Streptococcus parauberis is a major pathogen responsible for streptococcosis in both marine and freshwater fish species, causing substantial economic losses in aquaculture. The increasing prevalence of multidrug resistance has highlighted the urgent need for alternative disease control strategies. Interference with bacterial quorum sensing (QS) systems represents a promising approach. This study aimed to identify and biochemically characterize an Rgg-family transcriptional regulator and evaluate its potential as a target for quorum sensing-related regulatory interference in vitro. We hypothesized that this Rgg regulator may function as a quorum sensing-associated transcription factor capable of promoter binding and modulation by small molecules. Bioinformatic analyses were used to identify the rgg gene encoding an Rgg-family transcriptional regulator and predict its structural features. The gene was cloned, heterologously expressed, and purified. Promoter binding activity was examined using electrophoretic mobility shift assay (EMSA), and key amino acid residues were identified through site-directed mutagenesis. The inhibitory effect of the cyclic peptide cyclosporin A (CsA) on Rgg-promoter binding was further assessed. The rgg gene (864 bp) encoding a 287-amino-acid protein (34.1 kDa) was successfully identified and expressed. Purified Rgg specifically bound to its own promoter region in a concentration-dependent manner. Mutations at conserved arginine residues R12 and R15 within the helix-turn-helix DNA-binding domain abolished promoter binding activity. Furthermore, CsA disturbed Rgg-promoter binding in a dose-dependent manner. This study provides the first in vitro characterization of an Rgg-family transcriptional regulator in fish-derived S. parauberis. The findings expand current understanding of Rgg-family regulators potentially associated with quorum sensing in aquatic streptococci and provide a preliminary basis for further investigation of quorum sensing-related regulatory interference strategies for controlling streptococcal diseases in aquaculture.

We are interested in the mechanism and therapeutic strategy for the alteration of retinal neuronal integrity in diabetic retinopathy (DR) and other hypoxic retinal diseases. In this article, we will discuss our work in exploring the relationship between vascular endothelial growth factor (VEGF) and brain-derived neurotrophic factor (BDNF) in the maintenance of major retinal supporting cells, Müller cell (MC), and in investigating the potential role and mechanisms of BDNF-mediated MC viability through its signaling cascade under diabetic and hypoxic conditions. While our data suggest that BDNF is a positive regulator of MC viability through the classical survival and proliferation pathways under diabetic/hypoxic conditions, the biological significance of BDNF-mediated MC viability in diabetes/hypoxia remains unclear, which is a major challenge in designing BDNF-mediated neuroprotective strategy for DR and hypoxic retinal disorders.

Tight junction protein-1 and -2 (Tjp1/ZO-1 and Tjp2/ZO-2) function as scaffold proteins within the tight junction complexes of the blood-retinal barrier (BRB). Although the breakdown of the BRB is implicated in retinopathies, the contribution of ZO-1/2 in the pathogenesis of retinopathies is unknown. To understand their role, we generated RPE-specific conditional ZO-1/2 single KOs (T1KO/T2KO) and ZO-1/2 double knockout (DKO) mice. While T1KO and T2KO did not exhibit overt retinal phenotypes, DKO demonstrated a strong retinal phenotype from 1-month post-KO induction. This includes the loss of RPE integrity and retinal thinning. Furthermore, RPE in DKO re-entered the cell cycle with upregulated YAP. At 12 months, we observed severe structural and functional retinal deterioration. In response to laser-induced damage, RPE displayed persistent hyper-proliferation, delayed wound repair, and the up-regulation of YAP. These studies confirmed the critical role of ZO-1/2 in maintaining an intact BRB and revealed a role of ZO-1/2 in wound healing.

Unlike birds and mammals, many teleosts have homomorphic sex chromosomes, and changes in the chromosome carrying the sex-determining locus, termed "turnovers", are common. Recent turnovers allow studies of several interesting questions. One question is whether the new sex-determining regions evolve to become completely non-recombining, and if so, how and why. Another is whether (as predicted) evolutionary changes that benefit one sex accumulate in the newly sex-linked region. To study these questions, we analyzed the genome sequences of two seahorse species of the Syngnathidae, a fish group in which many species evolved a unique structure, the male brood pouch. We find that both seahorse species have XY sex chromosome systems, but their sex chromosome pairs are not homologs, implying that at least one turnover event has occurred. The Y-linked regions occupy 63.9% and 95.1% of the entire sex chromosome of the two species and do not exhibit extensive sequence divergence with their X-linked homologs. We find evidence for occasional recombination between the extant sex chromosomes that may account for their homomorphism. We argue that these Y-linked regions did not evolve by recombination suppression after the turnover, but by the ancestral nature of the low crossover rates in these chromosome regions. With such an ancestral crossover landscape, a turnover can instantly create an extensive Y-linked region. Finally, we test for adaptive evolution of male pouch-related genes after they became Y-linked in the seahorse.

Vascular endothelial growth factor (VEGF or VEGF-A), a major pathogenic factor for diabetic and hypoxic blood–retina barrier (BRB) diseases, has been shown to act as a direct functional regulator for neurons in the peripheral and central nerve systems. To determine if VEGF plays a direct role in regulating retinal neuronal function, we established specific experimental procedures and examined the effect of recombinant VEGF (rVEGF) on photoreceptor function with electroretinography (ERG) in mice. In our case, rVEGF caused a significant reduction of scotopic ERG a-wave and b-wave amplitudes and photopic ERG b-wave amplitudes in a dose-dependent manner in dark-adapted wild-type (WT) mice, shortly after the intravitreal delivery of rVEGF in dark. However, the effect of rVEGF on photoreceptor function was nullified in adult Akita diabetic mice. Our data strongly suggest that VEGF is a direct regulator of photoreceptor function and VEGF upregulation contributes significantly to the diabetes-induced reduction of photoreceptor function. In this chapter, we will discuss the relevant background, key experimental procedures and results, and clinical significance of our work.

In this study, we determined the complete mitochondrial genome of the Picasso panda clownfish for the first time.The length of the whole mitogenome is 16,652 bp long and contained 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes, 2 non-coding regions and 1 control region(D-lop).The mitochondrial genome of the Picasso panda clownfish consisted of A = 29.19%,T = 25.49%,G = 15.93% and C = 29.40%(AT-skew = 0.067 and GC-skew = -0.297).Molecular phylogenetic analysis suggested that Amphiprion ocellaris, Amphiprion percula, Premnas biaculeatus and the Picasso panda clownfish clustered in one branch of the phylogenetic tree.Picasso panda clownfishand and A. ocellaris are most closely related.

Since the release of the movie Finding Nemo, clownfish have been known and loved by many people. Clownfish have a very vivid skin color and extreme ornamental value. However, there are few species of clownfish, which greatly limits the breeding of new varieties. In this study, the Atmosphere and room temperature plasma (ARTP) method was used to treat clownfish-fertilized eggs and successfully screened out mutants that exhibited a skin color change from black to red in their offspring. To elucidate the molecular mechanism underlying this color mutation, more than 17,000 methylated peaks were obtained via m6A-specific methylated RNA immunoprecipitation with next-generation sequencing (MeRIP-seq). These modification sites were mainly distributed around stop codons, and the number of corresponding genes exceeded 10,000. Further RNA sequencing (RNA-seq) of clownfish before and after the mutation was performed identified 883 genes that exhibited significant differences between the two samples, among which 152 interaction genes showed significant differences in the m6A methylated level and gene expression level. The results of a functional analysis showed that the phosphatidylinositol-3-kinase (PI3K)-serine/threoninekinase (Akt) pathway and its related signal pathways may play an important role in skin color change. In particular, genes such as mitogen activated protein kinase kinase 1 ( MAP2K1) , insulin―like growth factor―l ( IGF1 ), and fibroblast growth factor 1 ( FGF1 ) may play key roles in the accumulation of melanin in clownfish, and the homeobox-protein-encoding empty spiracles homeobox 2 ( EMX2 ) and mesenchyme homeobox ( MEOX2 ) genes may be important for determining the regions of accumulation of this skin pigment. Our results provide a new reference for the genetic breeding of clownfish and lay a foundation for further understanding the molecular mechanism underlying body color changes in clownfish.

Abstract Background Betanodaviruses, members of the Nodaviridae family, are the causative agents of viral nervous necrosis in fish, resulting in great economic losses worldwide. Methods In this study, we isolated a virus strain named seahorse nervous necrosis virus (SHNNV) from cultured big-belly seahorses Hippocampus abdominalis in Xiamen city, Fujian Province, China. Virus isolation, PCR detection, phylogenetic analysis, qRT-PCR, fluorescence in situ hybridization and histology were used for virus identification and analysis of virus histopathology. Furthermore, an artificial infection experiment was conducted for virulence testing. Results Brain and eye tissue homogenates of diseased big-belly seahorses were inoculated onto a grouper spleen (GS) cell monolayer at 28 °C. Tissue homogenates induced obvious cytopathic effects in GS cells. PCR and sequencing analyses revealed that the virus belonged to Betanodavirus and shared high sequence identity with red-spotted grouper nervous necrosis virus isolates. qRT-PCR and fluorescence in situ hybridization revealed that SHNNV mainly attacked the brain and eye. Histopathological examination revealed that the virus led to cytoplasmic vacuolation in the brain and retinal tissues. Infection experiments confirmed that SHNNV was highly infectious, causing massive death in big-belly seahorses. Conclusion A novel seahorse betanodavirus from the big-belly seahorse cultured in China was discovered. This finding will contribute to the development of efficient strategies for disease management in aquaculture.

利用组织学切片及VASA抗体的免疫荧光技术解析了线纹海马的性腺发育与分化过程.结果显示:在早期性腺发育过程中,雌性线纹海马体长为12 mm时,体腔背部出现生殖嵴;体长为17 mm时,原始性腺开始分化,形成卵巢腔;随后卵巢开始发育,最后在体长为40 mm时,卵巢发育成熟.雄性线纹海马生殖嵴出现的时间与雌性相同,但是性腺分化比雌性时间略晚,雄性线纹海马在体长为22 mm时,原始性腺才开始分化,形成精小叶.最后在体长为45 mm时,精巢发育成熟.

为了提高膨腹海马(Hippocampus abdominalis)养殖效率,了解温度、配对方式与数量、养殖密度等对膨腹海马生长、存活及生殖的影响,本研究测定了不同温度、不同雌雄数量比对膨腹海马亲本产苗量的影响和不同培育密度对膨腹海马苗种生长、存活的影响.研究结果表明:膨腹海马亲鱼适宜的繁殖水温范围为16.0～19.0℃;膨腹海马雌雄鱼比例控制在1:1、2:3的范围内繁殖效果最佳,产苗量相对较高;膨腹海马苗种生产的早期阶段(30日龄前),最适培育密度为0.50～1.00尾/L,中期阶段(30～60日龄),最适培育密度为0.25～0.50尾/L.本研究为膨腹海马人工繁殖与育苗技术提供了科学数据.

赤点石斑鱼(Epinephelus akaara)是中国东南沿海地区一种重要的海水养殖鱼类,多年来其苗种生产深受神经坏死病的困扰,严重制约了其人工养殖业的发展.本研究采集230尾赤点石斑鱼神经坏死病毒(RGNNV)易感(染病死亡)和230尾抗性(最终健康存活)赤点石斑鱼苗进行基因组重测序,分析获得5412683个单核苷酸多态性(SNPs)位点的基因型,并以之对抗病性状进行了遗传评估和基因组选择研究,获得的估计遗传力均值为0.5662,预测基因组估计育种值(GEBV)均值为0.1543.随机选择不同数量标记对基因组选择准确性影响进行评估,结果表明,采用≥5000个(5 k)标记进行赤点石斑鱼抗神经坏死病性状遗传评估就可以获得比较理想的效果.本研究为开展赤点石斑鱼抗神经坏死病基因组选择育种实践提供了有用的理论参考.

Purpose:Tight junctions (TJs) form the structural basis of retinal pigment epithelium (RPE) barrier functions. Although oxidative stress contributes to age-related macular degeneration, it is unclear how RPE TJ integrity is controlled by redox balance. In this study, we investigated the protective roles of nuclear factor erythroid 2-related factor 2 (NRF2), a transcription factor, and heme oxygenase-1 (HO1), a heme-degrading enzyme encoded by the NRF2 target gene HMOX1. Methods:ARPE19 cell cultures and mice, including wild-type, Nrf2-/-, and RPE-specific NRF2-deficient mice, were treated with chemicals that impose oxidative stress or impact heme metabolism. In addition, NRF2 and HO1 expression in ARPE19 cells was knocked down by siRNA. TJ integrity was examined by anti-zonula occludens-1 staining of cultured cells or flatmount RPE tissues from mice. RPE barrier functions were evaluated by transepithelium electrical resistance in ARPE19 cells and immunofluorescence staining for albumin or dextran in eye histological sections. Results:TJ structures and RPE barrier functions were compromised due to oxidant exposure and NRF2 deficiency but were rescued by HO1 inducer. Furthermore, treatment with HO1 inhibitor or heme precursor is destructive to TJ structures and RPE barrier properties. Interestingly, both NRF2 and HO1 were upregulated under oxidative stress, probably as an adaptive response to mitigate oxidant-inflicted damages. Conclusions:Our data indicate that the NRF2-HO1 axis protects TJ integrity and RPE barrier functions by driving heme degradation.

为探究温度对不同体质量灰海马耗氧率、排氨率和窒息点的影响,设置13、18、23、28、32℃5个温度梯度,用大(L)、中(M)、小(S)3种体质量分别为(3.63±1.42)、(1.23±0.22)、(0.47±0.24)g的灰海马进行代谢试验.试验结果:在13～32℃范围内,随着水温的升高,灰海马的耗氧率和排氨率呈现先升高后降低的趋势,3种体质量的海马在水温28℃时耗氧率均达到最大值;同一温度条件下,灰海马的体质量越大,其耗氧率和排氨率越低;灰海马的窒息点随着温度的上升而提高,但在相同温度条件下,灰海马体质量越大,其窒息点越低.结果表明,灰海马养殖的最适水温在28℃左右,养殖和运输过程中应保证水体溶解氧在1.6 mg/L以上.

Seahorses belong to the teleost family Syngnathidae that evolved a distinct body plan and unique male pregnancy compared to other teleosts. As a classic model for studying evolution of viviparity and sexual selection of teleosts, seahorse species still lack a publicly available high-quality reference genome. Here, we generated the genome assembly of the big-belly seahorse, Hippocampus abdominalis with long-read and Hi-C technologies. We managed to place over 99% of the total length of 444.7 Mb of assembled genome into 21 linkage groups with almost no gaps. We reconstructed a phylogenomic tree with the big-belly seahorse genome and other representative Syngnathidae and teleost species. We also reconstructed the historical population dynamics of four representative Syngnathidae species. We found the gene families that underwent expansion or contraction in the Syngnathidae ancestor were enriched for immune-related or ion transporter gene ontology terms. Many of these genes were also reported to show a dynamic expression pattern during the pregnancy stages of H. abdominalis. We also identified putative positively selected genes in the Syngnathidae ancestor or in H. abdominalis, whose mouse mutants are enriched for abnormal craniofacial and limb morphological phenotypes. Overall, our study provides an important genome resource for evolutionary and developmental studies of seahorse species, and candidate genes for future experimental works.

毕加索小丑鱼(Picasso clownfish)因其皮肤中的白色斑块分布杂乱抽象而得名,同时也由于其白色斑块的形成无规律性和稀缺性,属于名贵的小丑鱼,因此,解析毕加索小丑鱼的皮肤白斑形成机制,可以为毕加索小丑鱼的人工育种提供理论依据.在本研究中,我们对3种体色毕加索小丑鱼的背鳍和臀鳍之间的身体相同部位3种色块(黑色、黄色、白色)的皮肤进行进行了转录组测序.研究结果显示,与黄色和黑色皮肤组织相比,在白色皮肤组织中存在大量差异表达基因.其中与黑色素产生相关的信号通路(例如黑色素生成、Hedgehog信号通路和Wnt信号通路)中的基因在白色皮肤组织中呈下调表达趋势.进一步对3种颜色皮肤组织中参与黑色素生成途径的基因进行详细分析,发现上游调控基因(如ednrba和mitfa)表达量从黑色到黄色至白色皮肤组织中逐步下调,但下游参与黑色素合成的核心基因(包括tyr、tyrp1b和dct)则在白色皮肤组织中表达出现显著下调.最后,本研究通过荧光定量PCR验证了转录组数据的可靠性.本研究结果将为今后人为干扰基因表达从而达到调控小丑鱼体色提供理论依据.

The main physiological function of 17β-estradiol (E2) in vertebrates is to regulate sexual development and reproduction. In fish, especially hermaphroditic fish, estrogen is often used to aid reproduction, but it also can trigger an inflammatory response. However, the molecular mechanism for this E2-induced inflammatory reaction is not clear. In this study, we found that the ERβ-CXCL19/CXCR4-NFκB cascade regulated the E2-induced inflammatory response in the orange-spotted grouper (Epinephelus coioides). Strikingly, E2 treatment resulted in significantly high expression of inflammatory cytokines and induced phosphorylation and degradation of IκBα and translocation of NFκB subunit p65 to the nucleus in grouper spleen cells. However, the E2-induced inflammatory response could be prevented by the broad estrogen receptor (ER) ligand ICI 182,780. Moreover, the luciferase assay showed that E2 induced the inflammatory response by activating the promotor of chemokine CXCL19 through ERβ1 and ERβ2. Knockdown of CXCL19 blocked the E2-induced inflammatory response and NFκB nucleus translocation. Additionally, knockdown of chemokines CXCR4a and CXCR4b together, but not alone, blocked the E2-induced inflammatory response. The immunofluorescence assay and co-immunoprecipitation analysis showed that CXCL19 mediated the E2-induced inflammatory response by activating CXCR4a or CXCR4b. Taken together, these results showed that the ERβ-CXCL19/CXCR4-NFκB pathway mediated the E2-induced inflammatory response in grouper. These findings are valuable for future comparative immunological studies and provide a theoretical basis for mitigating the adverse reactions that occur when using E2 to help fish reproduce.