Genetic evidence for multiple paternity in the critically endangered Cuban crocodile (Crocodylus rhombifer)

Conservation strategies can be most effective when factors influencing the persistence of populations are well-understood, including aspects of reproductive biology such as mating system. Crocodylians have been traditionally associated with a polygynous mating system, with genetic studies revealing multiple paternity of clutches in several species. The endemic Cuban crocodile, <i>Crocodylus rhombifer</i>, is currently listed as Critically Endangered, and is one of the least understood crocodylian species in terms of its mating behavior. Here, we tested a hypothesis of multiple paternity in the Cuban crocodile by collecting genotypic data at nine microsatellite loci for 102 hatchlings from five nests sampled at the Zapata Swamp captive breeding facility and analyzing them in relation to data previously collected for 137 putative parents. All five nests showed evidence of multiple paternity based on the numbers of alleles per locus, with sibship analyses reconstructing all nests as having four to six full-sib family groups. Accordingly, mean pairwise relatedness values per nest ranged from 0.21 to 0.39, largely intermediate between theoretical expected values for half-siblings (0.25) and full-siblings (0.50). It is not possible to differentiate whether the multiple paternity of a nest was due to multiple matings during the same breeding season, or a result of sperm storage. Our results reveal that the <i>C. rhombifer</i> mating system is likely best characterized as promiscuous and suggest that the standard practice of enforcing a 1:2 sex ratio at the captive breeding facility should be altered in order to better maintain a demographically and genetically healthy ex situ population