The yellowtail kingfish(Seriola aureovittata)is a large,fast-swimming fish species characterized by high and aggressive feeding activity,rapid growth rate,and tender flesh.Feed intake is a key factor that influences fish growth and production performance,and appetite is a major determinant of feed intake.To explore the role and potential mechanisms of orexin and its receptor ox2r in the regulation of feeding behavior in SS.aureovittata,we performed homologous cloning to obtain the open reading frame(ORF)sequences of the orexin and ox2r genes in S.aureovittata.Quantitative real-time PCR was used to analyze the expression of these genes across various tissues and developmental stages,as well as their regulatory roles in response to starvation and subsequent refeeding. The results showed that the ORF of orexin in S.aureovittata is 450 bp long,encoding 149 amino acids,including a 46-amino acid signal peptide,a 43-amino acid Orexin A mature peptide,and a 28-amino acid Orexin B mature peptide.The ox2r gene ORF is 1,269 bp,encoding 422 amino acids.Amino acid sequence alignment revealed that orexin and ox2r are highly conserved among teleosts.Phylogenetic analysis indicated that S.aureovittata orexin and ox2r are most closely related to those of Seriola dumerili.The mRNA expression of orexin and ox2r was detected in all 12 examined tissues,with the highest levels observed in the brain,followed by the pituitary gland and stomach.Expression of orexin and ox2r mRNA was also detected in the sperm and eggs,indicating parental inheritance.Both genes are involved in the embryonic and larval development of S.aureovittata,with significant expression changes observed during the transition from endogenous to exogenous nutrition.Starvation stimulated the upregulation of orexin and ox2r mRNA in the brain and pituitary,with expression levels returning to control group levels after 7 d of refeeding. These results suggest that the orexin system may be involved in organ development and feeding regulation during the early growth and feeding of S.aureovittata,thereby providing valuable insights into the mechanisms of feeding regulation in this species.
Often referred to as renowned"ecosystem engineers",oyster reefs perform crucial ecological functions,such as water purification,providing habitat and shelter for other organisms,and stabilizing coastlines.As important components of blue carbon sinks,oyster reefs sequester carbon by absorbing carbon dioxide from seawater and undergoing calcification,biological assimilation,and sedimentation,resulting in carbon fixation.However,global wild oyster resources have declined sharply,leading to the destruction or loss of the ecological structure and functions of oyster reef habitats.Currently,most studies on oyster reefs in China have focused on temperate regions,with relatively few studies on tropical waters,such as the coastal waters of Hainan Island.Investigating the community structure and carbon stocks of oyster reefs along the coast of Hainan Island will be conducive to the protection and restoration of coastal ecosystems in China. To fill this knowledge gap,we conducted sampling and environmental monitoring across 12 oyster reefs along the coast of Hainan Island.Based on preliminary surveys,reef areas were categorized into four habitat types:Estuarine oyster reefs,intertidal natural oyster reefs,mangrove oyster reefs,and coastal engineering area oyster reefs.The research methodology comprised the following steps to ensure comprehensive and accurate data collection.First,aerial imagery captured by drones was used to photograph the sampling sites,and specialized software was used to delineate the reef boundaries and areas.These measurements were integrated with satellite imagery to estimate total reef area and oyster coverage.For specific stations requiring validation,on-site surveys were conducted to verify spatial data.Second,a multi-parameter water quality analyzer was used to determine seawater temperature,salinity,pH,and chlorophyll-a concentration to characterize the aquatic environment of each reef area.Third,depending on habitat types,quadrats of appropriate sizes were established for quantitative sampling,supplemented by qualitative collection in the surrounding areas.After rinsing and screening,the samples were stored at low temperature and transported to the laboratory for further analysis.Fourth,all reef-dwelling organisms were identified and quantified using standard taxonomic guidelines.Oyster samples were identified to the species level using molecular methods,whereas other reef-dwelling organisms were identified at the family level.Species composition,oyster diversity,and proportional abundance,as well as the abundance and wet weight of live oysters and other reef-dwelling taxa were recorded for subsequent community analysis.Lastly,the dried shells,soft tissues,and shell debris from reef-dwelling shellfish were ground,sieved,and stored separately.Carbon content was measured using an elemental analyzer,and oyster reef carbon storage was quantified by applying the standard shellfish carbon sequestration equation. The results of this study were as follows:(1)A total of 53 species of reef-dwelling organisms were identified,belonging to seven phyla,nine classes,18 orders,and 28 families,with Bivalvia and Gastropoda being the dominant classes.(2)Eight oyster species were identified.Saccostrea malabonensis was the dominant species at most survey stations(except Longlou and Guoheyuan)with relative abundances ranging from 6.25％to 100％.Saccostrea mordax was primarily found in intertidal natural oyster reef areas,accounting for 93.75％and 50.00％of its relative abundance in Longlou and Huiwen Town,respectively.Crassostrea iredalei was distributed across all surveyed habitats,except for the intertidal natural oyster reefs.(3)Among the four habitats,the species diversity index(H')and richness index(d)of intertidal natural oyster reefs were higher than those of other habitats,with the highest values recorded at Haitou Town(1.83 for H'and 3.28 for d),followed by estuarine oyster reefs and mangrove oyster reefs,while coastal engineering area oyster reefs had the lowest indices,indicating that intertidal natural oyster reefs provide a more favorable habitat for reef-dwelling organisms.(4)The composition of reef-dwelling organisms and the content of shell remains varied significantly among stations:intertidal oyster reefs were mainly composed of live oysters(12.8％-77.9％)and shell remains(8.7％-86.7％);estuarine oyster reefs and coastal engineering area oyster reefs contained higher proportions of shell remains(67.2％-79.6％);and mangrove oyster reefs had higher biomass of other reef-dwelling shellfish(e.g.,mussels and beach snails,12.4％-28.9％).(5)In terms of biomass and carbon stock of reef-dwelling organisms in different oyster reef habitats,intertidal natural oyster reefs had significantly higher total biomass and total carbon stock than other habitats,with Haitou Town showing the highest value(total live biomass:49,760.86 t;shell remains:46,503.71 t;total carbon stock:14,844.48 tC). Through field surveys and measurements of carbon content in the shells and soft tissues of reef-dwelling shellfish,this study systematically investigated species composition,community structure,and their relationships with environmental factors across different oyster reef habitats.We also quantified the biomass and carbon storage capacities of reef-dwelling shellfish in these habitats.These results provide valuable data to support the protection and restoration of oyster reef ecosystems,particularly in understudied tropical coastal regions such as Hainan Island.The results of this study enhance our understanding of coastal ecosystem dynamics and strengthen the scientific basis for ecological conservation and restoration efforts in China's coastal zones.
Microplastics(MPs)are emerging contaminants and have become a prominent research focus in global environmental science.The Yellow River Estuary,a nationally significant estuarine wetland and ecological conservation zone in China,is a critical area for investigating MP pollution in intertidal environments.The study of contamination characteristics of MPs in this region advances the understanding of terrestrial MP transport processes in the marine environment and provides a scientific basis for ecological risk assessments of MPs in estuarine systems.This study systematically analyzed the distribution patterns,morphological characteristics,and sources of MPs(53-5 000 μm)within surface sediment samples collected from intertidal zones within and outside the Yellow River Delta,whilst assessing their ecological risks.The results demonstrate distinct spatial variations in MP abundance:The inner delta exhibited a decreasing gradient from east to west(range:60-520 items/kg;mean:257.5±166.6 items/kg),whereas the outer delta displayed an inverse pattern(range:160-880 items/kg;mean:360.0±249.8 items/kg).This distribution pattern is primarily governed by the dispersion of the Yellow River freshwater plumes under the influence of counterclockwise circulation in the southern Bohai Sea.Morphological analysis revealed that fibrous MPs were the predominant MP type(66.0％in the outer delta;50.6％in the inner delta),followed by granular fragments(17.2％in the outer delta;25.9％in the inner delta),with small MPs(＜1 mm)dominating both regions(57.5％in the outer delta;74.0％in the inner delta).The polymer composition showed regional differentiation;the outer delta was dominated by cellulose(43.7％)and polyethylene terephthalate(18.8％),primarily derived from domestic wastewater,whereas the inner delta exhibited comparable proportions of cellulose(23.5％),polyethylene terephthalate(23.5％),and polyvinyl chloride(23.5％).This indicates the predominance of industrial-source secondary MPs via fluvial transport.The pollution load index(PLI)assessment indicated mild overall contamination levels(PLI＜1)across the study area,with higher contamination in the inner delta(PLI=0.765)than in the outer delta(PLI=0.629).Stations B5 and B6-2 reached moderate levels of pollution.These findings have significant implications for ecological conservation strategies in the Yellow River Delta.
To investigate the function of the tumor necrosis factor receptor-associated factor(TRAF1)gene in black seabream Acanthopagrus schlegelii under low-temperature stress,the full-length cDNA sequence of TRAF1 in black seabream was obtained using RACE cloning technology.Gene structure was analyzed using bioinformatics tools,and the expression levels in various tissues of black seabream.Gene expression patterns under low-temperature stress were analyzed using qPCR.The SNP sites of TRAF1 related to low-temperature tolerance in black seabream were screened by direct sequencing.Genetic polymorphism analysis was conducted on the selected SNP loci,including genotype typing,correlation analysis for low-temperature tolerance,association analysis between SNP loci and the expression level of TRAF1 in black seabream,and protein structure prediction encoded by the coding region loci.The results showed that the full-length cDNA of TRAF1 in black seabream was 3,528 bp,including a 5'UTR of 93 bp,an ORF of 1,566 bp,and a 3'UTR of 1,869 bp,encoding 521 amino acids,including 10 exons and 9 introns,with a theoretical isoelectric point of 7.67 and predicted molecular weight of 58.91 kDa.A total of 65 negatively charged residues(Asp+Glu)and 67 positively charged residues(Arg+Lys)were identified.The fat and stability indices were 77.89 and 42.72,respectively,and were classified as unstable proteins.The hydrophobicity was-0.375 and the protein domain was 373-498 bp.Subcellular localization was observed in the extracellular matrix.This protein had no signal peptide or transmembrane region and the entire protein was located outside the membrane.In addition to one ring finger domain and one MATH domain,there were 29 serine phosphorylation,12 threonine phosphorylation,3 tyrosine phosphorylation,1 N-terminal glycosylation,and 1 C-terminal glycosylation sites.In the black seabream,TRAF1 expression was detected in various tissues,including the liver,brain,muscle,gills,kidney,heart,and intestine,with the highest expression level in the kidney tissue,which was significantly higher than that in other tissues(P＜0.05);the lowest expression was observed in the brain.Under low-temperature stress,compared to the control group,the expression level of TRAF1 was high in the low-temperature tolerant group and decreased in the tissues of the heart,spleen,muscle,brain,and liver in the low-temperature sensitive group.By comparing the full-length DNA sequences of TRAF1 between the tolerant and sensitive black seabream groups,16 SNP loci were identified through screening.Among them,two SNP loci were located in the exon region and 14 were located in the 3'UTR region.The SNP g.5862 locus in the exon region and the SNP g.7827 locus in the 3'UTR region were significantly correlated with the low temperature tolerance trait of black seabream(P＜0.05).At the SNP g.5862 and g.7827 loci,individuals with the GG genotype had significantly lower death temperatures than those with the AA and AG genotypes(P＜0.05).At the SNP g.8229 locus,individuals with the GG genotype had significantly lower death temperatures than those with the CC and CG genotypes(P＜0.05).Quantitative real-time fluorescence detection revealed that among the three SNP loci of TRAF1 in the black sea bream,the GG genotype at each locus exhibited significantly higher gene expression levels than did the other two genotypes(P＜0.05).The SNP at g.5862 was a non-synonymous mutation in which glycine was changed to serine,which in turn caused alterations in the secondary and tertiary structures of the protein.Specifically,it was characterized by a reduction inα--helices and β-sheets,and an increase in random coils.This SNP can be used as a candidate marker for the molecular marker-assisted breeding of black sea bream having low-temperature tolerance traits.
Black rockfish(Sebastes schlegelii),a key marine species cultivated in the Yellow Sea and Bohai Sea regions,is valued for its rapid growth,high-quality flesh,and robust resistance to diseases and cold environments.However,climate change has prolonged periods of high ambient temperatures in China during the summer,causing a sustained increase in coastal water temperatures,which has increased black rockfish mortality rates.Persistent high temperature stress can induce heat stress in fish,posing a significant challenge to the sustainable development of the aquaculture industry.Therefore,given its sensitivity to environmental stress,this study examined the hepatic stress response under gradient thermal stress in black rockfish. A total of 108 fish were used in the experiment,with an average total length of(20.63±0.72)cm and an average body weight of(117.96±10.93)g.Each group was equipped with three parallel tanks,with six fish per experimental tank(80 L).Except for the control group,the temperature of all five experimental groups was gradually increased by 1℃ every 12 h starting from 20℃ to 23℃,26℃,and 29 ℃.Temperature changes in the water were monitored and recorded,and the normal dissolved oxygen content in the water was maintained.Under gradient thermal stress,the experimental results showed that the livers of black rockfish underwent significant damage,including cellular vacuolization and nuclear migration.The increase in temperature leads to cumulative damage to liver tissue,with the most severe damage occurring at 29℃.The content of malondialdehyde and the activities of alanine aminotransferase and aspartate aminotransferase significantly increased(P＜0.05),reaching their maximum values at 29℃ with increasing temperature,which were 26.6 nmol/mg prot,67.6 U/g prot,and 59.6 U/g prot,respectively.Oxidative damage to the liver induces an increase in antioxidant enzyme activity,with significant increases in superoxide dismutase,catalase,and glutathione peroxidase activities(P＜0.05),reaching their maximum values at 29℃,which were 178.7 U/mg prot,3.8 U/mg prot,and 22 U/mg prot,respectively.With the increasing of temperature,the expression of liver heat shock protein genes(hsp70a and hsp90a)was significantly up-regulated(P＜0.05)to maintain cellular protein stability in the black rockfish.At the same time,gradient thermal stress activated the up-regulation of pro-apoptotic genes(p53,bax,and caspase3)expression(P＜0.05),and while the down-regulation of anti-apoptotic gene(bcl-2)expression(P＜0.05),leading to liver cell apoptosis.TUNEL staining of the liver tissue revealed a significant increase in the rate of cell apoptosis rate with increasing temperature(P＜0.05).Gradient thermal stress,the liver of black rockfish suffered severe damage and experienced oxidative stress.To resist high-temperature stress,antioxidant enzyme activity in the liver of black rockfish is enhanced to alleviate oxidative damage.The upregulation of heat shock protein genes(hsp70a and hsp90a)helps repair damaged proteins and maintain cellular environmental stability.However,under high-temperature stress,the liver undergoes apoptosis that intensifies with increasing temperature.After the water temperature was restored,the conditions of liver damage and oxidative stress,along with the expression of heat shock protein-and apoptosis-related genes gradually recovered,indicating that black rockfish have a certain degree of self-regulation and recovery ability.These results contribute to a deeper understanding of the physiological response mechanism of the liver in black rockfish under high-temperature stress and provide a theoretical basis and data support for sustainable and healthy aquaculture management of black rockfish.
Listeria monocytogenes is a significant foodborne pathogen responsible for zoonotic diseases and is considered one of the most important contaminants in raw aquatic products.Infection with L.monocytogenes can cause listeriosis,a serious disease that primarily affects pregnant women,newborns,the elderly,and other immunocompromised individuals,with a high mortality rate of 20％-30％.Salmon,an important raw aquatic product,is a high-risk food often contaminated with L.monocytogenes.Raw salmon products are often transported and stored at low temperatures.L.monocytogenes can survive refrigeration,and if raw salmon products are directly consumed without heat treatment,L.monocytogenes may be present,posing a significant health risk to consumers.In recent years,there have been multiple outbreaks of listeriosis worldwide,caused by the consumption of raw salmon products,resulting in severe illness and economic losses.Sequence type 18(ST18)L.monocytogenes has been associated with outbreaks of listeriosis;however,there are few reports of ST 18 strains.Thus,it is important to clarify the characteristics and genetic diversity of ST18 L.monocytogenes in raw salmon products,which is crucial for listeriosis prevention and control and for enhancing the food safety of raw salmon products.Whole-genome sequencing(WGS)technology is a powerful tool for investigating microbial disease outbreaks,offering high efficiency,accuracy,and comprehensive genomic data.In this study,WGS was applied to elucidate the characteristics,genomic features,molecular types,virulence genes,and antimicrobial resistance gene distribution of ST 18 L.monocytogenes in raw salmon products.Phenotypic characterization,including cytotoxicity assays,antimicrobial susceptibility testing,and serotyping of L.monocytogenes strain LMSUR02 isolated from raw salmon products,was performed.The cytotoxicity and antimicrobial susceptibility of the isolates were evaluated by determining lactate dehydrogenase released from Caco-2 cells and by the disc diffusion method,respectively,and the serogroups were identified using multiple PCR technologies.Subsequently,WGS of LMSUR02 was conducted.Sequencing data were predicted and annotated using the relevant software,and comparative genomic analyses with other L.monocytogenes isolates from different regions and years were conducted.The results indicated that LMSUR02 exhibited significantly higher cytotoxicity(66.53％)than the reference strain ATCC19115.LMSUR02 contained two major Listeria pathogenicity islands(LIPI-1 and LIPI-2),whereas LIPI-3 and LIPI-4 were absent.LMSUR02 was susceptible to most antimicrobials tested,showed intermediate resistance to ofloxacin and ciprofloxacin,and displayed resistance to cefuroxime sodium(CXM),ceftriaxone(CRO),cefepime(FEP),and nalidixic acid(NA).The strain carried antimicrobial resistance genes norB,lin,and FosX.Comparison of the antimicrobial resistance phenotype and genotype showed that LMSUR02 is resistant to CXM,CRO,FEP,and NA;however,the corresponding resistance genes were not detected.This suggests that nonspecific efflux mechanisms may contribute to resistance.Serotyping classified LMSUR02 as serotype I.1(1/2a,3a).Sporadic and outbreak listeriosis cases are predominantly caused by serotypes 4b and 1/2a,respectively.Given the increasingly strong association between serotype 1/2a and listeriosis,it has been suggested that LMSUR02 carries a relatively high risk of listeriosis and should be considered seriously.The assembled LMSUR02 genome was 2,945,085 bp,harbored one CRISPR sequence,three genomic islands,and two prophages.LMSUR02 belongs to sequence type CC18(ST 18).In previous studies,CC18 strains were detected in both environmental and clinical isolates,indicating that LMSUR02 has a potential risk of inducing listeriosis,which should be given more attention.L.monocytogenes strains can be phylogenetically categorized into four evolutionary lineages(Ⅰ,Ⅱ,Ⅲ,and Ⅳ),with lineage Ⅱ being the most prevalent among isolates from food and environmental sources.LMSUR02 originated from the evolutionary lineage Ⅱ,consistent with previously reported findings.Comparative genomics indicated a close evolutionary relationship between LMSUR02 and nine other strains-IZSAM_Lm_15_17439_A144(GCA_001513695.1),Lm_N1546(GCA_001483445.1),HPB913(GCA_001913195.1),LM26(GCA_016757715.1),J2-031(GCA_000438645.1),2015TE19005-1355(GCA_001551855.1),B-33260(GCA_016801005.1),FW040025(GCA_001644625.1),and Lm_3136(GCA_001483425.1)—with strong genomic synteny among all ten strains.The genomic module compositions of the ten strains were similar,all containing eight collinear modules;however,the specific module positions and arrangement sequences in the genome were different.Among the nine strains,Lm_3136(GCA_001483425.1)showed the closest evolutionary relationship with LMSUR02.In summary,this study clarified the characteristics of an ST18 L.monocytogenes strain from raw salmon products and enriched the genome database of L.monocytogenes,providing an important theoretical basis for the prevention and control of L.monocytogenes infections associated with raw aquatic products,including raw salmon products.
Closed shallow lakes represent unique and fragile aquatic ecosystems characterized by limited hydrological exchange, which notably shapes their community structure and functional processes. Scientifically elucidating the structural characteristics of such ecosystems and evaluating the ecological carrying capacity for key species are fundamental prerequisites for sustainable fisheries management and aquatic ecological restoration. Huangtai Lake, as a typical closed shallow lake, has undergone marked anthropogenic influences, including long-term stocking practices. However, a systematic understanding of its food web structure, energy flow pathways, and the sustainable thresholds for stocked species has been lacking, hindering the development of science-based management strategies. Thus, this study aimed to reveal the structural characteristics of the Huangtai Lake ecosystem, particularly its trophic level distribution and energy flow patterns; quantify the ecological carrying capacity for major stocked species, including silver carp (Hypophthalmichthys molitrix), bighead carp (Aristichthys nobilis), mandarin fish (Siniperca chuatsi), oriental river prawn (Macrobrachium nipponense), and crabs (Eriocheir sinensis); and assess the interspecific resource competition, especially among filter-feeding fishes, to provide a scientific basis for optimized stocking regimes and ecosystem-based management. Based on comprehensive environmental and fishery resource survey data collected from Huangtai Lake in 2024, a mass-balanced Ecopath model was constructed. The model comprised 17 functional groups, encompassing primary producers (phytoplankton), primary consumers (zooplankton, benthic organisms), secondary and tertiary consumers (various fish species, including key stocked species), and detritus. The model was employed to analyze the trophic structure, quantify energy flows between trophic levels, calculate key ecosystem indices, and estimate the ecological carrying capacities of the target species. Scenario simulations were further conducted to assess the impacts of the coexistence of multi-species on carrying capacities. For trophic structure and energy flow, trophic levels of the biological community in Huangtai Lake ranged from 1.000 to 3.287. Filter-feeding fishes, namely silver carp and bighead carp, served as critical nodes in energy transfer, highlighting the characteristic of the ecosystem of being dominated by lower trophic levels and possessing relatively short food chains. The total primary production was estimated at 8,726 t/(km2 ·a); however, the energy transfer efficiencies were notably low: only 1.57% from trophic level Ⅱ to Ⅲ and 3.79% from Ⅲ to Ⅳ—indicating severe constraints on energy delivery to higher trophic levels. For ecosystem state indicator, the system-level indices derived from the model suggested that Huangtai Lake is at an early developmental stage. The ratio of total primary production to total respiration was 1.856, Finn's cycling index was low at 4.506%, the calculatedconnectance index for the system was 0.263, and the system omnivory index was 0.116, reflecting a weak capacity for energy recycling within the system. These metrics, combined with a relatively low system omnivory index, depict an ecosystem with a simple structure, high dependence on primary production, and limited internal complexity. For ecological carrying capacity, the estimated capacities, under the modeled conditions, were 0.203, 10.906, 30.653, 2.802, and 0.794 t/km2 for mandarin fish, silver carp, bighead carp, oriental river prawn, and E.sinensis, respectively. For the impact of species interaction, scenario simulations considering multi-species coexistence revealed notable resource competition between silver carp and bighead carp due to substantial dietary overlap. When coexisting, their maximum sustainable biomasses decreased to 7.993 and 12.199 t/km2 , respectively, underscoring the necessity of considering interspecific competition in stocking plans. The Huangtai Lake ecosystem is characterized by energy concentration at lower trophic levels, inefficient energy transfer to apex consumers, and structural simplicity indicative of an immature and potentially vulnerable state. Filter-feeding fishes play a pivotal but competitive role; therefore, the management of stock enhancement must strictly adhere to the ecological carrying capacities estimated under realistic multi-species scenarios. Particular attention should be paid to controlling the combined biomass of silver carp and bighead carp to prevent excessive grazing pressure on plankton communities, which could destabilize the entire ecosystem. This study provides the first quantitative Ecopath model for Huangtai Lake, offering a systematic and holistic analysis of its ecosystem structure and function. The estimated species-specific ecological carrying capacities and the revealed competitive dynamics offer concrete, science-based benchmarks for fisheries management. The findings advocate for a shift from traditional empirical stocking towards an ecosystem-based approach that respects ecological thresholds and species interactions. The findings deliver direct, actionable quantitative benchmarks for the management of Huangtai Lake and analogous water bodies. Regarding precision management, the estimated ecological carrying capacities for each species, especially the adjusted capacities for silver and bighead carp considering competition, establish clear biomass ceilings for formulating science-based annual stocking plans. This facilitates a shift from traditional, experience-based stocking towards a precision management paradigm constrained by ecological carrying capacity. Regarding optimized species composition, the findings unambiguously highlight marked competition between filter-feeding fishes, serving as a critical warning for managers to coordinately plan the stocking ratio and total biomass of silver carp and bighead carp. Overstocking driven by the pursuit of a single-species yield must be avoided to prevent plankton resource depletion, interspecific imbalance, and diminished overall stocking efficacy. Concurrently, capacity estimates for species like mandarin fish, prawns, and crabs reveal opportunities for developing diversified, complementary eco-fishery models. Regarding informing ecological restoration goals: silver carp and bighead carp are classic species used in biomanipulation to control algal blooms. This study defines their sustainable carrying capacity, implying that their algae-control function must be harnessed within ecologically safe limits to prevent ecosystem distortion. The low Finn's cycling index and identified simple structure call for management strategies that enhance ecosystem complexity and resilience, such as the conservative introduction of appropriate omnivorous or benthic species to strengthen detrital pathways and internal recycling. This work serves as a crucial scientific foundation for achieving sustainable fisheries yield, maintaining water quality (through biomanipulation via filter feeders), and promoting the overall health and stability of the ecosystem of Huangtai Lake. The methodology and insights revealed are also relevant for the management of similar closed shallow lake ecosystems elsewhere.
Microplastics(MPs),widely detected in marine environments worldwide,have emerged as a new class of pollutants that pose a major threat to the stability of marine ecosystems and the security of fisheries.MPs can accumulate in marine organisms through ingestion,gill filtration,and trophic transfer,thereby directly compromising the edible quality of marine seafood.Although several reviews have previously addressed the potential health risks to humans associated with MP accumulation in seafood,systematic analyses of how MPs affect the nutritional value and flavor quality of marine aquatic products are still lacking.This review comprehensively elucidates the major sources of MPs in marine environments and their routes of bioaccumulation,and focuses on the mechanisms by which MPs affect key nutritional components(proteins,fatty acids,and vitamins)of aquatic products through the core pathway of"oxidative stress-energy redistribution-gut microbiota dysbiosis."Furthermore,from the perspectives of exogenous introduction of off-flavors,endogenous metabolic disorder,and physiological imbalance of organisms,this review reveals the potential routes through which MPs can cause the deterioration of flavor characteristics in seafood.These processes synergistically lead to a decline in both nutritional value and flavor quality,ultimately reducing the market value of marine products.Finally,this review proposes future research directions regarding quality and safety standards,and prevention and control technologies for MP contamination in marine aquatic products.This review aims to provide theoretical support for the development of effective MP risk prevention strategies and to promote the sustainable development of the"blue granary."
Microplastics(MPs)and engineered nanoparticles(ENPs)are two typical types of emerging marine pollutants that have attracted widespread attention in recent years.However,the specific mechanisms through which MPs influence the bioavailability,absorption,distribution,metabolism,and final in vivo fate of ENPs in marine organisms remain unclear and require further investigation.In this study,the filter-feeding bivalve Scapharca subcrenata,which is widely distributed in the coastal waters of China.Non-spherical polyethylene microplastics(PE-MPs)with a size range of 53-106 μm were prepared by mechanical crushing and sieving from commercially available common red plastic bags.Titanium dioxide nanoparticles(TiO2NPs)with an average particle size of 21 nm were used as the representative of ENPs.To explore the combined effects of the two pollutants,four experimental groups were established:A blank control group without any pollutant exposure,a single TiO2NPs exposure group,a single PE-MPs exposure group,and a combined exposure group with PE-MPs and TiO2NPs.Histological section and transmission electron microscopy observations together with detection and analysis of key oxidative stress biomarkers-such as superoxide dismutase,catalase,and malondialdehyde-were used to evaluate the effects of combined exposure on the physiological and biochemical status of S.subcrenata.The results showed that TiO2 accumulated significantly in the body of S.subcrenata after exposure,and the digestive glands and gills,which are the main organs for material absorption and gas exchange in bivalves,were identified as the main target organs of TiO2 accumulation.In addition,the presence of PE-MPs significantly slowed the accumulation rate and accumulation of TiO2 in the body of S.subcrenata during the exposure period.Meanwhile,it significantly inhibited the excretion and depuration of TiO2 accumulated in the body during the depuration stage.Histopathological observations revealed that the combined exposure to PE-MPs and TiO2NPs caused structural damage to the digestive gland and gill tissues of S.subcrenata.This manifested as tissue looseness,cell deformation,and cell necrosis and was accompanied by a significant increase in the number of lysosomes in the cells and obvious damage to the lysosomal membrane structure.Analysis of oxidative stress markers indicated that long-term exposure to the combined pollutants led to an imbalance in the antioxidant defense system in S.subcrenata,which further induced serious lipid peroxidation damage in the tissues.Collectively,these findings reveal a complex interaction mode in which PE-MPs adsorb TiO2NPs on their surface,thereby affecting the bioavailability and TiO2NPs fate in vivo and ultimately exacerbating the physiological and histological damage caused by S.subcrenata.This study provides an important scientific basis for the accurate assessment of the ecological risks caused by the combined pollution of MPs and ENPs in marine environments and lays a foundation for subsequent in-depth research on the toxicological mechanisms of marine composite pollutants.
Surimi is a source of myogenic fibrous protein derived from processed fish flesh,the fish species in the final product cannot be identified using morphological and sensory methods,determining the species of fish used in surimi is essential for quality assurance.Black carp,grass carp,silver carp,and bighead carp used in marine fish surimi production can enhance whiteness and texture,increasing surimi's grade and price.Some manufacturers add them without labeling,misleading consumers and disrupting market order.Therefore,developing an accurate,sensitive,and rapid method to identify these species in marine fish surimi is crucial. In this study,we designed a pair of specific primers/probes based on the mitochondrial NADH dehydrogenase subunit 5 gene for simultaneous detection of four freshwater fishes and developed a crystal digital PCR method.This method enabled quantitative detection of black carp,grass carp,silver carp,and bighead carp in marine fish surimi.The reaction conditions were optimized via an orthogonal experimental design,and the method's specificity and sensitivity were validated.Fifty commercially available surimi products were tested to verify its practicality in detecting target species. The results indicated that our method was highly specific.Furthermore,amplification of sample fish DNA(0.2 ng/μL)generated copy numbers of 638.8,402.5,785.3,and 627.5 copies/μL for black carp,grass carp,silver carp,and bighead carp,respectively,with no cross-reactivity to 24 other fish species in the crystal digital reaction.Determination of the lower limit of quantification demonstrated that the method could detect sample DNA at a minimum concentration of 0.32 pg/μL for black carp,grass carp,and silver carp and 0.064 pg/μL for bighead carp.Minimum of copy numbers of 1.43 copies/μL were detected for black carp,2.87 copies/μL for grass carp,2.69 copies/μL for silver carp,and 1.72 copies/μL for bighead carp.Among 50 commercial samples,31 tested positive and 19 tested negative for these species in dPCR analysis.The method could quantify the copy numbers of components derived from each of the four species.The qualitative results were consistent with those obtained using DNA barcoding technology.We developed a sensitive and rapid crystal digital PCR method for quantifying ingredients derived from black carp,grass carp,silver carp,and bighead carp in marine fish surimi.This study provided an effective technical method for industry authorities and testing agencies to monitor the compliance labeling of freshwater fish components in marine fish surimi,ensuring transparency in surimi product authentication.
Lithophaga has significant ecological value in coral reef ecosystems.However,publicly available mitochondrial genome information for this genus remains extremely limited.In this study,the mitochondrial genomes of Lithophaga teres and Lithophaga zitteliana were newly measured,and their applications in phylogenetic and population genetics were investigated.The mitochondrial genomes of L.teres and L.zitteliana are both double-stranded circular molecules,which are 35,469 bp and 38,333 bp,respectively,and both are composed of 2 rRNAs,22 tRNAs and 13 PCGs.Phylogenetic analyses of Mytilidae revealed that Lithophaga forms a monophyletic clade and is distantly related to Leiosolenus lischkei,which clusters closely with Limnoperna fortunei.Substantial differences were observed between Lithophaga and Leiosolenus in PCG composition(with L.lischkei lacking atp8)and in the arrangement of functional genes,supporting the removal of Leiosolenus from the subfamily Lithophaginae.Furthermore,selection pressure analyses of mitochondrial PCGs in L.teres and L.zitteliana indicated that the nad2 gene is subject to relatively weak selective pressure(Ka/Ks＞0.5)and possesses a relatively long sequence(921 bp),providing abundant informative sites and making it a suitable molecular marker for population genetic studies.Using 748 bp fragments of cox1 and 425 bp fragments of nad2,we assessed the genetic diversity of three wild populations of L.teres from the South China Sea.The Danzhou population exhibited higher genetic diversity and greater genetic divergence from the other populations,indicating pronounced population differentiation.Neutrality tests and mismatch distribution analyses further suggested that all three populations have undergone recent population expansion.
The intestinal microbiota is instrumental in the nutritional absorption,immune regulation,and overall health of aquatic animals.However,limited information exists on the gut microbiota of economically important marine fish such as greenfin horse-faced filefish(Thamnaconus septentrionalis),especially across different developmental time points.Understanding microbiota succession in this species is crucial for improving larval rearing success and optimizing feed strategies. This study systematically investigated the temporal changes and community structure of the intestinal microbiota of T septentrionalis larvae and juveniles at eight representative developmental stages(G0-G50),from fertilized eggs to advanced juvenile stages.The primary objective of this study was to explore the dynamic succession of the gut microbiota and evaluate its relationships with host age,physiological state,and feed transitions(rotifer,Artemia nauplii,and frozen Artemia).We also aimed to identify the dominant and stage-specific bacterial taxa associated with different gut ecological environments during development.High-throughput 16S rRNA gene sequencing was performed on 24 gut samples at eight time points.Alpha diversity was assessed using Chao1 and Shannon indices to quantify richness and diversity.Beta diversity analysis,including non-metric multidimensional scaling and unweighted UniFrac distances,was used to evaluate community dissimilarities across stages.Linear discriminant analysis effect size(LEfSe)was employed to identify significantly enriched bacterial taxa at different stages.In addition,heat maps and cluster analyses were used to visualize the taxonomic shifts and abundance gradients of the key genera.The results revealed that the gut microbial community of T.septentrionalis underwent distinct compositional shifts during development and exhibited a phased and directed successional pattern.Alpha diversity indices showed that microbial richness was the lowest in the G0 stage(fertilized eggs),characterized by sparse colonization,mostly originating from the egg surface and surrounding water.Following the initiation of exogenous feeding at G1(rotifer ingestion),both the Chao1 and Shannon indices increased sharply,peaking at G3.This increase corresponded to microbial input from live feeds.Diversity began to fluctuate during the G9-G30 stages and gradually stabilized by G40-G50(frozen Artemia stage),suggesting the establishment of a selective and stable gut microenvironment.Beta diversity analysis further supported these findings.The samples from G0 were clearly separated from those from other stages,reflecting a unique early stage microbial signature.A transitional shift was observed from G3 to G30,and samples from G40 to G50 clustered more closely,indicating microbiota convergence and ecological stabilization during the later stages.These trends align with previous findings in other marine fish species and emphasize the interaction between gut microbiota and host development.At the genus level,the dominant bacterial taxa changed substantially during development.At G0,Vibrio and Tenacibaculum were dominant and likely originated from the environmental or egg-associated communities.At G1-G9,genera such as Acinetobacter,Cupriavidus,and Rubritalea increased in relative abundance,indicating microbial adaptation to rotifer ingestion and environmental changes in the gut.At G30,Cupriavidus,Pseudobacteriovorax,and Legionella were significantly enriched,suggesting a strong selection pressure from Artemia nauplii and associated metabolites.At G50,Photobacterium became the predominant genus(up to 63.8％relative abundance),likely because of the prolonged feeding of frozen Artemia and altered gut nutrient availability.The enrichment of species,such as P.aphoticum and P.damselae,may indicate potential health risks related to pathogenicity,as reported in other marine fish.LEfSe analysis confirmed the presence of stage-specific genera.Tenacibaculum was a biomarker for G0,Epibacterium and Xanthomarina for G3,Lactobacillus and Streptococcus for G9,and Massilia and Pseudobacteriovorax were indicators for G20-G30.These findings suggest that different feeding regimes and developmental stages exert selective pressures that shape the unique microbial niches within the gut.The dynamic pattern of the appearance and disappearance of key taxa implies an adaptive microbial response to host nutritional status and gut maturation.This study provides the first comprehensive profile of intestinal microbial succession in T.septentrionalis during its early ontogeny.These results highlight the essential role of diet and age in shaping gut microbial ecology.Maintaining microbial diversity,particularly during critical feed transitions,enhances host resistance and survival.Moreover,insights into the dominant and beneficial taxa lay the groundwork for future development of probiotic or microbiome-targeted functional feeds.
Accurate assessment of fishery resources is essential for sustainable management and conservation.In the southern offshore waters of Zhejiang Province,an ecologically and economically important area where the Kuroshio and coastal currents converge,fishery resources have experienced considerable fluctuations owing to intensive fishing,pollution,and climate change.This study aimed to evaluate and compare the performance of three common sampling designs,Simple Random Sampling(SRS),Systematic Sampling(SyS),and Stratified Random Sampling(StS),to estimate the relative biomass of fish communities in the southern offshore waters of Zhejiang.We used bottom trawl survey data collected over four seasons(winter,spring,summer,and autumn)in 2019.The catch per unit effort(CPUE)served as an indicator of relative fish biomass.A reference"true"spatial distribution of biomass was established using Ordinary Kriging with a Gaussian model to interpolate CPUE values across a 3'x3'grid,yielding 643 potential sampling sites after excluding non-surveyable areas.Computer simulations were used to compare the sampling performance of SRS,SyS,and StS under two sampling scenarios:small(14-35 stations)and large(30-120 stations)sample sizes.For each sample size and sampling design,1,000 resampling simulations were performed.The sampling performance of each design was assessed using the coefficient of variation(CV),relative estimation error(REE),and relative bias(RB). The results revealed marked seasonal and spatial heterogeneity in the relative fish abundance.Abundance was the highest in summer,followed by spring,and was substantially lower in autumn and winter,with high-abundance areas displaying distinct spatial patterns across seasons.With small sample sizes,SyS exhibited high variability in REE and RB,with particularly pronounced estimation errors in spring and at low station numbers.In contrast,the SRS and StS showed RB values close to zero,and the StS demonstrated only a slight negative bias in some seasons,indicating better stability.As the sample size increased,the CV and REE generally decreased across all methods,although the REE for SyS continued to fluctuate for certain sample sizes.For large sample sizes,CV decreased with increasing station numbers for all methods,with SyS typically yielding lower,more stable CV values.The REE content for SyS and StS(approximately 1％-4％)was generally lower than that for SRS(approximately 1％-7.5％).However,SyS exhibited a consistent negative bias(underestimation)in spring,summer,and winter and a pronounced positive bias(overestimation)in autumn.In comparison,the RB for SRS and StS remained close to zero,with StS a more stable performance in terms of both precision and accuracy. In conclusion,this study demonstrated that Stratified Random Sampling is the most reliable and accurate method for estimating relative fish biomass in the southern offshore waters of Zhejiang,particularly under varying seasonal conditions and across different sample sizes.Owing to its high stability and low bias,StS is recommended as the core approach for future fishery-independent surveys in the region.To enhance the survey efficiency,a seasonally adaptive sampling strategy incorporating dynamic spatial stratification based on historical distribution patterns is recommended.These findings provide a scientific basis for optimizing resource survey designs,improving the precision of stock assessments,and supporting targeted fishery management measures in Zhejiang and other similar marine ecosystems.
Drifting seaweeds refer to certain higher aquatic plants and macroalgae that float on the water surface.They typically originate from seagrass beds and macroalgal beds,where large algae are broken off or dislodged by typhoons,waves,or natural abscission during specific growth stages,and are subsequently transported over long distances by wind and currents.When drifting seaweeds proliferate and accumulate in large quantities,they can trigger ecological disaster events.In recent years,"golden tides"caused by drifting Sargassum horneri have been observed in the East China Sea and the Yellow Sea,with their frequency,scale,and severity showing an increasing trend annually.However,current domestic research on marine ecological anomalies related to macroalgae in coastal waters primarily focuses on the traceability and ecological control of green tides(Ulva prolifera),while studies on the outbreak mechanisms and ecological effects of S.horneri golden tides remain limited.To investigate the impact of golden tides on marine ecosystems and understand the characteristics of nekton community structure in coastal areas affected by drifting seaweeds,this study was conducted in the coastal waters of Zhoushan,Zhejiang-a major outbreak area of drifting S.horneri in China.In April 2023,bottom trawl surveys were carried out to analyze the diversity of the nekton community in both drifting seaweed patches and adjacent control areas.Community structure,species composition,resource density,and biodiversity indices were compared between the two areas.A total of 69 nekton species were identified in the survey area,belonging to 12 orders,37 families,and 59 genera.The total number of species recorded in the drifting seaweed area and the control area was similar,with 11 orders,28 families,45 genera and 54 species in the drifting seaweed area,and 11 orders,32 families,49 genera and 56 species in the control area.Two dominant species were identified in the drifting seaweed area,compared to one in the control area.The common dominant species in both areas was Lophius litulon.However,significant differences were observed in the size of L.litulon between the two areas.Smaller juveniles were more abundant in the drifting seaweed area,while larger juveniles were more distributed in the control area,suggesting that L.litulon may prefer sheltering within drifting seaweed litter during early developmental stages.In terms of resource density,the average numerical density(62,194 ind./km2)and biomass density(592.19 kg/km2)of nekton in the drifting seaweed area were higher than those in the control area(22,461 ind./km2 and 525.16 kg/km2).This indicates that drifting seaweeds enhance the spatial heterogeneity of surface and bottom waters to some extent,providing favorable aggregation or conservation effects on nekton and functioning similarly to"floating fish reefs"or"benthic fish reefs,"thereby positively promoting the conservation of coastal fishery resources.However,it should be noted that the single trawl survey method used in this study only covered the nekton community in the water layer beneath the edge of the algal patches.The aggregation of organisms in this area may be more influenced by the shading effect of drifting seaweed or the attraction of algal litter.The abundance characteristics of nekton within the algal patches require further investigation using more appropriate sampling techniques.Regarding community diversity,the Shannon-Wiener index(2.632)and Pielou's evenness index(0.660)were higher in the drifting seaweed area than in the control area(2.136 and 0.531),while the Margalef richness index(6.771)was lower than in the control area(7.421).This suggests that species distribution evenness improved in the drifting seaweed area,with a more balanced abundance distribution among nekton groups,thereby enhancing overall community diversity.The decrease in species richness in the drifting seaweed area reflects significant habitat differences between the drifting seaweed patches and the surrounding waters,which may be closely related to the sheltering effect of drifting seaweeds and the abundant organic matter provided by algal litter.In conclusion,the presence of drifting seaweeds in the coastal waters of Zhejiang increases habitat heterogeneity and alters the dominant species composition compared to adjacent areas.The existence of drifting seaweeds has a positive impact on the diversity of the nekton community structure and the biomass of fishery resources in the affected areas.Therefore,during the harvesting,utilization,or control of drifting seaweeds,their potential effects on the ecological environment and fishery resource conservation should be fully considered to achieve a balance between resource utilization and ecological protection.
Antibiotics have been used to control diseases in sea cucumber hatcheries,which can result in antibiotic residues in adults.Prophylactic administration of probiotics is a promising alternative strategy to control sea cucumber diseases by strengthening their immunity.Clostridium butyricum is widely used as a feed additive to promote growth,improve immunity,and enhance digestive capacity in aquatic animals.However,its use in sea cucumber has rarely been reported.This study evaluated the potential of C.butyricum as a dietary probiotic in the sea cucumber Apostichopus japonicus.A total of 750 healthy juvenile sea cucumbers with an average body weight of(5.92±0.11)g were randomly divided into 5 groups with 3 replicates in each group.The animals were reared in aquaria and fed diets supplemented with C.butyricum at concentrations of 0 CFU/g(control group,G0),106 CFU/g(G1),107 CFU/g(G2),108 CFU/g(G3),and 109 CFU/g(G4)for 90 days.The water temperature was maintained at 11-23 ℃,salinity at 30-32,dissolved oxygen at a minimum of 6 mg/L,and pH at 7.8-8.2.At the end of the feeding trial,all sea cucumbers in each aquarium were weighed and counted.Five sea cucumbers from each aquarium were sampled,starved for 24 h,and their coelomic fluid and intestine were collected for immune and digestive indices analysis,respectively.Coelomocyte counts were determined using a hemocytometer and expressed as cells per mL.Phagocytosis activity was assessed using a modified neutral red method,whereas intracellular reactive oxygen species production was measured using the nitroblue tetrazolium method.Phenoloxidase(PO)activity was determined spectrophotometrically.Acid phosphatase(ACP)and alkaline phosphatase(AKP)activity were measured using commercial kits.Lysozyme activity was measured using a spectrophotometric method.Digestive enzyme(amylase,trypsin,and lipase)activities were determined using enzyme activity assay kits.Dietary supplementation with C.butyricum at concentrations of 108 CFU/g or higher significantly improved the specific growth and weight gain rates of sea cucumbers(P＜0.05).Phagocytosis and respiratory burst activity were significantly improved in the coelomocytes of sea cucumbers fed with C.butyricum at 109 CFU/g diet(P＜0.05).Additionally,PO,ACP,and AKP activity in coelomocytes were significantly higher in the G3 and G4 groups than in the other three groups(P＜0.05).Lysozyme activity in coelomocytes was also significantly higher in the G3 and G4 groups than in the G1 and control groups(P＜0.05).Furthermore,C.butyricum significantly improved the digestive enzyme activity of A.japonicus.Sea cucumbers in the G3 and G4 groups showed significantly higher amylase,trypsin,and lipase activities compared to those in the G0 and G1 groups.These results show that C.butyricum as a feed additive can promote growth,activate the immune system,and enhance digestive ability in sea cucumber,with an optimal supplemental level of 108 CFU/g.These findings provide valuable insights for developing sustainable and antibiotic-free strategies in the sea cucumber industry.
Microplastic pollution in aquatic environments has become increasingly severe,and effective control remains a pressing challenge.Conventional flocculants such as polyaluminum chloride(PAC)exhibit limited microplastic removal efficiency and often require relatively high dosages,which may pose potential ecological risks.Enteromorpha polysaccharide(EP),an anionic natural polysaccharide,shows promising flocculation potential.In this study,based on a systematic comparison of the removal performance of PAC for microplastics with different types and particle sizes,polyethylene(PE)microplastics with a particle size of 100 pm,which are relatively difficult to remove,were selected as the target pollutant.The removal efficiency and underlying mechanisms of PE microplastics by PAC combined with EP were systematically investigated,and the effects of environmental factors on flocculation performance were also examined.The results showed that adding an appropriate amount of EP significantly enhanced the flocculation efficiency of PAC for PE microplastics.When the dosages of PAC and EP were 300 and 20 mg/L,respectively,the removal efficiencies reached 76.3％for PAC alone and 94.0％for the PAC-EP composite system.Multiple characterization techniques,including zeta potential analysis and Fourier transform infrared(FTIR)spectroscopy,revealed that the enhanced removal of PE microplastics was achieved through the synergistic effects of charge neutralization by polymeric aluminum species,adsorption-bridging by the polysaccharide,and the reconstruction of the aluminum coordination environment.Environmental factors such as pH,the concentrations of coexisting anions,and natural organic matter were found to influence the removal performance of PE microplastics by the PAC-EP composite flocculant.Further evaluation using lake and aquaculture water demonstrated that the PAC-EP system maintained removal efficiencies exceeding 80％for PE microplastics at PAC and EP dosages of 300 mg/L and 20 mg/L,respectively.This study provides a theoretical basis for enhancing microplastic removal using bio-based polysaccharides in combination with conventional flocculants.It provides valuable data to support the broader application of polysaccharide materials for controlling emerging contaminants in aquatic environments.
The farming yield of Penaeus vannamei has been steadily increasing in recent years,positioning it as the highest-producing economic crustacean in global aquaculture.However,with the ongoing depletion of fishmeal resources and the fluctuation of its prices,the need for alternative protein sources has become more urgent.Consequently,plant protein is a promising and sustainable replacement for fishmeal,with the potential to be the primary protein source in aquaculture feeds.Despite its potential,the absorption and utilization of plant protein by aquatic animals,including shrimp,has been challenging because they are not naturally adapted to efficiently digest plant-based proteins.To overcome this,probiotic fermentation of plant protein has become the primary method for enhancing the digestibility and bioavailability of plant-based protein for aquatic species,ultimately improving the overall feed efficiency and growth performance of aquaculture animals.In this study,a Bacillus subtilis strain that is Gram-positive bacterium,non-hemolytic,and sensitive to most antibiotics,named SQVGB8,was isolated and identified from the intestine of P.vannamei,with a 99.20％similarity to Bacillus subtilis subsp.subtilis strain 168.The characteristic analysis results showed that the strain can efficiently produce protease,lipase,amylase,cellulase,and phytase;inhibit Vibrio harveyi,V alginolyticus,and V.parahaemolyticus;secrete active substances such as short-chain fatty acids(SCFAs)and extracellular polymeric substances(EPS);and exhibit high intestinal adhesion properties. The whole genome analysis results showed that SQVGB8 can produce digestive enzymes,phytase,SCFAs,and EPS.Additionally,SQVGB8 can secrete various substances with antibacterial and antimicrobial potential,including surfactin,fengycin,and bacillibactin.These active substances have significant application prospects in antibacterial and pathogen control.The suitable fermentation conditions for SQVGB8 fermented plant meals(soybean meal∶cottonseed meal=3∶1)were determined using a single-factor experiment.The optimal conditions are as follows:inoculum amount 3％-12％,fermentation temperature 35-45 ℃,fermentation time 36-72 h,and solid-to-liquid ratio 1∶1.2-1∶0.6 g/mL.Therefore,a response surface methodology with a four-factor,three-level analysis was used,and the optimal fermentation conditions for SQVGB8 fermented plant meals were an inoculum amount of 7.70％,a fermentation temperature of 35.70 ℃,a fermentation time of 55.00 h,and a solid-to-liquid ratio of 1∶0.78 g/mL.The acid soluble protein and acidic polysaccharide contents were 10.30％and 12 mg/mL,respectively.Compared to that of the unfermented plant meals,the antinutritional factor content was significantly reduced.The trypsin inhibitor,soybean globulin,β-conglycinin,gossypol,and phytic acid contents were reduced by 91.67％,61.57％,76.75％,59.00％,and 48.72％,respectively.In conclusion,SQVGB8 is an efficient enzyme-producing,active substance-secreting,and gut-colonizing probiotic,with high efficacy in fermented plant meals.It has great potential for development and application in aquaculture feed.
Anti-Lipopolysaccharide Factor (ALF)-like is an anti-microbial peptide that plays a crucial role in the innate immune system and is widely present in crustaceans and other organisms. However, the molecular mechanisms underlying disease resistance regulated by this factor in the intestinal tissues of Penaeus vannamei remain unclear. Based on this, we screened an ALF-like gene from the transcriptome databases of different disease-resistant materials (highly and poorly resistant) infected with white spotsyndrome virus (WSSV). This study was conducted to investigate the molecular mechanisms underlying disease resistance in shrimp intestinal tissues. Specifically, previous experiments on WSSV infection have shown that WSSV can induce the upregulation of this gene. In this study, RNA interference (RNAi) was used to knock down ALF-like, and the interference efficiency of ALF-like in shrimp intestinal tissue was detected using real-time fluorescence quantitative PCR. The relative expression levels of apoptosis and antioxidant pathway genes as well as the proliferation changes in WSSV (detected using IE1 and VP28) in the shrimp tissue were analyzed. The SNP sites in this gene were simultaneously explored and analyzed. Subsequently, kompetitive allele-specific PCR (KASP) technology was used to verify the genotyping of the selected SNP sites. The experimental results indicated that the expression of ALF-like in the intestinal tissue showed a significant downward trend at 24, 36, and 48 h after dsRNA injection, with the interference efficiency decreasing by 79.3%, 80.9%, and 63.2%, respectively, compared to that in the control group. After knocking down the target gene, pathogen detection showed that knockdown of ALF-like promoted the up-regulation of IE1 and VP28. ALF-like regulated the expression of genes involved in apoptosis (downregulation of Bax, upregulation of Bcl2, and downregulation of Caspase8) and antioxidant enzyme (upregulation of CAT and SOD) pathways. Subsequently, through SNP site screening in ALF-like, 19 SNP sites were screened from the exon region of this gene. Among these, SNP18 was moderately polymorphic and stably expressed in both the resistant and sensitive groups. Finally, KASP genotyping technology was used to conduct genotyping research on the DNA/cDNA of samples from the resistant and sensitive groups, and results indicated that this technology could be applied to SNP genotyping of shrimp. This approach provided key candidate markers for the subsequent research on “gene variation-disease resistance phenotype-disease resistance breeding.” In summary, this study revealed the key mechanism of ALF-like in the disease-resistant immunity of the P. vannamei intestinal tract. RNAi experiments revealed that after knockdown of ALF-like, the expression of key WSSV genes (IE1, VP28) was upregulated, indicating that ALF-like can inhibit viral proliferation and participate in immune defense by regulating cell apoptosis and antioxidant pathways. In the SNP screening study of ALF-like (related to disease resistance) in P. vannamei, 19 SNP loci were screened in the exon region. The KASP genotyping technique was successfully used to genotype SNP loci, confirming that this method is applicable for the study of SNP loci in shrimp. In shrimp, ALF exhibits extensive antipathogenic effects, including antibacterial and antiviral functions. Compared with the intestines of higher mammals, the intestinal folds of shrimp are relatively smooth, and the invasion of microorganisms has a particularly severe effect on beneficial bacteria in the intestine and intestinal immunity. Intestinal immune imbalance not only affects shrimp feeding but can also lead to intestinal lesions. Therefore, exploring the intestinal anti-disease immune system in the shrimp pathogen defense system is crucial. This research has preliminarily clarified the immune regulatory mechanisms of ALF-like molecules, providing a theoretical basis and new ideas for aquatic genetic improvement, disease prevention and control. In future research, multi-omics technologies can be combined to analyze the immune regulatory network of ALF-like molecules and explore their synergistic effects with other immune molecules simultaneously, the findings can promote their application in shrimp disease breeding (such as gene editing) and green aquaculture (such as feed additives), providing more precise technical support for aquatic disease prevention and control.
China,a major global shellfish aquaculture country,had a national shellfish aquaculture output of 16,659,000 t in 2024,of which marine aquaculture accounted for 98.81％.As the main farmed species,oysters have an annual output of 6,671,200 t,accounting for 40.52％of the total production of marine shellfish farming.Due to the over-expansion of certain traditional aquaculture areas,the ecological carrying capacity has been exceeded and the phytoplankton biomass available for oyster feeding has been reduced,resulting in lower fattening,quality,and market price,and higher mortality rates;therefore,exploring new oyster fattening sites and methods has become integral in solving the current problems in the industry.To investigate the feasibility of Crassostrea gigas pond fattening,a self-developed culture device of oyster ponds was used to enhance the water exchange around C.gigas by using the flow generated by the natural wind disturbance on the surface of the pond to increase the bait delivery efficiency to provide sufficient bait for the C.gigas,and to try to fertilize the diploid C.gigas,which have poor fattening degree,cultured in the sea area in the shrimp and crab ponds.We fattened diploid C.gigas with poor fattening in marine culture in shrimp and crab culture ponds.From November 2023 to May 2024(the experiment was suspended during the winter freezing period),we visited the experimental ponds every 20 days to collect samples and used the WTW multi-parameter water quality analysis to measure the water temperature,salinity,pH,and dissolved oxygen.We used a water collector to collect 2 L of water samples from each of the five points at the four corners of the ponds(5-10 m from the shore)and the center of the ponds and then stored them in water sample bottles.Next,referring to the Code of Practice for Marine Surveys,the water samples were fixed with 5％formaldehyde solution on site and transported back to the laboratory to measure nutrient salt,chlorophyll a,and particulate organic matter(POM)contents.Three sizes(S,M,L)of C.gigas were set up to determine the growth and physiological indices,with average wet weights of(54.86±3.26)g,(83.81±3.94)g,and(127.46±8.64)g,respectively,and were cultured in three mixed groups in oyster pond culture devices.For the physiological indices,eight replicates and one control were established for each specification treatment group,and a flow-through device comprising a buffer box,flow-through tank,and rectangular plastic box was used for feeding physiology and respiratory metabolism measurements.During the experimental period,a grapple-type mud collector was used to collect pond sediment underneath and around the aquaculture devices at the aquaculture site,artificial feed,suspended particles,and feces of C.gigas,which were used to determine the sediment of aquaculture ponds.The pond sediment was collected in different months.No significant differences(P＞0.05)were noted in water temperature,salinity,pH,dissolved oxygen content,chlorophyll a,and POM content of the culture ponds in different months;however,there were significant differences(P＜0.05)in PO43--P,NO2--N,NO3--N,and NH4+-N in the culture ponds in different months.The differences in fecundity of the three oyster sizes in the same month were not significant(P＞0.05);however,the differences in the water filtration,droppings,oxygen consumption,and ammonia discharge rates were all significant(P＜0.05),whereas those in fecundity of the same sizes of oysters in different months were significant(P＜0.05).The energy balance equation showed that all three oyster sizes used the most energy for growth in May,with the highest percentage of 48.64％;after 6 months of cultivation,the fattening degree of oysters increased by 31.7％,and the fastest growth rate of large-size oysters was 55.1％.Comparing the same batch of oysters from ponds and the sea area in the same period of time showed that the survival and fattening degree of oysters from pond culture were higher than those of oyster cultivation in the sea area(P＜0.05).An analysis of organic matter sources in the pond sediments showed that the further away from the aquaculture unit,the higher the contribution of suspended particles,whereas the opposite was true for artificial feed and feces.This study showed that C.gigas are more effective for fattening in marine shrimp and crab ponds,and it is recommended to begin the pond fattening production in autumn from mid-September to mid-October,and in spring from mid-March to mid-May,and to select C.gigas with larger sizes,so that the desired degree of fattening can be reached after 1-2 months.
The intestine is the primary site of nutrient digestion and absorption in fish,in addition to serving as a crucial immune organ.Salinity and temperature are key environmental factors that significantly influence fish physiology and survival.As a specific ecological factor in aquatic environments,salinity can markedly affect fish intestinal structure and function,thus posing a threat to fish health.Temperature is considered a major environmental factor affecting fish physiology,with temperature changes able to influence the activity of digestive,antioxidant,and other enzymes in the intestine,as well as the expression of growth-related genes.Temperature can also disrupt the intestinal morphology and induce intestinal cell apoptosis.The combined effects of salinity and temperature changes on fish physiology are often complex and additive.Their combined negative effects can far exceed their individual contributions,thereby compromising the adaptive capacity of fish.Alternative splicing(AS),as a key regulatory mechanism in gene expression,not only increases protein diversity,but also dynamically modulates gene expression programs,thereby enhancing the adaptability of organisms to environmental stress.In fish,AS plays a critical role in the response to various environmental stresses,including temperature and salinity changes.Rainbow trout(Oncorhynchus mykiss),a euryhaline fish with both economic and ecological research significance,exists in anadromous and resident forms.Resident rainbow trouts live in freshwater environments and thrive at temperatures between 12-18 ℃.In China,most cultured rainbow trout are of the resident type,and their production has been constrained by inland cold-water resources,remaining at around 40,000 tons.Marine farming is a viable strategy to increase rainbow trout production.However,despite their relatively high tolerance to salinity,rainbow trout exhibit significantly reduced intestinal digestive enzyme activity and growth performance in high-salinity compared to freshwater environments.Additionally,global warming has led to increased seawater temperatures,exacerbating the dual stress of salinity and temperature on rainbow trout and intensifying the environmental challenges for marine farming.To investigate the interactive effects of high salinity and high temperature on gene expression in the rainbow trout intestine and their dynamic AS responses,we conducted an in-depth analysis of RNA-seq data from rainbow trout intestines under high salinity and high temperature stress.Data were preprocessed using methods identical to those used in the original study,including data filtering,quality control,transcript assembly,gene counts,and FPKM calculation.Differentially expressed genes(DEGs)were identified in the high-salinity stress(SI),high-temperature stress(TI),and dual high-salinity high-temperature stress(DI)groups,with counts of 339,1 350,and 3 581,respectively.Subsequently,based on the gene count values,the DESeq2(v1.44.0)R package was used to construct a generalized linear model(GLM)for interaction effect analysis.The design matrix included the main effects of salinity(0/30)and temperature(16/25 ℃),as well as the salinity-temperature interaction term(～Salinity+Temperature+Salinity:Temperature).A likelihood ratio test(LRT)was used to compare the full model(with interaction terms)and the reduced model(with only the main effects of Salinity+Temperature).Low expression genes(filtered based on total expression,removing genes with counts totaling less than 10 times the sample number)were excluded before model fitting to enhance the accuracy and stability of model construction.The plotDispEsts function was then used to generate a dispersion estimate plot,validating the model and reliability of the data.Interaction analysis revealed 113 interacting genes,comprising six synergistic genes(two DEGs)and 107 antagonistic genes(51 DEGs).Although the interaction genes accounted for only 0.31％of the background genes,they exhibited strong interaction effect intensity.Using the freshwater control group(FI)as a reference,rMATS-turbo software(v4.1.2)was used to detect and analyze AS events in SI,TI,and DI,including five common splicing types:Skipped Exon(SE),Alternative 3'Splice Site(A3SS),Alternative 5'Splice Site(A5SS),Retained Intron(RI),and Mutually Exclusive Exons(MXE).In comparison with the freshwater temperature-suitable group(FI),AS events were abundant in SI,TI,and DI,comprising 32.33％,33.96％,and 33.71％of the total events,respectively,with a near 1∶1∶1 ratio.The local composition of SE∶A3SS∶A5SS∶RI∶MXE was approximately 29∶30∶22∶14∶5 with A3SS and SE being the most common.DAS events(FDR＜0.05,and difference with FI＞10％)comprised 1.62％,45.01％,and 53.37％of the events in the SI,TI,and DI,respectively,showing significant differences.The SE:A3SS∶A5SS∶RI∶MXE ratios varied distinctly across the conditions:SI(46∶11∶17∶23∶3),TI(54∶15∶20∶5∶6),and DI(54∶13∶20∶7∶6),with SE being the most common.Notably,SI exhibited fewer DAS events than TI and DI.Further analysis identified 31,699(30 DEGs),and 761(64 DEGs)genes in the DAS events for SI,TI,and DI,respectively,with substantial overlap between TI and DI genes.Seven DAS genes were shared across all three stressors,displaying diverse AS events.The TI and DI DAS genes were significantly enriched in three KEGG pathways(Padj＜0.05):Spliceosome(ko03040),Polycomb Repressive Complex(ko03083),and ATP-Dependent Chromatin Remodeling(ko03082),with high gene overlap.Only five antagonistic genes were DAS genes,and DAS events were exclusive to TI and DI.The results of this study indicate that the interactive effects of high salinity and high temperature stress on overall gene expression in the intestine of rainbow trouts may be minimal but have significant impacts on specific genes.Additionally,SE events may play an important role in the response of rainbow trout to different environmental stressors,whereas RI events may play a unique role in the response of fish to salinity stress.These findings reveal the complex mechanisms underlying gene expression changes for coping with environmental stress,thus providing new insights into the environmental adaptation strategies of rainbow trouts.This study offers an important theoretical foundation for the further development of marine farming and strain improvement efforts for this species.
