Unique multiple paternity in the endangered big-headed turtle (Platysternon megacephalum) in an ex situ population in South China

Understanding the mating system and reproductive strategies of an endangered species is critical to the success of captive breeding. The big-headed turtle (Platysternon megacephalum) is one of the most threatened turtle species in the world. Captive breeding and reintroduction are necessary to re-establish wild populations of P. megacephalum in some of its historical ranges in China, where the original populations have been extirpated. However, the captive breeding of P. megacephalum is very difficult and this may be due to its mysterious reproductive strategies and special behavior (e.g., aggressive temperament and territoriality). In this study, we achieved successful captive breeding of P. megacephalum by creating a habitat that mimics natural conditions and then investigated its mating system using microsatellite makers. A total of 16 clutches containing 79 eggs of P. megacephalum were collected, and 52 were hatched successfully over two breeding seasons. Of the 15 effective clutches, 6 clutches (40%) exhibited multiple paternity. There was no significant correlation between clutch size and multiple paternity, and no significant difference in hatching success between multiple-sired and single-sired clutches. However, there was significant correlation between male body size and the number of offspring, with higher-ranked males contributing to more clutches. Our results provide the first evidence of multiple paternity and male hierarchy in P. megacephalum. These findings suggest that multiple paternity and male hierarchy should be considered in captive breeding programs for P. megacephalum, and creating a habitat that mimics natural conditions is an effctive way to achieve successful captive breeding and investigate the mating systems of this species.