Freshwater Prawn Macrobrachium Nipponense Research Articles

The Dmrt1 gene: Identification and functional analysis in testicular development and spermatogenesis in the freshwater prawn, Macrobrachium nipponense

Products of the Dmrt gene family are known to harbor a zinc-finger DM domain and play crucial roles as transcription factors in various physiological processes, including sexual differentiation, gonadal development, and spermatogenesis. Despite their significance, research on the Dmrt family in Crustacea remains limited. In the present study, we identified a member of this gene family in the freshwater prawn Macrobrachium nipponense and designated it as MnDmrt1. We investigated its expression patterns across different tissues, potential target genes, and its influence on testicular development and protein interactions within the testes. Our findings revealed that MnDmrt1 encodes a protein composed of 252 amino acids that is predominantly expressed in the testes and localized in spermatogonia, spermatocytes, and spermatozoa. Silencing MnDmrt1 disrupts spermatogenesis and alters the expression levels of several key genes associated with testicular development, spermatogenesis, and sex reversal, such as Dmrt11E, Insulin receptor, and Masculinizer. Additionally, we identified 59 proteins that interact with MnDmrt1 in the testis using pull-down assays and liquid chromatography-tandem mass spectrometry. Notably, the eyestalk negatively regulates the expression level of MnDmrt1 in the testis. Collectively, these results underscore the essential role of MnDmrt1 in testicular development and spermatogenesis in M. nipponense.

EDNA metabarcoding of decapod crustaceans across Indonesian seas has implications for biodiversity conservation and fisheries sustainability

Environmental DNA (eDNA) methods are increasingly viewed as alternate or complementary approaches to conventional capture-based surveys for marine conservation and fisheries management purposes, especially at large spatial scales in mega-biodiversity regions such as Indonesia. Decapod crustacean distribution and diversity across Indonesia are still poorly known, even for economically important fisheries commodities. This study assessed coral reef associated decapod diversity and distribution by sampling 40 sites in three regions (West, Central, East), representing 17 provinces and 10 Fisheries Management Areas (FMAs) across Indonesia, with a special focus on the blue swimming crab Portunus pelagicus. DNA sequencing (Illumina iSeq100) data were analysed in mBRAVE (Multiplex Barcode Research And Visualization Environment) yielded 406 OTUs belonging to 32 families, with 47 genera and 51 species identified. The number of families identified was highest in the Central region (25), while the most genera (31) and species (36) were identified in the West region. Alpha diversity did not differ significantly between regions or provinces, while Beta diversity differed significantly between provinces but not between regions. Our results also showed 31 species are possibility native based on the distribution meanwhile 12 species do not appear to have been recorded based of SeaLifeBase or WorMS. While providing a reference for further exploration of Indonesian coastal and small island decapod biodiversity, the high proportion of unidentified taxa calls for concerted efforts to develop and maintain reference specimen and sequence repositories and expand species conservation status assessments. The economically important decapod crustaceans identified in this study included three crabs (Charybdis anisodon, Charybdis japonica, Portunus pelagicus), a freshwater prawn (Macrobrachium nipponense), a lobster (Panulirus stimpsoni) and two penaeid shrimps (Mierspenaeopsis hardwickii and Trachysalambria aspera). For most decapod taxa, observed patterns indicate management under existing provincial and/or FMA level management structures is appropriate. Furthermore, the data can inform science-based fisheries management strategies, in particular for P. pelagicus.

Polystyrene nanoplastics decrease molting and induce oxidative stress in adult Macrobrachium nipponense

The widespread occurrence and accumulation of plastic waste have been globally recognized as a critical issue. However, few researches have evaluated the adverse effects of nanoplastics to freshwater organisms. Thus, here, the effects of polystyrene nanoplastics (PS-NP) on the physiological changes (i.e., molting) and enzyme activity of oxidative stress were investigated in the adult freshwater prawn Macrobrachium nipponense. Based on a previous study and environmental microplastic concentrations, the prawn was exposed to 0, 0.04, 0.4, 4, and 40 mg/L waterborne PS-NP for 21 days. The results showed that growth and survival-related parameters were not affected by all PS-NP groups, while the molting rate were significantly decreased in the 4 and 40 mg/L PS-NP group. Meanwhile, the expression of molting-related gene (calcium–calmodulin-dependent protein kinase I, ecdysteroid receptor, and leucine-rich repeat-containing G-protein-coupled receptor 2) were significantly decreased. H2O2 content was significantly increased in all PS-NP groups relative to the control. Lower concentrations of PS-NP increased the activity of superoxide dismutase (SOD), glutathione S-transferase (GST), and glutathione peroxidase (GSH-Px), whereas increased concentrations, decreased SOD, GST, and GSH-Px activity. These results suggest that chronic exposure to PS-NP at an environmental concentration impaired molting and induced oxidative stress in the adult river prawn Macrobrachium nipponense. The findings provided basic information for assessing the risk assessment of nanoplastics and revealing the molecular mechanisms of nanoplastics toxicity.

Pond culture of the freshwater prawn Macrobrachium nipponense in China

This datasheet on Pond culture of the freshwater prawn Macrobrachium nipponense in China covers Identity, Overview, Environmental Requirements, Growout Systems, Related Content, Further Information.

The alleviative effects of taurine supplementation on growth, antioxidant enzyme activities, hepatopancreas morphology and mRNA expression of heat shock proteins in freshwater prawn Macrobrachium nipponense (De Haan) exposed to dietary lead stress

To investigate the potential alleviative effect of taurine supplementation on growth and oxidant stress in freshwater prawn Macrobrachium nipponense (De Haan) exposed to dietary lead, pelleted feeds were prepared with two concentrations of lead (0 and 10 mg/kg diet) and two levels of taurine (0 and 15 g/kg diet). Healthy prawns were randomly distributed between the four treatment groups, with three tanks per group and 50 prawns per tank. Dietary lead decreased weight gain and the activities of superoxide dismutase, catalase and glutathione peroxidase; increased malondialdehyde content; altered hepatopancreas morphology; and decreased the expression of heat shock protein 90 (HSP90; p < .05). A significant interactive effect was found between dietary lead and taurine on the malondialdehyde level, and taurine supplementation significantly decreased the malondialdehyde level under lead stress (p < .05). No significant effect on growth, hepatopancreas morphology and HSP90 expression was observed with taurine supplementation under lead stress. Overall, dietary lead affected growth, induced oxidative stress and impaired hepatopancreas morphology of the prawns, and dietary taurine significantly alleviated oxidative stress through a decreased malondialdehyde level, whereas the capacity of taurine to alleviate the decreased growth rate, hepatopancreas damage and low HSP90 expression under lead stress was limited.

A newly identified Hippo homologue from the oriental river prawn Macrobrachium nipponense is involved in the antimicrobial immune response

The Hippo signalling pathway plays a vital role in organ size control, cell proliferation, apoptosis, and immune regulation. In this study, a Hippo homologue with three isoforms (named MnHippo-a, MnHippo-b, and MnHippo-c) was isolated and characterized for the first time from the freshwater prawn Macrobrachium nipponense. The deduced amino acid sequences of MnHippo-a (698 aa), MnHippo-b (688 aa), and MnHippo-c (656 aa) were highly similar, and they all contained an N-terminal S_TKc (serine/threonine protein kinase catalytic) domain and a C-terminal Mst1_SARAH (Sav/Rassf/Hpo) domain. MnHippo-a and MnHippo-c were derived from alternative splicing. Phylogenetic analysis was performed, and the results revealed that MnHippo was a member of the clade containing STPK4 and Hippo of Penaeus vannamei. The expression distribution showed that MnHippo was constitutively expressed in various tissues of uninfected prawns and highly expressed in the hepatopancreas and intestine. In prawns challenged with Vibrio parahaemolyticus and Staphylococcus aureus, the expression of MnHippo in haemocytes was significantly upregulated. Furthermore, in MnHippo-knockdown prawns injected with V. parahaemolyticus or S. aureus, the transcription levels of five antimicrobial peptides were downregulated. MnHippo silencing weakened the clearance of V. parahaemolyticus and S. aureus in prawns. The survival rate of the MnHippo-dsRNA group was obviously decreased from 2 to 6 days post-injection with V. parahaemolyticus or S. aureus. Hence, MnHippo might be involved in the antibacterial immune defence of M. nipponense.

Anthraquinone extract from Rheum officinale Bail improves growth performance and Toll-Relish signaling-regulated immunity and hyperthermia tolerance in freshwater prawn Macrobrachium nipponense.

This study was aimed to investigate the facilitation of anthraquinone extract on growth performance, immunity, and antioxidant capacity of the oriental river prawn (Macrobrachium nipponense), and whether it could ameliorate the hyperthermia stress. A 12-week rearing experiment was conducted with 0, 125, 250, 500, and 1000mgkg-1 anthraquinone extract from Rheum officinale Bail-supplemented diets (AE0, AE125, AE250, AE500, and AE1000), and followed a 48-h thermal stress with 32℃ incubation. Results indicate AE250 and AE500 significantly improved the growth performance and feed utilization, the optimum level was estimated to be 251.22mgkg-1 based on the regression analysis of special growth ratio (SGR). Meanwhile, AE250 and AE500 improved antioxidant enzyme activity and immune-related protein concentration of iNOS-NO signaling. Under thermal stress, AE250 and AE500 improved the heat tolerance, and Toll-Relish signaling was active to the resistance. These results indicate anthraquinone extract could be used as an effective immunostimulant to improve growth performance, physiological balance and protect organism form environmental hyperthermia stress. This may provide insights for immunostimulant development in aquaculture production.

Recombinant expression of an oriental river prawn anti-lipopolysaccharide factor gene in Pichia pastoris and its characteristic analysis

Anti-lipopolysaccharide factors (ALFs), as main host-defense molecules of crustaceans, form a unique family of antimicrobial peptides (AMPs). MnALF4 is one isoform of ALFs isolated from the freshwater prawn Macrobrachium nipponense. In the present study, MnALF4 gene was successfully expressed in the yeast Pichia pastoris and the recombinant MnALF4 protein exhibited efficient and broad-spectrum antimicrobial activities against both Gram-positive bacteria and Gram-negative bacteria in vitro. When prawns were injected with rMnALF4 before bacterial challenge with E. coli, the recombinant protein effectively promote the elimination of bacteria by the host. It manifested that rMnALF4 could effectively kill the invading bacteria in vivo. Treatment with rMnALF4 led to remarkable changes in bacterial morphology, such as spheroidization, oversized bacteria, and cell lysis. In addition, rMnALF4 showed weak hemolysis activity to the rabbit red blood cells. Our work suggests that MnALF4 plays an important role in Macrobrachium immunity and is worthy of further investigation as a potential antibacterial agent with high efficacy against bacterial infection and low toxicity to host cells.

Effects of temperature and tank color on post larvae release and survival of freshwater Prawn Macrobrachium nipponense

Berried females of Macrobrachium nipponense were incubated under four different temperatures (20,23,26 and 28 ºC) and two types of tanks with different background coloration (dark and transparent tanks) to determine the best larval release and the post larvae survival were observed. The results show best larval release were 124 ind./L achieved at 26 ºC at dark color tanks, while 68 ind. / L larvae were released at the transparent color tank at the same temperature, while only 16 ind./L were released from berried females incubated at 20 ºC at dark color tanks and 6 ind./L at the transparent tanks. The best survival of larvae was 80 % at 23 ºC at dark color tanks and 75 % at transparent color tanks. Four stages for eggs development were discriminated based on color, size and eyes.

Effects of sodium thiosulfate on survival and molting of larvae of the freshwater prawn Macrobrachium nipponense

Effects of sodium thiosulfate on survival and molting of larvae of the freshwater prawn Macrobrachium nipponense

Molecular cloning and expression analysis of Fem1b from oriental river prawn Macrobrachium nipponense.

Feminization-1 homolog b (Fem1b) is one of the genes essential for male development and play central roles in sex determination of Caenorhabditis elegans. In this study, we cloned and characterized the full-length Fem1b cDNA from the freshwater prawn Macrobrachium nipponense (MnFem1b) in different tissues and at different developmental stages. Real-time quantitative reverse polymerase chain reaction (RT-qPCR) showed that the MnFem1b gene was expressed in all investigated tissues, with the highest expression level found in the testes. The results revealed that the MnFem1b gene might play roles in aspects of development of the male prawn phenotype. The RT-qPCR also revealed that MnFem1b mRNA expression was significantly increased at 10 days after metamorphosis. The expression levels in all investigated tissues showed a certain degree of sexually dimorphism, the expression levels in males were significantly higher than those in females (P < 0.05). Notably, the highest expression of MnFem1b was found in the testes. The expression of MnFem1b in different tissues indicates that it plays multiple biological functions in M. nipponense.

Characterization of the male-specific lethal 3 gene in the oriental river prawn, Macrobrachium nipponense.

In this study, male-specific lethal 3 homolog (Mnmsl3) was cloned and characterized from the freshwater prawn Macrobrachium nipponense (Crustacea: Decapoda: Palaemonidae) by rapid amplification of cDNA ends. The deduced amino acid sequences of Mnmsl3 showed high-sequence homology to the insect Msl3 and contained a conserved chromatin organization modifier domain and an MORF4-related gene domain. Real-time quantitative reverse transcription-polymerase chain reaction showed that the Mnmsl3 gene was expressed in all the investigated tissues, with the highest level of expression in the testis. The expression level of Mnmsl3 between males and females was different in the gonad (testis or ovary), abdominal ganglion, and heart. The results revealed that the Mnmsl3 gene might play roles in regulating chromatin and in dosage compensation of M. nipponense. Real-time quantitative reverse transcription-polymerase chain reaction also revealed that Mnmsl3 mRNA expression was significantly increased in both 5 and 20 days post-larvae after metamorphosis, suggesting that Mnmsl3 plays complex and important roles in the early embryonic development and sex differentiation of M. nipponense.

A clip-domain serine proteinase homolog (SPH) in oriental river prawn, Macrobrachium nipponense provides insights into its role in innate immune response

In this study, a clip-domain serine proteinase homolog designated as MnSPH was cloned and characterized from a freshwater prawn Macrobrachium nipponense. The full-length cDNA of MnSPH was 1897 bp and contained a 1701 bp open reading frame (ORF) encoding a protein of 566 amino acids, a 103 bp 5′-untranslated region, and a 93 bp 3′-untranslated region. Sequence comparison showed that the deduced amino acids of MnSPH shared 30–59% identity with sequences reported in other animals. Tissue distribution analysis indicated that the MnSPH transcripts were present in all detected tissues with highest in the hepatopancreas and ovary. The MnSPH mRNA levels in the developing ovary were stable at the initial three developmental stages, then increased gradually from stage IV (later vitellogenesis), and reached a maximum at stage VI (paracmasis). Furthermore, the expression of MnSPH mRNA in hemocytes was significantly up-regulated at 1.5 h, 6 h, 12 h and 48 h post Aeromonas hydrophila injection. The increased phenoloxidase activity also demonstrated a clear time-dependent pattern after A. hydrophila challenge. These results suggest that MnSPH participates in resisting to pathogenic microorganisms and plays a pivotal role in host defense against microbe invasion in M. nipponense.

Calcium–calmodulin dependent protein kinase I from Macrobrachium nipponense: cDNA cloning and involvement in molting

Calcium–calmodulin dependent protein kinase I is a component of a calmodulin-dependent protein kinase cascade and involved in many physiological processes. The full-length cDNA of calcium–calmodulin dependent protein kinase I (MnCaMKI) was cloned from the freshwater prawn Macrobrachium nipponense and its expression pattern during the molt cycle and after eyestalk ablation is described. The full-length cDNA of MnCaMKI is 3262bp in length and has an open reading frame (ORF) of 1038bp, encoding a 345 amino acid protein. The expression of MnCaMKI in three examined tissues was upregulated in the premolt stage of the molt cycle. Its expression was induced after eyestalk ablation (ESA): the highest expression level was reached 1day after ESA in hepatopancreas, and 3days after ESA in muscle. By dsRNA-mediated RNA interference assay, expression of MnCaMKI and ecydone receptor gene (MnEcR) was significantly decreased in prawns treated by injection of dsMnCaMKI, while expression of these two genes was also significantly decreased in prawns treated by injection of dsMnEcR, demonstrating a close correlation between the expression of these two genes. These results suggest that CaMKI in M. nipponense is involved in molting.

Artificial infection of the native Korean freshwater prawn Macrobrachium nipponense (De Haan, 1849) (Decapoda, Palaemonidae) with white spot syndrome virus (WSSV)

White spot syndrome virus (WSSV), an important shrimp pathogen that can affect both salt- and freshwater shrimp, has a high mortality rate that can reach up to 100% in 3-10 days. Therefore, we assessed the susceptibility of Macrobrachium nipponense (De Haan, 1849), a freshwater prawn native to Korea, to artificial WSSV infection. Although rearing in water containing WSSV-infected carcasses did not result in infection, M. nipponense was infected with WSSV after being fed a WSSV-containing diet, per os infection, or injection with the purified virus. In the feeding test, the first mortality was observed on the 7th day and cumulative mortality reached 30% by 3 weeks post-infection. After per os infection, experimental animals started dying on the second day and within 3 weeks mortality had reached 90%. In the injection test, the mortality was found to be dose-dependent. Prawn injected with 2.5 × 104, 900 and 80 copies/g body weight exhibited the first mortality at 1, 2 and 4 days post inoculation, with 100, 100 and 50% final mortality at 2, 10 and 12 days post inoculation, respectively. Histological observation revealed inclusion bodies in the gills, antennal glands and other tissues of ectodermal origin. Real-time polymerase chain reaction revealed the increase of viral titers, indicating viral replication in the inoculated host. This forms the first report of the experimental susceptibility of M. nipponense to WSSV.

Identification of spliced mRNA isoforms of retinoid X receptor (RXR) in the Oriental freshwater prawn Macrobrachium nipponense.

Retinoid X receptors (RXR) are members of the nuclear receptor family that are conserved from invertebrates to vertebrates, and they play an essential role in regulating reproductive maturation, molting, and embryo development. In this study, five RXR isoforms, named RXRL2 (L, long form), RXRL3, RXRS1 (S, short form), RXRS2, and RXRS3, containing six domains from A to F, were cloned from the prawn Macrobrachium nipponense using 5'- and 3'- rapid amplification of cDNA ends. Differences among their structures were observed not only in the D and E domains but also in the A/B domain, which were previously found in insects but not in crustaceans. This is the first report to show that differences occur in the A/B domain of RXR in crustaceans. RXR expressions were also examined in various tissues including the ovary, testis, muscle, hepatopancreas, heart, gill, stomach, intestine, and cuticle. Expression pattern investigations indicated that the five isoforms were differentially expressed. RXRS3 was only detected in the ovary, and the other RXRs were abundant in the ovary and testis. These data suggested that RXR mediates a series of processes related to reproduction.

Molecular cloning, characterization and expression analysis of the fatty acid-binding protein (MnFABP), involved in dietary lipid sources response in oriental river prawn,Macrobrachium nipponense

A fatty acid-binding protein (FABP) gene designated as MnFABP10 was cloned and characterized from the freshwater prawn Macrobrachium nipponense. The full-length cDNA of MnFABP10 was 646 bp encoding a 130 amino acid. Real-time quantitative RT-PCR showed that the MnFABP10 gene was expressed in various tissues with the highest expression in the hepatopancreas. The MnFABP10 mRNA levels in the hepatopancreas and ovary of M. nipponense were dependent on the stages of ovarian development. Western blot results revealed a single immunoreactive band with an estimated molecular mass of approximate 14 kDa in the developmental ovary. Then, M. nipponense with an initial body weight of 0.090 ± 0.0010 g were fed with four isonitrogenous and isocaloric diets with different oils, that is, beef tallow (BT), soybean oil (SO), pollack fish oil (FO) and a mixture of fish oil and soybean oil (FO/SO 2 : 1 w/w) for 52 days. The mRNA levels of MnFABP10 in the hepatopancreas were influenced by different lipid sources, with a peak expression observed in prawns fed SO. This study suggests that MnFABP10 may have a putative function in ovary maturation, and its mRNA expression in the hepatopancreas can be regulated by the source of dietary lipids in M. nipponense.

Effect of nitrite exposure on metabolic response in the freshwater prawn Macrobrachium nipponense

Abstract The metabolic response of the freshwater prawn, Macrobrachium nipponense to nitrite toxicity was evaluated. The prawns were exposed to 0, 1, 2, 3 and 4 mg L−1 NO2-N concentrations for 48 h. The metabolic parameters in muscle were measured after 12, 24 and 48 h. Glucose level significantly increased after 24 h. Exposure to lower nitrite concentrations (1 and 2 mg L−1) resulted in significant increases in alanine aminotransferase (ALT) activities after 24 and 48 h. Aspartate aminotransferase (AST) activities treated with 2 and 3 mg L−1 nitrite-N at 48 h were significantly higher than those at 12 and 24 h. Intermediate sublethal nitrite concentrations produced significant elevations in lactate dehydrogenase (LDH) activities from 12 h up to 48 h. No significant changes were detected in any of the groups for triglycerides and creatine kinase (CK). To satisfy the increased energy demands caused by acute nitrite exposure, mobilization of lipids is not the main reason while utilization of amino acids seems to play a more important role. The results would be helpful for aquaculture farmers to prevent a potential depression of productivity caused by elevated nitrite levels.

Ecdysone receptor gene from the freshwater prawn Macrobrachium nipponense: Identification of different splice variants and sexually dimorphic expression, fluctuation of expression in the molt cycle and effect of eyestalk ablation

The full-length cDNA of an ecdysone receptor gene (MnEcR) from Macrobrachium nipponense was cloned and the expression of the gene was investigated. MnEcR maintained a relatively low expression level in the early stages of embryos, but from nauplius stage, a steady increase in MnEcR expression was detected, it had the highest expression level in zoea stage. MnEcR was highly expressed in the hepatopancreas and gills among ten different tissues examined. MnEcR was rapidly upregulated in the premolt stage and rapidly downregulated in the postmolt stage. The expression of MnEcR was remarkably downregulated after eyestalk ablation in M. nipponense. An 18-amino-acid insertion/deletion and a 49-amino-acid substitution were found in the coding region of MnEcR, resulting in four splice variants: MnEcR-L1, -L2, -S1 and-S2. The expression of four splice variants of MnEcR in gonads was investigated using RT-PCR. Interestingly, the expression patterns of these splice variants differed between males and females. The dominant splice variants in testis were MnEcR-S1 and -S2, while in ovary they were MnEcR-L1 and -S2, indicating specific roles for these splice variants in male and female individuals.

The retinoid-X receptor gene from the freshwater prawn Macrobrachium nipponense: cloning, expression pattern and different responses of two splice variants during the molting cycle

The retinoid-X receptor (RXR) is among the most conserved members of the nuclear receptor superfamily and is widely studied in vertebrate and invertebrate families. RXR plays an important role in regulation of molting and/or metamorphosis, development and reproduction. We cloned the full-length cDNA sequence of the RXR from the freshwater prawn Macrobrachium nipponense (De Haan, 1849) (MnRXR) and investigated the expression profile of MnRXR in different developmental stages of embryos, in different tissues and in the molting cycle. Two MnRXR splice variants were identified: One, MnRXR-L, the full length of which was 2472 bp, encoded a 449-amino-acid protein; the second, MnRXR-S, the full length of which was 1832 bp, encoded a 420-amino-acid protein, in which the first 29 amino-acid residues of MnRXR-L were absent. MnRXR was observed in all developmental stages of embryos and had the highest expression level in the embryonised-zoea stage, it was highly expressed in hepatopancreas, gill and intestine among the ten tissues examined. The expression of MnRXR was rapidly up-regulated in the premolt stage and rapidly down-regulated after molting. Moreover, of the two MnRXR splice variants, only MnRXR-S was induced during the molting cycle, suggesting that the two splice variants play different roles in the molting cycle.