Identification of Candidate Genes Associated With Hypoxia Tolerance in Trachinotus blochii Using Bulked Segregant Analysis and RNA-Seq

Golden Pompano (Trachinotus blochii) has rapidly developed into the one of the main valuable fish species in Chinese marine aquaculture. Due to its rapid growth, active metabolism, and high oxygen consumption, hypoxia will increase its mortality and cause serious economic losses. We constructed two experimental groups of fish with different degrees of tolerance to hypoxia, used BSR-Seq analysis based on genome and genetic linkage groups to locate SNPs and genes that were related to the differences in hypoxia tolerance. The results showed that hypoxia tolerance SNPs of golden pompano may be jointly determined by multiple linkage groups, especially linkage groups 18 and 22. There were 768 and 348 candidate genes located in the candidate regions of the brain and liver, respectively. These genes were mainly involved in anaerobic energy metabolism, stress response, immune response, waste discharge, and cell death. The prostaglandin-endoperoxide synthase 2 (PTGS2) on LG8, which is involved in the metabolism of arachidonic acid, has a G/A nonsynonymous mutation at position 20641628, and the encoded amino acid was changed from hydrophobic aspartic acid to asparaginate. The specific pathway of the RIG-I-like receptor signaling pathway in the liver may mediate the metabolic system and the immune system, linking glucose metabolism with immune regulation. The death of the hypoxia-intolerant group may be due to the accumulation of lactic acid caused by the activation of anaerobic glycolysis during the early stage of hypoxia stress, and the activation of type I interferon was inhibited, which resulted in decreased immunity. Among the genes involved in the RIG-I-like receptor signaling pathway, the CYLD Lysine 63 Deubiquitinase (CYLD) located on LG16 had a G/T nonsynonymous mutation at position 13629651, and the encoded amino acid was changed from alanine acid to valine. The interferon induced with helicase C domain 1 (Ifih1) located on LG18 has a G/C nonsynonymous mutation at position 16153700, and the encoded hydrophilic glycine was changed to hydrophobic alanine. Our findings suggest these SNPs may assist in the molecular breeding of hypoxia-tolerant golden pompano, and speculate that the balance of glucose and lipid metabolism plays a key role in Trachinotus blochii under acute hypoxia.

Transcriptome Analysis of Gills Provides Insights Into Translation Changes Under Hypoxic Stress and Reoxygenation in Golden Pompano, Trachinotus ovatus (Linnaeus 1758)

Golden pompano (Trachinotus ovatus) is one of the most economically critical marine fish in South China. Low oxygen stress has resulted in substantial economic losses to the aquaculture of T. ovatus. However, the molecular responses of fish gills to hypoxia challenge remain unclear. To understand the mechanism underlying adaption to hypoxia, we analyzed the transcriptome of T. ovatus gills in response to hypoxic stress in the normal oxygen group, hypoxic group, and hypoxia treatment after oxygen recovery group. This study found that hypoxia for 8 h was the critical time of hypoxic stress and corresponded to the largest number of differentially expressed genes. After hypoxic stress, genes for chemokines, chemokine receptors, interleukins, complement factors, and other cytokines were significantly downregulated, which may be why fish are vulnerable to pathogen infection in a hypoxic environment. According to a Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, many downregulated genes were significantly enriched in the steroid biosynthesis, focal adhesion, and the extracellular matrix (ECM)-receptor interaction signal pathways, which affected cell signal transduction, adhesion, and apoptosis. Compared with the hypoxic group, the amounts of upregulated genes related to phagocytosis and protein degradation were upregulated in the dissolved oxygen recovery group. These results indicated that after the recovery of dissolved oxygen, the fish body repaired the stress-induced damage by rapidly removing misfolded proteins. These findings provide a better understanding of the hypoxia response mechanism of fish and represent a useful resource for the genetic breeding of T. ovatus.

Changes in Growth Performance, Nutrient Metabolism, Antioxidant Defense and Immune Response After Fishmeal Was Replaced by Low-Gossypol Cottonseed Meal in Golden Pompano (Trachinotus ovatus)

The present study was designed to elucidate the changing pattern of growth performance, nutrient metabolism, antioxidant defense, and immune response after fishmeal (FM) was replaced by cottonseed meal (CSM). Four isonitrogenous and isolipidic experimental diets (42.5% crude protein, 14.0% crude lipid) were formulated to replace 0% (CSM0 diet), 20% (CSM20 diet), 40% (CSM40 diet), and 60% (CSM60 diet) of the FM protein with CSM. Juvenile golden pompano (Trachinotus ovatus) with an initial body weight of 24.8 ± 0.02 g were fed each diet for 6 weeks. The results showed that compared with the control diet, significant differences were not found in the weight gain ratio, specific growth performance, and apparent digestibility of dry matter and protein after 20% FM was replaced by CSM. Higher CSM replacement markedly decreased the growth performance (P < 0.05). Moreover, the CSM20 and CSM40 diets did not depress the feed efficiency ratio and protein efficiency ratio compared with the CSM0 diet. Further exploring the physio-biochemical and molecular responses, the present study also showed that dietary 20–40% CSM replacement had no significant effect on altering the whole body, plasma nutrient contents, free amino acids, or whole-body fatty acid contents. Additionally, the CSM20 diet did not change the mRNA and protein phosphorylation levels of the key enzymes and regulators involved in target of rapamycin (TOR) and amino acid response (AAR) signaling, nutrient metabolism, immune response, and antioxidant defense. Higher CSM inclusion significantly varied the TOR and AAR signaling response, nutrient metabolism, immune response, and antioxidant defense. Based on the results, CSM could successfully replace 20% FM in the formulated diets for golden pompano. This study proposes the optimal inclusion level of CSM in the diet, which has an important significance in optimizing aquafeeds formulation.

Turnover of sex defining mutation provides an insight into evolution of sex chromosomes in the golden pompano (Trachinotus ovatus)

The golden pompano (Trachinotus ovatus) is a marine fish species in the family Carangidae. We constructed a chromosome-level genome assembly of a male golden pompano. QTL-mapping and GWAS analysis showed that this species has a ZZ/ZW sex determination system and a sex defining SNP (Chr16:18219150:G/A), located on the splice donor site (GT-AG) of the first intron of Hsd17b1, was exclusively associated with the phenotypic sex. The W-linked coding sequences of Hsd17b1 were conserved across vertebrates, while Z-linked coding sequences introduced extra 64 bases and were malfunctional. The golden pompano and the greater amberjack (Seriola dumerili), divergent in 57 million years ago in the same family, share the same features of sex determination, including the same sex determining gene, malfunctional Z-linked haplotypes, undifferentiated sex chromosomes except that the sex defining SNPs are different. In simulation analysis, turnover of sex determining mutation, single mutation dominating sex determination and undifferentiated sex chromosomes were also observed. We proposed a hypothesis that W-linked haplotypes of the sex determining gene of Hsd17b1 were under purifying selection, Z-linked haplotypes may evolve near neutrally, recurrent and directional transformations from W-linked haplotypes to Z-linked haplotypes caused by inactivating mutations, relatively strong forces of drift and recombination comprehensively contributed to turnover of sex defining mutation and undifferentiation of sex chromosome. We also established zebrafish mutants and homozygous mutants were “all male”, which indirectly supported this hypothesis.