Abstract
Pheromones are an important component of sexual communication in courting salamanders, but the number of species in which their use has been demonstrated with behavioral evidence remains limited. Here we developed a behavioral assay for demonstrating courtship pheromone use in the aquatically courting Iberian ribbed newt Pleurodeles waltl. By performing an in-depth study of the courtship behavior, we show that females invariably open their cloaca (cloacal gaping) before engaging in pinwheel behavior, the circling movement that is the prelude to spermatophore uptake. In contrast, cloacal gaping was not observed in failed courtships, where females escaped or displayed thanatosis. Since gaping mainly occurred during male amplexus and cloacal imposition, which is the obvious period of pheromone transfer, we next investigated whether male courtship water (i.e., water holding courtship pheromones) alone was able to induce this reaction in females. These tests showed that courtship water induced cloacal gaping significantly more than water, even in the absence of a male. Cloacal gaping thus provides a simple and robust test for demonstrating courtship pheromone use in the Iberian ribbed newt. Since opening the cloaca is an essential prerequisite for spermatophore pick-up in all internally fertilizing salamanders, we hypothesize that variations on this assay will also be useful in several other species.
Highlights
Courtship pheromones are chemical signals that elicit a specific reaction in conspecifics of the opposite gender during courtship [1]
Successful insemination in Pleurodeles waltl always results from pinwheel behavior (PW), i.e. the couple going into a circular movement that leads to spermatophore deposition by the male, and subsequent uptake by the female cloaca [3,24,25,30]
When the female immediately responded positively to the nudging male (2/42 observations), the male was triggered to pivot around the female, while she followed him in pinwheel behavior without interlocked forelimbs (Fig 1A, outcome 1; S1 Video)
Summary
Courtship pheromones are chemical signals that elicit a specific reaction in conspecifics of the opposite gender during courtship [1]. In the terrestrially courting lungless salamanders (Plethodontidae), courtship behavior among species is remarkably homogeneous [3], and consists of a tail-straddling walk in which the female holds her chin on the male’s tail base, while both move forward until the spermatophore transfer has occurred. For this family, a behavioral test has been designed in which the pheromone-producing mental glands of the male were removed and courtship duration of couples was compared with and without the application of pheromones [7,8]. PLOS ONE | DOI:10.1371/journal.pone.0144985 January 15, 2016
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