Taxonomic Sampling and Rare Genomic Changes Overcome Long-Branch Attraction in the Phylogenetic Placement of Pseudoscorpions.

Long-branch attraction is a systematic artifact that results in erroneous groupings of fast-evolving taxa. The combination of short, deep internodes in tandem with long-branch attraction artifacts has produced empirically intractable parts of the Tree of Life. One such group is the arthropod subphylum Chelicerata, whose backbone phylogeny has remained unstable despite improvements in phylogenetic methods and genome-scale data sets. Pseudoscorpion placement is particularly variable across data sets and analytical frameworks, with this group either clustering with other long-branch orders or with Arachnopulmonata (scorpions and tetrapulmonates). To surmount long-branch attraction, we investigated the effect of taxonomic sampling via sequential deletion of basally branching pseudoscorpion superfamilies, as well as varying gene occupancy thresholds in supermatrices. We show that concatenated supermatrices and coalescent-based summary species tree approaches support a sister group relationship of pseudoscorpions and scorpions, when more of the basally branching taxa are sampled. Matrix completeness had demonstrably less influence on tree topology. As an external arbiter of phylogenetic placement, we leveraged the recent discovery of an ancient genome duplication in the common ancestor of Arachnopulmonata as a litmus test for competing hypotheses of pseudoscorpion relationships. We generated a high-quality developmental transcriptome and the first genome for pseudoscorpions to assess the incidence of arachnopulmonate-specific duplications (e.g., homeobox genes and miRNAs). Our results support the inclusion of pseudoscorpions in Arachnopulmonata (new definition), as the sister group of scorpions. Panscorpiones (new name) is proposed for the clade uniting Scorpiones and Pseudoscorpiones.

Sperm competitive advantage of a rare mitochondrial haplogroup linked to differential expression of mitochondrial oxidative phosphorylation genes.

Maternal inheritance of mitochondria creates a sex-specific selective sieve through which mitochondrial mutations harmful to males but not females accumulate and contribute to sexual differences in longevity and disease susceptibility. Because eggs and sperm are under disruptive selection, sperm are predicted to be particularly vulnerable to the genetic load generated by maternal inheritance, yet evidence for mitochondrial involvement in male fertility is limited and controversial. Here, we exploit the coexistence of two divergent mitochondrial haplogroups (A and B2) in a Neotropical arachnid to investigate the role of mitochondria in sperm competition. DNA profiling demonstrated that B2-carrying males sired more than three times as many offspring in sperm competition experiments than A males, and this B2 competitive advantage cannot be explained by female mitochondrial haplogroup or male nuclear genetic background. RNA-Seq of testicular tissues implicates differential expression of mitochondrial oxidative phosphorylation (OXPHOS) genes in the B2 competitive advantage, including a 22-fold upregulation of atp8 in B2 males. Previous comparative genomic analyses have revealed functionally significant amino acid substitutions in differentially expressed genes, indicating that the mitochondrial haplogroups differ not only in expression but also in DNA sequence and protein functioning. However, mitochondrial haplogroup had no effect on sperm number or sperm viability, and, when females were mated to a single male, neither male haplogroup, female haplogroup nor the interaction between male/female haplogroup significantly affected female reproductive success. Our findings therefore suggest that mitochondrial effects on male reproduction may often go undetected in noncompetitive contexts and may prove more important in nature than is currently appreciated.

An epigenetic resolution of the lek paradox.

Female choice for traits signaling male genetic quality is expected to erode heritable variation in fitness, undermining the benefits of choice. Known as the lek paradox, this contradiction has motivated extensive population genetic theory, yet remains unresolved. Recent modeling by Bonduriansky and Day concludes that costly female preference is best maintained when male condition is determined by environmentally induced factors transmitted across single generations. Here, we reformulate their model in explicitly epigenetic terms, and review evidence that environmentally induced paternal effects are mediated through epigenetic changes in sperm. Noncoding RNA expression, DNA methylation and histone modifications are highly sensitive to diet, stress, toxicants and stochastic events. Epigenetic variation renews each generation and cannot be exhausted by selection. By choosing well-endowed males that produce gametes in epigenetically good states, females can increase their fitness directly through increased fertilization success or indirectly through epigenetic effects on the fitness of offspring and potentially subsequent generations. Also watch the video abstract.

Sisters' curse: sexually antagonistic effects constrain the spread of a mitochondrial haplogroup superior in sperm competition.

Maternal inheritance of mitochondria creates a sex-specific selective sieve with implications for male longevity, disease susceptibility and infertility. Because males are an evolutionary dead end for mitochondria, mitochondrial mutations that are harmful or beneficial to males but not females cannot respond directly to selection. Although the importance of this male/female asymmetry in evolutionary response depends on the extent to which mitochondrial mutations exert antagonistic effects on male and female fitness, few studies have documented sex-specific selection acting on mitochondria. Here, we exploited the discovery of two highly divergent mitochondrial haplogroups (A and B2) in central Panamanian populations of the pseudoscorpion Cordylochernes scorpioides. Next-generation sequencing and phylogenetic analyses suggest that selection on the ND4 and ND4L mitochondrial genes may partially explain sexually antagonistic mitochondrial effects on reproduction. Males carrying the rare B2 mitochondrial haplogroup enjoy a marked advantage in sperm competition, but B2 females are significantly less sexually receptive at second mating than A females. This reduced propensity for polyandry is likely to significantly reduce female lifetime reproductive success, thereby limiting the spread of the male beneficial B2 haplogroup. Our findings suggest that maternal inheritance of mitochondria and sexually antagonistic selection can constrain male adaptation and sexual selection in nature.

Outbred embryos rescue inbred half-siblings in mixed-paternity broods of live-bearing females.

Females commonly mate with more than one male, and polyandry has been shown to increase reproductive success in many species. Insemination by multiple males shifts the arena for sexual selection from the external environment to the female reproductive tract, where sperm competition or female choice of sperm could bias fertilization against sperm from genetically inferior or genetically incompatible males. Evidence that polyandry can be a strategy for avoiding incompatibility comes from studies showing that inbreeding cost is reduced in some egg-laying species by postcopulatory mechanisms that favour fertilization by sperm from unrelated males. In viviparous (live-bearing) species, inbreeding not only reduces offspring genetic quality but might also disrupt feto-maternal interactions that are crucial for normal embryonic development. Here we show that polyandry in viviparous pseudoscorpions reduces inbreeding cost not through paternity-biasing mechanisms favouring outbred offspring, but rather because outbred embryos exert a rescuing effect on inbred half-siblings in mixed-paternity broods. The benefits of polyandry may thus be more complex for live-bearing females than for females that lay eggs.

Transposable elements and an epigenetic basis for punctuated equilibria.

Evolution is frequently concentrated in bursts of rapid morphological change and speciation followed by long-term stasis. We propose that this pattern of punctuated equilibria results from an evolutionary tug-of-war between host genomes and transposable elements (TEs) mediated through the epigenome. According to this hypothesis, epigenetic regulatory mechanisms (RNA interference, DNA methylation and histone modifications) maintain stasis by suppressing TE mobilization. However, physiological stress, induced by climate change or invasion of new habitats, disrupts epigenetic regulation and unleashes TEs. With their capacity to drive non-adaptive host evolution, mobilized TEs can restructure the genome and displace populations from adaptive peaks, thus providing an escape from stasis and generating genetic innovations required for rapid diversification. This "epi-transposon hypothesis" can not only explain macroevolutionary tempo and mode, but may also resolve other long-standing controversies, such as Wright's shifting balance theory, Mayr's peripheral isolates model, and McClintock's view of genome restructuring as an adaptive response to challenge.

Historically Underrepresented Graduate Students' Experiences During the COVID-19 Pandemic.

Objective: The purpose of this study was to understand the experiences of historically underrepresented graduate students, more than half of whom were enrolled in science, technology, engineering, and mathematics (STEM) disciplines, during the COVID-19 pandemic. This focus group study represents an initial stage in developing an intervention for historically underrepresented graduate students and their families. Background: Underrepresentation of graduate students of color in STEM has been attributed to a myriad of factors, including a lack of support systems. Familial support is an endorsed reason for persisting in graduate school. It is unclear what historically underrepresented graduate students' experiences are during uncertain times, such as a pandemic. Method: Focus groups were conducted online using a videoconferencing platform during the COVID-19 pandemic. Five focus groups included: historically underrepresented doctoral students (n = 5), historically underrepresented master's students (n = 6), academic faculty (n = 7), administrator, administrative faculty, and academic faculty (n = 6), and families of historically underrepresented doctoral students (n = 6). Data were analyzed using thematic analysis. Results: Historically underrepresented graduate students experienced difficulties in accessing resources, adjustments to home and family life, amplification of existing nonfinancial issues, and expressed both fears of and hopes for the future. Conclusion: The COVID-19 pandemic exacerbated existing inequalities in access to resources as well as nonfinancial family support. Implications: This study may help normalize historically underrepresented graduate students' experiences during the COVID-19 pandemic. The findings include ideas for informing families about graduate school that can enlighten family support efforts for historically underrepresented graduate students and their families.

Reproductive compensation favours male-killing Wolbachia in a live-bearing host.

Wolbachia are maternally inherited, cellular endosymbionts that can enhance their fitness by biasing host sex ratio in favour of females. Male killing (MK) is an extreme form of sex-ratio manipulation that is selectively advantageous if the self-sacrifice of Wolbachia in males increases transmission through females. In live-bearing hosts, females typically produce more embryos than can be carried to term, and reproductive compensation through maternal resource reallocation from dead males to female embryos could increase the number of daughters born to infected females. Here, we report a new strain of MK Wolbachia (wCsc2) in the pseudoscorpion, Cordylochernes scorpioides, and present the first empirical evidence that reproductive compensation favours the killing of males in a viviparous host. Females infected with the wCsc2 strain produced 26 per cent more and significantly larger daughters than tetracycline-cured females. In contrast to the previously described wCsc1 MK Wolbachia strain in C. scorpioides, wCsc2 infection was not accompanied by an increase in the rate of spontaneous brood abortion. Characterization of the wCsc1 and wCsc2 strains by multi-locus sequence typing and by Wolbachia surface protein (wsp) gene sequencing indicates that the marked divergence between these two MK strains in their impact on host reproductive success, and hence in their potential to spread, has occurred in association with homologous recombination in the wsp gene.

Maternal inheritance, sexual conflict and the maladapted male.

Females differ from males in transmitting not only nuclear genes but also cytoplasmic genetic elements (CGEs), including DNA in mitochondria, chloroplasts and microorganisms that are present in the cell. Until recently, evolutionary research has adopted a nucleocentric approach in which organelles have been viewed as subservient energy suppliers. In this article, we propose that a more equitable view of nuclear genes and organelle genomes will lead to a better understanding of the dynamics of sexual selection and the constraints on male adaptation. Maternal inheritance of CGEs intensifies sexually-antagonistic coevolution and provides a parsimonious explanation for the relatively high frequency in males of such apparently maladaptive traits as infertility, homosexuality and baldness.

Viviparity-driven conflict: more to speciation than meets the fly.

Equipped with Mendel's laws and only rudimentary knowledge of genes and genomes, the architects of the Modern Synthesis provided key insights into the dynamics of gene frequency change within populations. Extension of population genetic models to speciation identified Dobzhansky-Muller incompatibilities (negative epistatic interactions between genes from isolated populations) as the primary cause of hybrid inviability and sterility, a view consistent with empirical findings on the genetics of reproductive isolation in Drosophila. Although speciation models have become increasingly mathematically sophisticated, many remain based on an overly static concept of the genome, grounded in Mendelian genetics and devoid of potentially important biological details. A unifying theory of speciation therefore remains elusive, with debate over the relative importance of natural selection, sexual selection, sexual conflict, genetic drift, and selfish genetic elements in the evolution of reproductive isolation. Drawing on recent findings in molecular genetics and comparative genomics, we revisit, update, and extend the theory that reproductive mode plays a crucial role in shaping the speciation process. By providing a direct conduit for manipulation of the mother's physiology by genes expressed in the embryo, viviparity creates a postfertilization arena for genomic conflicts absent in species that lay eggs. In polyandrous species, viviparity-driven conflict (VDC) is likely to generate perpetual antagonistic coevolution between genes expressed during embryonic development and those involved in maternal reproductive physiology, thereby accelerating the rate at which postzygotic isolation evolves between populations. Moreover, in mammals and flowering plants, VDC has favored the evolution of genomic imprinting and a central role for epigenetic mechanisms in the regulation of antagonistic patterns of gene expression by maternally and paternally inherited genomes. VDC can account for the rapid rate at which mammals and viviparous fishes lose their ability to hybridize; the key role of the triploid endosperm in postzygotic reproductive isolation in flowering plants; and the kinds of traits, genes, and gene regulatory systems most critical to the evolution of postzygotic reproductive isolation in live-bearing species.

Simulated climate warming and mitochondrial haplogroup modulate testicular small non-coding RNA expression in the neotropical pseudoscorpion, Cordylochernes scorpioides.

Recent theory suggests that tropical terrestrial arthropods are at significant risk from climate warming. Metabolic rate in such ectothermic species increases exponentially with environmental temperature, and a small temperature increase in a hot environment can therefore have a greater physiological impact than a large temperature increase in a cool environment. In two recent studies of the neotropical pseudoscorpion, Cordylochernes scorpioides, simulated climate warming significantly decreased survival, body size and level of sexual dimorphism. However, these effects were minor compared with catastrophic consequences for male fertility and female fecundity, identifying reproduction as the life stage most vulnerable to climate warming. Here, we examine the effects of chronic high-temperature exposure on epigenetic regulation in C. scorpioides in the context of naturally occurring variation in mitochondrial DNA. Epigenetic mechanisms, including DNA methylation, histone modifications and small non-coding RNA (sncRNA) expression, are particularly sensitive to environmental factors such as temperature, which can induce changes in epigenetic states and phenotypes that may be heritable across generations. Our results indicate that exposure of male pseudoscorpions to elevated temperature significantly altered the expression of >60 sncRNAs in testicular tissue, specifically microRNAs and piwi-interacting RNAs. Mitochondrial haplogroup was also a significant factor influencing both sncRNAs and mitochondrial gene expression. These findings demonstrate that chronic heat stress causes changes in epigenetic profiles that may account for reproductive dysfunction in C. scorpioides males. Moreover, through its effects on epigenetic regulation, mitochondrial DNA polymorphism may provide the potential for an adaptive evolutionary response to climate warming.

Constant diurnal temperature regime alters the impact of simulated climate warming on a tropical pseudoscorpion.

Recent theory suggests that global warming may be catastrophic for tropical ectotherms. Although most studies addressing temperature effects in ectotherms utilize constant temperatures, Jensen's inequality and thermal stress considerations predict that this approach will underestimate warming effects on species experiencing daily temperature fluctuations in nature. Here, we tested this prediction in a neotropical pseudoscorpion. Nymphs were reared in control and high-temperature treatments under a constant daily temperature regime, and results compared to a companion fluctuating-temperature study. At constant temperature, pseudoscorpions outperformed their fluctuating-temperature counterparts. Individuals were larger, developed faster, and males produced more sperm, and females more embryos. The greatest impact of temperature regime involved short-term, adult exposure, with constant temperature mitigating high-temperature effects on reproductive traits. Our findings demonstrate the importance of realistic temperature regimes in climate warming studies, and suggest that exploitation of microhabitats that dampen temperature oscillations may be critical in avoiding extinction as tropical climates warm.

From father to son: transgenerational effect of tetracycline on sperm viability.

The broad-spectrum antibiotic tetracycline is used in animal production, antimicrobial therapy, and for curing arthropods infected with bacterial endosymbionts such as Wolbachia. Tetracycline inhibits mitochondrial translation, and recent evidence indicates that male reproductive traits may be particularly sensitive to this antibiotic. Here, we report the first multi-generation investigation of tetracycline's effects on ejaculate traits. In a study of the pseudoscorpion, Cordylochernes scorpioides, in which siblings were randomly assigned to control and tetracycline treatments across replicate full-sibling families, tetracycline did not affect body size in either sex, female reproduction or sperm number. However, tetracycline-treated males exhibited significantly reduced sperm viability compared to control males, and transmitted this toxic effect of tetracycline on sperm to their untreated sons but not to their F2 grandsons. These results are consistent with tetracycline-induced epigenetic changes in the male germline, and suggest the need for further investigation of transgenerational effects of tetracycline on male reproductive function.

Maternal inheritance, epigenetics and the evolution of polyandry.

Growing evidence indicates that females actively engage in polyandry either to avoid genetic incompatibility or to bias paternity in favor of genetically superior males. Despite empirical support for the intrinsic male quality hypothesis, the maintenance of variation in male fitness remains a conundrum for traditional "good genes" models of sexual selection. Here, we discuss two mechanisms of non-Mendelian inheritance, maternal inheritance of mitochondria and epigenetic regulation of gene expression, which may explain the persistence of variation in male fitness traits important in post-copulatory sexual selection. The inability of males to transmit mitochondria precludes any direct evolutionary response to selection on mitochondrial mutations that reduce or enhance male fitness. Consequently, mitochondrial-based variation in sperm traits is likely to persist, even in the face of intense sperm competition. Indeed, mitochondrial nucleotide substitutions, deletions and insertions are now known to be a primary cause of low sperm count and poor sperm motility in humans. Paradoxically, in the field of sexual selection, female-limited response to selection has been largely overlooked. Similarly, the contribution of epigenetics (e.g., DNA methylation, histone modifications and non-coding RNAs) to heritable variation in male fitness has received little attention from evolutionary theorists. Unlike DNA sequence based variation, epigenetic variation can be strongly influenced by environmental and stochastic effects experienced during the lifetime of an individual. Remarkably, in some cases, acquired epigenetic changes can be stably transmitted to offspring. A recent study indicates that sperm exhibit particularly high levels of epigenetic variation both within and between individuals. We suggest that such epigenetic variation may have important implications for post-copulatory sexual selection and may account for recent findings linking sperm competitive ability to offspring fitness.

AGGRESSION, DENSITY, AND SEXUAL DIMORPHISM IN CHERNETID PSEUDOSCORPIONS (ARACHNIDA: PSEUDOSCORPIONIDA).

The determinants of sexual dimorphism in a family of false scorpions (Pseudoscorpionida: Chernetidae) were investigated experimentally and with a literature analysis of comparative morphometric and habitat data. Species vary in the extent to which males and females differ in size of the pedipalps, and, in particular, in size of the pedipalpal chelae. A statistical analysis of dimorphism patterns within the Chernetidae suggests that dimorphism is a highly variable condition, relatively unconstrained by phylogenetic influences. The evolution of species with enlarged male pedipalps appears to be associated with a change from nonpairing to pairing sperm-transfer behavior and with aggressive mate acquisition by males. Experiments with Dinocheirus arizonensis demonstrate a strong correlation between male combat ability and chela size. Manipulations also suggest that superior combat ability of large males results in increased mating success only under high-density conditions. The greater success in spermatophore transfer among large males can be attributed to increased opportunity at high density for large males to interrupt matings and aggressively displace smaller males. Comparative analysis showed a positive correlation between density and enlarged male chela size across chernetid species.

WHEN MORPHOLOGY MISLEADS: INTERPOPULATION UNIFORMITY IN SEXUAL SELECTION MASKS GENETIC DIVERGENCE IN HARLEQUIN BEETLE-RIDING PSEUDOSCORPION POPULATIONS.

Differences in secondary sexual characteristics of males often provide the most conspicuous means of distinguishing between closely related species. Does this therefore imply that the absence of differentiation in exaggerated male traits between allopatric populations provides evidence of a single, genetically cohesive species? We addressed this question with a comprehensive investigation of two populations (French Guiana and Panama) of the harlequin beetle-riding pseudoscorpion, Cordylochernes scorpioides. This highly sexually dimorphic pseudoscorpion is currently described as a single species, ranging throughout the Neotropics. Our morphometric analyses detected minimal differentiation between the two populations in all nine external morphological characters measured, including sexually dimorphic traits in males. Only in traits of the spermatophore was there any appreciable level of differentiation. Behavior differentiation and prezygotic reproductive isolation were also limited: 78.3% of males successfully transferred sperm to "foreign" females, and in 63.9% of these cases, females' eggs were successfully fertilized. By contrast, extensive divergence existed in two of nine electrophoretic loci, including an essentially fixed-allele difference at the Ldh locus. Most significantly, postzygotic reproductive isolation was complete, with heteropopulation zygotes invariably aborting early in development. These results strongly suggest that the two populations are, in fact, sibling species, a conclusion supported by our recently published findings on their marked divergence in minisatellite DNA. How can such interpopulation homogeneity in male sexually dimorphic traits exist in the face of strong genetic divergence? We propose that sexual selection, oscillating between favoring small and then large males, maintains such high levels of male variability within each population that it has obscured a speciation event in which genetic divergence and postzygotic incompatibility have clearly outpaced the evolution of prezygotic reproductive isolation.