Growing up together or apart: Rearing experience effects on opposite-sex sibling discrimination in Chilean brush tailed mouse (Octodon degus)

Growing up together or apart: Rearing experience effects on opposite-sex sibling discrimination in Chilean brush tailed mouse (Octodon degus) | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Growing up together or apart: Rearing experience effects on opposite-sex sibling discrimination in Chilean brush tailed mouse (Octodon degus) Natalia I. Márquez, Camila P. Villavicencio, Jaime Martínez-Harms, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7725339/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 12 Apr, 2026 Read the published version in Journal of Comparative Physiology A → Version 1 posted 9 You are reading this latest preprint version Abstract Kin recognition refers to the discrimination and subsequent behavioral bias toward close relatives. Kin recognition is an important modulator of social interaction. Among relatives, reduced exploration, lower aggressiveness, and decreased inbreeding are typically expected. Two main mechanisms influence kin discrimination: prior association (familiarity) and genetic relatedness. In this study, we investigated kin discrimination between opposite-sex siblings in degus ( Octodon degus ), a diurnal, communally nesting rodent endemic to central Chile. While in this species discrimination among same-sex siblings has been shown to be mainly influenced by rearing experience, it remains unknown whether degus can discriminate their opposite-sex siblings at all, and if so, what mechanisms are at play. We used a cross-fostering protocol to experimentally separate the effects of familiarity and genetic relatedness, producing four groups that combined siblings and non-siblings reared together or apart. Behavior was assessed in two experimental setups: (i) a y-maze to test olfactory preferences and (ii) a dyadic encounter arena to evaluate social interactions. In the y-maze, each experimental subject was simultaneously exposed to one sibling and one non-sibling (stimuli pair, SP), both of the opposite sex. Exploration increased significantly when the subject had no rearing experience with either SP member. Females explored more than males the y-maze. Dyadic encounter experiments showed that rearing experience influenced exploratory and agonistic interactions but not sexual behaviors. Social contact, on the other hand, was more frequent among siblings. Taken together, our results indicate that rearing experience plays a major role in modulating opposite-sex social behaviors in this species. Kin discrimination sibling discrimination Octodon degus opposite sex rearing experience cross fostering Figures Figure 1 Figure 2 Figure 3 Introduction Kin recognition refers to the ability of animals to discriminate between close relatives from unrelated conspecifics, often reflected in behavioral biases toward kin (Hepper 1991 ). Some authors distinguish between kin recognition and kin discrimination , the former describes the cognitive mechanism allowing animals to classify conspecifics as kin, and the latter represents the behavioral bias toward their kin (see Tang-Martinez 2001 ). Considering this difference, it is worth noting that most studies on kin recognition have focused on the behavioral aspect of the problem, thus from a kin discrimination perspective. Kin discrimination can be manifested in different forms: siblings can be less aggressive among them (Holmes and Sherman 1982 ), explore each other less (Holmes 1984 ; Villavicencio et al. 2009 ), perform alarm calls more frequently when close relatives form part of the social group (Sherman 1977 ), or perform affiliative interactions which are crucial for the social cohesion of the group (e.g. association, communal feeding and playing) (Wey and Blumstein 2010 ). Additionally, low levels of inbreeding in natural or captive populations -such as those reported in black-tailed prairie dogs (Hoogland 1982 ), spiral-horned antelopes (Apio et al. 2010 ), meadow voles (Bollinger et al. 1991 ), and mice (Sherborne et al. 2007 )- have also been associated with behavioral kin bias. These examples show the biological context in which kin discrimination is a significant aspect of mammalian sociality. Studies on kin discrimination mechanisms have revealed cases where individuals display behavioral biases toward conspecifics based on prior association rather than true kin recognition (see Mateo 2003 ), highlighting the need to separate genetic relatedness from prior association experiences when studying the mechanisms underlying this phenomenon. To address this issue cross-fostering protocols have been successfully implemented (Holmes 1984 ; Mateo and Holmes 2004 ) leading to the proposal of two main mechanisms: (i) recognition by prior association (i.e. familiarity), where common living rather than genetic similarity accounts for behavioral biases, and (ii) recognition by phenotype matching, where genetically close individual sharing specific cues (e.g. olfactory signals) are treated as kin (Heth et al. 1998 ; Holmes 1984 , 1986 ; Holmes and Sherman 1982 ; Paz y Miño and Tang-Martinez 1999b ; Tang-Martinez 2001 ). It is worth noting that even when cross-fostering is crucial to separate the effects of prior association and genetic relatedness, it fails to account for learning during early development inside the womb (Robinson and Smotherman 1991 ). Olfaction seems to be the predominant sense mediating kin recognition in rodents (Halpin 1991 ; Holmes 1984 ; Sherborne et al. 2007 ). Rodents produce different sources of olfactory signals (e.g. body scent, urine) that can mediate mating, individual identification, and kin discrimination (Hare 1998 ; Hurst et al. 2001 ; Johnston 2003 ). Evidence suggests that some of these signals are genetically determined, while others may be influenced by environmental factors such as diet (Beauchamp et al. 1988 ; Green et al. 2015 ; Radwan et al. 2008 ; Yamazaki et al. 1988 ). Furthermore, prenatal olfactory experience can shape the connectivity of the olfactory system (Liu et al. 2016 ); however, the extent to which these processes affect kin recognition has not been fully explored. Taken together, these characteristics make rodents valuable models to study their olfactory discrimination abilities and olfactory preferences in the context of social interactions and kin discrimination, especially in social rodents. In contrast to the well-studied infraorders Myomorpha (mouse-like rodents) and Sciuromorpha (squirrel like rodents) from the Northern hemisphere (Mateo 2003 ; Tang-Martinez 2003 ), only few studies have focused on the Hystricognathi infraorder, in particular on the Caviomorpha family (cavia like rodents) (See Jesseau et al. 2008 ; Márquez et al. 2015 ; Villavicencio et al. 2009 ). One of the few studies on hystricognath rodents from East African shows that kin discrimination and female mate choice in naked mole-rats ( Heterocephalus glaber ) are based solely on familiarity through prior association (Clarke and Faulkes 1999 ). To compensate for the underrepresentation of this group, we used the South American rodent degus ( Octodon degus ) as model to study kin discrimination. Degus are diurnal caviomorph rodents, endemic from central Chile. They are a highly social species that displays different kinds of social behaviors, including group foraging, group vigilance (Vásquez 1997 ; Vásquez et al. 2002 ), communal nesting (Ebensperger et al. 2004 ; Fulk 1976 ; Jesseau et al. 2008 ), and kin discrimination (Jesseau et al. 2008 ; Márquez et al. 2015 ; Villavicencio et al. 2009 ). Degus’ social system consists of one or two adult males living with two to six females, which might or might not be genetically related (Ebensperger et al. 2009 ; Ebensperger et al. 2004 ; Fulk 1976 ; Quirici et al. 2011 ). The mating system in degus is predominantly polygynandry, often with multiple paternity occurring within litters (Ebensperger et al. 2019 ). Degus are considered semifossorial and have a complex burrow system where communal nesting takes place (Ebensperger et al. 2019 ; Ebensperger et al. 2004 ), although many of their activities take place above ground (Ebensperger et al. 2002 ; Fulk 1976 ; Kenagy et al. 2002 ). Youngs remain underground during the communal nesting period, when they are around 1 month old they start going out the burrow and become active above the ground (Ebensperger et al. 2004 ). Unlike northern hemisphere species with short gestation period that produce altricial offspring, degus have long gestational period (three months), and their pups are precocial (Long and Ebensperger 2010 ; Reynolds and Wright 1979 ), born fully furred and open their eyes at postnatal day 1 (PN1). These characteristics may play an important role in kin discrimination in this species. Olfactory learning during gestation, together with early experiences acquired in communal nesting, likely contribute to this process. Together, these features make degus an excellent model for studying kin discrimination in greater depth. Here, we assessed the behavioral mechanisms underlying kin discrimination among opposite sex siblings in degus. Male-female interactions carry a different social connotation compared to same-sex behaviors. In degus, kin discrimination between same sex siblings is strongly mediated by prior experiences (Villavicencio et al. 2009 ). Furthermore, early olfactory experiences are essential for the development of the behavioral biases observed in adulthood (Márquez et al. 2015 ). However, to the best of our knowledge, male-female sibling discrimination on degus, or on any other Caviomorph rodent, has not yet been explored. By implementing the cross-fostering strategy (Mateo and Holmes 2004 ), we sought to determine whether degus exhibit behavioral biases toward kin of the opposite sex based on prior association (i.e. familiarity) or genetically determined phenotypic similarities (i.e. phenotype matching). Two different behavioral assays using sexually mature degus during the reproductive period were performed to i) characterize olfactory preferences toward conspecifics of the opposite sex in a y-maze and ii) quantify social behaviors during dyadic encounters between male and female degus that differed in prior association and/or genetic relatedness. If kin discrimination between degus of opposite sex is similar to kin discrimination in siblings of the same sex, a major effect of prior association on exploration, aggression, and sexual behaviors toward conspecifics would be expected (Halpin 1986 ; Holmes 1984 ; Johnston 1993 ; Villavicencio et al. 2009 ). Alternatively, if kin discrimination between siblings of the opposite sex is mainly driven by genetically related olfactory cues, no influence of prior association on the above-mentioned behaviors will be observed. Material and Methods Subjects Adult Octodon degus were captured and maintained as described by Villavicencio et al. ( 2009 ). To assure coordination of females’ estrus, we used wild animals captured during summer (January and February), from three populations in central Chile: la Campana (32º 55’S, 71º 05’W), Lampa (33º 17’S, 70º 53’W) and Rinconada de Maipú (33º 29’S, 70º 53’W). Within each population animals were captured using Sherman live traps from different sites separated at least 400 m to assure the collection of non-related females (see Ebensperger et al. 2004 ). Once at the University of Chile they were housed in metal cages (50 x 40 x 35 cm) with wood shavings in a climatized room under natural photoperiod in groups of 3 to 4 degus of the same sex. Animals were fed with rabbit pellet and alfalfa and provided with water ad libitum . Males and females were kept separated for four months’ prior the mating protocol (see below) to ensure using females that were not pregnant by the time they were captured. All procedures of capture, maintenance, and experimentation were approved by the ethics committee of the Faculty of Sciences of the University of Chile and followed Chilean regulations on wildlife management. Animal capture was authorized by the Chilean Agricultural and Livestock Service (SAG) Nr. 5193. Mating design To assess the role of genetic relatedness and/or prior association on kin discrimination, we implemented a cross-fostering protocol (see below). Wild degus mate during fall (May-June), undergo a three month gestation period and litters are born in early spring (September) (Ebensperger et al. 2004 ; Fulk 1976 ; Quispe et al. 2014 ). This seasonality in reproduction can be maintained during the first years of captivity by keeping the natural photoperiod (Villavicencio et al. 2009 ). We set up 16 mating groups during the reproductive period (May-June), consisting of one male and three females from the same population, but different capture sites, to assure the pregnancy of at least one female and the availability of half-siblings. Parturition was inspected daily after a 3-month gestation period, any female that gave birth was housed separately with her litter. This enabled us to later manipulate litter members and create cross-fostered groups (see below). Cross fostering Newly-born degus were cross-fostered within a time window from the day they were born (P0) to postnatal day 1 (P1) (see Villavicencio et al. 2009 ). Within this window period, when the number of pups was even, we took half of the littler members from one female and exchanged them with the pups from another litter. In the case of odd number of pups, we took half of the litter plus one. We always maintained the sex proportions of the litters. All pups were accepted by their foster mothers. For permanent identification each pup was marked with an eartag (National Band & Tag Co., Newport, KY, U.S.A). Through this cross-fostering protocol, we produced 14 litters where animals were housed and grew up with the nursing female, the sibs and the genetically unrelated conspecifics from early ontogeny (P0-P1) through adulthood, until experiments were performed. Experimental groups As a result of the mating group design and the cross fostering manipulation, we could create five experimental groups that differed in familiarity (i.e. animals reared together or apart) and in genetic relatedness (i.e. siblings or non-siblings): (1) siblings reared together (S.RT), (2) siblings reared apart (S.RA), (3) non-siblings reared together (NS.RT), (4) non-siblings reared apart (NS.RA), and (5) half-siblings reared apart (HS.RA). Each group consisted of 7 males and 7 females. Considering that cross-fostering cannot exclude the effects of fetal learning (Robinson and Smotherman 1991 ), the HS.RA group is relevant to account for genetic relatedness effects. Degus reach sexual maturity at 6 weeks of age (Lee 2004 ), and are considered adults at 6 month old (Davis 1975 ). Male-female sibling discrimination was assessed in sexually mature adult degus (7 to 8-month-old) during the breeding period (May-June). We performed two kinds of behavioral experiments: (i) olfactory exploration in a y-maze labyrinth where animals could show olfactory preferences toward two opposite sex conspecifics and (ii) pair encounters in an experimental arena which allows social interactions. Therefore, both experiments were conducted during degus’ reproductive season. Experimental degus participated in both experiments; however, we made sure that all individuals were tested only once with a particular conspecific to avoid any familiarity effect due to experience during the experiments. Degus are diurnal rodents, with activity peaks in the field occurring during daylight hours (Fulk 1976 ; Kenagy et al. 2002 ), therefore all experiments were performed under artificial light conditions during the day. Y-maze Social preferences and olfactory exploration of social conspecifics were assessed in a y-maze labyrinth (see Márquez et al. 2015 , Fig. 1 for a picture). The y-maze consisted of three Plexiglas arms forming angles of 120°, each arm was 50 cm long x 15 cm width x 15 cm high. At the end of each arm there was a chamber where a degu was placed. Two chambers (stimuli chambers) were designated to the conspecifics used as stimuli. These chambers had a perforated Plexiglas plate that prevented direct contact between the focal individual and the stimuli, but allowed the airflow produced by a fan behind each stimulus chamber to circulate throughout the labyrinth. The third chamber (start chamber) was used to keep the focal individual restrained before the start of the experiment. Two degus of the opposite sex respect to the focal individual were used as olfactory stimuli (hereafter stimuli pair, SP). Each degu from a SP was placed inside their respective stimulus chamber. The SPs were always a sib with a non-sib of the focal individual, that differed on the rearing condition (i.e. reared together or reared apart). Thus, we create four combinations of stimuli pairs: (1) sibling reared together (S.RT) versus non-sibling reared together (NS.RT), (2) S.RT versus non-sibling reared apart (NS.RA), (3) sibling reared apart (S.RA) versus NS.RT and (4) S.RA versus NS.RA. Focal individuals (males and females) were tested only once in the y-maze with only one kind of the four SPs ( N = 56; 7 females and 7 males per SP). The experiment consisted of 1 min of acclimatization and 3 min of testing time. This experimental period was chosen following Johnston et al. ( 1997 )d rquez et al. ( 2015 ). During the acclimatization time, Plexiglas plates (block plates) were placed in front of each the three chamber, preventing the movement of the focal individual and preventing volatiles from the stimulus chambers to reach the focal individual at the start chamber. After the acclimatation time, once the experiment began, the block plates were removed and the fans behind the stimulus chambers were switched on. At this point, focal individuals were able to freely explore the whole labyrinth. To control for any preference for a specific side of the labyrinth, the position of the sib and non-sib was randomized between experiments. No such preference was found (Paired t test: t (d.f :83) = -0.97, P = 0.33). To avoid scent contamination between experiments, we used different sets of gloves to manipulate animals in the maze, and disposable gloves to clean the y-maze after each experiment using 95° Ethanol. All experiments were video recorded (color CCTV camera connected to a Sony video recorder) from above the y-maze . A single observer blind to the treatment and to the subjects analyzed the video recordings using the JWatcher 1.0 software (Dan Blumstein, University of California, Los Angeles, U.S.A). Social preferences were evaluated by quantifying the frequency and time of the following behaviors: a) arm exploration: animals walk from the start chamber and enters an arm, while walking and sniffing; b) direct smelling of the division plate: animals reach the stimulus chamber and actively smell the plate that separates them from the stimulus animal, and c) standstill in each arm: animal enter a particular arm and remain still. In addition, we assessed the exploration of social conspecifics by comparing the total exploration of the y-maze between the four combinations of SPs. Y-maze statistics To evaluate social preference toward a particular conspecific in the y-maze we compared, for each experimental group, exploration time and frequency of the described behaviors using the paired t -test (Sokal and Rohlf 2012 ). On the other hand, to evaluate the exploration of social conspecifics, we analyzed the total exploration of the y -maze (stimuli arms) with SPs and sex as factors using a two-way ANOVA test, followed by a Tukey’s honestly significant difference (HSD) post-hoc test (Sokal and Rohlf 2012 ). For all the analysis we used the software R (R-Core-Team 2020 ) with the nortest package (Gross and Ligges 2015 ) to test for normality. Pair encounters Behavioral discrimination was assessed in dyadic encounter experiments of male-female pairs that differed in rearing experience and/or genetic relatedness. Combining rearing experiences (reared together or reared apart) and genetic relatedness (sib or non-sib) we formed four experimental groups of degus of the opposite sex in a balanced model: (1) siblings reared together (S.RT), (2) siblings reared apart (S.RA), (3) non-siblings reared together (NS.RT) and (4) non-siblings reared apart (NS.RA). To control for prenatal learning between cross-fostered siblings, a fifth group of half-siblings reared apart (HS.RA) was used for statistical comparison when a genetic relatedness effect was found. Animals were carried in individual plastic cages from the housing rooms to the experimental room. The experimental arenas consisted of a 80x80x50 cm metal arenas that was divided by a division plate (see Márquez et al. 2015 ; Villavicencio et al. 2009 ), that allowed degus to acclimate before the beginning of the experiment. The floor of the arena was a removable white-painted metal plate that was cleaned with detergent between experiments to remove any trace of scent. Seven pairs of animals were tested for each group (total pairs tested = 35). During each experiment one of the animals was painted with non-toxic paint in the back for individualization. We found no effects of marking on exploratory (Kruskal-Wallis test: H 1 = 0.59, P = 0.44), agonistic (Kruskal-Wallis test: H 1 = 0.046, P = 0.83) or any other measured behavior, in agreement with previous studies in degus (Márquez et al. 2015 ; Vásquez et al. 2002 ; Villavicencio et al. 2009 ). After a 10 min acclimatization period, the division plate was removed. The quantification of a 20 min trial began when any of the two experimental subjects started to explore the arena. If subjects did not interact and remained still for more than 5 min, the test was repeated on a different day (8 of 35 pairs). If the tested pairs did not interact in the second attempt (5 pairs), they were not considered in the analysis. All experiments were video recorded (color CCTV camera connected to a Sony video recorder) from above the arena. An observer blind to the treatment and animals’ sex analyzed the different behaviors using the JWatcher 1.0 software. Four behavioral categories were defined as social exploration, social contact, agonistic and sexual behavior, based on behavioral descriptions of intraspecific interactions in degus (see Fulk 1976 ; Kleiman 1975 ; Márquez et al. 2015 ; Wilson and Kleiman 1974 ), prairie vole (Paz y Miño and Tang-Martinez 1999a) and mice (Baudoin et al. 1991 ). (1) Social exploratory behavior, consisted of olfactory approaches to the nose/mouth, and/or anogenital area of its partner. (2) Social contact was considered when degus were in contact either side by side, on right angle to each other, grooming, or huddling one over the other. (3) Agonistic encounters were considered when degus showed either (i) evasive behaviors: an experimental subject avoided the partner in hostile contexts by turning aside or running away, or (ii) aggressive behavior: when the experimental subject performed hostile confrontation by tail wagging, hindleg kick, foreleg push, chasing or fighting against its partner. (4) Sexual behaviors were evaluated separately for males and females: (i) male sexual behaviors were defined whenever a male mounted the female and/or when attempted to mount the female, (ii) female sexual behaviors were defined when the female remained passive under the male during mounting. Due to our experimental setup, mating and copulation were difficult to quantified, therefore, not assessed. Pair encounters statistics The effects of rearing experience and genetic relatedness were assessed for the four behavioral categories described above. Social exploration was analyzed using a two-way ANOVA after square-root transformation to accomplish parametric assumptions. The other three behavioral categories, social contact, agonistic and sexual behavior , did not accomplish parametric assumptions, even after being transformed, therefore we used the Scheirer-Ray-Hare test (extension of the Kruskal-Wallis test) (Sokal and Rohlf 2012 ). We found no effect of sex on the mentioned behaviors; therefore, sex was not included as a factor. However, sexual behaviors between males and females differ (see Table 1 ), thus, they were analyzed separately. For behaviors affected by genetic relatedness, we added an extra group to control for fetal learning: the half-siblings reared apart group (HS.RA). Therefore, in those cases, we compared the three sibling groups (siblings reared together, sibling reared apart, and half-siblings reared apart) using the Kruskal-Wallis test. All statistical analysis were performed using the software R (R-Core-Team 2020 ) along with the nortest package (Gross and Ligges 2015 ) to test for normality. Table 1 Behavioral categories for social interaction quantified during dyadic encounters between degus adult pairs of the opposite sex*. Social interaction categories for degus of the opposite sex Social-olfactory exploration Nose/mouth Anogenital Both animals approach and sniff the nose/mouth area. One animal sniffs the anogenital area. Social contact Side by side Grooming Huddling Both animals are in close contact with each other either side-by-side or at a right angle. One animal grooms the other one. One animal lies on top of the other one on close contact. Agonistic (i) Evasive (ii) Aggressive Tail wagging Fight One animal avoids the other by turning aside or run away. Hostile confrontation. Movement of the tail from side to side at ground level. One animal gives a hindleg kick, foreleg push, chases or attacks the other. Sexual behaviors (i) Male (ii) Female Male mounts or attempts to mount the female. Female remains passive when the male is mounting or attempts mounting her. *Modified from (Baudoin et al. 1991 ; Fulk 1976 ; Kleiman 1975 ; Márquez et al. 2015 ; Paz y Miño and Tang-Martinez 1999a; Wilson and Kleiman 1974 ) Results Y-maze Degus social preferences toward kin were assessed by measuring arm visit frequency and social-olfactory exploration time in the y-maze. Focal individuals were always exposed to an opposite sex sib and to an opposite sex non-sib that differed on the rearing conditions (reared together or reared apart). We found no significant differences in the arms visit frequency, nor in the time of social exploration toward any of the individuals conforming the stimuli pair (SP) irrespectively of the genetic relatedness or rearing condition (see Table 2 for statistics). Table 2 In the y-maze focal degus (N = 56) showed no social preference toward opposite sex sibs or non-sibs irrespectively of the rearing conditions. Arm visit frequency Social-olfactory exploration time (s) r = 0.5 M (SE) r = 0 M (SE) t P N r = 0.5 M (SE) r = 0 M (SE) t P Stimuli pair S.RT vs. NS.RT 13.9 (1.1) 13.6 (1.2) -0.135 0.89 14 44.2 (4.2) 51.2 (3.6) 1.25 0. 24 S.RT vs. NS.RA 12.3 (1.4) 14.9 (1.4) 1.26 0.23 14 57 (5.4) 41 (4) -1.90 0.08 S.RA vs. NS.RT 19.1 (1.6) 20.2 (1.8) 0.299 0.77 14 63 (5.9) 62.2 (5.3) 0.055 0.96 S.RA vs. NS.RA 26.8 (2.1) 26.4 (2.2) 0.225 0.83 14 67.8 (5.2) 61.9 (4) -0.894 0.39 Data expressed as mean frequency and mean time (standard error), Mean (SE). t indicates Paired t- test statistic (d.f. = 13). r is the relatedness coefficient (Wright 1922 ), r = 0.5 stands for siblings and r = 0 for subjects that are not genetically related. Stimuli pairs : siblings reared together (S.RT) versus non-siblings rear together (NS.RT); S.RT versus non-siblings rear apart (NS.RA); siblings reared apart (S.RA) versus NS.RT; S.RA versus NS.RA. However, we found that the total visits of both arms were affected by the kind of stimuli pair presented (ANOVA: F [3,48] = 4.77, P = 0.0055). Focal individuals exposed to a stimuli pair with both unfamiliar conspecifics (S.RA vs. NS.RA, see methods or Table 2 for nomenclature), visited significantly more both arms of the y-maze compared to focal individuals exposed to two familiarized degus (S.RT/NS.RT; post hoc Tukey’s HSD test, P = 0.012) and to the S.RT/NS.RA pair (post hoc Tukey’s HSD test, P = 0.011). Degus exposed to a S.RA/NS.RT pair did not differ statistically to any other group (p > 0.05, Fig. 1 A). In a similar way, we found that the social exploratory frequency of the division plates was affected by the kind of SP presented (ANOVA: F [3,48] = 4.72, P = 0.0058). Focal degus exposed to both unfamiliar conspecifics (S.RA/NS.RA) explored significantly more the division plates than degus exposed to conspecifics reared together (S.RT/NS.RT) (post hoc Tukey’s HSD test, P = 0.008) and compared to animals exposed to S.RT/NS.RA stimuli pairs (post hoc Tukey’s HSD test, P = 0.018). Degus exposed to S.RA/NS.RT did not differ in the social exploration of the division plate to any other experimental group (Fig. 1 B). In addition, we found that females socially explored significantly longer both stimuli pairs than males (ANOVA: F [1,48] = 5.43, P = 0.024). In summary, degus socially explored unfamiliar pairs more frequently than pairs that included a sibling with previous rearing experience. Dyadic encounters We further assessed opposite sex sibling discrimination quantifying social interactions in an experimental arena. Social interactions were considered as any behavior involving both degus. These behaviors were further divided into four categories (see Table 1 ): social-olfactory exploration, social contact, agonistic encounters and sexual behaviors. Nose/mouth social-olfactory exploration time was significantly lower in pairs of opposite sex reared together than unfamiliar pairs with no previous contact (ANOVA: F [1, 40] = 6.243, P = 0.017; Fig. 2 A). We did not observe any effects of genetic relatedness (ANOVA: F [1, 40] = 0.071, P = 0.79) nor of the interaction between familiarity and genetic relatedness (ANOVA: F [1, 40] = 0.44, P = 0.51) on this behavior. In addition, male-female pairs reared apart explored each other’s anogenital area significantly more than pairs reared together (ANOVA: F [1, 40] = 5.72, P = 0.02; Fig. 2 B). We found no effect of genetic relatedness (ANOVA: F [1, 40] = 0.032, P = 0.86), nor an effect of the interaction between the two factors (ANOVA: F [1, 40] = 0.68, P = 0.41) in this behavior. We found that tail wagging, an agonistic behavior, was displayed significantly more by unfamiliar pairs (S.RA and NS.RA; Scheirer-Ray-Hare test: H 1 = 5.18, P = 0.023; Fig. 2 C), irrespectively of their degree of genetic relatedness (Scheirer-Ray-Hare test: H 2 = 0.036, P = 0.84) and we found no interaction effects (Scheirer-Ray-Hare test: H 1 = 0.84, P = 0.36). On the other hand, we rarely observed fights, 11% (only in 3 of 28 couples), and therefore we did not consider it in the statistical analysis. We observed sexual behaviors in all four experimental groups. However, as males have different sexual behaviors than females (see Table 1 ), statistical analyses were done for males and females separately, which yield to a reduced sample sizes per group (N = 5). We found no effect of familiarity (males: Scheirer-Ray-Hare test: H 2 = 1.2, P = 0.3; females: Scheirer-Ray-Hare test: H 2 = 0.05, P = 0.8), genetic relatedness (males: Scheirer-Ray-Hare test: H 2 = 0.2, P = 0.7; females: Scheirer-Ray-Hare test: H 2 = 0.4, P = 0.5), nor the interaction of these independent variables (males: Scheirer-Ray-Hare test: H 2 = 0.03, P = 0.9; females: Scheirer-Ray-Hare test: H 2 = 0.03, P = 0.9) on the frequency of sexual behaviors performed. In addition, no differences were found in the time of sexual behaviors performance (data not shown). We found that social contact (see Table 1 ) was the only behavior affected by genetic relatedness (Scheirer-Ray-Hare test: H 2 = 4.22, P = 0.04) and not by familiarity (Scheirer-Ray-Hare test: H 1 = 0.19, P = 0.66), without an interaction effect (Scheirer-Ray-Hare test: H 1 = 1.29, P = 0.26; Fig. 3 ). In addition, we compared both sibling groups (S.RT and S.RA) with the half-sibling reared apart (HS.RA) experimental group, to further assess the possible effect of fetal learning on social contact behaviors among genetically related individuals. We found no significant difference on social contact behaviors between these three groups (Kruskal-Wallis test: H 1 = 1.13, P = 0.57). In summary, we found that rearing experience explains a wide range of social interactions in adult degus of the opposite sex. However, we found an effect of genetic relatedness on the social contact behavior. Discussion The present study aimed to investigate the behavioral mechanisms underlying kin discrimination between opposite sex siblings in degus. To evaluate the influence of both rearing experience and genetic relatedness on this social behavior, we used a cross-fostering protocol and conducted two behavioral tests: a Y-maze and dyadic interaction trials. Consistent with previous findings in same sex sibling degus, we found that prior association in both behavioral experiments had a strong effect on kin discrimination between opposite sex siblings. Yet, we detected an effect of genetic relatedness in one behavior during dyadic interactions (e.g. social contact). In addition, we found that females explored more than males when exposed to opposite sex conspecifics in the y-maze . Y-maze Our results showed that male and female degus did not show a preference for either of the two conspecifics of the opposite sex presented on the y-maze , regardless of the genetic relatedness or the rearing experience they shared with them. In fact, degus explored both conspecifics similarly. Even though no signs of preference for either conspecific were found, the previous experience with the stimuli pairs affected the total number exploration events toward both conspecifics of the opposite sex. Considering that olfactory exploration can be interpret as indicator of interest in an odor source (Johnston et al. 1997 ), and that degus could explore the labyrinth freely, our findings suggests that degus were more interested in the pairs of conspecifics of the opposite sex that were unknown to them. In agreement with previous studies on olfactory discriminations in degus and other species (Halpin and Hoffman 1987 ; Holmes 1984 ; Johnston 1993 ; Villavicencio et al. 2009 ), our results show that individuals explored familiar olfactory cues less frequently than novel ones, indicating that degus can recognize olfactory cues they have previously encountered. Based on this, it would be expected that the group exposed to two unfamiliarized degus (S.RA vs NS.RA) explored significantly more than both groups exposed to one familiarized and one non-familiarized conspecific (S.RA vs NS.RT and S.RT vs NS.RA). However, this was not the case. Instead, social exploratory behavior in the S.RA/NS.RA group differed significantly only with the group where the experimental subject had been reared with its sibling (S.RT/ NS.RA) [see Fig. 1 ]. This result shows that the presence of a sib reared together was sufficient to reduce social exploratory behaviors, resembling the responses observed when both conspecifics were “known” (S.RT/NS.RT), suggesting a synergistic effect of familiarity and genetic relatedness in this context. The y-maze experiment also revealed sex differences in social exploratory behavior, with females exploring the social stimuli -two opposite sex conspecifics- significantly more than males. These findings are consistent with studies in other rodent species across different contexts. For instance, females mound-building mice ( Mus spicilegus ) explore novel object more than males (Simeonovska-Nikolova 2016 ). In rats ( Rattus norvegicus ), exploratory behavior develops during adolescence in both open field and elevated plus maze paradigms, with females showing higher levels of exploration than males (Lynn and Brown 2009 ). This sex difference is also evident in social contexts, where females explore novel social arenas more than males (Cavigelli et al. 2011 ). In degus, different bioassays have yielded contrasting results regarding sex difference in social exploration. For example, females explore more than males in experiments assessing sex differences during social interactions between non-genetically related conspecifics (Fischer et al. 1986 ). However, a sibling discrimination study found no sex differences in olfactory exploration between same-sex siblings during dyadic encounters in degus (Villavicencio et al. 2009 ). Another study addressing the influence of early olfactory experience reported that males explored more than females in same-sex dyadic encounters when exposed to a conspecific impregnated with an artificial odorant (Márquez et al. 2015 ). In our study, sex differences in exploratory behaviors were observed only in the y-maze experiment, but not during dyadic encounter (see below). We propose that social exploratory behaviors in degus is context-dependent and may vary between sexes depending on the social context and the nature of the olfactory stimuli (i.e. direct contact with conspecifics, urine marks, bedding, etc.). Dyadic encounters Most social behaviors observed during male-female dyadic encounters were determined by rearing experience rather than genetic relatedness per se . Social-olfactory exploration time (including mouth/nose and anogenital investigation) was significantly higher between unfamiliar individuals than between degus reared together. In addition, degus reared together performed significantly fewer agonistic behaviors compared to non-familiarized pairs, indicating a behavioral bias based on rearing experience. Social contact was the only behavior affected by genetic relatedness and not rearing experience, suggesting that not all male-female social interactions are equally affected by prior association. Our results confirm earlier findings in same sex degus (Villavicencio et al. 2009 ), showing that opposite sex sibling discrimination is predominantly mediated by rearing experience, and to a lesser extent by genetic relatedness. Sexual behaviors, on the other hand, are expressed differently between males and females and therefore were analyzed separately. Males mount or attempt to mount females, while females remain passive during these interactions. We found no effect of rearing experience or genetic relatedness on sexual behaviors. Considering that separating male and female behaviors reduces the sample size for statistical analysis, to confirm this result it may be necessary to increase the sample size. In addition, our study specifically focused on sexual behaviors observed in the experimental arena, rather than on mating per se. In line with our results, a field study in degus reported that one female’s offspring can be sired by one to six males, who may or may not be genetically related (Ebensperger et al. 2019 ). Thus, litters can exhibit multiple paternity, suggesting that in degus sociality inbreeding avoidance is not an issue. Kin discrimination mechanisms and olfaction Studies on the behavioral mechanisms underlying kin discrimination (prior association and/or phenotype matching) have revealed that while in some species rearing experience plays a major role in kin discrimination (e.g. thirteen-lined ground squirrels (Holmes 1984 ), and prairie voles (Paz y Miño and Tang-Martinez 1999b )) in others would be less relevant. Mateo ( 2003 ) reviewed the mechanisms of kin recognition in rodents an found that 12 out of 32 species can recognize their genetically close conspecifics based on genetically related cues (Mateo 2003 ). Interestingly, this type of behavioral discrimination is not correlated with the social system, nor with the species’ mating system (Mateo 2003 ). Tang-Martinez ( 2001 ) in a deeper analysis of kin recognition mechanisms proposed that both behavioral mechanisms are indeed the result of learning that might occur at different moments. In this line, cross-fostering, cannot rule out intra-uterine olfactory learning (Robinson and Smotherman 1991 ). Prenatal olfactory learning occurs and can determine flavor and food preferences in mice and rats (Kamenetzky et al. 2018 ). In the context of kin discrimination, prenatal olfactory learning has not been tested. Nevertheless, it is possible that the mother and wombmates’ olfactory cues present in the intrauterine environment can be learned, which would account for kin discrimination without prior association (Robinson and Smotherman 1991 ). Giving the developmental characteristics of degus with a long gestation period (90 days) and a precocial offspring (Weir 1970 ), intra-uterine olfactory learning could be a relevant process for kin discrimination. Further studies are needed to test this hypothesis. In rodent species, kin discrimination in its various forms (i.e. exploration, aggressiveness, affiliative behaviors, or inbreeding avoidance) is mediated by olfactory cues (Halpin 1991 ; Holmes 1984 ; Sherborne et al. 2007 ). Furthermore, the olfactory identity of individuals can be determined by the expression of major histocompatibility complex (MHC) genes (Beauchamp et al. 1988 ; Yamazaki et al. 1988 ) and/or by the presence of major urinary proteins (MUPs) (Cheetham et al. 2007 ; Green et al. 2015 ; Hurst et al. 2001 ). Kin discrimination in degus is mediated by olfactory cues (Jesseau et al. 2008 ; Márquez et al. 2015 ; Villavicencio et al. 2009 ) present in littermates and learned during early ontogeny within the nest context (Márquez et al. 2015 ). Although the sources of olfactory identity in degus remain unknown, behavioral data show that cross-fostered degus can discriminate the scent of a sibling from that of a non-sibling from its litter, but not that of a pair of its siblings from its litter (Villavicencio et al. 2009 ). This suggests that the scent profile of degus is more similar between kin than between non-kin and is therefore possibly influenced by MHC genes and/or MUPs. We propose that these kinds of olfactory signals mediate behavioral biases between closely related adult degus even when reared apart. We consider that the study of male-female socio-sexual interactions is essential for understanding the conservation of a particular way of living, because reproduction is a systemic phenomenon that preserves not only the transmission of genetic material to the offspring, but also the medium in which living takes place (as proposed by Maturana and Mpodozis (2000)). Here, we show that social interactions and kin discrimination between male and female degus are rich and complex and are worth further investigation. Conclusions We confirmed that male-female sibling discrimination in degus is similar to same-sex sibling discrimination; both are mainly driven by rearing experience. We found that some behaviors are mediated by rearing experience (e.g., exploration, agonistic behaviors), others by genetic relatedness (e.g., social contact), and some were not affected by either factor (e.g., sexual behaviors). Thus, focusing solely on a single type of behavior, or only on the ability to distinguish individuals based on olfactory cues, may lead to incomplete or misleading conclusions. Our findings highlight the importance of examining multiple behavioral categories during social encounters to gain a better understanding of the mechanisms underlying kin discrimination. Declarations Author Contribution N.I.M and R.A.V conceived and designed research. N.I.M and C.P.V performed experiments.N.I.M analyzed data and prepared figures.N.I.M and C.P.V drafted manuscript.N.I.M, C.P.V, J.M-H and JM edited and revised manuscript.N.I.M., C.P.V, J.M-H, J.M and R.A.V approved final version of manuscript. Acknowledgments We gratefully acknowledge Dr. Claudia Cecchi, Dr. Daniela Parra, Ronny Zúñiga and Solano Henríquez for their valuable assistance. We would also like to thank the Pilot Scientific Productivity Workshop for their valuable input and motivation. Support for this work was provided by a Programa de Mejoramiento de la Calidad de la Educación Superior (MECESUP UCO-0214) fellowship, and a Comisión Nacional de Investigación Científica y Tecnológica de Chile (CONICYT AT-24050185) grant to N.I.M and by a Fondo Nacional de Desarrollo Científico y Tecnológico (FONDECYT) grant to J.M. (FONDECYT 1250880). Conflict of interest: Authors report no conflict of interest. Data Availability Data will be made available upon reasonable request. 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Cite Share Download PDF Status: Published Journal Publication published 12 Apr, 2026 Read the published version in Journal of Comparative Physiology A → Version 1 posted Editorial decision: Revision requested 20 Nov, 2025 Reviews received at journal 20 Nov, 2025 Reviews received at journal 31 Oct, 2025 Reviewers agreed at journal 13 Oct, 2025 Reviewers agreed at journal 09 Oct, 2025 Reviewers invited by journal 01 Oct, 2025 Editor assigned by journal 27 Sep, 2025 Submission checks completed at journal 27 Sep, 2025 First submitted to journal 26 Sep, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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13:00:20","extension":"html","order_by":10,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":165364,"visible":true,"origin":"","legend":"","description":"","filename":"earlyproof.html","url":"https://assets-eu.researchsquare.com/files/rs-7725339/v1/434f60396529b991fc7d6ccf.html"},{"id":93495199,"identity":"26fcc7a6-a1c0-4db1-bd72-590381a97269","added_by":"auto","created_at":"2025-10-14 13:00:20","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":48276,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003ePrevious\u003c/strong\u003e \u003cstrong\u003erearing experience reduces\u003c/strong\u003e \u003cstrong\u003esocial-olfactory exploration frequency of conspecifics of the opposite sex.\u003c/strong\u003e (\u003cstrong\u003eA\u003c/strong\u003e) Arms visit frequency (mean ± SE). (\u003cstrong\u003eB\u003c/strong\u003e) Social exploratory frequency of both division plates. Focal individuals were exposed to one of the following stimuli pair (see methods): (1) siblings reared together (S.RT) versus non-sibling reared apart (NS.RA), (2) S.RT versus non-sibling reared together (NS.RT), (3) sibling reared apart (S.RA) versus NS.RT and (4) S.RA versus NS.RA. Different letters represent statistically significant differences between stimuli pairs (Tukey’s HSD post hoc test, \u003cem\u003eP \u003c/em\u003e\u0026lt; 0.05, see text for\u003cem\u003e \u003c/em\u003eexact statistical values).\u003c/p\u003e","description":"","filename":"1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7725339/v1/8f707f2e558c2daf1ffc293b.jpg"},{"id":93495200,"identity":"8494820f-a19f-401a-8601-addc89493bfc","added_by":"auto","created_at":"2025-10-14 13:00:20","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":77701,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003ePrevious\u003c/strong\u003e \u003cstrong\u003erearing experience reduces\u003c/strong\u003e \u003cstrong\u003edegus social-olfactory exploration and agonistic behaviors between opposite sex couples during dyadic encounters. (A) \u003c/strong\u003eNose/mouth social-olfactory exploration time\u003cstrong\u003e \u003c/strong\u003ewas significantly longer in degus reared apart than degus reared together.\u003cstrong\u003e (B)\u003c/strong\u003e Anogenital social-olfactory exploration time was significantly longer in in degus reared apart than degus reared together. \u003cstrong\u003e(C) \u003c/strong\u003eTail wagging (agonistic behavior) frequency was performed significantly more by male-female pairs reared apart\u003cstrong\u003e.\u003c/strong\u003e Experimental groups: siblings reared together (S.RT), siblings reared apart (S.RA), non-siblings reared together (NS.RT) and non-siblings reared apart (NS.RA). Asterisk represents \u003cem\u003eP \u003c/em\u003e\u0026lt; 0.05 (See text for exact statistical values).\u003c/p\u003e","description":"","filename":"2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7725339/v1/aa4a7ea25ee3a3f7f13f6960.jpg"},{"id":93495201,"identity":"fea7176d-8c1d-4d14-a5a7-d3bf3489bc13","added_by":"auto","created_at":"2025-10-14 13:00:20","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":39968,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSiblings of the opposite sex showed more social contact behaviors during dyadic encounters than non-sibs. \u003c/strong\u003eMale-female siblings remained together significantly longer than non-sib pairs. Experimental groups: siblings reared together (S.RT), siblings reared apart (S.RA), non-siblings reared together (NS.RT) and non-siblings reared apart (NS.RA). Asterisk represents statistically significant differences between groups (see text for exact statistical values).\u003c/p\u003e","description":"","filename":"3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7725339/v1/a470edde0413d487c541b19f.jpg"},{"id":106809851,"identity":"43e41c02-8db5-49c0-9122-ddc3f5fcf782","added_by":"auto","created_at":"2026-04-13 16:13:11","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1249336,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7725339/v1/721d0137-cd52-4061-becc-d4a15ed51ee9.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"\u003cp\u003eGrowing up together or apart: Rearing experience effects on opposite-sex sibling discrimination in Chilean brush tailed mouse (Octodon degus)\u003c/p\u003e","fulltext":[{"header":"Introduction","content":"\u003cp\u003eKin recognition refers to the ability of animals to discriminate between close relatives from unrelated conspecifics, often reflected in behavioral biases toward kin (Hepper \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e1991\u003c/span\u003e). Some authors distinguish between \u003cem\u003ekin recognition\u003c/em\u003e and \u003cem\u003ekin discrimination\u003c/em\u003e, the former describes the cognitive mechanism allowing animals to classify conspecifics as kin, and the latter represents the behavioral bias toward their kin (see Tang-Martinez \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e2001\u003c/span\u003e). Considering this difference, it is worth noting that most studies on kin recognition have focused on the behavioral aspect of the problem, thus from a kin discrimination perspective. Kin discrimination can be manifested in different forms: siblings can be less aggressive among them (Holmes and Sherman \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e1982\u003c/span\u003e), explore each other less (Holmes \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e1984\u003c/span\u003e; Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e), perform alarm calls more frequently when close relatives form part of the social group (Sherman \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e1977\u003c/span\u003e), or perform affiliative interactions which are crucial for the social cohesion of the group (e.g. association, communal feeding and playing) (Wey and Blumstein \u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). Additionally, low levels of inbreeding in natural or captive populations -such as those reported in black-tailed prairie dogs (Hoogland \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e1982\u003c/span\u003e), spiral-horned antelopes (Apio et al. \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2010\u003c/span\u003e), meadow voles (Bollinger et al. \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e1991\u003c/span\u003e), and mice (Sherborne et al. \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2007\u003c/span\u003e)- have also been associated with behavioral kin bias. These examples show the biological context in which kin discrimination is a significant aspect of mammalian sociality.\u003c/p\u003e\u003cp\u003eStudies on kin discrimination mechanisms have revealed cases where individuals display behavioral biases toward conspecifics based on prior association rather than true kin recognition (see Mateo \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2003\u003c/span\u003e), highlighting the need to separate \u003cem\u003egenetic relatedness\u003c/em\u003e from \u003cem\u003eprior association\u003c/em\u003e experiences when studying the mechanisms underlying this phenomenon. To address this issue cross-fostering protocols have been successfully implemented (Holmes \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e1984\u003c/span\u003e; Mateo and Holmes \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2004\u003c/span\u003e) leading to the proposal of two main mechanisms: (i) recognition by prior association (i.e. familiarity), where common living rather than genetic similarity accounts for behavioral biases, and (ii) recognition by phenotype matching, where genetically close individual sharing specific cues (e.g. olfactory signals) are treated as kin (Heth et al. \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e1998\u003c/span\u003e; Holmes \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e1984\u003c/span\u003e, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e1986\u003c/span\u003e; Holmes and Sherman \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e1982\u003c/span\u003e; Paz y Mi\u0026ntilde;o and Tang-Martinez \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e1999b\u003c/span\u003e; Tang-Martinez \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e2001\u003c/span\u003e). It is worth noting that even when cross-fostering is crucial to separate the effects of prior association and genetic relatedness, it fails to account for learning during early development inside the womb (Robinson and Smotherman \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e1991\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eOlfaction seems to be the predominant sense mediating kin recognition in rodents (Halpin \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e1991\u003c/span\u003e; Holmes \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e1984\u003c/span\u003e; Sherborne et al. \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). Rodents produce different sources of olfactory signals (e.g. body scent, urine) that can mediate mating, individual identification, and kin discrimination (Hare \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e1998\u003c/span\u003e; Hurst et al. \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2001\u003c/span\u003e; Johnston \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2003\u003c/span\u003e). Evidence suggests that some of these signals are genetically determined, while others may be influenced by environmental factors such as diet (Beauchamp et al. \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e1988\u003c/span\u003e; Green et al. \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Radwan et al. \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e2008\u003c/span\u003e; Yamazaki et al. \u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e1988\u003c/span\u003e). Furthermore, prenatal olfactory experience can shape the connectivity of the olfactory system (Liu et al. \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2016\u003c/span\u003e); however, the extent to which these processes affect kin recognition has not been fully explored. Taken together, these characteristics make rodents valuable models to study their olfactory discrimination abilities and olfactory preferences in the context of social interactions and kin discrimination, especially in social rodents.\u003c/p\u003e\u003cp\u003eIn contrast to the well-studied infraorders Myomorpha (mouse-like rodents) and Sciuromorpha (squirrel like rodents) from the Northern hemisphere (Mateo \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2003\u003c/span\u003e; Tang-Martinez \u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e2003\u003c/span\u003e), only few studies have focused on the Hystricognathi infraorder, in particular on the Caviomorpha family (cavia like rodents) (See Jesseau et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2008\u003c/span\u003e; M\u0026aacute;rquez et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). One of the few studies on hystricognath rodents from East African shows that kin discrimination and female mate choice in naked mole-rats (\u003cem\u003eHeterocephalus glaber\u003c/em\u003e) are based solely on familiarity through prior association (Clarke and Faulkes \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e1999\u003c/span\u003e). To compensate for the underrepresentation of this group, we used the South American rodent degus (\u003cem\u003eOctodon degus\u003c/em\u003e) as model to study kin discrimination. Degus are diurnal caviomorph rodents, endemic from central Chile. They are a highly social species that displays different kinds of social behaviors, including group foraging, group vigilance (V\u0026aacute;squez \u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e1997\u003c/span\u003e; V\u0026aacute;squez et al. \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e2002\u003c/span\u003e), communal nesting (Ebensperger et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2004\u003c/span\u003e; Fulk \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e1976\u003c/span\u003e; Jesseau et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2008\u003c/span\u003e), and kin discrimination (Jesseau et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2008\u003c/span\u003e; M\u0026aacute;rquez et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). Degus\u0026rsquo; social system consists of one or two adult males living with two to six females, which might or might not be genetically related (Ebensperger et al. \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2009\u003c/span\u003e; Ebensperger et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2004\u003c/span\u003e; Fulk \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e1976\u003c/span\u003e; Quirici et al. \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). The mating system in degus is predominantly polygynandry, often with multiple paternity occurring within litters (Ebensperger et al. \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Degus are considered semifossorial and have a complex burrow system where communal nesting takes place (Ebensperger et al. \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Ebensperger et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2004\u003c/span\u003e), although many of their activities take place above ground (Ebensperger et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2002\u003c/span\u003e; Fulk \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e1976\u003c/span\u003e; Kenagy et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2002\u003c/span\u003e). Youngs remain underground during the communal nesting period, when they are around 1 month old they start going out the burrow and become active above the ground (Ebensperger et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2004\u003c/span\u003e). Unlike northern hemisphere species with short gestation period that produce altricial offspring, degus have long gestational period (three months), and their pups are precocial (Long and Ebensperger \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; Reynolds and Wright \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e1979\u003c/span\u003e), born fully furred and open their eyes at postnatal day 1 (PN1). These characteristics may play an important role in kin discrimination in this species. Olfactory learning during gestation, together with early experiences acquired in communal nesting, likely contribute to this process. Together, these features make degus an excellent model for studying kin discrimination in greater depth.\u003c/p\u003e\u003cp\u003eHere, we assessed the behavioral mechanisms underlying kin discrimination among opposite sex siblings in degus. Male-female interactions carry a different social connotation compared to same-sex behaviors. In degus, kin discrimination between same sex siblings is strongly mediated by prior experiences (Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). Furthermore, early olfactory experiences are essential for the development of the behavioral biases observed in adulthood (M\u0026aacute;rquez et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). However, to the best of our knowledge, male-female sibling discrimination on degus, or on any other Caviomorph rodent, has not yet been explored. By implementing the cross-fostering strategy (Mateo and Holmes \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2004\u003c/span\u003e), we sought to determine whether degus exhibit behavioral biases toward kin of the opposite sex based on prior association (i.e. familiarity) or genetically determined phenotypic similarities (i.e. phenotype matching). Two different behavioral assays using sexually mature degus during the reproductive period were performed to i) characterize olfactory preferences toward conspecifics of the opposite sex in a \u003cem\u003ey-maze\u003c/em\u003e and ii) quantify social behaviors during dyadic encounters between male and female degus that differed in prior association and/or genetic relatedness. If kin discrimination between degus of opposite sex is similar to kin discrimination in siblings of the same sex, a major effect of prior association on exploration, aggression, and sexual behaviors toward conspecifics would be expected (Halpin \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e1986\u003c/span\u003e; Holmes \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e1984\u003c/span\u003e; Johnston \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e1993\u003c/span\u003e; Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). Alternatively, if kin discrimination between siblings of the opposite sex is mainly driven by genetically related olfactory cues, no influence of prior association on the above-mentioned behaviors will be observed.\u003c/p\u003e"},{"header":"Material and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003eSubjects\u003c/h2\u003e\u003cp\u003eAdult \u003cem\u003eOctodon degus\u003c/em\u003e were captured and maintained as described by Villavicencio et al. (\u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). To assure coordination of females\u0026rsquo; estrus, we used wild animals captured during summer (January and February), from three populations in central Chile: la Campana (32\u0026ordm; 55\u0026rsquo;S, 71\u0026ordm; 05\u0026rsquo;W), Lampa (33\u0026ordm; 17\u0026rsquo;S, 70\u0026ordm; 53\u0026rsquo;W) and Rinconada de Maip\u0026uacute; (33\u0026ordm; 29\u0026rsquo;S, 70\u0026ordm; 53\u0026rsquo;W). Within each population animals were captured using Sherman live traps from different sites separated at least 400 m to assure the collection of non-related females (see Ebensperger et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2004\u003c/span\u003e). Once at the University of Chile they were housed in metal cages (50 x 40 x 35 cm) with wood shavings in a climatized room under natural photoperiod in groups of 3 to 4 degus of the same sex. Animals were fed with rabbit pellet and alfalfa and provided with water \u003cem\u003ead libitum\u003c/em\u003e. Males and females were kept separated for four months\u0026rsquo; prior the mating protocol (see below) to ensure using females that were not pregnant by the time they were captured. All procedures of capture, maintenance, and experimentation were approved by the ethics committee of the Faculty of Sciences of the University of Chile and followed Chilean regulations on wildlife management. Animal capture was authorized by the Chilean Agricultural and Livestock Service (SAG) Nr. 5193.\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eMating design\u003c/h3\u003e\n\u003cp\u003eTo assess the role of genetic relatedness and/or prior association on kin discrimination, we implemented a cross-fostering protocol (see below). Wild degus mate during fall (May-June), undergo a three month gestation period and litters are born in early spring (September) (Ebensperger et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2004\u003c/span\u003e; Fulk \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e1976\u003c/span\u003e; Quispe et al. \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). This seasonality in reproduction can be maintained during the first years of captivity by keeping the natural photoperiod (Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eWe set up 16 mating groups during the reproductive period (May-June), consisting of one male and three females from the same population, but different capture sites, to assure the pregnancy of at least one female and the availability of half-siblings. Parturition was inspected daily after a 3-month gestation period, any female that gave birth was housed separately with her litter. This enabled us to later manipulate litter members and create cross-fostered groups (see below).\u003c/p\u003e\n\u003ch3\u003eCross fostering\u003c/h3\u003e\n\u003cp\u003eNewly-born degus were cross-fostered within a time window from the day they were born (P0) to postnatal day 1 (P1) (see Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). Within this window period, when the number of pups was even, we took half of the littler members from one female and exchanged them with the pups from another litter. In the case of odd number of pups, we took half of the litter plus one. We always maintained the sex proportions of the litters. All pups were accepted by their foster mothers. For permanent identification each pup was marked with an eartag (National Band \u0026amp; Tag Co., Newport, KY, U.S.A). Through this cross-fostering protocol, we produced 14 litters where animals were housed and grew up with the nursing female, the sibs and the genetically unrelated conspecifics from early ontogeny (P0-P1) through adulthood, until experiments were performed.\u003c/p\u003e\n\u003ch3\u003eExperimental groups\u003c/h3\u003e\n\u003cp\u003eAs a result of the mating group design and the cross fostering manipulation, we could create five experimental groups that differed in familiarity (i.e. animals reared together or apart) and in genetic relatedness (i.e. siblings or non-siblings): (1) siblings reared together (S.RT), (2) siblings reared apart (S.RA), (3) non-siblings reared together (NS.RT), (4) non-siblings reared apart (NS.RA), and (5) half-siblings reared apart (HS.RA). Each group consisted of 7 males and 7 females. Considering that cross-fostering cannot exclude the effects of fetal learning (Robinson and Smotherman \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e1991\u003c/span\u003e), the HS.RA group is relevant to account for genetic relatedness effects.\u003c/p\u003e\u003cp\u003eDegus reach sexual maturity at 6 weeks of age (Lee \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2004\u003c/span\u003e), and are considered adults at 6 month old (Davis \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e1975\u003c/span\u003e). Male-female sibling discrimination was assessed in sexually mature adult degus (7 to 8-month-old) during the breeding period (May-June). We performed two kinds of behavioral experiments: (i) olfactory exploration in a \u003cem\u003ey-maze\u003c/em\u003e labyrinth where animals could show olfactory preferences toward two opposite sex conspecifics and (ii) pair encounters in an experimental arena which allows social interactions. Therefore, both experiments were conducted during degus\u0026rsquo; reproductive season. Experimental degus participated in both experiments; however, we made sure that all individuals were tested only once with a particular conspecific to avoid any familiarity effect due to experience during the experiments. Degus are diurnal rodents, with activity peaks in the field occurring during daylight hours (Fulk \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e1976\u003c/span\u003e; Kenagy et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2002\u003c/span\u003e), therefore all experiments were performed under artificial light conditions during the day.\u003c/p\u003e\n\u003ch3\u003eY-maze\u003c/h3\u003e\n\u003cp\u003eSocial preferences and olfactory exploration of social conspecifics were assessed in a \u003cem\u003ey-maze\u003c/em\u003e labyrinth (see M\u0026aacute;rquez et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2015\u003c/span\u003e, Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e for a picture). The \u003cem\u003ey-maze\u003c/em\u003e consisted of three Plexiglas arms forming angles of 120\u0026deg;, each arm was 50 cm long x 15 cm width x 15 cm high.\u003c/p\u003e\u003cp\u003eAt the end of each arm there was a chamber where a degu was placed. Two chambers (stimuli chambers) were designated to the conspecifics used as stimuli. These chambers had a perforated Plexiglas plate that prevented direct contact between the focal individual and the stimuli, but allowed the airflow produced by a fan behind each stimulus chamber to circulate throughout the labyrinth. The third chamber (start chamber) was used to keep the focal individual restrained before the start of the experiment. Two degus of the opposite sex respect to the focal individual were used as olfactory stimuli (hereafter stimuli pair, SP). Each degu from a SP was placed inside their respective stimulus chamber. The SPs were always a sib with a non-sib of the focal individual, that differed on the rearing condition (i.e. reared together or reared apart). Thus, we create four combinations of stimuli pairs: (1) sibling reared together (S.RT) versus non-sibling reared together (NS.RT), (2) S.RT versus non-sibling reared apart (NS.RA), (3) sibling reared apart (S.RA) versus NS.RT and (4) S.RA versus NS.RA. Focal individuals (males and females) were tested only once in the \u003cem\u003ey-maze\u003c/em\u003e with only one kind of the four SPs (\u003cem\u003eN\u003c/em\u003e\u0026thinsp;=\u0026thinsp;56; 7 females and 7 males per SP).\u003c/p\u003e\u003cp\u003eThe experiment consisted of 1 min of acclimatization and 3 min of testing time. This experimental period was chosen following Johnston et al. (\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e1997\u003c/span\u003e)d rquez et al. (\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). During the acclimatization time, Plexiglas plates (block plates) were placed in front of each the three chamber, preventing the movement of the focal individual and preventing volatiles from the stimulus chambers to reach the focal individual at the start chamber. After the acclimatation time, once the experiment began, the block plates were removed and the fans behind the stimulus chambers were switched on. At this point, focal individuals were able to freely explore the whole labyrinth.\u003c/p\u003e\u003cp\u003eTo control for any preference for a specific side of the labyrinth, the position of the sib and non-sib was randomized between experiments. No such preference was found (Paired \u003cem\u003et\u003c/em\u003e test: \u003cem\u003et\u003c/em\u003e\u003csub\u003e\u003cem\u003e(d.f\u003c/em\u003e:83)\u003c/sub\u003e = -0.97, \u003cem\u003eP\u0026thinsp;=\u003c/em\u003e\u0026thinsp;0.33). To avoid scent contamination between experiments, we used different sets of gloves to manipulate animals in the maze, and disposable gloves to clean the \u003cem\u003ey-maze\u003c/em\u003e after each experiment using 95\u0026deg; Ethanol. All experiments were video recorded (color CCTV camera connected to a Sony video recorder) from above the \u003cem\u003ey-maze\u003c/em\u003e. A single observer blind to the treatment and to the subjects analyzed the video recordings using the JWatcher 1.0 software (Dan Blumstein, University of California, Los Angeles, U.S.A). Social preferences were evaluated by quantifying the frequency and time of the following behaviors: a) arm exploration: animals walk from the start chamber and enters an arm, while walking and sniffing; b) direct smelling of the division plate: animals reach the stimulus chamber and actively smell the plate that separates them from the stimulus animal, and c) standstill in each arm: animal enter a particular arm and remain still. In addition, we assessed the exploration of social conspecifics by comparing the total exploration of the \u003cem\u003ey-maze\u003c/em\u003e between the four combinations of SPs.\u003c/p\u003e\u003cp\u003e\u003cb\u003eY-maze\u003c/b\u003e \u003cb\u003estatistics\u003c/b\u003e\u003c/p\u003e\u003cp\u003eTo evaluate social preference toward a particular conspecific in the \u003cem\u003ey-maze\u003c/em\u003e we compared, for each experimental group, exploration time and frequency of the described behaviors using the paired \u003cem\u003et\u003c/em\u003e-test (Sokal and Rohlf \u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e2012\u003c/span\u003e). On the other hand, to evaluate the exploration of social conspecifics, we analyzed the total exploration of the \u003cem\u003ey\u003c/em\u003e-maze (stimuli arms) with SPs and sex as factors using a two-way ANOVA test, followed by a Tukey\u0026rsquo;s honestly significant difference (HSD) post-hoc test (Sokal and Rohlf \u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e2012\u003c/span\u003e). For all the analysis we used the software R (R-Core-Team \u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e2020\u003c/span\u003e) with the nortest package (Gross and Ligges \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2015\u003c/span\u003e) to test for normality.\u003c/p\u003e\u003cdiv id=\"Sec8\" class=\"Section2\"\u003e\u003ch2\u003ePair encounters\u003c/h2\u003e\u003cp\u003eBehavioral discrimination was assessed in dyadic encounter experiments of male-female pairs that differed in rearing experience and/or genetic relatedness. Combining rearing experiences (reared together or reared apart) and genetic relatedness (sib or non-sib) we formed four experimental groups of degus of the opposite sex in a balanced model: (1) siblings reared together (S.RT), (2) siblings reared apart (S.RA), (3) non-siblings reared together (NS.RT) and (4) non-siblings reared apart (NS.RA). To control for prenatal learning between cross-fostered siblings, a fifth group of half-siblings reared apart (HS.RA) was used for statistical comparison when a genetic relatedness effect was found.\u003c/p\u003e\u003cp\u003eAnimals were carried in individual plastic cages from the housing rooms to the experimental room. The experimental arenas consisted of a 80x80x50 cm metal arenas that was divided by a division plate (see M\u0026aacute;rquez et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e), that allowed degus to acclimate before the beginning of the experiment. The floor of the arena was a removable white-painted metal plate that was cleaned with detergent between experiments to remove any trace of scent. Seven pairs of animals were tested for each group (total pairs tested\u0026thinsp;=\u0026thinsp;35). During each experiment one of the animals was painted with non-toxic paint in the back for individualization. We found no effects of marking on exploratory (Kruskal-Wallis test: \u003cem\u003eH\u003c/em\u003e1\u0026thinsp;=\u0026thinsp;0.59, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.44), agonistic (Kruskal-Wallis test: \u003cem\u003eH\u003c/em\u003e1\u0026thinsp;=\u0026thinsp;0.046, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.83) or any other measured behavior, in agreement with previous studies in degus (M\u0026aacute;rquez et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; V\u0026aacute;squez et al. \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e2002\u003c/span\u003e; Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). After a 10 min acclimatization period, the division plate was removed. The quantification of a 20 min trial began when any of the two experimental subjects started to explore the arena. If subjects did not interact and remained still for more than 5 min, the test was repeated on a different day (8 of 35 pairs). If the tested pairs did not interact in the second attempt (5 pairs), they were not considered in the analysis.\u003c/p\u003e\u003cp\u003eAll experiments were video recorded (color CCTV camera connected to a Sony video recorder) from above the arena. An observer blind to the treatment and animals\u0026rsquo; sex analyzed the different behaviors using the JWatcher 1.0 software. Four behavioral categories were defined as social exploration, social contact, agonistic and sexual behavior, based on behavioral descriptions of intraspecific interactions in degus (see Fulk \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e1976\u003c/span\u003e; Kleiman \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e1975\u003c/span\u003e; M\u0026aacute;rquez et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Wilson and Kleiman \u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e1974\u003c/span\u003e), prairie vole (Paz y Mi\u0026ntilde;o and Tang-Martinez 1999a) and mice (Baudoin et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e1991\u003c/span\u003e). (1) Social exploratory behavior, consisted of olfactory approaches to the nose/mouth, and/or anogenital area of its partner. (2) Social contact was considered when degus were in contact either side by side, on right angle to each other, grooming, or huddling one over the other. (3) Agonistic encounters were considered when degus showed either (i) evasive behaviors: an experimental subject avoided the partner in hostile contexts by turning aside or running away, or (ii) aggressive behavior: when the experimental subject performed hostile confrontation by tail wagging, hindleg kick, foreleg push, chasing or fighting against its partner. (4) Sexual behaviors were evaluated separately for males and females: (i) male sexual behaviors were defined whenever a male mounted the female and/or when attempted to mount the female, (ii) female sexual behaviors were defined when the female remained passive under the male during mounting. Due to our experimental setup, mating and copulation were difficult to quantified, therefore, not assessed.\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003ePair encounters statistics\u003c/h3\u003e\n\u003cp\u003eThe effects of rearing experience and genetic relatedness were assessed for the four behavioral categories described above. \u003cem\u003eSocial exploration\u003c/em\u003e was analyzed using a two-way ANOVA after square-root transformation to accomplish parametric assumptions. The other three behavioral categories, \u003cem\u003esocial contact, agonistic and sexual behavior\u003c/em\u003e, did not accomplish parametric assumptions, even after being transformed, therefore we used the Scheirer-Ray-Hare test (extension of the Kruskal-Wallis test) (Sokal and Rohlf \u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e2012\u003c/span\u003e). We found no effect of sex on the mentioned behaviors; therefore, sex was not included as a factor. However, sexual behaviors between males and females differ (see Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e), thus, they were analyzed separately. For behaviors affected by genetic relatedness, we added an extra group to control for fetal learning: the half-siblings reared apart group (HS.RA). Therefore, in those cases, we compared the three sibling groups (siblings reared together, sibling reared apart, and half-siblings reared apart) using the Kruskal-Wallis test. All statistical analysis were performed using the software R (R-Core-Team \u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e2020\u003c/span\u003e) along with the nortest package (Gross and Ligges \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2015\u003c/span\u003e) to test for normality.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eBehavioral categories for social interaction quantified during dyadic encounters between degus adult pairs of the opposite sex*.\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"2\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e\u003cp\u003eSocial interaction categories for degus of the opposite sex\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eSocial-olfactory exploration\u003c/p\u003e\u003cp\u003eNose/mouth\u003c/p\u003e\u003cp\u003eAnogenital\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eBoth animals approach and sniff the nose/mouth area.\u003c/p\u003e\u003cp\u003eOne animal sniffs the anogenital area.\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eSocial contact\u003c/p\u003e\u003cp\u003eSide by side\u003c/p\u003e\u003cp\u003eGrooming\u003c/p\u003e\u003cp\u003eHuddling\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eBoth animals are in close contact with each other either side-by-side or at a right angle.\u003c/p\u003e\u003cp\u003eOne animal grooms the other one.\u003c/p\u003e\u003cp\u003eOne animal lies on top of the other one on close contact.\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eAgonistic\u003c/b\u003e\u003c/p\u003e\u003cp\u003e(i) Evasive\u003c/p\u003e\u003cp\u003e(ii) Aggressive\u003c/p\u003e\u003cp\u003eTail wagging\u003c/p\u003e\u003cp\u003eFight\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eOne animal avoids the other by turning aside or run away.\u003c/p\u003e\u003cp\u003eHostile confrontation.\u003c/p\u003e\u003cp\u003eMovement of the tail from side to side at ground level.\u003c/p\u003e\u003cp\u003eOne animal gives a hindleg kick, foreleg push, chases or attacks the other.\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eSexual behaviors\u003c/b\u003e\u003c/p\u003e\u003cp\u003e(i) Male\u003c/p\u003e\u003cp\u003e(ii) Female\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eMale mounts or attempts to mount the female.\u003c/p\u003e\u003cp\u003eFemale remains passive when the male is mounting or attempts mounting her.\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003ctfoot\u003e\u003ctr\u003e\u003ctd colspan=\"2\"\u003e*Modified from (Baudoin et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e1991\u003c/span\u003e; Fulk \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e1976\u003c/span\u003e; Kleiman \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e1975\u003c/span\u003e; M\u0026aacute;rquez et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Paz y Mi\u0026ntilde;o and Tang-Martinez 1999a; Wilson and Kleiman \u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e1974\u003c/span\u003e)\u003c/td\u003e\u003c/tr\u003e\u003c/tfoot\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e\u003ch2\u003eY-maze\u003c/h2\u003e\u003cp\u003eDegus social preferences toward kin were assessed by measuring arm visit frequency and social-olfactory exploration time in the \u003cem\u003ey-maze.\u003c/em\u003e Focal individuals were always exposed to an opposite sex sib and to an opposite sex non-sib that differed on the rearing conditions (reared together or reared apart). We found no significant differences in the arms visit frequency, nor in the time of social exploration toward any of the individuals conforming the stimuli pair (SP) irrespectively of the genetic relatedness or rearing condition (see Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e for statistics).\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eIn the \u003cem\u003ey-maze\u003c/em\u003e focal degus (N\u0026thinsp;=\u0026thinsp;56) showed no social preference toward opposite sex sibs or non-sibs irrespectively of the rearing conditions.\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"10\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colspan=\"4\" nameend=\"c5\" namest=\"c2\"\u003e\u003cp\u003eArm visit frequency\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colspan=\"4\" nameend=\"c10\" namest=\"c7\"\u003e\u003cp\u003eSocial-olfactory exploration time (s)\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e\u003cem\u003er\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.5\u003c/p\u003e\u003cp\u003e\u003cem\u003eM\u003c/em\u003e (SE)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e\u003cem\u003er\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0\u003c/p\u003e\u003cp\u003e\u003cem\u003eM\u003c/em\u003e (SE)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003cem\u003et\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e\u003cem\u003eN\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e\u003cem\u003er\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.5\u003c/p\u003e\u003cp\u003e\u003cem\u003eM\u003c/em\u003e (SE)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e\u003cem\u003er\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0\u003c/p\u003e\u003cp\u003e\u003cem\u003eM\u003c/em\u003e (SE)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e\u003cem\u003et\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e\u003cem\u003eP\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cb\u003eStimuli pair\u003c/b\u003e\u003c/p\u003e\u003cp\u003eS.RT vs. NS.RT\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e13.9 (1.1)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e13.6 (1.2)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e-0.135\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e0.89\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e14\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e44.2 (4.2)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e51.2 (3.6)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e1.25\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e0. 24\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eS.RT vs. NS.RA\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e12.3 (1.4)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e14.9 (1.4)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e1.26\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e0.23\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e14\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e57 (5.4)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e41 (4)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e-1.90\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e0.08\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eS.RA vs. NS.RT\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e19.1 (1.6)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e20.2 (1.8)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e0.299\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e0.77\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e14\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e63 (5.9)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e62.2 (5.3)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e0.055\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e0.96\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eS.RA vs. NS.RA\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e26.8 (2.1)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e26.4 (2.2)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e0.225\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e0.83\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e14\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e67.8 (5.2)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e61.9 (4)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e-0.894\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e0.39\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003ctfoot\u003e\u003ctr\u003e\u003ctd colspan=\"10\"\u003eData expressed as mean frequency and mean time (standard error), Mean (SE). \u003cem\u003et\u003c/em\u003e indicates Paired \u003cem\u003et-\u003c/em\u003etest statistic (d.f. = 13). \u003cem\u003er\u003c/em\u003e is the relatedness coefficient (Wright \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e1922\u003c/span\u003e), \u003cem\u003er\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.5 stands for siblings and \u003cem\u003er\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0 for subjects that are not genetically related. \u003cem\u003eStimuli pairs\u003c/em\u003e: siblings reared together (S.RT) versus non-siblings rear together (NS.RT); S.RT versus non-siblings rear apart (NS.RA); siblings reared apart (S.RA) versus NS.RT; S.RA versus NS.RA.\u003c/td\u003e\u003c/tr\u003e\u003c/tfoot\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003eHowever, we found that the total visits of both arms were affected by the kind of stimuli pair presented (ANOVA: \u003cem\u003eF\u003c/em\u003e\u003csub\u003e[3,48]\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;4.77, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.0055). Focal individuals exposed to a stimuli pair with both unfamiliar conspecifics (S.RA vs. NS.RA, see methods or Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e for nomenclature), visited significantly more both arms of the \u003cem\u003ey-maze\u003c/em\u003e compared to focal individuals exposed to two familiarized degus (S.RT/NS.RT; post hoc Tukey\u0026rsquo;s HSD test, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.012) and to the S.RT/NS.RA pair (post hoc Tukey\u0026rsquo;s HSD test, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.011). Degus exposed to a S.RA/NS.RT pair did not differ statistically to any other group (p\u0026thinsp;\u0026gt;\u0026thinsp;0.05, Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA). In a similar way, we found that the social exploratory frequency of the division plates was affected by the kind of SP presented (ANOVA: \u003cem\u003eF\u003c/em\u003e\u003csub\u003e[3,48]\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;4.72, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.0058). Focal degus exposed to both unfamiliar conspecifics (S.RA/NS.RA) explored significantly more the division plates than degus exposed to conspecifics reared together (S.RT/NS.RT) (post hoc Tukey\u0026rsquo;s HSD test, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.008) and compared to animals exposed to S.RT/NS.RA stimuli pairs (post hoc Tukey\u0026rsquo;s HSD test, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.018). Degus exposed to S.RA/NS.RT did not differ in the social exploration of the division plate to any other experimental group (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eB). In addition, we found that females socially explored significantly longer both stimuli pairs than males (ANOVA: \u003cem\u003eF\u003c/em\u003e\u003csub\u003e[1,48]\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.43, \u003cem\u003eP\u0026thinsp;=\u003c/em\u003e\u0026thinsp;0.024).\u003c/p\u003e\u003cp\u003eIn summary, degus socially explored unfamiliar pairs more frequently than pairs that included a sibling with previous rearing experience.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec12\" class=\"Section2\"\u003e\u003ch2\u003eDyadic encounters\u003c/h2\u003e\u003cp\u003eWe further assessed opposite sex sibling discrimination quantifying social interactions in an experimental arena. Social interactions were considered as any behavior involving both degus. These behaviors were further divided into four categories (see Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e): social-olfactory exploration, social contact, agonistic encounters and sexual behaviors.\u003c/p\u003e\u003cp\u003eNose/mouth social-olfactory exploration time was significantly lower in pairs of opposite sex reared together than unfamiliar pairs with no previous contact (ANOVA: \u003cem\u003eF\u003c/em\u003e\u003csub\u003e[1, 40]\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;6.243, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.017; Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eA). We did not observe any effects of genetic relatedness (ANOVA: \u003cem\u003eF\u003c/em\u003e\u003csub\u003e[1, 40]\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.071, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.79) nor of the interaction between familiarity and genetic relatedness (ANOVA: \u003cem\u003eF\u003c/em\u003e\u003csub\u003e[1, 40]\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.44, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.51) on this behavior. In addition, male-female pairs reared apart explored each other\u0026rsquo;s anogenital area significantly more than pairs reared together (ANOVA: \u003cem\u003eF\u003c/em\u003e\u003csub\u003e[1, 40]\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;5.72, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.02; Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eB). We found no effect of genetic relatedness (ANOVA: \u003cem\u003eF\u003c/em\u003e\u003csub\u003e[1, 40]\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.032, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.86), nor an effect of the interaction between the two factors (ANOVA: \u003cem\u003eF\u003c/em\u003e\u003csub\u003e[1, 40]\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;0.68, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.41) in this behavior. We found that tail wagging, an agonistic behavior, was displayed significantly more by unfamiliar pairs (S.RA and NS.RA; Scheirer-Ray-Hare test: \u003cem\u003eH\u003c/em\u003e1\u0026thinsp;=\u0026thinsp;5.18, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.023; Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eC), irrespectively of their degree of genetic relatedness (Scheirer-Ray-Hare test: \u003cem\u003eH\u003c/em\u003e2\u0026thinsp;\u003cem\u003e=\u003c/em\u003e\u0026thinsp;0.036, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.84) and we found no interaction effects (Scheirer-Ray-Hare test: \u003cem\u003eH\u003c/em\u003e1\u0026thinsp;=\u0026thinsp;0.84, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.36). On the other hand, we rarely observed fights, 11% (only in 3 of 28 couples), and therefore we did not consider it in the statistical analysis. We observed sexual behaviors in all four experimental groups. However, as males have different sexual behaviors than females (see Table \u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e), statistical analyses were done for males and females separately, which yield to a reduced sample sizes per group (N\u0026thinsp;=\u0026thinsp;5). We found no effect of familiarity (males: Scheirer-Ray-Hare test: \u003cem\u003eH\u003c/em\u003e2\u0026thinsp;\u003cem\u003e=\u003c/em\u003e\u0026thinsp;1.2, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.3; females: Scheirer-Ray-Hare test: \u003cem\u003eH\u003c/em\u003e2\u0026thinsp;\u003cem\u003e=\u003c/em\u003e\u0026thinsp;0.05, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.8), genetic relatedness (males: Scheirer-Ray-Hare test: \u003cem\u003eH\u003c/em\u003e2\u0026thinsp;\u003cem\u003e=\u003c/em\u003e\u0026thinsp;0.2, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.7; females: Scheirer-Ray-Hare test: \u003cem\u003eH\u003c/em\u003e2\u0026thinsp;\u003cem\u003e=\u003c/em\u003e\u0026thinsp;0.4, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.5), nor the interaction of these independent variables (males: Scheirer-Ray-Hare test: \u003cem\u003eH\u003c/em\u003e2\u0026thinsp;\u003cem\u003e=\u003c/em\u003e\u0026thinsp;0.03, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.9; females: Scheirer-Ray-Hare test: \u003cem\u003eH\u003c/em\u003e2\u0026thinsp;\u003cem\u003e=\u003c/em\u003e\u0026thinsp;0.03, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.9) on the frequency of sexual behaviors performed. In addition, no differences were found in the time of sexual behaviors performance (data not shown).\u003c/p\u003e\u003cp\u003eWe found that social contact (see Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e) was the only behavior affected by genetic relatedness (Scheirer-Ray-Hare test: \u003cem\u003eH\u003c/em\u003e2\u0026thinsp;=\u0026thinsp;4.22, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.04) and not by familiarity (Scheirer-Ray-Hare test: \u003cem\u003eH\u003c/em\u003e1\u0026thinsp;=\u0026thinsp;0.19, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.66), without an interaction effect (Scheirer-Ray-Hare test: \u003cem\u003eH\u003c/em\u003e1\u0026thinsp;=\u0026thinsp;1.29, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.26; Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). In addition, we compared both sibling groups (S.RT and S.RA) with the half-sibling reared apart (HS.RA) experimental group, to further assess the possible effect of fetal learning on social contact behaviors among genetically related individuals. We found no significant difference on social contact behaviors between these three groups (Kruskal-Wallis test: \u003cem\u003eH\u003c/em\u003e1\u0026thinsp;=\u0026thinsp;1.13, \u003cem\u003eP\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.57).\u003c/p\u003e\u003cp\u003eIn summary, we found that rearing experience explains a wide range of social interactions in adult degus of the opposite sex. However, we found an effect of genetic relatedness on the social contact behavior.\u003c/p\u003e\u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eThe present study aimed to investigate the behavioral mechanisms underlying kin discrimination between opposite sex siblings in degus. To evaluate the influence of both rearing experience and genetic relatedness on this social behavior, we used a cross-fostering protocol and conducted two behavioral tests: a \u003cem\u003eY-maze\u003c/em\u003e and \u003cem\u003edyadic interaction\u003c/em\u003e trials. Consistent with previous findings in same sex sibling degus, we found that prior association in both behavioral experiments had a strong effect on kin discrimination between opposite sex siblings. Yet, we detected an effect of genetic relatedness in one behavior during dyadic interactions (e.g. social contact). In addition, we found that females explored more than males when exposed to opposite sex conspecifics in the \u003cem\u003ey-maze\u003c/em\u003e.\u003c/p\u003e\u003cdiv id=\"Sec14\" class=\"Section2\"\u003e\u003ch2\u003eY-maze\u003c/h2\u003e\u003cp\u003eOur results showed that male and female degus did not show a preference for either of the two conspecifics of the opposite sex presented on the \u003cem\u003ey-maze\u003c/em\u003e, regardless of the genetic relatedness or the rearing experience they shared with them. In fact, degus explored both conspecifics similarly. Even though no signs of preference for either conspecific were found, the previous experience with the stimuli pairs affected the total number exploration events toward both conspecifics of the opposite sex. Considering that olfactory exploration can be interpret as indicator of interest in an odor source (Johnston et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e1997\u003c/span\u003e), and that degus could explore the labyrinth freely, our findings suggests that degus were more interested in the pairs of conspecifics of the opposite sex that were unknown to them.\u003c/p\u003e\u003cp\u003eIn agreement with previous studies on olfactory discriminations in degus and other species (Halpin and Hoffman \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e1987\u003c/span\u003e; Holmes \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e1984\u003c/span\u003e; Johnston \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e1993\u003c/span\u003e; Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e), our results show that individuals explored familiar olfactory cues less frequently than novel ones, indicating that degus can recognize olfactory cues they have previously encountered. Based on this, it would be expected that the group exposed to two unfamiliarized degus (S.RA vs NS.RA) explored significantly more than both groups exposed to one familiarized and one non-familiarized conspecific (S.RA vs NS.RT and S.RT vs NS.RA). However, this was not the case. Instead, social exploratory behavior in the S.RA/NS.RA group differed significantly only with the group where the experimental subject had been reared with its sibling (S.RT/ NS.RA) [see Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e]. This result shows that the presence of a sib reared together was sufficient to reduce social exploratory behaviors, resembling the responses observed when both conspecifics were \u0026ldquo;known\u0026rdquo; (S.RT/NS.RT), suggesting a synergistic effect of familiarity and genetic relatedness in this context.\u003c/p\u003e\u003cp\u003eThe \u003cem\u003ey-maze\u003c/em\u003e experiment also revealed sex differences in social exploratory behavior, with females exploring the social stimuli -two opposite sex conspecifics- significantly more than males. These findings are consistent with studies in other rodent species across different contexts. For instance, females mound-building mice (\u003cem\u003eMus spicilegus\u003c/em\u003e) explore novel object more than males (Simeonovska-Nikolova \u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). In rats (\u003cem\u003eRattus norvegicus\u003c/em\u003e), exploratory behavior develops during adolescence in both open field and elevated plus maze paradigms, with females showing higher levels of exploration than males (Lynn and Brown \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). This sex difference is also evident in social contexts, where females explore novel social arenas more than males (Cavigelli et al. \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). In degus, different bioassays have yielded contrasting results regarding sex difference in social exploration. For example, females explore more than males in experiments assessing sex differences during social interactions between non-genetically related conspecifics (Fischer et al. \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e1986\u003c/span\u003e). However, a sibling discrimination study found no sex differences in olfactory exploration between same-sex siblings during dyadic encounters in degus (Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). Another study addressing the influence of early olfactory experience reported that males explored more than females in same-sex dyadic encounters when exposed to a conspecific impregnated with an artificial odorant (M\u0026aacute;rquez et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). In our study, sex differences in exploratory behaviors were observed only in the \u003cem\u003ey-maze\u003c/em\u003e experiment, but not during dyadic encounter (see below). We propose that social exploratory behaviors in degus is context-dependent and may vary between sexes depending on the social context and the nature of the olfactory stimuli (i.e. direct contact with conspecifics, urine marks, bedding, etc.).\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec15\" class=\"Section2\"\u003e\u003ch2\u003eDyadic encounters\u003c/h2\u003e\u003cp\u003eMost social behaviors observed during male-female dyadic encounters were determined by rearing experience rather than genetic relatedness \u003cem\u003eper se\u003c/em\u003e. Social-olfactory exploration time (including mouth/nose and anogenital investigation) was significantly higher between unfamiliar individuals than between degus reared together. In addition, degus reared together performed significantly fewer agonistic behaviors compared to non-familiarized pairs, indicating a behavioral bias based on rearing experience. Social contact was the only behavior affected by genetic relatedness and not rearing experience, suggesting that not all male-female social interactions are equally affected by prior association. Our results confirm earlier findings in same sex degus (Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e), showing that opposite sex sibling discrimination is predominantly mediated by rearing experience, and to a lesser extent by genetic relatedness.\u003c/p\u003e\u003cp\u003eSexual behaviors, on the other hand, are expressed differently between males and females and therefore were analyzed separately. Males mount or attempt to mount females, while females remain passive during these interactions. We found no effect of rearing experience or genetic relatedness on sexual behaviors. Considering that separating male and female behaviors reduces the sample size for statistical analysis, to confirm this result it may be necessary to increase the sample size. In addition, our study specifically focused on sexual behaviors observed in the experimental arena, rather than on mating per se. In line with our results, a field study in degus reported that one female\u0026rsquo;s offspring can be sired by one to six males, who may or may not be genetically related (Ebensperger et al. \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Thus, litters can exhibit multiple paternity, suggesting that in degus sociality inbreeding avoidance is not an issue.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec16\" class=\"Section2\"\u003e\u003ch2\u003eKin discrimination mechanisms and olfaction\u003c/h2\u003e\u003cp\u003eStudies on the behavioral mechanisms underlying kin discrimination (prior association and/or phenotype matching) have revealed that while in some species rearing experience plays a major role in kin discrimination (e.g. thirteen-lined ground squirrels (Holmes \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e1984\u003c/span\u003e), and prairie voles (Paz y Mi\u0026ntilde;o and Tang-Martinez \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e1999b\u003c/span\u003e)) in others would be less relevant. Mateo (\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2003\u003c/span\u003e) reviewed the mechanisms of kin recognition in rodents an found that 12 out of 32 species can recognize their genetically close conspecifics based on genetically related cues (Mateo \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2003\u003c/span\u003e). Interestingly, this type of behavioral discrimination is not correlated with the social system, nor with the species\u0026rsquo; mating system (Mateo \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2003\u003c/span\u003e). Tang-Martinez (\u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e2001\u003c/span\u003e) in a deeper analysis of kin recognition mechanisms proposed that both behavioral mechanisms are indeed the result of learning that might occur at different moments. In this line, cross-fostering, cannot rule out intra-uterine olfactory learning (Robinson and Smotherman \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e1991\u003c/span\u003e). Prenatal olfactory learning occurs and can determine flavor and food preferences in mice and rats (Kamenetzky et al. \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). In the context of kin discrimination, prenatal olfactory learning has not been tested. Nevertheless, it is possible that the mother and wombmates\u0026rsquo; olfactory cues present in the intrauterine environment can be learned, which would account for kin discrimination without prior association (Robinson and Smotherman \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e1991\u003c/span\u003e). Giving the developmental characteristics of degus with a long gestation period (90 days) and a precocial offspring (Weir \u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e1970\u003c/span\u003e), intra-uterine olfactory learning could be a relevant process for kin discrimination. Further studies are needed to test this hypothesis.\u003c/p\u003e\u003cp\u003eIn rodent species, kin discrimination in its various forms (i.e. exploration, aggressiveness, affiliative behaviors, or inbreeding avoidance) is mediated by olfactory cues (Halpin \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e1991\u003c/span\u003e; Holmes \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e1984\u003c/span\u003e; Sherborne et al. \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). Furthermore, the olfactory identity of individuals can be determined by the expression of major histocompatibility complex (MHC) genes (Beauchamp et al. \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e1988\u003c/span\u003e; Yamazaki et al. \u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e1988\u003c/span\u003e) and/or by the presence of major urinary proteins (MUPs) (Cheetham et al. \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2007\u003c/span\u003e; Green et al. \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Hurst et al. \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2001\u003c/span\u003e). Kin discrimination in degus is mediated by olfactory cues (Jesseau et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2008\u003c/span\u003e; M\u0026aacute;rquez et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e) present in littermates and learned during early ontogeny within the nest context (M\u0026aacute;rquez et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). Although the sources of olfactory identity in degus remain unknown, behavioral data show that cross-fostered degus can discriminate the scent of a sibling from that of a non-sibling from its litter, but not that of a pair of its siblings from its litter (Villavicencio et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). This suggests that the scent profile of degus is more similar between kin than between non-kin and is therefore possibly influenced by MHC genes and/or MUPs. We propose that these kinds of olfactory signals mediate behavioral biases between closely related adult degus even when reared apart.\u003c/p\u003e\u003cp\u003eWe consider that the study of male-female socio-sexual interactions is essential for understanding the conservation of a particular way of living, because reproduction is a systemic phenomenon that preserves not only the transmission of genetic material to the offspring, but also the medium in which living takes place (as proposed by Maturana and Mpodozis (2000)). Here, we show that social interactions and kin discrimination between male and female degus are rich and complex and are worth further investigation.\u003c/p\u003e\u003c/div\u003e"},{"header":"Conclusions","content":"\u003cp\u003eWe confirmed that male-female sibling discrimination in degus is similar to same-sex sibling discrimination; both are mainly driven by rearing experience. We found that some behaviors are mediated by rearing experience (e.g., exploration, agonistic behaviors), others by genetic relatedness (e.g., social contact), and some were not affected by either factor (e.g., sexual behaviors). Thus, focusing solely on a single type of behavior, or only on the ability to distinguish individuals based on olfactory cues, may lead to incomplete or misleading conclusions. Our findings highlight the importance of examining multiple behavioral categories during social encounters to gain a better understanding of the mechanisms underlying kin discrimination.\u003c/p\u003e"},{"header":"Declarations","content":"\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eN.I.M and R.A.V conceived and designed research. N.I.M and C.P.V performed experiments.N.I.M analyzed data and prepared figures.N.I.M and C.P.V drafted manuscript.N.I.M, C.P.V, J.M-H and JM edited and revised manuscript.N.I.M., C.P.V, J.M-H, J.M and R.A.V approved final version of manuscript.\u003c/p\u003e\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe gratefully acknowledge Dr. Claudia Cecchi, Dr. Daniela Parra, Ronny Z\u0026uacute;\u0026ntilde;iga and Solano Henr\u0026iacute;quez for their valuable assistance. We would also like to thank the Pilot Scientific Productivity Workshop for their valuable input and motivation. Support for this work was provided by a Programa de Mejoramiento de la Calidad de la Educaci\u0026oacute;n Superior (MECESUP UCO-0214) fellowship, and a Comisi\u0026oacute;n Nacional de Investigaci\u0026oacute;n Cient\u0026iacute;fica y Tecnol\u0026oacute;gica de Chile (CONICYT AT-24050185) grant to N.I.M and by a\u0026nbsp;Fondo Nacional de Desarrollo Cient\u0026iacute;fico y\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;Tecnol\u0026oacute;gico (FONDECYT) grant to J.M. (FONDECYT 1250880).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eConflict of interest: Authors report no conflict of interest.\u0026nbsp;\u003c/p\u003e\u003ch2\u003eData Availability\u003c/h2\u003e\u003cp\u003eData will be made available upon reasonable request.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eApio A, Kabasa JD, Ketmaier V, Schroeder C, Plath M, Tiedemann R (2010) Female philopatry and male dispersal in a cryptic, bush-dwelling antelope: a combined molecular and behavioural approach. 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Science 240 (4857): 1331-1332. https://doi.org/10.1126/science.3375818\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"journal-of-comparative-physiology-a","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jcpa","sideBox":"Learn more about [Journal of Comparative Physiology A](http://link.springer.com/journal/359)","snPcode":"359","submissionUrl":"https://submission.nature.com/new-submission/359/3","title":"Journal of Comparative Physiology A","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Kin discrimination, sibling discrimination, Octodon degus, opposite sex, rearing experience, cross fostering","lastPublishedDoi":"10.21203/rs.3.rs-7725339/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7725339/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eKin recognition refers to the discrimination and subsequent behavioral bias toward close relatives. Kin recognition is an important modulator of social interaction. Among relatives, reduced exploration, lower aggressiveness, and decreased inbreeding are typically expected. Two main mechanisms influence kin discrimination: prior association (familiarity) and genetic relatedness. In this study, we investigated kin discrimination between opposite-sex siblings in degus (\u003cem\u003eOctodon degus\u003c/em\u003e), a diurnal, communally nesting rodent endemic to central Chile. While in this species discrimination among same-sex siblings has been shown to be mainly influenced by rearing experience, it remains unknown whether degus can discriminate their opposite-sex siblings at all, and if so, what mechanisms are at play. We used a cross-fostering protocol to experimentally separate the effects of familiarity and genetic relatedness, producing four groups that combined siblings and non-siblings reared together or apart. Behavior was assessed in two experimental setups: (i) a \u003cem\u003ey-maze\u003c/em\u003e to test olfactory preferences and (ii) a dyadic encounter arena to evaluate social interactions. In the y-maze, each experimental subject was simultaneously exposed to one sibling and one non-sibling (stimuli pair, SP), both of the opposite sex. Exploration increased significantly when the subject had no rearing experience with either SP member. Females explored more than males the y-maze. Dyadic encounter experiments showed that rearing experience influenced exploratory and agonistic interactions but not sexual behaviors. Social contact, on the other hand, was more frequent among siblings. Taken together, our results indicate that rearing experience plays a major role in modulating opposite-sex social behaviors in this species.\u003c/p\u003e","manuscriptTitle":"Growing up together or apart: Rearing experience effects on opposite-sex sibling discrimination in Chilean brush tailed mouse (Octodon degus)","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-10-14 13:00:15","doi":"10.21203/rs.3.rs-7725339/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-11-20T20:06:27+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-11-20T19:18:03+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-31T16:58:24+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"149258399757259368430225605631972377114","date":"2025-10-14T02:38:12+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"200852298629956881683281997466976363975","date":"2025-10-09T10:48:26+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-10-01T07:10:00+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-09-27T13:53:56+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-09-27T13:51:52+00:00","index":"","fulltext":""},{"type":"submitted","content":"Journal of Comparative Physiology A","date":"2025-09-27T02:21:12+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"journal-of-comparative-physiology-a","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jcpa","sideBox":"Learn more about [Journal of Comparative Physiology A](http://link.springer.com/journal/359)","snPcode":"359","submissionUrl":"https://submission.nature.com/new-submission/359/3","title":"Journal of Comparative Physiology A","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"ae5e7b60-bfec-40ad-b0d7-c693e3937786","owner":[],"postedDate":"October 14th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2026-04-13T16:08:50+00:00","versionOfRecord":{"articleIdentity":"rs-7725339","link":"https://doi.org/10.1007/s00359-026-01806-4","journal":{"identity":"journal-of-comparative-physiology-a","isVorOnly":false,"title":"Journal of Comparative Physiology A"},"publishedOn":"2026-04-12 15:59:22","publishedOnDateReadable":"April 12th, 2026"},"versionCreatedAt":"2025-10-14 13:00:15","video":"","vorDoi":"10.1007/s00359-026-01806-4","vorDoiUrl":"https://doi.org/10.1007/s00359-026-01806-4","workflowStages":[]},"version":"v1","identity":"rs-7725339","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7725339","identity":"rs-7725339","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}