Biology Letters

Group-living animals often coordinate their behavior using "contact calls". Identifying the function of these calls requires testing whether they are intended for any group member or targeted to specific preferred associates. If contact calling is used to coordinate with preferred associates, then higher rates of contact calling are expected between group members with a history of more frequent affiliation and cooperation. To test this hypothesis, we constructed a contact-calling network using synchronized recordings of vocal interactions between all 28 possible pairs of 8 female common vampire bats with well-sampled histories of social grooming and regurgitated food sharing. Bayesian multilevel models show that pairwise rates of contact calling were clearly predicted by social grooming and cooperative allofeeding rates in ways not explained by kinship. These findings show that common vampire bats use contact calls to coordinate with specific same-sex associates, unlike other studied bat species where individuals produce contact calls at similar rates towards different group members. We also found that, compared to white-winged vampire bats, common vampire bats are ten times less likely to rapidly respond to a contact call; this suggests yet-to-be-discovered differences in social behavior between vampire bat species. Finally, we discuss implications for the vocal grooming hypothesis.

Locating, tracking, and intercepting objects is a fundamental behavior for many organisms. For instance, predators must track and capture erratically moving prey for their survival. Using the echolocating bat as a model species, we investigate how short-term changes in target motion predictability affect longer-term motor plans when tracking a prey item. We used a paradigm where prey motion is under experimental control, and then applied computational methods to characterize how target motion predictability influences short- and long-term behavioral control. We find that target motion predictability during the tracking phase of insect capture influences both short-term changes in sonar call control, as well as longer-term behavioral control for transitioning between hunting phases. For changes in immediate behavioral control, bats produce more bursts of calls at a higher rate when tracking unpredictable moving prey, an indication that the bat is collecting more information about the targets motion for unpredictable than predictable trials. In terms of longer-term behavioral control, target motion unpredictability delays the transition from tracking to capture phase behaviors. We suggest that the bat does this to collect more information about target motion to time the transition from tracking to capture behaviors for hunting success. Additionally, we find the effects of target motion unpredictability are first seen as changes in the vocal motor plan and then the auditory motor plan (ear motion), hinting at a sequencing of motor changes that warrant further investigation. SummaryWhen presented with a more challenging hunting task, bats will increase their production of bursts of calls at a higher rate and delay their transition into capture behaviors.

The population of Madagascar exhibits a globally unique combination of African and Asian genetic ancestries. Previous studies have described the admixture history of Madagascar at island-wide scales [1,2], but less focus has been paid to fine-scale population structure across the island. We present new genome-wide genetic data from 192 individuals sampled across five regions of Madagascar. We identify population structure at extremely fine spatial scales ([~]10 km) among the Merina of the central highlands. By analysing subpopulations separately, we found one Merina group exhibited similarity to coastal populations in f4 ratios, estimated admixture dates, and pairwise FST distances, while another group was similar to other highland individuals in the same measures. This fine-scale substructure is likely associated with historical coastal-to-highland migration during the 18th and 19th centuries. In contrast, we also observe macro-scale structure in estimated timing of admixture across the island, with southeastern coastal groups exhibiting the earliest estimated admixture timings, and northern groups exhibiting the latest. This pattern corroborates previous results [1,2], and may suggest differing histories of admixture timing among Malagasy populations. Our results emphasise the importance of deep micro-geographic sampling to complement macro-scale analysis when characterising demographic history.

The ability of animals to innovate - solve novel problems - can shape their ecology and evolution. Here we investigate how individual traits and environmental complexity relate to successful solving of a novel problem. We presented foraging bumble bees (Bombus impatiens) with artificial flowers of not-previously-encountered shapes and recorded the bees latency to access nectar. We measured individual foraging traits across multiple trips with simple flowers that did not require innovation, and bees were foraging either in a simple or complex environment (cluttered flight arena). Bees in complex environments took longer to find and were less likely to land on novel flowers, indicating that environmental complexity may take up cognitive resources and make search more difficult. However, we did not find an effect of environmental treatment on the ability or time to access reward in novel flowers once bees had landed on them. In contrast, behavioral traits significantly predicted how quickly bees solved novel flowers. In particular, overall foraging tempo as well as routine formation, i.e. how much bees followed a fixed route on known flowers, predicted innovation - faster bees innovated faster, and bees with more repetitive foraging sequences were slower to solve the novel tasks. Overall, while the degree of evolutionary novelty in tasks or solutions is always hard to evaluate, our findings demonstrate that environment and individual traits may affect innovation in different ways. Individuals in simple environments may be more likely to detect, and individuals that are generally faster and have a lower tendency to develop fixed routines may be more likely to solve, novel tasks.

BackgroundAlthough strict maternal transmission of mitochondria is a general feature of animals and humans for ensuring homogeneity in mitochondrial DNA (mtDNA) across generations, exceptions were reported in the recent past. For example, some extremely rare but spectacular cases of heteroplasmy and paternal transmission in humans have questioned the universal evolutionary principle. Hence, as an alternative, the Mega-NUMT concept was coined to explain this discovery and was thereafter partly proven to exist. This concept expands on the quite common transfer of mtDNA fragments to the nucleus (NUMTs) by considering the existence of multicopy mitochondrial nuclear insertions. Mega-NUMT reports are currently restricted to a few cases in animals, including humans. However, even in humans, their detailed genomic organization, natural prevalence, and potential biological functions remain unclear. Methodology/Principal FindingsHere, we discovered that up to 60 full-sized mitochondrial genomes are integrated into the nuclear genome of the neotropical fruit fly Drosophila paulistorum using long-read sequencing and confirmed their presence by in situ hybridization. The copies are organized in one cluster on chromosome 3, which we, due to its similarity with the Mega-NUMT concept, designated the "Dpau Mega-NUMT". Contrary to the rarity in humans, this Mega-NUMT is found at high prevalence (40%) in both long-term laboratory lines and natural D. paulistorum populations of different semispecies. Additionally, the mitochondrial copies in the Mega-NUMT cluster are phylogenetically separated from the current mitotypes of D. paulistorum. Together, these observations suggest long-term maintenance of the Mega-NUMT in nature. Hence, we propose that the Dpau Mega-NUMT may have been transferred to the nuclear genome before D. paulistorum semispecies radiation and maintained at relatively high prevalence in nature by balancing selection due to yet undetermined functions. Conclusions/SignificanceTo our knowledge, this is the first verified existence and detailed dissection of a Mega-NUMT outside cats and humans. We show that Mega-NUMTs can be persistent in nature, even at high prevalence, potentially due to balancing selection. Our findings strengthen the importance of high-quality long-read sequencing technologies for deciphering complex repeat-rich genomic regions to deepen our understanding of the dynamics of genome evolution within genomic "dark matter".

In asexual reproduction, meiosis must be bypassed or altered to maintain ploidy from mother to daughter without fertilization. Most of the ways meiosis can be modified to this end are expected to reduce heterozygosity within individuals; however, many asexual species are highly heterozygous. Asexual reproduction is especially common among species of microscopic, desiccation-tolerant animals such as rotifers, nematodes, and tardigrades, but the cellular and genetic mechanisms underlying asexual reproduction have not been definitively documented in any species of tardigrade. Here, we show that the asexual tardigrade Hypsibius exemplaris fails to complete the cell division of meiosis I, followed by a complete meiosis II-like division, and reproduction proceeds without detectable loss of heterozygosity. We used a combined cytological and genomic approach to characterize the mechanism of reproduction and pattern of allele inheritance in this species. Furthermore, we identified heterozygous variants in a subset of transcriptionally active genes consistent with loss of function in one allele, suggesting that maintained heterozygosity in this species allowed divergence between alleles over time. This work establishes the meiotic mechanism and inheritance pattern of reproduction in H. exemplaris, which provides a framework for interpreting genetic variation in this organism as a laboratory model. Additionally, our finding that meiosis is modified in H. exemplaris via a mechanism that maintains heterozygosity across the genome adds to a growing body of evidence that maintaining heterozygosity is not detrimental to the long-term survival of asexual eukaryotes. Article SummaryAnimals that reproduce asexually must alter meiosis, a highly conserved process of two cell divisions normally used to make eggs and sperm. This study represents the first combined cytological and genetic characterization of how meiosis is modified in a tardigrade. The authors found that the model tardigrade Hypsibius exemplaris modifies meiosis by skipping the first cell division, but completing the second. Additionally, they found that this species preserves heterozygosity across the genome and from generation to generation. Finally, some genes show evidence of sequence divergence between alleles, supporting a broader conclusion that maintaining heterozygosity influences how asexual species genomes evolve.

Species in stable social groups engage in diverse social interactions, where social competence - the ability to adjust behaviour using social information - can influence fitness. Yet, whether adaptive behavioural flexibility is expressed across contexts within individuals remains relatively untested. To address this, we exposed cooperatively breeding cichlids (Neolamprologus pulcher) to a role-reversal paradigm. In this species, the early social environment shapes social competence, with more competent individuals adjusting behaviour flexibly to social challenges, while individuals also show consistent differences in traits such as aggression. In the present study, individuals were successively assigned to two contrasting roles, smaller territory owners (TOs) and larger intruders (INTs). We predicted role-specific social competence metrics based on behaviours facilitating shelter acquisition. Social competence metrics correlated within, but not across the two roles. Competent TOs showed shorter latencies to submit, more submissive responses to received aggression, and low aggression after initial submission. Competent INTs escalated quickly and relied more on overt aggression rather than displays, allowing faster shelter acquisition. Across roles, individuals competent as TOs were not competent as INTs. In contrast, consistent individual differences in aggression across social roles suggest that stable behavioural tendencies ( animal personalities) may constrain how social competence shapes behavioural strategies.

Although predation is a major driver of group living across taxa and the antipredator benefits of grouping are well established, the energetic costs experienced by groups under predation remain largely unexplored. In the current study, we use wild, white mullet (Mugil curema, Valenciennes 1836), to provide the first real-time quantification of the energetic cost of escape in schooling fish using intermittent, closed-loop respirometry. We found that small groups exposed to predators showed a 53.8% increase in their organismal metabolic rate (MO2) as compared to groups without predator exposure. When we evaluated antipredator behaviors such as escape response, group cohesion, and displacement of the group centroid, we found a positive correlation to energetic costs. We then investigated whether escape responses are socially modulated by comparing the energetic costs of escape across solitary individuals, solitary individuals with visual access to a group, and groups. We found that escape frequency and energetic costs to predation were comparable across social contexts, indicating that escape may be an intrinsic survival response independent of cues from group members. Furthermore, we found that fish exposed to predators showed markedly reduced feeding, suggesting that predation constrains energy acquisition in addition to imposing direct energetic costs. Our results provide the first direct quantification of the energetic costs of escape in a schooling fish, offering new insights into the physiological trade-offs underlying collective antipredator defenses.

Small marsupials in the family Dasyuridae are a key component of Australias arid and semi-arid fauna, whose high species richness is proposed to reflect an opportunity-driven adaptive radiation. Despite growing interest in this group from both ecological and evolutionary perspectives, genomic data for most species is non-existent, or limited to a few marker loci. Here, we generated a chromosome-level reference genome and a de novo mitochondrial genome for the desert-dwelling Wongai ningaui (Ningaui ridei). The nuclear genome assembly is highly contiguous, with a scaffold N50 of 594.484 MB and high BUSCO gene recovery (93.84%). Additionally, we produced a draft assembly for the related, semi-arid slender-tailed dunnart (Sminthopsis murina). We then used these assemblies to explore the demographic histories of these species. We find evidence for contrasting patterns of population growth during the late Pleistocene and early Holocene, corresponding with differences in local climate, potentially consistent with differences in optimal habitat. The new genomic resources and demographic findings presented here provide a foundation for future studies on adaptive specialisation in this group of Australian marsupials. Significance StatementDasyurid marsupials are the primary carnivorous and insectivorous mammals in Australia. This diverse family includes species such as the endangered Tasmanian devil (Sarcophilus harrisii) and quolls (Genus Dasyurus), as well as an emerging laboratory model species, the fat-tailed dunnart (Sminthopsis crassicaudata). Despite the species richness within dasyurids, most species remain under-studied. This is particularly true of arid and semi-arid zone species, who are often small in size, live in remote habitats and are cryptic by nature. By creating genome assemblies for two dasyurid species, this study provides resources to support a variety of phylogenetic, population genetic and evolutionary developmental lines of research. Importantly, the studys finding that arid and semi-arid dasyurids show distinct trajectories of demographic change in response to historical climatic shifts may point to local adaptations with implications for the resilience of these species to ongoing and future climate change.

Despite growing evidence that many animals can evaluate quantities, the ecological relevance of numerical cognition remains debated, particularly outside vertebrates. Would individuals still rely on numerousness if less computationally demanding cues, visual features extracted at the early stage of visual processing, were available to assess quantity? In primates, individuals show a numerical bias as they tend to rely on the number of items rather than non-numerical cues, such as total area, to categorize quantities. In this study, we trained free-flying honeybees to discriminate between two and four items in conditions where numerosity covaried with the total area and perimeter (Experiment Size) or the convex hull (Experiment Space) cues, mimicking ecological contexts. Transfer tests assessed which numerical or non-numerical cues were learned and preferentially used by the bees. Bees primarily relied on numerousness over these non-numerical cues. Individual analyses revealed two consistent strategies: a "numerical bias" strategy, in which bees encoded numerical information while ignoring non-numerical cues, and a "generalist" strategy, where bees flexibly switched between cues and favored non-numerical information when cues conflicted. We further reported improved discrimination when smaller quantities appeared on the left and larger ones on the right, consistent with an oriented mental number line. Together, these findings demonstrate a spontaneous numerical bias in honeybees and reveal that individuals within the same species can adopt distinct strategies when evaluating quantity. Our findings also suggest that distantly related taxa like bees and primates may have independently evolved comparable mechanisms for quantity evaluation.

Nocturnal migration is a remarkable phenomenon observed in many insect species, including moths. Migratory moths are capable of maintaining precise directional orientation during migration, as demonstrated in both laboratory and field studies, suggesting that they use multiple environmental cues for orientation and navigation. Recent studies on Australian Bogong moths revealed that these animals can use stellar cues and likely the geomagnetic field (in conjunction with local visual cues) to select and maintain population-specific migratory direction. However, the underlying orientation mechanisms used by most other migratory moths are still largely unresolved. Further, it remains unclear whether migratory moths can adjust their orientation using Earths magnetic field parameters for determining their position relative to the goal (i.e. location or map information) - an ability clearly shown in some migratory birds which respond to virtual magnetic displacements by correcting their orientation (experiments when animals are exposed to magnetic cues corresponding to other geographic locations). Here, we present results from virtual magnetic displacement experiments conducted on red underwings (Catocala nupta). In addition, we tested their orientation under simulated overcast conditions and in a vertical magnetic field to get indications whether this species relies on geomagnetic or celestial cues to maintain its population-specific migratory direction. Our results show that (1) red underwings did not compensate for virtual magnetic displacement, indicating the absence of a magnetic map; (2) they remained significantly oriented in the absence of geomagnetic information, suggesting the use of a stellar compass; and (3) there was no evidence of magnetic compass orientation in absence of any visual cues.

Many perceptual skills improve with a few days of training. However, weeks or months of practice may be required to reach a level of expertise on complex tasks (Watson, 1980). Here, we explored how gerbils attain expertise on a difficult task: amplitude modulation (AM) rate discrimination at very shallow AM depths, similar to the depths used during vocal communication. Using an appetitive Go-Nogo procedure, we first trained 6 gerbils to perform an AM discrimination task (Nogo: 4 Hz; Go: 4.25-10 Hz) at a depth of 0 dB (re: 100% depth). Animals were then trained to perform AM discrimination at successively shallower depths, from -3 to -18 dB, requiring an average of 5-10 days of practice to reach a performance metric of d[≥]1 for each depth. Finally, we determined that AM discrimination thresholds were nearly identical between 0 to -12 dB, and only slightly elevated at -15 dB. Improvements in performance were accompanied by a large reduction in response time during procedural learning, and a gradual reduction of response time during perceptual learning, even as AM depth became shallower (i.e., more difficult). The shallowest depth at which gerbils displayed peak performance on the AM discrimination task is similar to their lowest AM depth detection thresholds. These results suggest performance on challenging auditory perceptual tasks require prolonged practice, and is accompanied by increased automaticity (i.e., lower response time) that stabilizes once expertise is achieved.

Collective behavior in animal societies can buffer individual costs and confer resilience to environmental challenges. However, the mechanisms by which groups sustain function when members are compromised remain poorly understood. In the presented study, we investigate how social context shapes collective fanning, a thermoregulatory behavior critical for colony function, in Western honeybees (Apis mellifera). Using oxytetracycline (OTC), a known physiologically disruptive antibiotic to honeybees, to selectively impair certain group members, we tested our hypothesis that the presence of untreated bees would rescue the fanning response in mixed-composition groups. We show that groups containing untreated individuals fan at levels comparable to fully untreated groups, despite the presence of OTC-impaired bees. This preservation of collective thermoregulatory function was correlated with both treated and untreated individuals in mixed groups shifting their interaction dynamics and social network positions. These findings reveal a decentralized mechanism of collective resilience, whereby behavioral compensation by individuals sustains group-level thermoregulation under partial disruption. Our results provide a framework for understanding how social insect colonies maintain function in the face of individual-level perturbations, with broader implications for predicting the limits of collective resilience in animal societies experiencing increasing environmental pressures.

Social living often confers substantial fitness benefits; however, close spatial association among individuals can also elevate opportunities for pathogen transmission, especially where the populations are dense. Despite this, the extent to which avoidance behaviours are expressed by wild reptiles facing fungal disease remains unclear. We examined Eastern Water Dragons (EWDs; Intellagama lesueurii) in Roma Street Parklands, Brisbane, Australia, where a population is affected by the emerging fungal pathogen Nannizziopsis barbatae (Nb). Using a five-year dataset (2018-2023) spanning 146 individuals, we quantified social distance, as the minimum distance to the nearest neighbour, in relation to the number of diseased conspecifics that overlapped each individuals seasonal core home area. Social distance decreased as diseased conspecifics became more numerous, indicating a strong crowding effect; however, this reduction was weaker for diseased individuals, which maintained larger distances than healthy individuals even under high disease pressure. Together, these patterns support partial social avoidance consistent with behavioural changes in infected individuals, suggesting that infection risk constrains density-driven proximity. Our findings provide new insights into how disease pressure shapes social spacing in reptiles and contribute to a broader understanding of behavioural responses to emerging infectious fungal diseases.

Many animals communicate using sequences of signals, but identifying recurrent, non-random signal combinations remains methodologically challenging. Collocation analyses are increasingly popular approaches for detecting which signals animals combine at rates greater than expected by chance. However, existing methods for animal collocation analysis face several limitations that reduce their statistical rigour: they lack uncertainty estimates, fail to control for non-independence in sampled data, and do not account for inflated family-wise error rates when identifying attraction among many different signal types. These limitations restrict the broader applicability of animal collocation analysis, including preventing robust comparisons of signal combination strength between cohorts (e.g. populations, sexes or age classes). We adapt a novel form of Multiple Distinctive Collocation Analysis using Pearson residuals (MDCA-Pr) that addresses these statistical limitations, and validate its use in animal communication research in three ways: first, using numerous simulated datasets of different sizes and levels of signal recombination; second, using simulated data to evaluate the performance of MDCA-Pr in intercohort comparisons, and third, by demonstrating how MDCA-Pr can be applied to compare the vocal sequences produced by male and female captive-living common marmosets (Callithrix jacchus). MDCA-Pr shows high sensitivity, including at small sample sizes, and generally low false-positive rates, which we further reduce by applying additional criteria for identifying attraction between signals. During intercohort comparisons, MDCA-Pr is conservative, with low false-positive rates, and statistical power increases with sample size. MDCA-Pr is a robust method for evaluating signal attraction in animal communication and enables accurate intercohort comparison of animal signal combinations. Significance StatementBy assessing the performance of MDCA-Pr on simulated animal-like data, we demonstrate that this method reliably detects signal combinations within and across animal cohorts, while overcoming statistical limitations of previous collocation analyses. We present an analytical pipeline for applying MDCA-Pr to animal signal data, including for intercohort comparisons, enabling identification and comparison of combinatorial strategies across entire signal repertoires. We illustrate this approach by comparing call combination strategies of male and female common marmosets when presented with food under experimental conditions, finding similar combinatorial strategies between sexes. MDCA-Pr therefore permits rigorous characterization of animal signal combinatoriality and opens avenues for investigating how demographic, social, and group-level factors influence combinatorial patterns.

In the search for universals shaping acoustic communication across species, we increasingly look for patterns known from human languages and music in non-human animals. These parallels are often explored separately and with limited ecological context. Here, we take a deep dive into the temporal structure of a complex call used by the little auk (Alle alle), a pelagic seabird with elaborate vocal behaviour and socially complex colonial life. Based on syllable durations, intervals and silences, we examine its conformance to linguistic laws, rhythmic structure and information content. This reveals intricate problems of temporal organisation: while the calls conform not only to linguistic laws of brevity but also to the initial and final lengthening known from human prosody, these effects interact with the internal structure of the call and information carried within it. To our knowledge, this is the first time that conformance to multiple linguistic laws, exceeding simple vocal efficiency, has been described for a non-human, non-vocal learning animal. The calls rhythmic structure shows a progressive rallentando -- a systematic slowing driven by changes in syllable and silence durations and the intervals between syllable onsets. The exact patterns of this rallentando are indicative of the callers sex and individually specific. These results reveal how seabird communication is shaped not only by efficiency universals, but also the specific pressures of colonial life. Our work highlights the temporal structure as an important axis of communication evolution, but also serves as a reminder to consider the species ecological reality and the function, not only presence, of temporal organisation. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=127 SRC="FIGDIR/small/713940v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@13de3a8org.highwire.dtl.DTLVardef@2d64adorg.highwire.dtl.DTLVardef@2ca53aorg.highwire.dtl.DTLVardef@113c38d_HPS_FORMAT_FIGEXP M_FIG C_FIG

Animals often aggregate information from multiple different sensory modalities to accurately assess and react to a stimulus. It is often assumed that cross-modality integration mostly occurs at high-level processing centers, such as the mammalian cortex or insect mushroom bodies. However, we hypothesized that integration could occur relatively early in the sensory pathways. The insect antennal lobe is one such location, receiving direct inputs from the antennae via the antennal nerve. These inputs are highly multimodal, including olfactory, mechanosesnory, and gustatory information, all of which are relevant to foraging honeybees (Apis mellifera). Here we assess integration by recording electrophysiological spike data within the honeybee antennal lobe while exposing the bee to various combinations of wind speed and odor concentration. This paper accompanies another publication by Joseph Reed and Mainak Patel approaching the same question from a modelling perspective, where their model corroborates our data and vice versa. Together, we show that integration occurs within this early layer of processing, while also demonstrating the complex relationship of these two closely-linked stimuli. SIGNIFICANCEAccurate perception depends on the brains ability to combine information from multiple senses, commonly thought belonging to high level of information processing. Using the European honeybee, Apis mellifera, we show strong evidence that olfactory and mechanosensory signals interact at an early stage of neural processing, within the antennal lobe, producing stimulus representations closely-linked to the animals navigation and decision-making. By identifying a tractable model for early multisensory processing, this work offers broader insight into how animals construct reliable representations of their environment.

Escape behaviour directly influences survival, yet individuals often vary substantially in escape performance. Laboratory studies have documented trade-offs between anti-predator responses and life-history traits, but it remains unclear whether such trade-offs occur under natural predation risk. We studied a natural population of the field cricket Gryllus campestris. Mortality risk and behavioural performance are known to change with age in this species. We aimed to determine whether individuals expressing a higher escape response pay a cost in terms of a faster increase in mortality risk with age or a shorter lifespan. We quantified escape speed in response to a vibrational predation cue. We found no clear evidence for a trade-off between escape performance and lifespan or age-specific mortality risk. The relationship between escape speed and the among-individual effect of age differed between sexes: older males showed faster escape speeds compared with younger males, whereas younger females were faster than older females. This pattern is consistent with sex-specific selective disappearance. Individual baseline mortality risk varied with sex and escape speed, but age-dependent mortality did not. It suggests that such trade-offs in the wild may be context- or condition-dependent rather than reflecting a universal life-history trade-off.

The expensive son hypothesis posits that mothers incur higher fitness costs when caring for sons versus daughters in species with male-biased size dimorphism. Evidence for maternal survival costs of sons in humans is limited to shortened overall lifespans; whether having more sons reduces short-term survival during reproductive years is unknown. Here, we utilised life-history data from 5,456 mothers from preindustrial Finland to examine whether mothers with more sons had reduced survival within one year of their last birth. While mothers with few children but more sons showed no differences in survival, at higher family sizes, mothers with more sons had increasingly lower survival. These differences peaked at [~]0.4% lower survival per son among mothers with five children, suggesting accumulated physiological costs of sons. These differences then declined and reversed among mothers with more children, potentially due to selective disappearance of frailer mothers. Our results suggest that studies focusing on post-menopausal mothers may bias estimates of the fitness costs of sons and reproductive costs more broadly. We recommend future research further examines the overlooked short-term fitness costs of sons during reproductive years, which is vital for understanding how life-history trade-offs, sexual dimorphism, and their interaction have shaped human evolution.

Conspicuous coloration in animals is generally thought to evolve and be maintained through inter- or intraspecific interactions such as mate choice, but this might not always be the case. The sight-line hypothesis proposes that conspicuous light-dark contrast in front of the eyes (hereafter, eyeline) evolves and is maintained due to viability selection, enhancing an individual visual acuity and thus evolutionarily associated with a particular foraging behavior that requires accurate aiming. However, empirical evidence that supports the sight-line hypothesis is virtually absent, with no studies demonstrating the key prediction that the direction of eyelines matters. Here, I tested the sight-line hypothesis using macroevolutionary analyses in terns and allies, which are a suitable study system, because they have variation in facial color patterns, including presence/absence and, if any, various angles of eyelines. They also have a large variation in foraging behavior, including picking, plunge diving, and skimming. As predicted by the sight-line hypothesis, tern lineages that require accurate aiming at foraging (e.g., plunge diving) are more likely to have eyelines. In addition, the evolutionary transition to the state with eyelines and these foraging behaviors was more likely to occur than the reverse transition. Furthermore, as expected by the fact that the direction of travel is upwardly deviated from the direction of the bills during skimming, the eyeline angle from bills was evolutionarily positively associated with the occurrence of skimming behavior. To my knowledge, the current study is the first to demonstrate that the direction of the eyeline matters, thereby strongly supporting the sight-line hypothesis.