Nondestructive and rapid method for sex identification of the tropical sea cucumber Holothuria scabra by anal swab sampling

Abstract

As one of the most economically significant sea cucumber species, Holothuria scabra are naturally distributed in the Indo-West Pacific region and are widely cultural in Southeast Asia. The appearances of female and male H. scabra have no obvious phenotypic dimorphism. Thus, identification of sex-specific markers and establishment of sex detection methods are beneficial to artificially induced reproduction and genetic breeding in this species. In this study, a total of 26 male-specific and 9 female-specific sequences were obtained by genomic sequencing with 3 male and 3 female independent samples and a female mixed pool with 10 individuals of H. scabra. After PCR confirmation, two primer pairs (HSM24 and HSM25) for male-specific sequences were found to be true positive for all male individuals. Based on a male-specific marker (GenBank: OP218794), primers were designed for PCR, and loop-mediated isothermal amplification (LAMP) methods were further tested on another 30 sex-unknown individuals with DNA samples collected from a nondestructive anal swab method. In this case, the accuracies for PCR and LAMP were approximately 100%. Altogether, this study provided a nondestructive and rapid method for sex identification of H. scabra, which may contribute to the aquaculture of this economic species.