Whole genome sequencing of a snailfish from the Yap Trench (~7,000 m) clarifies the molecular mechanisms underlying adaptation to the deep sea.

Hadal environments (depths below 6,000 m) are characterized by extremely high hydrostatic pressures, low temperatures, a scarce food supply, and little light. The evolutionary adaptations that allow vertebrates to survive in this extreme environment are poorly understood. Here, we constructed a high-quality reference genome for Yap hadal snailfish (YHS), which was captured at a depth of ~7,000 m in the Yap Trench. The final YHS genome assembly was 731.75 Mb, with a contig N50 of 0.75 Mb and a scaffold N50 of 1.26 Mb. We predicted 24,329 protein-coding genes in the YHS genome, and 24,265 of these genes were successfully functionally annotated. Phylogenetic analyses suggested that YHS diverged from a Mariana Trench snailfish approximately 0.92 million years ago. Many genes associated with DNA repair show evidence of positive selection and have expanded copy numbers in the YHS genome, possibly helping to maintain the integrity of DNA under increased hydrostatic pressure. The levels of trimethylamine N-oxide (TMAO), a potent protein stabilizer, are much higher in the muscles of YHS than in those of shallow-water fish. This difference is perhaps due to the five copies of the TMAO-generating enzyme flavin-containing monooxygenase-3 gene (fmo3) in the YHS genome and the abundance of trimethylamine (TMA)-generating bacteria in the YHS gut. Thus, the high TMAO content might help YHS adapt to high hydrostatic pressure by improving protein stability. Additionally, the evolutionary features of the YHS genes encoding sensory-related proteins are consistent with the scarce food supply and darkness in the hadal environments. These results clarify the molecular mechanisms underlying the adaptation of hadal organisms to the deep-sea environment and provide valuable genomic resources for in-depth investigations of hadal biology.

Extreme-Phenotype Genome-Wide Association Analysis for Growth Traits in Spotted Sea Bass (Lateolabrax maculatus) Using Whole-Genome Resequencing.

Spotted sea bass (Lateolabrax maculatus) is an important marine economic fish in China, ranking third in annual production among marine fish. However, a declined growth rate caused by germplasm degradation has severely increased production costs and reduced economic benefits. There is an urgent need to develop the fast-growing varieties of L. maculatus and elucidate the genetic mechanisms underlying growth traits. Here, whole-genome resequencing technology combined with extreme phenotype genome-wide association analysis (XP-GWAS) was used to identify candidate markers and genes associated with growth traits in L. maculatus. Two groups of L. maculatus, consisting of 100 fast-growing and 100 slow-growing individuals with significant differences in body weight, body length, and carcass weight, underwent whole-genome resequencing. A total of 4,528,936 high-quality single nucleotide polymorphisms (SNPs) were used for XP-GWAS. These SNPs were evenly distributed across all chromosomes without large gaps, and the average distance between SNPs was only 175.8 bp. XP-GWAS based on the Bayesian-information and Linkage-disequilibrium Iteratively Nested Keyway (Blink) and Fixed and random model Circulating Probability Unification (FarmCPU) identified 50 growth-related markers, of which 17 were related to body length, 19 to body weight, and 23 to carcass weight. The highest phenotypic variance explained (PVE) reached 15.82%. Furthermore, significant differences were observed in body weight, body length, and carcass weight among individuals with different genotypes. For example, there were highly significant differences in body weight among individuals with different genotypes for four SNPs located on chromosome 16: chr16:13133726, chr16:13209537, chr16:14468078, and chr16:18537358. Additionally, 47 growth-associated genes were annotated. These genes are mainly related to the metabolism of energy, glucose, and lipids and the development of musculoskeletal and nervous systems, which may regulate the growth of L. maculatus. Our study identified growth-related markers and candidate genes, which will help to develop the fast-growing varieties of L. maculatus through marker-assisted breeding and elucidate the genetic mechanisms underlying the growth traits.

Cloning and functional analysis of glutathione peroxidase gene in red swamp crayfish Procambarus clarkii.

Glutathione peroxidases (GPxs) are key enzymes in the antioxidant defense systems of living organisms, including crustaceans. The red swamp crayfish Procambarus clarkii is the most commonly farmed freshwater crayfish in Chinese inland nowadays due to its commercial value. However, high stocking density has resulted in adverse effects in growth performance and health. To investigate the function of GPxs in immune defense of the crayfish, we cloned and characterized a full length GPx (PcGPx) from P. clarkii by a reverse-transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). The 931 bp PcGPx cDNA contains a 38 bp 5'-untranslated region (UTR), a 519 bp coding sequence (CDS) and a 375 bp 3'-UTR with a selenocysteine insertion sequence (SECIS). The PcGPx was predicted to encode 172 amino acids, and its putative molecular mass was 20.9 kDa with a pI of 4.37. A selenocysteine (Sec) encoded by the unusual stop codon, TGA, was in the protein coding region. Phylogenetic analysis showed that PcGPx clustered with the GPxs from the penaeid shrimp Metapenaeus ensis and Caenorhabditis elegans, sharing much higher similarity with vertebrate GPx1 and GPx2 than with GPx3 and GPx5. Quantitative RT-PCR revealed that PcGPx was extremely highly expressed in ovary and early embryos. In addition, the levels of PcGPx mRNA and reactive oxygen species (ROS) significantly increased after challenge with gram-negative Vibrio harveyi, gram-positive Staphyloccocus aureus or white spot syndrome virus (WSSV). These results suggest that PcGPx may play important roles not only in immune defense, but also in oogenesis in the crayfish.

Comparative secretome analysis reveals cross-talk between type III secretion system and flagella assembly in Pseudomonas plecoglossicida.

The Gram-negative bacterium Pseudomonas plecoglossicida has caused visceral granulomas disease in several farmed fish species, including large yellow croaker (Larimichthys crocea), which results in severe economic losses. Type III secretion systems (T3SS) are protein secretion and translocation nanomachines widely employed by many Gram-negative bacterial pathogens for infection and pathogenicity. However, the exact role of T3SS in the pathogenesis of P. plecoglossicida infection is still unclear. In this study, a T3SS translocators deletion strain (△popBD) of P. plecoglossicida was constructed to investigate the function of T3SS. Then comparative secretome analysis of the P. plecoglossicida wild-type (WT) and △popBD mutant strains was conducted by label-free quantitation (LFQ) mass spectrometry. The results show that knockout of T3SS translocators popB and popD has an adverse effect on the effector protein ExoU secretion, flagella assembly, and biofilm formation. Further experimental validations also confirmed that popB-popD deletion could affect the P. plecoglossicida flagella morphology/formation, adherence, mobility, and biofilm formation. These data indicate that a cross-talk exists between the P. plecoglossicida T3SS and the flagella system. Our results, therefore, will facilitate the further under-standing of the pathogenic mechanisms leading to visceral granulomas disease caused by P. plecoglossicida.

The genome of a hadal sea cucumber reveals novel adaptive strategies to deep-sea environments.

How organisms cope with coldness and high pressure in the hadal zone remains poorly understood. Here, we sequenced and assembled the genome of hadal sea cucumber Paelopatides sp. Yap with high quality and explored its potential mechanisms for deep-sea adaptation. First, the expansion of ACOX1 for rate-limiting enzyme in the DHA synthesis pathway, increased DHA content in the phospholipid bilayer, and positive selection of EPT1 may maintain cell membrane fluidity. Second, three genes for translation initiation factors and two for ribosomal proteins underwent expansion, and three ribosomal protein genes were positively selected, which may ameliorate the protein synthesis inhibition or ribosome dissociation in the hadal zone. Third, expansion and positive selection of genes associated with stalled replication fork recovery and DNA repair suggest improvements in DNA protection. This is the first genome sequence of a hadal invertebrate. Our results provide insights into the genetic adaptations used by invertebrate in deep oceans.

Transcriptome analysis revealed multiple immune processes and energy metabolism pathways involved in the defense response of the large yellow croaker Larimichthys crocea against Pseudomonas plecoglossicida.

The large yellow croaker (Larimichthys crocea) aquaculture industry is suffering substantial financial losses caused by visceral white nodules disease resulting from Pseudomonas plecoglossicida infection. However, how L. crocea responds to P. plecoglossicida infection remains largely unknown. Here, we characterized the changes in the mRNA profile in the spleen of L. crocea upon P. plecoglossicida infection and explored the related defensive strategies. Sample clustering analysis and qRT-PCR indicated that P. plecoglossicida induced profound and reproducible transcriptome remodeling in the L. crocea spleen. Many innate immune-related genes, such as IL-17 signaling molecules, chemokines and chemokine receptors, complement components, TLR5 signaling molecules, and antimicrobial peptide hepcidins (Hamps), were upregulated by P. plecoglossicida and may play important roles in the L. crocea defense against P. plecoglossicida. The antibacterial activity of Hamp2-5 against P. plecoglossicida was further confirmed by using synthetic mature peptide of Hamp2-5. Additionally, significant enrichment of "Glycolysis/Gluconeogenesis", "Citrate cycle" and "Oxidative phosphorylation" pathways and a significant upregulation of all 6 rate-limiting enzyme genes (HK1, PFK, PKM, CS, IDH2, DLST) in the Glycolysis and Citrate cycle pathways in P. plecoglossicida-infected fish suggested that ATP synthesis may be accelerated to ensure energy supply in response to pathogenic infection. Altogether, our results not only identified the key immune-related genes and immune pathways that participated in the defense response of L. crocea against P. plecoglossicida, but also revealed a novel defensive strategy involving ATP synthesis in this species.

Whole Genome Incorporation and Epigenetic Stability in a Newly Synthetic Allopolyploid of Gynogenetic Gibel Carp.

Allopolyploidization plays an important role in speciation, and some natural or synthetic allopolyploid fishes have been extensively applied to aquaculture. Although genetic and epigenetic inheritance and variation associated with plant allopolyploids have been well documented, the relative research in allopolyploid animals is scarce. In this study, the genome constitution and DNA methylation inheritance in a newly synthetic allopolyploid of gynogenetic gibel carp were analyzed. The incorporation of a whole genome of paternal common carp sperm in the allopolyploid was confirmed by genomic in situ hybridization, chromosome localization of 45S rDNAs, and sequence comparison. Pooled sample-based methylation sensitive amplified polymorphism (MSAP) revealed that an overwhelming majority (98.82%) of cytosine methylation patterns in the allopolyploid were inherited from its parents of hexaploid gibel carp clone D and common carp. Compared to its parents, 11 DNA fragments in the allopolyploid were proved to be caused by interindividual variation, recombination, deletion, and mutation through individual sample-based MSAP and sequencing. Contrast to the rapid and remarkable epigenetic changes in most of analyzed neopolyploids, no cytosine methylation variation was detected in the gynogenetic allopolyploid. Therefore, the newly synthetic allopolyploid of gynogenetic gibel carp combined genomes from its parents and maintained genetic and epigenetic stability after its formation and subsequently seven successive gynogenetic generations. Our current results provide a paradigm for recurrent polyploidy consequences in the gynogenetic allopolyploid animals.