Characterization of the Apoptotic and Antimicrobial Activities of Two Initiator Caspases of Sea Cucumber Apostichopus japonicus.

Caspase (CASP) is a protease family that plays a vital role in apoptosis, development, and immune response. Herein, we reported the identification and characterization of two CASPs, AjCASPX1 and AjCASPX2, from the sea cucumber Apostichopus japonicus, an important aquaculture species. AjCASPX1/2 share similar domain organizations with the vertebrate initiator caspases CASP2/9, including the CARD domain and the p20/p10 subunits with conserved functional motifs. However, compared with human CASP2/9, AjCASPX1/2 possess unique structural features in the linker region between p20 and p10. AjCASPX1, but not AjCASPX2, induced marked apoptosis of human cells by activating CASP3/7. The recombinant proteins of AjCASPX2 and the CARD domain of AjCASPX2 were able to bind to a wide range of bacteria, as well as bacterial cell wall components, and inhibit bacterial growth. AjCASPX1, when expressed in Escherichia coli, was able to kill the host bacteria. Under normal conditions, AjCASPX1 and AjCASPX2 expressions were most abundant in sea cucumber muscle and coelomocytes, respectively. After bacterial infection, both AjCASPX1 and AjCASPX2 expressions were significantly upregulated in sea cucumber tissues and cells. Together, these results indicated that AjCASPX1 and AjCASPX2 were initiator caspases with antimicrobial activity and likely functioned in apoptosis and immune defense against pathogen infection.

Fucoidan from Apostichopus japonicus ameliorates alcoholic liver disease by regulating gut-liver axis homeostasis.

Long-term and excessive alcohol consumption can lead to the development of alcoholic liver disease (ALD), characterized by oxidative damage, intestinal barrier injury, and disruption of intestinal microbiota. In this study, we extracted fucoidan (Aj-FUC) from Apostichopus japonicus using enzymatic methods and characterized its structure. The ALD model was established in male Balb/c mice using 56° Baijiu, with silymarin as a positive control. Mice were orally administered 100 mg/kg·bw and 300 mg/kg·bw of Aj-FUC for 28 days to evaluate its effects on liver injury in ALD mice and explore its potential role in modulating the gut-liver axis. The results showed significant improvements in histopathological changes and liver disease in the Aj-FUC group. Aj-FUC treatment significantly increased the levels of glutathione (GSH) and glutathione peroxidase (GSH-Px) while weakly reduced the elevation of malondialdehyde (MDA) induced by ALD. It also regulated the Nrf2/HO-1 signaling pathway, collectively alleviating hepatic oxidative stress. Aj-FUC intervention upregulated the expression of ZO-1 and Occludin, thus contributing to repair the intestinal barrier. Additionally, Aj-FUC increased the content of short-chain fatty acids (SCFAs) and regulated the imbalance in gut microbiota. These results suggested that Aj-FUC alleviates ALD by modulating the gut-liver axis homeostasis. It may prove to be a useful dietary supplement in the treatment of alcoholic liver damage.

A multi-objective optimization approach for green supply chain network design for the sea cucumber (Apostichopus japonicus) industry.

In China, aquatic supply chain network design does not include the green concept or the coordination of environmental and economic performance. Sea cucumber (Apostichopus japonicus) is an aquatic product of high economic value; however, studies on sea cucumber supply chain network optimization are lacking. This study is the first to design the sea cucumber supply chain and construct an optimization model. Considering the characteristics of the sea cucumber industry, LCA for Experts software and the CML-IA-Aug. 2016-world method were used to assess each aquaculture model's global warming potential (GWP), as the environmental performance indicator. In addition, multi-objective genetic algorithm (MOGA) coupled with Modified Technique for Order of Preference by Similarity to Ideal Solution (M-TOPSIS) integrates yield production, economic benefits, and environmental performance. The results demonstrated that cage seed rearing (CSR) combined bottom sowing aquaculture (BSA) represents the best production strategy upstream of the sea cucumber supply chain. In the downstream, the best proportion of sales channels in supermarkets, boutique stores and online shops accounted for 14.79 %, 58.02 % and 27.19 % of the production, respectively. The proposed optimization scenario 4 (S4) can increase product profit by 27.88 % and reduce GWP by 56.89 %. The following improvement measures are proposed: using sea cucumber aquaculture industry standards (cleaner production and green supplier selection) to regulate the behavior of enterprises, adopting an ecological and green production strategy, eliminating high-energy consumption and high emission production practices, and promoting widespread adoption of green consumption concepts. Finally, these measures may improve the sea cucumber supply chain, achieve coordinated environmental and economic performance development in the sea cucumber industry, and provide guidance for green optimization of other aquatic product supply chains in China.

TMT-Based Quantitative Proteomic Analysis Reveals the Key Role of Cell Proliferation and Apoptosis in Intestine Regeneration of Apostichopus japonicus.

Sea cucumbers are widely known for their powerful regenerative abilities, which allow them to regenerate a complete digestive tract within a relatively short time following injury or autotomy. Recently, even though the histological changes and cellular events in the processes of intestinal regeneration have been extensively studied, the molecular machinery behind this faculty remains unclear. In this study, tandem mass tag (TMT)-based quantitation was utilized to investigate protein abundance changes during the process of intestine regeneration. Approximately 538, 445, 397, 1012, and 966 differential proteins (DEPs) were detected (p < 0.05) between the normal and 2, 7, 12, 20, and 28 dpe stages, respectively. These DEPs also mainly focus on pathways of cell proliferation and apoptosis, which were further validated by 5-Ethynyl-2'-deoxyuridine (EdU) or Tunel-based flow cytometry assay. These findings provide a reference for a comprehensive understanding of the regulatory mechanisms of various stages of intestinal regeneration and provide a foundation for subsequent research on changes in cell fate in echinoderms.

Comparative Analysis of Gut Bacterial Community Composition in Two Tropical Economic Sea Cucumbers under Different Seasons of Artificial Environment.

With the continuous rise of the sea cucumber aquaculture industry in China, the tropical sea cucumber aquaculture industry is also improving. However, research on the gut microorganisms of tropical sea cucumbers in captivity is scarce. In this study, high-throughput sequencing methods were used to analyze the gut microbial composition of Stichopus monotuberculatus and Holothuria scabra in the dry season and wet season of artificial environments. The results showed that 66 phyla were obtained in all samples, of which 59 phyla were obtained in the dry season, and 45 phyla were obtained in the wet season. The Tax4Fun analysis showed that certain gut bacterial communities affect the daily metabolism of two sea cucumber species and are involved in maintaining gut microecological balance in the gut of two sea cucumber species. In addition, compared with differences between species, PCoA and UPGMA clustering analysis showed the gut prokaryotes of the same sea cucumber species varied more in different seasons, indicating that the influence of environment was higher than the feeding choices of sea cucumbers under relatively closed conditions. These results revealed the gut bacterial community composition of S. monotuberculatus and H. scabra and the differences in gut bacterial structure between two sea cucumber species in different seasons were compared, which would provide the foundation for tropical sea cucumber aquaculture in the future.

Polyphenol oxidase mediates (-)-epigallocatechin gallate to inhibit endogenous cathepsin activity in Apostichopus japonicus.

Apostichopus japonicus (A. japonicus) has rich nutritional value and is an important economic crop. Due to its rich endogenous enzyme system, fresh A. japonicus is prone to autolysis during market circulation and storage, resulting in economic losses. In order to alleviate this phenomenon, we investigated the effect of polyphenol oxidase (PPO) mediated (-)-epigallocatechin gallate (EGCG) on the activity and structure of endogenous cathepsin series protein (CEP) from A. japonicus. Research on cathepsin activity showed that PPO mediated EGCG could significantly reduce enzyme activity, resulting in a decrease in enzymatic reaction rate. SDS-PAGE and scanning electron microscopy results showed that PPO mediates EGCG could induce CEP aggregation to form protein aggregates. Various spectral results indicated that EGCG caused changes in the structure of CEP. Meanwhile, the conjugates formed by PPO mediated EGCG had lower thermal stability. In conclusion, PPO mediated EGCG was an effective method to inhibit the endogenous enzyme activity.

Akirin2 enhances antibacterial ability via interacting with 14-3-3ζ in V. splendidus-challenged Apostichopus japonicus.

Akirin2 is pivotal for regulating host immunological responses in vertebrates, including antibacterial immunity and inflammation. However, the functional significance of Akirin2 in invertebrates remains largely unexplored. In this study, we cloned the complete cDNA sequence of Akirin2 from A. japonicus (AjAkirin2) and elucidated its immunological mechanism upon pathogen infection. The whole AjAkirin2 cDNA sequence spanned 1014 bp, which comprised a 630 bp open reading frame encoding 209 amino acids, a 230 bp 5'-untranslated region (UTR), and a 154 bp 3'-UTR. Spatial expression analysis displayed constitutive expression of AjAkirin2 in all examined tissues. Both mRNA and protein expression abundance of the AjAkirin2 showed considerably high in coelomocytes of sea cucumbers challenged with Vibrio splendidus or stimulated with lipopolysaccharide. In addition, we found that sea cucumbers with 107 CFU/mL V. splendidus infection had a lower survival rate upon AjAkirin2 knockdown. Mechanistically, the result of GST-pull down and co-IP assays indicated that AjAkirin2 directly interacted with Aj14-3-3ζ. Moreover, we also detected that AjAkirin2 positively regulated Aj14-3-3ζ expression in sea cucumber coelomocytes. Furthermore, the knockdown of AjAkirin2 or Aj14-3-3ζ resulted in increasing intracellular bacteria load and suppressed the expression of key genes of the NF-κB signaling pathway (p65 and p105) and inflammatory cytokines including IL-17, VEGF, and MMP-1. In summary, these results confirmed the critical role of AjAkirin2 in mediating innate immune responses against V. splendidus infection via interaction with Aj14-3-3ζ and thereby exerting antibacterial function.

Ferroptosis resists intracellular Vibrio splendidus AJ01 mediated by ferroportin in sea cucumber Apostichopus japonicus.

Ferroptosis, a kind of programmed cell death, is characterized with iron-dependent lipid ROS buildup, which is considered as an important cellular immunity in resisting intracellular bacterial infection in mammalian macrophages. In this process, lipid ROS oxidizes the bacterial biofilm to inhibit intracellular bacteria. However, the function of ferroptosis in invertebrate remains unknown. In this study, the existence of ferroptosis in Apostichopus japonicus coelomocytes was confirmed, and its antibacterial mechanism was investigated. First, our results indicated that the expression of glutathione peroxidase (AjGPX4) was significantly inhibited by 0.21-fold (p < 0.01) after injecting A. japonicus with the ferroptosis inducer RSL3, and the contents of MDA (3.93-fold, p < 0.01), ferrous iron (1.40-fold, p < 0.01), and lipid ROS (3.10-fold, p < 0.01) were all significantly increased under this condition and simultaneously accompanied with mitochondrial contraction and disappearance of cristae, indicating the existence of ferroptosis in the coelomocytes of A. japonicus. Subsequently, the contents of ferrous iron (1.40-fold, p < 0.05), MDA (2.10-fold, p < 0.01), ROS (1.70-fold, p < 0.01), and lipid ROS (2.50-fold, p < 0.01) were all significantly increased, whereas the mitochondrial membrane potential and GSH/GSSG were markedly decreased by 0.68-fold (p < 0.05) and 0.69-fold (p < 0.01) under Vibrio splendidus (AJ01) infection. This process could be reversed by the iron-chelating agent deferoxamine mesylate, which indicated that AJ01 could induce coelomocytic ferroptosis. Moreover, the results demonstrated that the intracellular AJ01 load was clearly decreased to 0.49-fold (p < 0.05) and 0.06-fold (p < 0.01) after treating coelomocytes with RSL3 and ferrous iron, which indicated that enhanced ferroptosis could inhibit bacterial growth. Finally, subcellular localization demonstrated that ferrous iron efflux protein ferroportin (AjFPN) and intracellular AJ01 were co-localized in coelomocytes. After AjFPN interference (0.58-fold, p < 0.01), the signals of ferrous iron and lipid ROS levels in intracellular AJ01 were significantly reduced by 0.38-fold (p < 0.01) and 0.48-fold (p < 0.01), indicating that AjFPN was an important factor in the introduction of ferroptosis into intracellular bacteria. Overall, our findings indicated that ferroptosis could resist intracellular AJ01 infection via AjFPN. These findings provide a novel defense mechanism for aquatic animals against intracellular bacterial infection.

Apoptosis-inducing factor 1 mediates Vibrio splendidus-induced coelomocyte apoptosis via importin ß dependent nuclear translocation in Apostichopus japonicus.

As is well known, apoptosis is an important form of immune response and immune regulation, particularly playing a crucial role in combating microbial infections. Apoptosis-inducing factor 1 (AIF-1) is essential for apoptosis to induce chromatin condensation and DNA fragmentation via a caspase-independent pathway. The nuclear translocation of AIF-1 is a key step in apoptosis but the molecular mechanism is still unclear. In this study, the homologous gene of AIF-1, named AjAIF-1, was cloned and identified in Apostichopus japonicus. The mRNA expression of AjAIF-1 was significantly increased by 46.63-fold after Vibrio splendidus challenge. Silencing of AjAIF-1 was found to significantly inhibit coelomocyte apoptosis because the apoptosis rate of coelomocyte decreased by 0.62-fold lower compared with the control group. AjAIF-1 was able to promote coelomocyte apoptosis through nuclear translocation under the V. splendidus challenge. Moreover, AjAIF-1 and Ajimportin ß were mainly co-localized around the nucleus in vivo and silencing Ajimportin ß significantly inhibited the nuclear translocation of AjAIF-1 and suppressed coelomocyte apoptosis by 0.64-fold compared with control. In summary, nuclear translocation of AjAIF-1 will likely mediate coelomocyte apoptosis through an importin ß-dependent pathway in sea cucumber.

A2M possesses anti-bacterial functions by recruiting and enhancing phagocytosis through GRP78 in an echinoderm.

Alpha-2-macroglobulin (A2M) is an extracellular macromolecule mainly known for its role as a broad-spectrum protease inhibitor in mammals. However, the immune recognition and regulation mechanisms of A2M in invertebrates are still not well investigated. In the current study, the role of sea cucumber Apostichopus japonicus A2M in the regulation of innate immune responses was explored. We found that AjA2M promotes phagocytosis of Vibrio splendidus in coelomocytes of sea cucumber. Then two major functional structural domains of AjA2M, the thioester domain (TED) and the receptor-binding structural domain (RBD) were cloned. It was found that the AjA2M-TED binds to pathogens while causing Vibrio splendidus aggregation; the AjA2M-RBD interacts with the Glucose Regulated Protein 78 (AjGRP78), subsequently AjGRP78 accelerates the degradation of Vibrio splendidus in lysosomes by facilitating polymerisation and rearrangement of the cytoskeleton. Collectively, the findings together suggest that A2M-GRP78 axis mediates immune signaling pathway of phagocytosis and AjA2M has been characterized to play an essential crucial role in antibacterial immune responses of invertebrates.

Investigation of the Presence of Fibrillin in Sea Cucumber (Apostichopus japonicus) Body Wall by Utilizing Targeted Proteomics and Visualization Strategies.

Fibrillin is an important structural protein in connective tissues. The presence of fibrillin in sea cucumber Apostichopus japonicus is still poorly understood, which limits our understanding of the role of fibrillin in the A. japonicus microstructure. The aim of this study was to clarify the presence of fibrillin in the sea cucumber A. japonicus body wall. Herein, the presence of fibrillin in sea cucumber A. japonicus was investigated by utilizing targeted proteomics and visualization strategies. The contents of three different isoforms of fibrillin with high abundance in A. japonicus were determined to be 0.96, 2.54, and 0.15 µg/g (wet base), respectively. The amino acid sequence of fibrillin (GeneBank number: PIK56741.1) that started at position 631 and ended at position 921 was selected for cloning and expressing antigen. An anti-A. japonicus fibrillin antibody with a titer greater than 1:64 000 was successfully obtained. It was observed that the distribution of fibrillin in the A. japonicus body wall was scattered and dispersed in the form of fibril bundles at the microscale. It further observed that fibrillin was present near collagen fibrils and some entangled outside the collagen fibrils at the nanoscale. Moreover, the stoichiometry of the most dominant collagen and fibrillin molecules in A. japonicus was determined to be approximately 250:1. These results contribute to an understanding of the role of fibrillin in the sea cucumber microstructure.

YTHDC1/CRM1 Facilitates m6A-Modified circRNA388 Nuclear Export to Induce Coelomocyte Autophagy via the miR-2008/ULK Axis in Apostichopus japonicus.

N 6-methyladenosine (m6A), the most prevalent internal modification in eukaryotic RNA, was able to mediate circular RNA (circRNA) function in many immune processes. Nevertheless, the functional role of m6A-modified circRNAs in innate immunity of invertebrates remained unclear. In this study, we identified m6A-modified circRNA388 from cultured sea cucumber (Apostichopus japonicus) coelomocytes, which was mainly detected in cytoplasm after Vibrio splendidus infection. A knockdown assay indicated that cytoplasm circRNA388 promoted coelomocyte autophagy and decreased the number of intracellular V. splendidus. Mechanistically, the circRNA388 in the cytoplasm directly sponged miR-2008 to block its interaction with Unc-51-like kinase 1 from A. japonicus (AjULK) and further promoted autophagy to resist V. splendidus infection. More importantly, we found that m6A modification was vital to circRNA388 nuclear export with YTH domain-containing protein 1 from A. japonicus (AjYTHDC1) as the reader. AjYTHDC1 facilitated the nuclear export of m6A-modified circRNA388 via interaction with exportin-1 (chromosomal maintenance 1) from A. japonicus (AjCRM1). Knockdown of AjCRM1 could significantly decrease the content of cytoplasm circRNA388. Overall, our results provide the first evidence that nuclear export of m6A-modified circRNA388 is dependent on the novel AjCRM1 to our knowledge, which was further promoted coelomocyte autophagy by miR-2008/AjULK axis to clear intracellular V. splendidus.

Evaluation of the chronic toxicity of bisphenol A and bisphenol AF to sea cucumber Apostichopus japonicus after long-term single and combined exposure at environmental relevant concentration.

Bisphenols are emerging endocrine disrupting pollutant, and several studies have reported that they are already ubiquitous in various environmental matrices and intend to deposit in sediment. The primary sources of bisphenols are river and sewage discharge. Sea cucumber (Apostichopus japonicus), a typical deposit feeder, is one of the most important commercial marine species in Aisa. However, the effects of the bisphenol A (BPA) and its analogues bisphenol AF (BPAF) on sea cucumber was unclear. In this study, we carried out field survey in major sea cucumber farming areas in northern China, with the aim of determining which bisphenol analogue is the major bisphenol contamination in this aquaculture area. The results showed that the presence of BPAF was detected in four sampling sites (Dalian, Tangshan, Laizhou, and Longpan). The mean level of BPAF in Laizhou sediment samples was the highest which reached to 9.007 ± 4.702 µ g/kg. Among the seawater samples, the BPAF only have been detected in the samples collected at Longpan. (0.011 ± 0.003 µ g/L). Furthermore, we conducted an experiment to evaluate the single and combined toxicity of BPA and BPAF on sea cucumbers. The concentrations were informed by the findings based on the results of field research. (0.1, 1.0, and 10 µ g/L). After exposure, the body weight gain, and specific growth rate showed no significant changes (P > 0.05). We observed the histological alterations in respiratory tree of treated sea cucumbers including the fusion and detachment of lining epithelial tissue, and increase of lumen space. However, the catalase (CAT), malondialdehyde (MDA), and glutathione (GSH) activity was not significantly changed (P > 0.05). We evaluated the effects of BPA and BPAF through calculating the integrated biomarker response index (IBR), and the results indicated that the toxicity of combined treatment was higher than single treatment. Additionally, BPAF exposure to A. japonicus was more toxic than BPA.

Sulfamethoxazole stress endangers the gut health of sea cucumber (Apostichopus japonicus) and affects host metabolism.

Sulfamethoxazole (SMZ) is a frequently detected antibiotic in the environment, and there is a growing concern about its potential toxic effects on aquatic organisms. sea cucumber (Apostichopus japonicas) is a benthic invertebrate whose gut acts as a primary immune defense and serves critical protective barrier. In this study, growth performance, histology, gut microbiota, and metabolomics analyses were performed to investigate the toxic response in the intestine of sea cucumber effects caused by SMZ stress for 56 d by evaluating with different concentrations of SMZ (0, 1.2×10-3, and 1.2 mg/L). The weight gain rate of sea cucumbers under SMZ stress showed significant decrease, indicating that the growth of sea cucumbers was hindered. Analysis of the intestinal morphological features indicated that SMZ stimulation resulted in atrophy of the sea cucumber gut. In the 1.2×10-3 mg/L concentration, the thickness of muscle and mucosal layers was reduced by 12.40% and 21.39%, while in the 1.2 mg/L concentration, the reductions were 35.08% and 26.98%. The abundance and diversity of sea cucumber intestinal bacteria decreased significantly (P < 0.05) under the influence of SMZ. Notably, the intestinal bacteria of sea cucumber became homogenized with the increase in SMZ concentration, and the relative abundance of Ralstonia reached 81.64% under the stress of 1.2 mg/L concentration. The SMZ stress significantly impacted host metabolism and disrupted balance, particularly in L-threonine, L-tyrosine, neuronic acid, piperine, and docosapentaenoic acid. SMZ leads to dysregulation of metabolites, resulting in growth inhibition and potential inflammatory responses that could adversely affect the normal activities of aquatic organisms. Further metabolic pathway enrichment analyses demonstrated that impaired biosynthesis of unsaturated fatty acids and aminoacyl-tRNA biosynthesis metabolic pathway were major reasons for SMZ stress-induced intestinal bacteria dysbiosis. This research aims to provide some theoretical evidence for the ecological hazard assessment of antibiotics in water.

Holotoxin A1 from Apostichopus japonicus inhibited oropharyngeal and intra-abdominal candidiasis by inducing oxidative damage in Candida albicans.

BACKGROUND AND PURPOSE: The holotoxin A1, isolated from Apostichopus japonicus, exhibits potent antifungal activities, but the mechanism and efficacy against candidiasis are unclear. In this study we have studied the antifungal effects and mechanism of holotoxin A1 against Candida albicans and in murine oropharyngeal and intra-abdominal candidiasis. EXPERIMENTAL APPROACH: The antifungal effect of holotoxin A1 against C. albicans was tested in vitro. To explore the antifungal mechanism of holotoxin A1, the transcriptome, ROS levels, and mitochondrial function of C. albicans was evaluated. Effectiveness and systematic toxicity of holotoxin A1 in vivo was assessed in the oropharyngeal and intra-abdominal candidiasis models in mice. KEY RESULTS: Holotoxin A1 was a potent fungicide against C. albicans SC5314, clinical strains and drug-resistant strains. Holotoxin A1 inhibited oxidative phosphorylation and induced oxidative damage by increasing intracellular accumulation of ROS in C. albicans. Holotoxin A1 induced dysfunction of mitochondria by depolarizing the mitochondrial membrane potential and reducing the production of ATP. Holotoxin A1 directly inhibited the enzymatic activity of mitochondrial complex I and antagonized with the rotenone, an inhibitor of complex I, against C. albicans. Meanwhile, the complex I subunit NDH51 null mutants showed a decreased susceptibility to holotoxin A1. Furthermore, holotoxin A1 significantly reduced fungal burden and infections with no significant systemic toxicity in oropharyngeal and intra-abdominal candidiasis in murine models. CONCLUSION AND IMPLICATIONS: Holotoxin A1 is a promising candidate for the development of novel antifungal agents against both oropharyngeal and intra-abdominal candidiasis, especially when caused by drug-resistant strains.

Pseudocryptic diversity and species boundaries in the sea cucumber Stichopus cf. horrens (Echinodermata: Stichopodidae) revealed by mitochondrial and microsatellite markers.

Morphologically cryptic and pseudo-cryptic species pose a challenge to taxonomic identification and assessments of species diversity and distributions. Such is the case for the sea cucumber Stichopus horrens, commonly confused with Stichopus monotuberculatus. Here, we used mitochondrial cytochrome oxidase subunit I (COI) and microsatellite markers to examine genetic diversity in Stichopus cf. horrens throughout the Philippine archipelago, to aid species identification and clarify species boundaries. Phylogenetic analysis reveals two recently diverged COI lineages (Clade A and Clade B; c. 1.35-2.54 Mya) corresponding to sequence records for specimens identified as S. monotuberculatus and S. horrens, respectively. Microsatellite markers reveal two significantly differentiated genotype clusters broadly concordant with COI lineages (Cluster 1, Cluster 2). A small proportion of individuals were identified as later-generation hybrids indicating limited contemporary gene flow between genotype clusters, thus confirming species boundaries. Morphological differences in papillae distribution and form are observed for the two species, however tack-like spicules from the dorsal papillae are not a reliable diagnostic character. An additional putative cryptic species was detected within Clade B-Cluster 2 specimens warranting further examination. We propose that these lineages revealed by COI and genotype data be referred to as Stichopus cf. horrens species complex.

Fucosylated chondroitin sulfate from sea cucumber Stichopus chloronotus alleviate the intestinal barrier injury and oxidative stress damage in vitro and in vivo.

This study aimed to investigate the alleviative effects of fucosylated chondroitin sulfate from sea cucumber Stichopus chloronotus (fCSSc) on the intestinal barrier injury and oxidative stress damage in vitro and in vivo. The results showed that fCS-Sc protected the intestinal barrier and improved the antioxidant function in H2O2 damaged Caco-2 cells via up-regulating the tight junction proteins and activating Keap1-Nrf2-ARE antioxidant pathway. Furthermore, administration fCS-Sc could ameliorate the weight loss and spleen index decrease in Cyclophosphamide (Cy) treated mice, improve the expressions of ZO-1, Claudin-1, Nrf2, SOD, and NQO-1 in Cy damaged colon tissue, showing significant protective effects against intestinal barrier damage and oxidative stress in vivo. fCS-Sc intervention also alleviated the gut microbiota disorder though increasing the richness and diversity of intestinal bacteria, regulating the structural composition of gut microbiota. fCS-Sc promoted the relative abundance of beneficial microbiota and inhibited the growth of harmful bacteria. This study provided a theoretical basis for the application of fCS-Sc as a prebiotic in chemotherapy.

Alterations in mitochondrial structure and function in response to environmental temperature changes in Apostichopus japonicus.

Global temperatures have risen as a result of climate change, and the resulting warmer seawater will exert physiological stresses on many aquatic animals, including Apostichopus japonicus. It has been suggested that the sensitivity of aquatic poikilothermal animals to climate change is closely related to mitochondrial function. Therefore, understanding the interaction between elevated temperature and mitochondrial functioning is key to characterizing organisms' responses to heat stress. However, little is known about the mitochondrial response to heat stress in A. japonicus. In this work, we investigated the morphological and functional changes of A. japonicus mitochondria under three representative temperatures, control temperature (18 °C), aestivation temperature (25 °C) and heat stress temperature (32 °C) temperatures using transmission electron microscopy (TEM) observation of mitochondrial morphology combined with proteomics and metabolomics techniques. The results showed that the mitochondrial morphology of A. japonicus was altered, with decreases in the number of mitochondrial cristae at 25 °C and mitochondrial lysis, fracture, and vacuolization at 32 °C. Proteomic and metabolomic analyses revealed 103 differentially expressed proteins and 161 differential metabolites at 25 °C. At 32 °C, the levels of 214 proteins and 172 metabolites were significantly altered. These proteins and metabolites were involved in the tricarboxylic acid (TCA) cycle, substance transport, membrane potential homeostasis, anti-stress processes, mitochondrial autophagy, and apoptosis. Furthermore, a hypothetical network of proteins and metabolites in A. japonicus mitochondria in response to temperature changes was constructed based on proteomic and metabolomic data. These results suggest that the dynamic regulation of mitochondrial energy metabolism, resistance to oxidative stress, autophagy, apoptosis, and mitochondrial morphology in A. japonicus may play important roles in the response to elevated temperatures. In summary, this study describes the response of A. japonicus mitochondria to temperature changes from the perspectives of morphology, proteins, and metabolites, which provided a better understanding the mechanisms of mitochondrial regulation under environment stress in marine echinoderms.

Gut microbiota dynamics interacting with gastrointestinal evacuation of Apostichopus japonicus: novel insights into promising strategies for environmental improvement.

As an important input of environmental micropollutants into aquaculture environment, feed is now considered to be a critical factor in shaping gastrointestinal evacuation characteristics of animals. We analyzed the gastrointestinal evacuation characteristics and gut bacteria of Apostichopus japonicus within 30 h after feeding in recirculating aquaculture system (RAS) and explored the evacuation mechanism interacting by bacteria. The Gauss model was the most precise gastrointestinal evacuation curve, and 80% of gastrointestinal evacuation time was 27.81 h after feeding. Linear discriminant analysis effect size analysis revealed that gut microbial abundance associated significantly with time (P < 0.05), and 42 biomarkers that could predict gastrointestinal evacuation were totally detected, such as Lutibacter and Vibrio. Biomarkers at 25 h after feeding were related to harmful bacteria. A dynamic response between gastrointestinal content ratio and gut microbial abundance was detected. Taken together, we could discharge sewage about 25 h after feeding and carry out the next round of feeding activities.

Dietary steroids promote body weight growth and induce gametogenesis by increasing the expressions of genes related to cell proliferation of sea cucumber (Apostichopus japonicus).

Steroids play a vital role in animal survival, promoting growth and development when administered appropriate concentration exogenously. However, it remains unclear whether steroids can induce gonadal development and the underlying mechanism. This study assessed sea cucumber weights post-culturing, employing paraffin sections and RNA sequencing (RNA-seq) to explore gonadal changes and gene expression in response to exogenous steroid addition. Testosterone and cholesterol, dissolved in absolute ethanol, were incorporated into sea cucumber diets. After 30 days, testosterone and cholesterol significantly increased sea cucumber weights, with the total weight of experimental groups surpassing the control. The testosterone-fed group exhibited significantly higher eviscerated weight than the control group. In addition, dietary steroids influenced gonad morphology and upregulated genes related to cell proliferation，such as RPL35, PC, eLF-1, MPC2, ADCY10 and CYP2C18. Thees upregulated differentially expressed genes were significantly enriched in the organic system, metabolism, genetic information and environmental information categories. These findings imply that steroids may contribute to the growth and the process of genetic information translation and protein synthesis essential for gonadal development and gametogenesis.