DOI: 10.5061/dryad.2v6wwpzvt
title: Experimentally-seeded social cues in the wild: Costs to bearers and potential benefits to receivers

description: Conspecifics can provide social cues about the presence of key features of
 the surrounding environment, such as food or predators. Attending to
 social cues may therefore potentially benefit receivers, or at least be
 worth following. Yet, bearing social cues could also be costly,
 particularly if it increases the likelihood of close-range interaction
 with non-kin. Here, we experimentally-seeded social cues in the wild onto
 focal individuals of the social hermit crab (Coenobita compressus),
 testing (1) the ‘potential benefits to receivers’ hypothesis, which
 predicts that receivers will follow social cues to orient towards valuable
 resources, and (2) the ‘costs to bearers’ hypothesis, which predicts that
 bearers of social cues will experience direct (physical) costs or indirect
 (constrained movement) costs due to interaction with receivers. Consistent
 with hypothesis (1), in natural encounters, conspecifics that crossed
 paths frequently made antennal contact, potentially gathering social
 information at close range. In experiments, naïve conspecifics followed
 focal individuals bearing ‘positive’ social cues (about a valuable food
 resource) significantly more often than they followed individuals bearing
 less attractive (‘neutral’ or ‘ambivalent’) social cues, pointing to a
 potential benefit. Consistent with hypothesis (2), individuals bearing
 positive social cues incurred greater direct and indirect costs, being
 physically flipped more often and achieving shorter displacements compared
 to individuals bearing other social cues. We conclude that
 experimentally-seeded social cues in the wild can confer costs to bearers
 and potentially benefit receivers. Broadly, the costs of bearing social
 cues, revealed here, underscore the importance of not overlooking that
 social cues may be costly.
description: <strong>Study site</strong> We
 conducted extended preliminary observations on highly social terrestrial
 hermit crabs (<em>Coenobita compressus</em>) in the wild from
 2018 to 2022, using these observations as a foundation for the present
 study. The present study was carried out between January and March 2022 at
 the beach-forest interface of our long-term study site (Laidre 2010) in
 Osa Peninsula, Costa Rica (8°23'55.6"N 83°20'52.8"W),
 where this species occurs naturally in abundance and displays high social
 activity levels. Our study had three different parts (see below): (1)
 antennal contact during natural encounters, (2) overall attraction to
 chemical stimuli in dishes, and (3) experimentally-seeded social cues. All
 data, both observational and experimental, were collected by the first
 author between 0500 and 1600. <strong>Antennal
 contact during natural encounters</strong> To
 quantify how frequently individuals wandering the beach gathered
 chemo-tactile social cues via antennal contact we carried out focal and
 behavioral sampling, distributed evenly across ten separate days. For
 focal sampling, the observer stood stationary and used binoculars from
 &gt; 3 m distance to observe specific, randomly-selected focal
 individuals on the surrounding beach. Each focal individual (N = 20 total
 individuals) was observed for 5 min. For behavioral sampling, a parallel
 methodology was used, with each observation period lasting 5 min (N = 20
 behavioral samples, for a total of 100 min of cumulative sampling). In
 behavioral sampling, instead of a specific focal individual being chosen,
 the observer scanned the entire surrounding beach through binoculars,
 waiting for any instances of encounters between pairs of
 individuals. An ‘encounter’ (Figure 1a) occurred, in
 both behavioral and focal sampling, when: two individuals, originating
 more than three body lengths apart, came within a body length of one
 another, with one or both individuals approaching the other head on. The
 two individuals that encountered one another therefore had equal
 opportunity to make ‘antennal contact’ (Figure 1a), which was defined as
 follows: one or both of an individual’s two antennae touched any part of
 the other individual (body or shell), at any time during the encounter,
 regardless of who initiated contact. Every encounter thus had three
 distinct outcomes: antennal contact could be made by (1) just one
 individual in the pair, (2) by both individuals, or (3) by neither
 individual. Note: any subsequent reference to ‘antennal contact’ generally
 (without specification of whether it was made by just ‘one’ or by ‘both’
 individuals in a pair) simply denotes these two separate outcomes being
 lumped. <strong>Overall attraction to chemical
 stimuli in dishes</strong> <em>Experimental
 setup</em> To determine levels of attraction to
 different chemical stimuli, we placed each chemical stimulus alone in a
 dish in the field, recording video from above the dish as free-roaming
 individuals in the wild moved to and fro. Three different types of
 chemicals (see below) were tested in randomized order each day across
 twenty days (with N = 20 trials of each of the three chemical conditions,
 for a total of N = 60 experimental trials). Each trial involved placing
 out only one shallow plastic circular dish (5 cm in diameter) and testing
 only one type of chemical at a time. In between trials the dish was
 retrieved, and a separate dish was deployed at a new, randomly designated
 location on the beach. The experimental area consisted of a 65 cm diameter
 circle drawn in the sand immediately around the dish (Figure 1b). To
 remove any barrier to access by crabs, the dish was pressed into the sand,
 with its sides and upper rim flush with the substrate.
 To video overhead during the experiments, a video camera (Canon
 Vixia HF R72) was attached to a long wooden pole driven into the sand.
 After the experimental area and video camera were setup, the entire
 experimental area was filmed continuously for twenty minutes (from t = -10
 min to t = +10 min). The first ten minute period (from t = -10 min to t =
 0 min) provided a baseline with an empty dish; then at t = 0 min the
 experimenter used a pipette to add 9 mL of chemical to the dish; after
 which the trial continued for another ten minutes with the filled dish
 (from t = 0 min to t = +10 min). A visual count of the number of
 individuals present in the experimental area was made just before the
 addition of the chemical stimulus at t = 0 min (‘start’ count) and once
 again at t = +10 min (‘end’ count). From the videos we also quantified the
 latency for the first individual to make contact with the dish after the
 chemical stimulus had been added. <em>Three
 conditions: ecologically-relevant chemical stimuli</em>
 Three ecologically-relevant chemical stimuli, all natural
 elements of the species’ environment, were chosen for their biological
 significance and were put into dishes in the field (see above) and also
 paired with conspecifics (see below): (1) coconut milk signifies a
 valuable food resource (Laidre 2013; Steele and Laidre 2019), therefore
 representing a ‘positive’ social cue when paired with a conspecific (for
 consistency across all our experiments, we used the same brand of coconut
 milk: ‘AROY-D’ San Jose, Costa Rica, for which the sole ingredient was
 coconut milk); (2) water from the adjacent lagoon that individuals
 regularly pass through (Laidre 2010, 2013) served as a control, therefore
 representing a ‘neutral’ social cue when paired with a conspecific; and
 (3) urine signifies both a potential predation risk (Laidre et al. 2012)
 and a potential shell opportunity (Valdes and Laidre 2019), therefore
 representing an ‘ambivalent’ social cue when paired with a conspecific.
 Note that nonhuman mammals (e.g., white-nosed coatis <em>Nasua
 narica</em> and raccoons <em>Procyon </em>spp.) can
 occasionally act as predators on our study system, but this is rare
 because generally their bite forces are below that required to break the
 crabs’ shells (Laidre et al. 2012). As a consequence, most nonhuman mammal
 predators are less dangerous than human predators (fishermen), which have
 the dexterity to directly pull crabs out of their shells, leaving behind
 the empty shells and using the crab body as fish bait. We therefore used
 human urine as an ecologically-relevant and readily collectable chemical
 that signifies both a predation risk and shell opportunity in this system.
 To collect urine, we used voluntary human donors at the field site
 (including the authors), all of whom consumed omnivorous diets. Urine was
 collected in the same receptacle at the same time each day. It is worth
 emphasizing that unambiguously ‘negative’ social cues are rare to
 non-existent in this species (cf. Doherty and Laidre 2020 on threat
 display), otherwise we would have included a fourth ‘negative’ chemical
 condition. <strong>Experimentally-seeded social
 cues: focal individuals doused in chemicals</strong>
 <em>Experimental setup</em> To
 experimentally seed social cues, the above three chemical stimuli (coconut
 milk, water, and urine) were paired with conspecifics, thereby generating
 positive, neutral, and ambivalent social cues, respectively. Each of these
 three social cue conditions was tested, in randomized order, each day
 across twenty days (with N = 20 trials per condition, for a total of N =
 60 experimental trials). For every trial, a randomly selected (new) focal
 individual was doused with one of the chemicals. After applying the
 chemical stimulus to the focal individual, thereby making it a bearer of
 social cues, we then released the focal individual at a randomly
 designated location on the beach and recorded all its subsequent
 interactions with conspecifics from a distance for 5 min using a handheld
 video camera (Canon Vixia HF R72). To test the impact of chemical-based
 social cues on bearer-receiver interactions, we quantified from video
 recordings: the number of encounters each focal individual had with
 conspecifics and all ensuing interactions (Figure 1c), which could either
 involve the first step toward a potential benefit to receivers (i.e.,
 receivers followed the bearer) or which could demonstrate a realized cost
 to the bearer (i.e., the bearer was flipped by receivers; see Table S1 for
 definitions of each behavior). Only encounters the
 focal individual had with moving conspecifics were counted to clearly
 distinguish conspecifics from stationary shell-like objects (pebbles) in
 the background of the video. All videos were coded by the first author and
 by an independent observer, who was blind to both the conditions and the
 hypotheses. To measure inter-observer reliability for the number of
 encounters, follows, and flips (see Figure 1 and Table S1), a random
 sample of videos (N = 30 total, including N = 10 of the experimental
 trials from each of the three conditions) were coded. There was strong
 inter-observer reliability (r<sup>2</sup> &gt; 0.75) for
 all behaviors (encounters: F<sub>1,28</sub> = 94.2, p &lt;
 0.0001, r<sup>2</sup> = 0.77; follows:
 F<sub>1,28</sub> = 88.53, p &lt; 0.0001,
 r<sup>2</sup> = 0.76; flips: F<sub>1,28</sub> =
 705.83, p &lt; 0.0001, r<sup>2</sup> = 0.96).
 <em>Focal individual selection</em>
 For each day of experiments, three focal individuals, among
 usually thousands of social hermit crabs wandering the beach, were
 randomly selected and then allocated randomly to each of the three
 chemical conditions. A crab was only included as a focal individual if it
 had all appendages intact and was occupying the most common (Nerita spp.)
 shell. Across the three conditions, there was no difference in shell
 diameter of focal individuals (one-way ANOVA: F<sub>2,57</sub>
 = 0.04, p = 0.96; positive condition: 11.48 ± 0.33 mm; neutral condition:
 11.33 ± 0.45 mm; ambivalent condition: 11.45 ± 0.47 mm).
 <em>Application of chemicals onto focal
 individual</em> Before the start of each trial,
 the experimenter’s hands and a set of forceps were thoroughly washed, and
 dried, removing any potentially confounding chemical cues. Also, to enable
 identification of focal individuals in the video, the dorsal side of each
 crab’s shell was marked with a small square of highly visible duct tape,
 which was removed at the end of the trial. After its shell was marked, the
 chemical stimulus was seeded onto the focal individual, by placing the
 individual (while it continuously remained occupying its original shell)
 into a temporary holding container and then using a pipette to douse the
 individual with 9 mL of the chemical (Figure 1c). Dousing ensured that the
 entire external surface of the crab’s body and shell were covered with the
 chemical cue, something which occurs commonly and naturally when crabs’
 forage on many foods, including coconut (Laidre 2013). The focal
 individual was then removed from the temporary holding container using
 forceps and held in the air while any excess chemical ceased dripping,
 leaving just a thin residue on its exterior surface for close-range
 detection by receivers. The focal individual was then
 carried with clean forceps to a randomized drop point on the beach, where
 it was placed on top of the sand, with its start position marked by the
 insertion of a vertical wooden dowel. The number of conspecifics within a
 30 cm radius of this start position was recorded, then the focal crab was
 videoed continuously for 5 min. After 5 min, the focal individual’s end
 position was marked by the insertion of another vertical wooden dowel,
 with the number of conspecifics within a 30 cm radius of this end point
 being recorded, and then the focal individual was recollected. Upon
 recollection, the mark on the focal individual’s shell was removed, and
 the individual was rinsed with water before being placed in a separate
 holding container until the end of the day’s experiments were completed.
 After all experiments were finished for the day, all focal individuals
 were released back into the wild. To test if there was a difference in how
 far focal individuals moved across the different social cue conditions, we
 measured the distance between their start and end positions (i.e., their
 displacement; Figure 1c). Displacement is a strong indicator of the extent
 to which individuals can explore their environment (Trinh and Laidre 2016;
 Doherty and Laidre 2022), with shorter displacements providing less
 opportunity for discovering new resources (Laidre 2010; Steele and Laidre
 2023a). <strong>Statistical
 analyses</strong> <em>Antennal contact
 during natural encounters</em> We tested whether
 natural encounters were differentially likely to lead to antennal contact
 by both individuals, just one individual, or neither individual. For focal
 sampling, we tested differences in this likelihood of antennal contact by
 comparing the number of encounters with each of the three different
 outcomes (note: no formal statistical test was performed, since these
 three outcomes were non-independent, all deriving from the same N= 20
 focal individuals). For behavioral sampling, in which the three outcomes
 were independent, we tested differences in the likelihood of antennal
 contact by performing a Chi-square test, which compared observed
 occurrences of each of the three outcomes to the null assumption of an
 equal likelihood of these three possible outcomes.
 <em>Overall attraction to chemical stimuli in
 dishes</em> To test for differences in latency to
 contact dishes across the three chemical conditions, we performed a
 one-way ANOVA. To test if the positive (coconut milk) condition attracted
 individuals faster than the neutral (water) or ambivalent (urine)
 conditions, we conducted post-hoc All-Pairs Tukey-Kramer tests, which
 simultaneously allowed us to explore any differences between the neutral
 and ambivalent conditions. Data for latency were square route transformed
 (which made these data normally distributed) and analyses were then
 conducted on the transformed data. To compare
 differences in the number of individuals present in the experimental area
 from before-to-after the addition of each chemical, we performed a
 Kruskal-Wallace test. To test if the positive condition attracted more
 individuals than the neutral or ambivalent conditions, we conducted
 post-hoc All-Pairs Dunn tests, which simultaneously allowed us to explore
 any differences between the neutral and ambivalent conditions.
 <em>Experimentally-seeded social cues: focal individuals
 doused in chemicals</em> To test for differences
 across the three social cue conditions, we used Kruskal-Wallace tests to
 compare the number of encounters focal individuals had with conspecifics
 and the number of conspecifics surrounding the focal individual at the end
 of each experiment. For any variables that showed a significant difference
 across conditions, we then used post-hoc All-Pairs Dunn tests to determine
 if the positive condition differed from the neutral or ambivalent
 conditions and if any difference existed between the neutral and
 ambivalent conditions. To test differences across the three social cue
 conditions in the proportion of encounters that led to receivers following
 the focal individual (the bearer), we arcsine transformed these data
 (which made these data normally distributed) and performed a one-way
 ANOVA. A post-hoc All-Pairs Tukey-Kramer test was then conducted to
 determine if individuals in the positive condition followed bearers more
 than individuals in the neutral or ambivalent conditions, and if any
 difference existed between the neutral and ambivalent conditions. To test
 differences across the three social cue conditions in how many times focal
 individuals were flipped by receivers, we used a generalized linear model
 (GLM), with a Poisson distribution and a Log link function. Finally, to
 test differences across the three social cue conditions in displacement
 achieved by focal individuals, we performed a one-way ANOVA. A post-hoc
 All-Pairs Tukey-Kramer test was then conducted to determine if individuals
 in the positive condition achieved shorter displacements than individuals
 in the neutral or ambivalent conditions, and if any difference existed
 between the neutral and ambivalent conditions. Analyses
 were performed using JMP® Pro version 16.0.0. For post-hoc analyses,
 All-Pairs Tukey-Kramer tests controlled for the overall error rate, and
 All-Pairs Dunn tests used the Bonferroni adjustment. All post-hoc tests
 are thus reported as above or below p = 0.05.
description: # Experimentally-seeded social cues in the wild: costs to bearers and
 potential benefits to receivers
 The dataset provided is a
 study conducted in Jan - March 2022 in Osa Peninsula, Costa Rica, at a
 long-term field site (Osa Conservation’s Piro Biological Station). The
 file includes a metadata section to help navigate the raw data from 3
 related datasets, as well as screenshots of results. Datasets include: 1\)
 Antennal contact * Including: Sampling type (Focal, Behavioural); Number
 of encounters that resulted in antennal contact by no individuals, number
 of encounters that resulted in antennal contact by one individual, and
 number of encounters that resulted in antennal contact by two individuals
 (during each 5-minute sampling period); Total number of encounters
 throughout each 5-minute observation; Antennal contact rate per minute.
 2\) Overall attraction to chemicals * Including: What triplicate the
 experiment was a part of; What chemical stimuli was being tested (coconut
 milk, water, urine); What condition that stimulus represented (positive,
 neutral, ambivalent - respectively); Number of individuals within 30cm of
 the stimulus at the start of the trial (t = 0 mins); Number of individuals
 within 30cm of the stimulus at the end of the trial (t = +10 mins); Change
 in number of individuals (end minus start); Latency for any individual to
 first contact the stimulus (in seconds). 3\) Experimentally-seeded social
 cues * Including: What triplicate the experiment was a part of; Shell
 species worn by the focal crab (Nerita); Shell diameter of focal crab
 (mm); What chemical stimuli was being tested (coconut milk, water, urine);
 What condition that stimulus represented (positive, neutral, ambivalent -
 respectively); Total number of encounters focal individual had with
 conspecifics throughout the duration of the 5-minute trial; Number of
 encounters that resulted in the focal crab being followed by a
 conspecific; the proportion of encounters that resulted in following by a
 conspecific; The arcsin square root of the proportion of encounters that
 resulted in following by a conspecific (for statistical analysis); The
 back transformation of the arcsin square root of the proportion of
 encounters that resulted in following by a conspecific (for graphing);
 Number of conspecifics within 30cm of the focal crabs end position; Number
 of times the focal crab is flipped over by a conspecific; displacement (in
 m, i.e., linear distance between the focal crabs position from the start
 to the end of the 5 minute trial) For further information, or if you have
 any questions or comments about the data - collection, or analysis -
 please contact the author.