We tested whether the leg condition (missing legs or not) of an arachnid species (Prionostemma sp.2) affects roosting location within the group by experimentally inducing the formation of aggregations overnight. Autotomized individuals roosted more frequently in the aggregation center than intact individuals. Additionally, this pattern was observed only for aggregations of more than 13 individuals.

We studied Opiliones of Prionostemma sp.2 at La Selva Research Station, in Puerto Viejo de Sarapiquí, Heredia, Costa Rica (10.4311 N, 84.0068 W, elevation: 50 m) in February 2024. We collected individuals at night on two the trails (STR and SUR) for lab experiments. We conducted this research following research permits of the Costa Rican Government (permit # SINAC-ACC-PI-re-003-2024). Additionally, we ensure the welfare of Opiliones throughout all stages of this work by following suggested guidelines and best practices.

To test the hypothesis of the location within the group to roost, we placed the collected Opiliones in an arena composed of a plastic shoe box (33 × 20 × 12 cm) at night (19:00 - 21:00 h) to induce the formation of overnight aggregations. We added similar numbers of individuals with both leg conditions: The ratio of autotomized to intact individuals across the 64 groups we studied was 1.19 ± 0.37 (mean ± standard deviation). We studied 858 individuals, 46% of whom had eight legs, 38% had seven, 13% had six, and 3% had five. We did this in the Environmental Lab at La Selva (an open-window room with ambient temperature). A 20 × 10 cm wooden surface or bark fragment in each box provided a natural substrate to aggregate (Fig. 2A). We kept each box vertical to mimic the tree buttress where Opiliones roost.

In each trial, we added a total of 5 to 22 individuals. We studied 32 small groups (5 - 11 individuals, n=308), 17 medium groups (13 - 16 individuals, n=255), and 15 large groups (18 - 22 individuals, n=304).

We compared the proportions of autotomized and intact Opiliones in an aggregation’s center and periphery. The following morning, we photographed each group with a GoPro Hero 8 placed 50 cm away. To identify individuals as in the “center” or the “periphery” of the group from the pictures, we drew circles centered on their bodies with a radius equal to approximately 2 body widths. Because these circles encompassed each individual, we labeled the circles as in the center if they were completely surrounded by other circles. Then we labeled circles as in the periphery if they were not fully surrounded by others.

To test if leg condition affects the roosting location choice, we ran a goodness-of-fit χ² test with two nominal variables: group position (center or periphery) and leg condition (autotomized or intact) using all data (858 individuals in 64 aggregations). To explore if the proportion of individuals across the group’s locations differed between group size classes, we ran three additional goodness-of-fit χ² tests, one for each group size (small n=308, medium n=255, and large n=304). We performed all analyses in R version 4.2.1.