Males of the hermit crab Pagurus filholi perform precopulatory guarding behavior, and solitary males often show aggressive behavior to take away guarded females. Males behave coercively while guarding females, so direct mate choice by females seems difficult in such a situation. By performing several experiments we examined possible indirect female choice of hermit crab. Males were attached to a shell by their left cheliped to look like guarding pairs (fake guarding pairs). The shells were filled with cotton containing either seawater or pheromone water. The fake guarding pair with only seawater caused malemale combat in 60% of trials whereas with pheromone water combats occurred in 88% of trials. Mean duration of male-male combat was significantly longer in trials with drops of seawater containing pheromones than in those without pheromones. These results suggest guarding pairs themselves cause male-male combat by visual stimulation, that female sex pheromones have further significant function in the recognition of guarding pairs and intensification of male-male combat, and that females release sex pheromones while they are guarded. As a result of the combat, the larger male ended up guarding a female. This strongly suggests that females choose males indirectly by exploiting male-male competition induced by sex pheromones under male coercive behavior.

Males of the hermit crab Pagurus filholi perform assessment behavior toward females, as a preliminary step of precopulatory guarding, during the reproductive season. It is known that such behavior is elicited by female sex pheromones, but the compounds involved have never been characterized in this species. Several experiments were conducted to develop a reliable bioassay along with purification procedures to identify potential compounds with pheromonal activity in Pagurus filholi. We developed a bioassay protocol to assess pheromonal activity by using an empty shell with cotton containing either artificial seawater (control) or test water. We measured and compared the time duration of male assessment behavior toward each shell if the test water contained female sex pheromones. Ultra-filtering of seawater samples potentially containing pheromones showed that the compound was <1 kDa in molecular weight. Males showed precopulatory assessment behavior toward “female conditioned” water samples treated with open column purification and eluted with MeOH, suggesting that compounds triggering male behavior were low polar molecules. Molecules with pheromonal activity were not volatile after freeze drying, effective even after heating to 90 °C, and remained active in seawater at 12 °C even after 6 days from sample collection, which suggests a rather stable characteristic of the female sex pheromones of this species.

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