Abstract:

Abstract:

Abstract:The distribution and evolution of nutrients in the Yellow River estuary and its adjacent waters have always been a hot research topic on the geochemical cycle of biogenic elements in the northwest continental margin of the west Pacific Ocean as well as the focus of research on the eutrophication, environmental quality supervision, and comprehensive pollution control in the Bohai Sea. We conducted a survey on the current status of nutrients and related environmental factors in the Yellow River estuary and its adjacent waters in May and August 2023, during which period the primary production was high. Historical data from the past 20 years were collected and summarized to analyze and reveal the potential trends of nutrient composition and structure in the Yellow River estuary and to elaborate the current status and main influencing factors of nutrients in the Yellow River estuary. The main results are presented as follows. The concentrations of dissolved inorganic nitrogen (DIN) varied wider and higher in August than that in May, 2023. The higher concentration was located outside the Yellow River mouth and west Laizhou Bay, and the DIN concentration gradually decreased toward the northwest and southeast from the Yellow River mouth. The lower value appeared in the northern waters of the Yellow River estuary. Although the average DIN concentrations in August were about half that in May, the variation of DIN concentrations in August was larger than that in May, with the highest value in August being about twice that of May and the lowest value being only about 1/10 that of May. Vertically, the surface layer showed the highest DIN concentrations, followed by the bottom layer, and the middle layer showed the lowest concentration. The concentration of DIP located at wider and higher ranges in August than that in May, which was similar to DIN. The DIP distribution was generally similar to that of DIN in tendency, whereas the average DIP concentration in May was comparable to that in August. Meanwhile, the difference in the surface, middle, and bottom concentrations in August was smaller than that in May. The concentration of DIN and DIP in the lower reaches of the Yellow River in May was about 4 and 2 times higher than that outside the mouth of Yellow River, whereas that in August was about 10 and 4 times higher, respectively. Nitrate was the main component of DIN, accounting for 50.7%–96.5% (average 85.2%) in May and 3.9%–99.1% (average 57.1%) in August, respectively. Although the DIN composition was consistent between May and August, the percentage variation in the DIN composition was significant, and the percentage variation between DIN compositions in August was much larger than that in May. The DIN to phosphate ratio (N/P) waved at a wide range in both May and August, with a high average N/P of far from Redfield Ratio, among which the lowest N/P of 43.7 was observed in May. The average N/P of the lower reaches of the Yellow River was 910.7 in May and 410.9 in August. In comparison, the surface N/P was relatively high, approximately 1.5 and 3 times that of the bottom and middle layers. The DIN concentration in the Yellow River estuary increased to 60 μmol/L before 2006, decreased to 30 μmol/L in 2009, and then remained under 30 μmol/L until 2023. The DIP concentration showed peak vales in 2005 and 2013 after increasing from those in 2006 to 2008 and were maintained at lower levels in the other years. The DIN was dominated mainly by nitrate in the past 20 years, followed by ammonia, and then nitrite. In episodic cases, ammonia and nitrite contributed to more than 80% of the total DIN, which may be related to strong terrestrial inputs and weak oxidizing conditions in the bottom water. The nitrogen/phosphorus ratio in the Yellow River estuary continued to increase in the past 20 years. Based on the developing trend of nutrient composition and structure in the Yellow River estuary and its adjacent waters, the imbalance of “more nitrogen and less phosphorus” may be further intensified. DIN and silicate both showed a significant linear correlation with salinity, indicating that surface runoff was the major input pathway for nutrients in the Yellow River estuary. The discrete point plots of DIN components and DO indicated a clustered distribution, indicating that DIN components and DO were influenced by some common factors, and the direct interaction between the two was weak or rapidly reached an equilibrium state. There was no obvious evidence of groundwater transport in study area, and the contribution and mechanism of dissolved organic matter conversion and primary production to nutrients must be understood through further investigation. The results of principal component analysis indicated that the composition and structure of nutrients may significantly characterize the water properties of the Yellow River estuary. The extreme variations in the composition and distribution of nutrients, as well as the high nitrogen/phosphorus ratio, could push the water environment quality situation into a more severe status in the Yellow River estuary.

Abstract:Coastal ecosystems are critical to biodiversity owing to the strong interactions between terrestrial and marine environments. Zooplankton are important secondary producers and major drivers of biological pumps in marine ecosystems and changes in their communities may affect coastal ecosystems. Therefore, it is important to comprehensively evaluate the effects of various environmental factors on the changes in zooplankton communities in coastal ecosystems. Previous research has been limited to a single study on the environmental quality or biological communities in Jinghai Bay, and there have been no relevant reports on the analysis and evaluation of the relationship between changes in zooplankton communities and environmental factors in Jinghai Bay. To study the relationship between zooplankton communities and environmental factors in Jinghai Bay, four survey voyages of environmental factors and zooplankton were conducted in May (spring) and August (summer) from 2021 to 2022. Redundancy analysis (RDA) and Pearson correlation analysis were used to explore the effects of environmental factors on dominant zooplankton species and communities. The results showed that the degree of eutrophication in Jinghai Bay increased significantly from 2021 to 2022, particularly during the summer of 2022 (average eutrophication index EI >200). The distribution of EI showed that the EI was higher in estuaries in the surveyed sea area, and the source of nutrients may be river-diluted water. In addition, there were significant seasonal changes in salinity, especially in the summer of 2022 (the average salinity dropped to 12.06). The average zooplankton abundance in 2021 was higher than that in 2022, and in the summer of 2022, when eutrophication was most severe, the average zooplankton abundance dropped to 54.07 ind./m3. However, during the summer of 2022, which had the lowest abundance, the biomass did not show the same proportion of decline, which requires further study. In addition, Principal Co-ordinates Analysis (PCoA) revealed significant differences between the 2022 summer zooplankton communities and the previous three voyages. A total of 47 species of 10 groups of zooplankton were identified during the four voyages, among which copepods were the most dominant species in spring and pelagic larvae in summer. Acartia pacifica and Zoea larva (Brachyura) were the dominant species on all four voyages. From 2021 to 2022, the dominance of Acartia pacifica showed a continuously decreasing trend. In the spring and summer of 2022, Zoea larva (Brachyura) became the dominant species in the sea area, but the relative abundance and dominance of Zoea larva (Brachyura) decreased in summer. The number of dominant species, across both years, reached an apex (7 species) in the summer of 2022. The interannual and seasonal variation trends of the zooplankton communities, as shown by the Shannon-Wiener diversity index (H′) and Pielou’s evenness index (J), in Jinghai Bay were similar. In terms of seasonal changes, H′ and J in spring 2021 were higher than those in summer. In 2022, the opposite trend was observed, with H′ and J higher in summer than in spring. From the perspective of inter-annual change, both the H′ and J in 2022 were higher than those in 2021. Generally, from 2021 to 2022, the trend of change of zooplankton biodiversity in the Jinghai Bay area gradually increased. RDA showed that the dominant species, Acartia pacifica and Zoea larva (Brachyura), which appeared in all four surveys, were negatively correlated with EI during spring and summer in both years. Eurytemora pacifica, which only appeared in spring, was positively correlated with salinity during the spring of the two years, whereas in the summer of 2022, the newly emerged dominant species Penilia avirostris, Gammaridae, and Schmackeria poplesia were positively correlated with EI, dissolved inorganic phosphate (DIP), and chemical oxygen demand (COD), and negatively correlated with salinity. Pearson correlation analysis showed that EI was negatively correlated with the abundance of zooplankton in all four voyages, positively correlated with abundance and biomass in spring, and negatively correlated with abundance and biomass in summer. In addition, Pearson correlation analysis showed that salinity had an impact on the zooplankton diversity index. Based on the correlation analysis of the RDA and Pearson correlation coefficient, eutrophication was found to be the main environmental factor affecting the community structure and abundance of zooplankton in Jinghai Bay, and salinity affected the seasonal variation in zooplankton. In addition, through PCoA analysis and comparison of data from 2021 and 2022, it was found that occasional events, such as typhoons, may also be an important reason for changes in the zooplankton community. The results of this study will be conducive to a follow-up study of the ecosystem of Jinghai Bay and understanding the factors controlling the dynamics of its zooplankton community, which in turn will contribute to the scientific management of the ecological environment of Jinghai Bay and an in-depth understanding of the mechanisms of zooplankton community change in the bay.

Abstract:Fishing ports play an essential role in fisheries development and supply chains, as all catches are landed through fishing ports to enter the seafood market. Frequent and intensive fishing activities inevitably affect the fishing port ecosystem by discharging sewage and waste oil from sources such as fishing vessels, leaching ship paint, and operating wharves. However, fishing ports are usually located in semi-enclosed seas, leading to the accumulation of land-based pollution in port environments. Fishing ports are known potential sinks for land-sourced pollutants, such as heavy metals, phthalates, polycyclic aromatic hydrocarbons, and petroleum hydrocarbons. Currently, comprehensive research on the distribution and ecological risk of heavy metals in fishing ports is very limited. With no environmental and scientific data available for reference, it is difficult to formulate appropriate pollution control and prevention strategies for fishing ports. Environmental risk assessment in aquatic ecosystems typically uses biomarkers to detect interactions between potential hazards and biological systems. Next to knowing environmental contaminant levels in tissues and the environment, it is important to link to potentially deleterious effects at higher levels of biological organization, such as biochemistry, physiology, and overall health status. Biochemical reactions are frequently used as biomarkers in sentinel model species sampled from reference sites, for monitoring xenobiotic pollution in coastal areas. However, multiple biomarkers were too complex and could not directly reflect the toxic effects of pollutants on organisms, while the integrated biomarker response (IBR) index could overcome this and comprehensively evaluate the aquatic ecosystem health status. The primary purpose of this study was to evaluate the overall pollution level and risks in fishing ports. Samples of green mussels (Perna viridis) and surface water were collected at 12 sampling points in four fishing ports located in Guangdong and Guangxi provinces, China. Six types of heavy metals were measured in the surface water and soft tissues of P. viridis by inductively coupled plasma mass spectrometry. Moreover, biomarkers of the antioxidant defense system, such as superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT), and malondialdehyde (MDA), were analyzed in the hepatopancreas of P. viridis to calculate the IBR index. The results showed that the range of concentrations of Cu, Zn, Pb, Cd, As, and Cr in seawater was 1.13–2.37, 6.67–33.43, 0.14–0.32, 0.03–0.32, 1.09–1.73, and 0.80–1.20 μg/L, respectively, while in the soft tissues of P. viridis, it was 8.31–11.93, 67.92–103.17, 0.88–3.07, 0.80–2.33, 9.61–12.70, and 2.54–4.75 mg/kg, respectively. The mean concentrations of Cu, Zn, Pb, Cd, As, and Cr in each fishing port were lower than the fourth grade of the National Seawater Quality Standard (GB 3097-1997). The concentrations of heavy metals in the surface water and soft tissues of P. viridis were generally at a low level, the comprehensive pollution indices (PI) of heavy metals in the surface water of all four fishing ports were within the natural background range, and the comprehensive pollution indices (Pin) of heavy metals in the soft tissues of P. viridis were in the no pollution category. The PI of heavy metals in the surface water was Shekou Port > Nanwan Port > Dahao Port > Yuwan Port, while the Pin of heavy metals in the soft tissues of P. viridis was Yuwan Port > Shekou Port > Nanwan Port > Dahao Port. In addition, Cu concentrations in both the surface water and soft tissues of P. viridis from Shekou Port were the highest among those from the four fishing ports. Furthermore, there was no significant difference in MDA content or SOD, CAT, and GPx enzyme activities, in the hepatopancreas of P. viridis from the four fishing ports. The IBR index of P. viridis was 4.50 in Shekou Port, 2.14 in Yuwan Port, 1.91 in Dahao Port, and 1.50 in Nanwan Port, which showed a decreasing trend from Shekou Port > Yuwan Port > Dahao Port > Nanwan Port. The highest IBR index was identified at Shekou Port, which agreed with the profiles of the comprehensive pollution index of seawater. According to the correlation analysis, the IBR index showed no significant correlation with the comprehensive pollution index of seawater and marine mussels. It was noteworthy that the IBR index displayed significant positive correlations with Cu concentrations in seawater and mussel tissues, which indicated that Cu pollution may be an important factor to consider in pollution assessments of fishing ports. Overall, this study provides the first evidence of using the IBR index to evaluate the heavy metal pollution status of fishing ports. The pollution level and potential risk of heavy metals in Shekou Port were higher than those of the other three fishing ports. The IBR index coupled with chemical analysis is useful to assess the environmental pollution status of fishing ports for pollution source control and management policy formulation.

Abstract:The Bohai Sea, which is China’s only semi-enclosed inland sea, has received significant inputs of heavy metals from the rapidly industrializing and urbanizing Bohai Sea Economic Rim. Because of the limited water exchange and self-purification capacity, heavy metal pollution in the Bohai Sea has been intensifying, making it a focal area of heavy metal pollution research. Furthermore, the Bohai Sea serves as one of China’s traditional fishing grounds, acting as both a spawning area and foraging ground for various economically significant fishery species, thereby providing abundant seafood for urban and rural communities in the Bohai Sea region. Heavy metals are considered one of the principal pollutants in the marine environment of the Bohai Sea due to their high accumulation and resistance to degradation. They accumulate within organisms and are subsequently transferred through the food chain, resulting in pollution of marine biological resources. The issue of heavy metal pollution in the Bohai Sea has raised concerns among researchers regarding food product safety and human health risks. To assess the status of heavy metal pollution in fish species from the Bohai Sea, we randomly selected 144 samples from 10 different fish species collected during a fisheries resource survey in the Bohai Sea region. All fish samples were stored frozen at –20℃ until further analysis. The collected samples were thawed at room temperature, dissected to obtain muscle tissues, freeze-dried, and subsequently ground into a fine powder using agate mortar. Approximately 0.2 to 0.5 g of the homogenized samples were digested using a microwave digestion system with 2.5 mL of concentrated H2SO4, 4.0 mL of concentrated HNO3, and 1.5 mL of H2O2. The concentrations of heavy metals, i.e., Cu, Zn, Pb, Cr, Cd, and As, were determined using inductively coupled plasma mass spectrometry (ICP-MS). To estimate inorganic As (iAs) in fish muscle from the Bohai Sea, we used the highest reported percentage of iAs in the total As, which was 6.88%, as observed in the literature. The study assessed the single-element pollution and comprehensive heavy metal pollution in Bohai Sea fish using the single factor pollution index, Nemerow comprehensive pollution index (PN), and metal pollution index (MPI). To evaluate the dietary risk posed by heavy metal-contaminated fish, we introduced daily intake (DI) and target hazard quotient (THQ) as assessment indicators. DI represents the daily intake of heavy metals from fish consumption, whereas THQ indicates the potential health risk of ingesting heavy metals. The fish samples collected for this study belonged to 10 different species: Setipinna taty, Scomberomorus niphonius, Pampus argenteus, Trichiurus lepturus, Eupleurogrammus muticus, Engraulis japonicus, Cynoglossus joyneri, Thryssa kammalensis, Liparis tanakae, and Enedrias fangi. The mean values of heavy metal concentrations in the muscle tissues of Bohai Sea fish were as follows: Cu (0.702±0.680) mg/kg, Zn (9.697±5.279) mg/kg, Pb (0.035±0.059) mg/kg, Cr (0.029±0.035) mg/kg, Cd (0.093±0.091) mg/kg, and As (0.959±0.813) mg/kg. When we compared the heavy metal content in fish from the Bohai Sea with those from other sea regions, our findings indicated that there are notable differences in heavy metal levels among fish species in various sea areas of China. Fish from the Bohai Sea exhibit relatively higher levels of Cu, Zn, and As. The single-element pollution index for Cu, Pb, Cd, Cr, and iAs ranged from 0.003 to 0.056, 0.003 to 0.165, 0.021 to 0.767, 0.017 to 0.085, and 0.059 to 0.373, respectively. Some fish samples displayed Cd or iAs contamination, indicating that Cd and iAs were the major contributors to heavy metal pollution in Bohai Sea fish. The PN for the 10 fish species in the Bohai Sea ranged from 0.185 to 1.210, with lionfish exhibiting the highest PN value, signifying severe contamination. The MPI ranged from 0.022 to 0.097, with bluefin trevally, lionfish, and tongue sole having similar and higher MPI values compared to other species. The results of the dietary risk assessment showed that the daily intake of heavy metals (Cu, Zn, Pb, Cd, Cr, and iAs) from consuming Bohai Sea fish was relatively low, and it was below the maximum allowable intake levels established by the Food and Agriculture Organization (FAO) of the United Nations, the World Health Organization (WHO), and the National Research Council (NRC) for Zn, Cu, Pb, Cd, iAs, or Cr. The THQ values for single heavy metals and THQS for multiple heavy metals were all less than 1.0, indicating that consuming Bohai Sea fish is unlikely to pose health risks to consumers. Different heavy metals had varying contributions to THQs, with the average THQ values for Bohai Sea fish species ranking as follows: Pb (0.0029) < Cu (0.0059) < Cd (0.0098) < Cr (0.0105) < Zn (0.0113) < iAs (0.0733). Our findings contribute to a deeper understanding of the heavy metal pollution status in Bohai Sea fishery biota. Additionally, they provide a crucial scientific foundation for ensuring the safety of seafood products from the Bohai Sea. Furthermore, this study offers valuable insights for consumers, enabling the provision of more informed dietary recommendations.

Abstract:Plastic products are widely used in human daily life, while facilitating human life, plastics have also produced many negative effects due to the lack of effective recovery measures, plastic pollution has become an important environmental issue of global concern. Marine plastics can be degraded into smaller microplastics (MPs) through various ways under the influence of environmental factors. They can be ingested by marine organisms mixed with food sources, and then accumulated in the body, causing serious negative effects on marine lifes and marine ecology. Recently, it has been proved that the Changshan Islands sediments contain a certain amount of MPs, the content reached 133.14 to 499.82 n/kg. Changshan Islands is one of the eight major islands in China. It is located at the confluence of the Yellow Sea and the Bohai Sea, and has a unique geographical location. The fish community between the islands is rich in species, especially in many migratory species, and high in species diversity. The seasonal change of fish species composition and dominant species is obvious. It has been proved MPs can cause a certain degree of harm to marine organisms. Therefore, the distribution of MPs in organisms in the Changshan Islands sea area deserves to be studied. In this study, the MPs in the gastrointestinal tracts and muscles of wild fish were digested and separated after collecting them from the marine culture zone of Changshan Islands. The results showed that MPs were detected existing in the gastrointestinal tracts of all fish, but not in muscle tissues. The reason may be that MPs are too large to be endocytosed by intestinal epithelial cells, and thus can not participate in the blood circulation of fish. The abundance of MPs in the gastrointestinal tracts of seven species of marine wild fish ranged from 0.19 to 3.79 items/individual. The abundance of MPs in Coilia nasus is the highest among all fish, this phenomenon may be related to the living environment and predation habits of C. nasus, which living in the bottom of the sea. The MPs in sediments will undergo a cyclic process of suspension, sedimentation and resuspension under the flow of seawater, thus greatly increasing the exposure and intake risk of MPs by bottom organisms, such as C. nasus. The shape of MPs was dominated by the fiber, and the color was mostly transparent, which size is mainly less than 300 μm. The reason may be that the individual size of fish collected in this survey is small, and larger-sized MPs cannot enter into the gastrointestinal tract of fish through feeding. Large plastics in the environment are broken under the photooxidation, wave action, physical wear and alternating freeze-thaw. The different shapes of MPs detected in this study may be derived from the decomposition of these large plastics. Fibers are the predominant form of MPs encountered in global wildlife studies. It could also be because fibrous MPs are the most abundant in the marine environment. Additionally, MPs Fibers can be bended or intertwined with food, possibly due to long-term accumulation for the slower fibers excretion, increasing the chance of being ingested by organisms. The types of polymers detected were cellophane, cellulose and polyethylene, among which cellophane had the highest content. Cellophane is an organic cellulose-based polymer that has been used in food packaging and cigarette packaging. It is also used as a release agent in the manufacture of glass fiber and rubber products, or as a coating in combination with synthetic polymers. There was no obvious toxic effect on cellulose and cellulite. The polymer hazard index (PHI) is an important criterion for risk assessment of MPs, and is based primarily on the percentage content of a given polymer and the polymer’s hazard fraction for ecological and health risk assessment. The potential risk of MPs to humans can be estimated by the polymer hazard index. The higher the polymer hazard index, the higher the ecological risk in the sea area. The toxicity coefficient of polyethylene is 10, the toxicity grade is grade II, the hazard index is low, and it belongs to the low risk polymer. Moreover, people usually discard the gastrointestinal tract of fish before eating, thus the MPs in the fishes of Changshan Islands, and the safety factor of fish products in Changshan Islands is higher. In this study, through the collection of wild fish in Changshan Islands, the MPs in gastrointestinal tract and muscle tissue were extracted, and the abundance and type of MPs were analyzed. The pollution status of MPs in fish in the adjacent waters of Changshan Islands was clarified, which provided basic data for exploring the impact of MPs on the ecological environment safety of Changshan Islands ecosystem. Further research on the distribution of MPs in other wild organisms, such as shellfish, and in other economically viable locations should be considered, which can provide a scientific basis for the analysis of MP pollution levels and the formulation of prevention and control strategies in the marine environment.

Abstract:Owing to the extensive production and utilization of plastic products, a substantial volume of plastic waste is inevitably generated, comprising plastic particles <5 mm in size, which are categorized as microplastics. Microplastics emanating from diverse sources can be classified as primary and secondary. Primary microplastics encompass plastic particles with a diameter <5 mm that are synthetically produced in industrial processes, such as plastic microbeads found in exfoliating scrubs, toothpaste, and facial cleansers. Secondary microplastics result from mechanical abrasion, ultraviolet radiation, biological fragmentation, and other processes acting on larger plastic items. Surveys have revealed the global distribution of microplastics, aptly termed the "PM2.5 of the ocean." Microplastics are a prominent environmental issue, paralleling challenges posed by climate change, ozone depletion, and ocean acidification as an emerging global concern. Consequently, their pollution characteristics and associated ecological and environmental risks have become focal points of global research. Marine microplastics originate predominantly from atmospheric deposition, land-based inputs, coastal tourism, maritime shipping, and fishery production. Seabed sediments serve as a significant reservoir or "sink" for microplastics in oceans. Disparities in microplastic abundance are evident in nearshore beaches, offshore sediments, and deep ocean sediments. The pollution status of microplastics in sediments may be exacerbated in urban nearshore areas owing to intense human activities. However, the status and distribution characteristics of microplastics in urban nearshore sediments remain unclear. Qingdao is a typical coastal and marine city. In this study, we investigated the abundance, morphology, polymer types, and color of microplastics in the surface sediments of the Laoshan coastal area of Qingdao. Microplastics were isolated from surface sediment samples and examined under a stereomicroscope equipped with a camera. Fourier-transform infrared spectroscopy was employed to identify the polymer types of microplastics. The results revealed that the abundance of microplastics in the surface sediments of the Laoshan coastal area of Qingdao ranged from 9.06–34.48 ind./kg with an average of (21.97±8.32) ind./kg. Microplastics with particle sizes <0.5 mm constituted 76% of the total microplastic count. Fiber, granule, and film shapes were predominant, accounting for 53%, 33%, and 14%, respectively. Transparent microplastics were more prevalent than other colors, accounting for 66% of the total. Four polymer types of microplastics were identified: Polypropylene (PP), polystyrene (PS), cellulose, and polyethylene. PP was the dominant polymer type, representing 45%, followed by PS (22%). The microplastic pollution levels in the surface sediments of the Laoshan coastal area of Qingdao were found to be comparatively lower than those in other domestic sea sediments. Considering the heightened human activity in coastal cities, further investigation of the microplastic pollution status and distribution characteristics in coastal areas is warranted. These findings contribute to an enhanced understanding of the environmental risks posed by microplastics to marine ecosystems and provide a scientific foundation for the assessment and management of microplastic pollution in coastal regions.

Abstract:Amidst the rapid expansion of cage aquaculture and marine ranching in Zhoushan, safeguarding the marine environment has emerged as a matter of paramount concern. Microplastics (MPs), recognized as a burgeoning environmental hazard, have been shown to imperil marine organisms by inducing intestinal damage, disrupting feeding patterns, impeding nutrient absorption, and stunting growth when present in sufficiently high concentrations. Tuanjishan Island, once uninhabited but transformed into a landfill site in the 1980s, now faces burgeoning garbage production and gradual saturation of its landfill capacity owing to the city's continuous development. The intricate landfill milieu, characterized by high salinity, fluctuating temperatures and pH, gas emissions, physical pressure, and biodegradation, fosters the breakdown of plastic waste, leading to MP production. This transformation makes Tuanjishan Island a potential source of MP pollution that harbors environmental threats. Mismanagement leading to MP leakage can substantially impact marine aquaculture, impair the fishing economy, and significantly endanger the marine ecosystem. Therefore, predicting the aftermath of MP seepage is imperative. The evolution and sophistication of numerical simulation techniques have emerged as pivotal tools for unraveling and predicting the complex pathways and behaviors of marine pollutants. In light of these advancements, our study meticulously amalgamated the cutting-edge Mike 21 FM hydrodynamic module with a meticulously designed particle-tracking module to investigate the intricate migration trajectories of MP subsequent to their release from Tuanjishan Island. The delineation of the study area leveraged an unstructured mesh encompassing an expansive geographic range, with coordinates extending from 120°E to 124°E and 28.5°N to 33°N. Precise water depth metrics sourced from comprehensive nautical charts and topographical data rooted in the 1985 National Elevation Datum formed the foundational elements of this model. The intricate grid framework of our model, meticulously crafted using surface-water simulation software, comprised an assemblage of 62,614 nodes and 118,040 grids, ensuring a robust representation of the study area. The resolution of this grid was meticulously tailored, standing 400 m near the shoreline and 34,500 m in the expansive outer sea. This meticulously formulated numerical model captured and replicated the complex three-dimensional hydrodynamic flow field across our study area, which was validated through rigorous verification exercises encompassing tide levels and velocities. A meticulously structured sequence of continuous experiments extending across the seasonal spectrum of spring, summer, autumn, and winter was orchestrated to examine the temporal distribution patterns of MPs subsequent to their inadvertent release from Tuanjishan Island. Our findings underscore the intricate migration pathways of MPs, which are intricately intertwined within the proximate surroundings of the Zhoushan Islands and are deeply influenced by the dynamic interplay of ever-changing seasonal ocean currents and the force of monsoons. This delineation highlighted distinctly nuanced shifts in distribution dynamics and migration characteristics across different seasons. Notable concentrations of MPs were detected within the confines of the deep-water channel and northern peripheries surrounding Zhoushan Island, coupled with discernible accumulations observed in various aquaculture locales. The concentration distribution of MPs within the northern region of Zhoushan Island fluctuated between 9%, 10%, 8%, and 6% during spring, summer, autumn, and winter, respectively. Meanwhile, within the deep-water channel bridging Zhoushan Island and Ningbo, MPs exhibited variations of 38%, 36%, 44%, and 42% during spring, summer, autumn, and winter, respectively, indicating inherent seasonally dependent fluctuations. In the hypothetical case of an MP leakage event, an intensified surveillance focus is imperative along the deep-water channel and adjacent coastal sectors in the northern expanse of Zhoushan Island. Alarmingly, predictive models suggest a potential migration of up to 10.11% of the leaked MPs toward aquaculture zones, representing an imminent threat to offshore aquaculture ventures, particularly in the regions situated northwest of Tuanjishan Island. Projections for the winter months anticipated an MP infiltration rate of nearly 2% into Xiangshan Bay, which would affect net cage aquaculture in the area. These intrusive MPs have demonstrated a disruptive capacity within marine ecosystems, disrupting the delicate marine food chain and posing a threat to its delicate balance. Furthermore, our study revealed a dominant concentration of MPs released in varied seasons within coastal precincts along 122.5°E, remaining conspicuously distant from the East China Sea and potentially attributable to the intricate oceanic dynamics. Therefore, while MPs released from Tuanjishan Island may not directly ingress into the Zhoushan fishing grounds, their inevitable impact on offshore net cage aquaculture is definitively substantiated. This comprehensive study represents a pivotal contribution to advancing our understanding of MP pollution within coastal areas and serves as a cornerstone for establishing a robust framework aimed at controlling and mitigating the adverse impacts of MP contamination. However, it is imperative to underscore the limitations of the current model, which regrettably overlooks the pivotal processes integral to comprehending the full scope of MP pollution. The absence of consideration for critical elements such as MP adsorption, biological pollution, sedimentation dynamics, and temporal alterations in the physical and chemical attributes of MPs signals an area demanding meticulous exploration in future research endeavors. Addressing these complexities will pave the way for more comprehensive and nuanced strategies for combating the multifaceted challenges posed by MPs in marine ecosystems.

Abstract:In the process of mariculture, a large number of toxic and harmful substances such as organic matter, ammonia, and nitrite are produced during the metabolism of cultured organisms and the decomposition of feed residuals. If such maricultural wastewater is discharged without purification treatment, it will aggravate the occurrence of eutrophication in the receiving sea area. Constructed wetlands (CW) have received widespread attention due to their low operating costs, simple maintenance, and management advantages. Using CW to treat maricultural wastewater has great prospects. Nitrogen removal is one of the main tasks of constructed wetlands. The characteristics of high salinity and low C/N of maricultural wastewater result in the unique treatment environment and operating mechanism of CW. The substrate can adsorb nitrogen in the constructed wetlands, and nitrogen-cycling microorganisms such as nitrifying bacteria and denitrifying bacteria can attach to the surface to form biofilms. The selection of suitable substrate materials, in addition to zeolite, cinder, sand, and other commonly used water purification materials, can strengthen water purification. Given the low C/N characteristics of maricultural wastewater, materials with slow-release carbon sources can be selected as the filling substrate of constructed marine wetlands. For example, biological carbon sources such as corncob and wood chips, and polymer materials such as PCL and PLC, have recently been used as substrates to fill constructed wetlands and release carbon sources. Meanwhile, substrates that can drive the autotrophic denitrification process of microorganisms such as sulfur autotrophic, hydrogen autotrophic, and iron autotrophic have also been used as a solution. Plants are an important component of constructed marine wetlands, supporting nitrogen removal in four aspects: Nitrogen absorption, oxygen transport, carbon source secretion, and root enrichment of microorganisms. The high salinity environment determines that the wetland plants should be salt-tolerant, and the screening of salt-tolerant plants is a key step in constructed marine wetlands. Currently, Spartina alterniflora, Suaeda salsa, Salicornia bigelovii, Kandelia candel, and similar plants are chosen as candidate plants for constructed marine wetlands. The selection of plants should also consider local conditions, choosing salt-tolerant plants suitable for growing in the local environment. The nitrogen cycle of microorganisms is the main path of biological nitrogen removal in CWs. Various nitrogen-metabolizing bacteria cooperate and restrict each other in CWs, including autotrophic and heterotrophic bacteria, as well as aerobic and anaerobic bacteria. In the process of nitrogen removal in constructed wetlands, dissolved oxygen (DO) is an important environmental factor affecting the distribution and functioning of nitrogen-removing microorganisms. The relatively high DO in the upper layer of the constructed wetland favors the growth and reproduction of aerobic microorganisms, promoting the traditional nitrification process dominated by AOA, AOB, and NOB. The relatively low DO in the bottom layer is more conducive to the growth and colonization of anoxic and anaerobic microorganisms, favoring anaerobic denitrification, Anammox, and DNRA. The occurrence of Comammox can be driven under low nutrient and low oxygen conditions. These bacteria with nitrogen metabolism functions are distributed in different areas, cooperating and restricting each other, forming a complex nitrogen cycle network. Clarifying the basic path of the nitrogen cycle in seawater constructed wetlands is the fundamental basis for regulating the operating parameters of constructed wetlands. The low C/N of mariculture wastewater is not favorable for denitrification by microorganisms. Carbon sources can be supplemented with additional liquid carbon sources, solid carbon sources, and plant litter. DO is the key control index of constructed marine wetlands. The dissolved oxygen content in constructed wetlands is significantly correlated with the community composition of denitrification microorganisms. Therefore, oxygen supply regulation modes, such as continuous aeration, intermittent aeration, and tidal flow, may be effective measures for mariculture wastewater constructed wetlands to improve the overall nitrogen removal performance of wetlands. Accurate regulation of the oxygen supply mode and oxygen supply in constructed wetlands and optimization of dissolved oxygen distribution in different times and spaces within the system are the development trends of nitrogen removal technology in constructed wetlands in the future. The hydraulic operation conditions of CW play an important role in its nitrogen removal effect. Too high or too low indices will affect the efficiency of nitrogen removal in wetlands. Therefore, the optimal control values of hydraulic retention time (HRT), hydraulic loading rate (HLR), and other hydraulic parameters of constructed wetlands also need to be studied. The hydraulic conditions of constructed wetlands also have a significant impact on plant growth, affecting the purification efficiency of plants. In this paper, recent research progress and perspectives on constructed wetlands for the purification of maricultural wastewater and its biological nitrogen removal process were reviewed from four aspects: Selection of substrate, screening of salt-tolerant plants, nitrogen cycling microorganisms, and operation regulation. It is expected to provide a theoretical basis and support for regulating the actual operation of maricultural wastewater constructed wetlands and improving the technical level of maricultural wastewater treatment.

Abstract:Dimethylsulfoniopropionate (DMSP) is one of the most important organic sulfur compounds on Earth, and its significance in global sulfur cycling and climate regulation cannot be overlooked, as it plays an indispensable role in these processes. Phytoplankton are the major producers of DMSP in the marine environment, and nitrogen nutrients are key factors influencing the production of DMSP in phytoplankton. This study focused on two algal species, Emiliania huxleyi (a high DMSP producer) and Phaeodactylum tricornutum (a medium DMSP producer), and conducted indoor culture experiments to compare and analyze the content of particulate DMSP (DMSPp) in the algal culture media under different nitrogen nutrient concentrations and types. The study investigated the relationships between overall DMSPp content, algal density, and DMSPp content per individual algal cell. The results indicated that different nitrogen nutrient concentrations and types had a minimal impact on the content of DMSPp per individual cell in E. huxleyi (P>0.05), suggesting that the DMSPp concentration in the culture media was mostly influenced by algal cell density. Conversely, different nitrogen nutrient concentrations and types had a significant impact on the content of DMSPp per individual cell in P. tricornutum (P<0.05), indicating that the DMSPp concentration in the culture media was mainly influenced by the content of DMSPp per individual algal cell. For instance, in the case of P. tricornutum, the average DMSPp content per individual cell in the low NO3– concentration (0 μmol/L) culture group was 11 times greater than that in the high NO3– concentration (1 764 μmol/L) culture group. Furthermore, under different nitrogen nutrient types, the average total DMSPp concentration in NaNO3 culture media was three and four times higher than that in the NH4Cl and CH4N2O culture groups, respectively. These differences may be attributed to variations in the physiological effects of DMSP on different algal species.

Abstract:Chaetomorpha valida is an invasive species in shrimp and sea cucumber ponds and shallow sea areas in north China. Several studies have investigated its growth characteristics and the environmental factors required to control its bloom and avoid any harmful effects on cultured shrimp or sea cucumber. However, the possibility of utilizing these algae to remove nutrients from eutrophic seawater for breeding and maricultural waste water treatment should be explored because eutrophication has become a serious problem in near-shore sea areas and is threatening the sustainable development of mariculture and the eco-environment of oceans. Nevertheless, no study has been conducted on the nutrient absorption ability or the utilization of live C. valida, except one report on utilizing C. valida as a bio-carbon to absorb phosphate. Temperature and light intensity plays important roles in the nutrient absorption, growth, and reproduction of macroalgae. Different macroalgae species exhibit different responses to changes in temperature and light intensity. To understand the effect of temperature and light intensity on nutrient absorption of C. valida, the ability of the algae to absorb NH4+-N, NO2–-N, and PO43–-P at different temperatures and light intensities was investigated and the best conditions for nutrient absorption were identified in this study. The experiments were carried out in a reformed lab with no natural light. C. valida was cultivated in nutrient-removed sea water after adding a single nutrient and under artificially controlled temperature (5, 15, and 25 ℃) and light intensity (90 000, 180 000, 270 000 μmol photons/(m2·s). Three parallel groups were set up for each environmental factor, and the groups without algae were set up as control. The concentrations of NO2–-N, NH4+-N, and PO43–-P were determined every hour for a total of 40 h. The results showed that C. valida exhibit significant absorption ability for NO2–-N, NH4+-N, and PO43–-P under temperatures of over 5 ℃ and light intensity of over 90 000 μmol photons/(m2·s). Temperature and light intensity was positively correlated with nutrient absorption of C. valida under the range of conditions studied. At 25, 15, and 5 ℃, ammonium was almost totally absorbed at 6 h, 21 h, and 34 h, respectively; nitrite was absorbed at 4 h, 5 h, and 7 h, respectively; and phosphate was absorbed at 5 h, 15 h, and 17 h, respectively, under a fixed light intensity of 180 000 μmol photons/(m2·s). Under light intensities of 270 000, 180 000, and 90 000 μmol photons/(m2·s) and at a fixed temperature of 15 ℃, ammonium was almost totally absorbed at 8 h, 21 h, and 38 h, respectively; nitrite at 4 h, 5 h, and 21 h, respectively; and phosphate at 7 h, 10 h, and 12 h, respectively. C. valida exhibited a high absorption rate in the first 9 h, and the highest absorption rates for ammonium, nitrite, and phosphate were 0.152 0, 0.044 3, and 0.006 5 mg/(g·h), respectively, at the light intensity of 270 000 μmol photons/(m2·s); subsequently, absorption slowed down with time. ANOVA results show that the influence of temperature and light intensity has a superposition effect. The highest absorption rates of NH4+-N, NO2–-N, and PO43–-P were obtained at the highest temperature and light intensity in this study [25 ℃ and 270 000 μmol photons/(m2·s)], which were 99.99%, 99.70%, and 91.31%, respectively. The result of range analysis showed that the values of R (range) of the three nutrients for temperature were 43.06, 51.19, and 55.18 and those for light intensity were 37.82, 43.78, and 21.69, respectively, suggesting the influence of temperature was greater than that of light intensity. Our results provide a new understanding of the nutrient absorption characteristics of C. valida at different temperatures and light intensities. C. valida can absorb nutrients effectively in a shorter time than other macroalgae, with the potential to be used as a tool to purify eutrophicated sea water under controlled conditions. Thus, we provide a new idea and theoretical basis for water purification for breeding and maricultural water treatment. Especially, the absorption ability of C. valida at low and high temperatures may be of great significance for the purification of seedling water and mariculture waste water in the coldest and hotest seasons in north China.

Abstract:Microplastic pollution and excessive discharge of aquaculture tailwater have become important global issues. The use of filamentous algae to remove nitrogen, phosphorus, and microplastics in the same system has not been reported. In this study, we constructed an algal water purification and removal system and investigated its ability to remove microplastics, nitrogen, and phosphorus from the water column. Three kinds of filamentous algae were used, Spirogyra, Enteromorpha, and Cladophora, in microplastic removal experiments in the system; the three species of filamentous algae were the most effective in removing fibrous microplastics (88.50%, 79.50%, 75.50%) and the least effective in removing granular microplastics (67.50%, 53.00%, 55.00%), respectively. Spirogyra removed microplastics better than the other two algae; therefore, it was used for nitrogen and phosphorus removal experiments in the water column. The density of algae per unit area was set to 0 (control), 2, 4, and 6 g/dm2. Spirogyra showed a Total Nitrogen removal rate of 91.88% (4 g/dm2), a Total Phosphorus removal rate of 90.33% (6 g/dm2), and a PO43–-P removal rate of 90.38% (6 g/dm2) after 15 d. The density of 4 g/dm2 showed the highest removal rate, 90.38%. There were no significant differences between the experimental results of the 4 and 6 g/dm2 groups (P>0.05). The results showed that the system effectively removed fibrous microplastics and absorbed nitrogen and phosphorus from the water body and that an algal density per unit area of 4 g/dm2 was the most suitable for removing nitrogen and phosphorus in this system.

Abstract:China is the largest producer of marine aquaculture, and in terms of the output of sea water products, China has ranked first in the world for many years. In the traditional aquaculture process, residual bait and feces produce ammonia nitrogen (NH4+-N) and nitrite nitrogen (NO2–-N), which are toxic to aquaculture organisms. Recirculation in aquaculture systems can be performed using biological filters to purify water and convert NH4+-N and NO2–-N into nitrate nitrogen (NO3–-N) which has a lower toxicity; however, its accumulation leads to chronic adverse effects on aquaculture organisms. Heterotrophic denitrification is a highly efficient biological denitrification technique that converts NO3–-N into harmless nitrogen. Because of the low carbon-to-nitrogen ratio of mariculture wastewater, additional carbon sources are required. Liquid carbon sources have the disadvantages of difficulty in controlling the dosage and the ease of the production of N2O. Solid carbon sources can effectively solve these problems. Biodegradable polymers have a high denitrification efficiency and long duration of action; but their cost is high. Agricultural-waste carbon sources have the advantages of low cost and large quantity, but they are associated with problems of slow start-up and high water color and turbidity in the early stages of operation. A mixture of polycaprolactone (PCL) and corn cob (CC) can overcome the problem of high cost and has excellent denitrification performance, so it is an excellent externally added carbon source. In this study, a denitrification system consisting of inlet tanks (180 L), peristaltic pumps, column reactors and outlet tanks (180 L) was constructed, and a composite carbon source comprising PCL mixed with CC at a mass ratio of 1:1 was used as the denitrification carbon source, and three types of influent nitrate concentrations (INC), 20 mg/L (N-20), 30 mg/L (N-30), and 40 mg/L (N-40), and three temperature conditions, 20℃ (T-20), 25℃ (T-25), and 30℃ (T-30), were set. A 90-d experiment was conducted to investigate the effects of these two factors on denitrification, including a 60-d start-up phase and a 30-d continuous phase. During this period, the inlet and outlet water samples were taken daily to determine the NO3–-N, NO2–-N, NH4+-N, and total nitrogen content using an automatic nutrient salt analyzer (QuAAtro, SEAL, Germany). The inlet and outlet water samples were taken at 61 d, 75 d, and 90 d to determine the chemical oxygen demand (COD), dissolved organic carbon (DOC), and short chain fatty acids (SCFAs). In addition, the biofilm on the carbon source was sampled at 90 d for Illumina MiSeq high-throughput sequencing. The experimental results showed that INC affected denitrification by changing the C/N ratio. In the pre-startup period of the denitrification system (0~10 d), a very high INC (40 mg/L) was detrimental to the operation of the denitrification system, and the nitrate removal efficiencies (NREs) were all lower at 19.52% (N-20), 32.67% (N-30), and 25.28% (N-40). After biofilm maturation, an increase in the INC resulted in a higher denitrification rate but not a significant increase in NRE. The effluent DOC concentration tended to decrease as the INC increased. The optimal INC was 30 mg/L, and the corresponding NRE reached 99.12% in the last 30 days. No obvious accumulation of NH4+-N and NO2–-N was observed during the process, and the decrease in its DOC and DOC concentrations was also stable and rapid. Temperature also had an important effect on denitrification. In a certain interval, the increase in temperature enhanced the denitrification performance and accelerated the nitrogen removal efficiency. During the reactor startup stage, the effluent NO3–-N concentration of each reactor decreased gradually, and the nitrate removal rate increased with an increase in temperature. The NRE of reactor T-30 (70.20%) was higher than that of reactors T-25 (69.96%) and T-20 (28.63%). The effluent DOC concentration of the system at T-30 was significantly higher than that of the other two groups, and the temperatures of 20℃ and 25℃ were more suitable. The optimum temperature was 25℃ when the microbial enzyme activity was higher, and the NRE of the system reached 99.21% in the last 30 days. In contrast, the NH4+-N and NO2–-N produced were also lower, and the organic matter utilization was high. The SCFAs produced by each system had the largest proportion of acetic acid (AC) and no detectable butyric acid, and the AC/PA(propanoic acid) were all >1, which was favorable for denitrification to proceed efficiently. The dominant phylum at different INCs was Proteobacteria, and its abundance decreased with the increasing of INC, which was 54.46%, 39.96% and 24.77% in N-20, N-30 and N-40 groups, respectively. Temperature had a significant influence on the microbial community, and the dominant phylum was Proteobacteria at T= 30℃ and 25℃, with abundances of 55.86% and 38.85%, respectively, whereas the dominant phylum was Bacteroidota (28.87%) at T = 20℃. The dominant genus in all systems was Rhodobacter. In addition, several other active phyla in the denitrification process existed in the system. The influent NO3–-N concentration of 30 mg/L and T = 25℃ were the optimal conditions for the denitrification system using CC+PCL as an additional carbon source. The combination of CC+PCL as a composite carbon source has excellent denitrification performance, which can provide a theoretical basis for the process optimization of solid phase denitrification of mariculture wastewater.

Abstract:Petrochemical wastewater contains heavy metals and petroleum-based pollutants and is a major environmental and biological health hazard. Zebrafish are sensitive to water quality changes and can be used as biological indicators for water quality monitoring. The type, concentration, and toxicity of pollutants in the water can be inferred by observing zebrafish survival, behavior, activity, and other parameters. However, the traditional method of monitoring zebrafish toxicity-response behavior by manual observation and analysis is subjective, labor-intensive, and inefficient. Therefore, automating the monitoring and identification of zebrafish toxicity-response behavior using computer vision technology is an important and challenging research goal. The common methods of computer vision technology in zebrafish toxicity-response behavior monitoring and recognition can be divided into three steps: Foreground extraction, target tracking, and behavior analysis. However, there are problems such as sensitivity to light changes, inability to deal with occlusion and overlapping phenomena, and low efficiency. Therefore, the aim of this study was to improve efficiency and detection accuracy in complex situations such as fish shading for the automated and real-time identification of zebrafish toxicity- response behavior. In this study, four typical pollutants (zinc, chromium, lead, and phenol) in petrochemical tail water were selected to experimentally observe the swimming behavior of zebrafish at different concentrations and exposure times. A multi-target tracking technique based on YOLOv8+ Bytetrack was used to extract the characteristic values of zebrafish movements (average velocity, maximum velocity, minimum velocity, and average number of collisions). YOLOv8 is a deep learning-based end-to-end target detection algorithm that enables efficient and accurate target detection. Bytetrack is a multi-target tracking algorithm based on target detection that can achieve real-time target tracking coupled with the use of low-scoring frames in the tracking algorithm for secondary matching, which can effectively optimize the problem of switching IDs due to occlusion in the tracking process. The convolutional neural network Resnet was used to analyze the motion trajectory maps of zebrafish. The bounding box and confidence level output from the YOLOv8 model were inputted into the algorithm to obtain a unique ID and trajectory for each zebrafish. Finally, zebrafish features such as position, speed, number of wall touches, and trajectory were extracted based on the tracking results. The experimental results showed that the algorithm's tracking accuracy, missing rate, and detection time (per 300 frames) reached 90.26%, 16.33%, and 0.19 min, respectively, which represented a considerable improvement in detection time and accuracy over those of traditional target-detection methods. The tracking accuracy of manual labeling was up to 100%, and the monitoring time was 125.62 min, which was 661.16 times greater than that of the multi-target tracking method in this study. Moreover, the detection times of the threshold segmentation-based Kalman filter, SOTMOT-based multi-target tracking, and FairMOT-based multi-target tracking were 3.59, 0.41, and 0.37 min, respectively, representing 18.89-, 2.16-, and 1.95-fold increases over that of the proposed method, and the tracking accuracies were 67.09%, 88.52%, and 90.10%, which represented only 74.32%, 98.07%, and 99.82%, respectively, of the accuracy of this method. Moreover, the missing detection rates were 72.80%, 20.69%, and 26.45%, which were 4.46, 1.27, and 1.62 times greater than the missing detection rate of this method. This method outperforms other multi-target tracking methods (SOTMOT and Deepsort) regarding target-tracking accuracy and precision. Meanwhile, the proposed method can accurately identify the corresponding movement status and trajectory changes in zebrafish based on specific pollutants. An increase and then a decrease in velocity were observed in zebrafish exposed to zinc sulfate and lead acetate as compared to that of the control group. A significant difference (P<0.05) exists between the effects of zinc sulfate and lead acetate on the increase in velocity of zebrafish at the beginning of the exposure. The velocity of zebrafish in the potassium dichromate-exposed group showed a fluctuating trend, with values slightly lower than those of the control group. In contrast, the proportion of abnormal trajectories was significantly higher (P<0.05) than that in the other experimental groups. Under phenol exposure conditions, the velocity of zebrafish tended to fluctuate over a wide range, while the number of wall touches was significantly higher than that in the other experimental groups (P<0.05). At the late stage of exposure, the velocity of zebrafish in zinc sulfate, lead acetate, and potassium dichromate exposure groups gradually stabilized. The velocity of zebrafish under zinc sulfate and lead acetate exposure conditions tended to decrease significantly. In the potassium dichromate group, the velocity of zebrafish under 1 and 2 TU phenol exposure increased sharply and then fluctuated within a certain range, and 4 TU phenol exposure resulted in partial mortality of zebrafish. In summary, the multi-target tracking method can quickly identify the type of pollutant to which zebrafish are exposed by setting thresholds for the speed, number of wall touches, and percentage of abnormal trajectories in zebrafish behavior. This method is simple, effective, performs accurate identification, and determines real-time responses, making it highly valuable for reference in fish toxicity-response behavior identification.

Abstract:To reveal the differences in the microbial composition of skin mucus and the habitat of Chinese sturgeon (Acipenser sinensis) in different physiological states, we compared the community structure of the skin mucus flora of healthy and sub-healthy Chinese sturgeons and explored the correlation between the structure of the bacterial community and physiological state of Chinese sturgeons. Skin mucus and aquaculture water of healthy and sub-healthy Chinese sturgeons were collected, and the bacterial flora composition was analyzed using high-throughput sequencing technology. The results showed that microbial diversity and richness were significantly higher in the sub-healthy group than in the healthy group (P<0.05). Statistical analysis of the sequences showed that many unique operational taxonomic units (OTUs) were present in the mucus samples compared with the aquaculture water, and the number of shared OTUs with the aquaculture water changed according to the health status. The principal component and principal coordinate analyses between different groups showed a strong separation tendency, and the relative distance between samples within groups was small. There were significant differences in the composition and dominant species of skin mucus and aquaculture water communities of Chinese sturgeons at the phylum and genus levels. The dominant phyla in the water were Proteobacteria (31.84%) and Firmicutes (24.37%); the dominant phyla in the healthy group were Proteobacteria (55.23%) and Bacteroidetes (21.00%); and the percentage of Proteobacteria (40.23%) in the sub-healthy group, but the proportion of Acidobacteria (18.29%), Gemmatimonadetes (10.08%), Actinobacteria, and Verrucomicrobia increased. The dominant flora in water samples were Sphaerochaeta (10.56%) and Cloacibacillus (7.95%). The dominant flora in the healthy group were Ralstonia (19.83%) and Mucilaginibacter (8.11%). The dominant flora in the sub-healthy group, compared to the healthy group, consisted mainly of Sphingomonas (16.71%) and WPS-1_genera_incertae_sedis (5.05%). Functional prediction revealed that the functional abundance of flora in the sub-healthy group showed a decreasing trend compared with that in the healthy group, including environmental adaptation, biological metabolism, and signal transduction. Similarity and difference analysis of the mucus microbiome of Chinese sturgeons in different physiological states and aquaculture water showed that it changed with the physiological state, and the genus Sphingomonas was the dominant genus of mucus flora in the sub-healthy group, which can be used as a health marker. By comparing the microbial communities in the skin mucus of healthy and sub-healthy Chinese sturgeons, different microbial community structures were observed in different health states, which provides a new perspective for their healthy culture and monitoring.

Abstract:The Argentine shortfin squid Illex argentinus is a pelagic neritic cephalopod species found in the southwest Atlantic Ocean. The squid is characterized by a short lifespan, fast growth, and semelparous reproduction. The squid is one of most important targeted species in global cephalopod fisheries and plays a key role as transient “biological pumps” in the southwest Atlantic ecosystem. The squid is a highly migratory species and can migrate hundreds or even thousands of miles between its spawning and feeding ground to complete its life cycle. The squid adopts a mixed income-capital breeding strategy, where the reproduction is mainly supported by energy acquired and mobilized rapidly during the breeding season, coupled with using energy reserves when energy demands for reproduction are robust. The squid shows spatial differences in feeding habits and also shift diets as they grow, leading to different trophic niches. In general, the squid diet shifts from crustaceans to fish and cephalopods as they mature. Given the fact that species in a higher trophic position have greater energy content, a diet shift to fish and cephalopods is expected to meet the energy demands for reproduction. In the present review, we summarized the characteristics of energy accumulated in soma and reproductive systems for I. argentinus, which shows a dramatic increase in energy in the reproductive tissues when the squid reaches maturation. In addition, we compiled previous studies that focused on diet composition and trophic roles in the southwest Atlantic ecosystem, where the squid feeds voraciously on many prey species and plays a key role as prey item for many top predators, including marine mammals. Notably, squid changes diets in relation to energy demands and resource availability. However, future research is still needed to address the functional mechanisms of the shifting dietary habits with growth. To justify the mechanism, we suggest the following priorities in the near future studies: (1) evaluate the relationships between energy acquisition and migratory habits; (2) clarify feeding strategies that potentially maximize the energy gain; and (3) address the potential effect of the ongoing global climate change and fishery activity on the role of energy transfer in the southwest Atlantic ecosystem. Such research would advance our understanding of the species and support the sustainability of resource exploitation.

Abstract:Global biodiversity loss is one of the most serious environmental crises of the 20th and 21st centuries. Fish diversity, an important component of biodiversity, is also in decline. The conservation of fish diversity is therefore essential for ecosystem management and the sustainable use of resources. Most current aquatic biodiversity surveys rely on conventional methods. The emergence and application of environmental DNA (eDNA) technology has triggered unprecedented innovations in aquatic organism investigation and monitoring. eDNA metabarcoding refers to the selection of appropriate universal primers for different target taxa and the identification of multiple target species in environmental samples via PCR amplification combined with high-throughput sequencing. The Zhoushan Archipelago, located in the northeastern waters of Zhejiang Province, consists of 2 085 islands with a total area of 22 216 square kilometers. Located at the confluence of the Yangtze River, the Qiantang River, the Yongjiang River, and the Taiwan Warm Current, the Zhoushan Archipelago forms a unique and complex hydrological environment, which provides a rich source of nutrients and a suitable habitat for fish. In recent years, the large-scale exploitation of marine resources and marine pollution have led to a serious loss of fish diversity near the Zhoushan Archipelago. To restore fish diversity, conservation efforts have been implemented by the government and researchers, including stricter fishing bans and ecological fish stock enhancement and release. However, the lack of reliable tools to understand the species and assess biodiversity is the most critical issue in conservation efforts. Trawl sampling and morphological identification are the primary fish diversity assessment methods. However, these conventional methods are often inefficient, time-consuming, selective, and destructive. The extensive rocky reefs in the Zhoushan Archipelago make it difficult to use conventional fishery resource survey methods efficiently. Therefore, simple and accurate methods for species diversity monitoring are needed. In this study, Xixuan Island, an offshore island in Zhoushan, was taken as a reference. Four different sampling stations were designed, and a total of 12 water samples were collected in February, May, and November 2019. The species composition and diversity characteristics of offshore fish communities around Xixuan Island were described, and the spatial and temporal differences in fish diversity were evaluated. A total of 33 fish species belonging to 12 orders, 26 families, and 32 genera were detected. Among them, Perciformes has the most abundant species, with 19 species, accounting for approximately 57.6% of all species detected. The diversity and evenness indices of the different sampling seasons were significantly different (P<0.05), indicating that season may be one of the factors affecting the diversity of offshore fish around Xixuan Island. The results of the combined temporal and spatial analysis showed that more fish species were detected during the breeding season at sampling sites far from the main island side of Zhoushan. By comparing the results with the conventional fishery resources survey, we found a large variation in the dominant species in different seasons, which may be related to the limited sampling site numbers. Temperature and longitude may be important factors affecting the fish diversity around Xixuan Island. The result of enriching the dominant species in each season by heat trees differed significantly from those of traditional surveys, indicating that eDNA cannot completely substitute the conventional survey methods; however, it can be applied as an auxiliary method to monitor fishery resources, improve detection efficiency, and reduce interference to ecosystems. This study not only provides novel methods for the fishery resources survey in offshore Zhoushan but also provides technical support for the subsequent investigation and evaluation of fish diversity and its protection and management.

Abstract:The Dongwuyang Bay of Fujian Province is the core production area for sea cucumber culture in South China; however, research on the environmental status and bacterial community structure in this area is still limited. In this study, we investigated and evaluated the seawater quality in January, which is the peak culture period for sea cucumber in this area, and analyzed the diversity and differential characteristics of seawater and sediment bacterial community structure using the high-throughput sequencing method and analyzed the correlation between environmental factors and bacterial community structure. The following results were obtained: The Makeng eutrophication index reached 21.60, which indicated that this area was experiencing serious eutrophication. Dong´an and Leijiang were experiencing severe eutrophication, and only the Shawan sea area was experiencing moderate eutrophication. The organic pollution assessment showed that Dong´an and Shawan had light organic pollution, while Makeng and Leijiang had moderate organic pollution. A total of 1 520 OTUs were obtained via high-throughput sequencing, belonging to 28 phyla, 57 orders, 163 families, 322 families, and 581 genera. The diversity analysis revealed that the level of bacterial community diversity in the sediment of each region was significantly higher than that of the corresponding seawater diversity and presented stronger spatial heterogeneity. The diversity and abundance in Shawan were the highest. The results of the principal coordinate analysis and similarity analysis showed that there were significant differences in the bacterial community composition between sediment and seawater. The cluster analysis revealed that the cluster was influenced by water exchange and site location. Vibrionaceae, a family with potential pathogenicity, was one of the dominant species in all seawater and sediment samples. LEFSe (line discriminant analysis effect size) analysis of seawater column samples and sediment samples screened 74 specific OTUs (operational taxonomic units) of bacteria at different taxonomic levels. Prediction of gene function and COG (clusters of orthologous groups of proteins) classification statistics showed significant differences in 12 metabolic pathways between seawater and sediment samples (P<0.05). LEFSe analysis of bacterial community structure from four sites screened 30 specific OTUs at different taxonomic levels. The RDA (redundancy analysis) of the seawater bacterial community structure with environmental factors revealed that temperature, dissolved oxygen, and chlorophyll-a concentration were the major environmental factors affecting the structure in the four sites. Related studies should provide basic data and scientific reference for the management of sea cucumber culture in Fujian Province.

Abstract:To understand the seasonal variations in shellfish community structure, the composition and diversity of shellfish in the intertidal zone of the Changshan Archipelago, China, were investigated. We investigated the composition and diversity of shellfish species in the intertidal zone (including NCD1, NCD2, BCD1, BCD2, TJD1, TJD2, DQD1, DQD2, NHD1, and NHD2) of five representative islands and 10 survey stations in the Changshan Archipelago during the low tide period in July and October 2021 and February and May 2022, and the water quality parameters of point water and interstitial water were analyzed. The results showed that the bottom material of the Changshan Archipelago was generally rock or gravel and the salinity fluctuated with the seasons. The salinity was high in summer and fall while low in spring and winter, with an average of 30.92±0.84. Compared with spring, summer, and fall, the temperature changed significantly in winter, with an average of (4.51±0.32) ℃ and the highest temperature being 23.48 ℃ during spring, summer, and fall. In general, the dissolved oxygen in the main islands of Changshan Archipelago (North and South Changshan Islands) was relatively low, with an average of (8.59±0.38) mg/L, while that in Tuoji Island, Daqin Island, and Nanhuangcheng Island was relatively high, with an average of (9.95±0.51) mg/L. A total of 12 species of shellfish were collected in this survey, including Crassostrea sp., Monodonta labio Linnaeus, Littorina brevicula, Mytilus galloprovincialis, Patelloida pygmaea, Chlorostoma rustica, Reishia clavigera, Thais luteostoma, Ocinebrellus inornata, Cellana toreuma, Ruditapes philippinarum, and Rapana venosa. The number of shellfish species in each season was the highest in spring, with an average of eight species, followed by in fall, with an average of six species. Winter had the least number of shellfish species collected, with an average of four species. In addition, the North and South Changshan Islands presented the most shellfish species at eight and nine species, respectively. The number of shellfish collected in DQD was the least (six species). There were differences in shellfish density at different survey sites, among which DQD2 had the highest density, with an average density of 41.26 ind./m2. The lowest density was in NCD2, and the average density was 13.71 ind./m2. In the same season, the shellfish density at different survey sites was different, the proportion of dominant species at different survey sites was also different. In spring, the density of Crassostrea sp., Cellana toreuma, and Chlorostoma rustica was high, while in summer, the density of Crassostrea sp., Mytilus galloprovincialis, Littorina brevicula, and Cellana toreuma was high. In fall, the density of Crassostrea sp., Littorina brevicula, and Mytilus galloprovincialis was the highest, while in winter, the density of Crassostrea sp., Patelloida pygmaea, and Cellana toreuma was the highest. The species diversity (H´) of shellfish in each tidal area of the survey site is shown as middle tidal zone (1.44) > low tidal zone (1.40) > high tidal zone (0.44). Overall, the average shellfish species diversity in the 10 survey sites was spring (1.26) > winter (1.21) > fall (1.11) > summer (0.95). The average evenness index of species (J´) in the 10 survey stations was 0.55, among which the evenness index of DQD2, NCD2, NHD2, and TJD1 shellfish was 0.67, 0.64, 0.64, and 0.55, respectively. The evenness index of shellfish in each tidal area of the survey site was in order of middle tidal zone (0.67) > low tidal zone (0.59) > high tidal zone (0.29). The seasonal change was not obvious. The average shellfish abundance at the survey site was 0.533, including 0.82, 0.68, 0.60, and 0.53 shellfish abundance at NCD2, NHD2, BCD2, and NCD1, respectively. The abundance index of shellfish in each tidal area of the survey site was in order of middle tidal zone (0.56) > low tidal zone (0.52) > high tidal zone (0.17). Overall, the average shellfish abundance in the 10 survey sites was spring (0.681) > fall (0.573) > summer (0.458) > winter (0.418). Seasons (temperatures) have the most obvious impact on the diversity and abundance of intertidal shellfish resources. Anthropogenic activities, such as aquaculture and tourism, also affect the distribution and quantity of shellfish species. The results of this study not only enrich the data of intertidal shellfish resources on islands in China but also provide basic information and reference for biodiversity and environmental protection of intertidal shellfish on the Changshan Islands.

Abstract:The morphological characteristics, ecological habits, and physiological structures of fish vary rapidly in the early growth and development stage. Understanding the changes in the postembryonic early development morphology of organs or tissues could provide basic information for the classification or identification of fish and the investigation or utilization of early resources. Morphological characteristic observation is one of the most commonly used methods. The existing studies on the early development of a certain trait in the late larval and juvenile stage are unclear. Most of the digital images are side views, resulting in a lack of dorsal views, ventral views, or local images, which is not conducive to the understanding of larval and juvenile. In this study, the variations in observable, measurable, and countable traits in postembryonic early development of Thamnaconus septentrionalis were explored using optical microscopy and stereomicroscopy. The observable traits included the developmental changes of yolk sac, scales, and notochord curvature. The measurable traits included the growth changes of total length, body length, body height, and oral fissure. The countable characters included the number of fins in the second dorsal fin, anal fin, caudal fin, and the number of pterygiophore. The results showed that at (23.0±1.0) ℃, the newly hatched larvae moved up and down in the water, with a phenomenon of convergent clustering under light irradiation; the pectoral fin appeared at 1 d, and the heart rate was approximately 50 beats/min; the optic capsule was completely black, the maxillar appeared, the oral fissure extended below the eyes, the yolk sac disappeared, and the first dorsal fin base appeared at 4 d; the girdle was single and colorless and began from the front edge of the belly, the oil globule disappeared, and the pelvic fin base appeared at 7 d; the first dorsal fin base and pelvic fin base broke membrane at 8 d; the first dorsal fin and pelvic fin were significantly longer than before, transparent rounded scales were observed at 9 d; swimming ability was improved, indicated by hovering back and forth gnawing pool wall, transversally conical barbs appeared at the base of the first dorsal fin at 11 d; the swim bladder and postcleithrum on both sides of the belly were clearly visible at 12 d; the second dorsal fin and anal fin pterygiophore appeared at 14 d, the second dorsal fin pterygiophore spans seven sarcomeres and the anal fin pterygiophore spans six sarcomeres. There are 10~12 melanin masses in the second dorsal fin and 5~6 in the anal fin, the conical teeth appeared on the upper and lower jaws at 16 d; the second dorsal fin and anal fin were differentiated at 19 d; the swim bladder fills about 25% of the abdominal cavity; notochord curves upward clearly and the stalk of caudal fin came out at 20 d; the fin strips of the second dorsal fin, anal fin, and caudal fin were well-developed at 24 d; the open mouth was round and changed from swallowing to sucking at 30 d, and the rainbow cells were colorful and the body surface near the gill was the most dense; the abdominal cavity has a strong metallic texture, the hard spine of the pelvic fin was degraded, close to the girdle at 39 d; 4~5 black bands were arranged regularly on the body, and the lateral line was consistent with the vertebral column at 50 d. The R2 values of the curve equations of measurable traits were all greater than 0.90. The height of the first dorsal fin increased exponentially 4~30 d. The visual sac increased linearly from 0~11 d to 11~20 d, and the increment of 0~11 d was less than that of 11~20 d, while the increment of 21~30 d was exponential. The oral fissure in 4~13 d and 22~30 d increased linearly, while that in 13~22 d decreased linearly. The total length, body length, and body height increased linearly before the inflection point and exponentially after the inflection point, which was concentrated in 19~23 d. The number of fin pterygiophore of the second dorsal fin was 31~36, while that in the anal fin was 33~35. The number of pectoral fins and caudal fins were 15~18 and 12, respectively. In conclusion, metamorphosis occurs in the postembryonic early development of T. septentrionalis, showing an allometric growth pattern, with a slow linear growth in the rotifer-feeding stage and a fast exponential growth in the artemia-feeding stage. The changes in the first dorsal fin and pelvic fin were correlated with age in days. The changes in the second dorsal and anal fins were correlated with the total length. This study enriches the early biological theory of T. septentrionalis and provides basic data for the investigation or utilization of its seedling breeding and resources. The partial enlarged detail is helpful for scholars to further understand the larval or juvenile fish and perform deep research.

Abstract:The insulin-like growth factor (IGF) system plays a key regulatory role in biological processes, such as growth, reproduction, early development, cell differentiation, and proliferation, in fish and other vertebrates. The IGF family system consists of two IGFs (IGF1 and IGF2), two IGF receptors (IGF-1R and IGF-2R), and six IGF-binding proteins (IGF-binding proteins IGFBP1–6) in mammals. By binding to the cell surface-specific receptor (IGF-Rs), IGFs activate the activity of related protein kinases, initiate the signaling cascade in the cell, and participate in the regulation of various biological reaction processes. The research on fish IGFs mainly focuses on IGF1 and IGF2, and several studies have confirmed that they are widely involved in multiple biological processes, such as growth regulation, immune response, gonadal development, and maturation. IGF3 is a fish-specific sub-type; it was first detected in zebrafish (Danio rerio) and Nile tilapia (Oreochromis niloticus) in 2008 and plays an important role in gender differentiation in many reports. However, it is unclear whether it affects ovarian development. Turbot (Scophthalmus maximus) is an economically valuable flatfish species that is widely farmed in Europe and China. It cannot spawn eggs naturally in captivity and needs to be hand-stripped and artificially fertilized. The IGF system plays an important role during turbot ovary maturation. In this study, the full-length turbot IGF3 sequence was obtained via RACE cloning technology, its bioinformatics characteristics were analyzed, and the tertiary structure and docking situation were predicted. The relative expression levels during the whole tissue and ovarian development were detected using qRT-PCR. Hematoxylin-eosin staining was used to determine the gonadal development stage. The results showed that the full-length cDNA sequence of turbot IGF3 was 1 255 bp, encoding 259 amino acids, and displayed the highest homology with halibut (Hippoglossus stenolepis). A total of 33 amino acid signal peptides and 59 amino acid IGF-specific domains were found. It was a hydrophilic protein with a molecular mass of 26.39 kDa and a hydrophobic isoelectric point of 8.13. The turbot IGF3 is highly homologous with other fish. Turbot IGF3 is a hydrophobic protein, and its tertiary structure consists of three α spirals connected in series, closely integrated with IGF-1R and IGF-2R. igf3 is expressed differently between female and male turbot. In the liver, kidney, stomach, gonads, and brain, the male IGF3 expression was significantly higher than in females. In the heart, intestines, and pituitary gland, female IGF3 expression was significantly higher than males; however, the difference was not significant in the spleen. In male turbot, igf3 is mostly expressed in the brain, while in female turbot, igf3 is mostly expressed in the brain and intestine. During the five stages of development of turbot ovaries (previtellogenesis, early vitellogenesis, late vitellogenesis, migratory nucleus, and atresia), igf3 expression increased significantly in late vitellogenesis and maintained high expression in the migratory nucleus stage and decreased in the atresia stage. The results showed that turbot IGF3 has typical IGF family structure characteristics: local endocrine factors, widely distributed in tissues, significant sex dimorphism, and participates in regulating the growth and maturation of oocytes. Therefore, these findings lay a foundation for an in-depth research on the effects of IGF3 on ovarian development and oocyte maturation in turbot and provides crucial ideas for exploring the novel functions of IGF3 in fish.

Abstract:The whitespotted conger (Conger myriaster) is an important fish species with high economic value in China and Japan owing to its high nutritional content. It occupies an important position in marine-eel export in China. C. myriaster has the potential to be artificially cultured and has promising economic prospects. Thus, there is growing international interest in C. myriaster research. Indeed, to meet the demands of domestic and foreign markets, artificial culturing of C. myriaster is urgently required. However, at present, there is no report on the research of C. myriaster reared in industrial recirculating aquaculture systems. Feeding frequency has been reported to affect feed intake and growth performance in a number of fish species. Feed management in terms of optimization of feeding frequency has emerged as one of the crucial areas of research in the field of aquaculture. Overfeeding and leftover food disrupts the water quality, while inadequate food supply has direct impact on production cost. Traditional flow through systems uses large volumes of water, and intensified aquaculture has drawbacks, such as large amount of discharged waste from effluent water. Among the excreted nutrients, nitrogen and phosphorus constitute the largest proportion. Sustainability with low environmental impacts is one of the concerns when developing aquaculture systems. Based on the concepts of reduced water consumption and efficient water management, recirculating aquaculture systems offer the possibility of creating a sustainable future for fish production. Thus, a 70-day experiment was conducted to investigate the effects of different feeding frequencies and water quality on the growth performance and physiological and biochemical indices of C. myriaster [body weight: (150.64±5.43) g] reared in industrial recirculating aquaculture systems; physiological and ecological methods were used in this study. The experiment comprised two feeding frequency groups, feeding once (group T1) and twice (group T2) per day, and each treatment group had three replicates. The average body weight of the fish in each culture pond and the water quality indices of the culture system were measured and analyzed every 14 d. At the end of the experiment, the blood, liver, and intestine samples of C. myriaster were collected to determine and analyze growth and physiological indices. The results indicated that (1) the growth performance of group T2 was better than that of group T1. The final body weight, final body length, weight gain, and specific growth rate of group T2 was significantly higher than those of group T1 (P<0.05). The feed coefficient of group T1 was significantly higher than that of group T2 (P<0.05). However, no significant differences were found in the final body height, liver body ratio, visceral body ratio, condition factor, or survival rate at different feeding frequencies (P>0.05). (2) The serum malondialdehyde of group T1 was significantly higher than that of group T2 (P<0.05). The hepatic alanine aminotransferase and hepatic aspartate transaminase of group T1 was significantly higher than those of group T2 (P<0.05). The intestinal trypsin of group T2 was significantly higher than that of group T1 (P<0.05). There were no significant differences in the levels of serum superoxide dismutase, serum catalase, intestinal amylase, or intestinal lipase between groups T1 and T2 (P>0.05). (3) In the 24 h changes of ammonia nitrogen (NH4+-N) and nitrite nitrogen (NO2–-N), the change patterns of the two indices in outlet of the T1 group were similar to those of the outlet of the bio-filter; the two indices of the outlet water in T1 and T2 groups were one (24:00) and two peaks (20:00 and 08:00) respectively; the difference in NH4+-N in the outlet water of the two test groups within 24 h was significant (P<0.05); at 04:00 and 08:00, the NO2–-N content of the outlet water of the biofilter was significantly lower than that of the other two test groups (P<0.05); the 24 h changes range of NH4+-N and NO2–-N in T2 group were smaller than those in T1 group. (4) During the whole test period, each water quality index was maintained within the safe breeding range, and no significant differences were found in NH4+-N, NO2–-N, chemical oxygen demand, and active phosphate concentration between T1 and T2 groups (P>0.05); at the end of the test, the total nitrogen and total phosphorus of T2 group were significantly higher than those of T1 group (P<0.05); the biofilter has different treatment effects on various water quality indices, among which the treatment effect on NH4+-N is the most efficient, up to 54.23%. Based on the above results, the feeding frequency of twice per day was better than that of once per day in the industrial recirculating aquaculture mode for C. myriaster (body weight: 150~250 g). Without affecting the survival rate and condition factor, the twice per day group can save costs and improve economic benefits. However, due to the limitations of the test conditions, we did not explore more times of feeding, and further tests are needed in the future. The predominance, which was the high efficiency of water qualities treatment and optimal effect of culture, was demonstrated obviously in closed recirculating aquaculture systems. This study aims to provide a theoretical reference for the appropriate feeding strategy in the industrial recirculating aquaculture model of C. myriaster.

Abstract:Hybrid F1 is a novel hybrid of Erythroculter ilishaeformis (♀) and Ancherythroculter nigrocauda (♂). Current studies have focused on the genetic features, evaluation of fillet quality, farming techniques, and so on. Meanwhile, few studies have addressed enhancing fillet quality. Traditional intensive pond aquaculture is the major culture method in China. A large amount of excreta and feed residuals remaining in the aquaculture water bodies restricts the critical nutrients, flesh texture, and taste of the fillet. In this study, juvenile hybrid F1 cultured in ponds was obtained from the seed farm (Wuhan Xianfeng Aquaculture Technology Company Limited, Wuhan, Hubei) as the study subject. The fish was treated with short-term micro-flow water (1.2 BL/s) to improve muscle quality. The effects of treatment time (0, 4, 8, and 12 d) on muscle quality in juvenile hybrid F1 flesh were investigated following short-term micro-flow water treatment by assessing the color, muscle fiber, texture, nutrient composition, and the contents of amino acid and fatty acid. The color of the samples was determined by a CR-400 automatic chromaticity meter (Konica Minolta, Japan), wherein the color was expressed by L, a*, and b*. The skin yellowness values were significantly higher at treated 8 d and 12 d, while the short-term micro-flow water treatment significantly influenced muscle redness (P<0.05). The texture property of fish is one of the keys to its quality. The juvenile hybrid F1 flesh showed a significant improvement in fiber density, hardness, chewiness, and springiness by 70.96%, 92.04%, 198.99%, and 35.71% compared with the control group, respectively after 8 d treatment. Frozen fresh samples were lyophilized for the following analysis. Kjeldahl and Soxhlet-Henkel methods were used for the determination of total, crude protein (N=6.25) and fat content respectively. The protein and fat content significantly decreased in the flesh following the extension of micro-flow water treatment time, while the ash content did not significantly change (P>0.05, the same below). The amino acid content presented the highest values of glutamic acid, followed by proline, and the lowest values in cystine among the 17 amino acids detected in the fillet of juvenile hybrid F1. The amino acid composition was assayed by a Hitachi L-8900 automatic amino acid analyzer. The amino acids EAA/TAA of all groups were about 40%, and the amino acids EAA/NEAA was over 60%, which were high-quality proteins complied with the Food and Agriculture Organization of the United Nations and the World Health Organization (FAO/WHO) standards. Umami flavors are closely related to the composition and ratio of umami amino acid. The content of total amino acids and essential amino acids showed a decreasing trend under the influence of micro-flow water. The total of delicious amino acid treated at 4 d were significantly higher than those of juvenile hybrid F1 treated at 8 d and 12 d. The TAA content and total EAA content did not dramatically change. The essential amino acid index (EAAI) is used to indicate how close the essential amino acid content is to the standard protein. EAAI were 77.33%, 74.91%, 76.56% and 76.75%, respectively. The fatty acid composition was assayed using gas chromatography (Agilent 7890A, USA). Nineteen kinds of fatty acids were detected in the muscle of juvenile hybrid F1, including 5 kinds of saturated fatty acids, 6 kinds of monounsaturated fatty acids, and 8 kinds of polyunsaturated fatty acids, respectively. The micro-flow water significantly affected the fatty acid content in muscle. The content of cis-15-tetracosenoic acid (6.85 mg/100 g), eicosatrienoic acid (41.05 mg/100 g), and arachidonate (53.84 mg/100 g) was significantly higher than that in the other groups after treatment for 8 d. Meanwhile, the content of α-linolenic acid (48.05 mg/100 g), monounsaturated fatty acid (MUFA) (1 336.43 mg/100 g), and n-6 polyunsaturated fatty acids (524.43 mg/100 g) were higher than that in the other groups at 4 d and 12 d. This study indicated that short-term micro-flow water treatment improves the color of skin and meat, and the meat texture at 8 d while ensuring the quality of the flesh. The short-term micro-flow water treatment can help to provide new ideas to improve the quality of aquatic products cultured in ponds.

Abstract:Aeromonas hydrophila (Family: Aeromonadaceae) is a traditional aquatic animal pathogen. It has been widely prevalent in our country since the 1980s and 1990s, causing diseases in a series of freshwater fish, leading to serious economic losses. It has been confirmed to be one of the main pathogens in freshwater aquaculture worldwide. The American shad (Alosa sapidissima) is one of the biggest shad in the world and grows much faster than other shad. Because of its delicious taste as reeves shad (Tenualosa reevesii), American shad were introduced into China from the USA by the Shanghai Fisheries Research Institute for local farming in 1998 and is widely welcomed in Shanghai, Jiangsu, and Zhejiang Provinces. In recent years, with the rapid development of the American shad culturing, fish diseases have become a major threat to fish farming. However, because most of the American shad are cultured in extensive ponds, few diseases have been reported in China. In this study, we reported a case of A. hydrophila infection in American shad. In 2021, the disease outbreak was observed in American shad cultured in Linyi City, Shandong Province, with severe mortality. The daily mortality could be up to 2.5%. The fish were cultured in indoor ponds for breeding and outdoor ponds when fish reached 300 g. The American shad were cultured with underground water and water was changed 1–3 times daily. The water temperature was 18–20 ℃. The fish were fed largemouth bass (Micropterus salmoides) commercial feed, and the daily feeding rate was approximately 2%. The disease broke out in the indoor ponds first, and then in outdoor ponds. The cumulative mortality was approximately 90% in 2 months. Enrofloxacin was administered orally, but no effects were observed and the disease continued to progress. The typical disease symptoms in the American shad were furunculosis or ulceration, with shedding scales and surface bleeding, especially on the tail, sometimes swollen and pus-filled. In autopsies of the diseased fish, ascites were found in the fish abdomen, and dark red necrosis on the liver, with sepsis and enteritis. The liver, spleen, and kidney of American shad with typical symptoms were collected and cut into about 1 cm3 tissue blocks, immersed in Davidson's Fixative (Davidson's AFA) for 24 h, and preserved in 70% ethanol. The tissues were mounted onto glass slides with hematoxylin and eosin staining for histological analysis. Histopathological results showed swollen liver cells, vacuolar degeneration, basophilia, and diffuse necrosis; the spleen showed hemorrhagic anemic necrosis, nuclear rupture, and atrophy. Glomerular atrophy of the renal corpuscle, cells in the proximal and distal tubules cytoarchitectural loss, and necrosis and shedding of kidney lymphocytes was also observed. No parasite was found on the fish surface, fins, in the gills, or internal organs with the naked eye and a light microscope. Freshwater viruses, such as Cyprinus herpesvirus type 2, largemouth bass ranavirus, megalocytivirus, and rhabdovirus, were checked by polymerase chain reaction (PCR), and no viruses were detected. The liver, spleen, and kidney of the diseased fish were sampled and cultured in tryptic soy agar medium (TSA) and Luria-Bertani agar medium (LB) plate medium at 28 ℃ for 24 h. Several pure and dominant colonies with the same morphology were observed on all the plates. These colonies were purified and cultured. The 16S rRNA gene sequencing results of all the purified colonies showed that the dominant strains were of the same species. The typical isolate was purified and named AS-AH2101. The results of biochemical identification with Biolog GenⅢ showed that the isolate AS-AH2101 was negative to gentiobiose, stachyose, D-raffinose, α-D-lactose, D-melibiose, 3-methyl glucose, D-fucose, D-sorbitol, D-arabitol, and Myo-inositol, while positive to dextrin, D-maltose, D-trehalose, D-cellobiose, sucrose, D-turanose, and β-methyl-D-glucoside. According to the Biolog GenⅢ identification system database, the biochemical characterization of AS-AH2101 was similar to that of A. hydrophila, with a confidence of 0.999. The 16S rRNA gene sequence of AS-AH2101 was submitted to GenBank databases under the accession number OP787967 and blasted in GenBank and EzTaxon. Comparison of the 16S rRNA gene sequences showed 99%100% identity with those of A. hydrophila. The phylogenetic tree was constructed using Mega 7 with the Aeromonas typical strains 16S rRNA gene sequences obtained from GenBank, and the phylogenetic analysis also clustered AS-AH2101 with A. hydrophila. Thus, the molecular analysis results identified the SC18032201 strains as A. hydrophila, and the phenotype also supported this result. Because of the strong stress response of American shad, it is difficult to perform the experimental culturing and infection in the laboratory. As a classic pathogenic infection model organism in aquatic animals, blue gourami (Trichogaster trichopterus) is a traditional model for fish pathogen study and has been widely used in the research of A. hydrophilia and E. piscicida. Therefore, in this study, blue gouramis were used as the model organism for virulence evaluation of AS-AH2101 in the experimental infection. The results of the challenge experiment showed that the death of the blue gourami infected via intramuscular injection was observed on the third day post infection. The infected fish showed redness, bleeding, and scale shedding at the injection site, congestion or bleeding at the base of the fin, abdominal ascites, and liver necrosis, which were similar to the naturally infected American shad. The isolate strain AS-AH2101 showed high virulence to blue gouramis, with the median lethal dose (LD50) of 3.23×104 CFU/fish. The virulence genes of A. hydrophilia were also detected by PCR, and results indicated that AS-AH2101 possessed six virulence genes, including aerolysin (aerA), hemolysin (hlyA), extracellular protease (ahpA), anti-metalloproteinases (ast), enterotoxin (altA), and quorum sensing gene (luxS). Antibiotic sensitivity studies showed that AS-AH2101 was resistant to cefradine, amoxicillin, ampicillin, and erythromycin. These results provided important information for disease control and A. hydrophila prevention and control of American shad culturing in China.