Effects of scent enrichment on behavioural and physiological indicators of stress in zoo primates – a preliminary study on red-ruffed lemurs and Lar gibbons

Almost half of the primate species recognized today worldwide are classified as endangered or critically endangered in the wild. Captive breeding is vital for primate conservation, with modern zoos serving a crucial role as breeders of populations acting as buffers against extinction, ambassadors of endangered species, and educators of the general public. However, captive populations may experience welfare issues that may also undermine their breeding success. In order to design and test a new scent enrichment programme to enhance the breeding success and well-being of critically endangered zoo primates, we carried out a preliminary study to assess the effects of lavender on captive red-ruffed lemurs (Varecia rubra) and Lar gibbons (Hylobates lar) as these species presently show a low success rate in captive breeding and are critically endangered in the wild. We combined behavioural observations and faecal endocrinology analyses to assess the effects of lavender on two captive troops (N = 8) housed at Dudley Zoo and Castle (UK). We recorded observations of natural species-specific and abnormal stress-related behaviours for 144 hr using instantaneous scan sampling. We collected 78 faecal samples and measured the faecal cortisol concentrations using radioimmunoassay. We found a significant effect by the scent enrichment on social interactions and stress-related behaviours (i.e., pacing and self-scratching), with both species reducing their rates of stress-related behaviours after they were exposed to lavender. We also found that both species displayed a significant increase in faecal glucocorticoids following the exposure to lavender. Our findings suggest that lavender may work as scent enrichment to decrease the stress levels of zoo primates across the major lineages, but its effects might change depending on the species-specific social lives and olfactory repertoires of primates.


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We carried out behavioural observations and faecal sampling from September to 128 December 2019 (red-ruffed lemurs) and from September to December 2018 (Lar gibbons).
129 Both troops lived in an indoor enclosure (heated to 28 °C) with access to an outdoor 130 enclosure (i.e., "visitor walkthrough" enclosure in the case of red-ruffed lemurs).

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We divided the overall study period in three periods: baseline, scent enrichment, post 135 enrichment. We collected behavioural data and faecal samples (see below "Assessment of 136 welfare") for four days per study period (12 days in total) over three months.

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138 Scent enrichment 139 140 We cut white cotton sheets into 75 cm long and 5 cm wide strips, which we then 141 soaked with 20 drops naissance 100% pure lavender essential oil diluted with 12 ml of cold 142 boiled water. We prepared the scent cotton strips at early morning of each sampling day over 143 the scent enrichment period. We positioned these strips around both indoor and outdoor 144 enclosures; particularly, and we tied them approximately 1 m from the ground around the 145 climbing frames as these were the most used areas of the enclosures.

Sleeping
Lying on back, front or side, eyes closed and whole body is relaxed Grooming Using fingers or mouth to pick through the coat, removing any foreign bodies from a conspecific Play Animal is engaging in activities such as chasing others, leaping around the enclosure etc. in a playful context Self-scratching An animal rubs their own body at a fast pace Pacing Animal walks back and forth in a distinct, unchanging pattern within the enclosure.

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The 2x2 ANOVA model that included resting rates as the outcome variable revealed 212 only a significant main effect of Species, with red-ruffed lemurs resting more frequently than 213 Lar gibbons, but there was no significant main effect of Enrichment condition or the 214 interaction between Enrichment and Species on resting rates (Table 2). By contrast, we found a significant effect of the interaction between enrichment 222 condition and species on rates of social interaction (Table 3). Follow-up post-hoc t-tests 223 revealed that, while Lar gibbons increased their rates of social interactions following their 224 exposure to lavender (t(3.2) = 7, p = 0.005, d = 5.7, Figure 1), red-ruffed lemurs showed a 225 reduction in rates of social interactions after being exposed to lavender (t(8.3) = 2.8, p = 0.02, Fig. 1).

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228  234 Rates of stress-related behaviours were significantly predicted by enrichment 236 condition but not by species or the interaction between species and enrichment condition 237 (Table 4). More specifically, both species reduced their rates of stress-related behaviours 238 after they were exposed to lavender (Fig. 2).

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Finally, we found that enrichment condition significantly predicted FGCs (Table 5): 248 following the exposure to lavender, both species displayed a significant increase in FGC 249 concentration, compared to before being exposed to the scent enrichment (Fig. 3).

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In this study, we found a significant effect of the scent enrichment on social 289 showed that play rates were higher in individuals who experienced low stress levels [34][35][36].
290 In this context, our findings that Lar gibbon spent more time playing after the introduction of 291 the scent enrichment compared to before the exposure to lavender might reflect the fact that 292 this species might have experienced lower stress levels after the exposure to the scent, which 293 is consistent with our prediction. On the other hand, social grooming, a common affiliative 294 behaviour in primates, has long been shown to have a stress-reducing effect. Work conducted 295 on macaques has shown, for example, that either receiving or giving grooming lowers stress 296 levels in the individuals involved in the interaction [36-42]. We, therefore, suggest that red-297 ruffed lemurs reduced their rates of social grooming following the exposure to the scent 298 enrichment because the presence of lavender might have decreased their overall stress levels.

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As we predicted, we found a significant effect on the scent enrichment on stress-300 related behaviours, with both red-ruffed lemurs and Lar gibbons reducing their rates of stress-301 related behaviours after they were exposed to lavender. This is consistent with our prediction 302 that scent enrichment reduces behavioural indicators of stress (i.e., pacing and self-

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In conclusion, our preliminary study provides strong evidence of a beneficial effect of 369 scent enrichment on animal welfare. Following exposure to the scent enrichment, animals 370 spend more time play, and less time grooming and exhibit lower rates of stress-related 371 behaviours. Our finding that both red-ruffed lemurs and Lar gibbon increased their levels of 372 FGCs after exposure to lavender might be associated to an increase in activity after animals 373 were exposed to the scent enrichment. Future work will need to expand the investigation of 374 the effect of scent enrichment on animal welfare by focusing on a wider range of primate 375 species and on a bigger sample size.