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The sea cucumber (phylum Echinodermata) body wall is the first line of defense and is well known for its production of secondary metabolites; including vitamins and triterpenoid glycoside saponins that have important ecological functions and potential benefits to human health. The genes involved in the various biosynthetic pathways are unknown. To gain insight into these pathways in an echinoderm, we performed a comparative transcriptome analysis and functional annotation of the body wall and the radial nerve of the sea cucumber Holothuria scabra; to define genes associated with body wall metabolic functioning and secondary metabolite biosynthesis. We show that genes related to signal transduction mechanisms were more highly represented in the H. scabra body wall, including genes encoding enzymes involved in energy production. Eight of the core triterpenoid biosynthesis enzymes were found, however, the identity of the saponin specific biosynthetic pathway enzymes remains unknown. We confirm the body wall release of at least three different triterpenoid saponins using solid phase extraction followed by ultra-high-pressure liquid chromatography-quadrupole time of flight-mass spectrometry. The resource we have established will help to guide future research to explore secondary metabolite biosynthesis in the sea cucumber.

Wild sea cucumber resources have been rapidly exhausted and therefore there is an urgent need to develop approaches that will help restocking. Currently, there is a lack of information regarding the genes involved in sea cucumber reproductive processes. The neurohormone relaxin-like gonad-stimulating peptide (RGP) has been identified as the active gonad-stimulating peptide in sea stars (Asteroidea), which could also be present in other echinoderm groups. In this study, a sea cucumber RGP was identified and confirmed by phylogenetic analysis. A recombinant Holothuria scabra RGP was produced in the yeast Pichia pastoris and confirmed by mass spectrometry. To assess bioactivity, four levels of purification were tested in an in vitro germinal vesicle breakdown (GVBD) bioassay. The most pure form induced 98.56 ± 1.19% GVBD in H. scabra and 89.57 ± 1.19% GVBD in Holothuria leucospilota . Cruder levels of purification still resulted in some GVBD. Upon single injection into female H. scabra , the recombinant RGP induced head waving behavior followed by spawning within 90–170 min. Spawned oocytes were fertilized successfully, larvae settled and developed into juveniles. Our results provide a key finding for the development of a break-through new artificial breeding approach in sea cucumber aquaculture.

We isolated 18 new, polymorphic, microsatellite markers from Holothuria scabra, a commercially important species of sea cucumber found throughout the Asia-Pacific region. Next-generation sequencing was used to identify 206 unique loci for which primers were designed. Of these unique loci, we trialled 65 primer pairs in the target species, and 48 (74%) amplified a product of the expected size. Eighteen loci were found to be polymorphic and reliable, and were screened for variation in 50 individuals, from a single population from Croker Island, Northern Territory, Australia. Observed heterozygosity ranged from 0.00 to 0.96 (mean = 0.46) and the number of alleles per locus from 2 to 28 (mean = 9.61). These loci will be useful for the investigation of population structure and mating systems in H. scabra and may also be of use in other holothurian species.

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