogen vapour. Heterologous IVF of in vitro-matured domestic cat oocytes was used to test the fertilising ability of cryopreserved spermatozoa. Electroejaculates from five males were obtained. Characterisation of the electroejaculates revealed mean (+/- s.e.m.) values of 3.3 +/- 0.6 x 10(6) total spermatozoa, 73.6 +/- 4.6% motile spermatozoa, 23.7 +/- 4.0% morphologically normal spermatozoa and 40.7 +/- 2.3% spermatozoa with intact acrosomes. After thawing a higher percentage of motile spermatozoa was seen in TEST than in Biladyl (34.0 +/- 6.2% v. 7.5 +/- 4.8%, respectively; P < 0.05); however, there were no differences in the percentage of intact acrosomes between the two diluents. Iberian lynx spermatozoa fertilised domestic cat oocytes in vitro, with higher fertilisation rates observed for spermatozoa cryopreserved in TEST than in Biladyl, although the difference did not reach statistical significance (20.5 +/- 4.5% v. 11.5 +/- 6.8%, respectively). There were positive significant relations between the fertilisation rates and both the percentage of normal spermatozoa and the percentage of spermatozoa with an intact acrosome before cryopreservation (P = 0.04). This first report of the collection and cryopreservation of Iberian lynx semen and analysis of fertilising ability is an important step in the development of assisted reproductive techniques for this critically endangered felid species.

Sperm competition and reproductive mode influence sperm dimensions and structure among snakes.

The role of sperm competition in increasing sperm length is a controversial issue, because findings from different taxa seem contradictory. We present a comparative study of 25 species of snakes with different levels of sperm competition to test whether it influences the size and structure of different sperm components. We show that, as levels of sperm competition increase, so does sperm length, and that this elongation is largely explained by increases in midpiece length. In snakes, the midpiece is comparatively large and it contains structures, which in other taxa are present in the rest of the flagellum, suggesting that it may integrate some of its functions. Thus, increases in sperm midpiece size would result in more energy as well as greater propulsion force. Sperm competition also increases the area occupied by the fibrous sheath and outer dense fibers within the sperm midpiece, revealing for the first time an effect upon structural elements within the sperm. Finally, differences in male-male encounter rates between oviparous and viviparous species seem to lead to differences in levels of sperm competition. We conclude that the influence of sperm competition upon different sperm components varies between taxa, because their structure and function is different.

Sexual selection drives weak positive selection in protamine genes and high promoter divergence, enhancing sperm competitiveness.

Phenotypic adaptations may be the result of changes in gene structure or gene regulation, but little is known about the evolution of gene expression. In addition, it is unclear whether the same selective forces may operate at both levels simultaneously. Reproductive proteins evolve rapidly, but the underlying selective forces promoting such rapid changes are still a matter of debate. In particular, the role of sexual selection in driving positive selection among reproductive proteins remains controversial, whereas its potential influence on changes in promoter regions has not been explored. Protamines are responsible for maintaining DNA in a compacted form in chromosomes in sperm and the available evidence suggests that they evolve rapidly. Because protamines condense DNA within the sperm nucleus, they influence sperm head shape. Here, we examine the influence of sperm competition upon protamine 1 and protamine 2 genes and their promoters, by comparing closely related species of Mus that differ in relative testes size, a reliable indicator of levels of sperm competition. We find evidence of positive selection in the protamine 2 gene in the species with the highest inferred levels of sperm competition. In addition, sperm competition levels across all species are strongly associated with high divergence in protamine 2 promoters that, in turn, are associated with sperm swimming speed. We suggest that changes in protamine 2 promoters are likely to enhance sperm swimming speed by making sperm heads more hydrodynamic. Such phenotypic changes are adaptive because sperm swimming speed may be a major determinant of fertilization success under sperm competition. Thus, when species have diverged recently, few changes in gene-coding sequences are found, while high divergence in promoters seems to be associated with the intensity of sexual selection.

Pedigrees and microsatellites among endangered ungulates: what do they tell us?

Relationships between pedigree coefficients of inbreeding and molecular metrics are generally weak, suggesting that measures of heterozygosity estimated using microsatellites may be poor surrogates of genome-wide inbreeding. We compare three endangered species of gazelles (Gazella) with different degrees of threat in their natural habitats, for which captive breeding programmes exist. For G. dorcas, the species with the largest founding population, the highest and most recent number of founding events, the correlation between pedigree coefficient of inbreeding and molecular metrics was higher than for outbred populations of mammals, probably because it has both higher mean f and variance. For the two species with smaller founding populations, conventional assumptions about founders, i.e. outbred and unrelated, are unrealistic. When realistic assumptions about the founders were made, clear relationships between pedigree coefficients of inbreeding and molecular metrics were revealed for G. cuvieri. This population had a small founding population, but it did experience admixture years later; thus, the relationship between inbreeding and molecular metrics in G. cuvieri is very similar to the expected values but lower than in G. dorcas. In contrast, no relationship was found for G. dama mhorr which had a much smaller founding population than had been previously assumed, which probably had high levels of inbreeding and low levels of genetic variability, and no admixture. In conclusion, the strength of the association between pedigree coefficient of inbreeding and molecular metrics among endangered species depends on the level of inbreeding and genetic variability present in the founding population, its size and its history.