Oriental River Prawn Macrobrachium Nipponense Research Articles

Prospects for the Spread of the Invasive Oriental River Prawn Macrobrachium nipponense: Potentials and Risks for Aquaculture in Europe

Climate change has amplified the threat posed by aquatic invasive species as potential disruptors of biodiversity and ecosystem functioning. Species Distribution Models (MaxEnt) based on original data and ecological variables have identified contemporary seven global centers of the oriental rivel prawn Macrobrachium nipponense distribution: the native range in East Asia, Northern, Western and Eastern Europe, the Irano-Turanian region, and North and South America. By 2050, further expansion in Europe is expected, likely due to climate change, particularly temperature changes (Bio1) and rain precipitation during the warmest quarter (Bio18). However, the species may see a range reduction in southern Europe due to lower precipitation and increased droughts related to climate change. Therefore, a northward shift in the range of the species is also predicted. In the context of global change, and especially biological invasions, this study highlights the risks of introducing aquaculture based on M. nipponense and recommends controlling such economic activities, which are associated with a high risk for native species and ecosystems. Further, long-term monitoring is needed to assess impacts and to efficiently manage M. nipponense populations that are already present in their non-native habitats, for mitigating their negative effects on native species and ecosystems worldwide.

Safe concentration, unsafe effects: Impact of BPA on antioxidant function in the hepatopancreas and ovarian gene expression in oriental river prawns (Macrobrachium nipponense)

This study investigated the effects of Bisphenol A (BPA), a common endocrine-disrupting chemical, on the antioxidant enzyme activities in the hepatopancreas and the expression of genes related to ovarian development in oriental river prawns (Macrobrachium nipponense). The 24hLC50 and 48hLC50 values for BPA were 80.59 mg/L and 63.90 mg/L, respectively, with a safe concentration of 12.06 mg/L. Prawns were exposed to low (4.85 mg/L), safe (12.06 mg/L), and high (30.00 mg/L) concentrations of BPA for 10 days to measure enzyme activities, and for 20 days followed by 7 days in BPA-free water to measure gene expression.Short-term exposure (12 h, 1d, 3d) to low concentration BPA did not significantly affect superoxide dismutase (SOD) activity in the hepatopancreas (P > 0.05), but long-term exposure (6d, 10d) significantly reduced SOD activity (P < 0.05). Catalase (CAT) activity showed no significant changes throughout the low concentration exposure period (P > 0.05). At safe and high concentrations, SOD and CAT activities significantly decreased after 12 h of exposure (P < 0.05).BPA affected heat shock protein 90 (HSP90) expression in the ovary, with low concentration BPA significantly upregulating HSP90 after 1 day (P < 0.05), but returning to normal levels after 10 and 20 days. At the safe concentration, HSP90 was significantly upregulated at all three sampling points (1d, 10d, 20d) (P < 0.05), while high concentration exposure led to significant upregulation only on day 10 (P < 0.05).Low concentration BPA had no significant effect on Cathepsin B (CB) and Cathepsin L (CL) gene expression in the ovaries (P > 0.05). However, safe concentration exposure promoted CB expression on days 1, 10, and 20 (P < 0.05), while high concentration exposure significantly increased CB expression on day 1 (P < 0.05), with levels returning to normal on days 10 and 20. CL expression significantly increased after 20 days of exposure to both safe and high concentrations (P < 0.05). Gene expression levels in the ovaries returned to normal after transfer to BPA-free water, with HSP90 and CB normalizing by day 1, and CL by day 7.These results indicate that even safe concentrations of BPA impose stress on the hepatopancreas and increase the expression of HSP90, CB, and CL genes in the ovaries, affecting ovarian development. And, these effects are reversible within a certain period after the removal of BPA.

Comparative transcriptome analysis reveals the different responding mechanisms of ovary and hepatopancreas following polyI:C challenge in Macrobrachium nipponense

The ovary in mammals has developed specialized mechanisms for protection against pathogen infections; however, the understanding of the innate immune system in the ovary of crustaceans is still limited. To elucidate the ovary's defense mechanisms in response to viral challenges, we subjected oriental river prawns (Macrobrachium nipponense) to poly I:C, a double-stranded RNA analog that emulates viral dsRNA, and analyzed the ovary's transcriptome profiles. Concurrently, RNA-seq analysis was performed on the hepatopancreas, a well-recognized immune-related tissue, following poly I:C challenge to investigate the distinct response mechanisms of the ovary and hepatopancreas and to gain a comprehensive understanding of the immune responses in both tissues. The results indicate that 1368 genes are differentially expressed in the ovary, with 903 genes upregulated and 465 genes downregulated. Subsequent analysis reveals that these differentially expressed genes (DEGs) include numerous genes associated with innate immunity, such as members of the C-type lectin, fibrinogen-related protein (Frep), Toll-like receptor, and NOD-like receptor (NLR) gene families, as well as acid phosphatase, scavenger receptor, crustin, Down syndrome cell adhesion molecule (Dscam), hemocyanin, and lipopolysaccharide and beta-1,3-glucan binding protein (LGBP). Furthermore, the DEGs include several genes related to ovary development, such as sox8, vitellogenin, progranulin, cyclin-dependent kinase, ecdysone receptor, frizzled, and members of the Fox gene family. In the hepatopancreas, a total of 729 DEGs were identified. Comparison of the DEGs in both tissues indicates that only 91 genes are common to both groups, highlighting significant tissue-specific responses to poly I:C stimulation. This study aims to enhance our understanding of the immune protective mechanisms employed by the ovary in response to pathogen exposure and establishes a foundation for investigating ovarian reproductive immunity in crustaceans.

Dietary fishmeal substitution with Antarctic krill meal improves the growth performance, lipid metabolism, and health status of oriental river prawn (Macrobrachium nipponense)

Aims to assess the effects of dietary inclusion of Antarctic krill meal (KM), five experimental diets supplemented with graded levels of KM (0 %, FM; 5 %, KM5; 10 %, KM10; 15 %, KM15; 20 %, KM20; 0 %, 13.3 %, 26.7 %, 40.0 %, and 56.7 % replacement fishmeal levels, respectively), were prepared to feed oriental river prawn (Macrobrachium nipponense) (0.20 ± 0.01 g) for 8 weeks. The results revealed significant improvements in weight gain rate (WGR), specific growth rate (SGR), feed conversion rate (FCR), and protein efficiency ratio (PER) at the inclusion of 5 %-20 % KM (P < 0.05). In the KM10 and KM15 groups, the concentrations of hemolymph total protein (TP), albumin (ALB), triglycerides (TG), high-density lipoprotein (HDL), and hepatopancreatic glutathione (GSH) were observed to be significantly higher compared to the FM group (P < 0.05). Conversely, the contents of hepatopancreatic malondialdehyde (MDA), total nitric oxide synthase (TNOS), and inducible nitric oxide synthase (iNOS) were significantly lower in the KM10 and KM15 groups compared to the FM group (P < 0.05). The mRNA expression levels of dorsal protein (dorsal), heat shock protein 60 (hsp60), and myeloid differentiation factor 88 (myd88) in the hepatopancreas in the KM10 group exhibited the lowest level. Additionally, compared to the FM group, the expression levels of fatty acid synthase (fas) and acetyl CoA carboxylase (acc) were significantly upregulated (P < 0.05), while the acyl CoA bindingprotein (acbp), carnitine palmitoyltransferase I (cpt I), and scavenger receptor B I (sr-b I) were significantly downregulated in the KM10 group (P < 0.05). In addition, the results of histopathology sections showed that 5–15 % KM could improve the health status of hepatopancreas and was a safe alternative to fishmeal. Overall, the optimal KM inclusion level in feed was 12.96 % (substituting for 34.60 % fishmeal) based on the quadratic fitting curve calculations of WGR and SGR.

Molecular characterization and functional identification of the insulin-like peptides receptor gene in the oriental river prawn Macrobrachium nipponense

The insulin-like signaling pathway plays diverse biological roles in regulating metabolism, immune responses, growth, and development in invertebrates. The insulin-like peptides (ILP) receptor (ILPR) is the pivotal protein in any insulin-like signaling pathway. In our previous study, we cloned the Mn-ILP gene from the oriental river prawn Macrobrachium nipponense and elucidated its function. However, the ILPR has not yet been described in M. nipponense. In the present study, we further examined the function of Mn-ILPR in M. nipponense. The full length of the Mn-ILPR mRNA was 4790 bp, encoding 1412 amino acids. Mn-ILPR was a transmembrane receptor under a subfamily of receptor tyrosine kinases. The Mn-ILPR mRNA was predominantly expressed in muscle and hepatopancreas. To elucidate the function of Mn-ILPR, we investigated its expression in different growth stages of prawns of both sexes, in prawns of different sizes at the same time, and in prawns of the same size at different stages of the molt cycle using qRT-PCR. The expression of Mn-ILPR was markedly higher: (1) in the rapid growth stage than in other stages of male individuals; (2) in the normal growth stage than in the gonad development stage of female individuals; (3) in large male individuals than in small male individuals; and (4) in the intermolt stage than in other stages of the molt cycle. Furthermore, the long-term knockdown of Mn-ILPR by RNAi effectively delayed the growth speed of M. nipponense. Meanwhile, the short-term knockdown of Mn-ILPR revealed that the ILP-ILPR signal pathway played a part in the process of growth of M. nipponense. These findings provide a greater understanding of the molecular mechanisms underlying growth and development in crustaceans.

A novel exoskeletal-derived C-type lectin facilitates phagocytosis of hemocytes in the oriental river prawn Macrobrachium nipponense

C-type lectins (CTLs) execute critical functions in multiple immune responses of crustaceans as a member of pattern recognition receptors (PRRs) family. In this study, a novel CTL was identified from the exoskeleton of the oriental river prawn Macrobrachium nipponense (MnLec3). The full-length cDNA of MnLec3 was 1150 bp with an open reading frame of 723 bp, encoding 240 amino acids. MnLec3 protein contained a signal peptide and one single carbohydrate-recognition domain (CRD). MnLec3 transcripts were widely distributed at the exoskeleton all over the body. Significant up-regulation of MnLec3 in exoskeleton after Aeromonas hydrophila challenged suggested the involvement of MnLec3 as well as the possible function of the exoskeleton in immune response. In vitro tests with recombinant MnLec3 protein (rMnLec3) manifested that it had polysaccharide binding activity, a wide spectrum of bacterial binding activity and agglutination activity only for tested Gram-negative bacteria (Escherichia coli, Vibrio anguillarum and A. hydrophila). Moreover, rMnLec3 significantly promoted phagocytic ability of hemocytes against A. hydrophila in vivo. What's more, MnLec3 interference remarkably impaired the survivability of the prawns when infected with A. hydrophila. Collectively, these results ascertained that MnLec3 derived from exoskeleton took an essential part in immune defense of the prawns against invading bacteria as a PRR.

The physiological response of oriental river prawn Macrobrachium nipponense to starvation-induced stress

Environmental stresses play critical roles in the physiology of crustaceans. Food deprivation is an important environmental factor and a regular occurrence in both natural aquatic habitats and artificial ponds. However, the underlying physiological response mechanisms to starvation-caused stress in crustaceans are yet to be established. In the present study, the hepatopancreas tissue of Macrobrachium nipponense was transcriptome analyzed and examined for starvation effects on oxidative stress, DNA damage, autophagy, and apoptosis across four fasting stages (0 (control group), 7, 14, and 21 days). These results indicated that a ROS-mediated regulatory mechanism is critical to the entire fasting process. At the initial stage of starvation (fasting 0 d ~ 7 d), ROS concentration increased gradually, activating antioxidant enzymes to protect the cellular machinery from the detrimental effects of oxidative stress triggered by starvation-induced stress. ROS content production (hydrogen peroxide and superoxide anion) then rose continuously with prolonged starvation (fasting 7 d ~ 14 d), reaching peak levels and resulting in autophagy in hepatopancreas cells. During the final stages of starvation (fasting 14 d ~ 21 d), excessive ROS induced DNA damage and cell apoptosis. Furthermore, autophagolysosomes and apoptosis body were further identified with transmission electron microscopy. These findings lay a foundation for further scrutiny of the molecular mechanisms combating starvation-generated stress in M. nipponense and provide fishermen with the theoretical guidance for adopting fasting strategies in M. nipponense aquaculture.

A pattern recognition receptor interleukin-1 receptor is involved in reproductive immunity in Macrobrachium nipponense ovary

The family of TIR domain-containing receptors includes numerous proteins involved in innate immunity. In this study, a member of this family was characterized from the ovary of the oriental river prawn Macrobrachium nipponense and identified as interleukin-1 receptor (MnIL-1R). Meanwhile, to elucidate the conservation of IL-1R, its orthologous were identified in several crustacean species as well. In addition, the expression pattern of MnIL-1R in various adult tissues and post different pathogen-associated molecular patterns (PAMPs) challenge in ovary was analyzed with qRT-PCR technology. Finally, the roles of MnIL-1R in the ovary were analyzed by RNAi technology. The main results are as follows: (1) MnIL-1R comprises a 1785 bp ORF encoding 594 amino acids and is structurally composed of five domains: a signal peptide, two immunoglobulin (IG) domains, a transmembrane region, and a TIR-2 domain; (2) the TIR domain showed a high conservation among analyzed crustacean species; (3) MnIL-1R is widely detected in all tested tissues including ovary; (4) MnIL-1R showed a positive response to challenges with LPS, PGN, and polyI:C in the ovary; (5) its IG domain showed strong binding ability to LPS and PGN, confirming its role as a pattern recognition receptor; (6) the expression patterns of several members of the Toll signaling pathway (Myd88, TRAF-6, Dorsal, and Relish) was similar to that of MnIL-1R after challenges with LPS, PGN, and polyI:C in the ovary; (7) the silencing of MnIL-1R resulted in down-regulation of theses gene’ (Myd88, TRAF-6, Dorsal, and Relish) expression level in the ovary. These results suggest that MnIL-1R can activate the Toll signaling pathway in the ovary by directly recognizing LPS and PGN through its IG domain, thereby contributing to the immune response in the ovary of M. nipponense.

First report of Metschnikowia bicuspidata infection in the oriental river prawn (Macrobrachium nipponense, de Haan) in China.

During breeding, some oriental river prawns (Macrobrachium nipponense, de Haan), an important aquaculture species in China, exhibit yellowish-brown body colouration, reduced appetite, and vitality. Diseased prawns revealed characteristic emulsifying disease signs, including whitened musculature, hepatopancreatic tissues, milky haemolymph, and non-coagulation. The present study investigated the causative agent of M. nipponense infection through isolation, histopathology, molecular sequencing, and infection experiments. The pathogenic strain exhibited distinctive white colonies on Bengal red medium, with microscopic examination confirming the presence of yeast cells. Histopathological analysis revealed prominent pathological alterations and yeast cell infiltration in muscles, hepatopancreas and gills. Additionally, 26S rDNA sequencing of the isolated yeast strain LNMN2022 revealed Metschnikowia bicuspidata (GenBank: OR518659) as the causative agent. This strain exhibited a 98.28% sequence homology with M. bicuspidata LNMB2021 (GenBank: OK094821) and 96.62% with M. bicuspidata LNES0119 (GenBank: OK073903). The pathogenicity test confirmed that M. bicuspidata elicited clinical signs in M. nipponense consistent with those observed in natural populations, and the median lethal concentration was determined to be 3.3 × 105 cfu/mL. This study establishes a foundation for further investigations into the host range and epidemiological characteristics of the pathogen M. bicuspidata in aquatic animals and provides an empirical basis for disease management in M. nipponense.

The identification of a serpin with immune defense role in oriental river prawn Macrobrachium nipponense

Serpins are a protein superfamily of serine protease inhibitors. One of their functions is to participate in immune responses by inhibiting the activation of prophenoloxidase. To elucidate the immune role of serpin in Macrobrachium nipponense, a serpin gene (Mnserpin) was cloned from M. nipponense in this study. Mnserpin protein has an N-terminal signal peptide and a serpin domain that contains a hinge region, a signature sequence of serpin and a P1(arginine)-P1’ scissile bond, and evolutionally closely related to the crustacean serpins. Mnserpin highly expressed in the hepatopancreas and gill. Mnserpin expression increased first and then decreased after Vibrio parahaemolyticus and Aeromonas hydrophila infection, and was knocked down by dsMnserpin injection with a maximum knockdown efficiency of 92 %. Mnserpin knockdown increased the expression of the clip domain serine protease and prophenoloxidase genes and phenoloxidase activity of M. nipponense as well as its mortality rate after V. parahaemolyticus and A. hydrophila infection. The recombinant Mnserpin (rMnserpin) showed bacteria-binding and bacteriostatic activity in vitro. Moreover, rMnserpin injection decreased the bacterial number and the mortality rate of M. nipponense post V. parahaemolyticus and A. hydrophila infection. These results suggested that Mnserpin plays a major role in the innate immune response of M. nipponense.

Effects of Alkalinity Exposure on Antioxidant Status, Metabolic Function, and Immune Response in the Hepatopancreas of Macrobrachium nipponense.

The oriental river prawn Macrobrachium nipponense is an important freshwater economic species in China, producing huge economic benefits. However, M. nipponense shows lower alkali tolerance than fish species, thus genetic selection is urgently needed in order to improve alkali tolerance in this species. In the present study, the effects of alkalinity exposure on the hepatopancreas of M. nipponense were measured under the alkali concentrations of 0 (control), 4, 8, and 12 mmol/L with the exposure time of 96 h through histological observations, measurement of antioxidant enzymes, metabolic profiling analysis, and transcriptome profiling analysis. The present study identified that the low concentration of alkali treatment (<4 mmol/L) did not result in morphological changes in the hepatopancreas and activity changes in antioxidant enzymes, while high-alkali treatment (>8 mmol/L) damaged the normal structures of the lumen and vacuoles and significantly stimulated the levels of superoxide dismutase, catalase, and total antioxidant capacity, indicating these antioxidant enzymes play essential roles in the protection of the body from the damage caused by the alkali treatment. Metabolic profiling analysis revealed that the main enriched metabolic pathways of differentially expressed metabolites in the present study were consistent with the metabolic pathways caused by environmental stress in plants and other aquatic animals. Transcriptome profiling analysis revealed that the alkali concentration of <8 mmol/L did not lead to significant changes in gene expression. The main enriched metabolic pathways were selected from the comparison between 0 mmol/L vs. 12 mmol/L, and some significantly up-regulated genes were selected from these metabolic pathways, predicting these selected metabolic pathways and genes are involved in the adaptation to alkali treatment in M. nipponense. The expressions of Ras-like GTP-binding protein, Doublesex and mab-3 related transcription factor 1a, and Hypothetical protein JAY84 are sensitive to changes in alkali concentrations, suggesting these three genes participated in the process of alkali adaptation in M. nipponense. The present study identified the effects of alkalinity exposure on the hepatopancreas of M. nipponense, including the changes in antioxidant status and the expressions of metabolites and genes, contributing to further studies of alkali tolerance in this species.

Залежність лінійного зростання та виживання личинок субтропiчної прiсноводної креветки (Мacrobrachium nipponense Dе Haan, 1849) від температури і солоності

Purpose. Studies of the growth and survival of Oriental river prawn (Macrobrachium nipponense) larvae depending on water temperature and salinity during cultivation. Methodology. Experimental studies were carried out at the Aquatic Department of Aquatic Bioresources and Aquaculture of the Odesa State Ecological University in 2020-2021. Brood Oriental river prawns (Macrobrachium nipponense) were caught in the lower Dniester river and placed into a 0.8 m3 RAS. The larvae were stocked in separate aquariums with automatically maintained specified temperature regime. The first series of experiments in fresh water investigated the influence of water temperatures of 20–22, 22–24, 24–26, and 28–31°C on the growth of shrimp larvae (from the 1st stage to the Pl stage). Cultivation in water with a salinity of 5, 7, 12 ‰ was carried out at a temperature of 20–22, 22–24, 28–31 °C. Ten prawn larvae were measured daily under a binocular microscope (MBS-10) using an eyepiece-micrometer. The stage of larval development and the survival rate were determined. Water salinity was determined using an ATAGO-100 refractometer, and an Azha-101M thermal oximeter was used to determine the oxygen content in water. Statistical processing of data was carried out in Microsoft Excel. Findings. It was established that at a temperature of 24–26°C, the duration of the transition from one stage of larval development to another in Oriental river prawn (Macrobrachium nipponense) increased as the larvae grew and developed. It lasted 4–5 days in the first stages (1-3), then stages from 6 to 9 days in the later stages. High temperature stimulated the growth of larvae and accelerated their development. In fresh water at a temperature of 29–31°C, the length of postlarvae (Pl) reached 6.69±1.15 mm in 28 days of rearing. Cultivation at high temperature resulted in a significant variation in larval sizes (from 4.4 to 8.6 mm CV–25,27). At lower temperatures (25-27, 22-24 and 20-22°C), the average sizes of prawn postlarvae were smaller (5.83±0.78, 4.56±0.15, 4.43±0.15), and the survival rates increased (46, 49 and 54%, respectively). Survival of larvae was inversely dependent on temperature r = –0.89. It was minimal (32%) at a temperature of 29–31°C. The higher the growing temperature, the lower the output of postlarvae. Water salinity, like temperature, significantly affected the growth, survival and size uniformity of postlarvae. In water with a salinity of 5‰, postlarvae reached their maximum length (6.56±0.15 mm) at a temperature of 29-31 °C. At temperatures of 20-22 and 22-24 °C, the size of the postlarvae did not reliably differ among themselves. The maximum survival of postlarvae (52%) was noted at a temperature of 22-24°. Regardless of water temperature, salinity of 5‰ ensured high uniformity of larvae size (CV: 6.11–9.09). At a salinity of 7‰, the maximum length of postlarvae was reached at a temperature of 29-31oC, and the highest survival rate of prelarvae (34%) was noted at a temperature of 20-22oC. The size of the larvae and their survival at other growing temperature conditions did not differ significantly (Р&lt;95). A salinity of 7‰ also ensured the uniformity of postlarvae sizes (CV: 6.12-8.97). At a salinity of 12‰, high water temperature stimulated the growth of larvae with relative uniformity of their linear sizes, but in all variants of the experiment the survival rate of postlarvae was very low (4–9%). Originality. Experimental data on the growth and survival of larvae of the Oriental river prawn (Macrobrachium nipponense) depending on temperature and salinity are presented for the first time. The influence of the salinity of the environment on the size uniformity of the larvae was established. Practical value. The obtained results can be used for the development and improvement of methods of artificial reproduction and cultivation of the Oriental river prawn (Macrobrachium nipponense) under controlled conditions in recirculation aquaculture systems (RAS).

Dietary postbiotics supplementation improves growth, survival rate, antioxidant capacity, non-specific immunity and gut health of juvenile oriental river prawn (Macrobrachium nipponense)

Antibiotics use in aquaculture cause multiple adverse effects on aquatic animal health. Accordingly, studies on alternative approaches to antibiotics in aquaculture are urgently required. This study investigated the effects of dietary postbiotics as an alternative to florfenicol antibiotic on growth performance, survival rate, antioxidant capacity, non-specific immunity, and gut health of juvenile oriental river prawn (Macrobrachium nipponense). Seven isonitrogenous and isocaloric experimental diets were formulated containing 0% (control), 0.25%, 0.5%, 1%, 2% and 4% postbiotics. These experimental diets were fed to three replicates of 70 juvenile prawns (initial weight 0.110 ± 0.010 g) for eight weeks. In addition, a diet containing 0.2% florfenicol (FFC0.2) was included as the antibiotic treatment, which was fed to the prawns for two weeks. Results showed that feeding the prawns on the postbiotics diets improved weight gain and survival rate. The prawns fed on the antibiotics or postbiotics diets increased antioxidant capacity in the hepatopancreas. Moreover, the prawns fed on the postbiotics diets increased significantly non-specific immunity. The optimum amount of postbiotics required for maximum non-specific immunity was 0.76%, 0.86% and 2.85% based on acid phosphatase, alkaline phosphatase and phenoloxidase activities, respectively. The prawns fed on the postbiotics diets upregulated significantly the mRNA expression of immunity related genes, which were downregulated significantly in the prawns fed on the FFC0.2 diet. Furthemore, the prawns fed on the 1% and 2% postbiotics diets had longer microvilli with closely packed intestinal epithelial cell and basement membrane. In summary, supplementing 0.76 to 2.66% postbiotics in the diets for M. nipponense improves growth performance, survival rate, antioxidant capacity, non-specific immunity and gut health compared with florfenicol antibiotic.

Krill oil showed a good application potential as the dietary lipid source on the growth, physiology, nutritional quality, and flavor of the edible meat of the oriental river prawns (Macrobrachium nipponense)

An 8-week feeding trial was conducted to evaluate the effects of dietary krill oil (KO), fish oil (FO) and soybean oil (SO) on growth, nutritional quality and flavor of the edible meat of the oriental river prawns (Macrobrachium nipponense). The results revealed that SO and KO diets promoted the growth performance (P < 0.05). The activity of alanine aminotransferase in hemolymph of the FO group was significantly lower than that in the SO group (P < 0.05). The contents of total cholesterol in hemolymph in the SO and FO groups were significantly lower than those in the KO group (P < 0.05). Superoxide dismutase activity and glutathione content in the hepatopancreas were the lowest in the SO group (P < 0.05). Lipoprtein lipase and trypsin activities of the hepatopancreas decreased significantly in the KO group (P < 0.05). Among the fatty acids in meat, the contents of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) increased significantly in M. nipponense after feeding on animal lipid sources (FO and KO) (P < 0.05). The contents of umami and bitter free amino acids in the SO and KO groups were significantly higher than those in the FO group (P < 0.05), and the contents of adenosine monophosphate and inosine monophosphate were highest in the KO group (P < 0.05). The contents of bitter nucleotides hypoxanthine and hypoxanthine ribonucleoside were lowest in the KO group (P < 0.05). In general, compared with FO, the use of KO and SO as lipid sources in feed promoted the growth of M. nipponense, and KO significantly improved the antioxidant index of the hepatopancreas. FO and KO increased the contents of EPA and DHA in the edible meat of M. nipponense, and KO further enhanced the contents of amino acids and nucleotides responsible for flavor in the meat.

Identification of Male Sex-Related Genes Regulated by SDHB in Macrobrachium nipponense Based on Transcriptome Analysis after an RNAi Knockdown.

The oriental river prawn (Macrobrachium nipponense) is a commercially important species in Asia. A previous study showed that the succinate dehydrogenase complex iron sulfur subunit B (SDHB) gene participates in testes development in this species through its effect on the expression of the insulin-like androgenic gland hormone gene. This study knocked-down the Mn-SDHB genes in M. nipponense using RNAi. A transcriptome analysis of the androgenic gland and testes was then performed to discover the male sex-related genes regulated by SDHB and investigate the mechanism of male sexual development in this species. More than 16,623 unigenes were discovered in each sample generated. In the androgenic gland, most of the differentially expressed genes were enriched in the hypertrophic cardiomyopathy pathway, while in the testes, they were enriched in the citrate cycle pathway. In addition, after Mn-SDHB knockdown, five genes were found to be downregulated in the androgenic gland in a series of biological processes associated with phosphorylated carbohydrate synthesis and transformations in the glycolysis/gluconeogenesis pathway. Moreover, a total of nine male sex-related genes were identified including Pro-resilin, insulin-like androgenic gland hormone, Protein mono-ADP-ribosyltransferase PAPR11, DNAJC2, C-type Lectin-1, Tyrosine-protein kinase Yes, Vigilin, and Sperm motility kinase Y-like, demonstrating the regulatory effects of Mn-SDHB, and providing a reference for the further study of the mechanisms of male development in M. nipponense.

Combined effects of dietary carbohydrate levels and ammonia stress on growth, antioxidant capacity and glucose metabolism in juvenile oriental river prawn (Macrobrachium nipponense).

Ammonia is a common environmental stress factor that constrains aquaculture industry development. This study evaluated the effect of carbohydrate levels and ammonia stress in oriental river prawn (Macrobrachium nipponense). The experiment had six treatments containing two water ammonia levels (0 and 5 mg/L) and three dietary carbohydrate levels (low carbohydrate diet (LCD, 10%), medium carbohydrate diet [MCD, 20%], and high carbohydrate diet [HCD, 30%]), and lasted six weeks. The results showed that the prawns fed on MCD had higher weight gain than those fed on LCD and HCD during ammonia stress. Moreover, the prawns fed on MCD had significantly lower acid phosphatase and alkaline phosphatase activities during ammonia stress. Feeding the prawns on the MCD increased B cells in the hepatopancreas during ammonia stress. Interestingly, the prawns fed on MCD had significantly lower superoxide dismutase activity compared to LCD and HCD during ammonia stress. Moreover, the prawns fed on MCD had significantly lower pyruvate kinase activity and pyruvate and lactic acid contents, while those fed on LCD had significantly higher succinic dehydrogenase, 6-phosphogluconic dehydrogenase, and phosphoenol pyruvate carboxykinase activities during ammonia stress. The prawns fed on the MCD increased significantly glutaminase activity and decreased the ammonia content in the serum during ammonia exposure. In addition, feeding the prawns on MCD decreased significantly the expression of apoptosis and inflammation-related genes. Taken together, the MCD supplied energy required to counteract ammonia stress, which increased growth, improved antioxidant capacity, facilitated ammonia excretion, and alleviated inflammation and apoptosis of the oriental river prawn.

Effects of copper on gill function of juvenile oriental river prawn (Macrobrachium nipponense): Stress and toxic mechanism

As an important trace element and the accessory factor of many enzymatic processes, heavy metal copper is essential to aquatic animals. The toxic mechanism of copper on gill function of M. nipponense was clarified for the first time in terms of histopathological analysis, physiology, biochemistry and the expression of important genes. The results obtained by present in present research showed that heavy metal copper could affect normal respiratory and metabolic activities in M. nipponense. Copper stress could cause damage to the mitochondrial membrane of gill cells in M. nipponense, and the activity of mitochondrial respiratory chain complex could be inhibited by copper. Copper could affect normal electron transport and mitochondrial oxidative phosphorylation, resulting in the inhibition of energy production. High concentrations of copper could disrupt intracellular ion balance and induce cytotoxicity. The oxidative stress could be induced by copper, leading to excessive ROS. Copper could reduce the mitochondrial membrane potential, lead to the leakage of apoptotic factors, and induce apoptosis. Copper could damage structure of gill, affect normal respiration of gill. This study provided fundamental data for exploring impacts of copper on gill function in aquatic organisms and potential mechanisms of copper toxicity.

Characterization and immune functional analysis of two new type I crustins in the oriental river prawn Macrobrachium nipponense

Crustins are cationic antimicrobial peptides that are categorized into five types and play a key role in the nonspecific immunity of crustaceans. In the present study, full-length cDNA sequences of two Macrobrachium nipponense crustin genes (MnCrustin1 and MnCrustin2) were cloned and identified. The full-length cDNAs of MnCrustin1 and MnCrustin2 were 715 and 881 bp, including 351 and 330 bp open reading frames (ORFs) encoding 117 and 110 amino acids, respectively. A signal peptide, a cysteine (Cys)-rich region and a whey acidic protein (WAP) domain existed in both MnCrustin1 and MnCrustin2. Phylogenetic analysis and multiple sequence alignment indicated that MnCrustin1 and MnCrustin2 belonged to the type I crustin family. The MnCrustin1 and MnCrustin2 mRNAs were expressed in various tissues of M. nipponense and were highly expressed in hemocytes and gills. Recombinant MnCrustin1 and MnCrustin2 inhibited the growth of both gram-positive and gram-negative bacteria in vitro. After Aeromonas hydrophila and white spot syndrome virus (WSSV) infection, MnCrustin1 and MnCrustin2 expression levels were elevated in the hepatopancreas, hemocytes and gills of M. nipponense. Tests of RNA interference (RNAi) and recombinant protein injection were employed to further study the roles of MnCrustin1 and MnCrustin2 in pathogen infection. The results indicated that silencing MnCrustin1 and MnCrustin2 accelerated the mortality rate of M. nipponense after challenge with A. hydrophila or WSSV, while recombinant MnCrustin1 and MnCrustin2 decreased the mortality rate under these conditions. These results suggest that MnCrustin1 and MnCrustin2 both play prominent roles in the immune response of M. nipponense to bacterial and viral infection and are important immune effectors of this prawn against pathogen infection.

Polystyrene nanoplastics decrease nutrient accumulation, disturb sex hormones, and inhibit reproductive development in juvenile Macrobrachium nipponense

The biological effects of nanoplastics has grown exponentially over the past few years. However, little is known about the effects of nanoplastic exposure on gonadal development in crustaceans. Thus, juvenile oriental river prawns (Macrobrachium nipponense) were exposed to different concentrations of 75-nm polystyrene nanoplastics (0, 5, 10, 20, and 40 mg/L) for 28 days to study the effects of exposure to nanoplastics on gonadal development. The genes encoding extracellular signal-regulated kinase (ERK) and mitogen-activated protein kinase kinase (MEK) were selected and the nutrient composition, sex hormone level, and gonad development-related gene expression were determined. Crude lipid and crude protein decreased with exposure to higher concentrations of nanoplastics, whereas there were no significant differences in levels of ash or moisture (P > 0.05). Full-length Mn-ERK and Mn-MEK cDNAs were cloned from M. nipponense and homologous comparisons showed that the genes had conserved functional sequences and had evolved consistently in invertebrates. With nanoplastics concentration increased, the serum sex hormone (estradiol, progesterone, and testosterone) levels of juvenile shrimp first increased and then decreased. In addition, the expression of gonad development-related genes (Vitellogenin, Vitellogenin receptor, Cyclin B, Gametocyte Specific Factor 1, Vasa, and PL10), MEK and ERK initially increased and then decreased with increasing nanoplastic concentration. This suggests that polystyrene nanoplastics reduce the accumulation of nutrients and lead to suppression of gonadal development in juvenile M. nipponense and, thus, provides basic information on the toxic effects of nanoplastics that could extend to other crustaceans.

RNAi Analysis of Potential Functions of Cyclin B3 in Reproduction of Male Oriental River Prawns (Macrobrachium nipponense).

Cyclin B3 (CycB3) is involved in the metabolic pathway of the cell cycle, playing essential roles in the regulation of cell proliferation and mitosis. CycB3 is also predicted to be involved in the reproduction of male oriental river prawns (Macrobrachium nipponense). In this study, the potential functions of CycB3 in M. nipponense were investigated using quantitative real-time PCR, RNA interference, and histological observations. The full-length DNA sequence of CycB3 in M. nipponense was 2147 base pairs (bp) long. An open reading frame of 1500 bp was found, encoding 499 amino acids. A highly conserved destruction box and two conserved cyclin motifs were found in the protein sequence of Mn-CycB3. Phylogenetic tree analysis revealed that this protein sequence was evolutionarily close to that of CycB3s of crustacean species. Quantitative real-time PCR analysis results suggested that CycB3 was involved in the process of spermiogenesis, oogenesis, and embryogenesis in M. nipponense. RNA interference analysis showed that CycB3 had a positive regulatory relationship with insulin-like androgenic gland hormone (IAG) in M. nipponense. In addition, sperm were rarely observed in the testis of double-stranded CycB3-injected prawns after 14 days of treatment, and sperm abundance was dramatically lower than that in the double-stranded GFP-injected prawns on the same day. This result indicated that CycB3 can regulate the testis reproduction in M. nipponense through inhibiting the IAG expressions. Overall, these results indicated that CycB3 plays essential roles in the regulation of male reproduction in M. nipponense, which may promote the studies of male reproduction in other crustacean species.