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Abstract:Aquatic stock is the basis of cultivation of good strains and the core of biodiversity conservation. To comprehensively grasp of the core Breed Conservation area of China's first national sea cucumber native species farm, this study conducted an annual survey of the water body and sediment environment of the core breed conservation area from July 2022 to May 2023. Simultaneously, the annual change in microbial community structure in seawater and sediment was analyzed using high-throughput sequencing, and the correlation between ecological environment quality and microbial community structure was further completed. The results showed that the water quality of seawater was good, except for the reactive phosphate content in spring and the lead content in summer, autumn, and winter, which reached the water quality standard of ClassⅡand the heavy metal nickel, which met the water quality standard of ClassⅡin autumn. All other water quality indices met the water quality standard of ClassⅠ and the water body of the Breed Conservation area belongs to the poor trophic level. In the sediments, except for the chromium content in autumn which did not reach the standard of ClassⅠ, all other indices met the standard of ClassⅠ, indicating that the ecological environment of the core area is good. The results of microbial community structure analysis of seawater and sediments in four seasons showed that 3 796 OTUs were obtained in the water body of the core area and the Shannon index of the water body in the four seasons ranged from 5.90±0.04 to 6.84±0.01, with the highest in autumn and the lowest in summer; 4 151 OTUs were obtained in the sediment, and the Shannon index of the sediment in the four seasons was between 6.06±1.44 and 7.88±0.22, with the highest in summer and the lowest in spring. PcoA showed that the microbial community structure of the water body and sediments differed in the four seasons and the microbial community structure of the water body varied more by seasonal influences than that of sediments. The results of LEfSe analyses of water and sediment flora in different seasons showed that 73 and 66 significantly different bacteria were screened in water and sediment samples in different seasons, respectively (P<0.05), among which the representative bacteria in the winter sediment contained Vibrionaceae and Psychromonadaceae, which highlights the potential threat of vibrio to the health of sea cucumber in winter. The correlation analysis between environmental factors and microbial community structure showed that temperature, pH, and salinity were the major environmental factors affecting the microbial community structure in the water body, whereas organic carbon, sulfide, heavy metal copper, and lead were the chief environmental factors affecting the microbial community structure in the sediments. These results provide support for assessing the ecological environment of the core area for Breed Conservation, analyzing its risk factors and influencing factors, and providing scientific basis for the efficient preservation of the native sea cucumber in China.

Abstract:The intertidal zone, a dynamic interface between the ocean and land, is one of the most diverse ecological habitats and harbors some of the richest biological communities. However, due to its unique location at the water's edge, it also bears the brunt of human activities in coastal areas. Intertidal biological resources serve as a foundation for the survival of coastal residents and plays a crucial role in coastal economic development. However, the intertidal zone serves as a primary area for coastal zone development and utilization, facing unprecedented environmental pressure resulting from rapid industrialization, urbanization, and agricultural intensification along coastlines. Consequently, safeguarding the intertidal ecological environment and biodiversity has become imperative for China's marine ecological civilization construction efforts. The coastal substrates in Qingdao encompass a diverse range of types， with predominantly rocky, sandy, and silt-muddy coasts constituting its coastline features. As an important marine center city encompassing coastal resorts and international port cities in China, Qingdao has witnessed rapid growth in its marine industry sector, while experiencing mounting pressure on its intertidal ecosystem. Shellfish are the predominant biological group in Qingdao’s intertidal zone, exhibiting significant variation in species composition and community characteristics across different coastal types, thus serving as crucial ecological indicators for this zone. Despite a relatively early start to research on shellfish resources in Qingdao's intertidal zone, recent surveys focusing on shellfish distribution across various substrates are limited. To comprehensively assess the resource status and community structure of intertidal shellfish along the Qingdao coast, this study investigated intertidal shellfish resources at eight representative stations, including rocky, sandy, silt-muddy, and mixed sandy-rocky coastal types in May and October 2022. The composition of shellfish species, dominance patterns, and community structure parameters, including the Shannon-Wiener diversity index, Margalef richness index, and Pielou evenness index, were compared and analyzed at each station. Community clustering analyses were performed using similarity coefficient clustering and non-metric multi-dimensional scaling techniques. Our findings revealed in total 40 identified species of intertidal shellfish along the Qingdao coast. Among the stations surveyed for species richness, Canyon Street exhibited the highest number, with 18 species recorded, whereas Jinkou had the lowest, with only six species observed. The dominant species composition varied among survey stations and showed seasonal variations in type and dominance. During spring, Littorina brevicula, Mya arenaria, and Patelloida pygmaea emerged as the dominant species, whereas Crassostrea gigas, Musculista senhousia, and Ruditapes philippinarum dominated during fall. The Shannon-Wiener diversity index ranged from 0.85 to 1.84 and 0.66 to 2.00, Margalef richness index ranged from 0.86 to 1.97 and 0.62 to 2.96, and Pielou evenness index ranged from 0.41 to 0.84 and 0.31 to 0.98 in spring and autumn, respectively. Qiantaoyuan in Langya, Aoshanwei, and Cangkou exhibited the highest Shannon-Wiener diversity and Margalef richness indices among all stations. Community clustering results varied between spring and autumn at each station. Hongshiyi and Jinkou consistently formed separate branches, Cangkou and Wanggezhuang always clustered together, and the former Taoyuan in Langya and Aoshanwei were consistently grouped together. This study revealed that substrate type significantly influenced the composition and community structure characteristics of intertidal shellfish in Qingdao. This study identifies the status of shellfish resources on various substrates within Qingdao's intertidal zone, providing a foundation for resource development, protection, and ecological restoration efforts.

Abstract:Investigating the effects of different diets on carbon and nitrogen stable isotope turnover and fractionation coefficients in juvenile green crabs (Scylla paramamosain), we used 200 tanks measuring 50 × 40 × 40 cm. Fish (Callionymus richardsoni), shrimp (Penaeus latisulcatus), clam (Ruditapes philippinarum), and polychaeta worms (Perinereis aibuhitensis) were fed to the juvenile crabs whose initial weight is 1.5 g. Samples were collected at 0, 30, 60, 90, 135, and 180 d to determine the δ13C and δ15N and calculate turnover parameters as well as fractionation coefficients. Our findings described that the differences in the juvenile crabs’ δ13C and δ15N during the experimental period were demonstrated by δ13C = a × lnt + b and δ15N = a × lnt + b, where the value of "a" varied significantly with the different diets. Growth is the main factor driving carbon and nitrogen-stable isotopic turnover in juvenile crabs. The contributions of the four diets (fish, shrimp, clam, and crab) to the 13C turnover of the crabs were 79.83%, 83.65%, 84.88%, and 63.80%, respectively, whereas their contributions to the 15N turnover were 81.97%, 82.88%, 75.27%, and 59.80%, respectively. Our findings indicated a clear difference in the stable isotope turnover of juvenile green crabs, which was primarily metabolism-induced. The significant difference in growth rates between the two studies inevitably led to differences in the assimilation and deposition rates of carbon and nitrogen in their body, causing differences in the contributions of growth and metabolism to the turnover of stable isotopes. The time to complete 50% turnover (t50) of 13C and the time to complete 95% turnover (t95) for crabs fed the four diets were 36.30, 24.56, 27.96, 21.17 d and 156.86, 106.16, 120.83, 91.64 d, respectively. The t50 of 15N and t95 for crabs fed the four diets were 37.60, 24.34, 24.77, 20.17 d and 162.49, 105.22, 107.05, 86.99 d, respectively. The results of this experiment indicate that the t50 values of 13C and 15N in juvenile fiddler crabs fed the same diet were similar, and the t95 values were more than four times higher than the t50 values, indicating that the turnover rates of stable isotopes were high in the early stages of the experiment and decreased significantly in the later stages. The fractionation coefficients Δ13C180d for crabs fed the four diets at 180 d ranged from 0.71‰ to 1.64‰, and the Δ15N180d ranged from 2.15‰ to 2.66‰. Overall, the measured value of Δ13C180d is closer to the literature-cited value of 1.3‰, while the measured value of Δ15N180d was incongruent with the literature-cited value of 3.4‰. Since the large range of fractionation factors for stable isotopes in bait materials among consumers in nearshore and estuarine ecosystems, caution should be exercised when citing fractionation factors in related ecological studies, and actual measurements or data from identical or similar species under similar environmental conditions should be used whenever possible. The relationship between Δ13C and mass growth rate (MGR, %) followed the formula Δ13C = a × ln(MGR) + b, and the relationship between Δ15N and MGR followed the formula Δ15N = a × ln(MGR) + b, with significant variations in the values of "a" and "b" when consuming different diets. Δ13C showed a negative linear correlation with the δ13C of the diets, and a positive linear correlation with the difference in δ13C values between the initial juvenile crabs and diets (δ13CIC–δ13CD). Δ15N demonstrated a negative linear correlation with the δ15N of the diets, and a positive linear correlation with the difference in δ15N values between the initial juvenile crabs and diets (δ15NIC–δ15ND). The diet quality and stable isotope content were the main factors affecting fractionation. Although the δ13C and δ15N of the crabs during the experiment approach and ultimately maintain a level higher than that of the diet, the fractionation factor remains influenced by the δ13C and δ15N of the diet. We concluded that growth was the main driving factor for stable isotope turnover in juvenile green crabs and that the stable isotope abundance of the diets was an important factor affecting the fractionation coefficients. The findings of this study provide reference data for nutritional ecology research on green crabs.

Abstract:The leopard coral grouper (Plectropomus leopardus) exhibits captivating chromatic patterns and boasts substantial nutritional content. This species garners significant consumer preference owing to its aesthetic appeal and nutritional richness, thereby conferring considerable economic and aquacultural significance. The seedling and intermediate breeding of the P. leopardus, much like many other groupers, undergoes a process involving the screening of fries and acclimatization to artificial feed. During this process, repeated occurrences of growth differentiation among the same batch of fries, juveniles, and fingerlings are observed, which constitutes one of the principal factors contributing to prolonged cultivation cycles and impacting the economic efficiency of aquaculture. To address this issue, a majority of grouper species undergo a “size grading” process during seedling cultivation, wherein fries are segregated according to different size categories post-screening, thereby enhancing fry survival rates and shortening the overall cultivation cycle. However, “size grading” effectiveness is not only significantly influenced by human factors but also by inherent variations among fry themselves. Despite the considerable influence of environmental factors on this phenomenon of growth differentiation, undefined potential genetic factors remain. Yet, there is a noticeable absence of comprehensive research addressing this pressing issue. However, the quality of growth traits serves as a critical indicator for evaluating the economic value of fish aquaculture. Influenced by both genetic predispositions and environmental factors, these traits warrant investigation into the genetic mechanisms and regulatory strategies governing growth-related characteristics in farmed fish. Such research offers insights into variety improvement and breeding practices within the field. Here, during the artificial breeding and intermediate rearing process of P. leopardus, early juveniles exhibited relatively minor susceptibility to non-genetic effects such as water temperature and quality changes, along with significantly differentiated growth and shorter intervals of divergence. Consequently, we selected muscle tissues of individuals exhibiting differential divergence within three consecutive time points of size screening for early juveniles (42, 70 and 91 days post-hatching) for whole transcriptome sequencing, aiming to elucidate genetic mechanisms. The expression profiles of lncRNAs and mRNAs were acquired, and a Weighted Gene Co-expression Network Analysis (WGCNA) method was used to construct a co-expression network of growth differentiation related lncRNAs and mRNAs. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses of differentially expressed genes within specific gene modules were conducted to analyze the biological functions of the modules. Additionally, Cytoscape software was used to construct a gene interaction networks to identify key candidate genes associated with growth differential differentiation in P. leopardus. The results showed that a total of 6,252 differentially expressed mRNAs and 367 differentially expressed lncRNAs were identified through gene differential expression analysis. Co-expression network analysis revealed three growth differentiation specific gene modules (MEmagenta, MEgreenyellow, and MEblack). GO and KEGG analyses indicated that differentially expressed genes in these specific modules were significantly enriched in biological processes related to muscle protein formation, regulation of skeletal muscle tissue development, and axial tissue formation, participating in metabolic pathways associated with growth differential differentiation such as proteasome, cytoskeletal regulation by actin proteins, and inositol phosphate metabolism. High-weight and high-connectivity lncRNAs and mRNAs were screened from the three specific modules to construct a gene interaction network, revealing key genes associated with growth differentiation including psmd6, nmt1, als2, brcc3, kank1, ada12, at131, hdac3, as well as 20 lncRNA transcription factors such as MSTRG.15660, MSTRG.8694, and MSTRG.1896. Wherein, Psmd6 in zebrafish is encoded by the Volvox mutant, which regulates cellular function by degrading polyubiquitinated proteins, impacting the proliferation of zebrafish lens epithelial cells and the differentiation of lens fiber cells. Mmt1 is an essential gene for mammalian development, serving as a major N-myristoyltransferase during early embryogenesis. Als2 plays a significant role in normal muscle development, and mutations in this gene can lead to autosomal recessive amyotrophic lateral sclerosis and related disorders. The Brcc3, as a cell cycle regulatory gene, participates in the generation of phosphorylated cyclin-dependent kinase activator and biological processes such as cell proliferation. Kank1 regulates actin polymerization, actin stress fiber formation, and cell migration through RhoA signaling. Ada12 (Adam12) plays a crucial role in murine myogenesis and adipogenesis. Adam12 synthesis sequence and amino acid sequence in zebrafish are consistent with mammals and is strongly expressed in the cardiovascular system, erythroid progenitor cells, brain, and jaw cartilage. Zebrafish with knocked out Ada12 genes exhibited reduced body size in infancy without significant morphological defects. At131 (Atp13a1) plays a crucial role in stabilizing the MAVS virus protein in mice. At131 knockout in mice resulted in issues such as growth retardation and embryonic lethality. Hdac3 is essential for liver formation in zebrafish, primarily by inhibiting the growth differentiation factor 11 (Gdf11), which is a negative regulator of cell proliferation and a specific transcription target of Hdac3. Ada12 and At131 knockout leads to defects in growth in zebrafish and mice, directly affecting body size and growth rate. Therefore, Ada12 and At131 are considered as important candidate genes for further research on the molecular regulatory mechanisms of growth differentiation in P. leopardus. This study provides insights into the molecular mechanisms underlying growth differentiation in P. leopardus.

Abstract:Skeletal muscle contraction, which generates movement by pulling on the internal skeleton, is a distinctive mode of movement in vertebrates. Renowned for its flexibility, diversity, and efficiency, this mode of movement is significant for the individual survival and reproductive success of animals. Being the most ancient vertebrates, fish inhabit aquatic environments, where their skeletal muscles serve as structural and locomotor organs and as a crucial source of high-quality protein for human consumption. Based on the contraction characteristics, the skeletal muscles in teleost fishes are primarily categorized into fast-twitch and slow-twitch muscles, which play distinct roles, supporting burst swimming and prolonged endurance swimming, respectively. Preliminary analyses have been conducted on the structural, metabolic, and functional differences between the fast-twitch and slow-twitch muscles in fish at histological, enzymatic activity, and molecular regulatory levels. Proteins, amino acids, fat, fatty acids, and minerals constitute the material basis for the swimming function of fish skeletal muscles, providing a more intuitive and accurate reflection of the distinct physiological characteristics of fast-twitch and slow-twitch muscles. However, reported research on the comparative analysis of the material constituents comprising fast-twitch and slow-twitch muscles is scarce. To comprehend the chemical composition characteristics and elucidate the material basis for the functional differences between fast-twitch and slow-twitch muscles, this study used biochemical analysis to determine the chemical components of the two muscle types in Pseudocaranx dentex and Liza haematocheila. We integrated data from the literature on tuna, including Thunnus tonggol, T. albacares, Auxis rochei, A. thazard, Euthynnus affinis, and Katsuwonus pelamisi. These fishes have different swimming habits, which can provide a more comprehensive perspective on the differences between fast-twitch and slow-twitch muscles. First, the fast-twitch muscles exhibited a substantial enrichment in protein and 12 types of amino acids, particularly histidine. Notably, histidine is pivotal as a proton-buffering substance and for maintaining pH stability. The relative content difference of histidine was pronounced, ranging from 1.22 to 3.83 times higher in fast-twitch muscles than in slow-twitch muscles. Regarding the amino acid compositions, fast-twitch and slow-twitch muscles displayed similarities, with essential amino acids constituting approximately 40% of the total amino acid content. Glutamate and aspartate were the predominant amino acids, playing essential roles in eliminating ammonia during exercise and serving as crucial energy substrates for muscle function. Lysine and leucine, the two essential amino acids with the highest content, were instrumental in ketone body formation, glucose metabolism, and fat metabolism, and provided an essential energy supply. Further analysis of the fat content and fatty acid composition revealed intriguing differences. Slow-twitch muscles exhibited significantly higher levels of fat and each fatty acid than their fast-twitch counterparts. The aerobic oxidation metabolism of fatty acids was characterized by a prolonged energy supply duration and substantial ATP generation. This unique metabolic profile suggests that slow-twitch muscles rely on fatty acids as their primary energy substrate during swimming for extended periods. Examining the fatty acid composition in detail, the proportion of saturated fatty acids (SFA) was higher in slow-twitch muscles, whereas fast-twitch muscles had a higher proportion of polyunsaturated fatty acids (PUFA). This divergence could be attributed to the specific requirements of each muscle type. Slow-twitch muscles, engaged in long-distance movements, necessitate more SFA and monounsaturated fatty acids (MUFA) for oxidative energy supply. Conversely, fast-twitch muscles, responsible for burst swimming, require more PUFA to maintain the structural integrity and functionality of cell membranes. The main fatty acid composition types of SFA, PUFA, and MUFA in the fast-twitch and slow-twitch muscles are the same. C16:0, C18:0, and C14:0 were the main SFA types. C18:1 and C16:1 were the main MUFA types. C22:6n3 and C20:5n3 were the main PUFA types. Finally, the mineral element analysis revealed that slow-twitch muscles possess higher iron and zinc concentrations, which are critical in oxygen transportation and catalyzation of oxidation processes. The potassium, magnesium, and calcium contents showed no significant correlation with muscle types. Potassium was identified as the most abundant constant element, magnesium exhibited minimal content fluctuation across diverse species, and calcium was the most abundant metallic element. In summary, our comprehensive investigation into the chemical composition of fast-twitch and slow-twitch muscles in marine teleost fishes uncovered significant distinctions in proteins, amino acids, fats, fatty acids, and mineral elements. These differences form the foundation for executing diverse swimming functions, shedding light on the intricate interplay between muscle composition and swimming performance in teleost fishes.

Abstract:The pearl gentian grouper (Epinephelus fuscoguttatus ♀×Epinephelus lanceolatus ♂) is the main cultured species of marine fish in factory recirculating aquaculture system (RAS). High density presents the main characteristic of intensive farming model represented by RAS, and density stress is a fundamental factors affecting fish welfare. Density stress causes a series of changes in fish growth performance, digestive and metabolic capacity, oxidative stress states, and endocrine homeostasis. However, research on how the pearl gentian grouper copes with density stress, and the effects of density stress on the growth, digestion, metabolism, oxidative stress, and endocrine production requires further research. Seeking a suitable stocking density can improve the culture efficiency of the pearl gentian grouper and avoid culture risks. Selecting an appropriate stocking density is an important problem to be solved in the factory recirculating aquaculture model. Therefore, this study investigated the changes of growth performance, digestive and metabolic capacity, oxidative stress, and Hypothalamus - Pituitary - Interrenal (HPI) axis related parameters of pearl gentian grouper under different density conditions. Three density gradient groups were set: low-density (LD), medium-density (MD), and high-density (HD). In the LD, MD, and HD groups, 100, 200, and 300 pearl gentian groupers were added per barrel with a density of (3.14±0.13), (6.31±0.13), and (9.56±0.24) kg/m3, respectively. The three experiments were performed in parallel for 60 days. Fish were fed a formula feed of 2.5% of their body weight in the morning and evening (08:00, 17:00) daily. The density stress test was performed for 60 days in a factory recirculating aquaculture system [temperature (27±2) ℃, dissolved oxygen (8.0±1.0) mg/L, pH 7.8 and salinity 24±1]. The body weight, body length, and total length of all the fish were measured every 20 days. A total of 18 juvenile pearl gentian groupers were randomly collected from each density group, for a total of 54. Anesthesia was administered (MS-222, 80 mg/L), tail vein blood was drawn with a 2 mL disposable syringe, and centrifuged (4 000 r/min, 4 ℃, 10 min) to obtain a supernatant that was stored at –20 ℃ for testing. Before testing the corresponding indicators, the brain, liver, intestine, stomach, and kidney were frozen in liquid nitrogen. The results showed that the optimum stocking density of pearl gentian grouper was (15.48–29.67) kg/m3 under the conditions of water [temperature (27±2) ℃, dissolved oxygen (8.0±1.0) mg/L, pH 7.8, and salinity 24±1]. In terms of growth, the specific growth rate and condition factor in the HD group were significantly lower than those in the other groups (P<0.05), and the feed coefficient and coefficient of variation were significantly increased with the increase of density (P<0.05). In addition, the survival rate of the HD group was significantly lower than that of the other groups (P<0.05). Density stress results indicated that the growth retarded, weight difference increased, and mortality increased with increasing density. In terms of digestion and metabolism, the digestive enzyme activities (pepsin, trypsin, lipase, and amylase) in the HD group were significantly lower than those in the other groups (P<0.05). Density stress results in a significant decrease in the digestive performance of the fish. The metabolic enzyme activities (PK, SDH, HK and LDH) in the HD group were significantly higher than those in the other groups (P<0.05). In terms of oxidative stress, the activities of antioxidant enzymes (MDA, SOD, CAT and GSH-Px) in the HD group were significantly higher than those in other groups (P<0.05). In terms of HPI axis, the levels of HPI axis related hormones (CRH, ACTH and CORT) in the HD group were significantly higher than those in other groups (P<0.05). qRT-PCR was used to determine the expression levels of HPI axis-related genes, and the density stress upregulated crhr 1, nr3c 1, and nr3c 2 (P<0.05), while crh-bp was downregulated (P<0.05). In summary, the study revealed that the effects of density stress on growth performance, digestive and metabolic capacity, stress, and HPI axis of the pearl gentian grouper. Density was negatively correlated with the growth performance and caused a significant decrease in digestive enzyme activity, significantly increased metabolic enzyme activity, intensified oxidative stress, and dysregulated HPI axis hormones and related genes. The results of this experiment can provide theoretical reference for the establishment of stocking density for juvenile pearl gentian grouper in the production process. Our findings provide scientific evidence for further understanding the effects of density stress on the growth, digestive metabolism, oxidative stress, and endocrine production of the pearl gentian grouper.

Abstract:Rainbow trout (Oncorhynchus mykiss) is a hypoxia-sensitive fish, and its growth, behavior, metabolism, and immunity are affected in hypoxic environments. To elucidate the effects of hypoxic stress on the heart of rainbow trout, biochemical indices and hypoxia-related gene expression were measured during moderate (4.5±0.1 mg/L) and severe hypoxia (3.0±0.1 mg/L) stress for 4 h, 8 h, 12 h, and 24 h; moderate hypoxia for 1 month (TMM); severe hypoxia for 1 month (TMM); and reoxygenation (8.5±0.1 mg/L) for 12 h and 24 h using enzyme activity assays and quantitative real time polymerase chain reaction (RT-qPCR). The results showed that the pyruvate kinase (PK), total cholesterol (TC), lactic acid (LD), and glutamine aminotransferase (GPT) reactivates increased at 8 h, decreased at 24 h, and were significantly higher than the control levels after reoxygenation (P＜0.05) under moderate hypoxic stress. Under severe hypoxic stress, the succinate dehydrogenase (SDH) activities gradually increased and peaked at 8 h (P＜0.05). No significant difference was observed between the control at 24 h and after reoxygenation (P＞0.05). The adenosine triphosphatase (ATPase), lipase (LPS), TC, glutamic transaminase (GOT), and GPT reactivates decreased at 12 h and recovered to normal levels after reoxygenation (P＞0.05). The PK, TC, lactate dehydrogenase (LDH), and GPT reactivates were significantly higher in the TMM and TMS groups than those in the control group (P＜0.05). The expressions of succinate dehydrogenase gene (sdh), factor inhibiting hypoxia-inducible factor-1 (fih1), and hypoxia-inducible factor-1α (hif-1α) were significantly increased at 8 h compared to those of the control (P＜0.05) and returned to normal reactivates after reoxygenation. Under severe hypoxic stress, lactate dehydrogenase gene (ldh), pyruvate kinase gene (pk), sdh, hif-1α, proline hydroxylase domain protein 2 (egln-1), and von Hippel-Lindau (vhl) expressions were significantly increased at 24 h (P＜0.05) compared to those under moderate hypoxic stress. Compared with those of the control group, ldh, pk, sdh, hif-1α, egln-1, and vhl expressions were significantly decreased in the TMM and TMS groups under moderate hypoxic stress with no significant differences (P＞0.05). pk, sdh, and vhl were significantly increased under severe hypoxic stress (P＜0.05). This study indicated that varying levels of dissolved oxygen led to changes in biochemical indices and in the expression of hypoxic-related genes in the heart of rainbow trout. Hypoxic stress affected the cardiac metabolism of rainbow trout, which affected the normal metabolism level and caused damage. Under hypoxic stress, the rainbow trout were able to provide feedback regulation of hypoxic stress through the high expression pattern of hypoxic-related, which prevents the heart from being in a constant state of hypoxic stress, allowing the organism to quickly return to a stable state and perform its normal physiological function. This study provides basic data to further elucidate the regulatory mechanism of cardiac metabolism in rainbow trout under hypoxic stress, it has guiding significance for the intensive and healthy breeding of this fish and the selection and breeding of new hypoxia-tolerant species in the future.

Abstract:Oxyeleotris lineolata is a valuable economic fish cultured in Australia and countries of Southern Asia. It exhibits sex-related growth dimorphism; the males grow faster and have larger body size than females. Therefore, the efficient development of sex control breeding will be helpful to increase the yield and output value. However, the molecular mechanism regarding sex determination and gonadal differentiation of O. lineolata is poorly understood. Doublesex and mab-3 related transcription (DMRT) factors are known for their contributions in sex determination and differentiation. In this study, the Oxldmrt1 and Oxldmrt3 cDNA sequence was obtained from gonad transcriptome of O. lineolata, identified using PCR. The open reading frame (ORF) of Oxldmrt1 and Oxldmrt3 were 903 bp and 1 363 bp, encoding 300 and 453 amino acids, respectively. OxlDMRT1 belongs to basic proteins, whereas OxlDMRT3 belongs to acidic proteins. The two genes contain a highly conserved DM domain, and OxlDMRT3 also has a DMA domain. Phylogenetic analysis showed that OxlDMRT1 belonged to the DMRT1 family and OxlDMRT3 belonged to the DMRT3 family. The different DMRT families in vertebrates were clustered independently, with the DMRT1 family clustering first followed by the DMRT3 family. The expression levels of two genes in eight tissues of male and female fish were analyzed using RT-qPCR. The results showed that the expression levels of two dmrt genes were most highly expressed in the testis. The expression profiles of two dmrt genes at different developmental stages were analyzed using RT-qPCR. The results showed that the expression levels of two genes were highest in fertilized eggs, the expression level of Oxldmrt1 was the lowest in the optic capsule stage, and Oxldmrt3 was the lowest in the seven days post hatching. The expression and localize of the two genes in the testis were analyzed using fluorescence in situ hybridization (FISH). The results showed that the expression location of the two genes in the testis were identical, and both had strong expression signals in the spermatogonia. In conclusion, Oxldmrt1 and Oxldmrt3 are important in the early embryonic stage and testis development. Oxldmrt1 may also participate in regulating sex determination and differentiation in the late embryonic stage. Oxldmrt3 may also be involved in nervous system development. This study lays a foundation for the molecular mechanism of sex determination and sex differentiation in O. lineolata.

Abstract:Grass carp belongs to Cypriniformes, Cyprinidae, and Cyprinus family, and is typically referred to as the four famous domestic fishes, black carp, silver carp, and bighead carp, respectively. The annual production of Chinese grass carp is consistently among the highest, with an output of 5.905 million tons in 2022, accounting for 24.3% of total of freshwater fish production. Reproduction is the core process of fish reproduction. Normal gonad development is a prerequisite for fish reproduction. At present, there are few reports on the sex differentiation of Chinese grass carp. Studying the differentiation and development of grass carp gonads can provide a typical model for gonadal development of cyprinid fish. In contrast, the start and completion time of gonadal differentiation are one of the key factors inducing sex reversal in fish using hormones. Research on gonadal differentiation and development in grass carp can also lay the foundation for the establishment of a unisexual population of grass carp. Sex-related genes play important roles in biological processes such as sex determination, gonadal differentiation, and reproduction and development in fish. In most differentiated teleost fish, genes such as dmrt1, sox9, foxl2, amh, and cyp19a1a, are considered the key genes involved in sex regulation. cyp19a1a plays a key role in the gonadal differentiation and ovarian development of teleost fish by aromatizing androgens into estrogen via aromatase activity. This gene is specifically expressed in the gonads and is used served as a characteristic gene for sex differentiation towards females. Anti-Müllerian hormone (amh) regulates the structure of reproductive organs and the differentiation and development of reproductive cells. In fish such as carp, rainbow trout, and grass carp, the expression level of the amh gene in the testes is significantly higher than that in the ovaries, and is often used as a characteristic gene for male differentiation. To explore the sex differentiation and developmental patterns of grass carp gonads, this study analyzed the tissue structure and expression differences of sex characteristic genes cyp19a1a and amh in the gonads of grass carp at 1, 2, 3, 4, 5, 6, 12, 24, 36, and 48 months of age using tissue sectioning technology and real-time fluorescence quantification technology. The results of tissue sectioning showed that grass carp exhibited reproductive ridges at 1-month-old. At 2 months of age, primitive germ cells were first observed in the reproductive crest, marking the formation of their primitive gonads. At 3-month- old, ovarian cavities and lobules were observed in the gonads of female grass carp. At 4-month-old, oogonia was observed, indicating anatomical differentiation at 3-month-old and cytological differentiation at 4-month-old. At the age of 4 and 5 months, ductus deferens and spermatogonia were respectively observed in male grass carp gonads, indicating anatomical differentiation at 4 months and cytological differentiation at 5 months. The ovaries of 24, 36, and 48-month-old grass carp were in the first, second, third, and fourth stages of development, respectively, while the testes were in the second, second, third, and fourth stages of development, respectively. At 48 months of age, the gonads of grass carp have matured. The fluorescence quantitative results showed that the expression level of the female characteristic gene cyp19a1a in the ovary showed an overall trend of first increasing, then decreasing, and then increasing again. The expression level of this gene was significantly upregulated at 2 months of age (P<0.05), and reached its peak at 3, 6, and 48 months of age. During these three periods, anatomical changes were observed in the female gonads in the tissue sections, with a large proliferation of oocytes and the gonads tending to mature. The expression level of the male characteristic gene amh in the testes showed an overall trend of first increasing and then decreasing. At 2 months of age, the expression level of this gene was significantly upregulated (P<0.05). At 5 months of age, it reached the peak when there were a large proliferation of spermatogonia in the gonads, and then remained low expression until gonadal maturity. In summary, this study confirmed that the initiation time of grass carp gonadal development is approximately 2 months old, and the differentiation time of male and female gonads is approximately 3 and 4 months old, respectively. By the age of 6 months, both male and female grass carp gonads completely differentiated. These results not only enrich the reproductive physiology data of grass carp but also provide a reference basis for the study of hormone-induced sex reversal technology in grass carp.

Abstract:The thick-shelled mussel Mytilus coruscus belongs to Mollusca, Lanellibranchia, Anisomyaria, Mytilidae, and Mytilus. Due to its flavor and nutritional and high economic values, M. coruscus is a common commercial shellfish in the coastal area of Zhejiang and Fujian in China, and is an important cultured mussel species in China. With the increasing development of coastal areas and the changing habitat conditions, aquaculture industry of M. coruscus is facing problems such as slow growth of juvenile mussels, low meat production rate, and individual miniaturization of adult mussels, which affects the sustainable development of the mussel industry and economic income. Conversely, prebiotics are commonly used exogenous additives in aquaculture. Prebiotics help the organism to absorb nutrients by changing the morphology of the gastrointestinal tract and regulating the composition of the microbial community. They can promote the growth and development of aquatic organisms, enhance resistance to pathogenic bacteria, and improve the absorption and utilization rate of feed to increase the output of aquatic products. As a protein hydrolysate, mussel peptide is also a probiotic element with high nutritional value and multiple functions such as antimicrobial, antioxidant, and immune enhancement. In particular, low molecular weight mussel peptide has the function of anti-lipid peroxidation protective activity and scavenging of excess free radicals, which can be used as a natural antimicrobial food additive. In aquaculture, mussel peptides are considered to be natural active ingredients for treating infectious diseases in marine species. Till date, the relationship between mussel peptides and the growth and development of the thick-shelled mussel M. coruscus remains unclear. Therefore, the optimal concentration of mussel peptide for the potential application of mussel peptides in the M. coruscus aquaculture industry must be determined. In this study, we focused on the effects of mussel peptides on the growth and development of the thick-shelled mussel M. coruscus and on the structural composition of microbial communities. The aim was to provide a theoretical basis for green and efficient aquaculture of the thick-shelled mussel M. coruscus. Here, the thick-shell mussel plantigrades used in the experiment were provided by Donghai Mussel Technology Innovation Service Co., LTD., Shengsi County, Zhejiang Province, China. It was used in the experiment after 1 week of temporary cultivation at 18 ℃ and a salinity of 30. Before the bioassays, these mussel plantigrades were cultured in the lab at 18 ℃ and 30 for 7 d. Five feeding treatment groups and one non-feeding control group were set up in the experiment. The treatment groups were fed mussel peptides at concentrations of 7, 9, 10, 70 and 90 mg/L, and each group was set up with three replicates. On days 7, 14, 21, 28, 42, and 56; 50 plantigrades were randomly selected from different treatment groups for growth measurement including shell length, shell height, and wet body weight. In addition, samples of plantigrades from different treatment groups were collected on days 28 and 56 to analyze the change in microbial communities before and after feeding. The results showed that, compared with the control group, feeding 9 mg/L of mussel peptides could significantly promote plantigrade growth and the shell length, shell height, and wet body weight were increased by 27.37%, 32.35%, and 115.49%, respectively. However, the mussel peptide concentration was too high (70 mg/L and 90 mg/L), which could cause lethal effects on the thick-shelled mussel M. coruscus plantigrades. The microbiome of the thick-shelled mussel plantigrades in the 28-day and 56-day treatment and control groups was analyzed by 16S rRNA gene amplification and sequencing. The results of the study showed that feeding 9 mg/L mussel peptides could alter the structural composition of the thick-shelled mussel plantigrade microbial community, such as increase in the abundance of Bacteroidota and Proteobacteria, and decrease in the abundance of Actinobacteriota. Simultaneously, an increase was observed in the diversity of beneficial bacteria such as Ruegeria, Tenacibaculum, Maribacter, Arenibacter, Octadecabacter, and Shewanella and reduction in the potentially pathogenic bacteria such as Rhodococcus and Aeromonas. Therefore, the appropriate amount of mussel peptides is useful for promoting the growth and development of the thick-shelled mussel M. coruscus plantigrades and optimizing their microbial community structure. In summary, mussel peptides have potential prebiotic functions and have the advantage of easy absorption of small molecules. By changing and adjusting the structure composition of the microbial community, mussel peptides increase the relative abundance of probiotics and reduce the relative abundance of potential pathogenic bacteria in the mussel microbial community of thick-shell mussel plantigrades, and promotes the growth and development of thick-shell mussel plantigrades. The current findings provide a basis and data support for the subsequent cultivation of marine bioactive peptides such as mussel peptides for enhancing shellfish juvenile aquaculture and the sustainable development of marine shellfish aquaculture.

Abstract:In shellfish farms, to guide selection, it is important to rapidly determine the gonadal condition of the parents based on the appearance and morphology of the shells. For the intuitive shell size traits of shellfish, without the intuitive weight traits of the soft body and gonadal wet weights, the shellfish must be dissected and weighed pre-data collection in the laborious shellfish death after dissection is a significant loss to the breeding plant. It is crucial to determine weight trait growth (gonadal wet, soft body wet, and total wet weights) by measuring visual data (shell length, width, and height). Pathway analysis, which was developed by the quantitative geneticist Wright in the 1920s, identifies the correlations between parameters and categorizes their correlation coefficients into direct and indirect influences through other parameters to create the optimal regression equations. Pathway analysis to guide selective breeding of aquatic organisms was achieved for many species using the morphological traits of body mass and soft weight; however, pathway analysis of soft body and gonad wet weights, which are not readily available, was not reported. Mya japonica has a flavor comparable to that of a Crassostrea gigas whose soft body wet weight is heavier than that of an oyster of the same size, with a high meat yield and economic value. The relationship between shell size and weight traits of M. japonica during the breeding period was explored to guide seed shell selection during breeding. In this study, the shell size traits (shell length X1, width X2, and height X3) and weight traits (total wet weight Y1, soft wet weight Y2, and gonad wet weight Y3) of 185 M. japonica from Jiaozhou Bay, Qingdao, were measured, and a path analysis of shell size traits on weight traits was conducted. The results demonstrated that the correlation coefficients of six traits of the six breeding M. japonica traits reached a highly significant level (0.01), with correlation coefficients ranging from 0.891 to 0.966. The direct effects of shell width on the total, soft, and gonad wet weights were 0.462, 0.519, and 0.537, respectively. The influence of the shell width on the total, soft, and gonadal wet weights was the greatest, with values of 21.34%, 26.94%, and 28.84%, respectively. Shell width and height had the greatest degree of co-determination for total, soft, and gonadal wet weights, with values of 32.88%, 24.93%, and 21.34%, respectively. Using the multiple regression analysis method, the optimal regression equation of shell size trait to weight trait was established as Y1=0.295X1+ 1.73X2+1.128X3–72.554, R2=0.954; Y2=0.117X1+0.56X2+0.219X3–19.240, R2=0.927; Y3=0.055X1+ 0.362X2+0.082X3–9.402, R2=0.891. The results demonstrated that when weight were the primary breeding targets, indirect selection could be performed using shell width, and the synergistic effect of shell height could be considered. Our findings provide a theoretical basis for brood selection for breeding M. japonica.

Abstract:The study aimed to investigate the optimal light and temperature conditions, and physiological and biochemical mechanisms for the growth of young sporophytes (3–4 cm) of "Huangguan No.2". We conducted corresponding studies on the growth and physiological and biochemical conditions of young sporophytes under different light intensities and temperatures, including the relative growth rate (RGR), fluorescence parameter of chlorophyll, contents of pigments/proteins, reactive oxygen, antioxidants, and activity of antioxidant enzymes. The results showed that RGR was significantly affected by light, temperature, and their interaction. The degree of influence from large to small is temperature, light intensity and their interaction. They exhibited the highest RGR at 10–13 ℃ and 60–90 μmol photons/(m2·s), whereas the RGR of the high temperature (16 ℃ and 19 ℃) group was highest at 30 μmol photons/(m2·s). At the same temperature, the chlorophyll-a and carotenoid contents of young sporophytes decreased with increasing light intensity. Additionally, the SOD activities and ascorbic acid (AsA) contents tended to increase. Under the same light intensity, compared to 10–13 ℃, the contents of soluble protein decreased in the high temperature group; whereas the contents of MDA and H2O2 significantly increased. Additionally, the SOD, GSH-Px, and APX activities and antioxidant (AsA and carotenoids) contents were all higher. The maximum photochemical efficiency of photosystem Ⅱ (Fv/Fm) was significantly lower in the high temperature and high light [(120 μmol photons/(m2·s)] groups, indicating that the light energy conversion efficiency of the alga was reduced under high temperature and high light stress. In the high light group, the quantum yield of regulated non-photochemical energy loss in photosystem Ⅱ [Y(NPQ)] increased significantly and the initial slope (α) of the fast light curve decreased significantly. This suggests that the photoprotective system of young sporophytes of "Huangguan No.2" responded positively to the high light stress and reduced the absorption of light energy by lowering the efficiency of light energy utilization to reduce the photodamage. The activities of key antioxidant enzymes and the antioxidant contents in the high light and high temperature groups were significantly higher, indicating that the antioxidant systems of young sporophytes responded positively to high light and high temperature stresses to minimize the damage caused by reactive oxygen species. Under low light [10 μmol photons/(m2·s)] group, Fv/Fm, α, actual quantum yield of photosystem Ⅱ [Y(Ⅱ)], the chlorophyll-a contents significantly increased. This indicates that the light energy conversion efficiency and light energy utilization efficiency significantly increased in the low light environment, which led to an increase in the absorption of light energy. These results on the physiological and ecological adaptations of young sporophytes under the conditions of temperature and light intensity can provide a theoretical basis for further optimizing the light and temperature parameters of young sporophytes of "Huangguan No.2" during intermediate culture of young sporelings.

Abstract:As an intracellular protein, tumor necrosis factor receptor-associated factor 7 (TRAF7) is involved in signal transduction and regulates host stress defense. In a previous transcriptome sequencing analysis of the high-temperature-stressed sea cucumber body wall, TRAF7 was significantly differentially expressed between healthy sea cucumbers and ulcered skin in high-temperature-stressed sea cucumbers. However, the structure of the TRAF7 gene and its specific expression pattern under high-temperature stress have not been reported. In this study, based on the results of sea cucumber genome sequencing, the full-length sequence of sea cucumber TRAF7 (named AjTRAF7) cDNA was obtained using RACE technology. Its structural features and distribution in the genome were analyzed, and gene expression was examined in different tissues and under high-temperature stress. AjTRAF7 has a total length of 2,576 bp, an ORF length of 1,770 bp, a 5´UTR length of 345 bp, a 3´UTR length of 461 bp, and encodes 589 amino acids; the molecular weight of the predicted protein is 65.6 kDa, the theoretical isoelectric point (pI) is 7.52, the instability coefficient is 32.34, and the total average hydrophilicity is –0.150. The AjTRAF7 protein contains a RING finger structural domain, a coiled-coil structural domain, six WD40 repeats, and the N-terminus contains a helical region. The predicted protein contained 30.05% α-helices, 6.62% β-turns, 36.84% irregular coils, and 26.49% extended strands and was predicted as a cytoplasmic protein. It had 41, 20, and 3 serine, threonine, and tyrosine phosphorylation sites, respectively, and the N-terminus of the protein had 3 asparagine glycosylation sites. Aligned with the sea cucumber genome sequences, evm.model.chr8.1363 and evm.model.chr8.1742 served as two copies of the gene. Evm.model.chr8.1363 contained 18 exons and 15 introns, and evm.model.chr8.1742 contained 17 exons and 14 introns. The evolutionary tree was constructed with the amino acid sequence of TRAF7. The results showed that the sea cucumber, bat starfish (Patiria miniata), and purple globular sea urchin (Strongylocentrotus purpuratus) clustered into one branch, and the sea cucumber was more similar to the bat starfish and the purple globular sea urchin, at 40.58% and 38.37%, respectively. RT-qPCR showed that AjTRAF7 was expressed in all tested tissues of healthy sea cucumbers. Its expression was highest in female gonads, followed by coelomocytes; it decreased in respiratory trees, body walls, intestines, male gonads, and longitudinal muscles in that order, and the difference in expression among tissues was significant (P<0.05). Under high-temperature stress, AjTRAF7 expression in body wall tissues increased during the first four d and decreased in the subsequent four d. The results indicated that AjTRAF7 might be involved in regulating expression in response to high-temperature stress in sea cucumbers, and the results provide a scientific basis for analyzing the molecular mechanism of high-temperature stress in sea cucumbers.

Abstract:This article reviews the importance of biosecurity in aquaculture and its role in promoting sustainable and high-quality development. As a strategic and comprehensive method based on risk analysis, the core objective of biosecurity is to prevent and control disease risks, which ensures the health and sustainable development of the industry. This article elaborates on the concept and historical development of biosecurity, distinguishing it from biosafety, disease prevention, aquaculture, and animal health while exploring the connections and differences between these concepts. The authors proposed that animal health needs three approaches, including biosecurity, animal welfare, and ecological intervention. Six elements for constructing a biosecurity system and implementing the biosecurity plan are identified. Subsequently, the article outlines the key scientific and technological issues involved in the biosecurity framework and introduces the current state of research and development in the fields of science, technology, and policy, both internationally and domestically. Furthermore, this article discusses the role of global and regional organizations in promoting aquaculture biosecurity strategies and the practices of different countries and regions in constructing aquaculture biosecurity regulatory systems. In particular, it introduces how international organizations such as the Food and Agriculture Organization of the United Nations (FAO) and the World Organization for Animal Health (WOAH) emphasize the application of the biosecurity concept to promote the sustainable development of aquaculture. This article also discusses the evaluation of biosecurity in aquaculture enterprises and its technical content. Using the cultivation of specific pathogen-free (SPF) shrimp seedlings as an example, this article introduces the development of biosecurity in international enterprises and the practices employed by the authors in shrimp hatcheries. Finally, this article highlights the challenges and opportunities for China in the field of aquaculture biosecurity and discusses the future direction of biosecurity strategies. This includes constructing a roadmap for the long-term development strategy of the national biosecurity plan; strengthening research, education, and awareness of biosecurity; prioritizing the development of biosecurity for the aquatic seed industry; and building an aquatic seed industry system with a high standard of biosecurity to ensure the sustainable and high-quality development of aquaculture industry in China.

Abstract:Infectious muscle necrosis (IMN) caused by infectious myonecrosis virus (IMNV) broke out for the first time in Chinese shrimp in 2020, causing substantial economical loss to the shrimp farming industry. To understand the prevalence of Chinese IMNV in recent years, an epidemiological survey of IMNV was conducted in major Chinese shrimp farming areas between 2022–2023. Samples were analyzed using molecular biology and histopathology. A total of 829 samples were collected from Shandong, Jiangsu, Zhejiang, Hainan, Tianjin, Guangxi, Fujian, and Hebei. These samples included major farmed shrimp species, such as Penaeus vannamei, P. japonicas, P. chinensis, and Macrobrachium rosenbergii, as well as live feed, other aquatic economic species, and culture water. TaqMan real-time quantitative fluorescence RT-PCR (TaqMan RT-qPCR) was used for molecular detection. The skeletal muscle of diseased shrimp exhibited the typical plaque or diffuse white necrosis symptom of IMNV infection. IMNV was detected in major farmed shrimp species such as P. vannamei, P. japonicus, and P. chinensis. The positive samples were mainly distributed in Shandong, Hebei, Tianjin, and other provinces or cities around the Bohai Sea. In addition to farmed shrimp, IMNV-positive shrimp live bait (mainly Artemia) and offshore seawater filtered close to the farm was also detected. The IMNV-positive rates of samples collected in 2022 and 2023 were 6.27% (23/367) and 15.80% (73/462), respectively. The IMNV-positive TaqMan RT-qPCR samples were further analyzed using histopathological and in situ hybridization methods. The characteristic coagulation necrosis of IMNV infection was found in the white muscle tissue sections of the ventral and caudal segments of the diseased shrimp, with a clear blue-violet hybridization signal of the IMNV probe in the muscle tissue with pathological damage. The results of this study indicated a high positive rate of IMNV in cultured shrimp, biological live bait, and offshore seawater in many Chinese provinces between 2022–2023. Detection, monitoring, and early warning of IMNV infection in the shrimp farming process should be strengthened to reduce the risk of further spread and IMN epidemics.

Abstract:In a preliminary study conducted in our laboratory, the parkin co-regulated gene (PACRG) was identified as a candidate for white spot syndrome virus (WSSV) resistance using a genome-wide association approach. PACRG is genetically closely linked to the Parkinson´s disease-associated gene parkin, both of which are regulated by a bidirectional promoter. The PACRG and parkin genes have been found to interact with each other, associate with autophagy, and participate in cellular protection. Therefore, the functions of PACRG and parkin in WSSV resistance in Penaeus vannamei were investigated. The mRNA and amino acid sequences were analyzed, and the expression levels in shrimp infected with WSSV at different times and tissues were detected by real-time PCR. Spatial localization was performed using fluorescence in situ hybridization. PCR and Sanger sequencing were employed to obtain single nucleotide polymorphisms (SNPs) and conduct an association analysis of these SNPs with resistance to WSSV. Our findings illustrated that the complete open reading frame (ORF) sequence of PACRG was 600 bp, encoded 199 amino acids, and was predicted to contain the ParcG structural domain. The complete sequence of parkin mRNA was 2,329 bp, comprising a 1,653 bp ORF, 100 bp 5′-untranslated region (UTR), and a 576 bp 3′-UTR, encoding 550 amino acids. Parkin is predicted to contain UBQ and IBR structural domains and a signal peptide structure. Amino acid sequence alignment and phylogenetic tree analysis showed that the homology of PACRG between P. vannamei and Penaeus japonicus was the highest at 89.70% similarity. The phylogenetic relationship of P. vannamei was the closest to Penaeus chinensis and P. japonicus. Thus, PACRG may exhibit high evolutionary conservation. The parkin homology between P. vannamei and P. chinensis was the highest, with a similarity of 93.45%. It has been speculated that the parkin protein exhibits a high degree of evolutionary conservation. Herein, real-time PCR results suggested that PACRG and parkin were expressed in the hepatopancreas, gill, muscle, and eyestalk of healthy P. vannamei, with no significant difference. Following the challenge with WSSV, the PACRG and parkin expression levels in the hepatopancreas, gill, muscle, and eyestalk of P. vannamei were significantly altered. Post-WSSV infection for 48, 96, 192, and 228 h, the PACRG and parkin expression levels in the hepatopancreas of P. vannamei were significantly downregulated. At 48, 72, 96, 144, 192, and 228 h post-WSSV infection, PACRG expression in the gill of P. vannamei were significantly downregulated. However, at 48, 96, and 228 h post-WSSV infection, the parkin expression levels in the gill of P. vannamei were significantly upregulated. Post-WSSV infection at 96, 192, and 228 h, the PACRG and parkin expression levels in P. vannamei muscle were significantly upregulated. Post-WSSV infection, PACRG and parkin exhibited similar expression patterns in the eyestalk. The location of PACRG mRNAs mostly overlapped with the WSSV replication site in the shrimp muscle, suggesting that PACRG plays a functional role in the interaction between P. vannamei and WSSV. Two SNPs were identified within the ORF of the PACRG, One SNP was identified within the ORF of parkin, and one SNP was identified in the UTR of parkin. After conducting association analyses of these SNPs with WSSV resistance, SNPs located in the UTR of parkin-specific SNP3, SNP4, SNP5, SNP7, and SNP9 were significantly associated with resistance to WSSV. This study provides a theoretical reference for future research on the molecular mechanisms underlying P. vannamei’s resistance to WSSV.

Abstract:The golden pomfret (Trachinotus ovatus) is distributed in tropical and subtropical waters such as those of the East China Sea, South China Sea, and the Chinese Yellow and Bohai Seas. The golden pomfret grows rapidly and is the most modernized and intensive marine aquaculture fish in China. Furthermore, the golden pomfret is also the preferred variety of fish for expanding sea aquaculture spaces. Presently, the market mainly includes three methods: Freezing, processing, and live sales. If fish can be processed in multiple ways while maintaining freshness, fresh fish is the best choice. However, the existing transportation of live fish suffers from various problems such as low survival rate, nutrient loss, and short transportation time due to stress, hypoxia, and water quality deterioration. T. ovatus is a warm temperature-loving, omnivorous migratory fish with high oxygen consumption and vigorous metabolism. Fishing exerts high stress on the organism, and they easily die in low temperature environments (<13 ℃). The difficulty of keeping the fish alive and transporting is also the main reason live fish are difficult to find in markets. To solve the above problems, chemical anesthesia, physical dormancy, and other methods are generally used to improve the survival rate of fish and maintain good nutritional quality in the process of keeping them alive and for transportation. Chemical anesthesia may pose risks of drug residue, and there are certain restrictions on the drug withdrawal period for the fish to be transported alive. Among the physical dormancy methods, the ecological ice temperature induced dormancy method is widely used, but it needs low temperature acclimation before treatment, which consumes long time periods. Therefore, an efficient, green and safe way of keeping alive transportation technology is particularly urgent, and corona dormancy presents an environment friendly, safe, new, and efficient way of physical dormancy that meets consumer needs, with broad application prospects. Chemical anesthesia and low temperature-induced dormancy are mostly used in the pretreatment technology of survival and transportation of marine fish, while the research on corona dormancy technology is less, and the research and application of corona dormant T. ovatus have not been reported locally or abroad. In this study, T. ovatus were placed in an electric shock box after 6 h of temporary rearing. The T. ovatus were shocked by pulsed DC currents. The recovery phase was recorded by stages through behavioral observation. The optimal treatment conditions of pulsed DC corona dormancy were optimized by using the dormancy rate, dormancy time, 72 h survival rate and survival time as evaluation indexes through single factor and orthogonal experiments, and the biochemical parameters of serum, brain tissue, muscle and liver are determined indexes of oxidative stress, metabolism, and basic nutrients. The results showed that under the conditions of 20 ℃ water temperature, 140 V voltage, and 4 s treatment time, the dormancy rate and 72 h survival rate of fish could reach 100%, and the survival time was (165.6±42.7) h. After corona dormancy treatment, the contents of glucose (GLU), glutamic oxaloacetic transaminase (GOT), and cortisol (COR) in fish serum significantly increased (P<0.05), and return to normal levels within 4–12 hours of survival, indicating that electrical stimulation can make the life activities of fish become violent, resulting in the rise of stress indicators in a short time. The content of heat shock protein 70 (Hsp70), glutathione S-transferase (GST-S) activity, and catalase (CAT) activity in liver and brain tissues significantly increased (P<0.05), while the content of malondialdehyde (MDA) in brain tissues was significantly decreased compared with that in the control group within 4–72 hours (P<0.05), indicating that the technology can improve the tolerance of fish to environmental stress and reduce the degree of brain damage, reduce the lipid peroxidation in the brain and the accumulation of hydrogen peroxide in the liver, so as to reduce the damage cause by environmental stress and short-term damage to tissues. The content of liver glycogen (Gly) show a downward trend during the preservation process, while the content of lactic acid (LD) in liver and muscle increase significantly (P<0.05), indicating that anaerobic metabolism occurs during the preservation process of fasting, which consume glycogen and produced lactic acid. Crude ash, protein, and fat in fish meat show a downward trend during the preservation process, with the crude fat content decreased the most significantly (P<0.05), and the proportion of decline in the experimental group was reduced compared with that of the control group. The research shows that the appropriate conditions of pulsed DC can induce the dormancy of T. ovatus, and after corona dormancy fish exhibit less stress in the face of external factors. From the index point of view, the technology can improve the release of Hsp70 and the activity of antioxidant enzymes to slow the stress response of the fish under survival stress, reduce tissue damage, and maintain a low metabolic level after survival, reducing the consumption of inorganic matter, fat, and protein. Thus, the efficiency and quality in the process of keeping alive transportation are improving, which is convenient for breeding and transportation. Finally, these findings lay a theoretical foundation for maintaining the vitality and quality of T. ovatus.

Abstract:Scomberomorus niphonius, an important marine economic fish in China, is widely distributed in the Bohai, Yellow, and East China Seas. In 2022, the total catch of Scomberomorus niphonius in China was 356,100 tons, representing 3.75% of China's overall marine fish catch. Shandong Province accounted for a significant portion, with a catch of 175,300 tons, constituting 49.22% of the national total. The annual catch of S. niphonius in Shandong ranks first in the country. S. niphonius is rich in protein, unsaturated fatty acids, essential amino acids, minerals, and other nutrients. It has a delicious flavor and is popular with consumers. S. niphonius, during processing and storage, is susceptible to microorganisms, temperature, and other external environmental influences of corruption deterioration and loss of commodity value; therefore, its storage quality must be improved. Among various fish products, vacuum-packed ready-to-eat soft canned products are popular and widely used with high commercial value due to their short sterilization time, low nutritional loss, and easy portability. Sterilization is an important way of ensuring product quality and extending shelf-life. However, the irrationality of the sterilization process, which focuses mainly on the effect of a single sterilization temperature and time on these quality indicators, often results in a reduction in the quality of the product in terms of flavor, texture, and nutritional value. Lipids are important components of seafood and essential biomolecules. They are also involved in organismal processes essential in the nutritional value, texture, and overall organoleptic properties of seafood. For flavor modification, they act as solvents and precursors of volatile compounds. In addition, seafood contains essential nutritional lipids, including phospholipids, triglycerides, and essential fatty acids, which is an important feature distinguishing it from other species. In addition, most of the volatile flavor compounds in meat are produced by the Melad reaction and the thermal degradation of lipids (oxidation) and thiamine. Oxidative degradation of lipids is the main pathway for forming the characteristic flavor of aquatic products. S. niphonius, due to its high content of unsaturated fatty acids, releases unsaturated fatty acids from the lipids during sterilization to further oxidize them into volatile compounds such as aldehydes, ketones, and alcohols, which have a low threshold value and can significantly impact the flavor quality of the product. Currently, the processing of S. niphonius is focused on dry-cured products, and relatively few studies have been conducted on the quality changes during the processing and sterilization of its soft canning. Based on this, the present study took the S. niphonius as a research object and used conventional physicochemical tests combined with the electronic nose, gas chromatography-ion mobility spectrometry (GC-IMS), and other technical means to compare the effects of different sterilization conditions on the physicochemical indices and volatile substances of S. niphonius, to extend its shelf life while reducing the effects of sterilization on its flavor, sensory, and nutritional qualities. This will provide a reference for quality control in the processing of prepared fish dishes and enrich the basic theory of flavor formation in thermally sterilized aquatic products. The results showed that the sensory scores of texture, taste, and odor of S. niphonius changed significantly after sterilization. The total lipid, triglyceride, and phospholipid contents in the samples were significantly decreased after sterilization (P<0.05), and the free fatty acid content was significantly increased (P<0.05). Differences were observed in lipid composition between different sterilization groups, with the highest degree of lipid loss in Group C samples. The degree of lipid oxidation was significantly affected by sterilization conditions (P<0.05), peroxidevalue (POV) content was significantly increased, and thiobarbituric acid reactive substances (TBARS) content was significantly decreased (P<0.05). The results of electronic nose analysis showed that the odor characteristics of fish meat after different sterilization conditions were significantly different. Twenty-two volatile compounds were identified from the four groups of samples using GC-IMS, including seven aldehydes, five ketones, four alcohols, three esters, two acids, and one furan. Among these compounds, heptyl aldehyde, hexaldehyde, 3-hydroxybutane-2-ketone, acetic acid, and 1-octene-3-alcohol were identified as the primary volatile substances in the control group. The signal intensity of valeraldehyde, isovaleraldehyde, butyraldehyde, 3-hydroxy-butanone, and ethyl propionate significantly increased after the fish underwent various sterilization conditions. This observation suggests that sterilization promotes lipid oxidation and decomposition in fish while leading to significant variations in volatile substance profiles among different sterilization processes. Hexanal, pentanal, 2-methylbutanal, 3-pentanone, cyclohexanone, 3-hydroxybutan-2-one, and ethyl acetate were the major aroma compounds in the fish, and the highest levels were found in the samples from Group B. In conclusion, the organoleptic and flavor qualities of the samples at 115 ℃ for 14.7 min (Group B) were significantly better than those of the other sterilization conditions, and the degree of lipid oxidation was lower, which provides technical support for the fine processing of S. niphonius products.