Effects of female-specific selection for reproductive investment on male fertility traits.

Despite sharing an autosomal genome, the often divergent reproductive strategies of males and females cause the selection to act in a sex-specific manner. Selection acting on one sex can have negative, positive, or neutral fitness consequences on the opposite sex. Here, we test how female-limited selection on reproductive investment in Japanese quail (Coturnix japonica) affects male fertility-related traits. Despite there being no difference in the size of males' testes from lines selected for high female reproductive investment (H-line) or low female reproductive investment (L-line), in both lines, the left testis had a greater volume of sperm-producing tissue. Since H-line females have a larger left-side restricted oviduct, this suggests a positive genetic correlation between male and female gonad function and that internal testis structure is a target of sexual selection. However, despite H-line males having previously been found to have greater fertilization success in a competitive scenario, we found little evidence of a difference between the lines in sperm number, motility, velocity, length, or the number of sperm that reached the ova. Precopulatory cues and/or the role of seminal fluid in sperm motility may thus be more likely to contribute to the H-line male fertilization advantage in this species.

Inbreeding accelerates reproductive senescence, but not survival senescence, in a precocial bird.

Inbreeding depression is predicted to increase with age, because natural selection is less efficient at purging deleterious alleles that are only expressed later in life. However, empirical results are scarce, and equivocal between studies. Here we performed controlled matings between related and unrelated individuals of domesticated Japanese quail (Coturnix japonica), and monitored the performance of their offspring for all fitness components over their complete life course. We found rapid senescence in adult survival and egg laying performance, and inbreeding depression at all life stages (reduced embryo viability, increased age at maturity, as well as reduced adult survival and reproduction). Inbreeding depression did not increase at later ages for survival, but did so for egg laying, thereby accelerating reproductive senescence. Moreover, the effect of inbreeding on egg laying persisted after correcting for lifespan, indicating that both survival and reproduction were independently affected by inbreeding. We suggest that in heterogeneous populations intra-generational purging may at earlier ages already select out the individuals that are homozygous for the specific alleles responsible for depressed survival, preventing the appearance of increased inbreeding depression in survival with age. Given that inbreeding affects reproduction independent of survival this should not apply to reproductive senescence or homogeneous populations, which may explain equivocal results between traits and studies.

Increased male-induced harm in response to female-limited selection: interactive effects between intra- and interlocus sexual conflict?

Interlocus sexual conflict (IRSC) occurs because of shared interactions that have opposite effects on male and female fitness. Typically, it is assumed that loci involved in IRSC have sex-limited expression and are thus not directly affected by selective pressures acting on the other sex. However, if loci involved in IRSC have pleiotropic effects in the other sex, intersexual selection can shape the evolutionary dynamics of conflict escalation and resolution, as well as the evolution of reproductive traits linked to IRSC loci, and vice versa. Here we used an artificial selection approach in Japanese quail (Coturnix japonica) to test if female-limited selection on reproductive investment affects the amount of harm caused by males during mating. We found that males originating from lines selected for high female reproductive investment caused more oxidative damage in the female reproductive tract than males originating from lines selected for low female reproductive investment. This male-induced damage was specific to the oviduct and not found in other female tissues, suggesting that it was ejaculate-mediated. Our results suggest that intersexual selection shapes the evolution of IRSC and that male-induced harm may contribute to the maintenance of variation in female reproductive investment.

Reproductive Strategies Affect Telomere Dynamics across the Life Course.

AbstractBecause parental care has a heritable basis, the benefits of receiving increased parental provisioning early in life are genetically linked to the costs of providing increased parental provisioning at adulthood. Reproductive strategies thus result in distinct cost-benefit syndromes across the life course that may shape individual health and aging trajectories. Here we used an artificial selection approach in Japanese quail (Coturnix japonica) to test how reproductive strategies affect telomere length, a biomarker of somatic state, at different life stages. We show that males but not females from lines selected for low maternal investment (i.e., developing in a relatively small egg) had shorter telomeres at birth. These patterns were still weakly present at the end of the juvenile growth period. In contrast, significantly shorter telomeres were found in reproductively active adult birds from the high-investment lines, suggesting that telomere attrition was accelerated in these individuals once they had become reproductively active. Our study shows that reproductive strategies differentially affect telomere dynamics across the life course, highlighting the role of cross-generational constraints in shaping individual aging trajectories.

Maternally transferred thyroid hormones and life-history variation in birds.

In vertebrates, thyroid hormones (THs) play an important role in the regulation of growth, development, metabolism, photoperiodic responses and migration. Maternally transferred THs are important for normal early phase embryonic development when embryos are not able to produce endogenous THs. Previous studies have shown that variation in maternal THs within the physiological range can influence offspring phenotype. Given the essential functions of maternal THs in development and metabolism, THs may be a mediator of life-history variation across species. We tested the hypothesis that differences in life histories are associated with differences in maternal TH transfer across species. Using birds as a model, we specifically tested whether maternally transferred yolk THs covary with migratory status, developmental mode and traits related to pace-of-life (e.g. basal metabolic rate, maximum life span). We collected un-incubated eggs (n = 1-21 eggs per species, median = 7) from 34 wild and captive bird species across 17 families and six orders to measure yolk THs [both triiodothyronine (T3) and thyroxine (T4)], compiled life-history trait data from the literature and used Bayesian phylogenetic mixed models to test our hypotheses. Our models indicated that both concentrations and total amounts of the two main forms of THs (T3 and T4) were higher in the eggs of migratory species compared to resident species, and total amounts were higher in the eggs of precocial species, which have longer prenatal developmental periods, than in those of altricial species. However, maternal yolk THs did not show clear associations with pace-of-life-related traits, such as fecundity, basal metabolic rate or maximum life span. We quantified interspecific variation in maternal yolk THs in birds, and our findings suggest higher maternal TH transfer is associated with the precocial mode of development and migratory status. Whether maternal THs represent a part of the mechanism underlying the evolution of precocial development and migration or a consequence of such life histories is currently unclear. We therefore encourage further studies to explore the physiological mechanisms and evolutionary processes underlying these patterns.

Elevational Changes in Bacterial Microbiota Structure and Diversity in an Arthropod-Disease Vector.

Environmental conditions change rapidly along elevational gradients and have been found to affect community composition in macroscopic taxa, with lower diversity typically observed at higher elevations. In contrast, microbial community responses to elevation are still poorly understood. Specifically, the effects of elevation on vector-associated microbiota have not been studied to date, even though the within-vector microbial community is known to influence vector competence for a range of zoonotic pathogens. Here we characterize the structure and diversity of the bacterial microbiota in an important zoonotic disease vector, the sheep tick Ixodes ricinus, along replicated elevational gradient (630-1673 m) in the Swiss Alps. 16S rRNA sequencing of the whole within-tick bacterial microbiota of questing nymphs and adults revealed a decrease in Faith's phylogenetic microbial alpha diversity with increasing elevation, while beta diversity analyses revealed a lower variation in microbial community composition at higher elevations. We also found a higher microbial diversity later in the season and significant differences in microbial diversity among tick life stages and sexes, with lowest microbial alpha diversity observed in adult females. No associations between tick genetic diversity and bacterial diversity were observed. Our study demonstrates systematic changes in tick bacterial microbiota diversity along elevational gradients. The observed patterns mirror diversity changes along elevational gradients typically observed in macroscopic taxa, and they highlight the key role of environmental factors in shaping within-host microbial communities in ectotherms.

Sex-specific effects of experimental ectoparasite infestation on telomere length in great tit nestlings.

Telomere length is a biomarker of biological ageing and lifespan in various vertebrate taxa. Evidence is accumulating that telomeres shorten more rapidly when an individual is exposed to environmental stressors. Parasites are potent selective agents that can cause physiological stress directly or indirectly through the activation of the host's immune system. Yet to date, empirical evidence for a role of parasites in telomere dynamics in natural populations is limited. Here, we show experimentally that exposure to ectoparasitic hen fleas (Ceratophyllus gallinae) during growth results in shorter telomeres in female, but not male, great tit (Parus major) nestlings. Females had longer telomeres than males when growing up in experimentally deparasitized nests but, likely because of the sex-specific effects of ectoparasitism on telomere length, this sexual dimorphism was absent in birds growing up in experimentally infested nests. Our results provide the first experimental evidence for a role of ectoparasitism in telomere dynamics in a natural vertebrate population, and suggest that the costs of infection manifest in sex-specific ways.

Combining genome-wide association study and FST -based approaches to identify targets of Borrelia-mediated selection in natural rodent hosts.

Recent advances in high-throughput sequencing technologies provide opportunities to gain novel insights into the genetic basis of phenotypic trait variation. Yet to date, progress in our understanding of genotype-phenotype associations in nonmodel organisms in general and natural vertebrate populations in particular has been hampered by small sample sizes typically available for wildlife populations and a resulting lack of statistical power, as well as a limited ability to control for false-positive signals. Here we propose to combine a genome-wide association study (GWAS) and FST -based approach with population-level replication to partly overcome these limitations. We present a case study in which we used this approach in combination with genotyping-by-sequencing (GBS) single nucleotide polymorphism (SNP) data to identify genomic regions associated with Borrelia afzelii resistance or susceptibility in the natural rodent host of this Lyme disease-causing spirochete, the bank vole (Myodes glareolus). Using this combined approach we identified four consensus SNPs located in exonic regions of the genes Slc26a4, Tns3, Wscd1 and Espnl, which were significantly associated with the voles' Borrelia infectious status within and across populations. Functional links between host responses to bacterial infections and most of these genes have previously been demonstrated in other rodent systems, making them promising new candidates for the study of evolutionary host responses to Borrelia emergence. Our approach is applicable to other systems and may facilitate the identification of genetic variants underlying disease resistance or susceptibility, as well as other ecologically relevant traits, in wildlife populations.

Selection for Divergent Reproductive Investment Affects Neuron Size and Foliation in the Cerebellum.

The cerebellum has a highly conserved internal circuitry, but varies greatly in size and morphology within and across species. Despite this variation, the underlying volumetric changes among the layers of the cerebellar cortex or their association with Purkinje cell numbers and sizes is poorly understood. Here, we examine intraspecific scaling relationships and variation in the quantitative neuroanatomy of the cerebellum in Japanese quail (Coturnix japonica) selected for high or low reproductive investment. As predicted by the circuitry of the cerebellum, the volumes of the constituent layers of the cerebellar cortex were strongly and positively correlated with one another and with total cerebellar volume. The number of Purkinje cells also significantly and positively co-varied with total cerebellar volume and the molecular layer, but not the granule cell layer or white matter volumes. Purkinje cell size and cerebellar foliation did not significantly covary with any cerebellar measures, but differed significantly between the selection lines. Males and females from the high-investment lines had smaller Purkinje cells than males and females from the low-investment lines and males from the high-investment lines had less folded cerebella than quail from the low-investment lines. These results suggest that within species, the layers of the cerebellum increase in a coordinated fashion, but Purkinje cell size and cerebellar foliation do not increase proportionally with overall cerebellum size. In contrast, selection for differential reproductive investment affects Purkinje cell size and cerebellar foliation, but not other quantitative measures of cerebellar anatomy.

Small-scale spatial variation in infection risk shapes the evolution of a Borrelia resistance gene in wild rodents.

Spatial variation in pathogen-mediated selection is predicted to influence the evolutionary trajectory of host populations and lead to spatial variation in their immunogenetic composition. However, to date few studies have been able to directly link small-scale spatial variation in infection risk to host immune gene evolution in natural, nonhuman populations. Here, we use a natural rodent-Borrelia system to test for associations between landscape-level spatial variation in Borrelia infection risk along replicated elevational gradients in the Swiss Alps and Toll-like receptor 2 (TLR2) evolution, a candidate gene for Borrelia resistance, across bank vole (Myodes glareolus) populations. We found that Borrelia infection risk (i.e., the product of Borrelia prevalence in questing ticks and the average tick load of voles at a sampling site) was spatially variable and significantly negatively associated with elevation. Across sampling sites, Borrelia prevalence in bank voles was significantly positively associated with Borrelia infection risk along the elevational clines. We observed a significant association between naturally occurring TLR2 polymorphisms in hosts and their Borrelia infection status. The TLR2 variant associated with a reduced likelihood of Borrelia infection was most common in rodent populations at lower elevations that face a high Borrelia infection risk, and its frequency changed in accordance with the change in Borrelia infection risk along the elevational clines. These results suggest that small-scale spatial variation in parasite-mediated selection affects the immunogenetic composition of natural host populations, providing a striking example that the microbial environment shapes the evolution of the host's immune system in the wild.

In ovo yolk carotenoid and testosterone levels interactively influence female transfer of yolk antioxidants to her eggs.

Mothers can influence prenatal conditions by varying the amount of nutrients, hormones or antioxidants they provide to their developing young. Some of these substances even affect the transfer of these compounds in the next generation, but it is less clear how different maternally transmitted compounds interact with each other to shape reproductive resource allocation in their offspring. Here, we found that female Japanese quails (Coturnix japonica) that were exposed to high carotenoid levels during embryonic development transferred lower concentrations of yolk antioxidants to their own eggs later in life. This effect disappeared when both testosterone and carotenoid concentrations were manipulated simultaneously, showing long-term and interactive effects of these maternally derived egg components on a female's own egg composition. Given that exposure to high levels of testosterone during embryo development stimulates the production of reactive oxygen species (ROS) and impairs antioxidant defenses, we propose that carotenoids act as in ovo antioxidants in an oxidatively stressful environment (i.e. when levels of testosterone are high) but might have prooxidant properties in an environment where they are not used to counteract an increased production of ROS. In line with this hypothesis, we previously showed that prenatal exposure to increased concentrations of yolk carotenoids leads to a rise of oxidative damage at adulthood, but only when yolk testosterone concentrations were not experimentally increased as well. As a consequence, antioxidants in the body may be used to limit oxidative damage in females exposed to high levels of carotenoids during development (but not in females exposed to increased levels of both carotenoids and testosterone), resulting in lower amounts of antioxidants being available for deposition into eggs. Since prenatal antioxidant exposure is known to influence fitness-related traits, the effect detected in this study might have transgenerational consequences.

Increased prenatal maternal investment reduces inbreeding depression in offspring.

Inbreeding depression refers to the reduction of fitness that results from matings between relatives. Evidence for reduced fitness in inbred individuals is widespread, but the strength of inbreeding depression varies widely both within and among taxa. Environmental conditions can mediate this variation in the strength of inbreeding depression, with environmental stress exacerbating the negative consequences of inbreeding. Parents can modify the environment experienced by offspring, and have thus the potential to mitigate the negative consequences of inbreeding. While such parental effects have recently been demonstrated during the postnatal period, the role of prenatal parental effects in influencing the expression of inbreeding depression remains unexplored. To address this gap, we performed matings between full-sibs or unrelated individuals in replicated lines of Japanese quail (Coturnix japonica) experimentally selected for high and low maternal egg provisioning. We show that in the low maternal investment lines hatching success was strongly reduced when parents were related. In the high maternal investment lines, however, this negative effect of inbreeding on hatching success was absent, demonstrating that prenatal maternal provisioning can alleviate the negative fitness consequences of inbreeding.

Disentangling Genetic and Prenatal Maternal Effects on Offspring Size and Survival.

Organizational processes during prenatal development can have long-term effects on an individual's phenotype. Because these early developmental stages are sensitive to environmental influences, mothers are in a unique position to alter their offspring's phenotype by differentially allocating resources to their developing young. However, such prenatal maternal effects are difficult to disentangle from other forms of parental care, additive genetic effects, and/or other forms of maternal inheritance, hampering our understanding of their evolutionary consequences. Here we used divergent selection lines for high and low prenatal maternal investment and their reciprocal line crosses in a precocial bird-the Japanese quail (Coturnix japonica)-to quantify the relative importance of genes and prenatal maternal effects in shaping offspring phenotype. Maternal but not paternal origin strongly affected offspring body size and survival throughout development. Although the effects of maternal egg investment faded over time, they were large at key life stages. Additionally, there was evidence for other forms of maternal inheritance affecting offspring phenotype at later stages of development. Our study is among the first to successfully disentangle prenatal maternal effects from all other sources of confounding variation and highlights the important role of prenatal maternal provisioning in shaping offspring traits closely linked to fitness.

Matrilineal inheritance of a key mediator of prenatal maternal effects.

Sex-linkage is predicted to evolve in response to sex-specific or sexually antagonistic selection. In line with this prediction, most sex-linked genes are associated with reproduction in the respective sex. In addition to traits directly involved in fertility and fecundity, mediators of maternal effects may be predisposed to evolve sex-linkage, because they indirectly affect female fitness through their effect on offspring phenotype. Here, we test for sex-linked inheritance of a key mediator of prenatal maternal effects in oviparous species, the transfer of maternally derived testosterone to the eggs. Consistent with maternal inheritance, we found that in Japanese quail (Coturnix japonica) granddaughters resemble their maternal (but not their paternal) grandmother in yolk testosterone deposition. This pattern of resemblance was not due to non-genetic priming effects of testosterone exposure during prenatal development, as an experimental manipulation of yolk testosterone levels did not affect the females' testosterone transfer to their own eggs later in life. Instead, W chromosome and/or mitochondrial variation may underlie the observed matrilineal inheritance pattern. Ultimately, the inheritance of mediators of maternal effects along the maternal line will allow for a fast and direct response to female-specific selection, thereby affecting the dynamics of evolutionary processes mediated by maternal effects.

Artificial selection reveals the energetic expense of producing larger eggs.

BACKGROUND: The amount of resources provided by the mother before birth has important and long-lasting effects on offspring fitness. Despite this, there is a large amount of variation in maternal investment seen in natural populations. Life-history theory predicts that this variation is maintained through a trade-off between the benefits of high maternal investment for the offspring and the costs of high investment for the mother. However, the proximate mechanisms underlying these costs of reproduction are not well understood. Here we used artificial selection for high and low maternal egg investment in a precocial bird, the Japanese quail (Coturnix japonica) to quantify costs of maternal reproductive investment. RESULTS: We show that females from the high maternal investment lines had significantly larger reproductive organs, which explained their overall larger body mass, and resulted in a higher resting metabolic rate (RMR). Contrary to our expectations, this increase in metabolic activity did not lead to a higher level of oxidative damage. CONCLUSIONS: This study is the first to provide experimental evidence for metabolic costs of increased per offspring investment.

Long-term effect of yolk carotenoid levels on testis size in a precocial bird.

Conditions experienced during prenatal development can have long-lasting organizational effects on offspring. Maternal carotenoids deposited in the eggs of birds and other oviparous species play an important role during fast embryonic growth and chick development through their antioxidant properties. However, the long-term consequences of variation in maternal carotenoid transfer for the offspring have seldom been considered. Since plasma carotenoid levels at adulthood are known to influence testis size and yolk carotenoid levels influence the ability to extract carotenoids later in life, we hypothesized that maternally transmitted carotenoids might influence gonad size at adulthood. Here, we showed that male Japanese quail (Coturnix japonica) originating from a carotenoid-enriched egg had smaller testes than control individuals at adulthood. This result shows that yolk carotenoids have long-term organizational effects. In addition, given that carotenoid intake at sexual maturity increases sperm quality and that a decreased testis size is associated with a lower sperm production, we propose that carotenoid exposure during embryo development might influence a trade-off between ejaculate size and sperm quality.

A trade-off between reproductive investment and maternal cerebellum size in a precocial bird.

Natural selection favours increased investment in reproduction, yet considerable variation in parental investment is observed in natural populations. Life-history theory predicts that this variation is maintained by a trade-off between the benefits of increased reproductive investment and its associated costs for the parents. The nature of these costs of reproduction, however, remains poorly understood. The brain is an energetically highly expensive organ and increased reproductive investment may, therefore, negatively affect brain maintenance. Using artificial selection lines for high and low prenatal maternal investment in a precocial bird, the Japanese quail (Coturnix japonica), we provide experimental evidence for this hypothesis by showing that increased prenatal provisioning negatively affects the size of a particular brain region of the mother, the cerebellum. Our finding suggests that cognitive demands may constrain the evolution of parental investment, and vice versa, contributing to the maintenance of variation in reproductive behaviour in animal populations.

Borrelia burgdorferi sensu lato infection pressure shapes innate immune gene evolution in natural rodent populations across Europe.

Although parasite-mediated selection is assumed to be the main driver of immune gene evolution, empirical evidence that parasites induce allele frequency changes at host immune genes in time and/or space remains scarce. Here, I show that the frequency of a protective gene variant of the innate immune receptor Toll-like receptor 2 in natural bank vole (Myodes glareolus) populations is positively associated with the strength of Borrelia burgdorferi sensu lato infection risk across the European continent. Thereby, this study provides rare evidence for the role of spatially variable infection pressures in moulding the vertebrate immune system.

Natural selection acts in opposite ways on correlated hormonal mediators of prenatal maternal effects in a wild bird population.

Maternal hormones are important mediators of prenatal maternal effects. Although many experimental studies have demonstrated their potency in shaping offspring phenotypes, we know remarkably little about their adaptive value. Using long-term data on a wild collared flycatcher (Ficedula albicollis) population, we show that natural selection acts in opposite ways on two maternally derived androgens, yolk androstenedione (A4) and yolk testosterone (T). High yolk A4 concentrations are associated with higher fitness, whereas high yolk T concentrations are associated with lower fitness. Natural selection thus favours females that produce eggs with high A4 and low T concentrations. Importantly, however, there exists a positive (non-genetic) correlation between A4 and T, which suggests that females are limited in their ability to reach this adaptive optimum. Thereby, these results provide strong evidence for an adaptive value of differential maternal androgen deposition, and a mechanistic explanation for the maintenance of variation in maternal investment in the wild.

The multivariate egg: quantifying within- and among-clutch correlations between maternally derived yolk immunoglobulins and yolk androgens using multivariate mixed models.

Egg components are important mediators of prenatal maternal effects in birds and other oviparous species. Because different egg components can have opposite effects on offspring phenotype, selection is expected to favour their mutual adjustment, resulting in a significant covariation between egg components within and/or among clutches. Here we tested for such correlations between maternally derived yolk immunoglobulins and yolk androgens in great tit (Parus major) eggs using a multivariate mixed-model approach. We found no association between yolk immunoglobulins and yolk androgens within clutches, indicating that within clutches the two egg components are deposited independently. Across clutches, however, there was a significant negative relationship between yolk immunoglobulins and yolk androgens, suggesting that selection has co-adjusted their deposition. Furthermore, an experimental manipulation of ectoparasite load affected patterns of covariance among egg components. Yolk immunoglobulins are known to play an important role in nestling immune defence shortly after hatching, whereas yolk androgens, although having growth-enhancing effects under many environmental conditions, can be immunosuppressive. We therefore speculate that variation in the risk of parasitism may play an important role in shaping optimal egg composition and may lead to the observed pattern of yolk immunoglobulin and yolk androgen deposition across clutches. More generally, our case study exemplifies how multivariate mixed-model methodology presents a flexible tool to not only quantify, but also test patterns of (co)variation across different organisational levels and environments, allowing for powerful hypothesis testing in ecophysiology.