Transition to siblinghood causes a substantial and long-lasting increase in urinary cortisol levels in wild bonobos
Abstract
In animals with slow ontogeny and long-term maternal investment, immatures are likely to experience the birth of a younger sibling before reaching maturity. In these species, the birth of a sibling marks a major event in an offspring’s early life as the older siblings experience a decrease in maternal support. The transition to siblinghood (TTS) is often considered to be stressful for the older offspring, but physiological evidence is lacking. To explore the TTS in wild bonobos, we investigated physiological changes in urinary cortisol (stress response), neopterin (cell-mediated immunity), and total triiodothyronine (T3, metabolic rate), as well as changes in behaviors that reflect the mother–offspring relationship. Following a sibling’s birth, urinary cortisol levels of the older offspring increased fivefold, independent of their age, and remained elevated for 7 months. The cortisol level increase was associated with declining neopterin levels; however, T3 levels and behavioral measures did not change. Our results indicate that the TTS is accompanied by elevated cortisol levels and that this change does not coincide with nutritional weaning and attainment of physical independence. Our results suggest that bonobos and humans experience TTS in similar ways and that this developmental event may have emerged in the last common ancestor.
Editor's evaluation
This article examines the behavioral and physiological responses of wild bonobos to the birth of a younger sibling. The findings contribute to our understanding of the effects of a major life history transition in a primate species that is closely related to humans. An important strength of this article is the novel use of a longitudinal dataset that incorporates both behavioral and physiological measures.
https://doi.org/10.7554/eLife.77227.sa0Introduction
In mammals, weaning refers to the transition from nutritional dependency to a stage when immatures are independent of maternal food provisioning. The term weaning is often used for the attainment of nutritional independence, but also comprises the process of social independence and behavioral maturation, which can occur at different ages. Weaning age varies within and across species, and is an important developmental stage in the life history of mother and offspring (Smith, 2013; Weary et al., 2008). While the dependency on post-weaning maternal support can be inferred from behavioral observations, the putative fitness effects are rarely explored. A reduction or complete loss of maternal support has substantial fitness costs throughout an individual’s life span (Zipple et al., 2021). However, while maternal loss is a dramatic event, there are normative events such as sibling birth that affect the life of the older offspring. In vertebrate species with a slow development, many immatures grow up with siblings, and sibling relationships can have profound influences on fitness (Berger et al., 2021; Nitsch et al., 2013). The younger sibling may benefit from an older sibling in terms of survival, reproductive maturation, and socialization (Berger et al., 2021; Nitsch et al., 2013; Stanton et al., 2017). However, the older offspring must share maternal care, which may influence its social behavior as well as its physiological constitution.
Primates differ from most other social mammals in having remarkably slow life histories (Charnov and Berrigan, 1993; Jones, 2011). Immatures grow slowly, social maturation extends well into adulthood, and to a certain degree, beneficial mother–offspring relationships can last a lifetime (Jones, 2011; Pereira and Fairbanks, 1993; Surbeck et al., 2019). Therefore, female primates may give birth to another infant before the older offspring reaches physical or social maturity, or even before being weaned. For the older offspring, this transition to siblinghood (TTS) marks the onset of considerable changes, including the sudden emergence of a competitor for maternal resources (sibling rivalry; Dettwyler, 2017; Myers and Bjorklund, 2018) and a decline in maternal support (Kramer, 2011). Accordingly, in humans, TTS is considered to be a stressful life event for the older sibling even under favorable conditions, a perspective that seems to be supported by TTS-related behaviors of the older offspring such as aggression, clinginess, and depressive syndromes. However, sibling birth also presents opportunities for the older offspring, such as social and emotional growth through interacting with the newborn. Individuals vary in how they adjust to the birth of a younger sibling; some children have difficulties while others cope well (reviewed in Volling, 2012; Volling et al., 2017). In any case, the birth of a sibling is linked to a time of change the older child must cope with. Evidence from nonhuman primates is scarce, but the available information resembles reports from humans (Devinney et al., 2003; Schino and Troisi, 2001). However, whether behavioral changes during TTS are actually associated directly with sibling birth, or are rather simply a result of age-related withdrawal of maternal support, remains to be resolved (Volling, 2012; Volling et al., 2017).
TTS could overlap with and/or accelerate weaning and attainment of physical independence, which, on its own, is known to be stressful in primates and other mammals (e.g., Hau and Schapiro, 2007; Mandalaywala et al., 2014). As a result, it is difficult to differentiate between the effects of sibling birth and weaning (Volling, 2012; Weary et al., 2008). Nutritional weaning refers to the termination of an offspring’s consumption of maternal milk, though they may still continue nipple contact (without milk transfer) – this is assumed to be a social comfort behavior (Bădescu et al., 2017; Berghänel et al., 2016; Matsumoto, 2017). It is common that females give birth to another infant before the older offspring reaches full independence, resulting in an overlap of dependency in siblings of different ages (Achenbach and Snowdon, 1998). In nonhuman primates, sibling birth affects the quality and quantity of interactions between the older offspring and the mother (Schino and Troisi, 2001; van Noordwijk and van Schaik, 2005), and may affect the fitness of the older offspring throughout its life (Alberts, 2019; Bădescu et al., 2022; Thompson et al., 2016; Tung et al., 2016; Zipple et al., 2019).
Apes offer a particularly suitable model to explore developmental changes in an evolutionary context (Sayers, 2015): maternal support is intense and persists for a long time (Stanton et al., 2020; van Noordwijk et al., 2018), and extended periods of parental care of two dependent offspring of different ages are common (Achenbach and Snowdon, 1998). Juvenile apes associate with their mother for several years after nutritional weaning. While data on mother–offspring relationships are abundant, little is known about the interactions between immatures and infants born to the same female (Watts and Pusey, 1993). Wild orangutans have the longest known mammalian inter-birth interval (7–9 years), and sibling rivalry is likely modest or less intense since the close association of the mother with the older offspring ends before the next infant is born (van Noordwijk et al., 2018; van Noordwijk and van Schaik, 2005). In gorillas, inter-birth intervals range from 4 to 6 years (Stoinski et al., 2013). In male mountain gorillas, sibling bonds may last into adulthood (Robbins, 1995), and following maternal loss, siblings may provide social support (Morrison et al., 2021), indicating that siblings are strong partners in this species. In wild chimpanzees, inter-birth intervals range from 2 to 11 years (Thompson, 2013). There is one anecdotal report of an older offspring responding to sibling birth with increasing attempts to establish physical contact with the mother and the emergence of signs of depression (Clark, 1977). Based on this, it can be assumed that depending on the species, immature apes experiencing the birth of a sibling are exposed to different social environments: Because a greater difference in sibling ages may correspond to less conflict in terms of their maternal support needs, it is likely that species with shorter inter-birth intervals (gorillas and chimpanzees) experience stronger effects of TTS than those with longer intervals (orangutans).
Immature bonobos depend heavily on their mothers and maintain close spatial and physical contact during the first 2 years of life (De Lathouwers, 2004; Kuroda, 1989; Lee et al., 2020). After the age of 5 years, spatial distance to the mother increases (Kuroda, 1989; Toda et al., 2021), but in the case of sons, associations between mothers and offspring persist even when sons reach adulthood (Hohmann et al., 1999; Surbeck et al., 2019). Nutritional weaning occurs between 4 and 5 years of age (Kuroda, 1989; Oelze et al., 2020), and behavioral observations and urinary cortisol measures indicate that nutritional weaning is less stressful in bonobos than in chimpanzees (De Lathouwers and Van Elsacker, 2006; Tkaczynski et al., 2020). Notably, monitoring changes in urinary cortisol levels during weaning revealed the first evidence that the older offspring may respond physiologically to the birth of a sibling (Tkaczynski et al., 2020).
Here, we investigate TTS-related changes in physiological responses in wild habituated juvenile bonobos (Pan paniscus) at LuiKotale in the Democratic Republic of Congo. We used multiple physiological and behavioral measures to investigate the responses of older siblings to the birth of their younger sibling. We sought to disentangle the effects of changes in mother–offspring relationships and energetics that are associated with nutritional and social weaning, from the specific effects of a younger sibling’s birth. We leveraged the large variation in inter-birth intervals in bonobos (Knott, 2001; Tokuyama et al., 2021) to differentiate between the effects of TTS versus nutritional and social weaning. In our study population, inter-birth intervals ranged from 2.3 to 8.6 years (mean ± SD = 5.4 ± 1.5 years) (Tkaczynski et al., 2020), and thus the developmental status of older siblings at the time when their mothers gave birth to another infant ranged from highly dependent in terms of travel support and foraging skills (i.e., time carried and nursed) to mostly independent. This discordance between inter-birth interval lengths and the developmental timelines of the older offspring enabled us to explore the specific effects of TTS on older siblings’ (a) physiological stress response (cortisol), (b) immunity (neopterin), (c) energetic change (total T3), (d) relationships with their mothers, and (e) changes in foraging and travel competence, while controlling for (f) offspring sex and age.
Changes in cortisol are widely accepted as a physiological marker to quantify stress responses in humans and other mammals because after exposure to a stressor – an event that challenges homeostasis – cortisol is secreted to restore homeostasis (Karatsoreos and McEwen, 2010; Romero and Beattie, 2022). Cortisol is produced in response to physical as well as psychosocial stressors (Kirschbaum and Hellhammer, 1994; McEwen, 2017). In children, salivary cortisol levels increase during traumatic family events and/or in anticipation of important positive or negative events, indicating that cortisol measurements are a valuable tool to assess children’s stress responses to family and social interactions (Flinn et al., 2012; Flinn et al., 2011). We expected that TTS is experienced by the older offspring as a challenging event. Therefore, we predicted a sudden increase in cortisol levels at the time of sibling birth in reaction to this event.
Neopterin is produced by macrophages, monocytes, and dendritic cells after activation. Therefore, an increase in neopterin levels reflects the activation of cell-mediated immune response after an infection with intracellular pathogens (Murr et al., 2002). Immune responses are linked to changes in cortisol levels. While a short increase in cortisol levels can support immune functions, long-term elevation of cortisol levels suppresses immune function (Dhabhar, 2014). Therefore, if TTS stimulates a short increase in cortisol levels, we expect increasing or unchanged neopterin levels, whereas if TTS causes a long-lasting cortisol response, we expect a decline in neopterin levels.
Triiodothyronine (T3) is a thyroid hormone that influences metabolic rate. T3 levels decline during times of energy restriction to conserve energy (reviewed in Behringer et al., 2018). Measuring total T3 levels allows for disentangling the effects of energetic and social stressors, which may both occur around the age of nutritional weaning and/or TTS (Maestripieri, 2018; Mandalaywala et al., 2014). If sibling birth induces metabolic issues in the older offspring, we would expect a decline in total T3 levels following sibling birth.
We complemented physiological measures with behavioral scores of nipple contact and riding, the time offspring spent in body contact with their mothers, 5 m proximity to the mother, and independent foraging. Changes in these parameters can indicate nutritional weaning and attainment of physical and social independence around TTS. We compared measures of these parameters in the older offspring before versus after the birth of a sibling to investigate whether TTS-related changes in cortisol can be linked to similar changes in behavior and thus to weaning patterns and changes in the mother–offspring relationship.
Results
Our main results are summarized in Table 1, and model structures can be derived from Tables 2 and 3, and from Supplementary file 1. We applied nonlinear generalized additive mixed models (GAMM) to investigate continuous changes in our parameters of interest around the time of sibling birth and compared those models to identical ones in which we added a categorical distinction between before and after sibling birth to allow noncontinuous, sudden changes at sibling birth. We considered that our response variables may naturally change with offspring age. Age-related changes in our response variables might (a) directly mediate potential changes during TTS in case of strong temporal overlap and (b) moderate these effects as the impact of TTS may decline with decreasing dependency of older offspring from maternal support. To control for potential mediation, we ran a model with age for all our response variables. If TTS has effects beyond weaning, we would expect sudden changes at the time of sibling birth also after controlling for age-related changes. To investigate whether continuous and sudden effects of sibling birth decrease with increasing age at sibling birth, we split individuals along the median (5.11 years old at sibling birth) and ran additional models that allowed for different trajectories around sibling birth for the two age cohorts. Finally, we generated continuous two-way interaction plots to visually inspect whether and how the trajectories around sibling birth changed with increasing offspring age (for more details see ‘Methods’).
Summary of the main findings of analyses of physiological markers and scores of older offspring behavioral during the transition to siblinghood (TTS).
Cortisol | Neopterin | Total T3 | Nursing | Riding | 5m-proximity with mother | Body contact with mother | Independent foraging | |
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Sudden change at sibling birth | Yes, increase | Yes, decrease | No | No | Yes | No | Yes, but increase | No |
Effect of TTS attenuates with offspring age | No | No | No | No: all changes before sibling birth | Yes, effect exists only up to 5 years old | No | No | No: all changes finished before sibling birth |
General additive mixed model results for physiological changes (urinary cortisol, urinary neopterin, and urinary total T3 levels; all log-transformed) in the older offspring 7 years before and after sibling birth.
Reference | Log cortisol | Log neopterin | Log total T3 | ||||||||||
Factor variables: | Category | Est. | SE | t | p | Est. | SE | t | p | Est. | SE | t | p |
(intercept) | 0.85 | 0.05 | 16.23 | 2.41 | 0.04 | 58.68 | 0.89 | 0.05 | 17.26 | ||||
Males | Females | 0.11 | 0.05 | 2.09 | 0.037 | –0.03 | 0.04 | –0.69 | 0.488 | –0.11 | 0.05 | –2.19 | 0.030 |
After S-birth* | Before† | 0.43 | 0.08 | 5.27 | <0.001 | –0.19 | 0.06 | –3.01 | 0.002 | 0.13 | 0.08 | 1.61 | 0.114 |
Smooth term variables: | edf | Ref. df | F | p | edf | Ref. df | F | p | edf | Ref. df | F | p | |
Time-S-birth: males | 2.65 | 3.17 | 1.17 | 0.262 | 1.00 | 1.00 | 0.53 | 0.469 | 1.00 | 1.00 | 3.76 | 0.054 | |
Time-S-birth: females | 1.77 | 2.12 | 0.69 | 0.433 | 1.50 | 1.83 | 0.56 | 0.603 | 1.00 | 1.00 | 0.01 | 0.922 | |
Time-S-birth: males | Females | 1.00 | 1.00 | 0.05 | 0.818 | 1.00 | 1.00 | 0.23 | 0.585 | 1.00 | 1.00 | 2.85 | 0.093 |
Age: males | 1.00 | 1.00 | 3.22 | 0.074 | 3.19 | 3.74 | 4.25 | 0.002 | - | - | - | - | |
Age: females | 1.00 | 1.00 | 1.37 | 0.243 | 1.00 | 1.00 | 0.03 | 0.874 | - | - | - | - | |
Age: males | Females | 1.00 | 1.00 | 0.43 | 0.513 | 2.17 | 2.63 | 1.08 | 0.243 | - | - | - | - |
Daytime | 1.20 | 1.37 | 29.27 | <0.001 | 1.00 | 1.00 | 4.82 | 0.029 | 2.10 | 2.56 | 1.77 | 0.142 | |
Seasonal effect | 2.38 | 3.00 | 11.18 | <0.001 | 0.51 | 3.00 | 0.22 | 0.278 | 0.00 | 3.00 | 0.00 | 0.723 | |
Random effects: | |||||||||||||
Time-S-birth per ID (smooth) | 0.00 | 111.0 | 0 | 0.238 | 0.00 | 112.0 | 0 | 0.850 | 7 | 148.0 | 0 | 0.015 | |
Age per ID (smooth) | 0.00 | 109.0 | 0 | 0.291 | 0.00 | 108.0 | 0 | 0.737 | - | - | - | - | |
Mother ID (intercept) | 0.00 | 13.0 | 0 | 0.175 | 0.00 | 13.0 | 0 | 0.313 | 0 | 13.0 | 0 | 0.230 | |
Year (intercept) | 0.00 | 1.0 | 0 | 0.012 | 0.00 | 1.0 | 0 | 0.277 | 0 | 1.0 | 0 | 0.850 | |
R2adj (deviance explained) | 0.311 (33.8%) | 0.169 (19.6%) | 0.117 (15.3%) | ||||||||||
N (p-value, full/null comp) | 319 (<0.001) | 314 (<0.001) | 319 (0.020) |
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Green indicates classic interaction term derived from a separate model calculation (see ‘Methods’). Data points are physiological measures corrected for specific gravity (SG). All smooths are not controlled for age to show cumulative pattern.
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ID = individual; T3 = total triiodothyronine; S-birth = sibling birth. .
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*
After sibling birth.
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†
Before sibling birth.
Generalized additive mixed model (GAMM) results of behavioral changes (nipple contact, riding, and body contact and 5 m proximity with the mother) in the older offspring around sibling birth (±2 years).
Binomial GAMMs on proportions of time per day and individual.
Reference | Nipple contact | Riding | Proximity | Body contact with mother | |||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Factor variables: | Category | Est. | SE | z | p | Est. | SE | z | p | Est. | SE | z | p | Est. | SE | z | p |
(intercept) | –7.34 | 0.84 | –8.73 | –1.36 | 0.52 | –2.65 | 0.21 | 0.16 | 1.30 | –2.79 | 0.19 | –14.50 | |||||
Males | Females | 0.76 | 0.61 | 1.24 | 0.21 | 1.18 | 0.36 | 3.22 | 0.001 | –0.08 | 0.13 | –0.66 | 0.51 | 0.22 | 0.11 | 2.05 | 0.040 |
After YS-birth* | Before† | 1.39 | 0.88 | 1.55 | 0.12 | –2.00 | 0.55 | –3.64 | <0.001 | 0.02 | 0.10 | 0.18 | 0.85 | 0.47 | 0.11 | 4.17 | <0.001 |
Year | - | - | - | - | - | - | - | - | - | - | - | - | - | - | - | - | |
Smooth term Variables: | edf | Ref. df | Chi² | p | edf | Ref. df | Chi² | p | edf | Ref. df | Chi² | p | edf | Ref. df | Chi² | p | |
T-S-birth: males | 4.45 | 4.95 | 12.38 | 0.002 | 1.00 | 1.00 | 0.03 | 0.86 | 1.00 | 1.00 | 6.91 | 0.009 | 1.00 | 1.00 | 12.48 | <0.001 | |
T-S-birth: females | 3.72 | 4.19 | 12.38 | 0.017 | 1.00 | 1.00 | 5.24 | 0.022 | 3.28 | 3.37 | 8.03 | 0.032 | 1.00 | 1.00 | 28.70 | <0.001 | |
T-S-birth: males | Females | 1.00 | 1.00 | 0.20 | 0.66 | 1.00 | 1.00 | 2.78 | 0.095 | 1.00 | 1.00 | 0.05 | 0.83 | 1.00 | 1.00 | 1.04 | 0.31 |
Age: males | 1.00 | 1.00 | 0.55 | 0.46 | 1.00 | 1.00 | 19.89 | <0.001 | - | - | - | - | - | - | - | - | |
Age: females | 1.00 | 1.00 | 0.12 | 0.71 | 1.00 | 1.00 | 4.39 | 0.036 | - | - | - | - | - | - | - | - | |
Age: males | Females | 1.00 | 1.00 | 0.00 | 0.99 | 1.00 | 1.00 | 4.38 | 0.036 | - | - | - | - | - | - | - | - |
Daytime | 3.64 | 3.92 | 14.31 | 0.012 | 3.51 | 3.85 | 7.46 | 0.09 | 3.81 | 3.98 | 170.67 | <0.001 | 3.96 | 4.00 | 456.4 | <0.001 | |
Seasonal effect | 1.10 | 3.00 | 1.89 | 0.021 | 8.89 | 3.00 | 1.91 | 0.063 | 0.00 | 3.00 | 0.00 | 0.05 | 2.64 | 3.00 | 61.22 | <0.001 | |
Random effects: | |||||||||||||||||
Time-S-birth per ID (smooth) | 3.13 | 113.00 | 82.07 | <0.001 | 33.32 | 70.00 | 251.9 | <0.001 | 62.11 | 76.00 | 1323.3 | <0.001 | 58 | 76.0 | 1569 | <0.001 | |
Age per ID (smooth) | 8.05 | 94.00 | 34.07 | <0.001 | 0.00 | 61.00 | 0.00 | 0.010 | - | - | - | - | - | - | - | - | |
Mother ID (intercept) | 2.45 | 10.00 | 0.00 | <0.001 | 0.00 | 10.00 | 0.00 | 0.001 | 6.28 | 12.00 | 0.00 | <0.001 | 0.01 | 12.0 | 0.01 | <0.001 | |
Date (intercept) | 3.65 | 1.00 | 0.00 | 0.23 | 0.00 | 1.00 | 0.00 | 0.28 | 0.00 | 1.00 | 0.00 | <0.001 | 0.00 | 1.00 | 0.00 | <0.001 | |
R2adj (deviance explained) | 0.39 (62.4%) | 0.827 (81.7%) | 0.226 (29.3%) | 0.319 (39.7%) | |||||||||||||
N (p-value, full/null comp) | 545 (<0.001) | 301 (<0.001) | 545 (<0.001) | 545 (<0.001) |
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Green: classic interaction term derived from a separate model calculation (see ‘Methods’). Statistics for year (categorical control variable) not shown for clarity.
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ID: individual; S-birth = sibling birth; ‘:’ = interaction term.
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*
After sibling birth.
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†
Before sibling birth.
Physiological changes during TTS
Urinary cortisol level changes in response to TTS
At the time of sibling birth, older offspring’s cortisol levels showed a significant and sudden, noncontinuous, up to fivefold increase from the level prior to this event (cortisol model with one sudden change; Figure 1A–C, Table 2). Compared to a model that allowed for nonlinear, but only continuous, fitting of the data (cortisol model without sudden change, Figure 1—figure supplement 1A and B), allowing for discontinuity (i.e., a sudden change) in cortisol levels at the time of sibling birth (cortisol model with sudden change), significantly improved model fit (Figure 1—figure supplement 1A and B; Chi2(1) = 9.30, p<0.001), even if the continuous model was allowed to be wiggly and to overfit the data (Figure 1—figure supplement 2A).

Physiological changes in cortisol (A–C), neopterin (D–F), and total T3 (triiodothyronine) (G–I) levels in the older offspring, 7 years before and after sibling birth (sibling birth at 0).
Data points are physiological measures corrected for specific gravity (SG). All smooths are not controlled for age to show cumulative pattern. Axes for physiological variables are log-transformed. 95% confidence intervals are plotted. Left-hand plots (A, D, G): sex-specific trajectories around sibling birth (blue: males; red: females). Middle plots (B, E, H): age-specific trajectories around sibling birth for offspring that were older (purple) or younger (yellow) than the median value of 5.1 years at sibling birth. Right-hand plots (C, F, I): interaction plots visualizing how trajectories around sibling birth change with increasing offspring age at sibling birth (scale from dark green [lowest levels] to brown [highest levels]; white space: extrapolation would be unreliable due to lacking data) for the respective perspective plots, see Figure 1—figure supplement 5. (A) Urinary cortisol levels showed a significant, sudden rise to fivefold values at sibling birth (dotted line); no sex differences or age effects. (B, C) The sudden rise in cortisol levels was independent of the age of the older offspring at sibling birth. (D) Urinary neopterin levels decreased by 1/3 at sibling birth (dotted line; no sex differences or age effects). (E, F) The sudden decrease in neopterin levels was independent of the age of the older offspring at sibling birth. (G–I). Urinary total T3 levels increased around sibling birth, but this effect was indistinguishable from a general age effect. There was no significant sudden change at sibling birth in total T3 levels (G), and there was no significant effect of the age at sibling birth (H, I).
Post-hoc visual inspection of urinary cortisol levels indicated that urinary cortisol remained high for a long time. None of the older offspring’s samples collected during the months after sibling birth had low cortisol levels (Figure 1A–C, Figure 1—figure supplement 3); cortisol measures in all samples collected within 7 months following sibling birth were above the upper 99.9% confidence interval of the values from before sibling birth. Lower cortisol values appeared later, only after 7 months post-birth (Figure 1—figure supplement 3). To verify this unexpected pattern, we ran another model allowing for an additional discontinuity in cortisol levels, that is, one at sibling birth and another one 7 months later. This model (Supplementary file 1) significantly improved model fit (cortisol model with two sudden changes compared to the model with only one sudden change at sibling birth, Figure 1—figure supplement 4A: Chi2(1) = 18.36, p<0.001; compared with the continuous model without sudden change, Figure 1—figure supplement 1A: Chi2(2) = 27.65, p<0.001). Cortisol levels in samples collected after the 7-month period were not different from before sibling birth (Supplementary file 1). While the model with the two discontinuities describes our data better mathematically, there is no obvious biological explanation for the second change (i.e., the sudden decline in cortisol) after 7 months. However, also in the model with only one discontinuity at sibling birth and a smooth continuous decline thereafter (Figure 1A–C), the cortisol levels took over 7 months to return to previous levels. Hence, the absence of low cortisol levels after sibling birth was evident in both models.
Cortisol trajectories around sibling birth were independent of the age of the older sibling. Allowing for different levels and trajectories in older and younger individuals did not improve the model (Figure 1B, Figure 1—figure supplement 4B; Chi2(3) = 0.28, p=0.91), suggesting that the cortisol level changes were not moderated by offspring age, a finding that was also apparent from visual inspection of continuous interaction plots (see Figure 1C, Figure 1—figure supplement 5A, B for perspective plots). Hence, the effect of TTS did not decrease with increasing age of the older sibling. Introducing two sudden changes, cortisol trajectories around sibling birth did not decrease with increasing age of the older sibling (Figure 1B, Figure 1—figure supplement 4B, C; Chi2(3) = 1.12, p=0.53) and sex of older sibling did not affect the results (Figure 1A, Table 2).
Urinary neopterin level changes in response to TTS
Just after sibling birth, urinary neopterin levels of older offspring decreased significantly and discontinuously (neopterin model with one sudden change, Figure 1D–F, Table 2). Compared to the model allowing for nonlinear but continuous fitting of the data (neopterin model without sudden changes, Figure 1—figure supplement 1D), a model with discontinuity in neopterin levels at the time of sibling birth significantly increased model fit (neopterin model with one sudden change, Figure 1D; Chi2(1) = 4.28, p=0.003), even if the continuous model allowed for extreme wiggliness and overfitting of the data (Figure 1—figure supplement 2B). Post-hoc visual inspection of neopterin data suggested a 4.5-month post-birth period with particularly low neopterin levels (all values during the 4.5-month post-birth period were below the mean from before or after sibling birth). Running an additional model, allowing a second discontinuity in neopterin levels at 4.5 months, slightly improved model fit (neopterin model with two sudden changes, Figure 1—figure supplement 4D–F; Chi2(1) = 1.95, p=0.048). However, even when allowing for a second discontinuous change, neopterin levels in samples collected after sibling birth remained significantly lower than before sibling birth (Supplementary file 1).
Model fit did not improve when we allowed moderation of this effect by the age of the older offspring at sibling birth (allowing for different pattern in older and younger individuals: Chi2(3) = 1.19, p=0.50, Figure 1E, F, Figure 1—figure supplement 4E, F), and again, there was no sex difference in neopterin levels before or after sibling birth (Figure 1D, Table 2).
Total T3 levels during TTS
Urinary total T3 levels increased around the time of sibling birth (Figure 1G–I), but this change could neither be attributed to the age of the older siblings nor to the event of sibling birth. The model including both variables was not significantly different from the null model (p=0.096). A reduced model including only the event of sibling birth but not age was significantly better than the null model (p=0.020, Figure 1G, Table 2). There was neither a significant sex effect on urinary total T3 levels during TTS nor a significant and sudden change in total T3 levels at sibling’s birth (Figure 1G, Table 2; allowing for sudden change: Chi2(1) = 1.27, p=0.11). Adding interaction terms with age of the older offspring did not improve the model, nor did it if allowed for differences between older and younger individuals (Chi2(3) = 0.40, p=0.85; Figure 1I).
Behavioral changes during TTS
Nipple contact during TTS
The proportion of time the older offspring was observed in nipple contact showed a continuous decrease prior to sibling birth in both males and females, and reached zero about 2 months before sibling birth (Figure 2A–C, Table 3). Consequently, there was no sudden change at sibling birth in terms of nipple contact (Figure 2A–C, Table 3; Chi2(1) = 0.81, p=0.20). Allowing for different trajectories depending on the age categories of the older offspring at sibling birth (younger or older than 5.11 years old at sibling birth) significantly improved the model (allowing for different pattern in older and younger individuals: Chi2(3) = 4.99, p=0.019) and visual inspection of the data indicated that nipple contact persisted mainly in younger offspring (Figure 2B and C, Figure 2—figure supplement 1A).

Behavioral changes in nipple contact (A–C), riding (D–F), body contact (G–I) and 5 m proximity (J–L) with the mother of the older sibling in relation to sibling birth (sibling birth is set to 0).
Vertical dotted lines = time of putative conception (left dotted line) and sibling birth (right dotted line). Data points represent the proportion of time and circle size the underlying sample size (square-rooted; ranges: riding 3–44, all other behaviors 3–303). All smooths are not controlled for age to show cumulative pattern. 95% confidence intervals are plotted. Left-hand plots (A, D, G, J): sex-specific trajectories around sibling birth (blue: males; red: females). Middle plots (B, E, H, K): age-specific trajectories around sibling birth for offspring that were older (purple) or younger (yellow) than the median value of 5.1 years at sibling birth. Right-hand plots (C, F, I, L): interaction plots visualizing how trajectories around sibling birth change with increasing offspring age at sibling birth (scale from dark green [lowest levels] to brown [highest levels]; white space: extrapolation would be unreliable due to lacking data) for the respective perspective plots, see Figure 2—figure supplement 1. (A–C) Proportion of time spent suckling decreased to zero already before sibling birth (A) and was largely absent in older offspring (B, C), without a sudden change at sibling birth. (D–F) The proportion of time riding on the mother showed a significant and sudden decline at sibling birth (D), but this cut was evident only in offspring younger than 5 years old at sibling birth and not anymore in older offspring (E, F). (G–I) The proportion of time spent in body contact with the mother showed a significant and sudden increase at sibling birth, irrespective of the sex or age of the offspring. (J–L) The proportion of time in 5 m proximity to the mother decreased around sibling birth, but this effect was indiscernible from a general age effect. There was no significant sudden change at sibling birth (J), and there was no significant effect of offspring age at sibling birth (K, L).
Riding on the mother during TTS
The proportion of time the older offspring was riding on the mother during travel continuously decreased before sibling birth, then showed a significant and sudden decline at the time of sibling birth, and remained low thereafter (Figure 2D–F, Table 3; allowing for discontinuity at sibling birth: Chi2(1) = 6.06, p<0.001). Overall, sons spent significantly more time riding on their mothers than daughters, and the continuous decline before sibling birth was only significant in daughters, whereas the sudden decline at sibling birth appeared to be stronger in sons (Figure 2D, Table 3).
Adding the older offspring’s age categories significantly improved the model (allowing for different trajectories in younger and older individuals: Chi2(3) = 9.32, p=0.001; Figure 2E). Visual inspection of the data showed that the sudden decline in riding at sibling birth was only evident in older siblings belonging to the younger age cohort (less than 5.11 years old at sibling birth), whereas older siblings in the older age cohort were completely independent from maternal carrying before sibling birth (Figure 2E and F, Figure 2—figure supplement 1B). Hence, the effect of TTS on riding disappeared with increasing age of the older sibling.
Independent foraging during TTS
There was no effect of TTS on the proportion of time that offspring spent foraging on their own at times when mothers were foraging, and none of the full or reduced models was significantly different from the corresponding null models. Visual inspection of model results revealed that the proportion of time spent foraging independently reached high levels before sibling birth and did not change during the time window around sibling birth that was considered in our models (Figure 2—figure supplement 1A–D). In fact, all subjects were rather independent in terms of foraging at the time of sibling birth, irrespective of their age. In particular, there was no significant discontinuity at sibling birth (Figure 2—figure supplement 2A–D).
Body contact and 5 m proximity with the mother during TTS
The proportion of time that older offspring spent in body contact with or in proximity (within 5 m) to their mothers showed similar trajectories relative to sibling birth (Figure 2G–L). Both variables decreased before and around the time of sibling birth, reaching low levels at the time of gestation (Figure 2G–L, Table 3). This pattern could be attributed neither to the age of older offspring nor to the event of sibling birth. The models including both age and time around sibling birth were not significantly different from corresponding null models (body contact: p=0.055, 5 m proximity: p=0.062) but models without age were significantly different from the respective null models (both p<0.001, Table 3).
For 5 m proximity, there was no sudden change at sibling birth (Figure 2J–L; allowing for discontinuity at sibling birth: Chi2(1) = 0.016, p=0.86), nor did the pattern change with the age categories of the older sibling at sibling birth (allowing for different trajectories in younger and older individuals: Chi2(3) = 0.33, p=1; Figure 2K).
For body contact, there was a significant sudden change at sibling birth (Chi2(1) = 7.60, p<0.001), but in contrast to what one would expect to see in case of social weaning, this change was a sudden increase in body contact with the mother (Figure 2G, Table 3). Allowing moderation of the TTS effect by the age of the older offspring did not improve the model (allowing for different levels and trajectories in younger and older individuals: Chi2(3) = 1.86, p=0.29; Figure 2H and I), and the sudden increase in body contact at the time of sibling birth was independent of the age of the older offspring at sibling birth.
Discussion
Our data from wild bonobos demonstrate that the birth of a sibling induced a sudden increase in urinary cortisol levels in the older offspring, a physiological response that occurred in all subjects regardless of their age. Upon birth of a sibling, urinary cortisol levels in the older offspring increased fivefold and remained at this level for about 7 months. Simultaneously, neopterin levels declined at the time of the birth of a sibling and remained at low levels for about 5 months. This suggests that the birth of a sibling induced a cortisol response and reduced or suppressed cell-mediated immunity in the older offspring. Older offsrping’s physiological changes around sibling birth did not decrease with increasing age of the older sibling and were independent of behavioral measures of weaning and attainment of physical independence. At sibling birth, weaning-related behavioral changes were either already completed (independent foraging and nipple contact), did not change discontinuously (urinary total T3, nipple contact, time in spatial proximity to mother, and independent foraging), changed suddenly in directions opposite of our expectation (increasing body contact time with the mother), or were significant only in subjects belonging to the younger age cohort (riding).
The fivefold increase in cortisol levels in our study is an unusually strong physiological response. For comparison, captive bonobos exposed to an experimental stress test exhibited a twofold increase in cortisol levels (Verspeek et al., 2021). A similar cortisol response occurred in bonobos in response to a group member giving birth, but in this case, the individual’s cortisol levels returned to previous values within 1 day (Behringer et al., 2009). In wild chimpanzees, urinary cortisol levels were found to increase by a factor of 1.5 when subjects encounter a neighboring group, an event that exposes all group members to potentially lethal aggression (Samuni et al., 2019). Changes in cortisol that exceeded the magnitude of the changes observed in our study occurred in a population of wild chimpanzees who experienced a tenfold cortisol increase during a respiratory disease, which killed a number of group members (Behringer et al., 2020). The intensity of a stress response is generally determined by the severity, controllability, and predictability of the stressor (Seiler et al., 2020); TTS is novel, severe, uncontrollable, and relatively unpredictable for the older offspring, all characteristics that likely contributed to the comparably high cortisol response that we observed in our study.
In addition to the age-independent, sudden, and substantial physiological response that we observed, a post-hoc analysis revealed that cortisol levels remained elevated for 7 months after sibling birth. Anecdotal reports indicate that, in wild chimpanzees, it may take up to 1 year until the older offspring adapts behaviorally to the presence of a younger sibling (Clark, 1977). While the physiological effects of sibling birth in human children are still unknown, behavioral data suggest that it may take up to 8 months until the older sibling adapts to the novel situation (Oh et al., 2017; Stewart et al., 1987). This indicates that humans, bonobos, and chimpanzees respond similarly to the challenge deriving from the arrival of a sibling.
The sudden decline of cortisol and neopterin levels to pre-sibling birth levels after 7 and 5 months, respectively, was unexpected. It is important to note that in the model with only one sudden change the cortisol levels needed many months to decline. While this result requires explanation, it is important to differentiate what our data can show and what remains to be explored in future studies. Regarding cortisol levels, our results do show that for a period of about 7 months none of the older siblings had low or average cortisol levels, but all had values within a narrow range of extremely high levels until they returned to their typical wide distribution. However, our data resolution does not allow the exact tracking of individual cortisol trajectories and it remains unclear at which time and speed different individuals return to ‘normal’ levels following the 7-month period. This aspect was even more pronounced in the case of neopterin levels. After the 5-month period of almost exclusively low neopterin levels, some individuals returned to previous levels but others remained low. Therefore, the time at which individuals return to ‘normal’ level, and the factors determining this shift remain to be investigated in future studies aiming at higher sampling rates per individual.
Cortisol levels are known to increase in response to psychological and social stressors, like predation risk or social instability, as well as energetic and physiological events (McEwen and Karatsoreos, 2020). Our study indicates that the sudden and persistent increase in cortisol levels in the older sibling was not related to energetic stress. Neither urinary total T3 levels, nor nipple contact, nor time spent foraging independently from the mother showed a sudden change at the onset of TTS. Similarly, if the cortisol increase at sibling birth would have been triggered by energetic challenges, the intensity of the cortisol level change should decline with the age of the older offspring as nutritional dependency on the mother decreases with age. In our study, the age of the older offspring at sibling birth ranged from 2.3 to 8.6 years, and preliminary analyses of stable isotopes in fecal samples collected from the same population suggest that nutritional weaning terminates at the age of 4.5 years (Oelze et al., 2020). However, the age of the older sibling at sibling birth had no effect on the strength of the cortisol response and the behavioral changes (body contact, nipple contact, riding, and independent foraging) did not follow the sudden shift in cortisol levels at the time of sibling birth.
In conjunction, our results indicate that the sudden increase in cortisol levels is independent from nutritional weaning effects and resembles behavioral responses of human children to the birth of a sibling (Dunn and Kendrick, 1980; Stewart et al., 1987). In human children, changes at sibling birth can be age-dependent. In response to sibling birth, scores for, for example, clinging and other gestures of reassurance were negatively correlated with the age of the older sibling (Dunn et al., 1981; Nadelman and Begun, 1982; Volling, 2012). Thus, in children, age seems to affect the behavioral response toward, or the perception of, the arrival of a sibling. Based on the results of our study, a sibling birth event is perceived similarly and independently of age. Hence, within the scope of our behavioral metrics, cortisol patterns did not match changes in single or cumulative behavioral changes around or after sibling birth.
Sibling birth is likely to cause multiple changes in the relationship between the mother and the older offspring, and only a few of them were considered in our study. For example, cortisol levels increase in response to positive arousal in children (Flinn et al., 2011), and while the newborn attracts the full attention of the mother it may also attract the older sibling’s interest. Accordingly, it is not possible to exclude that the response of older siblings was influenced by affiliative intentions. Mothers may not always tolerate interactions between siblings and might prevent the older one from initiating interactions, which can also result in frustration and a concomitant increase in cortisol (Gunnar et al., 2010; Stroud et al., 2000). At the time of sibling birth, the social environment of the older offspring is likely to change. For example, during the first weeks after birth, female bonobos tend to avoid large parties and forage alone or associate with a few other females (Douglas, 2014). This may lead to reduced rates of interactions with similar aged immatures and increased demand for social interactions with the mother who may not always be responsive to the needs of the older offspring. Another source affecting cortisol levels is aggression from group members. In bonobos, aggression against infants is rare but juveniles of both sexes can be exposed to physical aggression from adult males. Rates of aggression were found to increase with age of the immature target and were particularly high at times when mothers of targets had given birth (Hohmann et al., 2019; ). Thus, when females give birth, the older offspring is likely to be exposed to multiple challenges that may affect allostatic load and require the development of coping mechanisms, an achievement that requires time.
Although body contact between the older offspring and the mother decreased with age, it also suddenly increased for a short period after sibling birth. This response is not unknown: during TTS, juvenile marmosets increase proximity to parents (Achenbach and Snowdon, 1998), infant rhesus macaques intensify their effort to maintain contact with their mothers (Mandalaywala et al., 2014), and human children exhibit increased rates of clinging behavior (Volling et al., 2017). In our study, we did not find consistent effects of TTS on proximity within 5 m. If such changes in proximity and body contact reflect reduced maternal attention, the older offspring may aim to regain more attention from their mothers or other caregivers (Baydar et al., 1997). Reduced maternal attention could contribute to the increase in cortisol levels that we found, but it is still unclear why this change persists for several months. Moreover, the most consistent effect of TTS on offspring behavior in humans was a decrease in affection and responsiveness to the mother (Volling, 2012), which seems to contradict this interpretation. Alternatively, young female primates are known to show a high interest in new babies (Maestripieri and Pelka, 2002) and the increase in body contact may reflect the interest of the older offspring in the younger sibling.
The sudden increase in cortisol and the abrupt decline in neopterin levels in our study emphasize the homeostatic challenges affecting the older offspring during TTS. It is possible that the increase in cortisol levels negatively affected cell-mediated immunity. In other mammals, stress responses to weaning had a negative effect on immunity (Kick et al., 2012; Kim et al., 2011), and stressful events were associated with changes in immune function in humans (Herbert and Cohen, 1993). While short-term increases in cortisol levels enhance immune functions in humans, long-lasting elevations of cortisol levels – such as those found in our study – dysregulate immune responses (Dhabhar, 2014). In our study, urinary cortisol and neopterin levels recovered several months after sibling birth, indicating that individuals can cope with TTS to some degree, for example, by becoming habituated to the new conditions or by recruiting social support from other group members.
Persistent early-life cortisol elevations can affect an individual’s ontogeny, with long-lasting consequences for its fitness, affecting its growth trajectory, metabolism, social behavior, immunity, stress reactivity, reproduction, and life history strategies (Berghänel et al., 2017; Maestripieri, 2018; Seiler et al., 2020). In view of our results, such effects may contribute to the observed negative effects of sibling birth on the fitness of the older offspring in nonhuman primates (Thompson et al., 2016; Tung et al., 2016; Zipple et al., 2019). However, the impact of sibling birth is not necessarily that strong. For example, the presence of a sibling did not affect the hypothalamic-pituitary-adrenal axis later in life in baboons, but other early-life adversities had lasting consequences (Rosenbaum et al., 2020). The physiological effects caused by a normative stressor that affects most individuals, such as the birth of a sibling, should be under negative selection and would therefore be considered to be a nonadaptive trait. Alternatively, it has been suggested that early-life events of ‘tolerable stress’ (McEwen and Karatsoreos, 2020) may serve to prime subjects to develop stress resistance later in life. Moreover, TTS may accelerate acquisition of motor, social, and cognitive skills (Azmitia and Hesser, 1993; Maestripieri, 2018; Song et al., 2016). Siblings are not only rivals but also important social partners, and the presence of an older sibling can buffer behavioral and physiological changes in response to stressful events like TTS (Hrdy, 2011). Having an older sibling may enhance the development and survival of the younger sibling that contributes to the inclusive fitness of both the older sibling and the mother (Salmon and Hehman, 2015; Stanton et al., 2017). Returning to our study, future studies should integrate behavior and physiological measures to estimate the impact of TTS for the older sibling and explore the long-term effects of increasing cortisol levels. The combination of physiological and behavioral measures could help to disentangle why immature bonobos show such an intense cortisol response. This would allow testing the hypotheses that the novel mother–infant constellation is an expression of positive valence arousal or a normative change of maturation.
To our knowledge, our study on wild bonobos is the first to investigate the physiological response during TTS and, along with other studies on nonhuman primates. In many human cultures, inter-birth intervals are shorter and children are weaned at a younger age than in wild apes (Humphrey, 2010; Robson et al., 2006), despite humans having slower development and longer ontogeny. However, parental effort varies tremendously across human cultures and is often supplemented by intense allomaternal care (Hrdy and Burkart, 2020). Thus, it is possible that human children do not necessarily experience such extreme and long-lasting cortisol elevation. In some families in Western societies and traditional societies, allomaternal caregivers provide nutritional, physical, and mental support to older children (Baydar et al., 1997; Kramer and Veile, 2018), which may buffer physiological responses. However, when such social buffering systems are absent or weakly developed, as in some Western societies, older children may experience the birth of a sibling as a particularly stressful time. Studies in humans are generally biased toward middle-class families in Western industrialized countries (Fouts and Bader, 2016; Volling, 2012), and our study expands research on TTS to a nonhuman primate.
The results of our study showed that bonobos, one of humans’ closest living relatives, had high cortisol levels during TTS. Together with anecdotal evidence from chimpanzees (Clark, 1977), the information obtained in our study may shed light on the evolutionary history of the behavioral and physiological changes associated with TTS. More detailed comparisons are required to identify the emergence of behavioral and physiological traits related to TTS, their interactions, and fitness consequences. Yet, the results obtained from wild bonobos render support to the long-standing but untested and recently questioned assumption that the birth of a sibling is a notable event for the older offspring (Volling, 2012; Volling et al., 2017). It highlights the ubiquity of this pattern across individuals and age classes, and indicates that emergence of this developmental period may not be a derived trait. Interpretation of data of nonhuman primates in an evolutionary context can lead to unjustified generalization (Sayers et al., 2012), and it is important to note that behavioral responses to TTS in human children are highly variable and individual- and age-dependent, ranging from aggression, emotional blackmailing, and psychological disturbances, to positive attitudes toward the new family constellation (Volling, 2012; Volling et al., 2017). This raises questions regarding the coping strategies and how they are (a) influenced by the socioecological conditions including actual parent–offspring and other caretaker relationships, (b) effective in modulating and buffering the shown physiological stress response, and (c) their phylogenetic history (Hrdy, 2011; Lonsdorf et al., 2018).
Methods
Study site and species
Data were collected from wild bonobos (P. paniscus) of the Bompusa West and East communities, at LuiKotale, Democratic Republic of the Congo. This bonobo population was never provisioned with food and lives in an intact, natural forest habitat. All subjects were habituated to human presence before the start of the study, were genotyped, and were individually known. We considered every offspring only for the next sibling birth; therefore, all older offspring in our study experienced the birth event for the first time. At the time of sibling birth, the older siblings were between 2.3 and 8.6 years old. Behavioral sampling included 397.17 hr of focal data on 11 immature females (mean = 36.11, SD = 14.70) and 253.95 hr on six immature males (mean = 42.33, SD = 27.62). Physiological measurements were performed using 319 (220 females, 99 males) urine samples of 20 females and 6 males (see Supplementary file 2).
Behavioral data collection and analysis
Behavioral data were collected between July 2015 and July 2018 via focal animal sampling (Altmann, 1974) whereby an infant was observed for 1 hr and its instantaneous behavior recorded at 1 min intervals (a detailed description in Lee et al., 2020). Data points were only included when focal subjects were continuously visible throughout the focal interval. Behaviors included nipple contact, defined as the infant applying its mouth to the nipple of the mother in a suckling manner, and riding, defined as the infant being transported as it clings ventrally or dorsally to its mother. For riding, we only considered data where the mother was traveling for at least three consecutive minutes to exclude situations where the mother was likely traveling for short distances only and riding on the mother would not have been important for the offspring. We recorded when the offspring was in body contact or within 5 m proximity to the mother and when it was foraging independently (i.e., searching for its own food instead of being food provisioned by the mother). For independent foraging of the offspring, we only considered scans where also the mother was foraging to cover typical foraging situations and reduce the influence of potential sampling bias, with foraging encompassing handling and ingesting food. For all other behaviors, all scores were considered and we calculated the proportion of instantaneous records per observation day.
Urine sample collection and analyses
Urine samples were collected between July 2008 and August 2018. Samples were collected opportunistically throughout the day between 5 am and 6 pm capturing urine directly from leaves or pipetting urine from the vegetation. Samples that were contaminated with feces were excluded. Samples were protected from direct sunlight to avoid degradation and stored in liquid nitrogen upon arrival at camp on the same day. Samples were shipped frozen to the Max Planck Institute for Evolutionary Anthropology in Leipzig, Germany, for cortisol and total triiodothyronine analysis, and later to the German Primate Center, Göttingen, Germany, for neopterin measurement.
Our urine dataset consisted of 16.0 ± 5.6 samples per individual (mean ± SD), with on average 7.5 samples before and 8.4 samples after sibling birth. Urine samples were temporally normally distributed around the day of sibling birth. Urine samples were collected from all individuals also during the first year after sibling birth, though one male and two females did not contribute samples during the first 7 months after sibling birth, and therefore, contributed only to the estimates of the urinary cortisol levels before and after the elevated cortisol period (see ‘Results’).
Frozen samples were first thawed at room temperature, shaken for 10 s (VX-2500 Multi-tube Vortexer), and centrifuged for 5 min at 2.000 × g (Multifuge Heraeus), after which specific gravity (SG) was measured using a refractometer. All results were corrected for SG to adjust the concentration of the physiological marker for urine concentration of the specimen, which depends on an individual’s hydration status and time since last urination (Miller et al., 2004). Aliquots of samples were prepared at this time for later neopterin and total T3 analyses. In order to exclude a methodological effect concerning the order of the samples, for example, that all post-sibling birth samples are run together, all samples were randomly assigned to the measurements.
Urinary cortisol analyses
We extracted and measured urinary cortisol in 319 (220 females, 99 males) urine samples of 20 females and 6 males. Cortisol extraction from urine samples was performed following the protocol described in Hauser et al., 2008 for liquid chromatography–tandem mass spectrometry (LC-MS/MS) analyses. Each urine sample was mixed with an internal standard (prednisolone, methyltestosterone, d3-testosterone, d4-estrone, and d9-progesterone). Prednisolone was used as an internal standard to assess sample recovery and quantify urinary cortisol levels. We performed hydrolysis using β‐glucuronidase from Escherichia coli (activity: 200 U/40 μl). Extracts were purified by solid-phase extractions (Chromabond HR-X SPE cartridges: 1 ml, 30 mg), followed by a solvolysis with 2.5 ml ethyl acetate and 200 mg sulfuric acid. The extraction of cortisol was carried out with methyl tert-butyl ether. Finally, we reconstituted evaporated extracts in 30% acetonitrile.
For urinary cortisol measurement, we used a LC-MS/MS with a Waters Acquity UPLC separation module equipped with a binary solvent manager, sample manager, and a column oven (Waters, Milford, MA). A Waters Acquity BEH C18 column (2.1 × 100 mm, 1.7 μm particle diameter) was used for chromatographic separation. Eluent A was water with 0.1% formic acid and eluent B was acetonitrile. We injected 10 μl of sample extract. The quantitative analysis of cortisol levels was realized in the range of 0.01–100 pg/μl. For cortisol quantification, we used MassLynx (version 4.1; QuanLynx Software). Final urinary cortisol results are represented in ng/ml corrected for SG. We accepted measurements of a batch if quality control measurements deviated less than 15% from the true cortisol concentration. Seventeen samples in which internal standard recovery deviated by more than 60% of the internal standard were remeasured via reinjection. In two samples, measurements were above the limit of the calibration curve, and were reinjected at a 1:10 dilution.
Urinary neopterin analyses
We measured urinary neopterin in 314 (215 females, 99 males) aliquots of 20 females and 6 males with a commercial neopterin ELISA for humans, previously validated to determine neopterin in bonobo urine (Behringer et al., 2017). Prior to neopterin measurement, urine samples were diluted (1:10–1:200 depending on SG) with the assay buffer provided by the supplier. We added to each well on the plate 20 µl of the diluted urine, 100 µl of the provided enzyme conjugate, and 50 µl of the neopterin antiserum. The plate was covered and incubated on an orbital shaker at 500 rpm in the dark for 90 min. The plate was then washed four times with 300 µl washing buffer, and 150 µl of tetramethylbenzidine substrate (TMB) solution was added. The plate was incubated again for 10 min, and the reaction was stopped by adding 150 µl of the provided stop solution. Optical density was measured photometrically at 450 nm.
All samples were measured in duplicates according to the supplier’s instructions. Inter-assay variation for high- and low-value quality controls was 4.2 and 1.7% (N = 17 assays), respectively. Intra-assay variation was 8.9%. Final neopterin concentrations are expressed in ng/ml corrected for SG.
Urinary total T3 analyses
We measured total T3 in 319 (220 females, 99 males) urine aliquots of 20 females and 6 males with a commercial, competitive total triiodothyronine (T3) ELISA (Ref. RE55251, IBL International GmbH, Hamburg, Germany). Samples were measured with a 1:2, 1:5, or without dilution depending on SG. Then, 50 µl of the diluted sample with 50 µl of the provided assay reagent was pipetted into a well. We shook the plate for 10 s and incubated the plate afterward for 30 min at room temperature. We then added 50 µl of the provided triiodothyronine-enzyme conjugate to each well, shacked the plate again for 10 s, and incubated it again at room temperature for 30 min. We then washed the plate five times with 300 µl of the washing buffer and added 100 µl of TMB substrate. After 10 min of incubation, we stopped the reaction with 100 µl of the provided stop solution and read the plate at 450 nm with a microplate reader.
All samples were also measured in duplicates. Inter-assay variation for high- and low-value quality controls was 6.3 and 5.6% (N = 25 assays), respectively. Intra-assay variation was 7.2%. Final total T3 concentrations are expressed in ng/ml corrected for SG.
Statistical analysis
All statistical analyses were performed with R 4.1.3 (R Core Development Team, 2020), and all R-codes can be found in the data depository. We applied GAMM, which allow for the detection and analysis of complex nonlinear relationships (termed ‘smooths’) that are typical for developmental trajectories. We used function gam for all models (package mgcv; Wood, 2017), with smooth estimation based on penalized cubic regression splines. We checked for model assumptions and appropriate model settings using functions gam.check (package mgcv), and all models were inspected for and showed negligible autocorrelation (function acf_resid, package itsadug; van Rij et al., 2020) and overdispersion (functions testDispersion and testZeroInflation, package DHARMa; Hartig, 2021). Model comparisons were conducted using the function compareML (package itsadug). GAMM smooths were plotted using package itsadug (van Rij et al., 2020) with removed random effects. As typical for GAMMs, interaction terms with factor variables were calculated in two ways, first analyzing whether significant changes occur within each level of the grouping factor, and second whether the smooths of the different levels differ significantly from each other (the classic interaction term statistic) (Wieling, 2018; Wood, 2017).
Urinary physiological data (urinary cortisol, total T3, and neopterin) were normally distributed after log-transformation, and Gaussian GAMMs were applied. The GAMMs on mother–offspring relationship (nipple contact, riding, independent foraging, body contact, and proximity) were based on single minute-by-minute focal scan records that were summed to time proportion values per day and individual, hence we applied GAMMs with a binomial logit-link error structure on proportion data and the underlying number of scans per proportion value as weight-argument. The main predictor variable of all analyses was the temporal change of the respective response variable around sibling birth, allowing for potential sex differences (Behringer et al., 2014; Leigh and Shea, 1996).
Time around sibling birth was added in two ways into the model, first as a continuous smooth term across time, and second as a factor variable coding for the time before and after sibling birth, thereby allowing for a sudden, noncontinuous and unconnected change right at sibling birth. This combination allowed us to model a discontinuity at sibling birth in response values (though not in the first derivative and thus the slope of the smooth) while at the same time avoiding the pitfalls of calculating separate smooths for before and after sibling birth. Significance of the discontinuity was estimated through model comparison.
Additionally, these models included potential mediating effects of age to control whether apparent TTS effects were in fact mere general age effects irrespective of TTS. Age and time around sibling birth were naturally 100% correlated within individuals and highly correlated within the entire datasets (range r = 0.659–0.855).
In a further step, we expanded these two terms of time around sibling birth to interaction terms incorporating offspring age at sibling birth to investigate a potential moderation effect of offspring age on the intensity and pattern of potential TTS effects. For this purpose, we ran two different models. First, we ran the above model but replaced the continuous age terms with a binomial variable differentiating between offspring that was older or younger at sibling birth than the median age at sibling birth (5.11 years old). We estimated the significance of a potential difference between these two age groups in trajectories around sibling birth by comparing this model with a model without this differentiation. Second, we ran a model including an interaction term between age at sibling birth and time around sibling birth, to show visually how trajectories in the response variable around sibling birth change with increasing age at sibling birth, and in particular whether specific pattern and discontinuity around sibling birth ceased with increasing offspring age.
All statistical GAMMs were controlled for repeated measurements per individual via (a) two random smooth effects (factor-smooth-interactions; for details, see Wood, 2017), one for individual changes over time relative to sibling birth and the other for individual changes with age (for those models that included age as predictor variable), and (b) a random intercept per mother since some mothers contributed multiple offspring. All GAMMs were controlled for year (as random intercept for hormonal data but as control variable for the 3 years of behavioral data), seasonal effects via a cyclic smooth term over the year, and for daytime effects via a smooth term over daytime. The binomial models on behavioral time proportion data included an additional random intercept of date to control for multiple measurements per day. Due to the structure of the interaction models combining age at and time around sibling birth into one interaction term, we did not additionally control for a general mediating age effect, but merged the random effects on age and mother ID to one random smooth term of age at sibling birth per mother ID.
In all models, the number of basis functions (k) was always set equal for all predictors and random smooths of time around sibling birth and of age. The number of basis functions was generally set to 10, but needed to be reduced to 6 in some cases for the full models including both a term for age and for time around sibling birth due to sample size (for all physiological variables and for riding). Additionally, k needed to be reduced to 6 also for all models on body contact and 5 m proximity to the mother since higher values often led to strong overfitting and uncertainty. We further tested for robustness of the estimated smooth parameters by setting the number of basis functions to the respective maximum value (for models without continuous age terms), which was k = 12 for all physiological responses, k = 15 for riding, and k = 25 for all other response variables. Patterns of smooth trajectories remained the same (also for body contact in this case), though naturally the parallel increase of k for both the predictor and the associated random smooth terms led to increasing identifiability constraints and thus increasing estimation uncertainty.
Data availability
Source data for statistics and figures in the paper is permanently stored at GRO (https://doi.org/10.25625/O1OD2I).
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Göttingen Research OnlineReplication Data for: Transition to siblinghood.https://doi.org/10.25625/O1OD2I
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Decision letter
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Stacy RosenbaumReviewing Editor; University of Michigan, United States
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Christian RutzSenior Editor; University of St Andrews, United Kingdom
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Iulia BadescuReviewer
Our editorial process produces two outputs: (i) public reviews designed to be posted alongside the preprint for the benefit of readers; (ii) feedback on the manuscript for the authors, including requests for revisions, shown below. We also include an acceptance summary that explains what the editors found interesting or important about the work.
Decision letter after peer review:
Thank you for submitting your article "Transition to siblinghood causes substantial and long-lasting physiological stress reactions in wild bonobos" for consideration by eLife. Your article has been reviewed by 3 peer reviewers, and the evaluation has been overseen by a Reviewing Editor and Christian Rutz as the Senior Editor. The following individual involved in the review of your submission has agreed to reveal their identity: Iulia Badescu (Reviewer #3).
The reviewers have discussed their reviews with one another, and the Reviewing Editor has drafted this to help you prepare a revised submission.
Essential revisions:
1) The link to the data and code needs to be fixed so that the reviewers can evaluate the data for themselves.
2) There are two potential options for addressing the interpretation issues raised by reviewer #2. The authors may either supplement their analyses with additional behavioral data that speak more directly to stress (the preferred approach) or adjust the framing and interpretation to better suit the ambiguities of the data. If the authors choose to exercise this option, I would suggest steering away from the developmental psychology perspective the paper currently relies on, and more towards physiological mediators of life history transitions. The comments of reviewers #1 and #3 can both help to guide this since they offer related advice on framing.
3) The authors need to directly address the somewhat puzzling discontinuity in cort concentrations that occurs around 7 months post-sibling birth; what is the biological plausibility of this, and what factors might explain it? How does the non-recovery of neopterin fit with the return of cort to pre-birth baselines?
4) Please pay careful attention to the specific suggestions offered by all three reviewers. The paper would benefit greatly from additional details about the analyses and the motivations for specific components of them, to better suit a general-interest audience.
Reviewer #1 (Recommendations for the authors):
Introduction
Lines 51-52: What does mere behavioral adjustments mean? This requires some further explanation. I assume that it is referring to e.g. the withdrawal of maternal support, etc., but it could be interpreted in different ways.
Lines 65-67: Are there really no data available on this for non-human primates? I find it hard to believe that no one has quantified the changes in mother-older offspring relationships after the birth of a new infant. But I could certainly be wrong!
Line 86: It sounds like Schino and Troisi fulfil what I was asking for above, re: a non-human primate reference-this would probably be a good reference to add above, depending on the specifics of the findings.
Line 121: What is meant by 'general changes' in cortisol levels?
Line 125: It would be helpful to quantify what rather independent (versus highly dependent) means. Spending X% of time within X meters of mom? Is it a nursing measure? Some combination?
Line 131: This is a place in which it feels like the cart is coming before the horse. It seems odd to frame this as disentangling something which we haven't established IS entangled. This would feel appropriate if, for example, this paper were building on human studies that showed a cort increase after the birth of a sibling but did not control for changes in time near mother.
Line 132: When I hear the term stress response I think of short-term, acute changes in cort. Is it more accurate to characterize this as changes in baseline cort, given the duration of something like a transition to siblinghood? To avoid this issue entirely, you could just say 'to assess physiological stress before and during TSS.'
Paragraph on lines 132-145: This paragraph is a complicated mix of background info, theoretical justification, and methods. It would be easier to read if these were separated out. W/r/t the methods here, the T3 and neopterin measures do not feel well-justified. Due to my specific background knowledge, I understand why these measures are included in the study, but for a general interest journal, this feels like it needs more explanation. It assumes a fair amount about what the reader already knows.
Lines 158-159: I'm not sure that I am a fan of the term 'age-related weaning.' Cortisol changes with age. Cort might also change due to weaning. While obviously the specific age-related changes should be trivial in the window in which weaning is occurring, this term feels like it conflates two separate processes that could each impact cort concentrations.
Results
I personally would like to see the behavioral results come before the physiological results, since potential behavioral changes are part of what the authors are proposing might drive any physiological changes.
How is independent foraging defined? This should be clarified in the results themselves, since the methods come later.
Line 251: I would say before the sibling was conceived, instead of the conception of the mother. This could be read as the mother herself being conceived.
Generally: is there any possibility that there are 'batch effects' going on in the cort data? I.e., were all of the post-birth to 7 month samples part of the same run?
Discussion
Paragraph on lines 342-357: Potentially bolstering this argument, Rosenbaum et al. 2020 (PNAS, 117(33), 20052-20062) found that having a close-in-age sibling did not predict higher adult fecal glucocorticoid levels in the Amboseli baboons. Since they feature so heavily in these citations this seems like a good way to connect the specifics of this study to the Amboseli findings.
Line 368: I would avoid the use of the word 'modern' here. I think the point is that there is a lot of variability in levels of allomaternal care amongst living humans, and that this variation may play an important role in how stressful (or not) children find TTS to be.
Line 373: I cringe at the idea that humans are not natural…we may be an unusual animal species, but this isn't the same thing 'unnatural' (a vague and here-undefined term that means different things in different parts of the literature). It would be more accurate to say something about it contributing to the comparative literature by showing that this transition happens in a closely related species, if you want to turn the focus back to humans.
Line 380: What exactly is meant by 'constellation and co-parenting?' The sibling isn't co-parenting.
Methods
I'm still not sure how independent foraging differs from just foraging. Is there such a thing as non-independent foraging? If so, what does that look like?
Lines 437-462: Unfortunately, I am not well-versed enough in LC-MS methods to evaluate this description, so I am taking it on faith that this is all standard and reasonable. My hope is that other reviewers will be better qualified to speak to this than I am.
Line 483: I assume shanked is meant to be shookϑ.
Figures and Tables
Figure 1 in general: I think there are typos in the legend for this figure. It never refers to the right-hand panels. In general, the figure captions need to be better explained. Figures should be stand-alone, and as presented these are not.
Reviewer #2 (Recommendations for the authors):
Lines 135-136: This same group of researchers also found that cortisol is high leading up to the days before Christmas, when children were excitedly anticipating presents from Santa (Flinn et al., 2011; https://doi.org/10.1016/j.neubiorev.2011.01.005)-not exactly traumatic. Flinn et al. interpret these rises in both kinds of situations as "arousal to social opportunities", which might be a valuable perspective for the authors to consider in their own dataset.
Analyses: For the GAMs, it would be good to see some information on how robust the presented splines are to alternative numbers of basis functions and alternative smoothing parameters.
Relatedly, I'm a little sceptical of the approach of estimating separate splines before and after sibling birth, for all of the causal inference considerations that regression discontinuity entails. A useful companion to these 'discontinuous' splines, especially for Figure 1 and Figure S1, would be estimates of a single, continuous spline for time relative to sibling birth. It's hard to know from eyeballing it, but my guess is a jump in cortisol would still be apparent, for example, but it would be more gradual and less striking than the current figures suggest.
Line 213: Is this Chi-squared value supposed to be negative?
Discussion: My public review mentions that it is difficult to interpret the finding of increased cortisol following sibling birth. At some points, the authors appear to recognize this, as they raise a number of good arguments in the discussion about potential factors leading to a TTS-timed cortisol increase. Some, like a sibling birth coinciding with the beginning of juveniles experiencing greater male aggression, have nothing to do with TTS per se. Other arguments imply a role for beneficial behavioral interactions between the older and younger sibling. Yet, the authors appear to dismiss those concerns and conclude that the cortisol increase indeed implies stress. First, I recommend at minimum addressing why numerous alternative interpretations-some that the authors identify, and some that I have identified-should be disregarded in favor of the "cortisol = stress" conclusion. Second, as I state in the public review, having data on behavioral interactions thought to be stressful would go a long way towards solidifying the interpretative elements of this paper. I don't know whether this sort of information is available, but if it is, I recommend integrating it into a revision of this manuscript. One of the main lessons from the developmental psychology literature is that there is substantial heterogeneity in TTS adjustments, which may be attributable to family dynamics or larger ecological factors. Being able to test these kinds of theories in bonobos would significantly boost the impact of this paper.
Line 304-305: The authors claim that "findings from human children show behavioral responses to sibling birth [are] independent of their actual age". True, some show that, but others do not; many claim younger children exhibit larger behavioral disruptions (e.g. Nadelman and Begun, 1982; Volling, 2012). Some expansion on why age might matter in some aspects of adjustment, but not others, would provide useful context to the discussion.
Figure 1: The right panels of Figure 1 (B, D, and F) will be very hard to interpret for the average reader: the figure captions are sparse, the contours have labels that are so small as to be invisible, no justification/explanation is given for the extrapolation parameter (and thus it won't be clear to the vast majority of people why the figure is splotchy), etc. I understand the objective of contour plots for visualizing non-linear interactions, but as they stand the figures and/or captions need to be changed to form a self-contained explanation, as figures should. I leave edits up to the authors' discretion: they could do some combination of beef up their figure captions; present a more traditional interaction plot of linear effects (i.e., present marginal trends of -1, 0, and +1 SE on offspring age), since all the splines in Figure 1 are very close to linear; or present a different kind of plot, like a perspective plot, or a plot of predicted splines using the get_predictions function in itsadug.
Figure 2: I think the caption for supplementary Figure 2 is incorrect-there is no depiction of time relative to sibling birth in this figure, just the age of the older sibling. It appears the caption was incorrectly copied over from part of Figure 2.
Data: The DOI linked for the source data doesn't work. I would need to re-evaluate the paper after having access to the raw data, so I can verify the analytical reproducibility of the key claims.
Reviewer #3 (Recommendations for the authors):
Abstract:
I would not identify the birth of a sibling as a developmental transition. Perhaps would be better to say a major life transition or major transition in the early life of an individual. Or if you stick with the word development, perhaps it would help to elaborate a bit to make clear why it is a developmental transition. Maybe it applied to nutritional development so that the mother-infant nutritional relationship must end with the birth of a new sibling? Perhaps you are referring to social development? Elaborating a bit might make clearer what you mean.
Should be: "Studying the transition", "in the mother-infant relationship".
Change to "evolutionarily old"- although evolutionarily old is a bit vague. Perhaps you can narrow it down to a specific time based on comparisons with other primates or mammals. For instance, you might say that this effect was likely present in the common ancestor of all the great apes. Something more specific to your study.
Line 43: change "while still being dependent" to "while they are still dependent"
This sentence also needs a reference- also do you mean children in humans only? I guess that is what you mean but perhaps you should extend this to all primates or long-lived, slow developing mammals. You could say that in mammals with slow rates of growth and development that give birth to single offspring at a time, most offspring are exposed to the birth of a younger sibling.
Extending this to other species could make it more relevant to more researchers.
Line 45: competitor in what sense? competitor for maternal attention and resources, or for food in the environment, or for reproduction or for social partners etc. I think this needs to be more specific.
Line 49: Do you mean behaviors shown by the older sibling? this is not clear. make clearer that it is the older sibling who could show increased aggression, clinginess, and depression. Otherwise, could be perceived to be in the mother or in the new sibling.
Line 51: not sure what you mean by mere behavioral adjustments. I think the point you are making here is important, but couldn't behavioral adjustments be in response to stress. I guess you are trying to say that these are behavioral changes that could occur in response to new siblings that are not associated to stress for the older sibling. Make clearer.
Line 94-97: I do think this statement needs references to support it.
Line 98: I think this statement also needs some references and a range in inter-birth interval lengths for bonobos, including a variety of sites, since you have the ones for your site specifically, below.
Line 101: should be social dependency on the mother.
Line 106: would co-dependency on the mother be better than co-residence? Co-residence sounds like one offspring needs to leave the natal group at some point, which is not always the case. Co-residence with siblings can occur for their entire lives if they are the philopatric sex.
Line 110-111: Sexual maturation at 4 years of age is very young… Many individuals might still be nursing at 4 years old. Perhaps your definition of the onset of sexual maturation needs to be explained. Also, unclear how the onset of sexual maturation is different from the onset of menarche, and how these two milestones are different, so this should be cleared up as well.
Introduction in general:
You've made many comparisons between bonobos and humans, which is good, but I wonder if it would be good to add in data and comparisons with other apes as well (chimps, gorillas, orangutans). I know you mention chimpanzees once or twice, but I wonder if it would help to make more explicit comparisons throughout with the other great apes. I say this because eventually in your discussion, you are going to be making some preliminary conclusions about when various developmental and stress mechanisms would have evolved and saying that the timing would have been in the last common ancestor between bonobos and humans may be incomplete. Inherently we would want to hear about the chimpanzee data at least, to know if we should think about the last common ancestor with Pan, but of course we would wonder about any evidence in the other great apes. If the other great apes show similar patterns in life history, development and maternal investment that you are pointing out for humans and bonobos, then the findings of your research here could also apply to them (pending studies in these other apes). Alternatively, there might be some key differences in the life history patterns and infant development of different apes, so it would be good to know about them to later understand why you may not think that the findings extend to a certain genus.
I suggest you try to keep the tense for your article in the past tense as much as possible. As it is now, you switch quite a bit between present and past tense, and at times, you use present tense but this is a bit awkward. Past tense would be best.
Line 123: You should mention the name and place of your study population here since you are bringing it up to set up your study.
Line 125: this statement on the dependency levels of the offspring needs to be supported with a reference and it needs to be quantified in some way. Is dependency based on nipple contacts, proximity, sleeping in the same nest, being carried from place to place, all the above?
Also, unclear if this statement refers to past research or if this is something that is being done in this paper. Did someone already investigate the nutritional development of these individuals, or is the level of dependency something you will be measuring in the present study and measured relative to TTS? This needs to be made clearer.
Lines 139-140: I think this sentence needs some references. Also, you explain T3 in greater detail but not the neopterin hormone. I think a sentence specifically on neopterin and how it works or what it means can help here.
Lines 140-141: I am not quite understanding T3 can help disentangle effect of energetic stress versus social stress. Can you please make this more obvious/clearer?
Line 143: unclear why neopterin levels should decrease because we do not have enough information about neopterin.
Line 146: you say nursing here but later you define suckling as contact with the nipple. Might be better to stick with one of the two terms, especially since people are going to keep an eye out for the term nursing to see how this was defined later in the text.
Line 148-150: Are you saying that social weaning is weaning from the mother more generally, in terms of proximity and access to carrying, etc.? This is very confusing because weaning in most infant development/maternal investment literature refers specifically to nursing or feeding behavior. Until this point in the article, I was thinking you were trying to make a distinction between nutritive and non-nutritive nursing.
If you would like to talk about changes in the mother-infant relationship outside of nursing and foraging behavior, I strongly encourage you NOT to use the term weaning for that. Perhaps switch to "attainment of physical independence" or some other similar phrase since the only other measures outside of nursing behavior that you are looking at are more physical measures (so the infant being carried versus moving on its own/ infant in certain proximity to the mother). Instead of nutritional and social weaning, you could say weaning and attainment of physical independence.
Line 193: With increasing age, urinary…
Also, you say that they significantly changed in males but not females, but then you say that this change was not significant… I am confused.
Line 210-211: same issue here. you say significantly declined but then in brackets you said sex difference not significant. Which is it?
Line 75, 262 and elsewhere: You keep talking about nutritional versus social weaning (or as others have said, nutritional versus behavioral weaning), but this concept is fairly recent. The idea that there are two components to weaning, the milk-transfer and the psycho-social relationship between the mother and the infant through continued nipple contacts, regardless of whether or not milk transfer occurs, is a fairly new and innovative idea that needs to be supported with literature in your paper, and that needs to be explained a little bit somewhere in the text. For example, research on wild chimps and other primates has shown that comfort nursing, without milk transfer, can occur for years after lactation has ended. This sets up a situation where you have these two separate weaning periods: weaning from milk and weaning from nipple contact- so the nutritional versus social/behavioral weaning. Thus, the mother-infant behavioral relationship can develop separately from the mother-infant nutritional relationship, despite considerable overlap between the two. This distinction, and its importance for the infant and mother, should be explained.
Line 276-279: Are you saying that TTS is severe, uncontrollable for older offspring, and was unpredictable or moderately unpredictable? Make clearer how this statement relates to your results.
Line 347-349: change sentence to
"However, strong negative effects caused by a highly predictable and normative stressor that invariably affects most individuals, such as the birth of a sibling in apes, should be under negative selection, and would thus be expected to XX XX (explain what this would mean)."
Line 298: I would change this to
"…suggest that nutritional weaning is usually completed by around 4.5 years of age".
Lines 372 to 372, or more broadly for this whole paragraph: Similar to my comment for the abstract, the statement that your results highlight the evolutionary history of stress response and TTS is vague and almost seems like a throw away idea because it is not specific enough. Can you go a step further and talk about the possible timing of an evolutionary link between stress and TTS? Presumably, this important interaction would have appeared with the great ape transition? Or would it be with the transition between genus Pan and Homo? Use the comparisons in life history traits and infant development of humans and the other great apes to infer when this mechanism that you found could have become more important or more prominent.
Methods:
Line 397: I would like to see a table with a breakdown, by infant age and sex, of the sample sizes and numbers for the behavioral and urine data.
Lines 399 to 411: Did you exclude from the calculations time out of view, or time when the infant was ventral, but it was unclear if they were in contact with the nipple, and excluded time in a nest with the mother since you can't see the infant? If yes, say so, if not, justify why you did not.
I am not familiar with any of the urine analyses so cannot comment on these.
Please explain how the proportions of the behavioral measures were calculated somewhere in the methods.
[Editors' note: further revisions were suggested prior to acceptance, as described below.]
Thank you for resubmitting your work entitled "Transition to siblinghood causes substantial and long-lasting increase in urinary cortisol levels in wild bonobos" for further consideration by eLife. Your revised article has been evaluated by Christian Rutz (Senior Editor) and a Reviewing Editor.
The manuscript has been greatly improved, but there are some remaining issues that need to be addressed before it can be accepted for publication.
– First, and most importantly, I (the reviewing editor) strongly urge you to consider the comments of reviewer #2 w/r/t the plausibility of the two-intercept model. While I understand that this model provides the best fit statistic, in my opinion, fit statistics should not override biological plausibility. There is no biologically plausible explanation for why cort would suddenly drop at 7 months (or at least, none that have been relayed in this text), and basic data visualization -- i.e., the raw scatterplot of the data--does not suggest this is what is happening. I personally feel that the single-cut discontinuous spline is a good middle-ground choice (due to the suddenness of sibling birth), followed by a continuous spline, followed by the two-cut discontinuous. The last two options would certainly be worth including in the supplementary materials.
This issue is the primary reason that I am recommending a revise and resubmit, rather than an acceptance. Please note that my acceptance of the manuscript is not contingent upon making this change. I respect that authors may have different points of view surrounding the interpretation of fit statistics. However, if it is not changed, the final evaluation summary will reflect the fact that myself and one of the other reviewers disagreed with the analysis strategy. I want to be transparent about the source of the disagreement so that the evaluation summary would not come as a surprise should you choose not to change which model you prioritize in the main text.
– Please include a scatterplot of the raw data in the supplementary materials.
– Reviewer #2 is correct that though the language around 'stress' has been considerably improved, there are still places where there are ambiguities in how it is used/what it is implying. The manuscript will be stronger with these ambiguities removed. These data are quite interesting and do not need to lean on 'stress' in order to be noteworthy and important. Similarly, I agree with reviewer #3s concerns about characterizing TTS as a developmental stage. Both of these are wording issues that should be simple to resolve.
– Please follow reviewer #3s request for a thorough proofread. There are a considerable number of spelling and grammar mistakes that remain, which is distracting as a reader. They have many helpful suggestions for places where the language could use additional clarification, which again, will help strengthen the final product.
Reviewer #2 (Recommendations for the authors):
Most of the issues I raised have been sufficiently addressed. Most prominently, the data and code are now available, and I was able to check that the authors' results are reproducible (Side note: I recommend further commenting the code so it is clear e.g. which models correspond to which figures, and I also recommend sharing the code in a different format than a Word document-these steps would help with accessibility).
I have a few remaining concerns:
Lines 75 – 78: "Accordingly, in humans, TTS is considered to be a stressful life event or even a disruptive crisis for the older sibling even under favorable conditions, a perspective that seems to be supported by TTS-related behaviors of the older offspring such as aggression, clinginess, and depressive syndromes (reviewed in Volling, 2012; Volling et al., 2017)."
As I wrote in my initial review, this claim is an oversimplification of these papers. Volling (2012), in particular, clearly concludes that there is substantial evidence for either of two quite different perspectives: that TTS is stressful, OR that it is an occasion for ecological adjustment that does not manifest in stress. This section of the intro should be edited to better reflect the ambiguity of the background literature, and the many different ways in which "stress" might be operationalized (see also my last comment below).
Lines 360 – 361: "The sudden recovery of cortisol and neopterin levels to pre-sibling birth levels after seven and five months, respectively, is puzzling and requires explanation."
I appreciate the authors expressing some caution here, but I would go further, which raises a somewhat larger point. The authors' statement I quoted is only warranted if the "two-cut" discontinuous model is taken at face value, and I am skeptical that should be done. The scatter plot of cort values, ignoring any splines, shows an apparent increase in the period right after sibling birth. That's interesting and worth trying to understand. But there are plenty of cort values just as high right before and right after the post hoc seven-month window. The "sudden-ness" of the increase and decrease is a function of specifying different intercepts at two different points. I wrote in my last review that I favored a single continuous spline for figures. The authors have basically responded by saying that "a one-cut discontinuous spline is a better fit than continuous, and a two-cut model is better still." But we have to consider biological plausibility, not just fit statistics. Do we really think it's realistic to expect such a uniform and sudden change seven months in? If we consider plausibility, I think one can make a reasonable case for a continuous spline, or one with a discontinuity at sibling birth (which is a prominent and biologically meaningful event), but not so much for the "two-cut" model. I recommend the authors foreground more biologically plausible models, and if they really still wish to include the "two-cut" model, that it occupies a less prominent position.
Lines 488 – 490: "Yet, the results obtained from wild bonobos renders support to the long-standing but untested and recently questioned assumption that the birth of a sibling is a stressful event for the older offspring (Volling, 2012; Volling et al., 2017)".
Once again, how are we to conclude that a cortisol increase equates to an individual experiencing a "stressful event"? The authors have become more careful in their language in many parts of the manuscript, but not here. The authors might say that they are simply using "stressful" to mean "a deviation from homeostasis", but my point all along has been that the authors need to be very, very clear about operationalizations if they want to use the term "stressful" in a different manner than its widely understood everyday definition. I recommend being super explicit about the usage and definition of the term "stress" in the intro, as I mention above. I mean literally provide a clear definition, upfront, so all readers are on the same page. The brief link between cortisol and homeostatic functioning, which isn't given until p. 5, is too indirect and vague. I also recommend removing statements like the one I quoted above. The authors already state in the discussion what we can actually determine here: cortisol goes up after the birth of a sibling, and it doesn't seem to be related to their age when it happens. Whether this event is "stressful" or not (which the authors claim without contextualization) cannot be determined by this dataset.
Reviewer #3 (Recommendations for the authors):
I am satisfied with the changes made by the authors based on my first review.
General comment: I would suggest a thorough proof-read of the text, just to make sure there aren't any little vocab or grammar mistakes and to polish it up. I caught many of the mistakes and pointed them out in my review, but I'm sure I missed some.
Title: would be better to say "causes a substantial and long-lasting increase in…"?
Abstract:
Second sentence could be worded better, as it is hard to understand. Perhaps "In these species, the birth of a sibling marks a major transition in early life, as maternal investment is constrained, and older siblings experience a decrease in maternal support."
Following sentence could be worded better:
"In the older offspring independent of its age, with siblings' birth, urinary cortisol levels increased fivefold and remained elevated for seven months."
Maybe better would be: "Following the births of siblings, urinary cortisol levels of older offspring, independent of their age, increased fivefold and remained elevated for seven months."
Change "did not show corresponding change" to "did not change". Otherwise, it is confusing for a few reasons: first, should be changes, second, make one wonder what corresponding means. I suggest you just simplify and cut out the unnecessary "show corresponding" part.
I am not a fan of the word syndrome. Sounds like a disease or physical problem. Could you say:
"Our results suggest that bonobos and humans experience TTS in similar ways and that this…".
Also, in this sentence, I do not like the idea of TTS as a developmental stage. I mentioned this in my last review and the authors made the change, but perhaps the idea got lost in the revising and then it was added back in here. TTS is more of a life stage or a life history stage… or a life transition. Would you say that offspring whose mothers never make a sibling are underdeveloped because they were not able to undergo the developmental stage of TTS? No because TTS is not a developmental stage, like weaning is, for example. So, I would reframe the idea of TTS that does not imply that it is a stage of development.
Line 50-51: but maternal provisioning through food sharing and feeding of young non-milk foods occurs after weaning. Revise this sentence to make clearer you are talking about milk and/or nursing.
Line 64-66: This sentence should be simplified and cleaned up. It is unnecessarily complicated and wordy, right now. Also, what are you trying to say exactly? That there are negative effects associated with having to share maternal care? Be more specific.
Line 69: Maybe change mother-offspring dependency to something less extreme, since dependency almost sounds like the offspring will be completely reliant on the mother for its whole life.
Line 91: change to "to be a social comfort behavior"
Also, the Badescu reference should be 2017, as it came out in early view in 2016 but then received an official issue in 2017. It is always very confusing.
Line 107-108: the last part of this sentence may not be grammatically correct.
Line 137: Not sure what constitution means. Can you replace with a more specific term?
Line 144-145: Hmm. I guess I understand what you mean but is this something measurable, that you determine from certain variables? It seems a bit speculative to say assume that the infant would die or would survive. Perhaps you can qualify these statements by offering as examples the variables you used to determine this. For example, you likely mean nursing behavior. So, the highly dependent were still regularly nursing, for prolonged periods, whereas the independent hadn't been seen nursing X focal hours of observation. Or maybe you mean the time that infants spent in body contact. Whatever you used to identify independence, give us that information.
Line: Visual inspection of?
Line 194: like here for example, should be "were" not "was". A small error but there are several like this throughout the text so go through carefully to polish up the writing.
Line 334: Do you mean "or showed a sudden change but in the opposite direction…"?
Line 340-341: levels returned to the levels before within one day. This phrase could be worded better or constructed a bit better to make easier to understand
Line 361: Wait, why is this puzzling. You just told us in the previous sentence that it takes human children 8 months to recover after sibling birth. Then the recovery you found after seven and five months is right in line with what you expect. Why puzzling?
Line 378: Unless I missed this somewhere, it would be good to add a note about whether some of the siblings continued to make nipple contacts after the birth of a new baby. And maybe tell us how many of the total number of infants were observed to continue nursing after birth of baby. This is interesting information for others. Maybe in results would be good to add?
Line 382: Say what kind of samples. Fecal samples? Hair samples? Urine samples? Bone samples?
Line 403: Affiliative intends? Not sure what you mean here.
Line 449: In baboons'? Maybe in baboons,…? Something is off here. Also, baboons repeats a few times. Some errors need fixing
Line 486: I don't like the word syndrome in this context. Makes it sound like TTS is a disease.
In methods, might be good to mention which authors collected which data, which authors did which analyses, etc. For example, the behavioral data were collected by focal animal sampling. Was this done by all the authors together, or just one? Give the initials of the author(s).
https://doi.org/10.7554/eLife.77227.sa1Author response
Essential revisions:
1) The link to the data and code needs to be fixed so that the reviewers can evaluate the data for themselves.
We apologize, we have now fixed this issue. Data as well as code are available.
2) There are two potential options for addressing the interpretation issues raised by reviewer #2. The authors may either supplement their analyses with additional behavioral data that speak more directly to stress (the preferred approach) or adjust the framing and interpretation to better suit the ambiguities of the data. If the authors choose to exercise this option, I would suggest steering away from the developmental psychology perspective the paper currently relies on, and more towards physiological mediators of life history transitions. The comments of reviewers #1 and #3 can both help to guide this since they offer related advice on framing.
We do not have behaviour data to clearly show that the increase in cortisol levels was a stressor. Throughout the manuscript we describe cortisol increase as a response to homeostatic threat. We also followed the suggestion of reviewer 1 and 3 not to label it stress response and we discuss more options, also positive homoeostatic threats.
3) The authors need to directly address the somewhat puzzling discontinuity in cort concentrations that occurs around 7 months post-sibling birth; what is the biological plausibility of this, and what factors might explain it? How does the non-recovery of neopterin fit with the return of cort to pre-birth baselines?
We have now included a whole paragraph in the discussion (line 351-363).
4) Please pay careful attention to the specific suggestions offered by all three reviewers. The paper would benefit greatly from additional details about the analyses and the motivations for specific components of them, to better suit a general-interest audience.
We are grateful for all the comments and help the reviewer provided. See the specifications below.
Reviewer #1 (Recommendations for the authors):
Introduction
Lines 51-52: What does mere behavioral adjustments mean? This requires some further explanation. I assume that it is referring to e.g. the withdrawal of maternal support, etc., but it could be interpreted in different ways.
We have now explained in detail- also following reviewer 3: “However, whether behavioral changes during TTS are associated with sibling birth rather than age-related behavioral adjustments related to withdrawal of maternal support remains to be resolved” (lines 79-81)
Lines 65-67: Are there really no data available on this for non-human primates? I find it hard to believe that no one has quantified the changes in mother-older offspring relationships after the birth of a new infant. But I could certainly be wrong!
We have now re-worded the introduction to show that this is not a human only topic and following also the suggestion of the editor more in the direction of a life history event (lines 50-79).
Line 86: It sounds like Schino and Troisi fulfil what I was asking for above, re: a non-human primate reference-this would probably be a good reference to add above, depending on the specifics of the findings.
See answer above.
Line 121: What is meant by 'general changes' in cortisol levels?
We have now re-worded the sentence to better explain what was shown: Notably, monitoring changes of urinary cortisol levels revealed first evidence that older offspring may respond physiologically to the birth of a sibling (lines 129-130)
Line 125: It would be helpful to quantify what rather independent (versus highly dependent) means. Spending X% of time within X meters of mom? Is it a nursing measure? Some combination?
Following also reviewer 3, we have now included two definitions for the terms and that this was used for our study: “… and the state of the older siblings at the time when mothers gave birth to another infant ranged from highly dependent (i.e., infant would die without mother) to rather independent (i.e., offspring would likely survive without mother)”. (lines 140-1452)
Line 131: This is a place in which it feels like the cart is coming before the horse. It seems odd to frame this as disentangling something which we haven't established IS entangled. This would feel appropriate if, for example, this paper were building on human studies that showed a cort increase after the birth of a sibling but did not control for changes in time near mother.
We have now re-written the whole part. This specific part does not exist anymore.
Line 132: When I hear the term stress response I think of short-term, acute changes in cort. Is it more accurate to characterize this as changes in baseline cort, given the duration of something like a transition to siblinghood? To avoid this issue entirely, you could just say 'to assess physiological stress before and during TSS.'
Following the suggestion by reviewer 2 we avoided to label it stress response and named it cortisol increase.
Paragraph on lines 132-145: This paragraph is a complicated mix of background info, theoretical justification, and methods. It would be easier to read if these were separated out. W/r/t the methods here, the T3 and neopterin measures do not feel well-justified. Due to my specific background knowledge, I understand why these measures are included in the study, but for a general interest journal, this feels like it needs more explanation. It assumes a fair amount about what the reader already knows.
We have now separated literature background from methods. We also have now added more information to explain why a marker was measured. We further structured the paragraph by cortisol, neopterin, and total T3 (lines 147-169).
Lines 158-159: I'm not sure that I am a fan of the term 'age-related weaning.' Cortisol changes with age. Cort might also change due to weaning. While obviously the specific age-related changes should be trivial in the window in which weaning is occurring, this term feels like it conflates two separate processes that could each impact cort concentrations.
We have now changed the terminology: If TTS had effects beyond weaning effects, then we expected sudden changes at sibling birth also after controlling for age-related changes. (lines 187-189)
Results
I personally would like to see the behavioral results come before the physiological results, since potential behavioral changes are part of what the authors are proposing might drive any physiological changes.
We thank the reviewer for the suggestion. We prefer to start with cortisol followed by the other two markers, because cortisol was the focus of the study and the two others as well as the behaviors were analysed to provide information about the circumstances and physiological state of the offspring.
How is independent foraging defined? This should be clarified in the results themselves, since the methods come later.
We have now added a definition in the method section: For independent foraging of the offspring, we only considered scans where also the mother was foraging to cover typical foraging situations and reduce the influence of potential sampling bias, with foraging encompassing handling and ingesting food (lines 518-520).
We have now added in the results when the term was used for the first time: There was no effect of TTS on the proportion of time spent foraging on their own (during maternal feeding time, to ensure foraging opportunity), … (lines 283-284)
Line 251: I would say before the sibling was conceived, instead of the conception of the mother. This could be read as the mother herself being conceived.
We thank the reviewer for the wording suggestion. The results were re-written for independent foraging. (lines 283-291)
Generally: is there any possibility that there are 'batch effects' going on in the cort data? I.e., were all of the post-birth to 7 month samples part of the same run?
We have now added: In order to exclude a methodological effect concerning the order of the samples e.g., that all post sibling birth samples are run together, all samples were randomly assigned to the measurements. (lines 544-546)
Discussion
Paragraph on lines 342-357: Potentially bolstering this argument, Rosenbaum et al. 2020 (PNAS, 117(33), 20052-20062) found that having a close-in-age sibling did not predict higher adult fecal glucocorticoid levels in the Amboseli baboons. Since they feature so heavily in these citations this seems like a good way to connect the specifics of this study to the Amboseli findings.
We are thankful that the reviewer mentioning this interesting reference. We have now added: However, the impact of sibling birth is not necessarily that strong. For example, in baboons’ presence of a sibling did not affect the HPA-axis later in life in baboons, but other early life adversities had lasting consequences (Rosenbaum et al., 2020). (lines 439-442)
Line 368: I would avoid the use of the word 'modern' here. I think the point is that there is a lot of variability in levels of allomaternal care amongst living humans, and that this variation may play an important role in how stressful (or not) children find TTS to be.
We agree with the reviewer and have now deleted modern. (line 469)
Line 373: I cringe at the idea that humans are not natural…we may be an unusual animal species, but this isn't the same thing 'unnatural' (a vague and here-undefined term that means different things in different parts of the literature). It would be more accurate to say something about it contributing to the comparative literature by showing that this transition happens in a closely related species, if you want to turn the focus back to humans.
We have now reworded the sentence following the reviewer’s suggestion: The results of our study showed that bonobos, one of humans closest living relatives, had high cortisol levels during TTS. (lines 474-475)
Line 380: What exactly is meant by 'constellation and co-parenting?' The sibling isn't co-parenting.
We agree with the reviewer and deleted co-parenting, ad added “family”: It is therefore important to note that, behavioral responses to TTS in human children are highly variable and individual- and age-dependent, and range from aggression, emotional blackmailing and psychological disturbances, to positive attitudes towards the new family constellation (Volling, 2012; Volling et al., 2017). (lines 484487)
Methods
I'm still not sure how independent foraging differs from just foraging. Is there such a thing as non-independent foraging? If so, what does that look like?
We have now added a clear definition the method section: … and when it was foraging independently (i.e., searching for its own food instead of being food provisioned by the mother). For independent foraging of the offspring, we only considered scans where also the mother was foraging to cover typical foraging situations and reduce the influence of potential sampling bias, with foraging encompassing handling and ingesting food. (lines 516-520).
Independent foraging is a term commonly used in animal behaviour publications:
For example:
Brown GR, Almond REA, Bergen Y van. 2004. Begging, stealing, and offering: food transfer in nonhuman primates Advances in the Study of Behavior. Elsevier. pp. 265–295. doi:10.1016/S0065-3454(04)34007-6
Burns JM, Clark CA, Richmond JP. 2004. The impact of lactation strategy on physiological development of juvenile marine mammals: implications for the transition to independent foraging. International Congress Series 1275:341–350. doi:10.1016/j.ics.2004.09.032
Jeglinski J, Werner C, Robinson P, Costa D, Trillmich F. 2012. Age, body mass and environmental variation shape the foraging ontogeny of Galapagos sea lions. Mar Ecol Prog Ser 453:279– 296. doi:10.3354/meps09649
Soulsbury CD, Iossa G, Baker PJ, Harris S. 2008. Environmental variation at the onset of independent foraging affects full-grown body mass in the red fox. Proc R Soc B 275:2411–2418.
doi:10.1098/rspb.2008.0705
Line 483: I assume shanked is meant to be shookϑ.
Absolutely, we have now changed shanked to shook (line 594)
Figures and Tables
Figure 1 in general: I think there are typos in the legend for this figure. It never refers to the right-hand panels. In general, the figure captions need to be better explained. Figures should be stand-alone, and as presented these are not.
We have now added additional information and have re-written the figure descriptions. See also comments of reviewer 2.
Reviewer #2 (Recommendations for the authors):
Lines 135-136: This same group of researchers also found that cortisol is high leading up to the days before Christmas, when children were excitedly anticipating presents from Santa (Flinn et al., 2011; https://doi.org/10.1016/j.neubiorev.2011.01.005)-not exactly traumatic. Flinn et al. interpret these rises in both kinds of situations as "arousal to social opportunities", which might be a valuable perspective for the authors to consider in their own dataset.
We are thankful for the reviewers thought, and unfortunately, we gave the impression that the cortisol stress reaction is something negative “stress”. We added therefore the suggested reference in the introduction and a sentence to show that the cortisol release is a reaction of stressor threatening homeostasis. (lines 147-155).
Analyses: For the GAMs, it would be good to see some information on how robust the presented splines are to alternative numbers of basis functions and alternative smoothing parameters.
We have now added a paragraph to the methods section where we discuss this in detail:
"Throughout models, the number of basis functions (k) was always set equal for all predictor and random smooths of time around sibling birth and of age. The number of basis functions was generally set to 10, but needed to be reduced to 6 in some cases for the full models including both a term for age and for time around sibling birth due to sample size (for al physiological variables and for riding). Additionally, k needed to be reduced to 6 also for all models on body contact and 5m- proximity to the mother since higher values often led to strong overfitting and uncertainty. We further tested for robustness of the estimated smooths parameters by setting the number of basis functions to the respective maximum value (for models without continuous age terms), which was k = 12 for all physiological, k = 15 for riding, and k = 25 for all other response variables. Patterns of smooth trajectories remained the same (also for body contact in this case), though naturally, the parallel increase of k for both the predictor and the associated random smooth terms led to increasing identifiability constraints and thus increasing estimation uncertainty". (lines 663-674).
In addition, we also run a series of models for our main result on discontinuities at sibling birth in cortisol and neopterin. Here, we allowed the model with the continuous smooth only for high wiggliness and thus higher potential to sufficiently fit the sudden change at sibling birth, by increasing k of the predictor smooth term to 50 and reducing k of the respective random smooth term to 6, plus introducing varying low smoothing parameters. We then compared these models with our unmodified discontinuity models and reported the results in the Results section and in supplemental figure S3.
Relatedly, I'm a little sceptical of the approach of estimating separate splines before and after sibling birth, for all of the causal inference considerations that regression discontinuity entails. A useful companion to these 'discontinuous' splines, especially for Figure 1 and Figure S1, would be estimates of a single, continuous spline for time relative to sibling birth. It's hard to know from eyeballing it, but my guess is a jump in cortisol would still be apparent, for example, but it would be more gradual and less striking than the current figures suggest.
We thank the reviewer for highlighting this insufficient communication in the previous version of the manuscript. Indeed, what we primarily did was a model comparison between a model with a continuous smooth only and a model with the same continuous plus the categorical before/after variable allowing for additional intercept differences between the time before and after sibling birth. We have now made this point much more explicit in the methods and the Results section.
It might be important to emphasize here that we did not calculate separate smooths for before and after sibling birth, to avoid the associated pitfalls and also misleading statistical results, also since predictions would still be calculated ( = extremely extrapolated) for the entire time period. Therefore, we decided to use the cumulative nature and allow the model to estimate both a continuous smooth for the entire time period plus an intercept difference in values. This approach comes at the small inconvenience that the model allows only for discontinuity of the predicted values but not the first and second derivatives, i.e. there will be no discontinuity in the slope of the smooth at sibling birth. Although this leads to a slight constraint on optimal smooth estimation (most pronounced in Figure 1A), we think that this is the best possible approach.
Line 213: Is this Chi-squared value supposed to be negative?
No, thank you for highlighting this mistake. We checked and corrected now throughout the manuscript.
Discussion: My public review mentions that it is difficult to interpret the finding of increased cortisol following sibling birth. At some points, the authors appear to recognize this, as they raise a number of good arguments in the discussion about potential factors leading to a TTS-timed cortisol increase. Some, like a sibling birth coinciding with the beginning of juveniles experiencing greater male aggression, have nothing to do with TTS per se. Other arguments imply a role for beneficial behavioral interactions between the older and younger sibling.
We thank the reviewer for highlighting this. We have now added other factors that may influence cortisol changes in the older offspring. (lines 389-408)
Yet, the authors appear to dismiss those concerns and conclude that the cortisol increase indeed implies stress. First, I recommend at minimum addressing why numerous alternative interpretations-some that the authors identify, and some that I have identified-should be disregarded in favor of the "cortisol = stress" conclusion.
Throughout the manuscript we claim that cortisol is a stress response or indicating a homeostatic challenge. As alternative interpretations we tried to exclude that the found increase in cortisol is related to food stress challenges (urinary total T3). We further found that the proxy for the function of the immune system (urinary neopterin) was affected. With this two finding we came to conclusion that the homeostasis of the youngsters must be challenged by a non-food related stressor, but in a way that is affects the immune function.
We now added in the introduction that cortisol is released to a threat of homeostasis. We avoided to use the world “stress” without any further description. We also changed the title.
Second, as I state in the public review, having data on behavioral interactions thought to be stressful would go a long way towards solidifying the interpretative elements of this paper. I don't know whether this sort of information is available, but if it is, I recommend integrating it into a revision of this manuscript. One of the main lessons from the developmental psychology literature is that there is substantial heterogeneity in TTS adjustments, which may be attributable to family dynamics or larger ecological factors. Being able to test these kinds of theories in bonobos would significantly boost the impact of this paper.
Unfortunately, we do not have data describing behaviour in detail during TTS in wild bonobos. We have now added:
“To clarify that our study is describing a physiological stress response but only discuss the potential reasons for the response we add: Whether the increase in cortisol levels in the bonobos in our study had positive or negative long-term consequences was not assessed. Future studies should therefore integrate behavior and physiological measures to estimate the impact of TTS for the older sibling. Such a combination of measures might help to disentangle why young bonobos show such an intense cortisol response, for example as a response to the novel mother-infant constellation, as a measure of positive valence arousal, or with TTS as a normative maturing experience. (lines 452-458)”.
Line 304-305: The authors claim that "findings from human children show behavioral responses to sibling birth [are] independent of their actual age". True, some show that, but others do not; many claim younger children exhibit larger behavioral disruptions (e.g. Nadelman and Begun, 1982; Volling, 2012). Some expansion on why age might matter in some aspects of adjustment, but not others, would provide useful context to the discussion.
We have now in cooperated these thoughts.
“However, changes at sibling birth can be age dependent. In response to sibling birth, scores for e.g., clinging and other gestures of reassurance were negatively correlated with the age of the older sibling (Dunn et al., 1981; Nadelman and Begun, 1982; Volling, 2012). Thus, in children, age seems to affect the behavioral response towards, or the perception of, the arrival of a sibling.” (lines 381-385)
Figure 1: The right panels of Figure 1 (B, D, and F) will be very hard to interpret for the average reader: the figure captions are sparse, the contours have labels that are so small as to be invisible, no justification/explanation is given for the extrapolation parameter (and thus it won't be clear to the vast majority of people why the figure is splotchy), etc. I understand the objective of contour plots for visualizing non-linear interactions, but as they stand the figures and/or captions need to be changed to form a self-contained explanation, as figures should. I leave edits up to the authors' discretion: they could do some combination of beef up their figure captions; present a more traditional interaction plot of linear effects (i.e., present marginal trends of -1, 0, and +1 SE on offspring age), since all the splines in Figure 1 are very close to linear; or present a different kind of plot, like a perspective plot, or a plot of predicted splines using the get_predictions function in itsadug.
Figure 2: I think the caption for supplementary Figure 2 is incorrect-there is no depiction of time relative to sibling birth in this figure, just the age of the older sibling. It appears the caption was incorrectly copied over from part of Figure 2.
We have now – also following rev1 – revised all figure captions to improve the understandings and self-standing of the figures.
Data: The DOI linked for the source data doesn't work. I would need to re-evaluate the paper after having access to the raw data, so I can verify the analytical reproducibility of the key claims.
We are really sorry and apologize that the doi link was not functioning. We have now solved this issue.
Reviewer #3 (Recommendations for the authors):
Abstract:
I would not identify the birth of a sibling as a developmental transition. Perhaps would be better to say a major life transition or major transition in the early life of an individual. Or if you stick with the word development, perhaps it would help to elaborate a bit to make clear why it is a developmental transition. Maybe it applied to nutritional development so that the mother-infant nutritional relationship must end with the birth of a new sibling? Perhaps you are referring to social development? Elaborating a bit might make clearer what you mean.
Should be: "Studying the transition", "in the mother-infant relationship".
We thank the reviewer for the suggestion. We have now changed the sentence to:
“In animals with slow ontogeny and long-term maternal effort, immatures are likely to experience sibling birth before reaching maturation. The birth of a sibling marks a major transition in early life, maternal investment is constrained, and it deprives the older offspring from maternal support.” (lines 30-33)
Change to "evolutionarily old"- although evolutionarily old is a bit vague. Perhaps you can narrow it down to a specific time based on comparisons with other primates or mammals. For instance, you might say that this effect was likely present in the common ancestor of all the great apes. Something more specific to your study.
We have now speculated: Our results suggest that bonobos and humans share the syndrome of TTS and that this developmental stage may have emerged in the last common ancestor. (lines 42-44)
Line 43: change "while still being dependent" to "while they are still dependent".
We thank the reviewer for the suggestion. This part does not exist anymore.
This sentence also needs a reference- also do you mean children in humans only? I guess that is what you mean but perhaps you should extend this to all primates or long-lived, slow developing mammals. You could say that in mammals with slow rates of growth and development that give birth to single offspring at a time, most offspring are exposed to the birth of a younger sibling.
Extending this to other species could make it more relevant to more researchers.
We thank the reviewer for that great idea and started the introduction with:
“In animals with slow ontogeny and long-term maternal effort, immatures are likely to experience sibling birth before reaching maturation. The birth of a sibling marks a major transition in early life, maternal investment is constrained, and it deprives the older offspring from maternal support. Transition to siblinghood (TTS) is often considered to be stressful for the older offspring, but physiological evidence for this is lacking.” (lines 30-34)
Line 45: competitor in what sense? Competitor for maternal attention and resources, or for food in the environment, or for reproduction or for social partners etc. I think this needs to be more specific.
We have now added that it is about resources in general. (line 72)
Line 49: Do you mean behaviors shown by the older sibling? This is not clear. Make clearer that it is the older sibling who could show increased aggression, clinginess, and depression. Otherwise, could be perceived to be in the mother or in the new sibling.
We now wrote: “Accordingly, in humans, TTS is considered to be a stressful life event or even a disruptive crisis for the older sibling even under favorable conditions, a perspective that seems to be supported by TTS-related behaviors of the older offspring such as aggression, clinginess, and depressive syndromes (reviewed in Volling, 2012; Volling et al., 2017).” (lines 74-77)
Line 51: not sure what you mean by mere behavioral adjustments. I think the point you are making here is important, but couldn't behavioral adjustments be in response to stress. I guess you are trying to say that these are behavioral changes that could occur in response to new siblings that are not associated to stress for the older sibling. Make clearer.
We have now – also following the suggestion of reviewer 1 changed it to: “However, whether behavioral changes during TTS are associated with sibling birth rather than age-related behavioral adjustments related to withdrawal of maternal support remains to be resolved (Volling, 2012; Volling et al., 2017).” (lines 79-81).
Line 94-97: I do think this statement needs references to support it.
We have rewritten this part.
Line 98: I think this statement also needs some references and a range in inter-birth interval lengths for bonobos, including a variety of sites, since you have the ones for your site specifically, below.
We have now included the range of inter-birth intervals of wild bonobos, also including other sites.
We leveraged the large variation in inter-birth intervals in bonobos ranging from three to nine years (Knott, 2001; Tokuyama et al., 2021) to differentiate between the effects of TTS on one hand, and nutritional and social weaning on the other. (lines 136-138)
Line 101: should be social dependency on the mother.
This was also part of the re-written part.
Line 106: would co-dependency on the mother be better than co-residence? Co-residence sounds like one offspring needs to leave the natal group at some point, which is not always the case. Co-residence with siblings can occur for their entire lives if they are the philopatric sex.
We have now changed it to: “Maternal support is intense and persists for a long time (Stanton et al., 2020; van Noordwijk et al., 2018), and extended periods of parental care of two dependent offspring of different ages is common (Achenbach and Snowdon, 1998).” (line 100-102)
Line 110-111: Sexual maturation at 4 years of age is very young… Many individuals might still be nursing at 4 years old. Perhaps your definition of the onset of sexual maturation needs to be explained. Also, unclear how the onset of sexual maturation is different from the onset of menarche, and how these two milestones are different, so this should be cleared up as well.
This part is now re-written.
Introduction in general:
You've made many comparisons between bonobos and humans, which is good, but I wonder if it would be good to add in data and comparisons with other apes as well (chimps, gorillas, orangutans). I know you mention chimpanzees once or twice, but I wonder if it would help to make more explicit comparisons throughout with the other great apes. I say this because eventually in your discussion, you are going to be making some preliminary conclusions about when various developmental and stress mechanisms would have evolved and saying that the timing would have been in the last common ancestor between bonobos and humans may be incomplete. Inherently we would want to hear about the chimpanzee data at least, to know if we should think about the last common ancestor with Pan, but of course we would wonder about any evidence in the other great apes. If the other great apes show similar patterns in life history, development and maternal investment that you are pointing out for humans and bonobos, then the findings of your research here could also apply to them (pending studies in these other apes). Alternatively, there might be some key differences in the life history patterns and infant development of different apes, so it would be good to know about them to later understand why you may not think that the findings extend to a certain genus.
We have now added information about sibling interaction or the effect of sibling birth in other apes (lines 102-119)
I suggest you try to keep the tense for your article in the past tense as much as possible. As it is now, you switch quite a bit between present and past tense, and at times, you use present tense but this is a bit awkward. Past tense would be best.
We now carefully checked to have the manuscript in past tense. We apologize if there are still mistakes.
Line 123: You should mention the name and place of your study population here since you are bringing it up to set up your study.
We have now added: “Our study investigated TTS-related changes in physiological responses in juvenile individuals of two habituated groups of wild bonobos (Pan paniscus) at the site of LuiKotale in the Democratic Republic of Congo.” (lines 131-133)
Line 125: this statement on the dependency levels of the offspring needs to be supported with a reference and it needs to be quantified in some way. Is dependency based on nipple contacts, proximity, sleeping in the same nest, being carried from place to place, all the above?
Also, unclear if this statement refers to past research or if this is something that is being done in this paper. Did someone already investigate the nutritional development of these individuals, or is the level of dependency something you will be measuring in the present study and measured relative to TTS? This needs to be made clearer.
Also following reviewer 1 we have now included definitions and this was the case in our study.
“… and the state of the older siblings at the time when mothers gave birth to another infant ranged from highly dependent (i.e. infant would die without mother) to rather independent (i.e. offspring would likely survive without mother).” (lines 140-142)
Lines 139-140: I think this sentence needs some references. Also, you explain T3 in greater detail but not the neopterin hormone. I think a sentence specifically on neopterin and how it works or what it means can help here.
Following also suggestions of reviewer1 we have restructured the whole paragraph and have added more information on neopterin and references (lines 147-169).
Lines 140-141: I am not quite understanding T3 can help disentangle effect of energetic stress versus social stress. Can you please make this more obvious/clearer?
Following also suggestions of reviewer1 we have restructured the whole paragraph and have added more information about T3 and energetic deficiency (lines 164-169)
Line 143: unclear why neopterin levels should decrease because we do not have enough information about neopterin.
We apologise for being less clear. We hope it is now better explained better with the changes we added (lines 156-163)
Line 146: you say nursing here but later you define suckling as contact with the nipple. Might be better to stick with one of the two terms, especially since people are going to keep an eye out for the term nursing to see how this was defined later in the text.
We have now changed nursing to suckling (line 170).
Line 148-150: Are you saying that social weaning is weaning from the mother more generally, in terms of proximity and access to carrying, etc.? This is very confusing because weaning in most infant development/maternal investment literature refers specifically to nursing or feeding behavior. Until this point in the article, I was thinking you were trying to make a distinction between nutritive and non-nutritive nursing.
If you would like to talk about changes in the mother-infant relationship outside of nursing and foraging behavior, I strongly encourage you NOT to use the term weaning for that. Perhaps switch to "attainment of physical independence" or some other similar phrase since the only other measures outside of nursing behavior that you are looking at are more physical measures (so the infant being carried versus moving on its own/ infant in certain proximity to the mother). Instead of nutritional and social weaning, you could say weaning and attainment of physical independence.
We are grateful for this suitable wording. We have now changed the terms to weaning and attainment of physical independence. Many, many thanks for the wording.
Line 193: With increasing age, urinary…
Also, you say that they significantly changed in males but not females, but then you say that this change was not significant… I am confused.
We have now re-worded the result section to be more clear and precise following also suggestions by reviewer2.
Line 210-211: same issue here. you say significantly declined but then in brackets you said sex difference not significant. Which is it?
See above, now it is re-written, and not contradictive anymore.
Line 75, 262 and elsewhere: You keep talking about nutritional versus social weaning (or as others have said, nutritional versus behavioral weaning), but this concept is fairly recent. The idea that there are two components to weaning, the milk-transfer and the psycho-social relationship between the mother and the infant through continued nipple contacts, regardless of whether or not milk transfer occurs, is a fairly new and innovative idea that needs to be supported with literature in your paper, and that needs to be explained a little bit somewhere in the text. For example, research on wild chimps and other primates has shown that comfort nursing, without milk transfer, can occur for years after lactation has ended. This sets up a situation where you have these two separate weaning periods: weaning from milk and weaning from nipple contact- so the nutritional versus social/behavioral weaning. Thus, the mother-infant behavioral relationship can develop separately from the mother-infant nutritional relationship, despite considerable overlap between the two. This distinction, and its importance for the infant and mother, should be explained.
We thank the reviewer for these thoughts. We have now added:
“The term weaning is often used for the attainment of nutritional independence, but also comprises the process of social independence and behavioral maturation, which can occur at different ages. Nutritional weaning refers to the termination of the consumption of maternal milk, but nipple contact of the offspring without milk transfer may exceed lactation and is assumed to be social comfort behavior (Bădescu et al., 2016; Berghänel et al., 2016; Matsumoto, 2017).” (lines 85-90)
Line 276-279: Are you saying that TTS is severe, uncontrollable for older offspring, and was unpredictable or moderately unpredictable? Make clearer how this statement relates to your results.
Yes, this was indeed what we think that could be the scenario. We reworded the sentence to:
“The intensity of a stress response is generally determined by the severity, controllability, and predictability of the stressor (Seiler et al., 2020), all of which probably apply to TTS as a stressor, being novel, sever, uncontrollable and relatively unpredictable for the older offspring, and therefore contribute to the comparably high cortisol response that we observed in our study.” (lines 337-341)
Line 347-349: change sentence to
"However, strong negative effects caused by a highly predictable and normative stressor that invariably affects most individuals, such as the birth of a sibling in apes, should be under negative selection, and would thus be expected to XX XX (explain what this would mean)."
We thank the reviewer for the wording, now we added:
“Moreover, strong negative effects caused by a highly predictable and normative stressor that invariably affects most individuals, such as the birth of a sibling, should be under negative selection, and would therefore be considered to be a non-adaptive trait.” (lines 442-444)
Line 298: I would change this to
"…suggest that nutritional weaning is usually completed by around 4.5 years of age".
We have now changed the sentence to: “…, and preliminary analyses of stable isotopes in samples collected from the same population suggest that nutritional weaning is completed by around 4.5 years of age (Oelze et al., 2020).” (lines 372-374)
Lines 372 to 372, or more broadly for this whole paragraph: Similar to my comment for the abstract, the statement that your results highlight the evolutionary history of stress response and TTS is vague and almost seems like a throw away idea because it is not specific enough. Can you go a step further and talk about the possible timing of an evolutionary link between stress and TTS? Presumably, this important interaction would have appeared with the great ape transition? Or would it be with the transition between genus Pan and Homo? Use the comparisons in life history traits and infant development of humans and the other great apes to infer when this mechanism that you found could have become more important or more prominent.
We have now added a speculation about when TTS might have occurred (last paragraph of the discussion) (lines 474-491)
Methods:
Line 397: I would like to see a table with a breakdown, by infant age and sex, of the sample sizes and numbers for the behavioral and urine data.
We agree that a table is helpful, bit while the data are not presented by individuals age, we thought it might be helpful to present them in relation to sibling birth. We now added a table in the supplement presenting the data in relation to sibling birth. If the age of the bonobo is still important, we can change the table or the reader might use the raw data available. (line 504, supplement table 1)
Lines 399 to 411: Did you exclude from the calculations time out of view, or time when the infant was ventral, but it was unclear if they were in contact with the nipple, and excluded time in a nest with the mother since you can't see the infant? If yes, say so, if not, justify why you did not.
Yes, we excluded all data points where the infant was not sufficiently visible, and also state this now in the methods section.
I am not familiar with any of the urine analyses so cannot comment on these.
Please explain how the proportions of the behavioral measures were calculated somewhere in the methods.
Has been added to the methods section (line 515ff, especially line 526f).
[Editors' note: further revisions were suggested prior to acceptance, as described below.]
The manuscript has been greatly improved, but there are some remaining issues that need to be addressed before it can be accepted for publication.
– First, and most importantly, I (the reviewing editor) strongly urge you to consider the comments of reviewer #2 w/r/t the plausibility of the two-intercept model. While I understand that this model provides the best fit statistic, in my opinion, fit statistics should not override biological plausibility. There is no biologically plausible explanation for why cort would suddenly drop at 7 months (or at least, none that have been relayed in this text), and basic data visualization -- i.e., the raw scatterplot of the data--does not suggest this is what is happening. I personally feel that the single-cut discontinuous spline is a good middle-ground choice (due to the suddenness of sibling birth), followed by a continuous spline, followed by the two-cut discontinuous. The last two options would certainly be worth including in the supplementary materials.
We have now focused our results on the cortisol and neopterin model with one cut at sibling birth in the results as well as in the discussion. We have now the three physiological markers with one cut as the main figure 1. As suggested, we now added the models and the figures for cortisol and neopterin with the second, unexpected sudden change, as supplement material. We further have now reworded also the part in the result section to already highlight that the results are unexpected and that we do not have an obvious explanation for.
This issue is the primary reason that I am recommending a revise and resubmit, rather than an acceptance. Please note that my acceptance of the manuscript is not contingent upon making this change. I respect that authors may have different points of view surrounding the interpretation of fit statistics. However, if it is not changed, the final evaluation summary will reflect the fact that myself and one of the other reviewers disagreed with the analysis strategy. I want to be transparent about the source of the disagreement so that the evaluation summary would not come as a surprise should you choose not to change which model you prioritize in the main text.
We thank the editor for her honest words. We could understand the argumentation of reviewer2 and the editor, and therefore, made the adjustments as described above.
– Please include a scatterplot of the raw data in the supplementary materials.
We have now included a scatter plot of the cortisol data in the supplementary materials, Supplementary Figure 3.
– Reviewer #2 is correct that though the language around 'stress' has been considerably improved, there are still places where there are ambiguities in how it is used/what it is implying. The manuscript will be stronger with these ambiguities removed. These data are quite interesting and do not need to lean on 'stress' in order to be noteworthy and important. Similarly, I agree with reviewer #3s concerns about characterizing TTS as a developmental stage. Both of these are wording issues that should be simple to resolve.
We have now carefully re-worded the MS. Describing sibling birth as an event and cortisol changes as cortisol changes.
– Please follow reviewer #3s request for a thorough proofread. There are a considerable number of spelling and grammar mistakes that remain, which is distracting as a reader. They have many helpful suggestions for places where the language could use additional clarification, which again, will help strengthen the final product.
Alison Ashbury (see Acknowledgement) edited the manuscript.
Reviewer #2 (Recommendations for the authors):
Most of the issues I raised have been sufficiently addressed. Most prominently, the data and code are now available, and I was able to check that the authors' results are reproducible (Side note: I recommend further commenting the code so it is clear e.g. which models correspond to which figures, and I also recommend sharing the code in a different format than a Word document-these steps would help with accessibility).
We thank the reviewer for the suggestions. We have now uploaded the code as txt files. We will write additional comments on the final code, if we all agree for example on figure order and files.
I have a few remaining concerns:
Lines 75 – 78: "Accordingly, in humans, TTS is considered to be a stressful life event or even a disruptive crisis for the older sibling even under favorable conditions, a perspective that seems to be supported by TTS-related behaviors of the older offspring such as aggression, clinginess, and depressive syndromes (reviewed in Volling, 2012; Volling et al., 2017)."
As I wrote in my initial review, this claim is an oversimplification of these papers. Volling (2012), in particular, clearly concludes that there is substantial evidence for either of two quite different perspectives: that TTS is stressful, OR that it is an occasion for ecological adjustment that does not manifest in stress. This section of the intro should be edited to better reflect the ambiguity of the background literature, and the many different ways in which "stress" might be operationalized (see also my last comment below).
We absolutely agree, we have now added in the introduction:
“However, sibling birth also presents opportunities for the older offspring, such as social and emotional growth through interacting with the newborn. Individuals vary in how they adjust to the birth of younger sibling; some children have difficulties while others cope well (reviewed in Volling, 2012; Volling et al., 2017). In any case, the birth of a sibling is linked to a time of change the older child must cope with.” (lines 75-79)
Lines 360 – 361: "The sudden recovery of cortisol and neopterin levels to pre-sibling birth levels after seven and five months, respectively, is puzzling and requires explanation."
I appreciate the authors expressing some caution here, but I would go further, which raises a somewhat larger point. The authors' statement I quoted is only warranted if the "two-cut" discontinuous model is taken at face value, and I am skeptical that should be done. The scatter plot of cort values, ignoring any splines, shows an apparent increase in the period right after sibling birth. That's interesting and worth trying to understand. But there are plenty of cort values just as high right before and right after the post hoc seven-month window. The "sudden-ness" of the increase and decrease is a function of specifying different intercepts at two different points. I wrote in my last review that I favored a single continuous spline for figures. The authors have basically responded by saying that "a one-cut discontinuous spline is a better fit than continuous, and a two-cut model is better still." But we have to consider biological plausibility, not just fit statistics. Do we really think it's realistic to expect such a uniform and sudden change seven months in? If we consider plausibility, I think one can make a reasonable case for a continuous spline, or one with a discontinuity at sibling birth (which is a prominent and biologically meaningful event), but not so much for the "two-cut" model. I recommend the authors foreground more biologically plausible models, and if they really still wish to include the "two-cut" model, that it occupies a less prominent position.
We agree with the view that the second cut is unexpected and without an obvious biological expectation. We have now focused our results on the cortisol and neopterin model with one cut at sibling birth in the result section, because we agree that we here had an event.
Figure 1 which was before showing the two-cut model is now the three physiological markers with one cut. As suggested by the editor, we added the models and the figures for cortisol and neopterin with the second unexpected sudden changes as supplement material. We further re-worded also the part in the result section to already clarify that the results are unexpected. We further added the cortisol scatter plot for the reader as supplementary material as suggested by the editor.
“While the model with the two discontinuities describes our data better mathematically, there is no obvious biological explanation for the second change (i.e., the sudden decline in cortisol) after seven months. However, in the model with only one discontinuity at sibling birth and a smooth continuous decline thereafter (Figure 1A-C), the cortisol levels took over 7 months to return to previous levels.
Hence, the absence of low cortisol levels after sibling birth was evident in both models.” (lines 217222)
Lines 488 – 490: "Yet, the results obtained from wild bonobos renders support to the long-standing but untested and recently questioned assumption that the birth of a sibling is a stressful event for the older offspring (Volling, 2012; Volling et al., 2017)".
Once again, how are we to conclude that a cortisol increase equates to an individual experiencing a "stressful event"? The authors have become more careful in their language in many parts of the manuscript, but not here. The authors might say that they are simply using "stressful" to mean "a deviation from homeostasis", but my point all along has been that the authors need to be very, very clear about operationalizations if they want to use the term "stressful" in a different manner than its widely understood everyday definition. I recommend being super explicit about the usage and definition of the term "stress" in the intro, as I mention above. I mean literally provide a clear definition, upfront, so all readers are on the same page. The brief link between cortisol and homeostatic functioning, which isn't given until p. 5, is too indirect and vague. I also recommend removing statements like the one I quoted above. The authors already state in the discussion what we can actually determine here: cortisol goes up after the birth of a sibling, and it doesn't seem to be related to their age when it happens. Whether this event is "stressful" or not (which the authors claim without contextualization) cannot be determined by this dataset.
We have now added in the introduction that we expect an increase in cortisol as a response to the challenging situation of sibling birth. We also re-formulated many sentences in the discussion. E.g., lines 78-79, lines 151-153. Line 362, lines 347-341, line 352 and so on.
Reviewer #3 (Recommendations for the authors):
I am satisfied with the changes made by the authors based on my first review.
General comment: I would suggest a thorough proof-read of the text, just to make sure there aren't any little vocab or grammar mistakes and to polish it up. I caught many of the mistakes and pointed them out in my review, but I'm sure I missed some.
We thank the reviewer for all her helpful comments and corrections so far. We had sent out our manuscript for proof reading, and we think it sounds now way better. We hope that the reviewer agrees.
Title: would be better to say "causes a substantial and long-lasting increase in…"?
We have now changed the title to: Transition to siblinghood causes a substantial and long-lasting increase in urinary cortisol levels in wild bonobos
Abstract:
Second sentence could be worded better, as it is hard to understand. Perhaps "In these species, the birth of a sibling marks a major transition in early life, as maternal investment is constrained, and older siblings experience a decrease in maternal support."
We have now changed it to: In these species, the birth of a sibling marks a major event in an offspring’s early life, as the older siblings experience a decrease in maternal support. Lines 31-33
Following sentence could be worded better:
"In the older offspring independent of its age, with siblings' birth, urinary cortisol levels increased fivefold and remained elevated for seven months."
Maybe better would be: "Following the births of siblings, urinary cortisol levels of older offspring, independent of their age, increased fivefold and remained elevated for seven months."
Change "did not show corresponding change" to "did not change". Otherwise, it is confusing for a few reasons: first, should be changes, second, make one wonder what corresponding means. I suggest you just simplify and cut out the unnecessary "show corresponding" part.
We have now changed it to: The cortisol level increase was associated with declining neopterin levels, however T3 levels and behavioral measures did not change (lines 39-40).
I am not a fan of the word syndrome. Sounds like a disease or physical problem. Could you say:
"Our results suggest that bonobos and humans experience TTS in similar ways and that this…".
Also, in this sentence, I do not like the idea of TTS as a developmental stage. I mentioned this in my last review and the authors made the change, but perhaps the idea got lost in the revising and then it was added back in here. TTS is more of a life stage or a life history stage… or a life transition. Would you say that offspring whose mothers never make a sibling are underdeveloped because they were not able to undergo the developmental stage of TTS? No because TTS is not a developmental stage, like weaning is, for example. So, I would reframe the idea of TTS that does not imply that it is a stage of development.
We absolutely agree, we have now changed it to: Our results suggest that bonobos and humans experience TTS in similar ways and that this developmental event may have emerged in the last common ancestor (lines 442-43).
Line 50-51: but maternal provisioning through food sharing and feeding of young non-milk foods occurs after weaning. Revise this sentence to make clearer you are talking about milk and/or nursing.
We have now revised the sentence to: In mammals, weaning refers to the transition from nutritional dependency to a stage when immatures are independent of maternal food provisioning (lines 49-50)
Line 64-66: This sentence should be simplified and cleaned up. It is unnecessarily complicated and wordy, right now. Also, what are you trying to say exactly? That there are negative effects associated with having to share maternal care? Be more specific.
We now changed it to: However, the older offspring must share maternal care, which may influence its social behavior as well as its physiological constitution (lines 62-63).
Line 69: Maybe change mother-offspring dependency to something less extreme, since dependency almost sounds like the offspring will be completely reliant on the mother for its whole life.
We have now changed it to something less extreme: Immatures grow slowly, social maturation extends well into adulthood, and to a certain degree, beneficial mother-offspring relationships can last a lifetime (lines 65-67).
Line 91: change to "to be a social comfort behavior"
Also, the Badescu reference should be 2017, as it came out in early view in 2016 but then received an official issue in 2017. It is always very confusing.
We are sorry for the wrong citation. We now changed it to Bădescu et al. 2017 in lines 89-90 as well as in the references (lines 720-722).
Line 107-108: the last part of this sentence may not be grammatically correct.
We have now changed it to: While data on mother-offspring relationships are abundant, little is known about interactions between immatures and infants born to the same female (lines 101.103).
Line 137: Not sure what constitution means. Can you replace with a more specific term?
This part was changed during editing to: We used multiple physiological and behavioral measures to investigate the responses of older siblings to the birth of their younger sibling. We sought to disentangle the effects of changes in mother-offspring relationships and energetics that are associated with nutritional and social weaning, from the specific effects of a younger sibling’s birth (lines 129-133).
Line 144-145: Hmm. I guess I understand what you mean but is this something measurable, that you determine from certain variables? It seems a bit speculative to say assume that the infant would die or would survive. Perhaps you can qualify these statements by offering as examples the variables you used to determine this. For example, you likely mean nursing behavior. So, the highly dependent were still regularly nursing, for prolonged periods, whereas the independent hadn't been seen nursing X focal hours of observation. Or maybe you mean the time that infants spent in body contact. Whatever you used to identify independence, give us that information.
We have now added this information: … and thus the developmental status of older siblings at the time when their mothers gave birth to another infant ranged from highly dependent in terms of travel support and foraging skills (i.e. time carried and nursed) to mostly independent (lines 136-139).
Line: Visual inspection of?
We added now: Visual inspection of the data showed that the sudden decline in riding at sibling birth was only evident in older siblings belonging to the younger age cohort… (lines 285-287).
Line 194: like here for example, should be "were" not "was". A small error but there are several like this throughout the text so go through carefully to polish up the writing.
We thank the reviewer for detecting all the little mistakes. We hope that the others we all changed by the editing service.
Line 334: Do you mean "or showed a sudden change but in the opposite direction…"?
The sentence was completely revised to: At sibling birth, weaning-related behavioral changes were either already completed (independent foraging and suckling), did not change discontinuously (urinary total T3, suckling, time in spatial proximity to mother, and independent foraging), changed suddenly in directions opposite of our expectation (increasing body contact time with the mother), or were significant only in subjects belonging to the younger age cohort (riding) (lines 332-337).
Line 340-341: levels returned to the levels before within one day. This phrase could be worded better or constructed a bit better to make easier to understand
We have now changed it to: A similar cortisol response occurred in bonobos in response to a group member giving birth, but in this case, the individual’s cortisol levels returned to previous values within one day (lines 340-342).
Line 361: Wait, why is this puzzling. You just told us in the previous sentence that it takes human children 8 months to recover after sibling birth. Then the recovery you found after seven and five months is right in line with what you expect. Why puzzling?
We have now explained – hopefully better – that it is not puzzling that the levels take a long time to decline, what is puzzling is that the model with the two cuts would suggest that cortisol levels decline at a time suddenly, similar to the sudden increase. We do not have an obvious explanation for why they should suddenly decline. When levels suddenly increase the sibling birth occurred, however, nothing we are aware of happens after 7-month period.
Line 378: Unless I missed this somewhere, it would be good to add a note about whether some of the siblings continued to make nipple contacts after the birth of a new baby. And maybe tell us how many of the total number of infants were observed to continue nursing after birth of baby. This is interesting information for others. Maybe in results would be good to add?
In the results we had written: The proportion of time the older offspring was observed in nipple contact showed a continuous decrease prior to sibling birth in both males and females, and reached zero about two months before sibling birth (Figure 2A-C, Table 3) (line 266 and 268). We hope these answers the question.
Line 382: Say what kind of samples. Fecal samples? Hair samples? Urine samples? Bone samples?
We now have added fecal (line 383).
Line 403: Affiliative intends? Not sure what you mean here.
We have now changed it to: intentions, as suggested by reviewer 2 (line 403).
Line 449: In baboons'? Maybe in baboons,…? Something is off here. Also, baboons repeats a few times. Some errors need fixing
We have now changed it to: For example, the presence of a sibling did not affect the HPA-axis later in life in baboons, but other early life adversities had lasting consequences (lines 448-449)
Line 486: I don't like the word syndrome in this context. Makes it sound like TTS is a disease.
We have now changed it to: physiological changes (line 484).
In methods, might be good to mention which authors collected which data, which authors did which analyses, etc. For example, the behavioral data were collected by focal animal sampling. Was this done by all the authors together, or just one? Give the initials of the author(s).
We thank the reviewer for this suggestion; however, this information is provided in the authors contribution.
https://doi.org/10.7554/eLife.77227.sa2Article and author information
Author details
Funding
Deutsche Forschungsgemeinschaft (BE 5511/4-1)
- Verena Behringer
Max Planck Institute for Evolutionary Anthropology (Open access funding)
- Gottfried Hohmann
Max Planck Institute of Animal Behavior (Open access funding)
- Barbara Fruth
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Acknowledgements
We thank the Institut Congolais pour la Conservation de la Nature (ICCN) for granting permission to conduct fieldwork on bonobos and issuing export permits for urine samples. We are grateful to the people of Lompole for hosting the LuiKotale Bonobo Project and all assistants contributing to the long-term data collection at LuiKotale. Thanks to Róisín Murtagh and Vera Schmeling for lab assistance. We are also grateful to Alison Ashbury for editorial advice. We thank Stacy Rosenbaum, Iulia Bădescu, and two anonymous referees for helpful comments and suggestions that greatly improved this article. The study was supported by funding by the German Research Foundation (Deutsche Forschungsgemeinschaft; grant number BE 5511/4-1). Long-term data collection at LuiKotale is funded by the Max-Planck-Society, Centre for Research and Conservation of the Royal Zoological Society of Antwerp, Federal Ministry of Education and Research (Germany), Leakey Foundation, Wenner-Gren Foundation, the George Washington University, and Bonobo Alive. Funding for laboratory analyses was provided by MPI-EVA and the German Primate Center.
Ethics
All samples were collected noninvasively and with permission of the Institut Congolais pour la Conservation de la Nature (ICCN).
Senior Editor
- Christian Rutz, University of St Andrews, United Kingdom
Reviewing Editor
- Stacy Rosenbaum, University of Michigan, United States
Reviewer
- Iulia Badescu
Publication history
- Received: January 20, 2022
- Preprint posted: February 17, 2022 (view preprint)
- Accepted: August 29, 2022
- Accepted Manuscript published: August 30, 2022 (version 1)
- Version of Record published: September 20, 2022 (version 2)
Copyright
© 2022, Behringer, Berghänel et al.
This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.
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