Cynoglossus semilaevis, commonly known as the Chinese tongue sole, is a popular aquaculture flatfish species. [1] They are native to China's northern coast but have experienced overfishing these past three decades. [1] [2] Tongue sole farming began in 2003, and they have since become a popular, expensive seafood. [2] Farmers in this practice encounter issues related to the tongue sole's pathogen susceptibility and uneven sex ratio. [2]
Cynoglossus semilaevis has a female heterogametic sex-determination system, with females being ZW and males being ZZ. [3] Female tongue soles grow up to 2-4 times larger and faster than males, making them more desirable for aquaculture. To maximize yields, efforts have been made to produce all-female stocks through artificial gynogenesis. [4] However, the sex ratio of C. semilaevis populations in both the wild and aquaculture environments is male-skewed due to the occurrence of pseudomales [5] or individuals with female ZW chromosomes that become physiologically male. [6] Offspring of pseudomales tend to grow more slowly than those of genetic males and are more likely to become pseudomales themselves, further skewing the sex ratio. [3] [5] This phenomenon is influenced by a combination of genotypic sex determination and temperature-dependent sex determination. [7] Higher temperatures during the gonadal sex differentiation stage (approximately 56–62 days post-hatching) significantly increase the likelihood of sex reversal. For example, one study found that the likelihood of females becoming pseudomales increased to 73% when reared at a higher temperature (28 °C) as opposed to a 14% sex-reversal rate when reared at an ambient temperature (22 °C). [3] [8] Furthermore, the offspring of the pseudomales reared at 28 °C had a sex-reversal rate of 94% despite being reared at 22 °C. [3] This high rate of females becoming pseudomales in the F2 generation was attributed to the offspring inheriting their Z chromosome from their pseudomale father and retaining paternal methylation patterns. [9]
A single SNP, Cyn_Z_6676874, allows certain females to become pseudomales. [10] Females with a thymine at this site can undergo sex reversal, but females with an adenine cannot. [10] There are also three known female-specific molecular markers that can be used to distinguish between the various tongue sole sexes. [5] Two SNPs, SNP_chr_8935925_C_T and SNP_chr_8936186_C_G, and one indel were found to enable accurate differentiation between males, females, and pseudomales. [5]
Gene expression in the gonads of males and females are different before sex determination, after sex determination, and after exposure to higher temperatures. [11] Prior to sex determination, males and females have differentially expressed genes (DEGs) related to muscle development. [11] Following sex determination, females have upregulated female-specific genes, figla and foxl2, and downregulation of the male-specific genes, dmrt1 and amh, under both ambient and high-temperature conditions. [11] When exposed to higher temperatures, males, females, and pseudomales have hundreds of DEGs in common that are mainly involved in biological processes and molecular functions. [11]
Pseudomale gonads have more similar gene expression patterns to males than females. [7] DEGs between males and pseudomales played a role in spermatogenesis and energy metabolism. [11] Whereas, DEGS between females and pseudomales were related to steroid hormones, helicase activity, sexual differentiation, and development. [11]