No abstract

Studies using genome-wide single nucleotide polymorphisms (SNPs) have become commonplace in genetics and genomics, due to advances in high-throughput sequencing technologies. Since the numbers of required SNPs and samples vary depending on each research goal, genotyping technologies with high flexibility in the number of SNPs/samples and high repeatability have been intensively investigated. For example, the ultrahigh-multiplexed amplicon sequencing, Ion AmpliSeq, has been used as a high-throughput genotyping method mainly for diagnostic purposes. Here, we designed a custom panel targeting 3,187 genome-wide SNPs of fugu, Takifugu rubripes , and applied it for genotyping farmed fugu to test its feasibility in aquaculture studies. We sequenced two libraries consisting of different pools of individuals ( n = 326 each) on the Illumina MiSeq sequencer. Consequently, over 99% target regions (3,178 SNPs) were amplified and 2,655 SNPs were available after filtering steps. Strong correlation was observed in the mean depth of coverage of each SNP between duplicate runs ( r = 0.993). Genetic analysis using these genotype data successfully detected the known population structure and the sex determining locus of fugu. These results show the method is superior in repeatability and flexibility, and suits genetic studies including molecular breeding, such as marker assisted and genomic selection.

Tuna (genus Thunnus), particularly Pacific bluefin tuna (T. orientalis; PBT), are commercially important fish in the aquaculture industry worldwide. The objective of this study was to investigate sexual dimorphism in the growth performance of aquaculture-produced PBT and develop techniques for its sex manipulation, for the first time in tuna. A comparison of the body size between sexes revealed that male-cultured PBTs were larger than females at harvest. We also confirmed that cyp19a1a (encoding a gonadal aromatase) expression increased specifically in the genotypic female PBT gonads during sex differentiation. This suggested that aromatase plays an important role in ovarian differentiation and that the suppression of aromatase activity may effectively induce masculinization in genotypic females. Therefore, we administered letrozole—an aromatase inhibitor (AI)—into sexually undifferentiated PBT through the oral route. AI administration resulted in a 100% sex reversal of genotypic females into phenotypic males at the molecular level. Our results provide the basis for future studies on the establishment of mono-sex male production technology in PBT, which would help improve the productivity of closed-cycle PBT aquaculture. Furthermore, this study offers important insights into the understanding of the sex-wise growth of tuna species in aquacultural settings, and developing sex manipulation techniques.

Testes of the tiger pufferfish Takifugu rubripes are a delicacy in Japan, and selective breeding for a male precocious phenotype, i.e., with early initiation of testes development, is desirable. However, it is unknown if precocious gonad development in this species is under genetic control. Here, we investigated genetic involvement in precociousness by using progeny tests with sires from two cultured populations, including a family line anecdotally known for its precociousness, and a wild population. Progeny derived from the “precocious” line consistently had greater testes weight than that from the other lines, even after accounting for effects of body weight, which indicates that precociousness is truly heritable. We also compared chronological changes in plasma steroid hormones between progenies sired by males from the precocious line and a wild population, and found that the precocious family line had higher levels of plasma estradiol-17β (E2) prior to the initiation of testicular development. Our findings suggest that selective breeding for testes precociousness in the tiger pufferfish is feasible, and that plasma E2 may be an indicator of this phenotype, which would allow for phenotype evaluation without the need to sacrifice specimens.