Abstract:Mandarin fish is one of the most commercially important freshwater fish in Chinese aquaculture. Once the fry of mandarin fish start feeding, in the wild they exclusively eat live prey fish; therefore in cultivation they refuse dead prey fish or artificial diets, contributing to the challenges of breeding this fish. Currently, the source of live feed for the intensively cultured mandarin fish is also cultivated. The challenges in cultivating live feed for mandarin fish include extremely high cultivation costs, difficulties in maintaining stock supply all year round, and disease prevention, which is very difficult in the cultivation process. Therefore introducing artificial diets in domesticated mandarin fish could resolve these challenges. Based on the existing research, this study analyzed the transcriptome data of mandarin fish, and the differences in expression of pepsinogen (pep and are considered as candidate genes for food habit traits in domesticated mandarin fish. In this study, single nucleotide polymorphisms (SNPs) on candidate genes were sequenced and typed in a feeder group fed dead prey fish diets and a non-feeder control group, and evaluated for associations with food habit domestication traits in 103 mandarin fish. The feeder means the fish accepts dead prey fish, and the non-feeder means the fish refuses to eat dead prey fish. In total, five SNPs were identified in genes. The effective allele numbers ranged from 1.1959 to 1.7001, with a mean of 1.4274 alleles per locus. The observed () values ranged from 0.1800 to 0.3585, and from 0.1655 to 0.4160, respectively. The value of polymorphism information content (PIC) was 0.2477, which is a classification of the median polymorphism level in the mandarin fish population. These results showed that the two genotypes TT and CT in the -A T/C of the SNP locus were significantly correlated with the high level of the food habit domestication traits in mandarin fish (<0.05). The three genotypes of genotype 1 (CT, CC/CT/TT, AA, AA, TT), genotype 2 (TT, CC/CT/TT, AA, AA, TT) and genotype 3 (TT, CC, AA, AA, TT) were also significantly associated with high levels of food habit domestication traits (<0.05). These genotypes affect the food habit domestication traits in mandarin fish, and genotype 2 had the highest correlation, which can be selected as the optimal genotype among individuals. In conclusion, our results demonstrated that these effective SNP molecular markers in genes could influence food habit traits in domestication of mandarin fish and could be used for accelerating the molecular marker-assisted selective breeding in mandarin fish. Based on the study of the SNPs association analysis of candidate genes with food habit domestication traits in mandarin fish, our findings could provide theoretical foundation and methodology for selectively breeding mandarin fish with increased tolerance to dead prey fish or artificial diets.

Abstract:The aim of this study was to explore the role of in stress-responsive regulation of roughskin sculpin () under low salinity stress. Firstly, sequences of these genes were obtained from transcriptome data of , and phylogenetic analysis was performed. In this study, fish were subjected to two different acute osmotic treatments (salinity changing rate at 27 and 1.1 ppt/h), expression patterns of the three genes in the four target tissues (gills, intestines, kidney, and liver) were examined using qRT-PCR. Phylogenetic analysis revealed that genes respectively formed an independent cluster, in which protein shared high identity with those of Perciformes, Cypriniformes, and Cyprinodontiformes species, teleost proteins formed a single lineage distinct from those of other vertebrates. Tissue-specific gene expression patterns in all three target genes showed that each tissue had a specific gene expression pattern in response to salinity changes. In the gills, the expression of increased significantly at 12 h, but different expression patterns were identified in the other tissues. In the intestines, the expression of decreased significantly at 48 h under the relative chronic salinity stress, while the expression of significantly increased; under the acute salinity stress, mRNA levels of increased significantly at 24 h, and the expression of HSPB1, HSPB7, and was significantly upregulated at 24 h in response to the relative chronic salinity stress; under the acute salinity stress, the expression of HSPB11 were significantly upregulated. The expression of HSPB1 expression was detected under chronic salinity stress, but a significantly upregulated expression was found under the acute low salinity stress; the expression of was significantly upregulated under both treatments. These results demonstrate the differences among expression profiling in the stress-responsive regulation activity of , these findings could provide a theoretical foundation for revealing the role of small heat shock proteins in fish stress-responsive regulation and the molecular mechanism of migratory fish salinity adaptation.

Abstract:The aims of this study were to examine the structural features of the amino acid sequence encoded by genes and to analyze the expression patterns of these genes during gonadal development, to understand the role of hypothalamic neuropeptide kisspeptin in reproductive regulation of Amur sturgeon (). We used reverse transcription PCR and rapid-amplification of cDNA ends (RACE) technology to obtain the full-length cDNA of kiss1 gene of gene was 518 bp, encoding 149 amino acids. Among them, the α-helix and irregular distortion in Kiss1 and Kiss2 are regarded as the main components in the secondary structure, both containing signal peptide, transmembrane structure, and unstable hydrophilic protein secreted outside the cell. Amino acid sequence alignment and phylogenetic tree analysis showed that had highly conserved regions, which had the highest consistency (and the closest relationship) with the Acipenser dabryanus). The results of real-time PCR showed that the expression of kiss1 had the highest expression in the gonads, while expression was the highest in the brain. In the early gonadal development of was lower at 0-49 d after hatching, while the expression of was higher, reaching the maximum at 21 d after hatching. With the development of gonads in gradually increased and reached the highest at 139 d, followed by a decline. In contrast, the expression level of reached the maximum at 184 days after hatching. In summary, the kisspeptin gene plays an important role in the early gonadal development of , with differing regulatory functions. This study lays a theoretical foundation for further elucidating the physiological functions and molecular regulation mechanisms of genes.

Abstract:The mantle is an indispensable tissue for pearl cultivation, but the biological activity (cell proliferation and biomineralization) of donor mussel mantle of different ages has not been systematically explored. In this study, was used to investigate the proliferative activity and biomineralization capacity in mantle cells in different years. In this experiment, five groups of were selected by age, at six months and one, two, three, and four years. Flow cytometry was used to detect the cell cycle of mantle cells of different ages and to calculate the cell proliferation index. Fluo-3/AM, a calcium fluorescence probe, combines with intracellular Ca²⁺ to produce fluorescence. The average fluorescence intensity of mantle cells of different ages was measured by flow cytometry fluorescein isothiocyanate (FITC) channel, which represented intracellular calcium concentration. Quantitative real-time PCR (qRT-PCR) was used to analyze expression of biomineralization related genes. Biomineralization-related genes included carbonic anhydrase gene (). The enzyme activities of ALP and CA were determined chemically and by ELISA, respectively. The aim of this study was to determine the suitable age of with fast proliferation and strong biomineralization ability of mantle cells. The results showed that:(a) Under the same environment, the proliferation index of two-year-old mantle cells was the largest (i.e., fastest proliferation). (b) The average fluorescence intensity of the two-year-old was the highest, in other words, the concentration of calcium ion in the mantle cells of the two-year-old gene in six-month-old mussels was significantly higher than that in older mussels (<0.05), followed by the one-year-old mussels and the two-year-old mussels. There was no significant difference between the three and four-year-old mussels, with the lowest gene expression level (<0.05). (d) The expression level of was different in mantle tissues in mussels of different ages. The highest expression level (<0.05) was found in one-year-old mussels, followed by the two-year-old mussels, and the lowest expression level (>0.05) was found in six-month-old, three and four-year-old mussels. The activity of CA was different in mantle tissue of different mussels and the activity of CA was the highest in one-year-old mussels (<0.05). Finally, it was concluded that the proliferation of mantle cells of the two-year-old mussels was the strongest and the intracellular calcium concentration was the highest. This study provides a foundation for further research on the proliferation of mantle cells in and the selection of donor mussel age for pearl culture.

Abstract:This study was conducted to evaluate the effects of replacement of fish meal (FM) by defatted black soldier fly larvae meal (DBSFLM) on growth, body composition, serum biochemical parameters, and antioxidant capacity of juvenile large yellow croakers (). Six isonitrogenous (45% crude protein) and isolipidic (10% crude lipid) diets were formulated by replacing 0, 20%, 40%, 60%, 80%, and 100% of FM protein with DBSFLM (named as G0, G20, G40, G60, G80, and G100, respectively). A total of 2160 juvenile large yellow croakers with an initial body weight of (50.08±3.31) g were randomly divided into six groups with three replicates, and 120 individuals for each replicate. Fish were reared in an indoor cement tank (2 m×1 m×1 m) and were hand-fed each diet to apparent satiation twice daily for 7 weeks. The results showed that survival rate of G100 was significantly lower than the other experimental groups (<0.05). Weight gain rates, specific growth rate, and protein efficiency ratio in the G40 group were all significantly higher than those in the other groups (<0.05). The muscle crude protein content decreased as the DBSFLM replacement level was increased, particularly when the replacement level exceeded 40% (<0.05). Crude lipid and crude ash content significantly increased when replacement level reached over 60% (<0.05). The activity of serum ALT and AST significantly increased, and serum TG and cholesterol levels significantly decreased with the increase of the replacement level (<0.05). Total antioxidant capacity (T-AOC) of fish liver in the G20 group and superoxide dismutase activity (SOD) in the G40 group were the highest among all groups. The lowest liver malondialdehyde (MDA) content was found in the G40 group. The catalase (CAT) activity decreased and was significantly lower when replacement level exceeded 40% (<0.05). In conclusion, up to 40% of DBSFLM may successfully replace FM in diets for juvenile large yellow croakers, without adverse effects on growth performance, body composition, and health status.

Abstract:Target strength (TS) is a pivotal parameter for converting echo integral values into absolute fish abundance. It is also a hot research topic in the research and application of fishery acoustic technology. However, the TS-length relationships have been studied in very few freshwater species. Most studies focused on commercially important species, while TS in fish of different physiological structures is investigated less frequently. In this study, cage method and KRM (the Kirchhoff ray-mode approximation model) method were used to measure TS of 4 fish species (Acipenser dabryanus) which represent different morphological and ecotypic characteristics. Thirty-one individuals were measured by a calibrated 199-kHz BioSonics DT-X split beam echosounder in the vertical direction in a net cage, using the Visual Acquisition software. The cage method experiment was conducted in the Chinese sturgeon farm base which is located in the Three Gorges Reservoir, in December 2012 and from June to July in 2013. During the cage experiment, one fish was measured each time. Then, the body length and weight of each species were measured after the fish were euthanized in Tricaine-S (MS-222). The acoustic data were processed by Sonar-5 Pro software. Two X-ray images were taken from the dorsal and side of fish to understand the internal physiology and the shape of the swim bladder of the fish. The TS of six individuals was calculated using KRM model, and the results were verified by cage method. The results showed that it was feasible to determine TS of freshwater fishes using the cage method, and the results of this method were not significantly different from the model method. The linear regression equations for mean TS with fish BL were derived in dorsal aspect were:TS_pp=20lg(BL)-70.1 (=0.80), TS_sa=20lg(BL)-74.5 (=0.94), respectively, where BL is fish body length, pp is , and ad is . The linear regression equations for mean TS with fish TL in dorsal aspect were:TS_pp=20lg(TL)-71.4 (=0.80), TS_sa=20lg(TL)-75.1 (=0.94), respectively, where TL is total fish length. Under the same body length conditions, the TS of the four fish species was compared:S. asotus. The results of KRM model support that swim bladder is the primary contributor to backscattered energy. The values of TS and their distribution calculated by the cage method and model method were affected by the morphological characteristics and the swim bladder structure of fish species. Maximum TS of occured between -10° and 0° at 38 kHz, while the maximum TS of appeared between -20° and -10°. The position of maximum TS occurred differently for each fish species. At the low frequency of 38 kHz, the model method results showed that there was significant difference in the distribution of TS with the angle of incidence of sound waves between fish with a one-chambered swim bladder and those with a multi-chambered swim bladder. Compared to immobilized and unconscious fish, the fish in the net cage can swim freely. However, the narrow beam lowers the probability of the fish being detected, which increases the time of the experiment. In the future, it is necessary to appropriately reduce the size of the cage to optimize the implementation conditions. Our study enriches the TS of freshwater fishes and accumulates basic data for acoustic identification.

Abstract:Six surveys were carried out to investigate the fishery resources and their habitat environments from May to October 2011 in the Laizhou Bay. Phytoplankton samples were collected using a standard vertical towing net (mesh size 77 μm), followed by the addition of formalin (approximately 5% final concentration in each sample) for preservation. Habitat environment was measured with a Multiparameter Sondes (YSI 6600). Phytoplankton species identification and cell enumeration were performed with an optical microscope. Phytoplankton community succession from spring to fall was clarified and its relationship with the density of major pelagic fishes was revealed by generalized additive models (GAM). A total of 33 genera and 78 taxa were recorded, with diatoms and dinoflagellates accounting for 85.9% and 14.1% in the total taxa numbers, respectively. Most of the species were eurythermal, and some were warm-temperate ecotypes, such as Rhizosolenia styliformis. Others were warm-water and oceanic species, such as . Two abundance peaks formed in the spring and fall seasons, with the values of 265×10⁴ and 111×10⁴ ind/m³, respectively. Monthly abundance from May to October was 265×10⁴, 53.1×10⁴, 27.1×10⁴, 27.7×10⁴, 111×10⁴, and 92.9×10⁴ ind/m³, respectively, with the diatoms accounting for 80.8%, 76%, 54.5%, 87.8%, 86.6%, and 68.7%. The monthly averaged ratios of dinoflagellate to diatoms abundance was 1.15, 1.13, 2.54, 0.23, 0.17, and 1.42, peaking in July. The dominant taxa were from late spring to early summer, from middle summer to middle fall. Monthly , 31.8×10³, 124×10³, 219×10³, 398×10³, and 188×10³ ind/m³, respectively, with the diatoms accounting for 32.7%, 18.7%, 34.8%, 51.7%, 49.3%, and 32.3%. , 8.57×10³, 19.9×10³, 22.4×10³, 250×10³, and 63.7×10³ ind/m³, respectively, accounting for 0.24%, 4.29%, 5.74%, 12.1%, 15.7%, and 13.3%. , 23.8×10³, 1.31×10³, 10.5×10³, 0.23×10³, and 1.24×10³ ind/m³, respectively, accounting for 4.14%, 9.64%, 5.49%, 15.9%, 0.045%, and 0.42%. 3, 0.57×10³, 3.76×10³, 68×10³, and 124×10³ ind/m³, respectively, accounting for 0.24%, 4.29%, 5.74%, 12.1%, 15.7%, and 13.3%. , 0.93×10³, 7.13×10³, 7.46×10³, 20.4×10³, and 33.7×10³ ind/m³, respectively, accounting for 0.032%, 0.39%, 3.36%, 5.42%, 2.8%, and 7.77%. The species richness, diversity and evenness increased markedly from late summer to middle fall. They had 72%, 70%, and 29.6% increases after August, respectively. The GAM analysis was fulfilled in the mgcv-1.8 package by R software. The stepwise optimized final model, as explained by environment and phytoplankton factors, clarified a sum of 76.5% deviance to the total CPUE (Catch Per Unit Effort) of four pelagic fishes (Engraulis japonicus). There were significant differences in the five predictor variables of month, sea surface temperature, abundance, and the ratio of dinoflagellate to diatom abundance, with stepwise deviation explanation of 52.3%, 8.6%, 9.3%, 0.5%, and 1.8%. It is clear that the long-term variations of the phytoplankton community structure and abundance level have made positive effects on the sustainability of the current status of fish communities and fishery resources in the Laizhou Bay. This study provides the baseline data and evidence from the view of phytoplankton diets, for further discussion of the factors that influence on the recruitment and food web processes of the fishery populations in the key habitats.

Abstract:With the current situation of resource depletion and environmental changes, artificial reefs may be used to promote the fish trapping and breeding to proliferate and conserve fishery resources and restore the ecosystems. The main idea of artificial reef construction was to stimulate the production and transmission of nutrients, sediment size, and primary productivity through the unique flow field generated around the reef, and to proliferate plankton, pelagic swimming organisms, and benthic resources in the form of food supply and habitat transformation. Therefore, the flow field effect of artificial reefs was based on ecological-biological interactions. The strength of the flow field effect is affected by the scale of artificial reef unit. The measurement of the artificial reef construction mode was one of the most important factors for planning the scale of artificial reef construction. The flow field volume was an important indicator to characterize the flow field effect. Based on the numerical experimental method, this paper analyzed the variation of the flow field volume of Mi-zi artificial reefs in 28 kinds of artificial reef scale units in four laying modes, and established the multivariate nonlinear model for the flow field volume of upwelling and back eddy and artificial reef construction model indicators. The results showed that in the primary indicators of artificial reef construction, the throwing amount () and target velocity ratio () and upwelling volume were linear, cubic, and power function, respectively. Combining the relative length (R) of the secondary index to establish the upwelling volume regression model was -0.055_u^-1.833+0.227_b=r)-51.779d× were 0.957 and 0.938, and the relative errors were 18.61% and 10.09%, respectively. The flow field volume regression model can be used to plan the construction of artificial reefs that guide the uniform laying mode, and gradually realize the artificial reef construction strategy of "reducing mass and improving efficiency".

Abstract:As a commercially important tuna in the high seas, bigeye tuna () has become one of the most important target species in Chinese distant-water fisheries. The research and management of is receiving more interest, especially for its life history, stock assessment, and fishery management. The fish life history traits including growth, is the fundamental and key process of population dynamics and stock assessment, with an increasing attention in recent years. Based on the data collected by Chinese observers onboard from 2013 to 2018, the growth of the bigeye tuna was analyzed, with the spatial-temporal variations. A total of 8806 individuals were measured, including the wet weight (kg; after removing gills, gut, and tail) and fork length (cm). The predicted power length-weight function indicated that the estimated condition factor , while the estimated allometric growth parameter is 3.08 (3.07-3.10). Mixed effects models were established to estimate the variations from different years, quarters, and regions, while 7 model candidates were considered with different random effect sources. AIC (Akaike Information Criterion) and Root Mean Square Error values revealed that the mixed effect model performed best with all variations from years, quarters, and regions. Results from the best model indicated that:(1) there were no substantial differences between bigeye tuna in the north or south Indian Ocean divided by 15°S; (2) individuals collected in the first and second quarters tended to gain more weight than those collected in the third and fourth quarters at the same longitude; and (3) individuals collected in 2015 and 2016 grew better, while those collected in 2014 and 2017 gained much less weight at the same fork length. The outcome from this study could assist the stock assessment and fishery management for this important tuna species in the Indian Ocean, and the methodology used in this paper can also be applied to the heterogeneity studies of other species in both coast water and far ocean.

Abstract:Bigeye tuna () is one of the most valuable tropical tuna species targeted by most longline fisheries. stock assessments have always been the focus for regional tuna fishery management organizations worldwide. Based on the catch from 1950 to 2016 and Catch Per Unit Effort (CPUE) from 1955 to 2016, the stock of the Indian Ocean bigeye tuna was assessed by the Bayesian state space surplus production model in an open environment, JABBA (Just Another Bayesian Biomass Assessment), and the implications on the effects of fishing boat and CPUE data scale was explored. The results showed that the stock assessment was sensitive to different CPUE, and the scenario using CPUE considering vessel effect from 1979 to 2016 was revealed to perform best with the lowest Root-Mean-Squared-Error (RMSE) and Deviance Information Criterion (DIC), and selected to be the base case. The median estimate for bigeye tuna biomass in 2016 was 812 kt, and the Maximum Sustainable Yield (MSY) was estimated to be 163 kt, which was much higher than the catch (86.81 kt) in 2016, indicating that the stock was not overfished, with 81% in the green zone of the Kobe plot. The biomass of bigeye tuna would be higher than the biomass that can produce the maximum sustainable yield () in the next 10-year projection when the total permissible catch was set to 69.45-104.17 kt (80%-120% of catch in 2016). There were some retrospective errors in the stock assessment results, with underestimated fishing rate and overestimated biomass. Therefore, the stock assessments should be improved by updating the model structure, CPUE standardization, and setting for prior distribution of model parameters.

Abstract:This study aimed to assess stock enhancement of the bighead carp ( is a fish of ecological and scientific significance, inhabiting most of Chinese freshwaters. Moreover, it is one of the most released species in the lower reaches of the Changjiang River. The primary focus of stock enhancement was to release juveniles into natural environments and enhance the stocks of interest. However, the genetic quality of large-scale stock fish is difficult to quantify. Consequently, it is essential to examine the genetic quality of the brood stocks in the hatchery, in order to provide a scientific reference for genetic resource management as well as living resources database construction of brood stocks in the lower reaches of the Changjiang River. During our study, the genetic diversity and population structure of 8 populations were studied using SSR markers. A total of 638 individuals from 7 parent populations and 1 candidate parent population were assayed by 10 pairs of SSR primers. Genetic diversity parameters, principal coordinate analysis (PCoA), and analysis of molecular variance (AMOVA) were generated using Cervus 3.0, GenAlEx 6.501 and Arlequin 3.1 software. Structure and CLUMP software were used to analyze the genetic structure of the populations. The results revealed that the genetic diversity was superior despite inbreeding risk within populations. The average number of alleles was 14.83±1.45, expected heterozygosity and observed heterozygosity were 0.82±0.02 and 0.76±0.05, respectively, while Shannon information index was 2.08±0.09, and inbreeding coefficient was 0.08±0.05. The highest level of genetic diversity was in the candidate parent population. Furthermore, the genetic diversity of parent populations was different. According to structure and PCoA, all individuals could be divided into two genetic clusters, which showed that the genetic differentiation among parent populations were not significantly different, in addition the genetic differentiation between parent and candidate parent populations was significantly different. AMOVA analysis showed that 11% of genetic variation was among the populations and 89% of genetic variation was within populations. The genetic differences between candidate parent and the parent of bighead carp suggest the importance for the introduction and protection of natural germplasm. The results from this study further suggest that it is possible to improve hatchery practices and mitigate the negative effects on wild stocks. However, theoretical support and data reference are required for constructing a healthy and sustainable stock enhancement strategy.

Abstract:Aptamers are oligonucleotide molecules obtained by systematic evolution of ligands by exponential enrichment, which have a high affinity and specificity towards their target. Aptamers are applied in many fields, such as life science research, drug screening, target identification, and environmental monitoring. Affinity determination of an aptamer and its specificity are necessary for the research and development of the aptamer. Therefore, a novel affinity determination of aptamer, called ssDNA concentration method, was developed to study the specificity and affinity constant of an aptamer against . The affinity and affinity constant of the aptamer were determined by measuring the ssDNA concentration of the aptamer binding to the V. harveyi detected by the ssDNA concentration method, was 15.2 times higher than that of the non-target bacteria. The affinity and specificity of the aptamer were also verified by fluorescence microscopy. There are two main methods of fluorescence microscopy, the direct observation method and the fluorescence blocking method. The direct observation method is observing the binding of aptamer and target bacteria under fluorescent microscope after bacteria were incubated with fluorescent labeled aptamer. The results verified by the direct observation method under fluorescence microscopy showed that the target bacteria emitted significant yellow-red fluorescence after binding with the FAM-labeled aptamer, but little fluorescence was found in other four kinds of non-target bacteria (Escherichia coli), proving that the bacteria have no spontaneous fluorescence. Furthermore, the fluorescence blocking method involves incubating bacteria with no fluorescent label aptamer first, followed by incubating with fluorescently labeled aptamer, and observing whether the fluorescence is blocked under the microscope. We found that all bacteria including the target bacteria had no significant fluorescence. These results of the fluorescence blocking method proved that the aptamer had an excellent affinity and specificity towards the target bacteria , which also verified the determination of the ssDNA concentration method. The affinity constant () of the aptamer measured by the ssDNA concentration method is equal to 33.70±7.83 nmol/L, and its corresponding fitting coefficient is equal to 0.960, which showed that the ssDNA concentration method is accurate, reliable, and feasible in the determination of affinity and affinity constant of an aptamer. The ssDNA concentration method used in this study is practical due to the simple equipment, short measurement time, and convenient operation.