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STUDY ON THE IN-SITE MITIGATION OF BIO-CLOGGING OF CONSTRUCTED WETLANDS BY TWO BIO-ELECTROCHEMICAL SYSTEMS
LI Zhi-Wei, LIN Li-Li, LIU Yong-Hong, ZHANG Bo, LU Rui, WU Zhen-Bin, XIAO En-Rong
 Available online   , doi: 10.7541/2022.2022.0154
[Abstract](0) [FullText HTML](0) [PDF 1728KB](0)
Abstract:
Constructed wetland bio-clogging is caused by the massive secretion of Extracellular Polymeric Substances (EPS) by microorganisms, which is more common and unavoidable. Bio-electrochemical Systems (BES) using additional electric fields or self-generated electric fields can promote the dispersion, degradation and utilization of EPS, and can control and alleviate the bio-clogging of constructed wetlands to a certain extent. Therefore, in this study, two bio-electrochemical systems, CW-MEC and CW-MFC, were used to intervene in constructed wetlands in different bio-clogging periods. removal) and the content of stratified EPS in the vertical direction, to evaluate the mitigation ability of the two BES systems to bio-clogging at different intervention timings. The results show that the two BES systems can show good clogging mitigation effect. Intervention before clogging, the CW-MFC system can well maintain the removal effect of pollutants COD and TN (91.4%—61.3% and 65%—40%) and increase the water output rate (the change rate is 67.6% in the CW group). The CW-MEC system can significantly reduce the EPS content of the upper layer, restore the porosity of the system, and have a better effect of relieving clogging. Both of these two BES have good application potential in alleviating bio-clogging in constructed wetlands in situ, which provides reference for the effective prevention and control of bio-clogging in constructed wetlands during different periods.
DEGRADATION AND TRANSFORMATION OF E2-3S IN HORIZONTAL FLOW CONSTRUCTED WETLAND
FU Jing-Miao, LI Rui, YANG Yang, ZHANG Xiao-Meng, CHEN Shan-Xu, LU Hui-Lin, WANG Xue-Li, HUANG Meng-Na, WANG Xi-Yan, DAI Yu-Nü
 Available online   , doi: 10.7541/2022.2022.0157
[Abstract](0) [FullText HTML](0) [PDF 3248KB](0)
Abstract:
Conjugated estrogens (CEs) are an important existing form of natural estrogens, which can be hydrolyzed into free forms (Free estrogens, FEs) producing strong endocrine interference effects in the environment. Therefore, they have potential environmental risks. The degradation and transformation behavior of CEs with complex structure is not clear in the sewage treatment system, which is the key to accurately assessing the comprehensive removal of estrogens. This study adopted the method of simultaneous analysis and detection of 15 kinds of FEs and CEs, to study the degradation law of 17β-estradiol-3-sulfate (E2-3S) in planted or unplanted horizontal flow constructed wetlands (HFCWs) and the transformation relationship between various estrogens. The results showed that under the condition of hydraulic residence time of 1.5 days, the removal efficiency of E2-3S reached 98% in the substrate depth of 15 cm at the inlet end of HFCWs. E2-3S could be transformed into other estrogens, which were the most abundant in FEs (accounting for more than 70%). Plants could significantly increase the wetland DO concentration and reduce the residual concentration of FEs. The removal efficiency of total estrogens by planted CW(P-CW) and unplanted CW(U-CW) were 84% and 56%, respectively. The major transformation pathway of E2-3S in HFCWs was to break the thioester bond to E2 and then oxidize to E1, followed by direct oxidation to E1-3S and then hydrolyzing to E1, and a small amount of path is hydroxylation to form E3-3S and then hydrolyzing to E3. In addition, E2-3S could also produce trace D-CEs (< 5% of total estrogen), and the residual quantity in P-CW is lower.
RECYCLING EFFECT OF WASTEWATER FROM FISHPOND BY THE PADDY FIELD WETLAND
TAO Ling, PENG Ge-Ge, CHEN Si-Yuan, HAO Liu-Liu, DAI Li-Li, PENG Liang, LI Gu
 Available online   , doi: 10.7541/2022.2022.0178
[Abstract](1) [FullText HTML](1) [PDF 1303KB](0)
Abstract:
The environmental concern and safety of pond aquaculture has become an important factor restricting the development of freshwater aquaculture in China. Research and development of efficient, ecological, safe and healthy culture technology and model has become an important development direction of domestic and foreign fishery production. By the ecological engineering of paddy fields, the integrated paddy-pond recirculating ecological system was constructed. In the system, the paddy field acted as the production unit, as well as water and nutrient recycling purification unit. In this paper, the treatment efficiencies of the wastewater from pond aquaculture by the paddy field wetland and food production potential of the rice field and fishpond were evaluated for two consecutive years. Biolog Eco micro-plate technology was used to compare the metabolic structure and diversity of microbial community in the inflow and outflow of the paddy field wetland. The results showed that under the hydraulic loads of 0.51 m/d and 0.74 m/d, the percent removal rates of NH4+-N, NO2 --N, TN, TP and CODMn by the paddy field wetland were 2.94%—33.38%, 12.60%—92.45%, 16.06%—32.36%, 23.34%—48.50% and 15.37%—61.12% respectively. The content of DO in outflow of the paddy field wetland was significantly increased. Paddy wetland retained nitrogen and phosphorus from pond culture wastewater, therefore, the pollution discharge coefficient of TN, TP and CODMn of wastewater from the pond culture decreased by 0.71—3.03、0.27—0.31和4.19—8.81 kg/t, respectively. The carbon source utilization characteristics of microbial communities in the inflow and outflow of the paddy wetland were changed, and the metabolic functional diversity index of microbial communities in the outflow were significantly higher than that in the inflow which was beneficial for the fish pond culture. Fish yield in the pond of the recirculating aquaculture system increased by more than 22% than that in the control pond and rice yield reached 9239 kg/hm2 in the system. These results probably implied that the rice field exhibited efficient treatment effect and nutrient recycling effect of wastewater from aquaculture pond and the integrated paddy-pond system could provide more food for people. The new recirculating planting-culture ecosystem integrating rice and fish production could be regarded as a clean and sustainable food production system from an environmental point of view and be applied in rural areas of China where crop breeding and fish culture coexist.
EFFECTS OF OVERWINTERING ON ANTIOXIDANT CAPACITY AND FATTY ACID COMPOSITION IN GRASS CARP (CTENOPHARYNGODON IDELLUS)
WU Wen-Yi, JI Hong
 Available online   , doi: 10.7541/2022.2020.213
[Abstract](144) [FullText HTML](54) [PDF 1247KB](6)
Abstract:
Grass carp Ctenopharyngodon idellus often faces the stress of overwintering in natural environment. It has been reported that fish can induce excessive production of reactive oxygen species (ROS) and cause oxidative stress during overwintering. Excessive ROS may destroy the balance and stability of antioxidant system, resulting in cell damage, nuclear apoptosis and fatty acid peroxidation. Especially during the period of overwintering, the fatty acids as the main energy supply material, have been greatly threatened and challenged because of the fatty acids maybe influenced by ROS. The most direct effect is to reduce the level of energy supply, the antioxidant capacity and immunity of the body, and increase the mortality rate, and to explore the relationship between oxidative stress and the change of fatty acid composition in tissues, In order to explore the metabolic adaptation mechanism of energy utilization of grass carp during overwintering, the initial body weight of (1053.33±16.11) g was investigated, 45 fish were randomly divided into three repetitions. When the water temperature (15℃) naturally decreased to the point that the grass carp ceased ingestion, the experiment began. When the water temperature (15℃) naturally rose to the point that the grass carp refeeding commenced, the overwintering period and our experiments were concluded. Samples were collected after 0, 1, 2, 4, 8, 12 and 16 weeks of natural overwintering. Biological indexes, antioxidant capacity of hepatopancreas, muscle, foregut, adipose tissue and serum and the fatty acid composition of hepatopancreas, muscle and adipose tissue were measured. The relationship between antioxidant capacity index and fatty acid composition was also studied association analysis. The results showed that the BW, CF, HSI, VSI, IFI and LW decreased significantly (P<0.05), while KI and SI increased significantly (P<0.05). Adipose tissue, hepatopancreas and muscle are the three tissues with the largest oxidative stress. During the overwintering period, the proportion of PUFA in hepatopancreas had a major effect on the total fatty acid composition (principal component load characteristic value >0.5), the proportion of C18﹕2n-6 and C16﹕0 in muscle had a major impact on the total fatty acid composition, and the proportion of PUFA, n-6 PUFA, SFA and MUFA in adipose tissue had a major impact on the total fatty acid composition. At the same time, association analysis showed that SFA in adipose tissue of grass carp was mainly positively correlated with oxidative stress and the fish body damage during the period of overwintering. PUFA and MUFA in muscle were positively correlated with oxidative stress and the fish body damage, while MUFA in hepatopancreas was mainly positively correlated with oxidative stress and the fish body damage. This study showed that the grass carp was subjected to strong oxidative stress during the period of overwintering, especially in adipose tissue; the fatty acid ratio of hepatopancreas, muscle and adipose tissue changed significantly, the correlation analysis with antioxidant index showed that SFA in adipose tissue, MUFA in hepatopancreas, PUFA and MUFA in muscle were related to oxidative stress have a more direct relationship between stimulation and injury. The baseline information provided in this study can be used to formulate effective feeding strategies before overwintering, and to make appropriate management and feeding decisions during the period of overwintering, and the recovery stage after overwintering and improving the survival rate and production efficiency of grass carp after overwintering.
THE CHARACTERISTICS, DEVELOPMENT AND APPLICATION OF THE BIOACTIVE PRODUCT SQUALENE
CHENG Peng-Fei, WANG Hai-Xia, BO Ya-Hui, ZHOU Cheng-Xu, YAN Xiao-Jun
 Available online   , doi: 10.7541/2022.2021.0206
[Abstract](232) [FullText HTML](160) [PDF 1042KB](9)
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Squalene is a precursor of several common sterols, which has important biological activities and food and drug value. Due to the protection of marine wildlife and fishery resources, the commercial production of squalene is greatly limited. It is very important to find new sustainable sources for the production of squalene. This research mainly expounded the biological activity, source and application of squalene, analyzed the biosynthetic pathways and extraction methods of squalene from different biological sources, and put forward the economic feasibility for green production of valuable squalene by microorganisms (especially for microalgae), which aim to provide a theoretical basis for the industrial production and application of squalene.
TRANSCRIPTOME ANALYSIS OF TWO DIFFERENT MORPHOLOGICAL POPULATIONS OF SCYLLA PARAMAMOSAIN UNDER COLD STRESS
CHEN Jie, DONG Meng, CUI Yue, FANG Cui-Lian, CHEN Xu-Tang, LIU Zi-Ming
 Available online   , doi: 10.7541/2022.2021.129
[Abstract](576) [FullText HTML](320) [PDF 1405KB](12)
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Scylla paramamosain is an economically important crab species mainly distribute in the southern reaches of the Yangtze River. Based on many years of breeding experience, it is speculated that different morphological populations of Scylla paramamosain have acquired different adaptations to temperature changes. In this study, Sp1 populations, with “one spine on the outer margin of the carpus of cheliped ” and Sp2 populations, with “two spines” were designated research objects, and 8℃ low temperature stress and 20℃ control groups were established. The hepatopancreas was used as a transcriptome source, and an Illumina Hiseq4000 platform was used for sequencing. A total of 81853 Unigenes were obtained by sequencing, with an average length of 420 bp, and an N50 of 1460 bp. Among these, 22.33% were known genes. Screening for differentially expressed genes identified 15773 Unigenes displaying significant differential expression in the Sp1 population under low temperature stress. Of these, 15628 Unigenes were down-regulated, and 145 Unigenes were up-regulated. Functional enrichment analysis revealed that the differentially expressed genes encode abundant cytosolic, RNA binding, and ribosome structural constituent proteins, and were further enriched in signal pathway components including ribosome, spliceosome, and protein processing components in the endoplasmic reticulum. 323 Unigenes were significantly differentially expressed in the Sp2 population under low temperature stress, of which, 209 Unigenes were down-regulated, and 114 Unigenes were up-regulated. Functional enrichment analysis revealed that the differentially expressed genes include abundant contributors to steroid hydroxylase activity, DNA integration, and bicarbonate transmembrane transporter activity, and were also enriched in signal pathways including lysine degradation, glycine, serine, and threonine metabolism, and arginine and proline metabolism. The above results indicated that as water temperature droped from 20℃ to 8℃, expression of a large number of genes was down-regulated in the Sp1 population, and physiological activities were drastically reduced. This may correspond to winter behaviors of Scylla paramamosain, including fasting and dwelling in burrows. By contrast, the Sp2 population mainly adjusted its material metabolism, indicating that the Sp2 population was relatively slow to respond to sudden temperature drops. Transcriptome data has been verified by RT-qPCR, supporting the feasibility of comparing responses of two morphological populations to low temperature stress based on transcriptome analysis. This study will inform selection of cold-tolerant and high-quality varieties for breeding of Scylla paramamosain.
ANTIOXIDANT AND NON-SPECIFIC IMMUNE RESPONSES OF FOUR SPECIES OF FISH LARVAE UNDER AMMONIA STRESS
WU Xue-Yang, GUO Hong-Hui, KUANG Yu, ZHANG Ce, YANG Hui, TANG Rong, ZHANG Xi, LI Da-Peng, LI Li
 Available online   , doi: 10.7541/2022.2021.0180
[Abstract](251) [FullText HTML](158) [PDF 1265KB](10)
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Ammonia, a ubiquitous pollutant in the aquatic system, has been proved to be high toxic to fish. The early life state of fish is acturally more sensitive to the influence of external environmental factors than adult fish since its antioxidant and immune systems are in lower level. However, there are limited information on difference responses of different species of fish larvae to ammonia stress. Aim to explore the effects of ammonia on the antioxidant and non-specific immune response of different species of fish larvae, four different feed-habits species of fish larvae (omnivory silver carp Hypophthalmichthys molitrix, herbivorous grass carp Ctenopharyngodon idella, Wuchang bream Megalobrama amblycephala and predacity yellow catfish Pelteobagrus fulvidraco) were selected and exposed to different concentrations of total ammonia nitrogen (0, 1 mg/L, 2 mg/L and 3 mg/L) for 96h. The results showed that acute ammonia exposue caused significant decrease of body length on four species of fish larvae in a dependent-concentration manner (P<0.05). Meantime, ammonia exposure significant decreased levels of total antioxidant capacity (T-AOC), catalase (CAT) and glutathione peroxidase (GPx) in four species of fish larvae (P<0.05). Also, 2 and 3 mg/L ammonia significantly reduced the activities of superoxde dismutase (SOD) in the larves of grass carp, Wuchang bream and yellow catfish (P<0.05). As for antioxidative-related genes, significant decrease of transcriptional levels were only detected in yellow catfish gpx and silver carp sod after ammonia exposure (P<0.05). In terms of immune parmameters, ammonia exposure significantly up-regulated transcriptional levels of immune-related genes in four species of fish larvae except silver carp interleukin1β (il1β) (P<0.05). By contrast, significant decrease of lysozyme contents were observed in grass carp larvae after ammonia exposure, and significant increase of lysozyme contents were detected in yellow catfish larvae exposed to 2 mg/L ammonia (P<0.05). The results of two-way ANOVA also confirmed that ammonia could caused to varying degree changes for all parameters including antioxidant enzymes, immune indexes and immune-related gene expression in four species of fish larvae (P<0.05). There were significant differences on levels of T-AOC, CAT, GPx, C3 and the gene expression levels of cuznsod, gpx, il1β, c3 among four species of fish larvae. However, interactive effects of ammonia and species were only observed on C3 contents and the expression of genes gpx, tnfα, il1β and c3 (P<0.05). In summary, ammonia exposure caused oxidative stress, decreased the antioxidant capacities and disturbed innate immune responses in four species of fish larvae, which led to growth retardation. Among four species of fish larvae, the predacity yellow catfish was the weakest to ammonia tolerance, the herbivorous Wuchang bream and grass carp is the next, and omnivory silver carp were the strongest.
EFFECTS OF HYPOXIA STRESS AND SPIROPLASMA ERIOCHEIRIS INFECTION ON THE SURVIVAL AND APOPTOSIS OF ERIOCHEIR SINENSIS
WANG Ya-Qin, HUANG Chen, GENG Chao, MIAO Yan-Yang, LU Yin-Yue, GU Wei, WANG Wen, MENG Qing-Guo
 Available online   , doi: 10.7541/2022.2021.0272
[Abstract](670) [FullText HTML](388) [PDF 4633KB](7)
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The Chinese mitten crab (Eriocheir sinensis) is often caused by hypoxia to impact growth and immunity. E. sinensis tremor disease is caused by the pathogen Spiroplasma eriocheiris, which is highly contagious with a high mortality rate. To explore the effect of hypoxia stress and the S. eriocheiris infection on E. sinensis, the mortality rate and the histopathology of crab, the Spiroplasma copy number, apoptosis & necrosis and mitochondrial membrane potential in hemocytes were studied. The results showed that long-term hypoxia stress damaged hepatopancreas and gill tissues. Multiple vesicles appeared in the crab hepatopancreas after 8d hypoxic stress. The crab gill tissue was diffused after 8d hypoxia stress. The mortality of E. sinensis in hypoxia group under S. eriocheiris infection at 10d was 100%, meanwhile, the mortalities in normoxia group under S. eriocheiris infection at 10d and 12d were 50% and 70%, respectively. The mortality of E. sinensis in hypoxia group at day 12 (60%) was higher than the mortality in normoxia group (30%). These results reveal that the hypoxia could accelerate the crab dead under S. eriocheiris infection with increased copy number of S. eriocheiris in hemocytes and enhanced hemocytes necrosis rate (PI stain) and apoptosis rate (FITC/PI stain). Hypoxia increased the mitochondrial membrane potential in hemocytes under S. eriocheiris infection at 4d, 6d and 8d, and S. eriocheiris infection induced the mitochondrial membrane potential in hemocytes. These results indicate that hypoxia stress and S. eriocheiris infection have a synergistic harmful effects on the physiology and biochemistry of E. sinensis.
REQUIREMENT LEVEL OF DIETARY PROTEIN FOR MONOPTERUS ALBUS
JIANG Wen-Hao, YANG Xin, ZHOU Qiu-Bai, WANG Zi-Rui, GAO Miao, WANG Ying, HU Zhong-Hua, ZHANG Wen-Ping
 Available online   , doi: 10.7541/2022.2020.238
[Abstract](450) [FullText HTML](268) [PDF 1230KB](11)
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The study was aimed to determine the requirement level of dietary protein for ricefield eel (Monopterus albus). The experiment was divided into 7 groups, with 4 replicates in each group. Each replicate included 25 ricefield eels with an initial weight of (17.1±0.2) g. The ricefield eels were fed seven isoenergetic diets respectively for 60d containing different levels of crude protein (32.74%, 38.50%, 42.26%, 48.35%, 52.65%, 57.87% and 62.31%) with fish meal and casein as the main protein source. Then the production performance, protein retention rate, physiological and biochemical indexes were evaluated to determine the requirement level of dietary protein. The results showed that 1) with the increase of feed protein level, the feed conversion ratio was decreased and then increased, but the weight gain rate, specific growth rate, protein efficiency ratio and protein retention rate increased first and then decreased. The growth performance was the highest in 57.87% of dietary protein group, and the protein retention rate was the best in 52.65% of dietary protein group. 2) with the increase of dietary protein level, the BUN in ricefield eel serum increased, the activities of GPT and GOT in ricefield eel serum decreased and then remained unchanged, the contents of TC, TG, HDL-C, LDL-C and blood glucose decreased, and the MDA content in ricefield eel serum decreased first and then increased, which was the lowest in 52.65% of dietary protein level. 3) With the increase of dietary protein level, the protease activities in liver, stomach and intestine of ricefield eel increased significantly and then increased slowly, the lipase activity in liver decreased significantly and then decreased slowly, the lipase activities in stomach and intestinal showed no significant difference, and the amylase activities showed no significant difference in stomach which decreased significantly first and then decreased slowly in liver and intestinal. These results indicated that ricefield eels will increase the feeding to supplement the protein requirement with low level of dietary protein. While the level of dietary protein was too high, the oxidative stress increased, thus ricefield eels will reduce the protein intake by reducing food intake. With the regression analysis of weight gain rate, protein efficiency ratio, protein retention rate and feed conversion ratio, the appropriate requirement level of dietary protein for ricefield eels is 48.51%—52.60%.
DIFFERENT EFFECTS OF VARIOUS DIETARY STARCHES ON THE GROWTH AND METABOLISM OF RAINBOW TROUT JUVENILES
DANG Jiang-Yu, CAI You-Wang, ZHANG Chun-Yan, CAO Kai-Lin, LI Xiao-Qin, LENG Xiang-Jun
 Available online   , doi: 10.7541/2022.2021.0150
[Abstract](575) [FullText HTML](333) [PDF 957KB](7)
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To investigate the effects of various starches on the growth, serum biochemical indices, and metabolism of rainbow trout juveniles, cassava starch, wheat starch, pea starch and corn starch were included in diets at the same level of 15% to form four isonitrogenous and isolipidic diets to feed rainbow trout with an initial body weight of (7.7±0.1) g for 56 days. The results showed that the cassava starch group had the highest weight gain (1049.3%) and the lowest feed coefficient ratio (0.83). There were no significant difference in viscera-body ratio, liver-body ratio, and proximate composition of whole body and liver among all the groups (P>0.05). The liver glycogen content of pea starch group was significantly lower than that of other groups (P<0.05). There were no significant differences in serum triglycerides, total cholesterol, aspartate aminotransferase and alanine aminotransferase activity among all the groups (P>0.05). The liver hexokinase activity of cassava starch and wheat starch groups was significantly higher than that of pea starch and corn starch groups (P<0.05), and the liver phosphofructokinase activity in corn starch group was significantly lower than that in cassava starch and wheat starch groups (P<0.05). Serum glucose content reached to the highest value at the 7th hour in all groups, and cassava starch group showed significantly higher serum glucose content than the other three groups from the 1h to 7h (P<0.05). No significant difference in liver tissue morphology was observed among all the groups. In conclusion, cassava starch is a suitable starch source for rainbow trout juveniles.
APPARENT DIGESTIBILITY OF SIX NEW NON-GRAIN PROTEIN INGREDIENTS FOR LARGEMOUTH BASS (MICROPTERUS SALMOIDES)
SHI Yu-Hui, ZHU Shu-Jie, TAN Bei-Ping, ZHANG Wen-Bin, ZHOU Xiao-Qiu, GAO Wei-Hua, CHI Shu-Yan
 Available online   , doi: 10.7541/2022.2021.0350
[Abstract](90) [FullText HTML](51) [PDF 800KB](4)
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Apparent digestibility of dry matter (ADMD), crude protein (ACPD), crude lipid (ACLD), gross energy (AED) and amino acids (AACA) in methanotroph (Methylococcus capsulatus, Bath) bacteria meal (MBM), cottonseed Clostridium autoethanogenum protein (CAP), Tenebrio molitor meal (TMM), enzyme-treated soy protein (ETSP), Chlorell meal (CM) and cottonseed protein concentrate (CPC) were determined respectively for largemouth bass (Micropterus salmoides). Apparent digestibility coefficients were determined by using a reference diet with 0.1% Yttrium oxide indicator and test diets that contained 70% reference diet and 30% of the test ingredient being evaluated. The largemouth bass with an average body weight of (19.28±0.10) g were randomly divided into 7 groups with 3 replicates per group and 30 fish per replicate. ADMD coefficients of test ingredients ranged from 37.27% to 86.43%, and ADMD coefficients for CAP and MBM (82.77%—86.43%) were significantly higher than TMM, ETSP, CM, and CPC (P<0.05). ACPD coefficients of test ingredients ranged from 79.97% to 88.45%. The higher ACPD coefficients were observed in CAP and MBM (87.44%—88.45%), followed by TMM, which were significantly higher than those of CPC (79.97%; P<0.05); AACA coefficients of test ingredients ranged from 70.52%—90.51%. The variation trend of the AACA coefficients was basically consistent with that of ACPD coefficients. ACLD coefficients of test ingredients ranged from 51.19%—97.48%. The highest ACLD coefficients were observed in CAP and MBM (94.79%—97.48%). ACLD coefficients for TMM, ETSP and CM (63.11%—88.98%) were significantly higher than those of CPC (51.19%; P<0.05). AED coefficients of test ingredients ranged from 43.25% to 85.43%. The highest AED coefficients were observed in CAP and MBM (83.91%—85.43%). AED coefficients for TMM, ETSP and CM (72.09%—80.53%) were significantly higher than those of CPC (43.25%; P<0.05). These results indicate that CAP and MBM are the best protein source for largemouth bass, and TMM, ETSP and CM are also high-quality protein source for largemouth bass.
MOLECULAR CLONING OF MCCASPASE 3-4 AND ITS FUNCTIONS IN MYTILUS CORUSCUS LARVAL METAMORPHOSIS
LIU Zhi-Xian, LI Jia-Zheng, LIANG Lin-Li, FENG Dan-Dan, LIANG Xiao, YANG Jin-Long, ZHU You-Ting
 Available online   , doi: 10.7541/2022.2021.0179
[Abstract](380) [FullText HTML](251) [PDF 1155KB](11)
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As an essential way to remove redundant tissues or cells, apoptosis plays a crucial role in regulating the metamorphosis of vertebrates and insects. However, little is known about the function of apoptosis and its executioner genes caspase-3 in the metamorphosis of marine mollusks. In the present work, a full-length caspase-3 cDNA had 1269 bp full length and 855 bp sequences of Open Reading Frame (ORF), which coded a polypeptide of 1486 amino acids, was cloned from the Mytilus coruscus. It encodes a predicted protein containing conserved caspase p20 and p10 domains and has the caspase family cysteine active site (KPKIFIFQCSRR) in the p20 domains. However, compared with conserved caspase 3 in other species, three amino acids sites of the pentapeptide active motif (QACXG, where X is R, Q, or D) at the end of the cysteine active site is mutated. Multi-sequence alignment and phylogenetic analysis showed that this gene had the highest similarity with the Caspase 3/7-4 gene in Mytilus galloprovincialis and was named McCaspase 3-4. The mRNA expression level of McCaspase 3-4 in different tissues of the adult and larval metamorphosis were analyzed by real-time quantitative PCR. Results showed that McCaspase 3-4 mRNA expression in the mantle and labial palp was significantly higher than that in other tissues and was 9.34 and 7.37 times higher than that in the adductor muscle, which had the lowest expression level, suggesting this gene might be involved in host immune defense or tissue renewal and repair of labial palp. The McCaspase 3-4 mRNA expression also increased at 12 to 24h after the epinephrine inducing and peaked at 24h after inducing when the expression level was 3.0 times higher than that at 0h, indicating the McCaspase 3-4 might play a role in the early stage of larval metamorphosis (larval metamorphosis usually begin at 48 to 72h after epinephrine inducing). Furthermore, after the pediveligers were transfected with the specific McCaspase 3-4 siRNA by electroporation, the metamorphosis rate of pediveligers at 48h induced by epinephrine was decreased to 5%, which were significantly lower than the epinephrine inducing after non-target gene siRNA transfected group (25.7%). These results indicated that although the pentapeptide active motif of McCaspase 3-4 is mutated, this molecular still has the function of metamorphosis regulation and plays a role in the early stage of larval metamorphosis. However, we do not know whether the active site mutation will affect its metamorphosis regulation effect. These results will contribute to understanding the role of apoptosis in mussel larval metamorphosis and the molecular mechanism of marine mollusks’ metamorphosis.
EFFECTS OF SODIUM CHOLATE ON GROWTH AND GLUCOSE METABOLISM OF LARGEMOUTH BASS (MICROPTERUS SALMOIDES)
DING Fei-Fei, ZHOU Nan-Nan, ZHANG Le, QIAO Fang, DU Zhen-Yu, ZHANG Mei-Ling
 Available online   , doi: 10.7541/2022.2021.043
[Abstract](900) [FullText HTML](502) [PDF 1493KB](12)
Abstract:
Cholic acid (CA) is the main bile acid (BAs) in fish. To explore the effects of cholic acid on the growth and glucose metabolism of largemouth bass (Micropterus salmoides), we formulated an experimental diet by adding 300 mg/kg sodium cholate (CAS) in the control diet for an 8-week trial before analyzing the effects of CAS on the growth, intestinal microbiota, gene expression of glucose metabolism and bile acid receptor Farnesoid X receptor (FXR) of largemouth bass. The results showed that CAS had no significant effects in the growth index, including weight gain rate (WGR), condition factor (CF), carcass ratio (CR), viscerosomatic index (VSI), hepatosomatic index (HSI) and mesenteric fat index (MFI). CAS also did not affect the body composition including moisture, crude protein, crude lipid and crude ash of the largemouth bass, and it did not regulate the intestinal microbiota of the largemouth bass. Sodium cholate promoted glycogen synthesis and activity of glycogen synthase (GCS) in liver but not in the muscle without impacting the activity of glycogen phosphorylase a (GPa) in the liver. Sodium cholate significantly promoted the expression of fructose-1,6-bisphosphatase (FBPase), glucose-6-phosphatase (G6Pase), and pyruvate kinase (PK) in liver, and it significantly decreased hexokinase (HK) genes in muscle. Moreover, sodium cholate significantly inhibited the expression of the bile acid receptor Farnesoid X receptor (FXR) in the liver, without changing the expression of intestinal FXR. These results suggest that 300 mg/kg CAS did not affect growth and body index of largemouth bass, and that CAS may regulate glucose metabolism via liver FXR expression.
CLONING OF CDNA OF BCL-2 GENE OF MARSUPENAEUS JAPONICUS AND ITS ROLE IN COLD RESISTANCE
SHAO Hui-Xin, REN Xian-Yun, YU Zhen-Xing, LI Jian, LIU Ping
 Available online   , doi: 10.7541/2022.2021.131
[Abstract](443) [FullText HTML](290) [PDF 1455KB](5)
Abstract:
As an important apoptosis regulatory gene, Bcl-2 gene plays an important role in the endogenous apoptosis pathway. In this study, the full-length cDNA sequence of MjBcl-2 gene was cloned by rapid amplification of cDNA ends (RACE) and analyzed by bioinformatics; Real time quantitative PCR (qPCR) was used to analyze the expression level of MjBcl-2 in different tissues of Marsupenaeus japonicus after different temperature stresses and RNA interferences; TUNEL technique was used to detect the apoptosis after the interference of MjBcl-2. The results showed that the full length of MjBcl-2 cDNA was 2432 bp, the open reading frame was 726 bp, encoding 241 amino acids with a molecular weight of 26.80 kD; Domain prediction analysis showed that MjBcl-2 contained typical conserved domains of Bcl-2 family. The results of multiple sequence alignment and phylogenetic tree construction showed that MjBcl-2 was highly conserved and similar to other species. Quantitative PCR results showed that MjBcl-2 was expressed in all tissues of M. japonicus, with the highest expression in muscle, the second in gill and the lowest in heart. Under the stress of low temperature (10℃ and 16℃), the expression of Bcl-2 gene in gill and hepatopancreas of M. japonicus increased gradually, and reached the highest point at 72h; After interference with Bcl-2 gene, the expression of Bcl-2 gene was down-regulated and Caspase-3 gene was up-regulated at all temperatures (P<0.05); There were only a few apoptotic cells in the 28℃ treatment group. TUNEL results showed that the number of apoptotic cells increased with the decrease of temperature in RNAi group and NC group (control group). There were a small number of apoptotic cells in 28℃ treatment group, but there was no significant change in the number of apoptotic cells in RNAi group and NC group. The number of apoptotic cells in 10℃ hypothermia treatment group was significantly higher than that in NC group; After 12h, the apoptosis rates of gill tissue in 28℃ NC group, 28℃ RNAi group, 10℃ NC group and 10℃ RNAi group were 1.73%, 2.35%, 21.59% and 33.70% respectively. The results showed that MjBcl-2 may play an important role in the immune defense response of M. japonicus to low temperature stress.
WHOLE-GENOME SEQUENCING AND PHOSPHORUS UPTAKE AND TRANSPORT PATHWAY COMPARATIVE ANALYSIS OF CYLINDROSPERMOPSIS RACIBORSKII N8
CHEN Zhi-Jiang, RUAN Zi-Xi, CHENG Nan, XIAO Li-Juan, PENG Liang, HAN Bo-Ping, LEI La-Mei
 Available online   , doi: 10.7541/2022.2021.0197
[Abstract](227) [FullText HTML](140) [PDF 3246KB](4)
Abstract:
Tropical cyanobacterium Cylindrospermopsis raciborskii have attracted much attention due to their rapid expansion into temperate regions. Studies on understanding the genomic characteristics will help analyze their environmental adaptation mechanisms at the molecular level. The genome of C. raciborskii N8 isolated from Zhenhai Reservoir in Guangdong Province was sequenced using PacBio single-molecule real-time sequencing technology, SMRT, and then performed comparative analysis of the whole genomic characteristics. It was shown that genome size of the strain N8 was 3.857 Mb with 40.13% GC content, similar to other 27 C. raciborskii strains in NCBI databases. Further annotation predicted 3598 protein coding genes, representing the largest number among the published genome data. The number of genes annotated in COG, KEGG and GO databases were 2429, 1664 and 2244 respectively. The phylogenetic tree constructed with a single copy gene of the whole genome indicated that C. raciborskii N8 was the most closely related to strain CR12 from Singapore. We inferred that C. raciborskii N8 did not produce toxin cylindrospermopsin (CYN) and saxitoxin (STX) due to the absence of biosynthesis gene cluster for both toxins in its genome. Through examining the genomes of N8 and other 7 strains, we found that these strains had relatively complete phosphorus (P) uptake and transport pathway genes (two-component regulation system, low affinity inorganic P transport gene, high affinity inorganic P transport system, organic phosphonate transport complex, C-P lyase and alkaline phosphatase). This finding demonstrated that C. raciborskii has the potential to utilize different P forms in the ambient environment. However, differences in gene copy number and arrangement among these strains were observed, for example, two C-P lyase complex proteins genes (phnF and phnM) were found in the genome of C. raciborskii N8, while only one in the other 7 strains, indicating variation in C. raciborskii strains. This study represented the first reported whole genome of Chinese C. raciborskii strain. The analysis of its genomic characteristics and P metabolism pathway will help clarify the winning strategies of C. raciborskii in South China.
THE CLONE OF FOXO1 GENE AND ITS SPATIOTEMPORAL EXPRESSION IN GDNAD COMPLEX OF NEWLY HATCHED ALLIGATOR SINENSIS
WEN Yue, HU Cheng-Cheng, WANG Ren-Ping, XIA Yong, XIA Tong-Sheng, HE Ming-Yue, ZHOU Yong-Kang, ZHANG Tong-Xin, ZHANG Jun, PANG Yu, WANG Hao, SHEN Xin-Yu, NIE Hai-Tao, WU Xiao-Bing
 Available online   , doi: 10.7541/2022.2021.122
[Abstract](446) [FullText HTML](313) [PDF 1150KB](4)
Abstract:
The molecular cloning and sequence characteristic analysis of Forkhead Box protein-1 (FoxO1) CDS region were investigated using the gonad tissues of Alligator sinensis in the resent research. Immunohistochemistry was used to detect its expression patterns in in peripheral tissues such as stomach, intestine, and lung. Immunofluorescence (IF), RT-PCR and Western blot were used to investigate the temporal and spatial expression patterns in the gonadal tissues of Chinese Alligator at different days of age (17, 63 and 96 days of age), and to investigate the biological functions in the gonadal development of Chinese alligator. The complete coding region of FoxO1 gene and predicted coding amino acid were 1941 bp and 646 aa, respectively. Homology comparison and phylogenetic tree analysis showed that FoxO1 of Chinese alligator was more closely to birds than reptiles such as Pelodiscus sinesis and Protobothrips mucrisquamatus. Other vertebrates such as mammals, teleost, and amphibians were also grouped into subgroups, suggesting that FoxO1 is both functionally conserved and species-specific. FoxO1 protein expressed in the newly hatched crocodile gonadal complex, lung, stomach and intestine. FoxO1 expressed significantly higher in the ovarian tissue at the 17th days post-hatched than that at the 63rd and 96th days post-hatched individuals. FoxO1 mainly expressed in the adrenal and mesonephrons but weakly expressed in the cortex of ovarian collected at the 17th days post-hatched, and its irregular and randomly distribution was gradually upregulated in the cortex at the 63rd days post-hatched individual. FoxO1strongly expressed in the cortex of the 96th days post-hatched individual ovarian tissues where oocytes were more developed. FoxO1 mRNA level was no significant difference in the gonadal tissues of Chinese alligator at different periods. These results provide important reference for enriching the regulation mechanism of oogenesis of Chinese Alligator.
CONSTRUCTION OF A SINGLE NUCLEOTIDE POLYMORPHISM GENOTYPING SYSTEM FOR SALMONID SPECIES BASED ON MICROFLUIDIC CHIP
ZHAO Zi-Xia, XU Jian, WU Bi-Yin, CAO Ding-Chen, BAI Qing-Li, XU Peng, MA Zhuo-Jun
 Available online   , doi: 10.7541/2022.2021.118
[Abstract](442) [FullText HTML](316) [PDF 1111KB](5)
Abstract:
To develop reliable single nucleotide polymorphism (SNP) genotyping technologies suitable for large sample number and limited marker number, a microfluidic platform, namely Fluidigm 96.96 dynamic chip, was used to conduct the SNP genotyping system for identification of released individuals from stock enhancement projects. This study screened 96 high-quality polymorphic markers shared among salmonid species based on genotyping results of fish samples from 4 genera of Salmonidae using rainbow trout 57K SNP array. The microfluidic system was proved with high accuracy using Brachymystax lenok as testing populations, with a call rate of 98.63%, and the consistency with the previous Affymetrix 57K array reached 97.92%. Parentage assignments for 96 offspring individuals to their candidate parents and interfering parents were performed using CERVUS 3.0.7 software and the 96 SNP genotyping data. The results matched perfectly with the true pedigrees among complicated families, proving high accuracy in pedigree assignment application for the constructed microfluidic system. The non-exclusion probability for first parent (NE-1P) was 4.362×10–4 during a single parent assignment, while the non-exclusion probability for parent pair (NE-PP) was as low as 6.538×10–12. These results fully met the needs for molecular identification of released individuals, to determine if one captured fish was an offspring of documented broodstocks, which meant it’s hatched and released from certain stock enhancement projects. STRUCTURE analysis was also performed based on the 96 SNP genotyping data. The genetic components of 4 wild populations clearly distinguished, and the genetic composition of testing individuals were well demonstrated, suggesting that the constructed system is suitable for preliminary population genetic analysis for captured salmonid species. Thus, this study has great application potentials in fishery stock enhancement, to monitor the survival rate of released individuals, and to evaluate the supplemental effects on natural fishery resources.
ROLE OF TREMATOMUS BERNACCHII DUSP1 GENE IN CELL COLD RESISTANCE
WANG Ying, HU Ling-Hong, WANG Hua-Min, CHEN Liang-Biao
 Available online   , doi: 10.7541/2022.2021.114
[Abstract](473) [FullText HTML](270) [PDF 1359KB](6)
Abstract:
Dual specificity phosphatase 1 (dusp1) is an enzyme to inactivate MAPKs by selectively dephosphorylating critical serine/threonine residues. Dusp1 plays an important role in many environmental stress including hypoxia, oxidative stress, UV and heat shock. However, the role of dusp1 in response to cold stress has been rarely reported in fish species, especially in polar fish. To address this, we cloned a dusp1 homolog gene encoding 376 amino acids from Trematomus bernacchii. The open reading frame encoding dusp1 was amplified by PCR with homologous recombination approach cloned into pcDNA3.1 vector, referred to as pcDNA3.1-dusp1. The constructed eukaryotic expression vector pcDNA3. 1-dusp1 were transferred into the human embryonic kidney 293T (HEK293T) cells, and pcDNA3.1 (empty vector)-transfected cells were used as a negative control. The reactive oxygen species (ROS) was assayed using fluorescence probe DCFH-DA and the survival rate under low temperature stress was detected by flow cytometry, respectively. A Western-blot analysis was applied to determine the phosphorylation of P38/MAPK levels and the apoptosis effector gene caspase-3 mRNA expression levels were measured using RT-qPCR. The results showed the T. bernacchii DUSP1 protein in the HEK293T cells with the subcellular localization at nucleus. Dusp1 over-expression significantly decreased ROS content and apoptosis rate in cells under cold stress, and it also suppressed hyperphosphorylation of the pro-apoptotic gene p38 and decreased caspase-3 transcriptional activity. These results indicated that dusp1 from T. bernacchii can protect cells from the damage under cold stress, and this protection was achieved by inhibiting of the p38MAPK and caspase-3 activation. This study provide new insights of the cold temperature adaptive evolution of polar fish and lay a foundation for further functional studies of dusp1 in teleosts under cold stress.
MOLECULAR CLONING OF TRPV1 IN SPINIBARBUS SINENSIS AND THE RESPONSES TO STIMULATION OF HOT TEMPERATURE AND LPS
LIU Yao-Hui, XIE Yun-Long, ZHANG Zi-Heng, YANG Xing-Yue, LU Min-Fang, PU De-Yong, WANG Zhi-Jian, ZHANG Yao-Guang, LIU Xiao-Hong
 Available online   , doi: 10.7541/2022.2021.0177
[Abstract](367) [FullText HTML](273) [PDF 1053KB](5)
Abstract:
TRPV1 is a multifunctional ion channel in mammals, and one of its most critical functions is to sense high temperature. According to the available data, the function of TRPV1 in fish temperature perception is controversial: TRPV1 of zebrafish (Danio rerio) and rainbow trout (Oncorhynchus mykiss) can sense heat, while TRPV1 of other fish like ice-fish (Chionodraco hamatus) and Antarctic fish (Trematomus bernacchii) are reported to be insensitive to temperature changes. In order to test whether the expression of TRPV1 in freshwater fish in China is sensitive to temperature, Spinibarbus sinensis was selected as the experimental animal in the present study. TRPV1 gene was cloned by RT-PCR, and then bioinformatic analyses were carried out to predict the secondary and tertiary structure or TRPV1 protein. Tissue expression profile of this gene was also studied by RT-qPCR. Since mammalian TRPV1 can sense both the environmental temperature and the inner body inflammation, we detected the expression of TRPV1 mRNA in the diencephalon and the medulla oblongata of S. sinensis, which were treated by PBS or LPS injection and then put in water of different temperatures (25℃ and 35℃) for 1h, 6h and 24h.  In the present study, the cloned fragment of TRPV1 gene is 2023 bp in length, encoding a peptide consisting of 673 amino acid residues. The predicted TRPV1 protein of S. sinensis shares a highest similarity of 88.80% with that of common carp (Cyprinus carpio), and it contains a predicted Ankyrin repeat domain (ANK), 6 transmembrane domains, and an ion transmembrane domain (Ion_trans). Based on the online SWISS-MODEL, the functional protein of TRPV1 in S. sinensis might be a homotetramer. The TRPV1 gene of S. sinensis is widely expressed in various tissues, including all parts of brain (telencephalon, mesencephalon, diencephalon, cerebellum, medulla oblongata), and internal organs such as kidney, liver, spleen and gut. However the expression levels in the skin and muscle are very low. High temperature and LPS injection can induce the significant changes of this gene in both the diencephalon and the medulla oblongata, however there are opposite alteration patterns in these two tissues, in a time specific manner. Taking all the above-mentioned information together, it is clear that the expression of TRPV1 in S. sinensis is temperature and inflammation sensitive, with a tissue and time specific pattern.
OPTIMAL LEVEL OF FISH OIL SUPPLEMENTATION IN DIET OF JUVENILE AMERICAN EELS (ANGUILLA ROSTRATA)
QU Qian, ZHAO Pan-Yue, CAI Guo-He, ZHAI Shao-Wei
 Available online   , doi: 10.7541/2022.2021.0284
[Abstract](685) [FullText HTML](469) [PDF 924KB](10)
Abstract:
This experiment was conducted to achieve the optimal dietary fish oil supplementation level of juvenile American eels (Anguilla rostrata) by evaluating the parameters of growth performance, digestive enzymes activities, body composition, and lipid metabolism enzymes in the liver. 800 juvenile American eels with an average body weight of (8.34±0.12) g per fish were randomly divided into 4 treatment groups with 5 replicates per group and 40 fish per replicate. The basal diets were supplemented with 0 (control group), 3% (FO3 group), 6% (FO6 group), 9% (FO9 group), and 12% (FO12 group) fish oil, respectively. The trial lasted for 56 days. Dietary fish oil supplementation significantly affected the growth performance of juvenile American eels. The weight gain rate, feeding rate, and feed efficiency of juvenile American eels in FO6 group were significantly higher than those in other groups (P<0.05). Compared with the control group, intestinal lipase activities increased significantly in fish oil supplementation groups (P<0.05). The activities of protease and amylase decreased significantly by fish oil supplementation (P<0.05), and there was no significant difference in the activities of protease and amylase among different fish oil supplementation groups (P>0.05). Compared with the control group, the contents of crude lipid of the whole body increased significantly in FO6 group, FO9 group, and FO12 group (P<0.05). The content of crude protein decreased significantly only in FO12 group (P<0.05). The contents of moisture and ash were similar among all the fish oil supplementation groups (P>0.05). The activities of fatty acid synthase decreased significantly only in FO9 and FO12 groups (P<0.05), the activities of lipoprotein esterase and liver lipase increased significantly only in FO12 group (P<0.05). In conclusion, the adequate supplementation level of fish oil in diet might improve the growth performance by regulating the intestinal digestive enzymes activities, lipid content in whole fish and lipid metabolism enzymes levels or activities in the liver. The recommended level of fish oil supplementation in the diet was 6.43% to 6.78% for the highest weight gain rate and feed efficiency of juvenile American eels.
SCREENING OF TRANSCRIPTOME SSR MARKER OF MACROBRACHIUM ROSENBERGII AND ANALYSIS OF ITS CORRELATION WITH BODY WEIGHT
CHEN Xue-Feng, WANG Chun-Lin, LI Rong-Hua, GU Zhi-Min, XU Bin-Peng, CHENG Hai-Hua, PENG Fei
 Available online   , doi: 10.7541/2022.2021.121
[Abstract](459) [FullText HTML](266) [PDF 816KB](4)
Abstract:
In this study, the SSR markers of Macrobrachium rosenbergii were excavated by transcriptome sequencing, and 18592 SSR loci were obtained. 63 SSR loci were selected and 10 individuals were utilized to screen the polymorphic loci, and 31 SSR markers with polymorphism were obtained finally. Among which, 19 SSR markers were used to analyze the population genetic structure of 119 shrimps from the full-sib family group (FS) and 199 shrimps from the F3 wild group of Bangladesh (MJL), and the correlation between SSR loci and body mass traits was analyzed. The results showed that 40 alleles were detected in FS population, with an average alleles number of 2.1053, an average observed heterozygosity of 0.4525, an average expected heterozygosity of 0.3804, an average polymorphism information content of 0.3076, which was moderate genetic polymorphism; A total of 65 alleles were detected in MJL population, with an average alleles number of 3.4211, an average observed heterozygosity of 0.4105, an average expected heterozygosity of 0.4496, and an average polymorphism information content of 0.3882, which was also moderate genetic polymorphism. There was no correlation between 19 SSR loci and body weight in FS population (P>0.05), but 4 SSR loci were significantly correlated with body weight in MJL population (P<0.05). For the loci with significant differences, body weight traits of different genotypes were analyzed by multiple comparison, which showed that the mean body weight of MR28 277/285 genotype was significantly higher than that of 277/289 and 285/285 genotypes (P<0.01), and significantly higher than that of 285/289, 273/289, 270/273 and 285/293 genotypes (P<0.05); The mean body mass of genotypes 266/266 and 266/270 at MR32 was significantly higher than that of genotypes 270/270 (P<0.05) . The mean body mass of genotypes of 210/214 at MR34 was significantly higher than that of genotypes 210/210 and 214/214 (P<0.05). The mean body mass of genotypes of 174/190 at MR45 was significantly higher than that of genotypes 182/182 and 182/190 (P<0.05). The results lay a foundation for marker-assisted breeding of M. rosenbergii.
THE ROLE OF FUNCTIONAL AMINO ACIDS IN AQUATIC CRUSTACEANS
CHENG Kai-Min, ZHANG Zhen, LI Pei-Yang, ZHENG Shi-Xuan, LI Xin-Yu
 Available online   , doi: 10.7541/2022.2021.0228
[Abstract](131) [FullText HTML](82) [PDF 857KB](9)
Abstract:
Traditionally, amino acids are classified as essential amino acids and non-essential amino acids based on their ability of syntheses from metabolic intermediates. Currently, many scientists gave a new concept of functional amino acids, which are defined as those amino acids that participate in and regulate key metabolic pathways and immune systems. These amino acids not only provide blocks for protein synthesis, but also as substrates for many functional compounds. It has been reported that these amino acids can improve health, survival, growth, development, and reproduction of organisms. Crustaceans (e.g., shrimp and crabs) are the second largest aquaculture species worldwide, their feed development play a vital role in industry development. Functional amino acids hold great promise in prevention and treatment of metabolic diseases, intrauterine growth restriction, and infectious diseases (including viral infections). However, there are still some technical difficulties and challenges in the use of crystalline amino acids for research and practical production. At present, the knowledge about amino acid metabolism and functions in crustaceans is still limited, and much of them are based on studies of mammals and fish species. This article highlights current knowledge about the functions of amino acids on molt, adaptation of stress, immune, and antioxidant responses, spawning and larval development of crustaceans. It will help to advance the field of protein nutrition and guide the development of future crustacean feeds.
GENETIC STRUCTURE AND DIVERSITY OF TRICHODINA RETICULATA HIRSCHMAN & PARTSCH, 1955 POPULATIONS BASED ON SSU rDNA SEQUENCE
XU Zhi-Xiang, TANG Fa-Hui, ZHAO Yuan-Jun
 Available online   , doi: 10.7541/2022.2021.0328
[Abstract](129) [FullText HTML](81) [PDF 839KB](7)
Abstract:
Trichodinid ciliates are well known as ectoparasites of fishes, mollusks, amphibians, as well as crustaceans, sometimes they can cause serious diseases for the maricultured and freshwater animals. They are diverse and widely distributed in the world, and about 400 nominal Trichodina species have been reported in different environments. So far, the shortage of molecular data of trichodinids in GenBank has inhibited the development of those studies on their population genetic structure and population evolutionary history, which resulted in very little valuable information of the genetic diversity about this special group. So, in order to learn about more genetic knowledge about trichodinids, the present work was carried out. Based on the SSU rDNA sequences, the population genetic structure and genetic diversity of Trichodina reticulata Hirschman & Partsch, 1955 in China were explored. The research results of genetic structure indicated that totally nine haplotypes were detected from twenty samples, including four shared haplotypes (Hap1, Hap2, Hap3 and Hap7) and five peculiar haplotypes (Hap4, Hap5, Hap6, Hap8 and Hap9). Among them, the Hap3 from Carassius auratus was the largest shared haplotype. The Hap8 from Ctenopharyngodon idellus and Hap9 from Micropercops swinhonis were temporarily regarded as the peculiar haplotype in Wuhan and Tibet Autonomous Region, respectively. At the same time, the Hap1 from Carassius auratus was supposed to be the ancestral haplotype. Combining with the ML phylogenetic tree, Trichoina reticulata from Carassius auratus was probably the first differentiated group during the multi-host evolutionary progress, and maybe Trichoina reticulata parasitic on Ctenopharyngodon idellus was originated from Carassius auratus. The research results of genetic diversity indicated that all populations showed high haplotype diversity (Hd≥0.5) and low nucleotide diversity (Pi<0.005), and the Hd of Carassius auratus source was significantly higher than that of Ctenopharyngodon idellus source, but the nucleotide diversity of Carassius auratus source was significantly lower than the latter. The research results of genetic differentiation (Fst) and gene flow (Nm) revealed that the group A (from Carassius auratus) and the group B (from Ctenopharyngodon idellus) were relatively independent and reached the extremely differentiated extent because of the lesser genetic communication within the group. Combining the neutral test with the nucleotide mismatch analysis, it was concluded that the group A (from Carassius auratus) has never gone through the population expansion, whereas the population expansion has probably existed during the early history of the group B (from Ctenopharyngodon idellus).
DIVERSITY INVESTIGATION AND PHYLOGENETIC ANALYSIS OF CLADOPHORA IN CHISHUI RIVER BASIN IN WINTER
ZHAO Zhi-Juan, YANG Xiao-Xia, ZHONG Qiao-Jun, CHEN Guan-Hai, PENG Xiang-Ying, LIU Ben-Wen, ZHU Huan, LIU Guo-Xiang
 Available online   , doi: 10.7541/2022.2021.0176
[Abstract](237) [FullText HTML](160) [PDF 1142KB](8)
Abstract:
The genus Cladophora Kützing is one of the largest genera of filamentous green algae (Cladophorales, Chlorophyta) and plays an important role in the aquatic ecosystem. The geographical distribution of Cladophora is cosmopolitan, from tropical to polar water, from freshwater to marine. On the one hand, Cladophora can provide important habitats for aquatic micro-animals and plants and be used in aquaculture and business as a valuable biological resource, they also can cause algal blooms in eutrophic waters frequently. Due to phenotypic plasticity of some morphological characters, such as branching pattern, filament dimensions and attachment organs, classification and identification of Cladophora have challenged taxonomists for years. With the development of technology, molecular methods demonstrate the effectiveness of Cladophora classification and reveal cryptic species diversity in several morphospecies. Although ribosomal DNA (rDNA) sequences are widely used in the phylogeny of Cladophorales, evolutionary analyses only based on single molecular marker, such as the small-subunit (SSU) rDNA or the large-subunit (LSU) rDNA, are difficult to resolve the phylogenetic relationships of Cladophora. An approach combining multiple molecular markers and increasing sample collection will be needed to arrive at a well-supported phylogeny. Chishui River, as a large and valuable tributary on the south bank of the upper Yangtze, has the feature of diverse landscape types and rich biodiversity. However, the diversity investigation of Cladophora in this area is still absent so far. In order to assess Cladophora diversity of Chishui River basin, a total of 38 sampling sites were established along the upper, middle and lower regions of the river. The field-collected filamentous samples were preserved in 10% formalin and 95% ethanol, respectively. These materials were taken back to the laboratory for DNA extraction and polymerase chain reaction (PCR) amplification. We sequenced three molecular markers including SSU rDNA, LSU rDNA and the internal transcribed spacer (ITS, ITS1-5.8S-ITS2) sequences to determine the phylogenetic position of these algae. All DNA sequence data generated for this study were submitted and deposited in GenBank. We used two datasets in the present study: dataset 1, a concatenated alignment of SSU+LSU from 80 cladophoracean samples (Chaetomorpha as an outgroup); dataset 2, a combined dataset of SSU+ITS+LSU from 51 freshwater Cladophora samples (plus 5 brackish Cladophora as an outgroup). Sequence matrices for phylogenetic analyses were aligned using MAFFT 7.0. All datasets were analysed using Bayesian Inference (BI) and Maximum Likelihood (ML). One thousand non-parametric boot strap replicates were inferred from 10 distinct alternative runs. Posterior probability values were calculated for 1×106 generations, with sampling conducted every 1000 generations. The results showed that: (1) Cladophora diversity is higher in Chishui River Basin, where the widespread species C. glomerta and the rare species C. rigida both distributed. Cladophorales (including Cladophoraceae and Pithophyceae) samples were obtained at a total of 21 sampling sites, 19 of which existed Cladophora samples. The distribution of these algae covered the upper, middle and lower regions of Chishui River, at the same time, Cladophora species diversity showed a decreasing trend from upstream to downstream. (2) Sequence analysis showed that the current freshwater Cladophora taxa contained at least ten phylogenetic clades, six clades of which contained samples collected from this area. These six clades are as follows: clade 1, clade 2, clade 4, clade 7, clade 9 and clade 10, respectively. (3) The phylogeny based on SSU+ITS+LSU divides freshwater Cladophora into two major groups (group Ⅰ and group Ⅱ). Clade 1, clade 2 and clade 4 formed a group I, as did clade 5, clade 6, clade 7, clade 9 and clade 10 (group Ⅱ). Compared with the phylogeny based on SSU+LSU, the support value of each clade (except clade 4) of the phylogeny based on SSU+ITS+LSU is higher (BI=1.00; BP>95), which further supported that using multiple molecular markers, and particularly, adding the rapidly evolving molecular marker ITS, are useful and effective in analyzing the taxonomy and phylogeny of Cladophora. Due to the lack of typical morphological characteristics, the species names of most clades are not determined for the time being, except clade 1 (C. glomerta) and clade 9 (C. rigida). Notably, C. rigida, differed from other freshwater Cladophora, had some unique characteristics, such as being unbranched, with a large cell diameter (180—429 µm) and smaller length: diameter (L﹕D) ratio (0.5—3.3) than other species. This species is now only reported in Chishui River basin and Hubei Province of China.  In conclusion, our results not only revealed the Cladophora species diversity of Chishui River based on multiple molecular markers, but also enriched the classification of freshwater filamentous green algae of China, as well as provided basic data for the ecological environment protection of Chishui River. Next, continued investigation and exploration of other regions of China that have not yet been covered will be needed, which likely yield additional records and findings.
GROWTH, MORTAILY AND OPTIMUM CATCHABLE SIZE OF COILIA MYSTUS IN OUJIANG RIVER ESTUARY
LI Xia-Fang, JIANG Ri-Jin, RUI Yin, WANG Ya-Li, SUN Hao-Qi, YIN Rui, LI Kai, HU Zhong-Jian
 Available online   , doi: 10.7541/2022.2021.03211
[Abstract](44) [FullText HTML](19) [PDF 1968KB](2)
Abstract:
Coilia mystus is one of the main fishing species in Oujiang River Estuary and it has high economic value. Based on the fishery resources monitoring data collected in Oujiang River Estuary in Wenzhou from 2015 and 2018 to 2020, the present study analyzed the biological characteristics (n=1359) of C. mystus, including body length, total length and body weight. The population growth and death parameters were estimated by body length frequency distribution. The results indicated that the average body length of C. mystus in Oujiang River Estuary was (15.1±3.0) cm, with the group of predominant body length 14.0—18.0 cm, accounting for 54.3%; the average body weight was (16.8±10.1) g, with the group of predominant body weight 5.0—25.0 g, accounting for 72.0%. The distribution of body length and body weight differed significantly between males and females (P<0.05). The average body length and body weight of females were greater than those of males, but the body length-weight relationship was not significantly different (P>0.05). The relationship between body length and weight was expressed as: W=0.0035×L3.0783 (R2=0.9509). The power exponent b was 3.0783, which is consistent with uniform growth characteristics. The growth parameters of von Bertalanffy formula estimated by ELEFAN were L=26.60 cm, k=0.47, and t0=−0.55 a. The total mortality coefficient (Z) was calculated using a length-converted catch curve as 2.30. The natural mortality coefficient (M) estimated referring to the Pauly empirical formula, was 1.00. Therefore, fishing mortality coefficient (F) was 1.30, and the exploitation rate (E) equaled 0.567, demonstrating that the stock had been over-exploited. The catch curve also determined that the current first capture L50 was 14.50 cm, corresponding to an age of 1.13, which was lower than both the critical age (1.36 a) and the turning point age (1.85 a). This indicated that the main fishing objectives were juveniles and included the recruitment group of C. mystus. Under the current resource status in Oujiang River Estuary, great amount of effort should be made to reduce fishing mortality and protect C. mystus habitat. It was suggested that the control of catchable length should be the main management strategy for the trend of miniaturization and low age of C. mystus resources. According to the Beverton-Holt dynamic model, the minimum capture size for C. mystus should be 15.50 cm (age 1.31 years).
COMMUNITY STRUCTURE OF ICHTHYOPLANKTON IN THE SECTIONS OF THE CENTRAL AND NORTH YELLOW SEA
YANG Yan-Yan, LI Huan-Jun
 Available online   , doi: 10.7541/2022.2021.0199
[Abstract](215) [FullText HTML](164) [PDF 2218KB](5)
Abstract:
In order to understand the community structure characteristics of ichthyoplankton at different sections in the coastal waters of the central and north Yellow Sea, three surveys were conducted in May, June and July 2017, the species composition, quantity distribution, and diversity of ichthyoplankton in this sea area were analyzed. The results showed that a total of 5493 fish eggs and 43 larvae were collected during the three surveys by horizontal trawl with plankton net. The obtained ichthyoplankton belonged to 20 species, 20 genera, 16 families, and 7 orders, the main group was perciformes. The number of ichthyoplankton collected varied in different months and there were differences between sections. The collected fish eggs and larvae were mainly composed of warm-temperate species and warm-water species, with only a few cold-temperate species, and the number of species of each ecological type varied with different sections. The T1section was dominated by warm-temperate species, and the T2section had slightly more warm-water species than warm-temperate species, Warm-temperate species showed a decreasing trend from north to south in the surveyed sea area, while warm-water species showed an increasing trend. The composition of dominant and important species in ichthyoplankton changed in different months and sections, Engraulis japonicus is the dominant species of fish eggs in the surveyed waters of the May voyage and sections T1 and T2, and no dominant species appeared in larvae. The average trophic levels of ichthyoplankton collected during the three investigations were 3.58, 3.51 and 3.41, respectively, showing a gradual decrease. The richness index (D), diversity index (H') and evenness index (J') of ichthyoplankton were the highest in May and the lowest in June. In addition, there were some differences in the community diversity index of ichthyoplankton between the two sections, which were closely related to the depth.
ACUTE TOXIC EFFECT OF ARSENATES ON CHLORELLA VULGARIS
LI Li-Jie, PENG Meng, YANG Fan, YU Jia-Ni, JIN Wei, TAN Feng-Xia, CHAI Yi
 Available online   , doi: 10.7541/2022.2021.0306
[Abstract](189) [FullText HTML](100) [PDF 1835KB](2)
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In order to explore the toxicity and mechanism of heavy metals arsenic in aquatic ecosystem, Chlorella vulgaris, a typical phytoplankton indicator species, was used as the experimental object. Different concentrations of As5+ (0, 0.1, 0.5, 1, 1.5 and 2.0 mg/L) were used to study the effects of As5+ on the density, chlorophyll a content, cell membrane permeability, reactive oxygen species (ROS) content, malondialdehyde (MDA) content, total antioxidant capacity (T-AOC), total protein (TP) content, total superoxide dismutase (T-SOD) activity, ATP activity and ultrastructure of C. vulgaris. The results showed that all concentrations of As5+ could inhibit the growth of C. vulgaris within 96h, and the inhibition rate of 2 mg/L treatment group was the highest (52.9%). The IC50 of 96h was 1.94 mg/L. The Chl.a content of algae cells was also inhibited in all treatment groups, among which, the treatment group of 2 mg/L was the lowest which was only 31.4% of the control group. When the concentration of As5+ was 1.5 and 2 mg/L, the content of TP and ATP decreased significantly. The content of MDA was 2 times and 2.6 times higher than that of the control group, and the cell membrane permeability was 1.7 times and 2 times higher than that of the control group. The content of ROS in 2 mg/L treated group increased significantly, which was 4.0 times of that in control group. The activities of T-AOC and T-SOD in algal cells stimulated by oxidative stress were 2.8 times and 1.6 times higher than those in control group. The results of transmission electron microscope showed that the group treated with 2 mg/L As5+ could destroy the internal structure of algae cells, resulting in cytoplasmic vacuolation, lamellar structure fracture of thylakoids and chloroplast structure disorder. The results showed that high concentration of As5+ was destructive to the structure and function of cell membrane and main organelles of C. vulgaris. At the same time, C. vulgaris produced oxidative stress in response to pollutant stress. The concentration of As5+ below 1.5 mg/L had no significant effect on physiological and biochemical indexes of C. vulgaris. This study reveals the physiological and biochemical reactions of the typical indicator species C. vulgaris under As5+ environmental stress, and provides basic data for the remediation of heavy metal pollution in the water environment and the formulation of relevant environmental standards, thereby realizing the sustainable development and utilization of the aquatic ecosystem.
ACUTE TOXIC EFFECTS OF POLYCYCLIC AROMATIC HYDROCARBONS (NAPHTHALENE) COMBINED WITH POLYSTYRENE MICROSPHERES ON CHLORELLA VULGARIS
ZHANG Xiao-Qun, LI Li-Jie, PENG Meng, CHAI Yi, ZHANG Heng, XI You-Jun, ZHANG Hui, LUO Ming-Zhong
 Available online   , doi: 10.7541/2022.2021.0187
[Abstract](140) [FullText HTML](71) [PDF 1492KB](3)
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Naphthalene (Nap), as one of the main types of polycyclic aromatic hydrocarbons pollutants, widely exists in China's aquatic environment. Micro-plastics (MPs) can easily become an excellent carrier of persistent organic pollutants and form compound toxic effects on aquatic organisms. However, there are few studies on the acute toxic effects of these two pollutants combined. In order to understand the joint toxic effects of polycyclic aromatic hydrocarbons (PAHs) and microplastics, the algae density, chlorophyll a (Chl.a), Soluble protein (TP), malondialdehyde content (MDA), catalase activity (CAT) and superoxide dismutase activity (T-SOD) were measured for studying the acute toxic effects of Nap combined with MPS on Chlorella vulgaris, which is an indicator of ecological environment. The results showed that C. vulgaris could be inhibited by both nap and MPs. The highest inhibition rates were 68.26% and 78.36% respectively under single stress, and 52.05% under combined stress. The inhibitory effect on Chl.a was consistent with the algae density. Under single and combined stress, TP content decreased and MDA content increased significantly. The activities of CAT and T-SOD decreased significantly under Nap stress. The activities of CAT and T-SOD increased first and then decreased under MPs and combined stress. The final activities of CAT and T-SOD of the combined stress experimental group were significantly higher than the control group. The EC50-96h of Nap and MPs on C. vulgaris were 81.35 and 383.3 mg/L, respectively. The toxicity of Nap was stronger than MPs. The EC50-96h of Nap combined with MPs (toxic unit ratio 1﹕1) on C. vulgaris was 1.320 TU. The results showed that joint toxic effect of the two pollutants hasn’t been more significant than single pollutant, and the joint toxic effect on C. vulgaris was antagonistic. MPs played an adsorption polymerization role on Nap, which greatly reduced the probability of contact between both pollutants and C. vulgaris. The existence of MPs showed a certain protective effect in the short term.
ADJUSTMENT MECHANISM OF LIGHT INTENSITY AND TEMPERATURE ON PHOTOSYNTHESIS OF ULVA FLEXUOSA
ZHA Min-Jun, ZHOU Ling-Jie, CAI Chun-Er, DONG Ao, HE Pei-Min
 Available online   , doi: 10.7541/2022.2021.0209
[Abstract](231) [FullText HTML](150) [PDF 1054KB](2)
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The excessive growth of green algae Ulva, including Ulva flexuosa, causes the outbreak of green tide in coastal area. These green algae with great fertility grow in the intertidal zone and are often affected by light, temperature and other ecological factors. At the same time, they have high tolerance to various abiotic factors. The growth and reproduction of Ulva flexuosa require the accumulation of organic matter, and the efficient accumulation of organic matter is the internal cause of its eruption. In this study, the response of photosynthetic system of U. flexuosa to the changes of light and temperature was investigated at molecular level. The adjustment mechanism of Ulva flexuosa was explained on these aspects: electron transfer of the initial reaction, photophosphorylation and finally carbon fixation. The whole experiment involved two factors, a total of nine experimental samples Nine sets of transcriptome data were obtained and the differential expression genes were mainly analyzed. First, the chlorophyll synthesis related genes down-regulated under high temperature and light [30℃, 400 μmol/(m2·s)]. Second, the carotenoid synthesis related genes down-regulated under low temperature and high light [4℃, 400 μmol/(m2·s)]. Third, electron transport chain, CF1F0-ATP synthase and other genes had weak ability in resistance to temperature and light, wherein the expression levels of related genes in all experimental groups showed a downward trend. Fourth, under high temperature environment, the influence of high light on U. flexuosa was more significantly (Q1). In low temperature environment, the effect of low light was more significantly (Q9). Finally, the expression of light harvesting complex gene up-regulated. The results showed that U. flexuosa had a certain tolerance to temperature and light changes, wherein temperature had a great influence on the photosynthetic gene of U. flexuosa, especially at high temperature and strong light. In this experiment, two ecological factors with the most significant effects on photosynthesis were integrated to conduct the experiment, which is helpful to fully understand the response mechanism of the photosynthesis of U. flexuosa, to accurately understand U. flexuosa outbreak in the light of the different sea temperature under the condition of the photosynthesis system coping mechanisms, for further analysis in the process of the green tide outbreaks physiological activity to support the main species.
EFFECT OF MICROCYSTIS AERUGINOSA ON COMPETITION BETWEEN BRACHIONUS CALYCIFLORUS AND DAPHNIA MAGNA
LI Hai-Lu, YAO Hui, HE Yu-Han, GENG Hong, XU Jun, HE Liang, ZHANG Huan
 Available online   , doi: 10.7541/2022.2021.0181
[Abstract](358) [FullText HTML](270) [PDF 817KB](2)
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To investigate the determinants of zooplankton community structure in aquatic ecosystems under cyanobacterial bloom and the effects of Microcystis aeruginosa density on zooplankton competition relationship, the interspecific competition between Brachionus calyciflorus and Daphnia magna was evaluated by conducting experiments at three concentration gradients of M. aeruginosa. Our results showed that the concentrations of M. aeruginosa had significant effects on the growth and competition between B. calyciflorus and D. magna, it did not change the dominance of D. magna. The results also showed that a low level of addition of M. aeruginosa (5×104 cells/mL) promoted D. magna population growth (P<0.01) and increased dominance in D. magna in the mixing culture. The growth of both D. magna and B. calyciflorus was significantly (P<0.01) affected by the medium concentration (1×105 cells/mL) level of M. aeruginosa. However, population growth of both species was suppressed (P<0.01) by the high concentration of M. aeruginosa (5×105 cells/mL) and only D. magna population survived in the co-exist system. Thus, in the absence of other external influences, our results suggested that D. magna dominated under various concentration levels of M. aeruginosa co-exist with B. calyciflorus, indicating that the concentration of M. aeruginosa may not be the main factor affecting the competitive position of D. magna and B. calyciflorus. The study on zooplankton community dynamics under different concentrations of M. aeruginosa will contribute our understanding the ecological consequences of cyanobacterial bloom.
BIOACCUMULATION OF CHROMIUM IN DIFFERENT BRAIN REGIONS OF PLECTROPOMUS LEOPARDUS DURING DIETARY EXPOSURE
LI Yu, GUO Zhi-Qiang, HAN Dong, WEI Lu
 Available online   , doi: 10.7541/2022.2021.110
[Abstract](44) [FullText HTML](22) [PDF 1649KB](4)
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Excessive intake of chromium (Cr) cause neurotoxicity to fish, though, the information on the bioaccumulation of Cr in fish brain under dietary exposure is still lacking. To evaluate the effects of dietary Cr exposure on the brain of coral trout (Plectropomus leopardus), fish were fed with a gradient levels of organic Cr (as chromium picolinate (CrPic)) and inorganic Cr (CrCl3 (Cr(Ⅲ), K2Cr2O7 (Cr(Ⅵ))) at 0, 10, 50, and 250 mg/kg for 66 days. Samples were collected to measure Cr accumulation in hypothalamus, forebrain, midbrain, hindbrain regions of brain at 7, 12, 18, 40 and 66 days using atomic absorption spectrometry (AAS). The results showed that Cr bioaccumulation in the four brain tissues increased significantly from day 7 to day 40, but decreased significantly after day 66 under both organic and inorganic Cr treatment. However, the total bioaccumulated Cr in four regions of brain did not increase with the increasing dietary Cr level and exposure time, indicating a dose- and time-independent pattern. The accumulated amount of Cr in the brain after dietary exposure followed the order CrPic > Cr(Ⅵ) > Cr(Ⅲ). These findings demonstrated that the dietary organic Cr had significantly higher bioaccumulation than inorganic Cr, providing a theoretical knowledge for further evaluation of the toxic effect of form-specific Cr in food on central nervous system and evaluation of its toxicological mechanism.
ANALYSIS OF INTESTINAL MICROBIOTA STRUCTURE IN DIFFERENT HEALTH CONDITIONS OF PAA SPINOSA
JIANG Yun, HU Ya-Zhou, XIANG Jian-Guo, PAN Wang-Chen
 Available online   , doi: 10.7541/2022.2021.094
[Abstract](635) [FullText HTML](382) [PDF 2436KB](8)
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Paa spinosa (David) is an important amphibious economic animal distributed in the rolling mountains in the south of China. In the process of artificial farming, diseases have become an important factor on its yield reduction. Intestinal microbiota is considered to be involved in organism’s immune response. However, it is unclear whether the composition and diversity of intestinal microbiota are related to the health condition of P. spinosa. To explore the structure of intestinal flora of P. spinosa under different health conditions, 13 healthy (Health), 19 wonky-head disease (WHD) and 18 blue-eye disease (BED) P. spinosa were sampled. Total intestinal DNA were extracted following manufacture’s procedures, and V4-V5 region of 16S rRNA were amplified and sequenced using Illumina Miseq sequencing platform. A total of 3984 operational taxonomic units (OTUs) were obtained, belonging to 35 phyla, 80 classes, 155 orders, 283 families, and 501 genera. The results showed that the microbiota abundance of the healthy group was significantly higher than that of the sick groups with no significant difference between group WHD and BED (Chao1 index: Health: 1232.92; WHD: 975.57; BED: 1048.76), and the microbial community diversity exhibited no significant difference between healthy and sick groups (Shannon index: 5.27; 5.20; 5.41). The dominant bacterial phyla included Firmicutes (52.08%; 49.57%; 26.48%), Bacteroidetes (30.55%; 21.93%; 43.02%) and Proteobacteria (15.55%; 10.42%; 5.25%). As a consequence, Deferribacteres increased significantly in sick groups (0.18%; 14.05%; 22.16%). The dominant bacterial genera included Bacteroides (29.70%; 16.09%; 21.76%), an unclassified Erysipelotrichaceae (21.38%; 6.92%; 4.94%), an unclassified Lachnospiraceae (12.23%; 15.98%; 4.43%), Citrobacter (10.31%; 6.19%; 1.10%) and Eubacterium (9.09%; 4.88%; 0.54%). Mucispirillum (0.17; 13.89%; 21.94%) of Deferribacteres increased significantly in sick groups. The analysis of Beta diversity showed that samples from healthy and sick groups were clustered separately. These results reveal that the structure and diversity of intestinal microbiota were related to the health conditions of P. spinosa. This study provides reliable data and theoretical guidance for the subsequent improvement of the production and quality of P. spinosa, and the selection of intestinal probiotics and the study of probiotic preparations.
CHARACTERISTICS OF MICROPLASTICS IN DIFFERENT CULTURE STAGES OF TYPICAL CRAB PONDS
XIE Shen-Hao, XIONG Xiong, HU Hong-Juan, WU Chen-Xi
 Available online   , doi: 10.7541/2022.2021.0191
[Abstract](209) [FullText HTML](147) [PDF 1191KB](8)
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Microplastics in freshwater has received wide concern recently. Microplastics in natural waters have been the subject of extensive research. However, research on microplastics in the aquaculture water is still limited. China has the largest aquaculture industry in the world. Large area of aquaculture ponds not only provide abundant aquatic products but are also considered to be an important source of pollutants. Ropes, nets, fences, boats, and packaging, among other plastic goods, are widely used in the fishery and aquaculture industries, and they are a potential source of microplastics in the aquaculture environment. Microplastics tend to be mistaken by organisms and possibly cause damage. Research has shown that the abundances of microplastics of several aquaculture areas in China are relatively high. From the previous research, crab ponds tended to have relatively high abundance of microplastics. The goal of this study is to learn about the features of microplastics in crab ponds during various culture periods in order to gain a better understanding of how they vary throughout the culture process. In this study, the abundance and characteristics of microplastics in the water of crab ponds at different culture stages were studied. The samples were collected from 5 crab ponds around Honghu Lake in May, July, and October 2020. Surface water was collected using a stainless-steel bucket and filtered with a 500-mesh stainless steel mesh. The microplastics were filtered on a membrane for analysis. The suspected plastic particles on the membrane were first observed in a stereo microscope to record their number, shape, size and color. Several particles were randomly selected to be tested by a Raman spectrometer to identify their polymer types. The results showed that the abundance of microplastics in crab ponds is at the level of 200—1640 items/m3, which is at a medium level compared with other water bodies. The abundance of microplastics of May, July and October were ranged from 200—880 items/m3 (average 532 items/m3), 280—880 items/m3 (average 528 items/m3), and 280—1640 items/m3 (average 976 items/m3), respectively. No significant difference in the abundance of microplastics in the three samples from different month was shown by the results of the one-way analysis of variance (P=0.175), but according to the average and maximum values, the abundance of microplastics was relatively low during the early and middle stage of cultivation but high at the end of cultivation. The water diversion and drainage processes, the use and aging of plastic products, and the changes in the water environment during the culture process of crab might all have impacts on the change in the abundance of microplastics in the crab ponds. The culture process not only affected the abundance of microplastics, but also other features of microplastics in crab ponds. The shape, size, color and polymer type of microplastics all changed in different cultivation periods. The particle size composition of the microplastics with 0.5 mm as the boundary was similar and evenly distributed in different month. The shape of microplastics were classified as fiber, sheep and fragment with fibrous microplastics being most abundant. Fibrous microplastics accounted for a relatively low proportion in the early stage of cultivation, and a relatively high proportion in the middle and end stages. The proportion of transparent microplastics continued to increase while the proportion of colored microplastics dropped significantly during the process of aquaculture. Seven different types of polymers including polypropylene (PP), polyethylene terephthalate (PET), polyethylene (PE), polyvinyl chloride (PVC), polystyrene (PS), polyamide (nylon, PA) and polymethyl methacrylate (acrylic, PMMA) were detected in the samples with PP, PET, and PE being most abundant. The polymer types were more various in the end of cultivation compared with the early and middle stage. The higher abundance of microplastics at the end of aquaculture suggested that attention might need to be paid to the impact of aquaculture tail water discharge on the occurrence of microplastics in natural water bodies. At the same time, the influence of the characteristics of microplastics in different culture periods on the ingestion of microplastics and ecological risks of cultured organisms also needs to be further explored in follow-up studies.
PROKARYOTIC EXPRESSION, PURIFICATION AND POLYCLONAL ANTIBODY PREPARATION OF CHLAMYDOMONAS REINHARDTII BBS8
LI Wen-Juan, JIA Hang, XUE Bin, FAN Zhen-Chuan
 Available online   , doi: 10.7541/2022.2021.0146
[Abstract](229) [FullText HTML](151) [PDF 1061KB](3)
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The purpose of this study is to prepare a highly specific rabbit polyclonal antibody against Chlamydomonas reinhardtii bbs8, and lay a material foundation for the follow-up study of BBS8 protein function. To achieve this, N-terminal 6×His-tagged BBS8 fusion protein (6×His::BBS8) was expressed in E. coli and purified with Ni-column. New Zealand white rabbits were immunized three times with 6×His::BBS8. Antiserum were used for ELISA at 1:102400 dilution. The IgG subtypes were enriched with protein A resin, which were affinity-purified with the purified E. coli-expressed N-terminal MBP-tagged BBS8 (MBP::BBS8) protein. The anti-BBS8 were validated using whole cell extracts of C. reinhardtii CC-125 and BBS8-null mutant bbs8 strains with high specificity, thus it is suitable for investigating the function of BBS8.
CLONING, BIOINFORMATICS AND INDUCED EXPRESSION ANALYSIS OF DXS GENE IN PHAEODACTYLUM TRICORNUTUM
SHU Ming-Yu, GONG Yi-Fu, LÜ Xin-Meng, JIA Zhe-Ming, WANG Bo, LIU Xuan-Xuan, WANG He-Yu
 Available online   , doi: 10.7541/2022.2021.0383
[Abstract](493) [FullText HTML](377) [PDF 1208KB](7)
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1-deoxy-D-xylulose-5-phosphate synthase(DXS)is the first rate limiting enzyme in carotenoids MEP biosynthesis pathway of algae. In this study, the full-length cDNA sequence of dxs was gained by Next-generation sequencing technology and bioinformatics analysis was carried out to investigate the effects of six kinds of exogenous factors on the dxs gene expression of Phaeodactylum tricornutum. The result showed that the dxs cDNA was 2476 bp long, contained an open reading frame (ORF) of 2193 bp with highly conservative ThDP-binding and transketolase domain, encoding 730 amino acids. With 79.31 kD in relative molecular mass and 6.65 in the theoretical isoelectric point, The DXS protein that contains signal peptide, transmembrane domain, coiled helix and TM-helix, is a hydrophilic stable protein. Phylogenetic tree analysis indicated that the DXS protein was divided into two clades, plant and algae. Furthermore, DXS protein of plant were classified as DXS1and DXS2 while there were three clusters in algae, namely Bacillariophyta, Rhodophyta and Chlorophyta, with Phaeodactylum tricornutum belonging to Bacillariophyta. The results of induced expression showed that dxs gene of Phaeodactylum tricornutum was regulated by six exogenous factors, including MeJA, AA, ACS, PIF, DCMU and light quality. The expression of dxs gene was the highest under the treatment of 100 μmol/L MeJA, 62.5 mg/L AA, 1.6 mg/L ACS, 1.00 μg/L PIF, 0.2 mg/L DCMU and purple light, respectively. With the induction of MeJA and light quality, the expression changes of dxs and fucoxanthin content in Phaeodactylum tricornutum showed a high consistency, which indicated that dxs is one of the key genes that promotes the fucoxanthin enrichment in Phaeodactylum tricornutum. High quality genetic resources for the future use of metabolic engineering is provided in this study to improve the content of fucoxanthin in cells of algae, and also a theoretical basis for deeply exploring the molecular regulatory principle for the biosynthesis of fucoxanthin in Phaeodactylum tricornutum.
PURIFICATION OF N-ACETYL-β-D-GLUCOSAMINIDASE FROM INTESTINE OF ANGUILLA JAPONICA AND ITS ENZYMATIC CHARACTERISTICS
LIN Jian-Cheng, HU Jian-Hui, WU Qin-Duan
 Available online   , doi: 10.7541/2022.2021.0169
[Abstract](434) [FullText HTML](264) [PDF 1164KB](3)
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In order to investigate the purification and its enzymatic characteristics of N-acetyl-β-D-glucosaminidase (EC3.2.1.52, NAGase) from intestinal tract of Japanese eel, Anguilla japonica, NAGase was purified by extraction with ammonium sulfate fractionation, then chromatographed on Sephadex G-100 followed by DEAE-cellulose (DE-32) columns. The purified enzyme was determined to be homogeneous by polyacrylamide gel electrophoresis (PAGE) and SDS-PAGE. The kinetic parameters of NAGase for the hydrolysis of pNP-β-D-GlcNAc (enzyme substrate) and enzymatic characteristics were also determined. The specific activity of the purified enzyme was 2517.40 U/mg. The molecular weight of enzyme was 69.98 kDa. The optimum pH and optimum temperature of the enzyme were 6.0 and 60℃, respectively. The Km value was 0.336 mmol/L and the Vmax value was 7.634 µmol/(L·min), respectively. The enzyme was stable with pH of 4.8 to 7.2 and temperature of 4—60℃. The enzyme lost its activity rapidly when temperature was >65℃. The effects of metal ions on the enzyme were also studied. Mg2+, Ca2+, Mn2+, Cu2+ and Fe3+ showed different degrees of activation effects on the NAGase. Na+, Li+ and Ba2+ had no influence on enzyme activity. Zn2+, Fe2+, Pb2+ and Hg2+ showed various degrees of inhibitory effects on the NAGase. Hg2+ inhibited the enzyme the most, and the enzyme activity decreased by 83.69% when its concentration reached 1.0 µmol/L. The essential groups of the NAGase were investigated using chemical modification method. The results demonstrated that essential groups of NAGase included lysine's ԑ-amidogen group, cysteine's sulfhydryl group, histidine's imidazolyl group, serine's hydroxyl group and tryptophan's indole group, while guanidyl of arginine was not an essential group of enzyme. Disulfide bond was essential for the catalytic activity of the enzyme. In conclusion, the purification scheme of NAGase from intestine of Anguilla japonica was effective and feasible. The activity of enzyme was affected easily by acidity-alkalinity, temperature and metal ions. The enzyme had similar essential groups to the NAGase from other animal sources.
PRODUCTION AND APPLICATION OF MONOCLONAL ANTIBODIES AGAINST VP7 CAPSID PROTEIN OF GRASS CARP REOVIRUS
JI Ning, WEI Wei, GUO Jia-Hong, WANG Jun-Ya, LI Yao-Guo, XIAO Tiao-Yi, ZOU Jun
 Available online   , doi: 10.7541/2022.2021.0227
[Abstract](122) [FullText HTML](69) [PDF 2276KB](3)
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Grass carp reovirus (GCRV) poses great threat to the grass carp aquaculture industry and causes huge economic losses. Grass carp reovirus (GCRV) is the pathogen of this disease. GCRV consists of three types, type Ⅰ, Ⅱ and Ⅲ. GCRV-873 is the most well studied stain of type I GCRV, with its complete genome sequenced in the 1980s. It has a segmented double stranded RNA genome with 11 fragments, encoding 7 structural proteins. VP5 and VP7 proteins form virus capsid. In this study, the capsid protein VP7 (GenBank: AF403396) of GCRV-873 strain was expressed in the E. coli cells. The cDNA fragment encoding the extracellular fragment of VP7 was cloned into pRSET-A vector and the resultant plasmid transformed into the E. coli BL21 (DE3) cells for prokaryotic expression. The recombinant protein was purified by size exclusion chromatography and used to immunize BALB/c mice for generation of monoclonal antibodies. It has been shown that the VP7 protein was highly expressed as inclusion bodies and the size was confirmed to be approximately 40 kD by SDS-PAGE analysis. After immunization, five IgG positive hybridoma cell lines were obtained, among which three were IgG1 subtype and two were IgG2a subtype. Western blotting and direct immunofluorescence analysis showed that the antibody could specifically recognize GCRV-I in the infected CIK cells, and the titer was high, reaching an affinity constant of 4.04×109. The present study provides a practical approach for the development of diagnostic tools for detecting type I GCRV virus and in-depth investigation on the GCRV infection mechanism.
PROTEIN EXPRESSION OF RAB5 AND RAB6 IN PROCAMBARUS CLARKII AND ANTIBODY PREPARATION ON PHAGOCYTIC ACTIVITY OF HAEMOCYTES
ZHU Lei, ZHAO Tong, WANG Xin-Ru, JIANG Xin-Yu, HOU Li-Bo, KONG Xiang-Hui
 Available online   , doi: 10.7541/2022.2021.0210
[Abstract](189) [FullText HTML](139) [PDF 1027KB](5)
Abstract:
Rab5 and Rab6 are important regulatory factors in regulating trafficking organelles, especially in phagosome formation. In our previous study, the Rab5 and Rab6 gene were cloned from Procambarus clarkii (PcRab5 and PcRab6), and they were proved involving in the immune response to pathogenic infection. In this study, homologous recombination technology was applied to produce prokaryotic expression protein of PcRab5 and PcRab6, then the expressed protein was purified and used to immunize rabbits for the preparation of polyclonal antibodies. The expression proteins and polyclonal antibodies of PcRab5 and PcRab6 were injected into Procambarus clarkii to study the effects on phagocytosis activity of haemocytes. The results showed that the constructed prokaryotic expression vector pET-B2m-Rab5 and pET-B2m-TGF-Rab6 were successfully induced to expression, and the molecular weight of rPcRab5 and rPcRab6 were both 67 kDa. The anti-PcRab5 and anti-PcRab6 antiserum with potency of 1:2048K and 512K were obtained from the immunized rabbits, respectively. In addition, the antibodies could specifically recognize the prokaryotically expressed protein, respectively. After the PcRab5 and PcRab6 protein were injected into healthy Procambarus clarkii, the phagocytosis rate of haemocytes increased to 38% and 30%, respectively (P<0.01). On the other hand, both the proportion of phagocytes in haemocytes decreased (P<0.05) after the purified Rab5 and Rab6 polyclonal antibodies which were injected into Procambarus clarkii, respectively. The above results indicated that the PcRab5 and PcRab6 prokaryotic proteins and polyclonal antibodies were prepared in this study, and it was proved that Rab5 and Rab6 were involved in the phagocytosis of haemocytes in Procambarus clarkii. The results of this study could lay a foundation for the further study of the molecular functions of Rab5 and Rab6 in Procambarus clarkii, and also help to understand the role of Rab5 and Rab6 during the phagocytosis of haemocytes in crustaceans.
ANDROGEN RECEPTOR DEFICIENCY CAUSES OVARIAN DYSGENESIS VIA REDUCING THE HEPATIC VITELLOGENIN PRODUCTION IN ZEBRAFISH
LI Tian-hui, ZHAI Gang, HE Jiang-yan, LOU Qi-yong, JIN Xia, DU Zhen-yu, XIAO Wu-han, YIN Zhan
 Available online   , doi: 10.7541/2023.2022.0207
[Abstract](186) [FullText HTML](159) [PDF 1260KB](5)
Abstract:
It has been observed the ovarian dysgenesis exhibited in androgen receptor (ar) knockout female zebrafish, with significant decreased gonadosomatic index (GSI) and expression levels of lhcgr and foxl2, reflecting reduced proportion of mature oocytes. However, the mechanism relating to the androgen receptor in ovarian development is still unknown. The mechanism of ar-deficiency for the ovarian dysgenesis in female zebrafish has been further explored in this study. The significantly reduced levels of vitellogenin (Vtg) production and estrogen receptor expressions have been seen in the liver of ar–/– females. The ovarian mass, lipid content and carotenoid content decreased significantly, indicates a reduced supply of nutrients such as lipid and carotenoids transferred via Vtg to the ovary in ar–/– females. Besides, the correlation between the significantly downregulated transcriptional expression levels of several key genes related with the ovarian development and the stagnation of ovarian development has been observed in the ar–/– females. Our results demonstrate that the ar-deficiency downregulates the estrogen receptors in the liver, impairs the Vtg transportation and yolk formation in the ovaries, and ultimately disrupts ovarian development. This study shed light on the connection between the androgen signaling and hepatic Vtg production in vivo.
GEOMETRIC MORPHOMETRIC ANALYSIS OF SILVER CARP LARVAE AFTER EXPOSURE TO TDCIPP
YANG Hao, PU Yan, GAO Lei, DUAN Xin-Bin, LIU Shao-Ping, CHEN Da-Qing, LI Yun
 Available online   , doi: 10.7541/2023.2022.0162
[Abstract](103) [FullText HTML](49) [PDF 1493KB](3)
Abstract:
Tris(1,3-dichloro-2-propyl) phosphate (TDCIPP), widely used as a kind of organophosphorus flame retardant, has been detected in the Yangtze River water environments. Many toxicological assessments have shown that TDCIPP could change morphology of fish. Silver carp (Hypophthalmichthys molitrix) lives in the Yangtze River for its entire life story. However, the effects of TDCIPP on silver carp is unclear. In order to clarify the main morphological characters of growth inhibition of silver carp larvae caused by TDCIPP, the present study analyzed the morphological traits between four environmentally relevant concentrations (0.05, 0.5, 5 and 50 μg/L) and the control group by geometric morphometric analysis. After the image information of larvae simple was obtained, the body length and body weight were measured. Then, digitization of landmarks was carried out with the TPS series software. Finally, principal component analysis (PCA), canonical variates analysis (CVA) and results visualization were carried out with Morpho J software. The body length and body weight of silver carp larvae decreased significantly under exposure to 0.5、5 and 50 μg/L of TDCIPP compared with the control group, but no effects were observed in 0.05 μg/L. This indicated that environmentally relevant concentrations of TDCIPP induced growth inhibition in silver carp larvae. The results of PCA and CVA indicated that the first principal component (PC1) and the second principal component (PC2) together accounted for 62.15% of the overall variables (47.36% and 14.51%, respectively). The first canonical variates (CV1) and the second canonical variates (CV2) together accounted for 79.48% (54.55% and 24.93%, respectively), which satisfied the requirement of morphological analysis of silver carp larvae. The results of grid profile analysis indicated that the average morphology of silver carp larvae in different concentrations was significantly different with the control group(P < 0.05), which identified by the growth retardation of the head, longitudinal axis of body and tail. As a conclusion, TDCIPP could induce the growth retardation of head, longitudinal axis of body and tail in silver carp larvae. Therefore, attentions should be paid to the environmental concentrations of TDCIPP in the Yangtze River Basin, and the ecological risk of TDCIPP to the replenishment of silver carp population resources should be assessed.
REMOVAL OF PBDES FROM SIMULATED TAIL WATER OF SEWAGE TREATMENT PLANT BY DEEP INTEGRATED VERTICAL FLOW CONSTRUCTED WETLAND
WU Pei, GAO Ke-Ji, LIU Tao, WANG Ya-Fen
 Available online   , doi: 10.7541/2023.2022.0156
[Abstract](94) [FullText HTML](47) [PDF 1159KB](2)
Abstract:
Two pilot-scale deep integrated vertical flow constructed wetland systems (IVCW, total matrix height=180 cm) were constructed with tetrabrominated diphenyl ethers (BDE-47) as target pollutants. Under the condition of fixed hydraulic retention time (HRT=3d) and hydraulic load (0.3m/d), the step-by-step removal rate of BDE-47 (influent concentration=25 μg/L) was analyzed, and the effects of PBDEs on the conventional purification efficiency and substrate microorganisms of IVCW systems were discussed. The batch experimental results showed that after three months of operation, the removal rate of BDE-47 in simulated sewage by IVCW reached 99.9%. Compared with the control group, the removal rate of ammonia nitrogen in BDE-47 treatment group increased from 72.3% to 82.9%, but the removal rate of nitrate nitrogen decreased from 53.0% to 28.1%, and the removal rate of chemical oxygen demand (COD) decreased from 88.1% to 82.3%. The residual amount of BDE-47 in the substrate of each unit of IVCW decreased gradually along the flow direction, and the content of BDE-47 was highest in the upper unit of the downstream flow nearest to the inflow (0.01 μg/g). A significant decrease in substrate microbial dehydrogenase activity was resulted due to BDE-47. Correspondingly, microbial fatty acid profile analysis showed that there was a significant difference in microbial community composition of the down-flow units between the experimental and the control groups, which were dominated by more tolerant anaerobic Gram-negative bacteria and fast-growing aerobic Gram-negative bacteria, respectively. The removal potential of BDE-47 and its migration and transformation mechanisms in IVCW could provide useful reference and a feasible scheme for the removal of refractory organic pollutants by ecological engineering systems.
ESTABLISHMENT AND CHARACTERIZATION OF A TESTICULAR CELL LINE FROM CHINESE STURGEON (ACIPENSER SINENSIS)
YE Huan, WANG Yi-Zhou, DU Hao, YUE Hua-Mei, RUAN Rui, LUO Jiang, LI Chuang-Ju
 Available online   , doi: 10.7541/2023.2022.0139
[Abstract](81) [FullText HTML](59) [PDF 1365KB](7)
Abstract:
The Chinese sturgeon, Acipenser sinensis, is a large anadromous fish, with an average 14 years for males and 21 years for females. It was listed as critically endangered by the International Union for Conservation of Nature (IUCN) since 2010, and its spawning activity was not observed during the past five spawning seasons. Therefore, there is an urgent need to preserve the genetic resources of Chinese sturgeon. To investigate the germplasm conservation of Chinese sturgeon and its spermatogonia stem cell culture in vitro, a new cell line (designated as AST) derived from the testis of Chinese sturgeon was established and characterized. The AST cell line are fibroblast-like cells, and have been stably subcultured for more than 80 passages. The optimal growth conditions for the AST cell line are DMEM medium, 25℃, and 15% FBS. After cryopreservation in liquid nitrogen, the viability of AST cells was about (81.36±1.13)%, as measured by trypan blue staining. Chromosome analysis of AST cells at the 30th passage showed that the number of chromosomes ranged from 142 to 310, and the modal number was 264. To detect the expression characterization of Sertoli cell-related genes (amh and gsdf), Leydig cell-related genes (cyp17a1), and germ cell-related genes (dazl, dnd, and vasa) in AST cells by RT-PCR, all of the genes were found in the P0 and P1 cells, where their amounts were similar with those of in testis; however, in P15, P30, and P60 AST cells, only amh and vasa genes were detected with very low level, suggesting only a few number of Sertoli cells and germ cells in the late passage of AST cells. The signal of enhanced green florescence protein (GFP) were detected in a few of AST cells after transfection with pEGFP-N3 plasmid. The establishment of the AST cell line could provide important experimental material for the conservation genetic resources of Chinese sturgeon, the in vitro study of proliferation and differentiation of spermatogonia stem cell, and function analysis of testicular related genes.
ENVIRONMENTAL COPPER ON GROWTH PERFORMANCE, LIPID METABOLISM, HEPATOPANCREAS AND SPLEEN HISTOLOGY OF GIFT TILAPIA (OREOCHROMIS NILOTICUS)
YAN Zi-Yuan, DENG Zhen-De, ZHANG Jing, LIANG Zhen-Ye, XU Chang
 Available online   , doi: 10.7541/2022.2021.0237
[Abstract](102) [FullText HTML](68) [PDF 2929KB](4)
Abstract:
Excessive environmental heavy metals can induce lipid accumulation and cause hepatopancreas and spleen damage of fish. In order to explore the effects of different concentrations of Copper (Cu) on growth performance, hepatopancreas lipid metabolism and spleen immunity, tilapia [initial weight of (0.45±0.02) g] were randomly assigned to 0 (control), 0.2, 0.4 and 0.8 mg/L Cu concentration, respectively. After four-week cultivation, hepatopancreas, serum, and spleen were sampled. The results showed that: 1. Under waterborne Cu exposure, the survival, weight gain, and specific growth rate of the tilapia showed a significant decline. The hepatosomatic index of tilapia in the 0.4 and 0.8 mg/L Cu concentration groups were significantly higher than those in control and 0.2 mg/L group (P<0.05). 2. Compared to control, triglycerides contents in hepatopancreas of tilapia under 0.4 and 0.8 mg/L groups increased significantly (P<0.05), and no significant difference was found in 0.2 mg/L group. There was no significant difference of total cholesterol content (T-CHO) and 3-hydroxy-3-methylglutaryl-Coenzyme A reductase activity in hepatopancreas among all groups (P>0.05). 3. Under Cu exposure, serum low-density lipoprotein cholesterol, T-CHO content and aspartate aminotransferase activity (P<0.05) were significantly higher than that of control group. Serum triglyceride content decreased significantly thereafter increased with the highest value in 0.8 mg/L group (P<0.05). The alanine aminotransferase activity of tilapia under 0.8 mg/L group was significantly higher than 0.2 and 0.4 mg/L groups (P<0.05), but had no significant difference of control. 4. The histological analyses indicated that the hepatopancreases tissue of tilapia showed vacuolization seriously, and the lipid droplets content was significantly higher than that in control. Environmental copper also caused more and larger melano-macrophage centers (MMCs) in spleen, and lipofuscin increased remarkably within MMCs. The results suggested that waterborne Cu exposure significantly reduced the growth performance of tilapia, and caused lipid deposition in hepatopancreas and serum, further induced hepatopancreas and spleen damage. This study provided basic date for clarifying abdominal lipid accumulation and spleen histological injury in aquaculture species under heavy metal environment.
THRESHOLD OF THE JUVENILE ACIPENSER DABRYANUS DUMERIL
XIANG Jie, ZOU Qiao-Lin, DU Hao, WANG Cheng-You, WEI Qi-Wei
 Available online   , doi: 10.7541/2022.2021.0239
[Abstract](117) [FullText HTML](69) [PDF 960KB](3)
Abstract:
As an important "golden waterway" in China, the Yangtze River has suffered more and more destruction by human activities, such as ship transportation, channel improvement, bridge construction, hydraulic operations, dock loading and unloading, all of those caused noise pollution and influenced the fish lived in the river, even caused fish death. Acipenser dabryanus Dumeril as an important fish in the Yangtze River, its necessary to evaluate how the noise effects it. Based on this, in order to simulate the natural noise environment in the wild, the indoor noise control was used to this experiment, a stimulation frequency of 100—500 Hz was set, the Auditory Evoked Potential was used to test the threshold of juvenile Yangtze sturgeon (Acipenser dabryanus Dumeril). The results showed that the most sensitive frequency of the Acipenser dabryanus Dumeril is 300 Hz, the sound pressure is 133±0.5 dB, the hearing curve of Acipenser dabryanus Dumeril is a "V"-shaped. Different from other fishes, the Acipenser dabryanus Dumeril has a higher hearing threshold, but it cannot hear frequency above 500 Hz. Compared with the Acipenser fulvescens and Aspiolucius merzbacheri, the hearing thresholds between the three are similar. When compared with the common freshwater fish in the Yangtze River, the results showed that Acipenser dabryanus Dumeril has a higher threshold and narrower range of hearing frequency. Results of the experiment will provide important basic data for the protection and population reconstruction of the Acipenser dabryanus Dumeril, which also can provide basic data support for evaluating the impact of the fish-related projects of the fish lived in the Yangtze River.
EFFECTS OF OVERWINTER STARVATION ON BIOCHEMICAL COMPOSITION AND TRANSCRIPTIONAL LEVEL OF GLUCOSE-LIPID-PROTEIN METABOLISM RELATED GENES IN GRASS CARP (CTENOPHARYNGODON IDELLUS)
WU Wen-Yi, JI Hong
 Available online  
[Abstract](101) [FullText HTML](63) [PDF 1439KB](3)
Abstract:
Overwinterstarvation is one of the environmental stress factors in grass carp Ctenopharyngodon idellus culture. During the overwinter starvation process, the energy mobilization mechanism of grass carp may be different from that under normal conditions. Although someone have studied the metabolism of protein and lipid in grass carp under starvation at a normal situation, the response of glucose, lipid and protein metabolism to overwinter starvation has not been reported. In order to explore the metabolic adaptation mechanism of energy utilization of grass carp during overwinter starvation, 45 fish with the initial body weight of (1053.33±16.11) g were randomly distributed into the three repetitions for natural overwinter starvation. When the water temperature naturally decreased to the point, the grass carp ceased ingestion (water temperature: 15 °C), the experiment began. When the water temperature naturally rose to the point, the grass carp refeeding commenced (water temperature: 15 °C), the overwinter starvation period, and thus our experiments, concluded. Samples were collected after 0, 1, 2, 4, 8, 12 and 16 weeks of natural overwinter starvation. The contents of muscle routine components, serum energy metabolites, tissue glycogen, triglyceride, and transcription levels of AMP activated protein kinase and glucose-lipid-protein metabolism related genes were detected. The results showed that the content of various conventional components in grass carp muscles changed significantly after overwinter starvation for 1 week (P<0.05); with the extension of overwinter starvation time, serum triglycerides (TG), glycerol, total protein (TP), total cholesterol (TCHO) and blood sugar (GLU) first decreased significantly (P<0.05), then remained stable, and the content of free fatty acids increased significantly (P<0.05); hepatopancreas glycogen and muscle glycogen and the content of TG in hepatopancreas, muscle and adipose tissue reduced significantly (P<0.05); the content of serum ATP, ADP and AMP reduced significantly, and the ratio of ADP+AMP/ATP increased significantly(P<0.05); the expression ofAMPKα1 and AMPKα2 genes in the hepatopancreas, muscle and adipose tissue increased significantly (P<0.05), and the transcription level of downstream glucose-lipid and protein metabolism related genes increased significantly (including ATGL, HSL, CPT1α, CD36 and other lipolysis related genes; GK, PFK, PK and other glycolysis related genes; GLDH, IGF-1 and other proteolysis-related genes) or significantly down-regulated (ACC, FAS and other lipid synthesis-related genes; CREB, FoxO1, PGC-1α, PEPCK, G6Pase, GLUT2 and other gluconeogenesis related genes; TOR, S6K and other related to protein synthesis genes) (P<0.05). The results revealed that grass carp utilized energy storage to maintain its energy homeostasis during the overwinter starvation period with significant changes in the biochemical composition of serum, hepatopancreas, muscle and adipose tissue of grass carp during the overwinter starvation period has undergone significant changes. At the same time, the overwinter starvation activates the AMPK pathway and promotes glycolysis, lipid decomposition, fatty acid β-oxidation, the process of fatty acid transport and protein decomposition inhibits the process of glycogen synthesis, lipid synthesis and protein synthesis, thereby maintaining the body's energy homeostasis. This study showed that the changes of biochemical components and the regulation of transcription level of glucose-lipid-protein metabolism genes based on AMPK pathway in grass carp during overwinter starvation, and revealed the regulatory content and mechanism of adaptive changes of grass carp during overwinter starvation.
POPULATION DYNAMICS AND PARASITISM PREFERENCE OF DIPLOSTOMUM SPATHACEUM IN GYMNOCYPRIS SELINCUOENSES FROM SELINCO LAKE, XIZANG AUTONOMOUS REGION, CHINA
PAN Ying-Zi
 Available online   , doi: 10.7541/2023.2022.0143
[Abstract](102) [FullText HTML](63) [PDF 0KB](3)
Abstract:
Through the population dynamics, study the temporal and spatial changes of Diplostomum spathaceum infection, and study whether the infection has parasitic preference for host gender, left and right eyes and different parts of eyeball, find out the infection of D. spathaceum in Gymnocypris selincuoenses from Selinco Lake, analyze the causes of population growth and decline, and explore its life history strategy. Catching the G. namensis in different seasons over the year, and recorded the total length, weight and gender. Collected and counted numbers of D. spathaceum, and calculated the prevalence and mean abundance in different seaons and of different total-length groups of G. namensis. Through independent sample nonparametric test, it was judged whether there were significant differences in the number of infections in different gender hosts, left and right eyes and different parts of eyeball, so as to test whether there were parasitic preferences of D. spathaceum. A total of 165 G. namensis (28.7—49.5 cm in length, 37.9±4.0 cm in average length, 196.9—827.2 g in weight, 473.3±127.9 g in average, including 82 females and 83 males) were dissected. A total of 515 D. spathaceum were detected, and the maximum parasitic amount was 32 per fish. We found D. spathaceum can infect D. spathaceum all year round. In the summer and autumn of 2020, the prevalence and mean abundance showed a downward trend. In 2021, the prevalence was high in spring, decreased in summer, and the highest in autumn. The mean abundance showed an upward trend. According to the total-length range of G. namensis, they were divided into five total-length groups at an interval of 5 cm. The prevalence and mean abundance were the lowest in the total-length group of 25 cm≤TL<30 cm, close in the total-length groups of 30 cm≤TL<35 cm, 35 cm≤TL<40 cm and 40 cm≤TL<45 cm, and increased greatly in the total-length group of 45 cm≤TL<50 cm. There was no preference in the host of different genders, and left and right eyes of the hosts, but there was an obvious preference in the lens and vitreous humours, and more parasitic in the lens. The seasonal dynamics of the population growth and decline of D. spathaceum are closely related to environmental factors such as water temperature, the migration time of migratory birds and the existence of snails. With the increase of the total length of G. namensis, the eyeball volume increases, which can accommodate more eye flukes, and the long-term cumulative contact also makes the number of infections more. In the 45 cm≤TL<50 cm total-length group, the prevalence and mean abundance increased sharply, may be because of the larger body size represents a larger surface area, which can not only be invaded by more cercariae, but also can help G. namensis to survive under the increasing infection level to a certain extent, so as to accumulate more D. spathaceum. The parasitic preference of D. spathaceum in Gymnocypris selincuoenses from Selinco Lake is a life history strategy to adapt to the environment, and its purpose is to be conducive to the spread and reproduction of the population.
ANALYSIS OF MOLLUSCS COMMUNITY STRUCTURE IN THE ZHEJIANG COAST
LU Zhan-Hui, ZHOU Yong-Dong, ZHU Wen-Bin, XU Kai-Da
 Available online   , doi: 10.7541/2023.2022.0203
[Abstract](107) [FullText HTML](59) [PDF 0KB](3)
Abstract:
Based on survey data collected in 4seasons of 2019 by single bottom trawling of Zhejiang surrounding waters, the paper analysed species composition, dominant species and the distribution of resources density of molluscs. Ecosystem diversity index, Species similarity index(Js), Alternate index (AI), Migration index (MI) and abundance/biomass comparison curve (ABC curve) were adopted to analyse species diversity of community and its dynamic changes. The results showed that 62 molluscs species were captured belonging to 3 classes 10 orders, and 32 families, and the dominant species were Octopus variabilis, Loligo edulis, Turritella bacillum, Abralia multihamatai, and Sepiola birostrata all the year. The seasonal variation of dominant species was quite different. The annual average resource density was 204.94 kg/km2, The resource density in summer was the highest value in the whole year, and Winter was the lowest. The average resource density was increasing gradually from north to south roughly. The annual average values of species abundance index (D), species diversity index (H’) and species evenness index (J’) were 0.70, 0.88 and 0.61 respectively. Three indices values indicated that molluscs community diversity was on a low level. The values of Species similarity index (Js), Alternate index (AI) and Migration index (MI) indicated that the community stability was higher in spring and winter than in summer and autumn. According to the ABC curve, molluscs community all the year were moderately disturbed respectively.
TAXONOMY OF MICROCOLEACEAE IN BIOLOGICAL SOIL CRUSTS IN THE GURBANTUNGGUT DESERT
LIU Yan-Jin, LI Kai-Kai, ZHANG Ya-Li, ZHAO Kang, ZHANG Bing-Chang
 Available online   , doi: 10.7541/2023.2022.0160
[Abstract](123) [FullText HTML](76) [PDF 1578KB](2)
Abstract:
Biological soil crusts (BSCs) play critical ecological functions in desert ecosystem. Microcoleus sp. are key filamentous cyanobacteria and play vital roles in BSCs. More and more strains of Microcoleaceae were found in desert areas. However, it is difficult to distinguish them in species level due to similar morphological characteristic. In this manuscript, 11 filamentous cyanobacterial strains with similar morphology to Microcoleus were isolated and purified from BSCs in Gurbantonggute desert. The experimental cyanobacterial strains were examined morphologically as well as phylogenetically using 16S rRNA gene and the 16S—23S internal transcribed spacer (ITS) rigion. The results show that the experimental cyanobacterial strains belong to the genera Microcoleus and Symplocastrum, including two newly recorded species in China: M. steenstrupii and S. flechtnerii, as well as M. vaginatus and a suspect species similar to M. steenstrupii. The number and alignment of cyanobacterial filaments, cell size and the shape of apical cells, and phylogenetic relationship based on 16S rRNA are key evidence to identity different species of Microcoleaceae. Secondary structure of ITS are also vital reference to distinguish to different species in same genus.
COMMUNITY ECOLOGY OF INTESTINAL HELMINTHS IN GYMNOCYPRIS WADDELLII FROM YAMDROK LAKE, XIZANG AUTONOMOUS REGION, CHINA
PAN Ying-Zi, TASHILHAMO
 Available online   , doi: 10.7541/2023.2022.0144
[Abstract](130) [FullText HTML](75) [PDF 1330KB](3)
Abstract:
In order to reveal the diversity and distribution of parasites, as well as the interaction and coexistence mechanism among parasites, analyze the relationship among parasites, hosts and environment, study the community ecology of intestinal helminths in Gymnocypris waddellii from Yamdrok Lake, understand the species diversity of aquatic organisms in Xizang Autonomous Region, explore the community characteristics and interspecific relations, and accumulate data for studying the relationship among parasites, Tibetan plateau environment and unique fish hosts, 180 G. waddellii (120 females and 60 males, with a total length of 22.20—49.20 cm, an average length of 36.76±4.18 cm, a body weight of 77.3—896.7 g, and an average weight of 425.92±148.27 g) were dissected in July 2020. The contents of community ecology such as community structure and interspecific relationship were analyzed. The results showed that the intestinal helminth community of G. waddellii in Yamdrok Lake were composed of five species: Parabreviscolex niepin, Contracaecum eudyptulae, Streptocara sp., Neoechinorhynchus sp. and Allocreadium sp.. From high to low, the prevalence of populations were Neoechinorhynchus sp., P. niepin, Allocreadium sp., C. eudyptulae and Streptocara sp.; the infection intensity of populations were P. niepin, Neoechinorhynchus sp., C. eudyptulae, Allocreadium sp. and Streptocara sp.; the average abundance of populations were P. niepin, Neoechinorhynchus sp., Streptocara sp., Allocreadium sp. and C. eudyptulae. The Margalef index of community was 0.59, Shannon-Wiener index was 1.26, Pielou index was 0.83 and Berger- Parker index was 0.50. The dominant species was P. niepin. There were positive correlations among four populations, and the order of correlation from high to generation were as follows: Neoechinorhynchus sp.and Allocreadium sp, C. eudyptulae and Neoechinorhynchus sp., C. eudyptulae and Allocreadium sp., C. eudyptulae and Streptocara sp.. There were no interspecific association between other parasitic worm populations. In terms of infection or not, the number of infected hosts was more than half of that of the sampled population. Among them, the host frequency with one parasite was the most, the host frequency with two parasites was also more, the host frequency with three and four parasites were significantly reduced, and no hosts infected with five parasites at the same time was found. Compared with Lake Chugutso, which is also located in southern Xizang and once connected with Yamdrok Lake, although the two lakes are geographically similar, the host fish species are the same, and the intestinal helminth species composition is the same, intestinal helminths in G. waddellii from Yamdrok Lake has its own characteristics, that is, higher average abundance, and most parasitic worm populations also have higher prevalence, and the dominant species in the community are also different from Chugutso Lake. Interspecific association is used to judge whether there is coexistence or exclusion between various groups in intestinal helminths community, that is, interspecific affinity. In the intestinal helminths community of G. waddellii from Yamdrok Lake, the affinity between the undetermined species of Neoechinorhynchus sp. and Allocreadium sp. were the highest. However, this relationship is not stable and will change with the changes of water ecological environment and species composition in the community. The frequency of co-infected hosts suggests that with the increase of species in a sub community, the greater the interaction between species, the more difficult it is to maintain coexistence.
REMOVAL EFFICIENCY AND MECHANISM OF BDE-47 BY THE ZN-FE LDHS MODIFIED QUARTZ SAND-BIOFILM SYSTEM
CAI Qin, LI Yan-Ni, WANG Chang, WANG Ya-Fen
 Available online   , doi: 10.7541/2023.2022.0158
[Abstract](118) [FullText HTML](66) [PDF 2261KB](2)
Abstract:
In this paper, Zn-Fe layered double hydroxides (LDHs) modified quartz sand was prepared by coprecipitation method and cultivated with inoculation of Shewanella putrefaciens CN32 under aerobic, anaerobic and alternating conditions, to examine the biofilm formation process on LDHs modified matrix and its removal efficiency of BDE-47 as the target pollutant. The biotic and abiotic removal mechanisms of BDE-47 were further explored by monitoring variations of Fe2+ and H2O2 concentrations in the reaction systems. The results showed that LDHs coating would not affect the formation of biofilm on the surface of the modified quartz sand, but Zn-Fe LDHs modified matrix showed certain inhibitory effect on electron transport system activity of CN32 under aerobic condition, while the composition of extracellular polymer substance (EPS) of matrix biofilm changed with an increase in the proportion of polysaccharide under anaerobic condition. Total concentrations of EPS in the matrix biofilm reaction systems were significantly higher than those in the pure CN32system under both aerobic and anaerobic conditions.The formation of matrix biofilm significantly improved the removal efficiency of BDE-47 in the reaction system (about 25%) under aerobic and anaerobic alternating condition. Under the alternating condition, the removal of BDE-47 in the first three cycles (within 72h) depended mainly on matrix adsorption; while 72h later, biofilm adsorption and biodegradation contributed together, and LDHs modified matrix exhibited greater boost potential in the later stage. This study demonstrated the biofilm formation characteristics of LDHs modified matrix and its potential for the removal of PBDEs from the aqueous phase, and provided new ideas for enhancing the biodegradation of PBDEs in constructed wetlands.
IDENTIFICATION AND INDUCED EXPRESSION OF THE METALLOTHIONEIN GENE OF A HIGH CD-TOLERANCE COLPODA INFLATA POPULATION
HOU Sen, ZHAO Xue, GAO Rong, WANG Ying, ZHENG Wei-Bin, WANG Li, PAN Xu-Ming, REN Nan-Qi, CHEN Ying
 Available online   , doi: 10.7541/2023.2022.0165
[Abstract](95) [FullText HTML](65) [PDF 1711KB](2)
Abstract:
Cadmium is the first of the heavy metal pollutants to be controlled in water. Metallothionein is one of the key proteins that can bind to cadmium and regulate the oxidative stress response of organisms. The protozoan MT has been reported in two ciliates, Tetrahymena and Paramecium. In this study, we obtained a northeast population of Colpoda inflata with high cadmium tolerance. Metallothionein content of C. inflata showed a positive correlation with cadmium concentration and growth rate of population in five gradient concentration of cadmium stress experimental groups from 24—96h. The highest cd-tolerance concentration was 10 mg/L in 96h. Metallothionein gene of Colpoda inflata was cloned and named Col-MT1. The gene sequence and amino-acid sequence were analyzed. The results showed that Col-MT1 had high homology with the amino acid sequences of other ciliates, and contains two conserved sites XXCXX and XCCX. It was a new member of subtype 7a of metallothionein gene family. TASSER protein model predicted that the secondary structure of Col-MT1 protein was composed of α-helix, β-folding and random crimp, accounting for 67.90%, 11.11% and 20.98%, respectively. SWISS-MODEL predicted that the 3D structure of Col-MT1 protein was the most similar to that of cyanobacteria metallothionein. qRT-PCR experiment confirmed that at 60h, 84h and 108h, the expression of Col-MT1 gene was up-regulated to different concentration cadmium and showed a dose-response relationship with cadmium concentration. The molecular mechanism of gene expression still needs further study. Above results supplemented the MT gene database of protozoa and lay a foundation for revealing the mechanism of C. inflata MT gene. As well as provide reference for monitoring and remediation of cadmium pollution.
EFFECT OF DIETARY BLACK SOLDIER FLY (HERMETIA ILLUCENS) LARVAE MEAL ON GROWTH PERFORMANCE, NON-SPECIFIC IMMUNITY AND LIPID METABOLISM OF LITOPENAEUS VANNAMEI
CHEN Yong-Kang, ZHENG Chao-Zhong, ZHANG Shuang, CHI Shu-Yan, DONG Xiao-Hui, YANG Qi-Hui, LIU Hong-Yu, TAN Bei-Ping, XIE Shi-Wei
 Available online   , doi: 10.7541/2023.2022.0201
[Abstract](101) [FullText HTML](70) [PDF 765KB](2)
Abstract:
This experiment was conducted to investigate the effects of dietary black soldier fly (Hermetia illucens) larvae meal on growth performance, non-specific immunity and lipid metabolism of Litopenaeus vannamei. The control diet (FM) contained 25% fishmeal and then 10% (BSF10), 20% (BSF20) and 30% (BSF30) of fishmeal protein were replaced by black soldier fly larvae meal to formulate four isonitrogenous and isolipidic diets. Shrimps with an initial body weight of (0.88±0.01) g were fed four times daily for 7 weeks. Results showed that there were no significant differences in growth performance of shrimp fed BSF10 and BSF20 diets compared to FM group (P>0.05), but the growth performance of the shrimp in the BSF30 group significantly reduced (P<0.05). The whole shrimp crude lipid content was significantly lower in the BSF20 and BSF30 groups, and the hemolymph triglyceride and total cholesterol levels were significantly lower in the BSF30 group compared to FM group (P<0.05), but there were no significantly different in crude lipid of hepatopancreas among the four groups (P>0.05). The activities of hemolymph alanine aminotransferase and aspartate aminotransferase significantly decreased in the BSF10, BSF20 and BSF30 groups compared to FM group (P<0.05). The total antioxidant capacity of BSF20 group was significantly higher than that of the other groups, and superoxide dismutase activity of BSF10 group was significantly higher than that of FM group (P<0.05). The hepatopancreas acid phosphatase and alkaline phosphatase activities were significantly lower in the BSF30 group than in the FM group (P<0.05). The lipase activity of BSF30 group was significantly lower than BSF10 group (P<0.05). The fatty acid synthase activity in BSF10 and BSF20 groups was significantly higher than that in FM group, the carnitine palmitoyl transferase activity in BSF20 group was significantly higher than the other three groups, and the activity of isocitrate dehydrogenase in BSF30 group was significantly higher than that in FM group (P<0.05). The activities of hemolymph lipid triglyceride lipase and α-ketoglutarate dehydrogenase in BSF10, BSF20 and BSF30 groups were significantly higher than those in FM group (P<0.05). To conclude, replacing 10% or 20% of fishmeal with black soldier fly larvae meal had no negative effect on the growth performance of Litopenaeus vannamei, improved the antioxidant capacity of shrimp and reduced the hemolymph glutamate transaminase and glutamic acid transaminase activities. The crude lipid of whole shrimp significantly reduced in BSF20 group, and when the replacing level reached to 30%, the activity of enzymes related to lipid catabolism and tricarboxylic acid cycle was significantly increased, which further promoted the lipid metabolism of shrimp.
SCREENING OF PEPTIDES INTERACTING WITH GRASS CARP REOVIRUS VP39 PROTEIN BY PHAGE DISPLAY TECHNOLOGY
LONG Chen, XU Ning, XIE Ya Qing, LÜ Li Qun
 Available online   , doi: 10.7541/2023.2022.0179
[Abstract](109) [FullText HTML](67) [PDF 968KB](6)
Abstract:
VP39 is a protein encoded by S9 gene of type Ⅲ grass carp reovirus (GCRV-Ⅲ). In order to study the biological function of VP39 protein in the process of GCRV infection of grass carp cells, the sequence of VP39 gene was cloned and the prokaryotic expression vector PET32A-VP39 was constructed. The fusion protein VP39-HIS was obtained by using prokaryotic expression method. Mouse anti-VP39 polyclonal antibody was prepared by immunizing mice with VP39solution protein, and the antibody was evaluated by Western Blot. The polyclonal antibody was used to investigate the expression dynamics of VP39 protein in GCRV infected cells. The results of SDS-PAGE showed that the fusion protein of vp39-his was well soluble in PBS and the protein size was about 39kDa. Western Blot analysis showed that the prepared VP39 polyclonal antibody could recognize both the prokaryotic expression of VP39-HIS fusion protein and the expression of VP39 protein after GCRV infection with CIK cells at the dilution ratio of 1:10000, showing good titer and specificity. In the process of virus infection, the expression of VP39 was low in the early stage and high in the middle and late stage. Two peptides were screened by phage display technology for specific binding to VP39 protein, and further bioinformatics analysis also found that 7 genes in grass carp genome had homology with the peptide, indicating that these genes may interact with VP39. In this study, mouse anti-VP39 polyclonal antibody was prepared, which provided a new immunological method for GCRV-Ⅲ detection. The screening of binding peptides also laid a foundation for the study of the biological function of VP39 in the process of GCRV infection.
APPARENT DIGESTIBILITY COEFFICIENTS OF SEVEN PROTEIN SOUCES FOR JUVENILE HYBRID GROUPER (EPINEPHELUS FUSCOGUTTATUS♀ × EPINEPHELUS LANCEOLATUS♂)
ZHENG Chao-Zhong, CAO Jun-Ming, DONG Xiao-Hui, CHI Shu-Yan, ZHANG Shuang, YANG Qi-Hui, LIU Hong-Yu, DENG Jun-Ming, ZHANG Wei, TAN Bei-Ping, XIE Shi-Wei
 Available online   , doi: 10.7541/2023.2022.0172
[Abstract](109) [FullText HTML](65) [PDF 776KB](2)
Abstract:
The apparent digestibility coefficients (ADCs) of Tenebrio molitor meal (TMM), Hermetia illucens meal (HIM), Clostridium autoethanogenum protein (CAP), Methylococcus capsulatus meal (MCM), Chlorella vullgaris meal (CVM), Cottonseed protein concentrate (CPC) and Peruvian fishmeal (PFM) were determined in juvenile hybrid grouper (Epinephelus fuscoguttatus♀×Epinephelus lanceolatu♂). A basal diet (including 50% fishmeal) and seven test diets (700 g/kg of the basal diet and 300 g/kg of each test ingredient) were formulated with 0.1% yttrium oxide (Y2O3) as an inert marker. The juvenile hybrid groupers, with initial average body weight of (9.95±0.50) g, were randomly distributed into 0.3 m³ fiberglass tanks, each tank with 30 fish. The faeces samples were collected twice-daily by siphoning following feeding fish after five days of domestication. The ADCs of dry matter of seven test ingredients were ranked as CVM>TMM=CAP=CPC>HIM=MCM=PFM (P<0.05). CVM showed the highest ADCs of dry matter (DM), crude protein (CP) and most amino acids (including methionine and threonine) except crude lipids (CL), whereas HIM had the relatively lower ADCs of DM, CP and most amino acids except CL. CAP had a higher lysine digestibility than the other six test ingredients, and was only lower than CVM in the ADC of CP. The ADC of DM in PFM was significantly lower than that in CVM (P<0.05), and showed no differences with that in CAP (P>0.05). Besides, PFM showed a lower ADC of CP than the ADCs of CP in CVM, CAP and MCM (P<0.05), and showed a lower ADC of lysine than that in CAP as well as a lower ADC of threonine than those in CAP and CVM (P<0.05). Overall, this study showed the advantage of CVM and CAP among the seven protein sources on the digestibility of feed available in hybrid grouper.
COMPARISON OF IMMUNE CHARACTERISTICS OF CHINESE STURGEON CULTURED IN SEA AND FRESH WATER
CHEN Xue-Hui, LENG Xiao-Qian, LUO Jiang, DU Hao, LIU Zhi-Gang, QIAO Xin-Mei, XIONG Wei, WEI Qi-Wei
 Available online   , doi: 10.7541/2023.2022.0129
[Abstract](136) [FullText HTML](65) [PDF 5661KB](2)
Abstract:
Chinese sturgeon (Acipenser sinensis) is a large anadromous migratory fish. In order to understand the immune characteristics of freshwater cultured Chinese sturgeon in seawater, a 5-month mariculture trial was conducted with 4-year-old Chinese sturgeons, and the adaptive changes of blood physiology, biochemistry and immune tissue were explored. The results showed that the white blood cell counts were significantly increased (P<0.05), and there was no significant difference on red blood cell count. But the hemoglobin and hematocrit increased significantly in the seawater from (46.50±10.59) g/L, (13.7±3.23)%, (9.19±1.10)×109/L to (74.5±1.10) 11.05) g/L, (21.80±3.33)% and (10.88±3.73)×109/L, respectively. In the differential count of white blood cells, lymphocytes accounted for the largest proportion, followed by neutrophils, and there was no significant difference in the percentages of various types of white blood cells. Blood biochemical indexes such as SOD, MDA, LZM, IgM, AKP and ACP have no significant changes in freshwater and seawater. Observation of immune tissue structure showed that there was no obvious change in the thymus in seawater, but the cells were aggregated, the shape distribution was more orderly and the center of melanin macrophages increased in the head kidney tissue; Lymphocytes and red blood cells are more densely distributed in the spleen tissue. The results showed that marine aquaculture can enhance the immunity and hematopoietic function of Chinese sturgeon to a certain extent, and maintain a good physiological state.
PURGING TIME ON MUSCLE QUALITY AND NUTRITIONAL VALUE OF YELLOW CATFISH (PELTEOBAGRUS FULVIDRACO) UNDER OPEN FLOWING WATER MODE
FU Yun-Yin, ZHEN Wei-You, ZHANG Heng, RUAN Guo-Liang, LIU Yu-Lin, CHAI Yi, FANG Liu
 Available online   , doi: 10.7541/2023.2022.0075
[Abstract](103) [FullText HTML](68) [PDF 764KB](1)
Abstract:
This experiment was conducted to investigate the effect of purging time on muscle quality and nutritional value of yellow catfish (Pelteobagrus fulvidraco) under open flowing water mode. Yellow catfish of (15.69±2.28) g at the rapid growth stage were randomly divided into 4 groups with 3 replicates in each group. The yellow catfish were temporarily fed for 0 (control), 20, 30 and 40 days respectively. During the experiment, Pelteobagrus fulvidraco were fed with a compound diet, and the water quality was regularly measured during the breeding process. After the experiment, the growth performance and serum biochemical indexes were measured; the composition of amino acids, fatty acids and textural properties in muscle were compared. The results showed as follows: 1) During the purging time period, the content of ammonia nitrogen in the water of the test tank was between 0.03—0.05 mg/L, the content of nitrite nitrogen was 0.01 mg/L, and the dissolved oxygen level exceeded 9.0 mg/L. 2) With the extension of purging time, the final body weight (Wt) and weight gain rate (WGR) of yellow catfish showed an increasing trend, while the specific growth rate (SGR) showed a decreasing trend, with significant differences among groups (P<0.05). There were no significant differences in survival rate (SR), condition factor (CF), hepatosomatic index (HSI) and viscerosomatic index (VSI) of purging time groups for 20, 30 and 40 days (P>0.05). 3) The contents of albumin (ALB) and total protein (TP) in serum of purging time 30 and 40 days group were significantly higher than those of control group and purging time of 20 days group (P<0.05), but the activities of gamma-glutamyltransferase (γ-GGT), alkaline phosphatase (ALP), the contents of total cholesterol (TC) and total bile acids (TBA) were significantly lower than those of the control group and the purging time of 20 days group; serum total bilirubin (TBIL) content and aspartate aminotransferase (AST) activity in the control group, the purging time of 20 days and 30 days groups were significantly higher than those of the purging time group for 40 days (P<0.05). The content of serum creatinine (CRE) and urine creatinine ratio (UCR) of Pelteobagrus fulvidraco in the purging time groups for 30 and 40 days were significantly lower than those in the control group and the purging time for 20 days group (P<0.05). 4) The crude protein content in muscle of purging time groups for 30 and 40 days was significantly higher than that of control group and purging time group for 20 days (P<0.05), but the crude lipid content was significantly lower than that of control group and purging time group for 20 days (P<0.05). 5) The total amount of amino acids (ΣAA), total essential amino acids (ΣEAA), total non-essential amino acids (ΣNEAA), total umami amino acids (ΣDAA) and essential amino acid index (EAAI) in the 30d and 40d purging time groups were significantly higher than those in the control group and the 20d purging time group (P<0.05). The first limiting amino acid in muscle of four groups was all phenylalanine+tyrosine. 6) The contents of C20:5n-3+C22:6n-3 (EPA+DHA), monounsaturated fatty acid (MUFA) and polyunsaturated fatty acid (PUFA) in muscle of yellow catfish in purging time group for 40 days were significantly higher than those in other groups (P<0.05). 7) The muscle hardness and gumminess properties of purging time group for 40d was significantly higher than those in other groups (P<0.05), and the muscle springiness, chewiness and resilience were significantly higher than those of control group and purging time group for 20d (P<0.05). In this experiment, under the conditions of temporary cultivation, purging time for 40 days can improve the muscle quality and nutritional value of yellow catfish, increase the contents of amino acids and fatty acids, reduce the earthen smell of pond cultured fish, and increase the aquaculture benefit of Pelteobagrus fulvidraco.
EXHAUSTION EXERCISE STRESS ON HYPOXIA AND THERMAL TOLERANCES OF THREE CYPRINID SPECIES
ZHANG Yong-Fei, HUANG Ke-Ren, LUO Yu-Lian, LIU Qian-Ying, PANG Xu, FU Shi-Jian
 Available online   , doi: 10.7541/2023.2022.0193
[Abstract](103) [FullText HTML](74) [PDF 1082KB](2)
Abstract:
The hypoxia and thermal tolerances of fish are important physiological characteristics that determine their distribution, habitat change, and adaptability to climate change. While in the nature, fish are always in the process of swimming or recovery of post-exercise, whether the hypoxia and thermal tolerances change during swimming or immediately after exhaustive recovery process is unknown for fish. Thus, to study the effects of exhaustion exercise stress on fish hypoxia and thermal tolerances, we investigated three cyprinid fish species (i.e. Carassius auratus, Spinibarbus sinensis and Cyprinus carpio) living in different habitats as study cases. Hypoxia and the thermal tolerance indicators of the three fish species were measured after exhaustion exercise, respectively, to determine whether exhaustion exercise stress would affect the stress resistance of fish. In the present study, we found that body weight only affected significantly on minimal critical temperature (CTmin), and the indicators of hypoxia and thermal tolerances were different significantly between species. Moreover, exhaustion exercise stress led to a significant increase in critical oxygen tension (Pcrit) of common carp and a significant increase in critical metabolic rate (CMR) of all the three species as well, but a significant decrease in point of oxygen tension for loss of equilibrium (LOE) of qingbo. Meanwhile, it also resulted in a significant decrease in maximal critical temperature (CTmax) of goldfish and qingbo. However, there was no significant effect on the species and other related measured parameters besides the fish species and their corresponding experimental parameters mentioned above. It could be said based on the results that changes in the hypoxia and thermal tolerances of fish living in different habitats are different after exhaustion exercise stress, and that fish species vary in physiological mechanisms responding to other environmental stressors following exhaustion exercise stress, which may be related to difference in their energy metabolism patterns.
CHARACTERISTICS OF FISH ASSEMBLAGES OF POYANG LAKE AT THE INITIAL STAGE OF THE FISHING BAN
YANG Min, SHENG Piao, ZHANG Yan-Ping, GUO Long-Gen, WANG Hai-Hua, GAO Xiao-Ping, CHEN Yu-Shun
 Available online   , doi: 10.7541/2023.2022.0161
[Abstract](97) [FullText HTML](77) [PDF 1917KB](3)
Abstract:
To systematically investigate fish assemblages of Poyang Lake at the initial stage of the fishing ban, quantitative field investigation on fish communities from a total of 21sampling sites was carried out in four consecutive seasons represented by April as spring, July as summer, October as autumn in 2020 and January as winter in 2021. A total of 57 fish species was collected throughout the study, which belonged to 8 orders, 12 families, and 43 genera. There were 13 dominant species, including Hemiculter bleekeri, Coilia brachygnathus, Saurogobio dabryi, Pseudobrama simoni, Carassius auratus, and others. Migratory and riverine fish accounted for 45.61% of total species. The number of species with an average body weight of less than 20 g per individual represented 88.47% of the total catch. Non-metric multidimensional scaling based on fish abundance divided the samples into two spatial groups, the channel and the main lake of Poyang Lake. Fish abundance of sampling sites in the channel was significantly lower than that in the main lake, while fish diversity was higher in the channel. Fish abundance and biomass varied significantly in four seasons. Diversity during summer and autumn seasons which witnessed high water levels was found higher than those during spring and winter seasons which had low water levels. A new euryhaline fish species, Mugil cephalus, which has never been recorded in this area, was collected both at the channel (S20) and the main lake (S9) sites during the October sampling. This possibly reflected the fishing ban effects on reducing the fishing pressure and promoting the fish resource recovery and migration, and other potential human related factors. Longer term monitoring and tracking is needed. This study provided basic information for assessing the ecological effects of the fishing ban and references for developing fish resources conservation strategies in Poyang Lake.
COMPARISON OF FISHERIES BIOLOGICAL CHARACTERISTICS OF STHENOTEUTHIS OUALANIENSIS IN THE NORTHWEST INDIAN OCEAN IN DIFFERENT CLIMATIC YEARS
LU Hua-Jie, HE Jing-Ru, CHEN Jing, WANG Hong-Hao, CHEN Xuan-Yu, LIU Kai, CHEN Xin-Jun
 Available online   , doi: 10.7541/2023.2022.0159
[Abstract](100) [FullText HTML](65) [PDF 1823KB](3)
Abstract:
According to the 1896samples of Sthenoteuthis oualaniensis collected by Chinese light falling-net fishery during the same months (February-May) in the northwest Indian Ocean, this study presents the differences of fisheries biological characteristics of Sthenoteuthis oualaniensis between 2019 (Ei-Niño) and 2020 (normal). The result indicated that the mantle length (ML) range and dominant ML group of 2019 were larger than that of 2020, especially for females. The ML range of females in both years were 129—347and 284—582 mm, and the dominant ML group were 171—220 and 481—530 mm, respectively. The ML range of males were 138—273 and 94—235 mm, and both of the dominant ML group were 171—220 mm. The analysis of covariance (ANCOVA) showed that the relationships between body weight (BW) and ML of Sthenoteuthis oualaniensis in different years was significantly different between the sexes. By fitting and optimizing the equations and comparing the AIC (Akaike’s information criterion), the relationship between ML and BW of males were most suitable to be expressed by exponential functions, except for the males of 2020 when the relationship between ML and BW of males was most suitable to be expressed by a exponential function. The relationships between ML and BW for both the 2019 and 2020 were the best expressed as power functions, except for the relationship between ML and BW for the 2020 males, which were the best expressed by a exponential function. The gonad maturity composition varied between different years, the females were mostly at immature stages (I and II) in 2019 but mature stages (stages III and IV) in 2020, while the males were mostly at stages II and III in 2019 but mature stages (III and IV) in 2020. The age composition and hatching characteristics differed between years, the females ranged from 97—263d and 148—267d, and dominant age groups were 171—200d and 231—260d with hatching dates from June to December and May to October, and peaked in August-September and July-August, respectively. The males age ranged from 131—253d and 133—221d, with a dominant age 171—200d for both years, and the hatching dates were from August to December and September to December, and the peak periods were September-November and October-November, respectively. This study shows that the fisheries biology of Sthenoteuthis oualaniensisthe in the northwest Indian Ocean varies from climate year to climate.
CHANGES IN CAROTENOID COMPOSITION AND ANTIOXIDANT CAPACITY OF PORTUNUS TRITUBERCULATUS DURING OVARIAN DEVELOPMENT
SUN Qiu-Feng, LIU Mei-Mei, HE Jie, ZHANG Min, TAO Xian-Ji, WU Xu-Gan
 Available online   , doi: 10.7541/2023.2022.0152
[Abstract](98) [FullText HTML](73) [PDF 1021KB](1)
Abstract:
Female P. trituberculatus at different stages of ovarian development (stage I—V) were investigated for their color, carotenoids contents, antioxidant and non-specific immune parameters. The results indicated that: (1) the gonadosomatic index (GSI) increased significantly (P<0.05), while the hepatosomatic index (HSI) rose slightly, then declined gradually. The redness (a*) and yellowness (b*) values in ovaries showed an increasing trend, as did lightness (L*) and b* values in hepatopancreas, but L* values in ovaries showed a decreasing trend. (2) Total carotenoids, astaxanthin, zeaxanthin and β-carotene content in ovaries showed an increasing trend followed by a decreasing trend, with the highest astaxanthin content in stage IV. Astaxanthin content in hepatopancreas was on the rise, while β-carotene was on a significant decline. Regarding the content of carotenoids in different tissues, the epithelium contained the highest amount of astaxanthin. L* values in the ovary and hepatopancreas exhibited a significant negative correlation with total carotenoids, and a* and b* values in the ovary both showed a significant positive correlation with total carotenoids, astaxanthin, luteolin, echinenone and β-carotene. There was no significant correlation between a* values and various carotenoids in hepatopancreas, and b* values only showed a significant positive correlation with astaxanthin. (3) For antioxidant indexes, T-AOC in hepatopancreas showed a significant decrease in stage I—V (P<0.05), while SOD, CAT and GPX showed an "increasing and then decreasing" trend. The T-AOC in hemolymph showed a trend of "high-low-high-low". SOD and GPX showed an increasing trend and then decreasing trend, while CAT and MDA showed a "low-high-low-high" trend. In terms of immune indexes, ACP, ALP and NO in the hepatopancreas showed an overall decreasing trend during ovarian development, while ACP, ALP and Hc in the hemolymph showed an increasing trend in stage I—IV and a slight decrease in stage V. (4) Correlation analysis showed that astaxanthin content in hepatopancreas was significantly negatively correlated with T-AOC, lutein content was significantly positively correlated with GPX (P<0.01), and no significant correlation was found between the other carotenoids content and antioxidant indexes. In conclusion, the increase of total carotenoids, astaxanthin and β-carotene in the ovaries of P. trituberculatus occurred mainly in stage II-IV, while most of the carotenoids in the epithelium showed a significant decreasing trend and only astaxanthin in the hepatopancreas showed an increasing trend.
GROWTH AND BIOENERGETICS OF THE SEA CUCUMBER APOSTICHOPUS JAPONICUS (ECHINODERMATA: HOLOTHUROIDEA) IN RESPONSE TO SEAWATER ACIDIFICATION AND WARMING
QU Liang, XIE Xi, LU Yu-Jie, YANG Xiao-Long, ZHANG An-Guo, WANG Qing-Zhi, YUAN Xiu-Tang
 Available online   , doi: 10.7541/2023.2022.0073
[Abstract](89) [FullText HTML](71) [PDF 765KB](2)
Abstract:
In recent years, threat to nearshore marine ecosystems and marine organisms caused by seawater acidification and seawater warming are becoming increasingly serious. In order to study the effects of CO2-driven ocean acidification and warming on growth and energy budget of the sea cucumber Apostichopus japonicus, which is a key species in the coastal ecosystem of East Asia, four experimental treatments were set up, namely, control (seawater temperature of the Dalian coast, pCO2 400 µatm), ocean warming (OW, seawater temperature of the Dalian coast plus 3℃, pCO2 400 µatm), ocean acidification (OA, seawater temperature of the Dalian coast, pCO2 1100 µatm) and Ocean acidification and warming (OAW, seawater temperature of the Dalian coast plus 3℃, pCO2 1100 µatm). The result showed that A. japonicus in OW were not significantly affected in contrast to control. However, the specific growth rate (SGR) of A. japonicus in OA was the lowest, which decreased by 0.19 %/d compared with the control treatment, and the body weight Coefficient variations of A. japonicus in OA was the largest. The final body weight and SGR of A. japonicus in OAW showed no significantly difference with those in control, bur ingestion rate and feces production rate were both significantly higher than those in the control. The bioenergetic pattern of A. japonicus in OW and OA did not change significantly compared with that in the control, but it changed significantly in OAW, with the percentage of the FCE being significantly higher than the other three treatments. Our study suggests that seawater acidification inhibited the growth of A. japonicus versus change its energy distribution pattern. The decrease of growth in OA mainly depended on the decrease of food ingestion. The combined effect of seawater acidification and rising temperature may compensate for the negative effect of seawater acidification on growth by changing the energy distribution pattern of A. japonicus.
COVALENT MODIFICATIONS OF ENOLASE 1 POLYPEPTIDE CHANGE CELL GROWTH AND METABOLISM PROFILE
SONG Xue-wen, LI Wen-Ling, ZHU Li-Wen, PRITHIVIRAJ Nagarajan, WANG Jun, YAN Ji-Zhou
 Available online   , doi: 10.7541/2022.2021.0221
[Abstract](123) [FullText HTML](69) [PDF 11434KB](1)
Abstract:
Enolase 1 (ENO1) is a rate limiting enzyme of glycolysis, and participates in a variety of physiological and pathological processes according to its cell localization. In this study, zebrafish embryonic fibroblasts (PAC2 cell line) were used to study biochemical functions of ENO1 polypeptide and its covalent modifications. At first, methylated, acetylated, phosphorylated and unmodified eno1 peptides were artificially synthesized, and separately transduced into PAC2 cells. Then the treated cells were collected and examined by using CCK-8 assay, intracellular and extracellular lactate dehydrogenase (LDH) assay, mitochondrial and lysosomal integrity detection kits, as well as quantitative PCR Array. The results showed that the four polypeptides differentially changes cell proliferation, morphology of lysosomes and mitochondria, and glucose metabolic pathways. Furthermore, RNA-sequencing were conducted for the cells treated by acetylated Eno1 polypeptide and vehicle control Transcriptomal analysis indicated that acetylation modified Eno1 polypeptide significantly changed metabolism profile of sugar, lipid and protein, and is closely associated with carcinogenesis, viral infection and cardiomyopathy. These results suggest that multifaceted functional diversity of Enolase 1 may result from its various post-translational modifications.
FISH COMMUNITY STRUCTURE AND ITS RELATIONSHIP WITH ENVIRONMENTAL FACTORS IN JUMA RIVER IN HAIHE RIVER BASIN
WANG Meng, WANG Yin-Xiao, TAN Hui-Min, WANG Min, MU Shu-Mei, Kang Xian-Jiang, CHEN Yong-Xia
 Available online   , doi: 10.7541/2023.2022.0034
[Abstract](108) [FullText HTML](74) [PDF 1285KB](2)
Abstract:
In order to understand the fish community structure in Juma River and its relationship with the environmental factors, 15sampling stations in Juma River were surveyed in May, August and October, 2019, August and October, 2020 and May, 2021. A total of 37 fish were collected, belonging to 11 families and 5 orders. Cypriniformes were the most, accounting for 64.86% of the total species captured. Ecologically, the fish community in Juma River is dominated by demersal and omnivores species and there are fewer carnivorous fish. Compared with the historical data, the number of fish species has decreased. Index Relative Importance (IRI) shows that 10species had IRI≥500, Carassius auratus, Pseudorasbora parva, Zacco platypus were the dominant species. C. auratus, P. parva and Micropercops swinhonis were collected in all seasons and all sampling sites. One-way analysis of variance (ANOVA) showed that the number of individuals (N), species (S), Shannon-Wiener diversity index (H') and Margalef richness index (DMa) of the fish community had significant differences between different seasons, Pielou evenness index (J') showed no significant differences. SNK multiple comparison test showed that the number of individuals and species was the highest in May 2019, Shannon-Wiener diversity index (H') and Margalef richness index (DMa)was the highest in August 2019. The numbers of individuals showed significant differences between different altitudes, an average of 143.91 fishes can be collected at sampling station above 500 meters and only 48.19 fishes can be collected at sampling station below 500 meters. Cluster analysis and Non-metric Multidimensional Scaling (NMDS) divides the source of the Juma River and the Diaojianghui into group Ⅰ, the Zijingguan Bridge and Qingliangjian into group Ⅱ, remaining sites are divided into group Ⅲ. The abundance biomass comparison curve (ABC) showed that the fish community was undisturbed only in August 2020 and was moderately disturbed in the remaining months. Redundancy analysis (RDA) showed that altitude, chlorophyll a, and temperature were the main environmental factors affecting the fish community in Juma River. Altitude is the environmental factor that greatestly affects the structure of fish communities in the Juma River, it has a great influence on Triplophysa cuneicephala, Triplophysa dalaica and Rhynchocypris oxycephalus. Chlorophyll a was highly correlated with the density of phytoplankton and it has a great influence on Micropercops swinhonis, Zacco platypus and other omnivorous fish.
THE OCCURRENCE OF AHPND IN POND CULTURED LITOPENAEUS VANNAMEI AND ITS INFLUENCE ON ENVIRONMENT, PATHOGEN AND IMMUNE FACTORS
YU Yong-Xiang, WANG Yin-Geng, CAI Xin-Xin, ZHANG Zheng, LIAO Mei-Jie, LI Bin, RONG Xiao-Jun, ZHU Hong-Yang, TANG Miao-Miao, WANG Chun-Yuan
 Available online   , doi: 10.7541/2022.2021.0217
[Abstract](128) [FullText HTML](70) [PDF 1043KB](1)
Abstract:
Acute hepatopancreatic necrosis disease (AHPND) is an important limiting factor affecting the healthy cultivation of shrimp in recent years, which has brought huge economic losses to the global shrimp industry. In order to explore the relationship between the occurrence of AHPND and environment, pathogen and shrimp immunity factors, the pond cultured Litopenaeus vannamei systems were continuously monitored. Based on the 35 days continuously monitoring showed that, the air temperature, water temperature, dissolved oxygen (DO), pH, salinity, ammonia nitrogen (NH4-N) and nitrite (NO2-N) were fluctuated in the range of 21—29℃, 24.8—31℃, 1.4—8.32 mg/L, 8—8.91, 34—50, 0.01—0.26 mg/L and 0.005—0.212 mg/L, respectively; the culturable bacteria and Vibrio sp. in water and shrimp hepatopancreas were fluctuated in the range of 3×103—2.4×105 CFU/mL, 2×102—1.8×104 CFU/mL, 9.8×104—8.8×106 and 3.9×103—3.61×106 CFU/g, respectively; 135 isolates of Vibrio sp. were detected based on 16S rDNA, and the main Vibrio species was V. owensii, V. campbellii, V. parahaemolyticus, V. alginolyticus and V. harveyi; the activities of ACP, AKP, SOD, LZM and PO were 7.5—75, 1.0—8.5, 2.4—11.07, 1.3—43 and 23—28 U/mg, respectively. Combined the physical and chemical factors before and after the occurrence of AHPND, the results showed that, water temperature, culturable bacteria and Vibrio sp. in hepatopancreas, DO, LZM and PO could be used as the early warning candidate factors for the occurrence of AHPND in pond culture. The results provide a scientific basis for the healthy culture of L. vannamei and the prevention and control of AHPND disease.
TEMPERATURE ON BIOLOGICAL CHARACTERISTICS AND ADAPTIVE MECHANISMS OF THE APPLE SNAILS
YANG Qian-Qian, LI Jia-Nan, ZHAO Xing-Xing, PU Jia-Jia, QIAO Dong-Dong, YU Xiao-Ping
 Available online   , doi: 10.7541/2022.2021.0224
[Abstract](168) [FullText HTML](81) [PDF 752KB](2)
Abstract:
Apple snails from the family Ampullariidae are large freshwater gastropods that are native to South, North and Central America. Some ampullariids, mainly from the genus Pomacea have been introduced out of their native areas, and subsequently caused serious damage to the freshwater habitats in such as Asia. The strong resistance and rapid adaptation to ecological stressors play key roles for the rapid spread and outbreak of the Pomacea species in their non-native ranges. With the efforts from researchers both in China and abroad, the invasive mechanisms on the resistance to various ecological factors of apple snails have been extensively investigated. Among which, the mechanisms on tolerance to cold and hot temperatures have brought the most research interests. Environmental temperature is crucial for the survival, growth, development, and reproduction of apple snails that serves as one of the key factors affecting their geographic distribution. We reviewed the ranges of survival temperature of both Pomacea canaliculata (Lamarck, 1822) and Pomacea maculata Perry, 1810, because both the species were the most widely distributed and economically important ones. Based on which, we summarized the physiological, biochemical, and molecular mechanisms in adaptive to high and low temperatures of P. canaliculata and P. maculata. The physiological and biochemical mechanisms on adaption to temperature of apple snails have currently focused on changes of the chemical compounds and enzyme activities. While the molecular mechanisms have mainly focused on the expression differences of several heat shock proteins. The chromosome level genomes of P. canaliculata has been sequenced in 2018, which will promote the ecological adaptive mechanisms of apple snails with the application of advanced genetic manipulation techniques.
RESPONSE OF PERIPHYTON IN AQUACULTURE WATER TO SULFAMETHOXAZOLE STRESS AND ITS DEGRADATION
ZHANG Xin-Yi, MIN Fen-Li, ZHANG Shu-Xian, ZHANG Hao-Kung, LI Zhu-Xi, PENG Xue, WU Zhen-Bin, LIU Bi-Yun
 Available online   , doi: 10.7541/2023.2022.0171
[Abstract](152) [FullText HTML](100) [PDF 1263KB](1)
Abstract:
The extensive use and accumulation of sulfamethoxazole (SMX) in the aquaculture contributed to its residues in aquaculture water, which may cause various negative impacts on the aquatic ecological environment. Furthermore, periphyton is a common micro-ecosystem in water bodies composed of autotrophs and heterotrophs, including algae, fungi, bacteria, protozoa, metazoans, extracellular polymers (EPS) and debris, etc. Nevertheless, there is a lack of research on the impact of SMX on the growth and development of periphyton and its purification function in aquaculture water. In this survey, the effect of SMX on periphyton was researched in simulating aquaculture water, and the response of periphyton to SMX was explored through the biomass and total antioxidant capacity of the periphyton. The removal effect and the main degradation intermediates of SMX by periphyton were investigated by high-performance liquid chromatography (HPLC) and ultra-performance liquid chromatography quadrupole-time-of-flight mass spectrometry (UPLC-QTOF-MS). This research demonstrated that under 5 and 10 mg/L SMX, the secretion of extracellular polymer (EPS) of the periphyton increased by 31.3% and 21.5% compared with the control group, and the antioxidant capacity diminished by 20.6% and 31.8%, but there was no significant difference in biomass accumulation. And it showed that SMX decreased the total antioxidant capacity of the mature periphyton, their oxidative systems were damaged, and more extracellular polymers were secreted to enhance tolerance to SMX. The results of the removal experiment indicated that the periphyton would promote the reduction of SMX, and the removal rate of SMX at 1 g/L was 25.10% on the 14th day. It is supposed that SMX was used by periphyton as a carbon source to promote biodegradation and shorten the half-life of SMX degradation. Thus, periphyton had a certain potential for SMX bioremediation in aquaculture water. The main routes of SMX biodegradation by the periphyton are the cleavage and activation of the sulfonamide group (N-S bond) and the amine group (-NH2) on the benzene ring. The primary degradation intermediates were N4-acetyl-SMX, p-Benzoquinoneimine, Desulphone-SMX and Desamino-SMX. Among these degradation products, N4-acetyl-SMX, p-Benzoquinoneimide and Desamino-SMX were fewer toxic than SMX, and Desulphone-SMX was somewhat more toxic than SMX. The research provides theoretical support for assessing the ecological risk of SMX in aquaculture water and the engineering application of periphyton to remove SMX.
PURIFICATION PERFORMANCE OF RURAL LIVESTOCK AND POULTRY BREEDING TAIL WATER BY CONSTRUCTED WETLAND UNDER CU AND SMZ COMBINED POLLUTION
YANG Ling-Li, MA Lin, LIU Wei, CHU Yi-Fan, TAN Qi-Yang, ZHOU Qiao-Hong, WU Zhen-Bin, HE Feng
 Available online  
[Abstract](128) [FullText HTML](96) [PDF 2546KB](3)
Abstract:
Some heavy metals and antibiotics are widely used as feed additives in livestock breeding in rural areas. However, the bioavailability of the additives is quite low, while 40%~80% are discharged in the forms of prototypes or metabolites in feces, which causes a series of environmental pollution problems. Constructed wetlands (CWs) are sewage treatment technology with advantages of low costs, high efficiency and easy maintenance. Earlier studies have been reported that CWs have efficient removal effect on conventional pollutants such as N, P and COD, but there are few studies on the simultaneous removal of heavy metals and antibiotics in the tail water of livestock and poultry breeding in CWs. The influence and mechanism of combined pollution on the treatment effect of CWs is not clear yet. In this study, based on comprehensively considering the additives in aquaculture feed and their toxic effects, copper (Cu) and sulfamethoxazole (SMZ) were chosen as target pollutants. Agroup of five small-scale vertical flow constructed wetlands (VFCWs) were constructed, while different concentrations of Cu and SMZ wastewater were prepared to simulate different combined pollution levels (A: blank, B: 2 mg/L Cu, C: 2 mg/L Cu and 5mg/L SMZ, D: 20 mg/L Cu and 0.2 mg/L SMZ, E: 20 mg/L Cu and 5 mg/L SMZ). The effects of combined pollution Cu and SMZ on the purification capacity of wetland system were then explored. Results showed that VFCWs had high removal rate and relatively stable removal effect of total phosphorus (TP), chemical oxygen demands (COD) and nitrate nitrogen (NO3-N) under different combined pollution levels, while the average removal rate was higher than 99%. However, combined pollution showed inhibitory effect on the removal of NH3-N and TN. This effect became strong with the increased addition of the Cu and SMZ. The average NH3-N removal rate of 5 groups were 81.75%, 87.59%, 79.50%, 74.45%, respectively, and 65.41%, and the average TP removal rate of 5 groups were 81.59%, 87.63%, 80.82%, 74.15% and 67.56%, respectively. When Cu and SMZ were added at the same time, the dry weight of plant foot end were 22.22%, 30.58% and 30.64% lower than that of group A, respectively. Combined pollution of Cu and SMZ had a negative effect on canna as well as a selective effect on microorganisms. It implies that the combined pollution may affect the removal of NH3-N and TN by affecting plant functions and microbial community structure.
CORRELATION BETWEEN FIVE MICROSATELLITE MARKERS AND FOUR GROWTH TRAITS IN PTERIA PENGUIN
LIU Er-Tian, CHEN Yi, ZHAN Xin, GU Zhi-Feng, WANG Ai-Min
 Available online   , doi: 10.7541/2022.2021.0214
[Abstract](155) [FullText HTML](90) [PDF 895KB](1)
Abstract:
The winged pearl oyster, Pteria penguin, is a widely distributed large pearl oyster species, which can produce large-sized mabé (half-pearl), and exhibit strong tolerance to severe environment. However, genetic improvement of P. penguin was still in its infancy. In this study, the correlation between five microsatellite loci and four growth traits (shell height, shell length, shell width and total weight) of 500 winged pearl oysters were analyzed by One-way ANOVA. Multiple comparisons between different genotypes of microsatellite markers and growth traits were performed. It was shown that the 5 loci in this study are all highly polymorphic loci. Among them, QEB-D15 and CL-232 were extremely significantly associated with the shell width of the winged pearl oyster (P<0.01). The individuals with genotype 239/263 and 239/273 at QEB-D15had the maximum and the minimum shell width, respectively. The 263bp allele might positively correlate with the shell width, while the 273bp allele might negatively correlate with the shell width. The average values of shell length, shell width and total weight of individuals with genotype 157/174 at locus CL-232 were larger than that of other genotypes, suggested that this genotype was the dominant genotype. Individuals with genotype of 177/192 at locus CL-232had the smallest shell length, shell width, shell height and total weight, indicated that the 177/192 may be an inferior genotype. In addition, the shell width is correlated with two SSR loci, which indicates that the shell width is a phenotypic trait that may not be controlled by a single gene. This study could provide valuable information for molecular marker-assisted selection of P. penguin.
HYDROLOGICAL IMROVEMENT AND ENVIRONMENTAL EFFECT OF TRANSFORMING HYDROLOGICAL ELEMENTS OF THE ESTUARINE FLUCTUATION ZONE
SU Guang-Yu, XU Shi-Guo, LI Yi-Jian, XIE Chu-Yi
 Available online  
[Abstract](143) [FullText HTML](94) [PDF 2182KB](1)
Abstract:
Estuarine fluctuation zone is the key area for pollutant reduction in tributaries in reservoir. Affected by runoff scouring, river channel downcutting is a common problem in estuarine fluctuation zone, which makes the low purification efficiency of the runoff. Aimed at the weak ability of pollutant reduction of the estuarine fluctuation zone, the technology of improving purification efficiency by transforming hydrological elements with multi-stage submerged dam, which can form a surface flow constructed wetland, was proposed. The estuarine fluctuation zone of the Bajia River in the Biliuhe Reservoir was taken as a case study in this paper. By comparing the water quality and Normalized Difference Vegetation Index (NDVI), acquired through on-site monitoring and satellite imagery of Sentinel-2A and the situation before and after the construction, the influence of the project was analyzed. The water surface area and the water level as well as the hydraulic retention time in the estuary were all effectively improved. With the change of hydrological elements, the emergent plants grew well without human intervention, while the vegetation on the floodplain was significantly improved due to the increase of underground water level. Due to the influence of flooding, the pollutants on the land were released. The precipitation of the original pollutants in the flooded area has a significant impact on wetland water quality. The maximum removal rates of TN and NO3-N reached 72.61% and 75.95% in July 2020, while those in October 2019 were 22.06% and 27.95%, respectively. As a whole, the removal rates were good in surface flow constructed wetlands in China, while the improvement of TP and NH3-N removal efficiencies was not obvious, which may be caused by the precipitation of original pollutants. The hydrological elements transformation technology can effectively improve the purification capacity of the estuarine fluctuation zone and promote the reduction of pollutants in the basin.
ENHANCED NITROGEN REMOVAL OVER SATURATED WATER LEVEL REGIME IN VERTICAL FLOW CONSTRUCTED WETLAND
ZUO Shang-Wu, WANG Yue-Yuan, YU Shao-Le, LI Zi-Chen, WU Juan, CHENG Shui-Ping
 Available online  
[Abstract](157) [FullText HTML](98) [PDF 988KB](2)
Abstract:
The partially saturated vertical flow constructed wetland (VFCW) has been reported to exhibit better nitrogen removal performance than unsaturated VFCW when treating domestic wastewater. However, it is unclear how saturated water level affects the nitrogen transforming and its removal in VFCW. Herein, three VFCWs were operated under 90, 70 and 20 cm saturated water level regime, respectively. The physicochemical parameters of influent and effluent, i.e., chemical oxygen demand (CODCr), total nitrogen (TN), ammonia (NH4+-N), nitrate (NO3-N), nitrite (NO2-N), total phosphorus (TP), as well as the abundance of nitrogen transforming functional genes were investigated. The results showed that the purification performance of VFCW was improved by optimizing the distribution of unsaturated and saturated zones through saturated water level adjustment. The best nitrogen removal performance of 53.6% was obtained when the saturated water level height was 70 cm (media bed depth was 90 cm). The abundance of functional genes suggested that the unsaturated zone had no negative influence on denitrification community. The enriching nitrite oxidation bacteria in the unsaturated zone promoted the simultaneous nitrification and denitrification processes. However, too much low saturated water level (20 cm) might cause the unnecessary loss of carbon sources, resulting in the limited denitrification process and decreasing nitrogen removal efficiency. Hence, the saturated water level regime could be employed to improve the nitrogen removal performance in VFCW.
GLYCYRRHIZA URALENSIS ON ANTI-AEROMONAS HYDROPHILA INFECTION OF GIANT SALAMANDER (ANDRIAS DAVIDIANUS)
WANG Wen-Bo, CAO Juan, LI Chun-Ying, SUN Sheng-Yan, HUANG Yu-Ying, LIU Yi-Nan, LU Yi-Shan
 Available online   , doi: 10.7541/2023.2022.0198
[Abstract](124) [FullText HTML](96) [PDF 788KB](1)
Abstract:
Chinese giant salamander (Andrias davidianus) is a national second-class key protected species. Wild giant salamanders mostly inhabit in mountain streams. They like cave dwelling, afraid of light and sound, and feed on aquatic animals such as fish and crab. They are widely distributed in Qinling Mountains of Shaanxi Province, China. There are three species of giant salamander namely Chinese giant salamander (A. davidianus), American giant salamander (Cryptobranchus alleganiensis) and Japanese giant salamander (A. japonicus). The latter two species can not be bought or sold in the local areas, but can only be protected. Therefore, there is no artificial breeding industry in USA and Japan. In China, the second generation of artificially bred offspring of Chinese giant salamander is allowed to trade, which has a certain industrial scale. However, the occurrence of diseases in the breeding process has brought heavy economic losses to farmers. Among them, the septicemia of giant salamander caused by Aeromonas hydrophila is one of the common diseases. A. hydrophila is a common aquatic bacterium, which exists in natural water, but usually does not cause diseases. Only when the immunity of aquatic animals decreases, it is possible to infect the body through the digestive tract or skin wounds. Therefore, if it can enhance the activity of immune cells and immune molecules and improve its autoimmunity, it can effectively block the infection pathway of A. hydrophila and reduce the incidence of sepsis. Chinese herbal medicine is rich in resources, low cost and relatively environmentally friendly in China, and has been more and more used in aquaculture. In addition to Asian countries, European and American countries have also used herbal medicine to successfully control fish and shrimp diseases. However, in the above studies, most of the used methods of Chinese herbal medicine in aquaculture are direct feeding of crude drugs, and there is no extraction and concentration of immunoactive components. Reports from the poultry industry believe that licorice has immunomodulatory effect on the body, but there is no report on the effect of licorice on the anti-disease ability of amphibian giant salamander. This study carried out this exploration after targeted extraction of the effective components of licorice.The effect of licorice on the anti-A. hydrophila infection of giant salamander was studied by feeding licorice extract to giant salamander. The results showed that from day 28 after continuous feeding, the serum lysozyme activity in the drug group increased firstly and then decreased, showing a parabolic trend. The maximum value was (158.4±34.7) U/mL in the low-dose group on day 42 and (178.3±28.8) U/mL in the high-dose group on day 35, both of which were significantly higher than those in the control group in the same period (P<0.05). The phagocytic activity of renal macrophages in the drug group was significantly higher than that in the control group at the last two sampling periods, i.e. 42d and 56d (P<0.05). The maximum values of low dose group and high dose group appeared at 42 days, which were (59.4±8.5)% and (58.4±5.2)% respectively. Compared with the same period, the leukocyte volume of the drug group was higher than that of the control group. Among them, the leukocyte specific volume of the high-dose group was (5.8±1.7)%, the low-dose group was (5.5±0.8)%, and the high-dose group was (5.9±1.7)%, which were significantly higher than those of the control group at the same time (P<0.05). There was no significant difference in spleen organ coefficient between the drug group and the control group. After the last sampling (56d), the mortality of the control group was 90%, and that of the low-dose group and the high-dose group were 60%, which were lower than that of the control group, while the immune protection rate of the drug group was 33.3%, which was also higher than that of the control group. The results showed that feeding licorice extract could improve the resistance of giant salamander to A. hydrophila to a certain extent.Why can Chinese herbal medicine effectively inhibit bacterial growth? We know that protein is the material basis of life and participates in almost all life activities. The production of proteins is guided by genetic material DNA or RNA. They have certain stability. They control the life metabolic activities of the body by controlling the synthesis of proteins, including life processes such as growth, development, heredity and variation. Therefore, Chinese herbal medicine can indirectly affect the synthesis of protein by affecting the self replication of bacterial DNA and inhibiting its reproduction and growth, and finally inhibit the growth of bacteria. On the other hand, in terms of nonspecific immune defense function of the body, lysozyme, leukocytes and macrophages are extremely important immune molecules and immune cells. They can kill and effectively protect the body by dissolving or swallowing foreign pathogens. Then, in this experiment, licorice improved the survival rate of giant salamander infected by A. hydrophila. At the same time, the activity of serum lysozyme, the number of leukocytes and the phagocytic activity of macrophages increased. Is there a correlation between them? Is the improvement of the survival rate of giant salamander caused by the improvement of these indicators, or is it caused by the fact that Chinese herbal medicine affects the synthesis of bacterial DNA and thus the synthesis of bacterial protein? Further research is needed.
DISTRIBUTION CHARACTERISTICS OF MICROPLASTICS IN THE SEDIMENTARY ENVIRONMENT IN ADJACRNT SEA AREAS OF CHANGSHAN ARCHIPELAGO
SUN Xue-Mei, MENG Di, ZHENG Xin-Yan, CHEN Bi-Juan, QU Ke-Ming, XIA Bin
 Available online   , doi: 10.7541/2023.2022.0170
[Abstract](128) [FullText HTML](89) [PDF 1375KB](1)
Abstract:
As an emerging pollutant, microplastics have spread all over the marine environment. Studies have shown that the marine sedimentary environment is the sink for marine microplastics, The Changshan Archipelago (Changdao) are located at the throat of the Bohai Sea and belong to a typical island ecosystem. In recent years, the coastal area of ​​Long Island has been affected by human factors such as the construction of breeding ponds and port terminals, and the ecological environment is relatively fragile. In this study, the sediments of Changshan Archipelago were collected, and the microplastics in these sediments were analyzed by microscope observation and fourier transform infrared spectroscopy. The results showed that the abundance of microplastics in the Changshan Archipelago sediments ranged from 133.14 to 499.82 N/kg, with an average abundance of 252.59 N/kg. The size of microplastics is mainly <500 μm, accounting for more than 70% of the total number of microplastics; the shape of microplastics is mainly granular, followed by fragments, fibers and small spheres; the color of microplastics is mainly transparent. The polymer type of plastic is mainly cellophane, followed by polyethylene, polyethylene terephthalate, cellulose, etc. Because Changdao is located at the intersection of the Yellow Sea and the Bohai Sea, the overall hydrodynamic exchange capacity is well. Although the water mobility of the southern waters of Changdao is weaker than that of the northern waters, and the southern waters are closer to the rivers entering the sea, and are more affected by the input of land sources, the abundance of microplastics in the sediments of the southern water of Changdao is not significantly higher than that of the northern water. This study provides a scientific basis for the scientific assessment and management of microplastic pollution in the marine ecological environment of Long Island.
FISH BACKGROUND INVESTIGATION AND COMMUNITY STRUCTURE ANALYSIS OF FIVE BACKBONE ARTIFICIAL RIVERS IN SHANGHAI
CHEN Zhen-Feng, TANG Wen-Qiao, ZHAO Zhen-Guan, ZHANG Yan-Yan, GONG Long, TANG Zhen, ZHANG Ya, GUO Hong-Yi, LIU Dong, YANG Jin-Quan
 Available online   , doi: 10.7541/2023.2022.0164
[Abstract](114) [FullText HTML](87) [PDF 1292KB](1)
Abstract:
In this study, a total of 13379 fish specimens were collected from 20sections using gill nets in June (summer) and November (autumn), 2021 to reveal the fish species resources formed in five backbone artificial rivers of Shanghai Huangpu River in the past half-century. Sixty fish species were identified, belonging to 45 genera, 17 families, and 8 orders, comprising 41species of Cypriniformes belonging to 28 genera and 2 families, 8species of Perciformes belonging to 8 genera and 7 families, 2species of Migratory fish, 8species of estuarine fish, and freshwater species. There were 9 dominant species with IRI≥1000 in total, and Coilia nasus was the dominant species in all 5 rivers. ABC curve demonstrated that except for the Jinhui River, small and medium-sized fishes dominated other rivers, and fish communities were severely disturbed. ΒC index and ΒR index reflected that the Chuanyang River and the Dazhi River in Pudong had the highest difference in fish composition; the difference in fish composition between the Jinhui River and the Longquan River in Punan was the least. The 20sections were divided into three groups; group I ( D1 and Z2sections), group II (the Jinhui River, the Longquan River, and the Dazhi River except for the Z2section), and group III (the Chuanyang River and the Dianpu River except for the D1section). Pseudobrama simony, Coilia nasus, Carassius auratus, and Hemiculter leucisculus were the main diverging species causing the differences in fish community structure among groups. As suggested in the study, more fish species were preserved in these five artificial backbone tributaries than in the headstream, mainstream, and natural tributaries of the Huangpu River, and population density would be a crucial reason for the significant spatial differences in fish community structure in these rivers.
BIOLOGICAL CHARACTERISTICS OF CLUPANODON THRISSA IN PEARL RIVER ESTUARY
ZHANG Cui-Ping, YUAN Li-Mei, WU Yu-Xin, YE Zhi-Quan, CHEN Xiao-Ying, LAI Xing-Xing, LI Qiang, SHU Hu
 Available online   , doi: 10.7541/2023.2022.0036
[Abstract](118) [FullText HTML](91) [PDF 2464KB](2)
Abstract:
In order to understand the biological characteristics and resource status of Clupanodon thrissa in the Pearl River, a total of 408samples of C. thrissa were collected monthly from October 2020 to September 2021 in Guangzhou section of the Pearl River Estuary. The age composition, growth and reproductive characteristics of C. thrissa were analyzed according to the methods such as biological measurement, age structure analysis and histological observation. The results showed that the average standard length and body weight of females were (173.60±17.10) mm and (92.30±24.37) g, respectively. While the average standard length and body weight of males were (155.94±15.10) mm and (65.81±19.97) g, respectively. The population age was ranged from 0+ to 5+, and the dominant age was 1+—3+, accounting for 89.03 % of the total number of samples. There was power function relationship between standard length and body weight of C. thrissa: W=1×10–5L3.0525 (R2=0.9057), indicating that it was uniform growth pattern. Von Bertalanffy growth equation was used to describe the growth characteristics of C. thrissa with the growth parameters: L=176.14 mm, W=71.70 g, k=0.62, t0=−0.27, φ=4.28, and ti=1.53. The sex ratio (female/male) was 2.28﹕1, and the number of female individuals was significantly higher than that of male. The trend of gonadosomatic index and hepatosomatic index is opposite, indicating that the liver may provide energy for gonad development. It was speculated that the propagation period was from March to July. The absolute fecundity ranged from 1625 to 72882 eggs, with a mean of (20361±2601) eggs, and the relative fecundity ranged from 19 to 602 eggs/g, with a mean of (190±23) eggs/g. The distribution of egg diameter frequency was unimodal, which indicates it was one-time spawning fish. Compared with privious data, the fecundity of C. thrissa showed a decline tendency, so it is necessary to strengthen the protection and utilization of its resources.
EFFECTS OF DIFFERENT WATER FLOW RATES ON THE GROWTH PERFORMANCE, ANTIOXIDANT CAPACITY, ENERGY METABOLISM AND TISSUE STRUCTURE OF MICROPTERUS SALMOIDES UNDER AN IN-POND RECIRCULATING AQUACULTURE SYSTEM
LIU Mei, YUAN Ju-Lin, LIAN Qing-Ping, NI Meng, GU Zhi-Min
 Available online   , doi: 10.7541/2022.2021.0167
[Abstract](181) [FullText HTML](95) [PDF 0KB](4)
Abstract:
The aim of this study is to investigate the effects of different water velocities on the growth performance, antioxidant capacity, energy metabolism and tissue structure of Micropterus salmoides kept in an in-pond recirculation aquaculture (IPRA) system. 192 well-fed, healthy and uniformly sized largemouth bass [average weight: (8.13±0.82) g] were randomly assigned to 4simulation tanks with 3 duplicates containing 16 fish each. The experiment was set up with a control group (a stagnant water group) and three water flow rate groups, namely, a low flow rate group (0.2 m/s), a medium flow rate group (0.4 m/s) and a high flow rate group (0.6 m/s), respectively, with a culture period of 154 days. The results were as follows: (1) WGR and SGR of largemouth bass in the medium flow rate group were significantly higher than those of the other three groups (P<0.05), while the SGR of the high flow rate group was significantly lower than that of the other three treatment groups. The CF was significantly lower in the medium and high flow rate groups than that in the stagnant water and low flow rate groups. The visceral adiposity index (VAI) of largemouth bass decreased significantly with the increasing of flow rate (P<0.05). (2) The crude protein levels of muscle were significantly higher in the medium flow rate group than that in the other three groups. The crude fat content of muscle was significantly higher in the stagnant water and low flow rate groups than that in the medium and high low rate groups. The amount of crude fat in the specimens decreased significantly with an increased flow rate of water (P<0.05). (3) Serum triglycerides and the total amount of cholesterol in the specimens were significantly lower in the medium and high flow rate groups than that in the stagnant water and low flow rate groups (P<0.05). The concentration of free fatty acids and total amount of protein were significantly lower in the high flow rate group than that in the other three groups (P<0.05). (4) SOD and CAT increased with the increasing of the water flow rate, with the highest SOD level in the livers of the specimens in the medium flow rate group and there was a significantly higher concentration of serum CAT in the high flow rate group than the other three groups (P<0.05). Serum and liver tissue MDA levels were significantly lower in the medium flow rate group than that in the other three groups (P<0.05). The AOC of serum and liver tissues were significantly higher in the medium flow rate group than that in the other three groups (P<0.05). (5) LPS activity increased significantly in the specimens tested with an increased flow rate (P<0.05). AMS and PTS activities were the highest in the medium flow rate group. Hepatic LPS activity was the highest in the medium flow rate group, which was significantly higher than that in the stagnant water and low flow rate groups (P<0.05). PTS activity was significantly higher in the medium and high flow rate groups than that in the low flow rate groups. GDH increased significantly with the increased flow rate in both the muscles and liver (P<0.05). (6) The diameter of muscle fibres in the low flow rate group was significantly higher than that in the other three treatment groups (P<0.05). The density of muscle fibres increased significantly with an increase of water velocity. The medium flow group improved the tissue structure, while the high flow group caused some damage to their liver cells. In conclusion, the medium flow rate group with a flow rate of 0.4 m/s can promote the growth of largemouth bass, boost the immune systems, improve the functioning of their metabolisms, increase the crude protein content and inhibit fat deposition, and should be used as the optimal water flow rate for IPRA.
DIFFERENT CULTURE MODES ON GROWTH, SERUM BIOCHEMICAL INDEXES AND MUSCLE NUTRIENT OF YOUNG SALMO TRUTTA
WANG Wan-Liang, ZHANG Bian-Bian, TANG Da-Ming
 Available online   , doi: 10.7541/2023.2022.0120
[Abstract](148) [FullText HTML](100) [PDF 0KB](10)
Abstract:
To explore the diversified culture mode of Salmon trutta, a study was conducted to compare the growth performance, serum biochemical indexes, and muscle nutrient composition between recirculating water and open water culture modes with an initial body mass of juvenile S. trutta (100.05g±1.12 g) for 180 days. The results showed that weight gain rate, fatness, liver body index, specific growth rate, and feed conversion rate were significantly higher in open flow water mode than in recirculating water mode (P<0.05), but there was no significant difference in survival rate between them (P >0.05). Meanwhile, complement C4, total protein, and growth hormone were significantly lower in the circulating water mode than in the open running water mode (P<0.05), but the opposite was true for lysozyme, ghrelin, and glutamate (P<0.05), while complement C3, alkaline phosphatase, acid phosphatase, and globulin were not significantly different between the two modes (P>0.05). In addition, the crude protein and crude fat contents of the main muscle nutrients were higher in the open running water mode than in the circulating water mode, the amino acid composition was not significantly different between the two (P>0.05), and the fatty acid contents of oleic acid, α-linolenic acid, C20:1, C20:2, and MUFA were significantly lower than in the open running water (P<0.05), while the contents of EPA+DHA, SAFA, and PUFA were significantly different (P<0.05). Furthermore, mineral magnesium content was significantly higher in the recycled water model than in the open running water model (P<0.05), calcium content was the opposite (P<0.05), and the differences in zinc, iron, and selenium content were not significant (P>0.05). In conclusion, the circulating water culture mode of S. trutta in Tibet can make full use of the abundant local solar and geothermal resources or rely on the greenhouse for water temperature control, while further optimizing the micro-ecological purification capacity of the water body to make its growth conditions more similar to the open water culture environment, thus improving the efficiency of S. trutta culture and ensuring the nutritional quality.
SUSTAINED SWIMMING ON THE ENDURANCE TIME AND PHYSIOLOGICAL METABOLISM OF ACANTHOPAGRUS SCHLEGELI AND SCIAENOPS OCELLATUS
CHAI Ruo-Yu, YIN Heng, HUO Run-Ming, WANG Han-Ying, HUANG Ling, WANG Ping
 Available online   , doi: 10.7541/2023.2022.0186
[Abstract](139) [FullText HTML](96) [PDF 1232KB](2)
Abstract:
It has become possible to set up deep-water cages in the open ocean in response to the further development of the marine aquaculture industry. In these conditions, fish encounter strong currents and waves, and ensuring their wellness becomes a crucial part of the farming process, so studying the endurance of farmed fish species is necessary. However, there are few studies on the ability of fish to swim continuously in constant current mode. Breeding of black snapper and American redfish is currently expanding into more open waters, so its endurance swimming ability needs to be assessed to ensure farmed yields and fish farming welfare. At 20°C, we tested the endurance swimming ability of two fish with no significant differences in body length (P>0.05). First, determine the endurance swimming time at different flow rates, and then choose the speed when the endurance swimming time is 150min to carry out the endurance swimming experiment. Black snapper and American redfish were tested for 0, 30 minutes, 60 minutes, 90 minutes, 120 minutes, and 150 minutes at constant swimming speeds of 3.15 and 4.32 BL/s, respectively. Measure the concentrations of metabolites in the muscle, blood, and liver of the fish at six-time points, ensuring that each time point produces three sets of valid data. At 0 and 150 minutes, the concentrations of fish liver glycogen, back muscle lactic acid, and blood glucose were significantly different between the experimental groups (P<0.05), but the concentrations of muscle glycogen was not (P>0.05). A bivariate correlation analysis revealed that liver glycogen concentration decreased and dorsal muscle lactate and blood glucose increased with increasing fatigue. Gray-scale correlation analysis and principal component analysis showed that blood glucose and liver glycogen concentrations were the main factors affecting fatigue. However, the concentration range of black sea bream was larger than that of American redfish. Our experimental results concluded that: (1) American redfish have stronger swimming abilities than black snapper, and black snapper and American redfish are unsuitable for culture at flow rates exceeding 3.15 and 4.32 BL/s, respectively. (2) Liver glycogen concentration limits the endurance swimming ability of both fishes, and the results also provide a reference for the study on swimming and metabolism in other fishes.
FASTING AND REFEEDING ON THE EXPRESSION OF CHOLECYSTOKININ AND ITS RECEPTOR GENE IN LARGEMOUTH BASS
SHAO Jia-Qi, LI Sheng-Jie, DU Jin-Xing, DONG Chuan-Ju, LEI Cai-Xia, SONG Hong-Mei, JIANG Peng
 Available online   , doi: 10.7541/2023.2022.0175
[Abstract](168) [FullText HTML](125) [PDF 1069KB](2)
Abstract:
CCK as brain-gut peptide, binds to its receptor (CCKR) and participates in physiological processes such as feeding and digestion in animals. In order to study the function of CCKs and CCKRs in largemouth bass (Micropterus salmoides) in feeding activities, the coding sequences of CCK1, CCK2, CCK1R and CCK2R were cloned in this study. Their lengths were 414, 387, 1368 and 1359 bp, and their encodes were 137, 128, 455 and 452 amino acids, respectively. Fluorescence quantitative results showed that both CCK1 and CCK2 were highly expressed in brain, followed by intestinal, while CCK1R and CCK2R were highly expressed in gallbladder and brain, respectively. Within 24h after ingestion, the relative expressions of CCK1, CCK2, CCK1R and CCK2R increased first and then decreased. The relative expression of CCK1, CCK1R and CCK2R reached the highest value at 3h after ingestion, while the relative expression of CCK2 was at 12h after ingestion. The relative expression reached the highest value (P<0.05). During fasting, the relative expression of CCK1, CCK1R and CCK2R was significantly increased on the 14th day of fasting (P<0.05). The relative expression trends of CCK1, CCK1R and CCK2R after refeeding were similar to those after meals, showing a trend of first increasing and then decreasing. However, there was no significant change in the relative expression of CCK2 during fasting and refeeding. In conclusion, the results of this study indicated that CCK1 can be combined with CCK1R and CCK2R to act as a satiety signal factor to regulate physiological processes such as feeding and digestion of largemouth bass by suppressing appetite; and CCK2 may act as a short-term appetite factor to regulate feeding. Activity. The results of this study can provide a theoretical basis for the regulation of feeding activities of largemouth bass.
REMOVAL OF TYPICAL PERFLUOROALKYL SUBSTANCES BY MICROBIAL FUEL CELL COUPLING CONSTRUCTED WETLAND SYSTEM
JI Bin, ZHAOYA-QIAN, YANG Yang, TAI Yi-Ping, LI Qi-Wen, WEI Ting
 Available online   , doi: 10.7541/2023.2022.0114
[Abstract](168) [FullText HTML](152) [PDF 2374KB](3)
Abstract:
Per- and polyfluoroalkyl substances (PFASs) are widespread and persistent in aquatic environments. Their toxicity and bioaccumulation are the great concern among the scientific community and the public. Wastewater is regarded as a significant sink for PFASs. However, the effluents of wastewater treatment plants (WWTPs) can still cause aquatic problems due to the low removal efficiency of PFASs in the WWTPs. As an efficient and eco-friendly wastewater treatment and environmental cleanup technology, constructed wetland (CW) has been popularly used worldwide for treating various wastewaters. Several studies have investigated wetlands (natural or/and treatment wetlands) to remove PFASs from aqueous environments. The conventional CW system works on the principles of plant uptake, bioaccumulation, and substrate sorption to remove PFASs. The newly developed CW-microbial fuel technology (CW-MFC), as an intensified CW system, can simultaneously achieve enhanced wastewater treatment and bioenergy recovery and could open an integrated while eco-friendly way for PFASs removal. However, so far, no study has investigated the PFASs removal and their effects on the performance of the CW-MFC system. Therefore, the objective of this study was to explore the removal performance of PFASs by CW-MFC system under the coexistence of conventional contaminants in wastewater. Two CW-MFC systems were designed in closed- and/or open-circuit operating modes, respectively. Firstly, the removal performance of PFASs under different configurations was investigated and compared. The effects of PFASs on conventional contaminants' removal and bioelectrochemical performance of CW-MFC systems were then explored. When the concentrations of PFOA and PFOS in the influent were 6.46±0.52 μg/L and 9.34±0.87 μg/L, respectively, both closed-circuit and open-circuit operation of the CW-MFC systems demonstrated over 96% removal performance of PFAS. These results indicate that the CW-MFC system can effectively remove PFASs from wastewater, regardless of whether it is operated in an open or closed circuit. Moreover, the results showed that the removals of COD and TP in CW-MFC systems were less affected by PFASs under the influent COD and TP concentrations of 400 mg/L and 2.5 mg/L, respectively. However, with the efficient removal of PFASs, organic matter, and TP, some adverse effects on system performance were observed. For example, the NH4+-N and TN removal rates were decreased by approximately 7.22% and 2.45%, respectively, in the closed-circuit system, while those were sharply dropped by 13.51% and 13.98%, respectively, in the open-circuit system. Specifically, the average output voltage of the closed-circuit CW-MFC system decreased by 7.32% under the stress of PFASs. Anyhow, the outlook of CW-MFC treatment of PFASs shows promising from the aspects of the economy and potential degradation mechanism. Our study highlights that the CW-MFC system owns the potential to remove PFASs from the aqueous phase in an eco-friendly and cost-effective approach and provides important insights into the effects of the conventional contaminants' removal and bioenergy variations before and during the PFASs co-existence. Overall, this study provides useful information for PFASs remediation in the aquatic environment for further study.
GROWTH DIFFERENCES BETWEEN HYDRILLA VERTICILLATA AND MYRIOPHYLLUM SPICATUM AND THE EFFECTS ON WATER ENVIRONMENT IN NEWLY BUILT WATER SUPPLY LAKES IN CHANGSHU CITY
LI Shi-Qi, SHI Xue-Feng, LI Jun-Feng, MAI Zhan, GUO Chao, CHU Ji-Yang, DI Chun-Hua, LI Wei, LIU Jia-Shou
 Available online   , doi: 10.7541/2023.2022.0100
[Abstract](161) [FullText HTML](149) [PDF 1390KB](3)
Abstract:
In order to explore the effect of submerged plant community construction on the water environment of newly built water supply lakes, a comparative study was conducted on the growth of Hydrilla verticillata and Myriophyllum spicatum under artificial planting conditions and their effects on water quality and chlorophyll-a in the newly built emergency water supply lake from May to November 2019 in Changshu City, Jiangsu Province, China. The results showed that the two kinds of submerged plants could grow normally in the experimental lake, and their biomass showed an upward trend from May to September, but H. verticillata was in a rapid growth period from May to July, while M. spicatum was in a rapid growth period from July to September. The decay period of H. verticillata was earlier than that of M. spicatum, and the decay process was shorter. In November, H. verticillata withered completely, while M. spicatum was still in the middle stage of decay. The concentration of Chl.a can be effectively reduced during growth period of two submerged plants, inhibiting phytoplankton growth, while the results are opposite during the period of decline in Autumn. The comprehensive membership function value calculated by TP, TN, CODMn, Chl.a, DO and SD showed that the water quality of submerged plants group was better than that of the control group during the period of vigorous growth, while deteriorating during decline period. Pearson analysis showed that the biomass of H. verticillata had a significantly negative correlation with Chl.a, and the biomass of H. verticillata and M. spicatum had a significantly negative correlation with TP. This study can provide theoretical support for selecting, planting and harvesting of submerged plants during lake restoration in the middle and lower Yangtze River Basin.
CHARACTERISTICS OF PHYTOPLANKTON COMMUNITY STRUCTURE IN NEWLY-BUILT RESERVOIRS IN MOUNTAINOUS AREA GUIZHOU PROVINCE
ZHANG Chao-Shuo, LUO Bin, ZHOU Xian-Jun, LI Wei, LI Zhong-Jie, LIU Jia-Shou, YU Gong-Liang, DUAN Ming
 Available online   , doi: 10.7541/2023.2022.0108
[Abstract](182) [FullText HTML](158) [PDF 1141KB](6)
Abstract:
Phytoplankton, as an important primary producer in aquatic ecosystem, is very sensitive to the changes of water nutrient status and environment. At the initial stage of operation of the new reservoir, the changes of hydrological situation may improve the interception ability of the water body to exogenous pollutants, which easily leads to abnormal phytoplankton reproduction and even blooms. Therefore, the study on the characteristics of phytoplankton community during this period is helpful to understand the succession trend of phytoplankton community in the reservoir and provide reference for the management. Guanyinyan Reservoir and Wanying Reservoir in Liupanshui city, Guizhou Province, were newly built in 2018 and 2017, respectively. Based on field investigation, this paper studied the phytoplankton community structure and its relationship with environmental factors in the two reservoirs. A quarterly survey was conducted on both reservoirs, and the results showed that diatoms and green algae dominated the phytoplankton in both reservoirs. A total of 32species of phytoplankton were detected in Guanyinyan reservoir, among which diatoms and chlorophyta accounted for 31.3% and 28.1%, respectively. The average annual density of phytoplankton was 3.46×106 cells/L. The total species of phytoplankton was 28 in Wanying reservoir, diatoms and chlorophyta accounted for 35.7% and 39.3%, respectively. The average annual density of phytoplankton was 4.79×106 cells/L. According to the results of redundancy analysis (RDA), water temperature, total phosphorus and transparency were the main environmental factors affecting the changes of phytoplankton communities in the two reservoirs. In general, the newly built reservoirs in this study have a low risk of algal eruption in the early stage of operation. In subsequent management, attention should be paid to the influence of temperature, exogenous nutrition and seasonal changes on phytoplankton density.
TEMPORAL AND SPATIAL VARIATION CHARACTERISTICS AND DRIVING FACTORS OF BACTERIAL COMMUNITY STRUCTURE IN BAO’AN LAKE
XUE Lu, XIANG Dong-Fang, XIAN Bo, BAO Shao-Pan, TANG Wei, LUO Shuai, XIONG Jie, FANG Tao
 Available online   , doi: 10.7541/2023.2022.0112
[Abstract](153) [FullText HTML](170) [PDF 6600KB](6)
Abstract:
To explore the temporal and spatial variation characteristics and driving factors of bacterial community structure in Bao’an Lake, water samples were collected in spring, summer, autumn and winter of 2019 in Bao’an Lake, and the composition and diversity of bacterial community in surface water of Bao’an Lake were studied using metagenomic sequencing. The results showed that: (1) There was no significant difference in bacterial community structure among different lake areas. The Richness index, Pielou index, Shannon index and Simpson index in summer and autumn were significantly higher than those in spring and winter (P<0.05). The dominant phyla in summer and autumn were Proteobacteria, Actinobacteria and Cyanobacteria, while in spring and winter, the dominant plyla were Proteobacteria and Actinobacteria; (2) Temperature, transparency, pH, dissolved oxygen, CODMn, Chl.a, and total phosphorus were main driving factors accounting for the temporal and spatial of bacterial community structure in Bao’an Lake; (3) The assembly process of bacterial community in Bao’an Lake was dominated by stochastic process in spring, summer and autumn, while deterministic process dominated in winter; (4) The bacterial network interactions in Bao’an Lake had obvious seasonal characteristics: the bacterial interspecific interaction in Bao’an Lake was becoming closer and more complex from spring to winter. In conclusion, the seasonal variation characteristics of the bacterial community structure of Bao’an lake is obvious, and the factors such as temperature, transparency, pH, dissolved oxygen and CODMn have the potential to shape the bacterial community structure. This study provides the basis for understanding the spatial-temporal characteristics of bacterial community and driving factors in freshwater lakes.
NITROGEN REMOVAL EFFECT OF AQUACULTURE WATER THROUGH IMPROVED CONSTRUCTED WETLAND USING RECIRCULATING AQUACULTURE SYSTEM SOLID WASTE AS CARBON SOURCE
CAO Tao-Tao, XU Dong, BAI Guo-Liang, HU Ze, CHENG Di-Song, WU Zhen-Bin
 Available online   , doi: 10.7541/2023.2022.0190
[Abstract](173) [FullText HTML](158) [PDF 1288KB](2)
Abstract:
The recirculating aquaculture system has the problem of discharging high-nutrient aquaculture solid waste and high-concentration nitrate wastewater. For this reason, this study constructed an improved constructed wetland treatment device utilizing the aquaculture solid waste as the additional carbon, in order to strengthen its nitrogen removal effect of the recirculating aquaculture water. The wetland microbial community was analyzed by high-throughput sequencing. The results show that the increase in the amount of aquaculture solid waste would increase the N load in the wetland influent. When the influent TCOD/TN was 8.78±0.41, and the anaerobic zone depths were 30 cm and 50 cm, the improved constructed wetland system had better removal effect on TN, NO3-N and NH4+-N. At the phylum level, the dominant bacterial phylum in each wetland system was Proteobacteria, with a relative abundance of 44.44%—62.87% in A zone and 61.93%—78.64% in B zone. The abundance of denitrifying bacteria in the improved constructed wetland system was high, mainly Silanimonas (1.66%—23.42%) and Rivibacter (4.74%—12.75%), and the relative abundance of denitrifying bacteria in the wetland B zone was higher than that in the lower anaerobic A zone. In addition, an uncultured Anammox bacteria genus (uncultured_c_Anammox_3, 0.22%—0.33%) was also found in the A zone (30 cm and 50 cm). When the improved constructed wetland uses aquaculture solid waste as the carbon source, nitrogen removal was mainly through denitrification, and there was a certain anammox process.
VISUAL DETECTION OF POLY-L-LYSINE ON THE COVER GLASS
LIU Tao, ZHOU Fang, YE Ding
 Available online   , doi: 10.7541/2023.2022.0185
[Abstract](191) [FullText HTML](167) [PDF 1169KB](3)
Abstract:
Poly-L-Lysine (PLL) with a molecular weight of 150000-300000 is often used in cell culture and histology to promote cell or tissue adhesion. After treating the cover glass with 0.05—0.1% PLL solution, PLL adhered to the surface of the cover glass to form a monolayer. The density of the PLL directly affects the adhesion effect of the cover glass. However, because PLL is colorless and transparent, it is difficult to distinguish the PLL density and distribution on the surface of PLL-treated cover glass with the naked eye, so it is crucial to establish a simple and effective PLL detection method. However, there are still no relevant methods reported worldwide. In this study, a laboratory visual and quanitative detection method of PLL was developed. In this method, the fluorescein isothiocyanate (FITC) is added to the side chain amino group (-NH2) of the PLL on the surface of the cover glass in sodium bicarbonate solution, and fluorescence imaging is applied to detect the FITC-PLL.In this study, we firstly coated cover glass of 0.1% PLL of three different brands (S, M and A), and found that only the brand S PLL-coated cover glass still had intact tissue section after several washes with phosphate buffer. Accordingly, we found that the FITC-PLL of brand S on the cover glass was bright and uniform, while both the FITC-PLL of brand M and A were dim (3.4% for M, and 4.5% for A compared with S). Therefore, this method is suitable for the quality control of the homemade or commercial PLL-coated cover glass. Furthermore, the method also provides a reference for the establishment of a visual and quantitative detection method of material with a -NH2 group attached to glass or plastic surface.
PURIFICATION PERFORMANCE OF CONSTRUCTED WETLAND FOR LOW CONCENTRATION SEWAGE TREATMENT PLANT EFFLUENT
LI Qing, ZHANG Qiong-Hua, ZHOU Wei-Dong, ZHU Rui-Ting, DZAKPASU Mawuli, XIONG Jia-Qing, WANG Xiao-Chang
 Available online   , doi: 10.7541/2023.2022.0134
[Abstract](185) [FullText HTML](166) [PDF 1642KB](2)
Abstract:
Constructed wetlands (CWs) have many advantages including high impact resistance, outstanding ecological value and low cost. Consequently, it is gradually becoming more widely used and promoted in wastewater treatment plant (WWTP) effluent upgrading projects. Hydraulic load, water depth, and hydraulic residence are three important factors that influence the operational stability of CWs. When the hydraulic load increased suitably, the area of the system can be effectively reduced. Therefore, given the complexity of construction and land deprivation, the application of high-load constructed wetlands needs further research and exploration. In addition, most of the studies on constructed wetlands are based on small pilot tests. Analysis and investigation of the purification performance and operation mechanisms of full-scale systems are lacking. Furthermore, the lack of carbon in the WWTP effluents leads to poor denitrification in constructed wetlands, whereas the accumulation of biofilm in large wetland systems over a long period can easily cause the wetland system to clog. So, the use of constructed wetlands on a large scale in practical engineering is limited. In response to these problems, a constructed wetland system was designed, featuring a bi-directional cross-flow filtration unit, which used WWTP effluent as feed water. A study of the purification performance of the constructed wetland system for low concentration of the effluent showed a highly efficient performance. Analysis of the bi-directional cross-flow filtration unit planted with different plants (Canna indica, Thalia dealbata and Arundo donax var versicolor) showed that: (1) All three different types of planted wetland units had great water quality enhancement effects on the low load wastewater treatment plant effluent. Nevertheless, the different types of aquatic plants showed significant variability in the purification performance for COD, TN and NH4+-N. The average removal efficiencies of COD, TN, NH4+-N and TP in the planted unit with Arundo donax var versicolor were the highest, at 20.11%, 17.17%, 28.08% and 18.12%, respectively. (2) The bi-directional cross-flow filtration wetland allowed for the effective retention of carbon in the substrate, which was utilized to enhance denitrification by switching the inlet and outlet water flow directions. Consequently, the TN removal rate increased by approximately 50%. (3) From the phylum classification level, the main microbial communities of the wetland system did not show significant changes between the forward feed influent stage and the reverse feed influent stage. Nonetheless, the wetland system had a high proportion of Proteobacteria and Bacteroidetes (around 65.00%) at the influent end of the system. This phenomenon also explained the rapid transition of the wetland system after switching the intake direction, restoring its efficacy in treating pollutants. The findings can provide theoretical guidance and technical reference for the operation of large-scale constructed wetlands.
RESPONSE OF CYANOBACTERIA WITH DIFFERENT INORGANIC CARBON TRANSPORT GENOTYPES TO ENVIRONMENTAL CO2 CHANGES
ZENG Jia-Ying, JIANG Jing-Yu, ZUO Jun, LI Shu-Xin, ZHANG Si-Long, SONG Li-Rong, GAN Nan-Qin
 Available online   , doi: 10.7541/2023.2022.0078
[Abstract](175) [FullText HTML](170) [PDF 1371KB](3)
Abstract:
The continuous rise of atmospheric CO2 concentration will alter the physiological and ecological response in cyanobacteria. The CO2 concentration mechanism (CCM) of cyanobacteria is one of the important competitive advantages in the formation of water bloom. Different transport systems in the CCM of cyanobacteria have different characteristics. For example, bicA is a bicarbonate transporter with high throughput and low affinity, while sbtA with high affinity but low throughput. In order to explore the relationship between different inorganic carbon (Ci) transport genotypes of cyanobacteria and pH changes in lake water, this paper optimized the detection protocol of the relative abundance of cyanobacteria with different inorganic carbon transport genotypes in water. We measured the relative abundance of various cyanobacteria species with different Ci transport genotypes in Taihu Lake, Dianchi Lake and 18 lakes in Wuhan, and combined with the pH value in the water body to analyze the response of different Microcystis genotypes to CO2 changes. It was showed that bicA strains, sbtA strains and bicA+sbtA strains all exist simultaneously in the sampled lakes, and the sbtA strains was the most widely distributed. With the increase of pH in water, the dominance of sbtA strains increased. In order to further analyze the response mechanism of different Ci transport genotypes of cyanobacteria to the change of CO2 concentration, we studied the competition of bicA strains, sbtA strains and bicA+sbtA strains under high concentration (1000 ppm) and low concentration (100 ppm) carbon dioxide respectively. The results showed that sbtA strains had obvious competitive advantage at low Ci level, while bicA strains occupied a dominant position at high Ci level. Our study showed that with the increase of CO2 concentration, bicA strains in cyanobacteria bloom would have a competitive advantage. Therefore, we predict that the increase of atmospheric CO2 concentration will affect the community composition of bloom cyanobacteria.
TETRAPLOID INDUCTION OF THE PACIFIC OYSTER “HAIDA NO. 3” USING CYTOCHALASIN B AND LOW SALINITY
ZHOU Jian-Min, JIANG Gao-Wei, XU Cheng-Xun, LI Yong-Guo, LI Qi
 Available online   , doi: 10.7541/2023.2022.0044
[Abstract](222) [FullText HTML](230) [PDF 1060KB](3)
Abstract:
To investigate the optimal conditions for the tetraploid induction of the Pacific oyster Crassostrea gigas “Haida No. 3” by cytochalasin B (CB) and low salinity, the effects of CB concentration (0.2, 0.4, 0.6 and 0.8 mg/L), low salinity (6, 8, 10 and 12) and induction duration (10, 15, 20 and 25min) on the cleavage rate, hatching rate, tetraploid rate and the efficiency of tetraploid induction were estimated by inhibiting the first polar body of fertilized eggs. At the same time, the growth and survival of the larvae were analyzed. The results showed that the maximum point of tetraploid rate (65.69±2.47)% and the efficiency of tetraploid induction was found at CB concentration of 0.6 mg/L and induction duration of 15min. In low-salt induction, the maximum point of tetraploid rate (38.77±2.69)% and the efficiency of tetraploid induction was found at salinity of 8 and induction duration of 15min. The shell height of CB and low-salt treatment groups were larger at the early stage and smaller at the later stage compared with the control group. The shell height of the CB treatment group was significantly greater than the low-salt treatment group (P<0.05), and the mean daily growth of larvae in the CB and the low-salt treatment groups was (14.2±1.08 μm/d) and (10.49±0.60 μm/d), respectively, which were smaller than the control group (15.43±1.08 μm/d). The survival rate of the two induction treatment groups was consistently lower than the control group, and the survival rate of the low-salt treatment group was higher at the early stage and lower at the late stage compared with the CB treatment group. In general, the CB induction method showed better results in terms of tetraploid rate, the efficiency of tetraploid induction, 12-day survival rate and growth rate, and has better applicability for the tetraploid induction of the Pacific oyster “Haida No.3”.
PATERNITY TEST TECHNOLOGY BASED ON MICROSATELLITE MARKER ISLET
XIE Min-Min, WANG Ya-Kun, WEI Cheng-Qing, CHEN Chen, LIU Xiao-Li, LI Wei, HONG Xiao-You, ZHU Xin-Ping
 Available online   , doi: 10.7541/2023.2022.0051
[Abstract](257) [FullText HTML](230) [PDF 853KB](2)
Abstract:
The Asian giant soft-shelled turtles (Pelochelys cantorii) belong to Testudines, Trionychidae, and Pelochelys. As one of the largest aquatic turtles in China, it is also an important indicator species for the health of the river ecosystem in the Pearl River Basin and even the southern part of China. Because of the long history of P. cantorii, it is of great scientific value in the study of environmental changes and biological evolution. P. cantorii was widely distributed in Southeast Asia of China in history. However, due to unfair trade, excessive harvesting and killing of turtles and water conservancy projects, the population of P. cantorii has been greatly reduced. Currently, P. cantorii have become an extremely endangered species. There are only 13 P. cantorii in captivity in China, which have been listed as national first-class protected animals and taken seriously by scientific researchers. With the success of artificial domestication and breeding in recent 3 years, the genetic information of the preserved population is still unknown. In this paper, we use transcriptome data design trinucleotide repeat and repeat four nucleotides microsatellite primers 30000 on, the choice of 230 pairs of primer synthesis, received 10 after two screening amplification with high efficiency and high specificity of microsatellite marker, and build 2 multiple PCR system. Genotypes and genetic diversity of 56soft-shelled islet were detected by ABI3130 genetic analyzer and PopGene3.2 and CERVUS software. The results showed that the average number of alleles was 2.3 and the average content of polymorphic information was 0.3829. The average observed and expected heterozygosity were 0.6305 and 0.4767, respectively. Paternity analysis of 56 progenies showed that the cumulative exclusion probabilities of 10 microsatellite loci were 73.14% (NE-1P), 90.98% (NE-2P) and 98% (NE-3p), Which can meet the needs of paternity testing. Respectively, and the four parents showed differences in reproductive selection. The analysis of the technology can help identify the captive population of genetic information, assist researchers to family management of groups. And more importantly, in the process of F3 generations of breeding in the future, the technology can be regarded as parental choice of technical means, to get healthier, higher genetic diversity, and their offspring, for the entire group of proliferation and endangered wild work to lay a solid foundation.
INDUCTION OF XX PSEUDO-MALES, AND SUCCESSFUL REPRODUCTION AND CULTIVATION OF ALL-FEMALE POPULATION IN YELLOW CATFISH
LIU Ya, YU Yue, LU Zi-Yi, DING Yang, FAN Qi-Xue, SHEN Zhi-Gang
 Available online  
[Abstract](673) [FullText HTML](242) [PDF 4628KB](18)
Abstract:
The shortage of female has impeded the development of aquaculture industry of yellow catfish (Tachysurus fulvidraco) in China in recent years. Therefore, large-scale production system of all-female is urgently needed. To produce an all-female population in large-scale of yellow catfish, 12-65 days post-hatching larvae were treated with the combination of three male-promoting factors, 17 α-Methyltestosterone (MT, 5mg/kg), Letrozole (LZ, 300 mg/kg), and high temperature (33.5℃). XX pseudo-males were mated with normal XX females and large-scale production of all-female population was conducted. Results showed that the combination of MT, LZ, and high temperature-induced masculinization of XX individuals, and the completely sex-reversed XX males displayed a similar percentage of motile sperm with XY males. Histological observation found that there are a large number of spermatids in the testis of XX males, indicating normal reproductive ability. Subsequently, we mate XX males and XX females, produced 570000 genotypic XX larvae, and successfully reared them to large fingerlings in a pond. The results indicated that 2.8% and 12.0% XX individuals reversed to XX males which is probably due to the high temperature during the critical time of sex differentiation in the rearing pond. The ovary developed very well for the rest of XX females and can be used as female parents in the coming spawning season. The current work successfully conducted large-scale production of the all-female population in yellow catfish, and the results will provide an important foundation for the large-scale production system of the all-female population, and important safeguard for female selection in genetic breeding of new varieties.
MAJOR PREDATORS AND PREDATION PRESSURE OF FIVE KEYSTONE PREY SPECIES IN HAIZHOU BAY
GAO Ming-Wei, LIU Shu-De, XU Cong-Jun, XU Bin-Duo, ZHANG Chong-Liang, JI Yu-Peng, REN Yi-Ping, XUE Ying
 Available online   , doi: 10.7541/2023.2022.0020
[Abstract](307) [FullText HTML](297) [PDF 1726KB](2)
Abstract:
This study aims to understand the fluctuation of predation pressure and natural mortality of keystone prey species in the food web of Haizhou Bay. Keystone prey species play a crucial role in the energy flow and material transference in the marine food webs. Researches on the importance of keystone prey species in the food web can provide a scientific basis for the restoration of ecosystem and conservation of fishery resources. Based on the bottom trawl survey data in Haizhou Bay and stomach contents analysis, the predation pressure index (PPI) of five keystone prey species (including Leptochela gracilis, Alpheus japonicus, Loligo sp., Larimichthys polyactis and Oratosquilla oratoria) were used to analyze the major predators of these prey species and their predation pressure. Results showed that the PPI of Chelidonichthys kumu to L. gracilis and Loligo sp. was the highest, being 168.89 and 75.77, respectively. The PPI of Miichthys miiuy to A. japonicus and O. oratoria was the highest, being 39.41 and 9.85, respectively. The PPI of Saurida elongata to L. polyactis was the highest (109.65). Anova showed that there was no significant difference in PPI in different years (P>0.05), and there were significant differences among predators (P<0.05). The natural mortality coefficient varied with the fluctuation of predation pressure index. There were differences in the feeding habits of these predators, which will help to reduce the food competition among them and benefit the ecosystem stability. Trophic interactions between species are instrumental in maintaining the stability of ecosystems. These keystone prey species play a vital part in the food web and can directly affect the structure and function of ecosystems, providing energy for the high trophic level predators and contributing to the energy flow and material circulation in the food web. Based on the findings of the present study, It is suggested to strengthen the research in the field of keystone prey species, explore the relationship between species, and analyze in-depth the ecological function and regulation mechanism of keystone prey species. This is pivotal in maintaining ecosystem stability and species diversity and developing ecosystem-based fishery management strategies.
SPECIES DIVERSITY AND ASSEMBLAGE STRUCTURE OF FISHES BY-CATCH IN ELVER NETS IN YANGTZE RIVER ESTUARY
DENG Yan, TANG Wen-Qiao, ZHANG Yuan-Yuan, ZHAO Zhen-Guan
 Available online   , doi: 10.7541/2023.2022.0016
[Abstract](364) [FullText HTML](277) [PDF 1282KB](3)
Abstract:
The Yangtze River Estuary is the major production regions of larval eels in China, and it is also the most important place for feeding and nursing place for fishes. In order to clarify the species diversity and damage rate of fishes by-catch in elver nets in the Yangtze Estuary, this article analyzed 11870 fish samples collected by elver nets during the eel fry flood season from 2017 to 2019. The results showed that 43samples contained 114388 fishes by-catch in elver nets, 54species were identified, belonging to 13 orders, 27 families and 46 genera. Among them, Perciformes accounted for 40.74% of the total species, and marine, carnivorous, demersal and small fishes accounted for 44.44%, 74.07%, 70.37% and 48.15%, respectively. Odontamblyopus lacepedii was the first dominant species, accounting for 77.07%; Cynoglossus joyneri, Coilia nasus and Konosirus punctatus were also the dominant species in individual years. According to the difference of fish assemblage, March and April, January and February can each be combined into one group. The major characteristic species of two groups were 5 and 6, and the major divergent species were 10. The average body length was (73.77±38) mm of 54species. Fish resources of the early life history stages account for 96.65% of the total samples, many of which are larvae of local economic fishes. There were 16470 eel elvers in the sample, and the average damage rate of fishes by-catch in 3 years was 7.58, which was higher than 6 years ago. Attention should be paid to the damage caused by fishing eel elvers to other fishes’ larvae.
MORPHOLOGICALCHARACTERS OF FIVE HYPOTRICHOUS CILIATES FROM THE YANGTZE ESTUARY AREA
ZHANG Zi-Hui, ZHU Xiao-Xuan, JIANG Jia-Mei
 Available online   , doi: 10.7541/2022.2021.0174
[Abstract](366) [FullText HTML](269) [PDF 2881KB](2)
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Hypotrichs, the most complex and evolved ciliate group, are widely found in various habitats. The morphological studies on the ciliates have mainly focused on freshwater of limited provinces, the Yellow and Bohai Seas and the South China Sea coast, and few reports are available from the Yangtze Estuary area. The morphologies of five hypotrichous ciliates isolated from Yangtze River area, Bakuella incheonensis, Histriculus histrio, Pseudokeronopsis rubra, Pseudokeronopsis flava, and Pseudourostyla cristata, were investigated using living observation and protargol staining methods. Bakuella incheonensis and H. histrio were new record species in China. Detailed redescriptions including the living characteristics and the infraciliature were supplemented. Results showed that the Jiangsu population B. (B.) incheonensis had a bigger size, more adoral membranelles and frontoterminal cirri than the original Korean population; The Shanghai population H. histrio differed from some foreign populations in body length; The Jiangsu population P. rubra presented a larger size range than the Qingdao population; The Shanghai population P. flava had more frontal cirri compared with the Zhanjiang population; The Shanghai population P. cristata owned more macronuclear nodules than the Japanese population. This work enriches our understanding of the hypotrichs diversity in China.
COMPARATIVE STUDY ON REPRODUCTIVE BIOLOGY OF TWO CO-INVASION TILAPIA SPECIES IN THE SHANMEI RESERVOIR
LI Jun-Feng, WANG Jiang-Bin, LIU Zhi-Yang, LIU Qiao, GUO Chao, LIAO Chuan-Song, LI Wei, GUO Chuan-Bo, ZHANG Tang-Lin, LIU Jia-Shou
 Available online   , doi: 10.7541/2023.2022.0089
[Abstract](350) [FullText HTML](266) [PDF 4055KB](5)
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The redbelly tilapia Coptodon zillii and the mango tilapia Sarotherodon galilaeus are two invasive fish species in the Shanmei Reservoir, which have become dominant species of the fish community in the reservoir. In order to compare their reproductive traits for developing invasive control measures for the two tilapia species, we monthly sampled fishes from March to October 2021 in the Shanmei Reservoir, and analyzed their reproductive time, sex ratio, absolute fecundity, relative fecundity and egg size. We also analyzed the reasons for their stable coexistence after co-invasion through the reproductive biology of two species of tilapia. Results showed that both populations started to spawn in April, but the spawning activity was peaked at July and June, and ended at October and September for C. zillii and S. galilaeus, respectively. The absolute fecundity of C. zillii was (4009.85±1305.69) eggs, the body weight relative fecundity was (67.32±15.63) eggs/g, and the body length relative fecundity was (31.31±5.03) eggs/mm for C. zillii, which were significantly higher than those of S. galilaeus [(1701.85±591.29) eggs, (6.46±0.87) eggs/g, and (8.22±2.33) eggs/mm]. However, the mature egg size of C. zillii was significantly smaller than that of S. galilaeus. The sex ratios (female/male) of C. zillii (1.59) and S. galilaeus (1.83) showed non-significantly difference and females outnumbered males in these two species but both ratios were significantly biased with 1﹕1. This study suggested that the two co-invasive tilapia exhibit different reproductive strategies, which should be key factors explaining their dominant abundance and stable coexistence after successful co-invasion. This study can provide implications for the management of the two co-invasive tilapia populations, which contribute to the sustainable development of fisheries resources management in the Shanmei Reservoir.
HIGH DIETARY DOSES OF ORGANIC AND INORGANIC SELENIUM ON GROWTH PERFORMANCE, TISSUE SELENIUM ACCUMULATION AND BLOOD BIOCHEMICAL PARAMETERS OF GIBEL CARP (CARASSIUS AURATUS GIBELIO VAR. CAS V)
LI Yuan-Ze, LIU Hao-Kun, GONG Yu-Long, GAO Wei-Ye, ZHU Xiao-Ming, HAN Dong, YANG Yun-Xia, JIN Jun-Yan, XIE Shou-Qi,
 Available online  
[Abstract](397) [FullText HTML](279) [PDF 1075KB](6)
Abstract:
To investigate the effects of high doses of organic and inorganic selenium in feed on the growth performance, selenium accumulation and plasma biochemical parameters of gibel carp, organic selenium and inorganic selenium were used as different types of selenium sources, and gibel carp with an initial body weight of 62.95±0.23 g was used for a 90d feeding experiment. The results indicated that the addition of 0, 10 and 20 mg/kg organic and inorganic selenium to the feed did not have a significant effect on the survival rate and the digestibility of the dry matter. Moreover, the addition of organic selenium significantly increased the selenium digestibility of the organic selenium treatment group (P<0.05), while the addition of inorganic selenium to the feed had no significant effect on the selenium digestibility (P>0.05). The addition of organic selenium to the feed increased the growth of gibel carp (P<0.05), reaching the highest level in the 20 mg/kg treatment group (P<0.05). The addition of inorganic selenium to the feed significantly reduced the specific growth rate of the gibel carp (P<0.05), and there was no significant difference in the specific growth rate between the 10 mg/kg inorganic selenium treatment group and the control group (P>0.05). The addition of organic selenium to the feed did not have any significant effect on the hepatosomatic index of gibel carp, while the addition of 10 mg/kg inorganic selenium significantly reduced the hepatosomatic index (P<0.05). The addition of organic and inorganic selenium to the feed did not have any significant effect on the renosomatic index of gibel carp. The addition of both organic and inorganic selenium significantly increased the selenium content of whole fish, liver, kidney, muscle, bone and gonads, and the organic selenium addition group had significantly higher selenium content in bone, muscle, gonads and whole fish than the inorganic selenium addition group (P<0.05). The addition of organic selenium and inorganic selenium to the diet significantly increased the plasma glucose content, decreased the total plasma bilirubin content and reduced the activity of glutamate and ghrelin (P<0.05), and the addition of 20 mg/kg organic selenium and 10 mg/kg inorganic selenium to the diet significantly reduced the plasma iron ion content (P<0.05). The results showed that the high levels of organic and inorganic selenium in the feed were well tolerated by the gibel carp and that the high levels of organic selenium in the feed had no toxic effect on the gibel carp after 90 d of the feeding trial, which showed a boost in growth performance and a high level of bioaccumulation in various tissues.
THE INFLUENCE OF STOCKING DENSITY ON THE GROWTH, BLOOD BIOCHEMISTRY, AND ANTIOXIDANT CAPACITY INDICATORS OF CHINESE PERCH (SINIPERCA CHUATSI)
LU Ke, ZHANG Yan-Peng, LIANG Xu-Fang, TANG Shu-Lin
 Available online   , doi: 10.7541/2022.2021.0395
[Abstract](952) [FullText HTML](268) [PDF 967KB](4)
Abstract:
The present study was to explore the influence of stocking density on the growth, blood biochemistry, and antioxidant capacity indicators of Chinese perch (Siniperca chuatsi) cultured in the bank (60×60×40 cm) based recirculating systems for 40d. Fish (average weight 175.76±15.85 g) were reared in triplicate under five densities: extra-low stocking density (ELD; 3.34 kg/m3); low stocking density (LD; 9.51 kg/m3); medium stocking density (MD; 15.82 kg/m3); high stocking density (HD; 21.05 kg/m3) and extra-high stocking density (EHD; 25.99 kg/m3). After 40 days, growth performance, biochemical parameters, appetite-related genes expression and stress indicators of fish were evaluated. The results showed that the ELD and LD groups showed better productive performance for weight gain (WG) and feed conversion ratio (FCR) (P<0.05), and increased plasma superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) activities, and decreased the formation of malondialdehyde (MDA) (P<0.05). Additionally, the EHD group performed higher glucose levels, cholesterol levels and lower triglyceride levels in plasma (P<0.05). The mRNA expression of agouti gene-related protein (agrp) was riesd in LD group, whereas gene expressions of pro-opiomelanocortin (pomc) resulted in a down-regulation in ELD group. In sum, the suitable stocking density for culturing Chinese perch in recirculating systems is recommended to be between 3.34 and 9.51 kg/m3. It is of great significance to select the best stocking density for improving the economic benefit of fish culture.
IDENTIFICATION OF HC-GSK3Β GENE AND SCREENING OF SNPS RELATED TO INNER SHELL COLOR OF HYRIOPSIS CUMINGII
GUO Bai-Ying, LI Xing, WANG He, YAN Ling, LU Gao-Lun, BAI Zhi-Yi
 Available online   , doi: 10.7541/2023.2022.0107
[Abstract](298) [FullText HTML](267) [PDF 1708KB](1)
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Melanin is an important factor for the formation of purple shells and pearls of Hyriopsis cumingii. Glycogen synthesis kinase-3β (GSK3β) is a key gene of animal melanin synthesis pathway. In order to explore the effect of Hc-GSK3β gene on shell color of H. cumingii, the full-length cDNA of Hc-GSK3β gene of 1867 bp was successfully cloned by RACE technology, including the ORF region of 1261 bp encoding 420 amino acids. The ORF contained a S_TKc domain, whose sequence is highly conserved. Tissue expression(qRT-PCR)analysis showed that the expression of Hc-GSK3β gene in purple mussel was significantly higher than that of white mussel in gill, foot, hepatopancreas and fringe mantle tissues (P<0.05), and the expression existed extremely significant difference in foot and fringe mantle tissues (P<0.01). The expression in the adductor muscle tissue of purple mussel was significantly lower than that of white mussel (P<0.05). The results of in situ hybridization (ISH) showed that positive signals appeared in the outer fold, middle fold, inner fold, dorsal mantle area and ventral mantle area of H. cumingii, and the signal expression in the outer fold was stronger. 6 SNPs locus were identified by re-sequencing and comparison. The distribution frequencyof genotype CA at C+185A locus was significantly higher in purple mussels than that of white mussels (P<0.05). At the T+341G locus, the value of color parameter b of the genotype TT was significantly lower than genotype TG (P<0.05). The study showed that Hc-GSK3β gene was involved in shell color formation of H. cumingii, and SNP markers could be used for the shell color breeding of H. cumingii.
A PILOT STUDY ON SPECIES IDENTIFICATION OF BRYDE’S WHALES INHABITING IN THE WEIZHOU ISLAND WATERS, BEIBU GULF
HE Rui-Lin, CHEN Mo, ZHANG Yao-Yao, MEI Zhi-Gang, FAN Fei, WANG Ding, ZHENG Jin-Song
 Available online   , doi: 10.7541/2023.2022.0105
[Abstract](329) [FullText HTML](266) [PDF 1337KB](3)
Abstract:
The Bryde’s whale is a medium sized baleen whale that is widely distributed in the tropical and subtropical waters. However, the taxonomy of the Bryde’s whale has long been controversial. Currently, both the IUCN (the International Union for Conservation of Nature and Natural Resources), and the IWC (the International Whaling Commission) assign the whale as two subspecies, Balaenoptera edeni edeni and B. e. brydei. Although, several studies indicate that they should be regarded as two separate species, B. e. edeni and B. e. brydei are so similar in their external morphology, and are sometimes sympatric, which make it is difficult to correctly identify them in the field just through observation. Therefore, accurate species delimitation usually requires molecular methods. Recently, a Bryde’s whale population was reported inhabiting in the Weizhou Island waters, Beibu Gulf. However, it is unclear which species these animals belong to. In this study, we collected two fecal samples from two individuals in this population, and tried to identify them using molecular methods. Genomic DNA was extracted from one of the two fecal samples. Fragments of the mitochondrial genes Cyt b and COⅠ were PCR amplified and sequenced. Based on the mitochondrial gene sequences, the whale was identified as B. e. edeni. In addition, a baleen whale that died in 2019 in the same waters was also identified as B. e. edeni using the same methods outline in this study. Thus, we infer that the population living in the Weizhou Island waters is B. e. edeni. This is the first study which successfully identified a living Bryde’s whale based on mitochondrial DNA present in fecal samples. This non-invasive sampling-based species identification method should be further developed and applied in the future. Additional studies, such as population genetics should be carried out to provide a scientific basis for the conservation of this Bryde’s whale population.
HARVESTING HAEMATOCOCCUS PLUVIALIS USING PH-INDUCED DISSOLVED AIR FLOTATION (DAF)
LI Jing, XU Cong, LIU Qing-Lin, LI Cheng, ZHANG Hang-Yang, ZHANG Xue-Zhi
 Available online   , doi: 10.7541/2023.2022.0076
[Abstract](282) [FullText HTML](263) [PDF 1258KB](1)
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As an important raw material for the extraction of natural astaxanthin, Haematococcus pluvialis is usually harvested by natural sedimentation or centrifugation. However, these harvesting methods either take a long time or consume a lot of energy. This study proposed an efficient method for the harvesting H. pluvialis using pH-induced induced dissolved air flotation (DAF). Harvesting efficiency and solid content, as well as their kinetic curves, were adopted to investigate the influence of pH on the pH-induced DAF harvesting. In addition, the harvesting performances of pH-induced DAF was compared with those of pH-induced sedimentation, chemical coagulant (aluminum sulfate)- and biological coagulant (chitosan)-induced DAF, based on the efficiency and residual metal contents in the harvested biomass. Whether pH treatment affects the astaxanthin content in the harvested biomass was also evaluated, comparing with the content in centrifugation harvested biomass. The results showed that auto-flocculation occurred when the pH of the media was decreased or increased, facilitating the flotation harvesting and sedimentation harvesting of H. pluvialis. pH-induced DAF obtained higher harvesting efficiency and required less separation time, comparing with pH-induced sedimentation. When pH was adjusted to less than 3 or greater than 11.5, the harvesting efficiency of 95% was achieved in 2 min using pH-induced DAF, while that took 30 min for pH-induced sedimentation to achieve 80%—90% harvesting efficiency. When the initial concentration was 3.2 g/L, the solid content of biomass harvested by pH-induced DAF can reach 17%, realizing 53 times concentration, which was much higher than those harvested by pH-induced sedimentation. Most importantly, pH-induced DAF performed as well as aluminum sulfate and chitosan coagulants induced DAF, while avoiding the impurity of biomass caused by chemical such as Al3+. The astaxanthin contents in H. pluvialis biomass harvested by pH-induced DAF were not affected by the pH treatment. Therefore, harvesting H. pluvialis by pH-induced DAF proved to be a rapid, efficient and environmental sustainable technique, which shed a light on the process development of mass production of H. pluvialis biomass.
INFLUENCE OF ASSISTED FILTRATION PRETREATMENT ON MEMBRANE FOULING DURING THE TREATMENT OF ALGAE-RICH WATER BY MICROFILTRATION
ZHANG Shi, MA Fang-kai, XU Wang-peng, WANG Jian, LI Ying-xi, WU Zhen-bin, ZHANG Xue-zhi
 Available online   , doi: 10.7541/2023.2022.0056
[Abstract](386) [FullText HTML](263) [PDF 2294KB](2)
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Membrane filtration technology has the advantages of well algae-water separation and sewage purification during the treatment of algae-rich water. Different pretreatment methods can affect the filtration characteristics and membrane fouling. In this study, three different pretreatment methods of diatomite, cotton plant and activated carbon were employed to filter Microcystis aeruginosa suspensions to compare their filtration characteristics, retention of algogenic organic matter and membrane fouling. The results revealed that the three pretreatment methods increased filtration flux and reduced membrane fouling. Among them, the diatomite pretreatment could increase the filtration flux by 915%, which was significantly better than other pretreatment. Activated carbon pretreatment could effectively adsorb aromatic protein-like compounds, and obviously decrease the resistance of irreversible chemical fouling. OCT and SEM analysis showed that no pretreatment caused the most serious membrane fouling, the lowest roughness and thickness of cake layer structure, while diatomite pretreatment could obviously mitigate membrane fouling by optimizing the cake layer structure. Meanwhile, the XDLVO theoretical results further confirmed that diatomite pretreatment had the best alleviating effect on membrane fouling. For further insight, the results provide valuable guidance for the development of membrane filtration of algae suspensions.
CHARACTERISTICS OF MTF-1 AND CADMIUM STRESS ON RHYTHMIC EXPRESSION IN CHINESE PERCH SINIPERCA CHUATSI
PAN Ya-Xiong, ZHOU Jun, ZHANG Yu, TAO Zheng-Sheng, PAN Jia-Lin, TANG Zhao-Yang, FAN Yi-Wei, HU Ming-Guang, LI Hui-Ju, ZHANG Jian-She, CHU Wu-Ying
 Available online   , doi: 10.7541/2023.2022.0131
[Abstract](420) [FullText HTML](299) [PDF 1058KB](3)
Abstract:
To study the characteristics of mtf-1 gene of Siniperca chuatsi and the effects of cadmium stress on the circadian rhythm of mtf-1 gene in brain, we analyzed cis-regulatory elements of mtf-1 gene promoter, sequence characteristics and tissues expression patterns of mtf-1 gene, and explored the circadian rhythm changes of mtf-1 gene expression in brain of Siniperca chuatsi under cadmium stress. The results showed that there were binding sites of zinc finger proteins including ZNF136, ZNF384 and ZNF143 in the mtf-1 promoter of S. chuatsi, indicating that their expression is potentially regulated by metal ions. In addition, RORβ binding sites of the core clock gene were also found in the mtf-1 promoter of S. chuatsi, indicating that the mtf-1 gene is potentially involved in circadian rhythm regulation. The analysis of function domain indicated that mtf-1 gene has a highly conserved DNA binding domain and a non-conserved trans-activation domain. mtf-1 gene is expressed in different tissues of S. chuatsi, among which the highest expression level is found in brain tissue, followed by kidney. Under natural conditions, mtf-1 gene showed a trend of high expression at night and low expression in the brain at day. The expression peak phase occurs at ZT 12.65h (nightfall), which is consistent with its life habits. Under cadmium stress, the mtf-1 gene in the brain of S. chuatsi presented continuous low expression level, and with no significant difference between day and night, indicating that the mtf-1 gene expression in the brain of S. chuatsi could be significantly inhibited by cadmium, and the circadian rhythm of mtf-1 in the brain tissue of S. chuatsi is damaged.
EFFECTS OF DIETARY BILE ACIDS SUPPLEMENTATION ON LIPID METABOLISM IN LIVER OF EUROPEAN EEL (ANGUILLA ANGUILLA) JUVENILES
MA Zhi-Ru, ZHAO Pan-Yue, ZHAI Shao-Wei
 Available online   , doi: 10.7541/2023.2022.0035
[Abstract](409) [FullText HTML](299) [PDF 1523KB](8)
Abstract:
The present study was conducted to investigate the effects of dietary bile acids (BA) on the lipid metabolism in the liver of European eel (Anguilla anguilla) juveniles. Nine cement tanks stocked with similar fish size (141.5 g/fish) and fish weight (681.7±22.7 kg/tank) were randomly divided into three treatment groups. The trial fish were fed diets with BA levels being 0 (control group), 500 mg/kg (BA1 group), and 1000 mg/kg (BA2 group), respectively. There were three tanks in each treatment group. The trial lasted for 15weeks.Compared with the control group, the numbers of the fat vacuole in hepatocytes of European eel juveniles were decreased in BA1 and BA2 groups. The decreased lipid content in the liver were found in the BA1 and BA2 groups in comparison with the control group (P<0.05). The levels of fatty acid synthetase in the liver were significantly increased in the BA1 and BA2 groups(P<0.05). The acetyl CoA carboxylase level in the liver of the BA1 group was significantly higher than that of the control group (P<0.05). The activities of hepatic lipase, lipoprotein lipase, and total lipase in the liver of BA1 and BA2 groups were significantly higher than those in the control group (P<0.05). There were no significant differences in lipid metabolism enzymes activities or levels in the liver between the BA1 group and BA2 group (P>0.05). The levels of phosphatidylcholine and lysophosphatidylcholine were up-regulated in the liver in European eel juveniles of BA1 group, which might mainly enhance theglycerophospholipid metabolism and glycerolipid metabolism. In conclusion, dietary BA supplementation might decrease lipid accumulation by decreasing the levels of enzymes related to lipid synthesis and increasing the activities of enzymes related to lipid decomposition with mainly up-regulating theglycerophospholipid metabolism and glycerolipid metabolism in the liver of European eel juveniles.
ACUTE HYPOXIA/REOXYGENATION STRESS AFFECT ANTIOXIDANT AND ENERGY METABOLISM OF PROCAMBARUS CLARKII
XU Yu, XU Zhi-Qiang, YAN Wei-Hui, LI Jia-Jia, HUANG Hong-Bing, SUN Meng-Ling, TANG Jian-Qing, PAN Jian-Lin
 Available online   , doi: 10.7541/2022.2021.0390
[Abstract](400) [FullText HTML](278) [PDF 2320KB](7)
Abstract:
Dissolved oxygen (DO) is vital in aquaculture, which influences the survival and growth of red swamp crayfish, Procambarus clarkii. To reveal the profiles of antioxidant and energy metabolism in response to acute hypoxia/reoxygenation stress, the activity of some key enzymes associated with antioxidant and non-specific immunity were measured and the ultrastructure changes in the hepatopancreas and gill were observed. Experimental crayfish with average body weight of (26.5±1.8) g were subjected to acute hypoxia stress (DO 1.0±0.1 mg/L), followed by reoxygenation (DO 6.8±0.2 mg/L). Hepatopancreas, gill and hemolymph from five groups of crayfish (six crayfish per group), including normoxia, hypoxia for one and six hours, and reoxygenation for one and 12h, were used to measure the changes of antioxidant and non-specific immunity enzymes. Cell ultrastructure of gill and hepatopancreas were also examined by transmission electron microscope. The results showed that the activity of superoxide dismutase (SOD) in hepatopancreas and hemolymph decreased significantly under hypoxia stress (P<0.05), while it increased significantly in hepatopancreas, hemolymph and gill in the reoxygenation stage (P<0.05). The significant increase of SOD activity may be related to the excessive production of superoxide anion (ROS) in the reoxygenation process. The content of malonaldehyde (MDA) in hemolymph and gill tissue was significantly higher than that of the control group (P<0.01), indicating lipid peroxidation occurred in cells under hypoxia/reoxygenation stress. Under hypoxic stress, the activities of acid phosphatase (ACP) and alkaline phosphatase (AKP) in hepatopancreas, gill and hemolymph were both significantly increased (P<0.05), and the activities of ACP in hepatopancreas and gill tissue were significantly decreased after reoxygenation for 12h (P<0.01). These results suggested that hypoxic-reoxygenation stress may affect the immune response of the crayfish. Compared with the control group, lactic dehydrogenase (LDH) content and total ATPase activity in hepatopancreas, hemolymph and gill tissue were significantly increased under acute hypoxic stress (P<0.05), indicating that the energy metabolism function of cells was seriously affected. Observation of mitochondria ultrastructure revealed that the organization structure of gill and hepatopancreas were seriously injured after acute hypoxic/reoxygenation stress. Mitochondrial damage, including mitochondria dissolved into vacuole, mitochondria swelled irregularly and their cristae fractured and become fuzzy. During hypoxia/reoxygenation stress, the number of mitochondria in hepatopancreas cells significantly increased while the number of lysosomes decreased significantly. The results showed that hypoxic-reoxygenation stress can cause great damage to the hepatopancreas and gill of P. clarkii, and affect the activities of antioxidant and non-specific immunity enzymes. Furthermore, the results indicated that hypoxic-reoxygenation stress has great influence on immune defense ability and energy metabolism function of P. clarkii.
EFFECTS OF SELENIUM-RICH CARDAMINE HUPINGSHANENSIS ON GROWTH, BIOCHEMICAL INDICES, SELENIUM METABOLISM, ANTIOXIDANT CAPACITIES AND INNATE IMMUNITIES IN JUVENILE BLACK CARP (MYLOPHARYNGODON PICEUS)
ZHANG Chen, JIA Xiao-Wei, QIAN Peng-Cheng, CHEN Yan-Ting, WU Cheng-Long, YE Jin-Yun
 Available online   , doi: 10.7541/2023.2022.0063
[Abstract](379) [FullText HTML](320) [PDF 1089KB](6)
Abstract:
In order to investigate the effects of different levels of dietary selenium-rich Cardamine hupingshanensis on the growth performance, physiological and biochemical parameters, selenium metabolism, antioxidant capacities and innate immunities in black carp (Mylopharyngodon piceus), 360 juvenile black carps with initial body weight of (5.51±0.02) g were randomly divided into four groups, and each group had 3 replicates. Four levels of selenium-rich C. hupingshanensis (0, 0.5, 1.0 and 2.0 g/kg) (selenium levels 0.04, 0.43, 0.75 and 1.57 mg/kg) were added and made four isonitrogenous and isoenergetic diets. The culture period was conducted for 60 days. The results showed that adequate selenium-rich C. hupingshanensis (0.5 and 1.0 g/kg) could significantly increase the weight gain (WG) and specific growth rate (SGR), while significantly decrease the feed conversion ratio (FCR), comparing with the control group and the excessive group (2.0 g/kg) (P<0.05). In the fish body composition, the crude protein levels were significantly increased in fish fed with 1.0 g/kg selenium-rich C. hupingshanensis, and then decreased at 2.0 g/kg selenium-rich C. hupingshanensis. Adequate supplemental level of selenium-rich C. hupingshanensis (0.5 and 1.0 g/kg) could significantly increase the serum albumin (ALB), triglycerides (TG) and total cholesterol (TCH), while decrease serum glucose (GLU) levels. Higher activities of superoxide dismutase (SOD), the catalase (CAT) and glutathione peroxidase (GSH-PX), levels of total antioxidant capacities (T-AOC), and the glutathione (GSH) (P<0.05) were shown in the liver of fish fed adequate supplemental level of selenium-rich C. hupingshanensis. Meanwhile, adequate supplemental level of selenium-rich C. hupingshanensis could significantly increase expression level of nuclear factor-erythroid-2-related factor 2 (Nrf2), Mn-superoxide dismutase (Mn-SOD), catalase (CAT), glutathione peroxidase 1 (GPX1) and glutathione peroxidase 4 (GPX4), while decrease the expression level of Kelch-like-ECH-associated protein 1 (Keap1) in the fish liver (P<0.05). In addition, adequate supplemental level of selenium-rich C. hupingshanensis significantly increase expression level of selenium-metabolic proteins, including selenocysteine lyase (SCLY), selenide, water dikinase 1 (SPS1), 15kDa selenoprotein (SEP15), selenoprotein T2 (SEPT2), selenoprotein H (SEPH), selenoprotein P (SEPP) in the liver of juvenile black carp compared with the control group (P<0.05). Furthermore, the expression level of innate immune defense molecules, including liver-expressed antimicrobial peptide 2 (Leap2), hepcidin (HEPC), lysozyme (LYS) and complement 3 (C3) were significantly up-regulated by adequate supplemental level of selenium-rich C. hupingshanensis (P<0.05). In conclusion, these results suggest that the adequate dietary supplemental level of selenium-rich C. hupingshanensis could significantly increase growth performances, promote the selenium metabolism, improve antioxidant capacities and enhance innate immunities in black carp.
EFFECT OF VITAMIN D3 ON POLARIZATION PHENOTYPE OF HEAD KIDNEY MACROPHAGES IN PELTEOBAGRUS FULVIDRACO
WANG Meng, CHENG Ke, MA Chun-Song, WANG Chun-Fang
 Available online   , doi: 10.7541/2023.2022.0037
[Abstract](385) [FullText HTML](304) [PDF 1546KB](3)
Abstract:
As a key class of immune cells, macrophages are not only the first line of defense against pathogen invasion, but also play different functions through polarization. Macrophages can secrete pro-inflammatory factors and cause excessive inflammatory response, resulting in a variety of inflammatory diseases when polarizing into M1 type. Macrophages can secrete anti-inflammatory factors and play an anti-inflammatory role when polarizing into M2 type. Macrophages are characterized by phenotypic heterogeneity and functional diversity, and it is important to target macrophage-type polarization through nutritional regulation. Vitamin D3 has been proved to play important role in fish anti-inflammatory response. In this study, the effects of 200pM vitamin D3 on polarization phenotype of head kidney macrophages of Pelteobagrus fulvidraco were studied. The primary macrophage cells of Pelteobagrus fulvidraco head kidney were isolated and cultured. After being challenged by LPS and cAMP to induce macrophage polarization, the cell morphological changes were observed by inverted light microscopy, and the functional changes were studied by measuring survival rate, phagocytosis, reactive oxygen species and nitric oxide production, superoxide anion radical and arginase activity, as well as the related gene expression charactering in different macrophages polarization states. The results showed that vitamin D3 reduced the mortality of M1 and M2 macrophages and enhanced the phagocytic activity of macrophages. In M1 macrophages, vitamin D3 inhibited the production of reactive oxygen species (ROS) and inflammatory mediator nitric oxide (NO), reduced the activity of superoxide anion free radical, and decreased the expression of interleukin-1β (IL-1β) and tumor necrosis factor (TNF-α)(P<0.05). The activity of arginase as well as the expression of interleukin-10 (IL-10) and transforming growth factor (TGF-β) in M2 cells was up-regulated by vitamin D3 (P<0.05). In conclusion, vitamin D3 inhibited the polarization of macrophages to M1 phenotype, promoted the polarization of macrophages to M2 phenotype, and played an anti-inflammatory role. In the current study, Nos-2 and arg-2 were found to be biomarker genes of M1 and M2 macrophages, respectively. These results preliminarily reveal the mechanism of vitamin D3 on the polarization of head kidney macrophages of Pelteobagrus fulvidraco, and favor the regulation of macrophage phenotypes in the treatment of inflammation, which provides useful information for further study on the polarization of fish macrophages and the effect of vitamin D3 on polarization regulation.
CHANGES OF FATTY ACID COMPONENTS AND CONTENT OF COILIA NASUS IN THE YANGTZE RIVER DURING SPAWNING MIGRATION
MA Feng-Jiao, GUO Wen-Jun, YING Cong-Ping, YANG Yan-Ping, XU Pao, LIU Kai, YIN Guo-Jun
 Available online   , doi: 10.7541/2023.2022.0047
[Abstract](461) [FullText HTML](304) [PDF 1520KB](1)
Abstract:
In order to elucidate the changes in the components and contents of fatty acid of female Coilia nasus in the Yangtze River during spawning migration process, the components and contents of various fatty acid in the hepatopancreas, muscle and ovary of Coilia nasus were determined from three aspects of migration distance, ovarian development and body length. The results showed that 28 kinds of fatty acids were detected in the three tissues of 62 female Coilia nasus, including 8saturated fatty acids (SFA), 6 monounsaturated fatty acids (MUFA) and 14 polyunsaturated fatty acids (PUFA). Among all the fatty acids, MUFA were the most abundant, accounting for more than 56.23% of the total fatty acids in each tissue. Specific fatty acids with higher levels included C18:1, C16:0, C16:1, DHA and EPA. During migration, the total amount of fatty acids in the hepatopancreas of Coilia nasus increased from (526.61±38.50) mg/g in Chongming to (587.21±124.72) mg/g in Anqing, but decreased by 33.03% and 57.09% in muscle and ovary respectively. The total amount of fatty acids, SFA, MUFA and PUFA in muscle were positively correlated with body length (P<0.05), but the contents of total fatty acids and fatty acids components in hepatopancreas and ovary had no significant correlation with body length (P>0.05). During the ovarian development from stages Ⅱ to Ⅳ, the contents of total fatty acids in hepatopancreas and muscle decreased by 47.56% and 22.40%, respectively, but increased from 364.17 to 418.95 mg/g in ovary. The contents of C18:1, EPA and C16:1 in hepatopantras and muscle decreased, but increased by 1.21, 1.36 and 1.31 times in ovary (stages Ⅱ to Ⅲ), respectively, indicating that these fatty acids were also significantly transferred during ovarian development. The present study also found that there were more fatty acids in muscle of large size Coilia nasus, the abundance of SFA and MUFA in each tissue, and EPA and DHA in the ovaries, which provided sufficient nutrition for migration activities and gonad development.
STUDIES ON RSPO1 IN MODULATING CONVERGENCE AND EXTENSION MOVEMENT IN ZEBRAFISH EMBRYOS
ZHANG Xin, ZHANG Qi, PAN Jun, ZHOU Jian-Feng, BAI Yan, RONG Xiao-Zhi
 Available online   , doi: 10.7541/2023.2022.0095
[Abstract](525) [FullText HTML](300) [PDF 2021KB](4)
Abstract:
In vertebrate embryos, gastrulation is the fundamental morphogenetic movements. It leads to the formation of the basic germ layers and the body axis. During gastrulation, convergence and extension (CE) movements narrow a group of cells mediolaterally and lengthen them to facilitate the elongation of the anteroposterior axis. In this process, the planar cell polarity (PCP) pathway, also called the noncanonical Wnt signaling pathway, is of particular importance to control CE movements. However, the precise cellular and molecular mechanisms underlying this process remains to be further studied.Rspo1 (R-spondin 1) is a secreted protein that has been implicated in activating the Wnt/β-catenin signaling levels with Wnt ligands, through which Rspo1 promotes angiogenesis and specifies hematopoietic stem cells, as well as promotes female development. Recently, it was reported that Rspo1 exhibits Wnt/β-catenin independent roles. For example, Rspo1-Lgr4-cAMP-Erα axis regulates estrogen receptor expression, and RSPO1-LGR5 activates TGFβ signaling in colon cancer. Given the complex role of Rspo1, we speculate that Rspo1 is involved in the early embryonic development.To investigate the developmental role of Rspo1 in zebrafish embryos, we examined the spatiotemporal expression pattern of rspo1 using RT-PCR and whole-mount in situ hybridization. The results showed that rspo1 mRNA is maternally deposited and expresses ubiquitously in early embryonic stages before 12hpf (hours post fertilization), implying that Rspo1 may play an important role in the regulation of embryonic development. Next, we carried out gain-of-function and loss-of function analysis of Rspo1. The results showed that either overexpression or knockdown of rspo1 abrogates the CE movements during gastrulation. In order to explore whether rspo1 affects CE movement by participating in Wnt/PCP signaling pathway, we used AP-1 luciferase reporter to monitor Wnt/PCP in zebrafish embryos. The results showed that forced expression of rspo1 decreases but knockdown of rspo1 increases Wnt/PCP signaling reporter activity, indicating that Rspo1 inhibits Wnt/PCP signaling pathway. Consistent with this result, the phosphorylated-JNK levels, an indicator of activity of Wnt/PCP signaling pathway, dramatically increased in rspo1 morphant embryos at gastrulation stage. Further analyses indicate that coinjection of rspo1 mRNA and dnJNK (Dominant negative JNK) mRNA, or coinjection of rspo1 MOs and wnt11/wnt5b mRNA synergistically enhanced CE defects. Taken together, these results suggest that Rspo1 regulates CE movements during gastrulation by negatively regulating the Wnt/PCP signaling in zebrafish embryos. Overall, our studies will provide novel insights into the regulation of Wnt/PCP signaling in vertebrates.
MOLECULAR SYSTEMATICS OF XENOCYPRINAE (CYPRINIFORMES, CYPRINIDAE)
LI Lin, CHEN Wei-Tao, YANG Yong-Tao, ZHOU Chuan-Jiang, HE Shun-Ping, FENG Cheng-Guang
 Available online   , doi: 10.7541/2023.2022.0085
[Abstract](425) [FullText HTML](321) [PDF 1443KB](7)
Abstract:
Xenocypridinae, as one of the benthic fishes of Cyprinidae (Cypriniformes) in East Asia, contains three genera of about nine species and is, therefore, one of the smallest subfamilies in Cyprinidae. Despite the small number of species, the phylogenetic relationships of Xenocypridinae still lack a valid and extensive assessment. To this end, this study assessed the phylogenetic relationships and divergence times between members of the subfamily Xenocypridinae using a variety of phylogenetic approaches based on two mitochondrial genes and five nuclear genes. The results show that analyses based on maximum likelihood and Bayesian approaches harvest consistent topologies, but there are some conflicts in the results based on different datasets. In particular, the monophyly of each of the three genera was well supported in all analyses, but the evolutionary relationships between them were discordant between the results based on mitochondrial and nuclear genes. Analyses based on the coalescent algorithm yielded a third phylogenetic relationship among the genera. These results imply that both ancient gene flow and lineage sorting influenced the relationships between deep clades of the subfamily Xenocypridinae. Additionally, these analyses give insights into the classification of these species, for example, Xenocypris hupeinensis should belong to the genus Xenocypris; the nested relationships among species X. argentea, X. yunnanensis, and X. davidi suggest the possibility of gene flow and the need for taxonomic reconsideration. Finally, a fossil-based molecular clock assessment indicates that the divergence of key Xenocypridinae clades occurred mainly in the late Miocene (ca. 15—­12 Ma), which coincides with the intensification of the Asian monsoon and supports the hypothesis that the ichthyofauna of East Asia was established in the early Pliocene and flourished to date. In summary, this study has systematically assessed the phylogeny of Xenocypridinae and made recommendations for future work.
CHARACTERISTICS OF COILIA NASUS RESOURCES AFTER FISHING BAN IN THE YANGTZE RIVER
MA Feng-Jiao, YANG Yan-Ping, FANG Di-An, YING Cong-Ping, XU Pao, LIU Kai, YIN Guo-Jun
 Available online   , doi: 10.7541/2023.2022.0070
[Abstract](411) [FullText HTML](278) [PDF 2239KB](4)
Abstract:
Estuarine tapertail anchovy (Coilia nasus) is a valuable migratory fish species and the most representative species in the lower reaches of the Yangtze River, and its resource status have been attracted an increasing amount of attention. In order to effectively track and evaluate the recovery effects of C. nasus resource after the national fishing ban strategies, this study carried out an investigation on the spatial-temporal characteristics of biology and resource density of C. nasus in the Yangtze River estuary from 2019 to 2021 based on the survey results conducted before the withdrawal of productive fishing (2017—2018). The results showed that a total of 2895 C. nasus were collected, and 1960 were dissected in 2021, with a female to male ratio of 1.87:1. The mean body length of (272±32) mm and mean body weight of (91.4±33.4) g, increased by 4.41% and 37.55% compared with 2019—2020, and significantly increased by 5.84% and 22.85% compared with 2017—2018 (P<0.05), respectively. The mean fatness of 0.44±0.10, 18.28% higher than that in 2019—2020 and 12.82% higher than that in 2017—2018. In terms of resource density, the mean daily catch number (NB) and weight (WB) in 2021 were 170 ind./d and 15.56 kg/d, 1.09 times and 1.48 times higher than that in 2019—2020, respectively. The number (Nt) and weight (Wt) of single net were 50 ind./net and 4.56 kg/net, 1.24× and 1.69× increase compared with 2019—2020, respectively, and the resource density showed an increasing trend from 2019 to 2021. The migration time of C. nasus mainly occurred in March and April in 2020—2021, which was earlier than the peak of migration in 2019. The density of C. nasus in deep water was significantly higher than that in shallow water (P<0.05). Our study suggested that the recourses of C. nasus did not recover immediately in 2019 due to the influence of overfishing in the early stage and the fact that the other fishery production was not completely eliminated. Since January 1st, 2021, the implementation of fishing ban in key waters of the Yangtze River and the Yangtze River Estuary has reduced fishing pressure, and the population biological specifications and resource density of C. nasus in Yangtze River estuary have recovered significantly. The resource shows a good tendency towards recovery, highlighting the important role of the fishing ban management policy. This study suggested that long-term follow up monitoring should be carried out on representative migratory species such as C. nasus to obtain the resource characteristics during the period of fishing ban in the Yangtze River, and to support the assessment of fishing ban effect and biological integrity evaluation in the Yangtze River.
SCREENING OF CHROMOCHLORIS ZOFINGIENSIS HYPER ASTAXANTHIN-PRODUCING MUTANTS AND THE MECHANISMS UNDERLYING ASTAXANTHIN SYNTHESIS
XIAO Ling, HAN Dan Xiang
 Available online   , doi: 10.7541/2023.2022.0083
[Abstract](421) [FullText HTML](313) [PDF 1807KB](3)
Abstract:
Chromochloris zofingiensis can achieve ultra-high cell density and simultaneously accumulate astaxanthin under heterotrophic conditions, however, the relatively low cellular astaxanthin content of C. zofingiensis hinders its commercial application. With the aim to improve the ellular astaxanthin content of C. zofingiensis, this study screened a mutant 12C10 with enhanced astaxanthin content from the mutant library established by using ethylmethylsulfone mutagenesis. The astaxanthin content of 12C10 was 74% higher than that of the wild type under the heterotrophic normal conditions, and 25% higher than that of the wild type at day 4 under the nitrogen deficiency conditions. Additionally, the results of the plant primary metabolomics analysis revealed the differences in the metabolites level between 12C10 and wild type under the normal conditions. Specifically, the contents of fatty acids and amino acids except glutamate generally decreased, but the content of glutamate significantly increased in 12C10 compared to wild type. Reduced synthesis of amino acids and fatty acids in 12C10 can provide more carbon skeleton, NADPH, and ATP for astaxanthin synthesis. The accumulation of glutamate on the one hand may stimulate the activity of glucose 6-phosphate dehydrogenase in pentose-phosphate pathway to produce more NADPH, and cause production of reactive oxygen species to trigger astaxanthin synthesis. The metabolome analysis also showed that the enhanced astaxanthin synthesis in 12C10 may be related to the enhanced ethylene synthesis. This study lays the foundation for improving C. zofingiensis astaxanthin content through metabolic regulation in the future, and is of great significance for guiding the development of the new processes of C. zofingiensis astaxanthin accumulation.
FEEDING STRATEGIES ON GROWTH AND REPRODUCTION OF YELLOW CATFISH (PELTEOBAGRUS FULVIDRACO)
GAO Wei-Ye, FEI Shu-Zhan, LIU Hao-Kun, HAN Dong, YANG Yun-Xia, JIN Jun-Yan, ZHU Xiao-Ming, ZHANG Zhi-Min, XIE Shou-Qi
 Available online   , doi: 10.7541/2023.2022.0096
[Abstract](1244) [FullText HTML](305) [PDF 951KB](8)
Abstract:
Most of feeding strategies which maximize the feeding rate are just designed for fish farming but not suitable for broodfish. Overfeeding caused by these feeding strategies will even impair the reproductive performance. However, moderate feed restriction can maintain fecundity while lowering breeding costs. Feeding strategies based on compensatory growth may promote growth and fecundity of broodfish, or make them mature faster. To test the effects on growth and reproductive capacity of female yellow catfish Pelteobagrus fulvidraco, fish were fed with four groups for 67 days: satiety feeding (AS), 80% satiety (80AS), 50% satiety (50AS) and starvation for 35 days and then satiety feeding for 31 days (HAS). Samples were taken at the middle (34 days) and the end of the experiment and various indexes were measured in this time. Results showed that the total food intake of the 80AS, 50AS and AS group roughly conforms to the proportion of the experimental design. The daily food intake of the HAS group was the highest. No significant difference was observed in weight gain rate and final weight between the 80AS group and the AS group. Meanwhile, the final weight of the HAS group was significantly lower than that of AS group, which shows the growth of the HAS group failed to reach those who fed at satiety level in the same period. As the serum lipid and protein concentrations increased in each group, the retention of lipid and protein in gonad increased. It can be inferred that parent fish preferentially transmit nutrition to gonads rather than muscles. There was no significant difference in final egg diameter between the AS group and 80AS group, which were significantly higher than the 50AS group and lower than the HAS group, indicating that the refeeding has a compensatory effect on egg growth. Feeding strategy did not impact absolute fecundity and fecundity, but alterd the expression of genes which related to gonad development. The relative expression of StAR, SF-1, 3β-hsd and cyp19a1 in the 80AS, 50AS and HAS was significantly lower than that in AS group in the middle stage. Nevertheless, no significant difference was found between these group and AS group in the end stage. The trend of plasma estradiol and testosterone was consistent with the above expression of genes, indicating that the synthesis and secretion of sex hormones and gonadal development in 80AS, 50AS and HAS groups were only inhibited by feeding strategy in the early stage. Finally, the inhibition effect was relieved, and the relevant indexes of each group recovered. These may be the result of interaction between the stress response and reproduction of broodfish. Based on the data above, satisfying the material and energy needs of gonadal development has priority in yellow catfish’s resource allocation. Although the growth of HAS group was lower than that of AS group, there was no difference in reproductive ability between HAS group and AS group, indicating that parent Pelteobagrus fulvidraco mainly compensated for fecundity rather than growth after starvation. Therefore, it is necessary to redesign the feeding strategy based on compensatory growth according to the physiological characteristics which need further exploration to improve the fecundity of Pelteobagrus fulvidraco broodfish. The growth and reproduction indexes of 80AS group were not significantly different from those of AS group, and the food intake was significantly lower than that of AS group, which was a more suitable feeding strategy in the breeding period of yellow catfish.
RESEARCH PROGRESS ON NEUROBEHAVIORAL TOXICITY IN FISHES
LI Shu-Shen, ZHAO Zhi-Yuan, SUI Chang-Run, JI Mei-Xu, PANG Yue, LI Qing-Wei, LI Jun
 Available online   , doi: 10.7541/2022.2021.0324
[Abstract](585) [FullText HTML](344) [PDF 791KB](9)
Abstract:
Neurobehavioral toxicity is one of important research fields in neuroscience, neuropharmacology and neurotoxicology, which of growing importance for understanding the mechanisms of chemicals on nervous system and for evaluating the quality of ecosystem. Possessing well-developed central nervous system, fish is extremely sensitive to chemicals in the water environment, and the nervous system can produce a comprehensive and coordinated response to various stimuli, which resulted in complex, well-characterized behaviors, including its swimming behaviors and social behaviors. And fish behavior is now recognized as a complex, homologous to mammals, context specific, adaptive and highly variable. A variety of behavioral tests have been developed to assess motor function, stress response, social behavior and learning/memory in fish. And behavior of fish can be measured to determine the functional impact of chemicals. The elementary actions of a neurotoxicant can be followed in terms of disruptions of neural differentiation, proliferation, migration, outgrowth, synapse formation, and circuit development. Fish has been widely used as a tool to detect toxins in water samples and to investigate the mechanisms of action of environmental toxins and their related diseases in recent years. Fish offer many advantages that complement classic mammalian models for the study of normal development as well as for neurobehavioral effects of exposure to chemicals. And fish provide a key intermediate model of neurobehavioral toxicity between high throughput in vitro cell-based assays and the classic mammalian models as they have the accessibility of in vitro models and the complex functional capabilities of mammalian models. The present article reviews recent research progress on neurobehavioral toxicology studies using fish as a model , and we present and discuss the neurobehavioral toxicity of typical pollutants (microplastics and toxins absorbed to microplastics, organic pollutants, et al.) and drugs (alcohol, caffeine, benzodiazepines, selective serotonin reuptake inhibitors, et al.) on fish. And future research directions are proposed. The article is expected to provide a reference for researchers in neurobehavioral toxicity and its application.
TRANSCTIPTOMIC RESPONSES TO ACUTE LOW TEMPERATURE STRESS IN GIANT FRESHWATER PRAWN MACROBRACHIUM ROSENBERGII
YANG Min-Min, TU Hai-Hui, XING Qian-Qian, TANG Qiong-Ying, YI Shao-Kui, XIA Zheng-Long, CAI Miao-Ying, CHEN Guo-Zhu, LAN Xuan, ZHONG Zhen-Xiao, HUANG Xiao, GAO Quan-Xin, YANG Guo-Liang
 Available online   , doi: 10.7541/2022.2022.0033
[Abstract](399) [FullText HTML](298) [PDF 1229KB](3)
Abstract:
The giant freshwater prawn (GFP) Macrobrachium rosenbergii is one of the important economically freshwater shrimp species, and its annual production in China is dominant in the world. However, its tolerance to low temperatures is extremely poor, and acute low temperatures can lead to large-scale deaths and cause huge economic losses. In order to explore genes related to the GFP response to acute low temperature, comparative transcriptomic analyses were performed for the hepatopancreas of adult GFPs exposed to low temperature. The low temperature stress group (16℃) and the control group (24℃) were set up. The water temperature of the stress group was decreased from 24℃ to 16℃ at a rate of around 2℃/h through adding ice cubes. The hepatopancreas were collected for transcriptomic analyses from exposed shrimps respectively at 1, 3, 6, 12, 24h after acute cooling at 16℃, rewarming to 24℃ and control group. The results of differentially expressed genes (DEGs) showed that 1702 DEGs were identified between samples of cooling for 1h and the control group (M1 vs. C0); the number of DEGs between the stress group and the control group began to increase gradually with stress time, and reached the maximum at 6h (M3 vs. C0, a total of 2899), then gradually decreased, maintaining the homeostasis under low temperature stress. After rewarming to 24℃, the number of DEGs (M6 vs. C0, 1969) returned to the level of 1h stress. Additionally, the DEGs number (5062) between samples of stress for 3h and rewarming to 24℃ (M2 vs. M6) was almost 1.5 times of that (3516) between samples of stress for 1h and rewarming (M1 vs. M6). With the extension of time, the number of DEGs decreased gradually, suggesting that the homeostasis of the GFP had drastic changes in the first 3h of acute low temperature stress, but the adaptability to low temperature gradually increased with stress time, and a steady state under low temperature was established probably after being stressed from 3h to 6h, and after rewarming, the homeostasis returned to the equilibrium of short-term cold stress. KEGG enrichment analysis showed that DEGs was enriched in lysosome, starch and sucrose metabolism, antigen processing and presentation. Adhesion spots, ECM-receptor interaction, metabolism of cytochrome P450 to isobiotic substances, glutathione metabolism, oxidative phosphorylation, and p53signaling pathway were also involved in the regulation of acute cold stress in the GFP. The common DEGs among all groups were clustered as the cell function and immunity modules and energy metabolism modules. In addition, the up-regulated cytochrome P450 2L1-like gene in the arachidonic acid metabolic signaling pathway was screened, and its expression increased firstly and then decreased with the increase of cold stress time. NADP-specific isocitrate dehydrogenase, an upregulated gene in glutathione metabolic signaling pathway, was firstly reduced and then increased with time. Expression of hexokinase, a down-regulated gene in starch and sucrose metabolic signaling pathways, increased and then decreased over time. The above mentioned enriched pathways indicated that the metabolic function of the GFP might have been seriously affected under acute low temperature stress, with a large amount of reactive oxygen species being produced, the balance of energy circulation being disrupted, and the immune system being also damaged, which was corroborated with the phenomena of fasting, slow movement, susceptibility to diseases and even death of the GFP under acute low temperature. Consequently, these screened pathways and genes may play important roles in energy metabolism and immune regulation during acute cold stress in the GFP. The present study provides basic data for revealing the molecular regulatory mechanism of the response of the GFP to acute low temperature stress, also provides a theoretical basis for the selective breeding of new cold-tolerant GFP varieties.
DISTRIBUTION PATTERNS OF MACROZOOBENTHOS IN THE MID-LOWER MAINSTEM OF THE YANGTZE RIVER
ZHANG Meng-Na, GAO Wen-Juan, ZHANG Ting-Yu, ZHAO Yong-Jing, WU Xing-Hua, LI Yuan, WHANG Hong-Zhu, CUI Yong-De
 Available online   , doi: 10.7541/2022.2021.134
[Abstract](517) [FullText HTML](321) [PDF 2449KB](12)
Abstract:
To understand the status quo and variation trend of macrozoobenthos along the mid-lower mainstem of the Yangtze River, three field surveys were carried out from 2019 to 2020. A total of 129 taxa of macrozoobenthos were recorded. The mean density and biomass of macrozoobenthos were 174 ind./m2 and 0.645 g/m2 (dry weight, similarly below), respectively. From the middle to the lower reaches, the total taxa number of macrozoobenthos decreased; the density of aquatic insects decreased, while the density of other taxa increased; the density of collector-gatherers and scrapers decreased, while predators and shredders increased. Correlation analysis showed that water depth and mean velocity were two parameters significantly affecting the distribution of macrozoobenthos. The distribution patterns of different groups were various. Oligochaetes and chironomids prefer slow flow and deep water, and there is little difference in the optimum range of water depth and mean velocity between them. Gastropods prefer shallow and slow flow, and both the optimum range of water depth and mean velocity are narrow, while bivalves are opposite. Other aquatic insects, polychaetes and gammarids are suitable for a wide range of water depth and a relatively narrow range of mean velocity. The results provide basic information for the research of environmental flow requirement of macrozoobenthos, reservoir ecological regulation and other protection and management work in the mid-lower mainstem of the Yangtze River.
EFFECTS OF DIETARY SUPPLEMENTATION OF BOVINE LACTOFERRICIN ON GROWTH PERFORMANCE, DIGESTIVE ENZYMES ACTIVITY, INTESTINAL TISSUE STRUCTURE AND DISEASE RESISTANCE OF JUVENILE LARGEMOUTH BASS (MICROPTERUS SALMOIDES)
HUANG He, ZHUANG Yi, ZHONG Guo-Fang, TANG Xiang-Shan
 Available online   , doi: 10.7541/2023.2021.0386
[Abstract](517) [FullText HTML](362) [PDF 2387KB](10)
Abstract:
Largemouth bass (Micropterus salmoides), commonly known as California bass, has been introduced to China since the 1980s. After years of breeding development, it has now become one of the important species of freshwater aquaculture in China. However, with the rapid development of intensive aquaculture and irregular feeding methods, California bass diseases frequently occur. The traditional method to prevent and treat bacterial diseases in fish is to add antibiotics to aquafeeds. However, the abuse of antibiotics can lead to problems such as bacterial resistance, drug residues and environmental pollution. Therefore, the selection of suitable antibiotic substitutes for the aquaculture industry is an urgent matter, and it has far-reaching significance for the growing largemouth bass aquaculture industry. Research on alternatives to antibiotics has found that antimicrobial peptides are a kind of polypeptides produced by the non-specific immune system in organisms. Because of their unique biological activity, antibacterial and bactericidal mechanisms different from traditional antibiotics, they are not easy to produce drug resistance and no resistance. Pollution and other advantages are expected to be developed into a new type of high-efficiency antibacterial drugs, which have great potential to replace antibiotics in the field of feed additives. The bovine lactoferricin (LfcinB) used in this test is a cationic antimicrobial peptide containing 25 amino acid residues produced by pepsin hydrolysis of bovine lactoferrin under acidic conditions. It has a broad-spectrum and high-efficiency antibacterial ability and antiviral ability. However, its research as a feed additive has not been reported yet. This study investigated the effect of the dietary supplementation of bovine lactoferricin on growth performance, digestive enzymes activity, intestinal tissue structure and disease resistance against Aeromonas hydrophila in juvenile largemouth bass, aiming to evaluate its potential to replace antibiotics and provide a theoretical basis with the application of functional feed. A total of 450 tails largemouth bass with an average body weight of (19.88±0.03) g were randomly allocated into 5 groups with 3 replicates per group, and fed with basal diet (negative control), basal diet supplemented with 30 mg/kg florfenicol (positive control), basal diet supplemented with 1000, 1500, and 2000 mg/kg bovine lactoferrin peptide, respectively. The experiment lasted for 8 weeks. The results showed that: 1) With the increase of the addition of bovine lactoferrin peptide, final body weight (FBW), weight gain rate (WGR) and specific growth rate (SGR) of largemouth bass showed a trend of first increasing and then decreasing. The 1000 mg/kg bovine lactoferrin peptide group had the best growth performance of largemouth bass, which was significantly different from the negative and positive control groups (P<0.05). 2) Compared with the negative and positive control groups, the intestinal trypsin, α-amylase and lipase activities of the 1000 mg/kg bovine lactoferrin peptide group of largemouth bass increased significantly (P<0.05). 3) Compared with the negative and positive control groups, the addition of 1000 mg/kg bovine lactoferrin peptide to feed could significantly increase the height and width of villi of the foregut, midgut and hindgut of largemouth bass (P<0.05). 4) After challenge with Aeromonas hydrophila, the survival rates of largemouth bass in the bovine lactoferrin peptide groups were higher than that of the negative control group, but there were no significant differences from the positive control group (P>0.05). In summary, under the experimental conditions, the addition of bovine lactoferrin peptide to feed could improve growth performance of juvenile largemouth bass, and the appropriate dosage was 1000 mg/kg. At the same time, bovine lactoferrin peptide could increase the intestinal digestive enzyme activity of largemouth bass, improve intestinal tissue structure, and improve disease resistance.
MORPHOLOGICAL AND MOLECULAR IDENTIFICATION OF 7 SPECIES OF DACTYLOGYRUS ON GILLS OF THE GOLDFISH (CARASSIUS AURATUS)
CHENG Jiang-Wen, PAN Ying-Zi, MA Xing-Rong, WANG Gui-Tang, LI Wen-Xiang
 Available online   , doi: 10.7541/2023.2022.0061
[Abstract](620) [FullText HTML](395) [PDF 7455KB](10)
Abstract:
Dactylogyrus species has high pathogenicity to the goldfish (Carassius auratus). In order to investigate the species of Dactylogyrus on the goldfish, we identified specimens of dactylogyrids collected from gills of the goldfish in Liangzi Lake, Hubei province based on morphological and molecular data in the present study. There were seven Dactylogyrus species found on gills of the goldfish, including D. vastator, D. intermedius, D. arcuatus, D. baueri, D. formosus, D. inexpeatatus and D. dulkeiti. The morphometric characteristics of the sclerotized parts of the opisthaptor in these dactylogyrids were measured and redescribed. D. intermedius, D. formosus and D. dulkeiti were identified by the higher than 99.0% similarities of 18S+ITS1+5.8S partial sequences to the related Dactylogyrus species from the GenBank. The sequence of D. arcuatus was obtained for the first time. However, the sequence similarity between D. vastator and the related Dactylogyrus species from the GenBank was only 96.37%. In addition, high Kimura-two-parameter genetic distances (0.004—0.058) were detected among D. vastator using partial 18S rDNA and ITS1sequences, but lower than the interspecific variability (0.046—0.064) between D. vastator and D. intermedius. In the present study, the morphological characteristics of seven species of Dactylogyrus on the gills of the goldfish were redescribed comprehensively and molecular classification markers were sequenced, which provided the basic data and a reference for the identification of Dactylogyrus species on the gills of the goldfish.
CLONING AND EXPRESSION ANALYSIS OF HSPIF FROM HYRIOPSIS SCHLEGELII
XIONG Ya-Ping, MA Hui-Mei, HU Bei-Juan, QIU Qi-Jun, HONG Yi-Jiang
 Available online   , doi: 10.7541/2023.2022.0038
[Abstract](461) [FullText HTML](349) [PDF 1537KB](4)
Abstract:
At present, there are few researches on the pearl breeding mechanism of the high-quality freshwater mussel Hyriopsis schlegelii, and the Pif gene is closely related to the formation of shellfish pearls. In order to explore the preliminary function of Pif gene of Hyriopsis schlegelii, the full-length cDNA sequence of Pif gene was obtained for the first time in Hyriopsis schlegelii by RACE-PCR technique, which is named HsPif. The full-length untranslated region (5'UTR) of 5'- terminal untranslated region of Pif gene was 485 bp, 3' untranslated region (3'UTR) and 363 bp, open reading frame (ORF) 3072 bp, which encodes a total of 1023 amino acids. It was speculated that the bioinformatics analysis of two proteins encoding HsPif97 and HsPif80showed that HsPif protein contained one von-Willebrand factor A (VWA) domain and three chitin binding domains (ChtBD2). The results of amino acid composition showed that the content of aspartic acid was the highest, accounting for 12.5%, and the content of histidine was the lowest, accounting for 1.32%. The α helix content in the protein secondary structure accounted for 19.63%, the irregular crimp accounted for 55.09%, and the extension chain accounted for 16.86%. The hydrophilic index of HsPif protein was -0.566. Phylogenetic tree analysis showed that HsPif and Hyriopsis cumingii Pif were the most conservative in one branch and on the same large branch as other shellfish. The results of tissue fluorescence quantitative PCR showed that HsPif gene was mainly expressed in the mantle, and in situ hybridization showed that the hybridization reaction mainly occurred in the mantle epithelial cells. The changes of HsPif gene expression were detected by qPCR after grafting. The results showed that HsPif gene expression changed with the development of pearl sac. The expression level of HsPif gene decreased significantly on the 7th and 90th day after nuclear insertion, and significantly increased on the 15th, 60th and 120th day. There was no significant change between the 30th day and the control group. The above experimental results suggest that HsPif gene may be related to the secretion and formation of pearls, which is helpful to further understand the mechanism of pearl formation and provide reference for pearl culture.
A TAXONOMIC STUDY OF EUGLENARIA FROM CHINA
JIANG Xiao-Die, WAN Shu-Ran, PANG Wan-Ting, WANG-QUAN-XI
 Available online   , doi: 10.7541/2023.2022.0079
[Abstract](455) [FullText HTML](398) [PDF 1416KB](6)
Abstract:
Euglenaria Karnkowska, Linton & Kwiatowski is a new genus isolated from Euglena Ehrenberg in 2010. Currently, there are 4species reported in the world. In China's existing algal taxonomic data, the species of Euglenaria are still classified in Euglena, and their morphological description is vague, making it difficult to distinguish them from some species of Euglena. It is difficult to identify Euglenaria and Euglena, and there is no obvious difference between Euglenaria and some species of Euglena in morphology. With the development of molecular biology, new technical means have been brought for the identification of euglenoid. While further research on the systematic classification of euglenoid has been carried out, the classification status of many euglenoid species has been revised, and Euglenaria is a new genus separated from Euglena at this stage. Our study found that the cells of Euglenaria are generally fusiform without mucocysts, and the Chloroplasts are disc-shaped with pyrenoids and edges irregular with deep grooves. The morphological differences among the species of Euglenaria are mainly reflected in the shape, number of chloroplasts and size of cells. The cells morphology of Eu. clepsydroides are hourglass shaped. The cells of Eu. caudata [length (58.95±12.38) μm, width (28.57±5.82) μm] were the largest. The chloroplast number of Eu. anabaena and Eu. clavata was significantly different. Eu. anabaena contained fewer chloroplasts (only 4±1), while Eu. clavata cells contained 10±3 chloroplasts. The morphological differences among species of Euglenaria are very subtle, and it is difficult to correctly identify them. The phylogenetic tree showed that with the closely related Eutreptiaceae as the outgroup, the results indicated that the rest of the euglenoids were divided into two major clades, the Euglenaceae clade and the Phacaceae clade. Euglenaceae (0.96/597) included Euglenaformis, Strombomonas, Trachelomonas, Cryptoglena, Euglena, Monomorphina, Euglenaria and Colacium. Each genus formed a well-supported evolutionary clade, respectively. The genus Euglenaria is located in the branch of Euglenaceae (0.84/-), consisting 28strains of Euglenaria, which are divided into 3subbranches, and the support rate of each subbranch exceed 99%. Eu1 clade contained 13strains of Eu. anabaena, Eu2 (1.00/998) contained 8strains Eu. caudata, and Eu3 (1.00/1000) contained 6strains Eu. clavata. Eu. anabaena and Eu. caudata were sister branches. However, there are great difference in cell size and chloroplast number between the two species. In morphology, Eu. anabaena and Eu. clavata were more similar, with similar cell size and large difference in chloroplast number. There were significant differences in cell size between Eu. caudata and Eu. clavata, but the number of chloroplasts was similar. This suggests that the relationship between morphology and molecular phylogeny may be inconsistent. At present, there are only 4species of Euglenaria and 162species of Euglena, and 80% species of Euglena have no molecular data. It is unknown whether there are still Euglena species belonging to Euglenaria among these species without molecular data, and further research is still needed. In this study, 12strains of Euglenaria collected from China were used as materials to study the phylogeny of Euglenaria based on morphological and molecular biological methods. We provide the Euglenaria species in morphological characters and molecular data and discusse the phylogenetic position of Euglenaria. In order to improve the taxonomic description of this group in China, we revised the taxonomic description of Euglenaria.
INHIBITORY EFFECT OF SCENEDESMUS SP. ON MICROCYSTIS AERUGINOSA AND ITS EVALUATION
SONG Chan-Yuan, BAI Fang, LI Tian-Li, SONG Li-Rong
 Available online   , doi: 10.7541/2023.2022.0042
[Abstract](449) [FullText HTML](343) [PDF 986KB](10)
Abstract:
Cyanobacterial harmful algal blooms are a global environmental problem. Microcystis, which can produce microcystin, is one of the most pervasive bloomforming cyanobacteria in freshwater ecosystems. Blooms of microcystis have adverse effects on lake ecology, and may effect human health, so it is important to control microcystis blooms. In order to investigate the inhibitory effect of green algae on Microcystis aeruginosa based on allelopathy among different algae strains, we screened potential green algae to control M. aeruginosa blooms. One strain which has allelochemicals production properties was screened from 34strains of green algae. The results showed that the spent medium of Scenedesmus sp. FACHB-1229had the highest inhibition rate, for M. aeruginosa FACHB-3550 was 53.95% and for FACHB-905 was 48.85%, respectively. And an analysis result of gas chromatography mass spectrometry (GC-MS) indicated that 1,2-Benzenedicarboxylic acid, bis(2-methoxyethyl) ester might be the allelochemical substances. In addition, we also measured growth rates, high ammonia nitrogen tolerance and photosynthetic oxygen evolution rates of the strain. The results indicate that Scenedesmus sp. FACHB-1229had high specific growth rate of (0.38±0.06) /d. It also had the highest photosynthetic oxygen evolution rates of (229.91±10.49) μmol O2/(mg Chl.a·h). And the ammonia nitrogen tolerance of Scenedesmus sp. FACHB-1229 was significantly higher than other strains, its growth rate was (0.30±0.08) /d when the ammonia nitrogen concentration reached up to 1888.60 mg/L. Moreover, the proportion of FACHB-1229 was consistently increased when co-cultured with FACHB-3550. In summary, FACHB-1229has the superiority in interspecies competition with M. aeruginosa, its spent medium can inhibit the growth of M. aeruginosa, and it also has high ammonia nitrogen tolerance and photosynthetic oxygen evolution rate. Therefore, this study has provided the theory and application basis in prevention and control of cyanobacterial blooms using interspecific competition among algae. FACHB-1229has the potential to control harmful algae through biological methods, it can grow rapidly in aquaculture pond or other polluted water bodies with high ammonia nitrogen concentration, while it can inhibit M. aeruginosa through allelopathy and provide dissolved oxygen at the same time.
THE INDUCTION AND IDENTIFICATION OF THE TRIPLOID YELLOW DRUM (NIBEA ALBIFLORA, SCIAENIDEA) AND CHARACTERIZATION OF THEIR GONADAL DEVELOPMENT
LU Lei, CHEN Rui-Yi, YANG Yang, ZHU Qi-Hui, HU Wei-Hua, LI Hai-Dong, XU Dong-Dong
 Available online   , doi: 10.7541/2023.2022.0003
[Abstract](483) [FullText HTML](338) [PDF 4337KB](7)
Abstract:
In this study, the cold shock treatment was used to induce triploid yellow drum (Nibea albiflora) and the ploidy was determined. The histology and gene expression analysis of reproduction-related genes were further used to characterize the gonadal development in triploid yellow drum. Firstly, the fertilized eggs were subjected to the cold shock treatment implemented at 3℃ for a period of 10 min commencing at post-fertilization 2.5min, and the fertilization rate and hatching rate in the cold shock treatment groups were (70.31±4.49)% and (21.5±6.63)%, respectively, which were significantly lower than that of diploid. Secondly, we found that the DNA relative content of triploid was 1.5 times larger than that of diploid by flow cytometer analysis, and the number of the chromosome in the triploid was 72, whereas the number of the chromosome in the diploid was 48shown by karyotype analysis. The rate of the triploid was 100% in the present study. Thirdly, the gonadosomatic indexes of the triploids were significantly lower than that of the diploids. Histological results showed that the gonadal development of the triploids was slower than that of the diploids. At the age of 12 months, the testis and ovary of the diploids were both at the stage V, while the testis and ovary of the triploids were at stage Ⅲ and stage Ⅰ, respectively. Fourthly, the dmrt1 and vasa expression in the triploid testis and cyp19a expression in the triploid ovary were significantly lower than those in the diploid gonads (P<0.05), and the vasa expression in the triploid ovary was also lower than that in the diploids (P>0.05). Taken together, our results indicated that the triploid yellow drum could be successfully induced by the cold shock treatment, and the gonadal development of the triploids was retarded. The results of present study will lay a basis for the application of the triploid in germ cell transplantation and the genetic improvement of yellow drum.
CHEMOTACTIC ACTIVITY AND IMMUNE ADJUVANT EFFECT OF GRASS CARP ΒETA DEFENSIN 1
KUANG Rui, ZHOU Jian-Cheng, LIU Xiao-Ling, SU Jian-Guo, YUAN Gai-Ling
 Available online   , doi: 10.7541/2023.2022.0022
[Abstract](423) [FullText HTML](390) [PDF 0KB](0)
Abstract:
β-defensin (BD) is a small cationic peptide rich in cysteine, which plays an important role in host defense and innate immunity.. BDs exist widely in all kinds of organisms in nature, and were initially concerned and studied extensively due to its spectral antibacterial activity. With in-depth research, it has been found in recent years that BD not only has direct antiviral microbial activity, but also plays an important regulatory role in natural immunity. At present, there are few studies on the immunomodulatory function of fish defensin. The study on grass carp BD still remained in the analysis of antibacterial activity. In this study, the antibacterial function of grass carp β-defensin 1 (CiBD1) was verified, and its chemochemogenic activity and adjuvant effect on Aeromonas hydrophila inactivated vaccine were explored, which can provide new insights for the prevention and treatment of grass carp bacterial disease. Relevant experimental results are as follows:1. CiBD1 is a class of cationic amphiphilic proteins with highly conserved three β-folded structures.Through Clustal Omega multi-sequence alignment and I-Tasser secondary structure prediction, it is found that grass carp β-defensin 1 (Ctenopharyngodon idella β-defensin 1, CiBD1) is a kind of amphiphile cationic peptide rich in cysteine. Its structure is highly conserved in bony fishes.2. CiBD1 is constitutively distributed in all tissues of grass carp and highly expressed in some mucosal tissues.In order to explore the physiological role of CiBD1, we studied the transcription level of CiBD1 gene in different tissues of grass carp under normal physiological conditions, and the results showed that skin and eye were the two tissues with the highest level of CiBD1 transcription before infection. The transcription level of CiBD1 in all tissues of grass carp infected with Aeromonas hydrophila at 72h was significantly different from that before infection, which showed that the transcription level of CiBD1 in tissues related to mucosal immunity, such as skin, eyes, swim bladder and hindgut, was significantly higher.3. The recombinant CiBD1 protein was successfully expressed by the prokaryotic expression system of Escherichia coli, and the antibacterial activity of CiBD1 was verified in vitro.The prokaryotic expression vector PET-32A-CIBD1 was constructed and induced by IPTG. The recombinant CiBD1 protein was purified by Ni2+ affinity chromatography and digested by enterocyte kinase. The antibacterial activity of CiBD1 was verified by CFU plate method. It has significant inhibitory activity against gram-negative bacteria (G+) and Gram-positive bacteria (G-).4. CiBD1has chemotactic activity of leukocytes in vitro, which can enhance the protective effect of Aeromonas hydrophila inactivated vaccine on grass carp, and has the potential to be used as an immune adjuvant.Chemotactic experiments showed that CiBD1had the best chemotactic activity on primary white blood cells of grass carp at 50 ng/mL. The immune adjuvant effect of CiBD1 recombinant protein on Aeromonas hydrophila inactivated vaccine was investigated by individual experiment. The mortality rate of the group with CiBD1 adjuvant was significantly lower than that of the group with vaccine alone, and the levels of IgM and MHC Ⅱ transcription and the contents of complement 3 (C3) and lysozyme in serum were significantly higher than those of the control group.In conclusion, CiBD1 is a class of highly conserved small molecule cationic antimicrobial peptide. CiBD1 is distributed in various tissues of grass carp in a constituent form and is highly expressed in some mucosal tissues. When the body is stimulated by pathogens or environmental stimuli, CiBD1 transcription level expression in some tissues in close contact with the peripheral environment is more significant. CiBD1 recombinant protein not only had certain killing activity against bacteria in vitro, but also showed strong chemotactic activity against lymphocyte and myeloid cells of grass carp. CiBD1 can improve the protection rate and disease resistance of Aeromonas hydrophila inactivated vaccine, and has a good immune adjuvant effect. This study indicated that CiBD1 recombinant protein plays an important role in bacterial anti-infection immunity and has a certain application prospect in aquaculture.