Temporal dynamics and fitness consequences of coalition formation in male primates.

Coalition formation is one of the most striking forms of cooperation found in animals. Yet, there is substantial variation between taxa regarding the mechanisms by which coalitions can result in fitness consequences. Here, we investigate the influence of coalitions on dominance rank trajectories and subsequently on reproductive success in wild male crested macaques (Macaca nigra) at Tangkoko Nature Reserve (Sulawesi, Indonesia). We observed 128 coalition events involving 28 males and tested how a variety of coalition properties and factors related to the social environment influenced future male rank. We further used genetic paternity analysis of 19 infants conceived during the study to assess male reproductive success. Our results show that males participating in coalitions achieved higher-than-expected future ranks, while coalition targets had lower-than-expected future ranks. Additionally, all-up coalitions had stronger effects on rank than all-down and bridging coalitions, and these were modulated by the relative strength of coalition partners versus targets. Finally, higher ranking males were more likely to sire infants than lower ranking males. These results provide important insights regarding the mechanisms underlying coalition formation and support the idea that one major path by which coalitions can affect fitness is through influencing male dominance trajectories.

Morphological variants of silent bared-teeth displays have different social interaction outcomes in crested macaques (Macaca nigra).

OBJECTIVES: While it has been demonstrated that even subtle variation in human facial expressions can lead to significant changes in the meaning and function of expressions, relatively few studies have examined primate facial expressions using similarly objective and rigorous analysis. Construction of primate facial expression repertoires may, therefore, be oversimplified, with expressions often arbitrarily pooled and/or split into subjective pigeonholes. Our objective is to assess whether subtle variation in primate facial expressions is linked to variation in function, and hence to inform future attempts to quantify complexity of facial communication. MATERIALS AND METHODS: We used Macaque Facial Action Coding System, an anatomically based and hence more objective tool, to quantify "silent bared-teeth" (SBT) expressions produced by wild crested macaques engaging in spontaneous behavior, and utilized discriminant analysis and bootstrapping analysis to look for morphological differences between SBT produced in four different contexts, defined by the outcome of interactions: Affiliation, Copulation, Play, and Submission. RESULTS: We found that SBT produced in these contexts could be distinguished at significantly above-chance rates, indicating that the expressions produced in these four contexts differ morphologically. We identified the specific facial movements that were typically used in each context, and found that the variability and intensity of facial movements also varied between contexts. DISCUSSION: These results indicate that nonhuman primate facial expressions share the human characteristic of exhibiting meaningful subtle differences. Complexity of facial communication may not be accurately represented simply by building repertoires of distinct expressions, so further work should attempt to take this subtle variability into account.

Assessing female reproductive status of spectral tarsier (Tarsius tarsier) using fecal steroid hormone metabolite analysis.

The wild population of spectral tarsier is declining and attempts to breed the species in captivity have been of limited success. One possible reason for this is that information on the reproductive biology of Tarsius tarsier is extremely limited and data on the species reproductive physiology are completely lacking. We validated fecal estrogen (E-total) and progesterone metabolite (5-P-3OH) measurements for monitoring female ovarian activity and pregnancy. We used this approach to provide the first data on cycle and pregnancy length based on endocrine information in this species. We collected regular fecal samples in combination with observations on socio-sexual behaviors for a maximum of 15 months from three females maintained at Primate Research Center of Bogor Agricultural University, Indonesia. Hormonal profiles indicated that behavioral estrus was associated with marked elevations in fecal E-total concentrations followed by increases in 5-P-3OH levels indicating luteal function. Pregnancy was characterized by low levels of E-total and 5-P-3OH during the first month and markedly rising concentrations thereafter. An ovarian cycle length of 21.7 ± 5.7 days was found. Gestation length was 128d (live infant), 131d (stillbirth), and 164d (death of mother and infant due to dystocia). Despite the small sample size, the study demonstrates the overall validity of fecal sex hormone metabolite measurements for reproductive monitoring in female T. tarsier, as such, the methods described here may ultimately help to improve the breeding management of the species in captivity. They may also offer new opportunities for investigating basic questions of tarsier reproductive biology in the wild by using fecal hormone metabolite analysis to diagnose pregnant animals and determine reproductive rates in relation to ecological and other factors influencing tarsier reproduction. Thus, non-invasive assessment of female reproductive condition as described here may ultimately contribute to facilitate in and ex situ conservation efforts of this endangered primate species.

Sexual Behaviour of the Spectral Tarsier (Tarsius spectrum) in Captivity.

Tarsius spectrum is a primate species endemic to Sulawesi. Populations of the species have decreased due to habitat destruction and hunting. The sexual behaviour of T. bancanus and T. syrichta are known, but that of T. spectrum has not been reported until recently. The aim of this research was to study the sexual behaviour of T. spectrum in captivity. We observed 3 pairs of T. spectrum at the captive breeding facility of the IPB Primate Research Centre for 9 months using focal animal sampling. We showed that principal courtship behaviours were scent marking (36.7%) and genital marking for females (16.2%) and genital inspection for males (16.0%). Copulations lasted between 3 and 4 min, starting with the male mounting the female and thrusting quickly as many as 168-236 times followed by slow thrusting 9-20 times. When slow thrusting occurred, females vocalized up to 6 times. At the end of the copulation sequence, males remained motionless with their penis inserted within the female's genitalia for about 31 s. Copulation occurred only once for each pair during the observation period. Our results should be useful to support breeding programmes and conservation actions for tarsiers.

Is social dispersal stressful? A study in male crested macaques (Macaca nigra).

In gregarious species, dispersal events represent one of the most dramatic changes in social life and environment an animal will experience during life due to increased predation risk, aggression from unfamiliar conspecifics and the lack of social support. However, little is known about how individuals respond physiologically to dispersal and whether this process is stressful for the individuals involved. We therefore studied the physiological stress response during dispersal in the crested macaque, a primate species in which males often change groups. Over a period of 14months and 14 dispersal events in 4 groups, we determined faecal glucocorticoid metabolite (FGCM) levels during the process of immigration into a new group and examined a variety of factors (e.g. male age, rank achieved, number of males in the group) potentially affecting FGCM levels during this process. We found that FGCM levels were significantly elevated in the first few days upon immigration, after which levels returned quickly to baseline. FGCM response levels upon immigration were significantly and positively influenced by the number of males in the group. The rank a male achieved upon immigration, aggression received, as well as the proximity to other males did not significantly influence FGCM levels. Our data confirm previous findings on other species demonstrating that in crested macaques immigration into a new social group is associated with an acute endocrine stress response. However, given that stress hormone levels remained elevated only for a short period of time, we do not expect males to experience high physiological costs during immigration. Given our limited knowledge on the physiological responses to dispersal in animals, this study contributes to our understanding of dispersal more generally, and particularly inter-individual differences in the stress response and the potential physiological costs associated with these.

Alpha male replacements and delayed dispersal in crested macaques (Macaca nigra).

In species with a high male reproductive skew, competition between males for the top dominant position is high and escalated fights are common between competitors. As a consequence, challenges incur potentially high costs. Selection should favor males who time an alpha male challenge to maximize chances of a successful outcome minimizing costs. Despite the importance of alpha male replacements for individual males, we know little about the timing of challenges and the condition of the challenger. We investigated the timing and process of alpha male replacements in a species living in multi-male groups with high male reproductive skew, the crested macaque. We studied four wild groups over 6 years in the Tangkoko Reserve, North Sulawesi, Indonesia, during which 16 alpha male replacements occurred. Although unusual for cercopithecines, male crested macaques delayed their natal dispersal until they attained maximum body mass and therefore fighting ability whereupon they emigrated and challenged the alpha male in another group. Accordingly, all observed alpha male replacements were from outside males. Ours is the first report of such a pattern in a primate species living in multi-male groups. Although the majority of alpha male replacements occurred through direct male-male challenges, many also took place opportunistically (i.e., after the alpha male had already been injured or had left the group). Furthermore, alpha male tenures were very short (averaging ca. 12 months). We hypothesize that this unusual pattern of alpha male replacements in crested macaques is related to the species-specific combination of high male reproductive skew with a large number of males per group. Am. J. Primatol. 79:e22448, 2017. © 2015 The Authors. American Journal of Primatology Published by Wiley Periodicals, Inc.

Endocrinological correlates of male bimaturism in wild Bornean orangutans.

Among primates, orangutans are unique in having pronounced male bimaturism leading to two fully adult morphs that differ in both physical appearance and behavior. While unflanged males have a female-like appearance, flanged males have the full suite of secondary sexual characteristics, including cheek flanges and a large throat sac. So far, hormonal correlates of arrested development in unflanged males and the expression of secondary sexual characteristics in flanged males have only been studied in zoo-housed individuals. In this study, we investigated fecal androgen and glucocorticoid metabolites as hormonal correlates of male bimaturism in 17 wild adult Bornean orangutans (Pongo pygmaeus) in Central Kalimantan, Indonesia. We predicted and found higher androgen levels in flanged males compared to unflanged males, probably due to ongoing strong competition among flanged males who meet too infrequently to establish a clear linear dominance hierarchy. Furthermore, we found no difference in fecal glucocorticoid metabolite concentrations between flanged and unflanged males, indicating that social stress is unlikely to explain arrested development in unflanged wild orangutans. The only actively developing male in our study showed significantly higher androgen levels during the period of development than later as a fully flanged male. This supports earlier findings from zoo studies that elevated androgen levels are associated with the development of secondary sexual characteristics.

Costs of mate-guarding in wild male long-tailed macaques (Macaca fascicularis): physiological stress and aggression.

Mate-guarding is an important determinant of male reproductive success in a number of species. However, it is known to potentially incur costs. The aim of the present study was to assess the effect of mate-guarding on male physiological stress and aggression in long-tailed macaques, a species in which males mate-guard females to a lesser extent than predicted by the Priority of Access model (PoA). The study was carried out during two mating periods on three groups of wild long-tailed macaques in Indonesia by combining behavioral observations with non-invasive measurements of fecal glucocorticoid (fGC) levels. Mate-guarding was associated with a general rise in male stress hormone levels but, from a certain threshold of mate-guarding onwards, increased vigilance time was associated with a decrease in stress hormone output. Mate-guarding also increased male-male aggression rate and male vigilance time. Overall, alpha males were more physiologically stressed than other males independently of mating competition. Increased glucocorticoid levels during mate-guarding are most likely adaptive since it may help males to mobilize extra-energy required for mate-guarding and ultimately maintain a balanced energetic status. However, repeated exposure to high levels of stress over an extended period is potentially deleterious to the immune system and thus may carry costs. This potential physiological cost together with the cost of increased aggression mate-guarding male face may limit the male's ability to mate-guard females, explaining the deviance from the PoA model observed in long-tailed macaques. Comparing our results to previous findings we discuss how ecological factors, reproductive seasonality and rank achievement may modulate the extent to which costs of mate-guarding limit male monopolization abilities.

Bovine nucleoprotein transitions genes and protein abundance as valuable markers of sperm motility and the correlation with fertility.

Bovine nucleoprotein transitions (TNPs), specifically TNP1 and TNP2, are essential molecules in sperm nucleus rich in arginine and lysine. These molecules act in the phase between histone expulsion and before incorporation of protamine in the spermatid nucleus. Therefore, this study aimed to analyze genes and protein abundance of TNP1 and TNP2 in sperm to determine the potential as motility markers and correlation with fertility in the field. An objective evaluation method, CASA-Sperm Vision, was used to separate 22 bulls into two groups (mg-A and mg-B) based on their increasing motility. Sperm quality parameters were also examined including velocity, mitochondrial membrane potential (MMP) by the JC-1 method, head defects using William staining, and DNA fragmentation by Halomax. TNPs genes abundance was performed using the RT-qPCR method, and the protein abundance was examined with the EIA approach. The fertility rate was also analyzed based on the conception rate generated from each bull in the field, with the data obtained from iSIKHNAS. The results showed that TNPs genes and protein abundance were significantly higher (P < 0.05) in mg-A compared to mg-B, followed by various sperm quality parameters and fertility rates (P < 0.05). Positive correlations were found in TNPs genes and protein abundance with motility, velocity, MMP, and fertility (P < 0.01). Meanwhile, a negative correlation (P < 0.01) was found between head defects and DNA fragmentation. These results showed the potential of TNPs as sperm motility markers and bull fertility.

Comparative developmental competence of in vitro embryos recovered from Bali cattle with normal and poor sperm motility.

Background and Aim: Fertility is crucial for enhancing the efficiency of livestock production, as it directly impacts the reproductive rates. A comprehensive understanding of the relationship between sperm quality and embryo development is key to optimizing reproductive outcomes and improving the quality of livestock. This study analyzed the developmental competence of in vitro embryos recovered from Bali cattle with normal or poor sperm motility. Materials and Methods: Nine bulls with normal fresh semen (NFS) or poor fresh semen (PFS) motility were ejaculated for semen. Semen ejaculates, including volume, motility, and sperm concentration, were evaluated immediately after collection to measure the quality of the fresh semen. Frozen semen was evaluated using computer-assisted semen analysis (CASA) for motility, progressive sperm motility, distance curve path, distance curve linear, distance straight line, average path velocity, curvilinear velocity, linear velocity, straightness (STR), linearity of forward progression (LIN), wobble, and average lateral head displacement (ALH). Bull groups were used to determine in vitro embryo cleavage ability after fertilization of Bali cattle. Ovaries of Bali cattle were collected by slicing, and only cytoplasmic oocytes with compact cumulus cells were used in this study. The oocytes were matured, and in vitro fertilization was performed using fertilization media with a final sperm concentration of 1.5 × 106 spermatozoa/mL. After 48 h, the embryo cleavage ability of the cultured oocytes was evaluated. Results: There were significant differences in motility values between the NFS and PFS groups; however, there were no significant differences in the volume or sperm concentration. There was a significant difference in the LIN value between the groups but no significant differences in other CASA parameters. There were no significant differences in the cleavage rate and morula between the groups, but a positive correlation was observed between the cleavage rate and the morula and between the morula and ALH. A significant negative correlation was observed between the cleavage rate and STR and between the morula and STR; no significant differences were observed for other variables. Conclusion: Despite variations in sperm characteristics, both normal and poor sperm motility demonstrated comparable in vitro embryonic development competence. These findings provide important insights into the fertility potential of Bali bulls, providing valuable information that can enhance selection strategies to improve the quality of livestock production.

Social tolerance in wild female crested macaques (Macaca nigra) in Tangkoko-Batuangus Nature Reserve, Sulawesi, Indonesia.

In primates, females typically drive the evolution of the social system and present a wide diversity of social structures. To understand this diversity, it is necessary to document the consistency and/or flexibility of female social structures across and within species, contexts, and environments. Macaques (Macaca sp.) are an ideal taxon for such comparative study, showing both consistency and variation in their social relations. Their social styles, constituting robust sets of social traits, can be classified in four grades, from despotic to tolerant. However, tolerant species are still understudied, especially in the wild. To foster our understanding of tolerant societies and to assess the validity of the concept of social style, we studied female crested macaques, Macaca nigra, under entirely natural conditions. We assessed their degree of social tolerance by analyzing the frequency, intensity, and distribution of agonistic and affiliative behaviors, their dominance gradient, their bared-teeth display, and their level of conciliatory tendency. We also analyzed previously undocumented behavioral patterns in grade 4 macaques: reaction upon approach and distribution of affiliative behavior across partners. We compared the observed patterns to data from other populations of grade 4 macaques and from species of other grades. Overall, female crested macaques expressed a tolerant social style, with low intensity, frequently bidirectional, and reconciled conflicts. Dominance asymmetry was moderate, associated with an affiliative bared-teeth display. Females greatly tolerated one another in close proximity. The observed patterns matched the profile of other tolerant macaques and were outside the range of patterns of more despotic species. This study is the first comprehensive analysis of females' social behavior in a tolerant macaque species under natural conditions and as such, contributes to a better understanding of macaque societies. It also highlights the relevance of the social style concept in the assessment of the degree of tolerance/despotism in social systems.

Multicomponent and multimodal lipsmacking in crested macaques (Macaca nigra).

Primates' communicative signals are often dynamic and composed of multiple components, sometimes belonging to different sensory modalities. Such multicomponent signals are of crucial importance in the study of communication: the addition of extra features to a signal has the potential to modulate or change the meaning and message of the specific signal. Traditionally, however, components of compound signals are studied in isolation from each other, or communicative displays are studied as static and invariant wholes. Both approaches may not allow us to assess the full function of the signals. In crested macaques (Macaca nigra), the lipsmack (a display mainly used in affiliative interactions) can be produced alone or combined with other visual and acoustic communicative features. We investigated whether the composition of the lipsmack influenced the outcome of social interaction while controlling for relationship quality. Our results show that lipsmacks composed of both visual and vocal components (i.e. multimodal signals) increased the probability of affiliative contact, and therefore have enhanced signal value. The total number of visual components involved in the display had no effect but some visual components seemed more influential than others. By analyzing lipsmacking behavior as a composite, dynamic display, we were able to reveal a level of complexity that is not apparent when looking at each component separately. The results highlight the importance of a more integrative, multimodal approach to the study of primate communication.

Ten Years of Positive Impact of a Conservation Education Program on Children's Knowledge and Behaviour Toward Crested Macaques (Macaca nigra) in the Greater Tangkoko Area, North Sulawesi, Indonesia.

In areas where primates are threatened, environmental education interventions are a key way to increase the local population's knowledge of their environment and encourage positive attitudes and habits to preserve the environment and wildlife on a local and global scale. This study assesses the impact of Tangkoko Conservation Education (TCE), the Macaca Nigra Project's conservation education programme, running since 2011 in North Sulawesi for school children, teachers, and the local population. TCE's goal is to help people increase their knowledge of their environment and develop more positive habits and behaviours towards their local environment, especially the Critically Endangered and endemic crested macaques (Macaca nigra). We measured the efficacy of TCE's programme for pupils using questionnaires provided one month before and one month after a year-long conservation education programme at school. Pupils' knowledge and behaviour scores increased significantly after their participation in the programme. Their habits score also increased but this increase was not statistically significant. Girls scored significantly higher than boys in terms of positive behaviour towards wildlife. Children participating in the programme more than once seemed to obtain higher scores in their second participation, although the sample size was too small for formal analysis. Despite some limitations, this study demonstrates the positive impact of our programme on young people living in an area where primates are threatened. We hope that this research will inspire similar programmes in Sulawesi and elsewhere by providing methods and activities to help prevent primate extinction. Supplementary Information: The online version contains supplementary material available at 10.1007/s10764-023-00356-9.

Effects of Tourist and Researcher Presence on Fecal Glucocorticoid Metabolite Levels in Wild, Habituated Sulawesi Crested Macaques (Macaca nigra).

Ecotourism managers and researchers often assume that apparently habituated primate groups no longer experience adverse consequences of prolonged exposure to tourists or researchers. We examined the effects of tourists and researchers on fecal glucocorticoid metabolite output (FGCM) in three critically endangered, wild crested macaque (Macaca nigra) groups in Tangkoko Nature Reserve, Sulawesi, Indonesia. We assayed FGCM from 456 fecal samples collected from thirty-three adults. Tourists can walk through and among macaque groups freely. Hence, we examined the possible effects of tourists both (1) in the reserve when away and not interacting with the study groups and (2) when they were present within the macaque groups. Generalized Linear Mixed Model (GLMM) analysis indicated that when tourists were present in the forest, but not directly among the macaques, FGCM levels in the macaque tourism groups were higher in months with more tourists. When tourists were among the macaque groups, some female macaques experienced rises and subsequent postexposure decreases in FGCM levels, consistent with predictions for acute stress. Male FGCM levels increased with tourist numbers within the group. Nevertheless, they were not significantly different from levels during undisturbed or postexposure conditions. FGCM responses related to researchers in groups varied by group, sex, and tourist presence. However, the temporal patterning of FGCM responses showed little evidence of chronic stress from tourism at this site.

Sperm Head Morphology Alterations Associated with Chromatin Instability and Lack of Protamine Abundance in Frozen-Thawed Sperm of Indonesian Local Bulls.

This study aimed to analyze various alterations in the morphology of the sperm head and its association with nucleus instability and insufficient sperm protamine. Frozen-thawed semen from twenty local Indonesian bulls was used for all stages in this study. The results of sperm head defect assessments are used for bull grouping, high (HD) and low (LD). Sperm DNA damage was assessed using Acridine Orange and Halomax. The PRM1 protein abundance was carried out using an enzyme immunoassay, while PRM1 gene expression was carried out using the RT-qPCR. PRM deficiency was performed using CMA3. Several kinds of sperm head defects in the HD were significantly higher (p < 0.05) than in the LD bulls. Sperm DNA damage showed a significant (p < 0.05) difference between the HD and LD bulls. PRM1 abundance was significantly (p < 0.05) decreased in HD bulls. PRM deficiency was significantly (p < 0.05) higher in HD bulls than in LD bulls. PRM deficiency in bulls correlated significantly (p < 0.01) with sperm head defects, DNA damage, and PRM1 abundance. The lack of sperm protamine might affect the sperm nucleus's stability and induce morphological alterations in the sperm head.

Correlation between Post-Thaw Spermatozoa Quality of the Endangered Javan Banteng with OPN Gene Expression.

The role of ex situ conservation facilities or captivity through captive breeding programs is essential in the conservation of the endangered Javan banteng. The development of semen cryopreservation may assist on one side of the conservation plan. However, the male Javan banteng reproductive capability must be considered as it influences the targeted outputs. Studying the potential biomarker for fertility such as osteopontin gene expression is also expected to help predict male fertility. Therefore, this study aimed to analyze the quality of spermatozoa after thawing to help predict the male reproductive capability of Javan banteng. Furthermore, this study investigated the potential role of osteopontin gene expression in male Javan banteng fertility. A positive reinforcement approach was used to accustom the male and female animals as we focused on establishing a collection procedure using neither sedation nor anaesthesia. Semen samples were collected at Taman Safari Indonesia, Bogor, in accordance with the female banteng receptivity. Semen samples were then evaluated and then cryopreserved under field conditions. Our study showed the different predicted reproductive capability of the Javan banteng based on the post-thaw spermatozoa quality, which showed significant differences. The OPN gene showed positive correlations with the progressive motility (r = 0.711, p = 0.048), viability (r = 0.822, p = 0.012), and acrosomal integrity (r = 0.665, p = 0.072) of Javan banteng spermatozoa after thawing. Our study demonstrated the predicted Javan banteng reproductive capability based on various post-thaw spermatozoa variables. This finding is also the first report on the OPN gene potential to be developed as the assessment tool of post-thaw spermatozoa quality of the male Javan banteng. The findings in our study may help give recommendations for future breeding programs, especially in the ex situ conservation sites.

Sexual signalling in female crested macaques and the evolution of primate fertility signals.

BACKGROUND: Female signals of fertility have evolved in diverse taxa. Among the most interesting study systems are those of multimale multifemale group-living primates, where females signal fertility to males through multiple signals, and in which there is substantial inter-specific variation in the composition and reliability of such signals. Among the macaques, some species display reliable behavioural and/or anogenital signals while others do not. One cause of this variation may be differences in male competitive regimes: some species show marked sexual dimorphism and reproductive skew, with males fighting for dominance, while others show low dimorphism and skew, with males queuing for dominance. As such, there is variation in the extent to which rank is a reliable proxy for male competitiveness, which may affect the extent to which it is in females' interest to signal ovulation reliably. However, data on ovulatory signals are absent from species at one end of the macaque continuum, where selection has led to high sexual dimorphism and male reproductive skew. Here we present data from 31 cycles of 19 wild female crested macaques, a highly sexually dimorphic species with strong mating skew. We collected measures of ovarian hormone data from faeces, sexual swelling size from digital images, and male and female behaviour. RESULTS: We show that both sexual swelling size and female proceptivity are graded-signals, but relatively reliable indicators of ovulation, with swelling size largest and female proceptive behaviours most frequent around ovulation. Sexual swelling size was also larger in conceptive cycles. Male mating behaviour was well timed to female ovulation, suggesting that males had accurate information about this. CONCLUSION: Though probabilistic, crested macaque ovulatory signals are relatively reliable. We argue that in species where males fight over dominance, male dominance rank is surrogate for competitiveness. Under these circumstances it is in the interest of females to increase paternity concentration and assurance in dominants beyond levels seen in species where such competition is less marked. As such, we suggest that it may in part be variation in male competitive regimes that leads to the evolution of fertility signalling systems of different reliability.

Social bonds affect anti-predator behaviour in a tolerant species of macaque, Macaca nigra.

Enduring positive social bonds between individuals are crucial for humans' health and well being. Similar bonds can be found in a wide range of taxa, revealing the evolutionary origins of humans' social bonds. Evidence suggests that these strong social bonds can function to buffer the negative effects of living in groups, but it is not known whether they also function to minimize predation risk. Here, we show that crested macaques (Macaca nigra) react more strongly to playbacks of recruitment alarm calls (i.e. calls signalling the presence of a predator and eliciting cooperative mobbing behaviour) if they were produced by an individual with whom they share a strong social bond. Dominance relationships between caller and listener had no effect on the reaction of the listener. Thus, strong social bonds may improve the coordination and efficiency of cooperative defence against predators, and therefore increase chances of survival. This result broadens our understanding of the evolution and function of social bonds by highlighting their importance in the anti-predator context.

Crested macaque facial movements are more intense and stereotyped in potentially risky social interactions.

Ambiguity in communicative signals may lead to misunderstandings and thus reduce the effectiveness of communication, especially in unpredictable interactions such as between closely matched rivals or those with a weak social bond. Therefore, signals used in these circumstances should be less ambiguous, more stereotyped and more intense. To test this prediction, we measured facial movements of crested macaques (Macaca nigra) during spontaneous social interaction, using the Facial Action Coding System for macaques (MaqFACS). We used linear mixed models to assess whether facial movement intensity and variability varied according to the interaction outcome, the individuals' dominance relationship and their social bond. Movements were least intense and most variable in affiliative contexts, and more intense in interactions between individuals who were closely matched in terms of dominance rating. We found no effect of social bond strength. Our findings provide evidence for a reduction in ambiguity of facial behaviour in risky social situations but do not demonstrate any mitigating effect of social relationship quality. The results indicate that the ability to modify communicative signals may play an important role in navigating complex primate social interactions. This article is part of the theme issue 'Cognition, communication and social bonds in primates'.

PRM1 Gene Expression and Its Protein Abundance in Frozen-Thawed Spermatozoa as Potential Fertility Markers in Breeding Bulls.

Functional genes and proteins in sperm play an essential role in bulls' reproductive processes. They are more accurate in determining bull fertility than conventional semen quality tests. Protamine-1 (PRM1) is a gene or protein crucial for packaging and protecting sperm DNA until fertilization affects normal sperm function. This study analyzes the genes and proteins potential from PRM1 as fertility markers for different breeds of bulls utilized in the artificial insemination programs, expected to be an accurate tool in interpreting bull fertility in Indonesia. This study used Limousin, Holstein, and Ongole Grade bulls divided into two groups based on fertility, high-fertility (HF) and low fertility (LF). The semen quality assessment included progressive motility (computer-assisted semen analysis), viability (eosin-nigrosine), and plasma membrane integrity (HOS test). Sperm DNA fragmentation (SDF) was assessed using the acridine orange staining and the Halomax test. Sperm PRM deficiency was evaluated with the chromomycin A3 method. Moreover, PRM1 gene expression was measured using qRT-PCR, and the PRM1 protein abundance was measured with the enzyme immunoassay method. Semen quality values, relative expression of PRM1 gene, and quantity of PRM1 protein were significantly higher (p < 0.05) in HF bulls than in LF bulls. The SDF and PRM deficiency values in LF bulls were significantly higher (p < 0.05) than HF bulls. Additionally, PRM1 at the gene and protein levels correlated significantly (p < 0.01) with fertility. Therefore, PRM1 is a potential candidate for fertility markers in bulls in Indonesia.