Dissecting the dynamic cellular transcriptional atlas of adult teleost testis development throughout the annual reproductive cycle.

Teleost testis development during the annual cycle involves dramatic changes in cellular compositions and molecular events. In this study, the testicular cells derived from adult black rockfish at distinct stages - regressed, regenerating and differentiating - were meticulously dissected via single-cell transcriptome sequencing. A continuous developmental trajectory of spermatogenic cells, from spermatogonia to spermatids, was delineated, elucidating the molecular events involved in spermatogenesis. Subsequently, the dynamic regulation of gene expression associated with spermatogonia proliferation and differentiation was observed across spermatogonia subgroups and developmental stages. A bioenergetic transition from glycolysis to mitochondrial respiration of spermatogonia during the annual developmental cycle was demonstrated, and a deeper level of heterogeneity and molecular characteristics was revealed by re-clustering analysis. Additionally, the developmental trajectory of Sertoli cells was delineated, alongside the divergence of Leydig cells and macrophages. Moreover, the interaction network between testicular micro-environment somatic cells and spermatogenic cells was established. Overall, our study provides detailed information on both germ and somatic cells within teleost testes during the annual reproductive cycle, which lays the foundation for spermatogenesis regulation and germplasm preservation of endangered species.

Deciphering dynamic interactions between spermatozoa and the ovarian microenvironment through integrated multi-omics approaches in viviparous Sebastes schlegelii.

The ovarian microenvironment plays a crucial role in ensuring the reproductive success of viviparous teleosts. However, the molecular mechanism underlying the interaction between spermatozoa and the ovarian microenvironment has remained elusive. This study aimed to contribute to a better understanding of this process in black rockfish (Sebastes schlegelii) using integrated multi-omics approaches. The results demonstrated significant upregulation of ovarian complement-related proteins and pattern recognition receptors, along with remodeling of glycans on the surface of spermatozoa at the early spermatozoa-storage stage (1 month after mating). As spermatozoa were stored over time, ovarian complement proteins were progressively repressed by tryptophan and hippurate, indicating a remarkable adaptation of spermatozoa to the ovarian microenvironment. Before fertilization, a notable upregulation of cellular junction proteins was observed. The study revealed that spermatozoa bind to ZPB2a protein through GSTM3 and that ZPB2a promotes spermatozoa survival and movement in a GSTM3-dependent manner. These findings shed light on a key mechanism that influences the dynamics of spermatozoa in the female reproductive tract, providing valuable insights into the molecular networks regulating spermatozoa adaptation and survival in species with internal fertilization.

PGE2 functions in ovoviviparous teleost black rockfish (Sebastes schlegelii): evolutionary status between parturition and ovulation†.

Fish have evolved various reproductive strategies including oviparity, viviparity, and ovoviviparity, which undoubtedly affect the survival of the whole species continuity. As the final step in reproduction, parturition in viviparous vertebrate and ovulation in oviparous teleost seem to share a similar mechanism, when prostaglandins (PGs) act as the trigger to launch the whole process. In the present study, ovoviviparous teleost black rockfish (Sebastes schlegelii) is employed as the research object. Intraperitoneal injection showed that PGE2 (500 µg/kg) could activate the delivery reactions in perinatal black rockfish. RNA-seq data of ovary in perinatal period revealed transcriptional change in cell junction, inflammation, and apoptosis, which is related to mammal parturition and teleost ovulation. Further results proved the positive correlation between ptger EP2 and previous mentioned pathways. Subsequent experiment proved that PGE2 was able to induce the ovulation and spawning in unfertilized individuals, which had a bilayer follicular structure compared to monolayer follicular in perinatal period black rockfish. Both unfertilized and perinatal ovary matrix could response to PGE2 stimulation. In conclusion, the function of PGE2 in activating both parturition and ovulation in a relatively different pathways conserved with viviparity or oviparity provided novel evidence of the evolutionary status of ovoviviparous vertebrates.

The residue of salinomycin in the muscles of olive flounder (Paralichthys olivaceus) and black rockfish (Sebastes Schlegeli) after oral administration analyzed by LC-Tandem-MS.

BACKGROUND: Salinomycin, an antibiotic, have potential as a veterinary drug for fish due to its anti-parasitic activity against several fish parasites. Thus the residual levels of salinomycin in muscles of two significant aquaculture species in Korea, olive flounder and black rockfish, were analyzed using HPLC-MS-MS. RESULTS: The proper method to analyze the residual salinomycin in fish muscles using LC-MS-MS was settled and the method was validated according to CODEX guidelines. The residues in three distinct groups for two fish species were analyzed using the matrix match calibration curves at points of five different times following oral administration. After oral administration, salinomycin rapidly breaks down in both olive flounder and black rockfish. After 7th days, the average residue in all groups of two fish spp. decreased below limit of quantitation (LOQ). CONCLUSION: Due to low residue levels in fish muscles, salinomycin may therefore be a treatment that is safe for both fish and humans. This result could contribute to establishment of MRL (minimal residual limit) for approval of salinomycin for use in aquaculture.

Pro-inflammatory cytokine IL1ß1 participates in promoting parturition related pathways in the ovoviviparous teleost black rockfish (Sebastes schlegelii).

Along with the evolution process, the reproductive strategies evolved including oviparity, viviparity and ovoviviparity, to fit the residential environment maximize the survival rate of the off spring. In mammals, the key to the initiation of parturition is the inflammatory response at the maternal-fetal interface. As a pro-inflammatory cytokine, interleukin 1 beta (IL1ß) plays an important role in the process of human parturition. While less is known about IL1ß1 in teleost parturition, identification of the functions of IL1ß1 in inducing the parturition, black rockfish, an ovoviviparity teleost, which provides over 60% nutrition supply for over 50 000 embryos though a placenta like structure during pregnant, was employed as the research model. In the present study, based on the gene cloning, we detected the expression pattern of both Il1b1 and its receptor perinatal period, as well as the localization to the ovary by in situ hybridization. The different expression genes in transcriptomic data of perinatal primary ovarian cells treated with the recombinant IL1ß1 (rIL1ß1) obtained by prokaryotic expression system were analyzed. Differentially expressed genes, functional enrichment and pathway analysis mainly included immune response, signal transduction and cell death. In summary, our research provides novel insights into the potential role of IL1ß1 in the parturition of ovoviviparity teleost.

Rapid turnover and evolution of sex-determining regions in Sebastes rockfishes.

Nature has evolved a wealth of sex determination (SD) mechanisms, driven by both genetic and environmental factors. Recent studies of SD in fishes have shown that not all taxa fit the classic paradigm of sex chromosome evolution and diverse SD methods can be found even among closely related species. Here, we apply a suite of genomic approaches to investigate sex-biased genomic variation in eight species of Sebastes rockfish found in the northeast Pacific Ocean. Using recently assembled chromosome-level rockfish genomes, we leverage published sequence data to identify disparate sex chromosomes and sex-biased loci in five species. We identify two putative male sex chromosomes in S. diaconus, a single putative sex chromosome in the sibling species S. carnatus and S. chrysomelas, and an unplaced sex determining contig in the sibling species S. miniatus and S. crocotulus. Our study provides evidence for disparate means of sex determination within a recently diverged set of species and sheds light on the diverse origins of sex determination mechanisms present in the animal kingdom.

Identification of reactive oxygen species modulator 1 (Romo 1) from black rockfish (Sebastes schlegelii) and deciphering its molecular characteristics, immune responses, oxidative stress modulation, and wound healing properties.

Reactive oxygen species modulator 1 (Romo1) is a mitochondrial inner membrane protein that induces mitochondrial reactive oxygen species (ROS) generation. In this study, we identified the Romo1 homolog from the black rockfish (Sebastes schlegelii), named it as SsRomo1, and characterized it at the molecular as well as functional levels. An open reading frame consisting of 240 bp was identified in the SsRomo1 complementary DNA (cDNA) sequence that encodes a 79 amino acid-long polypeptide with a molecular weight of 8,293 Da and a theoretical isoelectric point (pI) of 9.89. The in silico analysis revealed the characteristic features of SsRomo1, namely the presence of a transmembrane domain and the lack of a signal peptide. Homology analysis revealed that SsRomo1 exhibits the highest sequence identity with its fish counterparts (>93%) and shares a similar percentage of sequence identity with mammals (>92%). Additionally, it is closely clustered together with the fish clade in the constructed phylogenetic tree. The subcellular localization analysis confirmed its mitochondrial localization within the fathead minnow (FHM) cells. Under normal physiological conditions, the SsRomo1 mRNA is highly expressed in the rockfish ovary, followed by the blood and testis, indicating the abundance of mitochondria in these tissues. Furthermore, the significant upregulation of SsRomo1 in cells treated with lipopolysachharide (LPS), polyinosinic:polycytidylic acid, and Streptococcus iniae suggest that the increased ROS production is induced by SsRomo1 to eliminate pathogens during infections. Incidentally, we believe that this study is the first to determine the involvement of SsRomo1 in LPS-mediated nitric oxide (NO) production in RAW267.4 cells, based on their higher NO production as compared to that in the control. Moreover, overexpression of SsRomo1 enhanced the wound healing ability of FHM cells, indicating its high invasion and migration properties. We also determined the hydrogen peroxide-mediated cell viability of SsRomo1-overexpressed FHM cells and observed a significant reduction in viability, which is possibly due to increased ROS production. Collectively, our observations suggest that SsRomo1 plays an important role in oxidative stress modulation upon immune stimulation and in maintenance of tissue homeostasis in black rockfish.

Molecular characterization and expression patterns of glucocorticoid receptors in the viviparous black rockfish Sebastes schlegelii.

Glucocorticoid receptors (GRs) are ligand-activated transcription factors associated with anti-inflammation, stress, metabolism and gonadal development. In this study, two gr genes (gr1 and gr2) were cloned and analyzed from a viviparous teleost, black rockfish (Sebastes schlegelii). The phylogenetic analysis of GRs showed that GR1 and GR2 clustered into teleost GR1 and GR2 separately and differed from the GRs of tetrapods or basal ray-finned fishes. Black rockfish GRs possess four modular domains of the nuclear receptor superfamily: an N-terminal domain (NTD), a DNA-binding domain (DBD), a hinge region (HR) and a ligand-binding domain (LBD). Nine conserved amino acid inserts were found in the GR1 DBD, and the ligand cavity-related amino acids of GR1 and GR2 LBD were slightly different. Tissue distribution analysis revealed that grs was widely expressed in various tissues, while cyp11b was mainly expressed in the testis and head kidney. The cyp11b transcripts were localized in the interrenal glands of the head kidney, the main source of cortisol; grs transcripts were detected in oocytes, the follicle layer and the ovarian wall. Histologically, significant blood vessel dilation was observed in the fetal membrane during or after parturition of black rockfish. The highest levels of serum cortisol and ovarian cyp11b mRNA were detected in parturition. In addition, the relative expression level of gr1 was upregulated significantly after delivery, while the levels of gr2 showed no significant change. In addition, in vitro GC treatment inhibited the expression of il1b but significantly upregulated the transcription of il1r1. These data provide evidence that GRs are likely to work as anti-inflammatory factors by inhibiting the functions of pro-inflammatory factors in the parturition of black rockfish.

Genome-Wide Identification, Evolutionary Analysis, and Expression Patterns of Cathepsin Superfamily in Black Rockfish (Sebastes schlegelii) following Aeromonas salmonicida Infection.

Cathepsins are lysosomal cysteine proteases belonging to the papain family and play crucial roles in intracellular protein degradation/turnover, hormone maturation, antigen processing, and immune responses. In the present study, 18 cathepsins were systematically identified from the fish S. schlegelii genome. Phylogenetic analysis indicated that cathepsin superfamilies are categorized into eleven major clusters. Synteny and genome organization analysis revealed that whole-genome duplication led to the expansion of S. schlegelii cathepsins. Evolutionary rate analyses indicated that the lowest Ka/Ks ratios were observed in CTSBa (0.13) and CTSBb (0.14), and the highest Ka/Ks ratios were observed in CTSZa (1.97) and CTSZb (1.75). In addition, cathepsins were ubiquitously expressed in all examined tissues, with high expression levels observed in the gill, intestine, head kidney, and spleen. Additionally, most cathepsins were differentially expressed in the head kidney, gill, spleen, and liver following Aeromonas salmonicida infection, and their expression signatures showed tissue-specific and time-dependent patterns. Finally, protein-protein interaction network (PPI) analyses revealed that cathepsins are closely related to a few immune-related genes, such as interleukins, chemokines, and TLR genes. These results are expected to be valuable for comparative immunological studies and provide insights for further functional characterization of cathepsins in fish species.

Dominance of Scorpaena maderensis among scorpaenids of littoral rocky reefs in Corsica (NW Mediterranean): Further evidence of Mediterranean Sea warming.

The population of Scorpaenidae was investigated in the Nature Reserve of Scàndula (Corsica, France, NW Mediterranean). While absent from Corsica a few decades ago, the Madeira rockfish Scorpaena maderensis is now the most abundant scorpaenid species in shallow rocky reef areas, far outnumbering Scorpaena notata and Scorpaena porcus. Considered as a subtropical species of Atlantic origin, the northward progression of the thermophilic S. maderensis and its dominance in Corsican waters is further evidence of the ongoing warming of the Mediterranean Sea.

Effects of Cryopreservation on Sperm with Cryodiluent in Viviparous Black Rockfish (Sebastes schlegelii).

Black rockfish is an economically important fish in East Asia. Little mention has been paid to the sperm cryopreservation in black rockfish. In this study, the optimal cryodiluent was selected from 48 combinations by detecting various sperm parameters. Transcriptome and methylome analysis were further performed to explore the molecular mechanism of inevitable cryoinjuries. The results showed that cryopreservation had negative effects on the viability, DNA integrity, mitochondrial activity, total ATPase and LDH of sperm even with optimal cryodiluent (FBS + 15% Gly). Transcriptome and methylome analysis revealed that the expression of 179 genes and methylation of 1266 genes were affected by cryopreservation. These genes were enriched in GO terms of death, G-protein coupled receptor signaling pathway, response to external stimulus and KEGG pathways of phospholipase D signaling pathway and xenobiotic and carbohydrate metabolism pathways. The role of PIK3CA and CCNA2 were highlighted in the protein-protein interaction network, and the sperm quality-related imprinted gene mest was identified among the 7 overlapping genes between transcriptome and methylome. Overall, the cryodiluent for black rockfish sperm was optimized, providing a feasible method for cryopreservation. The transcriptome and methylome data further demonstrated the underlying molecular mechanisms of cryoinjuries, proving clues for improvement of cryopreservation method of black rockfish.

Expression Analysis of ZPB2a and Its Regulatory Role in Sperm-Binding in Viviparous Teleost Black Rockfish.

Black rockfish is a viviparous teleost whose sperm could be stored in the female ovary for five months. We previously proposed that zona pellucida (ZP) proteins of black rockfish play a similar sperm-binding role as in mammals. In this study, SsZPB2a and SsZPB2c were identified as the most similar genes with human ZPA, ZPB1 and ZPB2 by Blastp method. Immunohistochemistry showed that ovary-specific SsZPB2a was initially expressed in the cytoplasm of oocytes at stage III. Then it gradually transferred to the region close to the cell membrane and zona pellucida of oocytes at stage IV. The most obvious protein signal was observed at the zona pellucida region of oocytes at stage V. Furthermore, we found that the recombinant prokaryotic proteins rSsZPB2a and rSsZPB2c could bind with the posterior end of sperm head and rSsZPB2a was able to facilitate the sperm survival in vitro. After knocking down Sszpb2a in ovarian tissues cultivated in vitro, the expressions of sperm-specific genes were down-regulated (p < 0.05). These results illustrated the regulatory role of ZP protein to the sperm in viviparous teleost for the first time, which could advance our understanding about the biological function of ZP proteins in the teleost.

Comparative transcriptomic analysis of gonadal development and renewal in the ovoviviparous black rockfish (Sebastes schlegelii).

BACKGROUND: The black rockfish (Sebastes schlegelii) has an ovoviviparous reproductive pattern and long-term sperm storage, resulting in asynchronous gonadal development between the sexes. However, the comprehensive understanding of gonadal development in black rockfish has not yet been achieved. Here, we studied gonadal development and germ cell renewal using histology and RNA-seq. RESULTS: In this study, RNA-seq was performed on testes and ovaries to characterize key pathways and genes that are active during development and gamete maturation in black rockfish. Differentially expressed genes (DEGs) were identified and annotated in 4 comparisons (F_III vs. F_IV, F_IV vs. F_V, M_III vs. M_IV and M_IV vs. M_V). Based on analysis of DEGs enriched in the testis, 11 and 14 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were mapped to the M_III vs. M_IV group and the M_IV vs. M_V group, respectively. DEGs in ovarian development were also classified into 10 groups according to their biological functions. The expression patterns of the selected genes determined by qPCR were significantly correlated with the RNA-Seq results, supporting the reliability and accuracy of the RNA-Seq analysis. E2 levels showed down regulation from previtellogenesis to mature stage in female and T level showed down regulation from spermatogenesis to regressed stage in the male. CONCLUSIONS: The categories "intercellular interaction and cytoskeleton", "molecule amplification" and "repair in the cell cycle" were revealed to be crucial in testis development and spermatogenesis, as was the biosynthesis of a series of metabolites. Our results provide comprehensive insight into black rockfish gonadal development and provide a basis for further study of reproductive physiology and molecular biology in ovoviviparity teleosts.

Moving on up: Vertical distribution shifts in rocky reef fish species during climate-driven decline in dissolved oxygen from 1995 to 2009.

Anthropogenic climate change has resulted in warming temperatures and reduced oxygen concentrations in the global oceans. Much remains unknown on the impacts of reduced oxygen concentrations on the biology and distribution of marine fishes. In the Southern California Channel Islands, visual fish surveys were conducted frequently in a manned submersible at three rocky reefs between 1995 and 2009. This area is characterized by a steep bathymetric gradient, with the surveyed sites Anacapa Passage, Footprint and Piggy Bank corresponding to depths near 50, 150 and 300 m. Poisson models were developed for each fish species observed consistently in this network of rocky reefs to determine the impact of depth and year on fish peak distribution. The interaction of depth and year was significant in 23 fish types, with 19 of the modelled peak distributions shifting to a shallower depth over the surveyed time period. Across the 23 fish types, the peak distribution shoaled at an average rate of 8.7 m of vertical depth per decade. Many of the species included in the study, including California sheephead, copper rockfish and blue rockfish, are targeted by commercial and recreational fisheries. CalCOFI hydrographic samples are used to demonstrate significant declines in dissolved oxygen at stations near the survey sites which are forced by a combination of natural multidecadal oscillations and anthropogenic climate change. This study demonstrates in situ fish depth distribution shifts over a 15-year period concurrent with oxygen decline. Climate-driven distribution shifts in response to deoxygenation have important implications for fisheries management, including habitat reduction, habitat compression, novel trophic dynamics and reduced body condition. Continued efforts to predict the formation and severity of hypoxic zones and their impact on fisheries dynamics will be essential to guiding effective placement of protected areas and fisheries regulations.

Characterization of a Bacillus sp. KRF-7 isolated from the intestine of rockfish and effects of dietary supplementation with mannan oligosaccharide in rockfish aquaculture.

The purpose of this study was to characterize the bacteria isolated from rockfish intestines and to investigate the effects of feed supplementation in rockfish aquaculture. Bacillus sp. KRF-7 isolated from the intestine of rockfish (Sebastes schlegelii) was demonstrated to be safe based on in vitro tests confirming the absence of hemolysis, cytotoxicity, and genes with toxigenic potential. In a feeding trial, providing a supplemental diet of 1 × 108 CFU g-1Bacillus sp. KRF-7 was observed to positively alter the weight gain, specific growth rate, feed conversion ratio, and protein efficiency ratio of juvenile rockfish. KRF-7 supplementation showed positive regulation of nonspecific immune parameters, such as superoxide dismutase, lysozyme activity, and myeloperoxidase activity. This analysis also revealed a change in the composition of the intestinal microbiota at the phylum level from Proteobacteria to Firmicutes. In both the kidney and spleen, the expression levels of IL-10, NF-κB, and B cell activating factors in the KRF-7-supplemented group were significantly increased compared to those in the control group. Therefore, this study verified the safety of KRF-7 isolated from the intestine of rockfish and suggests that dietary supplementation with KRF-7 enhances the growth performance of rockfish and has beneficial effects on the regulation of the intestinal microbiota and immune response.

Identification of tissue-specific expression of CXCL14 in black rockfish (Sebastes schlegelii).

CXCL14 is a chemokine which is orthologous in mammals and fish. CXCL14 has a functional role in different organs, with immunomodulatory functions in mammals, but its expression and function in fish is not well known. Moreover, it shows no effects related to immunity in the central nervous system or the reproductive tract in diverse species. Black rockfish (Sebastes schlegelii) is an economically important fish in Asian countries, whose CXCL14 expression pattern is yet to be understood. In this study, the homology of the CXCL14 amino acid sequence in S. schlegelii was compared with that in other species, including fish. Moreover, in situ hybridization analysis revealed that it was highly expressed in the brain and ovary of S. schlegelii. Taken together, we identified for the first time, the cell-specific expression of CXCL14 in S. schlegelii.

Effects of sulfated polysaccharides isolated from Codium fragile on inflammatory cytokine gene expression and Edwardsiella tarda infection in rockfish, Sebastes schlegelii.

Sulfated polysaccharides (SPs) derived from Codium fragile (sponge seaweed) can regulate cytokine expression in mammalian macrophages, NK cell lines and olive flounder head kidney primary cells in vitro. In this study, we found that SPs from C. fragile exhibited anti-bacterial activities against fish pathogenic bacteria including Streptococcus parauberis, Lactococcus garvieae, Aeromonas salmonicida and Edwardsiella tarda at a minimum inhibitory concentration of 2 mg/mL, but not against S. iniae or Vibrio anguillarum. Immunostimulatory effects of SPs from C. fragile on rockfish (Sebastes schlegelii) were evaluated by analyzing mRNA expression levels of inflammatory cytokines (interleukin (IL)-1ß, IL-8, IL-6 and tumor necrosis factor (TNF)-α) and anti-inflammatory cytokines (IL-10) both in vitro and in vivo. Results revealed that expression levels of all genes tested were upregulated in rockfish head kidney and spleen cells by SPs from C. fragile in a dose/time-dependent manner in vitro. By contrast, expression levels of these genes were significantly (p < 0.05) downregulated in the head kidney and spleen of rockfish in vivo at 1 and 3 days post intraperitoneal injection of SPs from C. fragile. In the liver, these genes were downregulated on day 1, but upregulated on day 3. Treatment with SPs downregulated the expression of these genes in spleen, but upregulated IL-10 gene expression in the intestine and liver. Meanwhile, when fish were fed with crude SPs for 4 weeks and challenged with E. tarda, infected fish started to die starting from 2 days after immune challenge. The cumulative mortality of the 0.1% group was significantly lower (p < 0.05) than that of the control group without feeding with SPs. Expression levels of IL-1ß and IL-6 genes were significantly (p < 0.05) upregulated in head kidney of the 0.5% group on day 1 while IL-1ß gene expression was downregulated on day 3 in the liver. These results indicate that SPs from C. fragile can regulate the immune gene expression in rockfish and that a diet containing 0.1% crude SPs can reduce the mortality of rockfish caused by E. tarda infection.

Changes in the Hepatic Expression of Three Vitellogenin Subtype Genes During Ovarian Development in Female White-Edged Rockfish (Sebastes taczanowskii).

Viviparous fish, including white-edged rockfish (Sebastes taczanowskii), accumulate substantial yolk mass in the oocytes; however, the details of the molecular mechanisms underlying yolk formation are not yet fully understood, especially concerning multiplicity in the yolk precursor vitellogenin (Vtg). The present study aimed to reveal the hepatic transcriptional profiles of multiple vtg gene transcripts (vtgAa, vtgAb, vtgC) during the reproductive cycle in captive female white-edged rockfish reared in an aquarium under natural photo-thermal conditions. The serum estradiol-17ß concentration and the hepatic transcript levels of all vtg subtypes increased with the progress of vitellogenesis; both levels decreased at the beginning of oocyte maturation and remained low during the gestation period. Considering the similarity in the transcriptional profiles of vtg subtypes between Sebastes and Oncorhynchus, along with the differences between Sebastes and Morone, it is suggested that the transcription patterns of multiple vtg genes relate to neither their reproductive modes (viviparity versus oviparity) nor to teleost phylogeny.

Characterization of CYP11A1 and its potential role in sex asynchronous gonadal development of viviparous black rockfish Sebastes schlegelii (Sebastidae).

Mitochondrial cytochrome P450 side-chain cleavage (P450scc), encoded by the cyp11a1 gene, initiates the first step of steroid biosynthesis. In this study, a 1554-bp open reading frame (ORF) of black rockfish (Sebastes schlegelii) cyp11a1 was cloned. The cyp11a1 gene is located on chromosome 5 and has 9 exons. The ORF encodes a putative precursor protein of 517 amino acids, and the predicted cleavable mitochondrial targeting peptide is located at amino acids 1-39. P450scc shares homology with other teleosts and tetrapods, which have relatively conserved binding regions with heme, cholesterol and adrenodoxin. Tissue distribution analysis revealed that the highest expression levels of cyp11a1 were detected in mature gonads and head kidney but that low levels were detected in gestational/regressed ovaries, regressed testes and other tissues. Immunostaining of P450scc was observed in testicular Leydig cells, ovarian theca cells, interrenal glands of head kidney, pituitary and multiple regions of brain. Particularly, two kinds of fish-specific P450scc-positive cells, including coronet cells of brain saccus vasculosus and hypophyseal somatolactin cells, were identified in black rockfish. Our results provide novel evidence for the potential role played by P450scc in reproduction behavior by mediating steroidogenesis in viviparous teleost.

Systemic phaeohyphomycosis in splitnose rockfish (Sebastes diploproa) caused by Devriesia sp.

A novel pathogen was documented after two wild-caught, juvenile, splitnose rockfish presented with buphthalmia, grey corneal endothelial plaques and evidence of uveitis. Cytologic evaluation of ocular contents revealed fungal hyphae. Histologic evaluation identified multiple fungal granulomas and granulomatous inflammation in the globes, periocular tissue and heart. Fungi were slender, hyphenated and branched at angles, had parallel cell walls and had brown pigmentation in haematoxylin- and eosin-stained sections. Both fish were diagnosed with phaeohyphomycosis. Culture with nuclear ribosomal RNA internal transcribed spacer (ITS) segment identification further classified the fungus as Devriesia sp., which has not been previously documented as a cause of disease in animals.