Mud crabs, Scylla spp., are commercially important large-size marine crustaceans in the Indo-West Pacific region. As females have the higher growth rate and economic value, the production of all female stocks is extremely essential in aquaculture. However, the sex determination mechanism is still unclear. Development of sex-specific genetic markers based on next-generation sequencing proved to be an effective tool for discovering sex determination system in various animals. Restriction-site associated DNA sequencing (RAD-seq) was employed to isolate sex-specific SNP markers for S. paramamosain. A total of 335.6 million raw reads were obtained from 20 individuals, of which 204.7 million were from 10 females and 130.9 million from 10 males. After sequence assembly and female-male comparison, 20 SNP markers were identified to be sex-specific. Furthermore, ten SNPs in a short sequence (285 bp) were confirmed heterozygous in females and homozygous in males in a large population by PCR amplification and sequencing. Subsequently, a female-specific primer was successfully designed according to the female-specific nucleotide which could amplify an expected band from females but not from males. Thus, a rapid and effective method for molecular sexing in S. paramamosain was developed, meanwhile, this method could successfully identify the sex of S. tranquebarica and S. serrata. Finally, nine and four female-specific SNP markers were detected in S. tranquebarica and S. serrata, respectively. Sex-specific SNP markers were firstly identified in crab species and showed female heterogamety and male homogamety, which provided strong genetic evidence for a WZ/ZZ sex determination system in mud crabs S. paramamosain, S. tranquebarica and S. serrata. These findings will lay a solid foundation for the study of sex determination mechanism, sex chromosome evolution, and the development of mono-sex population in crustaceans.

Mud crab, Scylla paramamosain is one of the most important crustacean species in global aquaculture. To determine the genetic basis of sex and growth-related traits in S. paramamosain, a high-density genetic linkage map with 16,701 single nucleotide polymorphisms (SNPs) was constructed using SLAF-seq and a full-sib family. The consensus map has 49 linkage groups, spanning 5,996.66 cM with an average marker-interval of 0.81 cM. A total of 516 SNP markers, including 8 female-specific SNPs segregated in two quantitative trait loci (QTLs) for phenotypic sex were located on LG32. The presence of female-specific SNP markers only on female linkage map, their segregation patterns and lower female: male recombination rate strongly suggest the conformation of a ZW/ZZ sex determination system in S. paramamosain. The QTLs of most (90%) growth-related traits were found within a small interval (25.18–33.74 cM) on LG46, highlighting the potential involvement of LG46 in growth. Four markers on LG46 were significantly associated with 10–16 growth-related traits. BW was only associated with marker 3846. Based on the annotation of transcriptome data, 11 and 2 candidate genes were identified within the QTL regions of sex and growth-related traits, respectively. The newly constructed high-density genetic linkage map with sex-specific SNPs, and the identified QTLs of sex- and growth-related traits serve as a valuable genetic resource and solid foundation for marker-assisted selection and genetic improvement of crustaceans.

This study was conducted to determine the effects of dietary cellulase addition on improving the nutritive value of Chlorella for juvenile crucian carp Carassius auratus (initial body weight: 2.99 ± 0.02 g, mean ± SEM). Five isonitrogenous and isoenergetic experimental diets were formulated to contain 0.0 (control), 0.5, 1.0, 1.5 and 2.0 g kg⁻¹ cellulase, respectively. Each experimental diet was randomly assigned to triplicate groups with 25 juvenile fish per fibreglass tank for 8 weeks. The results showed that weight gain, specific growth rate, feed intake and the trypsin activity in the anterior intestine increased with increasing dietary cellulase to 1.5 g kg⁻¹ and then declined with further addition. However, the mRNA expression levels of Mrf4 and Myf5, the apparent digestibility coefficients for dry matter, protein, energy and the majority of amino acids, and the activity of lipase in the anterior intestine were highest in fish fed the 1.0 g kg⁻¹ cellulase diet, and then tended to decline with further cellulase supplementation. In conclusion, the optimal dietary cellulase supplementation level was 1.0–1.5 g kg⁻¹, which can improve growth performance, digestive activities and nutrient digestibility in crucian carp.