Evolutionary trade-offs between testes size and parenting in Neotropical glassfrogs.

In males, large testes size signifies high sperm production and is commonly linked to heightened sperm competition levels. It may also evolve as a response to an elevated risk of sperm depletion due to multiple mating or large clutch sizes. Conversely, weapons, mate or clutch guarding may allow individuals to monopolize mating events and preclude sperm competition, thereby reducing the selection of large testes. Herein, we examined how paternal care, sexual size dimorphism (SSD), weaponry and female fecundity are linked to testes size in glassfrogs. We found that paternal care was associated with a reduction in relative testes size, suggesting an evolutionary trade-off between testes size and parenting. Although females were slightly larger than males and species with paternal care tended to have larger clutches, there was no significant relationship between SSD, clutch size and relative testes size. These findings suggest that the evolution of testes size in glassfrogs is influenced by sperm competition risk, rather than sperm depletion risk. We infer that clutch guarding precludes the risk of fertilization by other males and consequently diminishes selective pressure for larger testes. Our study highlights the prominent role of paternal care in the evolution of testes size in species with external fertilization.

What Amphibians Can Teach Us About the Evolution of Parental Care.

Parenting is considered a key evolutionary innovation that contributed to the diversification and expansion of vertebrates. However, we know little about how such diversity evolved. Amphibians are an ideal group in which to identify the ecological factors that have facilitated or constrained the evolution of different forms of parental care. Among, but also within, the three amphibian orders-Anura, Caudata, and Gymnophiona-there is a high level of variation in habitat use, fertilization mode, mating systems, and parental sex roles. Recent work using broad phylogenetic, experimental, and physiological approaches has helped to uncover the factors that have selected for the evolution of care and transitions between different forms of parenting. Here, we highlight the exceptional diversity of amphibian parental care, emphasize the unique opportunities this group offers for addressing key questions about the evolution of parenting, and give insights into promising novel directions of research.

Sex-specific discrimination of familiar and unfamiliar mates in the Tokay gecko.

Social animals need to keep track of other individuals in their group to be able to adjust their behaviour accordingly and facilitate group cohesion. This recognition ability varies across species and is influenced by cognitive capacities such as learning and memory. In reptiles, particularly Squamates (lizards, snakes, and worm lizards), chemical communication is pivotal for territoriality, reproduction, and other social interactions. However, the cognitive processes underlying these social interactions remain understudied. In our study, we examined the ability of male and female Tokay geckos (Gekko gecko) to chemically differentiate familiar and unfamiliar mating partners. Our findings suggest that both sexes can make this distinction, with males responding more to the odour of a familiar mate, and females responding more to unfamiliar mates. The lizards maintained their discriminatory abilities for two to three weeks but not up to six weeks after separation. This research highlights the efficacy of using odours as social stimuli for investigating social cognition in lizards, a promising avenue to better understand social cognition in these animals.

Water-borne testosterone levels predict exploratory tendency in male poison frogs.

Hormones play a fundamental role in mediating social behaviors of animals. However, it is less well understood to what extent behavioral variation between individuals can be attributed to variation in underlying hormonal profiles. The goal of the present study was to infer if individual androgen levels, and/or the modulation thereof, can explain among-individual variation in aggressiveness, boldness and exploration. We used as a model the dart-poison frog Allobates femoralis and took repeated non-invasive water-borne hormonal samples of individual males before (baseline) and after (experimental) a series of behavioral tests for assessing aggression, boldness, and exploratory tendency. Our results show that androgen levels in A. femoralis are quite stable across the reproductive season. Repeatability in wbT baseline levels was high, while time of day, age of the frog, and trial order did not show any significant impact on measured wbT levels. In general, experimental wbT levels after behavioral tests were lower compared to the respective baseline levels. However, we identified two different patterns with regard to androgen modulation in response to behavioral testing: individuals with low baseline wbT tended to have increased wbT levels after the behavioral testing, while individuals with comparatively high baseline wbT levels rather showed a decrease in hormonal levels after testing. Our results also suggest that baseline wbT levels are linked to the personality trait exploration, and that androgen modulation is linked to boldness in A. femoralis males. These results show that differences in hormonal profiles and/or hormonal modulation in response to social challenges can indeed explain among-individual differences in behavioral traits.

Personality traits differentially affect components of reproductive success in a Neotropical poison frog.

Individual reproductive success has several components, including the acquisition of mating partners, offspring production, and offspring survival until adulthood. While the effects of certain personality traits-such as boldness or aggressiveness-on single components of reproductive success are well studied, we know little about the composite and multifaceted effects behavioural traits can have on all the aspects of reproductive success. Behavioural traits positively linked to one component of reproductive success might not be beneficial for other components, and these effects may differ between sexes. We investigated the influence of boldness, aggressiveness, and exploration on the number of mating partners, mating events, and offspring surviving until adulthood in males and females of the Neotropical poison frog Allobates femoralis. Behavioural traits had different-even opposite-effects on distinct components of reproductive success in both males and females. For example, males who displayed high levels of aggressiveness and exploration (or low levels of aggressiveness and exploration) managed to attract high number of mating partners, while males with low levels of boldness, low levels of aggressiveness, and high levels of exploration had the most offspring surviving until adulthood. Our results therefore suggest correlational selection favouring particular combinations of behavioural traits.

Complement protein C3a enhances adaptive immune responses towards FVIII products.

The most serious complication in the treatment of hemophilia A (HA) is the development of factor (F)VIII inhibitors or antidrug antibodies (ADA) occurring in 25-35% of patients with severe HA. The immunological mechanisms underlying the development of ADA against FVIII products have not been completely understood yet. Immunological danger signals associated with events such as infection or surgery have been suggested to play a critical role. In previous studies, we demonstrated that plasma-derived (pd)FVIII but not recombinant (r)FVIII can activate human monocyte-derived dendritic cells (DC) in a danger signal-dependent manner, which subsequently mediate the proliferation of autologous CD4+ T cells. In this study, we investigated the ability of plasma components, naturally present in pdFVIII products, to mediate T-cell responses. In fact, we show that addition of plasma to rFVIII plus lipopolysaccharide (LPS)-stimulated DC induces proliferation of autologous CD4+ T cells. Interestingly, although DC pulsed with LPS plus plasma induce T-cell proliferation upon co-culture, the addition of FVIII significantly increases the number of proliferating as well as FVIII-specific CD4+ T cells. Total proliferating CD4+ T cells and FVIII-specific subsets were identified mainly as central memory T cells. Experiments using blocking antibodies and receptor antagonists revealed that the complement proteins C3a and, to a lesser extent, C5a are critically involved in these LPS-mediated T-cell responses. Collectively, our results indicate that complement proteins are potent drivers of T-cell responses to FVIII. Data presented provide a model how event-related substitution of FVIII in HA patients might contribute to inhibitor development.

Fear of the new? Geckos hesitate to attack novel prey, feed near objects and enter a novel space.

Neophobia, the fear of novelty, is an ecologically important response which enables animals to avoid potentially harmful situations. Neophobia is a cognitive process by which individuals distinguish novelty from familiarity. In this study, we aimed to quantify this cognitive process in captive tokay geckos (Gekko gecko) across three contexts: when encountering novel prey, foraging near novel objects and entering a novel space. We also investigated individual consistency across trials using different novel stimuli, and correlation of individual responses across the three contexts. We found that geckos hesitate to attack novel prey and prey close to objects (familiar and novel). Geckos hesitated the most when entering novel space. Repeatability of behaviour within and across contexts was low (R = 0.101-0.190) indicating that neophobia might not be expressed similarly across contexts. The strength of a neophobic response can indicate how anxious or curious an individual is. This test has great potential to help answer questions about how captivity, enrichment, rearing environment and cognition affect fear responses in different contexts in lizards. By studying reptiles, we can better understand the universality of what is known about the causes leading to difference in neophobia across individuals and species.

Geckos differentiate self from other using both skin and faecal chemicals: evidence towards self-recognition?

Self-recognition is the ability to recognise stimuli originating from oneself. Humans and some non-human animals show evidence of true visual self-recognition in the mirror test. They use their reflection to inspect themselves and to remove a mark that is only visible in the mirror. Not all animals, however, rely primarily on vision. In lizards, chemical cues are crucial in social interactions, and therefore, lizards would benefit from a chemical self-other distinction. Here, we test the tokay gecko (Gekko gecko), a social species, on their ability to discriminate their own skin and faecal chemicals from those of same-sex, unfamiliar conspecifics. We predicted that individuals would show more self-directed behaviour when confronted with the chemicals from unfamiliar individuals within their home enclosure as a sign of the need for increased comparison. Geckos showed higher self-directed responses towards chemicals from unfamiliar individuals compared to self-produced chemicals and a water control. Furthermore, scat and skin chemicals (regardless of origin) elicited similar but stronger responses than peppermint oil pointing towards a possible social function of scat piles. Although further tests and controls are needed, our study provides evidence towards chemical self-recognition and for a social function of scat piling in tokay geckos.

Androgen responsiveness to simulated territorial intrusions in Allobates femoralis males: Evidence supporting the challenge hypothesis in a territorial frog.

Territoriality has been widely described across many animal taxa, where the acquisition and defence of a territory are critical for the fitness of an individual. Extensive evidence suggests that androgens are involved in the modulation of territorial behaviours in male vertebrates. Short-term increase of androgen following a territorial encounter appears to favour the outcome of a challenge. The "Challenge Hypothesis" proposed by Wingfield and colleagues outlines the existence of a positive feedback relationship between androgen and social challenges (e.g., territorial intrusions) in male vertebrates. Here we tested the challenge hypothesis in the highly territorial poison frog, Allobates femoralis, in its natural habitat by exposing males to simulated territorial intrusions in the form of acoustic playbacks. We quantified repeatedly androgen concentrations of individual males via a non-invasive water-borne sampling approach. Our results show that A. femoralis males exhibited a positive behavioural and androgenic response after being confronted to simulated territorial intrusions, providing support for the Challenge Hypothesis in a territorial frog.

Naive poison frog tadpoles use bi-modal cues to avoid insect predators but not heterospecific predatory tadpoles.

For animals to survive until reproduction, it is crucial that juveniles successfully detect potential predators and respond with appropriate behavior. The recognition of cues originating from predators can be innate or learned. Cues of various modalities might be used alone or in multi-modal combinations to detect and distinguish predators but studies investigating multi-modal integration in predator avoidance are scarce. Here, we used wild, naive tadpoles of the Neotropical poison frog Allobates femoralis ( Boulenger, 1884) to test their reaction to cues with two modalities from two different sympatrically occurring potential predators: heterospecific predatory Dendrobates tinctorius tadpoles and dragonfly larvae. We presented A. femoralis tadpoles with olfactory or visual cues, or a combination of the two, and compared their reaction to a water control in a between-individual design. In our trials, A. femoralis tadpoles reacted to multi-modal stimuli (a combination of visual and chemical information) originating from dragonfly larvae with avoidance but showed no reaction to uni-modal cues or cues from heterospecific tadpoles. In addition, visual cues from conspecifics increased swimming activity while cues from predators had no effect on tadpole activity. Our results show that A. femoralis tadpoles can innately recognize some predators and probably need both visual and chemical information to effectively avoid them. This is the first study looking at anti-predator behavior in poison frog tadpoles. We discuss how parental care might influence the expression of predator avoidance responses in tadpoles.

Experience shapes accuracy in territorial decision-making in a poison frog.

The trade-off between speed and accuracy affects many behavioural processes like predator avoidance, foraging and nest-site selection, but little is known about this trade-off relative to territorial behaviour. Some poison frogs are highly territorial and fiercely repel calling male intruders. However, attacks need to be conducted cautiously, as they are energetically costly and bear the risk of own injury or accidentally targeting the wrong individual. In this study, we investigated the speed-accuracy trade-off in the context of male territoriality during the breeding season in the brilliant-thighed poison frog, Allobates femoralis. In our experiment, we presented the call of an invisible 'threatening' intruder together with a visible 'non-threatening' intruder, using acoustic playback and a frog model, respectively. Contrary to our prediction, neither reaction time nor approach speed of the tested frogs determined the likelihood of erroneous attacks. However, younger individuals were more likely to attack the non-threatening model than older ones, suggesting that experience plays an essential role in identifying and distinguishing rivalling individuals in a territorial context.

Developments in Amphibian Parental Care Research: History, Present Advances, and Future Perspectives.

Despite rising interest among scientists for over two centuries, parental care behavior has not been as thoroughly studied in amphibians as it has in other taxa. The first reports of amphibian parental care date from the early 18th century, when Maria Sibylla Merian went on a field expedition in Suriname and reported frog metamorphs emerging from their mother's dorsal skin. Reports of this and other parental behaviors in amphibians remained descriptive for decades, often as side notes during expeditions with another purpose. However, since the 1980s, experimental approaches have proliferated, providing detailed knowledge about the adaptive value of observed behaviors. Today, we recognize more than 30 types of parental care in amphibians, but most studies focus on just a few families and have favored anurans over urodeles and caecilians. Here, we provide a synthesis of the last three centuries of parental care research in the three orders comprising the amphibians. We draw attention to the progress from the very first descriptions to the most recent experimental studies, and highlight the importance of natural history observations as a source of new hypotheses and necessary context to interpret experimental findings. We encourage amphibian parental care researchers to diversify their study systems to allow for a more comprehensive perspective of the behaviors that amphibians exhibit. Finally, we uncover knowledge gaps and suggest new avenues of research using a variety of disciplines and approaches that will allow us to better understand the function and evolution of parental care behaviors in this diverse group of animals.

Oviposition and father presence reduce clutch cannibalism by female poison frogs.

BACKGROUND: The consumption of conspecific young by adult individuals is a common phenomenon across various animal taxa. Possible adaptive benefits of such behaviour include the acquisition of nutrients, decreased competition for one's own offspring, and/or increased mating opportunities. Clutch cannibalism has occasionally been observed in several species of Neotropical poison frogs, but the circumstances under which this behaviour occurs has rarely been investigated experimentally. Recent experiments with the poison frog Allobates femoralis have shown that males indiscriminately transport all clutches located inside their own territory to bodies of water, but become highly cannibalistic when taking over a new territory. Females are able to indirectly discriminate between their own and foreign clutches by location and take over transport duties of their own clutches only in the absence of the father. Cannibalism by A. femoralis females has not been previously observed. We thus asked if, and under which circumstances, cannibalism of unrelated clutches by female A. femoralis would occur, by manipulating the presence of the clutch's father, the female's own reproductive state, and the female's familiarity with the environment. RESULTS: Females clearly cannibalize foreign clutches. Cannibalism was most pronounced when the female had not recently produced her own clutch and the father of the foreign clutch was absent. The female's familiarity with the area had no significant influence on the likelihood of cannibalism to occur. CONCLUSIONS: Our data indicate that both previous oviposition and the father's presence reduce cannibalistic behaviour in A. femoralis females. Cannibalistic females may gain nutritional benefits or enhanced inclusive fitness by preying on other females' offspring. The finding that the father's presence at the clutch site/territory was sufficient to reduce cannibalism by females suggests a prominent role of male territoriality for the evolution of male parental care.

Attacked ravens flexibly adjust signalling behaviour according to audience composition.

A fundamental attribute of social intelligence is the ability to monitor third-party relationships, which has been repeatedly demonstrated in primates, and recently also in captive ravens. It is yet unknown how ravens make use of this ability when dealing with different types of social relationships simultaneously during complex real-life situations. Free-ranging non-breeder ravens live in societies characterized by high fission-fusion dynamics and structured by age, pair-bond status and kinship. Here, we show that free-ranging ravens modify communication during conflicts according to audience composition. When being attacked by dominant conspecifics, victims of aggression signal their distress via defensive calls. Victims increased call rates when their kin were in the bystander audience, but reduced call rates when the bystanders were bonding partners of their aggressors. Hence, ravens use social knowledge flexibly and probably based on their own need (i.e. alert nearby allies and avoid alerting nearby rivals).

Hierarchical decision-making balances current and future reproductive success.

Parental decisions in animals are often context-dependent and shaped by fitness trade-offs between parents and offspring. For example, the selection of breeding habitats can considerably impact the fitness of both offspring and parents, and therefore, parents should carefully weigh the costs and benefits of available options for their current and future reproductive success. Here, we show that resource-use preferences are shaped by a trade-off between parental effort and offspring safety in a tadpole-transporting frog. In a large-scale in situ experiment, we investigated decision strategies across an entire population of poison frogs that distribute their tadpoles across multiple water bodies. Pool use followed a dynamic and sequential selection process, and transportation became more efficient over time. Our results point to a complex suite of environmental variables that are considered during offspring deposition, which necessitates a highly dynamic and flexible decision-making process in tadpole-transporting frogs.

Calls during agonistic interactions vary with arousal and raise audience attention in ravens.

BACKGROUND: Acoustic properties of vocalizations can vary with the internal state of the caller, and may serve as reliable indicators for a caller's emotional state, for example to prevent conflicts. Thus, individuals may associate distinct characteristics in acoustic signals of conspecifics with specific social contexts, and adjust their behaviour accordingly to prevent escalation of conflicts. Common ravens (Corvus corax) crowd-forage with individuals of different age classes, sex, and rank, assemble at feeding sites, and engage in agonistic interactions of varying intensity. Attacked individuals frequently utter defensive calls in order to appease the aggressor. Here, we investigated if acoustic properties of defensive calls change with varying levels of aggression, and if bystanders respond to these changes. RESULTS: Individuals were more likely to utter defensive calls when the attack involved contact aggression, and when the attacker was higher in rank than the victim. Defensive calls produced during intense conflicts were longer and uttered at higher rates, and showed higher fundamental frequency- and amplitude-related measures than calls uttered during low-intensity aggression, indicating arousal-based changes in defensive calls. Playback experiments showed that ravens were more likely to react in response to defensive calls with higher fundamental frequency by orientating towards the speakers as compared to original calls and calls manipulated in duration. CONCLUSIONS: Arousal-based changes are encoded in acoustic parameters of defensive calls in attacked ravens, and bystanders in the audience pay attention to the degree of arousal in attacked conspecifics. Our findings imply that common ravens can regulate conflicts with conspecifics by means of vocalizations, and are able to gather social knowledge from conspecific calls.

Induced parental care in a poison frog: a tadpole cross-fostering experiment.

Understanding the external stimuli and natural contexts that elicit complex behaviours, such as parental care, is key in linking behavioural mechanisms to their real-life function. Poison frogs provide obligate parental care by shuttling their tadpoles from terrestrial clutches to aquatic nurseries, but little is known about the proximate mechanisms that control these behaviours. In this study, we used Allobates femoralis, a poison frog with predominantly male parental care, to investigate whether tadpole transport can be induced in both sexes by transferring unrelated tadpoles to the backs of adults in the field. Specifically, we asked whether the presence of tadpoles on an adult's back can override the decision-making rules preceding tadpole pick-up and induce the recall of spatial memory necessary for finding tadpole deposition sites. We used telemetry to facilitate accurate tracking of individual frogs and spatial analysis to compare movement trajectories. All tested individuals transported their foster-tadpoles to water pools outside their home area. Contrary to our expectation, we found no sex difference in the likelihood to transport or in the spatial accuracy of finding tadpole deposition sites. We reveal that a stereotypical cascade of parental behaviours that naturally involves sex-specific offspring recognition strategies and the use of spatial memory can be manipulated by experimental placement of unrelated tadpoles on adult frogs. As individuals remained inside their home area when only the jelly from tadpole-containing clutches was brushed on the back, we speculate that tactile rather than chemical stimuli trigger these parental behaviours.

Low reproductive skew despite high male-biased operational sex ratio in a glass frog with paternal care.

BACKGROUND: Reproductive skew, the uneven distribution of reproductive success among individuals, is a common feature of many animal populations. Several scenarios have been proposed to favour either high or low levels of reproductive skew. Particularly a male-biased operational sex ratio and the asynchronous arrival of females is expected to cause high variation in reproductive success among males. Recently it has been suggested that the type of benefits provided by males (fixed vs. dilutable) could also strongly impact individual mating patterns, and thereby affecting reproductive skew. We tested this hypothesis in Hyalinobatrachium valerioi, a Neotropical glass frog with prolonged breeding and paternal care. RESULTS: We monitored and genetically sampled a natural population in southwestern Costa Rica during the breeding season in 2012 and performed parentage analysis of adult frogs and tadpoles to investigate individual mating frequencies, possible mating preferences, and estimate reproductive skew in males and females. We identified a polygamous mating system, where high proportions of males (69 %) and females (94 %) reproduced successfully. The variance in male mating success could largely be attributed to differences in time spent calling at the reproductive site, but not to body size or relatedness. Female H. valerioi were not choosy and mated indiscriminately with available males. CONCLUSIONS: Our findings support the hypothesis that dilutable male benefits - such as parental care - can favour female polyandry and maintain low levels of reproductive skew among males within a population, even in the presence of direct male-male competition and a highly male-biased operational sex ratio. We hypothesize that low male reproductive skew might be a general characteristic in prolonged breeders with paternal care.

Odor cues rather than personality affect tadpole deposition in a neotropical poison frog.

Animals constantly need to evaluate available external and internal information to make appropriate decisions. Identifying, assessing, and acting on relevant cues in contexts such as mate choice, intra-sexual competition, and parental care is particularly important for optimizing individual reproductive success. Several factors can influence decision-making, such as external environmental cues and the animal's own internal state, yet, we have limited knowledge on how animals integrate available information. Here, we used an entire island population (57 males, 53 females, and 1,109 tadpoles) of the neotropical brilliant-thighed poison frog Allobates femoralis to investigate how 2 factors (olfactory cues and personality traits) influence the ability of males to find and use new resources for tadpole deposition. We experimentally manipulated the location of tadpole deposition sites and their associated olfactory cues, and repeatedly measured exploration and boldness in adult males. We further reconstructed tadpole deposition choices via inferred parent-offspring relationships of adult frogs and tadpoles deposited in our experimental pools using molecular parentage analysis. We found that the discovery and use of new rearing sites were heavily influenced by olfactory cues; however, we did not find an effect of the measured behavioral traits on resource discovery and use. We conclude that in highly dynamic environments such as tropical rainforests, reliable external cues likely take priority over personality traits, helping individuals to discover and make use of reproductive resources.

Odor cues rather than personality affect tadpole deposition in a neotropical poison frog.

Animals constantly need to evaluate available external and internal information to make appropriate decisions. Identifying, assessing, and acting on relevant cues in contexts such as mate choice, intra-sexual competition, and parental care is particularly important for optimizing individual reproductive success. Several factors can influence decision-making, such as external environmental cues and the animal's own internal state, yet, we have limited knowledge on how animals integrate available information. Here, we used an entire island population (57 males, 53 females, and 1,109 tadpoles) of the neotropical brilliant-thighed poison frog Allobates femoralis to investigate how 2 factors (olfactory cues and personality traits) influence the ability of males to find and use new resources for tadpole deposition. We experimentally manipulated the location of tadpole deposition sites and their associated olfactory cues, and repeatedly measured exploration and boldness in adult males. We further reconstructed tadpole deposition choices via inferred parent-offspring relationships of adult frogs and tadpoles deposited in our experimental pools using molecular parentage analysis. We found that the discovery and use of new rearing sites were heavily influenced by olfactory cues; however, we did not find an effect of the measured behavioral traits on resource discovery and use. We conclude that in highly dynamic environments such as tropical rainforests, reliable external cues likely take priority over personality traits, helping individuals to discover and make use of reproductive resources.