Morphological descriptions and morphometric discriminant function analysis reveal an additional four groups of Scylla spp.

There are four species of mud crabs within the genus Scylla, and most of them live sympatrically in the equatorial region. Apart from a report in Japan about the finding of a natural Scylla hybrid more than a decade ago after the division of genus Scylla into four species by Keenan, Davie & Mann (1998), no subsequent sighting was found. Thus, this study investigates the possible natural occurrence of potential hybridization among Scylla species in the wild. A total of 76,211 individuals from mud crab landing sites around the Malacca Straits, South China Sea and Sulu Sea were screened. In addition to the four-purebred species, four groups (SH 1, n = 2, 627; SH 2, n = 136; SH 3, n = 1; SH 4, n = 2) with intermediate characteristics were found, mostly at Sulu Sea. Discriminant Function Analysis revealed that all Scylla species, including SH 1 - 4, are distinguishable via their morphometric ratios. The most powerful discriminant ratios for each character and the top five discriminant ratios of males and females were suggested. The carapace width of SH 1 males and females were significantly smaller than pure species. Based on the discriminant ratios and the description of morphological characters, we hypothesize that the additional four groups of Scylla with intermediate characteristics could be presumed hybrids. Future work at the molecular level is urgently needed to validate this postulate.

Transcriptome Analysis and Differential Gene Expression on the Testis of Orange Mud Crab, Scylla olivacea, during Sexual Maturation.

Adequate genetic information is essential for sustainable crustacean fisheries and aquaculture management. The commercially important orange mud crab, Scylla olivacea, is prevalent in Southeast Asia region and is highly sought after. Although it is a suitable aquaculture candidate, full domestication of this species is hampered by the lack of knowledge about the sexual maturation process and the molecular mechanisms behind it, especially in males. To date, data on its whole genome is yet to be reported for S. olivacea. The available transcriptome data published previously on this species focus primarily on females and the role of central nervous system in reproductive development. De novo transcriptome sequencing for the testes of S. olivacea from immature, maturing and mature stages were performed. A total of approximately 144 million high-quality reads were generated and de novo assembled into 160,569 transcripts with a total length of 142.2 Mb. Approximately 15-23% of the total assembled transcripts were annotated when compared to public protein sequence databases (i.e. UniProt database, Interpro database, Pfam database and Drosophila melanogaster protein database), and GO-categorised with GO Ontology terms. A total of 156,181 high-quality Single-Nucleotide Polymorphisms (SNPs) were mined from the transcriptome data of present study. Transcriptome comparison among the testes of different maturation stages revealed one gene (beta crystallin like gene) with the most significant differential expression-up-regulated in immature stage and down-regulated in maturing and mature stages. This was further validated by qRT-PCR. In conclusion, a comprehensive transcriptome of the testis of orange mud crabs from different maturation stages were obtained. This report provides an invaluable resource for enhancing our understanding of this species' genome structure and biology, as expressed and controlled by their gonads.

A simple and efficient total genomic DNA extraction method for individual zooplankton.

Molecular approaches are widely applied in species identification and taxonomic studies of minute zooplankton. One of the most focused zooplankton nowadays is from Subclass Copepoda. Accurate species identification of all life stages of the generally small sized copepods through molecular analysis is important, especially in taxonomic and systematic assessment of harpacticoid copepod populations and to understand their dynamics within the marine community. However, total genomic DNA (TGDNA) extraction from individual harpacticoid copepods can be problematic due to their small size and epibenthic behavior. In this research, six TGDNA extraction methods done on individual harpacticoid copepods were compared. The first new simple, feasible, efficient and consistent TGDNA extraction method was designed and compared with the commercial kit and modified available TGDNA extraction methods. The newly described TGDNA extraction method, "Incubation in PCR buffer" method, yielded good and consistent results based on the high success rate of PCR amplification (82%) compared to other methods. Coupled with its relatively consistent and economical method the "Incubation in PCR buffer" method is highly recommended in the TGDNA extraction of other minute zooplankton species.